Molecular phylogenetic analysis of non-sexually transmitted strains of Haemophilus ducreyi.

PubMed

Gaston, Jordan R; Roberts, Sally A; Humphreys, Tricia L

2015-01-01

Haemophilus ducreyi, the etiologic agent of chancroid, has been previously reported to show genetic variance in several key virulence factors, placing strains of the bacterium into two genetically distinct classes. Recent studies done in yaws-endemic areas of the South Pacific have shown that H. ducreyi is also a major cause of cutaneous limb ulcers (CLU) that are not sexually transmitted. To genetically assess CLU strains relative to the previously described class I, class II phylogenetic hierarchy, we examined nucleotide sequence diversity at 11 H. ducreyi loci, including virulence and housekeeping genes, which encompass approximately 1% of the H. ducreyi genome. Sequences for all 11 loci indicated that strains collected from leg ulcers exhibit DNA sequences homologous to class I strains of H. ducreyi. However, sequences for 3 loci, including a hemoglobin receptor (hgbA), serum resistance protein (dsrA), and a collagen adhesin (ncaA) contained informative amounts of variation. Phylogenetic analyses suggest that these non-sexually transmitted strains of H. ducreyi comprise a sub-clonal population within class I strains of H. ducreyi. Molecular dating suggests that CLU strains are the most recently developed, having diverged approximately 0.355 million years ago, fourteen times more recently than the class I/class II divergence. The CLU strains' divergence falls after the divergence of humans from chimpanzees, making it the first known H. ducreyi divergence event directly influenced by the selective pressures accompanying human hosts.

Draft Genome Sequence of Lactobacillus delbrueckii Strain #22 Isolated from a Patient with Short Bowel Syndrome and Previous d-Lactic Acidosis and Encephalopathy.

PubMed

Domann, Eugen; Fischer, Florence; Glowatzki, Fabian; Fritzenwanker, Moritz; Hain, Torsten; Zechel-Gran, Silke; Giffhorn-Katz, Susanne; Neubauer, Bernd A

2016-07-28

d-Lactic acidosis with associated encephalopathy caused by overgrowth of intestinal lactic acid bacteria is a rarely diagnosed neurological complication of patients with short bowel syndrome. Here, we report the draft genome sequence of Lactobacillus delbrueckii strain #22 isolated from a patient with short bowel syndrome and previous d-lactic acidosis/encephalopathy. Copyright © 2016 Domann et al.

Phylogeny of Neoparamoeba strains isolated from marine fish and invertebrates as inferred from SSU rDNA sequences.

PubMed

Dyková, Iva; Nowak, Barbara; Pecková, Hana; Fiala, Ivan; Crosbie, Philip; Dvoráková, Helena

2007-02-08

We characterised 9 strains selected from primary isolates referable to Paramoeba/Neoparamoeba spp. Based on ultrastructural study, 5 strains isolated from fish (amoebic gill disease [AGD]-affected Atlantic salmon and dead southern bluefin tuna), 1 strain from netting of a floating sea cage and 3 strains isolated from invertebrates (sea urchins and crab) were assigned to the genus Neoparamoeba Page, 1987. Phylogenetic analyses based on SSU rDNA sequences revealed affiliations of newly introduced and previously analysed Neoparamoeba strains. Three strains from the invertebrates and 2 out of 3 strains from gills of southern bluefin tunas were members of the N. branchiphila clade, while the remaining, fish-isolated strains, as well as the fish cage strain, clustered within the clade of N. pemaquidensis. These findings and previous reports point to the possibility that N. pemaquidensis and N. branchiphila can affect both fish and invertebrates. A new potential fish host, southern bluefin tuna, was included in the list of farmed fish endangered by N. branchiphila. The sequence of P. eilhardi (Culture Collection of Algae and Protozoa [CCAP] strain 1560/2) appeared in all analyses among sequences of strain representatives of Neoparamoeba species, in a position well supported by bootstrap value, Bremer index and Bayesian posterior probability. Our research shows that isolation of additional strains from invertebrates and further analyses of relations between molecular data and morphological characters of the genera Paramoeba and Neoparamoeba are required. This complexity needs to be considered when attempting to define molecular markers for identification of Paramoeba/Neoparamoeba species in tissues of fish and invertebrates.

Sequencing of emerging canine distemper virus strain reveals new distinct genetic lineage in the United States associated with disease in wildlife and domestic canine populations.

PubMed

Riley, Matthew C; Wilkes, Rebecca P

2015-12-18

Recent outbreaks of canine distemper have prompted examination of strains from clinical samples submitted to the University of Tennessee College of Veterinary Medicine (UTCVM) Clinical Virology Lab. We previously described a new strain of CDV that significantly diverged from all genotypes reported to date including America 2, the genotype proposed to be the main lineage currently circulating in the US. The aim of this study was to determine when this new strain appeared and how widespread it is in animal populations, given that it has also been detected in fully vaccinated adult dogs. Additionally, we sequenced complete viral genomes to characterize the strain and determine if variation is confined to known variable regions of the genome or if the changes are also present in more conserved regions. Archived clinical samples were genotyped using real-time RT-PCR amplification and sequencing. The genomes of two unrelated viruses from a dog and fox each from a different state were sequenced and aligned with previously published genomes. Phylogenetic analysis was performed using coding, non-coding and genome-length sequences. Virus neutralization assays were used to evaluate potential antigenic differences between this strain and a vaccine strain and mixed ANOVA test was used to compare the titers. Genotyping revealed this strain first appeared in 2011 and was detected in dogs from multiple states in the Southeast region of the United States. It was the main strain detected among the clinical samples that were typed from 2011-2013, including wildlife submissions. Genome sequencing demonstrated that it is highly conserved within a new lineage and preliminary serologic testing showed significant differences in neutralizing antibody titers between this strain and the strain commonly used in vaccines. This new strain represents an emerging CDV in domestic dogs in the US, may be associated with a stable reservoir in the wildlife population, and could facilitate vaccine escape.

First Report of a Fatal Case Associated with EV-D68 Infection in Hong Kong and Emergence of an Interclade Recombinant in China Revealed by Genome Analysis.

PubMed

Yip, Cyril C Y; Lo, Janice Y C; Sridhar, Siddharth; Lung, David C; Luk, Shik; Chan, Kwok-Hung; Chan, Jasper F W; Cheng, Vincent C C; Woo, Patrick C Y; Yuen, Kwok-Yung; Lau, Susanna K P

2017-05-16

A fatal case associated with enterovirus D68 (EV-D68) infection affecting a 10-year-old boy was reported in Hong Kong in 2014. To examine if a new strain has emerged in Hong Kong, we sequenced the partial genome of the EV-D68 strain identified from the fatal case and the complete VP1, and partial 5'UTR and 2C sequences of nine additional EV-D68 strains isolated from patients in Hong Kong. Sequence analysis indicated that a cluster of strains including the previously recognized A2 strains should belong to a separate clade, clade D, which is further divided into subclades D1 and D2. Among the 10 EV-D68 strains, 7 (including the fatal case) belonged to the previously described, newly emerged subclade B3, 2 belonged to subclade B1, and 1 belonged to subclade D1. Three EV-D68 strains, each from subclades B1, B3, and D1, were selected for complete genome sequencing and recombination analysis. While no evidence of recombination was noted among local strains, interclade recombination was identified in subclade D2 strains detected in mainland China in 2008 with VP2 acquired from clade A. This study supports the reclassification of subclade A2 into clade D1, and demonstrates interclade recombination between clades A and D2 in EV-D68 strains from China.

