[Comparison of clonal architecture between two divergent Leymus chinensis types in Songnen grassland].

PubMed

He, Nianpeng; Wu, Ling; Zhou, Daowei

2004-12-01

This paper studied the clonal architecture of two divergent Leymus chinensis types (grey-green type and yellow-green type) in Songnen grassland, and compared their internode length, spacer length, interbranching length, interbranching angle, and ramet population density and height under the same habitat. The results showed that there was no significant difference in these clonal characteristics except spacer length and ramet population density between the two types of L. chinensis, and yellow-green type, with less spacer length and more ramet density than grey-green type, should be more adaptable to the resourceful habitat. Moreover, the V-indices of the clonal architecture of two divergent L. chinensis types were all close to 1, and the difference was not significant. Therefore, both of the two types belonged to typical guerilla clonal plant.

[Multilocus Sequence Typing analysis of human Campylobacter coli in Granada (Spain)].

PubMed

Carrillo-Ávila, J A; Sorlózano-Puerto, A; Pérez-Ruiz, M; Gutiérrez-Fernández, J

2016-12-01

Different subtypes of Campylobacter spp. have been associated with diarrhoea and a Multilocus Sequence Typing (MLST) method has been performed for subtyping. In the present work, MLST was used to analyse the genetic diversity of eight strains of Campylobacter coli. Nineteen genetic markers were amplified for MLST analysis: AnsB, DmsA, ggt, Cj1585c, CJJ81176-1367/1371, Tlp7, cj1321-cj1326, fucP, cj0178, cj0755/cfrA, ceuE, pldA, cstII, cstIII. After comparing the obtained sequences with the Campylobacter MLST database, the allele numbers, sequence types (STs) and clonal complexes (CCs) were assigned. The 8 C. coli isolates yielded 4 different STs belonging to 2 CCs. Seven isolates belong to ST-828 clonal complex and only one isolate belong to ST-21. Two samples came from the same patient, but were isolated in two different periods of time. MLST can be useful for taxonomic characterization of C. coli isolates.

Molecular genotyping of Toxoplasma gondii from Central and South America revealed highly diverse populations and suggested possible different origins of the three archetypal lineages

USDA-ARS?s Scientific Manuscript database

Most T. gondii strains in North America and Europe belong to three archetypal clonal lineages including the Type I, II and III but, isolates from Brazil are highly diverse. Here, we analyzed 164 T. gondii isolates from three countries in Central America (Guatemala, Nicaragua, Costa Rica), from one c...

Variation in biological properties of cauliflower mosaic virus clones.

PubMed

al-Kaff, N; Covey, S N

1994-11-01

Infectious clones were prepared from virion DNA of three cauliflower mosaic virus (CaMV) isolates, 11/3, Xinjiang (XJ), and Aust, to investigate pathogenic variation in virus populations. Of 10 infectious clones obtained for isolate 11/3, four pathotypes were identified, each producing symptoms in turnip that differed from those of the 11/3 wild-type. Virus from two clonal groups of 11/3 was transmissible by aphids whereas that from two others was not. Of the five infectious clones obtained from isolate XJ, two groups were identified, one of which differed symptomatically from the wild-type. Only one infectious clone was obtained from isolate Aust and this had properties similar to the wild-type. Restriction enzyme polymorphisms were found in some clonal groups and these correlated with symptoms. Other groups with different pathogenic properties could not be distinguished apart by restriction site polymorphisms. Further variation was observed in the nucleotide sequences of gene II (coding for aphid transmission factor) from these viruses as compared with other CaMV isolates. In the aphid non-transmissible clones of isolate 11/3, one had a Gly to Arg mutation in gene II similar to that of other non-deleted non-transmissible CaMV isolates. The second had a 322 bp deletion at the site of a small direct repeat similar to that of isolate CM4-184 although occurring in a different position. The gene II deletion of isolate 11/3 produced a frame-shift that separated genes II and III by 60 bp. Most CaMV clones studied remained biologically stable producing similar symptoms during subsequent passages. However, one clone (11/3-7) produced two new biotypes during its first passage suggesting that it was relatively unstable. Our results show that wild-type populations of CaMV contain a range of infectious genome variants with contrasting biological properties and differing stability. We suggest that a variety of significant viral phenotypic changes can occur during each infection cycle resulting from relatively small genome changes.

Shifts in the Clonal Distribution of Methicillin-Resistant Staphylococcus aureus in Kuwait Hospitals: 1992-2010

PubMed Central

Boswihi, Samar S.; Udo, Edet E.; Al-Sweih, Noura

2016-01-01

Background As the epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) is constantly changing globally, determining the prevailing MRSA clones in a local healthcare facility is important for better management of infections. This study investigated clonal composition and distribution of MRSA isolates in Kuwait’s hospitals using a combination of molecular typing methods. Materials and Methods In total, 400 non-repeat MRSA isolates were obtained between 1992 and 2010 in 13 public hospitals and were characterized using antibiogram, SCCmec typing, spa typing, and multilocus-sequence typing. Clonal assignment and detection of virulence factors and antibiotic resistance genes were performed by DNA microarray. Results The isolates were resistant to kanamycin (74.2%), erythromycin (69.5%), tetracycline (66.7%), gentamicin (61%), ciprofloxacin, (61%), fusidic acid (53.5%), clindamycin (41.5%), high-level mupirocin resistance (5.2%) and carried aphA3, aacA-aphD, ermA, ermC, mupA, tetK, tetM, fusC and far1. Molecular typing revealed 31 different MRSA clones consisting of ST239-MRSA-III (52.2%), ST22-MRSA-IV (9.2%), ST80-MRSA-IV (7.5%), ST5-MRSA-II/IV/V/VI (6.5%), ST30-MRSA-IV (3.5%), ST241-MRSA-III (2.7%), ST6-MRSA-IV (2.2%), ST36-MRSA-II (2%) and ST772-MRSA-V (1.75%). The isolates differed in the carriage of genes for enterotoxins, Panton–Valentine leukocidin (PVL), toxic shock syndrome toxin (tst-1), arginine catabolic mobile element (ACME) and exfoliative toxins. The number of clones increased from one (ST239-III-t037) in 1992 to 30 in 2010 including ST8-IV-t008 [PVL+] [ACME+] (USA300), ST772-V (Bengal Bay clone) and ST2816 identified for the first time in Kuwait. Conclusion The study revealed that the MRSA isolates belonged to diverse clones that changed in numbers and diversity overtime. Although ST239-MRSA-III, a healthcare-associated clone remained the dominant MRSA clone overtime, the newly emerged clones consisted mostly of community-associated. PMID:27631623

Extensive clonal spread and extreme longevity in saw palmetto, a foundation clonal plant.

PubMed

Takahashi, Mizuki K; Horner, Liana M; Kubota, Toshiro; Keller, Nathan A; Abrahamson, Warren G

2011-09-01

The lack of effective tools has hampered out ability to assess the size, growth and ages of clonal plants. With Serenoa repens (saw palmetto) as a model, we introduce a novel analytical framework that integrates DNA fingerprinting and mathematical modelling to simulate growth and estimate ages of clonal plants. We also demonstrate the application of such life-history information of clonal plants to provide insight into management plans. Serenoa is an ecologically important foundation species in many Southeastern United States ecosystems; yet, many land managers consider Serenoa a troublesome invasive plant. Accordingly, management plans have been developed to reduce or eliminate Serenoa with little understanding of its life history. Using Amplified Fragment Length Polymorphisms, we genotyped 263 Serenoa and 134 Sabal etonia (a sympatric non-clonal palmetto) samples collected from a 20 × 20 m study plot in Florida scrub. Sabal samples were used to assign small field-unidentifiable palmettos to Serenoa or Sabal and also as a negative control for clone detection. We then mathematically modelled clonal networks to estimate genet ages. Our results suggest that Serenoa predominantly propagate via vegetative sprouts and 10,000-year-old genets may be common, while showing no evidence of clone formation by Sabal. The results of this and our previous studies suggest that: (i) Serenoa has been part of scrub associations for thousands of years, (ii) Serenoa invasion are unlikely and (ii) once Serenoa is eliminated from local communities, its restoration will be difficult. Reevaluation of the current management tools and plans is an urgent task. © 2011 Blackwell Publishing Ltd.

Comparison of the DiversiLab Repetitive Element PCR System with spa Typing and Pulsed-Field Gel Electrophoresis for Clonal Characterization of Methicillin-Resistant Staphylococcus aureus▿

PubMed Central

Babouee, B.; Frei, R.; Schultheiss, E.; Widmer, A. F.; Goldenberger, D.

2011-01-01

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) has become an increasing problem worldwide in recent decades. Molecular typing methods have been developed to identify clonality of strains and monitor spread of MRSA. We compared a new commercially available DiversiLab (DL) repetitive element PCR system with spa typing, spa clonal cluster analysis, and pulsed-field gel electrophoresis (PFGE) in terms of discriminatory power and concordance. A collection of 106 well-defined MRSA strains from our hospital was analyzed, isolated between 1994 and 2006. In addition, we analyzed 6 USA300 strains collected in our institution. DL typing separated the 106 MRSA isolates in 10 distinct clusters and 8 singleton patterns. Clustering analysis into spa clonal complexes resulted in 3 clusters: spa-CC 067/548, spa-CC 008, and spa-CC 012. The discriminatory powers (Simpson's index of diversity) were 0.982, 0.950, 0.846, and 0.757 for PFGE, spa typing, DL typing, and spa clonal clustering, respectively. DL typing and spa clonal clustering showed the highest concordance, calculated by adjusted Rand's coefficients. The 6 USA300 isolates grouped homogeneously into distinct PFGE and DL clusters, and all belonged to spa type t008 and spa-CC 008. Among the three methods, DL proved to be rapid and easy to perform. DL typing qualifies for initial screening during outbreak investigation. However, compared to PFGE and spa typing, DL typing has limited discriminatory power and therefore should be complemented by more discriminative methods in isolates that share identical DL patterns. PMID:21307215

Identification of a high-virulence clone of type III Streptococcus agalactiae (group B Streptococcus) causing invasive neonatal disease.

PubMed

Musser, J M; Mattingly, S J; Quentin, R; Goudeau, A; Selander, R K

1989-06-01

Chromosomal genotypes of 128 isolates of six serotypes (Ia, Ib, Ic, II, Ic/II, and III) of Streptococcus agalactiae (group B Streptococcus) recovered predominantly from human infants in the United States were characterized by an analysis of electrophoretically demonstrable allelic profiles at 11 metabolic enzyme loci. Nineteen distinctive electrophoretic types (ETs), representing multilocus clonal genotypes, were identified. Mean genetic diversity per locus among ETs of isolates of the same serotype was, on average, nearly equal to that in all 19 ETs. Cluster analysis of the ETs revealed two primary phylogenetic divisions at a genetic distance of 0.65. A single clone (ET 1) represented by 40 isolates expressing type III antigen formed division I. Division II was composed of 18 ETs in three major lineages diverging from one another at distances greater than 0.35 and included strains of all six antigenic classes. The type III organisms in division I produce more extracellular neuraminidase and apparently are more virulent than the type III strains in division II, which are related to strains of other serotypes that cause disease much less frequently. The existence of this unusually virulent clone accounts, in major part, for the high morbidity and mortality associated with infection by type III organisms.

Molecular Characterization of Streptococcus agalactiae Isolated from Bovine Mastitis in Eastern China

PubMed Central

Yang, Yongchun; Liu, Yinglong; Ding, Yunlei; Yi, Li; Ma, Zhe; Fan, Hongjie; Lu, Chengping

2013-01-01

One hundred and two Streptococcus agalactiae (group B streptococcus [GBS]) isolates were collected from dairy cattle with subclinical mastitis in Eastern China during 2011. Clonal groups were established by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE), respectively. Capsular polysaccharides (CPS), pilus and alpha-like-protein (Alp) family genes were also characterized by molecular techniques. MLST analysis revealed that these isolates were limited to three clonal groups and were clustered in six different lineages, i.e. ST (sequence type) 103, ST568, ST67, ST301, ST313 and ST570, of which ST568 and ST570 were new genotypes. PFGE analysis revealed this isolates were clustered in 27 PFGE types, of which, types 7, 8, 14, 15, 16, 18, 23 and 25 were the eight major types, comprising close to 70% (71/102) of all the isolates. The most prevalent sequence types were ST103 (58% isolates) and ST568 (31% isolates), comprising capsular genotype Ia isolates without any of the detected Alp genes, suggesting the appearance of novel genomic backgrounds of prevalent strains of bovine S. agalactiae. All the strains possessed the pilus island 2b (PI-2b) gene and the prevalent capsular genotypes were types Ia (89% isolates) and II (11% isolates), the conserved pilus type providing suitable data for the development of vaccines against mastitis caused by S. agalactiae. PMID:23874442

United we stand, divided we fall: a meta-analysis of experiments on clonal integration and its relationship to invasiveness.

PubMed

Song, Yao-Bin; Yu, Fei-Hai; Keser, Lidewij H; Dawson, Wayne; Fischer, Markus; Dong, Ming; van Kleunen, Mark

2013-02-01

Many ecosystems are dominated by clonal plants. Among the most distinctive characteristics of clonal plants is their potential for clonal integration (i.e. the translocation of resources between interconnected ramets), suggesting that integration may play a role in their success. However, a general synthesis of effects of clonal integration on plant performance is lacking. We conducted a meta-analysis on the effects of clonal integration on biomass production and asexual reproduction of the whole clone, the recipient part (i.e. the part of a clone that imports resources) and the donor part (i.e. the part of a clone that exports resources). The final dataset contained 389 effect sizes from 84 studies covering 57 taxa. Overall, clonal integration increased performance of recipient parts without decreasing that of donor parts, and thus increased performance of whole clones. Among the studies and taxa considered, the benefits of clonal integration did not differ between two types of experimental approaches, between stoloniferous and rhizomatous growth forms, between directions of resource translocation (from younger to older ramet or vice versa), or among types of translocated resources (water, nutrients and carbohydrates). Clonal taxa with larger benefits of integration on whole-clone performance were not more invasive globally, but taxa in which recipient parts in unfavorable patches benefited more from integration were. Our results demonstrate general performance benefits of clonal integration, at least in the short term, and suggest that clonal integration contributes to the success of clonal plants.

MRSA Causing Infections in Hospitals in Greater Metropolitan New York: Major Shift in the Dominant Clonal Type between 1996 and 2014.

PubMed

Pardos de la Gandara, Maria; Curry, Marie; Berger, Judith; Burstein, David; Della-Latta, Phyllis; Kopetz, Virgina; Quale, John; Spitzer, Eric; Tan, Rexie; Urban, Carl; Wang, Guiqing; Whittier, Susan; de Lencastre, Herminia; Tomasz, Alexander

2016-01-01

A surveillance study in 1996 identified the USA100 clone (ST5/SCCmecII)-also known as the "New York/Japan" clone-as the most prevalent MRSA causing infections in 12 New York City hospitals. Here we update the epidemiology of MRSA in seven of the same hospitals eighteen years later in 2013/14. Most of the current MRSA isolates (78 of 121) belonged to the MRSA clone USA300 (CC8/SCCmecIV) but the USA100 clone-dominant in the 1996 survey-still remained the second most frequent MRSA (25 of the 121 isolates) causing 32% of blood stream infections. The USA300 clone was most common in skin and soft tissue infections (SSTIs) and was associated with 84.5% of SSTIs compared to 5% caused by the USA100 clone. Our data indicate that by 2013/14, the USA300 clone replaced the New York/Japan clone as the most frequent cause of MRSA infections in hospitals in Metropolitan New York. In parallel with this shift in the clonal type of MRSA, there was also a striking change in the types of MRSA infections from 1996 to 2014.

Clonal evolution of chemotherapy-resistant urothelial carcinoma.

PubMed

Faltas, Bishoy M; Prandi, Davide; Tagawa, Scott T; Molina, Ana M; Nanus, David M; Sternberg, Cora; Rosenberg, Jonathan; Mosquera, Juan Miguel; Robinson, Brian; Elemento, Olivier; Sboner, Andrea; Beltran, Himisha; Demichelis, Francesca; Rubin, Mark A

2016-12-01

Chemotherapy-resistant urothelial carcinoma has no uniformly curative therapy. Understanding how selective pressure from chemotherapy directs the evolution of urothelial carcinoma and shapes its clonal architecture is a central biological question with clinical implications. To address this question, we performed whole-exome sequencing and clonality analysis of 72 urothelial carcinoma samples, including 16 matched sets of primary and advanced tumors prospectively collected before and after chemotherapy. Our analysis provided several insights: (i) chemotherapy-treated urothelial carcinoma is characterized by intra-patient mutational heterogeneity, and the majority of mutations are not shared; (ii) both branching evolution and metastatic spread are very early events in the natural history of urothelial carcinoma; (iii) chemotherapy-treated urothelial carcinoma is enriched with clonal mutations involving L1 cell adhesion molecule (L1CAM) and integrin signaling pathways; and (iv) APOBEC-induced mutagenesis is clonally enriched in chemotherapy-treated urothelial carcinoma and continues to shape the evolution of urothelial carcinoma throughout its lifetime.

Clonal Evolution of Chemotherapy-resistant Urothelial Carcinoma

PubMed Central

Faltas, Bishoy M.; Prandi, Davide; Tagawa, Scott T.; Molina, Ana M.; Nanus, David M.; Sternberg, Cora; Rosenberg, Jonathan; Mosquera, Juan Miguel; Robinson, Brian; Elemento, Olivier; Sboner, Andrea; Beltran, Himisha; Demichelis, Francesca; Rubin, Mark A.

2017-01-01

Chemotherapy-resistant urothelial carcinoma (UC) has no uniformly curative therapy. Understanding how selective pressure from chemotherapy directs UC’s evolution and shapes its clonal architecture is a central biological question with clinical implications. To address this question, we performed whole-exome sequencing and clonality analysis of 72 UCs including 16 matched sets of primary and advanced tumors prospectively collected before and after chemotherapy. Our analysis provided several insights: (i) chemotherapy-treated UC is characterized by intra-patient mutational heterogeneity and the majority of mutations are not shared, (ii) both branching evolution and metastatic spread are very early events in the natural history of UC; (iii) chemotherapy-treated UC is enriched with clonal mutations involving L1-cell adhesion molecule (L1CAM) and integrin signaling pathways; (iv) APOBEC induced-mutagenesis is clonally-enriched in chemotherapy-treated UC and continues to shape UC’s evolution throughout its lifetime. PMID:27749842

Genetic diversity among sea otter isolates of Toxoplasma gondii

USGS Publications Warehouse

Sundar, N.; Cole, Rebecca A.; Thomas, N.J.; Majumdar, D.; Dubey, J.P.; Su, C.

2008-01-01

Sea otters (Enhydra lutris) have been reported to become infected with Toxoplasma gondiiand at times succumb to clinical disease. Here, we determined genotypes of 39 T. gondiiisolates from 37 sea otters in two geographically distant locations (25 from California and 12 from Washington). Six genotypes were identified using 10 PCR-RFLP genetic markers including SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico, and by DNA sequencing of loci SAG1 and GRA6 in 13 isolates. Of these 39 isolates, 13 (33%) were clonal Type II which can be further divided into two groups at the locus Apico. Two of the 39 isolates had Type II alleles at all loci except a Type I allele at locus L358. One isolate had Type II alleles at all loci except the Type I alleles at loci L358 and Apico. One isolate had Type III alleles at all loci except Type II alleles at SAG2 and Apico. Two sea otter isolates had a mixed infection. Twenty-one (54%) isolates had an unique allele at SAG1 locus. Further genotyping or DNA sequence analysis for 18 of these 21 isolates at loci SAG1 and GRA6 revealed that there were two different genotypes, including the previously identified Type X (four isolates) and a new genotype named Type A (14 isolates). The results from this study suggest that the sea otter isolates are genetically diverse.

Host-induced genome alterations in Phytophthora ramorum, I. NA1 lineage on coast live oak in California, II. EU1 lineage on Chamaecyparis lawsoniana in UK

Treesearch

Takao Kasuga; Mai Bui; Elizabeth Bernhardt; Tedmund Swiecki; Kamyar Aram; Lien Bertier; Jennifer Yuzon; Liliana M. Cano; Joan Webber; Clive Brasier; Caroline Press; Niklaus Grünwald; David Rizzo; Matteo Garbelotto

2017-01-01

Rapid phenotypic diversification in clonal invasive populations is often observed, although the underlying genetic mechanisms remain elusive. Lineages of the sudden oak death pathogen Phytophthora ramorum are exclusively clonal, yet isolates of the NA1 lineage from oak (Quercus spp.) frequently exhibit...

A genotyping protocol for multiple tissue types from the polyploid tree species Sequoia sempervirens (Cupressaceae)1

PubMed Central

Narayan, Lakshmi; Dodd, Richard S.; O’Hara, Kevin L.

2015-01-01

Premise of the study: Identifying clonal lineages in asexually reproducing plants using microsatellite markers is complicated by the possibility of nonidentical genotypes from the same clonal lineage due to somatic mutations, null alleles, and scoring errors. We developed and tested a clonal identification protocol that is robust to these issues for the asexually reproducing hexaploid tree species coast redwood (Sequoia sempervirens). Methods: Microsatellite data from four previously published and two newly developed primers were scored using a modified protocol, and clones were identified using Bruvo genetic distances. The effectiveness of this clonal identification protocol was assessed using simulations and by genotyping a test set of paired samples of different tissue types from the same trees. Results: Data from simulations showed that our protocol allowed us to accurately identify clonal lineages. Multiple test samples from the same trees were identified correctly, although certain tissue type pairs had larger genetic distances on average. Discussion: The methods described in this paper will allow for the accurate identification of coast redwood clones, facilitating future studies of the reproductive ecology of this species. The techniques used in this paper can be applied to studies of other clonal organisms as well. PMID:25798341

A genotyping protocol for multiple tissue types from the polyploid tree species Sequoia sempervirens (Cupressaceae).

PubMed

Narayan, Lakshmi; Dodd, Richard S; O'Hara, Kevin L

2015-03-01

Identifying clonal lineages in asexually reproducing plants using microsatellite markers is complicated by the possibility of nonidentical genotypes from the same clonal lineage due to somatic mutations, null alleles, and scoring errors. We developed and tested a clonal identification protocol that is robust to these issues for the asexually reproducing hexaploid tree species coast redwood (Sequoia sempervirens). Microsatellite data from four previously published and two newly developed primers were scored using a modified protocol, and clones were identified using Bruvo genetic distances. The effectiveness of this clonal identification protocol was assessed using simulations and by genotyping a test set of paired samples of different tissue types from the same trees. Data from simulations showed that our protocol allowed us to accurately identify clonal lineages. Multiple test samples from the same trees were identified correctly, although certain tissue type pairs had larger genetic distances on average. The methods described in this paper will allow for the accurate identification of coast redwood clones, facilitating future studies of the reproductive ecology of this species. The techniques used in this paper can be applied to studies of other clonal organisms as well.

Emergence of carbapenem non-susceptible multidrug resistant Acinetobacter baumannii strains of clonal complexes 103(B) and 92(B) harboring OXA-type carbapenemases and metallo-β-lactamases in Southern India.

PubMed

Saranathan, Rajagopalan; Vasanth, Vaidyanathan; Vasanth, Thamodharan; Shabareesh, Pidathala Raghavendra Venkata; Shashikala, P; Devi, Chandrakesan Sheela; Kalaivani, Ramakrishnan; Asir, Johny; Sudhakar, Pagal; Prashanth, K

2015-05-01

The molecular epidemiology and carbapenem resistance mechanisms of clinical isolates of Acinetobacter baumannii obtained from a south Indian tertiary care hospital were investigated by repetitive extragenic palindromic sequence PCR (REP-PCR) and multi-locus sequence typing (MLST). Analysis of resistant determinants was achieved by PCR screening for the presence of genes encoding OXA-carbapenemases, metallo-β-lactamases (MBLs) and efflux pumps. REP-PCR generated around eight clusters of high heterogeneity; of these, two major clusters (I and V) appeared to be clonal in origin. Analysis of representative isolates from different clusters by MLST revealed that most of the isolates belonged to sequence type 103 of CC103(B) . Second most prevalent ST belonged to clonal complex (CC) 92(B) which is also referred to as international clone II. Most of the isolates were multi-drug resistant, being susceptible only to polymyxin-B and newer quinolones. Class D β-lactamases such as blaOXA-51-like (100%), blaOXA-23-like (56.8%) and blaOXA-24-like (14.8%) were found to be predominant, followed by a class B β-lactamase, namely blaIMP-1 (40.7%); none of the isolates had blaOXA-58 like, blaNDM-1 or blaSIM-1 . Genes of efflux-pump adeABC were predominant, most of isolates being biofilm producers that were PCR-positive for autoinducer synthase gene (>94%). Carbapenem non-susceptible isolates were highly diverse and present throughout the hospital irrespective of type of ward or intensive care unit. Although previous reports have documented diverse resistant mechanisms in A. baumannii, production of MBL and OXA-type of carbapenamases were found to be the predominant mechanism(s) of carbapenem resistance identified in strains isolated from Southern India. © 2015 The Societies and Wiley Publishing Asia Pty Ltd.

Molecular epidemiology of Methicillin-resistant Staphylococcus aureus in Africa: a systematic review

PubMed Central

Abdulgader, Shima M.; Shittu, Adebayo O.; Nicol, Mark P.; Kaba, Mamadou

2015-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) infections are a serious global problem, with considerable impact on patients and substantial health care costs. This systematic review provides an overview on the clonal diversity of MRSA, as well as the prevalence of Panton-Valentine leukocidin (PVL)-positive MRSA in Africa. A search on the molecular characterization of MRSA in Africa was conducted by two authors using predefined terms. We screened for articles published in English and French through to October 2014 from five electronic databases. A total of 57 eligible studies were identified. Thirty-four reports from 15 countries provided adequate genotyping data. CC5 is the predominant clonal complex in the healthcare setting in Africa. The hospital-associated MRSA ST239/ST241-III [3A] was identified in nine African countries. This clone was also described with SCCmec type IV [2B] in Algeria and Nigeria, and type V [5C] in Niger. In Africa, the European ST80-IV [2B] clone was limited to Algeria, Egypt and Tunisia. The clonal types ST22-IV [2B], ST36-II [2A], and ST612-IV [2B] were only reported in South Africa. No clear distinctions were observed between MRSA responsible for hospital and community infections. The community clones ST8-IV [2B] and ST88-IV [2B] were reported both in the hospital and community settings in Angola, Cameroon, Gabon, Ghana, Madagascar, Nigeria, and São Tomé and Príncipe. The proportion of PVL-positive MRSA carriage and/or infections ranged from 0.3 to 100% in humans. A number of pandemic clones were identified in Africa. Moreover, some MRSA clones are limited to specific countries or regions. We strongly advocate for more surveillance studies on MRSA in Africa. PMID:25983721

Isolation and RFLP genotyping of Toxoplasma gondii from the gray wolf (Canis lupus).

PubMed

Dubey, J P; Choudhary, S; Ferreira, L R; Kwok, O C H; Butler, E; Carstensen, M; Yu, L; Su, C

2013-11-08

Little is known of the genetic diversity of Toxoplasma gondii circulating in wildlife. In the present study feral gray wolves (Canis lupus) from Minnesota were examined for T. gondii infection. Antibodies to T. gondii were detected in 130 (52.4%) of 248 wolves tested by the modified agglutination test (cut-off titer of 25). Tissues (hearts, brains or both) of 109 wolves were bioassayed in mice for protozoal isolation. Viable T. gondii was isolated from 25 and the isolates were further propagated in cell culture. T. gondii DNA from these isolates was characterized using 10 PCR-RFLP markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico). Four genotypes were detected. Twenty-one isolates were Type 12 (ToxoDB PCR-RFLP genotype #5), 2 were Type II clonal (ToxoDB #1), 1 was Type II variant (ToxoDB #3), and 1 was a new genotype designated as ToxoDB genotype #219. Published by Elsevier B.V.

Population structure of clinical Pseudomonas aeruginosa from West and Central African countries.

PubMed

Cholley, Pascal; Ka, Roughyatou; Guyeux, Christophe; Thouverez, Michelle; Guessennd, Nathalie; Ghebremedhin, Beniam; Frank, Thierry; Bertrand, Xavier; Hocquet, Didier

2014-01-01

Pseudomonas aeruginosa (PA) has a non-clonal, epidemic population with a few widely distributed and frequently encountered sequence types (STs) called 'high-risk clusters'. Clinical P. aeruginosa (clinPA) has been studied in all inhabited continents excepted in Africa, where a very few isolates have been analyzed. Here, we characterized a collection of clinPA isolates from four countries of West and Central Africa. 184 non-redundant isolates of clinPA from hospitals of Senegal, Ivory Coast, Nigeria, and Central African Republic were genotyped by MLST. We assessed their resistance level to antibiotics by agar diffusion and identified the extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) by sequencing. The population structure of the species was determined by a nucleotide-based analysis of the entire PA MLST database and further localized on the phylogenetic tree (i) the sequence types (STs) of the present collection, (ii) the STs by continents, (iii) ESBL- and MBL-producing STs from the MLST database. We found 80 distinct STs, of which 24 had no relationship with any known STs. 'High-risk' international clonal complexes (CC155, CC244, CC235) were frequently found in West and Central Africa. The five VIM-2-producing isolates belonged to CC233 and CC244. GES-1 and GES-9 enzymes were produced by one CC235 and one ST1469 isolate, respectively. We showed the spread of 'high-risk' international clonal complexes, often described as multidrug-resistant on other continents, with a fully susceptible phenotype. The MBL- and ESBL-producing STs were scattered throughout the phylogenetic tree and our data suggest a poor association between a continent and a specific phylogroup. ESBL- and MBL-encoding genes are borne by both successful international clonal complexes and distinct local STs in clinPA of West and Central Africa. Furthermore, our data suggest that the spread of a ST could be either due to its antibiotic resistance or to features independent from the resistance to antibiotics.

Staphylococcus aureus isolated from wastewater treatment plants in Tunisia: occurrence of human and animal associated lineages.

PubMed

Ben Said, Meriam; Abbassi, Mohamed Salah; Gómez, Paula; Ruiz-Ripa, Laura; Sghaier, Senda; Ibrahim, Chourouk; Torres, Carmen; Hassen, Abdennaceur

2017-08-01

The objective was to characterize Staphylococcus aureus isolated from two wastewater treatment plants (WWTPs) located in Tunis City (Tunisia), during the period 2014-2015. Genetic lineages, antibiotic resistance mechanisms and virulence factors were determined for the recovered isolates. S. aureus isolates were recovered from 12 of the 62 wastewater samples tested (19.35%), and one isolate/sample was characterized, all of them being methicillin-susceptible (MSSA). Six spa types (t587, t674, t224, t127, t701 and t1534) were found among the 12 isolates, and the spa-t587, associated with the new sequence type ST3245, was the most predominant one (7 isolates). The remaining isolates were assigned to five clonal complexes (CC5, CC97, CC1, CC6 and CC522) according to the sequence-type determined and/or the spa-type detected. S. aureus isolates were ascribed to agrI (n = 3), agrII (n = 7) and agrIII (n = 1); however, one isolate was non-typeable. S. aureus showed resistance to (number of isolates): penicillin (12), erythromycin (7), tetracycline (one) and clindamycin (one). Among the virulence factors investigated, only one isolate harboured the tst gene, encoding the TSST-1 (toxic shock syndrome toxin 1). Despite the low number of studied isolates, the present study reports the occurrence of both human- and animal-associated S. aureus clonal complexes in WWTPs in Tunisia.

Inferring a Population Structure for Staphylococcus epidermidis from Multilocus Sequence Typing Data▿

PubMed Central

Miragaia, M.; Thomas, J. C.; Couto, I.; Enright, M. C.; de Lencastre, H.

2007-01-01

Despite its importance as a human pathogen, information on population structure and global epidemiology of Staphylococcus epidermidis is scarce and the relative importance of the mechanisms contributing to clonal diversification is unknown. In this study, we addressed these issues by analyzing a representative collection of S. epidermidis isolates from diverse geographic and clinical origins using multilocus sequence typing (MLST). Additionally, we characterized the mobile element (SCCmec) carrying the genetic determinant of methicillin resistance. The 217 S. epidermidis isolates from our collection were split by MLST into 74 types, suggesting a high level of genetic diversity. Analysis of MLST data using the eBURST algorithm revealed the existence of nine epidemic clonal lineages that were disseminated worldwide. One single clonal lineage (clonal complex 2) comprised 74% of the isolates, whereas the remaining isolates were clustered into 8 minor clonal lineages and 13 singletons. According to our evolutionary model, SCCmec was acquired at least 56 times by S. epidermidis. Although geographic dissemination of S. epidermidis strains and the value of the index of association between the alleles, 0.2898 (P < 0.05), support the clonality of S. epidermidis species, examination of the sequence changes at MLST loci during clonal diversification showed that recombination gives rise to new alleles approximately twice as frequently as point mutations. We suggest that S. epidermidis has a population with an epidemic structure, in which nine clones have emerged upon a recombining background and evolved quickly through frequent transfer of genetic mobile elements, including SCCmec. PMID:17220222

Clonal population of adult stem cells: life span and differentiation potential.

PubMed

Seruya, Mitchel; Shah, Anup; Pedrotty, Dawn; du Laney, Tracey; Melgiri, Ryan; McKee, J Andrew; Young, Henry E; Niklason, Laura E

2004-01-01

Adult stem cells derived from bone marrow, connective tissue, and solid organs can exhibit a range of differentiation potentials. Some controversy exists regarding the classification of mesenchymal stem cells as bona fide stem cells, which is in part derived from the limited ability to propagate true clonal populations of precursor cells. We isolated putative mesenchymal stem cells from the connective tissue of an adult rat (rMSC), and generated clonal populations via three rounds of dilutional cloning. The replicative potential of the clonal rMSC line far exceeded Hayflick's limit of 50-70 population doublings. The high capacity for self-renewal in vitro correlated with telomerase activity, as demonstrated by telomerase repeat amplification protocol (TRAP) assay. Exposure to nonspecific differentiation culture medium revealed multilineage differentiation potential of rMSC clones. Immunostaining confirmed the appearance of mesodermal phenotypes, including adipocytes possessing lipid-rich vacuoles, chondrocytes depositing pericellular type II collagen, and skeletal myoblasts expressing MyoD1. Importantly, the spectrum of differentiation capability was sustained through repeated passaging. Furthermore, serum-free conditions that led to high-efficiency smooth muscle differentiation were identified. rMSCs plated on collagen IV-coated surfaces and exposed to transforming growth factor-beta1 (TGF-beta1) differentiated into a homogeneous population expressing alpha-actin and calponin. Hence, clonogenic analysis confirmed the presence of a putative MSC population derived from the connective tissue of rat skeletal muscle. The ability to differentiate into a smooth muscle cell (SMC) phenotype, combined with a high proliferative capacity, make such a connective tissue-derived MSC population ideal for applications in vascular tissue construction.

Quantitative stability of hematopoietic stem and progenitor cell clonal output in rhesus macaques receiving transplants

PubMed Central

Koelle, Samson J.

2017-01-01

Autologous transplantation of hematopoietic stem and progenitor cells lentivirally labeled with unique oligonucleotide barcodes flanked by sequencing primer targets enables quantitative assessment of the self-renewal and differentiation patterns of these cells in a myeloablative rhesus macaque model. Compared with other approaches to clonal tracking, this approach is highly quantitative and reproducible. We documented stable multipotent long-term hematopoietic clonal output of monocytes, granulocytes, B cells, and T cells from a polyclonal pool of hematopoietic stem and progenitor cells in 4 macaques observed for up to 49 months posttransplantation. A broad range of clonal behaviors characterized by contribution level and biases toward certain cell types were extremely stable over time. Correlations between granulocyte and monocyte clonalities were greatest, followed by correlations between these cell types and B cells. We also detected quantitative expansion of T cell–biased clones consistent with an adaptive immune response. In contrast to recent data from a nonquantitative murine model, there was little evidence for clonal succession after initial hematopoietic reconstitution. These findings have important implications for human hematopoiesis, given the similarities between macaque and human physiologies. PMID:28087539

Antiviral CD8+ T Cells Restricted by Human Leukocyte Antigen Class II Exist during Natural HIV Infection and Exhibit Clonal Expansion.

PubMed

Ranasinghe, Srinika; Lamothe, Pedro A; Soghoian, Damien Z; Kazer, Samuel W; Cole, Michael B; Shalek, Alex K; Yosef, Nir; Jones, R Brad; Donaghey, Faith; Nwonu, Chioma; Jani, Priya; Clayton, Gina M; Crawford, Frances; White, Janice; Montoya, Alana; Power, Karen; Allen, Todd M; Streeck, Hendrik; Kaufmann, Daniel E; Picker, Louis J; Kappler, John W; Walker, Bruce D

2016-10-18

CD8 + T cell recognition of virus-infected cells is characteristically restricted by major histocompatibility complex (MHC) class I, although rare examples of MHC class II restriction have been reported in Cd4-deficient mice and a macaque SIV vaccine trial using a recombinant cytomegalovirus vector. Here, we demonstrate the presence of human leukocyte antigen (HLA) class II-restricted CD8 + T cell responses with antiviral properties in a small subset of HIV-infected individuals. In these individuals, T cell receptor β (TCRβ) analysis revealed that class II-restricted CD8 + T cells underwent clonal expansion and mediated killing of HIV-infected cells. In one case, these cells comprised 12% of circulating CD8 + T cells, and TCRα analysis revealed two distinct co-expressed TCRα chains, with only one contributing to binding of the class II HLA-peptide complex. These data indicate that class II-restricted CD8 + T cell responses can exist in a chronic human viral infection, and may contribute to immune control. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Analysis of the type II-A CRISPR-Cas system of Streptococcus agalactiae reveals distinctive features according to genetic lineages

PubMed Central

Lier, Clément; Baticle, Elodie; Horvath, Philippe; Haguenoer, Eve; Valentin, Anne-Sophie; Glaser, Philippe; Mereghetti, Laurent; Lanotte, Philippe

2015-01-01

CRISPR-Cas systems (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) are found in 90% of archaea and about 40% of bacteria. In this original system, CRISPR arrays comprise short, almost unique sequences called spacers that are interspersed with conserved palindromic repeats. These systems play a role in adaptive immunity and participate to fight non-self DNA such as integrative and conjugative elements, plasmids, and phages. In Streptococcus agalactiae, a bacterium implicated in colonization and infections in humans since the 1960s, two CRISPR-Cas systems have been described. A type II-A system, characterized by proteins Cas9, Cas1, Cas2, and Csn2, is ubiquitous, and a type I–C system, with the Cas8c signature protein, is present in about 20% of the isolates. Unlike type I–C, which appears to be non-functional, type II-A appears fully functional. Here we studied type II-A CRISPR-cas loci from 126 human isolates of S. agalactiae belonging to different clonal complexes that represent the diversity of the species and that have been implicated in colonization or infection. The CRISPR-cas locus was analyzed both at spacer and repeat levels. Major distinctive features were identified according to the phylogenetic lineages previously defined by multilocus sequence typing, especially for the sequence type (ST) 17, which is considered hypervirulent. Among other idiosyncrasies, ST-17 shows a significantly lower number of spacers in comparison with other lineages. This characteristic could reflect the peculiar virulence or colonization specificities of this lineage. PMID:26124774

Turning rice meiosis into mitosis

PubMed Central

Mieulet, Delphine; Jolivet, Sylvie; Rivard, Maud; Cromer, Laurence; Vernet, Aurore; Mayonove, Pauline; Pereira, Lucie; Droc, Gaëtan; Courtois, Brigitte; Guiderdoni, Emmanuel; Mercier, Raphael

2016-01-01

Introduction of clonal reproduction through seeds (apomixis) in crops has the potential to revolutionize agriculture by allowing self-propagation of any elite variety, in particular F1 hybrids. In the sexual model plant Arabidopsis thaliana synthetic clonal reproduction through seeds can be artificially implemented by (i) combining three mutations to turn meiosis into mitosis (MiMe) and (ii) crossing the obtained clonal gametes with a line expressing modified CENH3 and whose genome is eliminated in the zygote. Here we show that additional combinations of mutations can turn Arabidopsis meiosis into mitosis and that a combination of three mutations in rice (Oryza sativa) efficiently turns meiosis into mitosis, leading to the production of male and female clonal diploid gametes in this major crop. Successful implementation of the MiMe technology in the phylogenetically distant eudicot Arabidopsis and monocot rice opens doors for its application to any flowering plant and paves the way for introducing apomixis in crop species. PMID:27767093

Poor clinical outcome for meningitis caused by Haemophilus influenzae serotype A strains containing the IS1016-bexA deletion.

PubMed

Lima, Josilene B T; Ribeiro, Guilherme S; Cordeiro, Soraia M; Gouveia, Edilane L; Salgado, Kátia; Spratt, Brian G; Godoy, Daniel; Reis, Mitermayer G; Ko, Albert I; Reis, Joice N

2010-11-15

Since the introduction of Haemophilus influenzae type b (Hib) conjugate vaccines, meningitis caused by serotypes other than Hib has gained in importance. We conducted active hospital-based surveillance for meningitis over an 11-year period in Salvador, Brazil. H. influenzae isolates were serotyped and analyzed by polymerase chain reaction, pulsed-field gel electrophoresis, and DNA sequencing to identify strains with a specific deletion (IS1016) in the bexA gene (IS1016-bexA). We identified 43 meningitis cases caused by non-type b H. influenzae: 28 (65%) were caused by type a (Hia), 9 (21%) were caused by noncapsulated strains, and 3 (7%) each were caused by types e and f. Hia isolates clustered in 2 clonal groups; clonal group A strains (n = 9) had the IS1016-bexA deletion. Among children <5 years of age, meningitis caused by Hia from clonal group A had higher case-fatality than meningitis caused by clonal group B. Despite small numbers, these results indicate that the presence of the IS1016-bexA deletion is associated with enhanced virulence in non-type b H. influenzae.

Distribution and Molecular Characterization of Acinetobacter baumannii International Clone II Lineage in Japan.

PubMed

Matsui, Mari; Suzuki, Masato; Suzuki, Masahiro; Yatsuyanagi, Jun; Watahiki, Masanori; Hiraki, Yoichi; Kawano, Fumio; Tsutsui, Atsuko; Shibayama, Keigo; Suzuki, Satowa

2018-02-01

Multidrug-resistant (MDR) Acinetobacter spp. have been globally disseminated in association with the successful clonal lineage Acinetobacter baumannii international clone II (IC II). Because the prevalence of MDR Acinetobacter spp. in Japan remains very low, we characterized all Acinetobacter spp. ( n = 866) from 76 hospitals between October 2012 and March 2013 to describe the entire molecular epidemiology of Acinetobacter spp. The most prevalent species was A. baumannii ( n = 645; 74.5%), with A. baumannii IC II ( n = 245) accounting for 28.3% of the total. Meropenem-resistant isolates accounted for 2.0% ( n = 17) and carried IS Aba1-bla OXA-23-like ( n = 10), bla IMP ( n = 4), or IS Aba1-bla OXA-51-like ( n = 3). Multilocus sequence typing of 110 representative A. baumannii isolates revealed the considerable prevalence of domestic sequence types (STs). A. baumannii IC II isolates were divided into the domestic sequence type 469 (ST469) ( n = 18) and the globally disseminated STs ST208 ( n = 14) and ST219 ( n = 4). ST469 isolates were susceptible to more antimicrobial agents, while ST208 and ST219 overproduced the intrinsic AmpC β-lactamase. A. baumannii IC II and some A. baumannii non-IC II STs (e.g., ST149 and ST246) were associated with fluoroquinolone resistance. This study revealed that carbapenem-susceptible A. baumannii IC II was moderately disseminated in Japan. The low prevalence of acquired carbapenemase genes and presence of domestic STs could contribute to the low prevalence of MDR A. baumannii A similar epidemiology might have appeared before the global dissemination of MDR epidemic lineages. In addition, fluoroquinolone resistance associated with A. baumannii IC II may provide insight into the significance of A. baumannii epidemic clones. Copyright © 2018 American Society for Microbiology.

[Molecular epidemiology of methicillin-resistant Staphylococcus aureus isolates with toxic shock syndrome toxin and staphylococcal enterotoxin C genes].

PubMed

Kim, Jae Seok; Kim, Han Sung; Song, Wonkeun; Cho, Hyoun Chan; Lee, Kyu Man; Kim, Eui Chong

2007-04-01

Many methicillin-resistant Staphylococcus aureus (MRSA) isolates in Korea possess a specific profile of staphylococcal enterotoxins in that the toxic shock syndrome toxin gene (tst) coexists with the staphylococcal enterotoxin C gene (sec). Because the analysis of staphylococcal cassette chromosome mec (SCCmec), a mobile genetic element mecA gene encoding methicillin resistance, showed that majority of these are SCCmec type II, these MRSA isolates with tst and sec may be genetically related with each other. This study was performed to investigate the genetic relatedness of tstand sec-harboring MRSA strains isolated in Korea by using pulsed-field gel electrophoresis (PFGE). A total of 59 strains of MRSA isolates of SCCmec type II possessing tst and sec were selected for PFGE and phylogenetic analyses. These isolates were collected from 13 health care facilities during nationwide surveillance of antimicrobial resistance in 2002. The 59 MRSA isolates were clustered into 11 PFGE types, including one major group of 26 strains (44.1%) isolated from 7 healthcare facilities. Seven PFGE types contained 2 or more isolates each, comprising 55 isolates in total. Most of SCCmec type II MRSA isolates containing tst and sec showed closely related PFGE patterns. Moreover, MRSA isolates collected from different healthcare facilities showed identical PFGE patterns. These findings suggested a clonal spread of MRSA strains possessing tst and sec in Korean hospitals.

Flower-deficient mice have reduced susceptibility to skin papilloma formation

PubMed Central

Petrova, Evgeniya; López-Gay, Jesús M.; Rhiner, Christa; Moreno, Eduardo

2012-01-01

SUMMARY Skin papillomas arise as a result of clonal expansion of mutant cells. It has been proposed that the expansion of pretumoral cell clones is propelled not only by the increased proliferation capacity of mutant cells, but also by active cell selection. Previous studies in Drosophila describe a clonal selection process mediated by the Flower (Fwe) protein, whereby cells that express certain Fwe isoforms are recognized and forced to undergo apoptosis. It was further shown that knock down of fwe expression in Drosophila can prevent the clonal expansion of dMyc-overexpressing pretumoral cells. Here, we study the function of the single predicted mouse homolog of Drosophila Fwe, referred to as mFwe, by clonal overexpression of mFwe isoforms in Drosophila and by analyzing mFwe knock-out mice. We show that clonal overexpression of certain mFwe isoforms in Drosophila also triggers non-autonomous cell death, suggesting that Fwe function is evolutionarily conserved. Although mFwe-deficient mice display a normal phenotype, they develop a significantly lower number of skin papillomas upon exposure to DMBA/TPA two-stage skin carcinogenesis than do treated wild-type and mFwe heterozygous mice. Furthermore, mFwe expression is higher in papillomas and the papilloma-surrounding skin of treated wild-type mice compared with the skin of untreated wild-type mice. Thus, we propose that skin papilloma cells take advantage of mFwe activity to facilitate their clonal expansion. PMID:22362363

Clonality and serotypes of Streptococcus mutans among children by multilocus sequence typing

PubMed Central

Momeni, Stephanie S.; Whiddon, Jennifer; Cheon, Kyounga; Moser, Stephen A.; Childers, Noel K.

2015-01-01

Studies using multilocus sequence typing (MLST) have demonstrated that Streptococcus mutans isolates are genetically diverse. Our laboratory previously demonstrated clonality of S. mutans using MLST but could not discount the possibility of sampling bias. In this study, the clonality of randomly selected S. mutans plaque isolates from African American children was examined using MLST. Serotype and presence of collagen-binding proteins (CBP) cnm/cbm were also assessed. One hundred S. mutans isolates were randomly selected for MLST analysis. Sequence analysis was performed and phylogenetic trees were generated using START2 and MEGA. Thirty-four sequence types (ST) were identified of which 27 were unique to this population. Seventy-five percent of the isolates clustered into 16 clonal groups. Serotypes observed were c (n=84), e (n=3), and k (n=11). The prevalence of S. mutans isolates serotype k was notably high at 17.5%. All isolates were cnm/cbm negative. The clonality of S. mutans demonstrated in this study illustrates the importance of localized populations studies and are consistent with transmission. The prevalence of serotype k, a recently proposed systemic pathogen, observed in this study is higher than reported in most populations and is the first report of S. mutans serotype k in a US population. PMID:26443288

Clonal multidrug-resistant Corynebacterium striatum within a nosocomial environment, Rio de Janeiro, Brazil

PubMed Central

Baio, Paulo Victor Pereira; Mota, Higor Franceschi; Freitas, Andréa D'avila; Gomes, Débora Leandro Rama; Ramos, Juliana Nunes; Sant'Anna, Lincoln Oliveira; Souza, Mônica Cristina; Camello, Thereza Cristina Ferreira; Hirata, Raphael; Vieira, Verônica Viana; Mattos-Guaraldi, Ana Luíza

2013-01-01

Corynebacterium striatum is a potentially pathogenic microorganism with the ability to produce outbreaks of nosocomial infections. Here, we document a nosocomial outbreak caused by multidrug-resistant (MDR) C. striatum in Rio de Janeiro, Brazil. C. striatum identification was confirmed by 16S rRNA and rpoB gene sequencing. Fifteen C. striatum strains were isolated from adults (half of whom were 50 years of age and older). C. striatum was mostly isolated in pure culture from tracheal aspirates of patients undergoing endotracheal intubation procedures. The analysis by pulsed-field gel electrophoresis (PFGE) indicated the presence of four PFGE profiles, including two related clones of MDR strains (PFGE I and II). The data demonstrated the predominance of PFGE type I, comprising 11 MDR isolates that were mostly isolated from intensive care units and surgical wards. A potential causal link between death and MDR C. striatum (PFGE types I and II) infection was observed in five cases. PMID:23440110

Prevalence, genetic relatedness and antibiotic resistance of hospital-acquired clostridium difficile PCR ribotype 018 strains.

PubMed

Seo, Mi-Ran; Kim, Jieun; Lee, Yangsoon; Lim, Dong-Gyun; Pai, Hyunjoo

2018-05-01

Clostridium difficile infection (CDI) is a major healthcare-associated infection. The aim of this study was to investigate the genetic relatedness of the endemic C. difficile PCR ribotype 018 strains in an institution and changes to their characteristics during a five-year period. A total of 207 isolates from inpatients at Hanyang University Hospital from 2009 to 2013 were analysed using multilocus variable-number tandem-repeat analysis (MLVA). Minimum inhibitory concentrations (MICs) of several antibiotics were determined. In total, 204 (98.6%) were genetically related, with a summed tandem-repeat distance (STRD) ≤ 10. Minimum-spanning-tree analysis identified 78 MLVA types, categorized into six clonal complexes (CCs). The largest cluster, CC-I, included 51 MLVA types from 148 isolates (71.5%) and the second largest cluster, CC-II, included 10 MLVA types from 36 isolates (17.4%). Resistance rates for antibiotics were: clindamycin (CLI), 97.6%; moxifloxacin (MXF), 98.6%; vancomycin (VAN), 1.4%; and rifaximin (RFX), 8.2%. All isolates were susceptible to piperacillin/tazobactam (TZP) and metronidazole (MTZ). Comparing the MICs of antibiotics for the isolates each year from 2009 to 2013, MICs of antibiotics that promote CDI, such as CLI, MXF, TZP and RFX, increased over the five-year period (P-value by Kruskal-Wallis test: < 0.0001, <0.0001, <0.0001, and <0.0001 respectively); however, MICs of VAN or MTZ, antibiotics for treatment of CDI, did not increase or decreased over the same time period (P-value by Kruskal-Wallis test: 0.166, <0.0001). C. difficile RT018 isolates in a tertiary hospital over a five-year period presented a close clonal relationship. MICs of antibiotics promoting CDI increased with this clonal expansion. Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Molecular Typing and Virulence Gene Profiles of Enterotoxin Gene Cluster (egc)-Positive Staphylococcus aureus Isolates Obtained from Various Food and Clinical Specimens.

PubMed

Song, Minghui; Shi, Chunlei; Xu, Xuebing; Shi, Xianming

2016-11-01

The enterotoxin gene cluster (egc) has been proposed to contribute to the Staphylococcus aureus colonization, which highlights the need to evaluate genetic diversity and virulence gene profiles of the egc-positive population. Here, a total of 43 egc-positive isolates (16.2%) were identified from 266 S. aureus isolates that were obtained from various food and clinical specimens in Shanghai. Seven different egc profiles were found based on the polymerase chain reaction (PCR) result for egc genes. Then, these 43 egc-positive isolates were further typed by multilocus sequence typing, pulsed-field gel electrophoresis (PFGE), multiple-locus variable-number tandem-repeat analysis (MLVA), and accessory gene regulatory (agr) typing. It showed that the 43 egc-positive isolates displayed 17 sequence types, 28 PFGE patterns, 29 MLVA types, and 4 agr types, respectively. Among them, the dominant clonal lineage was CC5-agr II (48.84%). Thirty toxin and 20 adhesion-associated genes were detected by PCR in egc-positive isolates. Notably, invasive toxin genes showed a high prevalence, such as 76.7% for Panton-Valentine leukocidin encoding genes, 27.9% for sec, and 23.3% for tsst-1. Most of the examined adhesion-associated genes were found to be conserved (76.7-100%), whereas the fnbB gene was only found in 8 (18.6%) isolates. In addition, 33 toxin gene profiles and 13 adhesion gene profiles were identified, respectively. Our results imply that isolates belonging to the same clonal lineage harbored similar adhesion gene profiles but diverse toxin gene profiles. Overall, the high prevalence of invasive virulence genes increases the potential risk of egc-positive isolates in S. aureus infection.

Population Structure in Nontypeable Haemophilus influenzae

PubMed Central

LaCross, Nathan C.; Marrs, Carl F.; Gilsdorf, Janet R.

2013-01-01

Nontypeable Haemophilus influenzae (NTHi) frequently colonize the human pharynx asymptomatically, and are an important cause of otitis media in children. Past studies have identified typeable H. influenzae as being clonal, but the population structure of NTHi has not been extensively characterized. The research presented here investigated the diversity and population structure in a well-characterized collection of NTHi isolated from the middle ears of children with otitis media or the pharynges of healthy children in three disparate geographic regions. Multilocus sequence typing identified 109 unique sequence types among 170 commensal and otitis media-associated NTHi isolates from Finland, Israel, and the US. The largest clonal complex contained only five sequence types, indicating a high level of genetic diversity. The eBURST v3, ClonalFrame 1.1, and structure 2.3.3 programs were used to further characterize diversity and population structure from the sequence typing data. Little clustering was apparent by either disease state (otitis media or commensalism) or geography in the ClonalFrame phylogeny. Population structure was clearly evident, with support for eight populations when all 170 isolates were analyzed. Interestingly, one population contained only commensal isolates, while two others consisted solely of otitis media isolates, suggesting associations between population structure and disease. PMID:23266487

Use of X-Chromosome Inactivation Pattern to Analyze the Clonality of 14 Female Cases of Kaposi Sarcoma.

PubMed

Yuan, Ding; XiuJuan, Wu; Yan, Zhang; JunQin, Liang; Fang, Xiang; Shirong, Yu; Xiaojing, Kang; Yanyan, Feng; Weidong, Wu; Dong, Luo; Qingli, Lu; DeZhi, Zhang; XiongMing, Pu

2015-06-16

Kaposi sarcoma (KS) has features of both neoplastic growth and hyperplastic proliferation. It is the most common tumor seen in patients with HIV infection. Whether KS is a real tumor or a benign hyperplastic disease is not known. Tissues from KS and cutaneous hemangioma lesion DNA were extracted, and then digested with methylation-sensitive restriction endonuclease HpaII. Human androgen receptor gene (HUMARA) was amplified with PCR method and the product was separated on 10% denaturing polyacrylamide gels and stained with ethylene dibromide (EB) to show the polymorphism of HUMARA. Phosphoglycerate kinase (PGK) was amplified and the product was digested by BStXI, agarose gel and EB stained to show the polymorphism of PGK. Finally, we analyzed the clonality of KS. In the 14 patients with KS, heterozygosity of the HUMARA gene was observed in 12 (85.7%) cases. Loss of heterozygosity of HUMARA gene on X-chromosome (without HpaII digestion there were 2 bands, after HpaII digestion there were just 1 of the bands), representing monoclonal origin, was present in 11 cases of Kaposi sarcoma. Heterozygosity of the PGK gene was observed in 5 (35.7%) cases, which all represent monoclonal origin. There was no significant difference according to country, stage, or HIV and HHV-8 (P>0.05). The current findings suggest that Kaposi sarcoma is a clonal neoplasm, not a reactive proliferation.

TEM Derivative-Producing Enterobacter aerogenes Strains: Dissemination of a Prevalent Clone

PubMed Central

Dumarche, P.; De Champs, C.; Sirot, D.; Chanal, C.; Bonnet, R.; Sirot, J.

2002-01-01

TEM-24 (CAZ-6) extended-spectrum β-lactamase (ESBL) was detected in 1988 in Clermont-Ferrand, France, in Klebsiella pneumoniae (blaTEM-24) and Enterobacter aerogenes (blaTEM-24b), and since 1994, a TEM-24-producing E. aerogenes clonal strain has been observed elsewhere in the country. To determine if the spread of this clonal strain was restricted to TEM-24-producing E. aerogenes strains, 84 E. aerogenes strains (non-TEM/SHV-producing strains, TEM-1- or -2-producing strains, and different ESBL-producing strains), isolated from 1988 to 1999 in Clermont-Ferrand (n = 59) and in 11 other French hospitals in 1998 (n = 25), were studied. A clonal strain was found for TEM-24- but also for TEM-3- and TEM-1- or 2-producing isolates. This study shows that there is a clonal strain dependent on acquisition of the TEM-type enzyme (TEM-24 and other TEM types). PMID:11897606

Population Structure of Clinical Pseudomonas aeruginosa from West and Central African Countries

PubMed Central

Cholley, Pascal; Ka, Roughyatou; Guyeux, Christophe; Thouverez, Michelle; Guessennd, Nathalie; Ghebremedhin, Beniam; Frank, Thierry; Bertrand, Xavier; Hocquet, Didier

2014-01-01

Background Pseudomonas aeruginosa (PA) has a non-clonal, epidemic population with a few widely distributed and frequently encountered sequence types (STs) called ‘high-risk clusters’. Clinical P. aeruginosa (clinPA) has been studied in all inhabited continents excepted in Africa, where a very few isolates have been analyzed. Here, we characterized a collection of clinPA isolates from four countries of West and Central Africa. Methodology 184 non-redundant isolates of clinPA from hospitals of Senegal, Ivory Coast, Nigeria, and Central African Republic were genotyped by MLST. We assessed their resistance level to antibiotics by agar diffusion and identified the extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) by sequencing. The population structure of the species was determined by a nucleotide-based analysis of the entire PA MLST database and further localized on the phylogenetic tree (i) the sequence types (STs) of the present collection, (ii) the STs by continents, (iii) ESBL- and MBL-producing STs from the MLST database. Principal Findings We found 80 distinct STs, of which 24 had no relationship with any known STs. ‘High-risk’ international clonal complexes (CC155, CC244, CC235) were frequently found in West and Central Africa. The five VIM-2-producing isolates belonged to CC233 and CC244. GES-1 and GES-9 enzymes were produced by one CC235 and one ST1469 isolate, respectively. We showed the spread of ‘high-risk’ international clonal complexes, often described as multidrug-resistant on other continents, with a fully susceptible phenotype. The MBL- and ESBL-producing STs were scattered throughout the phylogenetic tree and our data suggest a poor association between a continent and a specific phylogroup. Conclusions ESBL- and MBL-encoding genes are borne by both successful international clonal complexes and distinct local STs in clinPA of West and Central Africa. Furthermore, our data suggest that the spread of a ST could be either due to its antibiotic resistance or to features independent from the resistance to antibiotics. PMID:25187957

Disruptive chemicals, senescence and immortality

PubMed Central

Carnero, Amancio; Blanco-Aparicio, Carmen; Kondoh, Hiroshi; Lleonart, Matilde E.; Martinez-Leal, Juan Fernando; Mondello, Chiara; Ivana Scovassi, A.; Bisson, William H.; Amedei, Amedeo; Roy, Rabindra; Woodrick, Jordan; Colacci, Annamaria; Vaccari, Monica; Raju, Jayadev; Al-Mulla, Fahd; Al-Temaimi, Rabeah; Salem, Hosni K.; Memeo, Lorenzo; Forte, Stefano; Singh, Neetu; Hamid, Roslida A.; Ryan, Elizabeth P.; Brown, Dustin G.; Wise, John Pierce; Wise, Sandra S.; Yasaei, Hemad

2015-01-01

Carcinogenesis is thought to be a multistep process, with clonal evolution playing a central role in the process. Clonal evolution involves the repeated ‘selection and succession’ of rare variant cells that acquire a growth advantage over the remaining cell population through the acquisition of ‘driver mutations’ enabling a selective advantage in a particular micro-environment. Clonal selection is the driving force behind tumorigenesis and possesses three basic requirements: (i) effective competitive proliferation of the variant clone when compared with its neighboring cells, (ii) acquisition of an indefinite capacity for self-renewal, and (iii) establishment of sufficiently high levels of genetic and epigenetic variability to permit the emergence of rare variants. However, several questions regarding the process of clonal evolution remain. Which cellular processes initiate carcinogenesis in the first place? To what extent are environmental carcinogens responsible for the initiation of clonal evolution? What are the roles of genotoxic and non-genotoxic carcinogens in carcinogenesis? What are the underlying mechanisms responsible for chemical carcinogen-induced cellular immortality? Here, we explore the possible mechanisms of cellular immortalization, the contribution of immortalization to tumorigenesis and the mechanisms by which chemical carcinogens may contribute to these processes. PMID:26106138

Characterization of antibodies directed against the immunoglobulin light kappa chain variable chain region (VK) of hepatitis C virus-related type-II mixed cryoglobulinemia and B-cell proliferations.

PubMed

de Re, Valli; Simula, Maria Paola; Pavan, Alessandro; Garziera, Marica; Marin, Dolores; Dolcetti, Riccardo; de Vita, Salvatore; Sansonno, Domenico; Geremia, Silvano; Toffoli, Giuseppe

2009-09-01

Autoimmune type-II cryoglobulinemia (II-MC) is sustained by hepatitis C virus (HCV) infection and B-cell (oligo)clones. This is the reason why the disease may be considered an "indolent B-cell lymphoma (NHL)." B clones show a restricted use of immunoglobulin variable genes (BCR), in particular in the use of the variable kappa (VK)3-20/15 light chain, and show a homology between their BCR functional regions and those of autoimmune rheumatoid factors. We underlined the BCR unique repertoire with frequent rheumatoid factor activity also observed in other autoimmune disorders associated with NHL. The immunoglobulin idiotype is a clonal B-cell marker and an ideal target for immunotherapy. Five monoclonal antibodies were produced in our laboratory toward the VK3-20 of a subject with HCV infection and a II-MC-associated NHL. Epitope determination was performed using the epitope excision approach. Monoclonal antibody reactivity was tested in vitro in ELISA, Western blot, and cytofluorimetry. Data confirmed that a panel of antibodies, reactive against shared idiotypes, can be produced from patients with HCV-associated B-cell lymphoproliferative diseases, thus obviating the need to produce an anti-idiotype antibody for each patient.

Molecular Evidence of Toxoplasma gondii, Neospora caninum, and Encephalitozoon cuniculi in Red Foxes ( Vulpes vulpes).

PubMed

Lukášová, Radka; Marková, Jiřina; Bártová, Eva; Murat, Jean-Benjamin; Sedlák, Kamil

2018-05-07

Toxoplasma gondii, Neospora caninum, and Encephalitozoon cuniculi are important infectious agents, with T. gondii and E. cuniculi having zoonotic potential. There are two main clonal lineages (types I and II) of T. gondii in Europe, but little is known about genotypes of T. gondii in wild animals. The aim of our study was molecular detection of these three pathogens in tissues of wild red foxes ( Vulpes vulpes) from the Czech Republic. Using PCR (B1 gene), we detected T. gondii in 10% of the animals that we tested ( n=100); N. caninum and E. cuniculi were not detected. The T. gondii samples were genotyped by single multiplex PCR assay with 15 microsatellite markers. Five samples were successfully genotyped as genotype II, a unique finding for T. gondii isolated from red foxes from the Czech Republic.

Assessment of clonality and serotypes of Streptococcus mutans among children by multilocus sequence typing.

PubMed

Momeni, Stephanie S; Whiddon, Jennifer; Cheon, Kyounga; Moser, Stephen A; Childers, Noel K

2015-12-01

Studies using multilocus sequence typing (MLST) have demonstrated that Streptococcus mutans isolates are genetically diverse. Our laboratory previously demonstrated clonality of S. mutans using MLST but could not discount the possibility of sampling bias. In this study, the clonality of randomly selected S. mutans plaque isolates from African-American children was examined using MLST. Serotype and the presence of collagen-binding proteins (CBPs) encoded by cnm/cbm were also assessed. One-hundred S. mutans isolates were randomly selected for MLST analysis. Sequence analysis was performed and phylogenetic trees were generated using start2 and mega. Thirty-four sequence types were identified, of which 27 were unique to this population. Seventy-five per cent of the isolates clustered into 16 clonal groups. The serotypes observed were c (n = 84), e (n = 3), and k (n = 11). The prevalence of S. mutans isolates of serotype k was notably high, at 17.5%. All isolates were cnm/cbm negative. The clonality of S. mutans demonstrated in this study illustrates the importance of localized population studies and are consistent with transmission. The prevalence of serotype k, a recently proposed systemic pathogen, observed in this study, is higher than reported in most populations and is the first report of S. mutans serotype k in a United States population. © 2015 Eur J Oral Sci.

Whole-organism clone tracing using single-cell sequencing.

PubMed

Alemany, Anna; Florescu, Maria; Baron, Chloé S; Peterson-Maduro, Josi; van Oudenaarden, Alexander

2018-04-05

Embryonic development is a crucial period in the life of a multicellular organism, during which limited sets of embryonic progenitors produce all cells in the adult body. Determining which fate these progenitors acquire in adult tissues requires the simultaneous measurement of clonal history and cell identity at single-cell resolution, which has been a major challenge. Clonal history has traditionally been investigated by microscopically tracking cells during development, monitoring the heritable expression of genetically encoded fluorescent proteins and, more recently, using next-generation sequencing technologies that exploit somatic mutations, microsatellite instability, transposon tagging, viral barcoding, CRISPR-Cas9 genome editing and Cre-loxP recombination. Single-cell transcriptomics provides a powerful platform for unbiased cell-type classification. Here we present ScarTrace, a single-cell sequencing strategy that enables the simultaneous quantification of clonal history and cell type for thousands of cells obtained from different organs of the adult zebrafish. Using ScarTrace, we show that a small set of multipotent embryonic progenitors generate all haematopoietic cells in the kidney marrow, and that many progenitors produce specific cell types in the eyes and brain. In addition, we study when embryonic progenitors commit to the left or right eye. ScarTrace reveals that epidermal and mesenchymal cells in the caudal fin arise from the same progenitors, and that osteoblast-restricted precursors can produce mesenchymal cells during regeneration. Furthermore, we identify resident immune cells in the fin with a distinct clonal origin from other blood cell types. We envision that similar approaches will have major applications in other experimental systems, in which the matching of embryonic clonal origin to adult cell type will ultimately allow reconstruction of how the adult body is built from a single cell.

A Longitudinal 6-Year Study of the Molecular Epidemiology of Clinical Campylobacter Isolates in Oxfordshire, United Kingdom

PubMed Central

McCarthy, Noel M.; Wimalarathna, Helen L.; Colles, Frances M.; Clark, Lorraine; Bowler, Ian C. J. W.; Maiden, Martin C. J.; Dingle, Kate E.

2012-01-01

Temporal and seasonal trends in Campylobacter genotypes causing human gastroenteritis were investigated in a 6-year study of 3,300 recent isolates from Oxfordshire, United Kingdom. Genotypes (sequence types [ST]) were defined using multilocus sequence typing and assigned to a clonal complex (a cluster of related strains that share four or more identical alleles with a previously defined central genotype). A previously undescribed clonal complex (ST-464) was identified which, together with ST-42, ST-45, and ST-52 complexes, showed increasing incidence. Concurrently, the incidence of ST-574, ST-607, and ST-658 complexes declined. The relative frequencies of three clonal complexes (ST-45, ST-283, and ST-42) peaked during summer and those of two (ST-353 and ST-403) peaked during winter. Nine clonal complexes (ST-22, ST-45, ST-48, ST-61, ST-257, ST-283, ST-403, ST-658, and ST-677) were significantly associated with ciprofloxacin sensitivity (P < 0.05). Seven clonal complexes (ST-49, ST-206, ST-354, ST-446, ST-460, ST-464, and ST-607) were associated with ciprofloxacin resistance (P < 0.05). Clonal complexes exhibited changing incidence and differences in seasonality and antibiotic resistance phenotype. These data also demonstrated that detailed surveillance at a single site captures information which reflects that observed nationally. PMID:22814466

Long-term consequences of disturbances on reproductive strategies of the rare epiphytic lichen Lobaria pulmonaria: clonality a gift and a curse.

PubMed

Singh, Garima; Dal Grande, Francesco; Werth, Silke; Scheidegger, Christoph

2015-01-01

The effect of disturbance on symbiotic organisms such as lichens is particularly severe. In case of heterothallic lichen-forming fungi, disturbances may lead to unbalanced gene frequency and patchy distribution of mating types, thus inhibiting sexual reproduction and imposing clonality. The impact of disturbance on reproductive strategies and genetic diversity of clonal systems has so far received little attention. To infer the effects of disturbances on mating-type allele frequencies and population structure, we selected three populations in the Parc Jurassien Vaudois (Switzerland), which were affected by uneven-aged forestry, intensive logging and fire, respectively. We used microsatellite markers to infer genetic diversity, allelic richness and clonal diversity of the epiphytic lichen Lobaria pulmonaria and used L. pulmonaria-specific MAT1-1 and MAT1-2 markers to analyse the frequency and distribution of mating types of 889 individuals. Our study shows that stand-replacing disturbances affect the mating-type frequency and distribution, thus compromising the potential for sexual reproduction. The fire-disturbed area had a significantly lower genetic and genotypic diversity and a higher clonality. Furthermore, the majority of compatible mating pairs in this area were beyond the effective vegetative dispersal range of the species. We conclude that stand-replacing disturbances lead to lower chances of sex and symbiont reshuffling and thus have long-lasting negative consequences on the reproductive strategies and adaptive potential of epiphytic lichen symbioses. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Expression of lumican in hidroacanthoma simplex and clonal-type seborrheic keratosis as a potent differential diagnostic marker.

PubMed

Takayama, Ryoko; Ansai, Shin-Ichi; Ishiwata, Toshiyuki; Yamamoto, Tetsushi; Matsuda, Yoko; Naito, Zenya; Kawana, Seiji

2014-08-01

Lumican, a member of the small leucine-rich proteoglycan family, regulates the assembly and diameter of collagen fibers in the extracellular matrix of various tissues. The lumican expression correlates with pathological conditions and the growth and metastasis of various malignancies. In cutaneous neoplasms, the lumican expression is lower in advanced-stage malignant melanomas that invade the dermis than in early-stage melanomas. Furthermore, we have recently reported that the expression pattern of lumican is different from that of actinic keratosis and the Bowen disease. Lumican is positive in the poroid cells of intraepidermal sweat ducts; therefore, we examined the expression patterns of lumican in acanthotic-type seborrheic keratosis and Pinkus-type poroma followed by clonal-type seborrheic keratosis and hidroacanthoma simplex. The neoplastic cells of acanthotic-type seborrheic keratosis exhibited positive immunostaining in only 1 of 31 cases (3.23%), whereas the poroid cells of Pinkus-type poroma exhibited positive immunoreactivity in 26 of 28 patients (92.8%). In the hidroacanthoma simplex cases, lumican was expressed in poroid cells forming intraepidermal nests in 22 of 28 patients (78.6%), whereas the neoplastic cells in most cases of clonal-type seborrheic keratosis were negative for lumican. In some seborrheic keratosis cases that were positive for lumican in neoplastic cells, lumican was observed in squamoid cells but not in basaloid cells. Therefore, it is necessary to evaluate the immunoreactivity of lumican in seborrheic keratosis and in basaloid cells. These findings suggest that lumican is a potent differential diagnostic marker that distinguishes hidroacanthoma simplex from clonal-type seborrheic keratosis.

Wide geographical dissemination of the multiresistant Staphylococcus capitis NRCS-A clone in neonatal intensive-care units.

PubMed

Butin, M; Rasigade, J-P; Martins-Simões, P; Meugnier, H; Lemriss, H; Goering, R V; Kearns, A; Deighton, M A; Denis, O; Ibrahimi, A; Claris, O; Vandenesch, F; Picaud, J-C; Laurent, F

2016-01-01

Nosocomial late-onset sepsis represents a frequent cause of morbidity and mortality in preterm neonates. The Staphylococcus capitis clone NRCS-A has been previously described as an emerging cause of nosocomial bacteraemia in French neonatal intensive-care units (NICUs). In this study, we aimed to explore the possible unrecognized dissemination of this clone on a larger geographical scale. One hundred methicillin-resistant S. capitis strains isolated from neonates (n = 86) and adult patients (n = 14) between 2000 and 2013 in four different countries (France, Belgium, the UK, and Australia) were analysed with SmaI pulsed-field gel electrophoresis (PFGE) and dru typing. The vast majority of NICU strains showed the NRCS-A pulsotype and the dt11c type (96%). We then randomly selected 14 isolates (from neonates, n = 12, three per country; from adult patients, n = 2), considered to be a subset of representative isolates, and performed further molecular typing (SacII PFGE, SCCmec typing, and multilocus sequence typing-like analysis), confirming the clonality of the S. capitis strains isolated from neonates, despite their distant geographical origin. Whole genome single-nucleotide polymorphism-based phylogenetic analysis of five NICU isolates (from the different countries) attested to high genetic relatedness within the NRCS-A clone. Finally, all of the NRCS-A strains showed multidrug resistance (e.g. methicillin and aminoglycoside resistance, and decreased vancomycin susceptibility), with potential therapeutic implications for infected neonates. In conclusion, this study represents the first report of clonal dissemination of methicillin-resistant coagulase-negative Staphylococcus clone on a large geographical scale. Questions remain regarding the origin and means of international spread, and the reasons for this clone's apparent predilection for neonates. Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Endemic and Epidemic Lineages of Escherichia coli that Cause Urinary Tract Infections

PubMed Central

Tabor, Helen; Tellis, Patricia; Vincent, Caroline; Tellier, Pierre-Paul

2008-01-01

Women with urinary tract infections (UTIs) in California, USA (1999–2001), were infected with closely related or indistinguishable strains of Escherichia coli (clonal groups), which suggests point source dissemination. We compared strains of UTI-causing E. coli in California with strains causing such infections in Montréal, Québec, Canada. Urine specimens from women with community-acquired UTIs in Montréal (2006) were cultured for E. coli. Isolates that caused 256 consecutive episodes of UTI were characterized by antimicrobial drug susceptibility profile, enterobacterial repetitive intergenic consensus 2 PCR, serotyping, XbaI and NotI pulsed-field gel electrophoresis, multilocus sequence typing, and phylogenetic typing. We confirmed the presence of drug-resistant, genetically related, and temporally clustered E. coli clonal groups that caused community-acquired UTIs in unrelated women in 2 locations and 2 different times. Two clonal groups were identified in both locations. Epidemic transmission followed by endemic transmission of UTI-causing clonal groups may explain these clusters of UTI cases. PMID:18826822

Disruptive chemicals, senescence and immortality.

PubMed

Carnero, Amancio; Blanco-Aparicio, Carmen; Kondoh, Hiroshi; Lleonart, Matilde E; Martinez-Leal, Juan Fernando; Mondello, Chiara; Scovassi, A Ivana; Bisson, William H; Amedei, Amedeo; Roy, Rabindra; Woodrick, Jordan; Colacci, Annamaria; Vaccari, Monica; Raju, Jayadev; Al-Mulla, Fahd; Al-Temaimi, Rabeah; Salem, Hosni K; Memeo, Lorenzo; Forte, Stefano; Singh, Neetu; Hamid, Roslida A; Ryan, Elizabeth P; Brown, Dustin G; Wise, John Pierce; Wise, Sandra S; Yasaei, Hemad

2015-06-01

Carcinogenesis is thought to be a multistep process, with clonal evolution playing a central role in the process. Clonal evolution involves the repeated 'selection and succession' of rare variant cells that acquire a growth advantage over the remaining cell population through the acquisition of 'driver mutations' enabling a selective advantage in a particular micro-environment. Clonal selection is the driving force behind tumorigenesis and possesses three basic requirements: (i) effective competitive proliferation of the variant clone when compared with its neighboring cells, (ii) acquisition of an indefinite capacity for self-renewal, and (iii) establishment of sufficiently high levels of genetic and epigenetic variability to permit the emergence of rare variants. However, several questions regarding the process of clonal evolution remain. Which cellular processes initiate carcinogenesis in the first place? To what extent are environmental carcinogens responsible for the initiation of clonal evolution? What are the roles of genotoxic and non-genotoxic carcinogens in carcinogenesis? What are the underlying mechanisms responsible for chemical carcinogen-induced cellular immortality? Here, we explore the possible mechanisms of cellular immortalization, the contribution of immortalization to tumorigenesis and the mechanisms by which chemical carcinogens may contribute to these processes. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Analysis of the clonal repertoire of gene-corrected cells in gene therapy.

PubMed

Paruzynski, Anna; Glimm, Hanno; Schmidt, Manfred; Kalle, Christof von

2012-01-01

Gene therapy-based clinical phase I/II studies using integrating retroviral vectors could successfully treat different monogenetic inherited diseases. However, with increased efficiency of this therapy, severe side effects occurred in various gene therapy trials. In all cases, integration of the vector close to or within a proto-oncogene contributed substantially to the development of the malignancies. Thus, the in-depth analysis of integration site patterns is of high importance to uncover potential clonal outgrowth and to assess the safety of gene transfer vectors and gene therapy protocols. The standard and nonrestrictive linear amplification-mediated PCR (nrLAM-PCR) in combination with high-throughput sequencing exhibits technologies that allow to comprehensively analyze the clonal repertoire of gene-corrected cells and to assess the safety of the used vector system at an early stage on the molecular level. It enables clarifying the biological consequences of the vector system on the fate of the transduced cell. Furthermore, the downstream performance of real-time PCR allows a quantitative estimation of the clonality of individual cells and their clonal progeny. Here, we present a guideline that should allow researchers to perform comprehensive integration site analysis in preclinical and clinical studies. Copyright © 2012 Elsevier Inc. All rights reserved.

First Case of NDM-1-Producing Klebsiella pneumoniae in Annaba University Hospital, Algeria.

PubMed

Abderrahim, Amel; Djahmi, Nassima; Pujol, Charlotte; Nedjai, Sabina; Bentakouk, Mohamed Cherif; Kirane-Gacemi, Djamila; Dekhil, Mazouz; Sotto, Albert; Lavigne, Jean-Philippe; Pantel, Alix

2017-10-01

The aim of this study was to characterize two carbapenem-resistant Klebsiella pneumoniae isolates recovered from urine samples in a patient hospitalized at Annaba University hospital (Algeria) in 2014. Two K. pneumoniae isolates were studied because they proved resistant to almost all antibiotics tested with a high level resistance to ertapenem (minimum inhibitory concentration = 32 mg/L). The results of modified Hodge test and combined disk test (ROSCO Diagnostica, Taastrup, Denmark) were positive. The two isolates harbored the bla NDM-1 gene and one was also positive for bla CTX-M-15 . Screening of aminoglycoside-modifying enzymes and plasmid-mediated quinolone resistance contents detected aac(6')-Ib-cr, aac(3')-II, qnrB2, and oqxAB in both isolates. Multilocus sequence typing demonstrated that the two isolates belonged to sequence type 147. However, repetitive sequence-based PCR and pulsed-field gel electrophoresis showed that they were not clonally related. The bla NDM-1 gene and all other resistant genes were contained on an IncR plasmid of c.a. 85 kb. This study comprises the first identification of NDM-1-producing K. pneumoniae in Algeria. We thus confirm the concerning worldwide dissemination of this carbapenemase that involves the emergence of the IncR plasmid and the success of the ST147 clonal complex harboring it.

The role of weak selection and high mutation rates in nearly neutral evolution.

PubMed

Lawson, Daniel John; Jensen, Henrik Jeldtoft

2009-04-21

Neutral dynamics occur in evolution if all types are 'effectively equal' in their reproductive success, where the definition of 'effectively equal' depends on the population size and the details of mutations. Empirically observed neutral genetic evolution in extremely large clonal populations can only be explained under current models if selection is completely absent. Such models typically consider the case where population dynamics occurs on a different timescale to evolution. However, this assumption is invalid when mutations are not rare in a whole population. We show that this has important consequences for the occurrence of neutral evolution in clonal populations. In highly connected type spaces, neutral dynamics can occur for all population sizes despite significant selective differences, via the forming of effectively neutral networks connecting rare neutral types. Biological implications include an explanation for the high diversity of rare types that survive in large clonal populations, and a theoretical justification for the use of neutral null models.

The management of refractory coeliac disease

PubMed Central

2013-01-01

A significant proportion of patients with coeliac disease are ‘nonresponsive’ to gluten withdrawal. Most cases of nonresponsive coeliac disease are due to persisting gluten ingestion. Refractory coeliac disease (RCD) is currently defined by persistent symptoms and signs of malabsorption after gluten exclusion for 12 months with ongoing intestinal villous atrophy. Primary (without initial response to diet) and secondary (relapse following response to diet) RCD is recognized. RCD is further classified as type I or type II based on the absence or presence of a population of aberrant intestinal lymphocytes. Quality of dietetic advice and support is fundamental, and lack of objective corroboration of gluten exclusion may result in over-identification of RCD I, particularly in those cases with persisting antibody responses. Over-reliance on lymphocyte clonality similarly may result in over-diagnosis of RCD II which requires careful quantification of aberrant lymphocyte populations. Management of RCD should be undertaken in specialist centres. It requires initial intensive dietary supervision, strict gluten exclusion and subsequent re-evaluation. There is currently insufficient evidence to recommend specific treatments. Steroids are often used in both RCD I and II (albeit with little objective evidence of benefit in RCD II), and azathioprine as steroid-sparing therapy in RCD I. There is growing evidence for the use of cladribine in RCD II with autologous stem cell transplantation in nonresponders, but this requires further multicentre evaluation. There remains considerable controversy regarding the diagnosis, treatment and surveillance of RCD: international consensus in these areas is urgently required to facilitate future therapeutic advances. PMID:23556127

Changes in the population structure of canine methicillin-resistant Staphylococcus pseudintermedius in Poland.

PubMed

Kizerwetter-Świda, Magdalena; Chrobak-Chmiel, Dorota; Rzewuska, Magdalena; Binek, Marian

2017-09-01

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) is being reported with an increasing frequency in small animal veterinary practice. The molecular typing of MRSP isolates revealed that the dominating European multidrug-resistant lineage is the sequence type 71 (ST71), associated with staphylococcal chromosomal cassette SCCmec type II-III. However, the recent reports indicated the emergence of other clones. The study aimed to determine the genetic properties of MRSP isolates obtained from dogs in Poland over a ten-year period. A total of 42 clinical MRSP isolates were subjected to multilocus-sequence typing (MLST) and SCCmec typing. MLST typing of 42 MRSP isolates yielded six STs belonging to two major clonal complexes (CCs): CC71 and CC551, associated with SCCmec element II-III and V, respectively. CC71 comprising ST71 and its newly described single locus variant (SLV) ST680. The second dominating CC551was represented by ST551 and newly described SLV ST771. The other, ST258 and ST85 were detected in single MRSP isolates. This is the first report concerning MLST typing of MRSP isolates in Poland. The results confirmed the domination of ST71 among MRSP until 2015, and the emergence of ST551 in 2015. Furthermore, in 2016 ST551 was identified in the majority of the strains, indicating the changes in the population structure of MRSP in Poland. Polish clinical MRSP isolates showed a shift in the population structure during the period of 2007 and 2016. The dominating MRSP lineage until 2015 was multidrug-resistant ST71-SCCmecII-III. The other lineage ST551-SCCmecV emerged in Poland since 2015, and in 2016 was found in the majority of MRSP isolates. Copyright © 2017 Elsevier B.V. All rights reserved.

Multidisciplinary insight into clonal expansion of HTLV-1-infected cells in adult T-cell leukemia via modeling by deterministic finite automata coupled with high-throughput sequencing.

PubMed

Farmanbar, Amir; Firouzi, Sanaz; Park, Sung-Joon; Nakai, Kenta; Uchimaru, Kaoru; Watanabe, Toshiki

2017-01-31

Clonal expansion of leukemic cells leads to onset of adult T-cell leukemia (ATL), an aggressive lymphoid malignancy with a very poor prognosis. Infection with human T-cell leukemia virus type-1 (HTLV-1) is the direct cause of ATL onset, and integration of HTLV-1 into the human genome is essential for clonal expansion of leukemic cells. Therefore, monitoring clonal expansion of HTLV-1-infected cells via isolation of integration sites assists in analyzing infected individuals from early infection to the final stage of ATL development. However, because of the complex nature of clonal expansion, the underlying mechanisms have yet to be clarified. Combining computational/mathematical modeling with experimental and clinical data of integration site-based clonality analysis derived from next generation sequencing technologies provides an appropriate strategy to achieve a better understanding of ATL development. As a comprehensively interdisciplinary project, this study combined three main aspects: wet laboratory experiments, in silico analysis and empirical modeling. We analyzed clinical samples from HTLV-1-infected individuals with a broad range of proviral loads using a high-throughput methodology that enables isolation of HTLV-1 integration sites and accurate measurement of the size of infected clones. We categorized clones into four size groups, "very small", "small", "big", and "very big", based on the patterns of clonal growth and observed clone sizes. We propose an empirical formal model based on deterministic finite state automata (DFA) analysis of real clinical samples to illustrate patterns of clonal expansion. Through the developed model, we have translated biological data of clonal expansion into the formal language of mathematics and represented the observed clonality data with DFA. Our data suggest that combining experimental data (absolute size of clones) with DFA can describe the clonality status of patients. This kind of modeling provides a basic understanding as well as a unique perspective for clarifying the mechanisms of clonal expansion in ATL.

Inventory of Extended-Spectrum-β-Lactamase-Producing Enterobacteriaceae in France as Assessed by a Multicenter Study.

PubMed

Robin, F; Beyrouthy, R; Bonacorsi, S; Aissa, N; Bret, L; Brieu, N; Cattoir, V; Chapuis, A; Chardon, H; Degand, N; Doucet-Populaire, F; Dubois, V; Fortineau, N; Grillon, A; Lanotte, P; Leyssene, D; Patry, I; Podglajen, I; Recule, C; Ros, A; Colomb-Cotinat, M; Ponties, V; Ploy, M C; Bonnet, R

2017-03-01

The objective of this study was to perform an inventory of the extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae isolates responsible for infections in French hospitals and to assess the mechanisms associated with ESBL diffusion. A total of 200 nonredundant ESBL-producing Enterobacteriaceae strains isolated from clinical samples were collected during a multicenter study performed in 18 representative French hospitals. Antibiotic resistance genes were identified by PCR and sequencing experiments. The clonal relatedness between isolates was investigated by the use of the DiversiLab system. ESBL-encoding plasmids were compared by PCR-based replicon typing and plasmid multilocus sequence typing. CTX-M-15, CTX-M-1, CTX-M-14, and SHV-12 were the most prevalent ESBLs (8% to 46.5%). The three CTX-M-type EBSLs were significantly observed in Escherichia coli (37.1%, 24.2%, and 21.8%, respectively), and CTX-M-15 was the predominant ESBL in Klebsiella pneumoniae (81.1%). SHV-12 was associated with ESBL-encoding Enterobacter cloacae strains (37.9%). qnrB , aac(6 ' )-Ib-cr , and aac(3)-II genes were the main plasmid-mediated resistance genes, with prevalences ranging between 19.5% and 45% according to the ESBL results. Molecular typing did not identify wide clonal diffusion. Plasmid analysis suggested the diffusion of low numbers of ESBL-encoding plasmids, especially in K. pneumoniae and E. cloacae However, the ESBL-encoding genes were observed in different plasmid replicons according to the bacterial species. The prevalences of ESBL subtypes differ according to the Enterobacteriaceae species. Plasmid spread is a key determinant of this epidemiology, and the link observed between the ESBL-encoding plasmids and the bacterial host explains the differences observed in the Enterobacteriaceae species. Copyright © 2017 American Society for Microbiology.

Novel immortal human cell lines reveal subpopulations in the nucleus pulposus

PubMed Central

2014-01-01

Introduction Relatively little is known about cellular subpopulations in the mature nucleus pulposus (NP). Detailed understanding of the ontogenetic, cellular and molecular characteristics of functional intervertebral disc (IVD) cell populations is pivotal to the successful development of cell replacement therapies and IVD regeneration. In this study, we aimed to investigate whether phenotypically distinct clonal cell lines representing different subpopulations in the human NP could be generated using immortalization strategies. Methods Nondegenerate healthy disc material (age range, 8 to 15 years) was obtained as surplus surgical material. Early passage NP monolayer cell cultures were initially characterized using a recently established NP marker set. NP cells were immortalized by simian virus 40 large T antigen (SV40LTag) and human telomerase reverse transcriptase expression. Immortalized cells were clonally expanded and characterized based on collagen type I, collagen type II, α1 (COL2A1), and SRY-box 9 (SOX9) protein expression profiles, as well as on expression of a subset of established in vivo NP cell lineage markers. Results A total of 54 immortal clones were generated. Profiling of a set of novel NP markers (CD24, CA12, PAX1, PTN, FOXF1 and KRT19 mRNA) in a representative set of subclones substantiated successful immortalization of multiple cellular subpopulations from primary isolates and confirmed their NP origin and/or phenotype. We were able to identify two predominant clonal NP subtypes based on their morphological characteristics and their ability to induce SOX9 and COL2A1 under conventional differentiation conditions. In addition, cluster of differentiation 24 (CD24)–negative NP responder clones formed spheroid structures in various culture systems, suggesting the preservation of a more immature phenotype compared to CD24-positive nonresponder clones. Conclusions Here we report the generation of clonal NP cell lines from nondegenerate human IVD tissue and present a detailed characterization of NP cellular subpopulations. Differential cell surface marker expression and divergent responses to differentiation conditions suggest that the NP subtypes may correspond to distinct maturation stages and represent distinct NP cell subpopulations. Hence, we provide evidence that the immortalization strategy that we applied is capable of detecting cell heterogeneity in the NP. Our cell lines yield novel insights into NP biology and provide promising new tools for studies of IVD development, cell function and disease. PMID:24972717

Novel immortal human cell lines reveal subpopulations in the nucleus pulposus.

PubMed

van den Akker, Guus G H; Surtel, Don A M; Cremers, Andy; Rodrigues-Pinto, Ricardo; Richardson, Stephen M; Hoyland, Judith A; van Rhijn, Lodewijk W; Welting, Tim J M; Voncken, Jan Willem

2014-06-27

Relatively little is known about cellular subpopulations in the mature nucleus pulposus (NP). Detailed understanding of the ontogenetic, cellular and molecular characteristics of functional intervertebral disc (IVD) cell populations is pivotal to the successful development of cell replacement therapies and IVD regeneration. In this study, we aimed to investigate whether phenotypically distinct clonal cell lines representing different subpopulations in the human NP could be generated using immortalization strategies. Nondegenerate healthy disc material (age range, 8 to 15 years) was obtained as surplus surgical material. Early passage NP monolayer cell cultures were initially characterized using a recently established NP marker set. NP cells were immortalized by simian virus 40 large T antigen (SV40LTag) and human telomerase reverse transcriptase expression. Immortalized cells were clonally expanded and characterized based on collagen type I, collagen type II, α1 (COL2A1), and SRY-box 9 (SOX9) protein expression profiles, as well as on expression of a subset of established in vivo NP cell lineage markers. A total of 54 immortal clones were generated. Profiling of a set of novel NP markers (CD24, CA12, PAX1, PTN, FOXF1 and KRT19 mRNA) in a representative set of subclones substantiated successful immortalization of multiple cellular subpopulations from primary isolates and confirmed their NP origin and/or phenotype. We were able to identify two predominant clonal NP subtypes based on their morphological characteristics and their ability to induce SOX9 and COL2A1 under conventional differentiation conditions. In addition, cluster of differentiation 24 (CD24)-negative NP responder clones formed spheroid structures in various culture systems, suggesting the preservation of a more immature phenotype compared to CD24-positive nonresponder clones. Here we report the generation of clonal NP cell lines from nondegenerate human IVD tissue and present a detailed characterization of NP cellular subpopulations. Differential cell surface marker expression and divergent responses to differentiation conditions suggest that the NP subtypes may correspond to distinct maturation stages and represent distinct NP cell subpopulations. Hence, we provide evidence that the immortalization strategy that we applied is capable of detecting cell heterogeneity in the NP. Our cell lines yield novel insights into NP biology and provide promising new tools for studies of IVD development, cell function and disease.

Transcriptional analysis of murine macrophages infected with different Toxoplasma strains identifies novel regulation of host signaling pathways.