The Awesome Power of Yeast Evolutionary Genetics: New Genome Sequences and Strain Resources for the Saccharomyces sensu stricto Genus

PubMed Central

Scannell, Devin R.; Zill, Oliver A.; Rokas, Antonis; Payen, Celia; Dunham, Maitreya J.; Eisen, Michael B.; Rine, Jasper; Johnston, Mark; Hittinger, Chris Todd

2011-01-01

High-quality, well-annotated genome sequences and standardized laboratory strains fuel experimental and evolutionary research. We present improved genome sequences of three species of Saccharomyces sensu stricto yeasts: S. bayanus var. uvarum (CBS 7001), S. kudriavzevii (IFO 1802T and ZP 591), and S. mikatae (IFO 1815T), and describe their comparison to the genomes of S. cerevisiae and S. paradoxus. The new sequences, derived by assembling millions of short DNA sequence reads together with previously published Sanger shotgun reads, have vastly greater long-range continuity and far fewer gaps than the previously available genome sequences. New gene predictions defined a set of 5261 protein-coding orthologs across the five most commonly studied Saccharomyces yeasts, enabling a re-examination of the tempo and mode of yeast gene evolution and improved inferences of species-specific gains and losses. To facilitate experimental investigations, we generated genetically marked, stable haploid strains for all three of these Saccharomyces species. These nearly complete genome sequences and the collection of genetically marked strains provide a valuable toolset for comparative studies of gene function, metabolism, and evolution, and render Saccharomyces sensu stricto the most experimentally tractable model genus. These resources are freely available and accessible through www.SaccharomycesSensuStricto.org. PMID:22384314

Complete genome sequence of a Klebsiella pneumoniae strain isolated from a known cotton insect boll vector

USDA-ARS?s Scientific Manuscript database

Klebsiella pneumoniae (associated with bacterial pneumonia) was previously isolated from Nezara viridula, a significant vector of cotton boll-rot pathogens. We provide the first annotated genome sequence of the cotton opportunistic strain K. pneumoniae 5-1. This data provides guidance to study the...

Outbreak of poliomyelitis in Finland in 1984-85 - Re-analysis of viral sequences using the current standard approach.

PubMed

Simonen, Marja-Leena; Roivainen, Merja; Iber, Jane; Burns, Cara; Hovi, Tapani

2010-01-01

In 1984, a wild type 3 poliovirus (PV3/FIN84) spread all over Finland causing nine cases of paralytic poliomyelitis and one case of aseptic meningitis. The outbreak was ended in 1985 with an intensive vaccination campaign. By limited sequence comparison with previously isolated PV3 strains, closest relatives of PV3/FIN84 were found among strains circulating in the Mediterranean region. Now we wanted to reanalyse the relationships using approaches currently exploited in poliovirus surveillance. Cell lysates of 22 strains isolated during the outbreak and stored frozen were subjected to RT-PCR amplification in three genomic regions without prior subculture. Sequences of the entire VP1 coding region, 150 nucleotides in the VP1-2A junction, most of the 5' non-coding region, partial sequences of the 3D RNA polymerase coding region and partial 3' non-coding region were compared within the outbreak and with sequences available in data banks. In addition, complete nucleotide sequences were obtained for 2 strains isolated from two different cases of disease during the outbreak. The results confirmed the previously described wide intraepidemic variation of the strains, including amino acid substitutions in antigenic sites, as well as the likely Mediterranean region origin of the strains. Simplot and bootscanning analyses of the complete genomes indicated complicated evolutionary history of the non-capsid coding regions of the genome suggesting several recombinations with different HEV-C viruses in the past.

[Molecular-genetic characterization of shiga-toxin producing Escherichia coli isolated during a food-borne outbreak in St. Petersburg in 2013].

PubMed

Onishchenko, G G; Dyatlov, I A; Svetoch, E A; Volozhantsev, N V; Bannov, V A; Kartsev, N N; Borzenkov, V N; Fursova, N K; Shemyakin, I G; Bogun, A G; Kislichkina, A A; Popova, A V; Myakinina, V P; Teimurazov, M G; Polosenko, O V; Kaftyreva, L A; Makarova, M A; Matveeva, Z N; Grechaninova, T A; Grigor'eva, N S; Kicha, E V; Zabalueva, G V; Kutasova, T B; Korzhaev, Yu N; Bashketova, N S; Bushmanova, O N; Stalevskaya, A V; Tchinjeria, I G; Zhebrun, F B

2015-01-01

Shiga toxin-producing Escherichia coli (STEC) food-borne infections are reported worldwide and represent a serious problem for public healthcare. In the Russian Federation there is little information on epidemiology and etiology of STEC-infections as well as on molecular-genetic peculiarities of STEC pathogens. Our aim was to describe a food-borne outbreak as hemorrhagic colitis (HC) along with hemolytic uremic syndrome (HUS), enterocolitis, and acute gastroenteritis in children in St. Petersburg in 2013. Epidemiological, microbiological, molecular-genetic and bioinformatic methods were applied. Objects to study were clinical specimens, milk and food samples, as well as STEC strains isolated during the outbreak. The outbreak of food-borne infection was found to be caused by STEC-contaminated raw milk as confirmed by epidemiological analysis, detection of STEC DNA and isolation of relevant pathogens in milk and sick children fecal specimens. The whole-genome sequencing revealed two groups ofpathogens, E. coli O157:H7 and E. coli O101:H33 among collected strains. Group I strains were attributed to the previously known sequence type ST24, while group II strains belonged to the previously non-described sequence type ST145. In strain genomes of both groups there were identified nucleotide sequences of VT2-like prophage carrying stx2c gene, plasmid enterohemolysin gene, and gene of the STEC main adhesion factor intimin. Gene of intimin gamma was identified in E. coli O157:H7 strains and intimin iota 2 in E. coli O101:H33 strains. The latter previously was identified only in enteropathogenic E. coli (EPEC) strains. The additional knowledge of epidemiology and biology of STEC pathogens would assist clinicians and epidemiologists in diagnosing, treating and preventing hemorrhagic colitis.

'Candidatus Phytoplasma solani', a novel taxon associated with stolbur- and bois noir-related diseases of plants.

PubMed

Quaglino, Fabio; Zhao, Yan; Casati, Paola; Bulgari, Daniela; Bianco, Piero Attilio; Wei, Wei; Davis, Robert Edward

2013-08-01

Phytoplasmas classified in group 16SrXII infect a wide range of plants and are transmitted by polyphagous planthoppers of the family Cixiidae. Based on 16S rRNA gene sequence identity and biological properties, group 16SrXII encompasses several species, including 'Candidatus Phytoplasma australiense', 'Candidatus Phytoplasma japonicum' and 'Candidatus Phytoplasma fragariae'. Other group 16SrXII phytoplasma strains are associated with stolbur disease in wild and cultivated herbaceous and woody plants and with bois noir disease in grapevines (Vitis vinifera L.). Such latter strains have been informally proposed to represent a separate species, 'Candidatus Phytoplasma solani', but a formal description of this taxon has not previously been published. In the present work, stolbur disease strain STOL11 (STOL) was distinguished from reference strains of previously described species of the 'Candidatus Phytoplasma' genus based on 16S rRNA gene sequence similarity and a unique signature sequence in the 16S rRNA gene. Other stolbur- and bois noir-associated ('Ca. Phytoplasma solani') strains shared >99 % 16S rRNA gene sequence similarity with strain STOL11 and contained the signature sequence. 'Ca. Phytoplasma solani' is the only phytoplasma known to be transmitted by Hyalesthes obsoletus. Insect vectorship and molecular characteristics are consistent with the concept that diverse 'Ca. Phytoplasma solani' strains share common properties and represent an ecologically distinct gene pool. Phylogenetic analyses of 16S rRNA, tuf, secY and rplV-rpsC gene sequences supported this view and yielded congruent trees in which 'Ca. Phytoplasma solani' strains formed, within the group 16SrXII clade, a monophyletic subclade that was most closely related to, but distinct from, that of 'Ca. Phytoplasma australiense'-related strains. Based on distinct molecular and biological properties, stolbur- and bois noir-associated strains are proposed to represent a novel species level taxon, 'Ca. Phytoplasma solani'; STOL11 is designated the reference strain.

Genome Sequences of Multidrug-Resistant Salmonella enterica subsp. enterica Serovar Infantis Strains from Broiler Chicks in Hungary

PubMed Central

Wilk, Tímea; Szabó, Móni; Szmolka, Ama; Kiss, János; Barta, Endre; Nagy, Tibor

2016-01-01

Three strains of Salmonella enterica serovar Infantis isolated from healthy broiler chickens from 2012 to 2013 have been sequenced. Comparison of these and previously published S. Infantis genome sequences of broiler origin in 1996 and 2004 will provide new insight into the genome evolution and recent spread of S. Infantis in poultry. PMID:27979950

Diversity of 16S rRNA genes of new Ehrlichia strains isolated from horses with clinical signs of Potomac horse fever.