PubMed

Melo, Mariane B; Nguyen, Quynh P; Cordeiro, Cynthia; Hassan, Musa A; Yang, Ninghan; McKell, Renée; Rosowski, Emily E; Julien, Lindsay; Butty, Vincent; Dardé, Marie-Laure; Ajzenberg, Daniel; Fitzgerald, Katherine; Young, Lucy H; Saeij, Jeroen P J

2013-01-01

Most isolates of Toxoplasma from Europe and North America fall into one of three genetically distinct clonal lineages, the type I, II and III lineages. However, in South America these strains are rarely isolated and instead a great variety of other strains are found. T. gondii strains differ widely in a number of phenotypes in mice, such as virulence, persistence, oral infectivity, migratory capacity, induction of cytokine expression and modulation of host gene expression. The outcome of toxoplasmosis in patients is also variable and we hypothesize that, besides host and environmental factors, the genotype of the parasite strain plays a major role. The molecular basis for these differences in pathogenesis, especially in strains other than the clonal lineages, remains largely unexplored. Macrophages play an essential role in the early immune response against T. gondii and are also the cell type preferentially infected in vivo. To determine if non-canonical Toxoplasma strains have unique interactions with the host cell, we infected murine macrophages with 29 different Toxoplasma strains, representing global diversity, and used RNA-sequencing to determine host and parasite transcriptomes. We identified large differences between strains in the expression level of known parasite effectors and large chromosomal structural variation in some strains. We also identified novel strain-specifically regulated host pathways, including the regulation of the type I interferon response by some atypical strains. IFNβ production by infected cells was associated with parasite killing, independent of interferon gamma activation, and dependent on endosomal Toll-like receptors in macrophages and the cytoplasmic receptor retinoic acid-inducible gene 1 (RIG-I) in fibroblasts.

Clonal diversity and genetic profiling of antibiotic resistance among multidrug/carbapenem-resistant Klebsiella pneumoniae isolates from a tertiary care hospital in Saudi Arabia.

PubMed

Zaman, Taher Uz; Alrodayyan, Maha; Albladi, Maha; Aldrees, Mohammed; Siddique, Mohammed Ismail; Aljohani, Sameera; Balkhy, Hanan H

2018-05-03

The nexus between resistance determinants, plasmid type, and clonality appears to play a crucial role in the dissemination and survival of carbapenem-resistant Klebsiella pneumoniae (CRKP). The incidence of infections involving CRKP in Saudi Arabia is increasing and there is a need for detailed molecular profiling of this pathogen for CRKP surveillance and control. The resistance determinants of 71 non-redundant CRKP isolates were investigated by polymerase chain reaction (PCR) and sequencing. Plasmid typing was performed using PCR-based replicon typing and the clonality of isolates was determined by multilocus sequence typing. Capsular polysaccharide synthesis genes and other virulence factors were examined using multiplex PCR. Diversity was calculated using DIVEIN, clonal relationship was determined using eBURST, and phylogenetic analysis was performed using SplitsTree4. A polyclonal OXA-48 gene alone was the most common carbapenemase detected in 48/71 (67.6%) isolates followed by NDM-1 alone in 9/71 (12.7%) isolates. Coproduction of OXA-48 and NDM-1 was observed in 6/71 (8.5%) isolates. Both carbapenemase genes could be transferred into an Escherichia coli recipient. CTX-M-15 was the most abundant extended-spectrum β-lactamase gene detected in 47/71 (66.2%) isolates, whereas clone-specific CTX-M-14 (ST-199 and -709) was found in 15/71 (21%) isolates. Sixty-seven of 71 isolates were positive for one or more plasmid replicons. The replicons detected were: IncFII; IncFIIK; IncFIA; IncFIB; L/M; IncI1; and IncN. FIIK and L/M were predominant, with 69 and 67% positivity, respectively. All isolates were negative for the magA (K1), rmpA, and K2 genes and presented a non-hypermucoviscous phenotype. A polyclonal CRKP reservoir of sequence types (STs)-37, - 199, and - 152 was observed and ST-152 appeared to be a "frequent carrier" of the NDM-1 gene. ST-199, a singleton not previously reported, showed a sequence diversity suggestive of positive selection. A significant association was evident between resistance determinants and the clonal types of K. pneumoniae: all ST-152 isolates were positive for NDM-1 but negative for OXA-48; ST-199 isolates were positive for OXA-48 but negative for NDM-1; and ST-709 and -199 isolates were positive for CTX-M-14. The incidence of certain clonal types in large numbers predicts an outbreak-like situation and warrants stringent surveillance and infection control.

A Clonal Lineage of Fusarium oxysporum Circulates in the Tap Water of Different French Hospitals

PubMed Central

Sautour, Marc; Gautheron, Nadine; Laurent, Julie; Aho, Serge; Bonnin, Alain; Sixt, Nathalie; Hartemann, Philippe; Dalle, Frédéric; Steinberg, Christian

2016-01-01

ABSTRACT Fusarium oxysporum is typically a soilborne fungus but can also be found in aquatic environments. In hospitals, water distribution systems may be reservoirs for the fungi responsible for nosocomial infections. F. oxysporum was previously detected in the water distribution systems of five French hospitals. Sixty-eight isolates from water representative of all hospital units that were previously sampled and characterized by translation elongation factor 1α sequence typing were subjected to microsatellite analysis and full-length ribosomal intergenic spacer (IGS) sequence typing. All but three isolates shared common microsatellite loci and a common two-locus sequence type (ST). This ST has an international geographical distribution in both the water networks of hospitals and among clinical isolates. The ST dominant in water was not detected among 300 isolates of F. oxysporum that originated from surrounding soils. Further characterization of 15 isolates by vegetative compatibility testing allowed us to conclude that a clonal lineage of F. oxysporum circulates in the tap water of the different hospitals. IMPORTANCE We demonstrated that a clonal lineage of Fusarium oxysporum inhabits the water distribution systems of several French hospitals. This clonal lineage, which appears to be particularly adapted to water networks, represents a potential risk for human infection and raises questions about its worldwide distribution. PMID:27663024

Clonal mature adipocyte production of proliferative-competent daughter cells requires lipid export prior to cell division

USDA-ARS?s Scientific Manuscript database

Numerous in vitro observations have been published to show that mature adipocytes may resume proliferation and begin to populate the adipofibroblast fraction or form other cell types. In the present study, we evaluated clonal cultures of mature pig-derived adipocytes as they began to reestablish the...

Dispersion of Multidrug-Resistant Enterococcus faecium Isolates Belonging to Major Clonal Complexes in Different Portuguese Settings▿

PubMed Central

Freitas, Ana R.; Novais, Carla; Ruiz-Garbajosa, Patricia; Coque, Teresa M.; Peixe, Luísa

2009-01-01

The population structure of 56 Enterococcus faecium isolates selected from a collection of enterococci from humans, animals, and the environment in Portugal (1997 to 2007) was analyzed by multilocus sequence typing. We identified 41 sequence types clustering into CC17, CC5, CC9, CC22 and CC94, all clonal lineages comprising isolates from different hosts. Our findings highlight the role of community-associated hosts as reservoirs of enterococci able to cause human infections. PMID:19447948

Clonality analysis of lymphoid proliferations using the BIOMED-2 clonality assays: a single institution experience

PubMed Central

Kokovic, Ira; Novakovic, Barbara Jezersek; Cerkovnik, Petra; Novakovic, Srdjan

2014-01-01

Background Clonality determination in patients with lymphoproliferative disorders can improve the final diagnosis. The aim of our study was to evaluate the applicative value of standardized BIOMED-2 gene clonality assay protocols for the analysis of clonality of lymphocytes in a group of different lymphoid proliferations. Materials and methods. With this purpose, 121 specimens from 91 patients with suspected lymphoproliferations submitted for routine diagnostics from January to December 2011 were retrospectively analyzed. According to the final diagnosis, our series comprised 32 cases of B-cell lymphomas, 38 cases of non-Hodgkin’s T-cell lymphomas and 51 cases of reactive lymphoid proliferations. Clonality testing was performed using the BIOMED-2 clonality assays. Results The determined sensitivity of the TCR assay was 91.9%, while the sensitivity of the IGH assay was 74.2%. The determined specificity of the IGH assay was 73.3% in the group of lymphomas and 87.2% in the group of reactive lesions. The determined specificity of the TCR assay was 62.5% in the group of lymphomas and 54.3% in the group of reactive lesions. Conclusions In the present study, we confirmed the utility of standardized BIOMED-2 clonality assays for the detection of clonality in a routine diagnostical setting of non-Hodgkin’s lymphomas. Reactions for the detection of the complete IGH rearrangements and reactions for the detection of the TCR rearrangements are a good choice for clonality testing of a wide range of lymphoid proliferations and specimen types while the reactions for the detection of incomplete IGH rearrangements have not shown any additional diagnostic value. PMID:24991205

Significance of clonal rearrangements of lymphocyte antigen receptor genes on the prognosis of chronic enteropathy in 22 Shiba dogs.

PubMed

Ohmi, Aki; Ohno, Koichi; Uchida, Kazuyuki; Goto-Koshino, Yuko; Tomiyasu, Hirotaka; Kanemoto, Hideyuki; Fukushima, Kenjiro; Tsujimoto, Hajime

2017-09-29

Shiba dogs are predisposed to chronic enteropathy (CE) and have poorer prognosis than other dog breeds. The objective of this study was to investigate the significance of polymerase chain reaction for antigen receptor rearrangement (PARR) results on clinical findings and prognosis of Shiba dogs with CE. We retrospectively collected data on 22 Shiba dogs diagnosed as having CE. Fifty-nine percent of the dogs had clonality-positive results on PARR analysis. Furthermore, on histopathology, epitheliotropic behavior of small lymphocytes of the intestinal mucosa was observed significantly more frequently in dogs with clonal rearrangement of antigen receptor genes (P=0.027). The median overall survival time of clonality-positive dogs was 48 days (range, 4-239 days), compared to 271 days (range, 45-1,316+ days) in clonality-negative dogs. The median overall survival time of epitheliotropism-positive dogs was 76 days (range, 30-349 days) compared to 239 days (range, 4-1,316+ days) for epitheliotropism-negative dogs. Statistical analysis revealed that the clonality-positive result was associated with significantly shorter survival time (P=0.036). In contrast, presence or absence of epitheliotropism had no statistically significant effect on survival time (P=0.223). These cases might appropriately be diagnosed as small T-cell intestinal lymphoma; there are some common clinical and pathogenic features with human enteropathy-associated T-cell lymphoma type 2. The pathogenesis and poor prognosis for Shiba dogs with CE seem to be associated with this type of lymphoma, although further investigation is warranted.

Multilocus sequence type system for the plant pathogen Xylella fastidiosa and relative contributions of recombination and point mutation to clonal diversity.

PubMed

Scally, Mark; Schuenzel, Erin L; Stouthamer, Richard; Nunney, Leonard

2005-12-01

Multilocus sequence typing (MLST) identifies and groups bacterial strains based on DNA sequence data from (typically) seven housekeeping genes. MLST has also been employed to estimate the relative contributions of recombination and point mutation to clonal divergence. We applied MLST to the plant pathogen Xylella fastidiosa using an initial set of sequences for 10 loci (9.3 kb) of 25 strains from five different host plants, grapevine (PD strains), oleander (OLS strains), oak (OAK strains), almond (ALS strains), and peach (PP strains). An eBURST analysis identified six clonal complexes using the grouping criterion that each member must be identical to at least one other member at 7 or more of the 10 loci. These clonal complexes corresponded to previously identified phylogenetic clades; clonal complex 1 (CC1) (all PD strains plus two ALS strains) and CC2 (OLS strains) defined the X. fastidiosa subsp. fastidiosa and X. fastidiosa subsp. sandyi clades, while CC3 (ALS strains), CC4 (OAK strains), and CC5 (PP strains) were subclades of X. fastidiosa subsp. multiplex. CC6 (ALS strains) identified an X. fastidiosa subsp. multiplex-like group characterized by a high frequency of intersubspecific recombination. Compared to the recombination rate in other bacterial species, the recombination rate in X. fastidiosa is relatively low. Recombination between different alleles was estimated to give rise to 76% of the nucleotide changes and 31% of the allelic changes observed. The housekeeping loci holC, nuoL, leuA, gltT, cysG, petC, and lacF were chosen to form the basis of a public database for typing X. fastidiosa (www.mlst.net). These loci identified the same six clonal complexes using the strain grouping criterion of identity at five or more loci with at least one other member.

Clinical implications of somatic mutations in aplastic anemia and myelodysplastic syndrome in genomic age.

PubMed

Maciejewski, Jaroslaw P; Balasubramanian, Suresh K

2017-12-08

Recent technological advances in genomics have led to the discovery of new somatic mutations and have brought deeper insights into clonal diversity. This discovery has changed not only the understanding of disease mechanisms but also the diagnostics and clinical management of bone marrow failure. The clinical applications of genomics include enhancement of current prognostic schemas, prediction of sensitivity or refractoriness to treatments, and conceptualization and selective application of targeted therapies. However, beyond these traditional clinical aspects, complex hierarchical clonal architecture has been uncovered and linked to the current concepts of leukemogenesis and stem cell biology. Detection of clonal mutations, otherwise typical of myelodysplastic syndrome, in the course of aplastic anemia (AA) and paroxysmal nocturnal hemoglobinuria has led to new pathogenic concepts in these conditions and created a new link between AA and its clonal complications, such as post-AA and paroxysmal nocturnal hemoglobinuria. Distinctions among founder vs subclonal mutations, types of clonal evolution (linear or branching), and biological features of individual mutations (sweeping, persistent, or vanishing) will allow for better predictions of the biologic impact they impart in individual cases. As clonal markers, mutations can be used for monitoring clonal dynamics of the stem cell compartment during physiologic aging, disease processes, and leukemic evolution. © 2016 by The American Society of Hematology. All rights reserved.

A rare case of acute toxoplasmosis in a stray dog due to infection of T. gondii clonal type I: public health concern in urban settings with stray animals?

PubMed

Migliore, Sergio; La Marca, Salvatore; Stabile, Cristian; Di Marco Lo Presti, Vincenzo; Vitale, Maria

2017-08-17

Typing of Toxoplasma gondii strains is important in epidemiological surveys, to understand the distribution and virulence of different clones of the parasite among human and animal populations. Stray dogs can be consider sentinel animals for contaminated environments playing an important but probably under- evaluated role in the epidemiology of T. gondii. We reported a rare case of acute toxoplasmosis in a stray dog due to clonal type I infection. The clonal type I, sporadic in Europe, is frequently associated with severe toxoplasmosis in humans and the control of its circulation is particularly relevant for public health. The symptomatology suggested a potential infection with the high similar parasite Neospora caninum but differential diagnosis showed that only T. gondii was involved highlighting the importance of multiple diagnostic methods beyond the clinical signs. A female stray dog approximately six-month of age presented muscular atrophy of the femoral region and hyperextension of hind limbs. Body condition score (BCS) was 20% below ideal weight, ribs had almost no fat and the sensor state was depressed. Haematological values were normal and the dog did not show any neurological abnormalities. Serological analysis showed a positive response for T. gondii immunoglobulin G (IgG) antibodies and exclude N. caninum infection. To confirm T. gondii infection, a muscle biopsy was performed and genomic DNA was extracted. PCR analysis resulted positive to T. gondii and strain genotyping reveals clonal type I infection. The dog recovered after 4 weeks of treatment with clindamycin hydrochloride and aquatic physiotherapy. Our study reports a rare and severe case of T. gondii clonal type I infection in a stray dog feeding in garbage containers. The data confirm the importance of an in vivo early diagnosis for toxoplasmosis in dog. Clinical signs are often related to specific T. gondii genotype and parasite genotyping is important in the epidemiological survey of toxoplasmosis in public health. The detection of parasitic DNA in the tissue could be an useful diagnostic method in facilitating early treatment of the disease, which is important for a timely clinical recovery.

Phenotypic plasticity and specialization in clonal versus non-clonal plants: A data synthesis

NASA Astrophysics Data System (ADS)

Fazlioglu, Fatih; Bonser, Stephen P.

2016-11-01

Reproductive strategies can be associated with ecological specialization and generalization. Clonal plants produce lineages adapted to the maternal habitat that can lead to specialization. However, clonal plants frequently display high phenotypic plasticity (e.g. clonal foraging for resources), factors linked to ecological generalization. Alternately, sexual reproduction can be associated with generalization via increasing genetic variation or specialization through rapid adaptive evolution. Moreover, specializing to high or low quality habitats can determine how phenotypic plasticity is expressed in plants. The specialization hypothesis predicts that specialization to good environments results in high performance trait plasticity and specialization to bad environments results in low performance trait plasticity. The interplay between reproductive strategies, phenotypic plasticity, and ecological specialization is important for understanding how plants adapt to variable environments. However, we currently have a poor understanding of these relationships. In this study, we addressed following questions: 1) Is there a relationship between phenotypic plasticity, specialization, and reproductive strategies in plants? 2) Do good habitat specialists express greater performance trait plasticity than bad habitat specialists? We searched the literature for studies examining plasticity for performance traits and functional traits in clonal and non-clonal plant species from different habitat types. We found that non-clonal (obligate sexual) plants expressed greater performance trait plasticity and functional trait plasticity than clonal plants. That is, non-clonal plants exhibited a specialist strategy where they perform well only in a limited range of habitats. Clonal plants expressed less performance loss across habitats and a more generalist strategy. In addition, specialization to good habitats did not result in greater performance trait plasticity. This result was contrary to the predictions of the specialization hypothesis. Overall, reproductive strategies are associated with ecological specialization or generalization through phenotypic plasticity. While specialization is common in plant populations, the evolution of specialization does not control the nature of phenotypic plasticity as predicted under the specialization hypothesis.

Clonal growth and plant species abundance

PubMed Central

Herben, Tomáš; Nováková, Zuzana; Klimešová, Jitka

2014-01-01

Background and Aims Both regional and local plant abundances are driven by species' dispersal capacities and their abilities to exploit new habitats and persist there. These processes are affected by clonal growth, which is difficult to evaluate and compare across large numbers of species. This study assessed the influence of clonal reproduction on local and regional abundances of a large set of species and compared the predictive power of morphologically defined traits of clonal growth with data on actual clonal growth from a botanical garden. The role of clonal growth was compared with the effects of seed reproduction, habitat requirements and growth, proxied both by LHS (leaf–height–seed) traits and by actual performance in the botanical garden. Methods Morphological parameters of clonal growth, actual clonal reproduction in the garden and LHS traits (leaf-specific area – height – seed mass) were used as predictors of species abundance, both regional (number of species records in the Czech Republic) and local (mean species cover in vegetation records) for 836 perennial herbaceous species. Species differences in habitat requirements were accounted for by classifying the dataset by habitat type and also by using Ellenberg indicator values as covariates. Key Results After habitat differences were accounted for, clonal growth parameters explained an important part of variation in species abundance, both at regional and at local levels. At both levels, both greater vegetative growth in cultivation and greater lateral expansion trait values were correlated with higher abundance. Seed reproduction had weaker effects, being positive at the regional level and negative at the local level. Conclusions Morphologically defined traits are predictive of species abundance, and it is concluded that simultaneous investigation of several such traits can help develop hypotheses on specific processes (e.g. avoidance of self-competition, support of offspring) potentially underlying clonal growth effects on abundance. Garden performance parameters provide a practical approach to assessing the roles of clonal growth morphological traits (and LHS traits) for large sets of species. PMID:24482153

Cluster of Serogroup W135 Meningococci, Southeastern Florida, 2008–2009

PubMed Central

Mejia-Echeverry, Alvaro; Fiorella, Paul; Leguen, Fermin; Livengood, John; Kay, Robyn; Hopkins, Richard

2010-01-01

Recently, 14 persons in southeastern Florida were identified with Neisseria meningitidis serogroup W135 invasive infections. All isolates tested had matching or near-matching pulsed-field gel electrophoresis patterns and belonged to the multilocus sequence type 11 clonal complex. The epidemiologic investigation suggested recent endemic transmission of this clonal complex in southeastern Florida. PMID:20031054

Clonal origins of Vibrio cholerae O1 El Tor strains, Papua New Guinea, 2009-2011.

PubMed

Horwood, Paul F; Collins, Deirdre; Jonduo, Marinjho H; Rosewell, Alexander; Dutta, Samir R; Dagina, Rosheila; Ropa, Berry; Siba, Peter M; Greenhill, Andrew R

2011-11-01

We used multilocus sequence typing and variable number tandem repeat analysis to determine the clonal origins of Vibrio cholerae O1 El Tor strains from an outbreak of cholera that began in 2009 in Papua New Guinea. The epidemic is ongoing, and transmission risk is elevated within the Pacific region.

Genotypes and Mouse Virulence of Toxoplasma gondii Isolates from Animals and Humans in China

PubMed Central

Liu, Daohua; Huo, Xingxing; Gao, Jiangmei; Song, Xiaorong; Xu, Xiucai; Huang, Kaiquan; Liu, Wenqi; Wang, Yong; Lu, Fangli; Lun, Zhao-Rong; Luo, Qingli; Wang, Xuelong; Shen, Jilong

2013-01-01

Background Recent population structure studies of T. gondii revealed that a few major clonal lineages predominated in different geographical regions. T. gondii in South America is genetically and biologically divergent, whereas this parasite is remarkably clonal in North America and Europe with a few major lineages including Types I, II and III. Information on genotypes and mouse virulence of T. gondii isolates from China is scarce and insufficient to investigate its population structure, evolution, and transmission. Methodology/Principal Findings Genotyping of 23 T. gondii isolates from different hosts using 10 markers for PCR-restriction fragment length polymorphism analyses (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico) revealed five genotypes; among them three genotypes were atypical and two were archetypal. Fifteen strains belong to the Chinese 1 lineage, which has been previously reported as a widespread lineage from swine, cats, and humans in China. Two human isolates fall into the type I and II lineages and the remaining isolates belong to two new atypical genotypes (ToxoDB#204 and #205) which has never been reported in China. Our results show that these genotypes of T. gondii isolates are intermediately or highly virulent in mice except for the strain TgCtwh6, which maintained parasitemia in mice for 35 days post infection although it possesses the uniform genotype of Chinese 1. Additionally, phylogenetic network analyses of all isolates of genotype Chinese 1 are identical, and there is no variation based on the sequence data generated for four introns (EF1, HP2, UPRT1 and UPRT7) and two dense granule proteins (GRA6 and GRA7). Conclusion/Significance A limited genetic diversity was found and genotype Chinese 1 (ToxoDB#9) is dominantly circulating in mainland China. The results will provide a useful profile for deep insight to the population structure, epidemiology and biological characteristics of T. gondii in China. PMID:23308233

Clonal plasticity and diversity facilitates the adaptation of Rhododendron aureum Georgi to alpine environment

PubMed Central

Wang, Xiaolong; Zhao, Wei; Li, Lin; You, Jian; Ni, Biao

2018-01-01

Four small oval populations and five large intensive populations of Rhododendron aureum growing at the alpine in Changbai Mountain (China) were studied in two types of habitat (in the tundra and in Betula ermanii forest). Identification and delimitation of genets were inferred from excavation in small populations and from amplified fragment length polymorphism (AFLP) markers by the standardized sampling design in large populations. Clonal architecture and clonal diversity were then estimated. For the four small populations, they were monoclonal, the spacer length (18.6 ± 5.6 in tundra, 29.7 ± 9.7 in Betula ermanii forest, P < 0.05) was shorter and branching intensity (136.7 ± 32.9 in tundra, 43.4 ± 12.3 in Betula ermanii forest, P < 0.05) was higher in the tundra than that in Betula ermanii forest. For the five large populations, they were composed of multiple genets with high level of clonal diversity (Simpson’s index D = 0.84, clonal richness R = 0.25, Fager's evenness E = 0.85); the spatial distribution of genets showed that the clonal growth strategy of R. aureum exhibits both guerilla and phalanx. Our results indicate that the clonal plasticity of R. aureum could enhance exploitation of resource heterogeneity and in turn greatly contribute to maintenance or improvement of fitness and the high clonal diversity of R. aureum increase the evolutionary rates to adapt the harsh alpine environment in Changbai Mountain. PMID:29746526

Biomass Allocation of Stoloniferous and Rhizomatous Plant in Response to Resource Availability: A Phylogenetic Meta-Analysis

PubMed Central

Xie, Xiu-Fang; Hu, Yu-Kun; Pan, Xu; Liu, Feng-Hong; Song, Yao-Bin; Dong, Ming

2016-01-01

Resource allocation to different functions is central in life-history theory. Plasticity of functional traits allows clonal plants to regulate their resource allocation to meet changing environments. In this study, biomass allocation traits of clonal plants were categorized into absolute biomass for vegetative growth vs. for reproduction, and their relative ratios based on a data set including 115 species and derived from 139 published literatures. We examined general pattern of biomass allocation of clonal plants in response to availabilities of resource (e.g., light, nutrients, and water) using phylogenetic meta-analysis. We also tested whether the pattern differed among clonal organ types (stolon vs. rhizome). Overall, we found that stoloniferous plants were more sensitive to light intensity than rhizomatous plants, preferentially allocating biomass to vegetative growth, aboveground part and clonal reproduction under shaded conditions. Under nutrient- and water-poor condition, rhizomatous plants were constrained more by ontogeny than by resource availability, preferentially allocating biomass to belowground part. Biomass allocation between belowground and aboveground part of clonal plants generally supported the optimal allocation theory. No general pattern of trade-off was found between growth and reproduction, and neither between sexual and clonal reproduction. Using phylogenetic meta-analysis can avoid possible confounding effects of phylogeny on the results. Our results shown the optimal allocation theory explained a general trend, which the clonal plants are able to plastically regulate their biomass allocation, to cope with changing resource availability, at least in stoloniferous and rhizomatous plants. PMID:27200071

A dominant lineage of Mycoplasma bovis is associated with an increased number of severe mastitis cases in cattle.

PubMed

Bürki, Sibylle; Spergser, Joachim; Bodmer, Michèle; Pilo, Paola

2016-11-30

Mycoplasma bovis is the most frequent etiologic agent of bovine mycoplasmosis. It causes various diseases in bovines and considerable economic loss due to the lack of effective treatment or preventive measures such as vaccination. In contrast to the US, where M. bovis-mastitis has been reported for a long time, M. bovis infections in Switzerland and Austria were predominantly associated with pneumonia and subclinical mastitis. However, since 2007 the situation has changed with the emergence of severe M. bovis-associated mastitis cases in both countries. In order to evaluate the molecular epidemiology of the bacteria isolated from these infections, recent and old Swiss, along with recent Austrian M. bovis isolates were analyzed by a typing method displaying intermediate resolution of evolutionary relationships among isolates called Multi-Locus Sequence Typing (MLST). The analysis of Swiss and Austrian M. bovis isolates revealed two major lineages. Isolates collected since 2007 in both countries cluster in the lineage I including ST5, ST33, ST34, 36, and ST38-40 (clonal complex 1), while all Swiss isolates recovered before 2007 cluster in the lineage II comprising ST17 and ST35 (clonal complex 5). Further investigations are necessary to understand if lineage I has a higher predilection or virulence toward mammary gland cells than the old lineage or if other factors are involved in the increased number of severe mastitis cases. Copyright © 2016 Elsevier B.V. All rights reserved.

[Actinic keratosis, Bowen's disease, keratoacanthoma and squamous cell carcinoma of the skin].

PubMed

Majores, M; Bierhoff, E

2015-02-01

Actinic (solar) keratosis is an intraepidermal squamous neoplasm of sun-damaged skin and by far the most frequent neoplastic skin lesion. A subdivison into three grades has been proposed with increasing acceptance not least because of the therapeutic consequences. The transition to invasive squamous cell carcinoma is reported in 5-10 % and with immunosuppression in 30 % of patients.Bowen's disease is a variant of squamous cell carcinoma in situ of the skin and the mucocutaneous junction. The differentiation from bowenoid papulosis as a lesion associated with human papillomavirus (HPV), actinic (solar) keratosis grade III, intraepidermal poroid lesions and in cases of clonal type from clonal seborrhoic keratosis and Paget's disease is very important.Keratoacanthoma is currently uniformly interpreted as a variant of highly differentiated squamous cell carcinoma of the skin with clinical and histomorphological characteristics. Clinically keratoacanthoma erupts rapidly and is capable of resolving spontaneously. Histologically, there is a characteristic growth pattern and various stages of regression. The final histomorphological diagnosis needs the entire specimen.Squamous cell carcinoma of the skin is the second most common type of skin cancer following basal cell carcinoma. With respect to reccurrencies and risk of metastases the subtyping of cutaneous squamous cell carcinoma is very important. The classification system of the Union Internationale Contra le Cancer (UICC) is based solely on the anatomical spread and the classification system of the American Joint Committee on Cancer (AJCC) also considers so-called high-risk features in the staging between stages I and II.

Advances for Studying Clonal Evolution in Cancer

PubMed Central

Raphael, Benjamin J.; Chen, Feng; Wendl, Michael C.

2013-01-01

The “clonal evolution” model of cancer emerged and “evolved” amid ongoing advances in technology, especially in recent years during which next generation sequencing instruments have provided ever higher resolution pictures of the genetic changes in cancer cells and heterogeneity in tumors. It has become increasingly clear that clonal evolution is not a single sequential process, but instead frequently involves simultaneous evolution of multiple subclones that co-exist because they are of similar fitness or are spatially separated. Co-evolution of subclones also occurs when they complement each other’s survival advantages. Recent studies have also shown that clonal evolution is highly heterogeneous: different individual tumors of the same type may undergo very different paths of clonal evolution. New methodological advancements, including deep digital sequencing of a mixed tumor population, single cell sequencing, and the development of more sophisticated computational tools, will continue to shape and reshape the models of clonal evolution. In turn, these will provide both an improved framework for the understanding of cancer progression and a guide for treatment strategies aimed at the elimination of all, rather than just some, of the cancer cells within a patient. PMID:23353056

Advances for studying clonal evolution in cancer.

PubMed

Ding, Li; Raphael, Benjamin J; Chen, Feng; Wendl, Michael C

2013-11-01

The "clonal evolution" model of cancer emerged and "evolved" amid ongoing advances in technology, especially in recent years during which next generation sequencing instruments have provided ever higher resolution pictures of the genetic changes in cancer cells and heterogeneity in tumors. It has become increasingly clear that clonal evolution is not a single sequential process, but instead frequently involves simultaneous evolution of multiple subclones that co-exist because they are of similar fitness or are spatially separated. Co-evolution of subclones also occurs when they complement each other's survival advantages. Recent studies have also shown that clonal evolution is highly heterogeneous: different individual tumors of the same type may undergo very different paths of clonal evolution. New methodological advancements, including deep digital sequencing of a mixed tumor population, single cell sequencing, and the development of more sophisticated computational tools, will continue to shape and reshape the models of clonal evolution. In turn, these will provide both an improved framework for the understanding of cancer progression and a guide for treatment strategies aimed at the elimination of all, rather than just some, of the cancer cells within a patient. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Evolution of oesophageal adenocarcinoma from metaplastic columnar epithelium without goblet cells in Barrett's oesophagus.

PubMed

Lavery, Danielle L; Martinez, Pierre; Gay, Laura J; Cereser, Biancastella; Novelli, Marco R; Rodriguez-Justo, Manuel; Meijer, Sybren L; Graham, Trevor A; McDonald, Stuart A C; Wright, Nicholas A; Jansen, Marnix

2016-06-01

Barrett's oesophagus commonly presents as a patchwork of columnar metaplasia with and without goblet cells in the distal oesophagus. The presence of metaplastic columnar epithelium with goblet cells on oesophageal biopsy is a marker of cancer progression risk, but it is unclear whether clonal expansion and progression in Barrett's oesophagus is exclusive to columnar epithelium with goblet cells. We developed a novel method to trace the clonal ancestry of an oesophageal adenocarcinoma across an entire Barrett's segment. Clonal expansions in Barrett's mucosa were identified using cytochrome c oxidase enzyme histochemistry. Somatic mutations were identified through mitochondrial DNA sequencing and single gland whole exome sequencing. By tracing the clonal origin of an oesophageal adenocarcinoma across an entire Barrett's segment through a combination of histopathological spatial mapping and clonal ordering, we find that this cancer developed from a premalignant clonal expansion in non-dysplastic ('cardia-type') columnar metaplasia without goblet cells. Our data demonstrate the premalignant potential of metaplastic columnar epithelium without goblet cells in the context of Barrett's oesophagus. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

Genetic diversity of Toxoplasma gondii isolates from Ethiopian feral cats.

PubMed

Dubey, J P; Choudhary, S; Tilahun, G; Tiao, N; Gebreyes, W A; Zou, X; Su, C

2013-09-01

Recent studies indicate greater genetic variability among isolates of Toxoplasma gondii worldwide than previously thought. However, there is no information on genetic diversity of T. gondii from any host in Ethiopia. In the present study, genotyping was performed on viable T. gondii isolates by bioassays in mice from tissues and feces of 27 cats from Ethiopia. Viable T. gondii was isolated from hearts of 26 cats, feces alone of 1 cat, and feces and tissues of 6 cats; in total there were 33 isolates. Genotyping was performed on DNA from cell-cultured derived T. gondii tachyzoites and by using 10 PCR-restriction fragment length polymorphism markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico). Four genotypes were recognized, including ToxoDB #1 (Type II clonal, nine isolates), ToxoDB #2 (Type III, five isolates), Toxo DB #3 (Type II variant, ten isolates), and ToxoDB #20 (nine isolates). Of interest is the isolation of different genotypes from tissues and feces of two cats, suggesting re-infection or mixed strain T. gondii infection. These findings are of epidemiological significance with respect to shedding of oocysts by cats. This is the first report of genotyping of T. gondii from any host in Ethiopia. Published by Elsevier B.V.

Multilocus Sequence Typing Analysis of Staphylococcus lugdunensis Implies a Clonal Population Structure

PubMed Central

Chassain, Benoît; Lemée, Ludovic; Didi, Jennifer; Thiberge, Jean-Michel; Brisse, Sylvain; Pons, Jean-Louis

2012-01-01

Staphylococcus lugdunensis is recognized as one of the major pathogenic species within the genus Staphylococcus, even though it belongs to the coagulase-negative group. A multilocus sequence typing (MLST) scheme was developed to study the genetic relationships and population structure of 87 S. lugdunensis isolates from various clinical and geographic sources by DNA sequence analysis of seven housekeeping genes (aroE, dat, ddl, gmk, ldh, recA, and yqiL). The number of alleles ranged from four (gmk and ldh) to nine (yqiL). Allelic profiles allowed the definition of 20 different sequence types (STs) and five clonal complexes. The 20 STs lacked correlation with geographic source. Isolates recovered from hematogenic infections (blood or osteoarticular isolates) or from skin and soft tissue infections did not cluster in separate lineages. Penicillin-resistant isolates clustered mainly in one clonal complex, unlike glycopeptide-tolerant isolates, which did not constitute a distinct subpopulation within S. lugdunensis. Phylogenies from the sequences of the seven individual housekeeping genes were congruent, indicating a predominantly mutational evolution of these genes. Quantitative analysis of the linkages between alleles from the seven loci revealed a significant linkage disequilibrium, thus confirming a clonal population structure for S. lugdunensis. This first MLST scheme for S. lugdunensis provides a new tool for investigating the macroepidemiology and phylogeny of this unusually virulent coagulase-negative Staphylococcus. PMID:22785196

A Clonal Lineage of Fusarium oxysporum Circulates in the Tap Water of Different French Hospitals.

PubMed

Edel-Hermann, Véronique; Sautour, Marc; Gautheron, Nadine; Laurent, Julie; Aho, Serge; Bonnin, Alain; Sixt, Nathalie; Hartemann, Philippe; Dalle, Frédéric; Steinberg, Christian

2016-11-01

Fusarium oxysporum is typically a soilborne fungus but can also be found in aquatic environments. In hospitals, water distribution systems may be reservoirs for the fungi responsible for nosocomial infections. F. oxysporum was previously detected in the water distribution systems of five French hospitals. Sixty-eight isolates from water representative of all hospital units that were previously sampled and characterized by translation elongation factor 1α sequence typing were subjected to microsatellite analysis and full-length ribosomal intergenic spacer (IGS) sequence typing. All but three isolates shared common microsatellite loci and a common two-locus sequence type (ST). This ST has an international geographical distribution in both the water networks of hospitals and among clinical isolates. The ST dominant in water was not detected among 300 isolates of F. oxysporum that originated from surrounding soils. Further characterization of 15 isolates by vegetative compatibility testing allowed us to conclude that a clonal lineage of F. oxysporum circulates in the tap water of the different hospitals. We demonstrated that a clonal lineage of Fusarium oxysporum inhabits the water distribution systems of several French hospitals. This clonal lineage, which appears to be particularly adapted to water networks, represents a potential risk for human infection and raises questions about its worldwide distribution. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Clonal group distribution of fluoroquinolone-resistant Escherichia coli among humans and companion animals in Australia.

PubMed

Platell, Joanne L; Cobbold, Rowland N; Johnson, James R; Trott, Darren J

2010-09-01

To determine the phylogenetic group distribution and prevalence of three major globally disseminated clonal groups [clonal group A (CGA) and O15:K52:H1, associated with phylogenetic group D, and sequence type ST131, associated with phylogenetic group B2] among fluoroquinolone-resistant extra-intestinal Escherichia coli isolates from humans and companion animals in Australia. Clinical extra-intestinal fluoroquinolone-resistant E. coli isolates were obtained from humans (n = 582) and companion animals (n = 125), on Australia's east coast (October 2007-October 2009). Isolates were tested for susceptibility to seven antimicrobial agents, and for phylogenetic group, O type and clonal-group-specific single nucleotide polymorphisms by PCR. The fluoroquinolone-resistant isolates were typically resistant to multiple agents (median of four). Analysis revealed that clonal group ST131 accounted for a large subset of the human isolates (202/585, 35%), but for a much smaller proportion of the companion animal isolates (9/125, 7.2%; P

Vibrio parahaemolyticus isolates from southeastern Chinese coast are genetically diverse with circulation of clonal complex 3 strains since 2002.

PubMed

Yu, Ying; Hu, Weizhao; Wu, Beibei; Zhang, Peipei; Chen, Jianshun; Wang, Shuna; Fang, Weihuan

2011-11-01

Multilocus sequence typing (MLST) was used to examine the clonal relationship and genetic diversity of 71 Vibrio parahaemolyticus isolates from clinical and seafood-related sources in southeastern Chinese coast between 2002 and 2009. The tested isolates fell into 61 sequence types (STs). Of 17 clinical isolates, 7 belonged to ST3 of the pandemic clonal complex 3, with 3 strains isolated in 2002. Although there was no apparent clonal relationship found between clinical strains and those from seafood-related sources positive with pathogenic markers, there were clonal relationships between clinical strains from this study and those from environmental sources in other parts of China. Phylogenetic analysis showed that strains of 112 STs (61 STs from this study and 51 retrieved from PUBMLST database covering different continents) could be divided into four branches. The vast majority of our isolates and those from other countries were genetically diverse and clustered into two major branches of mixed distribution (of geographic origins and sample sources), whereas five STs representing six isolates split as two minor branches because of divergence of their recA genes, which had 80%-82% nucleotide identity to typical V. parahaemolyticus strains and 73.3%-76.9% identity to the CDS24 of a Vibrio sp. plasmid p23023, indicating that the recA gene might have recombined by lateral gene transfer. This was further supported by a high ratio of recombination to mutation (3.038) for recA. In conclusion, MLST with fully extractable database is a powerful system for analysis of clonal relationship for strains of a particular region in a national or global scale as well as between clinical and environmental or food-related strains.

Genetic Heterogeneity and Clonal Evolution of Tumor Cells and their Impact on Precision Cancer Medicine.

PubMed

Sabaawy, Hatem E

2013-11-18

The efficacy of targeted therapies in leukemias and solid tumors depends upon the accurate detection and sustained targeting of initial and evolving driver mutations and/or aberrations in cancer cells. Tumor clonal evolution of the diverse populations of cancer cells during cancer progression contributes to the longitudinal variations of clonal, morphological, anatomical, and molecular heterogeneity of tumors. Moreover, drug-resistant subclones present at initiation of therapy or emerging as a result of targeted therapies represent major challenges for achieving success of personalized therapies in providing meaningful improvement in cancer survival rates. Here, I briefly portray tumor cell clonal evolution at the cellular and molecular levels, and present the multiple types of genetic heterogeneity in tumors, with a focus on their impact on the implementation of personalized or precision cancer medicine.

Clonal Origins of Vibrio cholerae O1 El Tor Strains, Papua New Guinea, 2009–2011

PubMed Central

Collins, Deirdre; Jonduo, Marinjho H.; Rosewell, Alexander; Dutta, Samir R.; Dagina, Rosheila; Ropa, Berry; Siba, Peter M.; Greenhill, Andrew R.

2011-01-01

We used multilocus sequence typing and variable number tandem repeat analysis to determine the clonal origins of Vibrio cholerae O1 El Tor strains from an outbreak of cholera that began in 2009 in Papua New Guinea. The epidemic is ongoing, and transmission risk is elevated within the Pacific region. PMID:22099099

A comparative study of Cutibacterium (Propionibacterium) acnes clones from acne patients and healthy controls.

PubMed

Lomholt, H B; Scholz, C F P; Brüggemann, H; Tettelin, H; Kilian, M

2017-10-01

Cutibacterium (Propionibacterium) acnes is assumed to play an important role in the pathogenesis of acne. To examine if clones with distinct virulence properties are associated with acne. Multiple C. acnes isolates from follicles and surface skin of patients with moderate to severe acne and healthy controls were characterized by multilocus sequence typing. To determine if CC18 isolates from acne patients differ from those of controls in the possession of virulence genes or lack of genes conducive to a harmonious coexistence the full genomes of dominating CC18 follicular clones from six patients and five controls were sequenced. Individuals carried one to ten clones simultaneously. The dominating C. acnes clones in follicles from acne patients were exclusively from the phylogenetic clade I-1a and all belonged to clonal complex CC18 with the exception of one patient dominated by the worldwide-disseminated and often antibiotic resistant clone ST3. The clonal composition of healthy follicles showed a more heterogeneous pattern with follicles dominated by clones representing the phylogenetic clades I-1a, I-1b, I-2 and II. Comparison of follicular CC18 gene contents, allelic versions of putative virulence genes and their promoter regions, and 54 variable-length intragenic and inter-genic homopolymeric tracts showed extensive conservation and no difference associated with the clinical origin of isolates. The study supports that C. acnes strains from clonal complex CC18 and the often antibiotic resistant clone ST3 are associated with acne and suggests that susceptibility of the host rather than differences within these clones may determine the clinical outcome of colonization. Copyright © 2017 Elsevier Ltd. All rights reserved.

Pre-leukemic clonal hematopoiesis and the risk of therapy-related myeloid neoplasms: a case-control study

PubMed Central

Takahashi, Koichi; Wang, Feng; Kantarjian, Hagop; Doss, Denaha; Khanna, Kanhav; Thompson, Erika; Zhao, Li; Patel, Keyur; Neelapu, Sattva; Gumbs, Curtis; Bueso-Ramos, Carlos; DiNardo, Courtney D; Colla, Simona; Ravandi, Farhad; Zhang, Jianhua; Huang, Xuelin; Wu, Xifeng; Samaniego, Felipe; Garcia-Manero, Guillermo; Andrew Futreal, P.

2017-01-01

Background Therapy-related myeloid neoplasms (t-MNs) are often fatal secondary malignancies. Risk factors for t-MNs are not well understood. Recent studies suggested that individuals with clonal hematopoiesis have higher risk of developing hematological malignancies. We hypothesized that cancer patients with clonal hematopoiesis have increased risk of developing t-MNs. Methods We conducted a retrospective case-control study to compare the prevalence of clonal hematopoiesis between patients who developed t-MNs (cases) and who did not develop t-MNs (control). For cases, we studied14 patients with various types of cancers who developed t-MNs and whose paired samples of t-MN bone marrow (BM) and peripheral blood (PB) that were previously obtained at the time of primary cancer diagnosis were available. Fifty four patients with lymphoma who received combination chemotherapy and did not develop t-MNs after at least 5 years of follow up were studied as a control. We performed molecular barcode sequencing of 32 genes on the pre-treatment PB samples to detect clonal hematopoiesis. For the t-MN cases, we also performed targeted gene sequencing on t-MN BM samples and investigated clonal evolution from clonal hematopoiesis to t-MNs. To confirm association between clonal hematopoiesis and t-MN development, we also analyzed prevalence of clonal hematopoiesis in a separate cohort of 74 patients with lymphoma. All of these patients were treated under the prospective randomized trial of frontline chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) with or without melatonin and 5 (7%) of them had developed t-MNs. Findings In 14 patients with t-MNs, we detected pre-leukemic mutations in 10 of their prior PB samples (71%). In control, clonal hematopoiesis was detected in 17 patients (31%), and the cumulative incidence of t-MNs at 5 years was significantly higher in patients with clonal hematopoiesis (30% [95% CI: 16% – 51%] vs. 7% [95% CI: 2% – 21%], P = 0.016). In the separate cohort, 5 patients (7%) developed t-MNs and 4 (80%) of them had clonal hematopoiesis. The cumulative incidence of t-MNs at 10 years was significantly higher in patients with clonal hematopoiesis (29% [95% CI: 8%–53%] vs. 0% [95% CI: 0%–0%], P = 0.0009). Multivariate Fine and Gray model showed that having clonal hematopoiesis significantly increased the risk of t-MN development (HR = 13.7, P = 0.013). Interpretation Pre-leukemic clonal hematopoiesis is frequently detected in patients with t-MNs at the time of their primary cancer diagnosis and before patients were exposed to chemotherapy/radiation therapy. Detection of clonal hematopoiesis significantly increased the risk of t-MN development in patients with lymphoma. These data suggest potential approaches of screening clonal hematopoiesis in cancer patients to identify patients at risk of t-MNs and warrants a validation in prospective trial investigating a role of clonal hematopoiesis as a predictive marker for t-MNs. PMID:27923552

Stable MSAP markers for the distinction of Vitis vinifera cv Pinot noir clones.

PubMed

Ocaña, Juan; Walter, Bernard; Schellenbaum, Paul

2013-11-01

Grapevine is one of the most economically important fruit crops. Molecular markers have been used to study grapevine diversity. For instance, simple sequence repeats are a powerful tool for identification of grapevine cultivars, while amplified fragment length polymorphisms have shown their usefulness in intra-varietal diversity studies. Other techniques such as sequence-specific amplified polymorphism are based on the presence of mobile elements in the genome, but their detection lies upon their activity. Relevant attention has been drawn toward epigenetic sources of variation. In this study, a set of Vitis vinifera cv Pinot noir clones were analyzed using the methylation-sensitive amplified polymorphism technique with isoschizomers MspI and HpaII. Nine out of fourteen selective primer combinations were informative and generated two types of polymorphic fragments which were categorized as "stable" and "unstable." In total, 23 stable fragments were detected and they discriminated 92.5 % of the studied clones. Detected stable polymorphisms were either common to several clones, restricted to a few clones or unique to a single clone. The identification of these stable epigenetic markers will be useful in clonal diversity studies. We highlight the relevance of stable epigenetic variation in V. vinifera clones and analyze at which level these markers could be applicable for the development of forthright techniques for clonal distinction.

Genetic abnormalities in leukemia secondary to treatment in patients with Hodgkin's disease.

PubMed

Salas, Consuelo; Pérez-Vera, Patricia; Frías, Sara

2011-01-01

Hodgkin's disease has been treated mainly with two chemotherapy schedules, MOPP (nitrogen mustard, Oncovin, procarbazine and prednisone), which includes alkylating agents, and ABVD (adriamycin, bleomycin, vinblastine and dacarbazine), which includes topoisomerase II inhibitors, either with or without radiation therapy. Due to the types of agents used, patients with Hodgkin's disease often develop secondary leukemias. The alkylating agents included in the MOPP scheme were the first drugs associated with the development of therapy-related myelodysplastic syndrome (t-MDS) and acute myeloid leukemia (t-AML); both entities are the result of the clonal selection of cells with accumulated genomic lesions induced by antineoplastic therapy. In patients who developed t-MDS and t-AML, eight alternative routes with specific cytogenetic and molecular changes have been identified, and the routes are related to the type of therapy, alkylating agents or DNA topoisomerase II inhibitors. At the cytogenetic level, patients treated with alkylating agents show deletion 5q/monosomy 5 and deletion 7q/monosomy 7; in contrast, those who were treated with topoisomerase II inhibitors show 11q23 translocations involving the MLL gene. At the molecular level, there are two types of mutations: Class I, which alter the RAS-BRAF signal transduction pathways and increase cell proliferation; Class II, which disrupt genes that encode transcription factors and NPM1 that are involved in cell differentiation, and the inactivation of p53 tumor suppressor gene. Knowledge of the genetic alterations in these conditions is important for the classification, treatment and prognosis of patients as well as essential for increasing the knowledge of the biology of these diseases, which leads to identifying potential therapeutic targets.

[Study of androgen receptor and phosphoglycerate kinase gene polymorphism in major cellular components of the so-called pulmonary sclerosing hemangioma].

PubMed

Qi, Feng-jie; Zhang, Xiu-wei; Zhang, Yong-xing; Dai, Shun-dong; Wang, En-hua

2006-05-01

To study the clonality of polygonal cells and surface cuboidal cells in the so-called pulmonary sclerosing hemangioma (PSH). 17 female surgically resected PSH were found. The polygonal cells and surface cuboidal cells of the 17 PSH cases were microdissected from routine hematoxylin and eosin-stained sections. Genomic DNA was extracted, pretreated through incubation with methylation-sensitive restrictive endonuclease HhaI or HpaII, and amplified by nested polymerase chain reaction for X chromosome-linked androgen receptor (AR) and phosphoglycerate kinase (PGK) genes. The length polymorphism of AR gene was demonstrated by denaturing polyacrylamide gel electrophoresis and silver staining. The PGK gene products were treated with Bst XI and resolved on agarose gel. Amongst the 17 female cases of PSH, 15 samples were successfully amplified for AR and PGK genes. The rates of polymorphism were 53% (8/15) and 27% (4/15) for AR and PGK genes respectively. Polygonal cells and surface cuboidal cells of 10 cases which were suitable for clonality study, showed the same loss of alleles (clonality ratio = 0) or unbalanced methylation pattern (clonality ratio < 0.25). The polygonal cells and surface cuboidal cells in PSH demonstrate patterns of monoclonal proliferation, indicating that both represent true neoplastic cells.

Individual crypt genetic heterogeneity and the origin of metaplastic glandular epithelium in human Barrett’s oesophagus

PubMed Central

Leedham, S J; Preston, S L; McDonald, S A C; Elia, G; Bhandari, P; Poller, D; Harrison, R; Novelli, M R; Jankowski, J A; Wright, N A

2008-01-01

Objectives: Current models of clonal expansion in human Barrett’s oesophagus are based upon heterogenous, flow-purified biopsy analysis taken at multiple segment levels. Detection of identical mutation fingerprints from these biopsy samples led to the proposal that a mutated clone with a selective advantage can clonally expand to fill an entire Barrett’s segment at the expense of competing clones (selective sweep to fixation model). We aimed to assess clonality at a much higher resolution by microdissecting and genetically analysing individual crypts. The histogenesis of Barrett’s metaplasia and neo-squamous islands has never been demonstrated. We investigated the oesophageal gland squamous ducts as the source of both epithelial sub-types. Methods: Individual crypts across Barrett’s biopsy and oesophagectomy blocks were dissected. Determination of tumour suppressor gene loss of heterozygosity patterns, p16 and p53 point mutations were carried out on a crypt-by-crypt basis. Cases of contiguous neo-squamous islands and columnar metaplasia with oesophageal squamous ducts were identified. Tissues were isolated by laser capture microdissection and genetically analysed. Results: Individual crypt dissection revealed mutation patterns that were masked in whole biopsy analysis. Dissection across oesophagectomy specimens demonstrated marked clonal heterogeneity, with multiple independent clones present. We identified a p16 point mutation arising in the squamous epithelium of the oesophageal gland duct, which was also present in a contiguous metaplastic crypt, whereas neo-squamous islands arising from squamous ducts were wild-type with respect to surrounding Barrett’s dysplasia. Conclusions: By studying clonality at the crypt level we demonstrate that Barrett’s heterogeneity arises from multiple independent clones, in contrast to the selective sweep to fixation model of clonal expansion previously described. We suggest that the squamous gland ducts situated throughout the oesophagus are the source of a progenitor cell that may be susceptible to gene mutation resulting in conversion to Barrett’s metaplastic epithelium. Additionally, these data suggest that wild-type ducts may be the source of neo-squamous islands. PMID:18305067

Clonal growth and plant species abundance.

PubMed

Herben, Tomáš; Nováková, Zuzana; Klimešová, Jitka

2014-08-01

Both regional and local plant abundances are driven by species' dispersal capacities and their abilities to exploit new habitats and persist there. These processes are affected by clonal growth, which is difficult to evaluate and compare across large numbers of species. This study assessed the influence of clonal reproduction on local and regional abundances of a large set of species and compared the predictive power of morphologically defined traits of clonal growth with data on actual clonal growth from a botanical garden. The role of clonal growth was compared with the effects of seed reproduction, habitat requirements and growth, proxied both by LHS (leaf-height-seed) traits and by actual performance in the botanical garden. Morphological parameters of clonal growth, actual clonal reproduction in the garden and LHS traits (leaf-specific area - height - seed mass) were used as predictors of species abundance, both regional (number of species records in the Czech Republic) and local (mean species cover in vegetation records) for 836 perennial herbaceous species. Species differences in habitat requirements were accounted for by classifying the dataset by habitat type and also by using Ellenberg indicator values as covariates. After habitat differences were accounted for, clonal growth parameters explained an important part of variation in species abundance, both at regional and at local levels. At both levels, both greater vegetative growth in cultivation and greater lateral expansion trait values were correlated with higher abundance. Seed reproduction had weaker effects, being positive at the regional level and negative at the local level. Morphologically defined traits are predictive of species abundance, and it is concluded that simultaneous investigation of several such traits can help develop hypotheses on specific processes (e.g. avoidance of self-competition, support of offspring) potentially underlying clonal growth effects on abundance. Garden performance parameters provide a practical approach to assessing the roles of clonal growth morphological traits (and LHS traits) for large sets of species. © The Author 2014. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Wide spread of OXA-23-producing carbapenem-resistant Acinetobacter baumannii belonging to clonal complex II in different hospitals in Lebanon.

PubMed

Al Atrouni, Ahmad; Hamze, Monzer; Jisr, Tamima; Lemarié, Carole; Eveillard, Matthieu; Joly-Guillou, Marie-Laure; Kempf, Marie

2016-11-01

To investigate the molecular epidemiology of Acinetobacter baumannii strains isolated from different hospitals in Lebanon. A total of 119 non-duplicate Acinetobacter strains were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and partial rpoB gene sequencing. Antibiotic susceptibility testing was performed by disc diffusion method and all identified carbapenem-resistant isolates were investigated by PCR assays for the presence of the carbapenemase-encoding genes. Multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) were used for molecular typing. Of the 119 A. baumannii isolates, 76.5% were resistant to carbapenems. The most common carbapenemase was the OXA-23-type, found in 82 isolates. The study of population structure using MLST revealed the presence of 30 sequence types (STs) including 18 new ones, with ST2 being the most commonly detected, accounting for 61% of the isolates typed. PFGE performed on all strains of ST2 identified a major cluster of 53 isolates, in addition to three other minor clusters and ten unique profiles. This study highlights the wide dissemination of highly related OXA-23-producing carbapenem-resistant A. baumannii belonging to the international clone II in Lebanon. Thus, appropriate infection control measures are recommended in order to control the geographical spread of this clone in this country. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

The clonal and mutational evolution spectrum of primary triple-negative breast cancers.

PubMed

Shah, Sohrab P; Roth, Andrew; Goya, Rodrigo; Oloumi, Arusha; Ha, Gavin; Zhao, Yongjun; Turashvili, Gulisa; Ding, Jiarui; Tse, Kane; Haffari, Gholamreza; Bashashati, Ali; Prentice, Leah M; Khattra, Jaswinder; Burleigh, Angela; Yap, Damian; Bernard, Virginie; McPherson, Andrew; Shumansky, Karey; Crisan, Anamaria; Giuliany, Ryan; Heravi-Moussavi, Alireza; Rosner, Jamie; Lai, Daniel; Birol, Inanc; Varhol, Richard; Tam, Angela; Dhalla, Noreen; Zeng, Thomas; Ma, Kevin; Chan, Simon K; Griffith, Malachi; Moradian, Annie; Cheng, S-W Grace; Morin, Gregg B; Watson, Peter; Gelmon, Karen; Chia, Stephen; Chin, Suet-Feung; Curtis, Christina; Rueda, Oscar M; Pharoah, Paul D; Damaraju, Sambasivarao; Mackey, John; Hoon, Kelly; Harkins, Timothy; Tadigotla, Vasisht; Sigaroudinia, Mahvash; Gascard, Philippe; Tlsty, Thea; Costello, Joseph F; Meyer, Irmtraud M; Eaves, Connie J; Wasserman, Wyeth W; Jones, Steven; Huntsman, David; Hirst, Martin; Caldas, Carlos; Marra, Marco A; Aparicio, Samuel

2012-04-04

Primary triple-negative breast cancers (TNBCs), a tumour type defined by lack of oestrogen receptor, progesterone receptor and ERBB2 gene amplification, represent approximately 16% of all breast cancers. Here we show in 104 TNBC cases that at the time of diagnosis these cancers exhibit a wide and continuous spectrum of genomic evolution, with some having only a handful of coding somatic aberrations in a few pathways, whereas others contain hundreds of coding somatic mutations. High-throughput RNA sequencing (RNA-seq) revealed that only approximately 36% of mutations are expressed. Using deep re-sequencing measurements of allelic abundance for 2,414 somatic mutations, we determine for the first time-to our knowledge-in an epithelial tumour subtype, the relative abundance of clonal frequencies among cases representative of the population. We show that TNBCs vary widely in their clonal frequencies at the time of diagnosis, with the basal subtype of TNBC showing more variation than non-basal TNBC. Although p53 (also known as TP53), PIK3CA and PTEN somatic mutations seem to be clonally dominant compared to other genes, in some tumours their clonal frequencies are incompatible with founder status. Mutations in cytoskeletal, cell shape and motility proteins occurred at lower clonal frequencies, suggesting that they occurred later during tumour progression. Taken together, our results show that understanding the biology and therapeutic responses of patients with TNBC will require the determination of individual tumour clonal genotypes.

Purification of Bone Marrow Clonal Cells from Patients with Myelodysplastic Syndrome via IGF-IR

PubMed Central

He, Qi; Chang, Chun-Kang; Xu, Feng; Zhang, Qing-Xia; Shi, Wen-Hui; Li, Xiao

2015-01-01

Malignant clonal cells purification can greatly benefit basic and clinical studies in myelodysplastic syndrome (MDS). In this study, we investigated the potential of using type 1 insulin-like growth factor receptor (IGF-IR) as a marker for purification of malignant bone marrow clonal cells from patients with MDS. The average percentage of IGF-IR expression in CD34+ bone marrow cells among 15 normal controls was 4.5%, 70% of which also express the erythroid lineage marker CD235a. This indicates that IGF-IR mainly express in erythropoiesis. The expression of IGF-IR in CD34+ cells of 55 MDS patients was significantly higher than that of cells from the normal controls (54.0 vs. 4.5%). Based on the pattern of IGF-IR expression in MDS patients and normal controls, sorting of IGF-IR-positive and removal of CD235a-positive erythroid lineage cells with combination of FISH detection were performed on MDS samples with chromosomal abnormalities. The percentage of malignant clonal cells significantly increased after sorting. The enrichment effect was more significant in clonal cells with a previous percentage lower than 50%. This enrichment effect was present in samples from patients with +8, 5q-/-5, 20q-/-20 or 7q-/-7 chromosomal abnormalities. These data suggest that IGF-IR can be used as a marker for MDS bone marrow clonal cells and using flow cytometry for positive IGF-IR sorting may effectively purify MDS clonal cells. PMID:26469401

MRSA clonal complex 22 strains harboring toxic shock syndrome toxin (TSST-1) are endemic in the primary hospital in Gaza, Palestine.

PubMed

Al Laham, Nahed; Mediavilla, José R; Chen, Liang; Abdelateef, Nahed; Elamreen, Farid Abu; Ginocchio, Christine C; Pierard, Denis; Becker, Karsten; Kreiswirth, Barry N

2015-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in both community and healthcare-related settings worldwide. Current knowledge regarding the epidemiology of S. aureus and MRSA in Gaza is based on a single community-based carriage study. Here we describe a cross-sectional analysis of 215 clinical isolates collected from Al-Shifa Hospital in Gaza during 2008 and 2012. All isolates were characterized by spa typing, SCCmec typing, and detection of genes encoding Panton-Valentine leukocidin (PVL) and toxic shock syndrome toxin (TSST-1). Representative genotypes were also subjected to multilocus sequence typing (MLST). Antibiotic susceptibility testing was performed using VITEK2 and MicroScan. MRSA represented 56.3% of all S. aureus strains, and increased in frequency from 2008 (54.8%) to 2012 (58.4%). Aside from beta-lactams, resistance was observed to tetracycline, erythromycin, clindamycin, gentamicin, and fluoroquinolones. Molecular typing identified 35 spa types representing 17 MLST clonal complexes (CC), with spa 998 (Ridom t223, CC22) and spa 70 (Ridom t044, CC80) being the most prevalent. SCCmec types I, III, IV, V and VI were identified among MRSA isolates, while type II was not detected. PVL genes (lukF/S-PV) were detected in 40.0% of all isolates, while the TSST-1 gene (tst) was detected in 27.4% of all isolates, with surprisingly high frequency within CC22 (70.4%). Both PVL and TSST-1 genes were found in several isolates from 2012. Molecular typing of clinical isolates from Gaza hospitals revealed unusually high prevalence of TSST-1 genes among CC22 MRSA, which is noteworthy given a recent community study describing widespread carriage of a CC22 MRSA clone known as the 'Gaza strain'. While the latter did not address TSST-1, tst-positive spa 998 (Ridom t223) has been detected in several neighboring countries, and described as endemic in an Italian NICU, suggesting international spread of a 'Middle Eastern variant' of pandemic CC22 strain EMRSA-15.

Radiation-induced transmissable chromosomal instability in haemopoietic stem cells

NASA Astrophysics Data System (ADS)

Kadhim, M. A.; Wright, E. G.

Heritable radiation-induced genetic alterations have long been assumed to be ``fixed'' within the first cell division. However, there is a growing body of evidence that a considerable fraction of cells surviving radiation exposure appear normal, but a variety of mutational changes arise in their progeny due to a transmissible genomic instability. In our investigations of G-banded metaphases, non-clonal cytogenetic aberrations, predominantly chromatid-type aberrations, have been observed in the clonal descendants of murine and human haemopoietic stem cells surviving low doses (~1 track per cell) of alpha-particle irradiations. The data are consistent with a transmissible genetic instability induced in a stem cell resulting in a diversity of chromosomal aberrations in its clonal progeny many cell divisions later. Recent studies have demonstrated that the instability phenotype persists in vivo and that the expression of chromosomal instability has a strong dependence on the genetic characteristics of the irradiated cell. At the time when cytogenetic aberrations are detected, an increased incidence of hprt mutations and apoptotic cells have been observed in the clonal descendants of alpha-irradiated murine haemopoietic stem cells. Thus, delayed chromosomal abnormalities, delayed cell death by apoptosis and late-arising specific gene mutations may reflect diverse consequences of radiation-induced genomic instability. The relationship, if any, between these effects is not established. Current studies suggest that expression of these delayed heritable effects is determined by the type of radiation exposure, type of cell and a variety of genetic factors.

Hybrid origin of gynogenetic clones and the introgression of their mitochondrial genome into sexual diploids through meiotic hybridogenesis in the loach, Misgurnus anguillicuadatus.

PubMed

Yamada, Aya; Kodo, Yukihiro; Murakami, Masaru; Kuroda, Masamichi; Aoki, Takao; Fujimoto, Takafumi; Arai, Katsutoshi

2015-11-01

In a few Japanese populations of the loach Misgurnus anguillicaudatus (Teleostei: Cobitidae), clonal diploid lineages produce unreduced diploid eggs that normally undergo gynogenetic reproduction; however the origin of these clones remains elusive. Here, we show the presence of two diverse clades, A and B, within this loach species from sequence analyses of two nuclear genes RAG1 (recombination activating gene 1) and IRBP2 (interphotoreceptor retinoid-binding protein, 2) and then demonstrate heterozygous genotypes fixed at the two loci as the evidence of the hybrid nature of clonal lineages. All the clonal individuals were identified by clone-specific mitochondrial DNA haplotypes, microsatellite genotypes, and random amplified polymorphic DNA fingerprints; they commonly showed two alleles, one from clade A and another from clade B, whereas other wild-type diploids possessed alleles from either clade A or B. However, we also found wild-type diploids with clone-specific mitochondrial DNA and nuclear genes from clade B. One possible explanation is an introgression of a clone-specific mitochondrial genome from clonal to these wild-type loaches. These individuals likely arose by a cross between haploid sperm from bisexual B clade males and haploid eggs with clone-specific mtDNA and clade B nuclear genome, produced by meiotic hybridogenesis (elimination of unmatched A genome followed by meiosis after preferential pairing between two matched B genomes) in clone-origin triploid individual (ABB). © 2015 Wiley Periodicals, Inc.

Atypical Strain of Toxoplasma gondii Causing Fatal Reactivation after Hematopoietic Stem Cell Transplantion in a Patient with an Underlying Immunological Deficiency

PubMed Central

Štajner, Tijana; Vasiljević, Zorica; Vujić, Dragana; Marković, Marija; Ristić, Goran; Mićić, Dragan; Pašić, Srdjan; Ivović, Vladimir; Ajzenberg, Daniel

2013-01-01

In immunocompromized patients, including hematopoietic stem cell transplant (HSCT) recipients, life-threatening toxoplasmosis may result from reactivation of previous infection. We report a case of severe disseminated toxoplasmosis that developed early after allogeneic HSCT for T-cell lymphoblastic leukemia/lymphoma in a 15-year-old Toxoplasma gondii-seropositive boy with Nijmegen breakage syndrome, a rare genetic DNA repair disorder associated with immunodeficiency. The donor was the patient's HLA-identical brother. Prophylaxis with cotrimoxazole was discontinued a day before the HSCT procedure. Signs of lung infection appeared as early as day 14 post-HSCT. The presence of tachyzoite-like structures on Giemsa-stained bronchoalveolar lavage (BAL) fluid smears suggested toxoplasmosis. Real-time PCR targeted at the T. gondii AF146527 gene revealed extremely high parasite burdens in both blood and BAL fluid. Although immediate introduction of specific treatment resulted in a marked reduction of the parasite load and transient clinical improvement, the patient deteriorated and died of multiple organ failure on day 39 post-HSCT. Direct genotyping of T. gondii DNA from blood and BAL fluid with the PCR-restriction fragment length polymorphism method revealed type II alleles with SAG1, SAG2, and GRA6 markers but alleles of both type I and type II with GRA7. Additional analysis with 15 microsatellite markers showed that the T. gondii DNA was atypical and genetically divergent from that of the clonal type I, II, and III strains. This is the first report of increased clinical severity of toxoplasmosis associated with an atypical strain in the setting of immunosuppression, which emphasizes the need to diagnose and monitor toxoplasmosis by quantitative molecular methods in cases of reactivation risk. PMID:23761151

Prevalence and clonal analysis of Porphyromonas gingivalis in primary endodontic infections.

PubMed

Siqueira, José F; Rôças, Isabela N; Silva, Marlei G

2008-11-01

This study investigated the prevalence of Porphyromonas gingivalis in 62 teeth with primary endodontic infections by using a species-specific 16S rRNA gene-based nested polymerase chain reaction assay. P. gingivalis isolates recovered from 2 infected root canals were also analyzed for clonal diversity by using arbitrarily primed PCR. Overall, P. gingivalis was found in 48% of the samples. This species was specifically detected in 36% of canals of teeth with chronic apical periodontitis, in 46% of the cases of acute apical periodontitis, and in 67% of acute apical abscesses. P. gingivalis was significantly more frequent in abscess aspirates than in canals of teeth with chronic apical periodontitis (P < .05). Typing of colonies retrieved from 2 infected canals revealed 2 clones per individual. These findings confirmed that P. gingivalis can be an important endodontic pathogen, mostly associated with acute abscesses, and demonstrated that different clonal types of this species can colonize the root canal in the same individual.

Molecular characterization of Toxoplasma gondii in pork meat from different production systems in the Czech Republic.

PubMed

Slany, Michal; Reslova, Nikol; Babak, Vladimir; Lorencova, Alena

2016-12-05

Toxoplasmosis is a major public health issue, due to the presence of Toxoplasma gondii, mainly in pork. The aim of this study was to determine the occurrence of T. gondii in pigs and wild boars bred in different production systems in the Czech Republic using ELISA and qPCR methods. Our results show that T. gondii infection is widespread in pigs and wild boars bred and slaughtered in the Czech Republic and that there is a higher exposure to T. gondii in backyard slaughter operations and organic pig farming, indicating a potential risk for meat consumption. Additionally, genotyping of amplified loci for Type II suggests the presence of one clonal genotype circulating in these animals. Copyright © 2016 Elsevier B.V. All rights reserved.

Streptococcus mutans clonal variation revealed by multilocus sequence typing.

PubMed

Nakano, Kazuhiko; Lapirattanakul, Jinthana; Nomura, Ryota; Nemoto, Hirotoshi; Alaluusua, Satu; Grönroos, Lisa; Vaara, Martti; Hamada, Shigeyuki; Ooshima, Takashi; Nakagawa, Ichiro

2007-08-01

Streptococcus mutans is the major pathogen of dental caries, a biofilm-dependent infectious disease, and occasionally causes infective endocarditis. S. mutans strains have been classified into four serotypes (c, e, f, and k). However, little is known about the S. mutans population, including the clonal relationships among strains of S. mutans, in relation to the particular clones that cause systemic diseases. To address this issue, we have developed a multilocus sequence typing (MLST) scheme for S. mutans. Eight housekeeping gene fragments were sequenced from each of 102 S. mutans isolates collected from the four serotypes in Japan and Finland. Between 14 and 23 alleles per locus were identified, allowing us theoretically to distinguish more than 1.2 x 10(10) sequence types. We identified 92 sequence types in these 102 isolates, indicating that S. mutans contains a diverse population. Whereas serotype c strains were widely distributed in the dendrogram, serotype e, f, and k strains were differentiated into clonal complexes. Therefore, we conclude that the ancestral strain of S. mutans was serotype c. No geographic specificity was identified. However, the distribution of the collagen-binding protein gene (cnm) and direct evidence of mother-to-child transmission were clearly evident. In conclusion, the superior discriminatory capacity of this MLST scheme for S. mutans may have important practical implications.

CRISPR-cas loci profiling of Cronobacter sakazakii pathovars.

PubMed

Ogrodzki, Pauline; Forsythe, Stephen James

2016-12-01

Cronobacter sakazakii sequence types 1, 4, 8 and 12 are associated with outbreaks of neonatal meningitis and necrotizing enterocolitis infections. However clonality results in strains which are indistinguishable using conventional methods. This study investigated the use of clustered regularly interspaced short palindromic repeats (CRISPR)-cas loci profiling for epidemiological investigations. Seventy whole genomes of C. sakazakii strains from four clonal complexes which were widely distributed temporally, geographically and origin of source were profiled. All strains encoded the same type I-E subtype CRISPR-cas system with a total of 12 different CRISPR spacer arrays. This study demonstrated the greater discriminatory power of CRISPR spacer array profiling compared with multilocus sequence typing, which will be of use in source attribution during Cronobacter outbreak investigations.

A Livestock-Associated, Multidrug-Resistant, Methicillin-Resistant Staphylococcus aureus Clonal Complex 97 Lineage Spreading in Dairy Cattle and Pigs in Italy

PubMed Central

Feltrin, Fabiola; Alba, Patricia; Kraushaar, Britta; Ianzano, Angela; Argudín, María Angeles; Di Matteo, Paola; Porrero, María Concepción; Aarestrup, Frank M.; Butaye, Patrick; Franco, Alessia

2015-01-01

Pandemic methicillin-resistant Staphylococcus aureus (MRSA) clonal complex 97 (CC97) lineages originated from livestock-to-human host jumps. In recent years, CC97 has become one of the major MRSA lineages detected in Italian farmed animals. The aim of this study was to characterize and analyze differences in MRSA and methicillin-susceptible S. aureus (MSSA) mainly of swine and bovine origins. Forty-seven CC97 isolates, 35 MRSA isolates, and 6 MSSA isolates from different Italian pig and cattle holdings; 5 pig MRSA isolates from Germany; and 1 human MSSA isolate from Spain were characterized by macrorestriction pulsed-field gel electrophoresis (PFGE) analysis, multilocus sequence typing (MLST), spa typing, staphylococcal cassette chromosome mec (SCCmec) typing, and antimicrobial resistance pattern analysis. Virulence and resistance genes were investigated by PCR and microarray analysis. Most of the isolates were of SCCmec type V (SCCmec V), except for two German MRSA isolates (SCCmec III). Five main clusters were identified by PFGE, with the German isolates (clusters I and II) showing 60.5% similarity with the Italian isolates, most of which (68.1%) grouped into cluster V. All CC97 isolates were Panton-Valentine leukocidin (PVL) negative, and a few (n = 7) tested positive for sak or scn. All MRSA isolates were multidrug resistant (MDR), and the main features were erm(B)- or erm(C)-mediated (n = 18) macrolide-lincosamide-streptogramin B resistance, vga(A)-mediated (n = 37) pleuromutilin resistance, fluoroquinolone resistance (n = 33), tet(K) in 32/37 tet(M)-positive isolates, and blaZ in almost all MRSA isolates. Few host-associated differences were detected among CC97 MRSA isolates: their extensive MDR nature in both pigs and dairy cattle may be a consequence of a spillback from pigs of a MRSA lineage that originated in cattle as MSSA and needs further investigation. Measures should be implemented at the farm level to prevent spillover to humans in intensive farming areas. PMID:26590279

The Opportunistic Pathogen Propionibacterium acnes: Insights into Typing, Human Disease, Clonal Diversification and CAMP Factor Evolution

PubMed Central

McDowell, Andrew; Nagy, István; Magyari, Márta; Barnard, Emma; Patrick, Sheila

2013-01-01

We previously described a Multilocus Sequence Typing (MLST) scheme based on eight genes that facilitates population genetic and evolutionary analysis of P. acnes. While MLST is a portable method for unambiguous typing of bacteria, it is expensive and labour intensive. Against this background, we now describe a refined version of this scheme based on two housekeeping (aroE; guaA) and two putative virulence (tly; camp2) genes (MLST4) that correctly predicted the phylogroup (IA1, IA2, IB, IC, II, III), clonal complex (CC) and sequence type (ST) (novel or described) status for 91% isolates (n = 372) via cross-referencing of the four gene allelic profiles to the full eight gene versions available in the MLST database (http://pubmlst.org/pacnes/). Even in the small number of cases where specific STs were not completely resolved, the MLST4 method still correctly determined phylogroup and CC membership. Examination of nucleotide changes within all the MLST loci provides evidence that point mutations generate new alleles approximately 1.5 times as frequently as recombination; although the latter still plays an important role in the bacterium's evolution. The secreted/cell-associated ‘virulence’ factors tly and camp2 show no clear evidence of episodic or pervasive positive selection and have diversified at a rate similar to housekeeping loci. The co-evolution of these genes with the core genome might also indicate a role in commensal/normal existence constraining their diversity and preventing their loss from the P. acnes population. The possibility that members of the expanded CAMP factor protein family, including camp2, may have been lost from other propionibacteria, but not P. acnes, would further argue for a possible role in niche/host adaption leading to their retention within the genome. These evolutionary insights may prove important for discussions surrounding camp2 as an immunotherapy target for acne, and the effect such treatments may have on commensal lineages. PMID:24058439

Clonal diversity of extended-spectrum beta-lactamase producing Escherichia coli isolates in fecal samples of wild animals.

PubMed

Cristóvão, Filipe; Alonso, Carla Andrea; Igrejas, Gilberto; Sousa, Margarida; Silva, Vanessa; Pereira, José Eduardo; Lozano, Carmen; Cortés-Cortés, Gerardo; Torres, Carmen; Poeta, Patrícia

2017-03-01

The clonal diversity of extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli isolates from nine different species of wild animals from distinct regions of Portugal and Spain and their content in replicon plasmids were analyzed. Among the initial 53 ESBL-producing E. coli isolates that were studied (from previous studies), 28 were selected, corresponding to different animal origins with distinct ESBL types and pulsed-field gel electrophoresis (PFGE) patterns. These 28 isolates produced different ESBLs ascribed to the following families: CTX-M, SHV and TEM. The isolates were classified into three phylogenetic groups: B1 (n = 11), A (n = 10) and D (n = 7). The seven E. coli of phylogroup D were then typed by multilocus sequence typing and ascribed to four distinct sequence types: ST117, ST115, ST2001 and ST69. The clonal diversity and relationship between isolates was studied by PFGE. Lastly, the plasmids were analyzed according to their incompatibility group using the PCR-based-replicon-typing scheme. A great diversity of replicon types was identified, with up to five per isolate. Most of the CTX-M-1 and SHV-12 producing E. coli isolates carried IncI1 or IncN replicons. The diversity of ESBL-producing E. coli isolates in wild animals, which can be disseminated in the environment, emphasizes the environmental and health problems that we face nowadays. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Listeria monocytogenes source distribution analysis indicates regional heterogeneity and ecological niche preference among serotype 4b clones

USDA-ARS?s Scientific Manuscript database

Human illness due to the foodborne bacterial pathogen Listeria monocytogenes frequently involves certain widely disseminated clonal complexes (CCs), primarily of serotype 4b. CC1, CC2 and CC6, previously also designated epidemic clone (EC) I, Ia and II, respectively, have been frequently implicate...

Orthogonal typing methods identify genetic diversity among Belgian Campylobacter jejuni strains isolated over a decade from poultry and cases of sporadic human illness.

PubMed

Elhadidy, Mohamed; Arguello, Hector; Álvarez-Ordóñez, Avelino; Miller, William G; Duarte, Alexandra; Martiny, Delphine; Hallin, Marie; Vandenberg, Olivier; Dierick, Katelijne; Botteldoorn, Nadine

2018-06-20

Campylobacter jejuni is a zoonotic pathogen commonly associated with human gastroenteritis. Retail poultry meat is a major food-related transmission source of C. jejuni to humans. The present study investigated the genetic diversity, clonal relationship, and strain risk-analysis of 403 representative C. jejuni isolates from chicken broilers (n = 204) and sporadic cases of human diarrhea (n = 199) over a decade (2006-2015) in Belgium, using multilocus sequence typing (MLST), PCR binary typing (P-BIT), and identification of lipooligosaccharide (LOS) biosynthesis locus classes. A total of 123 distinct sequence types (STs), clustered in 28 clonal complexes (CCs) were assigned, including ten novel sequence types that were not previously documented in the international database. Sequence types ST-48, ST-21, ST-50, ST-45, ST-464, ST-2274, ST-572, ST-19, ST-257 and ST-42 were the most prevalent. Clonal complex 21 was the main clonal complex in isolates from humans and chickens. Among observed STs, a total of 35 STs that represent 72.2% (291/403) of the isolates were identified in both chicken and human isolates confirming considerable epidemiological relatedness; these 35 STs also clustered together in the most prevalent CCs. A majority of the isolates harbored sialylated LOS loci associated with potential neuropathic outcomes in humans. Although the concordance between MLST and P-BIT, determined by the adjusted Rand and Wallace coefficients, showed low congruence between both typing methods. The discriminatory power of P-BIT and MLST was similar, with Simpson's diversity indexes of 0.978 and 0.975, respectively. Furthermore, P-BIT could provide additional epidemiological information that would provide further insights regarding the potential association to human health from each strain. In addition, certain clones could be linked to specific clinical symptoms. Indeed, LOS class E was associated with less severe infections. Moreover, ST-572 was significantly associated with clinical infections occurring after travelling abroad. Ultimately, the data generated from this study will help to better understand the molecular epidemiology of C. jejuni infection. Copyright © 2018. Published by Elsevier B.V.

Surface antigens from Escherichia coli O2 and O78 strains of avian origin.

PubMed Central

Dho-Moulin, M; van den Bosch, J F; Girardeau, J P; Brée, A; Barat, T; Lafont, J P

1990-01-01

Fimbriae from O2 and O78 virulent strains of avian Escherichia coli were compared with type 1A fimbriae with regard to the apparent molecular weights of their subunits and their antigenic relationships. Under static broth culture conditions, most O78 strains expressed fimbriae closely related to those of type 1A. Under the same culture conditions, another type of fimbriae, sharing some common properties with type 1A fimbriae, was observed only on O2 strains; however, these fimbriae differed from type 1A fimbriae in the apparent molecular weights of their subunits and in the expression of specific epitopes. They were called type 1-like fimbriae. Homologies in lipopolysaccharide and outer membrane protein profiles were also demonstrated among the strains expressing type 1-like fimbriae, which suggests the existence of a clonal relationship among O2:K1 avian E. coli strains. The O78 strains studied did not appear to be clonally related. Images PMID:1968434

Genetic Diversity and Population Structure of Vibrio cholerae

PubMed Central

Beltrán, Pilar; Delgado, Gabriela; Navarro, Armando; Trujillo, Francisca; Selander, Robert K.; Cravioto, Alejandro

1999-01-01

Multilocus enzyme electrophoresis (MLEE) of 397 Vibrio cholerae isolates, including 143 serogroup reference strains and 244 strains from Mexico and Guatemala, identified 279 electrophoretic types (ETs) distributed in two major divisions (I and II). Linkage disequilibrium was demonstrated in both divisions and in subdivision Ic of division I but not in subdivision Ia, which includes 76% of the ETs. Despite this evidence of relatively frequent recombination, clonal lineages may persist for periods of time measured in at least decades. In addition to the pandemic clones of serogroups O1 and O139, which form a tight cluster of four ETs in subdivision Ia, MLEE analysis identified numerous apparent clonal lineages of non-O1 strains with intercontinental distributions. A clone of serogroup O37 that demonstrated epidemic potential in the 1960s is closely related to the pandemic O1/O139 clones, but the nontoxigenic O1 Inaba El Tor reference strain is not. A strain of serogroup O22, which has been identified as the most likely donor of exogenous rfb region DNA to the O1 progenitor of the O139 clone, is distantly related to the O1/O139 clones. The close evolutionary relationships of the O1, O139, and O37 epidemic clones indicates that new cholera clones are likely to arise by the modification of a lineage that is already epidemic or is closely related to such a clone. PMID:9986816

Genetic diversity and population structure of Vibrio cholerae.

PubMed

Beltrán, P; Delgado, G; Navarro, A; Trujillo, F; Selander, R K; Cravioto, A

1999-03-01

Multilocus enzyme electrophoresis (MLEE) of 397 Vibrio cholerae isolates, including 143 serogroup reference strains and 244 strains from Mexico and Guatemala, identified 279 electrophoretic types (ETs) distributed in two major divisions (I and II). Linkage disequilibrium was demonstrated in both divisions and in subdivision Ic of division I but not in subdivision Ia, which includes 76% of the ETs. Despite this evidence of relatively frequent recombination, clonal lineages may persist for periods of time measured in at least decades. In addition to the pandemic clones of serogroups O1 and O139, which form a tight cluster of four ETs in subdivision Ia, MLEE analysis identified numerous apparent clonal lineages of non-O1 strains with intercontinental distributions. A clone of serogroup O37 that demonstrated epidemic potential in the 1960s is closely related to the pandemic O1/O139 clones, but the nontoxigenic O1 Inaba El Tor reference strain is not. A strain of serogroup O22, which has been identified as the most likely donor of exogenous rfb region DNA to the O1 progenitor of the O139 clone, is distantly related to the O1/O139 clones. The close evolutionary relationships of the O1, O139, and O37 epidemic clones indicates that new cholera clones are likely to arise by the modification of a lineage that is already epidemic or is closely related to such a clone.

Genotypic and Phenotypic Characterization of Carriage and Invasive Disease Isolates of Neisseria meningitidis in Finland

PubMed Central

Saukkoriipi, Annika; Bratcher, Holly B.; Bloigu, Aini; Juvonen, Raija; Silvennoinen-Kassinen, Sylvi; Peitso, Ari; Harju, Terttu; Vainio, Olli; Kuusi, Markku; Maiden, Martin C. J.; Leinonen, Maija; Käyhty, Helena; Toropainen, Maija

2012-01-01

The relationship between carriage and the development of invasive meningococcal disease is not fully understood. We investigated the changes in meningococcal carriage in 892 military recruits in Finland during a nonepidemic period (July 2004 to January 2006) and characterized all of the oropharyngeal meningococcal isolates obtained (n = 215) by using phenotypic (serogrouping and serotyping) and genotypic (porA typing and multilocus sequence typing) methods. For comparison, 84 invasive meningococcal disease strains isolated in Finland between January 2004 and February 2006 were also analyzed. The rate of meningococcal carriage was significantly higher at the end of military service than on arrival (18% versus 2.2%; P < 0.001). Seventy-four percent of serogroupable carriage isolates belonged to serogroup B, and 24% belonged to serogroup Y. Most carriage isolates belonged to the carriage-associated ST-60 clonal complex. However, 21.5% belonged to the hyperinvasive ST-41/44 clonal complex. Isolates belonging to the ST-23 clonal complex were cultured more often from oropharyngeal samples taken during the acute phase of respiratory infection than from samples taken at health examinations at the beginning and end of military service (odds ratio [OR], 6.7; 95% confidence interval [95% CI], 2.7 to 16.4). The ST-32 clonal complex was associated with meningococcal disease (OR, 17.8; 95% CI, 3.8 to 81.2), while the ST-60 clonal complex was associated with carriage (OR, 10.7; 95% CI, 3.3 to 35.2). These findings point to the importance of meningococcal vaccination for military recruits and also to the need for an efficacious vaccine against serogroup B isolates. PMID:22135261

Evidence of substantial recombination among Trypanosoma cruzi II strains from Minas Gerais.

PubMed

Baptista, Rodrigo de Paula; D'Ávila, Daniella Alchaar; Segatto, Marcela; do Valle, Ítalo Faria; Franco, Glória Regina; Valadares, Helder Magno Silva; Gontijo, Eliane Dias; Galvão, Lúcia Maria da Cunha; Pena, Sérgio Danilo Junho; Chiari, Egler; Machado, Carlos Renato; Macedo, Andréa Mara

2014-03-01

Due to the scarcity of evidence of sexuality in Trypanosoma cruzi, the causative agent of Chagas disease, it has been general accepted that the parasite reproduction is essentially clonal with infrequent genetic recombination. This assumption is mainly supported by indirect evidence, such as Hardy-Weinberg imbalances, linkage disequilibrium and a strong correlation between independent sets of genetic markers of T. cruzi populations. However, because the analyzed populations are usually isolated from different geographic regions, the possibility of population substructuring as generating these genetic marker imbalances cannot be eliminated. To investigate this possibility, we firstly compared the allele frequencies and haplotype networks using seven different polymorphic loci (two from mitochondrial and five from different nuclear chromosomes) in two groups of TcII strains: one including isolates obtained from different regions in Latin America and the other including isolates obtained only from patients of the Minas Gerais State in Brazil. Our hypothesis was that if the population structure is essentially clonal, Hardy-Weinberg disequilibrium and a sharp association between the clusters generated by analyzing independent markers should be observed in both strain groups, independent of the geographic origin of the samples. The results demonstrated that the number of microsatellite loci in linkage disequilibrium decreased from 4 to 1 when only strains from Minas Gerais were analyzed. Moreover, we did not observed any correlation between the clusters when analyzing the nuclear and mitochondrial loci, suggesting independent inheritance of these markers among the Minas Gerais strains. Besides, using a second subset of five physically linked microsatellite loci and the Minas Gerais strains, we could also demonstrate evidence of homologous recombination roughly proportional to the relative distance among them. Taken together, our results do not support a clonal population structure for T. cruzi, particularly in TcII, which coexists in the same geographical area, suggesting that genetic exchanges among these strains may occur more frequently than initially expected. Copyright © 2013 Elsevier B.V. All rights reserved.

Molecular and phenotypic characteristics of methicillin-resistant Staphylococcus aureus isolated from hospitalized patients.

PubMed

de Oliveira, Caio Ferreira; Morey, Alexandre Tadachi; Santos, Jussevania Pereira; Gomes, Ludmila Vilela Pereira; Cardoso, Juscélio Donizete; Pinge-Filho, Phileno; Perugini, Márcia Regina Eches; Yamauchi, Lucy Megumi; Yamada-Ogatta, Sueli Fumie

2015-07-30

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of infections acquired in both community and hospital settings. In this study, MRSA isolated from different sources of hospitalized patients was characterized by molecular and phenotypic methods. A total of 123 S. aureus isolates were characterized according to their genetic relatedness by repetitive element sequence based-PCR (REP-PCR), in vitro antimicrobial susceptibility profile, SCCmec typing and presence of seven virulence factor-encoding genes. REP-PCR fingerprinting showed low relatedness between the isolates, and the predominance of one specific lineage or clonal group was not observed. All isolates were susceptible to teicoplanin and linezolide. All isolates were resistant to cefoxitin and penicillin, and the majority were also resistant to one or more other antimicrobials. Fifty isolates (41.7%) were intermediately resistant to vancomycin. Most isolates harbored SCCmec type II (53.7%), followed by type I (22.8%), type IV (8.1%) and type III (1.6%). All isolates harbored at least two virulence factor-encoding genes, and the prevalence was as follows: coa, 100%; icaA, 100%; hla, 13.0%; hlb, 91.1%, hld, 91.1%; lukS-PV and lukF-PV, 2.4%; and tst, 34.1%. A positive association with the presence of hla and SCCmec type II, and tst and SCCmec type I was observed. This study showed the high virulence potential of multidrug-resistant MRSA circulating in a teaching hospital. A high prevalence of MRSA showing intermediate vancomycin resistance was also observed, indicating the urgent need to improve strategies for controlling the use of antimicrobials for appropriate management of S. aureus infections.

Murine glomerulotropic monoclonal antibodies are highly oligoclonal and exhibit distinctive molecular features.

PubMed

Lefkowith, J B; Di Valerio, R; Norris, J; Glick, G D; Alexander, A L; Jackson, L; Gilkeson, G S

1996-08-01

We recently produced a panel of seven glomerular-binding mAbs from a nephritic MRL-lpr mouse that bind to histones/nucleosomes (group I) or DNA (group II) adherent to glomerular basement membrane. To elucidate the molecular basis of their binding and ontogeny, we sequenced their variable (V) regions, analyzed the apparent somatic mutations, and predicted their three-dimensional structures. There were two clonally related sets (3 of 4 in group I, 3 of 3 in group II) both of the VHJ1558 family, and one mAb of the VH 7183 family. V region somatic mutations within clonally related sets had little effect on glomerular binding and did not appear to be selected for based on glomerular binding. The VH regions were most homologous with those from autoantibodies to histones, DNA, or IgG (i.e., rheumatoid factors), the Vkappa regions, with those from autoantibodies to small nuclear ribonucleoproteins (snRNP). The VH regions also exhibited an unusual VD junction (in the group I clonally related set) and an overall high content of charged amino acids (arginine, aspartic acid) in complementarity-determining regions (CDRs), particularly in CDR3. Molecular modeling studies suggested that the Fv regions of these mAbs converge to form a flat, open surface with a net positive charge. The CDR arginines in group I mAbs; appear to be located in Ag contact regions of the binding cleft. In sum, these data suggest that glomerulotropic mAbs are a highly restricted set of Abs with distinctive molecular features that may mediate their binding to glomeruli.

Population structure of Lactobacillus helveticus isolates from naturally fermented dairy products based on multilocus sequence typing.

PubMed

Sun, Zhihong; Liu, Wenjun; Song, Yuqin; Xu, Haiyan; Yu, Jie; Bilige, Menghe; Zhang, Heping; Chen, Yongfu

2015-05-01

Lactobacillus helveticus is an economically important lactic acid bacterium used in industrial dairy fermentation. In the present study, the population structure of 245 isolates of L. helveticus from different naturally fermented dairy products in China and Mongolia were investigated using an multilocus sequence typing scheme with 11 housekeeping genes. A total of 108 sequence types were detected, which formed 8 clonal complexes and 27 singletons. Results from Structure, SplitsTree, and ClonalFrame software analyses demonstrated the presence of 3 subpopulations in the L. helveticus isolates used in our study, namely koumiss, kurut-tarag, and panmictic lineages. Most L. helveticus isolates from particular ecological origins had specific population structures. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Superclone Expansion, Long-Distance Clonal Dispersal and Local Genetic Structuring in the Coral Pocillopora damicornis Type β in Reunion Island, South Western Indian Ocean.

PubMed

Gélin, Pauline; Fauvelot, Cécile; Mehn, Vincent; Bureau, Sophie; Rouzé, Héloïse; Magalon, Hélène

2017-01-01

The scleractinian coral Pocillopora damicornis type β is known to present a mixed reproduction mode: through sexual reproduction, new genotypes are created, while asexual reproduction insures their propagation. In order to investigate the relative proportion of each reproduction mode in P. damicornis type β populations from Reunion Island, Indian Ocean, clonal propagation along the west coast was assessed through four sampling sites with increasing geographical distance between sites. Coral colonies were sampled either exhaustively, randomly or haphazardly within each site, and genotypic diversity was assessed using 13 microsatellite loci over a total of 510 P. damicornis type β determined a posteriori from their mtDNA haplotype (a 840 bp sequenced fragment of the Open Reading Frame). Overall, 47% of all the sampled colonies presented the same multi-locus genotype (MLG), a superclone, suggesting that asexual propagation is extremely important in Reunion Island. Within each site, numerous MLGs were shared by several colonies, suggesting local clonal propagation through fragmentation. Moreover, some of these MLGs were found to be shared among several sites located 40 km apart. While asexual reproduction by fragmentation seems unlikely over long distances, our results suggest a production of parthenogenetic larvae. Despite shared MLGs, two differentiated clusters were enclosed among populations of the west coast of Reunion Island, revealing the necessity to set up appropriate managing strategies at a local scale.

Predominance of Three Closely Related Methicillin-Resistant Staphylococcus aureus Clones Carrying a Unique ccrC-Positive SCCmec type III and the Emergence of spa t304 and t690 SCCmec type IV pvl+ MRSA Isolates in Kinta Valley, Malaysia.

PubMed

Ho, Wai-Yew; Choo, Quok-Cheong; Chew, Choy-Hoong

2017-03-01

We investigated the epidemiology and clonality of 175 nonrepetitive methicillin-resistant Staphylococcus aureus (MRSA) isolates from clinical specimens collected between 2011 and 2012 in Kinta Valley in Malaysia. Molecular tools such as polymerase chain reaction, pulsed-field gel electrophoresis, and staphylococcal protein A (spa) typing were used. Our study revealed the predominance of three closely related ermA + SCCmec type III pulsotypes belonging to spa type t037 (Brazilian-Hungarian clone), which were deficient in the locus F, but positive for the ccrC gene in majority (65.7%) of the MRSA infections in this region. The first evidence of SCCmec type II MRSA in the country, belonging to spa type t2460, was also noted. Although the carriage of pvl gene was uncommon (8.6%) and mostly confined to either SCCmec type IV or SCCmec type V isolates, most of these isolates belonged to spa types t345 or t657, which are associated with the Bengal-Bay CA-MRSA clone. Interestingly, spa t304 and t690 SCCmec type IV pvl + were also detected among the MRSA isolates. Data from this study show the rise of uncommon clones among MRSA isolates in Malaysia.

Temporal Changes in BEXSERO® Antigen Sequence Type Associated with Genetic Lineages of Neisseria meningitidis over a 15-Year Period in Western Australia

PubMed Central

Mowlaboccus, Shakeel; Perkins, Timothy T.; Smith, Helen; Sloots, Theo; Tozer, Sarah; Prempeh, Lydia-Jessica; Tay, Chin Yen; Peters, Fanny; Speers, David; Keil, Anthony D.; Kahler, Charlene M.

2016-01-01

Neisseria meningitidis is the causative agent of invasive meningococcal disease (IMD). The BEXSERO® vaccine which is used to prevent serogroup B disease is composed of four sub-capsular protein antigens supplemented with an outer membrane vesicle. Since the sub-capsular protein antigens are variably expressed and antigenically variable amongst meningococcal isolates, vaccine coverage can be estimated by the meningococcal antigen typing system (MATS) which measures the propensity of the strain to be killed by vaccinated sera. Whole genome sequencing (WGS) which identifies the alleles of the antigens that may be recognised by the antibody response could represent, in future, an alternative estimate of coverage. In this study, WGS of 278 meningococcal isolates responsible for 62% of IMD in Western Australia from 2000–2014 were analysed for association of genetic lineage (sequence type [ST], clonal complex [cc]) with BEXSERO® antigen sequence type (BAST) and MATS to predict the annual vaccine coverage. A hyper-endemic period of IMD between 2000–05 was caused by cc41/44 with the major sequence type of ST-146 which was not predicted by MATS or BAST to be covered by the vaccine. An increase in serogroup diversity was observed between 2010–14 with the emergence of cc11 serogroup W in the adolescent population and cc23 serogroup Y in the elderly. BASTs were statistically associated with clonal complex although individual antigens underwent antigenic drift from the major type. BAST and MATS predicted an annual range of 44–91% vaccine coverage. Periods of low vaccine coverage in years post-2005 were not a result of the resurgence of cc41/44:ST-146 but were characterised by increased diversity of clonal complexes expressing BASTs which were not predicted by MATS to be covered by the vaccine. The driving force behind the diversity of the clonal complex and BAST during these periods of low vaccine coverage is unknown, but could be due to immune selection and inter-strain competition with carriage of non-disease causing meningococci. PMID:27355628

A Prospective Cohort Multicenter Study of Molecular Epidemiology and Phylogenomics of Staphylococcus aureus Bacteremia in Nine Latin American Countries

PubMed Central

Reyes, Jinnethe; Carvajal, Lina Paola; Rincon, Sandra; Diaz, Lorena; Panesso, Diana; Ibarra, Gabriel; Rios, Rafael; Munita, Jose M.; Salles, Mauro J.; Alvarez-Moreno, Carlos; Labarca, Jaime; Garcia, Coralith; Luna, Carlos M.; Mejia-Villatoro, Carlos; Zurita, Jeannete; Guzman-Blanco, Manuel; Rodriguez-Noriega, Eduardo; Narechania, Apurva; Rojas, Laura J.; Planet, Paul J.; Weinstock, George M.; Gotuzzo, Eduardo; Seas, Carlos

2017-01-01

ABSTRACT Staphylococcus aureus is an important pathogen causing a spectrum of diseases ranging from mild skin and soft tissue infections to life-threatening conditions. Bloodstream infections are particularly important, and the treatment approach is complicated by the presence of methicillin-resistant S. aureus (MRSA) isolates. The emergence of new genetic lineages of MRSA has occurred in Latin America (LA) with the rise and dissemination of the community-associated USA300 Latin American variant (USA300-LV). Here, we prospectively characterized bloodstream MRSA recovered from selected hospitals in 9 Latin American countries. All isolates were typed by pulsed-field gel electrophoresis (PFGE) and subjected to antibiotic susceptibility testing. Whole-genome sequencing was performed on 96 MRSA representatives. MRSA represented 45% of all (1,185 S. aureus) isolates. The majority of MRSA isolates belonged to clonal cluster (CC) 5. In Colombia and Ecuador, most isolates (≥72%) belonged to the USA300-LV lineage (CC8). Phylogenetic reconstructions indicated that MRSA isolates from participating hospitals belonged to three major clades. Clade A grouped isolates with sequence type 5 (ST5), ST105, and ST1011 (mostly staphylococcal chromosomal cassette mec [SCCmec] I and II). Clade B included ST8, ST88, ST97, and ST72 strains (SCCmec IV, subtypes a, b, and c/E), and clade C grouped mostly Argentinian MRSA belonging to ST30. In summary, CC5 MRSA was prevalent in bloodstream infections in LA with the exception of Colombia and Ecuador, where USA300-LV is now the dominant lineage. Clonal replacement appears to be a common phenomenon, and continuous surveillance is crucial to identify changes in the molecular epidemiology of MRSA. PMID:28760895

A Prospective Cohort Multicenter Study of Molecular Epidemiology and Phylogenomics of Staphylococcus aureus Bacteremia in Nine Latin American Countries.

PubMed

Arias, Cesar A; Reyes, Jinnethe; Carvajal, Lina Paola; Rincon, Sandra; Diaz, Lorena; Panesso, Diana; Ibarra, Gabriel; Rios, Rafael; Munita, Jose M; Salles, Mauro J; Alvarez-Moreno, Carlos; Labarca, Jaime; Garcia, Coralith; Luna, Carlos M; Mejia-Villatoro, Carlos; Zurita, Jeannete; Guzman-Blanco, Manuel; Rodriguez-Noriega, Eduardo; Narechania, Apurva; Rojas, Laura J; Planet, Paul J; Weinstock, George M; Gotuzzo, Eduardo; Seas, Carlos

2017-10-01

Staphylococcus aureus is an important pathogen causing a spectrum of diseases ranging from mild skin and soft tissue infections to life-threatening conditions. Bloodstream infections are particularly important, and the treatment approach is complicated by the presence of methicillin-resistant S. aureus (MRSA) isolates. The emergence of new genetic lineages of MRSA has occurred in Latin America (LA) with the rise and dissemination of the community-associated USA300 Latin American variant (USA300-LV). Here, we prospectively characterized bloodstream MRSA recovered from selected hospitals in 9 Latin American countries. All isolates were typed by pulsed-field gel electrophoresis (PFGE) and subjected to antibiotic susceptibility testing. Whole-genome sequencing was performed on 96 MRSA representatives. MRSA represented 45% of all (1,185 S. aureus ) isolates. The majority of MRSA isolates belonged to clonal cluster (CC) 5. In Colombia and Ecuador, most isolates (≥72%) belonged to the USA300-LV lineage (CC8). Phylogenetic reconstructions indicated that MRSA isolates from participating hospitals belonged to three major clades. Clade A grouped isolates with sequence type 5 (ST5), ST105, and ST1011 (mostly staphylococcal chromosomal cassette mec [SCC mec ] I and II). Clade B included ST8, ST88, ST97, and ST72 strains (SCC mec IV, subtypes a, b, and c/E), and clade C grouped mostly Argentinian MRSA belonging to ST30. In summary, CC5 MRSA was prevalent in bloodstream infections in LA with the exception of Colombia and Ecuador, where USA300-LV is now the dominant lineage. Clonal replacement appears to be a common phenomenon, and continuous surveillance is crucial to identify changes in the molecular epidemiology of MRSA. Copyright © 2017 American Society for Microbiology.

RAPD- and ERIC-Based Typing of Clinical and Environmental Pseudomonas aeruginosa Isolates.

PubMed

Auda, Ibtesam Ghadban; Al-Kadmy, Israa M S; Kareem, Sawsan Mohammed; Lafta, Aliaa Khyuon; A'Affus, Mustafa Hussein Obeid; Khit, Ibrahim Abd Aloahd; Al Kheraif, Abdulaziz Abdullah; Divakar, Darshan Devang; Ramakrishnaiah, Ravikumar

2017-03-01

Pseudomonas aeruginosa is a major cause of nosocomial infection in children and adults, resulting in significant morbidity and mortality due to its ability to acquire drug resistance. The ability of P. aeruginosa in the environment to cause infection in individuals has been reported previously; henceforth, surveillance of the emergence and transmission of P. aeruginosa strains among patients is important for infection control in a clinical setup. Various gene-typing methods have been used for epidemiological typing of P. aeruginosa isolates for the purpose of surveillance. In this work, the suitability and comparability of two typing methods, enterobacterial repetitive intergenic consensus (ERIC)-PCR and random amplification of polymorphic DNA (RAPD)-PCR fingerprinting, were studied to characterize P. aeruginosa strains isolated from clinical and environmental sources. Forty-four clinical and environmental bacterial isolates of P. aeruginosa were collected between October 2015 and January 2016. DNA extraction, ERIC-PCR and RAPD-PCR, agarose gel electrophoresis, and phylogenetic analyses were carried using the unweighted pair-group method with mean. RAPD typing revealed less clonality among clinical isolates, whereas the ERIC method showed greater similarity in comparison with RAPD. Environmental isolates, however, showed greater similarity using RAPD compared with ERIC typing. With only a few exceptions, most clinical isolates were distinct from environmental isolates, irrespective of the typing method. In conclusion, both the RAPD and ERIC typing methods proved to be good tools in understanding clonal diversity. The results also suggest that there is no relationship between clinical and environmental isolates. The absence of clonality among the clinical isolates may indicate that most P. aeruginosa infection cases could be endemic and not epidemic and that endemic infections may be due to nonclonal strains of P. aeruginosa.

Genotypic and Phenotypic Characteristics of Corynebacterium diphtheriae Strains Isolated from Patients in Belarus during an Epidemic Period

PubMed Central

Titov, Leonid; Kolodkina, Valentina; Dronina, Alina; Grimont, Francine; Grimont, Patrick A. D.; Lejay-Collin, Monique; de Zoysa, Aruni; Andronescu, Constantin; Diaconescu, Angela; Marin, Byanca; Efstratiou, Androulla

2003-01-01

One hundred two Corynebacterium diphtheriae strains (93 of the gravis biotype and nine of the mitis biotype) isolated from clinical cases during the Belarus diphtheria epidemic were characterized by biotyping, toxigenicity testing by the Elek test and an indirect hemagglutination assay, phage typing, and ribotyping. The gravis biotype strains were characterized as high and medium toxin producers, and strains of biotype mitis were characterized as low and medium toxin producers. Most strains (82 of 102) were distributed among five phage types. Seventy-two strains (64 of the gravis biotype and 8 of the mitis biotype) belonged to phage type VI ls5,34add. Hybridization of genomic DNA digested with BstEII and PvuII revealed five ribotype patterns, namely, D1, D4, D6, D7, and D13. The majority of gravis biotype strains belonged to ribotypes D1 (49 of 93) and D4 (33 of 93) and included one clonal group of C. diphtheriae. This clone predominated in all regions in Belarus. There was a statistical association between ribotypes and phage types but not between ribotypes and levels of toxin production. PMID:12624069

Putative Panmixia in Restricted Populations of Trypanosoma cruzi Isolated from Wild Triatoma infestans in Bolivia

PubMed Central

Barnabe, Christian; Buitrago, Rosio; Bremond, Philippe; Aliaga, Claudia; Salas, Renata; Vidaurre, Pablo; Herrera, Claudia; Cerqueira, Frédérique; Bosseno, Marie-France; Waleckx, Etienne; Breniere, Simone Frédérique

2013-01-01

Trypanosoma cruzi, the causative agent of Chagas disease, is subdivided into six discrete typing units (DTUs; TcI–TcVI) of which TcI is ubiquitous and genetically highly variable. While clonality is the dominant mode of propagation, recombinant events play a significant evolutive role. Recently, foci of wild Triatoma infestans have been described in Bolivia, mainly infected by TcI. Hence, for the first time, we evaluated the level of genetic exchange within TcI natural potentially panmictic populations (single DTU, host, area and sampling time). Seventy-nine TcI stocks from wild T. infestans, belonging to six populations were characterized at eight microsatellite loci. For each population, Hardy-Weinberg equilibrium (HWE), linkage disequilibrium (LD), and presence of repeated multilocus genotypes (MLG) were analyzed by using a total of seven statistics, to test the null hypothesis of panmixia (H0). For three populations, none of the seven statistics allowed to rejecting H0; for another one the low size did not allow us to conclude, and for the two others the tests have given contradictory results. Interestingly, apparent panmixia was only observed in very restricted areas, and was not observed when grouping populations distant of only two kilometers or more. Nevertheless it is worth stressing that for the statistic tests of "HWE", in order to minimize the type I error (i. e. incorrect rejection of a true H0), we used the Bonferroni correction (BC) known to considerably increase the type II error ( i. e. failure to reject a false H0). For the other tests (LD and MLG), we did not use BC and the risk of type II error in these cases was acceptable. Thus, these results should be considered as a good indicator of the existence of panmixia in wild environment but this must be confirmed on larger samples to reduce the risk of type II error. PMID:24312410

Nonclonal emergence of colistin-resistant Klebsiella pneumoniae isolates from blood samples in South Korea.

PubMed

Suh, Ji-Yoeun; Son, Jun Seong; Chung, Doo Ryeon; Peck, Kyong Ran; Ko, Kwan Soo; Song, Jae-Hoon

2010-01-01

In vitro activities of colistin and other drugs were tested against 221 Klebsiella pneumoniae isolates that were collected between 2006 and 2007 in nine tertiary care South Korean hospitals from patients with bacteremia. The clonality of colistin-resistant K. pneumoniae (CRKP) isolates was assessed by multilocus sequence typing (MLST). We found that 15 isolates (6.8%) were resistant to colistin. MLST showed that CRKP isolates were nonclonal, with colistin resistance in K. pneumoniae occurring independently and not by clonal spreading.

Clonal success of piliated penicillin nonsusceptible pneumococci

PubMed Central

Sjöström, K.; Blomberg, C.; Fernebro, J.; Dagerhamn, J.; Morfeldt, E.; Barocchi, M. A.; Browall, S.; Moschioni, M.; Andersson, M.; Henriques, F.; Albiger, B.; Rappuoli, Rino; Normark, S.; Henriques-Normark, B.

2007-01-01

Antibiotic resistance in pneumococci is due to the spread of strains belonging to a limited number of clones. The Spain9V-3 clone of sequence type (ST)156 is one of the most successful clones with reduced susceptibility to penicillin [pneumococci nonsusceptible to penicillin (PNSP)]. In Sweden during 2000–2003, a dramatic increase in the number of PNSP isolates was observed. Molecular characterization of these isolates showed that a single clone of sequence type ST156 increased from 40% to 80% of all serotype 14, thus causing the serotype expansion. Additionally, during the same time period, we examined the clonal composition of two serotypes 9V and 19F: all 9V and 20% of 19F isolates belonged to the clonal cluster of ST156, and overall ≈50% of all PNSP belonged to the ST156 clonal cluster. Moreover, microarray and PCR analysis showed that all ST156 isolates, irrespective of capsular type, carried the rlrA pilus islet. This islet was also found to be present in the penicillin-sensitive ST162 clone, which is believed to be the drug-susceptible ancestor of ST156. Competitive experiments between related ST156 serotype 19F strains confirmed that those containing the rlrA pilus islet were more successful in an animal model of carriage. We conclude that the pilus island is an important biological factor common to ST156 isolates and other successful PNSP clones. In Sweden, a country where the low antibiotic usage does not explain the spread of resistant strains, at least 70% of all PNSP isolates collected during year 2003 carried the pilus islet. PMID:17644611

Diphtheria in the Republic of Georgia: Use of Molecular Typing Techniques for Characterization of Corynebacterium diphtheriae Strains

PubMed Central

Sulakvelidze, Alexander; Kekelidze, Merab; Gomelauri, Tsaro; Deng, Yingkang; Khetsuriani, Nino; Kobaidze, Ketino; De Zoysa, Aruni; Efstratiou, Androulla; Morris, J. Glenn; Imnadze, Paata

1999-01-01

Sixty-six Corynebacterium diphtheriae strains (62 of the gravis biotype and 4 of the mitis biotype) isolated during the Georgian diphtheria epidemic of 1993 to 1998 and 13 non-Georgian C. diphtheriae strains (10 Russian and 3 reference isolates) were characterized by (i) biotyping, (ii) toxigenicity testing with the Elek assay and PCR, (iii) the randomly amplified polymorphic DNA (RAPD) technique, and (iv) pulsed-field gel electrophoresis (PFGE). Fifteen selected strains were ribotyped. Six RAPD types and 15 PFGE patterns were identified among all strains examined, and 12 ribotypes were found among the 15 strains that were ribotyped. The Georgian epidemic apparently was caused by one major clonal group of C. diphtheriae (PFGE type A, ribotype R1), which was identical to the predominant epidemic strain(s) isolated during the concurrent diphtheria epidemic in Russia. A dendrogram based on the PFGE patterns revealed profound differences between the minor (nonpredominant) epidemic strains found in Georgia and Russia. The methodologies for RAPD typing, ribotyping, and PFGE typing of C. diphtheriae strains were improved to enable rapid and convenient molecular typing of the strains. The RAPD technique was adequate for biotype differentiation; however, PFGE and ribotyping were better (and equal to each other) at discriminating between epidemiologically related and unrelated isolates. PMID:10488190

Metallo-β-lactamase-producing Pseudomonas aeruginosa in the Netherlands: the nationwide emergence of a single sequence type.

PubMed

Van der Bij, A K; Van der Zwan, D; Peirano, G; Severin, J A; Pitout, J D D; Van Westreenen, M; Goessens, W H F

2012-09-01

Recently, the first outbreak of clonally related VIM-2 metallo-β-lactamase (MBL)-producing Pseudomonas aeruginosa in a Dutch tertiary-care centre was described. Subsequently, a nationwide surveillance study was performed in 2010-2011, which identified the presence of VIM-2 MBL-producing P. aeruginosa in 11 different hospitals. Genotyping by multiple-locus variable-number tandem-repeat analysis (MLVA) showed that the majority of the 82 MBL-producing isolates found belonged to a single MLVA type (n = 70, 85%), identified as ST111 by multilocus sequence typing (MLST). As MBL-producing isolates cause serious infections that are difficult to treat, the presence of clonally related isolates in various hospitals throughout the Netherlands is of nationwide concern. © 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

High prevalence and genotypes of Toxoplasma gondii isolated from organic pigs in northern USA.

PubMed

Dubey, J P; Hill, D E; Rozeboom, D W; Rajendran, C; Choudhary, S; Ferreira, L R; Kwok, O C H; Su, C

2012-08-13

The ingestion of undercooked pork infected with Toxoplasma gondii is considered an important source of transmission of this parasite. While T. gondii infection in confinement raised market pigs (market pigs are typically used for fresh, unprocessed pork products) in the USA has decreased significantly over the last 20 years, infection levels in pigs with access to the outdoors can be quite high. An upsurge in consumer demand for 'organically raised', 'humanely raised' and 'free range' pork products has resulted in increasing numbers of hogs being raised in non-confinement systems. To determine T. gondii infection rate in these organic pigs, prevalence of T. gondii in organically raised pigs in two establishments (Farm 1, Farm 2) in Michigan was investigated. Serum and tissue samples from 33 pigs on the farm were available for T. gondii evaluation at slaughter. Serological testing was performed using both ELISA and the modified agglutination test (MAT). Antibodies to T. gondii were detected by both ELISA and MAT in 30 of 33 animals with MAT titers of 1:25 in three, 1:50 in six, 1:100 in seven, 1:200 in 13, and 1:400 in one. Hearts of all 33 pigs were bioassayed for T. gondii in mice; T. gondii was isolated from 17 pigs including one from a seronegative (both ELISA and MAT) pig. Genetic typing of 16 of the 17 T. gondii isolates using the SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico loci revealed clonal Type II from Farm 1 and clonal Type III on Farm 2. These results revealed very high prevalence of T. gondii in organic pigs for the first time in USA, indicating potentially increased health risk of consuming organic swine products. Published by Elsevier B.V.

Molecular Characterization of Carbapenemase-Producing Escherichia coli and Klebsiella pneumoniae in the Countries of the Gulf Cooperation Council: Dominance of OXA-48 and NDM Producers

PubMed Central

Sartor, Anna L.; Balkhy, Hanan H.; Walsh, Timothy R.; Al Johani, Sameera M.; AlJindan, Reem Y.; Alfaresi, Mubarak; Ibrahim, Emad; Al-Jardani, Amina; Al-Abri, Seif; Al Salman, Jameela; Dashti, Ali A.; Kutbi, Abdullah H.; Schlebusch, Sanmarié; Sidjabat, Hanna E.; Paterson, David L.

2014-01-01

The molecular epidemiology and mechanisms of resistance of carbapenem-resistant Enterobacteriaceae (CRE) were determined in hospitals in the countries of the Gulf Cooperation Council (GCC), namely, Saudi Arabia, United Arab Emirates, Oman, Qatar, Bahrain, and Kuwait. Isolates were subjected to PCR-based detection of antibiotic-resistant genes and repetitive sequence-based PCR (rep-PCR) assessments of clonality. Sixty-two isolates which screened positive for potential carbapenemase production were assessed, and 45 were found to produce carbapenemase. The most common carbapenemases were of the OXA-48 (35 isolates) and NDM (16 isolates) types; 6 isolates were found to coproduce the OXA-48 and NDM types. No KPC-type, VIM-type, or IMP-type producers were detected. Multiple clones were detected with seven clusters of clonally related Klebsiella pneumoniae. Awareness of CRE in GCC countries has important implications for controlling the spread of CRE in the Middle East and in hospitals accommodating patients transferred from the region. PMID:24637692

Multilocus Sequence Types of Campylobacter jejuni Isolates from Different Sources in Eastern China.

PubMed

Zhang, Gong; Zhang, Xiaoyan; Hu, Yuanqing; Jiao, Xin-An; Huang, Jinlin

2015-09-01

Campylobacter jejuni is a major food-borne pathogen that causes human gastroenteritis in many developed countries. In our study, we applied multilocus sequence typing (MLST) technology to 167 C. jejuni isolates from diverse sources in Eastern China to examine their genetic diversity. MLST defined 94 sequence types (STs) belonging to 18 clonal complexes (CCs). Forty-five STs from 60 isolates (36%) and 22 alleles have not been previously documented in an international database. One hundred and two isolates, accounting for 61.1% of all isolates, belonged to eight clonal complexes. The eight major CCs were also the most common complexes from different sources. The most common ST type of isolates from human and food was ST-353. The dominant ST type in chicken and foods was ST-354. Among 21 STs that contained two or more different sources isolates, 15 STs contained human isolates and isolates from other sources, suggesting that potentially pathogenic strains are not restricted to specific lineages.

Reassessing the Role of Type II Toxin-Antitoxin Systems in Formation of Escherichia coli Type II Persister Cells.

PubMed

Goormaghtigh, Frédéric; Fraikin, Nathan; Putrinš, Marta; Hallaert, Thibaut; Hauryliuk, Vasili; Garcia-Pino, Abel; Sjödin, Andreas; Kasvandik, Sergo; Udekwu, Klas; Tenson, Tanel; Kaldalu, Niilo; Van Melderen, Laurence

2018-06-12

Persistence is a reversible and low-frequency phenomenon allowing a subpopulation of a clonal bacterial population to survive antibiotic treatments. Upon removal of the antibiotic, persister cells resume growth and give rise to viable progeny. Type II toxin-antitoxin (TA) systems were assumed to play a key role in the formation of persister cells in Escherichia coli based on the observation that successive deletions of TA systems decreased persistence frequency. In addition, the model proposed that stochastic fluctuations of (p)ppGpp levels are the basis for triggering activation of TA systems. Cells in which TA systems are activated are thought to enter a dormancy state and therefore survive the antibiotic treatment. Using independently constructed strains and newly designed fluorescent reporters, we reassessed the roles of TA modules in persistence both at the population and single-cell levels. Our data confirm that the deletion of 10 TA systems does not affect persistence to ofloxacin or ampicillin. Moreover, microfluidic experiments performed with a strain reporting the induction of the yefM-yoeB TA system allowed the observation of a small number of type II persister cells that resume growth after removal of ampicillin. However, we were unable to establish a correlation between high fluorescence and persistence, since the fluorescence of persister cells was comparable to that of the bulk of the population and none of the cells showing high fluorescence were able to resume growth upon removal of the antibiotic. Altogether, these data show that there is no direct link between induction of TA systems and persistence to antibiotics. IMPORTANCE Within a growing bacterial population, a small subpopulation of cells is able to survive antibiotic treatment by entering a transient state of dormancy referred to as persistence. Persistence is thought to be the cause of relapsing bacterial infections and is a major public health concern. Type II toxin-antitoxin systems are small modules composed of a toxic protein and an antitoxin protein counteracting the toxin activity. These systems were thought to be pivotal players in persistence until recent developments in the field. Our results demonstrate that previous influential reports had technical flaws and that there is no direct link between induction of TA systems and persistence to antibiotics. Copyright © 2018 Goormaghtigh et al.

Prevalence and genetic characterization of Toxoplasma gondii in free-range chickens from grocery stores and farms in Maryland, Ohio and Massachusetts, USA.

PubMed

Ying, Yuqing; Verma, Shiv K; Kwok, Oliver C H; Alibana, Fatima; Mcleod, Rima; Su, Chunlei; Dubey, Jitender P; Pradhan, Abani K

2017-05-01

Chickens are considered important in the epidemiology of Toxoplasma gondii. Chicken hearts (n = 1185) obtained from grocery stores were tested for T. gondii infection. Antibodies to T. gondii were assayed in fluid removed from the heart cavity using the modified agglutination test (MAT) at 1:5, 1:25, and 1:100 dilutions. MAT antibodies were detected in 222 hearts at 1:5 dilution and 8 hearts at 1:25 dilution, but none were positive at 1:100 dilution. Seropositive (n = 230, 19.4%) chicken hearts were bioassayed in mice and seronegative (n = 157) chickens were bioassayed in cats. Viable T. gondii was not isolated from any hearts by bioassays in mice. The 2 cats fed 60 and 97 hearts did not excrete T. gondii oocysts. The results indicate a low prevalence of viable T. gondii in chickens from grocery stores. Molecular typing of 23 archived T. gondii strains isolated from free-range chickens from Ohio and Massachusetts using the 10 PCR-RFLP markers including SAG1, SAG2 (5'-3'SAG2 and altSAG2), SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico revealed that seven were ToxoDB PCR-RFLP genotype #1, 11 were genotype #2, one was genotype #3, three were genotype #170, and one was mixed genotype. These results indicate that the clonal genotypes #1 (type II), #2 (type III), and #3 (type II variant) are common in free-range chickens.

Identification and removal of low-complexity sites in allele-specific analysis of ChIP-seq data.

PubMed

Waszak, Sebastian M; Kilpinen, Helena; Gschwind, Andreas R; Orioli, Andrea; Raghav, Sunil K; Witwicki, Robert M; Migliavacca, Eugenia; Yurovsky, Alisa; Lappalainen, Tuuli; Hernandez, Nouria; Reymond, Alexandre; Dermitzakis, Emmanouil T; Deplancke, Bart

2014-01-15

High-throughput sequencing technologies enable the genome-wide analysis of the impact of genetic variation on molecular phenotypes at unprecedented resolution. However, although powerful, these technologies can also introduce unexpected artifacts. We investigated the impact of library amplification bias on the identification of allele-specific (AS) molecular events from high-throughput sequencing data derived from chromatin immunoprecipitation assays (ChIP-seq). Putative AS DNA binding activity for RNA polymerase II was determined using ChIP-seq data derived from lymphoblastoid cell lines of two parent-daughter trios. We found that, at high-sequencing depth, many significant AS binding sites suffered from an amplification bias, as evidenced by a larger number of clonal reads representing one of the two alleles. To alleviate this bias, we devised an amplification bias detection strategy, which filters out sites with low read complexity and sites featuring a significant excess of clonal reads. This method will be useful for AS analyses involving ChIP-seq and other functional sequencing assays. The R package abs filter for library clonality simulations and detection of amplification-biased sites is available from http://updepla1srv1.epfl.ch/waszaks/absfilter

Population genetic structure of Phytophthora cinnamomi associated with avocado in California and the discovery of a potentially recent introduction of a new clonal lineage.

PubMed

Pagliaccia, D; Pond, E; McKee, B; Douhan, G W

2013-01-01

Phytophthora root rot (PRR) of avocado (Persea americana), caused by Phytophthora cinnamomi, is the most serious disease of avocado worldwide. Previous studies have determined that this pathogen exhibits a primarily clonal reproductive mode but no population level studies have been conducted in the avocado-growing regions of California. Therefore, we used amplified fragment length polymorphism based on 22 polymorphic loci and mating type to investigate pathogen diversity from 138 isolates collected in 2009 to 2010 from 15 groves from the Northern and Southern avocado-growing regions. Additional isolates collected from avocado from 1966 to 2007 as well as isolates from other countries and hosts were also used for comparative purposes. Two distinct clades of A2 mating-type isolates from avocado were found based on neighbor joining analysis; one clade contained both newer and older collections from Northern and Southern California, whereas the other clade only contained isolates collected in 2009 and 2010 from Southern California. A third clade was also found that only contained A1 isolates from various hosts. Within the California population, a total of 16 genotypes were found with only one to four genotypes identified from any one location. The results indicate significant population structure in the California avocado P. cinnamomi population, low genotypic diversity consistent with asexual reproduction, potential evidence for the movement of clonal genotypes between the two growing regions, and a potential introduction of a new clonal lineage into Southern California.

Clonality, virulence and antimicrobial resistance of enteroaggregative Escherichia coli from Mirzapur, Bangladesh.

PubMed

Chattaway, Marie Anne; Day, Michaela; Mtwale, Julia; White, Emma; Rogers, James; Day, Martin; Powell, David; Ahmad, Marwa; Harris, Ross; Talukder, Kaisar Ali; Wain, John; Jenkins, Claire; Cravioto, Alejandro

2017-10-01

This study investigates the virulence and antimicrobial resistance in association with common clonal complexes (CCs) of enteroaggregative Escherichia coli (EAEC) isolated from Bangladesh. The aim was to determine whether specific CCs were more likely to be associated with putative virulence genes and/or antimicrobial resistance. The presence of 15 virulence genes (by PCR) and susceptibility to 18 antibiotics were determined for 151 EAEC isolated from cases and controls during an intestinal infectious disease study carried out between 2007-2011 in the rural setting of Mirzapur, Bangladesh (Kotloff KL, Blackwelder WC, Nasrin D, Nataro JP, Farag TH et al.Clin Infect Dis 2012;55:S232-S245). These data were then analysed in the context of previously determined serotypes and clonal complexes defined by multi-locus sequence typing. Overall there was no association between the presence of virulence or antimicrobial resistance genes in isolates of EAEC from cases versus controls. However, when stratified by clonal complex (CC) one CC associated with cases harboured more virulence factors (CC40) and one CC harboured more resistance genes (CC38) than the average. There was no direct link between the virulence gene content and antibiotic resistance. Strains within a single CC had variable virulence and resistance gene content indicating independent and multiple gene acquisitions over time. In Bangladesh, there are multiple clonal complexes of EAEC harbouring a variety of virulence and resistance genes. The emergence of two of the most successful clones appeared to be linked to either increased virulence (CC40) or antimicrobial resistance (CC38), but increased resistance and virulence were not found in the same clonal complexes.

Non-canonical IDH1 and IDH2 mutations: a clonal and relevant event in an Italian cohort of gliomas classified according to the 2016 World Health Organization (WHO) criteria.

PubMed

Visani, Michela; Acquaviva, Giorgia; Marucci, Gianluca; Paccapelo, Alexandro; Mura, Antonella; Franceschi, Enrico; Grifoni, Daniela; Pession, Annalisa; Tallini, Giovanni; Brandes, Alba A; de Biase, Dario

2017-11-01

According to the 2016 World Health Organization (WHO) classification of tumors of the central nervous system, assessment of exon 4 mutations in isocitrate dehydrogenase 1 or 2 genes (IDH1 or IDH2) is an essential step in the characterization of gliomas. The p.R132H mutation is the most frequent alteration in IDH genes, however other non-canonical IDH mutations can be identified. The aim of this study is to investigate in depth the prevalence of non-R132H IDH ("non-canonical") mutations in brain tumors classified according to the 2016 WHO scheme and their clonal distribution in neoplastic cells. A total of 288 consecutive cases of brain gliomas (grade II-IV) were analyzed for exon 4 IDH1 and IDH2 mutations. IDH1 and IDH2 analysis was performed using next generation sequencing. Non-canonical IDH mutations were identified in 13/52 (25.0%) grade II gliomas (astrocytomas: 8/31, 25.8%; oligodendrogliomas: 5/21, 23.8%) and in 5/40 (12.5%) grade III gliomas (astrocytomas: 3/25, 12.0%; oligodendrogliomas: 2/15, 13.3%). They were not identified in 196 grade IV gliomas (192 glioblastomas, 4 gliosarcomas). In the large majority (>80%) of tumors IDH mutations, both IDH1-R132H and the non-canonical ones, were present in the large majority (>80%) of neoplastic cells. Our data highlight the importance of investigating not only the IDH1-R132H mutation but also the non-canonical ones. These mutations are clonally distributed, with proportions of mutated neoplastic cells overlapping with those of p.R132H, a finding consistent with their driver role in gliomagenesis.

Phenotype profiling and multivariate statistical analysis of Spur-pruning type Grapevine in National Clonal Germplasm Repository (NCGR, Davis)

USDA-ARS?s Scientific Manuscript database

Most Korean vineyards employed spur-pruning type modified-T trellis system. This produce system is suitable to spur-pruning type cultivars. But most European table grape is not adaptable to this produce system because their fruitfulness is sufficient to cane-pruning type system. Total 20 of fruit ch...

Genetic shifts in methicillin-resistant Staphylococcus aureus epidemic clones and toxin gene profiles in Japan: comparative analysis among pre-epidemic, epidemic and post-epidemic phases.

PubMed

Osaka, Shunsuke; Okuzumi, Katsuko; Koide, Shota; Tamai, Kiyoko; Sato, Tomoaki; Tanimoto, Koichi; Tomita, Haruyoshi; Suzuki, Masahiro; Nagano, Yukiko; Shibayama, Keigo; Arakawa, Yoshichika; Nagano, Noriyuki

2018-03-01

The decline in methicillin-resistant Staphylococcus aureus (MRSA) isolation rates has become a general observation worldwide, including Japan. We hypothesized that some genetic shift in MRSA might cause this phenomenon, and therefore we investigated the genetic profiles among MRSA clinical isolates obtained from three different epidemic phases in Japan. A total of 353 MRSA isolates were selected from 202 medical facilities in 1990 (pre-epidemic phase), 2004 (epidemic phase) and 2016 (post-epidemic phase). Molecular typing was performed by PCR detection of 22 genes using the polymerase chain reaction (PCR)-based ORF typing (POT) system, including an additional eight genes including small genomic islets and seven toxin genes. Isolates with a POT1 of score 93, identified as presumed clonal complex (pCC)5-staphylococcal cassette chromosome mec (SCCmec) type II including ST5-SCCmec type II New York/Japan clone, represented the major epidemic MRSA lineage in 1990 and 2004. In 2016, however, a marked decrease in isolates with a POT1 score of 93, along with changes in the epidemiology of toxin genes carried, was noted, where the carriers of tst genes including the tst-sec combination were markedly reduced, and those possessing the seb gene alone were markedly increased. Rather, isolates with a POT1 score of 106, including pCC1 or pCC8 among the isolates with SCCmec type IV, which often links to community-associated MRSA, were predominant. Interestingly, the pCC1 and pCC8 lineages were related to sea and tst-sec carriage, respectively. Over time, a transition in MRSA genetic profiles from a POT1 score of 93 in 1990 and 2004 to 106 in 2014 was found in Japan.

[Clinical and biological prognostic factors in relapsed acute myeloid leukemia patients].

PubMed

Yébenes-Ramírez, Manuel; Serrano, Josefina; Martínez-Losada, Carmen; Sánchez-García, Joaquín

2016-09-02

Acute myeloid leukemia (AML) is the most frequent type of acute leukemia in adults. Despite recent advances in the characterization of pathogenesis of AML, the cure rates are under 40%, being leukemia relapse the most common cause of treatment failure. Leukaemia relapse occurs due to clonal evolution or clonal escape. In this study, we aimed to analyze the clinical and biological factors influencing outcomes in patients with AML relapse. We included a total of 75 AML patients who experienced leukaemia relapse after achieving complete remission. We performed complete immunophenotyping and conventional karyotyping in bone marrow aspirates obtained at diagnosis and at leukemia relapse. Overall survival (OS) of the series was 3.7%±2.3, leukaemia progression being the most common cause of death. Patients relapsing before 12 months and those with adverse cytogenetic-molecular risk had statistically significant worse outcomes. A percentage of 52.5 of patients showed phenotypic changes and 50% cytogenetic changes at relapse. We did not find significant clinical factors predicting clonal evolution. The presence of clonal evolution at relapse did not have a significant impact on outcome. Patients with relapsed AML have a dismal prognosis, especially those with early relapse and adverse cytogenetic-molecular risk. Clonal evolution with phenotypic and cytogenetic changes occurred in half of the patients without predictive clinical factors or impact on outcome. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

Distinguishing between incomplete lineage sorting and genomic introgressions: complete fixation of allospecific mitochondrial DNA in a sexually reproducing fish (Cobitis; Teleostei), despite clonal reproduction of hybrids.

PubMed

Choleva, Lukas; Musilova, Zuzana; Kohoutova-Sediva, Alena; Paces, Jan; Rab, Petr; Janko, Karel

2014-01-01

Distinguishing between hybrid introgression and incomplete lineage sorting causing incongruence among gene trees in that they exhibit topological differences requires application of statistical approaches that are based on biologically relevant models. Such study is especially challenging in hybrid systems, where usual vectors mediating interspecific gene transfers--hybrids with Mendelian heredity--are absent or unknown. Here we study a complex of hybridizing species, which are known to produce clonal hybrids, to discover how one of the species, Cobitis tanaitica, has achieved a pattern of mito-nuclear mosaic genome over the whole geographic range. We appplied three distinct methods, including the method using solely the information on gene tree topologies, and found that the contrasting mito-nuclear signal might not have resulted from the retention of ancestral polymorphism. Instead, we found two signs of hybridization events related to C. tanaitica; one concerning nuclear gene flow and the other suggested mitochondrial capture. Interestingly, clonal inheritance (gynogenesis) of contemporary hybrids prevents genomic introgressions and non-clonal hybrids are either absent or too rare to be detected among European Cobitis. Our analyses therefore suggest that introgressive hybridizations are rather old episodes, mediated by previously existing hybrids whose inheritance was not entirely clonal. Cobitis complex thus supports the view that the type of resulting hybrids depends on a level of genomic divergence between sexual species.

Serogroup and Clonal Characterization of Czech Invasive Neisseria meningitidis Strains Isolated from 1971 to 2015

PubMed Central

Jandova, Zuzana; Musilek, Martin; Vackova, Zuzana; Kozakova, Jana; Krizova, Pavla

2016-01-01

Background This study presents antigenic and genetic characteristics of Neisseria meningitidis strains recovered from invasive meningococcal disease (IMD) in the Czech Republic in 1971–2015. Material and Methods A total of 1970 isolates from IMD, referred to the National Reference Laboratory for Meningococcal Infections in 1971–2015, were studied. All isolates were identified and characterized by conventional biochemical and serological tests. Most isolates (82.5%) were characterized by multilocus sequence typing method. Results In the study period 1971–2015, the leading serogroup was B (52.4%), most often assigned to clonal complexes cc32, cc41/44, cc18, and cc269. A significant percentage of strains were of serogroup C (41.4%), with high clonal homogeneity due to hyperinvasive complex cc11, which played an important role in IMD in the Czech Republic in the mid-1990s. Serogroup Y isolates, mostly assigned to cc23, and isolates of clonally homogeneous serogroup W have also been recovered more often over the last years. Conclusion The incidence of IMD and distribution of serogroups and clonal complexes of N. meningitidis in the Czech Republic varied over time, as can be seen from the long-term monitoring, including molecular surveillance data. Data from the conventional and molecular IMD surveillance are helpful in refining the antimeningococcal vaccination strategy in the Czech Republic. PMID:27936105

Clonal Distribution of Disease-Associated and Healthy Carrier Isolates of Neisseria meningitidis between 1983 and 2005 in Cuba ▿

PubMed Central

Climent, Yanet; Yero, Daniel; Martinez, Isabel; Martín, Alejandro; Jolley, Keith A.; Sotolongo, Franklin; Maiden, Martin C. J.; Urwin, Rachel; Pajón, Rolando

2010-01-01

In response to epidemic levels of serogroup B meningococcal disease in Cuba during the 1980s, the VA-MENGOC-BC vaccine was developed and introduced into the National Infant Immunization Program in 1991. Since then the incidence of meningococcal disease in Cuba has returned to the low levels recorded before the epidemic. A total of 420 Neisseria meningitidis strains collected between 1983 and 2005 in Cuba were analyzed by multilocus sequence typing (MLST). The set of strains comprised 167 isolated from disease cases and 253 obtained from healthy carriers. By MLST analysis, 63 sequence types (STs) were identified, and 32 of these were reported to be a new ST. The Cuban isolates were associated with 12 clonal complexes; and the most common were ST-32 (246 isolates), ST-53 (86 isolates), and ST-41/44 (36 isolates). This study also showed that the application of VA-MENGOC-BC, the Cuban serogroup B and C vaccine, reduced the frequency and diversity of hypervirulent clonal complexes ST-32 (vaccine serogroup B type-strain) and ST-41/44 and also affected other lineages. Lineages ST-8 and ST-11 were no longer found during the postvaccination period. The vaccine also affected the genetic composition of the carrier-associated meningococcal isolates. The number of carrier isolates belonging to hypervirulent lineages decreased significantly after vaccination, and ST-53, a sequence type common in carriers, became the predominant ST. PMID:20042619

Verifying Parentage and Confirming Identity in Blackberry with a Fingerprinting Set

USDA-ARS?s Scientific Manuscript database

Parentage and identity confirmation is an important aspect of clonally propagated crops outcrossing. Potential errors resulting misidentification include off-type pollination events, labeling errors, or sports of clones. DNA fingerprinting sets are an excellent solution to quickly identify off-type ...

Nonclonal Emergence of Colistin-Resistant Klebsiella pneumoniae Isolates from Blood Samples in South Korea ▿

PubMed Central

Suh, Ji-Yoeun; Son, Jun Seong; Chung, Doo Ryeon; Peck, Kyong Ran; Ko, Kwan Soo; Song, Jae-Hoon

2010-01-01

In vitro activities of colistin and other drugs were tested against 221 Klebsiella pneumoniae isolates that were collected between 2006 and 2007 in nine tertiary care South Korean hospitals from patients with bacteremia. The clonality of colistin-resistant K. pneumoniae (CRKP) isolates was assessed by multilocus sequence typing (MLST). We found that 15 isolates (6.8%) were resistant to colistin. MLST showed that CRKP isolates were nonclonal, with colistin resistance in K. pneumoniae occurring independently and not by clonal spreading. PMID:19752282

Genetic diversity and virulence profiles of Listeria monocytogenes recovered from bulk tank milk, milk filters, and milking equipment from dairies in the United States (2002 to 2014).

PubMed

Kim, Seon Woo; Haendiges, Julie; Keller, Eric N; Myers, Robert; Kim, Alexander; Lombard, Jason E; Karns, Jeffrey S; Van Kessel, Jo Ann S; Haley, Bradd J

2018-01-01

Unpasteurized dairy products are known to occasionally harbor Listeria monocytogenes and have been implicated in recent listeriosis outbreaks and numerous sporadic cases of listeriosis. However, the diversity and virulence profiles of L. monocytogenes isolates recovered from these products have not been fully described. Here we report a genomic analysis of 121 L. monocytogenes isolates recovered from milk, milk filters, and milking equipment collected from bovine dairy farms in 19 states over a 12-year period. In a multi-virulence-locus sequence typing (MVLST) analysis, 59 Virulence Types (VT) were identified, of which 25% were Epidemic Clones I, II, V, VI, VII, VIII, IX, or X, and 31 were novel VT. In a multi-locus sequence typing (MLST) analysis, 60 Sequence Types (ST) of 56 Clonal Complexes (CC) were identified. Within lineage I, CC5 and CC1 were among the most abundant, and within lineage II, CC7 and CC37 were the most abundant. Multiple CCs previously associated with central nervous system and maternal-neonatal infections were identified. A genomic analysis identified variable distribution of virulence markers, Listeria pathogenicity islands (LIPI) -1, -3, and -4, and stress survival island-1 (SSI-1). Of these, 14 virulence markers, including LIPI-3 and -4 were more frequently detected in one lineage (I or II) than the other. LIPI-3 and LIPI-4 were identified in 68% and 28% of lineage I CCs, respectively. Results of this analysis indicate that there is a high level of genetic diversity among the L. monocytogenes present in bulk tank milk in the United States with some strains being more frequently detected than others, and some being similar to those that have been isolated from previous non-dairy related outbreaks. Results of this study also demonstrate significant number of strains isolated from dairy farms encode virulence markers associated with severe human disease.

Clonal populations of amniotic cells by dilution and direct plating: evidence for hidden diversity.

PubMed

Wilson, Patricia G; Devkota, Lorna; Payne, Tiffany; Crisp, Laddie; Winter, Allison; Wang, Zhan

2012-01-01

Fetal cells are widely considered a superior cell source for regenerative medicine; fetal cells show higher proliferative capacity and have undergone fewer replicative cycles that could generate spontaneous mutations. Fetal cells in amniotic fluid were among the first normal primary cells to be cultured ex vivo, but the undefined composition of amniotic fluid has hindered advance for regenerative applications. We first developed a highly efficient method to generate clonal populations by dilution of amniocentesis samples in media and direct plating without intervening refrigeration, centrifugation, or exposure of cells to the paracrine effects in mixed cell cultures. More than 40 clonal populations were recovered from 4 amniocentesis samples and representative clones were characterized by flow cytometry, conventional assays for differentiation potential, immunofluorescence imaging, and transcript analysis. The results revealed previously unreported diversity among stromal and epithelial cell types and identified unique cell types that could be lost or undetected in mixed cell populations. The differentiation potential of amniotic cells proved to be uncoupled from expression of definitive cell surface or cytoplasmic markers for stromal and epithelial cells. Evidence for diversity among stromal and epithelial cells in amniotic fluid bears on interpretations applied to molecular and functional tests of amniotic cell populations.

Casein gene expression in mouse mammary epithelial cell lines: Dependence upon extracellular matrix and cell type

DOE Office of Scientific and Technical Information (OSTI.GOV)

Medina, D.; Oborn, C.J.; Li, M.L.

1987-09-01

The COMMA-D mammary cell line exhibits mammary-specific functional differentiation under appropriate conditions in cell culture. The cytologically heterogeneous COMMA-D parental line and the clonal lines DB-1, TA-5, and FA-1 derived from the COMMA-D parent were examined for similar properties of functional differentiation. In monolayer cell culture, the cell lines DB-1, TA-5, FA-1, and MA-4 were examined for expression of mammary-specific and epithelial-specific proteins by an indirect immunofluorescence assay. The clonal cell lines were relatively homogeneous in their respective staining properties and seemed to represent three subpopulations found in the heterogeneous parental COMMA-D lines. None of the four clonal lines appearedmore » to represent myoepithelial cells. The cell lines were examined for expression of {beta}-casein mRNA in the presence or absence of prolactin. The inducibility of {beta}-casein in the COMMA-D cell line was further enhanced by a reconstituted basement membrane preparation enriched in laminin, collagen IV, and proteoglycans. These results support the hypothesis that the functional response of inducible mammary cell populations is a result of interaction among hormones, multiple extracellular matrix components, and specific cell types.« less

Recurrent recovery of Pseudomonas oryzihabitans strains in a karstified chalk aquifer.

PubMed

Dussart-Baptista, L; Bodilis, J; Barray, S; Frébourg, N; Fournier, M; Dupont, J-P; Jouenne, T

2007-01-01

Pseudomonas oryzihabitans is an uncommon pathogen that may cause catheter-associated infections, particularly in immunocompromised patients. Although it has been isolated from environment, the source of human infection is not well documented. In the present study, 14 isolates of P. oryzihabitans were recovered over a 28-month period from a karstified chalk aquifer, allowing to advance that distributed natural water could be a source of contamination. Microbiological analyses showed that the bacterium was mainly associated with suspended particulate matters. To investigate the clonality of P. oryzihabitans environmental isolates, 16S rRNA gene sequencing, antibiogram and randomly amplified polymorphic DNA (RAPD) typings were performed. Results demonstrated (i) the presence of at least three clones within the aquifer and (ii) that the presence of the bacterium in groundwater is not only the result of a biofilm bloom but also of an exogenous contamination.

Multilocus sequence type profiles of Bacillus cereus isolates from infant formula in China.

PubMed

Yang, Yong; Yu, Xiaofeng; Zhan, Li; Chen, Jiancai; Zhang, Yunyi; Zhang, Junyan; Chen, Honghu; Zhang, Zheng; Zhang, Yanjun; Lu, Yiyu; Mei, Lingling

2017-04-01

Bacillus cereus sensu stricto is an opportunistic foodborne pathogen. The multilocus sequence type (MLST) of 74 B. cereus isolated from 513 non-random infant formula in China was analyzed. Of 64 sequence types (STs) detected, 50 STs and 6 alleles were newly found in PubMLST database. All isolates except for one singleton (ST-1049), were classified into 7 clonal complexes (CC) by BURST (n-4), in which CC1 with core ancestral clone ST-26 was the largest group including 86% isolates, and CC2, 3, 9, 10 and 13 were first reported in China. MLST profiles of the isolates from 8 infant formula brands were compared. It was found the brands might be potentially tracked by the variety of STs, such as ST-1049 of singleton and ST-1062 of isolate from goat milk source, though they could not be easily tracked just by clonal complex types of the isolates. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mutational landscape, clonal evolution patterns, and role of RAS mutations in relapsed acute lymphoblastic leukemia

PubMed Central

Oshima, Koichi; Khiabanian, Hossein; da Silva-Almeida, Ana C.; Tzoneva, Gannie; Abate, Francesco; Ambesi-Impiombato, Alberto; Sanchez-Martin, Marta; Carpenter, Zachary; Penson, Alex; Perez-Garcia, Arianne; Eckert, Cornelia; Nicolas, Concepción; Balbin, Milagros; Sulis, Maria Luisa; Kato, Motohiro; Koh, Katsuyoshi; Paganin, Maddalena; Basso, Giuseppe; Gastier-Foster, Julie M.; Devidas, Meenakshi; Loh, Mignon L.; Kirschner-Schwabe, Renate; Palomero, Teresa; Rabadan, Raul; Ferrando, Adolfo A.

2016-01-01

Although multiagent combination chemotherapy is curative in a significant fraction of childhood acute lymphoblastic leukemia (ALL) patients, 20% of cases relapse and most die because of chemorefractory disease. Here we used whole-exome and whole-genome sequencing to analyze the mutational landscape at relapse in pediatric ALL cases. These analyses identified numerous relapse-associated mutated genes intertwined in chemotherapy resistance-related protein complexes. In this context, RAS-MAPK pathway-activating mutations in the neuroblastoma RAS viral oncogene homolog (NRAS), kirsten rat sarcoma viral oncogene homolog (KRAS), and protein tyrosine phosphatase, nonreceptor type 11 (PTPN11) genes were present in 24 of 55 (44%) cases in our series. Interestingly, some leukemias showed retention or emergence of RAS mutant clones at relapse, whereas in others RAS mutant clones present at diagnosis were replaced by RAS wild-type populations, supporting a role for both positive and negative selection evolutionary pressures in clonal evolution of RAS-mutant leukemia. Consistently, functional dissection of mouse and human wild-type and mutant RAS isogenic leukemia cells demonstrated induction of methotrexate resistance but also improved the response to vincristine in mutant RAS-expressing lymphoblasts. These results highlight the central role of chemotherapy-driven selection as a central mechanism of leukemia clonal evolution in relapsed ALL, and demonstrate a previously unrecognized dual role of RAS mutations as drivers of both sensitivity and resistance to chemotherapy. PMID:27655895

Characterization of ROP18 alleles in human toxoplasmosis.

PubMed

Sánchez, Víctor; de-la-Torre, Alejandra; Gómez-Marín, Jorge Enrique

2014-04-01

The role of the virulent gene ROP18 polymorphisms is not known in human toxoplasmosis. A total of 320 clinical samples were analyzed. In samples positive for ROP18 gene, we determined by an allele specific PCR, if patients got the upstream insertion positive ROP18 sequence Toxoplasma strain (mouse avirulent strain) or the upstream insertion negative ROP18 sequence Toxoplasma strain (mouse virulent strain). We designed an ELISA assay for antibodies against ROP18 derived peptides from the three major clonal lineages of Toxoplasma. 20 clinical samples were of quality for ROP18 allele analysis. In patients with ocular toxoplasmosis, a higher inflammatory reaction on eye was associated to a PCR negative result for the upstream region of ROP18. 23.3%, 33% and 16.6% of serums from individuals with ocular toxoplasmosis were positive for type I, type II and type III ROP18 derived peptides, respectively but this assay was affected by cross reaction. The absence of Toxoplasma ROP18 promoter insertion sequence in ocular toxoplasmosis was correlated with severe ocular inflammatory response. Determination of antibodies against ROP18 protein was not useful for serotyping in human toxoplasmosis. © 2013.

Clonality, virulence and antimicrobial resistance of enteroaggregative Escherichia coli from Mirzapur, Bangladesh

PubMed Central

Chattaway, Marie Anne; Day, Michaela; Mtwale, Julia; White, Emma; Rogers, James; Day, Martin; Powell, David; Ahmad, Marwa; Harris, Ross; Talukder, Kaisar Ali; Wain, John; Jenkins, Claire; Cravioto, Alejandro

2017-01-01

Purpose This study investigates the virulence and antimicrobial resistance in association with common clonal complexes (CCs) of enteroaggregative Escherichia coli (EAEC) isolated from Bangladesh. The aim was to determine whether specific CCs were more likely to be associated with putative virulence genes and/or antimicrobial resistance. Methodology The presence of 15 virulence genes (by PCR) and susceptibility to 18 antibiotics were determined for 151 EAEC isolated from cases and controls during an intestinal infectious disease study carried out between 2007–2011 in the rural setting of Mirzapur, Bangladesh (Kotloff KL, Blackwelder WC, Nasrin D, Nataro JP, Farag TH et al. Clin Infect Dis 2012;55:S232–S245). These data were then analysed in the context of previously determined serotypes and clonal complexes defined by multi-locus sequence typing. Results Overall there was no association between the presence of virulence or antimicrobial resistance genes in isolates of EAEC from cases versus controls. However, when stratified by clonal complex (CC) one CC associated with cases harboured more virulence factors (CC40) and one CC harboured more resistance genes (CC38) than the average. There was no direct link between the virulence gene content and antibiotic resistance. Strains within a single CC had variable virulence and resistance gene content indicating independent and multiple gene acquisitions over time. Conclusion In Bangladesh, there are multiple clonal complexes of EAEC harbouring a variety of virulence and resistance genes. The emergence of two of the most successful clones appeared to be linked to either increased virulence (CC40) or antimicrobial resistance (CC38), but increased resistance and virulence were not found in the same clonal complexes. PMID:28945190

Breast tumor heterogeneity: cancer stem cells or clonal evolution?

PubMed

Campbell, Lauren L; Polyak, Kornelia

2007-10-01

Breast tumors are composed of a variety of cell types with distinct morphologies and behaviors. It is not clear how this tumor heterogeneity comes about. Two popular concepts that attempt to explain this are the cancer stem cell hypothesis and the clonal evolution model. Each of these ideas has been investigated for some time, leading to the accumulation of numerous findings that are used to support one or the other. Although the two views share some similarities, they are fundamentally different notions with very different clinical implications. Analysis of the research backing each concept, along with a review of the results of our recent study investigating putative breast cancer stem cells, suggests how the cancer stem cell hypothesis and the clonal evolution model may be involved in generating breast tumor heterogeneity. An understanding of this process will allow the development of more effective ways to treat and prevent breast cancer.

Comparative analysis of antibiotic resistance and phylogenetic group patterns in human and porcine urinary tract infectious Escherichia coli.

PubMed

Hancock, Viktoria; Nielsen, Eva Møller; Krag, Louise; Engberg, Jørgen; Klemm, Per

2009-11-01

Urinary tract infections (UTIs) are one of the most common infectious diseases in humans and domestic animals such as pigs. The most frequent infectious agent in such infections is Escherichia coli. Virulence characteristics of E. coli UTI strains range from highly virulent pyelonephritis strains to relatively benign asymptomatic bacteriuria strains. Here we analyse a spectrum of porcine and human UTI E. coli strains with respect to their antibiotic resistance patterns and their phylogenetic groups, determined by multiplex PCR. The clonal profiles of the strains differed profoundly; whereas human strains predominantly belonged to clonal types B2 and D, these were not seen among the porcine strains, which all belonged to the E. coli clonal groups A and B1. Contrary to the human strains, the majority of the porcine strains were multidrug resistant. The distinct profiles of the porcine strains suggest selective pressure due to extensive antibiotic use.

Effect of sugarcane ripener glyphosate (Roundup PowerMAX® II) on growth and biomass characteristics of energy cane

USDA-ARS?s Scientific Manuscript database

Energy cane is considered as one of the most promising feedstock for the emerging bioenergy industry. A random set of 42 energy cane clones bred by the USDA-ARS Sugarcane Research Unit at Houma, LA were taken from a second clonal stage trial and evaluated during 2015-16 at the LSU AgCenter’s Sugar R...

Molecular Characterization of Epithelial Ovarian Cancer: Implications for Diagnosis and Treatment.

PubMed

Rojas, Veronica; Hirshfield, Kim M; Ganesan, Shridar; Rodriguez-Rodriguez, Lorna

2016-12-15

Epithelial ovarian cancer is a highly heterogeneous disease characterized by multiple histological subtypes. Molecular diversity has been shown to occur within specific histological subtypes of epithelial ovarian cancer, between different tumors of an individual patient, as well as within individual tumors. Recent advances in the molecular characterization of epithelial ovarian cancer tumors have provided the basis for a simplified classification scheme in which these cancers are classified as either type I or type II tumors, and these two categories have implications regarding disease pathogenesis and prognosis. Molecular analyses, primarily based on next-generation sequencing, otherwise known as high-throughput sequencing, are allowing for further refinement of ovarian cancer classification, facilitating the elucidation of the site(s) of precursor lesions of high-grade serous ovarian cancer, and providing insight into the processes of clonal selection and evolution that may be associated with development of chemoresistance. Potential therapeutic targets have been identified from recent molecular profiling studies of these tumors, and the effectiveness and safety of a number of specific targeted therapies have been evaluated or are currently being studied for the treatment of women with this disease.

Molecular Characterization of Epithelial Ovarian Cancer: Implications for Diagnosis and Treatment

PubMed Central

Rojas, Veronica; Hirshfield, Kim M.; Ganesan, Shridar; Rodriguez-Rodriguez, Lorna

2016-01-01

Epithelial ovarian cancer is a highly heterogeneous disease characterized by multiple histological subtypes. Molecular diversity has been shown to occur within specific histological subtypes of epithelial ovarian cancer, between different tumors of an individual patient, as well as within individual tumors. Recent advances in the molecular characterization of epithelial ovarian cancer tumors have provided the basis for a simplified classification scheme in which these cancers are classified as either type I or type II tumors, and these two categories have implications regarding disease pathogenesis and prognosis. Molecular analyses, primarily based on next-generation sequencing, otherwise known as high-throughput sequencing, are allowing for further refinement of ovarian cancer classification, facilitating the elucidation of the site(s) of precursor lesions of high-grade serous ovarian cancer, and providing insight into the processes of clonal selection and evolution that may be associated with development of chemoresistance. Potential therapeutic targets have been identified from recent molecular profiling studies of these tumors, and the effectiveness and safety of a number of specific targeted therapies have been evaluated or are currently being studied for the treatment of women with this disease. PMID:27983698

Phage idiotype vaccination: first phase I/II clinical trial in patients with multiple myeloma

PubMed Central

2014-01-01

Background Multiple myeloma is characterized by clonal expansion of B cells producing monoclonal immunoglobulins or fragments thereof, which can be detected in the serum and/or urine and are ideal target antigens for patient-specific immunotherapies. Methods Using phage particles as immunological carriers, we employed a novel chemically linked idiotype vaccine in a clinical phase I/II trial including 15 patients with advanced multiple myeloma. Vaccines composed of purified paraproteins linked to phage were manufactured successfully for each patient. Patients received six intradermal immunizations with phage idiotype vaccines in three different dose groups. Results Phage idiotype was well tolerated by all study participants. A subset of patients (80% in the middle dose group) displayed a clinical response indicated by decrease or stabilization of paraprotein levels. Patients exhibiting a clinical response to phage vaccines also raised idiotype-specific immunoglobulins. Induction of a cellular immune response was demonstrated by a cytotoxicity assay and delayed type hypersensitivity tests. Conclusion We present a simple, time- and cost-efficient phage idiotype vaccination strategy, which represents a safe and feasible patient-specific therapy for patients with advanced multiple myeloma and produced promising anti-tumor activity in a subset of patients. PMID:24885819

Molecular characterization of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the countries of the Gulf cooperation council: dominance of OXA-48 and NDM producers.

PubMed

Zowawi, Hosam M; Sartor, Anna L; Balkhy, Hanan H; Walsh, Timothy R; Al Johani, Sameera M; AlJindan, Reem Y; Alfaresi, Mubarak; Ibrahim, Emad; Al-Jardani, Amina; Al-Abri, Seif; Al Salman, Jameela; Dashti, Ali A; Kutbi, Abdullah H; Schlebusch, Sanmarié; Sidjabat, Hanna E; Paterson, David L

2014-06-01

The molecular epidemiology and mechanisms of resistance of carbapenem-resistant Enterobacteriaceae (CRE) were determined in hospitals in the countries of the Gulf Cooperation Council (GCC), namely, Saudi Arabia, United Arab Emirates, Oman, Qatar, Bahrain, and Kuwait. Isolates were subjected to PCR-based detection of antibiotic-resistant genes and repetitive sequence-based PCR (rep-PCR) assessments of clonality. Sixty-two isolates which screened positive for potential carbapenemase production were assessed, and 45 were found to produce carbapenemase. The most common carbapenemases were of the OXA-48 (35 isolates) and NDM (16 isolates) types; 6 isolates were found to coproduce the OXA-48 and NDM types. No KPC-type, VIM-type, or IMP-type producers were detected. Multiple clones were detected with seven clusters of clonally related Klebsiella pneumoniae. Awareness of CRE in GCC countries has important implications for controlling the spread of CRE in the Middle East and in hospitals accommodating patients transferred from the region. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Polyclonal origin of parathyroid tumors is common and is associated with multiple gland disease in primary hyperparathyroidism.

PubMed

Shi, Yuhong; Azimzadeh, Pedram; Jamingal, Sarada; Wentworth, Shannon; Ferlitch, Janice; Koh, James; Balenga, Nariman; Olson, John A

2018-01-01

Parathyroid tumors are mostly considered monoclonal neoplasms, the rationale for focused parathyroidectomy in primary hyperparathyroidism. We reported that flow sorting parathyroid tumor cells and methylation-sensitive polymerase chain reaction (me-PCR) of polymorphic human androgen receptor gene and phosphoglycerate kinase gene alleles in deoxyribonucleic acid reveals that ≤35% of parathyroid tumors are polyclonal. We sought to confirm these findings and assess for clinical relevance. Parathyroid tumors from 286 female primary hyperparathyroidism patients were analyzed for clonal status. Tumor clonal status was compared with clinical variables and operative findings. Statistical analysis was performed and significance was established at P < .05. In the study, 176 (62%) patients were informative for human androgen receptor gene and/or phosphoglycerate kinase gene. Assignment of clonal status was made in 119 (68%) tumors, of which 64 (54%) were monoclonal and 55 (46%) were polyclonal. Comparison of tumor clonal status to clinical variables in patients with complete operative data (N = 82) showed that while clinical features were the same between tumor types, patients with polyclonal tumors more often had multiple gland disease (risk ratio 4.066, confidence interval, 1.016-16.26; P = .039) potentially missed at unilateral neck exploration. This work confirms that primary hyperparathyroidism is often the result of polyclonal tumors and that parathyroid tumor clonal status may be associated with multiple gland disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Distinguishing Intestinal Lymphoma From Inflammatory Bowel Disease in Canine Duodenal Endoscopic Biopsy Samples.

PubMed

Carrasco, V; Rodríguez-Bertos, A; Rodríguez-Franco, F; Wise, A G; Maes, R; Mullaney, T; Kiupel, M

2015-07-01

Inflammatory bowel disease (IBD) and intestinal lymphoma are intestinal disorders in dogs, both causing similar chronic digestive signs, although with a different prognosis and different treatment requirements. Differentiation between these 2 conditions is based on histopathologic evaluation of intestinal biopsies. However, an accurate diagnosis is often difficult based on histology alone, especially when only endoscopic biopsies are available to differentiate IBD from enteropathy-associated T-cell lymphoma (EATL) type 2, a small cell lymphoma. The purpose of this study was to evaluate the utility of histopathology; immunohistochemistry (IHC) for CD3, CD20, and Ki-67; and polymerase chain reaction (PCR) for antigen receptor rearrangement (T-cell clonality) in the differential diagnosis of severe IBD vs intestinal lymphoma. Endoscopic biopsies from 32 dogs with severe IBD or intestinal lymphoma were evaluated. The original diagnosis was based on microscopic examination of hematoxylin and eosin (HE)-stained sections alone followed by a second evaluation using morphology in association with IHC for CD3 and CD20 and a third evaluation using PCR for clonality. Our results show that, in contrast to feline intestinal lymphomas, 6 of 8 canine small intestinal lymphomas were EATL type 1 (large cell) lymphomas. EATL type 2 was uncommon. Regardless, in dogs, intraepithelial lymphocytes were not an important diagnostic feature to differentiate IBD from EATL as confirmed by PCR. EATL type 1 had a significantly higher Ki-67 index than did EATL type 2 or IBD cases. Based on the results of this study, a stepwise diagnostic approach using histology as the first step, followed by immunophenotyping and determining the Ki67 index and finally PCR for clonality, improves the accuracy of distinguishing intestinal lymphoma from IBD in dogs. © The Author(s) 2014.

The cacao pathogen Moniliophthora roreri (Marasmiaceae) possesses biallelic A and B mating loci but reproduces clonally

PubMed Central

Díaz-Valderrama, J R; Aime, M C

2016-01-01

The cacao pathogen Moniliophthora roreri belongs to the mushroom-forming family Marasmiaceae, but it has never been observed to produce a fruiting body, which calls to question its capacity for sexual reproduction. In this study, we identified potential A (HD1 and HD2) and B (pheromone precursors and pheromone receptors) mating genes in M. roreri. A PCR-based method was subsequently devised to determine the mating type for a set of 47 isolates from across the geographic range of the fungus. We developed and generated an 11-marker microsatellite set and conducted association and linkage disequilibrium (standardized index of association, IAs) analyses. We also performed an ancestral reconstruction analysis to show that the ancestor of M. roreri is predicted to be heterothallic and tetrapolar, which together with sliding window analyses support that the A and B mating loci are likely unlinked and follow a tetrapolar organization within the genome. The A locus is composed of a pair of HD1 and HD2 genes, whereas the B locus consists of a paired pheromone precursor, Mr_Ph4, and receptor, STE3_Mr4. Two A and B alleles but only two mating types were identified. Association analyses divided isolates into two well-defined genetically distinct groups that correlate with their mating type; IAs values show high linkage disequilibrium as is expected in clonal reproduction. Interestingly, both mating types were found in South American isolates but only one mating type was found in Central American isolates, supporting a prior hypothesis of clonal dissemination throughout Central America after a single or very few introductions of the fungus from South America. PMID:26932308

The cacao pathogen Moniliophthora roreri (Marasmiaceae) possesses biallelic A and B mating loci but reproduces clonally.

PubMed

Díaz-Valderrama, J R; Aime, M C

2016-06-01

The cacao pathogen Moniliophthora roreri belongs to the mushroom-forming family Marasmiaceae, but it has never been observed to produce a fruiting body, which calls to question its capacity for sexual reproduction. In this study, we identified potential A (HD1 and HD2) and B (pheromone precursors and pheromone receptors) mating genes in M. roreri. A PCR-based method was subsequently devised to determine the mating type for a set of 47 isolates from across the geographic range of the fungus. We developed and generated an 11-marker microsatellite set and conducted association and linkage disequilibrium (standardized index of association, IA(s)) analyses. We also performed an ancestral reconstruction analysis to show that the ancestor of M. roreri is predicted to be heterothallic and tetrapolar, which together with sliding window analyses support that the A and B mating loci are likely unlinked and follow a tetrapolar organization within the genome. The A locus is composed of a pair of HD1 and HD2 genes, whereas the B locus consists of a paired pheromone precursor, Mr_Ph4, and receptor, STE3_Mr4. Two A and B alleles but only two mating types were identified. Association analyses divided isolates into two well-defined genetically distinct groups that correlate with their mating type; IA(s) values show high linkage disequilibrium as is expected in clonal reproduction. Interestingly, both mating types were found in South American isolates but only one mating type was found in Central American isolates, supporting a prior hypothesis of clonal dissemination throughout Central America after a single or very few introductions of the fungus from South America.

Trends and molecular mechanisms of antimicrobial resistance in clinical staphylococci isolated from companion animals over a 16 year period.

PubMed

Couto, Natacha; Monchique, Cláudia; Belas, Adriana; Marques, Cátia; Gama, Luís T; Pomba, Constança

2016-06-01

The objective of this study was to investigate the evolution of resistance to antimicrobials, corresponding mechanisms and molecular characteristics of Staphylococcus spp., between 1999 and 2014. Susceptibility to 38 antimicrobials was determined for 632 clinical staphylococcal isolates obtained from companion animals (dogs, cats, horses and other animals). Twenty antimicrobial resistance genes, including mecA and mecC, were screened by PCR. Methicillin-resistant staphylococci were characterized by spa (Staphylococcus aureus), SCCmec, MLST and PFGE typing. Statistical analyses were performed using SAS v9.3 and differences were considered relevant if P ≤ 0.05. The mecA gene was identified in 74 staphylococcal isolates (11.6%): 11 MRSA (40.7%), 40 methicillin-resistant Staphylococcus pseudintermedius (MRSP; 8.7%) and 23 methicillin-resistant CoNS (26.7%). Resistance to the majority of antimicrobials and the number of mecA-positive isolates increased significantly over time. Eighteen spa types were identified, including two new ones. MRSA isolates were divided into three PFGE clusters that included ST22-IV, ST105-II, ST398-V and ST5-VI. Most methicillin-resistant Staphylococcus epidermidis isolates were of clonal complex (CC) 5, including a new ST, and clustered in eight PFGE clusters. MRSP were grouped into five PFGE clusters and included ST45-NT, ST71-II-III, ST195-III, ST196-V, ST339-NT, ST342-IV and the new ST400-III. Methicillin-resistant Staphylococcus haemolyticus clustered in two PFGE clusters. The significant increase in antimicrobial-resistant and mecA-positive isolates in recent years is worrying. Furthermore, several isolates are MDR, which complicates antimicrobial treatment and increases the risk of transfer to humans or human isolates. Several clonal lineages of MRSA and methicillin-resistant S. epidermidis circulating in human hospitals and the community were found, suggesting that companion animals can become infected with and contribute to the dissemination of highly successful human clones. Urgent measures, such as determination of clinical breakpoints and guidelines for antimicrobial use, are needed. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Whole genome sequencing analyses of Listeria monocytogenes that persisted in a milkshake machine for a year and caused illnesses in Washington State.

PubMed

Li, Zhen; Pérez-Osorio, Ailyn; Wang, Yu; Eckmann, Kaye; Glover, William A; Allard, Marc W; Brown, Eric W; Chen, Yi

2017-06-15

In 2015, in addition to a United States multistate outbreak linked to contaminated ice cream, another outbreak linked to ice cream was reported in the Pacific Northwest of the United States. It was a hospital-acquired outbreak linked to milkshakes, made from contaminated ice cream mixes and milkshake maker, served to patients. Here we performed multiple analyses on isolates associated with this outbreak: pulsed-field gel electrophoresis (PFGE), whole genome single nucleotide polymorphism (SNP) analysis, species-specific core genome multilocus sequence typing (cgMLST), lineage-specific cgMLST and whole genome-specific MLST (wgsMLST)/outbreak-specific cgMLST. We also analyzed the prophages and virulence genes. The outbreak isolates belonged to sequence type 1038, clonal complex 101, genetic lineage II. There were no pre-mature stop codons in inlA. Isolates contained Listeria Pathogenicity Island 1 and multiple internalins. PFGE and multiple whole genome sequencing (WGS) analyses all clustered together food, environmental and clinical isolates when compared to outgroup from the same clonal complex, which supported the finding that L. monocytogenes likely persisted in the soft serve ice cream/milkshake maker from November 2014 to November 2015 and caused 3 illnesses, and that the outbreak strain was transmitted between two ice cream production facilities. The whole genome SNP analysis, one of the two species-specific cgMLST, the lineage II-specific cgMLST and the wgsMLST/outbreak-specific cgMLST showed that L. monocytogenes cells persistent in the milkshake maker for a year formed a unique clade inside the outbreak cluster. This clustering was consistent with the cleaning practice after the outbreak was initially recognized in late 2014 and early 2015. Putative prophages were conserved among prophage-containing isolates. The loss of a putative prophage in two isolates resulted in the loss of the AscI restriction site in the prophage, which contributed to their AscI-PFGE banding pattern differences from other isolates. The high resolution of WGS analyses allowed the differentiation of epidemiologically unrelated isolates, as well as the elucidation of the microevolution and persistence of isolates within the scope of one outbreak. We applied a wgsMLST scheme which is essentially the outbreak-specific cgMLST. This scheme can be combined with lineage-specific cgMLST and species-specific cgMLST to maximize the resolution of WGS.

Genetic Diversity and Evolution of Salmonella enterica Serovar Enteritidis Strains with Different Phage Types

PubMed Central

Pettengill, James; Strain, Errol; Allard, Marc W.; Ahmed, Rafiq; Zhao, Shaohua; Brown, Eric W.

2014-01-01

Phage typing has been used for the epidemiological surveillance of Salmonella enterica serovar Enteritidis for over 2 decades. However, knowledge of the genetic and evolutionary relationships between phage types is very limited, making differences difficult to interpret. Here, single nucleotide polymorphisms (SNPs) identified from whole-genome comparisons were used to determine the relationships between some S. Enteritidis phage types (PTs) commonly associated with food-borne outbreaks in the United States. Emphasis was placed on the predominant phage types PT8, PT13a, and PT13 in North America. With >89,400 bp surveyed across 98 S. Enteritidis isolates representing 14 distinct phage types, 55 informative SNPs were discovered within 23 chromosomally anchored loci. To maximize the discriminatory and evolutionary partitioning of these highly homogeneous strains, sequences comprising informative SNPs were concatenated into a single combined data matrix and subjected to phylogenetic analysis. The resultant phylogeny allocated most S. Enteritidis isolates into two distinct clades (clades I and II) and four subclades. Synapomorphic (shared and derived) sets of SNPs capable of distinguishing individual clades/subclades were identified. However, individual phage types appeared to be evolutionarily disjunct when mapped to this phylogeny, suggesting that phage typing may not be valid for making phylogenetic inferences. Furthermore, the set of SNPs identified here represents useful genetic markers for strain differentiation of more clonal S. Enteritidis strains and provides core genotypic markers for future development of a SNP typing scheme with S. Enteritidis. PMID:24574287

Characterization and Exploitation of CRISPR Loci in Bifidobacterium longum

PubMed Central

Hidalgo-Cantabrana, Claudio; Crawley, Alexandra B.; Sanchez, Borja; Barrangou, Rodolphe

2017-01-01

Diverse CRISPR-Cas systems provide adaptive immunity in many bacteria and most archaea, via a DNA-encoded, RNA-mediated, nucleic-acid targeting mechanism. Over time, CRISPR loci expand via iterative uptake of invasive DNA sequences into the CRISPR array during the adaptation process. These genetic vaccination cards thus provide insights into the exposure of strains to phages and plasmids in space and time, revealing the historical predatory exposure of a strain. These genetic loci thus constitute a unique basis for genotyping of strains, with potential of resolution at the strain-level. Here, we investigate the occurrence and diversity of CRISPR-Cas systems in the genomes of various Bifidobacterium longum strains across three sub-species. Specifically, we analyzed the genomic content of 66 genomes belonging to B. longum subsp. longum, B. longum subsp. infantis and B. longum subsp. suis, and identified 25 strains that carry 29 total CRISPR-Cas systems. We identify various Type I and Type II CRISPR-Cas systems that are widespread in this species, notably I-C, I-E, and II-C. Noteworthy, Type I-C systems showed extended CRISPR arrays, with extensive spacer diversity. We show how these hypervariable loci can be used to gain insights into strain origin, evolution and phylogeny, and can provide discriminatory sequences to distinguish even clonal isolates. By investigating CRISPR spacer sequences, we reveal their origin and implicate phages and prophages as drivers of CRISPR immunity expansion in this species, with redundant targeting of select prophages. Analysis of CRISPR spacer origin also revealed novel PAM sequences. Our results suggest that CRISPR-Cas immune systems are instrumental in mounting diversified viral resistance in B. longum, and show that these sequences are useful for typing across three subspecies. PMID:29033911

Characterization and Exploitation of CRISPR Loci in Bifidobacterium longum.

PubMed

Hidalgo-Cantabrana, Claudio; Crawley, Alexandra B; Sanchez, Borja; Barrangou, Rodolphe

2017-01-01

Diverse CRISPR-Cas systems provide adaptive immunity in many bacteria and most archaea, via a DNA-encoded, RNA-mediated, nucleic-acid targeting mechanism. Over time, CRISPR loci expand via iterative uptake of invasive DNA sequences into the CRISPR array during the adaptation process. These genetic vaccination cards thus provide insights into the exposure of strains to phages and plasmids in space and time, revealing the historical predatory exposure of a strain. These genetic loci thus constitute a unique basis for genotyping of strains, with potential of resolution at the strain-level. Here, we investigate the occurrence and diversity of CRISPR-Cas systems in the genomes of various Bifidobacterium longum strains across three sub-species. Specifically, we analyzed the genomic content of 66 genomes belonging to B. longum subsp. longum, B. longum subsp. infantis and B. longum subsp. suis , and identified 25 strains that carry 29 total CRISPR-Cas systems. We identify various Type I and Type II CRISPR-Cas systems that are widespread in this species, notably I-C, I-E, and II-C. Noteworthy, Type I-C systems showed extended CRISPR arrays, with extensive spacer diversity. We show how these hypervariable loci can be used to gain insights into strain origin, evolution and phylogeny, and can provide discriminatory sequences to distinguish even clonal isolates. By investigating CRISPR spacer sequences, we reveal their origin and implicate phages and prophages as drivers of CRISPR immunity expansion in this species, with redundant targeting of select prophages. Analysis of CRISPR spacer origin also revealed novel PAM sequences. Our results suggest that CRISPR-Cas immune systems are instrumental in mounting diversified viral resistance in B. longum , and show that these sequences are useful for typing across three subspecies.

Molecular Types of Methicillin-Resistant Staphylococcus aureus and Methicillin-Sensitive S. aureus Strains Causing Skin and Soft Tissue Infections and Nasal Colonization, Identified in Community Health Centers in New York City

PubMed Central

Pardos de la Gandara, Maria; Raygoza Garay, Juan Antonio; Mwangi, Michael; Tobin, Jonathan N.; Tsang, Amanda; Khalida, Chamanara; D'Orazio, Brianna; Kost, Rhonda G.; Leinberger-Jabari, Andrea; Coffran, Cameron; Evering, Teresa H.; Coller, Barry S.; Balachandra, Shirish; Urban, Tracie; Parola, Claude; Salvato, Scott; Jenks, Nancy; Wu, Daren; Burgess, Rhonda; Chung, Marilyn; de Lencastre, Herminia

2015-01-01

In November 2011, The Rockefeller University Center for Clinical and Translational Science (CCTS), the Laboratory of Microbiology and Infectious Diseases, and Clinical Directors Network (CDN) launched a research and learning collaborative project with six community health centers in the New York City metropolitan area to determine the nature (clonal type) of community-acquired Staphylococcus aureus strains causing skin and soft tissue infections (SSTIs). Between November 2011 and March 2013, wound and nasal samples from 129 patients with active SSTIs suspicious for S. aureus were collected and characterized by molecular typing techniques. In 63 of 129 patients, the skin wounds were infected by S. aureus: methicillin-resistant S. aureus (MRSA) was recovered from 39 wounds and methicillin-sensitive S. aureus (MSSA) was recovered from 24. Most—46 of the 63–wound isolates belonged to the CC8/Panton-Valentine leukocidin-positive (PVL+) group of S. aureus clone USA300: 34 of these strains were MRSA and 12 were MSSA. Of the 63 patients with S. aureus infections, 30 were also colonized by S. aureus in the nares: 16 of the colonizing isolates were MRSA, and 14 were MSSA, and the majority of the colonizing isolates belonged to the USA300 clonal group. In most cases (70%), the colonizing isolate belonged to the same clonal type as the strain involved with the infection. In three of the patients, the identity of invasive and colonizing MRSA isolates was further documented by whole-genome sequencing. PMID:26063853

Genetic diversity of Flavobacterium psychrophilum isolates from three Oncorhynchus spp. in the United States, as revealed by multilocus sequence typing

USDA-ARS?s Scientific Manuscript database

Flavobacterium psychrophilum is an important pathogen of salmonids worldwide. Multilocus sequence typing (MLST) has identified a recombinogenic population structure from which emerged a few epidemic clonal complexes particularly threatening for salmonid aquaculture. To date, MLST genotypes for this ...

High prevalence and genotypes of Toxoplasma gondii isolated from goats, from a retail meat store, destined for human consumption in the USA.

PubMed

Dubey, J P; Rajendran, C; Ferreira, L R; Martins, J; Kwok, O C H; Hill, D E; Villena, I; Zhou, H; Su, C; Jones, J L

2011-07-01

Little information is available concerning the presence of viable Toxoplasma gondii in tissues of goats worldwide. In the present study, hearts of 234 goats obtained from a local USA grocery store were examined for T. gondii infection. Blood clot or fluid removed from each heart was tested for antibodies to T. gondii by using the modified agglutination test (MAT). Antibodies to T. gondii were found in 125 (53.4%) of 234 goats, with titers of 1:5 in 20, 1:10 in 44, 1:20 in 16, 1:40 in five, 1:160 in five, 1:320 in five, and 1:640 or higher in 30 goats. Hearts of 112 goats (46 goats <1:5, and 66 goats 1:10 or higher) were used for isolation of viable T. gondii by bioassays in mice. For bioassays, 50 g of the myocardium were digested in an acid pepsin solution and the digest inoculated into mice; the recipient mice were examined for T. gondii infection. Toxoplasma gondii was isolated from 29 goats; from hearts of one of 46 with titers of <1:5, one of nine with titers of 1:10, one of three with titers of 1:40, and 26 of 40 with titers of 1:160 or higher. Two isolates were highly virulent to outbred Swiss Webster mice; all infected mice died of toxoplasmosis, irrespective of the dose. All T. gondii isolates were subsequently grown in cell cultures. Genotyping of the 29 T. gondii isolates using 10 PCR-restriction fragment length polymorphism markers (SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico) from DNA obtained from cell culture grown tachyzoites revealed 12 genotypes. Nine isolates were clonal Type II lineage, four isolates had type II alleles at all loci except a type I allele at the Apico locus, and four isolates were clonal Type III. The remaining 12 strains were divided into nine atypical genotypes, including five new and four previously identified genotypes. DNA sequences of four introns (EF1, HP2, UPRT1 and UPRT7) and two genes (GRA6 and GRA7) were generated for the five new genotypes. Comparing these sequences with previously published data revealed no unique sequences in these goat strains. Taken together, these results indicate high parasite prevalence and moderate genetic diversity of T. gondii in goats, which have important implications in public health. We believe this is the first genetic analysis of T. gondii isolates from goats in the USA. Published by Elsevier Ltd.

Evolutionary perspectives on clonal reproduction in vertebrate animals

PubMed Central

Avise, John C.

2015-01-01

A synopsis is provided of different expressions of whole-animal vertebrate clonality (asexual organismal-level reproduction), both in the laboratory and in nature. For vertebrate taxa, such clonal phenomena include the following: human-mediated cloning via artificial nuclear transfer; intergenerational clonality in nature via parthenogenesis and gynogenesis; intergenerational hemiclonality via hybridogenesis and kleptogenesis; intragenerational clonality via polyembryony; and what in effect qualifies as clonal replication via self-fertilization and intense inbreeding by simultaneous hermaphrodites. Each of these clonal or quasi-clonal mechanisms is described, and its evolutionary genetic ramifications are addressed. By affording an atypical vantage on standard vertebrate reproduction, clonality offers fresh perspectives on the evolutionary and ecological significance of recombination-derived genetic variety. PMID:26195735

Clonal Integration Enhances the Performance of a Clonal Plant Species under Soil Alkalinity Stress

PubMed Central

Sun, Juanjuan; Chen, Jishan; Zhang, Yingjun

2015-01-01

Clonal plants have been shown to successfully survive in stressful environments, including salinity stress, drought and depleted nutrients through clonal integration between original and subsequent ramets. However, relatively little is known about whether clonal integration can enhance the performance of clonal plants under alkalinity stress. We investigated the effect of clonal integration on the performance of a typical rhizomatous clonal plant, Leymus chinensis, using a factorial experimental design with four levels of alkalinity and two levels of rhizome connection treatments, connected (allowing integration) and severed (preventing integration). Clonal integration was estimated by comparing physiological and biomass features between the rhizome-connected and rhizome-severed treatments. We found that rhizome-connected treatment increased the biomass, height and leaf water potential of subsequent ramets at highly alkalinity treatments but did not affect them at low alkalinity treatments. However, rhizome-connected treatment decreased the root biomass of subsequent ramets and did not influence the photosynthetic rates of subsequent ramets. The biomass of original ramets was reduced by rhizome-connected treatment at the highest alkalinity level. These results suggest that clonal integration can increase the performance of clonal plants under alkalinity stress. Rhizome-connected plants showed dramatically increased survival of buds with negative effects on root weight, indicating that clonal integration influenced the resource allocation pattern of clonal plants. A cost-benefit analysis based on biomass measures showed that original and subsequent ramets significantly benefited from clonal integration in highly alkalinity stress, indicating that clonal integration is an important adaptive strategy by which clonal plants could survive in local alkalinity soil. PMID:25790352

Serotype IV Sequence Type 468 Group B Streptococcus Neonatal Invasive Disease, Minnesota, USA.

PubMed

Teatero, Sarah; Ferrieri, Patricia; Fittipaldi, Nahuel

2016-11-01

To further understand the emergence of serotype IV group B Streptococcus (GBS) invasive disease, we used whole-genome sequencing to characterize 3 sequence type 468 strains isolated from neonates in Minnesota, USA. We found that strains of tetracycline-resistant sequence type 468 GBS have acquired virulence genes from a putative clonal complex 17 GBS donor by recombination.

Strain-specific transmission in an outbreak of ESBL-producing Enterobacteriaceae in the hemato-oncology care unit: a cohort study.

PubMed

Uemura, Makiko; Imataki, Osamu; Uchida, Shumpei; Nakayama-Imaohji, Haruyuki; Ohue, Yukiko; Matsuka, Harumi; Mori, Hatsune; Dobashi, Hiroaki; Kuwahara, Tomomi; Kadowaki, Norimitsu

2017-01-05

Extended-spectrum β-lactamase (ESBL)-producing bacteria are resistant to several types of antibiotics excluding carbapenems. A transmissibility of ESBL-producing Enterobacteriaceae would be depending on each bacterial property, however, that has not been elucidated in clinical setting. In this study, we attempted to identify the source of an outbreak of ESBL-producing bacteria in a medical oncology and immunology care unit. An ESBL-producing Enterobacteriaceae (ESBL-E) outbreak observed between July 2012 and August 2012 in Kagawa University Hospital was surveyed using various molecular microbiology techniques. We used Pulsed-field gel electrophoresis (PFGE), PCR-based ESBL gene typing, and direct sequence of ESBL gene as molecular microbiology typing method to distinguish each strain. The typical prevalence of ESBL-E isolation in the unit was 7.0 per month (1.7 per week). The prevalence of ESBL-E isolation during the target research period was 20.0 per month (5.0 per week). In total, 19 isolates (11 K. pneumoniae and 8 E. coli) were obtained from clinical samples, including four control strains (two each of both bacteria), that were physically different from those obtained from other inpatient units in our hospital. Pulsed-field gel electrophoresis (PFGE) for K. pneumoniae (digested by XbaI) produced similar patterns excluding one control strain. PCR classification of the ESBL gene for K. pneumoniae revealed that all strains other than the control strain carried SHV and CTX-M-9. This result was reconfirmed by direct DNA sequencing. Although the outbreak of K. pneumoniae was considered to be "clonal," PFGE and PCR classification of the ESBL genes for E. coli uncovered at least six different "non-clonal" strains possessing individual ESBL gene patterns. According to the result of an antibiogram, the pattern of antimicrobial susceptibility was more variable for K. pneumoniae than for E. coli. Typing by PFGE and ESBL gene PCR analysis is practical for discriminating various organisms. In our cohort, two outbreaks were concomitantly spread with different transmission strategies, namely clonal and non-clonal, in the same unit. This might represent clinical evidence that transmissibility differs according to the type of strain. We speculated that patient-to-patient transmission of ESBL-E occurred according to the properties of each individual strain.

Chlorhexidine whole-body washing of patients reduces methicillin-resistant Staphylococcus aureus and has a direct effect on the distribution of the ST5-MRSA-II (New York/Japan) clone.

PubMed

Velázquez-Meza, Maria Elena; Mendoza-Olazarán, Soraya; Echániz-Aviles, Gabriela; Camacho-Ortiz, Adrián; Martínez-Reséndez, Michel Fernando; Valero-Moreno, Vanessa; Garza-González, Elvira

2017-06-01

Methicillin-resistant Staphylococcus aureus (MRSA) colonizes the skin of hospitalized patients and is associated with high morbidity and mortality. To prevent colonization and infection by S. aureus, better disinfection practices are required. Therefore, we evaluated the effect of chlorhexidine whole-body washing on hospital-acquired S. aureus infections among intensive care unit (ICU) patients in a tertiary hospital in Mexico. The study was conducted over 18 months to evaluate the effect of 2 % chlorhexidine gluconate (CXG) whole-body washing of ICU adult patients on chlorhexidine and antibiotic resistance, biofilm production and clonal distribution of S. aureus in a tertiary care hospital. Minimum inhibitory concentrations for CXG, antibiotic susceptibility and biofilm production by S. aureus isolates were determined. Pulsed-field gel electrophoresis, multilocus sequence typing (MLST) and PCR for Panton-Valentine leucocidin (PVL) were used for molecular typing of MRSA isolates.Results/Key findings. We included 158 isolates. A reduction in antibiotic resistance in the study period was observed for clindamycin, levofloxacin, norfloxacin, oxacillin and trimethoprim/sulfamethoxazole. None of the isolates showed reduced susceptibility to CXG. Most of the isolates were non-biofilm producers (147/158). The most commonly identified clone was a descendant of the ST5-MRSA-II (New York/Japan) clone. This clone decreased during the intervention period and reappeared markedly in the post-intervention period. During the post-intervention period, two isolates were related with the clone ST8-MRSA-IV (also known as USA300). Our findings suggest that the CXG bathing favored the reduction of healthcare-associated MRSA isolates and a temporary reduction of the predominant ST5-MRSA-II (New York/Japan) clone.