PubMed

Wen, B; Rikihisa, Y; Fuerst, P A; Chaichanasiriwithaya, W

1995-04-01

Ehrlichia risticii is the causative agent of Potomac horse fever. Variations among the major antigens of different local E. risticii strains have been detected previously. To further assess genetic variability in this species or species complex, the sequences of the 16S rRNA genes of several isolates obtained from sick horses diagnosed as having Potomac horse fever were determined. The sequences of six isolates obtained from Ohio and three isolates obtained from Kentucky were amplified by PCR. Three groups of sequences were identified. The sequences of five of the Ohio isolates were identical to the sequence of the type strain of E. risticii, the Illinois strain. The sequence of one Ohio isolate, isolate 081, was unique; this sequence differed in 10 nucleotides from the sequence of the type strain (level of similarity, 99.3%). The sequences of the three Kentucky isolates were identical to each other, but differed by five bases from the sequence of the type strain (level of similarity, 99.6%). The levels of sequence similarity of isolate 081, the Kentucky isolates, and the type strain to the next most closely related Ehrlichia sp., Ehrlichia sennetsu, were 99.3, 99.2, and 99.2%, respectively. On the basis of the distinct antigenic profiles and the levels of 16S rRNA sequence divergence, isolate 081 is as divergent from the type strain of E. risticii as E. sennetsu is. Therefore, we suggest that strain 081 and the Kentucky isolates may represent two new distinct Ehrlichia species.

Burkholderia sp. induces functional nodules on the South African invasive legume Dipogon lignosus (Phaseoleae) in New Zealand soils.

PubMed

Liu, Wendy Y Y; Ridgway, Hayley J; James, Trevor K; James, Euan K; Chen, Wen-Ming; Sprent, Janet I; Young, J Peter W; Andrews, Mitchell

2014-10-01

The South African invasive legume Dipogon lignosus (Phaseoleae) produces nodules with both determinate and indeterminate characteristics in New Zealand (NZ) soils. Ten bacterial isolates produced functional nodules on D. lignosus. The 16S ribosomal RNA (rRNA) gene sequences identified one isolate as Bradyrhizobium sp., one isolate as Rhizobium sp. and eight isolates as Burkholderia sp. The Bradyrhizobium sp. and Rhizobium sp. 16S rRNA sequences were identical to those of strains previously isolated from crop plants and may have originated from inocula used on crops. Both 16S rRNA and DNA recombinase A (recA) gene sequences placed the eight Burkholderia isolates separate from previously described Burkholderia rhizobial species. However, the isolates showed a very close relationship to Burkholderia rhizobial strains isolated from South African plants with respect to their nitrogenase iron protein (nifH), N-acyltransferase nodulation protein A (nodA) and N-acetylglucosaminyl transferase nodulation protein C (nodC) gene sequences. Gene sequences and enterobacterial repetitive intergenic consensus (ERIC) PCR and repetitive element palindromic PCR (rep-PCR) banding patterns indicated that the eight Burkholderia isolates separated into five clones of one strain and three of another. One strain was tested and shown to produce functional nodules on a range of South African plants previously reported to be nodulated by Burkholderia tuberum STM678(T) which was isolated from the Cape Region. Thus, evidence is strong that the Burkholderia strains isolated here originated in South Africa and were somehow transported with the plants from their native habitat to NZ. It is possible that the strains are of a new species capable of nodulating legumes.

Complete genome sequence of Clostridium pasteurianum NRRL B-598, a non-type strain producing butanol.

PubMed

Sedlar, Karel; Kolek, Jan; Skutkova, Helena; Branska, Barbora; Provaznik, Ivo; Patakova, Petra

2015-11-20

The strain Clostridium pasteurianum NRRL B-598 is non-type, oxygen tolerant, spore-forming, mesophilic and heterofermentative strain with high hydrogen production and ability of acetone-butanol fermentation (ethanol production being negligible). Here, we present the annotated complete genome sequence of this bacterium, replacing the previous draft genome assembly. The genome consisting of a single circular 6,186,879 bp chromosome with no plasmid was determined using PacBio RSII and Roche 454 sequencing. Copyright © 2015 Elsevier B.V. All rights reserved.

Complete genome sequence of duck Tembusu virus, isolated from Muscovy ducks in southern China.

PubMed

Zhu, Wanjun; Chen, Jidang; Wei, Chunya; Wang, Heng; Huang, Zhen; Zhang, Minze; Tang, Fengfeng; Xie, Jiexiong; Liang, Huanbin; Zhang, Guihong; Su, Shuo

2012-12-01

We report here the complete genomic sequence of the duck Tembusu virus (DTMUV) WJ-1 strain, isolated from Muscovy ducks. This is the first complete genome sequence of DTMUV reported in southern China. Compared with the other strains (TA, GH-2, YY5, and ZJ-407) that were previously found in eastern China, WJ-1 bears a few differences in the nucleotide and amino acid sequences. We found that there are 47 mutations of amino acids encoded by the whole open reading frame (ORF) among these five strains. The whole-genome sequence of DTMUV will help in understanding the epidemiology and molecular characteristics of duck Tembusu virus in southern China.

Detection of Mixed Infection from Bacterial Whole Genome Sequence Data Allows Assessment of Its Role in Clostridium difficile Transmission

PubMed Central

Eyre, David W.; Cule, Madeleine L.; Griffiths, David; Crook, Derrick W.; Peto, Tim E. A.

2013-01-01

Bacterial whole genome sequencing offers the prospect of rapid and high precision investigation of infectious disease outbreaks. Close genetic relationships between microorganisms isolated from different infected cases suggest transmission is a strong possibility, whereas transmission between cases with genetically distinct bacterial isolates can be excluded. However, undetected mixed infections—infection with ≥2 unrelated strains of the same species where only one is sequenced—potentially impairs exclusion of transmission with certainty, and may therefore limit the utility of this technique. We investigated the problem by developing a computationally efficient method for detecting mixed infection without the need for resource-intensive independent sequencing of multiple bacterial colonies. Given the relatively low density of single nucleotide polymorphisms within bacterial sequence data, direct reconstruction of mixed infection haplotypes from current short-read sequence data is not consistently possible. We therefore use a two-step maximum likelihood-based approach, assuming each sample contains up to two infecting strains. We jointly estimate the proportion of the infection arising from the dominant and minor strains, and the sequence divergence between these strains. In cases where mixed infection is confirmed, the dominant and minor haplotypes are then matched to a database of previously sequenced local isolates. We demonstrate the performance of our algorithm with in silico and in vitro mixed infection experiments, and apply it to transmission of an important healthcare-associated pathogen, Clostridium difficile. Using hospital ward movement data in a previously described stochastic transmission model, 15 pairs of cases enriched for likely transmission events associated with mixed infection were selected. Our method identified four previously undetected mixed infections, and a previously undetected transmission event, but no direct transmission between the pairs of cases under investigation. These results demonstrate that mixed infections can be detected without additional sequencing effort, and this will be important in assessing the extent of cryptic transmission in our hospitals. PMID:23658511

A wide extent of inter-strain diversity in virulent and vaccine strains of alphaherpesviruses.

PubMed

Szpara, Moriah L; Tafuri, Yolanda R; Parsons, Lance; Shamim, S Rafi; Verstrepen, Kevin J; Legendre, Matthieu; Enquist, L W

2011-10-01

Alphaherpesviruses are widespread in the human population, and include herpes simplex virus 1 (HSV-1) and 2, and varicella zoster virus (VZV). These viral pathogens cause epithelial lesions, and then infect the nervous system to cause lifelong latency, reactivation, and spread. A related veterinary herpesvirus, pseudorabies (PRV), causes similar disease in livestock that result in significant economic losses. Vaccines developed for VZV and PRV serve as useful models for the development of an HSV-1 vaccine. We present full genome sequence comparisons of the PRV vaccine strain Bartha, and two virulent PRV isolates, Kaplan and Becker. These genome sequences were determined by high-throughput sequencing and assembly, and present new insights into the attenuation of a mammalian alphaherpesvirus vaccine strain. We find many previously unknown coding differences between PRV Bartha and the virulent strains, including changes to the fusion proteins gH and gB, and over forty other viral proteins. Inter-strain variation in PRV protein sequences is much closer to levels previously observed for HSV-1 than for the highly stable VZV proteome. Almost 20% of the PRV genome contains tandem short sequence repeats (SSRs), a class of nucleic acids motifs whose length-variation has been associated with changes in DNA binding site efficiency, transcriptional regulation, and protein interactions. We find SSRs throughout the herpesvirus family, and provide the first global characterization of SSRs in viruses, both within and between strains. We find SSR length variation between different isolates of PRV and HSV-1, which may provide a new mechanism for phenotypic variation between strains. Finally, we detected a small number of polymorphic bases within each plaque-purified PRV strain, and we characterize the effect of passage and plaque-purification on these polymorphisms. These data add to growing evidence that even plaque-purified stocks of stable DNA viruses exhibit limited sequence heterogeneity, which likely seeds future strain evolution.