Horizontal gene transfer of a ColV plasmid has resulted in a dominant avian clonal type of Salmonella enterica serovar Kentucky.

PubMed

Johnson, Timothy J; Thorsness, Jessica L; Anderson, Cole P; Lynne, Aaron M; Foley, Steven L; Han, Jing; Fricke, W Florian; McDermott, Patrick F; White, David G; Khatri, Mahesh; Stell, Adam L; Flores, Cristian; Singer, Randall S

2010-12-22

Salmonella enterica continues to be a significant cause of foodborne gastrointestinal illness in humans. A wide variety of Salmonella serovars have been isolated from production birds and from retail poultry meat. Recently, though, S. enterica subsp. enterica serovar Kentucky has emerged as one of the prominent Salmonella serovars isolated from broiler chickens. Recent work suggests that its emergence apparently coincides with its acquisition of a ColV virulence plasmid. In the present study, we examined 902 Salmonella isolates belonging to 59 different serovars for the presence of this plasmid. Of the serovars examined, the ColV plasmid was found only among isolates belonging to the serovars Kentucky (72.9%), Typhimurium (15.0%) and Heidelberg (1.7%). We demonstrated that a single PFGE clonal type of S. Kentucky harbors this plasmid, and acquisition of this plasmid by S. Kentucky significantly increased its ability to colonize the chicken cecum and cause extraintestinal disease. Comparison of the completed sequences of three ColV plasmids from S. Kentucky isolated from different geographical locales, timepoints and sources revealed a nearly identical genetic structure with few single nucleotide changes or insertions/deletions. Overall, it appears that the ColV plasmid was recently acquired by a single clonal type S. Kentucky and confers to its host enhanced colonization and fitness capabilities. Thus, the potential for horizontal gene transfer of virulence and fitness factors to Salmonella from other enteric bacteria exists in poultry, representing a potential human health hazard.

Analysis of Clonality and Antibiotic Resistance among Early Clinical Isolates of Enterococcus faecium in the United States

PubMed Central

Galloway-Peña, Jessica R.; Nallapareddy, Sreedhar R.; Arias, Cesar A.; Eliopoulos, George M.; Murray, Barbara E.

2009-01-01

Background The Enterococcus faecium genogroup, referred to as clonal complex 17 (CC17), seems to possess multiple determinants that increase its ability to survive and cause disease in nosocomial environments. Methods Using 53 clinical and geographically diverse US E. faecium isolates dating from 1971 to 1994 we determined the multi-locus sequence type, the presence of 16 putative virulence genes (hylEfm, espEfm and fms genes), resistance to ampicillin (AMPR), vancomycin (VANR) and high-levels of gentamicin and streptomycin. Results Overall, 16 different sequence types (STs), mostly CC17 isolates, were identified in 9 different regions of the US. The earliest CC17 isolates were part of an outbreak in 1982 in Richmond, VA. Characteristics of CC17 isolates included increases in AMPR, the presence of hylEfm and espEfm, emergence of VANR and the presence of at least 13/14 fms genes. Eight out of forty-one of the early AMPR isolates, however, were not within CC17. Conclusions While not all early US AMPR isolates were clonally related, E. faecium CC17 isolates have been circulating in the US since at least 1982 and appear to have progressively acquired additional virulence and antibiotic resistance determinants, perhaps explaining the recent success of this species in the hospital environment. PMID:19821720

Selection of Phage Display Peptides Targeting Human Pluripotent Stem Cell-Derived Progenitor Cell Lines.

PubMed

Bignone, Paola A; Krupa, Rachel A; West, Michael D; Larocca, David

2016-01-01

The ability of human pluripotent stem cells (hPS) to both self-renew and differentiate into virtually any cell type makes them a promising source of cells for cell-based regenerative therapies. However, stem cell identity, purity, and scalability remain formidable challenges that need to be overcome for translation of pluripotent stem cell research into clinical applications. Directed differentiation from hPS cells is inefficient and residual contamination with pluripotent cells that have the potential to form tumors remains problematic. The derivation of scalable (self-renewing) embryonic progenitor stem cell lines offers a solution because they are well defined and clonally pure. Clonally pure progenitor stem cell lines also provide a means for identifying cell surface targeting reagents that are useful for identification, tracking, and repeated derivation of the corresponding progenitor stem cell types from additional hPS cell sources. Such stem cell targeting reagents can then be applied to the manufacture of genetically diverse banks of human embryonic progenitor cell lines for drug screening, disease modeling, and cell therapy. Here we present methods to identify human embryonic progenitor stem cell targeting peptides by selection of phage display libraries on clonal embryonic progenitor cell lines and demonstrate their use for targeting quantum dots (Qdots) for stem cell labeling.

Clonal Populations of Amniotic Cells by Dilution and Direct Plating: Evidence for Hidden Diversity

PubMed Central

Wilson, Patricia G.; Devkota, Lorna; Payne, Tiffany; Crisp, Laddie; Winter, Allison; Wang, Zhan

2012-01-01

Fetal cells are widely considered a superior cell source for regenerative medicine; fetal cells show higher proliferative capacity and have undergone fewer replicative cycles that could generate spontaneous mutations. Fetal cells in amniotic fluid were among the first normal primary cells to be cultured ex vivo, but the undefined composition of amniotic fluid has hindered advance for regenerative applications. We first developed a highly efficient method to generate clonal populations by dilution of amniocentesis samples in media and direct plating without intervening refrigeration, centrifugation, or exposure of cells to the paracrine effects in mixed cell cultures. More than 40 clonal populations were recovered from 4 amniocentesis samples and representative clones were characterized by flow cytometry, conventional assays for differentiation potential, immunofluorescence imaging, and transcript analysis. The results revealed previously unreported diversity among stromal and epithelial cell types and identified unique cell types that could be lost or undetected in mixed cell populations. The differentiation potential of amniotic cells proved to be uncoupled from expression of definitive cell surface or cytoplasmic markers for stromal and epithelial cells. Evidence for diversity among stromal and epithelial cells in amniotic fluid bears on interpretations applied to molecular and functional tests of amniotic cell populations. PMID:23024659

Proposed categorization of pathological states of EBV-associated T/natural killer-cell lymphoproliferative disorder (LPD) in children and young adults: overlap with chronic active EBV infection and infantile fulminant EBV T-LPD.

PubMed

Ohshima, Koichi; Kimura, Hiroshi; Yoshino, Tadashi; Kim, Chul Woo; Ko, Young H; Lee, Seung-Suk; Peh, Suat-Cheng; Chan, John K C

2008-04-01

EBV-associated T/natural killer (NK)-cell lymphoproliferative disorder (EBV-T/NK LPD) of children and young adults is generally referred to with the blanket nosological term of severe chronic active EBV infection (CAEBV). This disease is rare, associated with high morbidity and mortality, and appears to be more prevalent in East Asian countries. But because there is no grading or categorization system for CAEBV, pathologists and clinicians often disagree regarding diagnosis and therapy. EBV-T/NK LPD includes polyclonal, oligoclonal, and monoclonal proliferation of cytotoxic T and/or NK cells. Moreover, a unique disease previously described as infantile fulminant EBV-associated T-LPD has been identified and overlaps with EBV-T/NK LPD. In the present review a clinicopathological categorization of EBV-T/NK LPD is proposed, based on pathological evaluation and molecular data, as follows: (i) category A1, polymorphic LPD without clonal proliferation of EBV-infected cells; (ii) category A2, polymorphic LPD with clonality; (iii) category A3, monomorphic LPD (T-cell or NK cell lymphoma/leukemia) with clonality; and (iv) category B, monomorphic LPD (T-cell lymphoma) with clonality and fulminant course. Categories A1, A2, and A3 possibly constitute a continuous spectrum and together are equivalent to CAEBV. Category B is the exact equivalent of infantile fulminant EBV-associated T-LPD. It is expected that this categorization system will provide a guide for the better understanding of this disorder. This proposal was approved at the third meeting of the Asian Hematopathology Association (Nagoya, 2006).

Panmictic and Clonal Evolution on a Single Patchy Resource Produces Polymorphic Foraging Guilds

PubMed Central

Getz, Wayne M.; Salter, Richard; Lyons, Andrew J.; Sippl-Swezey, Nicolas

2015-01-01

We develop a stochastic, agent-based model to study how genetic traits and experiential changes in the state of agents and available resources influence individuals’ foraging and movement behaviors. These behaviors are manifest as decisions on when to stay and exploit a current resource patch or move to a particular neighboring patch, based on information of the resource qualities of the patches and the anticipated level of intraspecific competition within patches. We use a genetic algorithm approach and an individual’s biomass as a fitness surrogate to explore the foraging strategy diversity of evolving guilds under clonal versus hermaphroditic sexual reproduction. We first present the resource exploitation processes, movement on cellular arrays, and genetic algorithm components of the model. We then discuss their implementation on the Nova software platform. This platform seamlessly combines the dynamical systems modeling of consumer-resource interactions with agent-based modeling of individuals moving over a landscapes, using an architecture that lays transparent the following four hierarchical simulation levels: 1.) within-patch consumer-resource dynamics, 2.) within-generation movement and competition mitigation processes, 3.) across-generation evolutionary processes, and 4.) multiple runs to generate the statistics needed for comparative analyses. The focus of our analysis is on the question of how the biomass production efficiency and the diversity of guilds of foraging strategy types, exploiting resources over a patchy landscape, evolve under clonal versus random hermaphroditic sexual reproduction. Our results indicate greater biomass production efficiency under clonal reproduction only at higher population densities, and demonstrate that polymorphisms evolve and are maintained under random mating systems. The latter result questions the notion that some type of associative mating structure is needed to maintain genetic polymorphisms among individuals exploiting a common patchy resource on an otherwise spatially homogeneous landscape. PMID:26274613

Investigation of the Virulence Factors and Molecular Characterization of the Clonal Relations of Multidrug-Resistant Acinetobacter baumannii Isolates.

PubMed

Ali, Hayssam M; Salem, Mohamed Z M; El-Shikh, Mohamed S; Megeed, Ahmed Abdel; Alogaibi, Yahya A; Talea, Ibrahim Ahmed

2017-01-01

Multidrug-resistant (MDR) Acinetobacter baumannii infections are a great public health concern and demand continuous surveillance and antibiotic stewardship. Virulence traits and the pathogenicity of Acinetobacter are less studied compared with the molecular epidemiological and antibiotic resistance profile of this organism. In our present study, we investigated the primary characteristics contributing to the virulence of MDR A. baumannii isolates and compared them with avirulent isolates. A total of 32 well-characterized MDR A. baumannii clinical isolates and 22 avirulent isolates from a healthy individual were subjected to multilocus sequence typing and polymerase chain reaction (PCR) for a variety of biofilm-associated genes. Additionally, a number of in vitro tests were performed to determine virulence properties. Isolates were found to relate to six sequence types (STs) in which the dominant sequence was ST557 in clinical isolates, followed by ST195 and ST208. However, ST557 and ST222 were absent in avirulent isolates. All STs belonged to clonal complex 2 and clonal lineage 2, which is considered to be a universal clone. PCR analysis showed that most clinical isolates were positive for biofilm-forming genes, such as csu and bap, and also carried pga and ompA genes, which were less common in avirulent isolates. Biofilm formation, phospholipase C production, hemolytic activity, and acinetobactin production occurred significantly more frequently in clinical isolates compared with avirulent isolates. Though A. baumannii clonal lineages showed common virulence traits, they differed in virulent phenotype expression. These findings further support previous studies indicating that A. baumannii is a versatile pathogen with an ability to acquire iron and survive in iron-limiting conditions, highlighting the acinetobactin-mediated iron acquisition mechanisms involved in the pathogenesis of A. baumannii infections.

Epidemiologic and Genotypic Review of Carbapenemase-Producing Organisms in British Columbia, Canada, between 2008 and 2014

PubMed Central

Sekirov, Inna; Croxen, Matthew A.; Ng, Corrinne; Azana, Robert; Chang, Yin; Mataseje, Laura; Boyd, David; Mangat, Chand; Mack, Benjamin; Tadros, Manal; Brodkin, Elizabeth; Kibsey, Pamela; Stefanovic, Aleksandra; Champagne, Sylvie; Mulvey, Michael R.

2015-01-01

Carbapenemase-producing organisms (CPOs) are a serious emerging problem for health care facilities worldwide. Owing to their resistance to most antimicrobial therapies, CPOs are difficult to treat and pose a challenge for infection prevention and control. Since 2010, lab-based surveillance for CPOs and PCR-based testing were implemented in British Columbia (BC), Canada. A review of CPOs in BC from 2008 to March 2014 was done to characterize the resistance mechanisms and possible clonal strain transmission and to compare pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and plasmid restriction fragment length polymorphism (RFLP) as molecular typing tools. During this study period, a total of 177 CPO cases were identified. Patient demographics and travel history were reviewed, and a descriptive analysis was carried out. PFGE profiles, MLST, and plasmid RFLP analysis for a subset of Escherichia coli, Klebsiella pneumoniae, and Enterobacter species isolates were obtained and analyzed. Our findings demonstrate that CPOs have been increasing in number in BC over time, from 1 isolate/year retrospectively identified in 2008 and 2009 to 82 isolates in 2013 and 30 isolates in the first quarter of 2014. Overall, K. pneumoniae isolates lack clonality, although some seemingly related clusters have been found. Plasmid analysis showed evidence of the spread of plasmids carrying carbapenemase-encoding genes between the examined isolates. Analysis of Enterobacter cloacae isolates revealed a more clonal nature of these CPOs in BC. The presence of related clusters provides evidence of interpatient organism transmission both within and between institutions. Although in our study, NDM-harboring E. cloacae isolates appeared to spread clonally, the spread of carbapenem resistance in K. pneumoniae seems to be plasmid mediated. PMID:26607987

Defining Clonal Color in Fluorescent Multi-Clonal Tracking

PubMed Central

Wu, Juwell W.; Turcotte, Raphaël; Alt, Clemens; Runnels, Judith M.; Tsao, Hensin; Lin, Charles P.

2016-01-01

Clonal heterogeneity and selection underpin many biological processes including development and tumor progression. Combinatorial fluorescent protein expression in germline cells has proven its utility for tracking the formation and regeneration of different organ systems. Such cell populations encoded by combinatorial fluorescent proteins are also attractive tools for understanding clonal expansion and clonal competition in cancer. However, the assignment of clonal identity requires an analytical framework in which clonal markings can be parameterized and validated. Here we present a systematic and quantitative method for RGB analysis of fluorescent melanoma cancer clones. We then demonstrate refined clonal trackability of melanoma cells using this scheme. PMID:27073117

Invasive alien plants benefit more from clonal integration in heterogeneous environments than natives.

PubMed

Wang, Yong-Jian; Müller-Schärer, Heinz; van Kleunen, Mark; Cai, Ai-Ming; Zhang, Ping; Yan, Rong; Dong, Bi-Cheng; Yu, Fei-Hai

2017-12-01

What confers invasive alien plants a competitive advantage over native plants remains open to debate. Many of the world's worst invasive alien plants are clonal and able to share resources within clones (clonal integration), particularly in heterogeneous environments. Here, we tested the hypothesis that clonal integration benefits invasive clonal plants more than natives and thus confers invasives a competitive advantage. We selected five congeneric and naturally co-occurring pairs of invasive alien and native clonal plants in China, and grew pairs of connected and disconnected ramets under heterogeneous light, soil nutrient and water conditions that are commonly encountered by alien plants during their invasion into new areas. Clonal integration increased biomass of all plants in all three heterogeneous resource environments. However, invasive plants benefited more from clonal integration than natives. Consequently, invasive plants produced more biomass than natives. Our results indicate that clonal integration may confer invasive alien clonal plants a competitive advantage over natives. Therefore, differences in the ability of clonal integration could potentially explain, at least partly, the invasion success of alien clonal plants in areas where resources are heterogeneously distributed. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

An invasive clonal plant benefits from clonal integration more than a co-occurring native plant in nutrient-patchy and competitive environments.

PubMed

You, Wenhua; Fan, Shufeng; Yu, Dan; Xie, Dong; Liu, Chunhua

2014-01-01

Many notorious invasive plants are clonal, however, little is known about the different roles of clonal integration effects between invasive and native plants. Here, we hypothesize that clonal integration affect growth, photosynthetic performance, biomass allocation and thus competitive ability of invasive and native clonal plants, and invasive clonal plants benefit from clonal integration more than co-occurring native plants in heterogeneous habitats. To test these hypotheses, two stoloniferous clonal plants, Alternanthera philoxeroides (invasive), Jussiaea repens (native) were studied in China. The apical parts of both species were grown either with or without neighboring vegetation and the basal parts without competitors were in nutrient- rich or -poor habitats, with stolon connections were either severed or kept intact. Competition significantly reduced growth and photosynthetic performance of the apical ramets in both species, but not the biomass of neighboring vegetation. Without competition, clonal integration greatly improved the growth and photosynthetic performance of both species, especially when the basal parts were in nutrient-rich habitats. When grown with neighboring vegetation, growth of J. repens and photosynthetic performance of both species were significantly enhanced by clonal integration with the basal parts in both nutrient-rich and -poor habitats, while growth and relative neighbor effect (RNE) of A. philoxeroides were greatly improved by clonal integration only when the basal parts were in nutrient-rich habitats. Moreover, clonal integration increased A. philoxeroides's biomass allocation to roots without competition, but decreased it with competition, especially when the basal ramets were in nutrient-rich sections. Effects of clonal integration on biomass allocation of J. repens was similar to that of A. philoxeroides but with less significance. These results supported our hypothesis that invasive clonal plants A. philoxeroides benefits from clonal integration more than co-occurring native J. repens, suggesting that the invasiveness of A. philoxeroides may be closely related to clonal integration in heterogeneous environments.

An Invasive Clonal Plant Benefits from Clonal Integration More than a Co-Occurring Native Plant in Nutrient-Patchy and Competitive Environments

PubMed Central

You, Wenhua; Fan, Shufeng; Yu, Dan; Xie, Dong; Liu, Chunhua

2014-01-01

Many notorious invasive plants are clonal, however, little is known about the different roles of clonal integration effects between invasive and native plants. Here, we hypothesize that clonal integration affect growth, photosynthetic performance, biomass allocation and thus competitive ability of invasive and native clonal plants, and invasive clonal plants benefit from clonal integration more than co-occurring native plants in heterogeneous habitats. To test these hypotheses, two stoloniferous clonal plants, Alternanthera philoxeroides (invasive), Jussiaea repens (native) were studied in China. The apical parts of both species were grown either with or without neighboring vegetation and the basal parts without competitors were in nutrient- rich or -poor habitats, with stolon connections were either severed or kept intact. Competition significantly reduced growth and photosynthetic performance of the apical ramets in both species, but not the biomass of neighboring vegetation. Without competition, clonal integration greatly improved the growth and photosynthetic performance of both species, especially when the basal parts were in nutrient-rich habitats. When grown with neighboring vegetation, growth of J. repens and photosynthetic performance of both species were significantly enhanced by clonal integration with the basal parts in both nutrient-rich and -poor habitats, while growth and relative neighbor effect (RNE) of A. philoxeroides were greatly improved by clonal integration only when the basal parts were in nutrient-rich habitats. Moreover, clonal integration increased A. philoxeroides's biomass allocation to roots without competition, but decreased it with competition, especially when the basal ramets were in nutrient-rich sections. Effects of clonal integration on biomass allocation of J. repens was similar to that of A. philoxeroides but with less significance. These results supported our hypothesis that invasive clonal plants A. philoxeroides benefits from clonal integration more than co-occurring native J. repens, suggesting that the invasiveness of A. philoxeroides may be closely related to clonal integration in heterogeneous environments. PMID:24816849

Listeria monocytogenes sequence type 1 is predominant in ruminant rhombencephalitis

PubMed Central

Dreyer, Margaux; Aguilar-Bultet, Lisandra; Rupp, Sebastian; Guldimann, Claudia; Stephan, Roger; Schock, Alexandra; Otter, Arthur; Schüpbach, Gertraud; Brisse, Sylvain; Lecuit, Marc; Frey, Joachim; Oevermann, Anna

2016-01-01

Listeria (L.) monocytogenes is an opportunistic pathogen causing life-threatening infections in diverse mammalian species including humans and ruminants. As little is known on the link between strains and clinicopathological phenotypes, we studied potential strain-associated virulence and organ tropism in L. monocytogenes isolates from well-defined ruminant cases of clinical infections and the farm environment. The phylogeny of isolates and their virulence-associated genes were analyzed by multilocus sequence typing (MLST) and sequence analysis of virulence-associated genes. Additionally, a panel of representative isolates was subjected to in vitro infection assays. Our data suggest the environmental exposure of ruminants to a broad range of strains and yet the strong association of sequence type (ST) 1 from clonal complex (CC) 1 with rhombencephalitis, suggesting increased neurotropism of ST1 in ruminants, which is possibly related to its hypervirulence. This study emphasizes the importance of considering clonal background of L. monocytogenes isolates in surveillance, epidemiological investigation and disease control. PMID:27848981

A Single-Cell Roadmap of Lineage Bifurcation in Human ESC Models of Embryonic Brain Development.

PubMed

Yao, Zizhen; Mich, John K; Ku, Sherman; Menon, Vilas; Krostag, Anne-Rachel; Martinez, Refugio A; Furchtgott, Leon; Mulholland, Heather; Bort, Susan; Fuqua, Margaret A; Gregor, Ben W; Hodge, Rebecca D; Jayabalu, Anu; May, Ryan C; Melton, Samuel; Nelson, Angelique M; Ngo, N Kiet; Shapovalova, Nadiya V; Shehata, Soraya I; Smith, Michael W; Tait, Leah J; Thompson, Carol L; Thomsen, Elliot R; Ye, Chaoyang; Glass, Ian A; Kaykas, Ajamete; Yao, Shuyuan; Phillips, John W; Grimley, Joshua S; Levi, Boaz P; Wang, Yanling; Ramanathan, Sharad

2017-01-05

During human brain development, multiple signaling pathways generate diverse cell types with varied regional identities. Here, we integrate single-cell RNA sequencing and clonal analyses to reveal lineage trees and molecular signals underlying early forebrain and mid/hindbrain cell differentiation from human embryonic stem cells (hESCs). Clustering single-cell transcriptomic data identified 41 distinct populations of progenitor, neuronal, and non-neural cells across our differentiation time course. Comparisons with primary mouse and human gene expression data demonstrated rostral and caudal progenitor and neuronal identities from early brain development. Bayesian analyses inferred a unified cell-type lineage tree that bifurcates between cortical and mid/hindbrain cell types. Two methods of clonal analyses confirmed these findings and further revealed the importance of Wnt/β-catenin signaling in controlling this lineage decision. Together, these findings provide a rich transcriptome-based lineage map for studying human brain development and modeling developmental disorders. Copyright © 2017 Elsevier Inc. All rights reserved.

Human Staphylococcus aureus lineages among Zoological Park residents in Greece

PubMed Central

Drougka, E.; Foka, A.; Posantzis, D.; Giormezis, N.; Anastassiou, E.D.; Petinaki, E.; Spiliopoulou, I.

2015-01-01

Staphylococcus aureus is a part of the microbiota flora in many animal species. The clonal spread of S. aureus among animals and personnel in a Zoological Park was investigated. Samples were collected from colonized and infected sites among 32 mammals, 11 birds and eight humans. The genes mecA, mecC, lukF/lukS-PV (encoding Panton-Valentine leukocidin, PVL) and tst (toxic shock syndrome toxin-1) were investigated by PCR. Clones were defined by Multilocus Sequence Typing (MLST), spa type and Pulsed-Field Gel Electrophoresis (PFGE). Seven S. aureus isolates were recovered from four animals and one from an employee. All were mecA, mecC and tst–negative, whereas, one carried the PVL genes and was isolated from an infected Squirrel monkey. Clonal analysis revealed the occurrence of seven STs, eight PFGE and five spa types including ones of human origin. Even though a variety of genotypes were identified among S. aureus strains colonizing zoo park residents, our results indicate that colonization with human lineages has indeed occurred. PMID:26623381

Clonal type I interferon-producing and dendritic cell precursors are contained in both human lymphoid and myeloid progenitor populations.

PubMed

Chicha, Laurie; Jarrossay, David; Manz, Markus G

2004-12-06

Because of different cytokine responsiveness, surface receptor, and transcription factor expression, human CD11c(-) natural type I interferon-producing cells (IPCs) and CD11c(+) dendritic cells were thought to derive through lymphoid and myeloid hematopoietic developmental pathways, respectively. To directly test this hypothesis, we used an in vitro assay allowing simultaneous IPC, dendritic cell, and B cell development and we tested lymphoid and myeloid committed hematopoietic progenitor cells for their developmental capacity. Lymphoid and common myeloid and granulocyte/macrophage progenitors were capable of developing into both functional IPCs, expressing gene transcripts thought to be associated with lymphoid lineage development, and into dendritic cells. However, clonal progenitors for both populations were about fivefold more frequent within myeloid committed progenitor cells. Thus, in humans as in mice, natural IPC and dendritic cell development robustly segregates with myeloid differentiation. This would fit with natural interferon type I-producing cell and dendritic cell activity in innate immunity, the evolutionary older arm of the cellular immune system.

Multiplexing clonality: combining RGB marking and genetic barcoding

PubMed Central

Cornils, Kerstin; Thielecke, Lars; Hüser, Svenja; Forgber, Michael; Thomaschewski, Michael; Kleist, Nadja; Hussein, Kais; Riecken, Kristoffer; Volz, Tassilo; Gerdes, Sebastian; Glauche, Ingmar; Dahl, Andreas; Dandri, Maura; Roeder, Ingo; Fehse, Boris

2014-01-01

RGB marking and DNA barcoding are two cutting-edge technologies in the field of clonal cell marking. To combine the virtues of both approaches, we equipped LeGO vectors encoding red, green or blue fluorescent proteins with complex DNA barcodes carrying color-specific signatures. For these vectors, we generated highly complex plasmid libraries that were used for the production of barcoded lentiviral vector particles. In proof-of-principle experiments, we used barcoded vectors for RGB marking of cell lines and primary murine hepatocytes. We applied single-cell polymerase chain reaction to decipher barcode signatures of individual RGB-marked cells expressing defined color hues. This enabled us to prove clonal identity of cells with one and the same RGB color. Also, we made use of barcoded vectors to investigate clonal development of leukemia induced by ectopic oncogene expression in murine hematopoietic cells. In conclusion, by combining RGB marking and DNA barcoding, we have established a novel technique for the unambiguous genetic marking of individual cells in the context of normal regeneration as well as malignant outgrowth. Moreover, the introduction of color-specific signatures in barcodes will facilitate studies on the impact of different variables (e.g. vector type, transgenes, culture conditions) in the context of competitive repopulation studies. PMID:24476916

Spa typing and identification of pvl genes of meticillin-resistant Staphylococcus aureus isolated from a Libyan hospital in Tripoli.

PubMed

Ahmed, Mohamed O; Baptiste, Keith E; Daw, Mohamed A; Elramalli, Asma K; Abouzeed, Yousef M; Petersen, Andreas

2017-09-01

The purpose of the study was to investigate the molecular characteristics of meticillin-resistant Staphylococcus aureus (MRSA) isolated from clinical sources in Tripoli, Libya. A total of 95 MRSA strains collected at the Tripoli medical Centre were investigated by spa typing and identification of the Panton-Valentine Leukocidin (pvl) genes. A total of 26 spa types were characterized and distributed among nine clonal complexes; CC5 (n=32), CC80 (n=18), CC8 (n=17) and CC22 (n=12) were the most prevalent clonal complexes. In total, 34% of the isolates were positive for PVL. This study demonstrated the presence of CA-MRSA and pvl positive strains in hospital settings and underlines the importance of using molecular typing to investigate the epidemiology of MRSA. Preventative measures and surveillance systems are needed to control and minimize the spread of MRSA in the Libyan health care system. Copyright © 2017 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

Molecular Gene Profiling of Clostridium botulinum Group III and Its Detection in Naturally Contaminated Samples Originating from Various European Countries

PubMed Central

Woudstra, Cedric; Le Maréchal, Caroline; Souillard, Rozenn; Bayon-Auboyer, Marie-Hélène; Anniballi, Fabrizio; Auricchio, Bruna; De Medici, Dario; Bano, Luca; Koene, Miriam; Sansonetti, Marie-Hélène; Desoutter, Denise; Hansbauer, Eva-Maria; Dorner, Martin B.; Dorner, Brigitte G.

2015-01-01

We report the development of real-time PCR assays for genotyping Clostridium botulinum group III targeting the newly defined C. novyi sensu lato group; the nontoxic nonhemagglutinin (NTNH)-encoding gene ntnh; the botulinum neurotoxin (BoNT)-encoding genes bont/C, bont/C/D, bont/D, and bont/D/C; and the flagellin (fliC) gene. The genetic diversity of fliC among C. botulinum group III strains resulted in the definition of five major subgroups named fliC-I to fliC-V. Investigation of fliC subtypes in 560 samples, with various European origins, showed that fliC-I was predominant and found exclusively in samples contaminated by C. botulinum type C/D, fliC-II was rarely detected, no sample was recorded as fliC-III or fliC-V, and only C. botulinum type D/C samples tested positive for fliC-IV. The lack of genetic diversity of the flagellin gene of C. botulinum type C/D would support a clonal spread of type C/D strains in different geographical areas. fliC-I to fliC-III are genetically related (87% to 92% sequence identity), whereas fliC-IV from C. botulinum type D/C is more genetically distant from the other fliC types (with only 50% sequence identity). These findings suggest fliC-I to fliC-III have evolved in a common environment and support a different genetic evolution for fliC-IV. A combination of the C. novyi sensu lato, ntnh, bont, and fliC PCR assays developed in this study allowed better characterization of C. botulinum group III and showed the group to be less genetically diverse than C. botulinum groups I and II, supporting a slow genetic evolution of the strains belonging to C. botulinum group III. PMID:25636839

Geographical distribution of Toxoplasma gondii genotypes in Asia: A link with neighboring continents.

PubMed

Chaichan, P; Mercier, A; Galal, L; Mahittikorn, A; Ariey, F; Morand, S; Boumédiène, F; Udonsom, R; Hamidovic, A; Murat, J B; Sukthana, Y; Dardé, M L

2017-09-01

Defining the pattern of genetic diversity of Toxoplasma gondii is important to understand its worldwide distribution. During the last decades, a large number of studies have been published on Toxoplasma genotypes circulating in Europe, in North and South America. Two continents are still largely unexplored, Africa and, to a less extent, Asia. In this last continent, an increasing number of publications reported genotypes circulating in diverse provinces of China, but very few data are available for other Asian countries. After a systematic database search, 47 papers related to T. gondii genotypes in Asia were analyzed. Genetic characterization of DNA was performed by microsatellite markers, or more usually by a multiplex PCR using 11 PCR-RFLP markers, allowing data comparison to draw a first global picture of the population structure of this parasite throughout Asia. Overall, 390 isolates or DNA extracts were completely typed by PCR-RFLP and/or microsatellite marker methods, revealing 36 different PCR-RFLP or equivalent microsatellite genotypes: 15 genotypes identified by a ToxoDB number and 21 atypical or unique genotypes. The most common genotype found in Asia is the genotype ToxoDB#9 (Chinese 1). The clonal types I, II and II variant, and III were also commonly found in Asia. The geographical distribution of these genotypes across Asia may reflect either a continuum with Europe for the western part of Asia (presence of Type II), or the circulation of strains through animal migration or human activities between Africa and the Southwestern part of Asia (Africa 1 genotype in Turkey or ToxoDB#20 both I Sri-Lanka and in Ethiopia or Egypt). Although there are some indications of a genetic population structure in Southeast Asian countries different from the rest of Asia, more studies in this tropical part of Asia will be necessary for a region which represent as well as Africa one of the missing links of the T. gondii genetic diversity. Copyright © 2017 Elsevier B.V. All rights reserved.

Clonal heterogeneity as a driver of disease variability in the evolution of myeloproliferative neoplasms.

PubMed

Prick, Janine; de Haan, Gerald; Green, Anthony R; Kent, David G

2014-10-01

Myeloproliferative neoplasms (MPNs) are clonal hematological diseases in which cells of the myelo-erythroid lineage are overproduced and patients are predisposed to leukemic transformation. Hematopoietic stem cells are the suspected disease-initiating cells, and these cells must acquire a clonal advantage relative to nonmutant hematopoietic stem cells to perpetuate disease. In 2005, several groups identified a single gain-of-function point mutation in JAK2 that associated with the majority of MPNs, and subsequent studies have led to a comprehensive understanding of the mutational landscape in MPNs. However, confusion still exists as to how a single genetic aberration can be associated with multiple distinct disease entities. Many explanations have been proposed, including JAK2V617F homozygosity, individual patient heterogeneity, and the differential regulation of downstream JAK2 signaling pathways. Several groups have made knock-in mouse models expressing JAK2V617F and have observed divergent phenotypes, each recapitulating some aspects of disease. Intriguingly, most of these models do not observe a strong hematopoietic stem cell self-renewal advantage compared with wild-type littermate controls, raising the question of how a clonal advantage is established in patients with MPNs. This review summarizes the current molecular understanding of MPNs and the diversity of disease phenotypes and proposes that the increased proliferation induced by JAK2V617F applies a selection pressure on the mutant clone that results in highly diverse clonal evolution in individuals. Copyright © 2014 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

Generation of diversity in Streptococcus mutans genes demonstrated by MLST.

PubMed

Do, Thuy; Gilbert, Steven C; Clark, Douglas; Ali, Farida; Fatturi Parolo, Clarissa C; Maltz, Marisa; Russell, Roy R; Holbrook, Peter; Wade, William G; Beighton, David

2010-02-05

Streptococcus mutans, consisting of serotypes c, e, f and k, is an oral aciduric organism associated with the initiation and progression of dental caries. A total of 135 independent Streptococcus mutans strains from caries-free and caries-active subjects isolated from various geographical locations were examined in two versions of an MLST scheme consisting of either 6 housekeeping genes [accC (acetyl-CoA carboxylase biotin carboxylase subunit), gki (glucokinase), lepA (GTP-binding protein), recP (transketolase), sodA (superoxide dismutase), and tyrS (tyrosyl-tRNA synthetase)] or the housekeeping genes supplemented with 2 extracellular putative virulence genes [gtfB (glucosyltransferase B) and spaP (surface protein antigen I/II)] to increase sequence type diversity. The number of alleles found varied between 20 (lepA) and 37 (spaP). Overall, 121 sequence types (STs) were defined using the housekeeping genes alone and 122 with all genes. However pi, nucleotide diversity per site, was low for all loci being in the range 0.019-0.007. The virulence genes exhibited the greatest nucleotide diversity and the recombination/mutation ratio was 0.67 [95% confidence interval 0.3-1.15] compared to 8.3 [95% confidence interval 5.0-14.5] for the 6 concatenated housekeeping genes alone. The ML trees generated for individual MLST loci were significantly incongruent and not significantly different from random trees. Analysis using ClonalFrame indicated that the majority of isolates were singletons and no evidence for a clonal structure or evidence to support serotype c strains as the ancestral S. mutans strain was apparent. There was also no evidence of a geographical distribution of individual isolates or that particular isolate clusters were associated with caries. The overall low sequence diversity suggests that S. mutans is a newly emerged species which has not accumulated large numbers of mutations but those that have occurred have been shuffled as a consequence of intra-species recombination generating genotypes which can be readily distinguished by sequence analysis.

Assessing the genome level diversity of Listeria monocytogenes from contaminated ice cream and environmental samples linked to a listeriosis outbreak in the United States.

PubMed

Chen, Yi; Luo, Yan; Curry, Phillip; Timme, Ruth; Melka, David; Doyle, Matthew; Parish, Mickey; Hammack, Thomas S; Allard, Marc W; Brown, Eric W; Strain, Errol A

2017-01-01

A listeriosis outbreak in the United States implicated contaminated ice cream produced by one company, which operated 3 facilities. We performed single nucleotide polymorphism (SNP)-based whole genome sequencing (WGS) analysis on Listeria monocytogenes from food, environmental and clinical sources, identifying two clusters and a single branch, belonging to PCR serogroup IIb and genetic lineage I. WGS Cluster I, representing one outbreak strain, contained 82 food and environmental isolates from Facility I and 4 clinical isolates. These isolates differed by up to 29 SNPs, exhibited 9 pulsed-field gel electrophoresis (PFGE) profiles and multilocus sequence typing (MLST) sequence type (ST) 5 of clonal complex 5 (CC5). WGS Cluster II contained 51 food and environmental isolates from Facility II, 4 food isolates from Facility I and 5 clinical isolates. Among them the isolates from Facility II and clinical isolates formed a clade and represented another outbreak strain. Isolates in this clade differed by up to 29 SNPs, exhibited 3 PFGE profiles and ST5. The only isolate collected from Facility III belonged to singleton ST489, which was in a single branch separate from Clusters I and II, and was not associated with the outbreak. WGS analyses clustered together outbreak-associated isolates exhibiting multiple PFGE profiles, while differentiating them from epidemiologically unrelated isolates that exhibited outbreak PFGE profiles. The complete genome of a Cluster I isolate allowed the identification and analyses of putative prophages, revealing that Cluster I isolates differed by the gain or loss of three putative prophages, causing the banding pattern differences among all 3 AscI-PFGE profiles observed in Cluster I isolates. WGS data suggested that certain ice cream varieties and/or production lines might have contamination sources unique to them. The SNP-based analysis was able to distinguish CC5 as a group from non-CC5 isolates and differentiate among CC5 isolates from different outbreaks/incidents.

Assessing the genome level diversity of Listeria monocytogenes from contaminated ice cream and environmental samples linked to a listeriosis outbreak in the United States

PubMed Central

Chen, Yi; Luo, Yan; Curry, Phillip; Timme, Ruth; Melka, David; Doyle, Matthew; Parish, Mickey; Hammack, Thomas S.; Allard, Marc W.; Brown, Eric W.; Strain, Errol A.

2017-01-01

A listeriosis outbreak in the United States implicated contaminated ice cream produced by one company, which operated 3 facilities. We performed single nucleotide polymorphism (SNP)-based whole genome sequencing (WGS) analysis on Listeria monocytogenes from food, environmental and clinical sources, identifying two clusters and a single branch, belonging to PCR serogroup IIb and genetic lineage I. WGS Cluster I, representing one outbreak strain, contained 82 food and environmental isolates from Facility I and 4 clinical isolates. These isolates differed by up to 29 SNPs, exhibited 9 pulsed-field gel electrophoresis (PFGE) profiles and multilocus sequence typing (MLST) sequence type (ST) 5 of clonal complex 5 (CC5). WGS Cluster II contained 51 food and environmental isolates from Facility II, 4 food isolates from Facility I and 5 clinical isolates. Among them the isolates from Facility II and clinical isolates formed a clade and represented another outbreak strain. Isolates in this clade differed by up to 29 SNPs, exhibited 3 PFGE profiles and ST5. The only isolate collected from Facility III belonged to singleton ST489, which was in a single branch separate from Clusters I and II, and was not associated with the outbreak. WGS analyses clustered together outbreak-associated isolates exhibiting multiple PFGE profiles, while differentiating them from epidemiologically unrelated isolates that exhibited outbreak PFGE profiles. The complete genome of a Cluster I isolate allowed the identification and analyses of putative prophages, revealing that Cluster I isolates differed by the gain or loss of three putative prophages, causing the banding pattern differences among all 3 AscI-PFGE profiles observed in Cluster I isolates. WGS data suggested that certain ice cream varieties and/or production lines might have contamination sources unique to them. The SNP-based analysis was able to distinguish CC5 as a group from non-CC5 isolates and differentiate among CC5 isolates from different outbreaks/incidents. PMID:28166293

Monitoring the dynamics of clonal tumour evolution in vivo using secreted luciferases.

PubMed

Charles, Joël P; Fuchs, Jeannette; Hefter, Mirjam; Vischedyk, Jonas B; Kleint, Maximilian; Vogiatzi, Fotini; Schäfer, Jonas A; Nist, Andrea; Timofeev, Oleg; Wanzel, Michael; Stiewe, Thorsten

2014-06-03

Tumours are heterogeneous cell populations that undergo clonal evolution during tumour progression, metastasis and response to therapy. Short hairpin RNAs (shRNAs) generate stable loss-of-function phenotypes and are versatile experimental tools to explore the contribution of individual genetic alterations to clonal evolution. In these experiments tumour cells carrying shRNAs are commonly tracked with fluorescent reporters. While this works well for cell culture studies and leukaemia mouse models, fluorescent reporters are poorly suited for animals with solid tumours--the most common tumour types in cancer patients. Here we develop a toolkit that uses secreted luciferases to track the fate of two different shRNA-expressing tumour cell clones competitively, both in vitro and in vivo. We demonstrate that secreted luciferase activities can be measured robustly in the blood stream of tumour-bearing mice to accurately quantify, in a minimally invasive manner, the dynamic evolution of two genetically distinct tumour subclones in preclinical mouse models of tumour development, metastasis and therapy.

Interfaces of Malignant and Immunologic Clonal Dynamics in Ovarian Cancer.

PubMed

Zhang, Allen W; McPherson, Andrew; Milne, Katy; Kroeger, David R; Hamilton, Phineas T; Miranda, Alex; Funnell, Tyler; Little, Nicole; de Souza, Camila P E; Laan, Sonya; LeDoux, Stacey; Cochrane, Dawn R; Lim, Jamie L P; Yang, Winnie; Roth, Andrew; Smith, Maia A; Ho, Julie; Tse, Kane; Zeng, Thomas; Shlafman, Inna; Mayo, Michael R; Moore, Richard; Failmezger, Henrik; Heindl, Andreas; Wang, Yi Kan; Bashashati, Ali; Grewal, Diljot S; Brown, Scott D; Lai, Daniel; Wan, Adrian N C; Nielsen, Cydney B; Huebner, Curtis; Tessier-Cloutier, Basile; Anglesio, Michael S; Bouchard-Côté, Alexandre; Yuan, Yinyin; Wasserman, Wyeth W; Gilks, C Blake; Karnezis, Anthony N; Aparicio, Samuel; McAlpine, Jessica N; Huntsman, David G; Holt, Robert A; Nelson, Brad H; Shah, Sohrab P

2018-05-07

High-grade serous ovarian cancer (HGSC) exhibits extensive malignant clonal diversity with widespread but non-random patterns of disease dissemination. We investigated whether local immune microenvironment factors shape tumor progression properties at the interface of tumor-infiltrating lymphocytes (TILs) and cancer cells. Through multi-region study of 212 samples from 38 patients with whole-genome sequencing, immunohistochemistry, histologic image analysis, gene expression profiling, and T and B cell receptor sequencing, we identified three immunologic subtypes across samples and extensive within-patient diversity. Epithelial CD8+ TILs negatively associated with malignant diversity, reflecting immunological pruning of tumor clones inferred by neoantigen depletion, HLA I loss of heterozygosity, and spatial tracking between T cell and tumor clones. In addition, combinatorial prognostic effects of mutational processes and immune properties were observed, illuminating how specific genomic aberration types associate with immune response and impact survival. We conclude that within-patient spatial immune microenvironment variation shapes intraperitoneal malignant spread, provoking new evolutionary perspectives on HGSC clonal dispersion. Copyright © 2018 Elsevier Inc. All rights reserved.

Monitoring the dynamics of clonal tumour evolution in vivo using secreted luciferases

PubMed Central

Charles, Joël P.; Fuchs, Jeannette; Hefter, Mirjam; Vischedyk, Jonas B.; Kleint, Maximilian; Vogiatzi, Fotini; Schäfer, Jonas A.; Nist, Andrea; Timofeev, Oleg; Wanzel, Michael; Stiewe, Thorsten

2014-01-01

Tumours are heterogeneous cell populations that undergo clonal evolution during tumour progression, metastasis and response to therapy. Short hairpin RNAs (shRNAs) generate stable loss-of-function phenotypes and are versatile experimental tools to explore the contribution of individual genetic alterations to clonal evolution. In these experiments tumour cells carrying shRNAs are commonly tracked with fluorescent reporters. While this works well for cell culture studies and leukaemia mouse models, fluorescent reporters are poorly suited for animals with solid tumours—the most common tumour types in cancer patients. Here we develop a toolkit that uses secreted luciferases to track the fate of two different shRNA-expressing tumour cell clones competitively, both in vitro and in vivo. We demonstrate that secreted luciferase activities can be measured robustly in the blood stream of tumour-bearing mice to accurately quantify, in a minimally invasive manner, the dynamic evolution of two genetically distinct tumour subclones in preclinical mouse models of tumour development, metastasis and therapy. PMID:24889111

Genetic Surveillance Detects Both Clonal and Epidemic Transmission of Malaria following Enhanced Intervention in Senegal

PubMed Central

Séne, Papa Diogoye; Park, Danny C.; Neafsey, Daniel E.; Schaffner, Stephen F.; Hamilton, Elizabeth J.; Lukens, Amanda K.; Van Tyne, Daria; Mboup, Souleymane; Sabeti, Pardis C.; Ndiaye, Daouda; Wirth, Dyann F.

2013-01-01

Using parasite genotyping tools, we screened patients with mild uncomplicated malaria seeking treatment at a clinic in Thiès, Senegal, from 2006 to 2011. We identified a growing frequency of infections caused by genetically identical parasite strains, coincident with increased deployment of malaria control interventions and decreased malaria deaths. Parasite genotypes in some cases persisted clonally across dry seasons. The increase in frequency of genetically identical parasite strains corresponded with decrease in the probability of multiple infections. Further, these observations support evidence of both clonal and epidemic population structures. These data provide the first evidence of a temporal correlation between the appearance of identical parasite types and increased malaria control efforts in Africa, which here included distribution of insecticide treated nets (ITNs), use of rapid diagnostic tests (RDTs) for malaria detection, and deployment of artemisinin combination therapy (ACT). Our results imply that genetic surveillance can be used to evaluate the effectiveness of disease control strategies and assist a rational global malaria eradication campaign. PMID:23593309

The effect of ozone on the yellowing process of magnesium-deficient clonal Norway spruce grown under defined conditions.

PubMed

Siefermann-Harms, Dorothea; Payer, Hans Dieter; Schramel, Peter; Lütz, Cornelius

2005-02-01

During two vegetation periods, young clonal spruce trees (Picea abies (L.) Karst.) with sufficient and poor magnesium (Mg) supply were exposed in the environmental chambers of the GSF phytotron to three levels of ozone (daily means: 18-22, 88-130, and 135-190 microg m(-3); 10% reduction at night). Previous year's needles were examined at 4-week intervals with respect to their contents of Mg, Ca, K, Mn, N, P, and chlorophyll (Chl), various parameters of Chl fluorescence, and the stability of the isolated light-harvesting Chl-a/b-protein complex LHC II. The needles of the two nutrition variants contained more than 0.53 or less than 0.27mg Mg g(-1) needle dry matter, respectively. The ratio of variable to maximal Chl-a fluorescence of the dark-adapted needles, Fv/Fm, and the photoinhibitory quenching of Fv after light treatment, SVi.v, were affected by the Mg content of the needles rather than the ozone levels. Changes of the Chl content and the behavior of the LHC II allowed differentiating between a slow process of needle yellowing occurring under Mg deficiency only, and a rapid process of needle yellowing occurring under the combined action of Mg deficiency and ozone pollution. Only the rapid yellowing process was accompanied by destabilization of the LHC II, and the degree of destabilization was correlated with the ozone concentration present in the days before sampling. The results are consistent with observations obtained at a research site in the Central Black Forest (J Plant Physiol 161 (2004) 423).

Clonality and distribution of clinical Ureaplasma isolates recovered from male patients and infertile couples in China

PubMed Central

Ruan, Zhi; Yang, Ting; Shi, Xinyan; Kong, Yingying; Xie, Xinyou

2017-01-01

Ureaplasma spp. have gained increasing recognition as pathogens in both adult and neonatal patients with multiple clinical presentations. However, the clonality of this organism in the male population and infertile couples in China is largely unknown. In this study, 96 (53 U. parvum and 43 U. urealyticum) of 103 Ureaplasma spp. strains recovered from genital specimens from male patients and 15 pairs of infertile couples were analyzed using multilocus sequence typing (MLST)/expanded multilocus sequence typing (eMLST) schemes. A total of 39 sequence types (STs) and 53 expanded sequence types (eSTs) were identified, with three predominant STs (ST1, ST9 and ST22) and eSTs (eST16, eST41 and eST82). Moreover, phylogenetic analysis revealed two distinct clusters that were highly congruent with the taxonomic differences between the two Ureaplasma species. We found significant differences in the distributions of both clusters and sub-groups between the male and female patients (P < 0.001). Moreover, 66.7% and 40.0% of the male and female partners of the infertile couples tested positive for Ureaplasma spp. The present study also attained excellent agreement of the identification of both Ureaplasma species between paired urine and semen specimens from the male partners (k > 0.80). However, this concordance was observed only for the detection of U. urealyticum within the infertile couples. In conclusion, the distributions of the clusters and sub-groups significantly differed between the male and female patients. U. urealyticum is more likely to transmit between infertile couples and be associated with clinical manifestations by the specific epidemic clonal lineages. PMID:28859153

Clonality and distribution of clinical Ureaplasma isolates recovered from male patients and infertile couples in China.

PubMed

Ruan, Zhi; Yang, Ting; Shi, Xinyan; Kong, Yingying; Xie, Xinyou; Zhang, Jun

2017-01-01

Ureaplasma spp. have gained increasing recognition as pathogens in both adult and neonatal patients with multiple clinical presentations. However, the clonality of this organism in the male population and infertile couples in China is largely unknown. In this study, 96 (53 U. parvum and 43 U. urealyticum) of 103 Ureaplasma spp. strains recovered from genital specimens from male patients and 15 pairs of infertile couples were analyzed using multilocus sequence typing (MLST)/expanded multilocus sequence typing (eMLST) schemes. A total of 39 sequence types (STs) and 53 expanded sequence types (eSTs) were identified, with three predominant STs (ST1, ST9 and ST22) and eSTs (eST16, eST41 and eST82). Moreover, phylogenetic analysis revealed two distinct clusters that were highly congruent with the taxonomic differences between the two Ureaplasma species. We found significant differences in the distributions of both clusters and sub-groups between the male and female patients (P < 0.001). Moreover, 66.7% and 40.0% of the male and female partners of the infertile couples tested positive for Ureaplasma spp. The present study also attained excellent agreement of the identification of both Ureaplasma species between paired urine and semen specimens from the male partners (k > 0.80). However, this concordance was observed only for the detection of U. urealyticum within the infertile couples. In conclusion, the distributions of the clusters and sub-groups significantly differed between the male and female patients. U. urealyticum is more likely to transmit between infertile couples and be associated with clinical manifestations by the specific epidemic clonal lineages.

Presence and mechanisms of acquired antimicrobial resistance in Belgian Brachyspira hyodysenteriae isolates belonging to different clonal complexes.

PubMed

Mahu, M; Pasmans, F; Vranckx, K; De Pauw, N; Vande Maele, L; Vyt, Philip; Vandersmissen, Tamara; Martel, A; Haesebrouck, F; Boyen, F

2017-08-01

Swine dysentery (SD) is an economically important disease for which antimicrobial treatment still occupies an important place to control outbreaks. However, acquired antimicrobial resistance is increasingly observed in Brachyspira hyodysenteriae. In this study, the Minimal Inhibitory Concentrations (MIC) of six antimicrobial compounds for 30 recent Belgian B. hyodysenteriae isolates were determined using a broth microdilution method. In addition, relevant regions of the 16S rRNA, 23S rRNA and the L3 protein encoding genes were sequenced to reveal mutations associated with acquired resistance. Finally, a phylogeny was reconstructed using minimal spanning tree analysis of multi locus sequence typing of the isolates. For lincomycin, doxycycline, tylosin and tylvalosin, at least 70% of the isolates did not belong to the wild-type population and were considered to have acquired resistance. For valnemulin and tiamulin, this was over 50%. In all isolates with acquired resistance to doxycycline, the G1058C mutation was present in their 16S rRNA gene. All isolates showing acquired resistance to lincomycin and both macrolides displayed the A2058T mutation in their 23S rRNA gene. Other mutations in this gene and the N148S mutation in the L3 protein were present in both wild-type isolates and isolates considered to have acquired resistance. Multi locus sequence analysis revealed a previously undescribed clonal complex, with 4 novel sequence types in which the majority of isolates showed acquired resistance to all tested antimicrobial products. In conclusion, acquired antimicrobial resistance is widespread among Belgian B. hyodysenteriae isolates. The emergence of multi-resistant clonal complexes can pose a threat to swine industry. Copyright © 2017 Elsevier B.V. All rights reserved.

The clonal distribution and diversity of extraintestinal Escherichia coli isolates vary according to patient characteristics.

PubMed

Banerjee, Ritu; Johnston, Brian; Lohse, Christine; Chattopadhyay, Sujay; Tchesnokova, Veronika; Sokurenko, Evgeni V; Johnson, James R

2013-12-01

The clonal distribution of Escherichia coli across an unselected population in the current era of widespread antimicrobial resistance is incompletely defined. In this study, we used a newly described clonal typing strategy based on sequencing of fumC and fimH (i.e., CH typing) to infer multilocus sequence types (STs) for 299 consecutive, nonduplicate extraintestinal E. coli isolates from all cultures submitted to Olmsted County, MN, laboratories in February and March 2011 and then compared STs with epidemiological data. Forty-seven different STs were identified, most commonly ST131 (27%), ST95 (11%), ST73 (8%), ST127 (6%), and ST69 (5%). Isolates from these five STs comprised two-thirds of health care-associated (HA) isolates but only half of community-associated (CA) isolates. ST131 was represented overwhelmingly (88%) by a single recently expanded H30 subclone, which was the most extensively antimicrobial-resistant subclone overall and was especially predominant in HA infections and among adults >50 years old. In contrast, among patients 11 to 50 years old, ST69, -95, and -73 were more common. Because of the preponderance of the H30 subclone of ST131, ST diversity was lower among HA than CA isolates, and among antimicrobial-resistant than antimicrobial-susceptible isolates, which otherwise had similar ST distributions. In conclusion, in this U.S. Midwest region, the distribution and diversity of STs among extraintestinal E. coli clinical isolates vary by patient age, type of infection, and resistance phenotype. ST131 predominates among young children and the elderly, HA infections, and antimicrobial-resistant isolates, whereas other well-known pathogenic lineages are more common among adolescents and young adults, CA infections, and antimicrobial-susceptible isolates.

Co-colonization and clonal diversity of methicillin-sensitive and methicillin-resistant Staphylococcus aureus in sows.

PubMed

Fetsch, Alexandra; Roesler, Uwe; Kraushaar, Britta; Friese, Anika

2016-03-15

Methicillin-susceptible Staphylococcus (S.) aureus (MSSA) and methicillin-resistant S. aureus (MRSA) are colonizers of skin and mucosa. In humans, MSSA and MRSA compete for colonization space in the anterior nares of pig farmers; however, it was also shown that MSSA/MRSA co-colonization is common and one clone can be found rather than differing types of MSSA and MRSA. We investigated the colonization and clonality of both, MSSA and MRSA in pigs over a longer time. Eighteen sows were nasally sampled three times every ten weeks. Additionally, environmental samples were taken. Samples were investigated for MSSA and MRSA, respectively. The spa type was defined from up to five MRSA and MSSA isolates found per sample and sampling time; selected isolates were further investigated by microarray. Three sows (16.7%) were completely negative for MSSA and MRSA. Twelve pigs (66.7%) were irregularly positive for both, MSSA and MRSA over the time, whereas seven out of them (38.9%) were simultaneously colonized. CC398 (t034, t011) MRSA and CC9 (t337, t1430, and t13816) MSSA associated spa types were exclusively found. In 44.4% (n=8) of sows up to two different types of MSSA were present at the same time and sample. Strains of the same clonal lineage showed a high genetic identity despite their origin. Highly identic clones were present in sows and their environment. As conclusion, MSSA/MRSA may not exclude each other in the anterior nares of pigs. Pigs may also carry different clones at the same time. Copyright © 2016 Elsevier B.V. All rights reserved.

Molecular Types of Methicillin-Resistant Staphylococcus aureus and Methicillin-Sensitive S. aureus Strains Causing Skin and Soft Tissue Infections and Nasal Colonization, Identified in Community Health Centers in New York City.

PubMed

Pardos de la Gandara, Maria; Raygoza Garay, Juan Antonio; Mwangi, Michael; Tobin, Jonathan N; Tsang, Amanda; Khalida, Chamanara; D'Orazio, Brianna; Kost, Rhonda G; Leinberger-Jabari, Andrea; Coffran, Cameron; Evering, Teresa H; Coller, Barry S; Balachandra, Shirish; Urban, Tracie; Parola, Claude; Salvato, Scott; Jenks, Nancy; Wu, Daren; Burgess, Rhonda; Chung, Marilyn; de Lencastre, Herminia; Tomasz, Alexander

2015-08-01

In November 2011, The Rockefeller University Center for Clinical and Translational Science (CCTS), the Laboratory of Microbiology and Infectious Diseases, and Clinical Directors Network (CDN) launched a research and learning collaborative project with six community health centers in the New York City metropolitan area to determine the nature (clonal type) of community-acquired Staphylococcus aureus strains causing skin and soft tissue infections (SSTIs). Between November 2011 and March 2013, wound and nasal samples from 129 patients with active SSTIs suspicious for S. aureus were collected and characterized by molecular typing techniques. In 63 of 129 patients, the skin wounds were infected by S. aureus: methicillin-resistant S. aureus (MRSA) was recovered from 39 wounds and methicillin-sensitive S. aureus (MSSA) was recovered from 24. Most-46 of the 63-wound isolates belonged to the CC8/Panton-Valentine leukocidin-positive (PVL(+)) group of S. aureus clone USA300: 34 of these strains were MRSA and 12 were MSSA. Of the 63 patients with S. aureus infections, 30 were also colonized by S. aureus in the nares: 16 of the colonizing isolates were MRSA, and 14 were MSSA, and the majority of the colonizing isolates belonged to the USA300 clonal group. In most cases (70%), the colonizing isolate belonged to the same clonal type as the strain involved with the infection. In three of the patients, the identity of invasive and colonizing MRSA isolates was further documented by whole-genome sequencing. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Effect of clonal reproduction on genetic structure in Pentaclethra macroloba (Fabaceae: Mimosoideae).

PubMed

Gaddis, Keith D; Zukin, Helen L; Dieterich, Inca A; Braker, Elizabeth; Sork, Victoria L

2014-06-01

The existence of monodominant forests on well-drained soils in tropical regions has been widely reported. Such forests most likely result from a combination of both ecological and evolutionary factors. Under conditions of high seed and seedling mortality, vegetative reproduction could create a reproductive advantage leading to forest dominance, and profoundly affect the distribution of genetic variation in a clonal species. We investigated these effects in a low diversity forest site in Northeastern Costa Rica dominated by the species Pentaclethra macroloba, which sprouts from the root mass of fallen trees and from snapped trunks. We examined the population structure of juvenile P. macroloba growing in different soil types and across an elevational gradient. Using seven molecular markers, we genotyped 173 juvenile P. macroloba from 18 plots (six plots in seasonally inundated swamps, and 12 plots in upland non-swamp) spanning 50-300m in elevation at La Selva Biological Station and the adjacent Reserva Ecológica Bijagual in Northeastern Costa Rica. We answered two specific questions: (1) How extensive is clonal reproduction? and (2) what is the distribution of genetic diversity and structure? We found that clonal reproduction occurred exclusively within inundated swamp areas. However, there was no significant difference between genetic diversity measures in swamp and non-swamp plots, which were both generally low when compared with other tropical forest species. Genetic structure was significant across all plots (F(ST) = -0.109). However, genetic structure among swamp plots (F(ST) = 0.128) was higher than among non-swamp upland plots (F(ST) = 0.093). Additionally, spatial autocorrelation among individuals within non-swamp upland plots was significant from the 25 to 100m spatial scale, but not within swamp plots. The degree of overall genetic structure we found in P. macroloba is high for a tropical forest tree. The incidence of clonal reproduction is a contributing factor in genetic differentiation, but the high structure among plots without clonal reproduction indicates that other factors contribute as well.

Molecular epidemiology of clonal diploids: a quick overview and a short DIY (do it yourself) notice.

PubMed

De Meeûs, Thierry; Lehmann, Laurent; Balloux, François

2006-03-01

In this short review we report the basic notions needed for understanding the population genetics of clonal diploids. We focus on the consequences of clonality on the distribution of genetic diversity within individuals, between individuals and between populations. We then summarise how to detect clonality in mainly sexual populations, conversely, how to detect sexuality in mainly clonal populations and also how genetic differentiation between populations is affected by clonality in diploids. This information is then used for building recipes on how to analyse and interpret genetic polymorphism data in molecular epidemiology studies of clonal diploids.

[Differentiation of spa types and staphylococcal cassette chromosome mec (SCCmec) in clinical methicillin-resistant Staphylococcus aureus isolated in medical sites of Gdańsk region].

PubMed

Kasprzyk, Joanna; Piechowicz, Lidia; Wiśniewska, Katarzyna; Dziewit, Łukasz; Bronk, Marek; Świeć, Krystyna

2015-01-01

Methicillin-resistant Staphylococcus aureus bacteria are one of the key etiological factors of hospital-acquired and community-acquired infections. MRSA strains have an ability of causing a broad spectrum infections: from a relatively mild skin infections to severe life-threatening systemic infections. They are characterized by multi-drug resistance, virulence of a number of factors, may clonally spread within the hospitals and between hospitals. The study embraced a number of 75 isolates of MRSA isolated from patients of 7 medical sites of the Gdansk region within the period of six months (June to December 2013). Strains have derived from various clinical materials, both of hospitalized patients (n=59) and outpatient (n=16). The isolates were tested for the susceptibility to antimicrobial agents accordance with the guidelines EUCAST. To estimate of the variability of occurrence of S. aureus clones used were standard spa gene, consisting in the amplified polymorphic region of the X gene encoding the protein A gene (spa). After receiving the results, a spa types were identified using international database Ridom Spa Server (www.spaserver.ridom.de). To determine the polymorphism cassette carrying the inecA gene from MRSA strains, used typing five major chromosomal cassette SCCmec (I-V) by multiplex PCR. MRSA population genetic analysis carried out on the basis of typing SCCmec cassettes and spa gene has showed a predominance of strains with SCCmec type II casette (46.7%) and SCCmec IV casette (38.7%). Less frequently detected were strains containing SCCmec I cassette (12.0%) and SCCmec III cassette (2.6%). Spa typing revealed the presence of 13 gene types in MRSA. The most frequently observed spa types were: t151 (24.0%), t003 (16.0%) in strains of the SCCmec II cassette and t437 (16.0%) and t008 (14.8%) in the isolates with SCCmec cassette IV, whereas staphylococcus with the type of spa t011 (12.0%) had SCCmec cassette I. In our population most frequent strains cassette SCCmec II (46.7%), in most representing types of spa t151 (51.4%) and t003 (34.3%), generally resistant not only to β-lactam antibiotics, but as erythromycin, clindamycin and norfloxacin (82.8%), the more frequently they were isolated from patients than a hospital outpatient centers. The strains SCCmec IV that represent the majority of outpatient centers (68.8%), the most represented type t437 (41.4%) and often occurred in hospital centers.

Genetic and molecular predictors of high vancomycin MIC in Staphylococcus aureus bacteremia isolates.

PubMed

Holmes, Natasha E; Turnidge, John D; Munckhof, Wendy J; Robinson, J Owen; Korman, Tony M; O'Sullivan, Matthew V N; Anderson, Tara L; Roberts, Sally A; Warren, Sanchia J C; Coombs, Geoffrey W; Tan, Hui-Leen; Gao, Wei; Johnson, Paul D R; Howden, Benjamin P

2014-09-01

An elevated vancomycin MIC is associated with poor outcomes in Staphylococcus aureus bacteremia (SAB) and is reported in patients with methicillin-susceptible S. aureus (MSSA) bacteremia in the absence of vancomycin treatment. Here, using DNA microarray and phenotype analysis, we investigated the genetic predictors and accessory gene regulator (agr) function and their relationship with elevated vancomycin MIC using blood culture isolates from a multicenter binational cohort of patients with SAB. Specific clonal complexes were associated with elevated (clonal complex 8 [CC8] [P < 0.001]) or low (CC22 [P < 0.001], CC88 [P < 0.001], and CC188 [P = 0.002]) vancomycin MIC. agr dysfunction (P = 0.014) or agr genotype II (P = 0.043) were also associated with an elevated vancomycin MIC. Specific resistance and virulence genes were also linked to an elevated vancomycin MIC, including blaZ (P = 0.002), sea (P < 0.001), clfA (P < 0.001), splA (P = 0.001), and the arginine catabolic mobile element (ACME) locus (P = 0.02). These data suggest that inherent organism characteristics may explain the link between elevated vancomycin MICs and poor outcomes in patients with SAB, regardless of the antibiotic treatment received. A consideration of clonal specificity should be included in future research when attempting to ascertain treatment effects or clinical outcomes. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Donor B cells in Transplants Augment Clonal Expansion and Survival of Pathogenic CD4+ T cells That Mediate Autoimmune-like Chronic GVHD

PubMed Central

Young, James S; Wu, Tao; Chen, Yuhong; Zhao, Dongchang; Liu, Hongjun; Yi, Tangsheng; Johnston, Heather; Racine, Jeremy; Li, Xiaofan; Wang, Audrey; Todorov, Ivan; Zeng, Defu

2013-01-01

We reported that both donor CD4+ T and B cells in transplants were required for induction of an autoimmune-like chronic graft versus host disease (cGVHD) in a murine model of DBA/2 donor to BALB/c recipient, but mechanisms whereby donor B cells augment cGVHD pathogenesis remain unknown. Here, we report that, although donor B cells have little impact on acute GVHD (aGVHD) severity, they play an important role in augmenting the persistence of tissue damage in the acute and chronic GVHD overlapping target organs (i.e. skin and lung); they also markedly augment damage in a prototypical cGVHD target organ- the salivary gland. During cGVHD pathogenesis, donor B cells are activated by donor CD4+ T cells to upregulate MHC II and co-stimulatory molecules. Acting as efficient APCs, donor B cells augment donor CD4+ T clonal expansion, autoreactivity, IL-7Rα expression, and survival. These qualitative changes markedly augment donor CD4+ T cells' capacity in mediating autoimmune-like cGVHD, so that they mediate disease in the absence of donor B cells in secondary recipients. Therefore, a major mechanism whereby donor B cells augment cGVHD is through augmenting the clonal expansion, differentiation and survival of pathogenic CD4+ T cells. PMID:22649197

Clonal growth: invasion or stability? A comparative study of clonal architecture and diversity in native and introduced lineages of Phragmites australis (Poaceae).

PubMed

Douhovnikoff, Vladimir; Hazelton, Eric L G

2014-09-01

• The characteristics of clonal growth that are advantageous in invasive plants can also result in native plants' ability to resist invasion. In Maine, we compared the clonal architecture and diversity of an invasive lineage (introduced Phragmites) and a noninvasive lineage (native Phragmites) present in much of North America. This study is the first on stand-scale diversity using a sample size and systematic spatial-sampling scheme adequate for characterizing clonal structure in Phragmites. Our questions included: (1) Does the structure and extent of clonal growth suggest that the potential for clonal growth contributes to the invasiveness of the introduced lineage? (2) Is clonal growth common in the native lineage, acting as a possible source of ecological resistance and resilience?• Microsatellite markers were used to measure clonal sizes, architecture, and diversity within each lineage in stands within four marshes in Maine.• Clonal diversity measures indicated that clonal growth was significantly greater in stands of the native lineage than in the introduced. While lineage was a consistent predictor of clonal diversity relative ranking, the marsh location was a much stronger predictor of the absolute range of these values.• Our results indicate an important role for clonal growth in the space consolidation of native Phragmites and could explain why the introduced lineage, with stronger competitive traits, has not replaced the native where they co-occur. These results with regard to clone size, size distributions, singleton occurrence, and clonal architecture provide some evidence for stand development that follows a genotypic initial floristics model. © 2014 Botanical Society of America, Inc.

Clonal Growth of Dermal Papilla Cells in Hydrogels Reveals Intrinsic Differences between Sox2-Positive and -Negative Cells In Vitro and In Vivo

PubMed Central

Driskell, Ryan R; Juneja, Vikram R; Connelly, John T; Kretzschmar, Kai; Tan, David W -M; Watt, Fiona M

2012-01-01

In neonatal mouse skin, two types of dermal papilla (DP) are distinguished by Sox2 expression: CD133+Sox2+ DP are associated with guard/awl/auchene hairs, whereas CD133+Sox2− DP are associated with zigzag (ZZ) hairs. We describe a three-dimensional hydrogel culture system that supports clonal growth of CD133+Sox2+, CD133+Sox2−, and CD133−Sox2− (non-DP) neonatal dermal cells. All three cell populations formed spheres that expressed the DP markers alkaline phosphatase, α8 integrin, and CD133. Nevertheless, spheres formed by CD133− cells did not efficiently support hair follicle formation in skin reconstitution assays. In the presence of freshly isolated P2 dermal cells, CD133+Sox2+ and CD133+Sox2− spheres contributed to the DP of both AA and ZZ hairs. Hair type did not correlate with sphere size. Sox2 expression was maintained in culture, but not induced significantly in Sox2− cells in vitro or in vivo, suggesting that Sox2+ cells are a distinct cellular lineage. Although Sox2+ cells were least efficient at forming spheres, they had the greatest ability to contribute to DP and non-DP dermis in reconstituted skin. As the culture system supports clonal growth of DP cells and maintenance of distinct DP cell types, it will be useful for further analysis of intrinsic and extrinsic signals controlling DP function. PMID:22189784

Horizontal Gene Transfer of a ColV Plasmid Has Resulted in a Dominant Avian Clonal Type of Salmonella enterica Serovar Kentucky

PubMed Central

Johnson, Timothy J.; Thorsness, Jessica L.; Anderson, Cole P.; Lynne, Aaron M.; Foley, Steven L.; Han, Jing; Fricke, W. Florian; McDermott, Patrick F.; White, David G.; Khatri, Mahesh; Stell, Adam L.; Flores, Cristian; Singer, Randall S.

2010-01-01

Salmonella enterica continues to be a significant cause of foodborne gastrointestinal illness in humans. A wide variety of Salmonella serovars have been isolated from production birds and from retail poultry meat. Recently, though, S. enterica subsp. enterica serovar Kentucky has emerged as one of the prominent Salmonella serovars isolated from broiler chickens. Recent work suggests that its emergence apparently coincides with its acquisition of a ColV virulence plasmid. In the present study, we examined 902 Salmonella isolates belonging to 59 different serovars for the presence of this plasmid. Of the serovars examined, the ColV plasmid was found only among isolates belonging to the serovars Kentucky (72.9%), Typhimurium (15.0%) and Heidelberg (1.7%). We demonstrated that a single PFGE clonal type of S. Kentucky harbors this plasmid, and acquisition of this plasmid by S. Kentucky significantly increased its ability to colonize the chicken cecum and cause extraintestinal disease. Comparison of the completed sequences of three ColV plasmids from S. Kentucky isolated from different geographical locales, timepoints and sources revealed a nearly identical genetic structure with few single nucleotide changes or insertions/deletions. Overall, it appears that the ColV plasmid was recently acquired by a single clonal type S. Kentucky and confers to its host enhanced colonization and fitness capabilities. Thus, the potential for horizontal gene transfer of virulence and fitness factors to Salmonella from other enteric bacteria exists in poultry, representing a potential human health hazard. PMID:21203520

Genetic variation among Staphylococcus aureus strains from Norwegian bulk milk.

PubMed

Jørgensen, H J; Mørk, T; Caugant, D A; Kearns, A; Rørvik, L M

2005-12-01

Strains of Staphylococcus aureus obtained from bovine (n = 117) and caprine (n = 114) bulk milk were characterized and compared with S. aureus strains from raw-milk products (n = 27), bovine mastitis specimens (n = 9), and human blood cultures (n = 39). All isolates were typed by pulsed-field gel electrophoresis (PFGE). In addition, subsets of isolates were characterized using multilocus sequence typing (MLST), multiplex PCR (m-PCR) for genes encoding nine of the staphylococcal enterotoxins (SE), and the cloverleaf method for penicillin resistance. A variety of genotypes were observed, and greater genetic diversity was found among bovine than caprine bulk milk isolates. Certain genotypes, with a wide geographic distribution, were common to bovine and caprine bulk milk and may represent ruminant-specialized S. aureus. Isolates with genotypes indistinguishable from those of strains from ruminant mastitis were frequently found in bulk milk, and strains with genotypes indistinguishable from those from bulk milk were observed in raw-milk products. This indicates that S. aureus from infected udders may contaminate bulk milk and, subsequently, raw-milk products. Human blood culture isolates were diverse and differed from isolates from other sources. Genotyping by PFGE, MLST, and m-PCR for SE genes largely corresponded. In general, isolates with indistinguishable PFGE banding patterns had the same SE gene profile and isolates with identical SE gene profiles were placed together in PFGE clusters. Phylogenetic analyses agreed with the division of MLST sequence types into clonal complexes, and isolates within the same clonal complex had the same SE gene profile. Furthermore, isolates within PFGE clusters generally belonged to the same clonal complex.

Genetic Variation among Staphylococcus aureus Strains from Norwegian Bulk Milk

PubMed Central

Jørgensen, H. J.; Mørk, T.; Caugant, D. A.; Kearns, A.; Rørvik, L. M.

2005-01-01

Strains of Staphylococcus aureus obtained from bovine (n = 117) and caprine (n = 114) bulk milk were characterized and compared with S. aureus strains from raw-milk products (n = 27), bovine mastitis specimens (n = 9), and human blood cultures (n = 39). All isolates were typed by pulsed-field gel electrophoresis (PFGE). In addition, subsets of isolates were characterized using multilocus sequence typing (MLST), multiplex PCR (m-PCR) for genes encoding nine of the staphylococcal enterotoxins (SE), and the cloverleaf method for penicillin resistance. A variety of genotypes were observed, and greater genetic diversity was found among bovine than caprine bulk milk isolates. Certain genotypes, with a wide geographic distribution, were common to bovine and caprine bulk milk and may represent ruminant-specialized S. aureus. Isolates with genotypes indistinguishable from those of strains from ruminant mastitis were frequently found in bulk milk, and strains with genotypes indistinguishable from those from bulk milk were observed in raw-milk products. This indicates that S. aureus from infected udders may contaminate bulk milk and, subsequently, raw-milk products. Human blood culture isolates were diverse and differed from isolates from other sources. Genotyping by PFGE, MLST, and m-PCR for SE genes largely corresponded. In general, isolates with indistinguishable PFGE banding patterns had the same SE gene profile and isolates with identical SE gene profiles were placed together in PFGE clusters. Phylogenetic analyses agreed with the division of MLST sequence types into clonal complexes, and isolates within the same clonal complex had the same SE gene profile. Furthermore, isolates within PFGE clusters generally belonged to the same clonal complex. PMID:16332822

Natural killer cell-type body cavity lymphoma following chronic active Epstein-Barr virus infection.

PubMed

Ogata, Masao; Imamura, Tomoyuki; Mizunoe, Syunji; Ohtsuka, Eiichi; Kikuchi, Hiroshi; Nasu, Masaru

2003-06-01

We describe a 69-year-old female who developed natural killer cell-type body cavity lymphoma following chronic active Epstein-Barr virus (CAEBV) infection. Examination of the patient's pleural effusion revealed large abnormal lymphocytes, which were CD2(+), CD7(+), CD30(+), CD56(+), CD3(-), and CD4(-). No rearrangement of T cell receptor genes was detected. Clonal proliferation of Epstein-Barr virus (EBV)-infected cells in pleural effusion was demonstrated by Southern blot hybridization analysis. Human herpesvirus type-8 (HHV-8) DNA was not detected in these cells. The patient achieved a complete remission with combination chemotherapy. Prior to the clinical onset of lymphoma, high fever of unknown origin had persisted for 21 months. IgG antibodies to EBV-viral capsid antigen and to EBV-early antigens, types D and R were not high (1:160 and less than 1:10, respectively). Two months after the onset of fever, however, retrospective quantitative PCR assay revealed a high EBV DNA load in plasma, indicating that CAEBV infection had been the cause of the patient's recurrent fever. The remarkable features of this case are (i) the development of lymphoma following CAEBV infection that demonstrated a normal pattern of EBV-specific antibodies, (ii) the development of HHV-8-negative body cavity lymphoma, and (iii) the effectiveness of combination chemotherapy. Copyright 2003 Wiley-Liss, Inc.

Based Upon Repeat Pattern (BURP): an algorithm to characterize the long-term evolution of Staphylococcus aureus populations based on spa polymorphisms.

PubMed

Mellmann, Alexander; Weniger, Thomas; Berssenbrügge, Christoph; Rothgänger, Jörg; Sammeth, Michael; Stoye, Jens; Harmsen, Dag

2007-10-29

For typing of Staphylococcus aureus, DNA sequencing of the repeat region of the protein A (spa) gene is a well established discriminatory method for outbreak investigations. Recently, it was hypothesized that this region also reflects long-term epidemiology. However, no automated and objective algorithm existed to cluster different repeat regions. In this study, the Based Upon Repeat Pattern (BURP) implementation that is a heuristic variant of the newly described EDSI algorithm was investigated to infer the clonal relatedness of different spa types. For calibration of BURP parameters, 400 representative S. aureus strains with different spa types were characterized by MLST and clustered using eBURST as "gold standard" for their phylogeny. Typing concordance analysis between eBURST and BURP clustering (spa-CC) were performed using all possible BURP parameters to determine their optimal combination. BURP was subsequently evaluated with a strain collection reflecting the breadth of diversity of S. aureus (JCM 2002; 40:4544). In total, the 400 strains exhibited 122 different MLST types. eBURST grouped them into 23 clonal complexes (CC; 354 isolates) and 33 singletons (46 isolates). BURP clustering of spa types using all possible parameter combinations and subsequent comparison with eBURST CCs resulted in concordances ranging from 8.2 to 96.2%. However, 96.2% concordance was reached only if spa types shorter than 8 repeats were excluded, which resulted in 37% excluded spa types. Therefore, the optimal combination of the BURP parameters was "exclude spa types shorter than 5 repeats" and "cluster spa types into spa-CC if cost distances are less than 4" exhibiting 95.3% concordance to eBURST. This algorithm identified 24 spa-CCs, 40 singletons, and excluded only 7.8% spa types. Analyzing the natural population with these parameters, the comparison of whole-genome micro-array groupings (at the level of 0.31 Pearson correlation index) and spa-CCs gave a concordance of 87.1%; BURP spa-CCs vs. manually grouped spa types resulted in 95.7% concordance. BURP is the first automated and objective tool to infer clonal relatedness from spa repeat regions. It is able to extract an evolutionary signal rather congruent to MLST and micro-array data.

Ecological Consequences of Clonal Integration in Plants

PubMed Central

Liu, Fenghong; Liu, Jian; Dong, Ming

2016-01-01

Clonal plants are widespread throughout the plant kingdom and dominate in diverse habitats. Spatiotemporal heterogeneity of environment is pervasive at multiple scales, even at scales relevant to individual plants. Clonal integration refers to resource translocation and information communication among the ramets of clonal plants. Due to clonal integration, clonal plant species possess a series of peculiar attributes: plasticity in response to local and non-local conditions, labor division with organ specialization for acquiring locally abundant resources, foraging behavior by selective placement of ramets in resource-rich microhabitats, and avoidance of intraclonal competition. Clonal integration has very profound ecological consequences for clonal plants. It allows them to efficiently cope with environmental heterogeneity, by alleviating local resource shortages, buffering environmental stresses and disturbances, influencing competitive ability, increasing invasiveness, and altering species composition and invasibility at the community level. In this paper, we present a comprehensive review of research on the ecological consequences of plant clonal integration based on a large body of literature. We also attempt to propose perspectives for future research. PMID:27446093

Sex-specific gene expression during asexual development of Neurospora crassa.

PubMed

Wang, Zheng; Kin, Koryu; López-Giráldez, Francesc; Johannesson, Hanna; Townsend, Jeffrey P

2012-07-01

The impact of loci that determine sexual identity upon the asexual, dominant stage of fungal life history has been well studied. To investigate their impact, expression differences between strains of different mating type during asexual development were assayed, with RNA sampled from otherwise largely isogenic mat A and mat a strains of Neurospora crassa at early, middle, and late clonal stages of development. We observed significant differences in overall gene expression between mating types across clonal development, especially at late development stages. The expression levels of mating-type genes and pheromone genes were assayed by reverse transcription and quantitative PCR, revealing expression of pheromone and receptor genes in strains of both mating types in all development stages, and revealing that mating type (mat) genes were increasingly expressed over the course of asexual development. Interestingly, among differentially expressed genes, the mat A genotype more frequently exhibited a higher expression level than mat a, and demonstrated greater transcriptional regulatory dynamism. Significant up-regulation of expression was observed for many late light-responsive genes at late asexual development stages. Further investigation of the impact of light and the roles of light response genes in asexual development of both mating types are warranted. Copyright © 2012 Elsevier Inc. All rights reserved.

Characterization of levofloxacin non-susceptible clinical Streptococcus pyogenes isolated in the central part of Italy.

PubMed

Petrelli, D; Di Luca, M C; Prenna, M; Bernaschi, P; Repetto, A; Vitali, L A

2014-02-01

We investigated the prevalence, genetics, and clonality of fluoroquinolone non-susceptible isolates of Streptococcus pyogenes in the central part of Italy. S. pyogenes strains (n = 197) were isolated during 2012 from patients with tonsillopharyngitis, skin, wound or invasive infections and screened for fluoroquinolone non-susceptibility (resistance to norfloxacin and levofloxacin minimum inhibitory concentration (MIC) = 2 mg/L) following EUCAST guidelines. First-step topoisomerase parC and gyrA substitutions were investigated using sequencing analysis. Clonality was determined by pulsed field gel electrophoresis (PFGE; SmaI digestion) and by emm typing. The fluoroquinolone non-susceptible phenotype was identified in 18 isolates (9.1 %) and correlated with mutations in parC, but not in gyrA, the most frequent leading to substitution of the serine at position 79 with an alanine. Most of the fluoroquinolone non-susceptible isolates belonged to the emm-type 6, even if other emm-types were also represented (emm75, emm89, and emm2). A significant level of association was measured between PFGE and both emm type and substitutions in parC. The prevalence of fluoroquinolone non-susceptible Streptococcus pyogenes isolates in Italy is of concern and, although the well-known emm type 6 is dominant, other types are appearing and spreading.

Detecting local establishment strategies of wild cherry (Prunus avium L.).

PubMed

Höltken, Aki M; Gregorius, Hans-Rolf

2006-10-04

P. avium, a pioneer tree species that colonizes early forest successional stages, is assumed to require an effective strategy allowing stably repeatable rounds of local establishment, dispersal and local extinction. Consequently, the early replacement of cherry by climax tree species makes the establishment of several local generations very unlikely, especially in central European continuous cover forests. This has to be seen in connection with the mixed reproduction system involving asexual reproduction as a complementary adaptational strategy. Tests of the local establishment of wild cherry must therefore consider the possibility of first generation establishment via seedling recruitment potentially followed by an asexual generation (root suckering). Successful establishment can therefore be determined only among adult individuals with the option of detecting vegetative reproduction at these stages. To test the implied suggestion about local establishment strategies of wild cherry, nuclear microsatellites were used to analyse patterns of asexual propagation among adult stages that have been subjected to one of two major types of forest management. These management types, the historical "coppice with standards system" (CWS) and the "high forest system" (HFS), can be reasonably assumed to have affected the reproduction system of P. avium. Clear differences were found in the reproduction pattern between two stands representing the two forest management types: 1) Clonal propagation is observed in both management systems, but with a distinctly higher frequency in the CWS. Hence, sexual recruitment as a first local generation is followed by a second asexual generation in both, whereas in the CWS there is evidence for an additional clonal generation. 2) The estimation of amounts of clonal reproduction critically depends on the assumptions about multilocus gene associations. This is revealed by the application of newly developed methods of quantifying gene associations. 3) Haplotype diversities are higher in the CWS and found to be associated with a large degree of heterozygosity for the second largest clonal group. 4) Seed set was sparse over the last eight years of observation in the CWS stand. This study provides useful guidelines for more comprehensive investigations, particularly on the interrelationships between degrees of cloning and capacity of sexual reproduction, amounts of multilocus gene associations, effects of heterozygosity on cloning success, and sustainability of different forest management types.

Whole-genome sequencing provides new insights into the clonal architecture of Barrett’s esophagus and esophageal adenocarcinoma

PubMed Central

Warren, Andrew; Cheetham, R. Keira; Northen, Helen; O’Donovan, Maria; Malhotra, Shalini; di Pietro, Massimiliano; Ivakhno, Sergii; He, Miao; Weaver, Jamie M.J.; Lynch, Andy G.; Kingsbury, Zoya; Ross, Mark; Humphray, Sean; Bentley, David; Fitzgerald, Rebecca C.

2015-01-01

The molecular genetic relationship between esophageal adenocarcinoma (EAC) and its precursor lesion, Barrett’s esophagus, is poorly understood. Using whole-genome sequencing on 23 paired Barrett’s esophagus and EAC samples, together with one in-depth Barrett’s esophagus case-study sampled over time and space, we have provided new insights on the following aspects: i) Barrett’s esophagus is polyclonal and highly mutated even in the absence of dysplasia; ii) when cancer develops, copy number increases and heterogeneity persists such that the spectrum of mutations often shows surprisingly little overlap between EAC and adjacent Barrett’s esophagus; and iii) despite differences in specific coding mutations the mutational context suggests a common causative insult underlying these two conditions. From a clinical perspective, the histopathological assessment of dysplasia appears to be a poor reflection of the molecular disarray within the Barrett’s epithelium and a molecular Cytosponge™ technique overcomes sampling bias and has capacity to reflect the entire clonal architecture. PMID:26192915

Clonal evolution following chemotherapy-induced stem cell depletion in cats heterozygous for glucose-6-phosphate dehydrogenase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Abkowitz, J.L.; Ott, R.M.; Holly, R.D.

The number of hematopoietic stem cells necessary to support normal hematopoiesis is not known but may be small. If so, the depletion or damage of such cells could result in apparent clonal dominance. To test this hypothesis, dimethylbusulfan (2 to 4 mg/kg intravenously (IV) x 3) was given to cats heterozygous for the X-linked enzyme glucose-6-phosphate dehydrogenase (G-6-PD). These cats were the daughters of domestic X Geoffroy parents. After the initial drug-induced cytopenias (2 to 4 weeks), peripheral blood counts and the numbers of marrow progenitors detected in culture remained normal, although the percentages of erythroid burst-forming cells (BFU-E) andmore » granulocyte/macrophage colony-forming cells (CFU-GM) in DNA synthesis increased, as determined by the tritiated thymidine suicide technique. In three of six cats treated, a dominance of Geoffroy-type G-6-PD emerged among the progenitor cells, granulocytes, and RBCs. These skewed ratios of domestic to Geoffroy-type G-6-PD have persisted greater than 3 years. No changes in cell cycle kinetics or G-6-PD phenotypes were noted in similar studies in six control cats. These data suggest that clonal evolution may reflect the depletion or damage of normal stem cells and not only the preferential growth and dominance of neoplastic cells.« less

Recombination-Driven Genome Evolution and Stability of Bacterial Species.

PubMed

Dixit, Purushottam D; Pang, Tin Yau; Maslov, Sergei

2017-09-01

While bacteria divide clonally, horizontal gene transfer followed by homologous recombination is now recognized as an important contributor to their evolution. However, the details of how the competition between clonality and recombination shapes genome diversity remains poorly understood. Using a computational model, we find two principal regimes in bacterial evolution and identify two composite parameters that dictate the evolutionary fate of bacterial species. In the divergent regime, characterized by either a low recombination frequency or strict barriers to recombination, cohesion due to recombination is not sufficient to overcome the mutational drift. As a consequence, the divergence between pairs of genomes in the population steadily increases in the course of their evolution. The species lacks genetic coherence with sexually isolated clonal subpopulations continuously formed and dissolved. In contrast, in the metastable regime, characterized by a high recombination frequency combined with low barriers to recombination, genomes continuously recombine with the rest of the population. The population remains genetically cohesive and temporally stable. Notably, the transition between these two regimes can be affected by relatively small changes in evolutionary parameters. Using the Multi Locus Sequence Typing (MLST) data, we classify a number of bacterial species to be either the divergent or the metastable type. Generalizations of our framework to include selection, ecologically structured populations, and horizontal gene transfer of nonhomologous regions are discussed as well. Copyright © 2017 by the Genetics Society of America.

Pandemic extra-intestinal pathogenic Escherichia coli (ExPEC) clonal group O6-B2-ST73 as a cause of avian colibacillosis in Brazil.

PubMed

Cunha, Marcos Paulo Vieira; Saidenberg, Andre Becker; Moreno, Andrea Micke; Ferreira, Antonio José Piantino; Vieira, Mônica Aparecida Midolli; Gomes, Tânia Aparecida Tardelli; Knöbl, Terezinha

2017-01-01

Extra-intestinal pathogenic Escherichia coli (ExPEC) represent an emerging pathogen, with pandemic strains increasingly involved in cases of urinary tract infections (UTIs), bacteremia, and meningitis. In addition to affecting humans, the avian pathotype of ExPEC, avian pathogenic E. coli (APEC), causes severe economic losses to the poultry industry. Several studies have revealed overlapping characteristics between APEC and human ExPEC, leading to the hypothesis of a zoonotic potential of poultry strains. However, the description of certain important pandemic clones, such as Sequence Type 73 (ST73), has not been reported in food sources. We characterized 27 temporally matched APEC strains from diverse poultry farms in Brazil belonging to the O6 serogroup because this serogroup is frequently described as a causal factor in UTI and septicemia in humans in Brazil and worldwide. The isolates were genotypically characterized by identifying ExPEC virulence factors, phylogenetically tested by phylogrouping and multilocus sequence type (MLST) analysis, and compared to determine their similarity employing the pulsed field gel electrophoresis (PFGE) technique. The strains harbored a large number of virulence determinants that are commonly described in uropathogenic E. coli (UPEC) and sepsis associated E. coli (SEPEC) strains and, to a lesser extent in neonatal meningitis associated E. coli (NMEC), such as pap (85%), sfa (100%), usp (100%), cnf1 (22%), kpsMTII (66%), hlyA (52%), and ibeA (4%). These isolates also yielded a low prevalence of some genes that are frequently described in APEC, such as iss (37%), tsh, ompT, and hlyF (8% each), and cvi/cva (0%). All strains were classified as part of the B2 phylogroup and sequence type 73 (ST73), with a cluster of 25 strains showing a clonal profile by PFGE. These results further suggest the zoonotic potential of some APEC clonal lineages and their possible role in the epidemiology of human ExPEC, in addition to providing the first description of the O6-B2-ST73 clonal group in poultry.

Enteroaggregative Escherichia coli O78:H10, the Cause of an Outbreak of Urinary Tract Infection

PubMed Central

Scheutz, Flemming; Andersen, Rebecca L.; Menard, Megan; Boisen, Nadia; Johnston, Brian; Hansen, Dennis S.; Krogfelt, Karen A.; Nataro, James P.; Johnson, James R.

2012-01-01

In 1991, multiresistant Escherichia coli O78:H10 strains caused an outbreak of urinary tract infections in Copenhagen, Denmark. The phylogenetic origin, clonal background, and virulence characteristics of the outbreak isolates, and their relationship to nonoutbreak O78:H10 strains according to these traits and resistance profiles, are unknown. Accordingly, we extensively characterized 51 archived E. coli O78:H10 isolates (48 human isolates from seven countries, including 19 Copenhagen outbreak isolates, and 1 each of calf, avian, and unknown-source isolates), collected from 1956 through 2000. E. coli O78:H10 was clonally heterogeneous, comprising one dominant clonal group (61% of isolates, including all 19 outbreak isolates) from ST10 (phylogenetic group A) plus several minor clonal groups (phylogenetic groups A and D). All ST10 isolates, versus 25% of non-ST10 isolates, were identified by molecular methods as enteroaggregative E. coli (EAEC) (P < 0.001). Genes present in >90% of outbreak isolates included fimH (type 1 fimbriae; ubiquitous in E. coli); fyuA, traT, and iutA (associated with extraintestinal pathogenic E. coli [ExPEC]); and sat, pic, aatA, aggR, aggA, ORF61, aaiC, aap, and ORF3 (associated with EAEC). An outbreak isolate was lethal in a murine subcutaneous sepsis model and exhibited characteristic EAEC “stacked brick” adherence to cultured epithelial cells. Thus, the 1991 Copenhagen outbreak was caused by a tight, non-animal-associated subset within a broadly disseminated O78:H10 clonal group (ST10; phylogenetic group A), members of which exhibit both ExPEC and EAEC characteristics, whereas O78:H10 isolates overall are phylogenetically diverse. Whether ST10 O78:H10 EAEC strains are both uropathogenic and diarrheagenic warrants further investigation. PMID:22972830

Enteroaggregative Escherichia coli O78:H10, the cause of an outbreak of urinary tract infection.