Draft Genome Sequence of Achromobacter sp. Strain AR476-2, Isolated from a Cellulolytic Consortium

PubMed Central

Kurth, Daniel; Romero, Cintia M.; Fernandez, Pablo M.; Ferrero, Marcela A.

2016-01-01

Achromobacter sp. AR476-2 is a noncellulolytic strain previously isolated from a cellulolytic consortium selected from samples of insect gut. Its genome sequence could contribute to the unraveling of the complex interaction of microorganisms and enzymes involved in the biodegradation of lignocellulosic biomass in nature. PMID:27340069

Draft Genome Sequence of Lactobacillus pobuzihii E100301T.

PubMed

Chiu, Chi-Ming; Chang, Chi-Huan; Pan, Shwu-Fen; Wu, Hui-Chung; Li, Shiao-Wen; Chang, Chuan-Hsiung; Lee, Yun-Shien; Chiang, Chih-Ming; Chen, Yi-Sheng

2013-05-09

Lactobacillus pobuzihii E100301(T) is a novel Lactobacillus species previously isolated from pobuzihi (fermented cummingcordia) in Taiwan. Phylogenetically, this strain is closest to Lactobacillus acidipiscis, but its phenotypic characteristics can be clearly distinguished from those of L. acidipiscis. We present the draft genome sequence of strain L. pobuzihii E100301(T).

Draft Genome Sequence of Xylella fastidiosa subsp. fastidiosa Strain Stag's Leap.

PubMed

Chen, J; Wu, F; Zheng, Z; Deng, X; Burbank, L P; Stenger, D C

2016-04-21

ITALIC! Xylella fastidiosasubsp. ITALIC! fastidiosacauses Pierce's disease of grapevine. Presented here is the draft genome sequence of the Stag's Leap strain, previously used in pathogenicity/virulence assays to evaluate grapevine germplasm bearing Pierce's disease resistance and a phenotypic assessment of knockout mutants to determine gene function. Copyright © 2016 Chen et al.

Taxonomic evaluation of putative Streptomyces scabiei strains held in the ARS (NRRL) Culture Collection using multi-locus sequence analysis

USDA-ARS?s Scientific Manuscript database

Multi-locus sequence analysis has been demonstrated to be a useful tool for identification of Streptomyces species and was previously applied to phylogenetically differentiate the type strains of species pathogenic on potatoes (Solanum tuberosum L.). The ARS Culture Collection (NRRL) contains 43 str...

Terminal region sequence variations in variola virus DNA.

PubMed

Massung, R F; Loparev, V N; Knight, J C; Totmenin, A V; Chizhikov, V E; Parsons, J M; Safronov, P F; Gutorov, V V; Shchelkunov, S N; Esposito, J J

1996-07-15

Genome DNA terminal region sequences were determined for a Brazilian alastrim variola minor virus strain Garcia-1966 that was associated with an 0.8% case-fatality rate and African smallpox strains Congo-1970 and Somalia-1977 associated with variola major (9.6%) and minor (0.4%) mortality rates, respectively. A base sequence identity of > or = 98.8% was determined after aligning 30 kb of the left- or right-end region sequences with cognate sequences previously determined for Asian variola major strains India-1967 (31% death rate) and Bangladesh-1975 (18.5% death rate). The deduced amino acid sequences of putative proteins of > or = 65 amino acids also showed relatively high identity, although the Asian and African viruses were clearly more related to each other than to alastrim virus. Alastrim virus contained only 10 of 70 proteins that were 100% identical to homologs in Asian strains, and 7 alastrim-specific proteins were noted.

High resolution identity testing of inactivated poliovirus vaccines

PubMed Central

Mee, Edward T.; Minor, Philip D.; Martin, Javier

2015-01-01

Background Definitive identification of poliovirus strains in vaccines is essential for quality control, particularly where multiple wild-type and Sabin strains are produced in the same facility. Sequence-based identification provides the ultimate in identity testing and would offer several advantages over serological methods. Methods We employed random RT-PCR and high throughput sequencing to recover full-length genome sequences from monovalent and trivalent poliovirus vaccine products at various stages of the manufacturing process. Results All expected strains were detected in previously characterised products and the method permitted identification of strains comprising as little as 0.1% of sequence reads. Highly similar Mahoney and Sabin 1 strains were readily discriminated on the basis of specific variant positions. Analysis of a product known to contain incorrect strains demonstrated that the method correctly identified the contaminants. Conclusion Random RT-PCR and shotgun sequencing provided high resolution identification of vaccine components. In addition to the recovery of full-length genome sequences, the method could also be easily adapted to the characterisation of minor variant frequencies and distinction of closely related products on the basis of distinguishing consensus and low frequency polymorphisms. PMID:26049003

Phylogeny of the family Moraxellaceae by 16S rDNA sequence analysis, with special emphasis on differentiation of Moraxella species.

PubMed

Pettersson, B; Kodjo, A; Ronaghi, M; Uhlén, M; Tønjum, T

1998-01-01

Thirty-three strains previously classified into 11 species in the bacterial family Moraxellaceae were subjected to phylogenetic analysis based on 16S rRNA sequences. The family Moraxellaceae formed a distinct clade consisting of four phylogenetic groups as judged from branch lengths, bootstrap values and signature nucleotides. Group I contained the classical moraxellae and strains of the coccal moraxellae, previously known as Branhamella, with 16S rRNA similarity of > or = 95%. A further division of group I into five tentative clusters is discussed. Group II consisted of two strains representing Moraxella atlantae and Moraxella osloensis. These strains were only distantly related to each other (93.4%) and also to the other members of the Moraxellaceae (< or = 93%). Therefore, reasons for reclassification of these species into separate and new genera are discussed. Group III harboured strains of the genus Psychrobacter and strain 752/52 of [Moraxella] phenylpyruvica. This strain of [M.] phenylpyruvica formed an early branch from the group III line of descent. Interestingly, a distant relationship was found between Psychrobacter phenylpyruvicus strain ATCC 23333T (formerly classified as [M.] phenylpyruvica) and [M.] phenylpyruvica strain 752/52, exhibiting less than 96% nucleotide similarity between their 16S rRNA sequences. The establishment of a new genus for [M.] phenylpyruvica strain 752/52 is therefore suggested. Group IV contained only two strains of the genus Acinetobacter. Strategies for the development of diagnostic probes and distinctive sequences for 16S rRNA-based species-specific assays within group I are suggested. Although these findings add to the classificatory placements within the Moraxellaceae, analysis of a more comprehensive selection of strains is still needed to obtain a complete classification system within this family.

Comparative pathogenomics of Clostridium tetani.

PubMed

Cohen, Jonathan E; Wang, Rong; Shen, Rong-Fong; Wu, Wells W; Keller, James E

2017-01-01

Clostridium tetani and Clostridium botulinum produce two of the most potent neurotoxins known, tetanus neurotoxin and botulinum neurotoxin, respectively. Extensive biochemical and genetic investigation has been devoted to identifying and characterizing various C. botulinum strains. Less effort has been focused on studying C. tetani likely because recently sequenced strains of C. tetani show much less genetic diversity than C. botulinum strains and because widespread vaccination efforts have reduced the public health threat from tetanus. Our aim was to acquire genomic data on the U.S. vaccine strain of C. tetani to better understand its genetic relationship to previously published genomic data from European vaccine strains. We performed high throughput genomic sequence analysis on two wild-type and two vaccine C. tetani strains. Comparative genomic analysis was performed using these and previously published genomic data for seven other C. tetani strains. Our analysis focused on single nucleotide polymorphisms (SNP) and four distinct constituents of the mobile genome (mobilome): a hypervariable flagellar glycosylation island region, five conserved bacteriophage insertion regions, variations in three CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) systems, and a single plasmid. Intact type IA and IB CRISPR/Cas systems were within 10 of 11 strains. A type IIIA CRISPR/Cas system was present in two strains. Phage infection histories derived from CRISPR-Cas sequences indicate C. tetani encounters phages common among commensal gut bacteria and soil-borne organisms consistent with C. tetani distribution in nature. All vaccine strains form a clade distinct from currently sequenced wild type strains when considering variations in these mobile elements. SNP, flagellar glycosylation island, prophage content and CRISPR/Cas phylogenic histories provide tentative evidence suggesting vaccine and wild type strains share a common ancestor.

Structure and Genetic Content of the Megaplasmids of Neurotoxigenic Clostridium butyricum Type E Strains from Italy

PubMed Central

Iacobino, Angelo; Scalfaro, Concetta; Franciosa, Giovanna

2013-01-01

We determined the genetic maps of the megaplasmids of six neutoroxigenic Clostridium butyricum type E strains from Italy using molecular and bioinformatics techniques. The megaplasmids are circular, not linear as we had previously proposed. The differently-sized megaplasmids share a genetic region that includes structural, metabolic and regulatory genes. In addition, we found that a 168 kb genetic region is present only in the larger megaplasmids of two tested strains, whereas it is absent from the smaller megaplasmids of the four remaining strains. The genetic region unique to the larger megaplasmids contains, among other features, a locus for clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated (cas) genes, i.e. a bacterial adaptive immune system providing sequence-specific protection from invading genetic elements. Some CRISPR spacer sequences of the neurotoxigenic C. butyricum type E strains showed homology to prophage, phage and plasmid sequences from closely related clostridia species or from distant species, all sharing the intestinal habitat, suggesting that the CRISPR locus might be involved in the microorganism adaptation to the human or animal intestinal environment. Besides, we report here that each of four distinct CRISPR spacers partially matched DNA sequences of different prophages and phages, at identical nucleotide locations. This suggests that, at least in neurotoxigenic C. butyricum type E, the CRISPR locus is potentially able to recognize the same conserved DNA sequence of different invading genetic elements, besides targeting sequences unique to previously encountered invading DNA, as currently predicted for a CRISPR locus. Thus, the results of this study introduce the possibility that CRISPR loci can provide resistance to a wider range of invading DNA elements than previously appreciated. Whether it is more advantageous for the peculiar neurotoxigenic C. butyricum type E strains to maintain or to lose the CRISPR-cas system remains an open question. PMID:23967192

Draft Genome Sequence of Burkholderia cenocepacia Strain CEIB S5-2, a Methyl Parathion- and p-Nitrophenol-Degrading Bacterium, Isolated from Agricultural Soils in Morelos, Mexico

PubMed Central

Martínez-Ocampo, Fernando; Fernández López, Maikel Gilberto; Lozano-Aguirre Beltrán, Luis Fernando; Popoca-Ursino, Elida Carolina; Ortiz-Hernández, M. Laura; Sánchez-Salinas, Enrique; Ramos Quintana, Fernando; Villalobos-López, Miguel A.

2016-01-01

Burkholderia cenocepacia is an opportunistic pathogen that belongs to Burkholderia cepacia complex (BCC). Burkholderia cenocepacia strain CEIB S5-2 was isolated from agricultural soils in Morelos, Mexico, and previously has shown its abilities for bioremediation. In this study, we report the draft genome sequence of Burkholderia cenocepacia strain CEIB S5-2. PMID:27125479

[The genetical evolution of the full length genes of 5 EV 71 strains from 5 Shenzhen patients with hand-food-mouth disease associated with EV71 infection].

PubMed

Liu, Wei-long; Yang, Gui-lin; Wei, Qing; Zhang, Ming-xia; Chen, Xin-chun; Liu, Ying-xia; Gao, Yang; Zhou, Bo-ping

2011-02-01

To investigate the characteristics of molecular epidemiology and molecular evolution of 5 EV 71 (enterovirus 71, EV71) strains from 5 Shenzhen patients with hand-food-mouth disease associated with EV 71 infection. 5 EV 71 strains were isolated, and sequenced to analyzed the full length gene sequences in order to compare nucleotide and amino acid homology with other EV71 strains from other regions and countries as well as previous strains across the world through bioinformatics software. 5 strains of EV 71 belonged to sub-genotype C4 by analysis of nucleotide sequences of VP1 and VP4 of EV 71. The differences of nucleotide and amino acid sequences were much small with nucleotide homology of 93% and amino acid homology of 98% among these 5 strains. A phylogenetic tree analysis indicated that 2008 Shenzhen epidemic strains were the most close to 2004 Shenzhen circulating strains, and also much close to 1998 Shenzhen epidemic strains and 2008 Fuyang Anhui strains. The dead strain was very close to 2008 Fuyang Anhui epidemic strains. It can be speculated that this epidemic strains of EV 71 probably originate from the same ancient strain in the history, may from 1998 Shenzhen strain.

Draft Genome Sequence of Achromobacter sp. Strain AR476-2, Isolated from a Cellulolytic Consortium.

PubMed

Kurth, Daniel; Romero, Cintia M; Fernandez, Pablo M; Ferrero, Marcela A; Martinez, M Alejandra

2016-06-23

Achromobacter sp. AR476-2 is a noncellulolytic strain previously isolated from a cellulolytic consortium selected from samples of insect gut. Its genome sequence could contribute to the unraveling of the complex interaction of microorganisms and enzymes involved in the biodegradation of lignocellulosic biomass in nature. Copyright © 2016 Kurth et al.

Draft Genome Sequence of Lactobacillus pobuzihii E100301T

PubMed Central

Chiu, Chi-ming; Chang, Chi-huan; Pan, Shwu-fen; Wu, Hui-chung; Li, Shiao-wen; Chang, Chuan-hsiung; Lee, Yun-shien; Chiang, Chih-ming

2013-01-01

Lactobacillus pobuzihii E100301T is a novel Lactobacillus species previously isolated from pobuzihi (fermented cummingcordia) in Taiwan. Phylogenetically, this strain is closest to Lactobacillus acidipiscis, but its phenotypic characteristics can be clearly distinguished from those of L. acidipiscis. We present the draft genome sequence of strain L. pobuzihii E100301T. PMID:23661478

Taxonomic evaluation of Streptomyces albus and related species using multilocus sequence analysis and proposals to emend the description of Streptomyces albus and describe Streptomyces pathocidini sp. nov.

PubMed Central

Doroghazi, J. R.; Ju, K.-S.; Metcalf, W. W.

2014-01-01

In phylogenetic analyses of the genus Streptomyces using 16S rRNA gene sequences, Streptomyces albus subsp. albus NRRL B-1811T forms a cluster with five other species having identical or nearly identical 16S rRNA gene sequences. Moreover, the morphological and physiological characteristics of these other species, including Streptomyces almquistii NRRL B-1685T, Streptomyces flocculus NRRL B-2465T, Streptomyces gibsonii NRRL B-1335T and Streptomyces rangoonensis NRRL B-12378T are quite similar. This cluster is of particular taxonomic interest because Streptomyces albus is the type species of the genus Streptomyces. The related strains were subjected to multilocus sequence analysis (MLSA) utilizing partial sequences of the housekeeping genes atpD, gyrB, recA, rpoB and trpB and confirmation of previously reported phenotypic characteristics. The five strains formed a coherent cluster supported by a 100 % bootstrap value in phylogenetic trees generated from sequence alignments prepared by concatenating the sequences of the housekeeping genes, and identical tree topology was observed using various different tree-making algorithms. Moreover, all but one strain, S. flocculus NRRL B-2465T, exhibited identical sequences for all of the five housekeeping gene loci sequenced, but NRRL B-2465T still exhibited an MLSA evolutionary distance of 0.005 from the other strains, a value that is lower than the 0.007 MLSA evolutionary distance threshold proposed for species-level relatedness. These data support a proposal to reclassify S. almquistii, S. flocculus, S. gibsonii and S. rangoonensis as later heterotypic synonyms of S. albus with NRRL B-1811T as the type strain. The MLSA sequence database also demonstrated utility for quickly and conclusively confirming that numerous strains within the ARS Culture Collection had been previously misidentified as subspecies of S. albus and that Streptomyces albus subsp. pathocidicus should be redescribed as a novel species, Streptomyces pathocidini sp. nov., with the type strain NRRL B-24287T. PMID:24277863

Viral forensic genomics reveals the relatedness of classic herpes simplex virus strains KOS, KOS63, and KOS79

PubMed Central

Bowen, Christopher D.; Renner, Daniel W.; Shreve, Jacob T.; Tafuri, Yolanda; Payne, Kimberly M.; Dix, Richard D.; Kinchington, Paul R.; Gatherer, Derek; Szpara, Moriah L.