PubMed

Olesen, Bente; Scheutz, Flemming; Andersen, Rebecca L; Menard, Megan; Boisen, Nadia; Johnston, Brian; Hansen, Dennis S; Krogfelt, Karen A; Nataro, James P; Johnson, James R

2012-11-01

In 1991, multiresistant Escherichia coli O78:H10 strains caused an outbreak of urinary tract infections in Copenhagen, Denmark. The phylogenetic origin, clonal background, and virulence characteristics of the outbreak isolates, and their relationship to nonoutbreak O78:H10 strains according to these traits and resistance profiles, are unknown. Accordingly, we extensively characterized 51 archived E. coli O78:H10 isolates (48 human isolates from seven countries, including 19 Copenhagen outbreak isolates, and 1 each of calf, avian, and unknown-source isolates), collected from 1956 through 2000. E. coli O78:H10 was clonally heterogeneous, comprising one dominant clonal group (61% of isolates, including all 19 outbreak isolates) from ST10 (phylogenetic group A) plus several minor clonal groups (phylogenetic groups A and D). All ST10 isolates, versus 25% of non-ST10 isolates, were identified by molecular methods as enteroaggregative E. coli (EAEC) (P < 0.001). Genes present in >90% of outbreak isolates included fimH (type 1 fimbriae; ubiquitous in E. coli); fyuA, traT, and iutA (associated with extraintestinal pathogenic E. coli [ExPEC]); and sat, pic, aatA, aggR, aggA, ORF61, aaiC, aap, and ORF3 (associated with EAEC). An outbreak isolate was lethal in a murine subcutaneous sepsis model and exhibited characteristic EAEC "stacked brick" adherence to cultured epithelial cells. Thus, the 1991 Copenhagen outbreak was caused by a tight, non-animal-associated subset within a broadly disseminated O78:H10 clonal group (ST10; phylogenetic group A), members of which exhibit both ExPEC and EAEC characteristics, whereas O78:H10 isolates overall are phylogenetically diverse. Whether ST10 O78:H10 EAEC strains are both uropathogenic and diarrheagenic warrants further investigation.

Processing-Dependent and Clonal Contamination Patterns of Listeria monocytogenes in the Cured Ham Food Chain Revealed by Genetic Analysis

PubMed Central

Morganti, Marina; Scaltriti, Erika; Cozzolino, Paolo; Bolzoni, Luca; Casadei, Gabriele; Pierantoni, Marco; Foni, Emanuela

2015-01-01

The quantitative and qualitative patterns of environmental contamination by Listeria monocytogenes were investigated in the production chain of dry-cured Parma ham. Standard arrays of surfaces were sampled in processing facilities during a single visit per plant in the three compartments of the food chain, i.e., ham production (19 plants) and postproduction, which was divided into deboning (43 plants) and slicing (25 plants) steps. The numbers of sampled surfaces were 384 in ham production, with 25 positive for L. monocytogenes, and 1,084 in postproduction, with 83 positives. Statistical analysis of the prevalence of contaminated surfaces showed that in ham production, contamination was higher at the beginning of processing and declined significantly toward the end, while in postproduction, prevalence rose toward the end of processing. Prevalence was higher in the deboning facilities than in slicing facilities and was dependent on the type of surface (floor/drainage > clothing > equipment). The qualitative pattern of contamination was investigated through an analysis of the survey isolates and a set of isolates derived from routine monitoring, including longitudinal isolations. Pulsed-field gel electrophoresis (PFGE) and whole-genome single-nucleotide polymorphism (SNP) analysis revealed a remarkable clonality of L. monocytogenes within plants, with the detection of 16 plant-specific clones out of 17 establishments with multiple isolates. Repeated detections of clonal isolates >6 months apart were also observed. Six was the maximum number of between-isolate differences in core SNPs observed within these clones. Based on the same six-SNP threshold, three clusters of clonal isolates, shared by six establishments, were also identified. The spread of L. monocytogenes within and between plants, as indicated by its clonal behavior, is a matter of concern for the hygienic management of establishments. PMID:26590278

Novel synthetic monoketone transmute radiation-triggered NFκB-dependent TNFα cross-signaling feedback maintained NFκB and favors neuroblastoma regression.

PubMed

Aravindan, Sheeja; Natarajan, Mohan; Awasthi, Vibhudutta; Herman, Terence S; Aravindan, Natarajan

2013-01-01

Recently, we demonstrated that radiation (IR) instigates the occurrence of a NFκB-TNFα feedback cycle which sustains persistent NFκB activation in neuroblastoma (NB) cells and favors survival advantage and clonal expansion. Further, we reported that curcumin targets IR-induced survival signaling and NFκB dependent hTERT mediated clonal expansion in human NB cells. Herein, we investigated the efficacy of a novel synthetic monoketone, EF24, a curcumin analog in inhibiting persistent NFκB activation by disrupting the IR-induced NFκB-TNFα-NFκB feedback signaling in NB and subsequent mitigation of survival advantage and clonal expansion. EF24 profoundly suppressed the IR-induced NFκB-DNA binding activity/promoter activation and, maintained the NFκB repression by deterring NFκB-dependent TNFα transactivation/intercellular secretion in genetically varied human NB (SH-SY5Y, IMR-32, SK-PN-DW, MC-IXC and SK-N-MC) cell types. Further, EF24 completely suppressed IR-induced NFκB-TNFα cross-signaling dependent transactivation/translation of pro-survival IAP1, IAP2 and Survivin and subsequent cell survival. In corroboration, EF24 treatment maximally blocked IR-induced NFκB dependent hTERT transactivation/promoter activation, telomerase activation and consequent clonal expansion. EF24 displayed significant regulation of IR-induced feedback dependent NFκB and NFκB mediated survival signaling and complete regression of NB xenograft. Together, the results demonstrate for the first time that, novel synthetic monoketone EF24 potentiates radiotherapy and mitigates NB progression by selectively targeting IR-triggered NFκB-dependent TNFα-NFκB cross-signaling maintained NFκB mediated survival advantage and clonal expansion.

Novel Synthetic Monoketone Transmute Radiation-Triggered NFκB-Dependent TNFα Cross-Signaling Feedback Maintained NFκB and Favors Neuroblastoma Regression

PubMed Central

Aravindan, Sheeja; Natarajan, Mohan; Awasthi, Vibhudutta; Herman, Terence S.; Aravindan, Natarajan

2013-01-01

Recently, we demonstrated that radiation (IR) instigates the occurrence of a NFκB-TNFα feedback cycle which sustains persistent NFκB activation in neuroblastoma (NB) cells and favors survival advantage and clonal expansion. Further, we reported that curcumin targets IR-induced survival signaling and NFκB dependent hTERT mediated clonal expansion in human NB cells. Herein, we investigated the efficacy of a novel synthetic monoketone, EF24, a curcumin analog in inhibiting persistent NFκB activation by disrupting the IR-induced NFκB-TNFα-NFκB feedback signaling in NB and subsequent mitigation of survival advantage and clonal expansion. EF24 profoundly suppressed the IR-induced NFκB-DNA binding activity/promoter activation and, maintained the NFκB repression by deterring NFκB-dependent TNFα transactivation/intercellular secretion in genetically varied human NB (SH-SY5Y, IMR-32, SK–PN–DW, MC-IXC and SK–N-MC) cell types. Further, EF24 completely suppressed IR-induced NFκB-TNFα cross-signaling dependent transactivation/translation of pro-survival IAP1, IAP2 and Survivin and subsequent cell survival. In corroboration, EF24 treatment maximally blocked IR-induced NFκB dependent hTERT transactivation/promoter activation, telomerase activation and consequent clonal expansion. EF24 displayed significant regulation of IR-induced feedback dependent NFκB and NFκB mediated survival signaling and complete regression of NB xenograft. Together, the results demonstrate for the first time that, novel synthetic monoketone EF24 potentiates radiotherapy and mitigates NB progression by selectively targeting IR-triggered NFκB-dependent TNFα-NFκB cross-signaling maintained NFκB mediated survival advantage and clonal expansion. PMID:23967300

Invasive clonal plant species have a greater root-foraging plasticity than non-invasive ones.

PubMed

Keser, Lidewij H; Dawson, Wayne; Song, Yao-Bin; Yu, Fei-Hai; Fischer, Markus; Dong, Ming; van Kleunen, Mark

2014-03-01

Clonality is frequently positively correlated with plant invasiveness, but which aspects of clonality make some clonal species more invasive than others is not known. Due to their spreading growth form, clonal plants are likely to experience spatial heterogeneity in nutrient availability. Plasticity in allocation of biomass to clonal growth organs and roots may allow these plants to forage for high-nutrient patches. We investigated whether this foraging response is stronger in species that have become invasive than in species that have not. We used six confamilial pairs of native European clonal plant species differing in invasion success in the USA. We grew all species in large pots under homogeneous or heterogeneous nutrient conditions in a greenhouse, and compared their nutrient-foraging response and performance. Neither invasive nor non-invasive species showed significant foraging responses to heterogeneity in clonal growth organ biomass or in aboveground biomass of clonal offspring. Invasive species had, however, a greater positive foraging response in terms of root and belowground biomass than non-invasive species. Invasive species also produced more total biomass. Our results suggest that the ability for strong root foraging is among the characteristics promoting invasiveness in clonal plants.

Diversification and Distribution of Ruminant Chlamydia abortus Clones Assessed by MLST and MLVA.

PubMed

Siarkou, Victoria I; Vorimore, Fabien; Vicari, Nadia; Magnino, Simone; Rodolakis, Annie; Pannekoek, Yvonne; Sachse, Konrad; Longbottom, David; Laroucau, Karine

2015-01-01

Chlamydia abortus, an obligate intracellular bacterium, is the most common infectious cause of abortion in small ruminants worldwide and has zoonotic potential. We applied multilocus sequence typing (MLST) together with multiple-locus variable-number tandem repeat analysis (MLVA) to genotype 94 ruminant C. abortus strains, field isolates and samples collected from 1950 to 2011 in diverse geographic locations, with the aim of delineating C. abortus lineages and clones. MLST revealed the previously identified sequence types (STs) ST19, ST25, ST29 and ST30, plus ST86, a recently-assigned type on the Chlamydiales MLST website and ST87, a novel type harbouring the hemN_21 allele, whereas MLVA recognized seven types (MT1 to MT7). Minimum-spanning-tree analysis suggested that all STs but one (ST30) belonged to a single clonal complex, possibly reflecting the short evolutionary timescale over which the predicted ancestor (ST19) has diversified into three single-locus variants (ST86, ST87 and ST29) and further, through ST86 diversification, into one double-locus variant (ST25). ST descendants have probably arisen through a point mutation evolution mode. Interestingly, MLVA showed that in the ST19 population there was a greater genetic diversity than in other STs, most of which exhibited the same MT over time and geographical distribution. However, the evolutionary pathways of C. abortus STs seem to be diverse across geographic distances with individual STs restricted to particular geographic locations. The ST30 singleton clone displaying geographic specificity and represented by the Greek strains LLG and POS was effectively distinguished from the clonal complex lineage, supporting the notion that possibly two separate host adaptations and hence independent bottlenecks of C. abortus have occurred through time. The combination of MLST and MLVA assays provides an additional level of C. abortus discrimination and may prove useful for the investigation and surveillance of emergent C. abortus clonal populations.

Effect of clonal integration on nitrogen cycling in rhizosphere of rhizomatous clonal plant, Phyllostachys bissetii, under heterogeneous light.

PubMed

Li, Yang; Chen, Jing-Song; Xue, Ge; Peng, Yuanying; Song, Hui-Xing

2018-07-01

Clonal integration plays an important role in clonal plant adapting to heterogeneous habitats. It was postulated that clonal integration could exhibit positive effects on nitrogen cycling in the rhizosphere of clonal plant subjected to heterogeneous light conditions. An in-situ experiment was conducted using clonal fragments of Phyllostachys bissetii with two successive ramets. Shading treatments were applied to offspring or mother ramets, respectively, whereas counterparts were treated to full sunlight. Rhizomes between two successive ramets were either severed or connected. Extracellular enzyme activities and nitrogen turnover were measured, as well as soil properties. Abundance of functional genes (archaeal or bacterial amoA, nifH) in the rhizosphere of shaded, offspring or mother ramets were determined using quantitative polymerase chain reaction. Carbon or nitrogen availabilities were significantly influenced by clonal integration in the rhizosphere of shaded ramets. Clonal integration significantly increased extracellular enzyme activities and abundance of functional genes in the rhizosphere of shaded ramets. When rhizomes were connected, higher nitrogen turnover (nitrogen mineralization or nitrification rates) was exhibited in the rhizosphere of shaded offspring ramets. However, nitrogen turnover was significantly decreased by clonal integration in the rhizosphere of shaded mother ramets. Path analysis indicated that nitrogen turnover in the rhizosphere of shaded, offspring or mother ramets were primarily driven by the response of soil microorganisms to dissolved organic carbon or nitrogen. This unique in-situ experiment provided insights into the mechanism of nutrient recycling mediated by clonal integration. It was suggested that effects of clonal integration on the rhizosphere microbial processes were dependent on direction of photosynthates transport in clonal plant subjected to heterogeneous light conditions. Copyright © 2018 Elsevier B.V. All rights reserved.

Disturbance Is an Important Driver of Clonal Richness in Tropical Seagrasses

PubMed Central

McMahon, Kathryn M.; Evans, Richard D.; van Dijk, Kor-jent; Hernawan, Udhi; Kendrick, Gary A.; Lavery, Paul S.; Lowe, Ryan; Puotinen, Marji; Waycott, Michelle

2017-01-01

Clonality is common in many aquatic plant species, including seagrasses, where populations are maintained through a combination of asexual and sexual reproduction. One common measure used to describe the clonal structure of populations is clonal richness. Clonal richness is strongly dependent on the biological characteristics of the species, and how these interact with the environment but can also reflect evolutionary scale processes especially at the edge of species ranges. However, little is known about the spatial patterns and drivers of clonal richness in tropical seagrasses. This study assessed the spatial patterns of clonal richness in meadows of three tropical seagrass species, Thalassia hemprichii, Halodule uninervis, and Halophila ovalis, spanning a range of life-history strategies and spatial scales (2.5–4,711 km) in Indonesia and NW Australia. We further investigated the drivers of clonal richness using general additive mixed models for two of the species, H. uninervis and H. ovalis, over 8° latitude. No significant patterns were observed in clonal richness with latitude, yet disturbance combined with sea surface temperature strongly predicted spatial patterns of clonal richness. Sites with a high probability of cyclone disturbance had low clonal richness, whereas an intermediate probability of cyclone disturbance and the presence of dugong grazing combined with higher sea surface temperatures resulted in higher levels of clonal richness. We propose potential mechanisms for these patterns related to the recruitment and mortality rates of individuals as well as reproductive effort. Under a changing climate, increased severity of tropical cyclones and the decline in populations of mega-grazers have the potential to reduce clonal richness leading to less genetically diverse populations. PMID:29259609

Change in genotype of methicillin-resistant Staphylococcus aureus (MRSA) affects the antibiogram of hospital-acquired MRSA.

PubMed

Harada, Dai; Nakaminami, Hidemasa; Miyajima, Eri; Sugiyama, Taku; Sasai, Nao; Kitamura, Yoshinobu; Tamura, Taku; Kawakubo, Takashi; Noguchi, Norihisa

2018-07-01

Recently, the dissemination of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) into hospitals has frequently been reported worldwide. Hospital-acquired MRSA (HA-MRSA) strains exhibit high-level resistance to multiple antimicrobial agents, whereas CA-MRSA strains are usually susceptible to non-β-lactams. Thus, it is predicted that the antibiogram of the HA-MRSA population would change along with the change in genotype of MRSA. Here, we investigated the changes in the MRSA population along with the MRSA antibiogram in a hospital between 2010 and 2016. Staphylococcal cassette chromosome (SCC) mec typing showed that the predominant HA-MRSA strains in the hospital dramatically changed from SCCmec type II, which is the major type of HA-MRSA, to SCCmec type IV, which is the major type of CA-MRSA. Multilocus sequence typing revealed that the predominant SCCmec type IV strain was a clonal complex (CC) 8 clone, which is mainly found among CA-MRSA. Furthermore, the CC1-SCCmec type IV (CC1-IV) clone significantly increased. Both the CC8-IV and CC1-IV clones exhibited high antimicrobial susceptibility. The antibiogram change of the HA-MRSA population was consistent with the antimicrobial susceptibilities and increased prevalence of the CC8-IV and CC1-IV clones. Our data reveal that the change in the genotypes of MRSA strains could impact the antibiogram of HA-MRSA population. Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Multilocus sequence typing (MLST) analysis of Propionibacterium acnes isolates from radical prostatectomy specimens.

PubMed

Mak, Tim N; Yu, Shu-Han; De Marzo, Angelo M; Brüggemann, Holger; Sfanos, Karen S

2013-05-01

Inflammation is commonly observed in radical prostatectomy specimens, and evidence suggests that inflammation may contribute to prostate carcinogenesis. Multiple microorganisms have been implicated in serving as a stimulus for prostatic inflammation. The pro-inflammatory anaerobe, Propionibacterium acnes, is ubiquitously found on human skin and is associated with the skin disease acne vulgaris. Recent studies have shown that P. acnes can be detected in prostatectomy specimens by bacterial culture or by culture-independent molecular techniques. Radical prostatectomy tissue samples were obtained from 30 prostate cancer patients and subject to both aerobic and anaerobic culture. Cultured species were identified by 16S rDNA gene sequencing. Propionibacterium acnes isolates were typed using multilocus sequence typing (MLST). Our study confirmed that P. acnes can be readily cultured from prostatectomy tissues (7 of 30 cases, 23%). In some cases, multiple isolates of P. acnes were cultured as well as other Propionibacterium species, such as P. granulosum and P. avidum. Overall, 9 of 30 cases (30%) were positive for Propionibacterium spp. MLST analyses identified eight different sequence types (STs) among prostate-derived P. acnes isolates. These STs belong to two clonal complexes, namely CC36 (type I-2) and CC53/60 (type II), or are CC53/60-related singletons. MLST typing results indicated that prostate-derived P. acnes isolates do not fall within the typical skin/acne STs, but rather are characteristic of STs associated with opportunistic infections and/or urethral flora. The MLST typing results argue against the likelihood that prostatectomy-derived P. acnes isolates represent contamination from skin flora. Copyright © 2012 Wiley Periodicals, Inc.

3′UTR-truncated Hmga2 cDNA causes MPN-like hematopoiesis by conferring a clonal growth advantage at the level of HSC in mice

PubMed Central

Ikeda, Kazuhiko; Mason, Philip J.

2011-01-01

Overexpression of high mobility group AT-hook 2 (HMGA2) is found in a number of benign and malignant tumors, including the clonal PIGA− cells in 2 cases of paroxysmal nocturnal hemoglobinuria (PNH) and some myeloproliferative neoplasms (MPNs), and recently in hematopoietic cell clones resulting from gene therapy procedures. In nearly all these cases overexpression is because of deletions or translocations that remove the 3′ untranslated region (UTR) which contains binding sites for the regulatory micro RNA let-7. We were therefore interested in the effect of HMGA2 overexpression in hematopoietic tissues in transgenic mice (ΔHmga2 mice) carrying a 3′UTR-truncated Hmga2 cDNA. ΔHmga2 mice expressed increased levels of HMGA2 protein in various tissues including hematopoietic cells and showed proliferative hematopoiesis with increased numbers in all lineages of peripheral blood cells, hypercellular bone marrow (BM), splenomegaly with extramedullary erythropoiesis and erythropoietin-independent erythroid colony formation. ΔHmga2-derived BM cells had a growth advantage over wild-type cells in competitive repopulation and serial transplantation experiments. Thus overexpression of HMGA2 leads to proliferative hematopoiesis with clonal expansion at the stem cell and progenitor levels and may account for the clonal expansion in PNH and MPNs and in gene therapy patients after vector insertion disrupts the HMGA2 locus. PMID:21460244

Whole-Genome Analysis Illustrates Global Clonal Population Structure of the Ubiquitous Dermatophyte Pathogen Trichophyton rubrum

PubMed Central

Persinoti, Gabriela F.; Martinez, Diego A.; Li, Wenjun; Döğen, Aylin; Billmyre, R. Blake; Averette, Anna; Goldberg, Jonathan M.; Shea, Terrance; Young, Sarah; Zeng, Qiandong; Oliver, Brian G.; Barton, Richard; Metin, Banu; Hilmioğlu-Polat, Süleyha; Ilkit, Macit; Gräser, Yvonne; Martinez-Rossi, Nilce M.; White, Theodore C.; Heitman, Joseph; Cuomo, Christina A.

2018-01-01

Dermatophytes include fungal species that infect humans, as well as those that also infect other animals or only grow in the environment. The dermatophyte species Trichophyton rubrum is a frequent cause of skin infection in immunocompetent individuals. While members of the T. rubrum species complex have been further categorized based on various morphologies, their population structure and ability to undergo sexual reproduction are not well understood. In this study, we analyze a large set of T. rubrum and T. interdigitale isolates to examine mating types, evidence of mating, and genetic variation. We find that nearly all isolates of T. rubrum are of a single mating type, and that incubation with T. rubrum “morphotype” megninii isolates of the other mating type failed to induce sexual development. While the region around the mating type locus is characterized by a higher frequency of SNPs compared to other genomic regions, we find that the population is remarkably clonal, with highly conserved gene content, low levels of variation, and little evidence of recombination. These results support a model of recent transition to asexual growth when this species specialized to growth on human hosts. PMID:29467168

Fatal toxoplasmosis in an immunosuppressed domestic cat from Brazil caused by Toxoplasma gondii clonal type I.

PubMed

Pena, Hilda Fátima de Jesus; Evangelista, Camila Mariellen; Casagrande, Renata Assis; Biezus, Giovana; Wisser, Claudia Salete; Ferian, Paulo Eduardo; Moura, Anderson Barbosa de; Rolim, Veronica Machado; Driemeier, David; Oliveira, Solange; Alves, Bruna Farias; Gennari, Solange Maria; Traverso, Sandra Davi

2017-01-01

The objective of the study was to report on a fatal case of feline toxoplasmosis with coinfection with the feline leukemia virus (FeLV). A domestic cat (Felis silvestris catus) presented intense dyspnea and died three days later. In the necropsy, the lungs were firm, without collapse and with many white areas; moderate lymphadenomegaly and splenomegaly were also observed. The histopathological examination showed severe necrotic interstitial bronchopneumonia and mild necrotic hepatitis, associated with intralesional cysts and tachyzoites of Toxoplasma gondii that were positive by anti-T. gondii immunohistochemical (IHC) evaluation. The bone marrow showed chronic myeloid leukemia and the neoplastic cells were positive by anti-FeLV IHC evaluation. DNA extracted from lungs was positive for T. gondii by PCR targeting REP-529. T. gondii was characterized by PCR-RFLP and by the microsatellites technique. ToxoDB-PCR-RFLP #10, i.e. the archetypal type I, was identified. Microsatellite analysis showed that the strain was a variant of type I with two atypical alleles. This was the first time that a T. gondii clonal type I genotype was correlated with a case of acute toxoplasmosis in a host in Brazil.

Clonal Type I Interferon–producing and Dendritic Cell Precursors Are Contained in Both Human Lymphoid and Myeloid Progenitor Populations

PubMed Central

Chicha, Laurie; Jarrossay, David; Manz, Markus G.

2004-01-01

Because of different cytokine responsiveness, surface receptor, and transcription factor expression, human CD11c− natural type I interferon–producing cells (IPCs) and CD11c+ dendritic cells were thought to derive through lymphoid and myeloid hematopoietic developmental pathways, respectively. To directly test this hypothesis, we used an in vitro assay allowing simultaneous IPC, dendritic cell, and B cell development and we tested lymphoid and myeloid committed hematopoietic progenitor cells for their developmental capacity. Lymphoid and common myeloid and granulocyte/macrophage progenitors were capable of developing into both functional IPCs, expressing gene transcripts thought to be associated with lymphoid lineage development, and into dendritic cells. However, clonal progenitors for both populations were about fivefold more frequent within myeloid committed progenitor cells. Thus, in humans as in mice, natural IPC and dendritic cell development robustly segregates with myeloid differentiation. This would fit with natural interferon type I–producing cell and dendritic cell activity in innate immunity, the evolutionary older arm of the cellular immune system. PMID:15557348

Analysis of clonality and HPV infection in benign, hyperplastic, premalignant, and malignant lesions of the vulvar mucosa.

PubMed

Ueda, Yutaka; Enomoto, Takayuki; Miyatake, Takashi; Shroyer, Kenneth R; Yoshizaki, Tatsuo; Kanao, Hiroyuki; Ueno, Yuko; Sun, Hongbo; Nakashima, Ryuichi; Yoshino, Kiyoshi; Kimura, Toshihiro; Haba, Tomoko; Wakasa, Kenichi; Murata, Yuji

2004-08-01

To elucidate the pathogenesis of vulvar carcinomas, we studied clonality and human papillomavirus (HPV) infection in vulvar epithelial diseases. Monoclonal composition was demonstrated in all 9 invasive tumors (squamous cell carcinoma [SCC], 6; basal cell carcinoma, 1; malignant melanoma, 2), 15 of 20 cases of vulvar intraepithelial neoplasia (VIN), 7 of 9 cases of Paget disease, 2 of 6 cases of lichen sclerosus (LS), and 2 of 3 cases of squamous cell hyperplasia (SCH); high-risk type HPV was revealed in 5 of 6 SCCs and 17 of 20 VINs. These observations might imply that a subset of cases of LS and SCH result from a neoplastic proliferation, similar to VINs but not related to infection with high-risk type HPV. In 1 case of SCC with concurrent VIN 3 in an adjacent lesion, both lesions showed the same pattern of X chromosome inactivation and the presence of HPV-16 in episomal and integrated forms, suggesting that monoclonal expansion triggered by high-risk type HPV integration is an early event for carcinogenesis of HPV-associated SCC.

An Improved MLVF Method and Its Comparison with Traditional MLVF, spa Typing, MLST/SCCmec and PFGE for the Typing of Methicillin-Resistant Staphylococcus aureus

PubMed Central

Du, Xue-Fei; Xiao, Meng; Liang, Hong-Yan; Sun, Zhe; Jiang, Yue-Hong; Chen, Guo-Yu; Meng, Xiao-Yu; Zou, Gui-Ling; Zhang, Li; Liu, Ya-Li; Zhang, Hui; Sun, Hong-Li; Jiang, Xiao-Feng; Xu, Ying-Chun

2014-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) has become an important nosocomial pathogen, causing considerable morbidity and mortality. During the last 20 years, a variety of genotyping methods have been introduced for screening the prevalence of MRSA. In this study, we developed and evaluated an improved approach capillary gel electrophoresis based multilocus variable-number tandem-repeat fingerprinting (CGE/MLVF) for rapid MRSA typing. A total of 42 well-characterized strains and 116 non-repetitive clinical MRSA isolates collected from six hospitals in northeast China between 2009 and 2010 were tested. The results obtained by CGE/MLVF against clinical isolates were compared with traditional MLVF, spa typing, Multilocus sequence typing/staphylococcal cassette chromosome mec (MLST/SCCmec) and pulse field gel electrophoresis (PFGE). The discriminatory power estimated by Simpson’s index of diversity was 0.855 (28 types), 0.855 (28 patterns), 0.623 (11 types), 0.517 (8 types) and 0.854 (28 patterns) for CGE/MLVF, traditional MLVF, spa typing, MLST/SCCmec and PFGE, respectively. All methods tested showed a satisfied concordance in clonal complex level calculated by adjusted Rand’s coefficient. CGE/MLVF showed better reproducibility and accuracy than traditional MLVF and PFGE methods. In addition, the CGE/MLVF has potential to produce portable results. In conclusion, CGE/MLVF is a rapid and easy to use MRSA typing method with lower cost, good reproducibility and high discriminatory power for monitoring the outbreak and clonal spread of MRSA isolates. PMID:24406728

Gene Loss and Lineage-Specific Restriction-Modification Systems Associated with Niche Differentiation in the Campylobacter jejuni Sequence Type 403 Clonal Complex

PubMed Central

Morley, Laura; McNally, Alan; Paszkiewicz, Konrad; Corander, Jukka; Méric, Guillaume; Sheppard, Samuel K.; Blom, Jochen

2015-01-01

Campylobacter jejuni is a highly diverse species of bacteria commonly associated with infectious intestinal disease of humans and zoonotic carriage in poultry, cattle, pigs, and other animals. The species contains a large number of distinct clonal complexes that vary from host generalist lineages commonly found in poultry, livestock, and human disease cases to host-adapted specialized lineages primarily associated with livestock or poultry. Here, we present novel data on the ST403 clonal complex of C. jejuni, a lineage that has not been reported in avian hosts. Our data show that the lineage exhibits a distinctive pattern of intralineage recombination that is accompanied by the presence of lineage-specific restriction-modification systems. Furthermore, we show that the ST403 complex has undergone gene decay at a number of loci. Our data provide a putative link between the lack of association with avian hosts of C. jejuni ST403 and both gene gain and gene loss through nonsense mutations in coding sequences of genes, resulting in pseudogene formation. PMID:25795671

Molecular characterization of endocarditis-associated Staphylococcus aureus.

PubMed

Nethercott, Cara; Mabbett, Amanda N; Totsika, Makrina; Peters, Paul; Ortiz, Juan C; Nimmo, Graeme R; Coombs, Geoffrey W; Walker, Mark J; Schembri, Mark A

2013-07-01

Infective endocarditis (IE) is a life-threatening infection of the heart endothelium and valves. Staphylococcus aureus is a predominant cause of severe IE and is frequently associated with infections in health care settings and device-related infections. Multilocus sequence typing (MLST), spa typing, and virulence gene microarrays are frequently used to classify S. aureus clinical isolates. This study examined the utility of these typing tools to investigate S. aureus epidemiology associated with IE. Ninety-seven S. aureus isolates were collected from patients diagnosed with (i) IE, (ii) bloodstream infection related to medical devices, (iii) bloodstream infection not related to medical devices, and (iv) skin or soft-tissue infections. The MLST clonal complex (CC) for each isolate was determined and compared to the CCs of members of the S. aureus population by eBURST analysis. The spa type of all isolates was also determined. A null model was used to determine correlations of IE with CC and spa type. DNA microarray analysis was performed, and a permutational analysis of multivariate variance (PERMANOVA) and principal coordinates analysis were conducted to identify genotypic differences between IE and non-IE strains. CC12, CC20, and spa type t160 were significantly associated with IE S. aureus. A subset of virulence-associated genes and alleles, including genes encoding staphylococcal superantigen-like proteins, fibrinogen-binding protein, and a leukocidin subunit, also significantly correlated with IE isolates. MLST, spa typing, and microarray analysis are promising tools for monitoring S. aureus epidemiology associated with IE. Further research to determine a role for the S. aureus IE-associated virulence genes identified in this study is warranted.

Molecular Characterization of Endocarditis-Associated Staphylococcus aureus

PubMed Central

Nethercott, Cara; Mabbett, Amanda N.; Totsika, Makrina; Peters, Paul; Ortiz, Juan C.; Nimmo, Graeme R.; Coombs, Geoffrey W.

2013-01-01

Infective endocarditis (IE) is a life-threatening infection of the heart endothelium and valves. Staphylococcus aureus is a predominant cause of severe IE and is frequently associated with infections in health care settings and device-related infections. Multilocus sequence typing (MLST), spa typing, and virulence gene microarrays are frequently used to classify S. aureus clinical isolates. This study examined the utility of these typing tools to investigate S. aureus epidemiology associated with IE. Ninety-seven S. aureus isolates were collected from patients diagnosed with (i) IE, (ii) bloodstream infection related to medical devices, (iii) bloodstream infection not related to medical devices, and (iv) skin or soft-tissue infections. The MLST clonal complex (CC) for each isolate was determined and compared to the CCs of members of the S. aureus population by eBURST analysis. The spa type of all isolates was also determined. A null model was used to determine correlations of IE with CC and spa type. DNA microarray analysis was performed, and a permutational analysis of multivariate variance (PERMANOVA) and principal coordinates analysis were conducted to identify genotypic differences between IE and non-IE strains. CC12, CC20, and spa type t160 were significantly associated with IE S. aureus. A subset of virulence-associated genes and alleles, including genes encoding staphylococcal superantigen-like proteins, fibrinogen-binding protein, and a leukocidin subunit, also significantly correlated with IE isolates. MLST, spa typing, and microarray analysis are promising tools for monitoring S. aureus epidemiology associated with IE. Further research to determine a role for the S. aureus IE-associated virulence genes identified in this study is warranted. PMID:23616460

Characterization of in vivo-acquired resistance to macrolides of Mycoplasma gallisepticum strains isolated from poultry

PubMed Central

2011-01-01

The macrolide class of antibiotics, including tylosin and tilmicosin, is widely used in the veterinary field for prophylaxis and treatment of mycoplasmosis. In vitro susceptibility testing of 50 strains of M. gallisepticum isolated in Israel during the period 1997-2010 revealed that acquired resistance to tylosin as well as to tilmicosin was present in 50% of them. Moreover, 72% (13/18) of the strains isolated from clinical samples since 2006 showed acquired resistance to enrofloxacin, tylosin and tilmicosin. Molecular typing of the field isolates, performed by gene-target sequencing (GTS), detected 13 molecular types (I-XIII). Type II was the predominant type prior to 2006 whereas type X, first detected in 2008, is currently prevalent. All ten type X strains were resistant to both fluoroquinolones and macrolides, suggesting selective pressure leading to clonal dissemination of resistance. However, this was not a unique event since resistant strains with other GTS molecular types were also found. Concurrently, the molecular basis for macrolide resistance in M. gallisepticum was identified. Our results revealed a clear-cut correlation between single point mutations A2058G or A2059G in domain V of the gene encoding 23S rRNA (rrnA, MGA_01) and acquired macrolide resistance in M. gallisepticum. Indeed, all isolates with MIC ≥ 0.63 μg/mL to tylosin and with MIC ≥ 1.25 μg/mL to tilmicosin possess one of these mutations, suggesting an essential role in decreased susceptibility of M. gallisepticum to 16-membered macrolides. PMID:21810258

Campylobacter jejuni colonization and population structure in urban populations of ducks and starlings in New Zealand.

PubMed

Mohan, Vathsala; Stevenson, Mark; Marshall, Jonathan; Fearnhead, Paul; Holland, Barbara R; Hotter, Grant; French, Nigel P

2013-08-01

A repeated cross-sectional study was conducted to determine the prevalence of Campylobacter spp. and the population structure of C. jejuni in European starlings and ducks cohabiting multiple public access sites in an urban area of New Zealand. The country's geographical isolation and relatively recent history of introduction of wild bird species, including the European starling and mallard duck, create an ideal setting to explore the impact of geographical separation on the population biology of C. jejuni, as well as potential public health implications. A total of 716 starling and 720 duck fecal samples were collected and screened for C. jejuni over a 12 month period. This study combined molecular genotyping, population genetics and epidemiological modeling and revealed: (i) higher Campylobacter spp. isolation in starlings (46%) compared with ducks (30%), but similar isolation of C. jejuni in ducks (23%) and starlings (21%), (ii) significant associations between the isolation of Campylobacter spp. and host species, sampling location and time of year using logistic regression, (iii) evidence of population differentiation, as indicated by FST , and host-genotype association with clonal complexes CC ST-177 and CC ST-682 associated with starlings, and clonal complexes CC ST-1034, CC ST-692, and CC ST-1332 associated with ducks, and (iv) greater genetic diversity and genotype richness in ducks compared with starlings. These findings provide evidence that host-associated genotypes, such as the starling-associated ST-177 and ST-682, represent lineages that were introduced with the host species in the 19th century. The isolation of sequence types associated with human disease in New Zealand indicate that wild ducks and starlings need to be considered as a potential public health risk, particularly in urban areas. © 2013 The Authors. Microbiology Open published by John Wiley & Sons Ltd.

Group B streptococci causing neonatal infections in barcelona are a stable clonal population: 18-year surveillance.

PubMed

Martins, E R; Andreu, A; Correia, P; Juncosa, T; Bosch, J; Ramirez, M; Melo-Cristino, J

2011-08-01

We analyzed 212 group B streptococci (GBS) from newborns with invasive infections in the area of Barcelona, Spain, between 1992 and 2009, with the aim of documenting changes in the prevalences of serotypes, antimicrobial resistance, and genetic lineages and evaluating their associations with either early-onset disease (EOD) or late-onset disease (LOD). Serotypes III (n = 118) and Ia (n = 47) together accounted for nearly 78% of the isolates. All isolates carried an alpha or alpha-like protein gene, and specific associations between genes and serotypes, such as serotype Ib and bca, serotype II and bca, serotype III and rib, and serotype V and alp3, reflected the presence of particular genetic lineages. Macrolide resistance (14.2%) was significantly associated with serotype V. Pulsed-field gel electrophoresis (PFGE) clustering was an excellent predictor of serotype and antibiotic resistance. The combination of PFGE and multilocus sequence typing revealed a large number of genetically distinct lineages. Still, specific lineages were dominant in our collection, particularly the serotype III/ST17/rib lineage, which had enhanced potential to cause LOD. Serotype Ia was concentrated in a single PFGE cluster composed of two genetic lineages: ST23/eps and ST24/bca. The ST24/bca sublineage of serotype Ia, which is found infrequently elsewhere, may be emerging as an important cause of neonatal invasive infections in the Mediterranean region. In spite of the introduction of prophylaxis, resulting in a pronounced decline in the frequency of EOD, the study revealed a remarkably stable clonal structure of GBS causing neonatal infections in Barcelona over a period of 18 years.

Group B Streptococci Causing Neonatal Infections in Barcelona Are a Stable Clonal Population: 18-Year Surveillance▿

PubMed Central

Martins, E. R.; Andreu, A.; Correia, P.; Juncosa, T.; Bosch, J.; Ramirez, M.; Melo-Cristino, J.

2011-01-01

We analyzed 212 group B streptococci (GBS) from newborns with invasive infections in the area of Barcelona, Spain, between 1992 and 2009, with the aim of documenting changes in the prevalences of serotypes, antimicrobial resistance, and genetic lineages and evaluating their associations with either early-onset disease (EOD) or late-onset disease (LOD). Serotypes III (n = 118) and Ia (n = 47) together accounted for nearly 78% of the isolates. All isolates carried an alpha or alpha-like protein gene, and specific associations between genes and serotypes, such as serotype Ib and bca, serotype II and bca, serotype III and rib, and serotype V and alp3, reflected the presence of particular genetic lineages. Macrolide resistance (14.2%) was significantly associated with serotype V. Pulsed-field gel electrophoresis (PFGE) clustering was an excellent predictor of serotype and antibiotic resistance. The combination of PFGE and multilocus sequence typing revealed a large number of genetically distinct lineages. Still, specific lineages were dominant in our collection, particularly the serotype III/ST17/rib lineage, which had enhanced potential to cause LOD. Serotype Ia was concentrated in a single PFGE cluster composed of two genetic lineages: ST23/eps and ST24/bca. The ST24/bca sublineage of serotype Ia, which is found infrequently elsewhere, may be emerging as an important cause of neonatal invasive infections in the Mediterranean region. In spite of the introduction of prophylaxis, resulting in a pronounced decline in the frequency of EOD, the study revealed a remarkably stable clonal structure of GBS causing neonatal infections in Barcelona over a period of 18 years. PMID:21697333

Genetic Characterization of Toxoplasma gondii Isolates from Pigs in Jilin Province, Northeastern China.

PubMed

Jiang, Hai-Hai; Wang, Shu-Chao; Huang, Si-Yang; Zhao, Lei; Wang, Ze-Dong; Zhu, Xing-Quan; Liu, Quan

2016-02-01

Toxoplasma gondii is prevalent in humans and animals worldwide. The present study aimed to determine the genetic diversity of T. gondii isolates from pigs in Jilin province, northeastern China. A total of 100 DNA samples were extracted from the hilar lymph nodes of slaughtered pigs, and 9 (9.0%, 95% confidence interval: 3.4-14.6%) were detected positive for T. gondii B1 gene by a nested polymerase chain reaction (PCR). The positive DNA samples were typed at 11 genetic markers, including 10 nuclear loci (SAG1, 5'-SAG2, and 3'-SAG2, alternative SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, and PK1) and an apicoplast locus (Apico) using the multilocus PCR-restriction fragment length polymorphism technology. Only three isolates were completely typed at all loci, showing that they all belonged to the clonal type I. One isolate was typed at five loci, including 5' +3'-SAG2, SAG2, SAG3, GRA6, and L358, revealing the possible clonal type I. This is the first report of the genetic characterization of T. gondii isolates in pigs in Jilin province, northeastern China, which has implications for better understanding the population structure of T. gondii infection in China.

[Multilocus sequence-typing for characterization of Moscow strains of Haemophilus influenzae type b].

PubMed

Platonov, A E; Mironov, K O; Iatsyshina, S B; Koroleva, I S; Platonova, O V; Gushchin, A E; Shipulin, G A

2003-01-01

Haemophilius influenzae, type b (Hib) bacteria, were genotyped by multilocus sequence typing (MLST) using 5 loci (adk, fucK, mdh, pgi, recA). 42 Moscow Hib strains (including 38 isolates form cerebrospinal fluid of children, who had purulent meningitis in 1999-2001, and 4 strains isolated from healthy carriers of Hib), as well as 2 strains from Yekaterinburg were studied. In MLST a strain is characterized, by alleles and their combinations (an allele profile) referred to also as sequence-type (ST). 9 Sts were identified within the Russian Hib bacteria: ST-1 was found in 25 strains (57%), ST-12 was found in 8 strains (18%), ST-11 was found in 4 strains (9%) and ST-15 was found in 2 strains (4.5%); all other STs strains (13, 14, 16, 17, 51) were found in isolated cases (2.3%). A comparison of allelic profiles and of nucleotide sequences showed that 93% of Russian isolates, i.e. strain with ST-1, 11, 12, 13, 15 and 17, belong to one and the same clonal complex. 2 isolates from Norway and Sweden from among 7 foreign Hib strains studied up to now can be described as belonging to the same clonal complex; 5 Hib strains were different from the Russian ones.

Enterotoxin gene profile of methicillin-resistant Staphylococcus pseudintermedius isolates from dogs, humans and the environment.

PubMed

Phumthanakorn, Nathita; Fungwithaya, Punpichaya; Chanchaithong, Pattrarat; Prapasarakul, Nuvee

2018-06-01

This study aimed to detect and identify staphylococcal enterotoxin (SE) genes in methicillin-resistant Staphylococcus pseudintermedius (MRSP) strains from different sources, and to investigate the relationship between their sequence types (STs) and SE gene patterns. The profiles of 17 SE genes in 93 MRSP strains isolated from dogs (n=43), humans (n=18) and the environment (n=32) were detected by PCR. Multilocus sequence typing (MLST), SCCmec typing and pulsed-field gel electrophoresis (PFGE) were used to analyse the clonal relatedness between the molecular type and SE gene profiles.Results/Key findings. The human MRSP strains harboured the greatest number of SE genes (12/17; sea, sec, seg, sei, sek, sel, sem, sen, seo, sep, seq and tst-1) compared to those from dogs (5/17; sec, sel, sem, seq and tst-1) and environmental sources (3/17; sec, seq and tst-1). Using MLST and PFGE, different SE gene profiles were found within the same clonal type. We show that isolates of MRSP vary in their virulence gene profiles, depending on the source from which they have been isolated. This insight should encourage the development of appropriate monitoring and mitigation strategies to reduce the transmission of MRSP in veterinary hospitals and households.

Intraspecific competition and light effect on reproduction of Ligularia virgaurea, an invasive native alpine grassland clonal herb

PubMed Central

Xie, Tian-peng; Zhang, Ge-fei; Zhao, Zhi-gang; Du, Guo-zhen; He, Gui-yong

2014-01-01

The relationship between sexual reproduction and clonal growth in clonal plants often shows up at the ramet level. However, only a few studies focus on the relationship at the genet level, which could finally account for evolution. The sexual reproduction and clonal growth of Ligularia virgaurea, a perennial herb widely distributed in the alpine grasslands of the Qinghai-Tibetan Plateau of China, were studied under different competition intensities and light conditions at the genet level through a potted experiment. The results showed that: (1) sexual reproduction did not depend on density or light, and increasing clonal growth with decreasing density and increasing light intensity indicated that intraspecific competition and light intensity may affect the clonal life history of L. virgaurea; (2) both sexual reproduction and clonal growth show a positive linear relationship with genet size under different densities and light conditions; (3) a threshold size is required for sexual reproduction and no evidence of a threshold size for clonal growth under different densities and light conditions; (4) light level affected the allocation of total biomass to clonal and sexual structures, with less allocation to clonal structures and more allocation to sexual structures in full sunlight than in shade; (5) light determined the onset of sexual reproduction, and the genets in the shade required a smaller threshold size for sexual reproduction to occur than the plants in full sunlight; and (6) no evidence was found of trade-offs between clonal growth and sexual reproduction under different densities and light conditions at the genet level, and the positive correlation between two reproductive modes indicated that these are two integrated processes. Clonal growth in this species may be viewed as a growth strategy that tends to maximize genet fitness. PMID:24683463

Real-time polymerase chain reaction for detection of encapsulated Haemophilus influenzae using degenerate primers to target the capsule transport gene bexA.

PubMed

Law, Dennis K S; Tsang, Raymond S W

2013-05-01

A real-time polymerase chain reaction assay that uses degenerate primers and a dual-labelled probe was developed to detect the bexA gene of Haemophilus influenzae, including those belonging to non-b serotypes as well as clonal division II strains. This assay is sensitive and specific, detecting 20 copies of the gene, but negative with a variety of bacteria associated with meningitis and bacteremia or septicemia.

Contrasting clonal structure among Pocillopora (Scleractinia) communities at two environmentally distinct sites in the Gulf of California

NASA Astrophysics Data System (ADS)

Pinzón, J. H.; Reyes-Bonilla, H.; Baums, I. B.; LaJeunesse, T. C.

2012-09-01

The contributions of sexual versus asexual reproduction are thought to play an important role in the abundance and ecological success of corals, especially in marginal habitats. Pocillopora corals are distributed throughout the Indo-Pacific and dominate shallow hard-bottom communities in the eastern Pacific where broad seasonal fluctuations in temperature and water turbidity create suboptimal conditions for reef community development. Previous work had revealed three genetic clades in the eastern Pacific that show little correspondence with colony morphology; the broad distribution of type 1 extends into the subtropical southern Gulf of California. Here we examine genetic and clonal structure of two type 1 communities separated by 10 km with microsatellite data. Samples were collected randomly in six 10 m radius circular plots (20 colonies per plot, 3 plots per site). Sites differed in their relative clonality because clonemates (ramets) from a single clone (genet) dominated a large portion (90.9 m long) of the protected leeward side of Gaviota Island (Number of genets/Number of samples = 0.35; observed Genotypic diversity/expected Genotypic diversity = 0.087), while an exposed community at the entrance to La Paz Bay, Punta Galeras, exhibited high genotypic diversity ( N g / N = 0.85; G o / G e = 0.714). Gene flow was unrestricted between sites indicating these communities comprised a single population. The relative proportion of asexual colonies found between community aggregations of Pocillopora in the Gulf of California differed significantly and suggests factors at local, not regional, scales affect these patterns. The possibility that heterogeneity in clonal structure is common throughout the eastern Pacific and across the west Indo-Pacific requires further study. Finally, since morphological variation in Pocillopora has been underappreciated and is in need of taxonomic revision, the use of a consistent field-sampling protocol and high-resolution makers will advance ecological research and aid in the conservation of these corals.

Molecular Characterization of Invasive Meningococcal Isolates from Countries in the African Meningitis Belt before Introduction of a Serogroup A Conjugate Vaccine

PubMed Central

Caugant, Dominique A.; Kristiansen, Paul A.; Wang, Xin; Mayer, Leonard W.; Taha, Muhamed-Kheir; Ouédraogo, Rasmata; Kandolo, Denis; Bougoudogo, Flabou; Sow, Samba; Bonte, Laurence

2012-01-01

Background The serogroup A conjugate meningococcal vaccine, MenAfriVac, was introduced in mass vaccination campaigns in December 2010 in Burkina Faso, Mali and Niger. In the coming years, vaccination will be extended to other African countries at risk of epidemics. To document the molecular characteristics of disease-causing meningococcal strains circulating in the meningitis belt of Africa before vaccine introduction, the World Health Organization Collaborating Centers on Meningococci in Europe and United States established a common strain collection of 773 isolates from cases of invasive meningococcal disease collected between 2004 and 2010 from 13 sub-Saharan countries. Methodology All isolates were characterized by multilocus sequence typing, and 487 (62%) were also analyzed for genetic variation in the surface antigens PorA and FetA. Antibiotic susceptibility was tested for part of the collection. Principal Findings Only 19 sequence types (STs) belonging to 6 clonal complexes were revealed. ST-5 clonal complex dominated with 578 (74.8%) isolates. All ST-5 complex isolates were remarkably homogeneous in their PorA (P1.20,9) and FetA (F3-1) and characterized the serogroup A strains which have been responsible for most epidemics during this time period. Sixty-eight (8.8%) of the 773 isolates belonged to the ST-11 clonal complex which was mainly represented by serogroup W135, while an additional 38 (4.9%) W135 isolates belonged to the ST-175 complex. Forty-eight (6.2%) serogroup X isolates from West Africa belonged to the ST-181 complex, while serogroup X cases in Kenya and Uganda were caused by an unrelated clone, ST-5403. Serogroup X, ST-181, emerged in Burkina Faso before vaccine introduction. Conclusions In the seven years preceding introduction of a new serogroup A conjugate vaccine, serogroup A of the ST-5 clonal complex was identified as the predominant disease-causing strain. PMID:23029368

Epidemiological tracking and population assignment of the non-clonal bacterium, Burkholderia pseudomallei.

PubMed

Dale, Julia; Price, Erin P; Hornstra, Heidie; Busch, Joseph D; Mayo, Mark; Godoy, Daniel; Wuthiekanun, Vanaporn; Baker, Anthony; Foster, Jeffrey T; Wagner, David M; Tuanyok, Apichai; Warner, Jeffrey; Spratt, Brian G; Peacock, Sharon J; Currie, Bart J; Keim, Paul; Pearson, Talima

2011-12-01

Rapid assignment of bacterial pathogens into predefined populations is an important first step for epidemiological tracking. For clonal species, a single allele can theoretically define a population. For non-clonal species such as Burkholderia pseudomallei, however, shared allelic states between distantly related isolates make it more difficult to identify population defining characteristics. Two distinct B. pseudomallei populations have been previously identified using multilocus sequence typing (MLST). These populations correlate with the major foci of endemicity (Australia and Southeast Asia). Here, we use multiple Bayesian approaches to evaluate the compositional robustness of these populations, and provide assignment results for MLST sequence types (STs). Our goal was to provide a reference for assigning STs to an established population without the need for further computational analyses. We also provide allele frequency results for each population to enable estimation of population assignment even when novel STs are discovered. The ability for humans and potentially contaminated goods to move rapidly across the globe complicates the task of identifying the source of an infection or outbreak. Population genetic dynamics of B. pseudomallei are particularly complicated relative to other bacterial pathogens, but the work here provides the ability for broad scale population assignment. As there is currently no independent empirical measure of successful population assignment, we provide comprehensive analytical details of our comparisons to enable the reader to evaluate the robustness of population designations and assignments as they pertain to individual research questions. Finer scale subdivision and verification of current population compositions will likely be possible with genotyping data that more comprehensively samples the genome. The approach used here may be valuable for other non-clonal pathogens that lack simple group-defining genetic characteristics and provides a rapid reference for epidemiologists wishing to track the origin of infection without the need to compile population data and learn population assignment algorithms.

Microsatellites within the feline androgen receptor are suitable for X chromosome-linked clonality testing in archival material.

PubMed

Farwick, Nadine M; Klopfleisch, Robert; Gruber, Achim D; Weiss, Alexander Th A

2017-04-01

Objectives A hallmark of neoplasms is their origin from a single cell; that is, clonality. Many techniques have been developed in human medicine to utilise this feature of tumours for diagnostic purposes. One approach is X chromosome-linked clonality testing using polymorphisms of genes encoded by genes on the X chromosome. The aim of this study was to determine if the feline androgen receptor gene was suitable for X chromosome-linked clonality testing. Methods The feline androgen receptor gene was characterised and used to test clonality of feline lymphomas by PCR and polyacrylamide gel electrophoresis, using archival formalin-fixed, paraffin-embedded material. Results Clonality of the feline lymphomas under study was confirmed and the gene locus was shown to represent a suitable target in clonality testing. Conclusions and relevance Because there are some pitfalls of using X chromosome-linked clonality testing, further studies are necessary to establish this technique in the cat.

Aging, clonal hematopoiesis and preleukemia: not just bad luck?

PubMed

Shlush, Liran I; Zandi, Sasan; Itzkovitz, Shalev; Schuh, Andre C

2015-11-01

Chronological human aging is associated with a number of changes in the hematopoietic system, occurring at many levels from stem to mature cells, and the marrow microenvironment as well. This review will focus mainly on the aging of hematopoietic stem and progenitor cells (HSPCs), and on the associated increases in the incidence of hematological malignancies. HSPCs manifest reduced function and acquire molecular changes with chronological aging. Furthermore, while for many years it has been known that the human hematopoietic system becomes increasingly clonal with chronological aging (clonal hematopoiesis), only in the last few years has it become clear that clonal hematopoiesis may result from the accumulation of preleukemic mutations in HSPCs. Such mutations confer a selective advantage that leads to clonal hematopoiesis, and that may occasionally result in the development of leukemia, and define the existence of both preleukemic stem cells, and of 'preleukemia' as a clinical entity. While it is well appreciated that clonal hematopoiesis is very common in the elderly, several questions remain unanswered: why and how does clonal hematopoiesis develop? How is clonal hematopoiesis related to the age-related changes observed in the hematopoietic system? And why do only some individuals with clonal hematopoiesis develop leukemia?

Proposal of a Consensus Set of Hypervariable Mycobacterial Interspersed Repetitive-Unit–Variable-Number Tandem-Repeat Loci for Subtyping of Mycobacterium tuberculosis Beijing Isolates

PubMed Central

Allix-Béguec, Caroline; Wahl, Céline; Hanekom, Madeleine; Nikolayevskyy, Vladyslav; Drobniewski, Francis; Maeda, Shinji; Campos-Herrero, Isolina; Mokrousov, Igor; Niemann, Stefan; Kontsevaya, Irina; Rastogi, Nalin; Samper, Sofia; Sng, Li-Hwei; Warren, Robin M.

2014-01-01

Mycobacterium tuberculosis Beijing strains represent targets of special importance for molecular surveillance of tuberculosis (TB), especially because they are associated with spread of multidrug resistance in some world regions. Standard 24-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat (MIRU-VNTR) typing lacks resolution power for accurately discriminating closely related clones that often compose Beijing strain populations. Therefore, we evaluated a set of 7 additional, hypervariable MIRU-VNTR loci for better resolution and tracing of such strains, using a collection of 535 Beijing isolates from six world regions where these strains are known to be prevalent. The typeability and interlaboratory reproducibility of these hypervariable loci were lower than those of the 24 standard loci. Three loci (2163a, 3155, and 3336) were excluded because of their redundant variability and/or more frequent noninterpretable results compared to the 4 other markers. The use of the remaining 4-locus set (1982, 3232, 3820, and 4120) increased the number of types by 52% (from 223 to 340) and reduced the clustering rate from 58.3 to 36.6%, when combined with the use of the standard 24-locus set. Known major clonal complexes/24-locus-based clusters were all subdivided, although the degree of subdivision varied depending on the complex. Only five single-locus variations were detected among the hypervariable loci of an additional panel of 92 isolates, representing 15 years of clonal spread of a single Beijing strain in a geographically restricted setting. On this calibrated basis, we propose this 4-locus set as a consensus for subtyping Beijing clonal complexes and clusters, after standard typing. PMID:24172154

Proposal of a consensus set of hypervariable mycobacterial interspersed repetitive-unit-variable-number tandem-repeat loci for subtyping of Mycobacterium tuberculosis Beijing isolates.

PubMed

Allix-Béguec, Caroline; Wahl, Céline; Hanekom, Madeleine; Nikolayevskyy, Vladyslav; Drobniewski, Francis; Maeda, Shinji; Campos-Herrero, Isolina; Mokrousov, Igor; Niemann, Stefan; Kontsevaya, Irina; Rastogi, Nalin; Samper, Sofia; Sng, Li-Hwei; Warren, Robin M; Supply, Philip

2014-01-01

Mycobacterium tuberculosis Beijing strains represent targets of special importance for molecular surveillance of tuberculosis (TB), especially because they are associated with spread of multidrug resistance in some world regions. Standard 24-locus mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing lacks resolution power for accurately discriminating closely related clones that often compose Beijing strain populations. Therefore, we evaluated a set of 7 additional, hypervariable MIRU-VNTR loci for better resolution and tracing of such strains, using a collection of 535 Beijing isolates from six world regions where these strains are known to be prevalent. The typeability and interlaboratory reproducibility of these hypervariable loci were lower than those of the 24 standard loci. Three loci (2163a, 3155, and 3336) were excluded because of their redundant variability and/or more frequent noninterpretable results compared to the 4 other markers. The use of the remaining 4-locus set (1982, 3232, 3820, and 4120) increased the number of types by 52% (from 223 to 340) and reduced the clustering rate from 58.3 to 36.6%, when combined with the use of the standard 24-locus set. Known major clonal complexes/24-locus-based clusters were all subdivided, although the degree of subdivision varied depending on the complex. Only five single-locus variations were detected among the hypervariable loci of an additional panel of 92 isolates, representing 15 years of clonal spread of a single Beijing strain in a geographically restricted setting. On this calibrated basis, we propose this 4-locus set as a consensus for subtyping Beijing clonal complexes and clusters, after standard typing.

Subtyping of Clostridium difficile PCR ribotypes 591, 106 and 002, the dominant strain types circulating in Medellin, Colombia.

PubMed

Salazar, Clara Lina; Reyes, Catalina; Cienfuegos-Gallet, Astrid Vanessa; Best, Emma; Atehortua, Santiago; Sierra, Patricia; Correa, Margarita M; Fawley, Warren N; Paredes-Sabja, Daniel; Wilcox, Mark; Gonzalez, Angel

2018-01-01

We aimed to achieve a higher typing resolution within the three dominant Clostridium difficile ribotypes (591,106 and 002) circulating in Colombia. A total of 50 C. difficile isolates we had previously typed by PCR-ribotyping, representing the major three ribotypes circulating in Colombia, were analyzed. Twenty-seven isolates of ribotype 591, 12 of ribotype 106 and 11 of ribotype 002 were subtyped by multiple locus variable-number tandem-repeat analysis (MLVA). The presence of the PaLoc genes (tcdA/tcdB), toxin production in culture and antimicrobial susceptibility were also determined. From the total C. difficile ribotypes analyzed, 20 isolates (74%) of ribotype 591, nine (75%) of ribotype 106 and five (45.5%) of ribotype 002 were recovered from patients with Clostridium difficile infection (CDI). MLVA allowed us to recognize four and two different clonal complexes for ribotypes 591 and 002, respectively, having a summed tandem-repeat difference (STRD) <2, whereas none of the ribotype 106 isolates were grouped in a cluster or clonal complex having a STRD >10. Six ribotype 591 and three ribotype 002 isolates belonging to a defined clonal complex were isolated on the same week in two different hospitals. All ribotypes harbored either tcdA+/tcdB+ or tcdA-/tcdB+ PaLoc genes. Moreover, 94% of the isolates were positive for toxin in culture. All isolates were susceptible to vancomycin and metronidazole, while 75% to 100% of the isolates were resistant to clindamycin, and less than 14.8% of ribotype 591 isolates were resistant to moxifloxacina. No significant differences were found among ribotypes with respect to demographic and clinical patients' data; however, our results demonstrated a high molecular heterogeneity of C. difficile strains circulating in Colombia.

Subtyping of Clostridium difficile PCR ribotypes 591, 106 and 002, the dominant strain types circulating in Medellin, Colombia

PubMed Central

Salazar, Clara Lina; Reyes, Catalina; Cienfuegos-Gallet, Astrid Vanessa; Best, Emma; Atehortua, Santiago; Sierra, Patricia; Correa, Margarita M.; Fawley, Warren N.; Paredes-Sabja, Daniel; Wilcox, Mark

2018-01-01

We aimed to achieve a higher typing resolution within the three dominant Clostridium difficile ribotypes (591,106 and 002) circulating in Colombia. A total of 50 C. difficile isolates we had previously typed by PCR-ribotyping, representing the major three ribotypes circulating in Colombia, were analyzed. Twenty-seven isolates of ribotype 591, 12 of ribotype 106 and 11 of ribotype 002 were subtyped by multiple locus variable-number tandem-repeat analysis (MLVA). The presence of the PaLoc genes (tcdA/tcdB), toxin production in culture and antimicrobial susceptibility were also determined. From the total C. difficile ribotypes analyzed, 20 isolates (74%) of ribotype 591, nine (75%) of ribotype 106 and five (45.5%) of ribotype 002 were recovered from patients with Clostridium difficile infection (CDI). MLVA allowed us to recognize four and two different clonal complexes for ribotypes 591 and 002, respectively, having a summed tandem-repeat difference (STRD) <2, whereas none of the ribotype 106 isolates were grouped in a cluster or clonal complex having a STRD >10. Six ribotype 591 and three ribotype 002 isolates belonging to a defined clonal complex were isolated on the same week in two different hospitals. All ribotypes harbored either tcdA+/tcdB+ or tcdA-/tcdB+ PaLoc genes. Moreover, 94% of the isolates were positive for toxin in culture. All isolates were susceptible to vancomycin and metronidazole, while 75% to 100% of the isolates were resistant to clindamycin, and less than 14.8% of ribotype 591 isolates were resistant to moxifloxacina. No significant differences were found among ribotypes with respect to demographic and clinical patients’ data; however, our results demonstrated a high molecular heterogeneity of C. difficile strains circulating in Colombia. PMID:29649308

Clonal Spread of a Unique Strain of Macrolide-Resistant Mycoplasma Pneumoniae Within a Single Family in Italy.

PubMed

Chironna, Maria; Loconsole, Daniela; De Robertis, Anna Lisa; Morea, Anna; Scalini, Egidio; Quarto, Michele; Tafuri, Silvio; Germinario, Cinzia; Manzionna, Mariano

2016-03-01

Macrolide-resistant Mycoplasma pneumoniae (MR-MP) is an increasing problem worldwide. This study describes the clonal spread of a unique strain of MR-MP within a single family. On January 23, 2015, nasopharyngeal swabs and sputum samples were collected from the index case (a 9-year-old girl) in southern Italy. The patient had pneumonia and was initially treated with clarithromycin. MR-MP infection was suspected due to prolonged symptoms despite appropriate antibiotic therapy. Two further cases of pneumonia occurred in relatives (a 7-year-old cousin and the 36-year-old mother of the index case); therefore, respiratory samples were also collected from other family members. Sequence analysis identified mutations associated with resistance to macrolides. Both P1 major adhesion protein typing and multiple loci variable-number tandem repeat analysis (MLVA) typing were performed to assess the relatedness of the strains. The index case, the cousin, the mother, and another 4 family members (twin siblings of the index case, a 3-year-old cousin, and the grandmother) were positive for MR-MP. All strains harbored the mutation A2063G, had the same P1 subtype (1), and were MLVA (7/4/5/7/2) type Z. In addition, the index case's aunt (31 years of age and the probable source of infection) harbored an M pneumoniae strain with the same molecular profile; however, this strain was susceptible to macrolides. This cluster of MR-MP infection/carriage caused by a clonal strain suggests a high transmission rate within this family and highlights the need for increased awareness among clinicians regarding the circulation of MR-MP. Novel strategies for the treatment and prevention of M pneumoniae infections are required.

Molecular characterization of macrolide resistant Streptococcus pyogenes isolates from pharyngitis patients in Serbia.

PubMed

Opavski, Natasa; Gajic, Ina; Borek, Anna L; Obszańska, Katarzyna; Stanojevic, Maja; Lazarevic, Ivana; Ranin, Lazar; Sitkiewicz, Izabela; Mijac, Vera

2015-07-01

A steady increase in macrolide resistance in Streptococcus pyogenes, group A streptococci (GAS) was reported in Serbia during 2004-2009 (9.9%). However, there are no data on the molecular epidemiology of pharyngeal macrolide resistance GAS (MRGAS) isolates. Therefore, the aims of this first nationwide study were to examine the prevalence of macrolide resistance in Serbian GAS and to determine their resistance phenotypes, genotypes and clonal relationships. Overall 3893 non-duplicate pharyngeal S. pyogenes isolates from outpatients with GAS infection were collected throughout country during 2008 and 2009. Among 486 macrolide resistant pharyngeal isolates collected, 103 were further characterized. Macrolide resistance phenotypes and genotypes were determined by double-disk diffusion test and PCR, respectively. Strain relatedness was determined by emm typing, multilocus sequence typing (MLST), multilocus variable tandem repeat analysis (MLVA), phage profiling (PP) and virulence factor profiling (VFP). Overall, macrolide resistance among GAS isolates in Serbia was 12.5%. M phenotype was the most common (71.8%), followed by iMLS (18.4%) and cMLS (9.7%). Three clonal complexes--emm75/mefA/ST49, emm12/mefA/ST36 and emm77/ermA/tetO/ST63 comprised over 90% of the tested strains. Although MLVA, PP and VFP distinguished 10, 20 and 12 different patterns, respectively, cluster analysis disclosed only small differences between strains which belonged to the same emm/ST type. Our data indicate dominance of three major internationally widely disseminated macrolide resistant clones and a high genetic homogeneity among the Serbian MRGAS population. Continued surveillance of macrolide resistance and clonal composition in MRGAS in Serbia in future is necessary to determine stability of MRGAS clones and to guide therapy strategies. Copyright © 2015 Elsevier B.V. All rights reserved.

From homothally to heterothally: Mating preferences and genetic variation within clones of the dinoflagellate Gymnodinium catenatum

NASA Astrophysics Data System (ADS)

Figueroa, Rosa Isabel; Rengefors, Karin; Bravo, Isabel; Bensch, Staffan

2010-02-01

The chain-forming dinoflagellate Gymnodinium catenatum Graham is responsible for outbreaks of paralytic shellfish poisoning (PSP), a human health threat in coastal waters. Sexuality in this species is of great importance in its bloom dynamics, and has been shown to be very complex but lacks an explanation. For this reason, we tested if unreported homothallic behavior and rapid genetic changes may clarify the sexual system of this alga. To achieve this objective, 12 clonal strains collected from the Spanish coast were analyzed for the presence of sexual reproduction. Mating affinity results, self-compatibility studies, and genetic fingerprinting (amplified fragment length polymorphism, AFLP) analysis on clonal strains, showed three facts not previously described for this species: (i) That there is a continuous mating system within G. catenatum, with either self-compatible strains (homothallic), or strains that needed to be outcrossed (heterothallic), and with a range of differences in cyst production among the crosses. (ii) There was intraclonal genetic variation, i.e. genetic variation within an asexual lineage. Moreover, the variability among homothallic clones was smaller than among the heterothallic ones. (iii) Sibling strains (the two strains established by the germination of one cyst) increased their intra- and inter-sexual compatibility with time. To summarize, we have found that G. catenatum's sexual system is much more complex than previously described, including complex homothallic/heterothallic behaviors. Additionally, high rates of genetic variability may arise in clonal strains, although explanations for the mechanisms responsible are still lacking.

Molecular characterization of Ambler class A to D β-lactamases, ISAba1, and integrons reveals multidrug-resistant Acinetobacter spp. isolates in northeastern China.

PubMed

Sun, Xiaoyu; Liu, Bin; Chen, Yan; Huang, Honglan; Wang, Guoqing; Li, Fan; Ni, Zhaohui

2016-12-01

The prevalence of various Ambler class A to D β-lactamases, ISAba1, and class 1 and 2 integrons as well as the clonal relatedness in 105 Acinetobacter spp. isolates found in northeastern China was investigated. All 105 Acinetobacter spp. isolates were determined to be multidrug resistant (MDR), and the resistance rates to carbapenem agents were approximately 50%. PER, IMP, AmpC, and OXA-23 were found to be dominant β-lactamases belonging to different classes, respectively. This is the first report of the coexistence of bla PER , bla IMP , bla AmpC , and bla OXA-23-like genes in Acinetobacter spp. isolates from northeastern China. ISAba1 was found upstream of the bla OXA-23-like gene in 87.8% (36/41) strains and upstream of the bla OXA-51-like gene in 26.5% (13/49) strains. ISAba3-like element was found upstream of the bla OXA-58-like gene in one bla OXA-58-like -positive strain. The presence of IntI1 was detected in 63.8% (67/105) of the isolates and the most prevalent gene cassettes were aacA4, aadA1, and catB8. The highly prevalent isolates belong to international clonal lineage (ICL)-II. These results indicate that the wide horizontal and clonal spread of MDR Acinetobacter spp. isolates harbouring multiple β-lactamase genes has become a serious problem in northeastern China.

Diagnostic Utility of a Clonality Test for Lymphoproliferative Diseases in Koreans Using the BIOMED-2 PCR Assay

PubMed Central

Kim, Young; Choi, Yoo Duk; Choi, Chan

2013-01-01

Background A clonality test for immunoglobulin (IG) and T cell receptor (TCR) is a useful adjunctive method for the diagnosis of lymphoproliferative diseases (LPDs). Recently, the BIOMED-2 multiplex polymerase chain reaction (PCR) assay has been established as a standard method for assessing the clonality of LPDs. We tested clonality in LPDs in Koreans using the BIOMED-2 multiplex PCR and compared the results with those obtained in European, Taiwanese, and Thai participants. We also evaluated the usefulness of the test as an ancillary method for diagnosing LPDs. Methods Two hundred and nineteen specimens embedded in paraffin, including 78 B cell lymphomas, 80 T cell lymphomas and 61 cases of reactive lymphadenitis, were used for the clonality test. Results Mature B cell malignancies showed 95.7% clonality for IG, 2.9% co-existing clonality, and 4.3% polyclonality. Mature T cell malignancies exhibited 83.8% clonality for TCR, 8.1% co-existing clonality, and 16.2% polyclonality. Reactive lymphadenitis showed 93.4% polyclonality for IG and TCR. The majority of our results were similar to those obtained in Europeans. However, the clonality for IGK of B cell malignancies and TCRG of T cell malignancies was lower in Koreans than Europeans. Conclusions The BIOMED-2 multiplex PCR assay was a useful adjunctive method for diagnosing LPDs. PMID:24255634

Lentiviral and targeted cellular barcoding reveals ongoing clonal dynamics of cell lines in vitro and in vivo

PubMed Central

2014-01-01

Background Cell lines are often regarded as clonal, even though this simplifies what is known about mutagenesis, transformation and other processes that destabilize them over time. Monitoring these clonal dynamics is important for multiple areas of biomedical research, including stem cell and cancer biology. Tracking the contributions of individual cells to large populations, however, has been constrained by limitations in sensitivity and complexity. Results We utilize cellular barcoding methods to simultaneously track the clonal contributions of tens of thousands of cells. We demonstrate that even with optimal culturing conditions, common cell lines including HeLa, K562 and HEK-293 T exhibit ongoing clonal dynamics. Starting a population with a single clone diminishes but does not eradicate this phenomenon. Next, we compare lentiviral and zinc-finger nuclease barcode insertion approaches, finding that the zinc-finger nuclease protocol surprisingly results in reduced clonal diversity. We also document the expected reduction in clonal complexity when cells are challenged with genotoxic stress. Finally, we demonstrate that xenografts maintain clonal diversity to a greater extent than in vitro culturing of the human non-small-cell lung cancer cell line HCC827. Conclusions We demonstrate the feasibility of tracking and quantifying the clonal dynamics of entire cell populations within multiple cultured cell lines. Our results suggest that cell heterogeneity should be considered in the design and interpretation of in vitro culture experiments. Aside from clonal cell lines, we propose that cellular barcoding could prove valuable in modeling the clonal behavior of heterogeneous cell populations over time, including tumor populations treated with chemotherapeutic agents. PMID:24886633

Genetic characterization of Toxoplasma gondii isolates and toxoplasmosis seroprevalence in stray cats of İzmir, Turkey.

PubMed

Can, Hüseyin; Döşkaya, Mert; Ajzenberg, Daniel; Özdemir, H Gökhan; Caner, Ayşe; İz, Sultan Gülce; Döşkaya, Aysu Değirmenci; Atalay, Esra; Çetinkaya, Çağdaş; Ürgen, Saygun; Karaçalı, Sabire; Ün, Cemal; Dardé, Marie-Laure; Gürüz, Yüksel

2014-01-01

Currently, some Toxoplasma gondii genotypes are being associated with serious clinical presentations. A recent report showing the Africa 1 genotype in two local congenital toxoplasmosis cases acquired in Turkey formed the basis of this study because atypical Africa 1 genotype is most frequently detected in animals and patients from sub-Saharan Africa. Since stray cats are considered as the linkage between wild life and urban life in T. gondii transmission, the present study aimed to isolate and characterize T. gondii strains circulating in stray cats of İzmir (Western Turkey). A secondary objective was to determine toxoplasmosis seroprevalence in this cat population. Tissues obtained from 100 deceased stray cats were bioassayed and isolated strains were genotyped using 15 microsatellite markers. In addition, toxoplasmosis seroprevalence was analyzed in 1121 cat sera collected from several large veterinary clinics in İzmir. Among the 22 isolates, 19 were Type II (86.3%), two were Type III (9%) and one was Africa 1 genotype (4.5%). The overall seropositivity rates in cats were 42-48% and 33.4-34.4% according to IFA and ELISA, respectively. Seroprevalence in deceased cats was significantly higher than in healthy cats (P = 0.0033). Finding both the major clonal Type II lineage together with the Type III lineage also found in Middle East, and an atypical genotype, Africa 1 appears consistent with the specific geographic location of Turkey between three continents and raises the possibility of transportation of these strains between continents through trade routes or long distance migratory birds. In addition, the first large study of toxoplasma seroprevalence in a stray cat population was also reported. The relatively high seropositivity rates and the variety of T. gondii genotypes confirm the local stray cat population as a risk factor for human toxoplasmosis in İzmir.

Genetic Characterization of Toxoplasma gondii Isolates and Toxoplasmosis Seroprevalence in Stray Cats of İzmir, Turkey

PubMed Central

Can, Hüseyin; Döşkaya, Mert; Ajzenberg, Daniel; Özdemir, H. Gökhan; Caner, Ayşe; İz, Sultan Gülce; Döşkaya, Aysu Değirmenci; Atalay, Esra; Çetinkaya, Çağdaş; Ürgen, Saygun; Karaçalı, Sabire; Ün, Cemal; Dardé, Marie-Laure; Gürüz, Yüksel

2014-01-01

Currently, some Toxoplasma gondii genotypes are being associated with serious clinical presentations. A recent report showing the Africa 1 genotype in two local congenital toxoplasmosis cases acquired in Turkey formed the basis of this study because atypical Africa 1 genotype is most frequently detected in animals and patients from sub-Saharan Africa. Since stray cats are considered as the linkage between wild life and urban life in T. gondii transmission, the present study aimed to isolate and characterize T. gondii strains circulating in stray cats of İzmir (Western Turkey). A secondary objective was to determine toxoplasmosis seroprevalence in this cat population. Tissues obtained from 100 deceased stray cats were bioassayed and isolated strains were genotyped using 15 microsatellite markers. In addition, toxoplasmosis seroprevalence was analyzed in 1121 cat sera collected from several large veterinary clinics in İzmir. Among the 22 isolates, 19 were Type II (86.3%), two were Type III (9%) and one was Africa 1 genotype (4.5%). The overall seropositivity rates in cats were 42–48% and 33.4–34.4% according to IFA and ELISA, respectively. Seroprevalence in deceased cats was significantly higher than in healthy cats (P = 0.0033). Finding both the major clonal Type II lineage together with the Type III lineage also found in Middle East, and an atypical genotype, Africa 1 appears consistent with the specific geographic location of Turkey between three continents and raises the possibility of transportation of these strains between continents through trade routes or long distance migratory birds. In addition, the first large study of toxoplasma seroprevalence in a stray cat population was also reported. The relatively high seropositivity rates and the variety of T. gondii genotypes confirm the local stray cat population as a risk factor for human toxoplasmosis in İzmir. PMID:25127360

Sequence, distribution and chromosomal context of class I and class II pilin genes of Neisseria meningitidis identified in whole genome sequences

PubMed Central

2014-01-01

Background Neisseria meningitidis expresses type four pili (Tfp) which are important for colonisation and virulence. Tfp have been considered as one of the most variable structures on the bacterial surface due to high frequency gene conversion, resulting in amino acid sequence variation of the major pilin subunit (PilE). Meningococci express either a class I or a class II pilE gene and recent work has indicated that class II pilins do not undergo antigenic variation, as class II pilE genes encode conserved pilin subunits. The purpose of this work was to use whole genome sequences to further investigate the frequency and variability of the class II pilE genes in meningococcal isolate collections. Results We analysed over 600 publically available whole genome sequences of N. meningitidis isolates to determine the sequence and genomic organization of pilE. We confirmed that meningococcal strains belonging to a limited number of clonal complexes (ccs, namely cc1, cc5, cc8, cc11 and cc174) harbour a class II pilE gene which is conserved in terms of sequence and chromosomal context. We also identified pilS cassettes in all isolates with class II pilE, however, our analysis indicates that these do not serve as donor sequences for pilE/pilS recombination. Furthermore, our work reveals that the class II pilE locus lacks the DNA sequence motifs that enable (G4) or enhance (Sma/Cla repeat) pilin antigenic variation. Finally, through analysis of pilin genes in commensal Neisseria species we found that meningococcal class II pilE genes are closely related to pilE from Neisseria lactamica and Neisseria polysaccharea, suggesting horizontal transfer among these species. Conclusions Class II pilins can be defined by their amino acid sequence and genomic context and are present in meningococcal isolates which have persisted and spread globally. The absence of G4 and Sma/Cla sequences adjacent to the class II pilE genes is consistent with the lack of pilin subunit variation in these isolates, although horizontal transfer may generate class II pilin diversity. This study supports the suggestion that high frequency antigenic variation of pilin is not universal in pathogenic Neisseria. PMID:24690385

Dominance of multidrug-resistant Denmark(14)-32 (ST230) clone among Streptococcus pneumoniae serotype 19A isolates causing pneumococcal disease in Bulgaria from 1992 to 2013.

PubMed

Setchanova, Lena Petrova; Alexandrova, Alexandra; Dacheva, Daniela; Mitov, Ivan; Kaneva, Radka; Mitev, Vanio

2015-02-01

A pneumococcal conjugate vaccine (PCV10) was introduced in Bulgarian national immunization program since April 2010. Clonal composition based on pulsed-field gel electrophoresis and multilocus sequence typing genotyping of 52 serotype 19A Streptococcus pneumoniae isolates was analyzed. These were invasive and respiratory isolates collected between 1992 and 2013 from both children (78.8% <5 years) and adults with pneumococcal infections. Multidrug resistance was found in 82.7% of all 19A isolates. The most prevalent genotype (63.5%) among serotype 19A pneumococcal strains was the multidrug-resistant clonal complex CC230, which is a capsular switched variant of the Denmark(14)-32 (ST230) global clone. The most frequent sequence type (ST) was ST230 (48.1%) and together with four other closely related STs (15.4%), belonging to ST1611, ST276, ST7466, and ST2013, which were single- and double-locus variants; they were included in the main CC230. The disappearance of highly drug-resistant ST663 clone and emergence of new clones as CC320 and CC199 was also observed among the rest 19A isolates. A comparison of clonal composition between invasive and noninvasive isolates did not show a great genetic diversity among both kinds of isolates. Continuous surveillance of serotype 19A population following the introduction of PCV10 is essential to evaluate the impact of the vaccine on the epidemiology of this serotype.

Dissemination of the ST-103 clonal complex serogroup C meningococci in Salvador, Brazil.

PubMed

Cordeiro, Soraia Machado; Cardoso, Cristiane Wanderley; de Araújo, Lorena Galvão; Ribeiro, Luis Eduardo; Azevedo, Jailton; Silva, Rita de Cassia Vilasboas; Dos Reis, Mitermayer Galvão; Ko, Albert Icksang; Reis, Joice Neves

2018-01-01

Invasive meningococcal disease (IMD) is a major public health problem worldwide. An epidemic of serogroup C (NmC) IMD occurred in 2010 in the city of Salvador. In this study, we describe the antigenic and genetic characterization of meningococcal isolates collected from meningitis cases in Salvador from 2001 to 2012. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were performed for the analysis of IMD isolates. A total of 733 cases were identified, and the serogroup was determined for 391 (53.0%) of these. Most cases were caused by NmC (53%) or B (47%). The most prevalent strains were B:4,7:P1.19,15 (32.9%; 129/391) and C:23:P1.14-6 (28.6%; 112/391). Based on PFGE/MLST analysis, 71.3% (77/108 PFGE-tested isolates) clustered as two clones of sequence type ST-3779 and ST-3780, both belonging to the ST-103 clonal complex. ST-3779 has been detected in Salvador since 1996 and together with ST-3780 became predominant after 2005. There was a predominance of C:23:P1.14-6, ST-3779/3780 in Salvador during the period of 2007-2012, establishing a major clonal lineage, which remained in the community for a long time; this has serious implications for public health, particularly in terms of prevention and control strategies of IMD. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Cytoreductive conditioning intensity predicts clonal diversity in ADA-SCID retroviral gene therapy patients

PubMed Central

Lill, Georgia R.; Shaw, Kit; Carbonaro-Sarracino, Denise A.; Davila, Alejandra; Sokolic, Robert; Candotti, Fabio; Pellegrini, Matteo

2017-01-01

Retroviral gene therapy has proved efficacious for multiple genetic diseases of the hematopoietic system, but roughly half of clinical gene therapy trial protocols using gammaretroviral vectors have reported leukemias in some of the patients treated. In dramatic contrast, 39 adenosine deaminase–deficient severe combined immunodeficiency (ADA-SCID) patients have been treated with 4 distinct gammaretroviral vectors without oncogenic consequence. We investigated clonal dynamics and diversity in a cohort of 15 ADA-SCID children treated with gammaretroviral vectors and found clear evidence of genotoxicity, indicated by numerous common integration sites near proto-oncogenes and by increased abundance of clones with integrations near MECOM and LMO2. These clones showed stable behavior over multiple years and never expanded to the point of dominance or dysplasia. One patient developed a benign clonal dominance that could not be attributed to insertional mutagenesis and instead likely resulted from expansion of a transduced natural killer clone in response to chronic Epstein-Barr virus viremia. Clonal diversity and T-cell repertoire, measured by vector integration site sequencing and T-cell receptor β-chain rearrangement sequencing, correlated significantly with the amount of busulfan preconditioning delivered to patients and to CD34+ cell dose. These data, in combination with results of other ADA-SCID gene therapy trials, suggest that disease background may be a crucial factor in leukemogenic potential of retroviral gene therapy and underscore the importance of cytoreductive conditioning in this type of gene therapy approach. PMID:28351939

Stenotrophomonas maltophilia in Mexico: antimicrobial resistance, biofilm formation and clonal diversity.

PubMed

Flores-Treviño, Samantha; Gutiérrez-Ferman, Jessica Lizzeth; Morfín-Otero, Rayo; Rodríguez-Noriega, Eduardo; Estrada-Rivadeneyra, Diego; Rivas-Morales, Catalina; Llaca-Díaz, Jorge M; Camacho-Ortíz, Adrián; Mendoza-Olazarán, Soraya; Garza-González, Elvira

2014-11-01

Stenotrophomonas maltophilia is an important multidrug-resistant nosocomial pathogen associated with high mortality. Our aim was to examine antimicrobial susceptibility, biofilm production and clonal relatedness of clinical isolates of S. maltophilia. S. maltophilia isolates were collected between 2006 and 2013 from two tertiary care hospitals in Mexico. Antimicrobial susceptibility was evaluated by the broth microdilution method. PCR was used to determine the presence of β-lactamase genes L1 and L2. Biofilm formation was assessed with crystal violet staining. Clonal relatedness was determined by PFGE. Among the 119 collected S. maltophilia isolates, 73 (61.3%) were from the respiratory tract. Resistance levels exceeded 75% for imipenem, meropenem, ampicillin, aztreonam, gentamicin and tobramycin. Resistance to trimethoprim-sulfamethoxazole was 32.8%. L1 and L2 genes were detected in 77.1% (91/118) and 66.9% (79/118) of isolates, respectively. All S. maltophilia strains were able to produce biofilms. Strains were classified as weak (47.9%, 57/119), moderate (38.7%, 46/119), or strong (13.4%, 16/119) biofilm producers. A total of 89 distinct PFGE types were identified and 21.6% (22/102) of the isolates were distributed in nine clusters. This is the first study in Mexico to reveal characteristics of clinical isolates of S. maltophilia. Clonal diversity data indicate low cross-transmission of S. maltophilia in a hospital setting. The high antibiotic resistance underscores the need for continuous surveillance of S. maltophilia in hospital settings in Mexico. © 2014 The Authors.

Clonal evolution and tumor-initiating cells: New dimensions in cancer patient treatment.

PubMed

Apostoli, Anthony J; Ailles, Laurie

2016-01-01

Human cancer is not a uniform disease but a plethora of disparate tumor types and subtypes. The differences that exist between individual tumors (intertumoral heterogeneity) present a significant roadblock to the eradication of cancer. It has also become increasingly clear that variations across individual tumors (intratumoral heterogeneity) have important implications to cancer progression and treatment efficacy. Therefore, in order to improve patient care and develop novel chemotherapeutics, the evolving tumor landscape needs to be further explored. Next-generation sequencing (NGS) technologies are revolutionizing the cancer research arena by providing state-of-the-art, high-speed methods of genome sequencing at single-nucleotide resolution, thus enabling an unprecedented detection of tumor-specific genetic abnormalities. These anomalies can be quantified to reveal specific frequencies of DNA alterations that correspond to distinct clonal populations within a given tumor. As such, NGS approaches have also been utilized to explore the heterogeneous landscape of patient tumors as well as to match metastatic and/or recurrent growths and patient-derived engrafts. By sequencing in this manner--through time so to speak--cancer researchers can track shifting clonal populations, make important inferences about tumor evolution and potentially identify tumor subclones that could be viably targeted. This exciting new territory has important implications for the competing clonal evolution and cancer stem cell models of tumor heterogeneity, and also offers a new dimension for cancer treatment and profound hope for patients in the coming years.

Cytoreductive conditioning intensity predicts clonal diversity in ADA-SCID retroviral gene therapy patients.

PubMed

Cooper, Aaron R; Lill, Georgia R; Shaw, Kit; Carbonaro-Sarracino, Denise A; Davila, Alejandra; Sokolic, Robert; Candotti, Fabio; Pellegrini, Matteo; Kohn, Donald B

2017-05-11

Retroviral gene therapy has proved efficacious for multiple genetic diseases of the hematopoietic system, but roughly half of clinical gene therapy trial protocols using gammaretroviral vectors have reported leukemias in some of the patients treated. In dramatic contrast, 39 adenosine deaminase-deficient severe combined immunodeficiency (ADA-SCID) patients have been treated with 4 distinct gammaretroviral vectors without oncogenic consequence. We investigated clonal dynamics and diversity in a cohort of 15 ADA-SCID children treated with gammaretroviral vectors and found clear evidence of genotoxicity, indicated by numerous common integration sites near proto-oncogenes and by increased abundance of clones with integrations near MECOM and LMO2 These clones showed stable behavior over multiple years and never expanded to the point of dominance or dysplasia. One patient developed a benign clonal dominance that could not be attributed to insertional mutagenesis and instead likely resulted from expansion of a transduced natural killer clone in response to chronic Epstein-Barr virus viremia. Clonal diversity and T-cell repertoire, measured by vector integration site sequencing and T-cell receptor β-chain rearrangement sequencing, correlated significantly with the amount of busulfan preconditioning delivered to patients and to CD34 + cell dose. These data, in combination with results of other ADA-SCID gene therapy trials, suggest that disease background may be a crucial factor in leukemogenic potential of retroviral gene therapy and underscore the importance of cytoreductive conditioning in this type of gene therapy approach.

Molecular typing of avian pathogenic Escherichia coli colonies originating from outbreaks of E. coli peritonitis syndrome in chicken flocks.

PubMed

Landman, W J M; Buter, G J; Dijkman, R; van Eck, J H H

2014-01-01

Escherichia coli colonies isolated from the bone marrow of fresh dead hens of laying flocks with the E. coli peritonitis syndrome (EPS) were genotyped using pulsed-field gel electrophoresis (PFGE). Typing is important from an epidemiological point of view and also if the use of autogenous (auto)vaccines is considered. Birds with EPS originated from one house of each of three layer farms and one broiler breeder farm. Farms were considered as separate epidemiological units. In total, six flocks were examined including two successive flocks of one layer farm and the broiler breeder farm. E. coli colonies (one per bird) from nine to 16 hens of each flock were genotyped. The clonality of E. coli within birds was studied using five colonies of each of nine to 14 birds per flock. E. coli genotypes, which totalled 15, differed between farms and flocks except for two successive layer flocks that shared three genotypes. One to five genotypes were found per flock with one or two genotypes dominating each outbreak. Within hens, E. coli bacteria were always clonal. Colonies of the same PFGE type always had the same multilocus sequence type. However, four PFGE types shared sequence type 95. Neither PFGE types nor multilocus sequence types were unambiguously related to avian pathogenic E. coli from EPS. In cases where persistence of E. coli strains associated with EPS is found to occur frequently, routine genotyping to select strains for autovaccines should be considered.

Acute Leukemia with a Translocation T(4;11)(q21;q23): a Distinct Clinicopathological Entity: Report of a Case with Cytogenetic Clonal Evolution and Review of 146 Cases of the Literature.

PubMed

Léglise, M C; Rivière, D; Brière, J

1990-01-01

We present a cytogenetic clonal evolution that correlates morphological and immunological shifts in a case of a patient with a t(4;11) (q21;q23) acute leukemia. We take this opportunity to review 146 cases reported so far, with special reference to morphology, immunophenotyping, cytogenetics, clinical characteristics and evolution. Particular features are underlined, and prognosis, leukemic stem cell origin, chromosomal breakpoints and genes involved are discussed. A relationship between this type of leukemia and exposure to carcinogens is suggested by a high rate of secondary leukemia in adults and a high frequency in newborns and infants.

Molecular Epidemiology of Staphylococcus aureus in the General Population in Northeast Germany: Results of the Study of Health in Pomerania (SHIP-TREND-0)

PubMed Central

Holtfreter, Silva; Grumann, Dorothee; Balau, Veronika; Barwich, Annette; Kolata, Julia; Goehler, André; Weiss, Stefan; Holtfreter, Birte; Bauerfeind, Stephanie S.; Döring, Paula; Friebe, Erika; Haasler, Nicole; Henselin, Kristin; Kühn, Katrin; Nowotny, Sophie; Radke, Dörte; Schulz, Katrin; Schulz, Sebastian R.; Trübe, Patricia; Vu, Chi Hai; Walther, Birgit; Westphal, Susanne; Cuny, Christiane; Witte, Wolfgang; Völzke, Henry; Grabe, Hans Jörgen; Kocher, Thomas; Steinmetz, Ivo

2016-01-01

Population-based studies on Staphylococcus aureus nasal colonization are scarce. We examined the prevalence, resistance, and molecular diversity of S. aureus in the general population in Northeast Germany. Nasal swabs were obtained from 3,891 adults in the large-scale population-based Study of Health in Pomerania (SHIP-TREND). Isolates were characterized using spa genotyping, as well as antibiotic resistance and virulence gene profiling. We observed an S. aureus prevalence of 27.2%. Nasal S. aureus carriage was associated with male sex and inversely correlated with age. Methicillin-resistant S. aureus (MRSA) accounted for 0.95% of the colonizing S. aureus strains. MRSA carriage was associated with frequent visits to hospitals, nursing homes, or retirement homes within the previous 24 months. All MRSA strains were resistant to multiple antibiotics. Most MRSA isolates belonged to the pandemic European hospital-acquired MRSA sequence type 22 (HA-MRSA-ST22) lineage. We also detected one livestock-associated MRSA ST398 (LA-MRSA-ST398) isolate, as well as six livestock-associated methicillin-susceptible S. aureus (LA-MSSA) isolates (clonal complex 1 [CC1], CC97, and CC398). spa typing revealed a diverse but also highly clonal S. aureus population structure. We identified a total of 357 spa types, which were grouped into 30 CCs or sequence types. The major seven CCs (CC30, CC45, CC15, CC8, CC7, CC22, and CC25) included 75% of all isolates. Virulence gene patterns were strongly linked to the clonal background. In conclusion, MSSA and MRSA prevalences and the molecular diversity of S. aureus in Northeast Germany are consistent with those of other European countries. The detection of HA-MRSA and LA-MRSA within the general population indicates possible transmission from hospitals and livestock, respectively, and should be closely monitored. PMID:27605711

Cronobacter, the emergent bacterial pathogen Enterobacter sakazakii comes of age; MLST and whole genome sequence analysis.