2016-01-01

Herpes simplex virus 1 (HSV-1) is a widespread global pathogen, of which the strain KOS is one of the most extensively studied. Previous sequence studies revealed that KOS does not cluster with other strains of North American geographic origin, but instead clustered with Asian strains. We sequenced a historical isolate of the original KOS strain, called KOS63, along with a separately isolated strain attributed to the same source individual, termed KOS79. Genomic analyses revealed that KOS63 closely resembled other recently sequenced isolates of KOS and was of Asian origin, but that KOS79 was a genetically unrelated strain that clustered in genetic distance analyses with HSV-1 strains of North American/European origin. These data suggest that the human source of KOS63 and KOS79 could have been infected with two genetically unrelated strains of disparate geographic origins. A PCR RFLP test was developed for rapid identification of these strains. PMID:26950505

Viral forensic genomics reveals the relatedness of classic herpes simplex virus strains KOS, KOS63, and KOS79.

PubMed

Bowen, Christopher D; Renner, Daniel W; Shreve, Jacob T; Tafuri, Yolanda; Payne, Kimberly M; Dix, Richard D; Kinchington, Paul R; Gatherer, Derek; Szpara, Moriah L

2016-05-01

Herpes simplex virus 1 (HSV-1) is a widespread global pathogen, of which the strain KOS is one of the most extensively studied. Previous sequence studies revealed that KOS does not cluster with other strains of North American geographic origin, but instead clustered with Asian strains. We sequenced a historical isolate of the original KOS strain, called KOS63, along with a separately isolated strain attributed to the same source individual, termed KOS79. Genomic analyses revealed that KOS63 closely resembled other recently sequenced isolates of KOS and was of Asian origin, but that KOS79 was a genetically unrelated strain that clustered in genetic distance analyses with HSV-1 strains of North American/European origin. These data suggest that the human source of KOS63 and KOS79 could have been infected with two genetically unrelated strains of disparate geographic origins. A PCR RFLP test was developed for rapid identification of these strains. Copyright © 2016 Elsevier Inc. All rights reserved.

Revalidation and genetic characterization of new members of Group C (Orthobunyavirus genus, Peribunyaviridae family) isolated in the Americas.

PubMed

Nunes, Márcio Roberto Teixeira; de Souza, William Marciel; Acrani, Gustavo Olszanski; Cardoso, Jedson Ferreira; da Silva, Sandro Patroca; Badra, Soraya Jabur; Figueiredo, Luiz Tadeu Moraes; Vasconcelos, Pedro Fernando da Costa

2018-01-01

Group C serogroup includes members of the Orthobunyavirus genus (family Peribunyaviridae) and comprises 15 arboviruses that can be associated with febrile illness in humans. Although previous studies described the genome characterization of Group C orthobunyavirus, there is a gap in genomic information about the other viruses in this group. Therefore, in this study, complete genomes of members of Group C serogroup were sequenced or re-sequenced and used for genetic characterization, as well as to understand their phylogenetic and evolutionary aspects. Thus, our study reported the genomes of three new members in Group C virus (Apeu strain BeAn848, Itaqui strain BeAn12797 and Nepuyo strain BeAn10709), as well as re-sequencing of original strains of five members: Caraparu (strain BeAn3994), Madrid (strain BT4075), Murucutu (strain BeAn974), Oriboca (strain BeAn17), and Marituba (strain BeAn15). These viruses presented a typical genomic organization related to members of the Orthobunyavirus genus. Interestingly, all viruses of this serogroup showed an open reading frame (ORF) that encodes the putative nonstructural NSs protein that precedes the nucleoprotein ORF, an unprecedented fact in Group C virus. Also, we confirmed the presence of natural reassortment events. This study expands the genomic information of Group C viruses, as well as revalidates the genomic organization of viruses that were previously reported.

Complete Genome Sequence of a Novel Newcastle Disease Virus Strain Isolated from a Chicken in West Africa

PubMed Central

Kim, Shin-Hee; Nayak, Subhashree; Paldurai, Anandan; Nayak, Baibaswata; Samuel, Arthur; Aplogan, Gilbert L.; Awoume, Kodzo A.; Webby, Richard J.; Ducatez, Mariette F.; Collins, Peter L.

2012-01-01

The complete genome sequence of an African Newcastle disease virus (NDV) strain isolated from a chicken in Togo in 2009 was determined. The genome is 15,198 nucleotides (nt) in length and is classified in genotype VII in the class II cluster. Compared to common vaccine strains, the African strain contains a previously described 6-nt insert in the downstream untranslated region of the N gene and a novel 6-nt insert in the HN-L intergenic region. Genome length differences are a marker of the natural history of NDV. This is the first description of a class II NDV strain with a genome of 15,198 nt and a 6-nt insert in the HN-L intergenic region. Sequence divergence relative to vaccine strains was substantial, likely contributes to outbreaks, and illustrates the continued evolution of new NDV strains in West Africa. PMID:22997417

How close is close: 16S rRNA sequence identity may not be sufficient to guarantee species identity

NASA Technical Reports Server (NTRS)

Fox, G. E.; Wisotzkey, J. D.; Jurtshuk, P. Jr

1992-01-01

16S rRNA (genes coding for rRNA) sequence comparisons were conducted with the following three psychrophilic strains: Bacillus globisporus W25T (T = type strain) and Bacillus psychrophilus W16AT, and W5. These strains exhibited more than 99.5% sequence identity and within experimental uncertainty could be regarded as identical. Their close taxonomic relationship was further documented by phenotypic similarities. In contrast, previously published DNA-DNA hybridization results have convincingly established that these strains do not belong to the same species if current standards are used. These results emphasize the important point that effective identity of 16S rRNA sequences is not necessarily a sufficient criterion to guarantee species identity. Thus, although 16S rRNA sequences can be used routinely to distinguish and establish relationships between genera and well-resolved species, very recently diverged species may not be recognizable.

The genome sequence of 'Mycobacterium massiliense' strain CIP 108297 suggests the independent taxonomic status of the Mycobacterium abscessus complex at the subspecies level.

PubMed

Cho, Yong-Joon; Yi, Hana; Chun, Jongsik; Cho, Sang-Nae; Daley, Charles L; Koh, Won-Jung; Shin, Sung Jae

2013-01-01

Members of the Mycobacterium abscessus complex are rapidly growing mycobacteria that are emerging as human pathogens. The M. abscessus complex was previously composed of three species, namely M. abscessus sensu stricto, 'M. massiliense', and 'M. bolletii'. In 2011, 'M. massiliense' and 'M. bolletii' were united and reclassified as a single subspecies within M. abscessus: M. abscessus subsp. bolletii. However, the placement of 'M. massiliense' within the boundary of M. abscessus subsp. bolletii remains highly controversial with regard to clinical aspects. In this study, we revisited the taxonomic status of members of the M. abscessus complex based on comparative analysis of the whole-genome sequences of 53 strains. The genome sequence of the previous type strain of 'Mycobacterium massiliense' (CIP 108297) was determined using next-generation sequencing. The genome tree based on average nucleotide identity (ANI) values supported the differentiation of 'M. bolletii' and 'M. massiliense' at the subspecies level. The genome tree also clearly illustrated that 'M. bolletii' and 'M. massiliense' form a distinct phylogenetic clade within the radiation of the M. abscessus complex. The genomic distances observed in this study suggest that the current M. abscessus subsp. bolletii taxon should be divided into two subspecies, M. abscessus subsp. massiliense subsp. nov. and M. abscessus subsp. bolletii, to correspondingly accommodate the previously known 'M. massiliense' and 'M. bolletii' strains.