PubMed

Forsythe, Stephen J; Dickins, Benjamin; Jolley, Keith A

2014-12-16

Following the association of Cronobacter spp. to several publicized fatal outbreaks in neonatal intensive care units of meningitis and necrotising enterocolitis, the World Health Organization (WHO) in 2004 requested the establishment of a molecular typing scheme to enable the international control of the organism. This paper presents the application of Next Generation Sequencing (NGS) to Cronobacter which has led to the establishment of the Cronobacter PubMLST genome and sequence definition database (http://pubmlst.org/cronobacter/) containing over 1000 isolates with metadata along with the recognition of specific clonal lineages linked to neonatal meningitis and adult infections Whole genome sequencing and multilocus sequence typing (MLST) has supports the formal recognition of the genus Cronobacter composed of seven species to replace the former single species Enterobacter sakazakii. Applying the 7-loci MLST scheme to 1007 strains revealed 298 definable sequence types, yet only C. sakazakii clonal complex 4 (CC4) was principally associated with neonatal meningitis. This clonal lineage has been confirmed using ribosomal-MLST (51-loci) and whole genome-MLST (1865 loci) to analyse 107 whole genomes via the Cronobacter PubMLST database. This database has enabled the retrospective analysis of historic cases and outbreaks following re-identification of those strains. The Cronobacter PubMLST database offers a central, open access, reliable sequence-based repository for researchers. It has the capacity to create new analysis schemes 'on the fly', and to integrate metadata (source, geographic distribution, clinical presentation). It is also expandable and adaptable to changes in taxonomy, and able to support the development of reliable detection methods of use to industry and regulatory authorities. Therefore it meets the WHO (2004) request for the establishment of a typing scheme for this emergent bacterial pathogen. Whole genome sequencing has additionally shown a range of potential virulence and environmental fitness traits which may account for the association of C. sakazakii CC4 pathogenicity, and propensity for neonatal CNS.

Recent advances in understanding clonal haematopoiesis in aplastic anaemia

PubMed Central

Stanley, Natasha; Olson, Timothy S.; Babushok, Daria V.

2016-01-01

Summary Acquired aplastic anaemia (AA) is an immune-mediated bone marrow failure disorder inextricably linked to clonal haematopoiesis. The majority of AA patients have somatic mutations and/or structural chromosomal abnormalities detected as early as at diagnosis. In contrast to other conditions linked to clonal haematopoiesis, the clonal signature of AA reflects its immune pathophysiology. The most common alterations are clonal expansions of cells lacking glycophosphotidylinositol-anchored proteins, loss of human leucocyte antigen alleles, and mutations in BCOR/BCORL1, ASXL1 and DNMT3A. Here, we present the current knowledge of clonal haematopoiesis in AA as it relates to aging, inherited bone marrow failure, and the grey-zone overlap of AA and myelodysplastic syndrome (MDS). We conclude by discussing the significance of clonal haematopoiesis both for improved diagnosis of AA, as well as for a more precise, personalized approach to prognostication of outcomes and therapy choices. PMID:28107566

Recent advances in understanding clonal haematopoiesis in aplastic anaemia.

PubMed

Stanley, Natasha; Olson, Timothy S; Babushok, Daria V

2017-05-01

Acquired aplastic anaemia (AA) is an immune-mediated bone marrow failure disorder inextricably linked to clonal haematopoiesis. The majority of AA patients have somatic mutations and/or structural chromosomal abnormalities detected as early as at diagnosis. In contrast to other conditions linked to clonal haematopoiesis, the clonal signature of AA reflects its immune pathophysiology. The most common alterations are clonal expansions of cells lacking glycophosphotidylinositol-anchored proteins, loss of human leucocyte antigen alleles, and mutations in BCOR/BCORL1, ASXL1 and DNMT3A. Here, we present the current knowledge of clonal haematopoiesis in AA as it relates to aging, inherited bone marrow failure, and the grey-zone overlap of AA and myelodysplastic syndrome (MDS). We conclude by discussing the significance of clonal haematopoiesis both for improved diagnosis of AA, as well as for a more precise, personalized approach to prognostication of outcomes and therapy choices. © 2017 John Wiley & Sons Ltd.

Non-cell-autonomous driving of tumour growth supports sub-clonal heterogeneity.

PubMed

Marusyk, Andriy; Tabassum, Doris P; Altrock, Philipp M; Almendro, Vanessa; Michor, Franziska; Polyak, Kornelia

2014-10-02

Cancers arise through a process of somatic evolution that can result in substantial sub-clonal heterogeneity within tumours. The mechanisms responsible for the coexistence of distinct sub-clones and the biological consequences of this coexistence remain poorly understood. Here we used a mouse xenograft model to investigate the impact of sub-clonal heterogeneity on tumour phenotypes and the competitive expansion of individual clones. We found that tumour growth can be driven by a minor cell subpopulation, which enhances the proliferation of all cells within a tumour by overcoming environmental constraints and yet can be outcompeted by faster proliferating competitors, resulting in tumour collapse. We developed a mathematical modelling framework to identify the rules underlying the generation of intra-tumour clonal heterogeneity. We found that non-cell-autonomous driving of tumour growth, together with clonal interference, stabilizes sub-clonal heterogeneity, thereby enabling inter-clonal interactions that can lead to new phenotypic traits.

Genetic inactivation of Nrf2 prevents clonal expansion of initiated cells in a nutritional model of rat hepatocarcinogenesis.

PubMed

Orrù, Claudia; Szydlowska, Marta; Taguchi, Keiko; Zavattari, Patrizia; Perra, Andrea; Yamamoto, Masayuki; Columbano, Amedeo

2018-06-21

Dysregulation of the Keap1-Nrf2 pathway has been observed in experimental and human tumors, suggesting possible roles of the pathway in cancer development. Herein, we examined whether Nrf2 (Nfe2l2) activation occurs at early steps of rat hepatocarcinogenesis, to assess critical contributions of Nrf2 to the onset of hepatocellular carcinoma (HCC). We used wild-type (WT) and Nrf2 knockout (Nrf2KO) rats treated with a single injection of diethylnitrosamine (DENA) followed by choline-devoid methionine-deficient (CMD) diet. This experimental model causes massive fatty liver and steatohepatitis with fibrosis and enables identification of early stages of hepatocarcinogenesis. We found that Nrf2 activation takes place in early preneoplastic lesions identified by the marker glutathione S-transferase placental form (GSTP). Nrf2 missense mutations, known to disrupt the Keap1-Nrf2 binding, were present in 65.7% of GSTP-positive foci. Nrf2KO rats were used to directly investigate whether Nrf2 is critical for initiation and/or clonal expansion of DENA-damaged hepatocytes. While Nrf2 genetic inactivation did not alter DENA-induced initiation, it led to increased liver injury and chronic compensatory hepatocyte regeneration when rats were fed a CMD diet. However, in spite of such a permissive environment, the livers of Nrf2KO rats did not display any preneoplastic lesion unlike those of WT rats. These results demonstrate that, in a model of hepatocarcinogenesis resembling human non-alcoholic fatty liver disease: i) Nrf2 is activated at early steps of the tumorigenic process and ii) Nrf2 is mandatory for the clonal expansion of initiated cells, indicating that Nrf2 is critical in the onset of HCC. Dysregulation of the Keap1-Nrf2 molecular pathway has been observed in human tumors. In a nutritional model of hepatocarcinogenesis, the protein Nrf2 is frequently mutated/activated at early steps of the tumorigenic process. Herein, we show that Nrf2 is mandatory for the development of preneoplastic lesions. These results suggest that Nrf2 has a critical role in the onset of hepatocellular carcinoma. Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Clonality Testing in Veterinary Medicine: A Review With Diagnostic Guidelines.

PubMed

Keller, S M; Vernau, W; Moore, P F

2016-07-01

The accurate distinction of reactive and neoplastic lymphoid proliferations can present challenges. Given the different prognoses and treatment strategies, a correct diagnosis is crucial. Molecular clonality assays assess rearranged lymphocyte antigen receptor gene diversity and can help differentiate reactive from neoplastic lymphoid proliferations. Molecular clonality assays are commonly used to assess atypical, mixed, or mature lymphoid proliferations; small tissue fragments that lack architecture; and fluid samples. In addition, clonality testing can be utilized to track neoplastic clones over time or across anatomic sites. Molecular clonality assays are not stand-alone tests but useful adjuncts that follow clinical, morphologic, and immunophenotypic assessment. Even though clonality testing provides valuable information in a variety of situations, the complexities and pitfalls of this method, as well as its dependency on the experience of the interpreter, are often understated. In addition, a lack of standardized terminology, laboratory practices, and interpretational guidelines hinders the reproducibility of clonality testing across laboratories in veterinary medicine. The objectives of this review are twofold. First, the review is intended to familiarize the diagnostic pathologist or interested clinician with the concepts, potential pitfalls, and limitations of clonality testing. Second, the review strives to provide a basis for future harmonization of clonality testing in veterinary medicine by providing diagnostic guidelines. © The Author(s) 2016.

Synthesis of clonality and polyploidy in vertebrate animals by hybridization between two sexual species.

PubMed

Choleva, Lukáš; Janko, Karel; De Gelas, Koen; Bohlen, Jörg; Šlechtová, Věra; Rábová, Marie; Ráb, Petr

2012-07-01

Because most clonal vertebrates have hybrid genomic constitutions, tight linkages are assumed among hybridization, clonality, and polyploidy. However, predictions about how these processes mechanistically relate during the switch from sexual to clonal reproduction have not been validated. Therefore, we performed a crossing experiment to test the hypothesis that interspecific hybridization per se initiated clonal diploid and triploid spined loaches (Cobitis) and their gynogenetic reproduction. We reared two F1 families resulting from the crossing of 14 pairs of two sexual species, and found their diploid hybrid constitution and a 1:1 sex ratio. While males were infertile, females produced unreduced nonrecombinant eggs (100%). Synthetic triploid females and males (96.3%) resulted in each of nine backcrossed families from eggs of synthesized diploid F1s fertilized by haploid sperm from sexual males. Five individuals (3.7%) from one backcross family were genetically identical to the somatic cells of the mother and originated via gynogenesis; the sperm of the sexual male only triggered clonal development of the egg. Our reconstruction of the evolutionary route from sexuality to clonality and polyploidy in these fish shows that clonality and gynogenesis may have been directly triggered by interspecific hybridization and that polyploidy is a consequence, not a cause, of clonality. © 2012 The Author(s).

Phenotypic profile of expanded NK cells in chronic lymphoproliferative disorders: a surrogate marker for NK-cell clonality

PubMed Central

Bárcena, Paloma; Jara-Acevedo, María; Tabernero, María Dolores; López, Antonio; Sánchez, María Luz; García-Montero, Andrés C.; Muñoz-García, Noemí; Vidriales, María Belén; Paiva, Artur; Lecrevisse, Quentin; Lima, Margarida; Langerak, Anton W.; Böttcher, Sebastian; van Dongen, Jacques J.M.

2015-01-01

Currently, the lack of a universal and specific marker of clonality hampers the diagnosis and classification of chronic expansions of natural killer (NK) cells. Here we investigated the utility of flow cytometric detection of aberrant/altered NK-cell phenotypes as a surrogate marker for clonality, in the diagnostic work-up of chronic lymphoproliferative disorders of NK cells (CLPD-NK). For this purpose, a large panel of markers was evaluated by multiparametric flow cytometry on peripheral blood (PB) CD56low NK cells from 60 patients, including 23 subjects with predefined clonal (n = 9) and polyclonal (n = 14) CD56low NK-cell expansions, and 37 with CLPD-NK of undetermined clonality; also, PB samples from 10 healthy adults were included. Clonality was established using the human androgen receptor (HUMARA) assay. Clonal NK cells were found to show decreased expression of CD7, CD11b and CD38, and higher CD2, CD94 and HLADR levels vs. normal NK cells, together with a restricted repertoire of expression of the CD158a, CD158b and CD161 killer-associated receptors. In turn, NK cells from both clonal and polyclonal CLPD-NK showed similar/overlapping phenotypic profiles, except for high and more homogeneous expression of CD94 and HLADR, which was restricted to clonal CLPD-NK. We conclude that the CD94hi/HLADR+ phenotypic profile proved to be a useful surrogate marker for NK-cell clonality. PMID:26556869

Natural and Chemotherapy-Induced Clonal Evolution of Tumors.

PubMed

Ibragimova, M K; Tsyganov, M M; Litviakov, N V

2017-04-01

Evolution and natural selection of tumoral clones in the process of transformation and the following carcinogenesis can be called natural clonal evolution. Its main driving factors are internal: genetic instability initiated by driver mutations and microenvironment, which enables selective pressure while forming the environment for cell transformation and their survival. We present our overview of contemporary research dealing with mechanisms of carcinogenesis in different localizations from precancerous pathologies to metastasis and relapse. It shows that natural clonal evolution establishes intratumoral heterogeneity and enables tumor progression. Tumors of monoclonal origin are of low-level intratumoral heterogeneity in the initial stages, and this increases with the size of the tumor. Tumors of polyclonal origin are of extremely high-level intratumoral heterogeneity in the initial stages and become more homogeneous when larger due to clonal expansion. In cases of chemotherapy-induced clonal evolution of a tumor, chemotherapy becomes the leading factor in treatment. The latest research shows that the impact of chemotherapy can radically increase the speed of clonal evolution and lead to new malignant and resistant clones that cause tumor metastasis. Another option of chemotherapy-induced clonal evolution is formation of a new dominant clone from a clone that was minor in the initial tumor and obtained free space due to elimination of sensitive clones by chemotherapy. As a result, in ~20% of cases, chemotherapy can stimulate metastasis and relapse of tumors due to clonal evolution. The conclusion of the overview formulates approaches to tumor treatment based on clonal evolution: in particular, precision therapy, prediction of metastasis stimulation in patients treated with chemotherapy, methods of genetic evaluation of chemotherapy efficiency and clonal-oriented treatment, and approaches to manipulating the clonal evolution of tumors are presented.

ClonEvol: clonal ordering and visualization in cancer sequencing.

PubMed

Dang, H X; White, B S; Foltz, S M; Miller, C A; Luo, J; Fields, R C; Maher, C A

2017-12-01

Reconstruction of clonal evolution is critical for understanding tumor progression and implementing personalized therapies. This is often done by clustering somatic variants based on their cellular prevalence estimated via bulk tumor sequencing of multiple samples. The clusters, consisting of the clonal marker variants, are then ordered based on their estimated cellular prevalence to reconstruct clonal evolution trees, a process referred to as 'clonal ordering'. However, cellular prevalence estimate is confounded by statistical variability and errors in sequencing/data analysis, and therefore inhibits accurate reconstruction of the clonal evolution. This problem is further complicated by intra- and inter-tumor heterogeneity. Furthermore, the field lacks a comprehensive visualization tool to facilitate the interpretation of complex clonal relationships. To address these challenges we developed ClonEvol, a unified software tool for clonal ordering, visualization, and interpretation. ClonEvol uses a bootstrap resampling technique to estimate the cellular fraction of the clones and probabilistically models the clonal ordering constraints to account for statistical variability. The bootstrapping allows identification of the sample founding- and sub-clones, thus enabling interpretation of clonal seeding. ClonEvol automates the generation of multiple widely used visualizations for reconstructing and interpreting clonal evolution. ClonEvol outperformed three of the state of the art tools (LICHeE, Canopy and PhyloWGS) for clonal evolution inference, showing more robust error tolerance and producing more accurate trees in a simulation. Building upon multiple recent publications that utilized ClonEvol to study metastasis and drug resistance in solid cancers, here we show that ClonEvol rediscovered relapsed subclones in two published acute myeloid leukemia patients. Furthermore, we demonstrated that through noninvasive monitoring ClonEvol recapitulated the emerging subclones throughout metastatic progression observed in the tumors of a published breast cancer patient. ClonEvol has broad applicability for longitudinal monitoring of clonal populations in tumor biopsies, or noninvasively, to guide precision medicine. ClonEvol is written in R and is available at https://github.com/ChrisMaherLab/ClonEvol. © The Author 2017. Published by Oxford University Press on behalf of the European Society for Medical Oncology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Clustering of Tuberculosis Cases Based on Variable-Number Tandem-Repeat Typing in Relation to the Population Structure of Mycobacterium tuberculosis in the Netherlands

PubMed Central

Sloot, Rosa; Borgdorff, Martien W.; de Beer, Jessica L.; van Ingen, Jakko; Supply, Philip

2013-01-01

The population structure of 3,776 Mycobacterium tuberculosis isolates was determined using variable-number tandem-repeat (VNTR) typing. The degree of clonality was so high that a more relaxed definition of clustering cannot be applied. Among recent immigrants with non-Euro-American isolates, transmission is overestimated if based on identical VNTR patterns. PMID:23658260

Capsule null locus meningococci: typing of antigens used in an investigational multicomponent meningococcus serogroup B vaccine.

PubMed

Claus, Heike; Jördens, Markus S; Kriz, Pavla; Musilek, Martin; Jarva, Hanna; Pawlik, Marie-Christin; Meri, Seppo; Vogel, Ulrich

2012-01-05

The investigational multicomponent meningococcus serogroup B vaccine (4CMenB) targets the antigenetically variable population of serogroup B meningococci. Forty-one strains of capsule null locus (cnl) meningococci, which are frequent among healthy carriers, were selected from nine sequence types (ST), which belong to four clonal complexes (cc), and three countries. They were antigen sequence typed and analyzed for antigen expression to predict whether these strains harbor the genes and express the four vaccine antigens of 4CMenB as measured by the meningococcal antigen typing system (MATS). The PorA variant used in the vaccine was not found. The nadA gene was absent in all but one strain, which did not express the antigen in vitro. Only strains of clonal complex ST-198 harbored a factor H binding protein (FHBP) allele of the cross-reactive variant 1 family which is included in the vaccine. All these strains expressed the antigen. Five variants of the Neisserial heparin binding antigen (NHBA) gene were identified. Expression of NHBA was observed in all strains with highest levels in ST-198 cc and ST-845. The data suggest a potential impact of 4CMenB immunization at least on cnl meningococci of the ST-198 cc and ST-845. Copyright © 2011 Elsevier Ltd. All rights reserved.

Genetic diversity and antibiotic susceptibility of Staphylococcus aureus isolates from wild boars.

PubMed

Seinige, D; Von Altrock, A; Kehrenberg, C

2017-10-01

We here report the occurrence of S. aureus in wild boars and characterize isolates genotypically and phenotypically in order to get knowledge about the occurrence of clonal lineages and genotypes in free-living wild animals. Forty-one S. aureus isolates obtained from 111 wild boars hunted in Lower Saxony, Germany, were investigated and compared to human and livestock isolates. The S. aureus belonged to multilocus sequence types ST1, ST7, ST30, ST133, ST425, ST804, ST890 and to the new ST3237, ST3238, ST3255 and ST3369. The livestock associated CC398-MRSA lineage, however, was not found. In addition to well-known spa types, the new types t14999, t15000, t15001 and t15002 were detected. Macrorestriction analysis revealed a variety of different SmaI fragment patterns. Most isolates were susceptible to all antimicrobials tested, including methicillin, and resistance was detected only to ampicillin, penicillin and erythromycin. PCR analysis confirmed the presence of staphylococcal enterotoxin genes (seh) in all t127-ST1 isolates. A high degree of genetic diversity was detected with many spa types and clonal lineages previously reported in humans and livestock animals. Copyright © 2017 Elsevier Ltd. All rights reserved.

V beta-specific deletion of mature thymocytes induced by the plant superantigen Urtica dioica agglutinin.

PubMed

Delcourt, M; Peumans, W J; Wagner, M C; Truffa-Bachi, P

1996-03-15

Urtica dioica agglutinin (UDA), a plant protein, is a superantigen activating in a MHC class II-restricted manner the V beta 8. 3-bearing T-cells (Galelli and Truffa-Bachi, J. Immunol. 151, 1821, 1993). Administration of UDA to adult mice provokes the clonal expansion of the responding cells which is followed by the deletion of the major fraction of the UDA-sensitive cells, whereas the remaining cells become anergic (Galelli et al., J. Immunol. 154, 2600, 1995). We have analyzed the effect of UDA on thymocytes. Injection of UDA resulted in a rapid, but transient, deletion of a large fraction of the V beta 8.3-bearing mature T-cells. In contrast to other exogenous superantigens, this deletion was not preceded by the clonal expansion of the UDA-responding thymocytes. Moreover, the V beta 8.3-bearing mature T-cells escaping the deletion were not anergic to an in vitro UDA restimulation. UDA and the other superantigens also differ as the general, V beta-unrestricted, thymic atrophy induced by classical superantigens was not observed with UDA.

Diagnostic Applications of Next Generation Sequencing in Immunogenetics and Molecular Oncology

PubMed Central

Grumbt, Barbara; Eck, Sebastian H.; Hinrichsen, Tanja; Hirv, Kaimo

2013-01-01

Summary With the introduction of the next generation sequencing (NGS) technologies, remarkable new diagnostic applications have been established in daily routine. Implementation of NGS is challenging in clinical diagnostics, but definite advantages and new diagnostic possibilities make the switch to the technology inevitable. In addition to the higher sequencing capacity, clonal sequencing of single molecules, multiplexing of samples, higher diagnostic sensitivity, workflow miniaturization, and cost benefits are some of the valuable features of the technology. After the recent advances, NGS emerged as a proven alternative for classical Sanger sequencing in the typing of human leukocyte antigens (HLA). By virtue of the clonal amplification of single DNA molecules ambiguous typing results can be avoided. Simultaneously, a higher sample throughput can be achieved by tagging of DNA molecules with multiplex identifiers and pooling of PCR products before sequencing. In our experience, up to 380 samples can be typed for HLA-A, -B, and -DRB1 in high-resolution during every sequencing run. In molecular oncology, NGS shows a markedly increased sensitivity in comparison to the conventional Sanger sequencing and is developing to the standard diagnostic tool in detection of somatic mutations in cancer cells with great impact on personalized treatment of patients. PMID:23922545

Processing-Dependent and Clonal Contamination Patterns of Listeria monocytogenes in the Cured Ham Food Chain Revealed by Genetic Analysis.

PubMed

Morganti, Marina; Scaltriti, Erika; Cozzolino, Paolo; Bolzoni, Luca; Casadei, Gabriele; Pierantoni, Marco; Foni, Emanuela; Pongolini, Stefano

2016-02-01

The quantitative and qualitative patterns of environmental contamination by Listeria monocytogenes were investigated in the production chain of dry-cured Parma ham. Standard arrays of surfaces were sampled in processing facilities during a single visit per plant in the three compartments of the food chain, i.e., ham production (19 plants) and postproduction, which was divided into deboning (43 plants) and slicing (25 plants) steps. The numbers of sampled surfaces were 384 in ham production, with 25 positive for L. monocytogenes, and 1,084 in postproduction, with 83 positives. Statistical analysis of the prevalence of contaminated surfaces showed that in ham production, contamination was higher at the beginning of processing and declined significantly toward the end, while in postproduction, prevalence rose toward the end of processing. Prevalence was higher in the deboning facilities than in slicing facilities and was dependent on the type of surface (floor/drainage > clothing > equipment). The qualitative pattern of contamination was investigated through an analysis of the survey isolates and a set of isolates derived from routine monitoring, including longitudinal isolations. Pulsed-field gel electrophoresis (PFGE) and whole-genome single-nucleotide polymorphism (SNP) analysis revealed a remarkable clonality of L. monocytogenes within plants, with the detection of 16 plant-specific clones out of 17 establishments with multiple isolates. Repeated detections of clonal isolates >6 months apart were also observed. Six was the maximum number of between-isolate differences in core SNPs observed within these clones. Based on the same six-SNP threshold, three clusters of clonal isolates, shared by six establishments, were also identified. The spread of L. monocytogenes within and between plants, as indicated by its clonal behavior, is a matter of concern for the hygienic management of establishments. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

NetF-producing Clostridium perfringens: Clonality and plasmid pathogenicity loci analysis.

PubMed

Mehdizadeh Gohari, Iman; Kropinski, Andrew M; Weese, Scott J; Whitehead, Ashley E; Parreira, Valeria R; Boerlin, Patrick; Prescott, John F

2017-04-01

Clostridium perfringens is an important cause of foal necrotizing enteritis and canine acute hemorrhagic diarrhea. A major virulence determinant of the strains associated with these diseases appears to be a beta-sheet pore-forming toxin, NetF, encoded within a pathogenicity locus (NetF locus) on a large tcp-conjugative plasmid. Strains producing NetF also produce the putative toxin NetE, encoded within the same pathogenicity locus, as well as CPE enterotoxin and CPB2 on a second plasmid, and sometimes the putative toxin NetG within a pathogenicity locus (NetG locus) on another separate large conjugative plasmid. Previous genome sequences of two netF-positive C. perfringens showed that they both shared three similar plasmids, including the NetF/NetE and CPE/CPB2 toxins-encoding plasmids mentioned above and a putative bacteriocin-encoding plasmid. The main purpose of this study was to determine whether all NetF-producing strains share this common plasmid profile and whether their distinct NetF and CPE pathogenicity loci are conserved. To answer this question, 15 equine and 15 canine netF-positive isolates of C. perfringens were sequenced using Illumina Hiseq2000 technology. In addition, the clonal relationships among the NetF-producing strains were evaluated by core genome multilocus sequence typing (cgMLST). The data obtained showed that all NetF-producing strains have a common plasmid profile and that the defined pathogenicity loci on the plasmids are conserved in all these strains. cgMLST analysis showed that the NetF-producing C. perfringens strains belong to two distinct clonal complexes. The pNetG plasmid was absent from isolates of one of the clonal complexes, and there were minor but consistent differences in the NetF/NetE and CPE/CPB2 plasmids between the two clonal complexes. Copyright © 2017 Elsevier B.V. All rights reserved.

Staphylococcus aureus from the German general population is highly diverse.

PubMed

Becker, Karsten; Schaumburg, Frieder; Fegeler, Christian; Friedrich, Alexander W; Köck, Robin

2017-01-01

This prospective cohort study evaluates colonization dynamics and molecular characteristics of methicillin-susceptible and - resistant Staphylococcus aureus (MSSA/MRSA) in a German general population. Nasal swabs of 1878 non-hospitalized adults were screened for S. aureus. Participants were screened thrice in intervals of 6-8 months. Isolates were characterized by spa and agr typing, mecA and mecC possession, respectively, and PCRs targeting virulence factors. 40.9% of all participants carried S. aureus at least once while 0.7% of the participants carried MRSA (mainly spa t011). MSSA isolates (n=1359) were associated with 331 different spa types; t084 (7.7%), t091 (6.1%) and t012 (71, 5.2%) were predominant. Of 206 participants carrying S. aureus at all three sampling time points, 14.1% carried the same spa type continuously; 5.3% carried different spa types with similar repeat patterns, but 80.6% carried S. aureus with unrelated spa types. MSSA isolates frequently harboured genes encoding enterotoxins (sec: 16.6%, seg: 63.1%, sei: 64.5%) and toxic shock syndrome toxin (tst: 17.5%), but rarely Panton-Valentine leukocidin (lukS-PV/lukF-PV: 0.2%). MSSA colonizing human nares in the community are clonally highly diverse. Among those constantly carrying S. aureus, clonal lineages changed over time. The proportion of persistent S. aureus carriers was lower than reported elsewhere. Copyright © 2016 Elsevier GmbH. All rights reserved.

The Emergence and Spread of Multiple Livestock-Associated Clonal Complex 398 Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus Strains among Animals and Humans in the Republic of Ireland, 2010–2014

PubMed Central

Brennan, Gráinne I.; Abbott, Yvonne; Burns, Aisling; Leonard, Finola; McManus, Brenda A.; O’Connell, Brian; Coleman, David C.; Shore, Anna C.

2016-01-01

Clonal complex (CC) 398 methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) are associated with carriage and infection among animals and humans but only a single case of CC398 MRSA has been reported in the Republic of Ireland (ROI). The present study investigated the molecular epidemiology of CC398 MRSA (n = 22) and MSSA (n = 10) from animals and humans in the ROI from 2010–2014. Isolates underwent antimicrobial susceptibility testing, spa typing, DNA microarray profiling and PCR for CC398-associated resistance genes. All MRSA underwent SCCmec IV or V subtyping. Four distinct CC398-MRSA incidents were identified from (i) a man in a nursing home (spa type t011-SCCmec IVa, immune evasion complex (IEC) negative), (ii) a horse and veterinarian who had recently travelled to Belgium (t011-IVa, IEC positive), (iii) pigs (n = 9) and farm workers (n = 9) on two farms, one which had been restocked with German gilts and the other which was a finisher farm (t034-VT, IEC negative, 3/9 pigs; t011- VT, IEC negative, 6/9 pigs & 9/9 farm workers), and (iv) a child who had worked on a pig farm in the UK (t034-VT, IEC negative). Isolates also carried different combinations of multiple resistance genes including erm(A), erm(B), tet(K), tet(M) & tet(L), fexA, spc, dfrG, dfrK aacA-aphD and aadD further highlighting the presence of multiple CC398-MRSA strains. CC398 MSSA were recovered from pigs (n = 8) and humans (n = 2). CC398 MSSA transmission was identified among pigs but zoonotic transmission was not detected with animal and human isolates exhibiting clade-specific traits. This study highlights the importation and zoonotic spread of CC398 MRSA in the ROI and the spread of CC398 MSSA among pigs. Increased surveillance is warranted to prevent further CC398 MRSA importation and spread in a country that was considered CC398 MRSA free. PMID:26886749

Lamarck Will Not Lie Down.

ERIC Educational Resources Information Center

Lewin, Roger

1981-01-01

Describes recent research by Edward Steele appearing to support the Lamarckian theory of inheritance. Steele suggests that a mutant somatic cell favored by the environment will undergo clonal expansion. Altered genetic materials from these cells is then picked up by C-type viruses and inserted into the germ line genome. (CS)

Microsatellite alterations as clonal markers for the detection of human cancer.

PubMed Central

Mao, L; Lee, D J; Tockman, M S; Erozan, Y S; Askin, F; Sidransky, D

1994-01-01

Microsatellite instability has been reported to be an important feature of tumors from hereditary nonpolyposis colorectal carcinoma (HNPCC) patients. The recent discovery of genetic instability in small cell lung carcinoma, a neoplasm not associated with HNPCC, led us to investigate the possible presence of microsatellite alterations in other tumor types. We examined 52 microsatellite repeat sequences in the DNA of normal and tumor pairs from 100 head and neck, bladder, and lung cancer patients by the polymerase chain reaction. Although alterations were rare in dinucleotide repeats, larger (tri- or tetranucleotide) repeats were found to be more prone to expansion or deletion. We screened 100 tumors with a panel of nine tri- and tetranucleotide repeat markers and identified 26 (26%) that displayed alterations in at least one locus. This observation prompted us to examine the possibility of using microsatellite alterations as markers to detect clonal tumor-derived cell populations in pathologic samples. The identical microsatellite alterations detected in the primary tumors were successfully identified in corresponding urine, sputum, and surgical margins from affected patients. This study demonstrates that appropriately selected microsatellite loci are commonly altered in many cancers and can serve as clonal markers for their detection. Images PMID:7937908

Community-Associated Methicillin-Resistant Staphylococcus aureus Clonal Complex 80 Type IV (CC80-MRSA-IV) Isolated from the Middle East: A Heterogeneous Expanding Clonal Lineage

PubMed Central

Harastani, Houda H.; Tokajian, Sima T.

2014-01-01

Background The emergence of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) has caused a change in MRSA epidemiology worldwide. In the Middle East, the persistent spread of CA-MRSA isolates that were associated with multilocus sequence type (MLST) clonal complex 80 and with staphylococcal cassette chromosome mec (SCCmec) type IV (CC80-MRSA-IV), calls for novel approaches for infection control that would limit its spread. Methodology/Principal Findings In this study, the epidemiology of CC80-MRSA-IV was investigated in Jordan and Lebanon retrospectively covering the period from 2000 to 2011. Ninety-four S. aureus isolates, 63 (67%) collected from Lebanon and 31 (33%) collected from Jordan were included in this study. More than half of the isolates (56%) were associated with skin and soft tissue infections (SSTIs), and 73 (78%) were Panton-Valentine Leukocidin (PVL) positive. Majority of the isolates (84%) carried the gene for exofoliative toxin d (etd), 19% had the Toxic Shock Syndrome Toxin-1 gene (tst), and seven isolates from Jordan had a rare combination being positive for both tst and PVL genes. spa typing showed the prevalence of type t044 (85%) and pulsed-field gel electrophoresis (PFGE) recognized 21 different patterns. Antimicrobial susceptibility testing showed the prevalence (36%) of a unique resistant profile, which included resistance to streptomycin, kanamycin, and fusidic acid (SKF profile). Conclusions The genetic diversity among the CC80 isolates observed in this study poses an additional challenge to infection control of CA-MRSA epidemics. CA-MRSA related to ST80 in the Middle East was distinguished in this study from the ones described in other countries. Genetic diversity observed, which may be due to mutations and differences in the antibiotic regimens between countries may have led to the development of heterogeneous strains. Hence, it is difficult to maintain “the European CA-MRSA clone” as a uniform clone and it is better to designate as CC80-MRSA-IV isolates. PMID:25078407

Ex Vivo Analysis of Human T Lymphotropic Virus Type 1–Specific CD4+ Cells by Use of a Major Histocompatibility Complex Class II Tetramer Composed of a Neurological Disease–Susceptibility Allele and Its Immunodominant Peptide

PubMed Central

Nose, Hirohisa; Kubota, Ryuji; Seth, Nilufer P.; Goon, Peter K.; Tanaka, Yuetsu; Izumo, Shuji; Usuku, Koichiro; Ohara, Yoshiro; Wucherpfennig, Kai W.; Bangham, Charles R. M.; Osame, Mitsuhiro; Saito, Mineki

2015-01-01

HLA-DRB1*0101 is associated with susceptibility to human T lymphotropic virus type 1 (HTLV-1)–associated myelopathy/tropical spastic paraparesis (HAM/TSP). Here, we used a synthetic tetramer of DRB1*0101 and its epitope peptide to analyze HTLV-1–specific CD4+ T cells ex vivo. The frequency of tetramer+CD4+ T cells was significantly greater in patients with HAM/TSP than in healthy HTLV-1 carriers (HCs) at a given proviral load and correlated with HTLV-1 tax messenger RNA expression in HCs but not in patients with HAM/TSP. These cells displayed an early to intermediate effector memory phenotype and were preferentially infected by HTLV-1. T cell receptor gene analyses of 2 unrelated DRB1*0101-positive patients with HAM/TSP showed similar Vβ repertoires and amino acid motifs in complementarity-determining region 3. Our data suggest that efficient clonal expansion of virus-specific CD4+ T cells in patients with HAM/TSP does not simply reflect higher viral burden but rather reflects a rapid turnover caused by preferential infection and/or in vivo stimulation by major histocompatibility complex–peptide complexes. PMID:18190256

Diversity of Tn1546 in vanA-positive Enterococcus faecium clinical isolates with VanA, VanB, and VanD phenotypes and susceptibility to vancomycin.

PubMed

Cha, J O; Yoo, J I; Kim, H K; Kim, H S; Yoo, J S; Lee, Y S; Jung, Y H

2013-10-01

To investigate diversity in the vanA cluster in Enterococcus faecium isolates from nontertiary hospitals. We identified 43 vanA-positive Ent. faecium isolates, including two vancomycin-susceptible isolates, from hospitals between 2003 and 2006. Of these isolates, >85% were resistant to ampicillin, erythromycin and ciprofloxacin. The vanA cluster was classified into six types using overlapping PCR, but the prototype transposon Tn1546 was not found. Most vanA-positive vancomycin-resistant Enterococcus (VRE) carried IS1216V and belonged to Type III (58·1%) or Type II (20·9%). vanY, vanZ and IS1216V were observed in the left and right ends of Type III with long-range PCR. IS1216V was also observed within vanS and vanX in the two vancomycin-susceptible isolates and in two vancomycin-resistant isolates. No VRE isolates with VanB and VanD phenotypes contained point mutations in vanS, unlike in previous reports. Sequence types (STs) of all isolates belonged to clonal complex 17, and ST78 was predominant. Insertion sequences, especially IS1216V, cause structural variation in the vanA cluster. We report the first observation of vanY and vanZ at the left end of Tn1546 in clinical isolates. This is the first report of the frequency of vancomycin resistance and diversity of Tn1546 in vanA-positive Ent. faecium isolates from nontertiary hospitals. © 2013 The Society for Applied Microbiology.

Clonal diversity analysis using SNP microarray: a new prognostic tool for chronic lymphocytic leukemia.

PubMed

Zhang, Linsheng; Znoyko, Iya; Costa, Luciano J; Conlin, Laura K; Daber, Robert D; Self, Sally E; Wolff, Daynna J

2011-12-01

Chronic lymphocytic leukemia (CLL) is a clinically heterogeneous disease. The methods currently used for monitoring CLL and determining conditions for treatment are limited in their ability to predict disease progression, patient survival, and response to therapy. Although clonal diversity and the acquisition of new chromosomal abnormalities during the disease course (clonal evolution) have been associated with disease progression, their prognostic potential has been underappreciated because cytogenetic and fluorescence in situ hybridization (FISH) studies have a restricted ability to detect genomic abnormalities and clonal evolution. We hypothesized that whole genome analysis using high resolution single nucleotide polymorphism (SNP) microarrays would be useful to detect diversity and infer clonal evolution to offer prognostic information. In this study, we used the Infinium Omni1 BeadChip (Illumina, San Diego, CA) array for the analysis of genetic variation and percent mosaicism in 25 non-selected CLL patients to explore the prognostic value of the assessment of clonal diversity in patients with CLL. We calculated the percentage of mosaicism for each abnormality by applying a mathematical algorithm to the genotype frequency data and by manual determination using the Simulated DNA Copy Number (SiDCoN) tool, which was developed from a computer model of mosaicism. At least one genetic abnormality was identified in each case, and the SNP data was 98% concordant with FISH results. Clonal diversity, defined as the presence of two or more genetic abnormalities with differing percentages of mosaicism, was observed in 12 patients (48%), and the diversity correlated with the disease stage. Clonal diversity was present in most cases of advanced disease (Rai stages III and IV) or those with previous treatment, whereas 9 of 13 patients without detected clonal diversity were asymptomatic or clinically stable. In conclusion, SNP microarray studies with simultaneous evaluation of genomic alterations and mosaic distribution of clones can be used to assess apparent clonal evolution via analysis of clonal diversity. Since clonal evolution in CLL is strongly correlated with disease progression, whole genome SNP microarray analysis provides a new comprehensive and reliable prognostic tool for CLL patients. Copyright © 2011 Elsevier Inc. All rights reserved.

Protection, pathogenesis and phenotypic plasticity in Plasmodium falciparum malaria.

PubMed

Roberts, D J; Biggs, B A; Brown, G; Newbold, C I

1993-08-01

Why does Plasmodium falciparum cause severe illness in some but not all infections? How is clinical immunity acquired? These questions have intrigued investigators since the clinical epidemiology of malaria was first described. The search for answers to both questions has highlighted the changes that take place at the surface of infected red blood cells during the last half of the erythrocytic cycle. These changes specify the antigenic and adhesive or cytoadherence phenotypes for the infected cell. Now the antigenic and adhesive phenotypes appear to be linked and together undergo clonal variation. In this article David Roberts, Beverley-Ann Biggs, Graham Brown and Christopher Newbold explain how clonal phenotypic variation and the linkage between adhesive and antigenic types contribute to our understanding of naturally acquired immunity and of pathogenesis of severe malaria.

Demographic consequences of greater clonal than sexual reproduction in Dicentra canadensis.

PubMed

Lin, Chia-Hua; Miriti, Maria N; Goodell, Karen

2016-06-01

Clonality is a widespread life history trait in flowering plants that may be essential for population persistence, especially in environments where sexual reproduction is unpredictable. Frequent clonal reproduction, however, could hinder sexual reproduction by spatially aggregating ramets that compete with seedlings and reduce inter-genet pollination. Nevertheless, the role of clonality in relation to variable sexual reproduction in population dynamics is often overlooked. We combined population matrix models and pollination experiments to compare the demographic contributions of clonal and sexual reproduction in three Dicentra canadensis populations, one in a well-forested landscape and two in isolated forest remnants. We constructed stage-based transition matrices from 3 years of census data to evaluate annual population growth rates, λ. We used loop analysis to evaluate the relative contribution of different reproductive pathways to λ. Despite strong temporal and spatial variation in seed set, populations generally showed stable growth rates. Although we detected some pollen limitation of seed set, manipulative pollination treatments did not affect population growth rates. Clonal reproduction contributed significantly more than sexual reproduction to population growth in the forest remnants. Only at the well-forested site did sexual reproduction contribute as much as clonal reproduction to population growth. Flowering plants were more likely to transition to a smaller size class with reduced reproductive potential in the following year than similarly sized nonflowering plants, suggesting energy trade-offs between sexual and clonal reproduction at the individual level. Seed production had negligible effects on growth and tuber production of individual plants. Our results demonstrate that clonal reproduction is vital for population persistence in a system where sexual reproduction is unpredictable. The bias toward clonality may be driven by low fitness returns for resource investment in sexual reproduction at the individual level. However, chronic failure in sexual reproduction may exacerbate the imbalance between sexual and clonal reproduction and eventually lead to irreversible loss of sex in the population.

Evolution of proliferation and the angiogenic switch in tumors with high clonal diversity.

PubMed

Bickel, Scott T; Juliano, Joseph D; Nagy, John D

2014-01-01

Natural selection among tumor cell clones is thought to produce hallmark properties of malignancy. Efforts to understand evolution of one such hallmark--the angiogenic switch--has suggested that selection for angiogenesis can "run away" and generate a hypertumor, a form of evolutionary suicide by extreme vascular hypo- or hyperplasia. This phenomenon is predicted by models of tumor angiogenesis studied with the techniques of adaptive dynamics. These techniques also predict that selection drives tumor proliferative potential towards an evolutionarily stable strategy (ESS) that is also convergence-stable. However, adaptive dynamics are predicated on two key assumptions: (i) no more than two distinct clones or evolutionary strategies can exist in the tumor at any given time; and (ii) mutations cause small phenotypic changes. Here we show, using a stochastic simulation, that relaxation of these assumptions has no effect on the predictions of adaptive dynamics in this case. In particular, selection drives proliferative potential towards, and angiogenic potential away from, their respective ESSs. However, these simulations also show that tumor behavior is highly contingent on mutational history, particularly for angiogenesis. Individual tumors frequently grow to lethal size before the evolutionary endpoint is approached. In fact, most tumor dynamics are predicted to be in the evolutionarily transient regime throughout their natural history, so that clinically, the ESS is often largely irrelevant. In addition, we show that clonal diversity as measured by the Shannon Information Index correlates with the speed of approach to the evolutionary endpoint. This observation dovetails with results showing that clonal diversity in Barrett's esophagus predicts progression to malignancy.

Reproductive system, social organization, human disturbance and ecological dominance in native populations of the little fire ant, Wasmannia auropunctata.

PubMed

Foucaud, Julien; Orivel, Jérôme; Fournier, Denis; Delabie, Jacques H C; Loiseau, Anne; Le Breton, Julien; Cerdan, Philippe; Estoup, Arnaud

2009-12-01

The invasive ant species Wasmannia auropunctata displays both ecologically dominant and non-dominant populations within its native range. Three factors could theoretically explain the ecological dominance of some native populations of W. auropunctata: (i) its clonal reproductive system, through demographic and/or adaptive advantages; (ii) its unicolonial social organization, through lower intraspecific and efficient interspecific competition; (iii) the human disturbance of its native range, through the modification of biotic and abiotic environmental conditions. We used microsatellite markers and behavioural tests to uncover the reproductive modes and social organization of dominant and non-dominant native populations in natural and human-modified habitats. Microsatellite and mtDNA data indicated that dominant and non-dominant native populations (supercolonies as determined by aggression tests) of W. auropunctata did not belong to different evolutionary units. We found that the reproductive system and the social organization are neither necessary nor sufficient to explain W. auropunctata ecological dominance. Dominance rather seems to be set off by unknown ecological factors altered by human activities, as all dominant populations were recorded in human-modified habitats. The clonal reproductive system found in some populations of W. auropunctata may however indirectly contribute to its ecological dominance by allowing the species to expand its environmental niche, through the fixation over time of specific combinations of divergent male and female genotypes. Unicoloniality may rather promote the range expansion of already dominant populations than actually trigger ecological dominance. The W. auropunctata model illustrates the strong impact of human disturbance on species' ecological features and the adaptive potential of clonal reproductive systems.

Population genetics of reef coral endosymbionts (Symbiodinium, Dinophyceae).

PubMed

Thornhill, D J; Howells, E J; Wham, D C; Steury, T D; Santos, S R

2017-05-01

Symbiodinium is a diverse genus of unicellular dinoflagellate symbionts associating with various marine protists and invertebrates. Although the broadscale diversity and phylogenetics of the Symbiodinium complex is well established, there have been surprisingly few data on fine-scale population structure and biogeography of these dinoflagellates. Yet population-level processes contribute strongly to the biology of Symbiodinium, including how anthropogenic-driven global climate change impacts these symbionts and their host associations. Here, we present a synthesis of population-level characteristics for Symbiodinium, with an emphasis on how phylogenetic affinities, dynamics within and among host individuals, and a propensity towards clonality shape patterns on and across reefs. Major inferences include the following: (i) Symbiodinium populations within individual hosts are comprised mainly of cells belonging to a single or few genetic clones. (ii) Symbiont populations exhibit a mixed mode of reproduction, wherein at least one sexual recombination event occurs in the genealogy between most genotypes, but clonal propagation predominates overall. (iii) Mutualistic Symbiodinium do not perpetually persist outside their hosts, instead undergoing turnover and replacement via the continuous shedding of viable clonal cells from host individuals. (iv) Symbiont populations living in the same host, but on different reefs, are often genetically subdivided, suggesting low connectivity, adaptation to local conditions, or prolific asexual reproduction and low effective population sizes leading to disproportionate success within and among hosts. Overall, this synthesis forms a basis for future investigations of coral symbiosis ecology and evolution as well as delimitation of species boundaries in Symbiodinium and other eukaryotic microorganisms. © 2017 John Wiley & Sons Ltd.

Clonal integration facilitates spread of Paspalum paspaloides from terrestrial to cadmium-contaminated aquatic habitats.

PubMed

Luo, F-L; Xing, Y-P; Wei, G-W; Li, C-Y; Yu, F-H

2017-11-01

Cadmium (Cd) is a hazardous environmental pollutant with high toxicity to plants, which has been detected in many wetlands. Clonal integration (resource translocation) between connected ramets of clonal plants can increase their tolerance to stress. We hypothesised that clonal integration facilitates spread of amphibious clonal plants from terrestrial to Cd-contaminated aquatic habitats. The spread of an amphibious grass Paspalum paspaloides was simulated by growing basal older ramets in uncontaminated soil connected (allowing integration) or not connected (preventing integration) to apical younger ramets of the same fragments in Cd-contaminated water. Cd contamination of apical ramets of P. paspaloides markedly decreased growth and photosynthetic capacity of the apical ramets without connection to the basal ramets, but did not decrease these properties with connection. Cd contamination did not affect growth of the basal ramets without connection to the apical ramets, but Cd contamination of 4 and 12 mg·l -1 significantly increased growth with connection. Consequently, clonal integration increased growth of the apical ramets, basal ramets and whole clones when the apical ramets were grown in Cd-contaminated water of 4 and 12 mg·l -1 . Cd was detected in the basal ramets with connection to the apical ramets, suggesting Cd could be translocated due to clonal integration. Clonal integration, most likely through translocation of photosynthates, can support P. paspaloides to spread from terrestrial to Cd-contaminated aquatic habitats. Amphibious clonal plants with a high ability for clonal integration are particularly useful for re-vegetation of degraded aquatic habitats caused by Cd contamination. © 2017 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.

Clonal growth strategy, diversity and structure: A spatiotemporal response to sedimentation in tropical Cyperus papyrus swamps.

PubMed

Geremew, Addisie; Stiers, Iris; Sierens, Tim; Kefalew, Alemayehu; Triest, Ludwig

2018-01-01

Land degradation and soil erosion in the upper catchments of tropical lakes fringed by papyrus vegetation can result in a sediment load gradient from land to lakeward. Understanding the dynamics of clonal modules (ramets and genets) and growth strategies of plants on such a gradient in both space and time is critical for exploring a species adaptation and processes regulating population structure and differentiation. We assessed the spatial and temporal dynamics in clonal growth, diversity, and structure of an emergent macrophyte, Cyperus papyrus (papyrus), in response to two contrasting sedimentation regimes by combining morphological traits and genotype data using 20 microsatellite markers. A total of 636 ramets from six permanent plots (18 x 30 m) in three Ethiopian papyrus swamps, each with discrete sedimentation regimes (high vs. low) were sampled for two years. We found that ramets under the high sedimentation regime (HSR) were significantly clumped and denser than the sparse and spreading ramets under the low sedimentation regime (LSR). The HSR resulted in significantly different ramets with short culm height and girth diameter as compared to the LSR. These results indicated that C. papyrus ameliorates the effect of sedimentation by shifting clonal growth strategy from guerrilla (in LSR) to phalanx (in HSR). Clonal richness, size, dominance, and clonal subrange differed significantly between sediment regimes and studied time periods. Each swamp under HSR revealed a significantly high clonal richness (R = 0.80) as compared to the LSR (R = 0.48). Such discrepancy in clonal richness reflected the occurrence of initial and repeated seedling recruitment strategies as a response to different sedimentation regimes. Overall, our spatial and short-term temporal observations highlighted that HSR enhances clonal richness and decreases clonal subrange owing to repeated seedling recruitment and genets turnover.

Clonal growth strategy, diversity and structure: A spatiotemporal response to sedimentation in tropical Cyperus papyrus swamps

PubMed Central

Stiers, Iris; Sierens, Tim; Kefalew, Alemayehu; Triest, Ludwig

2018-01-01

Land degradation and soil erosion in the upper catchments of tropical lakes fringed by papyrus vegetation can result in a sediment load gradient from land to lakeward. Understanding the dynamics of clonal modules (ramets and genets) and growth strategies of plants on such a gradient in both space and time is critical for exploring a species adaptation and processes regulating population structure and differentiation. We assessed the spatial and temporal dynamics in clonal growth, diversity, and structure of an emergent macrophyte, Cyperus papyrus (papyrus), in response to two contrasting sedimentation regimes by combining morphological traits and genotype data using 20 microsatellite markers. A total of 636 ramets from six permanent plots (18 x 30 m) in three Ethiopian papyrus swamps, each with discrete sedimentation regimes (high vs. low) were sampled for two years. We found that ramets under the high sedimentation regime (HSR) were significantly clumped and denser than the sparse and spreading ramets under the low sedimentation regime (LSR). The HSR resulted in significantly different ramets with short culm height and girth diameter as compared to the LSR. These results indicated that C. papyrus ameliorates the effect of sedimentation by shifting clonal growth strategy from guerrilla (in LSR) to phalanx (in HSR). Clonal richness, size, dominance, and clonal subrange differed significantly between sediment regimes and studied time periods. Each swamp under HSR revealed a significantly high clonal richness (R = 0.80) as compared to the LSR (R = 0.48). Such discrepancy in clonal richness reflected the occurrence of initial and repeated seedling recruitment strategies as a response to different sedimentation regimes. Overall, our spatial and short-term temporal observations highlighted that HSR enhances clonal richness and decreases clonal subrange owing to repeated seedling recruitment and genets turnover. PMID:29338034

Hotspot mutation panel testing reveals clonal evolution in a study of 265 paired primary and metastatic tumors.

PubMed

Goswami, Rashmi S; Patel, Keyur P; Singh, Rajesh R; Meric-Bernstam, Funda; Kopetz, E Scott; Subbiah, Vivek; Alvarez, Ricardo H; Davies, Michael A; Jabbar, Kausar J; Roy-Chowdhuri, Sinchita; Lazar, Alexander J; Medeiros, L Jeffrey; Broaddus, Russell R; Luthra, Rajyalakshmi; Routbort, Mark J

2015-06-01

We used a clinical next-generation sequencing (NGS) hotspot mutation panel to investigate clonal evolution in paired primary and metastatic tumors. A total of 265 primary and metastatic tumor pairs were sequenced using a 46-gene cancer mutation panel capable of detecting one or more single-nucleotide variants as well as small insertions/deletions. Mutations were tabulated together with tumor type and percentage, mutational variant frequency, time interval between onset of primary tumor and metastasis, and neoadjuvant therapy status. Of note, 227 of 265 (85.7%) tumor metastasis pairs showed identical mutation calls. Of the tumor pairs with identical mutation calls, 160 (60.4%) possessed defining somatic mutation signatures and 67 (25.3%) did not exhibit any somatic mutations. There were 38 (14.3%) cases that showed at least one novel mutation call between the primary and metastasis. Metastases were almost two times more likely to show novel mutations (n = 20, 7.5%) than primary tumors (n = 12, 4.5%). TP53 was the most common additionally mutated gene in metastatic lesions, followed by PIK3CA and SMAD4. PIK3CA mutations were more often associated with metastasis in colon carcinoma samples. Clinical NGS hotspot panels can be useful in analyzing clonal evolution within tumors as well as in determining subclonal mutations that can expand in future metastases. PIK3CA, SMAD4, and TP53 are most often involved in clonal divergence, providing potential targets that may help guide the clinical management of tumor progression or metastases. ©2015 American Association for Cancer Research.

Identification of genuine primary pulmonary NK cell lymphoma via clinicopathologic observation and clonality assay.

PubMed

Gong, Li; Wei, Long-Xiao; Huang, Gao-Sheng; Zhang, Wen-Dong; Wang, Lu; Zhu, Shao-Jun; Han, Xiu-Juan; Yao, Li; Lan, Miao; Li, Yan-Hong; Zhang, Wei

2013-08-19

Extranodal natural killer (NK)/T-cell lymphoma, nasal type, is an uncommon lymphoma associated with the Epstein-Barr virus (EBV). It most commonly involves the nasal cavity and upper respiratory tract. Primary pulmonary NK/T cell lymphoma is extremely rare. If a patient with a NK or T-cell tumor has an unusual reaction to treatment or an unusual prognosis, it is wise to differentiate NK from T-cell tumors. The clinicopathologic characteristics, immunophenotype, EBV in situ hybridization, and T cell receptor (TCR) gene rearrangement of primary pulmonary NK cell lymphoma from a 73-year-old Chinese woman were investigated and the clonal status was determined using female X-chromosomal inactivation mosaicism and polymorphisms at the phosphoglycerate kinase (PGK) gene. The lesion showed the typical histopathologic characteristics and immunohistochemical features of NK/T cell lymphoma. However, the sample was negative for TCR gene rearrangement. A clonality assay demonstrated that the lesion was monoclonal. It is concluded that this is the first recorded case of genuine primary pulmonary NK cell lymphoma. The purpose of the present work is to recommend that pathologists carefully investigate the whole lesion to reduce the likelihood that primary pulmonary NK cell lymphoma will be misdiagnosed as an infectious lesion. In addition, TCR gene rearrangement and clonal analysis, which is based on female X-chromosomal inactivation mosaicism and polymorphisms at PGK and androgen receptor (AR) loci, were found to play important roles in differentiating NK cell lymphoma from T cell lymphoma. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/5205300349457729.

Clonal T-Cell Receptor γ-Chain Gene Rearrangements in Differential Diagnosis of Lymphomatoid Papulosis From Skin Metastasis of Nodal Anaplastic Large-Cell Lymphoma

PubMed Central

Akilov, Oleg E.; Pillai, Raju K.; Grandinetti, Lisa M.; Kant, Jeffrey A.; Geskin, Larisa

2012-01-01

Background In patients with a history of nodal anaplastic large-cell lymphoma (ALCL), differentiation of type C lymphomatoid papulosis from cutaneous involvement of systemic ALCL may be challenging because the 2 entities may exhibit identical histologic features. Although metastatic ALCL generally carries the same clone as the primary lymphoma, expression of a distinct clone likely represents a distinct process. Observations A 54-year-old white man had a history of anaplastic lymphoma kinase 1–negative ALCL in the right inguinal lymph node 6 years ago. A complete response was achieved after 6 cycles of CHOP (cyclophosphamide, doxorubicin, vincristine [Oncovin], and prednisone administered in 21-day cycles) and radiation therapy. After 3½ years, the patient observed waxing and waning papules and nodules. Examination of the biopsy specimen revealed a dense CD30+ lymphocytic infiltrate; no evidence of systemic malignancy was evident on positron emission tomography. Although clinically the presentation was consistent with lymphomatoid papulosis, metastatic ALCL had to be excluded. Polymerase chain reaction analysis with T-cell receptor γ-chain gene rearrangement (TCR-γR) was performed on the original lymph node and new skin lesions. Results of the TCR-γR analysis were positive for clonality in both lesions. However, separate clonal processes were identified. The identification of distinct clones supported the clinical impression of lymphomatoid papulosis. Conclusion Polymerase chain reaction analysis of TCR-γR is a useful method for distinguishing different clonal processes and is recommended when differentiation of primary and secondary lymphoproliferative disorders is required. PMID:21844453

Isolation and characterization of true mesenchymal stem cells derived from human term decidua capable of multilineage differentiation into all 3 embryonic layers.

PubMed

Macias, Maria I; Grande, Jesús; Moreno, Ana; Domínguez, Irene; Bornstein, Rafael; Flores, Ana I

2010-11-01

The objective of the study was to isolate and characterize a population of mesenchymal stem cells (MSCs) from human term placental membranes. We isolated an adherent cell population from extraembryonic membranes. Morphology, phenotype, growth characteristics, karyotype, and immunological and differentiation properties were analyzed. The isolated placental MSCs were from maternal origin and named as decidua-derived mesenchymal stem cells (DMSCs). DMSCs differentiated into derivatives of all germ layers. It is the first report about placental MSC differentiation into alveolar type II cells. Clonally expanded DMSCs differentiated into all embryonic layers, including pulmonary cells. DMSCs showed higher life span than placental cells from fetal origin and proliferated without genomic instability. The data suggest that DMSCs are true multipotent MSCs, distinguishing them from other placental MSCs. DMSCs could be safely used in the mother as a potential source of MSCs for pelvic floor dysfunctions and immunological diseases. Additionally, frozen DMSCs can be stored for both autologous and allogeneic tissue regeneration. Copyright © 2010 Mosby, Inc. All rights reserved.

Unique Features of Fish Immune Repertoires: Particularities of Adaptive Immunity Within the Largest Group of Vertebrates

PubMed Central

Sunyer, Oriol J.

2016-01-01

Fishes (i.e., teleost fishes) are the largest group of vertebrates. Although their immune system is based on the fundamental receptors, pathways, and cell types found in all groups of vertebrates, fishes show a diversity of particular features that challenge some classical concepts of immunology. In this chapter, we discuss the particularities of fish immune repertoires from a comparative perspective. We examine how allelic exclusion can be achieved when multiple Ig loci are present, how isotypic diversity and functional specificity impact clonal complexity, how loss of the MHC class II molecules affects the cooperation between T and B cells, and how deep sequencing technologies bring new insights about somatic hypermutation in the absence of germinal centers. The unique coexistence of two distinct B-cell lineages respectively specialized in systemic and mucosal responses is also discussed. Finally, we try to show that the diverse adaptations of immune repertoires in teleosts can help in understanding how somatic adaptive mechanisms of immunity evolved in parallel in different lineages across vertebrates. PMID:26537384

Clonal dissemination of multilocus sequence type ST15 KPC-2-producing Klebsiella pneumoniae in Bulgaria.

PubMed

Markovska, Rumyana; Stoeva, Temenuga; Schneider, Ines; Boyanova, Lyudmila; Popova, Valentina; Dacheva, Daniela; Kaneva, Radka; Bauernfeind, Adolf; Mitev, Vanyo; Mitov, Ivan

2015-10-01

A total of 36 consecutive clinical and two fecal-screening carbapenem-resistant Klebsiella pneumoniae isolates from two Bulgarian university hospitals (Varna and Pleven) were investigated. Susceptibility testing, conjugation experiments, and plasmid replicon typing were carried out. Beta-lactamases were characterized by isoelectric focusing, PCR, and sequencing. Clonal relatedness was investigated by RAPD and multilocus sequence typing (MLST). Most of the isolates demonstrated multidrug resistance profile. Amikacin and tigecycline retained good activity with susceptibility rates of 95 and 87%, respectively. The resistance rate to colistin was 63%. Six RAPD- and MLST-types were identified: the dominating MLST-type was ST15 (27 isolates), followed by ST76 (six isolates), and ST1350 (two isolates). ST101, ST258, and ST151 were detected once. All except one of the K. pneumoniae produced KPC-2, mostly in combination with CTX-M-15, while for one isolate (ST101) the enzymes OXA-48 and CTX-M-14 were found. All KPC-2-producing transconjugants revealed the presence of IncFII plasmid. The OXA-48- and CTX-M-14-producing isolate showed the presence of L/M replicon type. The dissemination of KPC-2-producing K.pneumoniae in Bulgaria is mainly due to the sustained spread of successful ST15 clone and to a lesser extent of ST76 clone. This is the first report of OXA-48 producing ST101 K. pneumoniae in Bulgaria. © 2015 APMIS. Published by John Wiley & Sons Ltd.

Molecular Epidemiology of Carbapenem-Resistant Acinetobacter baumannii Isolates in the Gulf Cooperation Council States: Dominance of OXA-23-Type Producers

PubMed Central

Sartor, Anna L.; Sidjabat, Hanna E.; Balkhy, Hanan H.; Walsh, Timothy R.; Al Johani, Sameera M.; AlJindan, Reem Y.; Alfaresi, Mubarak; Ibrahim, Emad; Al-Jardani, Amina; Al Salman, Jameela; Dashti, Ali A.; Johani, Khalid; Paterson, David L.

2015-01-01

The molecular epidemiology and mechanisms of resistance of carbapenem-resistant Acinetobacter baumannii (CRAB) were determined in hospitals in the states of the Cooperation Council for the Arab States of the Gulf (Gulf Cooperation Council [GCC]), namely, Saudi Arabia, United Arab Emirates, Oman, Qatar, Bahrain, and Kuwait. Isolates were subjected to PCR-based detection of antibiotic resistance genes and repetitive sequence-based PCR (rep-PCR) assessments of clonality. Selected isolates were subjected to multilocus sequence typing (MLST). We investigated 117 isolates resistant to carbapenem antibiotics (either imipenem or meropenem). All isolates were positive for OXA-51. The most common carbapenemases were the OXA-23-type, found in 107 isolates, followed by OXA-40-type (OXA-24-type), found in 5 isolates; 3 isolates carried the ISAba1 element upstream of blaOXA-51-type. No OXA-58-type, NDM-type, VIM-type, or IMP-type producers were detected. Multiple clones were detected with 16 clusters of clonally related CRAB. Some clusters involved hospitals in different states. MLST analysis of 15 representative isolates from different clusters identified seven different sequence types (ST195, ST208, ST229, ST436, ST450, ST452, and ST499), as well as three novel STs. The vast majority (84%) of the isolates in this study were associated with health care exposure. Awareness of multidrug-resistant organisms in GCC states has important implications for optimizing infection control practices; establishing antimicrobial stewardship programs within hospital, community, and agricultural settings; and emphasizing the need for establishing regional active surveillance systems. This will help to control the spread of CRAB in the Middle East and in hospitals accommodating transferred patients from this region. PMID:25568439

Molecular epidemiology of carbapenem-resistant Acinetobacter baumannii isolates in the Gulf Cooperation Council States: dominance of OXA-23-type producers.

PubMed

Zowawi, Hosam M; Sartor, Anna L; Sidjabat, Hanna E; Balkhy, Hanan H; Walsh, Timothy R; Al Johani, Sameera M; AlJindan, Reem Y; Alfaresi, Mubarak; Ibrahim, Emad; Al-Jardani, Amina; Al Salman, Jameela; Dashti, Ali A; Johani, Khalid; Paterson, David L

2015-03-01

The molecular epidemiology and mechanisms of resistance of carbapenem-resistant Acinetobacter baumannii (CRAB) were determined in hospitals in the states of the Cooperation Council for the Arab States of the Gulf (Gulf Cooperation Council [GCC]), namely, Saudi Arabia, United Arab Emirates, Oman, Qatar, Bahrain, and Kuwait. Isolates were subjected to PCR-based detection of antibiotic resistance genes and repetitive sequence-based PCR (rep-PCR) assessments of clonality. Selected isolates were subjected to multilocus sequence typing (MLST). We investigated 117 isolates resistant to carbapenem antibiotics (either imipenem or meropenem). All isolates were positive for OXA-51. The most common carbapenemases were the OXA-23-type, found in 107 isolates, followed by OXA-40-type (OXA-24-type), found in 5 isolates; 3 isolates carried the ISAba1 element upstream of blaOXA-51-type. No OXA-58-type, NDM-type, VIM-type, or IMP-type producers were detected. Multiple clones were detected with 16 clusters of clonally related CRAB. Some clusters involved hospitals in different states. MLST analysis of 15 representative isolates from different clusters identified seven different sequence types (ST195, ST208, ST229, ST436, ST450, ST452, and ST499), as well as three novel STs. The vast majority (84%) of the isolates in this study were associated with health care exposure. Awareness of multidrug-resistant organisms in GCC states has important implications for optimizing infection control practices; establishing antimicrobial stewardship programs within hospital, community, and agricultural settings; and emphasizing the need for establishing regional active surveillance systems. This will help to control the spread of CRAB in the Middle East and in hospitals accommodating transferred patients from this region. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Clonal diversity and detection of carbapenem resistance encoding genes among multidrug-resistant Acinetobacter baumannii isolates recovered from patients and environment in two intensive care units in a Moroccan hospital.

PubMed

Uwingabiye, Jean; Lemnouer, Abdelhay; Roca, Ignasi; Alouane, Tarek; Frikh, Mohammed; Belefquih, Bouchra; Bssaibis, Fatna; Maleb, Adil; Benlahlou, Yassine; Kassouati, Jalal; Doghmi, Nawfal; Bait, Abdelouahed; Haimeur, Charki; Louzi, Lhoussain; Ibrahimi, Azeddine; Vila, Jordi; Elouennass, Mostafa

2017-01-01

Carbapenem-resistant Acinetobacter baumannii has recently been defined by the World Health Organization as a critical pathogen. The aim of this study was to compare clonal diversity and carbapenemase-encoding genes of A. baumannii isolates collected from colonized or infected patients and hospital environment in two intensive care units (ICUs) in Morocco. The patient and environmental sampling was carried out in the medical and surgical ICUs of Mohammed V Military teaching hospital from March to August 2015. All A. baumannii isolates recovered from clinical and environmental samples, were identified using routine microbiological techniques and Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry. Antimicrobial susceptibility testing was performed using disc diffusion method. The carbapenemase-encoding genes were screened for by PCR. Clonal relatedness was analyzed by digestion of the DNA with low frequency restriction enzymes and pulsed field gel electrophoresis (PFGE) and the multi locus sequence typing (MLST) was performed on two selected isolates from two major pulsotypes. A total of 83 multidrug-resistant A. baumannii isolates were collected: 47 clinical isolates and 36 environmental isolates. All isolates were positive for the bla OXA51-like and bla OXA23-like genes. The coexistence of bla NDM-1 /bla OXA-23-like and bla OXA 24-like /bla OXA-23-like were detected in 27 (32.5%) and 2 (2.4%) of A. baumannii isolates, respectively. The environmental samples and the fecally-colonized patients were significantly identified ( p  < 0.05) as the most common sites of isolation of NDM-1-harboring isolates. PFGE grouped all isolates into 9 distinct clusters with two major groups (0007 and 0008) containing up to 59% of the isolates. The pulsotype 0008 corresponds to sequence type (ST) 195 while pulsotype 0007 corresponds to ST 1089.The genetic similarity between the clinical and environmental isolates was observed in 80/83 = 96.4% of all isolates, belonging to 7 pulsotypes. This study shows that the clonal spread of environmental A. baumannii isolates is related to that of clinical isolates recovered from colonized or infected patients, being both associated with a high prevalence of the bla OXA23-like and bla NDM-1 genes. These findings emphasize the need for prioritizing the bio-cleaning of the hospital environment to control and prevent the dissemination of A. baumannii clonal lineages.

Assignment of Streptococcus agalactiae isolates to clonal complexes using a small set of single nucleotide polymorphisms.

PubMed

Honsa, Erin; Fricke, Thomas; Stephens, Alex J; Ko, Danny; Kong, Fanrong; Gilbert, Gwendolyn L; Huygens, Flavia; Giffard, Philip M

2008-08-19

Streptococcus agalactiae (Group B Streptococcus (GBS)) is an important human pathogen, particularly of newborns. Emerging evidence for a relationship between genotype and virulence has accentuated the need for efficient and well-defined typing methods. The objective of this study was to develop a single nucleotide polymorphism (SNP) based method for assigning GBS isolates to multilocus sequence typing (MLST)-defined clonal complexes. It was found that a SNP set derived from the MLST database on the basis of maximization of Simpsons Index of Diversity provided poor resolution and did not define groups concordant with the population structure as defined by eBURST analysis of the MLST database. This was interpreted as being a consequence of low diversity and high frequency horizontal gene transfer. Accordingly, a different approach to SNP identification was developed. This entailed use of the "Not-N" bioinformatic algorithm that identifies SNPs diagnostic for groups of known sequence variants, together with an empirical process of SNP testing. This yielded a four member SNP set that divides GBS into 10 groups that are concordant with the population structure. A fifth SNP was identified that increased the sensitivity for the clinically significant clonal complex 17 to 100%. Kinetic PCR methods for the interrogation of these SNPs were developed, and used to genotype 116 well characterized isolates. A five SNP method for dividing GBS into biologically valid groups has been developed. These SNPs are ideal for high throughput surveillance activities, and combining with more rapidly evolving loci when additional resolution is required.

Assignment of Streptococcus agalactiae isolates to clonal complexes using a small set of single nucleotide polymorphisms

PubMed Central

Honsa, Erin; Fricke, Thomas; Stephens, Alex J; Ko, Danny; Kong, Fanrong; Gilbert, Gwendolyn L; Huygens, Flavia; Giffard, Philip M

2008-01-01

Background Streptococcus agalactiae (Group B Streptococcus (GBS)) is an important human pathogen, particularly of newborns. Emerging evidence for a relationship between genotype and virulence has accentuated the need for efficient and well-defined typing methods. The objective of this study was to develop a single nucleotide polymorphism (SNP) based method for assigning GBS isolates to multilocus sequence typing (MLST)-defined clonal complexes. Results It was found that a SNP set derived from the MLST database on the basis of maximisation of Simpsons Index of Diversity provided poor resolution and did not define groups concordant with the population structure as defined by eBURST analysis of the MLST database. This was interpreted as being a consequence of low diversity and high frequency horizontal gene transfer. Accordingly, a different approach to SNP identification was developed. This entailed use of the "Not-N" bioinformatic algorithm that identifies SNPs diagnostic for groups of known sequence variants, together with an empirical process of SNP testing. This yielded a four member SNP set that divides GBS into 10 groups that are concordant with the population structure. A fifth SNP was identified that increased the sensitivity for the clinically significant clonal complex 17 to 100%. Kinetic PCR methods for the interrogation of these SNPs were developed, and used to genotype 116 well characterized isolates. Conclusion A five SNP method for dividing GBS into biologically valid groups has been developed. These SNPs are ideal for high throughput surveillance activities, and combining with more rapidly evolving loci when additional resolution is required. PMID:18710585

Minimal Residual Disease Monitoring of Acute Myeloid Leukemia by Massively Multiplex Digital PCR in Patients with NPM1 Mutations.

PubMed

Mencia-Trinchant, Nuria; Hu, Yang; Alas, Maria Antonina; Ali, Fatima; Wouters, Bas J; Lee, Sangmin; Ritchie, Ellen K; Desai, Pinkal; Guzman, Monica L; Roboz, Gail J; Hassane, Duane C

2017-07-01

The presence of minimal residual disease (MRD) is widely recognized as a powerful predictor of therapeutic outcome in acute myeloid leukemia (AML), but methods of measurement and quantification of MRD in AML are not yet standardized in clinical practice. There is an urgent, unmet need for robust and sensitive assays that can be readily adopted as real-time tools for disease monitoring. NPM1 frameshift mutations are an established MRD marker present in half of patients with cytogenetically normal AML. However, detection is complicated by the existence of hundreds of potential frameshift insertions, clonal heterogeneity, and absence of sequence information when the NPM1 mutation is identified using capillary electrophoresis. Thus, some patients are ineligible for NPM1 MRD monitoring. Furthermore, a subset of patients with NPM1-mutated AML will have false-negative MRD results because of clonal evolution. To simplify and improve MRD testing for NPM1, we present a novel digital PCR technique composed of massively multiplex pools of insertion-specific primers that selectively detect mutated but not wild-type NPM1. By measuring reaction end points using digital PCR technology, the resulting single assay enables sensitive and specific quantification of most NPM1 exon 12 mutations in a manner that is robust to clonal heterogeneity, does not require NPM1 sequence information, and obviates the need for maintenance of hundreds of type-specific assays and associated plasmid standards. Copyright © 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Detecting truly clonal alterations from multi-region profiling of tumours

PubMed Central

Werner, Benjamin; Traulsen, Arne; Sottoriva, Andrea; Dingli, David

2017-01-01

Modern cancer therapies aim at targeting tumour-specific alterations, such as mutations or neo-antigens, and maximal treatment efficacy requires that targeted alterations are present in all tumour cells. Currently, treatment decisions are based on one or a few samples per tumour, creating uncertainty on whether alterations found in those samples are actually present in all tumour cells. The probability of classifying clonal versus sub-clonal alterations from multi-region profiling of tumours depends on the earliest phylogenetic branching event during tumour growth. By analysing 181 samples from 10 renal carcinoma and 11 colorectal cancers we demonstrate that the information gain from additional sampling falls onto a simple universal curve. We found that in colorectal cancers, 30% of alterations identified as clonal with one biopsy proved sub-clonal when 8 samples were considered. The probability to overestimate clonal alterations fell below 1% in 7/11 patients with 8 samples per tumour. In renal cell carcinoma, 8 samples reduced the list of clonal alterations by 40% with respect to a single biopsy. The probability to overestimate clonal alterations remained as high as 92% in 7/10 renal cancer patients. Furthermore, treatment was associated with more unbalanced tumour phylogenetic trees, suggesting the need of denser sampling of tumours at relapse. PMID:28344344

Heterogeneous water supply affects growth and benefits of clonal integration between co-existing invasive and native Hydrocotyle species.

PubMed

Wang, Yong-Jian; Bai, Yun-Fei; Zeng, Shi-Qi; Yao, Bin; Wang, Wen; Luo, Fang-Li

2016-07-21

Spatial patchiness and temporal variability in water availability are common in nature under global climate change, which can remarkably influence adaptive responses of clonal plants, i.e. clonal integration (translocating resources between connected ramets). However, little is known about the effects of spatial patchiness and temporal heterogeneity in water on growth and clonal integration between congeneric invasive and native Hydrocotyle species. In a greenhouse experiment, we subjected severed or no severed (intact) fragments of Hydrocotyle vulgaris, a highly invasive species in China, and its co-existing, native congener H. sibthorpioides to different spatial patchiness (homogeneous and patchy) and temporal interval (low and high interval) in water supply. Clonal integration had significant positive effects on growth of both species. In the homogeneous water conditions, clonal integration greatly improved the growth in fragments of both species under low interval in water. However, in the patchy water conditions, clonal integration significantly increased growth in both ramets and fragments of H. vulgaris under high interval in water. Therefore, spatial patchiness and temporal interval in water altered the effects of clonal integration of both species, especially for H. vulgaris. The adaptation of H. vulgaris might lead to invasive growth and potential spread under the global water variability.

Detecting truly clonal alterations from multi-region profiling of tumours

NASA Astrophysics Data System (ADS)

Werner, Benjamin; Traulsen, Arne; Sottoriva, Andrea; Dingli, David

2017-03-01

Modern cancer therapies aim at targeting tumour-specific alterations, such as mutations or neo-antigens, and maximal treatment efficacy requires that targeted alterations are present in all tumour cells. Currently, treatment decisions are based on one or a few samples per tumour, creating uncertainty on whether alterations found in those samples are actually present in all tumour cells. The probability of classifying clonal versus sub-clonal alterations from multi-region profiling of tumours depends on the earliest phylogenetic branching event during tumour growth. By analysing 181 samples from 10 renal carcinoma and 11 colorectal cancers we demonstrate that the information gain from additional sampling falls onto a simple universal curve. We found that in colorectal cancers, 30% of alterations identified as clonal with one biopsy proved sub-clonal when 8 samples were considered. The probability to overestimate clonal alterations fell below 1% in 7/11 patients with 8 samples per tumour. In renal cell carcinoma, 8 samples reduced the list of clonal alterations by 40% with respect to a single biopsy. The probability to overestimate clonal alterations remained as high as 92% in 7/10 renal cancer patients. Furthermore, treatment was associated with more unbalanced tumour phylogenetic trees, suggesting the need of denser sampling of tumours at relapse.

Effects of Clonal Reproduction on Evolutionary Lag and Evolutionary Rescue.

PubMed

Orive, Maria E; Barfield, Michael; Fernandez, Carlos; Holt, Robert D

2017-10-01

Evolutionary lag-the difference between mean and optimal phenotype in the current environment-is of keen interest in light of rapid environmental change. Many ecologically important organisms have life histories that include stage structure and both sexual and clonal reproduction, yet how stage structure and clonality interplay to govern a population's rate of evolution and evolutionary lag is unknown. Effects of clonal reproduction on mean phenotype partition into two portions: one that is phenotype dependent, and another that is genotype dependent. This partitioning is governed by the association between the nonadditive genetic plus random environmental component of phenotype of clonal offspring and their parents. While clonality slows phenotypic evolution toward an optimum, it can dramatically increase population survival after a sudden step change in optimal phenotype. Increased adult survival slows phenotypic evolution but facilitates population survival after a step change; this positive effect can, however, be lost given survival-fecundity trade-offs. Simulations indicate that the benefits of increased clonality under environmental change greatly depend on the nature of that change: increasing population persistence under a step change while decreasing population persistence under a continuous linear change requiring de novo variation. The impact of clonality on the probability of persistence for species in a changing world is thus inexorably linked to the temporal texture of the change they experience.

Clonal sets of a binary relation

NASA Astrophysics Data System (ADS)

Zedam, Lemnaouar; Pérez-Fernández, Raúl; Bouremel, Hassane; De Baets, Bernard

2018-05-01

In a recent paper, we have introduced the notion of clone relation of a given binary relation. Intuitively, two elements are said to be "clones" if they are related in the same way w.r.t. every other element. In this paper, we generalize this notion from pairs of elements to sets of elements of any cardinality, resulting in the introduction of clonal sets. We investigate the most important properties of clonal sets, paying particular attention to the introduction of the clonal closure operator, to the analysis of the (lattice) structure of the set of clonal sets and to a distance metric expressing how close two elements are to being clones.

Molecular Typing and Virulence Characteristic of Methicillin-Resistant Staphylococcus Aureus Isolates from Pediatric Patients in Bucaramanga, Colombia

PubMed Central

Machuca, Mayra Alejandra; Sosa, Luis Miguel; González, Clara Isabel

2013-01-01

Background Staphylococcus aureus is among the most common global nosocomial pathogens. The emergence and spread of methicillin-resistant Staphylococcus aureus (MRSA) is a public health problem worldwide that causes nosocomial and community infections. The goals of this study were to establish the clonal complexes (CC) of the isolates of MRSA obtained from pediatric patients in a university hospital in Colombia and to investigate its molecular characteristics based on the virulence genes and the genes of staphylococcal toxins and adhesins. Methods A total of 53 MRSA isolates from pediatric patients with local or systemic infections were collected. The MRSA isolates were typed based on the SCCmec, MLST, spa and agr genes. The molecular characterization included the detection of Panton-Valentine Leukocidin, superantigenic and exfoliative toxins, and adhesin genes. The correlation between the molecular types identified and the profile of virulence factors was determined for all isolates. Results Four CC were identified, including CC8, CC5, CC80 and CC78. The ST8-MRSA-IVc-agrI was the predominant clone among the isolates, followed by the ST5-MRSA-I-agrII and ST5-MRSA-IVc-agrII clones. Twelve spa types were identified, of which t10796 and t10799 were new repeat sequences. The isolates were carriers of toxin genes, and hlg (100%), sek (92%) and pvl (88%) were the most frequent. Ten toxin gene profiles were observed, and the most frequent were seq-sek-hlg (22.6%), sek-hlg (22.6%), seb-seq-sek-hlg (18.9%) and seb-sek-hlg (15.1%). The adhesion genes were present in most of the MRSA isolates, including the following: clf-A (89%), clf-B (87%), fnb-A (83%) and ica (83%). The majority of the strains carried SCCmec-IVc and were identified as causing nosocomial infection. No significant association between a molecular type and the virulence factors was found. Conclusion Four major MRSA clone complexes were identified among the isolates. ST8-MRSA-IVc-agrI pvl+ (USA300-LV) was the most frequent, confirming the presence of community-associated MRSA in Colombian hospitals. PMID:24058415

Bradykinin-mediated diseases.

PubMed

Kaplan, Allen P

2014-01-01

Diseases which have been demonstrated to be caused by increased plasma levels of bradykinin all have angioedema as the common major clinical manifestation. Angioedema due to therapy with angiotensin-converting enzyme (ACE) inhibitors is caused by suppressed bradykinin degradation so that it accumulates. This occurs because ACE metabolizes bradykinin by removal of Phe-Arg from the C-terminus, which inactivates it. By contrast, angioedema due to C1 inhibitor deficiency (either hereditary types I and II, or acquired) is caused by bradykinin overproduction. C1 inhibitor inhibits factor XIIa, kallikrein and activity associated with the prekallikrein-HK (high-molecular-weight kininogen) complex. In its absence, uncontrolled activation of the plasma bradykinin cascade is seen once there has been an initiating stimulus. C4 levels are low in all types of C1 inhibitor deficiency due to the instability of C1 (C1r, in particular) such that some activated C1 always circulates and depletes C4. In the hereditary disorder, formation of factor XIIf (factor XII fragment) during attacks of swelling causes C4 levels to drop toward zero, and C2 levels decline. A kinin-like molecule, once thought to be a cleavage product derived from C2 that contributes to the increased vascular permeability seen in hereditary angioedema (HAE), is now thought to be an artifact, i.e. no such molecule is demonstrable. The acquired C1 inhibitor deficiency is associated with clonal disorders of B cell hyperreactivity, including lymphoma and monoclonal gammopathy. Most cases have an IgG autoantibody to C1 inhibitor which inactivates it so that the presentation is strikingly similar to type I HAE. New therapies for types I and II HAE include C1 inhibitor replacement therapy, ecallantide, a kallikrein antagonist, and icatibant, a B2 receptor antagonist. A newly described type III HAE has normal C1 inhibitor, although it is thought to be mediated by bradykinin, as is an antihistamine-resistant subpopulation of patients with 'idiopathic' angioedema. The mechanism(s) for the formation of bradykinin in these disorders is unknown. © 2014 S. Karger AG, Basel.

Rise and fall of subclones from diagnosis to relapse in pediatric B-acute lymphoblastic leukaemia | Office of Cancer Genomics

Cancer.gov

There is incomplete understanding of genetic heterogeneity and clonal evolution during cancer progression. Here we use deep whole-exome sequencing to describe the clonal architecture and evolution of 20 pediatric B-acute lymphoblastic leukaemias from diagnosis to relapse. We show that clonal diversity is comparable at diagnosis and relapse and clonal survival from diagnosis to relapse is not associated with mutation burden.

Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade

PubMed Central

McGranahan, Nicholas; Furness, Andrew J. S.; Rosenthal, Rachel; Ramskov, Sofie; Lyngaa, Rikke; Saini, Sunil Kumar; Jamal-Hanjani, Mariam; Wilson, Gareth A.; Birkbak, Nicolai J.; Hiley, Crispin T.; Watkins, Thomas B. K.; Shafi, Seema; Murugaesu, Nirupa; Mitter, Richard; Akarca, Ayse U.; Linares, Joseph; Marafioti, Teresa; Henry, Jake Y.; Van Allen, Eliezer M.; Miao, Diana; Schilling, Bastian; Schadendorf, Dirk; Garraway, Levi A.; Makarov, Vladimir; Rizvi, Naiyer A.; Snyder, Alexandra; Hellmann, Matthew D.; Merghoub, Taha; Wolchok, Jedd D.; Shukla, Sachet A.; Wu, Catherine J.; Peggs, Karl S.; Chan, Timothy A.; Hadrup, Sine R.; Quezada, Sergio A.; Swanton, Charles

2016-01-01

As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we demonstrate a relationship between clonal neoantigen burden and overall survival in primary lung adenocarcinomas. CD8+ tumor-infiltrating lymphocytes reactive to clonal neoantigens were identified in early-stage non–small cell lung cancer and expressed high levels of PD-1. Sensitivity to PD-1 and CTLA-4 blockade in patients with advanced NSCLC and melanoma was enhanced in tumors enriched for clonal neoantigens. T cells recognizing clonal neoantigens were detectable in patients with durable clinical benefit. Cytotoxic chemotherapy–induced subclonal neoantigens, contributing to an increased mutational load, were enriched in certain poor responders. These data suggest that neoantigen heterogeneity may influence immune surveillance and support therapeutic developments targeting clonal neoantigens. PMID:26940869

Kin Recognition in a Clonal Fish, Poecilia formosa

PubMed Central

Makowicz, Amber M.; Tiedemann, Ralph; Schlupp, Ingo

2016-01-01

Relatedness strongly influences social behaviors in a wide variety of species. For most species, the highest typical degree of relatedness is between full siblings with 50% shared genes. However, this is poorly understood in species with unusually high relatedness between individuals: clonal organisms. Although there has been some investigation into clonal invertebrates and yeast, nothing is known about kin selection in clonal vertebrates. We show that a clonal fish, the Amazon molly (Poecilia formosa), can distinguish between different clonal lineages, associating with genetically identical, sister clones, and use multiple sensory modalities. Also, they scale their aggressive behaviors according to the relatedness to other females: they are more aggressive to non-related clones. Our results demonstrate that even in species with very small genetic differences between individuals, kin recognition can be adaptive. Their discriminatory abilities and regulation of costly behaviors provides a powerful example of natural selection in species with limited genetic diversity. PMID:27483372

Analysis of allelic expression patterns in clonal somatic cells by single-cell RNA-seq

PubMed Central

Ramsköld, Daniel; Deng, Qiaolin; Johnsson, Per; Michaëlsson, Jakob; Frisén, Jonas; Sandberg, Rickard

2016-01-01

Cellular heterogeneity can emerge from the expression of only one parental allele. However, it has remained controversial whether, or to what degree, random monoallelic expression of autosomal genes (aRME) is mitotically inherited (clonal) or stochastic (dynamic) in somatic cells, particularly in vivo. Here, we used allele-sensitive single-cell RNA-seq on clonal primary mouse fibroblasts and in vivo human CD8+ T-cells to dissect clonal and dynamic monoallelic expression patterns. Dynamic aRME affected a considerable portion of the cells’ transcriptomes, with levels dependent on the cells’ transcriptional activity. Importantly, clonal aRME was detected but was surprisingly scarce (<1% of genes) and affected mainly the most low-expressed genes. Consequently, the overwhelming portion of aRME occurs transiently within individual cells and patterns of aRME are thus primarily scattered throughout somatic cell populations rather than, as previously hypothesized, confined to patches of clonally related cells. PMID:27668657

Analysis of allelic expression patterns in clonal somatic cells by single-cell RNA-seq.

PubMed

Reinius, Björn; Mold, Jeff E; Ramsköld, Daniel; Deng, Qiaolin; Johnsson, Per; Michaëlsson, Jakob; Frisén, Jonas; Sandberg, Rickard

2016-11-01

Cellular heterogeneity can emerge from the expression of only one parental allele. However, it has remained controversial whether, or to what degree, random monoallelic expression of autosomal genes (aRME) is mitotically inherited (clonal) or stochastic (dynamic) in somatic cells, particularly in vivo. Here we used allele-sensitive single-cell RNA-seq on clonal primary mouse fibroblasts and freshly isolated human CD8 + T cells to dissect clonal and dynamic monoallelic expression patterns. Dynamic aRME affected a considerable portion of the cells' transcriptomes, with levels dependent on the cells' transcriptional activity. Notably, clonal aRME was detected, but it was surprisingly scarce (<1% of genes) and mainly affected the most weakly expressed genes. Consequently, the overwhelming majority of aRME occurs transiently within individual cells, and patterns of aRME are thus primarily scattered throughout somatic cell populations rather than, as previously hypothesized, confined to patches of clonally related cells.

Clonality: an R package for testing clonal relatedness of two tumors from the same patient based on their genomic profiles.

PubMed

Ostrovnaya, Irina; Seshan, Venkatraman E; Olshen, Adam B; Begg, Colin B

2011-06-15

If a cancer patient develops multiple tumors, it is sometimes impossible to determine whether these tumors are independent or clonal based solely on pathological characteristics. Investigators have studied how to improve this diagnostic challenge by comparing the presence of loss of heterozygosity (LOH) at selected genetic locations of tumor samples, or by comparing genomewide copy number array profiles. We have previously developed statistical methodology to compare such genomic profiles for an evidence of clonality. We assembled the software for these tests in a new R package called 'Clonality'. For LOH profiles, the package contains significance tests. The analysis of copy number profiles includes a likelihood ratio statistic and reference distribution, as well as an option to produce various plots that summarize the results. Bioconductor (http://bioconductor.org/packages/release/bioc/html/Clonality.html) and http://www.mskcc.org/mskcc/html/13287.cfm.

Insights in Anaphylaxis and Clonal Mast Cell Disorders.

PubMed

González-de-Olano, David; Álvarez-Twose, Iván

2017-01-01

The prevalence of anaphylaxis among patients with clonal mast cell disorders (MCD) is clearly higher comparing to the general population. Due to a lower frequency of symptoms outside of acute episodes, clonal MCD in the absence of skin lesions might sometimes be difficult to identify which may lead to underdiagnosis, and anaphylaxis is commonly the presenting symptom in these patients. Although the release of mast cell (MC) mediators upon MC activation might present with a wide variety of symptoms, particular clinical features typically characterize MC mediator release episodes in patients with clonal MCD without skin involvement. Final diagnosis requires a bone marrow study, and it is recommended that this should be done in reference centers. In this article, we address the main triggers for anaphylaxis, risk factors, clinical presentation, diagnosis, and management of patients with MC activation syndromes (MCASs), with special emphasis on clonal MCAS [systemic mastocytosis and mono(clonal) MC activations syndromes].

Insights in Anaphylaxis and Clonal Mast Cell Disorders

PubMed Central

González-de-Olano, David; Álvarez-Twose, Iván

2017-01-01

The prevalence of anaphylaxis among patients with clonal mast cell disorders (MCD) is clearly higher comparing to the general population. Due to a lower frequency of symptoms outside of acute episodes, clonal MCD in the absence of skin lesions might sometimes be difficult to identify which may lead to underdiagnosis, and anaphylaxis is commonly the presenting symptom in these patients. Although the release of mast cell (MC) mediators upon MC activation might present with a wide variety of symptoms, particular clinical features typically characterize MC mediator release episodes in patients with clonal MCD without skin involvement. Final diagnosis requires a bone marrow study, and it is recommended that this should be done in reference centers. In this article, we address the main triggers for anaphylaxis, risk factors, clinical presentation, diagnosis, and management of patients with MC activation syndromes (MCASs), with special emphasis on clonal MCAS [systemic mastocytosis and mono(clonal) MC activations syndromes]. PMID:28740494

Outbreak of Vibrio parahaemolyticus Sequence Type 120, Peru, 2009.

PubMed

Gonzalez-Escalona, Narjol; Gavilan, Ronnie G; Toro, Magaly; Zamudio, Maria L; Martinez-Urtaza, Jaime

2016-07-01

In 2009, an outbreak of Vibrio parahaemolyticus occurred in Piura, Cajamarca, Lambayeque, and Lima, Peru. Whole-genome sequencing of clinical and environmental samples from the outbreak revealed a new V. parahaemolyticus clone. All the isolates identified belonged to a single clonal complex described exclusively in Asia before its emergence in Peru.

Outbreak of Vibrio parahaemolyticus Sequence Type 120, Peru, 2009

PubMed Central

Gonzalez-Escalona, Narjol; Gavilan, Ronnie G.; Toro, Magaly; Zamudio, Maria L.

2016-01-01

In 2009, an outbreak of Vibrio parahaemolyticus occurred in Piura, Cajamarca, Lambayeque, and Lima, Peru. Whole-genome sequencing of clinical and environmental samples from the outbreak revealed a new V. parahaemolyticus clone. All the isolates identified belonged to a single clonal complex described exclusively in Asia before its emergence in Peru. PMID:27315090

Raman spectroscopy-based identification of nosocomial outbreaks of the clonal bacterium Escherichia coli.

PubMed

Kusters, J G; van Leeuwen, W B; Maquelin, K; Blok, H E M; Willemse, H F M; de Graaf-Miltenburg, L A M; Fluit, A C; Troelstra, A

2016-01-01

DNA-based techniques are frequently used to confirm the relatedness of putative outbreak isolates. These techniques often lack the discriminatory power when analyzing closely related microbes such as E. coli. Here the value of Raman spectroscopy as a typing tool for E. coli in a clinical setting was retrospectively evaluated.

Clonality, recombination, and hybridization in the plumbing-inhabiting human pathogen Fusarium keratoplasticum inferred from multilocus sequence typing

USDA-ARS?s Scientific Manuscript database

Recent work has shown that Fusarium species and genotypes most commonly associated with human infections, particularly of the cornea (mycotic keratitis), are the same as those most commonly isolated from plumbing systems. The species most dominant in plumbing biofilms is Fusarium keratoplasticum, a ...

Clonal structure in Ichthyobacterium seriolicida, the causative agent of bacterial haemolytic jaundice in yellowtail, Seriola quinqueradiata, inferred from molecular epidemiological analysis.

PubMed

Matsuyama, T; Fukuda, Y; Sakai, T; Tanimoto, N; Nakanishi, M; Nakamura, Y; Takano, T; Nakayasu, C

2017-08-01

Bacterial haemolytic jaundice caused by Ichthyobacterium seriolicida has been responsible for mortality in farmed yellowtail, Seriola quinqueradiata, in western Japan since the 1980s. In this study, polymorphic analysis of I. seriolicida was performed using three molecular methods: amplified fragment length polymorphism (AFLP) analysis, multilocus sequence typing (MLST) and multiple-locus variable-number tandem repeat analysis (MLVA). Twenty-eight isolates were analysed using AFLP, while 31 isolates were examined by MLST and MLVA. No polymorphisms were identified by AFLP analysis using EcoRI and MseI, or by MLST of internal fragments of eight housekeeping genes. However, MLVA revealed variation in repeat numbers of three elements, allowing separation of the isolates into 16 sequence types. The unweighted pair group method using arithmetic averages cluster analysis of the MLVA data identified four major clusters, and all isolates belonged to clonal complexes. It is likely that I. seriolicida populations share a common ancestor, which may be a recently introduced strain. © 2016 John Wiley & Sons Ltd.

Genotyping of clinical and environmental multidrug resistant Enterococcus faecium strains.

PubMed

Shokoohizadeh, Leili; Mobarez, Ashraf Mohabati; Alebouyeh, Masoud; Zali, Mohammad Reza; Ranjbar, Reza

2017-01-01

Multidrug resistant (MDR) Enterococcus faecium is a nosocomial pathogen and clonal complex 17 (CC17) is the main genetic subpopulation of E. faecium in hospitals worldwide. There has thus far been no report of major E. faecium clones in Iranian hospitals. The present study analyzed strains of MDR E. faecium obtained from patients and the Intensive Care Unit environments using pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) to determine the antibiotic resistance patterns and genetic features of the dominant. clones of E. faecium. PFGE and MLST analysis revealed the presence of 17and 15 different subtypes, respectively. Of these, 18 (86%) isolates belonged toCC17. Most strains in this clonal complex harbored the esp gene and exhibited resistance to vancomycin, teicoplanin, ampicillin, ciprofloxacin, gentamicin, and erythromycin. The MLST results revealed 12 new sequence types (ST) for the first time. Approximately 50% of the STs were associated with ST203. Detection of E. faecium strains belonging to CC17 on medical equipment and in clinical specimens verified the circulation of high-risk MDR clones among the patients and in hospital environments in Iran.

Evolution of Sequence Type 4821 Clonal Complex Meningococcal Strains in China from Prequinolone to Quinolone Era, 1972–2013

PubMed Central

Guo, Qinglan; Mustapha, Mustapha M.; Chen, Mingliang; Qu, Di; Zhang, Xi; Harrison, Lee H.

2018-01-01

The expansion of hypervirulent sequence type 4821 clonal complex (CC4821) lineage Neisseria meningitidis bacteria has led to a shift in meningococcal disease epidemiology in China, from serogroup A (MenA) to MenC. Knowledge of the evolution and genetic origin of the emergent MenC strains is limited. In this study, we subjected 76 CC4821 isolates collected across China during 1972–1977 and 2005–2013 to phylogenetic analysis, traditional genotyping, or both. We show that successive recombination events within genes encoding surface antigens and acquisition of quinolone resistance mutations possibly played a role in the emergence of CC4821 as an epidemic clone in China. MenC and MenB CC4821 strains have spread across China and have been detected in several countries in different continents. Capsular switches involving serogroups B and C occurred among epidemic strains, raising concerns regarding possible increases in MenB disease, given that vaccines in use in China do not protect against MenB. PMID:29553310

Sequence Typing Confirms that a Predominant Listeria monocytogenes Clone Caused Human Listeriosis Cases and Outbreaks in Canada from 1988 to 2010

PubMed Central

Reimer, Aleisha; Verghese, Bindhu; Lok, Mei; Ziegler, Jennifer; Farber, Jeffrey; Pagotto, Franco; Graham, Morag; Nadon, Celine A.

2012-01-01

Human listeriosis outbreaks in Canada have been predominantly caused by serotype 1/2a isolates with highly similar pulsed-field gel electrophoresis (PFGE) patterns. Multilocus sequence typing (MLST) and multi-virulence-locus sequence typing (MVLST) each identified a diverse population of Listeria monocytogenes isolates, and within that, both methods had congruent subtypes that substantiated a predominant clone (clonal complex 8; virulence type 59; proposed epidemic clone 5 [ECV]) that has been causing human illness across Canada for more than 2 decades. PMID:22337989

Haemophilus ducreyi Outer Membrane Determinants, Including DsrA, Define Two Clonal Populations

PubMed Central

White, Catherine Dinitra; Leduc, Isabelle; Olsen, Bonnie; Jeter, Chrystina; Harris, Chavala; Elkins, Christopher

2005-01-01

The Haemophilus ducreyi outer membrane component DsrA (for ducreyi serum resistance A) is necessary for complete resistance to normal human serum (NHS). When DsrA expression in 19 temporally and geographically diverse clinical isolates of H. ducreyi was examined by Western blotting, 5 of the strains expressed a different immunotype of the DsrA protein (DsrAII) than the well-characterized prototypical strain 35000HP (DsrAI). The predicted DsrA proteins expressed by the DsrAII strains were 100% identical to each other but only 48% identical to that of strain 35000HP. In addition to the DsrAII protein, class II strains also expressed variant forms of other outer membrane proteins (OMPs) including NcaA (necessary for collagen adhesion A), DltA (ducreyi lectin A), Hlp (H. ducreyi lipoprotein), major OMP, and/or OmpA2 (for OMP A2) and synthesized a distinct, faster-migrating lipooligosaccharide. Based on these data, strains expressing DsrAI were termed class I, and those expressing DsrAII were termed class II. Expression of dsrAII from strain CIP 542 ATCC in the class I dsrAI mutant FX517 (35000HP background), which does not express a DsrA protein, rendered this strain resistant to 50% NHS. This demonstrates that DsrAII protein is also critical to serum resistance. Taken together, these results indicate that there are two clonal populations of H. ducreyi. The implications of two classes of H. ducreyi strains differing in important antigenic outer membrane components are discussed. PMID:15784585

Parallel Evolution of Two Clades of an Atlantic-Endemic Pathogenic Lineage of Vibrio parahaemolyticus by Independent Acquisition of Related Pathogenicity Islands

PubMed Central

Xu, Feng; Drees, Kevin P.; Sebra, Robert P.; Jones, Stephen H.

2017-01-01

ABSTRACT Shellfish-transmitted Vibrio parahaemolyticus infections have recently increased from locations with historically low disease incidence, such as the Northeast United States. This change coincided with a bacterial population shift toward human-pathogenic variants occurring in part through the introduction of several Pacific native lineages (ST36, ST43, and ST636) to nearshore areas off the Atlantic coast of the Northeast United States. Concomitantly, ST631 emerged as a major endemic pathogen. Phylogenetic trees of clinical and environmental isolates indicated that two clades diverged from a common ST631 ancestor, and in each of these clades, a human-pathogenic variant evolved independently through acquisition of distinct Vibrio pathogenicity islands (VPaI). These VPaI differ from each other and bear little resemblance to hemolysin-containing VPaI from isolates of the pandemic clonal complex. Clade I ST631 isolates either harbored no hemolysins or contained a chromosome I-inserted island we call VPaIβ that encodes a type 3 secretion system (T3SS2β) typical of Trh hemolysin producers. The more clinically prevalent and clonal ST631 clade II had an island we call VPaIγ that encodes both tdh and trh and that was inserted in chromosome II. VPaIγ was derived from VPaIβ but with some additional acquired elements in common with VPaI carried by pandemic isolates, exemplifying the mosaic nature of pathogenicity islands. Genomics comparisons and amplicon assays identified VPaIγ-type islands containing tdh inserted adjacent to the ure cluster in the three introduced Pacific and most other emergent lineages that collectively cause 67% of infections in the Northeast United States as of 2016. IMPORTANCE The availability of three different hemolysin genotypes in the ST631 lineage provided a unique opportunity to employ genome comparisons to further our understanding of the processes underlying pathogen evolution. The fact that two different pathogenic clades arose in parallel from the same potentially benign lineage by independent VPaI acquisition is surprising considering the historically low prevalence of community members harboring VPaI in waters along the Northeast U.S. coast that could serve as the source of this material. This illustrates a possible predisposition of some lineages to not only acquire foreign DNA but also become human pathogens. Whereas the underlying cause for the expansion of V. parahaemolyticus lineages harboring VPaIγ along the U.S. Atlantic coast and spread of this element to multiple lineages that underlies disease emergence is not known, this work underscores the need to define the environment factors that favor bacteria harboring VPaI in locations of emergent disease. PMID:28687650

Preleukaemic clonal haemopoiesis and risk of therapy-related myeloid neoplasms: a case-control study.

PubMed

Takahashi, Koichi; Wang, Feng; Kantarjian, Hagop; Doss, Denaha; Khanna, Kanhav; Thompson, Erika; Zhao, Li; Patel, Keyur; Neelapu, Sattva; Gumbs, Curtis; Bueso-Ramos, Carlos; DiNardo, Courtney D; Colla, Simona; Ravandi, Farhad; Zhang, Jianhua; Huang, Xuelin; Wu, Xifeng; Samaniego, Felipe; Garcia-Manero, Guillermo; Futreal, P Andrew

2017-01-01

Therapy-related myeloid neoplasms are secondary malignancies that are often fatal, but their risk factors are not well understood. Evidence suggests that individuals with clonal haemopoiesis have increased risk of developing haematological malignancies. We aimed to identify whether patients with cancer who have clonal haemopoiesis are at an increased risk of developing therapy-related myeloid neoplasms. We did this retrospective case-control study to compare the prevalence of clonal haemopoiesis between patients treated for cancer who later developed therapy-related myeloid neoplasms (cases) and patients who did not develop these neoplasms (controls). All patients in both case and control groups were treated at MD Anderson Cancer Center (Houston, TX, USA) from 1997 to 2015. We used the institutional medical database to locate these patients. Patients were included as cases if they were treated for a primary cancer, subsequently developed therapy-related myeloid neoplasms, and had available paired samples of bone marrow from the time of therapy-related myeloid neoplasm diagnosis and peripheral blood from the time of primary cancer diagnosis. Patients were eligible for inclusion as age-matched controls if they were treated for lymphoma, received combination chemotherapy, and did not develop therapy-related myeloid neoplasms after at least 5 years of follow-up. We used molecular barcode sequencing of 32 genes on the pretreatment peripheral blood samples to detect clonal haemopoiesis. For cases, we also used targeted gene sequencing on bone marrow samples and investigated clonal evolution from clonal haemopoiesis to the development of therapy-related myeloid neoplasms. To further clarify the association between clonal haemopoiesis and therapy-related myeloid neoplasm development, we also analysed the prevalence of clonal haemopoiesis in an external cohort of patients with lymphoma who were treated in a randomised trial of front-line chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone, with or without melatonin. This trial was done at MD Anderson Cancer Center between 1999 and 2001 (protocol number 98-009). We identified 14 cases and 54 controls. Of the 14 cases, we detected clonal haemopoiesis in the peripheral blood samples of ten (71%) patients. We detected clonal haemopoiesis in 17 (31%) of the 54 controls. The cumulative incidence of therapy-related myeloid neoplasms in both cases and controls at 5 years was significantly higher in patients with clonal haemopoiesis (30%, 95% CI 16-51) than in those without (7%, 2-21; p=0·016). In the external cohort, five (7%) of 74 patients developed therapy-related myeloid neoplasms, of whom four (80%) had clonal haemopoiesis; 11 (16%) of 69 patients who did not develop therapy-related myeloid neoplasms had clonal haemopoiesis. In the external cohort, the cumulative incidence of therapy-related myeloid neoplasms at 10 years was significantly higher in patients with clonal haemopoiesis (29%, 95% CI 8-53) than in those without (0%, 0-0; p=0·0009). In a multivariate Fine and Gray model based on the external cohort, the presence of clonal haemopoiesis significantly increased the risk of therapy-related myeloid neoplasm development (hazard ratio 13·7, 95% CI 1·7-108·7; p=0·013). Preleukaemic clonal haemopoiesis is common in patients with therapy-related myeloid neoplasms at the time of their primary cancer diagnosis and before they have been exposed to treatment. Our results suggest that clonal haemopoiesis could be used as a predictive marker to identify patients with cancer who are at risk of developing therapy-related myeloid neoplasms. A prospective trial to validate this concept is warranted. Cancer Prevention Research Institute of Texas, Red and Charline McCombs Institute for the Early Detection and Treatment of Cancer, NIH through MD Anderson Cancer Center Support Grant, and the MD Anderson MDS & AML Moon Shots Program. Copyright © 2017 Elsevier Ltd. All rights reserved.

CLONAL EVOLUTION IN CANCER

PubMed Central

Greaves, Mel; Maley, Carlo C.

2012-01-01

Cancers evolve by a reiterative process of clonal expansion, genetic diversification and clonal selection within the adaptive landscapes of tissue ecosystems. The dynamics are complex with highly variable patterns of genetic diversity and resultant clonal architecture. Therapeutic intervention may decimate cancer clones, and erode their habitats, but inadvertently provides potent selective pressure for the expansion of resistant variants. The inherently Darwinian character of cancer lies at the heart of therapeutic failure but perhaps also holds the key to more effective control. PMID:22258609

Biological and Clinical Implications of Clonal Heterogeneity and Clonal Evolution in Multiple Myeloma.

PubMed

Bianchi, Giada; Ghobrial, Irene M

Clonal heterogeneity and clonal evolution have emerged as critical concepts in the field of oncology over the past four decades, largely thanks to the implementation of novel technologies such as comparative genomic hybridization, whole genome/exome sequencing and epigenetic analysis. Along with the identification of cancer stem cells in the majority of neoplasia, the recognition of intertumor and intratumor variability has provided a novel perspective to understand the mechanisms behind tumor evolution and its implication in terms of treatment failure and cancer relapse or recurrence. First hypothesized over two decades ago, clonal heterogeneity and clonal evolution have been confirmed in multiple myeloma (MM), an incurable cancer of plasma cells, almost universally preceded by a pre-malignant conditioned named monoclonal gammopathy of undetermined significance (MGUS). The genetic events and molecular mechanisms underlying such evolution have been difficult to dissect. Moreover, while a role for the bone marrow microenvironment in supporting MM cell survival, proliferation and drug-resistance has been well established, whether it is directly involved in driving evolution from MGUS to MM is at present unclear. We present in this review a historical excursus on the concepts of clonal heterogeneity and clonal evolution in MM with a special emphasis on their role in the progression from MGUS to MM; the contribution of the microenvironment; and the clinical implications in terms of resistance to treatment and disease relapse/recurrence.

Biological and Clinical Implications of Clonal Heterogeneity and Clonal Evolution in Multiple Myeloma

PubMed Central

Bianchi, Giada; Ghobrial, Irene M.

2015-01-01

Clonal heterogeneity and clonal evolution have emerged as critical concepts in the field of oncology over the past four decades, largely thanks to the implementation of novel technologies such as comparative genomic hybridization, whole genome/exome sequencing and epigenetic analysis. Along with the identification of cancer stem cells in the majority of neoplasia, the recognition of intertumor and intratumor variability has provided a novel perspective to understand the mechanisms behind tumor evolution and its implication in terms of treatment failure and cancer relapse or recurrence. First hypothesized over two decades ago, clonal heterogeneity and clonal evolution have been confirmed in multiple myeloma (MM), an incurable cancer of plasma cells, almost universally preceded by a pre-malignant conditioned named monoclonal gammopathy of undetermined significance (MGUS). The genetic events and molecular mechanisms underlying such evolution have been difficult to dissect. Moreover, while a role for the bone marrow microenvironment in supporting MM cell survival, proliferation and drug-resistance has been well established, whether it is directly involved in driving evolution from MGUS to MM is at present unclear. We present in this review a historical excursus on the concepts of clonal heterogeneity and clonal evolution in MM with a special emphasis on their role in the progression from MGUS to MM; the contribution of the microenvironment; and the clinical implications in terms of resistance to treatment and disease relapse/recurrence. PMID:25705146

Deep sequencing is an appropriate tool for the selection of unique Hepatitis C virus (HCV) variants after single genomic amplification.

PubMed

Guinoiseau, Thibault; Moreau, Alain; Hohnadel, Guillaume; Ngo-Giang-Huong, Nicole; Brulard, Celine; Vourc'h, Patrick; Goudeau, Alain; Gaudy-Graffin, Catherine

2017-01-01

Hepatitis C virus (HCV) evolves rapidly in a single host and circulates as a quasispecies wich is a complex mixture of genetically distinct virus's but closely related namely variants. To identify intra-individual diversity and investigate their functional properties in vitro, it is necessary to define their quasispecies composition and isolate the HCV variants. This is possible using single genome amplification (SGA). This technique, based on serially diluted cDNA to amplify a single cDNA molecule (clonal amplicon), has already been used to determine individual HCV diversity. In these studies, positive PCR reactions from SGA were directly sequenced using Sanger technology. The detection of non-clonal amplicons is necessary for excluding them to facilitate further functional analysis. Here, we compared Next Generation Sequencing (NGS) with De Novo assembly and Sanger sequencing for their ability to distinguish clonal and non-clonal amplicons after SGA on one plasma specimen. All amplicons (n = 42) classified as clonal by NGS were also classified as clonal by Sanger sequencing. No double peaks were seen on electropherograms for non-clonal amplicons with position-specific nucleotide variation below 15% by NGS. Altogether, NGS circumvented many of the difficulties encountered when using Sanger sequencing after SGA and is an appropriate tool to reliability select clonal amplicons for further functional studies.

Deep sequencing is an appropriate tool for the selection of unique Hepatitis C virus (HCV) variants after single genomic amplification

PubMed Central

Guinoiseau, Thibault; Moreau, Alain; Hohnadel, Guillaume; Ngo-Giang-Huong, Nicole; Brulard, Celine; Vourc’h, Patrick; Goudeau, Alain; Gaudy-Graffin, Catherine

2017-01-01

Hepatitis C virus (HCV) evolves rapidly in a single host and circulates as a quasispecies wich is a complex mixture of genetically distinct virus’s but closely related namely variants. To identify intra-individual diversity and investigate their functional properties in vitro, it is necessary to define their quasispecies composition and isolate the HCV variants. This is possible using single genome amplification (SGA). This technique, based on serially diluted cDNA to amplify a single cDNA molecule (clonal amplicon), has already been used to determine individual HCV diversity. In these studies, positive PCR reactions from SGA were directly sequenced using Sanger technology. The detection of non-clonal amplicons is necessary for excluding them to facilitate further functional analysis. Here, we compared Next Generation Sequencing (NGS) with De Novo assembly and Sanger sequencing for their ability to distinguish clonal and non-clonal amplicons after SGA on one plasma specimen. All amplicons (n = 42) classified as clonal by NGS were also classified as clonal by Sanger sequencing. No double peaks were seen on electropherograms for non-clonal amplicons with position-specific nucleotide variation below 15% by NGS. Altogether, NGS circumvented many of the difficulties encountered when using Sanger sequencing after SGA and is an appropriate tool to reliability select clonal amplicons for further functional studies. PMID:28362878

Strong but diverging clonality - climate relationships of different plant clades explain weak overall pattern across China.

PubMed

Ye, Duo; Liu, Guofang; Song, Yao-Bin; Cornwell, William K; Dong, Ming; Cornelissen, Johannes H C

2016-06-01

The clonal strategy should be relatively important in stressful environments (i.e. of low resource availability or harsh climate), e.g. in cold habitats. However, our understanding of the distribution pattern of clonality along environmental gradients is still far from universal. The weakness and inconsistency of overall clonality-climate relationships across taxa, as reported in previous studies, may be due to different phylogenetic lineages having fundamental differences in functional traits other than clonality determining their climate response. Thus, in this study we compared the clonality-climate relationships along a latitudinal gradient within and between different lineages at several taxonomic levels, including four major angiosperm lineages (Magnoliidae, Monocotyledoneae, Superrosidae and Superasteridae), orders and families. To this aim we used a species clonality dataset for 4015 vascular plant species in 545 terrestrial communities across China. Our results revealed clear predictive patterns of clonality proportion in relation to environmental gradients for the predominant representatives of each of the taxonomic levels above, but the relationships differed in shape and strength between the 4 major angiosperm lineages, between the 12 orders and between the 12 families. These different relationships canceled out one another when all lineages at a certain taxonomic level were pooled. Our findings highlight the importance of explicitly accounting for the functional or taxonomic scale for studying variation in plant ecological strategy across environmental gradients.

The clonal origin and clonal evolution of epithelial tumours

PubMed Central

Garcia, Sergio Britto; Novelli, Marco; Wright, Nicholas A

2000-01-01

While the origin of tumours, whether from one cell or many, has been a source of fascination for experimental oncologists for some time, in recent years there has been a veritable explosion of information about the clonal architecture of tumours and their antecedents, stimulated, in the main, by the ready accessibility of new molecular techniques. While most of these new results have apparently confirmed the monoclonal origin of human epithelial (and other) tumours, there are a significant number of studies in which this conclusion just cannot be made. Moreover, analysis of many articles show that the potential impact of such considerations as patch size and clonal evolution on determinations of clonality have largely been ignored, with the result that a number of these studies are confounded. However, the clonal architecture of preneoplastic lesions provide some interesting insights — many lesions which might have been hitherto regarded as hyperplasias are apparently clonal in derivation. If this is indeed true, it calls into some question our hopeful corollary that a monoclonal origin presages a neoplastic habitus. Finally, it is clear, for many reasons, that methods of analysis which involve the disaggregation of tissues, albeit microdissected, are far from ideal and we should be putting more effort into techniques where the clonal architecture of normal tissues, preneoplastic and preinvasive lesions and their derivative tumours can be directly visualized in situ. PMID:10762440

Consequences of clonality for sexual fitness: Clonal expansion enhances fitness under spatially restricted dispersal.

PubMed

Van Drunen, Wendy E; van Kleunen, Mark; Dorken, Marcel E

2015-07-21

Clonality is a pervasive feature of sessile organisms, but this form of asexual reproduction is thought to interfere with sexual fitness via the movement of gametes among the modules that comprise the clone. This within-clone movement of gametes is expected to reduce sexual fitness via mate limitation of male reproductive success and, in some cases, via the production of highly inbred (i.e., self-fertilized) offspring. However, clonality also results in the spatial expansion of the genetic individual (i.e., genet), and this should decrease distances gametes and sexually produced offspring must travel to avoid competing with other gametes and offspring from the same clone. The extent to which any negative effects of clonality on mating success might be offset by the positive effects of spatial expansion is poorly understood. Here, we develop spatially explicit models in which fitness was determined by the success of genets through their male and female sex functions. Our results indicate that clonality serves to increase sexual fitness when it is associated with the outward expansion of the genet. Our models further reveal that the main fitness benefit of clonal expansion might occur through the dispersal of offspring over a wider area compared with nonclonal phenotypes. We conclude that, instead of interfering with sexual reproduction, clonal expansion should often serve to enhance sexual fitness.

MLST typing of antimicrobial-resistant Propionibacterium acnes isolates from patients with moderate to severe acne vulgaris.

PubMed

Giannopoulos, Lambros; Papaparaskevas, Joseph; Refene, Eirini; Daikos, Georgios; Stavrianeas, Nikolaos; Tsakris, Athanassios

2015-02-01

Molecular typing data on antimicrobial-resistant Propionibacterium strains are limited in the literature. We examined antimicrobial resistance profiles and the underlying resistance mechanisms in Propionibacterium spp. isolates recovered from patients with moderate to severe acne vulgaris in Greece. The clonallity of the resistant Propionibacterium acnes isolates was also investigated. Propionibacterium spp. isolates were detected using Tryptone-Yeast Extract-Glucose (TYG) agar plates supplemented with 4% furazolidone. Erythromycin, clindamycin, vancomycin, penicillin, co-trimoxazole, doxycycline, minocycline and ciprofloxacin MICs were determined using the gradient strip method. Erythromycin, clindamycin and tetracycline mechanisms of resistance were determined using PCR and sequencing of the domain V of 23S rRNA and 16S rRNA, as well as the presence of the ermX gene. Typing was performed using the multi locus sequence typing (MLST) methodology. Seventy nine isolates from 76 patients were collected. Twenty-three isolates (29.1%) exhibited resistance to erythromycin and clindamycin, while two additional isolates (2.5%) were resistant only to erythromycin. Resistance to tetracycline was not detected. The underlying molecular mechanisms were point mutations A2059G and A2058G. MLST typing of the P. acnes resistant isolates revealed that lineage type IA1 (ST-1, 3 and 52) prevailed (12/18; 66.7%), whilst lineage type IA2 (ST-2 and 22) accounted for five more isolates (27.8%). Susceptible isolates were more evenly distributed between ST types. Propionibacterium spp. from moderate to severe acne vulgaris in Greece are frequently resistant to erythromycin/clindamycin but not to tetracyclines, mainly due to the point mutations A2059G and A2058G. P. acnes resistant isolates were more clonally related than susceptible ones and belonged to a limited number of MLST types. Copyright © 2014 Elsevier Ltd. All rights reserved.

Multilocus sequence typing analyses of Clostridium perfringens type A strains harboring tpeL and netB genes.

PubMed

Nakano, V; Ignacio, A; Llanco, L; Bueris, V; Sircili, M P; Avila-Campos, M J

2017-04-01

Clostridium perfringens is an anaerobic bacterium ubiquitous in various environments, especially in soil and the gastrointestinal tract of healthy humans and animals. In this study, multilocus sequence typing protocol was used to investigate genotypic relationships among 40 C. perfringens strains isolated from humans and broiler chicken with necrotic enteritis [NE]. The results indicated a few clonal populations, mainly observed in human strains, with 32.5% of all strains associated with one of three clonal complexes and 30 sequences types. The CC-1 cluster showed an interesting and unexpected result because it contained seven strains [six from animals and one of human origin]. Detection assays for toxin genes tpeL and netB were also performed. The netB gene was only observed in 7.5% of the strains from healthy human. The toxin gene tpeL was detected in 22.5% of the C. perfringens strains isolated from three individuals and in six broilers with NE. Our study describes the role of some C. perfringens strains of human origin acting as reservoirs of virulence genes and sources of infection. In addition, the strains of human and animal origin were found to be genetically distinct but phylogenetically close, and the human strains showed more diversity than the animal strains. Copyright © 2017 Elsevier Ltd. All rights reserved.

Population and evolutionary dynamics of Shiga-toxin producing Escherichia coli O157 in a beef herd: A longitudinal study.

PubMed

Jones, Meghan; Octavia, Sophie; Lammers, Geraldine; Heller, Jane; Lan, Ruiting

2017-05-01

Shiga toxin producing Escherichia coli O157:H7 (STEC O157) is naturally found in the gastrointestinal tract of cattle and can cause severe disease in humans. There is limited understanding of the population dynamics and microevolution of STEC O157 at herd level. In this study, isolates from a closed beef herd of 23 cows were used to examine the population turnover in the herd. Of the nine STEC O157 clades previously described, clade 7 was found in 162 of the 169 isolates typed. Multiple locus variable number tandem repeat analysis (MLVA) differentiated 169 isolates into 33 unique MLVA types. Five predominant MLVA types were evident with most of the remaining types containing only a single isolate. MLVA data suggest that over time clonal replacement occurred within the herd. Genome sequencing of 18 selected isolates found that the isolates were divided into four lineages, representing four different 'clones' in the herd. Genome data confirmed clonal replacement over time and provided evidence of cross transmission of strains between cows. The findings enhanced our understanding of the population dynamics of STEC O157 in its natural host that will help developing effective control measures to prevent the spread of the pathogen to the human population. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

Clonally related methicillin-resistant Staphylococcus aureus isolated from short-finned pilot whales (Globicephala macrorhynchus), human volunteers, and a bayfront cetacean rehabilitation facility.

PubMed

Hower, Suzanne; Phillips, Matthew C; Brodsky, Micah; Dameron, Adrienne; Tamargo, Manuel A; Salazar, Norma C; Jackson, Charlene R; Barrett, John B; Davidson, Maureen; Davis, Johnnie; Mukherjee, Sampa; Ewing, Ruth Y; Gidley, Maribeth L; Sinigalliano, Christopher D; Johns, Lisa; Johnson, Frank E; Adebanjo, Olufunmilola; Plano, Lisa R W

2013-05-01

In May of 2011, a live mass stranding of 26 short-finned pilot whales (Globicephala macrorhynchus) occurred in the lower Florida Keys. Five surviving whales were transferred from the original stranding site to a nearby marine mammal rehabilitation facility where they were constantly attended to by a team of volunteers. Bacteria cultured during the routine clinical care of the whales and necropsy of a deceased whale included methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MSSA and MRSA). In order to investigate potential sources or reservoirs of MSSA and MRSA, samples were obtained from human volunteers, whales, seawater, and sand from multiple sites at the facility, nearby recreational beaches, and a canal. Samples were collected on 3 days. The second collection day was 2 weeks after the first, and the third collection day was 2 months after the last animal was removed from the facility. MRSA and MSSA were isolated on each day from the facility when animals and volunteers were present. MSSA was found at an adjacent beach on all three collection days. Isolates were characterized by utilizing a combination of quantitative real-time PCR to determine the presence of mecA and genes associated with virulence, staphylococcal protein A typing, staphylococcal cassette chromosome mec typing, multilocus sequence typing, and pulsed field gel electrophoresis (PFGE). Using these methods, clonally related MRSA were isolated from multiple environmental locations as well as from humans and animals. Non-identical but genetically similar MSSA and MRSA were also identified from distinct sources within this sample pool. PFGE indicated that the majority of MRSA isolates were clonally related to the prototype human strain USA300. These studies support the notion that S. aureus may be shed into an environment by humans or pilot whales and subsequently colonize or infect exposed new hosts.

Intra-Species Genetic Diversity and Clonal Structure of Cryptosporidium parvum in Sheep Farms in a Confined Geographical Area in Northeastern Spain.

PubMed

Ramo, Ana; Monteagudo, Luis V; Del Cacho, Emilio; Sánchez-Acedo, Caridad; Quílez, Joaquín

2016-01-01

A multilocus fragment typing approach including eleven variable-number tandem-repeat (VNTR) loci and the GP60 gene was used to investigate the intra-farm and intra-host genetic diversity of Cryptosporidium parvum in sheep farms in a confined area in northeastern Spain. Genomic DNA samples of 113 C. parvum isolates from diarrheic pre-weaned lambs collected in 49 meat-type sheep farms were analyzed. Loci exhibited various degrees of polymorphism, the finding of 7-9 alleles in the four most variable and discriminatory markers (ML2, Cgd6_5400, Cgd6_3940, and GP60) being remarkable. The combination of alleles at the twelve loci identified a total of 74 multilocus subtypes (MLTs) and provided a Hunter-Gaston discriminatory index of 0.988 (95% CI, 0.979-0.996). The finding that most MLTs (n = 64) were unique to individual farms evidenced that cryptosporidial infection is mainly transmitted within sheep flocks, with herd-to-herd transmission playing a secondary role. Limited intra- host variability was found, since only five isolates were genotypically mixed. In contrast, a significant intra-farm genetic diversity was seen, with the presence of multiple MLTs on more than a half of the farms (28/46), suggesting frequent mutations or genetic exchange through recombination. Comparison with a previous study in calves in northern Spain using the same 12-loci typing approach showed differences in the identity of major alleles at most loci, with a single MLT being shared between lambs and calves. Analysis of evolutionary descent by the algorithm eBURST indicated a high degree of genetic divergence, with over 41% MLTs appearing as singletons along with a high number of clonal complexes, most of them linking only two MLTs. Bayesian Structure analysis and F statistics also revealed the genetic remoteness of most C. parvum isolates and no ancestral population size was chosen. Linkage analysis evidenced a prevalent pattern of clonality within the parasite population.

SIApopr: a computational method to simulate evolutionary branching trees for analysis of tumor clonal evolution.

PubMed

McDonald, Thomas O; Michor, Franziska

2017-07-15

SIApopr (Simulating Infinite-Allele populations) is an R package to simulate time-homogeneous and inhomogeneous stochastic branching processes under a very flexible set of assumptions using the speed of C ++. The software simulates clonal evolution with the emergence of driver and passenger mutations under the infinite-allele assumption. The software is an application of the Gillespie Stochastic Simulation Algorithm expanded to a large number of cell types and scenarios, with the intention of allowing users to easily modify existing models or create their own. SIApopr is available as an R library on Github ( https://github.com/olliemcdonald/siapopr ). Supplementary data are available at Bioinformatics online. michor@jimmy.harvard.edu. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Tracking Genomic Cancer Evolution for Precision Medicine: The Lung TRACERx Study

PubMed Central

Jamal-Hanjani, Mariam; Hackshaw, Alan; Ngai, Yenting; Shaw, Jacqueline; Dive, Caroline; Quezada, Sergio; Middleton, Gary; de Bruin, Elza; Le Quesne, John; Shafi, Seema; Falzon, Mary; Horswell, Stuart; Blackhall, Fiona; Khan, Iftekhar; Janes, Sam; Nicolson, Marianne; Lawrence, David; Forster, Martin; Fennell, Dean; Lee, Siow-Ming; Lester, Jason; Kerr, Keith; Muller, Salli; Iles, Natasha; Smith, Sean; Murugaesu, Nirupa; Mitter, Richard; Salm, Max; Stuart, Aengus; Matthews, Nik; Adams, Haydn; Ahmad, Tanya; Attanoos, Richard; Bennett, Jonathan; Birkbak, Nicolai Juul; Booton, Richard; Brady, Ged; Buchan, Keith; Capitano, Arrigo; Chetty, Mahendran; Cobbold, Mark; Crosbie, Philip; Davies, Helen; Denison, Alan; Djearman, Madhav; Goldman, Jacki; Haswell, Tom; Joseph, Leena; Kornaszewska, Malgorzata; Krebs, Matthew; Langman, Gerald; MacKenzie, Mairead; Millar, Joy; Morgan, Bruno; Naidu, Babu; Nonaka, Daisuke; Peggs, Karl; Pritchard, Catrin; Remmen, Hardy; Rowan, Andrew; Shah, Rajesh; Smith, Elaine; Summers, Yvonne; Taylor, Magali; Veeriah, Selvaraju; Waller, David; Wilcox, Ben; Wilcox, Maggie; Woolhouse, Ian; McGranahan, Nicholas; Swanton, Charles

2014-01-01

The importance of intratumour genetic and functional heterogeneity is increasingly recognised as a driver of cancer progression and survival outcome. Understanding how tumour clonal heterogeneity impacts upon therapeutic outcome, however, is still an area of unmet clinical and scientific need. TRACERx (TRAcking non-small cell lung Cancer Evolution through therapy [Rx]), a prospective study of patients with primary non-small cell lung cancer (NSCLC), aims to define the evolutionary trajectories of lung cancer in both space and time through multiregion and longitudinal tumour sampling and genetic analysis. By following cancers from diagnosis to relapse, tracking the evolutionary trajectories of tumours in relation to therapeutic interventions, and determining the impact of clonal heterogeneity on clinical outcomes, TRACERx may help to identify novel therapeutic targets for NSCLC and may also serve as a model applicable to other cancer types. PMID:25003521

Whole-genome sequencing reveals clonal expansion of multiresistant Staphylococcus haemolyticus in European hospitals.

PubMed

Cavanagh, Jorunn Pauline; Hjerde, Erik; Holden, Matthew T G; Kahlke, Tim; Klingenberg, Claus; Flægstad, Trond; Parkhill, Julian; Bentley, Stephen D; Sollid, Johanna U Ericson

2014-11-01

Staphylococcus haemolyticus is an emerging cause of nosocomial infections, primarily affecting immunocompromised patients. A comparative genomic analysis was performed on clinical S. haemolyticus isolates to investigate their genetic relationship and explore the coding sequences with respect to antimicrobial resistance determinants and putative hospital adaptation. Whole-genome sequencing was performed on 134 isolates of S. haemolyticus from geographically diverse origins (Belgium, 2; Germany, 10; Japan, 13; Norway, 54; Spain, 2; Switzerland, 43; UK, 9; USA, 1). Each genome was individually assembled. Protein coding sequences (CDSs) were predicted and homologous genes were categorized into three types: Type I, core genes, homologues present in all strains; Type II, unique core genes, homologues shared by only a subgroup of strains; and Type III, unique genes, strain-specific CDSs. The phylogenetic relationship between the isolates was built from variable sites in the form of single nucleotide polymorphisms (SNPs) in the core genome and used to construct a maximum likelihood phylogeny. SNPs in the genome core regions divided the isolates into one major group of 126 isolates and one minor group of isolates with highly diverse genomes. The major group was further subdivided into seven clades (A-G), of which four (A-D) encompassed isolates only from Europe. Antimicrobial multiresistance was observed in 77.7% of the collection. High levels of homologous recombination were detected in genes involved in adherence, staphylococcal host adaptation and bacterial cell communication. The presence of several successful and highly resistant clones underlines the adaptive potential of this opportunistic pathogen. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.

Whole-genome sequencing reveals clonal expansion of multiresistant Staphylococcus haemolyticus in European hospitals

PubMed Central

Cavanagh, Jorunn Pauline; Hjerde, Erik; Holden, Matthew T. G.; Kahlke, Tim; Klingenberg, Claus; Flægstad, Trond; Parkhill, Julian; Bentley, Stephen D.; Sollid, Johanna U. Ericson

2014-01-01

Objectives Staphylococcus haemolyticus is an emerging cause of nosocomial infections, primarily affecting immunocompromised patients. A comparative genomic analysis was performed on clinical S. haemolyticus isolates to investigate their genetic relationship and explore the coding sequences with respect to antimicrobial resistance determinants and putative hospital adaptation. Methods Whole-genome sequencing was performed on 134 isolates of S. haemolyticus from geographically diverse origins (Belgium, 2; Germany, 10; Japan, 13; Norway, 54; Spain, 2; Switzerland, 43; UK, 9; USA, 1). Each genome was individually assembled. Protein coding sequences (CDSs) were predicted and homologous genes were categorized into three types: Type I, core genes, homologues present in all strains; Type II, unique core genes, homologues shared by only a subgroup of strains; and Type III, unique genes, strain-specific CDSs. The phylogenetic relationship between the isolates was built from variable sites in the form of single nucleotide polymorphisms (SNPs) in the core genome and used to construct a maximum likelihood phylogeny. Results SNPs in the genome core regions divided the isolates into one major group of 126 isolates and one minor group of isolates with highly diverse genomes. The major group was further subdivided into seven clades (A–G), of which four (A–D) encompassed isolates only from Europe. Antimicrobial multiresistance was observed in 77.7% of the collection. High levels of homologous recombination were detected in genes involved in adherence, staphylococcal host adaptation and bacterial cell communication. Conclusions The presence of several successful and highly resistant clones underlines the adaptive potential of this opportunistic pathogen. PMID:25038069

Importance of whole genome sequencing for the assessment of outbreaks in diagnostic laboratories: analysis of a case series of invasive Streptococcus pyogenes infections.

PubMed

Tagini, F; Aubert, B; Troillet, N; Pillonel, T; Praz, G; Crisinel, P A; Prod'hom, G; Asner, S; Greub, G

2017-07-01

Outbreaks of Streptococcus pyogenes hypervirulent clones are constant public health threats. In western Switzerland, an increase of severe cases of S. pyogenes invasive infections was observed between December 2015 and March 2016. Our aim was (i) to investigate these cases by the use of Whole Genome Sequencing (WGS) and (ii) to determine the specific virulome and resistome of each isolate in order to undertake adequate public health measures. Eleven Streptococcus pyogenes strains isolated from 11 patients with severe invasive infections between December 13, 2015 and March 12, 2016 were included in our study. Practically, emm-typing, MLST and WGS were used to investigate the relatedness between the isolates. The presence of virulence and antibiotic resistance genes as well as mutations in transcriptional regulators of virulence and in genes encoding for antibiotic targets were assessed. Three and two groups of isolates shared the same emm-type and ST type, respectively. Single Nucleotide Polymorphism (SNP) analysis revealed 14 to 32 SNPs between the strains of the same emm-type group, ruling out the possibility of a clonal outbreak. Mutations found in covS and rocA could partially explain an increased virulence. As these reassuring results were obtained in less than 10 days, no specific hospital hygiene and no dedicated public health measures had to be undertaken. WGS is a powerful technique to discriminate between closely related strains, excluding an outbreak in less than 10 days. Moreover, WGS provided extensive data on the virulome and resistome of all these strains.

Ultraviolet B irradiation induces expansion of intraepithelial tumor cells in a tissue model of early cancer progression.

PubMed

Mudgil, Adarsh V; Segal, Nadav; Andriani, Frank; Wang, Youai; Fusenig, Norbert E; Garlick, Jonathan A

2003-07-01

Ultraviolet B irradiation is thought to enable skin cancer progression as clones of genetically damaged keratinocytes escape apoptosis and expand at the expense of adjacent normal cells. Mechanisms through which potentially malignant cells in human skin undergo clonal expansion, however, are not well understood. The goal of this study was to characterize the role of ultraviolet B irradiation on the intraepithelial expansion of early stage human tumor cells in organotypic skin cultures. To accomplish this, we have studied the effect of ultraviolet B irradiation on organotypic cultures that were fabricated by mixing normal human keratinocytes with beta-galactosidase-marked, intraepithelial tumor cells (HaCaT-ras, clone II-4), which bear mutations in both p53 alleles and harbor an activated H-ras oncogene. We found that when organotypic mixtures were exposed to an ultraviolet B dose of 50 mJ per cm2, intraepithelial tumor cells underwent a significant degree of proliferative expansion compared to nonirradiated cultures. To understand this response, organotypic cultures of nor-mal keratinocytes were exposed to ultraviolet B and showed a dose-dependent increase in numbers of sunburn cells and TUNEL-positive cells although their proliferation was suppressed. In contrast, neither the apoptotic nor the proliferative response of II-4 cells was altered by ultraviolet B in organotypic cultures. The differential response of these cell types suggested that II-4 cells were resistant to ultraviolet-B-induced alterations, which allowed these intraepithelial tumor cells to gain a selective growth and survival advantage relative to neighboring normal cells. These findings demonstrate that ultraviolet B exposure can induce the intraepithelial expansion of apoptosis-resistant, p53-mutant, and ras-activated keratinocytes, suggesting that this agent can act to promote the early stages of epithelial carcinogenesis.

Genome-wide-analyses of Listeria monocytogenes from food-processing plants reveal clonal diversity and date the emergence of persisting sequence types.

PubMed

Knudsen, Gitte M; Nielsen, Jesper Boye; Marvig, Rasmus L; Ng, Yin; Worning, Peder; Westh, Henrik; Gram, Lone

2017-08-01

Whole genome sequencing is increasing used in epidemiology, e.g. for tracing outbreaks of food-borne diseases. This requires in-depth understanding of pathogen emergence, persistence and genomic diversity along the food production chain including in food processing plants. We sequenced the genomes of 80 isolates of Listeria monocytogenes sampled from Danish food processing plants over a time-period of 20 years, and analysed the sequences together with 10 public available reference genomes to advance our understanding of interplant and intraplant genomic diversity of L. monocytogenes. Except for three persisting sequence types (ST) based on Multi Locus Sequence Typing being ST7, ST8 and ST121, long-term persistence of clonal groups was limited, and new clones were introduced continuously, potentially from raw materials. No particular gene could be linked to the persistence phenotype. Using time-based phylogenetic analyses of the persistent STs, we estimate the L. monocytogenes evolutionary rate to be 0.18-0.35 single nucleotide polymorphisms/year, suggesting that the persistent STs emerged approximately 100 years ago, which correlates with the onset of industrialization and globalization of the food market. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

Mice infected with low-virulence strains of Toxoplasma gondii lose their innate aversion to cat urine, even after extensive parasite clearance.

PubMed

Ingram, Wendy Marie; Goodrich, Leeanne M; Robey, Ellen A; Eisen, Michael B

2013-01-01

Toxoplasma gondii chronic infection in rodent secondary hosts has been reported to lead to a loss of innate, hard-wired fear toward cats, its primary host. However the generality of this response across T. gondii strains and the underlying mechanism for this pathogen-mediated behavioral change remain unknown. To begin exploring these questions, we evaluated the effects of infection with two previously uninvestigated isolates from the three major North American clonal lineages of T. gondii, Type III and an attenuated strain of Type I. Using an hour-long open field activity assay optimized for this purpose, we measured mouse aversion toward predator and non-predator urines. We show that loss of innate aversion of cat urine is a general trait caused by infection with any of the three major clonal lineages of parasite. Surprisingly, we found that infection with the attenuated Type I parasite results in sustained loss of aversion at times post infection when neither parasite nor ongoing brain inflammation were detectable. This suggests that T. gondii-mediated interruption of mouse innate aversion toward cat urine may occur during early acute infection in a permanent manner, not requiring persistence of parasite cysts or continuing brain inflammation.

Expression Analysis of a Highly Adherent and Cytotoxic Small Colony Variant of Pseudomonas aeruginosa Isolated from a Lung of a Patient with Cystic Fibrosis†

PubMed Central

von Götz, Franz; Häussler, Susanne; Jordan, Doris; Saravanamuthu, Senthil Selvan; Wehmhöner, Dirk; Strüßmann, André; Lauber, Joerg; Attree, Ina; Buer, Jan; Tümmler, Burkhard; Steinmetz, Ivo

2004-01-01

The heterogeneous environment of the lung of the cystic fibrosis (CF) patient gives rise to Pseudomonas aeruginosa small colony variants (SCVs) with increased antibiotic resistance, autoaggregative growth behavior, and an enhanced ability to form biofilms. In this study, oligonucleotide DNA microarrays were used to perform a genome-wide expression study of autoaggregative and highly adherent P. aeruginosa SCV 20265 isolated from a CF patient's lung in comparison with its clonal wild type and a revertant generated in vitro from the SCV population. Most strikingly, SCV 20265 showed a pronounced upregulation of the type III protein secretion system (TTSS) and the respective effector proteins. This differential expression was shown to be biologically meaningful, as SCV 20265 and other hyperpiliated and autoaggregative SCVs with increased TTSS expression were significantly more cytotoxic for macrophages in vitro and were more virulent in a mouse model of respiratory tract infection than the wild type. The observed cytotoxicity and virulence of SCV 20265 required exsA, an important transcriptional activator of the TTSS. Thus, the prevailing assumption that P. aeruginosa is subject to selection towards reduced cytotoxicity and attenuated virulence during chronic CF lung infection might not apply to all clonal variants. PMID:15175297

Enforced Clonality Confers a Fitness Advantage

PubMed Central

Martínková, Jana; Klimešová, Jitka

2016-01-01

In largely clonal plants, splitting of a maternal plant into potentially independent plants (ramets) is usually spontaneous; however, such fragmentation also occurs in otherwise non-clonal species due to application of external force. This process might play an important yet largely overlooked role for otherwise non-clonal plants by providing a mechanism to regenerate after disturbance. Here, in a 5-year garden experiment on two short-lived, otherwise non-clonal species, Barbarea vulgaris and Barbarea stricta, we compared the fitness of plants fragmented by simulated disturbance (“enforced ramets”) both with plants that contemporaneously originate in seed and with individuals unscathed by the disturbance event. Because the ability to regrow from fragments is related to plant age and stored reserves, we compared the effects of disturbance applied during three different ontogenetic stages of the plants. In B. vulgaris, enforced ramet fitness was higher than the measured fitness values of both uninjured plants and plants established from seed after the disturbance. This advantage decreased with increasing plant age at the time of fragmentation. In B. stricta, enforced ramet fitness was lower than or similar to fitness of uninjured plants and plants grown from seed. Our results likely reflect the habitat preferences of the study species, as B. vulgaris occurs in anthropogenic, disturbed habitats where body fragmentation is more probable and enforced clonality thus more advantageous than in the more natural habitats preferred by B. stricta. Generalizing from our results, we see that increased fitness yielded by enforced clonality would confer an evolutionary advantage in the face of disturbance, especially in habitats where a seed bank has not been formed, e.g., during invasion or colonization. Our results thus imply that enforced clonality should be taken into account when studying population dynamics and life strategies of otherwise non-clonal species in disturbed habitats. PMID:26858732

Differential influence of clonal integration on morphological and growth responses to light in two invasive herbs.

PubMed

Xu, Cheng-Yuan; Schooler, Shon S; Van Klinken, Rieks D

2012-01-01

In contrast to seeds, high sensitivity of vegetative fragments to unfavourable environments may limit the expansion of clonal invasive plants. However, clonal integration promotes the establishment of propagules in less suitable habitats and may facilitate the expansion of clonal invaders into intact native communities. Here, we examine the influence of clonal integration on the morphology and growth of ramets in two invasive plants, Alternanthera philoxeroides and Phyla canescens, under varying light conditions. In a greenhouse experiment, branches, connected ramets and severed ramets of the same mother plant were exposed under full sun and 85% shade and their morphological and growth responses were assessed. The influence of clonal integration on the light reaction norm (connection×light interaction) of daughter ramets was species-specific. For A. philoxeroides, clonal integration evened out the light response (total biomass, leaf mass per area, and stem number, diameter and length) displayed in severed ramets, but these connection×light interactions were largely absent for P. canescens. Nevertheless, for both species, clonal integration overwhelmed light effect in promoting the growth of juvenile ramets during early development. Also, vertical growth, as an apparent shade acclimation response, was more prevalent in severed ramets than in connected ramets. Finally, unrooted branches displayed smaller organ size and slower growth than connected ramets, but the pattern of light reaction was similar, suggesting mother plants invest in daughter ramets prior to their own branches. Clonal integration modifies light reaction norms of morphological and growth traits in a species-specific manner for A. philoxeroides and P. canescens, but it improves the establishment of juvenile ramets of both species in light-limiting environments by promoting their growth during early development. This factor may be partially responsible for their ability to successfully colonize native plant communities.

Low incidence of clonality in cold water corals revealed through the novel use of standardized protocol adapted to deep sea sampling

USGS Publications Warehouse

Becheler, Ronan; Cassone, Anne-Laure; Noel, Philippe; Mouchel, Olivier; Morrison, Cheryl L.; Arnaud-Haond, Sophie

2017-01-01

Sampling in the deep sea is a technical challenge, which has hindered the acquisition of robust datasets that are necessary to determine the fine-grained biological patterns and processes that may shape genetic diversity. Estimates of the extent of clonality in deep-sea species, despite the importance of clonality in shaping the local dynamics and evolutionary trajectories, have been largely obscured by such limitations. Cold-water coral reefs along European margins are formed mainly by two reef-building species, Lophelia pertusa and Madrepora oculata. Here we present a fine-grained analysis of the genotypic and genetic composition of reefs occurring in the Bay of Biscay, based on an innovative deep-sea sampling protocol. This strategy was designed to be standardized, random, and allowed the georeferencing of all sampled colonies. Clonal lineages discriminated through their Multi-Locus Genotypes (MLG) at 6–7 microsatellite markers could thus be mapped to assess the level of clonality and the spatial spread of clonal lineages. High values of clonal richness were observed for both species across all sites suggesting a limited occurrence of clonality, which likely originated through fragmentation. Additionally, spatial autocorrelation analysis underlined the possible occurrence of fine-grained genetic structure in several populations of both L. pertusa and M. oculata. The two cold-water coral species examined had contrasting patterns of connectivity among canyons, with among-canyon genetic structuring detected in M. oculata, whereas L. pertusa was panmictic at the canyon scale. This study exemplifies that a standardized, random and georeferenced sampling strategy, while challenging, can be applied in the deep sea, and associated benefits outlined here include improved estimates of fine grained patterns of clonality and dispersal that are comparable across sites and among species.

Propagule Pressure, Habitat Conditions and Clonal Integration Influence the Establishment and Growth of an Invasive Clonal Plant, Alternanthera philoxeroides.

PubMed

You, Wen-Hua; Han, Cui-Min; Fang, Long-Xiang; Du, Dao-Lin

2016-01-01

Many notorious invasive plants are clonal, spreading mainly by vegetative propagules. Propagule pressure (the number of propagules) may affect the establishment, growth, and thus invasion success of these clonal plants, and such effects may also depend on habitat conditions. To understand how propagule pressure, habitat conditions and clonal integration affect the establishment and growth of the invasive clonal plants, an 8-week greenhouse with an invasive clonal plant, Alternanthera philoxeroides was conducted. High (five fragments) or low (one fragment) propagule pressure was established either in bare soil (open habitat) or dense native vegetation of Jussiaea repens (vegetative habitat), with the stolon connections either severed from or connected to the relatively older ramets. High propagule pressure greatly increased the establishment and growth of A. philoxeroides, especially when it grew in vegetative habitats. Surprisingly, high propagule pressure significantly reduced the growth of individual plants of A. philoxeroides in open habitats, whereas it did not affect the individual growth in vegetative habitats. A shift in the intraspecific interaction on A. philoxeroides from competition in open habitats to facilitation in vegetative habitats may be the main reason. Moreover, clonal integration significantly improved the growth of A. philoxeroides only in open habitats, especially with low propagule pressure, whereas it had no effects on the growth and competitive ability of A. philoxeroides in vegetative habitats, suggesting that clonal integration may be of most important for A. philoxeroides to explore new open space and spread. These findings suggest that propagule pressure may be crucial for the invasion success of A. philoxeroides, and such an effect also depends on habitat conditions.

Low incidence of clonality in cold water corals revealed through the novel use of a standardized protocol adapted to deep sea sampling

NASA Astrophysics Data System (ADS)

Becheler, Ronan; Cassone, Anne-Laure; Noël, Philippe; Mouchel, Olivier; Morrison, Cheryl L.; Arnaud-Haond, Sophie

2017-11-01

Sampling in the deep sea is a technical challenge, which has hindered the acquisition of robust datasets that are necessary to determine the fine-grained biological patterns and processes that may shape genetic diversity. Estimates of the extent of clonality in deep-sea species, despite the importance of clonality in shaping the local dynamics and evolutionary trajectories, have been largely obscured by such limitations. Cold-water coral reefs along European margins are formed mainly by two reef-building species, Lophelia pertusa and Madrepora oculata. Here we present a fine-grained analysis of the genotypic and genetic composition of reefs occurring in the Bay of Biscay, based on an innovative deep-sea sampling protocol. This strategy was designed to be standardized, random, and allowed the georeferencing of all sampled colonies. Clonal lineages discriminated through their Multi-Locus Genotypes (MLG) at 6-7 microsatellite markers could thus be mapped to assess the level of clonality and the spatial spread of clonal lineages. High values of clonal richness were observed for both species across all sites suggesting a limited occurrence of clonality, which likely originated through fragmentation. Additionally, spatial autocorrelation analysis underlined the possible occurrence of fine-grained genetic structure in several populations of both L. pertusa and M. oculata. The two cold-water coral species examined had contrasting patterns of connectivity among canyons, with among-canyon genetic structuring detected in M. oculata, whereas L. pertusa was panmictic at the canyon scale. This study exemplifies that a standardized, random and georeferenced sampling strategy, while challenging, can be applied in the deep sea, and associated benefits outlined here include improved estimates of fine grained patterns of clonality and dispersal that are comparable across sites and among species.

Clonal Expansion of the Pseudogymnoascus destructans Genotype in North America Is Accompanied by Significant Variation in Phenotypic Expression

PubMed Central

Khankhet, Jordan; Vanderwolf, Karen J.; McAlpine, Donald F.; McBurney, Scott; Overy, David P.; Slavic, Durda; Xu, Jianping

2014-01-01

Pseudogymnoascus destructans is the causative agent of an emerging infectious disease that threatens populations of several North American bat species. The fungal disease was first observed in 2006 and has since caused the death of nearly six million bats. The disease, commonly known as white-nose syndrome, is characterized by a cutaneous infection with P. destructans causing erosions and ulcers in the skin of nose, ears and/or wings of bats. Previous studies based on sequences from eight loci have found that isolates of P. destructans from bats in the US all belong to one multilocus genotype. Using the same multilocus sequence typing method, we found that isolates from eastern and central Canada also had the same genotype as those from the US, consistent with the clonal expansion of P. destructans into Canada. However, our PCR fingerprinting revealed that among the 112 North American isolates we analyzed, three, all from Canada, showed minor genetic variation. Furthermore, we found significant variations among isolates in mycelial growth rate; the production of mycelial exudates; and pigment production and diffusion into agar media. These phenotypic differences were influenced by culture medium and incubation temperature, indicating significant variation in environmental condition - dependent phenotypic expression among isolates of the clonal P. destructans genotype in North America. PMID:25122221

Clonal expansion of the Pseudogymnoascus destructans genotype in North America is accompanied by significant variation in phenotypic expression.

PubMed

Khankhet, Jordan; Vanderwolf, Karen J; McAlpine, Donald F; McBurney, Scott; Overy, David P; Slavic, Durda; Xu, Jianping

2014-01-01

Pseudogymnoascus destructans is the causative agent of an emerging infectious disease that threatens populations of several North American bat species. The fungal disease was first observed in 2006 and has since caused the death of nearly six million bats. The disease, commonly known as white-nose syndrome, is characterized by a cutaneous infection with P. destructans causing erosions and ulcers in the skin of nose, ears and/or wings of bats. Previous studies based on sequences from eight loci have found that isolates of P. destructans from bats in the US all belong to one multilocus genotype. Using the same multilocus sequence typing method, we found that isolates from eastern and central Canada also had the same genotype as those from the US, consistent with the clonal expansion of P. destructans into Canada. However, our PCR fingerprinting revealed that among the 112 North American isolates we analyzed, three, all from Canada, showed minor genetic variation. Furthermore, we found significant variations among isolates in mycelial growth rate; the production of mycelial exudates; and pigment production and diffusion into agar media. These phenotypic differences were influenced by culture medium and incubation temperature, indicating significant variation in environmental condition--dependent phenotypic expression among isolates of the clonal P. destructans genotype in North America.

Clonal yeast biofilms can reap competitive advantages through cell differentiation without being obligatorily multicellular

PubMed Central

Hanghøj, Kristian Ebbesen; Andersen, Kaj Scherz; Boomsma, Jacobus J.

2016-01-01

How differentiation between cell types evolved is a fundamental question in biology, but few studies have explored single-gene phenotypes that mediate first steps towards division of labour with selective advantage for groups of cells. Here, we show that differential expression of the FLO11 gene produces stable fractions of Flo11+ and Flo11− cells in clonal Saccharomyces cerevisiae biofilm colonies on medium with intermediate viscosity. Differentiated Flo11+/− colonies, consisting of adhesive and non-adhesive cells, obtain a fourfold growth advantage over undifferentiated colonies by overgrowing glucose resources before depleting them, rather than depleting them while they grow as undifferentiated Flo11− colonies do. Flo11+/− colonies maintain their structure and differentiated state by switching non-adhesive cells to adhesive cells with predictable probability. Mixtures of Flo11+ and Flo11− cells from mutant strains that are unable to use this epigenetic switch mechanism produced neither integrated colonies nor growth advantages, so the condition-dependent selective advantages of differentiated FLO11 expression can only be reaped by clone-mate cells. Our results show that selection for cell differentiation in clonal eukaryotes can evolve before the establishment of obligate undifferentiated multicellularity, and without necessarily leading to more advanced organizational complexity. PMID:27807261

Escherichia coli ST131, an Intriguing Clonal Group

PubMed Central

Bertrand, Xavier; Madec, Jean-Yves

2014-01-01

SUMMARY In 2008, a previously unknown Escherichia coli clonal group, sequence type 131 (ST131), was identified on three continents. Today, ST131 is the predominant E. coli lineage among extraintestinal pathogenic E. coli (ExPEC) isolates worldwide. Retrospective studies have suggested that it may originally have risen to prominence as early as 2003. Unlike other classical group B2 ExPEC isolates, ST131 isolates are commonly reported to produce extended-spectrum β-lactamases, such as CTX-M-15, and almost all are resistant to fluoroquinolones. Moreover, ST131 E. coli isolates are considered to be truly pathogenic, due to the spectrum of infections they cause in both community and hospital settings and the large number of virulence-associated genes they contain. ST131 isolates therefore seem to contradict the widely held view that high levels of antimicrobial resistance are necessarily associated with a fitness cost leading to a decrease in pathogenesis. Six years after the first description of E. coli ST131, this review outlines the principal traits of ST131 clonal group isolates, based on the growing body of published data, and highlights what is currently known and what we need to find out to provide public health authorities with better information to help combat ST131. PMID:24982321

Tissue-specific and time-dependent clonal expansion of ENU-induced mutant cells in gpt delta mice.

PubMed

Nakayama, Takafumi; Sawai, Tomoko; Masuda, Ikuko; Kaneko, Shinya; Yamauchi, Kazumi; Blyth, Benjamin J; Shimada, Yoshiya; Tachibana, Akira; Kakinuma, Shizuko

2017-10-01

DNA mutations play a crucial role in the origins of cancer, and the clonal expansion of mutant cells is one of the fundamental steps in multistage carcinogenesis. In this study, we correlated tumor incidence in B6C3F1 mice during the period after exposure to N-ethyl-N-nitrosourea (ENU) with the persistence of ENU-induced mutant clones in transgenic gpt delta B6C3F1 mice. The induced gpt mutations afforded no selective advantage in the mouse cells and could be distinguished by a mutational spectrum that is characteristic of ENU treatment. The gpt mutations were passengers of the mutant cell of origin and its daughter cells and thus could be used as neutral markers of clones that arose and persisted in the tissues. Female B6C3F1 mice exposed for 1 month to 200 ppm ENU in the drinking water developed early thymic lymphomas and late liver and lung tumors. To assay gpt mutations, we sampled the thymus, liver, lung, and small intestine of female gpt delta mice at 3 days, 4 weeks, and 8 weeks after the end of ENU exposure. Our results reveal that, in all four tissues, the ENU-induced gpt mutations persisted for weeks after the end of mutagen exposure. Clonal expansion of mutant cells was observed in the thymus and small intestine, with the thymus showing larger clone sizes. These results indicate that the clearance of mutant cells and the potential for clonal expansion during normal tissue growth depends on tissue type and that these factors may affect the sensitivity of different tissues to carcinogenesis. Environ. Mol. Mutagen. 58:592-606, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Serotypes and Clonal Diversity of Streptococcus pneumoniae Causing Invasive Disease in the Era of PCV13 in Catalonia, Spain

PubMed Central

del Amo, Eva; Esteva, Cristina; Hernandez-Bou, Susanna; Galles, Carmen; Navarro, Marian; Sauca, Goretti; Diaz, Alvaro; Gassiot, Paula; Marti, Carmina; Larrosa, Nieves; Ciruela, Pilar; Jane, Mireia; Sá-Leão, Raquel; Muñoz-Almagro, Carmen

2016-01-01

The aim of this study was to study the serotypes and clonal diversity of pneumococci causing invasive pneumococcal disease in Catalonia, Spain, in the era of 13-valent pneumococcal conjugate vaccine (PCV13). In our region, this vaccine is only available in the private market and it is estimated a PCV13 vaccine coverage around 55% in children. A total of 1551 pneumococcal invasive isolates received between 2010 and 2013 in the Molecular Microbiology Department at Hospital Sant Joan de Déu, Barcelona, were included. Fifty-two serotypes and 249 clonal types—defined by MLST—were identified. The most common serotypes were serotype 1 (n = 182; 11.7%), 3 (n = 145; 9.3%), 19A (n = 137; 8.8%) and 7F (n = 122; 7.9%). Serotype 14 was the third most frequent serotype in children < 2 years (15 of 159 isolates). PCV7 serotypes maintained their proportion along the period of study, 16.6% in 2010 to 13.4% in 2013, whereas there was a significant proportional decrease in PCV13 serotypes, 65.3% in 2010 to 48.9% in 2013 (p<0.01). This decrease was mainly attributable to serotypes 19A and 7F. Serotype 12F achieved the third position in 2013 (n = 22, 6.4%). The most frequent clonal types found were ST306 (n = 154, 9.9%), ST191 (n = 111, 7.2%), ST989 (n = 85, 5.5%) and ST180 (n = 80, 5.2%). Despite their decrease, PCV13 serotypes continue to be a major cause of disease in Spain. These results emphasize the need for complete PCV13 vaccination. PMID:26953887

Clonal architecture of secondary acute myeloid leukemia defined by single-cell sequencing.

PubMed

Hughes, Andrew E O; Magrini, Vincent; Demeter, Ryan; Miller, Christopher A; Fulton, Robert; Fulton, Lucinda L; Eades, William C; Elliott, Kevin; Heath, Sharon; Westervelt, Peter; Ding, Li; Conrad, Donald F; White, Brian S; Shao, Jin; Link, Daniel C; DiPersio, John F; Mardis, Elaine R; Wilson, Richard K; Ley, Timothy J; Walter, Matthew J; Graubert, Timothy A

2014-07-01

Next-generation sequencing has been used to infer the clonality of heterogeneous tumor samples. These analyses yield specific predictions-the population frequency of individual clones, their genetic composition, and their evolutionary relationships-which we set out to test by sequencing individual cells from three subjects diagnosed with secondary acute myeloid leukemia, each of whom had been previously characterized by whole genome sequencing of unfractionated tumor samples. Single-cell mutation profiling strongly supported the clonal architecture implied by the analysis of bulk material. In addition, it resolved the clonal assignment of single nucleotide variants that had been initially ambiguous and identified areas of previously unappreciated complexity. Accordingly, we find that many of the key assumptions underlying the analysis of tumor clonality by deep sequencing of unfractionated material are valid. Furthermore, we illustrate a single-cell sequencing strategy for interrogating the clonal relationships among known variants that is cost-effective, scalable, and adaptable to the analysis of both hematopoietic and solid tumors, or any heterogeneous population of cells.

High Incidence of Invasive Group A Streptococcus Disease Caused by Strains of Uncommon emm Types in Thunder Bay, Ontario, Canada

PubMed Central

Athey, Taryn B. T.; Teatero, Sarah; Sieswerda, Lee E.; Gubbay, Jonathan B.; Marchand-Austin, Alex; Li, Aimin; Wasserscheid, Jessica; Dewar, Ken; McGeer, Allison; Williams, David

2015-01-01

An outbreak of type emm59 invasive group A Streptococcus (iGAS) disease was declared in 2008 in Thunder Bay District, Northwestern Ontario, 2 years after a countrywide emm59 epidemic was recognized in Canada. Despite a declining number of emm59 infections since 2010, numerous cases of iGAS disease continue to be reported in the area. We collected clinical information on all iGAS cases recorded in Thunder Bay District from 2008 to 2013. We also emm typed and sequenced the genomes of all available strains isolated from 2011 to 2013 from iGAS infections and from severe cases of soft tissue infections. We used whole-genome sequencing data to investigate the population structure of GAS strains of the most frequently isolated emm types. We report an increased incidence of iGAS in Thunder Bay compared to the metropolitan area of Toronto/Peel and the province of Ontario. Illicit drug use, alcohol abuse, homelessness, and hepatitis C infection were underlying diseases or conditions that might have predisposed patients to iGAS disease. Most cases were caused by clonal strains of skin or generalist emm types (i.e., emm82, emm87, emm101, emm4, emm83, and emm114) uncommonly seen in other areas of the province. We observed rapid waxing and waning of emm types causing disease and their replacement by other emm types associated with the same tissue tropisms. Thus, iGAS disease in Thunder Bay District predominantly affects a select population of disadvantaged persons and is caused by clonally related strains of a few skin and generalist emm types less commonly associated with iGAS in other areas of Ontario. PMID:26491184

High Incidence of Invasive Group A Streptococcus Disease Caused by Strains of Uncommon emm Types in Thunder Bay, Ontario, Canada.

PubMed

Athey, Taryn B T; Teatero, Sarah; Sieswerda, Lee E; Gubbay, Jonathan B; Marchand-Austin, Alex; Li, Aimin; Wasserscheid, Jessica; Dewar, Ken; McGeer, Allison; Williams, David; Fittipaldi, Nahuel

2016-01-01

An outbreak of type emm59 invasive group A Streptococcus (iGAS) disease was declared in 2008 in Thunder Bay District, Northwestern Ontario, 2 years after a countrywide emm59 epidemic was recognized in Canada. Despite a declining number of emm59 infections since 2010, numerous cases of iGAS disease continue to be reported in the area. We collected clinical information on all iGAS cases recorded in Thunder Bay District from 2008 to 2013. We also emm typed and sequenced the genomes of all available strains isolated from 2011 to 2013 from iGAS infections and from severe cases of soft tissue infections. We used whole-genome sequencing data to investigate the population structure of GAS strains of the most frequently isolated emm types. We report an increased incidence of iGAS in Thunder Bay compared to the metropolitan area of Toronto/Peel and the province of Ontario. Illicit drug use, alcohol abuse, homelessness, and hepatitis C infection were underlying diseases or conditions that might have predisposed patients to iGAS disease. Most cases were caused by clonal strains of skin or generalist emm types (i.e., emm82, emm87, emm101, emm4, emm83, and emm114) uncommonly seen in other areas of the province. We observed rapid waxing and waning of emm types causing disease and their replacement by other emm types associated with the same tissue tropisms. Thus, iGAS disease in Thunder Bay District predominantly affects a select population of disadvantaged persons and is caused by clonally related strains of a few skin and generalist emm types less commonly associated with iGAS in other areas of Ontario. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Molecular epidemiology of Staphylococcus aureus bacteremia in a single large Minnesota medical center in 2015 as assessed using MLST, core genome MLST and spa typing.

PubMed

Park, Kyung-Hwa; Greenwood-Quaintance, Kerryl E; Uhl, James R; Cunningham, Scott A; Chia, Nicholas; Jeraldo, Patricio R; Sampathkumar, Priya; Nelson, Heidi; Patel, Robin

2017-01-01

Staphylococcus aureus is a leading cause of bacteremia in hospitalized patients. Whether or not S. aureus bacteremia (SAB) is associated with clonality, implicating potential nosocomial transmission, has not, however, been investigated. Herein, we examined the epidemiology of SAB using whole genome sequencing (WGS). 152 SAB isolates collected over the course of 2015 at a single large Minnesota medical center were studied. Staphylococcus protein A (spa) typing was performed by PCR/Sanger sequencing; multilocus sequence typing (MLST) and core genome MLST (cgMLST) were determined by WGS. Forty-eight isolates (32%) were methicillin-resistant S. aureus (MRSA). The isolates encompassed 66 spa types, clustered into 11 spa clonal complexes (CCs) and 10 singleton types. 88% of 48 MRSA isolates belonged to spa CC-002 or -008. Methicillin-susceptible S. aureus (MSSA) isolates were more genotypically diverse, with 61% distributed across four spa CCs (CC-002, CC-012, CC-008 and CC-084). By MLST, there was 31 sequence types (STs), including 18 divided into 6 CCs and 13 singleton STs. Amongst MSSA isolates, the common MLST clones were CC5 (23%), CC30 (19%), CC8 (15%) and CC15 (11%). Common MRSA clones were CC5 (67%) and CC8 (25%); there were no MRSA isolates in CC45 or CC30. By cgMLST analysis, there were 9 allelic differences between two isolates, with the remaining 150 isolates differing from each other by over 40 alleles. The two isolates were retroactively epidemiologically linked by medical record review. Overall, cgMLST analysis resulted in higher resolution epidemiological typing than did multilocus sequence or spa typing.

Effects of complex life cycles on genetic diversity: cyclical parthenogenesis.

PubMed

Rouger, R; Reichel, K; Malrieu, F; Masson, J P; Stoeckel, S

2016-11-01

Neutral patterns of population genetic diversity in species with complex life cycles are difficult to anticipate. Cyclical parthenogenesis (CP), in which organisms undergo several rounds of clonal reproduction followed by a sexual event, is one such life cycle. Many species, including crop pests (aphids), human parasites (trematodes) or models used in evolutionary science (Daphnia), are cyclical parthenogens. It is therefore crucial to understand the impact of such a life cycle on neutral genetic diversity. In this paper, we describe distributions of genetic diversity under conditions of CP with various clonal phase lengths. Using a Markov chain model of CP for a single locus and individual-based simulations for two loci, our analysis first demonstrates that strong departures from full sexuality are observed after only a few generations of clonality. The convergence towards predictions made under conditions of full clonality during the clonal phase depends on the balance between mutations and genetic drift. Second, the sexual event of CP usually resets the genetic diversity at a single locus towards predictions made under full sexuality. However, this single recombination event is insufficient to reshuffle gametic phases towards full-sexuality predictions. Finally, for similar levels of clonality, CP and acyclic partial clonality (wherein a fixed proportion of individuals are clonally produced within each generation) differentially affect the distribution of genetic diversity. Overall, this work provides solid predictions of neutral genetic diversity that may serve as a null model in detecting the action of common evolutionary or demographic processes in cyclical parthenogens (for example, selection or bottlenecks).

Antimicrobial efficacy of preoperative skin antisepsis and clonal relationship to postantiseptic skin-and-wound flora in patients undergoing clean orthopedic surgery.

PubMed

Daeschlein, G; Napp, M; Layer, F; von Podewils, S; Haase, H; Spitzmueller, R; Assadian, O; Kasch, R; Werner, G; Jünger, M; Hinz, P; Ekkernkamp, A

2015-11-01

Nosocomial surgical site infections (SSI) are still important complications in surgery. The underlying mechanisms are not fully understood. The aim of this study was to elucidate the possible role of skin flora surviving preoperative antisepsis as a possible cause of SSI. We conducted a two-phase prospective clinical trial in patients undergoing clean orthopedic surgery at a university trauma center in northern Germany. Quantitative swab samples were taken from pre- and postantiseptic skin and, additionally, from the wound base, wound margin, and the suture of 137 patients. Seventy-four patients during phase I and 63 during phase II were investigated. Microbial growth, species spectrum, and antibiotic susceptibility were analyzed. In phase two, the clonal relationship of strains was additionally analyzed. 18.0 % of the swab samples were positive for bacterial growth in the wound base, 24.5 % in the margin, and 27.3 % in the suture. Only 65.5 % of patients showed a 100 % reduction of the skin flora after antisepsis. The microbial spectrum in all postantiseptic samples was dominated by coagulase-negative staphylococci (CoNS). Clonally related staphylococci were detected in ten patients [nine CoNS, one methicillin-susceptible Staphylococcus aureus (MSSA)]. Six of ten patients were suspected of having transmitted identical clones from skin flora into the wound. Ethanol-based antisepsis results in unexpected high levels of skin flora, which can be transmitted into the wound during surgery causing yet unexplained SSI. Keeping with the concept of zero tolerance, further studies are needed in order to understand the origin of this flora to allow further reduction of SSI.

Somatic HLA Mutations Expose the Role of Class I-Mediated Autoimmunity in Aplastic Anemia and its Clonal Complications.

PubMed

Babushok, Daria V; Duke, Jamie L; Xie, Hongbo M; Stanley, Natasha; Atienza, Jamie; Perdigones, Nieves; Nicholas, Peter; Ferriola, Deborah; Li, Yimei; Huang, Hugh; Ye, Wenda; Morrissette, Jennifer J D; Kearns, Jane; Porter, David L; Podsakoff, Gregory M; Eisenlohr, Laurence C; Biegel, Jaclyn A; Chou, Stella T; Monos, Dimitrios S; Bessler, Monica; Olson, Timothy S

2017-10-10

Acquired aplastic anemia (aAA) is an acquired deficiency of early hematopoietic cells, characterized by inadequate blood production, and a predisposition to myelodysplastic syndrome (MDS) and leukemia. Although its exact pathogenesis is unknown, aAA is thought to be driven by Human Leukocyte Antigen (HLA)-restricted T cell immunity, with earlier studies favoring HLA class II-mediated pathways. Using whole exome sequencing (WES), we recently identified two aAA patients with somatic mutations in HLA class I genes. We hypothesized that HLA class I mutations are pathognomonic for autoimmunity in aAA, but were previously underappreciated because the Major Histocompatibility Complex (MHC) region is notoriously difficult to analyze by WES. Using a combination of targeted deep sequencing of HLA class I genes and single nucleotide polymorphism array (SNP-A) genotyping we screened 66 aAA patients for somatic HLA class I loss. We found somatic HLA loss in eleven patients (17%), with thirteen loss-of-function mutations in HLA-A *33:03, HLA-A *68:01, HLA-B *14:02 and HLA-B *40:02 alleles. Three patients had more than one mutation targeting the same HLA allele. Interestingly, HLA-B *14:02 and HLA-B *40:02 were significantly overrepresented in aAA patients, compared to ethnicity-matched controls. Patients who inherited the targeted HLA alleles, regardless of HLA mutation status, had a more severe disease course with more frequent clonal complications as assessed by WES, SNP-A, and metaphase cytogenetics, and more frequent secondary MDS. The finding of recurrent HLA class I mutations provides compelling evidence for a predominant HLA class I-driven autoimmunity in aAA, and establishes a novel link between aAA patients' immunogenetics and clonal evolution.

Somatic HLA mutations expose the role of class I–mediated autoimmunity in aplastic anemia and its clonal complications

PubMed Central

Duke, Jamie L.; Xie, Hongbo M.; Stanley, Natasha; Atienza, Jamie; Perdigones, Nieves; Nicholas, Peter; Ferriola, Deborah; Li, Yimei; Huang, Hugh; Ye, Wenda; Morrissette, Jennifer J. D.; Kearns, Jane; Porter, David L.; Podsakoff, Gregory M.; Eisenlohr, Laurence C.; Biegel, Jaclyn A.; Chou, Stella T.; Monos, Dimitrios S.; Bessler, Monica; Olson, Timothy S.

2017-01-01

Acquired aplastic anemia (aAA) is an acquired deficiency of early hematopoietic cells, characterized by inadequate blood production, and a predisposition to myelodysplastic syndrome (MDS) and leukemia. Although its exact pathogenesis is unknown, aAA is thought to be driven by human leukocyte antigen (HLA)–restricted T cell immunity, with earlier studies favoring HLA class II-mediated pathways. Using whole-exome sequencing (WES), we recently identified 2 patients with aAA with somatic mutations in HLA class I genes. We hypothesized that HLA class I mutations are pathognomonic for autoimmunity in aAA, but were previously underappreciated because the major histocompatibility complex (MHC) region is notoriously difficult to analyze by WES. Using a combination of targeted deep sequencing of HLA class I genes and single nucleotide polymorphism array (SNP-A) genotyping, we screened 66 patients with aAA for somatic HLA class I loss. We found somatic HLA loss in 11 patients (17%), with 13 loss-of-function mutations in HLA-A*33:03, HLA-A*68:01, HLA-B*14:02, and HLA-B*40:02 alleles. Three patients had more than 1 mutation targeting the same HLA allele. Interestingly, HLA-B*14:02 and HLA-B*40:02 were significantly overrepresented in patients with aAA compared with ethnicity-matched controls. Patients who inherited the targeted HLA alleles, regardless of HLA mutation status, had a more severe disease course with more frequent clonal complications as assessed by WES, SNP-A, and metaphase cytogenetics, and more frequent secondary MDS. The finding of recurrent HLA class I mutations provides compelling evidence for a predominant HLA class I-driven autoimmunity in aAA and establishes a novel link between immunogenetics and clonal evolution of patients with aAA. PMID:28971166

Significant spread of extensively drug-resistant Acinetobacter baumannii genotypes of clonal complex 92 among intensive care unit patients in a university hospital in southern Iran.

PubMed

Saffari, Fereshteh; Monsen, Tor; Karmostaji, Afsaneh; Azimabad, Fahimeh Bahadori; Widerström, Micael

2017-11-01

Infections associated with Acinetobacter baumannii represent an increasing threat in healthcare settings. Therefore, we investigated the epidemiological relationship between clinical isolates of A. baumannii obtained from patients in a university hospital in Bandar Abbas in southern Iran. Sixty-four consecutive non-duplicate clinical isolates collected during 2014-2015 were subjected to susceptibility testing, clonal relationship analysis using PFGE, multilocus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST), and examined for the presence of carbapenemases and integrons. Almost all A. baumannii isolates were extensively drug-resistant (XDR; 98 %) and carried an OXA carbapenemase gene (blaOXA-23-like; 98 %) and class 1 integrons (48 %). PFGE and MLST analysis identified three major genotypes, all belonging to clonal complex 92 (CC92): sequence type 848 (ST848) (n=23), ST451 (n=16) and ST195 (n=8). CC92 has previously been documented in the hospital setting in northern Iran, and ST195 has been reported in Arab States of the Persian Gulf. These data suggest national and global transmission of A. baumannii CC92. This report demonstrates the occurrence and potential spread of closely related XDR genotypes of A. baumannii CC92 within a university hospital in southern Iran. These genotypes were found in the majority of the investigated isolates, showed high prevalence of blaOXA-23 and integron class 1, and were associated with stay in the intensive care unit. Very few treatment options remain for healthcare-adapted XDR A. baumannii, and hence effective measures are desperately needed to reduce the spread of these strains and resultant infections in the healthcare setting.

Comparative Analysis of the Orphan CRISPR2 Locus in 242 Enterococcus faecalis Strains

PubMed Central

Hullahalli, Karthik; Rodrigues, Marinelle; Schmidt, Brendan D.; Li, Xiang; Bhardwaj, Pooja; Palmer, Kelli L.

2015-01-01

Clustered, Regularly Interspaced Short Palindromic Repeats and their associated Cas proteins (CRISPR-Cas) provide prokaryotes with a mechanism for defense against mobile genetic elements (MGEs). A CRISPR locus is a molecular memory of MGE encounters. It contains an array of short sequences, called spacers, that generally have sequence identity to MGEs. Three different CRISPR loci have been identified among strains of the opportunistic pathogen Enterococcus faecalis. CRISPR1 and CRISPR3 are associated with the cas genes necessary for blocking MGEs, but these loci are present in only a subset of E. faecalis strains. The orphan CRISPR2 lacks cas genes and is ubiquitous in E. faecalis, although its spacer content varies from strain to strain. Because CRISPR2 is a variable locus occurring in all E. faecalis, comparative analysis of CRISPR2 sequences may provide information about the clonality of E. faecalis strains. We examined CRISPR2 sequences from 228 E. faecalis genomes in relationship to subspecies phylogenetic lineages (sequence types; STs) determined by multilocus sequence typing (MLST), and to a genome phylogeny generated for a representative 71 genomes. We found that specific CRISPR2 sequences are associated with specific STs and with specific branches on the genome tree. To explore possible applications of CRISPR2 analysis, we evaluated 14 E. faecalis bloodstream isolates using CRISPR2 analysis and MLST. CRISPR2 analysis identified two groups of clonal strains among the 14 isolates, an assessment that was confirmed by MLST. CRISPR2 analysis was also used to accurately predict the ST of a subset of isolates. We conclude that CRISPR2 analysis, while not a replacement for MLST, is an inexpensive method to assess clonality among E. faecalis isolates, and can be used in conjunction with MLST to identify recombination events occurring between STs. PMID:26398194

Population Genetics of Lactobacillus sakei Reveals Three Lineages with Distinct Evolutionary Histories

PubMed Central

Chaillou, Stéphane; Lucquin, Isabelle; Najjari, Afef; Zagorec, Monique; Champomier-Vergès, Marie-Christine

2013-01-01

Lactobacillus sakei plays a major role in meat fermentation and in the preservation of fresh meat. The large diversity of L. sakei strains represents a valuable and exploitable asset in the development of a variety of industrial applications; however, an efficient method to identify and classify these strains has yet to be developed. In this study, we used multilocus sequence typing (MLST) to analyze the polymorphism and allelic distribution of eight loci within an L. sakei population of 232 strains collected worldwide. Within this population, we identified 116 unique sequence types with an average pairwise nucleotide diversity per site (π) of 0.13%. Results from Structure, goeBurst, and ClonalFrame software analyses demonstrated that the L. sakei population analyzed here is derived from three ancestral lineages, each of which shows evidence of a unique evolutionary history influenced by independent selection scenarios. However, the signature of selective events in the contemporary population of isolates was somewhat masked by the pervasive phenomenon of homologous recombination. Our results demonstrate that lineage 1 is a completely panmictic subpopulation in which alleles have been continually redistributed through the process of intra-lineage recombination. In contrast, lineage 2 was characterized by a high degree of clonality. Lineage 3, the earliest-diverging branch in the genealogy, showed evidence of both clonality and recombination. These evolutionary histories strongly indicate that the three lineages may correspond to distinct ecotypes, likely linked or specialized to different environmental reservoirs. The MLST scheme developed in this study represents an easy and straightforward tool that can be used to further analyze the population dynamics of L. sakei strains in food products. PMID:24069179

Population genetics of Lactobacillus sakei reveals three lineages with distinct evolutionary histories.

PubMed

Chaillou, Stéphane; Lucquin, Isabelle; Najjari, Afef; Zagorec, Monique; Champomier-Vergès, Marie-Christine

2013-01-01

Lactobacillus sakei plays a major role in meat fermentation and in the preservation of fresh meat. The large diversity of L. sakei strains represents a valuable and exploitable asset in the development of a variety of industrial applications; however, an efficient method to identify and classify these strains has yet to be developed. In this study, we used multilocus sequence typing (MLST) to analyze the polymorphism and allelic distribution of eight loci within an L. sakei population of 232 strains collected worldwide. Within this population, we identified 116 unique sequence types with an average pairwise nucleotide diversity per site (π) of 0.13%. Results from Structure, goeBurst, and ClonalFrame software analyses demonstrated that the L. sakei population analyzed here is derived from three ancestral lineages, each of which shows evidence of a unique evolutionary history influenced by independent selection scenarios. However, the signature of selective events in the contemporary population of isolates was somewhat masked by the pervasive phenomenon of homologous recombination. Our results demonstrate that lineage 1 is a completely panmictic subpopulation in which alleles have been continually redistributed through the process of intra-lineage recombination. In contrast, lineage 2 was characterized by a high degree of clonality. Lineage 3, the earliest-diverging branch in the genealogy, showed evidence of both clonality and recombination. These evolutionary histories strongly indicate that the three lineages may correspond to distinct ecotypes, likely linked or specialized to different environmental reservoirs. The MLST scheme developed in this study represents an easy and straightforward tool that can be used to further analyze the population dynamics of L. sakei strains in food products.

Molecular analysis and distribution of multidrug-resistant Enterococcus faecium isolates belonging to clonal complex 17 in a tertiary care center in Mexico City

PubMed Central

2013-01-01

Background Enterococcus faecium has recently emerged as a multidrug-resistant nosocomial pathogen involved in outbreaks worldwide. A high rate of resistance to different antibiotics has been associated with virulent clonal complex 17 isolates carrying the esp and hyl genes and the purK1 allele. Results Twelve clinical vancomycin-resistant Enterococcus faecium (VREF) isolates were obtained from pediatric patients at the Hospital Infantil de México Federico Gómez (HIMFG). Among these VREF isolates, 58.3% (7/12) were recovered from urine, while 41.7% (5/12) were recovered from the bloodstream. The VREF isolates showed a 100% rate of resistance to ampicillin, amoxicillin-clavulanate, ciprofloxacin, clindamycin, chloramphenicol, streptomycin, gentamicin, rifampicin, erythromycin and teicoplanin. In addition, 16.7% (2/12) of the isolates were resistant to linezolid, and 66.7% (8/12) were resistant to tetracycline and doxycycline. PCR analysis revealed the presence of the vanA gene in all 12 VREF isolates, esp in 83.3% (10/12) of the isolates and hyl in 50% (6/12) of the isolates. Phylogenetic analysis via molecular typing was performed using pulsed-field gel electrophoresis (PFGE) and demonstrated 44% similarity among the VREF isolates. MLST analysis identified four different sequence types (ST412, ST757, ST203 and ST612). Conclusion This study provides the first report of multidrug-resistant VREF isolates belonging to clonal complex 17 from a tertiary care center in Mexico City. Multidrug resistance and genetic determinants of virulence confer advantages among VREF in the colonization of their host. Therefore, the prevention and control of the spread of nosocomial infections caused by VREF is crucial for identifying new emergent subclones that could be challenging to treat in subsequent years. PMID:24330424

Molecular Analysis of Mixed Endometrial Carcinomas Shows Clonality in Most Cases.

PubMed

Köbel, Martin; Meng, Bo; Hoang, Lien N; Almadani, Noorah; Li, Xiaodong; Soslow, Robert A; Gilks, C Blake; Lee, Cheng-Han

2016-02-01

Mixed endometrial carcinoma refers to a tumor that comprises 2 or more distinct histotypes. We studied 18 mixed-type endometrial carcinomas-11 mixed serous and low-grade endometrioid carcinomas (SC/EC), 5 mixed clear cell and low-grade ECs (CCC/EC), and 2 mixed CCC and SCs (CCC/SC), using targeted next-generation sequencing and immunohistochemistry to compare the molecular profiles of the different histotypes present in each case. In 16 of 18 cases there was molecular evidence that both components shared a clonal origin. Eight cases (6 EC/SC, 1 EC/CCC, and 1 SC/CCC) showed an SC molecular profile that was the same in both components. Five cases (3 CCC/EC and 2 SC/EC) showed a shared endometrioid molecular profile and identical mismatch-repair protein deficiency in both components. A single SC/EC case harbored the same POLE exonuclease domain mutation in both components. One SC/CCC and 1 EC/CCC case showed both shared and unique molecular features in the 2 histotype components, suggesting early molecular divergence from a common clonal origin. In 2 cases, there were no shared molecular features, and these appear to be biologically unrelated synchronous tumors. Overall, these results show that the different histologic components in mixed endometrial carcinomas typically share the same molecular aberrations. Mixed endometrial carcinomas most commonly occur through morphologic mimicry, whereby tumors with serous-type molecular profile show morphologic features of EC or CCC, or through underlying deficiency in DNA nucleotide repair, with resulting rapid accrual of mutations and intratumoral phenotypic heterogeneity. Less commonly, mixed endometrial carcinomas are the result of early molecular divergence from a common progenitor clone or are synchronous biologically unrelated tumors (collision tumors).

Molecular analysis of mixed endometrial carcinomas shows clonality in most cases

PubMed Central

Hoang, Lien N.; Almadani, Noorah; Li, Xiaodong; Soslow, Robert A; Gilks, C. Blake; Lee, Cheng-Han

2016-01-01

Mixed endometrial carcinoma refers to a tumor that is comprised of two or more distinct histotypes. We studied 18 mixed-type endometrial carcinomas - 11 mixed serous and low-grade endometrioid carcinomas (SC/EC), 5 mixed clear cell and low-grade endometrioid carcinomas (CCC/EC), and 2 mixed clear cell and serous carcinoma (CCC/SC), using targeted next generation sequencing and immunohistochemistry to compare the molecular profiles of the different histotypes present in each case. In 16 of 18 cases there was molecular evidence that both components shared a clonal origin. Eight cases (6 EC/SC, 1 EC/CCC and 1 SC/CCC) showed a serous carcinoma molecular profile that was the same in both components. Five cases (3 CCC/EC and 2 SC/EC) showed a shared endometrioid molecular profile and identical mismatch repair protein (MMR) deficiency in both components. A single SC/EC case harbored the same POLE exonuclease domain mutation in both components. One SC/CCC and one EC/CCC case showed both shared and unique molecular features in the two histotype components, suggesting early molecular divergence from a common clonal origin. In two cases, there were no shared molecular features and these appear to be biologically unrelated synchronous tumors. Overall, these results show that the different histologic components in mixed endometrial carcinomas typically share the same molecular aberrations. Mixed endometrial carcinomas most commonly occur through morphological mimicry, whereby tumors with serous-type molecular profile show morphological features of endometrioid or clear cell carcinoma, or through underlying deficiency in DNA nucleotide repair, with resulting rapid accrual of mutations and intratumoral phenotypic heterogeneity. Less commonly, mixed endometrial carcinomas are the result of early molecular divergence from a common progenitor clone or are synchronous biologically unrelated tumors (collision tumors). PMID:26492180

Prevalence of Complement-Mediated Cell Lysis-like Gene (sicG) in Streptococcus dysgalactiae subsp. equisimilis Isolates From Japan (2014-2016).

PubMed

Takahashi, Takashi; Fujita, Tomohiro; Shibayama, Akiyoshi; Tsuyuki, Yuzo; Yoshida, Haruno

2017-07-01

Streptococcus dysgalactiae subsp. equisimilis (SDSE; a β-hemolytic streptococcus of human or animal origin) infections are emerging worldwide. We evaluated the clonal distribution of complement-mediated cell lysis-like gene (sicG) among SDSE isolates from three central prefectures of Japan. Group G/C β-hemolytic streptococci were collected from three institutions from April 2014 to March 2016. Fifty-five strains (52 from humans and three from animals) were identified as SDSE on the basis of 16S rRNA sequencing data.; they were obtained from 25 sterile (blood, joint fluid, and cerebrospinal fluid) and 30 non-sterile (skin-, respiratory tract-, and genitourinary tract-origin) samples. emm genotyping, multilocus sequence typing, sicG amplification/sequencing, and random amplified polymorphic DNA (RAPD) analysis of sicG-positive strains were performed. sicG was detected in 30.9% of the isolates (16 human and one canine) and the genes from the 16 human samples (blood, 10; open pus, 3; sputum, 2; throat swab, 1) and one canine sample (open pus) showed the same sequence pattern. All sicG-harboring isolates belonged to clonal complex (CC) 17, and the most prevalent emm type was stG6792 (82.4%). There was a significant association between sicG presence and the development of skin/soft tissue infections. CC17 isolates with sicG could be divided into three subtypes by RAPD analysis. CC17 SDSE harboring sicG might have spread into three closely-related prefectures in central Japan during 2014-2016. Clonal analysis of isolates from other areas might be needed to monitor potentially virulent strains in humans and animals. © The Korean Society for Laboratory Medicine

Identification of Carbapenem-Resistant Klebsiella pneumoniae with Emphasis on New Delhi Metallo-Beta-Lactamase-1 (blaNDM-1) in Bandar Abbas, South of Iran.