Genome sequence of a serotype M3 strain of group A Streptococcus: phage-encoded toxins, the high-virulence phenotype, and clone emergence.

PubMed

Beres, Stephen B; Sylva, Gail L; Barbian, Kent D; Lei, Benfang; Hoff, Jessica S; Mammarella, Nicole D; Liu, Meng-Yao; Smoot, James C; Porcella, Stephen F; Parkins, Larye D; Campbell, David S; Smith, Todd M; McCormick, John K; Leung, Donald Y M; Schlievert, Patrick M; Musser, James M

2002-07-23

Genome sequences are available for many bacterial strains, but there has been little progress in using these data to understand the molecular basis of pathogen emergence and differences in strain virulence. Serotype M3 strains of group A Streptococcus (GAS) are a common cause of severe invasive infections with unusually high rates of morbidity and mortality. To gain insight into the molecular basis of this high-virulence phenotype, we sequenced the genome of strain MGAS315, an organism isolated from a patient with streptococcal toxic shock syndrome. The genome is composed of 1,900,521 bp, and it shares approximately 1.7 Mb of related genetic material with genomes of serotype M1 and M18 strains. Phage-like elements account for the great majority of variation in gene content relative to the sequenced M1 and M18 strains. Recombination produces chimeric phages and strains with previously uncharacterized arrays of virulence factor genes. Strain MGAS315 has phage genes that encode proteins likely to contribute to pathogenesis, such as streptococcal pyrogenic exotoxin A (SpeA) and SpeK, streptococcal superantigen (SSA), and a previously uncharacterized phospholipase A(2) (designated Sla). Infected humans had anti-SpeK, -SSA, and -Sla antibodies, indicating that these GAS proteins are made in vivo. SpeK and SSA were pyrogenic and toxic for rabbits. Serotype M3 strains with the phage-encoded speK and sla genes increased dramatically in frequency late in the 20th century, commensurate with the rise in invasive disease caused by M3 organisms. Taken together, the results show that phage-mediated recombination has played a critical role in the emergence of a new, unusually virulent clone of serotype M3 GAS.

Evolution and Diversity in Human Herpes Simplex Virus Genomes

PubMed Central

Gatherer, Derek; Ochoa, Alejandro; Greenbaum, Benjamin; Dolan, Aidan; Bowden, Rory J.; Enquist, Lynn W.; Legendre, Matthieu; Davison, Andrew J.

2014-01-01

Herpes simplex virus 1 (HSV-1) causes a chronic, lifelong infection in >60% of adults. Multiple recent vaccine trials have failed, with viral diversity likely contributing to these failures. To understand HSV-1 diversity better, we comprehensively compared 20 newly sequenced viral genomes from China, Japan, Kenya, and South Korea with six previously sequenced genomes from the United States, Europe, and Japan. In this diverse collection of passaged strains, we found that one-fifth of the newly sequenced members share a gene deletion and one-third exhibit homopolymeric frameshift mutations (HFMs). Individual strains exhibit genotypic and potential phenotypic variation via HFMs, deletions, short sequence repeats, and single-nucleotide polymorphisms, although the protein sequence identity between strains exceeds 90% on average. In the first genome-scale analysis of positive selection in HSV-1, we found signs of selection in specific proteins and residues, including the fusion protein glycoprotein H. We also confirmed previous results suggesting that recombination has occurred with high frequency throughout the HSV-1 genome. Despite this, the HSV-1 strains analyzed clustered by geographic origin during whole-genome distance analysis. These data shed light on likely routes of HSV-1 adaptation to changing environments and will aid in the selection of vaccine antigens that are invariant worldwide. PMID:24227835

Novel Molecular Method for Identification of Streptococcus pneumoniae Applicable to Clinical Microbiology and 16S rRNA Sequence-Based Microbiome Studies

PubMed Central

Scholz, Christian F. P.; Poulsen, Knud

2012-01-01

The close phylogenetic relationship of the important pathogen Streptococcus pneumoniae and several species of commensal streptococci, particularly Streptococcus mitis and Streptococcus pseudopneumoniae, and the recently demonstrated sharing of genes and phenotypic traits previously considered specific for S. pneumoniae hamper the exact identification of S. pneumoniae. Based on sequence analysis of 16S rRNA genes of a collection of 634 streptococcal strains, identified by multilocus sequence analysis, we detected a cytosine at position 203 present in all 440 strains of S. pneumoniae but replaced by an adenosine residue in all strains representing other species of mitis group streptococci. The S. pneumoniae-specific sequence signature could be demonstrated by sequence analysis or indirectly by restriction endonuclease digestion of a PCR amplicon covering the site. The S. pneumoniae-specific signature offers an inexpensive means for validation of the identity of clinical isolates and should be used as an integrated marker in the annotation procedure employed in 16S rRNA-based molecular studies of complex human microbiotas. This may avoid frequent misidentifications such as those we demonstrate to have occurred in previous reports and in reference sequence databases. PMID:22442329

High resolution identity testing of inactivated poliovirus vaccines.

PubMed

Mee, Edward T; Minor, Philip D; Martin, Javier

2015-07-09

Definitive identification of poliovirus strains in vaccines is essential for quality control, particularly where multiple wild-type and Sabin strains are produced in the same facility. Sequence-based identification provides the ultimate in identity testing and would offer several advantages over serological methods. We employed random RT-PCR and high throughput sequencing to recover full-length genome sequences from monovalent and trivalent poliovirus vaccine products at various stages of the manufacturing process. All expected strains were detected in previously characterised products and the method permitted identification of strains comprising as little as 0.1% of sequence reads. Highly similar Mahoney and Sabin 1 strains were readily discriminated on the basis of specific variant positions. Analysis of a product known to contain incorrect strains demonstrated that the method correctly identified the contaminants. Random RT-PCR and shotgun sequencing provided high resolution identification of vaccine components. In addition to the recovery of full-length genome sequences, the method could also be easily adapted to the characterisation of minor variant frequencies and distinction of closely related products on the basis of distinguishing consensus and low frequency polymorphisms. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Short, interspersed, and repetitive DNA sequences in Spiroplasma species.

PubMed

Nur, I; LeBlanc, D J; Tully, J G

1987-03-01

Small fragments of DNA from an 8-kbp plasmid, pRA1, from a plant pathogenic strain of Spiroplasma citri were shown previously to be present in the chromosomal DNA of at least two species of Spiroplasma. We describe here the shot-gun cloning of chromosomal DNA from S. citri Maroc and the identification of two distinct sequences exhibiting homology to pRA1. Further subcloning experiments provided specific molecular probes for the identification of these two sequences in chromosomal DNA from three distinct plant pathogenic species of Spiroplasma. The results of Southern blot hybridization indicated that each of the pRA1-associated sequences is present as multiple copies in short, dispersed, and repetitive sequences in the chromosomes of these three strains. None of the sequences was detectable in chromosomal DNA from an additional nine Spiroplasma strains examined.