PubMed

Shoja, Saeed; Ansari, Maryam; Faridi, Forogh; Azad, Mohsen; Davoodian, Parivash; Javadpour, Sedigheh; Farahani, Abbas; Mobarrez, Banafsheh Douzandeh; Karmostaji, Afsaneh

2018-05-01

The spread of carbapenem-resistant Klebsiella pneumoniae especially bla NDM-1 -carrying isolates is a great concern worldwide. In this study we describe the molecular basis of carbapenem-resistant K. pneumoniae in three teaching hospitals at Bandar Abbas, south of Iran. A total of 170 nonduplicate clinical isolates of K. pneumoniae were investigated. Antimicrobial susceptibility test was performed by disc diffusion method. PCR was carried out for detection of carbapenemase (bla KPC , bla IMP , bla VIM , bla NDM , bla SPM , bla OXA-48 , and bla OXA-181 ) and extended-spectrum β-lactamase (bla CTX-M , bla SHV , bla TEM , bla VEB , bla GES , and bla PER ). Clonal relatedness of bla NDM-1 -positive isolates was evaluated by multilocus sequence typing (MLST). Tigecycline was the most effective antimicrobial agent with 96.5% susceptibility. In addition, 6.5% of the isolates were carbapenem resistant. Bla NDM-1 was identified in four isolates (isolate A-D) and all of them were multidrug-resistant. MLST revealed that bla NDM-1 -positive isolates were clonally related and belonged to two distinct clonal complexes, including sequence type (ST) 13 and ST 392. In addition to bla NDM-1, isolate A coharbored bla SHV-11 , bla CTX-M-15 , and bla TEM-1 , isolate B harbored bla SHV-11 and bla CTX-M-15 , and isolates C and D contained both bla SHV-1 and bla CTX-M-15 . Our results indicate that NDM-1-producing K. pneumoniae ST 13 and ST 392 are disseminated in our region. Moreover, one of our major concerns is that these isolates may be more prevalent in the near future. Tracking and urgent intervention is necessary for control and prevention of these resistant isolates.

Eastern cottonwood clonal mixing study: intergenotypic competition effects

Treesearch

G. Sam Foster; R.J. Rousseau; W.L Nance

1998-01-01

Intergenotypic competition of seven clones of eastern cottonwood (Populus deltoides) was evaluated in a replacement series experiment. A partial diallel competition design was used to choose pairs (binary sets) of clones for plot type treatments. Two separate treatments were established for each pair of clones, namely (1) 75 percent clone A: 25 percent clone B and (2)...

Risk factors for Clostridium difficile infection in a hepatology ward.

PubMed

Vanjak, Dominique; Girault, Guillaume; Branger, Catherine; Rufat, Pierre; Valla, Dominique-Charles; Fantin, Bruno

2007-02-01

During 2001, Clostridium difficile infection was observed in 23 patients hospitalized in a hepatology ward (attack rate, 0.9%). Since strain typing ruled out a clonal dissemination, we performed a case-control study. In addition to antibiotic use as a risk factor, the C. difficile infection rate was higher among patients with autoimmune hepatitis (P<.01).

Orthogonal typing methods identify genetic diversity among Belgian Campylobacter jejuni strains isolated over a decade from poultry and cases of sporadic human illness

USDA-ARS?s Scientific Manuscript database

Campylobacter jejuni is a zoonotic pathogen commonly associated with human gastroenteritis. Retail poultry meat is a major food-related transmission source of C. jejuni to humans. The present study investigated the genetic diversity, clonal relationship, and strain risk-ranking of 403 representativ...

Genetic characterization of Toxoplasma gondii in wildlife from North America revealed widespread and high prevalence of the fourth clonal type

USDA-ARS?s Scientific Manuscript database

Little is known of the genetic diversity of Toxoplasma gondii circulating in wildlife. In the present study, wild animals, including dolphins from the USA were examined for T. gondii infection. Tissues of naturally exposed animals were bioassayed in mice for isolation of viable parasites. Viable T. ...

Alfalfa (Medicago sativa L.).

PubMed

Fu, Chunxiang; Hernandez, Timothy; Zhou, Chuanen; Wang, Zeng-Yu

2015-01-01

Alfalfa (Medicago sativa L.) is a high-quality forage crop widely grown throughout the world. This chapter describes an efficient protocol that allows for the generation of large number of transgenic alfalfa plants by sonication-assisted Agrobacterium-mediated transformation. Binary vectors carrying different selectable marker genes that confer resistance to phosphinothricin (bar), kanamycin (npt II), or hygromycin (hph) were used to generate transgenic alfalfa plants. Intact trifoliates collected from clonally propagated plants in the greenhouse were sterilized with bleach and then inoculated with Agrobacterium strain EHA105. More than 80 % of infected leaf pieces could produce rooted transgenic plants in 4-5 months after Agrobacterium-mediated transformation.

Antimicrobial Resistance Mechanisms and Genetic Diversity of Multidrug-Resistant Acinetobacter baumannii Isolated from a Teaching Hospital in Malaysia.

PubMed

Biglari, Shirin; Hanafiah, Alfizah; Mohd Puzi, Shaliawani; Ramli, Ramliza; Rahman, Mostafizur; Lopes, Bruno Silvester

2017-07-01

Multidrug-resistant (MDR) Acinetobacter baumannii has increasingly emerged as an important nosocomial pathogen. The aim of this study was to determine the resistance profiles and genetic diversity in A. baumannii clinical isolates in a tertiary medical center in Malaysia. The minimum inhibitory concentrations of carbapenems (imipenem and meropenem), cephalosporins (ceftazidime and cefepime), and ciprofloxacin were determined by E-test. PCR and sequencing were carried out for the detection of antibiotic resistance genes and mutations. Clonal relatedness among A. baumannii isolates was determined by REP-PCR. Sequence-based typing of OXA-51 and multilocus sequence typing were performed. One hundred twenty-five of 162 (77.2%) A. baumannii isolates had MDR phenotype. From the 162 A. baumannii isolates, 20 strain types were identified and majority of A. baumannii isolates (66%, n = 107) were classified as strain type 1 and were positive for ISAba1-bla OXA-23 and ISAba1-bla ADC and had mutations in both gyrA and parC genes at positions, 83 and 80, resulting in serine-to-leucine conversion. REP-PCR analysis showed 129 REP types that generated 31 clones with a 90% similarity cutoff value. OXA-66 variant of the bla OXA-51-like genes was predominantly detected among our A. baumannii clinical isolates belonging to ST195 (found in six clones: 1, 8, 9, 19, 27, and 30) and ST208 (found in clone 21). The study helps us in understanding the genetic diversity of A. baumannii isolates in our setting and confirms that international clone II is the most widely distributed clone in Universiti Kebangsaan Malaysia Medical Centre, Malaysia.

Phylogenetic analysis of Mycobacterium massiliense strains having recombinant rpoB gene laterally transferred from Mycobacterium abscessus.

PubMed

Kim, Byoung-Jun; Kim, Ga-Na; Kim, Bo-Ram; Shim, Tae-Sun; Kook, Yoon-Hoh; Kim, Bum-Joon

2017-01-01

Recent multi locus sequence typing (MLST) and genome based studies indicate that lateral gene transfer (LGT) events in the rpoB gene are prevalent between Mycobacterium abscessus complex strains. To check the prevalence of the M. massiliense strains subject to rpoB LGT (Rec-mas), we applied rpoB typing (711 bp) to 106 Korean strains of M. massiliense infection that had already been identified by hsp65 sequence analysis (603 bp). The analysis indicated 6 smooth strains in M. massiliense Type I (10.0%, 6/60) genotypes but no strains in M. massiliense Type II genotypes (0%, 0/46), showing a discrepancy between the 2 typing methods. Further MLST analysis based on the partial sequencing of seven housekeeping genes, argH, cya, glpK, gnd, murC, pta and purH, as well as erm(41) PCR proved that these 6 Rec-mas strains consisted of two distinct genotypes belonging to M. massiliense and not M. abscessus. The complete rpoB sequencing analysis showed that these 6 Rec-mas strains have an identical hybrid rpoB gene, of which a 478 bp partial rpoB fragment may be laterally transferred from M. abscessus. Notably, five of the 6 Rec-mas strains showed complete identical sequences in a total of nine genes, including the seven MLST genes, hsp65, and rpoB, suggesting their clonal propagation in South Korea. In conclusion, we identified 6 M. massiliense smooth strains of 2 phylogenetically distinct genotypes with a specific hybrid rpoB gene laterally transferred from M. abscessus from Korean patients. Their clinical relevance and bacteriological traits remain to be elucidated.

Phylogenetic analysis of Mycobacterium massiliense strains having recombinant rpoB gene laterally transferred from Mycobacterium abscessus

PubMed Central

Kim, Byoung-Jun; Kim, Ga-Na; Kim, Bo-Ram; Shim, Tae-Sun; Kook, Yoon-Hoh

2017-01-01

Recent multi locus sequence typing (MLST) and genome based studies indicate that lateral gene transfer (LGT) events in the rpoB gene are prevalent between Mycobacterium abscessus complex strains. To check the prevalence of the M. massiliense strains subject to rpoB LGT (Rec-mas), we applied rpoB typing (711 bp) to 106 Korean strains of M. massiliense infection that had already been identified by hsp65 sequence analysis (603 bp). The analysis indicated 6 smooth strains in M. massiliense Type I (10.0%, 6/60) genotypes but no strains in M. massiliense Type II genotypes (0%, 0/46), showing a discrepancy between the 2 typing methods. Further MLST analysis based on the partial sequencing of seven housekeeping genes, argH, cya, glpK, gnd, murC, pta and purH, as well as erm(41) PCR proved that these 6 Rec-mas strains consisted of two distinct genotypes belonging to M. massiliense and not M. abscessus. The complete rpoB sequencing analysis showed that these 6 Rec-mas strains have an identical hybrid rpoB gene, of which a 478 bp partial rpoB fragment may be laterally transferred from M. abscessus. Notably, five of the 6 Rec-mas strains showed complete identical sequences in a total of nine genes, including the seven MLST genes, hsp65, and rpoB, suggesting their clonal propagation in South Korea. In conclusion, we identified 6 M. massiliense smooth strains of 2 phylogenetically distinct genotypes with a specific hybrid rpoB gene laterally transferred from M. abscessus from Korean patients. Their clinical relevance and bacteriological traits remain to be elucidated. PMID:28604829

First Isolate of KPC-2-Producing Klebsiella pneumonaie Sequence Type 23 from the Americas

PubMed Central

Cejas, Daniela; Fernández Canigia, Liliana; Rincón Cruz, Giovanna; Elena, Alan X.; Maldonado, Ivana; Gutkind, Gabriel O.

2014-01-01

KPC-2-producing Klebsiella pneumoniae isolates mainly correspond to clonal complex 258 (CC258); however, we describe KPC-2-producing K. pneumoniae isolates belonging to invasive sequence type 23 (ST23). KPC-2 has scarcely been reported to occur in ST23, and this report describes the first isolation of this pathogen in the Americas. Acquisition of resistant markers in virulent clones could mark an evolutionary step toward the establishment of these clones as major nosocomial pathogens. PMID:25031447

Epidemic Typhoid in Vietnam: Molecular Typing of Multiple-Antibiotic-Resistant Salmonella enterica Serotype Typhi from Four Outbreaks

PubMed Central

Connerton, Phillippa; Wain, John; Hien, Tran T.; Ali, Tahir; Parry, Christopher; Chinh, Nguyen T.; Vinh, Ha; Ho, Vo A.; Diep, To S.; Day, Nicholas P. J.; White, Nicholas J.; Dougan, Gordon; Farrar, Jeremy J.

2000-01-01

Multidrug-resistant Salmonella enterica serotype Typhi isolates from four outbreaks of typhoid fever in southern Vietnam between 1993 and 1997 were compared. Pulsed-field gel electrophoresis, bacteriophage and plasmid typing, and antibiotic susceptibilities showed that independent outbreaks of multidrug-resistant typhoid fever in southern Vietnam are caused by single bacterial strains. However, different outbreaks do not derive from the clonal expansion of a single multidrug-resistant serotype Typhi strain. PMID:10655411

Sub-inhibitory concentrations of oxacillin modify the expression of agr locus in Staphylococcus aureus clinical strains belonging to different clonal complexes.

PubMed

Viedma, Esther; Pérez-Montarelo, Dafne; Villa, Jennifer; Muñoz-Gallego, Irene; Larrosa, Nieves; Fernández-Hidalgo, Nuria; Gavaldà, Joan; Almirante, Benito; Chaves, Fernando

2018-04-16

The ability of Staphylococcus aureus to invade tissues and cause an infectious disease is the result of a multi-factorial process supported by the huge number of virulence factors inherent to this microorganism tightly regulated by the accessory gene regulator (agr). During antimicrobial therapy bacteria may be exposed to sub-inhibitory concentrations (subMICs) of antibiotics that may trigger transcriptional changes that may have an impact on the pathogenesis of infection. The objective of this study was to investigate the effect of oxacillin sub-MICs on agr system expression as the key component in the regulation of virulence in methicillin-susceptible (MSSA) and -resistant S. aureus (MRSA) strains. Furthermore, we studied the genetic basis of the agr locus and their potential association with the expression levels. We have examined the expression of RNAIII and agrA mRNA as biomarkers for agr expression in the presence and absence of oxacillin subMICs in 10 MSSA and 4 MRSA clinical strains belonging to 5 clonal complexes (CC45-agrI, CC8-agrI, CC5-agrII, CC15-agrII and CC30-agrIII) causing endovascular complications. The DNA sequences of agr locus were obtained by whole genome sequencing. Our results revealed that exposure to subMICs of oxacillin had an impact on agr locus expression modifying the relative levels of expression with increases in 11 strains and with decreases in 3 strains. Thereby, the exposure to subMICs of oxacillin resulted in higher levels of expression of agr in CC15 and CC45 and lower levels in CC30. We also observed the presence of mutations in agrC and agrA in 13/14 strains with similar mutation profiles among strains within individual CCs except for strains of CC5. Although, agr expression levels differed among strains within CCs, the presence of these mutations was associated with differences in agr expression levels in most cases. Changes in agr expression induced by exposure to oxacillin subMICs should be considered because they could lead to changes in the virulence modulation and have an adverse effect on the course of infection, especially in certain clonal complexes.

Campylobacter jejuni colonization and population structure in urban populations of ducks and starlings in New Zealand

PubMed Central

Mohan, Vathsala; Stevenson, Mark; Marshall, Jonathan; Fearnhead, Paul; Holland, Barbara R; Hotter, Grant; French, Nigel P

2013-01-01

Abstract A repeated cross-sectional study was conducted to determine the prevalence of Campylobacter spp. and the population structure of C. jejuni in European starlings and ducks cohabiting multiple public access sites in an urban area of New Zealand. The country's geographical isolation and relatively recent history of introduction of wild bird species, including the European starling and mallard duck, create an ideal setting to explore the impact of geographical separation on the population biology of C. jejuni, as well as potential public health implications. A total of 716 starling and 720 duck fecal samples were collected and screened for C. jejuni over a 12 month period. This study combined molecular genotyping, population genetics and epidemiological modeling and revealed: (i) higher Campylobacter spp. isolation in starlings (46%) compared with ducks (30%), but similar isolation of C. jejuni in ducks (23%) and starlings (21%), (ii) significant associations between the isolation of Campylobacter spp. and host species, sampling location and time of year using logistic regression, (iii) evidence of population differentiation, as indicated by FST, and host-genotype association with clonal complexes CC ST-177 and CC ST-682 associated with starlings, and clonal complexes CC ST-1034, CC ST-692, and CC ST-1332 associated with ducks, and (iv) greater genetic diversity and genotype richness in ducks compared with starlings. These findings provide evidence that host-associated genotypes, such as the starling-associated ST-177 and ST-682, represent lineages that were introduced with the host species in the 19th century. The isolation of sequence types associated with human disease in New Zealand indicate that wild ducks and starlings need to be considered as a potential public health risk, particularly in urban areas. We applied molecular epidemiology and population genetics to obtain insights in to the population structure, host-species relationships, gene flow and evolution of Campylobacter jejuni in urban ducks and starlings. PMID:23873654

Richter transformation driven by Epstein-Barr virus reactivation during therapy-related immunosuppression in chronic lymphocytic leukaemia.

PubMed

García-Barchino, Maria J; Sarasquete, Maria E; Panizo, Carlos; Morscio, Julie; Martinez, Antonio; Alcoceba, Miguel; Fresquet, Vicente; Gonzalez-Farre, Blanca; Paiva, Bruno; Young, Ken H; Robles, Eloy F; Roa, Sergio; Celay, Jon; Larrayoz, Marta; Rossi, Davide; Gaidano, Gianluca; Montes-Moreno, Santiago; Piris, Miguel A; Balanzategui, Ana; Jimenez, Cristina; Rodriguez, Idoia; Calasanz, Maria J; Larrayoz, Maria J; Segura, Victor; Garcia-Muñoz, Ricardo; Rabasa, Maria P; Yi, Shuhua; Li, Jianyong; Zhang, Mingzhi; Xu-Monette, Zijun Y; Puig-Moron, Noemi; Orfao, Alberto; Böttcher, Sebastian; Hernandez-Rivas, Jesus M; Miguel, Jesus San; Prosper, Felipe; Tousseyn, Thomas; Sagaert, Xavier; Gonzalez, Marcos; Martinez-Climent, Jose A

2018-05-01

The increased risk of Richter transformation (RT) in patients with chronic lymphocytic leukaemia (CLL) due to Epstein-Barr virus (EBV) reactivation during immunosuppressive therapy with fludarabine other targeted agents remains controversial. Among 31 RT cases classified as diffuse large B-cell lymphoma (DLBCL), seven (23%) showed EBV expression. In contrast to EBV - tumours, EBV + DLBCLs derived predominantly from IGVH-hypermutated CLL, and they also showed CLL-unrelated IGVH sequences more frequently. Intriguingly, despite having different cellular origins, clonally related and unrelated EBV + DLBCLs shared a previous history of immunosuppressive chemo-immunotherapy, a non-germinal centre DLBCL phenotype, EBV latency programme type II or III, and very short survival. These data suggested that EBV reactivation during therapy-related immunosuppression can transform either CLL cells or non-tumoural B lymphocytes into EBV + DLBCL. To investigate this hypothesis, xenogeneic transplantation of blood cells from 31 patients with CLL and monoclonal B-cell lymphocytosis (MBL) was performed in Rag2 -/- IL2γc -/- mice. Remarkably, the recipients' impaired immunosurveillance favoured the spontaneous outgrowth of EBV + B-cell clones from 95% of CLL and 64% of MBL patients samples, but not from healthy donors. Eventually, these cells generated monoclonal tumours (mostly CLL-unrelated but also CLL-related), recapitulating the principal features of EBV + DLBCL in patients. Accordingly, clonally related and unrelated EBV + DLBCL xenografts showed indistinguishable cellular, virological and molecular features, and synergistically responded to combined inhibition of EBV replication with ganciclovir and B-cell receptor signalling with ibrutinib in vivo. Our study underscores the risk of RT driven by EBV in CLL patients receiving immunosuppressive therapies, and provides the scientific rationale for testing ganciclovir and ibrutinib in EBV + DLBCL. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Characterizing mutagenesis in the hprt gene of rat alveolar epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Driscoll, K.E.; Deyo, L.C.; Howard, B.W.

1995-12-31

A clonal selection assay was developed for mutation in the hypoxanthine-guanine phosphoribosyl transferase (hprt) gene of rat alveolar epithelial cells. Studies were conducted to establish methods for isolation and long-term culture of rat alveolar epithelial cells. When isolated by pronase digestion purified on a Nycodenz gradient and cultured in media containing 7.5% fetal bovine serum (FBS), pituitary extract, EGF, insulin, and IGF-1, rat alveolar epithelial cells could be maintained in culture for several weeks with cell doubling times of 2-4 days. The rat alveolar epithelial cell cultures were exposed in vitro to the mutagens ethylnitrosourea (ENU) and H{sub 2}O{sub 2},more » and mutation in the hprt gene was selected for by culture in the presence of the toxic purine analog, 6-thioguanine (6TG). In vitro exposure to ENU or H{sub 2}O produced a dose-dependent increase in hprt mutation frequency in the alveolar epithelial cells. To determine if the assay system could be used to evaluate mutagenesis in alveolar type II cells after in vivo mutagen or carcinogen exposure, cells were isolated from rats treated previously with ENU or {alpha}-quartz. A significant increase in hprt mutation frequency was detected in alveolar epithelial cells obtained from rats exposed to ENU or {alpha}-quartz; the latter observation is the first demonstration that crystalline silica exposure is mutagenic in vivo. In summary, these studies show that rat alveolar epithelial cells isolated by pronase digestion and Nycodenz separation techniques and cultured in a defined media can be used in a clonal selection assay for mutation in the hprt gene. This assay demonstrates that ENU and H{sub 2}O{sub 2} in vitro and ENU and {alpha}-quartz in vivo are mutagenic for rat alveolar epithelial cells. This model should be useful for investigating the genotoxic effects of chemical and physical agents on an important lung cell target for neoplastic transformation. 41 refs., 4 figs., 3 tabs.« less

Characterization of extended-spectrum-beta-lactamase-producing Escherichia coli strains involved in maternal-fetal colonization: prevalence of E. coli ST131.

PubMed

Birgy, André; Mariani-Kurkdjian, Patricia; Bidet, Philippe; Doit, Catherine; Genel, Nathalie; Courroux, Céline; Arlet, Guillaume; Bingen, Edouard

2013-06-01

Maternal-fetal Escherichia coli infections, such as neonatal bacteremia and meningitis, are important causes of morbidity and mortality. From 2006 to 2010, we studied newborns and their mothers who were colonized with E. coli in a French hospital in order to document (i) the epidemiology and genetic characteristics of extended-spectrum-beta-lactamase (ESBL)-producing E. coli strains, (ii) the prevalence of associated virulence genes, (iii) the prevalence of clone sequence type 131 (ST131), and (iv) the genetic relationship among ESBL-producing strains. Among the 2,755 E. coli cultures recovered from vaginal or neonatal samples, 68 were ESBL producers (2.46%). We found a wide diversity of ESBL genes, with the majority being bla(CTX-M-14), bla(CTX-M-1), and bla(CTX-M-15), distributed among the 4 main phylogenetic groups. Genes encoding virulence factors were found in 90.7% of the isolates, with ≥ 2 virulence genes present in 76% of cases. The prevalence of ST131 among ESBL-producing E. coli isolates was 9.4% (6/64). Five of these 6 ST131 isolates possessed bla(CTX-M-15) enzymes (and also were resistant to quinolones), and one possessed bla(CTX-M-2) enzymes. Two possessed virulence genes, suggesting the presence of pathogenicity island IIJ96 (PAI IIJ96)-like domains. Pulsed-field gel electrophoresis (PFGE) revealed a high level of genomic diversity overall, except for 3 closely related isolates belonging to clonal group ST131. Repetitive PCR showed that the six ST131 isolates were closely related to ST131 control strains (>95% similarity). This study shows a high prevalence of ESBL-producing E. coli strains and clonal group ST131 in the French maternal-fetal population. These results suggest a widespread distribution of ESBL enzymes in the community and highlight the early transmission between mothers and neonates. These findings are worrisome, especially for this particularly vulnerable population.

Presence of qnr gene in Escherichia coli and Klebsiella pneumoniae resistant to ciprofloxacin isolated from pediatric patients in China.

PubMed

Wang, Aihua; Yang, Yonghong; Lu, Quan; Wang, Yi; Chen, Yuan; Deng, Li; Ding, Hui; Deng, Qiulian; Zhang, Hong; Wang, Chuanqing; Liu, Lan; Xu, Xiwei; Wang, Li; Shen, Xuzhuang

2008-05-22

Quinolone resistance in Enterobacteriaceae results mainly from mutations in type II DNA topoisomerase genes and/or changes in the expression of outer membrane and efflux pumps. Several recent studies have indicated that plasmid-mediated resistance mechanisms also play a significant role in fluoroquinolone resistance, and its prevalence is increasing worldwide. In China, the presence of the qnr gene in the clinical isolates of Enterobacteriaceae has been reported, but this transmissible quinolone resistance gene has not been detected in strains isolated singly from pediatric patients. Because quinolones associated with a variety of adverse side effects on children, they are not authorized for pediatric use. This study therefore aimed to investigate the presence of the qnr gene in clinical isolates of E. coli and K. pneumoniae from pediatric patients in China. A total 213 of non-repetitive clinical isolates resistant to ciprofloxacin from E. coli and K. pneumoniae were collected from hospitalized patients at five children's hospital in Beijing, Shanghai, Guangzhou, and Chongqing. The isolates were screened for the plasmid-mediated quinolone resistance genes of qnrA, qnrB, and qnrS by PCR. Transferability was examined by conjugation with the sodium azide-resistant E. coli J53. All qnr-positive were analyzed for clonality by enterobacterial repetitive intergenic consensus (ERIC)-PCR. The study found that 19 ciprofloxacin-resistant clinical isolates of E. coli and K. pneumoniae were positive for the qnr gene, and most of the qnr positive strains were ESBL producers. Conjugation experiments showed that quinolone resitance could be transferred to recipients. Apart from this, different DNA banding patterns were obtained by ERIC-PCR from positive strains, which means that most of them were not clonally related. This report on transferable fluoroquinolone resistance due to the qnr gene among E. coli and K. pneumoniae strains indicated that plasmid-mediated quinolone resistance has emerged in pediatric patients in China.

Clonal propagation of eucalyptus in Brazilian nurseries

Treesearch

Ken McNabb; Natal Goncalves; Jose Goncalves

2002-01-01

Brazil has established extensive Eucalyptus plantations to support a growing forest products industry. During the past 25 years, the country has been a pioneer in developing clonal propagation systems to regenerate these highly productive plantations. Original clonal selections optimized disease resistance, coppicing ability, and volume growth, while recent priorities...

Virulence, sporulation, and elicitin production in three clonal lineages of Phytophthora ramorum

USDA-ARS?s Scientific Manuscript database

Phytophthora ramorum populations are clonal and consist of three lineages. Recent studies have shown that the clonal lineages may have varying degrees of aggressiveness on some host species, such as Quercus rubra. In this study, we examined virulence, sporulation and elicitin production of five P. ...

SNP-based differentiation of Phytophthora infestans clonal lineages using locked nucleic acid probes and high resolution melt analysis

USDA-ARS?s Scientific Manuscript database

Phytophthora infestans, the cause of the devastating late blight disease of potato and tomato, exhibits a clonal reproductive lifestyle in North America. Phenotypes such as fungicide sensitivity and host preference are conserved among individuals within clonal lineages, while substantial phenotypic ...

Novel R tools for analysis of genome-wide population genetic data with emphasis on clonality

USDA-ARS?s Scientific Manuscript database

To gain a detailed understanding of how plant microbes evolve and adapt to hosts, pesticides, and other factors, knowledge of the population dynamics and evolutionary history of populations is crucial. Plant pathogen populations are often clonal or partially clonal which requires different analytica...

Chromosome aberrations of clonal origin are present in astronauts' blood lymphocytes.

PubMed

George, K; Durante, M; Willingham, V; Cucinotta, F A

2004-01-01

Radiation-induced chromosome translocations remain in peripheral blood cells over many years, and can potentially be used to measure retrospective doses or prolonged low-dose rate exposures. However, several recent studies have indicated that some individuals possess clones of cells with balanced chromosome abnormalities, which can result in an overestimation of damage and, therefore, influence the accuracy of dose calculations. We carefully examined the patterns of chromosome damage found in the blood lymphocytes of twelve astronauts, and also applied statistical methods to screen for the presence of potential clones. Cells with clonal aberrations were identified in three of the twelve individuals. These clonal cells were present in samples collected both before and after space flight, and yields are higher than previously reported for healthy individuals in this age range (40-52 years of age). The frequency of clonal damage appears to be even greater in chromosomes prematurely condensed in interphase, when compared with equivalent analysis in metaphase cells. The individuals with clonal aberrations were followed-up over several months and the yields of all clones decreased during this period. Since clonal aberrations may be associated with increased risk of tumorigenesis, it is important to accurately identify cells containing clonal rearrangements for risk assessment as well as biodosimetry. Copyright 2003 S. Karger AG, Basel

Chromosome aberrations of clonal origin are present in astronauts' blood lymphocytes

NASA Technical Reports Server (NTRS)

George, K.; Durante, M.; Willingham, V.; Cucinotta, F. A.

2004-01-01

Radiation-induced chromosome translocations remain in peripheral blood cells over many years, and can potentially be used to measure retrospective doses or prolonged low-dose rate exposures. However, several recent studies have indicated that some individuals possess clones of cells with balanced chromosome abnormalities, which can result in an overestimation of damage and, therefore, influence the accuracy of dose calculations. We carefully examined the patterns of chromosome damage found in the blood lymphocytes of twelve astronauts, and also applied statistical methods to screen for the presence of potential clones. Cells with clonal aberrations were identified in three of the twelve individuals. These clonal cells were present in samples collected both before and after space flight, and yields are higher than previously reported for healthy individuals in this age range (40-52 years of age). The frequency of clonal damage appears to be even greater in chromosomes prematurely condensed in interphase, when compared with equivalent analysis in metaphase cells. The individuals with clonal aberrations were followed-up over several months and the yields of all clones decreased during this period. Since clonal aberrations may be associated with increased risk of tumorigenesis, it is important to accurately identify cells containing clonal rearrangements for risk assessment as well as biodosimetry. Copyright 2003 S. Karger AG, Basel.

Three-gene identity coefficients demonstrate that clonal reproduction promotes inbreeding and spatial relatedness in yellow-cedar, Callitropsis nootkatensis.

PubMed

Thompson, Stacey Lee; Bérubé, Yanik; Bruneau, Anne; Ritland, Kermit

2008-10-01

Asexual reproduction has the potential to promote population structuring through matings between clones as well as through limited dispersal of related progeny. Here we present an application of three-gene identity coefficients that tests whether clonal reproduction promotes inbreeding and spatial relatedness within populations. With this method, the first two genes are sampled to estimate pairwise relatedness or inbreeding, whereas the third gene is sampled from either a clone or a sexually derived individual. If three-gene coefficients are significantly greater for clones than nonclones, then clonality contributes excessively to genetic structure. First, we describe an estimator of three-gene identity and briefly evaluate its properties. We then use this estimator to test the effect of clonality on the genetic structure within populations of yellow-cedar (Callitropsis nootkatensis) using a molecular marker survey. Five microsatellite loci were genotyped for 485 trees sampled from nine populations. Our three-gene analyses show that clonal ramets promote inbreeding and spatial structure in most populations. Among-population correlations between clonal extent and genetic structure generally support these trends, yet with less statistical significance. Clones appear to contribute to genetic structure through the limited dispersal of offspring from replicated ramets of the same clonal genet, whereas this structure is likely maintained by mating among these relatives.

MLST genotypes of Campylobacter jejuni isolated from broiler products, dairy cattle and human campylobacteriosis cases in Lithuania.

PubMed

Ramonaite, Sigita; Tamuleviciene, Egle; Alter, Thomas; Kasnauskyte, Neringa; Malakauskas, Mindaugas

2017-06-15

Campylobacter (C.) jejuni is the leading cause of human campylobacteriosis worldwide. We performed a molecular epidemiological study to investigate the genetic relationship among C. jejuni strains isolated from human diarrhoeal patients, broiler products and dairy cattle in Lithuania. The C. jejuni isolates from human clinical cases, dairy cattle and broiler products were genotyped using multilocus sequence typing (MLST). Allele numbers for each housekeeping gene, sequence type (ST), and clonal complex (CC) were assigned by submitting the DNA sequences to the C. jejuni MLST database ( http://pubmlst.org/campylobacter ). Based on the obtained sequence data of the housekeeping genes a phylogenetic analysis of the strains was performed and a minimum spanning tree (MST) was calculated. Among the 262 C. jejuni strains (consisting of 43 strains isolated from dairy cattle, 102 strains isolated from broiler products and 117 clinical human C. jejuni strains), 82 different MLST sequence types and 22 clonal complexes were identified. Clonal complexes CC21 and CC353 predominated among the C. jejuni strains. On ST-level, five sequence types (ST-5, ST-21, ST-50, ST-464 and ST-6410) were dominating and these five STs accounted for 35.9% (n = 94) of our isolates. In addition, 51 (19.5%) C. jejuni strains representing 27 (32.9%) STs were reported for the first time in the PubMLST database ( http://pubmlst.org/campylobacter ). The highest Czekanowski index or proportional similarity index (PSI) was calculated for C. jejuni strains isolated from human campylobacteriosis cases and broiler products (PSI = 0.32) suggesting a strong link between broiler strains and human cases. The PSI of dairy cattle and human samples was lower (PSI = 0.11), suggesting a weaker link between bovine strains and human cases. The calculated Simpson's index of all C. jejuni isolates showed a high genetic diversity (D = 0.96). Our results suggest that broiler products are the most important source of human campylobacteriosis in Lithuania. The study provides information on MLST type distribution and genetic relatedness of C. jejuni strains from humans, broiler products and dairy cattle in Lithuania for the first time, enabling a better understanding of the transmission pathways of C. jejuni in this country.

Interleukin-6 inhibits early differentiation of ATDC5 chondrogenic progenitor cells.

PubMed

Nakajima, Shoko; Naruto, Takuya; Miyamae, Takako; Imagawa, Tomoyuki; Mori, Masaaki; Nishimaki, Shigeru; Yokota, Shumpei

2009-08-01

Interleukin (IL)-6 is a causative agent of systemic juvenile idiopathic arthritis (sJIA), a chronic inflammatory disease complicated with severe growth impairment. Recent trials of anti-IL-6 receptor monoclonal antibody, tocilizumab, indicated that tocilizumab blocks IL-6/IL-6 receptor-mediated inflammation, and induces catch-up growth in children with sJIA. This study evaluates the effects of IL-6 on chondrogenesis by ATDC5 cells, a clonal murine chondrogenic cell line that provides an excellent model for studying endochondral ossification at growth plate. ATDC5 cells were examined for the expression of IL-6 receptor and gp130 by fluorescence-activated cell sorting analysis. Recombinant murine IL-6 was added to ATDC5 cultures to observe cell differentiation, using a quantitative RT-PCR for the chondrogenic differentiation markers type II collagen, aggrecan, and type X collagen. To block IL-6, the anti-mouse IL-6 receptor monoclonal antibody MR16-1 was added. As a result, the cells expressed IL-6 receptor and gp130. The expression of chondrogenic differentiation marker gene was reduced by IL-6, but this was abrogated by MR16-1. We conclude that IL-6 inhibits early chondrogenesis of ATDC5 cells suggesting that IL-6 may affect committed stem cells at a cellular level during chondrogenic differentiation of growth plate chondrocytes, and that IL-6 may be a cellular-level factor in growth impairment in sJIA.

TOXOPLASMOSIS

PubMed Central

Halonen, Sandra K.; Weiss, Louis M.

2014-01-01

Toxoplasma gondii, an Apicomplexan, is a pathogen that can infect the central nervous system. Infection during pregnancy can result in a congenial infection with severe neurological sequela. In immune compromised individuals reactivation of latent neurological foci can result in encephalitis. Immune competent individuals infected with T. gondii are typically asymptomatic and maintain this infection for life. However, recent studies suggest that these asymptomatic infections may have effects on behavior and other physiological processes. T. gondii infects approximately one-third of the world population, making it one of the most successful parasitic organisms. Cats and other felidae serve as the definite host producing oocysts, an environmentally resistant life cycle stage found in cat feces, which can transmit the infection when ingested orally. A wide variety of warm-blooded animals, including humans, can serve as the intermediate host in which tissue cysts (containing bradyzoites) develop. Transmission also occurs due to ingestion of the tissue cysts. There are 3 predominant clonal lineages, termed types I, II and III and an association with higher pathogenicity with the Type I strains in humans has emerged. This chapter presents a review of the biology of this infection including the life cycle, transmission, epidemiology, parasite strains, and the host immune response. The major clinical outcomes of congenital infection, chorioretinitis, and encephalitis, and the possible association of infection of toxoplasmosis with neuropsychriatric disorders such as schizophrenia, are reviewed. PMID:23829904

Pathogenesis and Consequences of Uniparental Disomy in Cancer

PubMed Central

Makishima, Hideki; Maciejewski, Jaroslaw P.

2012-01-01

Systematic application of new genome-wide single nucleotide polymorphism arrays has demonstrated that somatically acquired regions of loss of heterozygosity (LOH) without changes in copy number frequently occur in many types of cancer. Until recently, the ubiquity of this type of chromosomal defect had remained unrecognized as it cannot be detected using routine cytogenetic technologies. Random and recurrent patterns of copy-neutral LOH, also referred to as uniparental disomy (UPD), can be found in specific cancer types and probably contribute to clonal outgrowth owing to various mechanisms. In this review we explore the types, topography, genesis, pathophysiological consequences and clinical implications of UPD. PMID:21518781

Multi-Virulence-Locus Sequence Typing of Staphylococcus lugdunensis Generates Results Consistent with a Clonal Population Structure and Is Reliable for Epidemiological Typing

PubMed Central

Didi, Jennifer; Lemée, Ludovic; Gibert, Laure; Pons, Jean-Louis

2014-01-01

Staphylococcus lugdunensis is an emergent virulent coagulase-negative staphylococcus responsible for severe infections similar to those caused by Staphylococcus aureus. To understand its potentially pathogenic capacity and have further detailed knowledge of the molecular traits of this organism, 93 isolates from various geographic origins were analyzed by multi-virulence-locus sequence typing (MVLST), targeting seven known or putative virulence-associated loci (atlLR2, atlLR3, hlb, isdJ, SLUG_09050, SLUG_16930, and vwbl). The polymorphisms of the putative virulence-associated loci were moderate and comparable to those of the housekeeping genes analyzed by multilocus sequence typing (MLST). However, the MVLST scheme generated 43 virulence types (VTs) compared to 20 sequence types (STs) based on MLST, indicating that MVLST was significantly more discriminating (Simpson's index [D], 0.943). No hypervirulent lineage or cluster specific to carriage strains was defined. The results of multilocus sequence analysis of known and putative virulence-associated loci are consistent with a clonal population structure for S. lugdunensis, suggesting a coevolution of these genes with housekeeping genes. Indeed, the nonsynonymous to synonymous evolutionary substitutions (dN/dS) ratio, the Tajima's D test, and Single-likelihood ancestor counting (SLAC) analysis suggest that all virulence-associated loci were under negative selection, even atlLR2 (AtlL protein) and SLUG_16930 (FbpA homologue), for which the dN/dS ratios were higher. In addition, this analysis of virulence-associated loci allowed us to propose a trilocus sequence typing scheme based on the intragenic regions of atlLR3, isdJ, and SLUG_16930, which is more discriminant than MLST for studying short-term epidemiology and further characterizing the lineages of the rare but highly pathogenic S. lugdunensis. PMID:25078912

Epidemiology and emm types of invasive group A streptococcal infections in Finland, 2008-2013.

PubMed

Smit, P W; Lindholm, L; Lyytikäinen, O; Jalava, J; Pätäri-Sampo, A; Vuopio, J

2015-10-01

Invasive Streptococcus pyogenes (group A streptococcus, GAS) infections are a major global cause of morbidity and mortality. We analysed the surveillance data on invasive GAS and the microbiological characteristics of corresponding isolates to assess the incidence and emm type distribution of invasive GAS infections in Finland. Cases defined as patients with isolations of blood and cerebrospinal fluid S. pyogenes are mandatorily notified to the National Infectious Disease Registry and sent to the national reference laboratory for emm typing. Antimicrobial data were collected through the network including all clinical microbiology laboratories. Pulsed-field gel electrophoresis (PFGE) analysis was performed to assess clonality. In total, 1165 cases of invasive GAS were reported in Finland during 2008-2013; the median age was 52 years (range, 0-100) and 54% were male. The overall day 7 case fatality rate was 5.1% (59 cases). The average annual incidence was 3.6 cases per 100,000 population. A total of 1122 invasive GAS isolates (96%) were analysed by emm typing; 72 different emm types were identified, of which emm28 (297 isolates, 26%), emm89 (193 isolates, 12%) and emm1 (132 isolates, 12%) were the most common types. During 2008-2013, an increase of erythromycin resistance (1.9% to 8.7%) and clindamycin (0.9% to 9.2%) was observed. This resistance increase was in parallel with the introduction of a novel clone emm33 into Finland. The overall incidence of invasive GAS infections remained stable over the study period in Finland. We identified clonal spread of macrolide-resistant invasive emm33 GAS type, highlighting the importance of molecular surveillance.

Lineage tracing reveals multipotent stem cells maintain human adenomas and the pattern of clonal expansion in tumor evolution

PubMed Central

Humphries, Adam; Cereser, Biancastella; Gay, Laura J.; Miller, Daniel S. J.; Das, Bibek; Gutteridge, Alice; Elia, George; Nye, Emma; Jeffery, Rosemary; Poulsom, Richard; Novelli, Marco R.; Rodriguez-Justo, Manuel; McDonald, Stuart A. C.; Wright, Nicholas A.; Graham, Trevor A.

2013-01-01

The genetic and morphological development of colorectal cancer is a paradigm for tumorigenesis. However, the dynamics of clonal evolution underpinning carcinogenesis remain poorly understood. Here we identify multipotential stem cells within human colorectal adenomas and use methylation patterns of nonexpressed genes to characterize clonal evolution. Numerous individual crypts from six colonic adenomas and a hyperplastic polyp were microdissected and characterized for genetic lesions. Clones deficient in cytochrome c oxidase (CCO−) were identified by histochemical staining followed by mtDNA sequencing. Topographical maps of clone locations were constructed using a combination of these data. Multilineage differentiation within clones was demonstrated by immunofluorescence. Methylation patterns of adenomatous crypts were determined by clonal bisulphite sequencing; methylation pattern diversity was compared with a mathematical model to infer to clonal dynamics. Individual adenomatous crypts were clonal for mtDNA mutations and contained both mucin-secreting and neuroendocrine cells, demonstrating that the crypt contained a multipotent stem cell. The intracrypt methylation pattern was consistent with the crypts containing multiple competing stem cells. Adenomas were epigenetically diverse populations, suggesting that they were relatively mitotically old populations. Intratumor clones typically showed less diversity in methylation pattern than the tumor as a whole. Mathematical modeling suggested that recent clonal sweeps encompassing the whole adenoma had not occurred. Adenomatous crypts within human tumors contain actively dividing stem cells. Adenomas appeared to be relatively mitotically old populations, pocketed with occasional newly generated subclones that were the result of recent rapid clonal expansion. Relative stasis and occasional rapid subclone growth may characterize colorectal tumorigenesis. PMID:23766371

Lineage tracing reveals multipotent stem cells maintain human adenomas and the pattern of clonal expansion in tumor evolution.

PubMed

Humphries, Adam; Cereser, Biancastella; Gay, Laura J; Miller, Daniel S J; Das, Bibek; Gutteridge, Alice; Elia, George; Nye, Emma; Jeffery, Rosemary; Poulsom, Richard; Novelli, Marco R; Rodriguez-Justo, Manuel; McDonald, Stuart A C; Wright, Nicholas A; Graham, Trevor A

2013-07-02

The genetic and morphological development of colorectal cancer is a paradigm for tumorigenesis. However, the dynamics of clonal evolution underpinning carcinogenesis remain poorly understood. Here we identify multipotential stem cells within human colorectal adenomas and use methylation patterns of nonexpressed genes to characterize clonal evolution. Numerous individual crypts from six colonic adenomas and a hyperplastic polyp were microdissected and characterized for genetic lesions. Clones deficient in cytochrome c oxidase (CCO(-)) were identified by histochemical staining followed by mtDNA sequencing. Topographical maps of clone locations were constructed using a combination of these data. Multilineage differentiation within clones was demonstrated by immunofluorescence. Methylation patterns of adenomatous crypts were determined by clonal bisulphite sequencing; methylation pattern diversity was compared with a mathematical model to infer to clonal dynamics. Individual adenomatous crypts were clonal for mtDNA mutations and contained both mucin-secreting and neuroendocrine cells, demonstrating that the crypt contained a multipotent stem cell. The intracrypt methylation pattern was consistent with the crypts containing multiple competing stem cells. Adenomas were epigenetically diverse populations, suggesting that they were relatively mitotically old populations. Intratumor clones typically showed less diversity in methylation pattern than the tumor as a whole. Mathematical modeling suggested that recent clonal sweeps encompassing the whole adenoma had not occurred. Adenomatous crypts within human tumors contain actively dividing stem cells. Adenomas appeared to be relatively mitotically old populations, pocketed with occasional newly generated subclones that were the result of recent rapid clonal expansion. Relative stasis and occasional rapid subclone growth may characterize colorectal tumorigenesis.

Effects of complex life cycles on genetic diversity: cyclical parthenogenesis

PubMed Central

Rouger, R; Reichel, K; Malrieu, F; Masson, J P; Stoeckel, S

2016-01-01

Neutral patterns of population genetic diversity in species with complex life cycles are difficult to anticipate. Cyclical parthenogenesis (CP), in which organisms undergo several rounds of clonal reproduction followed by a sexual event, is one such life cycle. Many species, including crop pests (aphids), human parasites (trematodes) or models used in evolutionary science (Daphnia), are cyclical parthenogens. It is therefore crucial to understand the impact of such a life cycle on neutral genetic diversity. In this paper, we describe distributions of genetic diversity under conditions of CP with various clonal phase lengths. Using a Markov chain model of CP for a single locus and individual-based simulations for two loci, our analysis first demonstrates that strong departures from full sexuality are observed after only a few generations of clonality. The convergence towards predictions made under conditions of full clonality during the clonal phase depends on the balance between mutations and genetic drift. Second, the sexual event of CP usually resets the genetic diversity at a single locus towards predictions made under full sexuality. However, this single recombination event is insufficient to reshuffle gametic phases towards full-sexuality predictions. Finally, for similar levels of clonality, CP and acyclic partial clonality (wherein a fixed proportion of individuals are clonally produced within each generation) differentially affect the distribution of genetic diversity. Overall, this work provides solid predictions of neutral genetic diversity that may serve as a null model in detecting the action of common evolutionary or demographic processes in cyclical parthenogens (for example, selection or bottlenecks). PMID:27436524

Neisseria meningitidis; clones, carriage, and disease.

PubMed

Read, R C

2014-05-01

Neisseria meningitidis, the cause of meningococcal disease, has been the subject of sophisticated molecular epidemiological investigation as a consequence of the significant public health threat posed by this organism. The use of multilocus sequence typing and whole genome sequencing classifies the organism into clonal complexes. Extensive phenotypic, genotypic and epidemiological information is available on the PubMLST website. The human nasopharynx is the sole ecological niche of this species, and carrier isolates show extensive genetic diversity as compared with hyperinvasive lineages. Horizontal gene exchange and recombinant events within the meningococcal genome during residence in the human nasopharynx result in antigenic diversity even within clonal complexes, so that individual clones may express, for example, more than one capsular polysaccharide (serogroup). Successful clones are capable of wide global dissemination, and may be associated with explosive epidemics of invasive disease. © 2014 The Author Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Clonal Occurrence of Salmonella Weltevreden in Cultured Shrimp in the Mekong Delta, Vietnam

PubMed Central

Noor Uddin, Gazi Md.; Larsen, Marianne Halberg; Barco, Lisa; Minh Phu, Tran; Dalsgaard, Anders

2015-01-01

This study investigated the occurrence, serovar and antimicrobial resistance of Salmonella spp. in shrimp samples from intensive and extensive farms located in three different provinces in the Mekong Delta, Vietnam. Shrimp from 11 of the 48 farms all contained S. Weltevreden, except for one farm yielding S. Agona, with no difference in Salmonella occurrence between the two production systems. Pulsed field gel electrophoresis (PFGE) of S. Weltevreden showed closely related XbaI pulse types, suggesting a clonal relationship despite the farms and shrimp samples being epidemiologically unrelated. S. Weltevreden was susceptible to most antimicrobials tested, with a few strains being resistant to florfenicol, chloramphenicol, sulfamethoxazole or trimethoprim. Future studies of the ecology of S. Weltevreden should establish if this serovar may survive better and even multiply in warm-water shrimp farm environments compared to other Salmonella serovars. PMID:26222547

Clonal Spread in Second Growth Stands of Coast Redwood, Sequoia sempervirens

Treesearch

Vladimir Douhovnikoff; Richard S. Dodd

2007-01-01

Coast redwood (Sequoia sempervirens) is one of the rare conifers to reproduce successfully through clonal spread. The importance of this mode of reproduction in stand development is largely unknown. Understanding the importance of clonal spread and the spatial structure of clones is crucial for stand management strategies that would aim to maximize...

Growth and stem form quality of clonal Pinus taeda following fertilization in the Virginia Piedmont

Treesearch

Jeremy P. Stovall; Colleen A. Carlson; John R. Seiler; Thomas R. Fox

2013-01-01

Clonal forestry offers the opportunity to increase yields, enhance uniformity, and improve wood characteristics. Intensive silvicultural practices, including fertilization, will be required to capture the full growth potential of clonal plantations. However, variation in nutrient use efficiency that exists among clones could affect growth responses. Our research...

GACD: Integrated Software for Genetic Analysis in Clonal F1 and Double Cross Populations.

PubMed

Zhang, Luyan; Meng, Lei; Wu, Wencheng; Wang, Jiankang

2015-01-01

Clonal species are common among plants. Clonal F1 progenies are derived from the hybridization between 2 heterozygous clones. In self- and cross-pollinated species, double crosses can be made from 4 inbred lines. A clonal F1 population can be viewed as a double cross population when the linkage phase is determined. The software package GACD (Genetic Analysis of Clonal F1 and Double cross) is freely available public software, capable of building high-density linkage maps and mapping quantitative trait loci (QTL) in clonal F1 and double cross populations. Three functionalities are integrated in GACD version 1.0: binning of redundant markers (BIN); linkage map construction (CDM); and QTL mapping (CDQ). Output of BIN can be directly used as input of CDM. After adding the phenotypic data, the output of CDM can be used as input of CDQ. Thus, GACD acts as a pipeline for genetic analysis. GACD and example datasets are freely available from www.isbreeding.net. © The American Genetic Association. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Diagnostic value of immunoglobulin κ light chain gene rearrangement analysis in B-cell lymphomas.

PubMed

Kokovic, Ira; Jezersek Novakovic, Barbara; Novakovic, Srdjan

2015-03-01

Analysis of the immunoglobulin κ light chain (IGK) gene is an alternative method for B-cell clonality assessment in the diagnosis of mature B-cell proliferations in which the detection of clonal immunoglobulin heavy chain (IGH) gene rearrangements fails. The aim of the present study was to evaluate the added value of standardized BIOMED-2 assay for the detection of clonal IGK gene rearrangements in the diagnostic setting of suspected B-cell lymphomas. With this purpose, 92 specimens from 80 patients with the final diagnosis of mature B-cell lymphoma (37 specimens), mature T-cell lymphoma (26 specimens) and reactive lymphoid proliferation (29 specimens) were analyzed for B-cell clonality. B-cell clonality analysis was performed using the BIOMED-2 IGH and IGK gene clonality assays. The determined sensitivity of the IGK assay was 67.6%, while the determined sensitivity of the IGH assay was 75.7%. The sensitivity of combined IGH+IGK assay was 81.1%. The determined specificity of the IGK assay was 96.2% in the group of T-cell lymphomas and 96.6% in the group of reactive lesions. The determined specificity of the IGH assay was 84.6% in the group of lymphomas and 86.2% in the group of reactive lesions. The comparison of GeneScan (GS) and heteroduplex pretreatment-polyacrylamide gel electrophoresis (HD-PAGE) methods for the analysis of IGK gene rearrangements showed a higher efficacy of GS analysis in a series of 27 B-cell lymphomas analyzed by both methods. In the present study, we demonstrated that by applying the combined IGH+IGK clonality assay the overall detection rate of B-cell clonality was increased by 5.4%. Thus, we confirmed the added value of the standardized BIOMED-2 IGK assay for assessment of B-cell clonality in suspected B-cell lymphomas with inconclusive clinical and cyto/histological diagnosis.

The maintenance of sex: Ronald Fisher meets the Red Queen.

PubMed

Green, David; Mason, Chris

2013-08-21

Sex in higher diploids carries a two-fold cost of males that should reduce its fitness relative to cloning, and result in its extinction. Instead, sex is widespread and clonal species face early obsolescence. One possible reason is that sex is an adaptation that allows organisms to respond more effectively to endless changes in their environment. The purpose of this study was to model mutation and selection in a diploid organism in an evolving environment and ascertain their support for sex. We used a computational approach to model finite populations where a haploid environment subjects a diploid host to endlessly evolving change. Evolution in both populations is primarily through adoption of novel advantageous mutations within a large allele space. Sex outcompetes cloning by two complementary mechanisms. First, sexual diploids adopt advantageous homozygous mutations more rapidly than clonal ones under conditions of lag load (the gap between the actual adaptation of the diploid population and its theoretical optimum). This rate advantage can offset the higher fecundity of cloning. Second, a relative advantage to sex emerges where populations are significantly polymorphic, because clonal polymorphism runs the risk of clonal interference caused by selection on numerous lines of similar adaptation. This interference extends allele lifetime and reduces the rate of adaptation. Sex abolishes the interference, making selection faster and elevating population fitness. Differences in adaptation between sexual and clonal populations increase markedly with the number of loci under selection, the rate of mutation in the host, and a rapidly evolving environment. Clonal interference in these circumstances leads to conditions where the greater fecundity of clones is unable to offset their poor adaptation. Sexual and clonal populations then either co-exist, or sex emerges as the more stable evolutionary strategy. Sex can out-compete clones in a rapidly evolving environment, such as that characterized by pathogens, where clonal interference reduces the adaptation of clonal populations and clones adopt advantageous mutations more slowly. Since all organisms carry parasitic loads, the model is of potentially general applicability.

Longevity of clonal plants: why it matters and how to measure it

PubMed Central

de Witte, Lucienne C.; Stöcklin, Jürg

2010-01-01

Background Species' life-history and population dynamics are strongly shaped by the longevity of individuals, but life span is one of the least accessible demographic traits, particularly in clonal plants. Continuous vegetative reproduction of genets enables persistence despite low or no sexual reproduction, affecting genet turnover rates and population stability. Therefore, the longevity of clonal plants is of considerable biological interest, but remains relatively poorly known. Scope Here, we critically review the present knowledge on the longevity of clonal plants and discuss its importance for population persistence. Direct life-span measurements such as growth-ring analysis in woody plants are relatively easy to take, although, for many clonal plants, these methods are not adequate due to the variable growth pattern of ramets and difficult genet identification. Recently, indirect methods have been introduced in which genet size and annual shoot increments are used to estimate genet age. These methods, often based on molecular techniques, allow the investigation of genet size and age structure of whole populations, a crucial issue for understanding their viability and persistence. However, indirect estimates of clonal longevity are impeded because the process of ageing in clonal plants is still poorly understood and because their size and age are not always well correlated. Alternative estimators for genet life span such as somatic mutations have recently been suggested. Conclusions Empirical knowledge on the longevity of clonal species has increased considerably in the last few years. Maximum age estimates are an indicator of population persistence, but are not sufficient to evaluate turnover rates and the ability of long-lived clonal plants to enhance community stability and ecosystem resilience. In order to understand the dynamics of populations it will be necessary to measure genet size and age structure, not only life spans of single individuals, and to use such data for modelling of genet dynamics. PMID:20880935

Regulation of the Prostate Cancer Tumor Microenvironment

DTIC Science & Technology

2016-04-01

adenocarcinomas than wild-type controls at 30 weeks of age. Analysis of tumor infiltrating cells revealed increased infiltration of macrophage lineage...angiogenesis and proliferation2. Conversely, inflammation can trigger the infiltration of cytotoxic immune effector cells, resulting in the production of...clonal CD8+ T cells3. However, the contribution of the tumor infiltrating lymphocytes (TILs) to prostate cancer development, growth, and metastasis

Methicillin-Resistant Staphylococcus aureus ST398 from Human Patients, Upper Austria

PubMed Central

Metz-Gercek, Sigrid; Mittermayer, Helmut

2009-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) clonal type ST398 is usually associated with animals. We examined 1,098 confirmed MRSA samples from human patients and found that 21 were MRSA ST398. Most (16) patients were farmers. Increasing prevalence from 1.3% (2006) to 2.5% (2008) shows emergence of MRSA ST398 in humans in Austria. PMID:19402964

Growth and wood properties of genetically improved loblolly pine: propagation type comparison and genetic parameters

Treesearch

Finto Antony; Laurence Schimleck; Lewis Jordan; Benjamin Hornsby; Joseph Dahlen; Richard Daniels; Alexander Clark; Luis Apiolaza; Dudley Huber

2013-01-01

The use of clonal varieties in forestry offers great potential to improve growth traits (quantity) and wood properties (quality) of loblolly pine (Pinus taeda L.). Loblolly pine trees established via somatic embryogenesis (clones), full-sib zygotic crosses, and half-sib zygotic open-pollinated families were sampled to identify variation in growth and wood properties...

Prevalence of three campylobacter species, C. jejuni, C. coli, and C. lari, using multilocus sequence typing in wild birds of the Mid-Atlantic region, USA.

PubMed

Keller, Judith I; Shriver, W Gregory

2014-01-01

Campylobacter jejuni is responsible for the majority of bacterial foodborne gastroenteritis in the US, usually due to the consumption of undercooked poultry. Research on which avian species transmit the bacterium is limited, especially in the US. We sampled wild birds in three families-Anatidae, Scolopacidae, and Laridae-in eastern North America to determine the prevalence and specific strains of Campylobacter. The overall prevalence of Campylobacter spp. was 9.2% for all wild birds sampled (n = 781). Campylobacter jejuni was the most prevalent species (8.1%), while Campylobacter coli and Campylobacter lari prevalence estimates were low (1.4% and 0.3%, respectively). We used multilocus sequence typing PCR specific to C. jejuni to characterize clonal complexes and sequence types isolated from wild bird samples and detected 13 novel sequence types, along with a clonal complex previously only associated with human disease (ST-658). Wild birds share an increasing amount of habitat with humans as more landscapes become fragmented and developed for human needs. Wild birds are and will remain an important aspect of public health due to their ability to carry and disperse emerging zoonotic pathogens or their arthropod vectors. As basic information such as prevalence is limited or lacking from a majority of wild birds in the US, this study provides further insight into Campylobacter epidemiology, host preference, and strain characterization of C. jejuni.

Population genetics of the wild yeast Saccharomyces paradoxus.

PubMed Central

Johnson, Louise J; Koufopanou, Vassiliki; Goddard, Matthew R; Hetherington, Richard; Schäfer, Stefanie M; Burt, Austin

2004-01-01

Saccharomyces paradoxus is the closest known relative of the well-known S. cerevisiae and an attractive model organism for population genetic and genomic studies. Here we characterize a set of 28 wild isolates from a 10-km(2) sampling area in southern England. All 28 isolates are homothallic (capable of mating-type switching) and wild type with respect to nutrient requirements. Nine wild isolates and two lab strains of S. paradoxus were surveyed for sequence variation at six loci totaling 7 kb, and all 28 wild isolates were then genotyped at seven polymorphic loci. These data were used to calculate nucleotide diversity and number of segregating sites in S. paradoxus and to investigate geographic differentiation, population structure, and linkage disequilibrium. Synonymous site diversity is approximately 0.3%. Extensive incompatibilities between gene genealogies indicate frequent recombination between unlinked loci, but there is no evidence of recombination within genes. Some localized clonal growth is apparent. The frequency of outcrossing relative to inbreeding is estimated at 1.1% on the basis of heterozygosity. Thus, all three modes of reproduction known in the lab (clonal replication, inbreeding, and outcrossing) have been important in molding genetic variation in this species. PMID:15020405

Genetic relationships and clonal population structure of serotype 2 strains of Neisseria meningitidis.

PubMed Central

Caugant, D A; Zollinger, W D; Mocca, L F; Frasch, C E; Whittam, T S; Frøholm, L O; Selander, R K

1987-01-01

Two hundred and thirty-four strains of Neisseria meningitidis, including 94 serotype 2a, 111 serotype 2b, and 19 serotype 2c isolates, together with 10 isolates that were serotyped as 2 with polyvalent antiserum but did not react with monoclonal antibodies, were characterized by the electrophoretic mobilities of 15 metabolic enzymes. Of these enzymes, 14 were polymorphic, and 56 distinctive combinations of alleles at the enzyme loci (electrophoretic types) were identified, among which the mean genetic diversity per locus was 0.413, or about 75% of that recorded for the species N. meningitidis as a whole. Mean genetic diversity among electrophoretic types of the same serotype (2a, 2b, or 2c) was, however, on average, less than half the total species diversity, and no multilocus genotypes were shared between isolates of the different serotypes, which belong to distinctive clonal lineages. Recent temporal changes in the frequencies of recovery of pathogenic strains of serotypes 2a and 2b in South Africa and North America resulted from clone replacement in these populations rather than evolutionary modification of the serotype protein of the initially dominant clones. PMID:3106223

Mitochondrial depolarization in yeast zygotes inhibits clonal expansion of selfish mtDNA.

PubMed

Karavaeva, Iuliia E; Golyshev, Sergey A; Smirnova, Ekaterina A; Sokolov, Svyatoslav S; Severin, Fedor F; Knorre, Dmitry A

2017-04-01

Non-identical copies of mitochondrial DNA (mtDNA) compete with each other within a cell and the ultimate variant of mtDNA present depends on their relative replication rates. Using yeast Saccharomyces cerevisiae cells as a model, we studied the effects of mitochondrial inhibitors on the competition between wild-type mtDNA and mutant selfish mtDNA in heteroplasmic zygotes. We found that decreasing mitochondrial transmembrane potential by adding uncouplers or valinomycin changes the competition outcomes in favor of the wild-type mtDNA. This effect was significantly lower in cells with disrupted mitochondria fission or repression of the autophagy-related genes ATG8 , ATG32 or ATG33 , implying that heteroplasmic zygotes activate mitochondrial degradation in response to the depolarization. Moreover, the rate of mitochondrially targeted GFP turnover was higher in zygotes treated with uncoupler than in haploid cells or untreated zygotes. Finally, we showed that vacuoles of zygotes with uncoupler-activated autophagy contained DNA. Taken together, our data demonstrate that mitochondrial depolarization inhibits clonal expansion of selfish mtDNA and this effect depends on mitochondrial fission and autophagy. These observations suggest an activation of mitochondria quality control mechanisms in heteroplasmic yeast zygotes. © 2017. Published by The Company of Biologists Ltd.

Clonal dominance among T-lymphocyte infiltrates in arthritis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stamenkovic, I.; Stegagno, M.; Wright, K.A.

1988-02-01

Synovial membranes in patients with rheumatoid arthritis as well as other types of chronic destructive inflammatory arthritis contain infiltrates of activated T lymphocytes that probably contribute to the pathogenesis of the disease. In an effort to elucidate the nature of these infiltrates, interleukin 2 (IL-2)-responsive T lymphocytes were grown out of synovial fragments from 14 patients undergoing surgery for advanced destructive inflammatory joint disease. Eleven of the samples examined were from patients with classical rheumatoid arthritis, while three others were obtained from individuals with clinical osteoarthritis. Southern blot analysis of T-cell receptor (TCR) ..beta..-chain genes in 13 of 14 culturesmore » showed distinct rearrangements, indicating that each culture was characterized by the predominance of a limited number of clones. T-cell populations from peripheral blood stimulated with a variety of activators and expanded with IL-2 did not demonstrate evidence of similar clonality in long-term culture. These results suggest that a limited number of activated T-cell clones predominate at the site of tissue injury in rheumatoid synovial membranes as well as in other types of destructive inflammatory joint disease. Further characterization of these T-cell clones may aid our understanding of the pathogenesis of these rheumatic disorders.« less

Clonal Dominance among T-Lymphocyte Infiltrates in Arthritis

NASA Astrophysics Data System (ADS)

Stamenkovic, Ivan; Stegagno, Michele; Wright, Kathryn A.; Krane, Stephen M.; Amento, Edward P.; Colvin, Robert B.; Duquesnoy, Rene J.; Kurnick, James T.

1988-02-01

Synovial membranes in patients with rheumatoid arthritis as well as other types of chronic destructive inflammatory arthritis contain infiltrates of activated T lymphocytes that probably contribute to the pathogenesis of the disease. In an effort to elucidate the nature of these infiltrates, interleukin 2 (IL-2)-responsive T lymphocytes were grown out of synovial fragments from 14 patients undergoing surgery for advanced destructive inflammatory joint disease. Eleven of the samples examined were from patients with classical rheumatoid arthritis, while three others were obtained from individuals with clinical osteoarthritis. Southern blot analysis of T-cell receptor (TCR) β -chain genes in 13 of 14 cultures showed distinct rearrangements, indicating that each culture was characterized by the predominance of a limited number of clones. T-cell populations from peripheral blood stimulated with a variety of activators and expanded with IL-2 did not demonstrate evidence of similar clonality in long-term culture. These results suggest that a limited number of activated T-cell clones predominate at the site of tissue injury in rheumatoid synovial membranes as well as in other types of destructive inflammatory joint disease. Further characterization of these T-cell clones may aid our understanding of the pathogenesis of these rheumatic disorders.

Detection of clonal aberrations by cytogenetic analysis after different culture methods and by FISH in 129 patients with Chronic Lymphocytic Leukemia.

PubMed

Jenderny, Jutta; Goldmann, Claudia; Thede, Rebekka; Ebrecht, Monika; Korioth, Frank

2014-01-01

There are only a few cytogenetic analysis (CA) studies that directly compare the novel cultivation technique using immunostimulatory CpG-oligonucleotide DSP30/interleukin-2 (DSP30/IL2) with other culture methods. Therefore, parallel cultures of peripheral blood of 129 chronic lymphocytic leukemia (CLL) patients were set up in unstimulated cultures, in the presence of pokeweed medium (PWM), and with DSP30/IL2. Furthermore, CA results were compared with data obtained by FISH. Clonal aberrations were observed by CA in 6% of the cases in unstimulated cultures, in 27% of the cases with PWM, and in 40% of the cases with DSP30/IL2. Some clonal aberrations were detected by CA only with one culture method. Using 3 different culture methods, clonal aberrations were detected in 41% of the cases by CA and in 71% of the cases by FISH. Altogether, 78% of the cases exhibited clonal aberrations discovered by CA and FISH. Also, CA detected clonal aberrations not targeted by FISH in 7% of the cases, and FISH identified clonal aberrations not detected by CA in 36% of the cases. Our study demonstrates that the combined use of CA with different culture methods together with FISH increases our knowledge of the genetic complexity and heterogeneity in CLL pathogenesis. © 2014 S. Karger AG, Basel.

Evolution of Tumor Clones in Adult Acute Lymphoblastic Leukemia.

PubMed

Smirnova, S Yu; Sidorova, Yu V; Ryzhikova, N V; Sychevskaya, K A; Parovichnikova, E N; Sudarikov, A B

2016-01-01

Clonal instability of a tumor cell population in acute lymphoblastic leukemia (ALL) may complicate the monitoring of a minimal residual disease (MRD) by means of patient-specific targets identified at the disease onset. Most of the data concerning the possible instability of rearranged clonal TCR and IG genes during disease recurrence were obtained for ALL in children. The appropriate features of adult ALL, which are known to differ from those of childhood ALL in certain biological characteristics and prognosis, remain insufficiently studied. The aim of this study was to assess the stability of IG and TCR gene rearrangements in adult ALL. Rearrangements were identified according to the BIOMED-2 protocol (PCR followed by fragment analysis). Mismatch in clonal rearrangements at onset and relapse was identified in 83% of patients, indicating clonal instability during treatment. Clonal evolution and diversity of IG and TCR gene rearrangements may be one of the tumor progression mechanisms. New rearrangements may emerge due to residual VDJ-recombinase activity in tumor cells. Also, many clonal IG and TCR gene rearrangements may be present at different levels at a diagnosis, but less abundant clones may be "invisible" due to limited detection sensitivity. Later, major clones may disappear in the course of chemotherapy, while others may proliferate. Investigation of clonal evolution and heterogeneity in ALL and their impact on the treatment efficacy will contribute to the identification of new prognostic factors and the development of therapeutic approaches.

Plant Clonal Integration Mediates the Horizontal Redistribution of Soil Resources, Benefiting Neighboring Plants.

PubMed

Ye, Xue-Hua; Zhang, Ya-Lin; Liu, Zhi-Lan; Gao, Shu-Qin; Song, Yao-Bin; Liu, Feng-Hong; Dong, Ming

2016-01-01

Resources such as water taken up by plants can be released into soils through hydraulic redistribution and can also be translocated by clonal integration within a plant clonal network. We hypothesized that the resources from one (donor) microsite could be translocated within a clonal network, released into different (recipient) microsites and subsequently used by neighbor plants in the recipient microsite. To test these hypotheses, we conducted two experiments in which connected and disconnected ramet pairs of Potentilla anserina were grown under both homogeneous and heterogeneous water regimes, with seedlings of Artemisia ordosica as neighbors. The isotopes [(15)N] and deuterium were used to trace the translocation of nitrogen and water, respectively, within the clonal network. The water and nitrogen taken up by P. anserina ramets in the donor microsite were translocated into the connected ramets in the recipient microsites. Most notably, portions of the translocated water and nitrogen were released into the recipient microsite and were used by the neighboring A. ordosica, which increased growth of the neighboring A. ordosica significantly. Therefore, our hypotheses were supported, and plant clonal integration mediated the horizontal hydraulic redistribution of resources, thus benefiting neighboring plants. Such a plant clonal integration-mediated resource redistribution in horizontal space may have substantial effects on the interspecific relations and composition of the community and consequently on ecosystem processes.

High prevalence of EMRSA-15 in Portuguese public buses: a worrisome finding.

PubMed

Simões, Roméo Rocha; Aires-de-Sousa, Marta; Conceição, Teresa; Antunes, Filipa; da Costa, Paulo Martins; de Lencastre, Hermínia

2011-03-02

The nosocomial prevalence of methicillin resistant Staphylococcus aureus (MRSA) in Portugal remains one of the highest in Europe and is currently around 50%. Transmission of S. aureus, including MRSA, occurs principally by direct human-to-human skin contact. However, S. aureus can survive for long periods on inanimate objects, which may represent an important reservoir for dissemination as well. Between May 2009 and February 2010, handrails of 85 public urban buses circulating in Oporto, Portugal, were screened for the occurrence of MRSA. Twenty-two (26%) buses showed MRSA contamination. The molecular characterization of a total of 55 MRSA, by pulsed-field gel electrophoresis (PFGE), staphylococcal cassette chromosome (SCC) mec typing, spa typing, and multilocus sequence typing (MLST), clustered the isolates into three clonal types. However, the overwhelming majority (n = 50; 91%) of the isolates belonged to a single clone (PFGE A, spa types t747, t032, t025 or t020, ST22, SCCmec type IVh) that exhibits the characteristics of the pandemic EMRSA-15, currently the major lineage circulating in Portuguese hospitals, namely in the Oporto region. Two additional clones were found but in much lower numbers: (i) PFGE B, ST5, spa type t002, SCCmec IVa (n = 3), and (ii) PFGE C, spa type t008, ST8, SCCmec IVa (n = 2). None of the 55 isolates was PVL positive. Public buses in Oporto seem to be an important reservoir of MRSA of nosocomial origin, providing evidence that the major hospital-associated MRSA clone in Portugal is escaping from the primary ecological niche of hospitals to the community environment. Infection control measures are urgently warranted to limit the spread of EMRSA-15 to the general population and future studies are required to assess the eventual increase of MRSA in the Portuguese community, which so far remains low.

Characterisation of Phytophthora capsici isolates from black pepper in Vietnam.

PubMed

Truong, Nguyen V; Liew, Edward C Y; Burgess, Lester W

2010-01-01

Phytophthora foot rot of black pepper caused by Phytophthora capsici is a major disease of black pepper (Piper nigrum) throughout Vietnam. To understand the population structure of P. capsici, a large collection of P. capsici isolates from black pepper was studied on the basis of mating type, random amplified microsatellites (RAMS) and repetitive extragenic palindromic (REP) fingerprinting. Two mating types A1 and A2 were detected in four provinces in two climatic regions, with A1:A2 ratios ranging from 1:3 to 1:5. In several instances A1 and A2 mating types were found to co-exist in the same farm or black pepper pole, suggesting the potential for sexual reproduction of P. capsici in the field in Vietnam although its contribution to disease epidemics is uncertain. RAMS and REP DNA fingerprinting analysis of 118 isolates of P. capsici from black pepper showed that the population was genetically more diverse where two mating types were found, although the overall genetic diversity was low with most of the isolates belonging to one clonal group. The implication of these findings is discussed. The low diversity among isolates suggests that the P. capsici population may have originated from a single source. There was no genetic differentiation of isolates from different climatic regions. In addition to the large clonal group, several isolates with unique RAMS/REP phenotypes were also detected. Most of these unique phenotypes belonged to the minority A1 mating type. This may have significant implications for a gradual increase in overall genetic diversity.

Dynamics of acquisition and loss of carriage of Staphylococcus aureus strains in the community: The effect of clonal complex☆☆☆

PubMed Central

Miller, Ruth R.; Walker, A. Sarah; Godwin, Heather; Fung, Rowena; Votintseva, Antonina; Bowden, Rory; Mant, David; Peto, Timothy E.A.; Crook, Derrick W.; Knox, Kyle

2014-01-01

Summary Background Staphylococcus aureus nasal carriage increases infection risk. However, few studies have investigated S. aureus acquisition/loss over >1 year, and fewer still used molecular typing. Methods 1123 adults attending five Oxfordshire general practices had nasal swabs taken. 571 were re-swabbed after one month then every two months for median two years. All S. aureus isolates were spa-typed. Risk factors were collected from interviews and medical records. Results 32% carried S. aureus at recruitment (<1% MRSA). Rates of spa-type acquisition were similar in participants S. aureus positive (1.4%/month) and negative (1.8%/month, P = 0.13) at recruitment. Rates were faster in those carrying clonal complex (CC)15 (adjusted (a)P = 0.03) or CC8 (including USA300) (aP = 0.001) at recruitment versus other CCs. 157/274 (57%) participants S. aureus positive at recruitment returning ≥12 swabs carried S. aureus consistently, of whom 135 carried the same spa-type. CC22 (including EMRSA-15) was more prevalent in long-term than intermittent spa-type carriers (aP = 0.03). Antibiotics transiently reduced carriage, but no other modifiable risk factors were found. Conclusions Both transient and longer-term carriage exist; however, the approximately constant rates of S. aureus gain and loss suggest that ‘never’ or truly ‘persistent’ carriage are rare. Long-term carriage varies by strain, offering new explanations for the success of certain S. aureus clones. PMID:24393651

Genetic diversity of Streptococcus suis isolated from three pig farms of China obtained by acquiring antibiotic resistance genes.

PubMed

Huang, Jinhu; Shang, Kexin; Kashif, Jam; Wang, Liping

2015-05-01

Acquiring antibiotic resistance genes may change an organism's genetic characteristics and the effect of antibiotics, resulting in a rapid transmission of microbial pathogens. The objectives of this experiment were to identify the features of Streptococcus suis (S. suis) isolated from three pig farms in China which are geographically isolated. Among the isolates, 56.52% were sequence type 7 (ST7), followed by ST1 (26.09%), indicating that ST7 prevails in China, as revealed by multi-locus sequence typing (MLST). Statistical analysis indicated an association between geography, sequence types and antibiotic resistance genotypes. 66.67% of the isolates in Sichuan province presented a (ermB(-) + mefA(-) + tetO(-) + tetM(-)) + ST7 type. The tetM(+) +ST7 type was the most prevalent in Jiangsu province, whereas the strains from Hebei province had a phenotype ermB(+) +tetO(+) +ST1 (63.64%). Pulsed-field gel electrophoresis (PGFE) pattern A2 with 100% similarity reflected the clonal dissemination between Sichuan and Jiangsu provinces. Strains carrying or not carrying antibiotic resistance genes presented different PFGE patterns in Hebei province. ST7 is widespread in many regions of China and a clonal dissemination occurred between Sichuan and Jiangsu provinces in diseased pigs. However, ST1 strains with macrolide and tetracycline resistance (ermB(+) +tetO(+) +ST1) isolated from a farm in Hebei province demonstrated that the genetic diversity was contributed by horizontal acquiring of ermB and tetO carrying elements. © 2014 Society of Chemical Industry.

MLST-Based Population Genetic Analysis in a Global Context Reveals Clonality amongst Cryptococcus neoformans var. grubii VNI Isolates from HIV Patients in Southeastern Brazil

PubMed Central

Ferreira-Paim, Kennio; Andrade-Silva, Leonardo; Fonseca, Fernanda M.; Ferreira, Thatiana B.; Mora, Delio J.; Andrade-Silva, Juliana; Khan, Aziza; Dao, Aiken; Reis, Eduardo C.; Almeida, Margarete T. G.; Maltos, Andre; Junior, Virmondes R.; Trilles, Luciana; Rickerts, Volker; Chindamporn, Ariya; Sykes, Jane E.; Cogliati, Massimo; Nielsen, Kirsten; Boekhout, Teun; Fisher, Matthew; Kwon-Chung, June; Engelthaler, David M.; Lazéra, Marcia; Meyer, Wieland; Silva-Vergara, Mario L.

2017-01-01

Cryptococcosis is an important fungal infection in immunocompromised individuals, especially those infected with HIV. In Brazil, despite the free availability of antiretroviral therapy (ART) in the public health system, the mortality rate due to Cryptococcus neoformans meningitis is still high. To obtain a more detailed picture of the population genetic structure of this species in southeast Brazil, we studied 108 clinical isolates from 101 patients and 35 environmental isolates. Among the patients, 59% had a fatal outcome mainly in HIV-positive male patients. All the isolates were found to be C. neoformans var. grubii major molecular type VNI and mating type locus alpha. Twelve were identified as diploid by flow cytometry, being homozygous (AαAα) for the mating type and by PCR screening of the STE20, GPA1, and PAK1 genes. Using the ISHAM consensus multilocus sequence typing (MLST) scheme, 13 sequence types (ST) were identified, with one being newly described. ST93 was identified from 81 (75%) of the clinical isolates, while ST77 and ST93 were identified from 19 (54%) and 10 (29%) environmental isolates, respectively. The southeastern Brazilian isolates had an overwhelming clonal population structure. When compared with populations from different continents based on data extracted from the ISHAM-MLST database (mlst.mycologylab.org) they showed less genetic variability. Two main clusters within C. neoformans var. grubii VNI were identified that diverged from VNB around 0.58 to 4.8 million years ago. PMID:28099434

Standardizing the Nomenclature for Clonal Lineages of the Sudden Oak Death Pathogen, Phytophthora ramorum

USDA-ARS?s Scientific Manuscript database

Phytophthora ramorum, the causal agent of sudden oak death and ramorum blight, is known to exist as three distinct clonal lineages based on a range of molecular marker systems. However, in the recent literature there exists no consensus on naming of lineages. Here we name clonal lineages of P. ramor...

Revealing hidden clonal complexity in Mycobacterium tuberculosis infection by qualitative and quantitative improvement of sampling.

PubMed

Pérez-Lago, L; Palacios, J J; Herranz, M; Ruiz Serrano, M J; Bouza, E; García-de-Viedma, D

2015-02-01

The analysis of microevolution events, its functional relevance and impact on molecular epidemiology strategies, constitutes one of the most challenging aspects of the study of clonal complexity in infection by Mycobacterium tuberculosis. In this study, we retrospectively evaluated whether two improved sampling schemes could provide access to the clonal complexity that is undetected by the current standards (analysis of one isolate from one sputum). We evaluated in 48 patients the analysis by mycobacterial interspersed repetitive unit-variable number tandem repeat of M. tuberculosis isolates cultured from bronchial aspirate (BAS) or bronchoalveolar lavage (BAL) and, in another 16 cases, the analysis of a higher number of isolates from independent sputum samples. Analysis of the isolates from BAS/BAL specimens revealed clonal complexity in a very high proportion of cases (5/48); in most of these cases, complexity was not detected when the isolates from sputum samples were analysed. Systematic analysis of isolates from multiple sputum samples also improved the detection of clonal complexity. We found coexisting clonal variants in two of 16 cases that would have gone undetected in the analysis of the isolate from a single sputum specimen. Our results suggest that analysis of isolates from BAS/BAL specimens is highly efficient for recording the true clonal composition of M. tuberculosis in the lungs. When these samples are not available, we recommend increasing the number of isolates from independent sputum specimens, because they might not harbour the same pool of bacteria. Our data suggest that the degree of clonal complexity in tuberculosis has been underestimated because of the deficiencies inherent in a simplified procedure. Copyright © 2014 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

VDJ-Seq: Deep Sequencing Analysis of Rearranged Immunoglobulin Heavy Chain Gene to Reveal Clonal Evolution Patterns of B Cell Lymphoma.

PubMed

Jiang, Yanwen; Nie, Kui; Redmond, David; Melnick, Ari M; Tam, Wayne; Elemento, Olivier

2015-12-28

Understanding tumor clonality is critical to understanding the mechanisms involved in tumorigenesis and disease progression. In addition, understanding the clonal composition changes that occur within a tumor in response to certain micro-environment or treatments may lead to the design of more sophisticated and effective approaches to eradicate tumor cells. However, tracking tumor clonal sub-populations has been challenging due to the lack of distinguishable markers. To address this problem, a VDJ-seq protocol was created to trace the clonal evolution patterns of diffuse large B cell lymphoma (DLBCL) relapse by exploiting VDJ recombination and somatic hypermutation (SHM), two unique features of B cell lymphomas. In this protocol, Next-Generation sequencing (NGS) libraries with indexing potential were constructed from amplified rearranged immunoglobulin heavy chain (IgH) VDJ region from pairs of primary diagnosis and relapse DLBCL samples. On average more than half million VDJ sequences per sample were obtained after sequencing, which contain both VDJ rearrangement and SHM information. In addition, customized bioinformatics pipelines were developed to fully utilize sequence information for the characterization of IgH-VDJ repertoire within these samples. Furthermore, the pipeline allows the reconstruction and comparison of the clonal architecture of individual tumors, which enables the examination of the clonal heterogeneity within the diagnosis tumors and deduction of clonal evolution patterns between diagnosis and relapse tumor pairs. When applying this analysis to several diagnosis-relapse pairs, we uncovered key evidence that multiple distinctive tumor evolutionary patterns could lead to DLBCL relapse. Additionally, this approach can be expanded into other clinical aspects, such as identification of minimal residual disease, monitoring relapse progress and treatment response, and investigation of immune repertoires in non-lymphoma contexts.

Validation of the REMA score for predicting mast cell clonality and systemic mastocytosis in patients with systemic mast cell activation symptoms.

PubMed

Alvarez-Twose, I; González-de-Olano, D; Sánchez-Muñoz, L; Matito, A; Jara-Acevedo, M; Teodosio, C; García-Montero, A; Morgado, J M; Orfao, A; Escribano, L

2012-01-01

A variable percentage of patients with systemic mast cell (MC) activation symptoms meet criteria for systemic mastocytosis (SM). We prospectively evaluated the clinical utility of the REMA score versus serum baseline tryptase (sBt) levels for predicting MC clonality and SM in 158 patients with systemic MC activation symptoms in the absence of mastocytosis in the skin (MIS). World Health Organization criteria for SM were applied in all cases. MC clonality was defined as the presence of KIT-mutated MC or by a clonal HUMARA test. The REMA score consisted of the assignment of positive or negative points as follows: male (+1), female (-1), sBt <15 μg/l (-1) or >25 μg/l (+2), presence (-2) or absence (+1) of pruritus, hives or angioedema and presence (+3) of presyncope or syncope. Efficiency of the REMA score for predicting MC clonality and SM was assessed by receiver operating characteristic (ROC) curve analyses and compared to those obtained by means of sBt levels alone. Molecular studies revealed the presence of clonal MC in 68/80 SM cases and in 11/78 patients who did not meet the criteria for SM. ROC curve analyses confirmed the greater sensitivity and a similar specificity of the REMA score versus sBt levels (84 vs. 59% and 74 vs. 70% for MC clonality and 87 vs. 62% and 73 vs. 71% for SM, respectively). Our results confirm the clinical utility of the REMA score to predict MC clonality and SM in patients suffering from systemic MC activation symptoms without MIS. Copyright © 2011 S. Karger AG, Basel.

Reconstruction of cartilage with clonal mesenchymal stem cell-acellular dermal matrix in cartilage defect model in nonhuman primates.

PubMed

Ma, Anlun; Jiang, Li; Song, Lijun; Hu, Yanxin; Dun, Hao; Daloze, Pierre; Yu, Yonglin; Jiang, Jianyuan; Zafarullah, Muhammad; Chen, Huifang

2013-07-01

Articular cartilage defects are commonly associated with trauma, inflammation and osteoarthritis. Mesenchymal stem cell (MSC)-based therapy is a promising novel approach for repairing articular cartilage. Direct intra-articular injection of uncommitted MSCs does not regenerate high-quality cartilage. This study explored utilization of a new three-dimensional, selected chondrogenic clonal MSC-loaded monkey acellular dermal matrix (MSC-ADM) scaffold to repair damaged cartilage in an experimental model of knee joint cartilage defect in Cynomolgus monkeys. MSCs were characterized for cell size, cell yield, phenotypes, proliferation and chondrogenic differentiation capacity. Chondrogenic differentiation assays were performed at different MSC passages by sulfated glycosaminoglycans (sGAG), collagen, and fluorescence activated cell sorter (FACS) analysis. Selected chondrogenic clonal MSCs were seeded onto ADM scaffold with the sandwich model and MSC-loaded ADM grafts were analyzed by confocal microscopy and scanning electron microscopy. Cartilage defects were treated with normal saline, clonal MSCs and clonal MSC-ADM grafts, respectively. The clinical parameters, and histological and immunohistochemical examinations were evaluated at weeks 8, 16, 24 post-treatment, respectively. Polyclonal and clonal MSCs could differentiate into the chondrogenic lineage after stimulation with suitable chondrogenic factors. They expressed mesenchymal markers and were negative for hematopoietic markers. Articular cartilage defects were considerably improved and repaired by selected chondrogenic clonal MSC-based treatment, particularly, in MSC-ADM-treated group. The histological scores in MSC-ADM-treated group were consistently higher than those of other groups. Our results suggest that selected chondrogenic clonal MSC-loaded ADM grafts could improve the cartilage lesions in Cynomolgus monkey model, which may be applicable for repairing similar human cartilage defects. Copyright © 2013 Elsevier B.V. All rights reserved.

T-cell stimuli independently sum to regulate an inherited clonal division fate

PubMed Central

Marchingo, J. M.; Prevedello, G.; Kan, A.; Heinzel, S.; Hodgkin, P. D.; Duffy, K. R.

2016-01-01

In the presence of antigen and costimulation, T cells undergo a characteristic response of expansion, cessation and contraction. Previous studies have revealed that population-level reproducibility is a consequence of multiple clones exhibiting considerable disparity in burst size, highlighting the requirement for single-cell information in understanding T-cell fate regulation. Here we show that individual T-cell clones resulting from controlled stimulation in vitro are strongly lineage imprinted with highly correlated expansion fates. Progeny from clonal families cease dividing in the same or adjacent generations, with inter-clonal variation producing burst-size diversity. The effects of costimulatory signals on individual clones sum together with stochastic independence; therefore, the net effect across multiple clones produces consistent, but heterogeneous population responses. These data demonstrate that substantial clonal heterogeneity arises through differences in experience of clonal progenitors, either through stochastic antigen interaction or by differences in initial receptor sensitivities. PMID:27869196

Multifocal clonal evolution characterized using circulating tumour DNA in a case of metastatic breast cancer

PubMed Central

Murtaza, Muhammed; Dawson, Sarah-Jane; Pogrebniak, Katherine; Rueda, Oscar M.; Provenzano, Elena; Grant, John; Chin, Suet-Feung; Tsui, Dana W. Y.; Marass, Francesco; Gale, Davina; Ali, H. Raza; Shah, Pankti; Contente-Cuomo, Tania; Farahani, Hossein; Shumansky, Karey; Kingsbury, Zoya; Humphray, Sean; Bentley, David; Shah, Sohrab P.; Wallis, Matthew; Rosenfeld, Nitzan; Caldas, Carlos

2015-01-01

Circulating tumour DNA analysis can be used to track tumour burden and analyse cancer genomes non-invasively but the extent to which it represents metastatic heterogeneity is unknown. Here we follow a patient with metastatic ER-positive and HER2-positive breast cancer receiving two lines of targeted therapy over 3 years. We characterize genomic architecture and infer clonal evolution in eight tumour biopsies and nine plasma samples collected over 1,193 days of clinical follow-up using exome and targeted amplicon sequencing. Mutation levels in the plasma samples reflect the clonal hierarchy inferred from sequencing of tumour biopsies. Serial changes in circulating levels of sub-clonal private mutations correlate with different treatment responses between metastatic sites. This comparison of biopsy and plasma samples in a single patient with metastatic breast cancer shows that circulating tumour DNA can allow real-time sampling of multifocal clonal evolution. PMID:26530965

An atlas of B-cell clonal distribution in the human body.

PubMed

Meng, Wenzhao; Zhang, Bochao; Schwartz, Gregory W; Rosenfeld, Aaron M; Ren, Daqiu; Thome, Joseph J C; Carpenter, Dustin J; Matsuoka, Nobuhide; Lerner, Harvey; Friedman, Amy L; Granot, Tomer; Farber, Donna L; Shlomchik, Mark J; Hershberg, Uri; Luning Prak, Eline T

2017-09-01

B-cell responses result in clonal expansion, and can occur in a variety of tissues. To define how B-cell clones are distributed in the body, we sequenced 933,427 B-cell clonal lineages and mapped them to eight different anatomic compartments in six human organ donors. We show that large B-cell clones partition into two broad networks-one spans the blood, bone marrow, spleen and lung, while the other is restricted to tissues within the gastrointestinal (GI) tract (jejunum, ileum and colon). Notably, GI tract clones display extensive sharing of sequence variants among different portions of the tract and have higher frequencies of somatic hypermutation, suggesting extensive and serial rounds of clonal expansion and selection. Our findings provide an anatomic atlas of B-cell clonal lineages, their properties and tissue connections. This resource serves as a foundation for studies of tissue-based immunity, including vaccine responses, infections, autoimmunity and cancer.

Multifocal clonal evolution characterized using circulating tumour DNA in a case of metastatic breast cancer.

PubMed

Murtaza, Muhammed; Dawson, Sarah-Jane; Pogrebniak, Katherine; Rueda, Oscar M; Provenzano, Elena; Grant, John; Chin, Suet-Feung; Tsui, Dana W Y; Marass, Francesco; Gale, Davina; Ali, H Raza; Shah, Pankti; Contente-Cuomo, Tania; Farahani, Hossein; Shumansky, Karey; Kingsbury, Zoya; Humphray, Sean; Bentley, David; Shah, Sohrab P; Wallis, Matthew; Rosenfeld, Nitzan; Caldas, Carlos

2015-11-04

Circulating tumour DNA analysis can be used to track tumour burden and analyse cancer genomes non-invasively but the extent to which it represents metastatic heterogeneity is unknown. Here we follow a patient with metastatic ER-positive and HER2-positive breast cancer receiving two lines of targeted therapy over 3 years. We characterize genomic architecture and infer clonal evolution in eight tumour biopsies and nine plasma samples collected over 1,193 days of clinical follow-up using exome and targeted amplicon sequencing. Mutation levels in the plasma samples reflect the clonal hierarchy inferred from sequencing of tumour biopsies. Serial changes in circulating levels of sub-clonal private mutations correlate with different treatment responses between metastatic sites. This comparison of biopsy and plasma samples in a single patient with metastatic breast cancer shows that circulating tumour DNA can allow real-time sampling of multifocal clonal evolution.

Epigenetic Memory as a Basis for Intelligent Behavior in Clonal Plants.

PubMed

Latzel, Vít; Rendina González, Alejandra P; Rosenthal, Jonathan

2016-01-01

Environmentally induced epigenetic change enables plants to remember past environmental interactions. If this memory capability is exploited to prepare plants for future challenges, it can provide a basis for highly sophisticated behavior, considered intelligent by some. Against the backdrop of an overview of plant intelligence, we hypothesize: (1) that the capability of plants to engage in such intelligent behavior increases with the additional level of complexity afforded by clonality, and; (2) that more faithful inheritance of epigenetic information in clonal plants, in conjunction with information exchange and coordination between connected ramets, is likely to enable especially advanced intelligent behavior in this group. We therefore further hypothesize that this behavior provides ecological and evolutionary advantages to clonal plants, possibly explaining, at least in part, their widespread success. Finally, we suggest avenues of inquiry to enable assessing intelligent behavior and the role of epigenetic memory in clonal species.