Bifidobacterium animalis subsp. lactis ATCC 27673 Is a Genomically Unique Strain within Its Conserved Subspecies

PubMed Central

Loquasto, Joseph R.; Barrangou, Rodolphe; Dudley, Edward G.; Stahl, Buffy; Chen, Chun

2013-01-01

Many strains of Bifidobacterium animalis subsp. lactis are considered health-promoting probiotic microorganisms and are commonly formulated into fermented dairy foods. Analyses of previously sequenced genomes of B. animalis subsp. lactis have revealed little genetic diversity, suggesting that it is a monomorphic subspecies. However, during a multilocus sequence typing survey of Bifidobacterium, it was revealed that B. animalis subsp. lactis ATCC 27673 gave a profile distinct from that of the other strains of the subspecies. As part of an ongoing study designed to understand the genetic diversity of this subspecies, the genome of this strain was sequenced and compared to other sequenced genomes of B. animalis subsp. lactis and B. animalis subsp. animalis. The complete genome of ATCC 27673 was 1,963,012 bp, contained 1,616 genes and 4 rRNA operons, and had a G+C content of 61.55%. Comparative analyses revealed that the genome of ATCC 27673 contained six distinct genomic islands encoding 83 open reading frames not found in other strains of the same subspecies. In four islands, either phage or mobile genetic elements were identified. In island 6, a novel clustered regularly interspaced short palindromic repeat (CRISPR) locus which contained 81 unique spacers was identified. This type I-E CRISPR-cas system differs from the type I-C systems previously identified in this subspecies, representing the first identification of a different system in B. animalis subsp. lactis. This study revealed that ATCC 27673 is a strain of B. animalis subsp. lactis with novel genetic content and suggests that the lack of genetic variability observed is likely due to the repeated sequencing of a limited number of widely distributed commercial strains. PMID:23995933

Detection and analysis of recombination in GII.4 norovirus strains causing gastroenteritis outbreaks in Alberta.

PubMed

Hasing, Maria E; Hazes, Bart; Lee, Bonita E; Preiksaitis, Jutta K; Pang, Xiaoli L

2014-10-01

Recombination is an important mechanism generating genetic diversity in norovirus (NoV) that occurs commonly at the NoV polymerase-capsid (ORF1/2) junction. The genotyping method based on partial ORF2 sequences currently used to characterize circulating NoV strains in gastroenteritis outbreaks in Alberta cannot detect such recombination events and provides only limited information on NoV genetic evolution. The objective of this study was to determine whether any NoV GII.4 strains causing outbreaks in Alberta are recombinants. Twenty stool samples collected during outbreaks occurring between July 2004 and January 2012 were selected to include the GII.4 variants Farmington Hills 2002, Hunter 2004, Yerseke 2006a, Den Haag 2006b, Apeldoorn 2007, New Orleans 2009, and Sydney 2012 based on previous NoV ORF2-genotyping results. Near full-length NoV genome sequences were obtained, aligned with reference sequences from GenBank and analyzed with RDPv4.13. Two sequences corresponding to Apeldoorn 2007, and Sydney 2012 were identified as recombinants with breakpoints near the ORF1/2 junction and putative parental strains as previously reported. We also identified, for the first time, a non-recombinant sequence resembling the ORF2-3 parent of the recombinant cluster Sydney 2012 responsible for the most recent pandemic. Our results confirmed the presence of recombinant NoV GII.4 strains in Alberta, and highlight the importance of including additional genomic regions in surveillance studies to trace the evolution of pandemic NoV GII.4 strains. Copyright © 2014 Elsevier B.V. All rights reserved.

Molecular analysis of the rRNA genes of Babesia spp and Ehrlichia canis detected in dogs from RibeirÃo Preto, Brazil

PubMed Central

Oliveira, L.P.; Cardozo, G.P.; Santos, E.V.; Mansur, M.A.B.; Donini, I.A.N.; Zissou, V.G.; Roberto, P.G.; Marins, M.

2009-01-01

The partial DNA sequences of the 18S rRNA gene of Babesia canis and the 16S rRNA gene of Ehrlichia canis detected in dogs from Ribeirão Preto, Brazil, were compared to sequences from other strains deposited in GenBank. The E. canis strain circulating in Ribeirão Preto is identical to other strains previously detected in the region, whereas the subspecies Babesia canis vogeli is the main Babesia strain circulating in dogs from Ribeirão Preto. PMID:24031351

Complete Deletion of the Fucose Operon in Haemophilus influenzae Is Associated with a Cluster in Multilocus Sequence Analysis-Based Phylogenetic Group II Related to Haemophilus haemolyticus: Implications for Identification and Typing

PubMed Central

de Gier, Camilla; Kirkham, Lea-Ann S.

2015-01-01

Nonhemolytic variants of Haemophilus haemolyticus are difficult to differentiate from Haemophilus influenzae despite a wide difference in pathogenic potential. A previous investigation characterized a challenging set of 60 clinical strains using multiple PCRs for marker genes and described strains that could not be unequivocally identified as either species. We have analyzed the same set of strains by multilocus sequence analysis (MLSA) and near-full-length 16S rRNA gene sequencing. MLSA unambiguously allocated all study strains to either of the two species, while identification by 16S rRNA sequence was inconclusive for three strains. Notably, the two methods yielded conflicting identifications for two strains. Most of the “fuzzy species” strains were identified as H. influenzae that had undergone complete deletion of the fucose operon. Such strains, which are untypeable by the H. influenzae multilocus sequence type (MLST) scheme, have sporadically been reported and predominantly belong to a single branch of H. influenzae MLSA phylogenetic group II. We also found evidence of interspecies recombination between H. influenzae and H. haemolyticus within the 16S rRNA genes. Establishing an accurate method for rapid and inexpensive identification of H. influenzae is important for disease surveillance and treatment. PMID:26378279

Draft Genome Sequence of Oil-Degrading Bacterium Gallaecimonas pentaromativorans Strain YA_1 from the Southwest Indian Ocean

PubMed Central

Xu, Yiyuan; Ren, Chong; Chen, Ruixuan

2016-01-01

Gallaecimonas pentaromativorans has been previously reported to be capable of degrading crude oil and diesel oil. G. pentaromativorans strain YA_1 was isolated from the southwest Indian Ocean and can degrade crude oil. This study reports the draft genome sequence of G. pentaromativorans, which can provide insights into the mechanisms of microbial oil biodegradation. PMID:27491993

Serovar distribution of a DNA sequence involved in the antigenic relationship between Leptospira and equine cornea.

PubMed

Lucchesi, Paula M A; Parma, Alberto E; Arroyo, Guillermo H

2002-01-01

Horses infected with Leptospira present several clinical disorders, one of them being recurrent uveitis. A common endpoint of equine recurrent uveitis is blindness. Serovar pomona has often been incriminated, although others have also been reported. An antigenic relationship between this bacterium and equine cornea has been described in previous studies. A leptospiral DNA fragment that encodes cross-reacting epitopes was previously cloned and expressed in Escherichia coli. A region of that DNA fragment was subcloned and sequenced. Samples of leptospiral DNA from several sources were analysed by PCR with two primer pairs designed to amplify that region. Reference strains from serovars canicola, icterohaemorrhagiae, pomona, pyrogenes, wolffi, bataviae, sentot, hebdomadis and hardjo rendered products of the expected sizes with both pairs of primers. The specific DNA region was also amplified from isolates from Argentina belonging to serogroups Canicola and Pomona. Both L. biflexa serovar patoc and L. borgpetersenii serovar tarassovi rendered a negative result. The DNA sequence related to the antigen mimicry with equine cornea was not exclusively found in serovar pomona as it was also detected in several strains of Leptospira belonging to different serovars. The results obtained with L. biflexa serovar patoc strain Patoc I and L. borgpetersenii serovar tarassovi strain Perepelicin suggest that this sequence is not present in these strains, which belong to different genomospecies than those which gave positive results. This is an interesting finding since L. biflexa comprises nonpathogenic strains and serovar tarassovi has not been associated clinically with equine uveitis.

Genome sequence and analysis of a stress-tolerant, wild-derived strain of Saccharomyces cerevisiae used in biofuels research

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

McIlwain, Sean J.; Peris, Davis; Sardi, Maria

The genome sequences of more than 100 strains of the yeast Saccharomyces cerevisiae have been published. Unfortunately, most of these genome assemblies contain dozens to hundreds of gaps at repetitive sequences, including transposable elements, tRNAs, and subtelomeric regions, which is where novel genes generally reside. Relatively few strains have been chosen for genome sequencing based on their biofuel production potential, leaving an additional knowledge gap. Here, we describe the nearly complete genome sequence of GLBRCY22-3 (Y22-3), a strain of S. cerevisiae derived from the stress-tolerant wild strain NRRL YB-210 and subsequently engineered for xylose metabolism. After benchmarking several genome assemblymore » approaches, we developed a pipeline to integrate Pacific Biosciences (PacBio) and Illumina sequencing data and achieved one of the highest quality genome assemblies for any S. cerevisiae strain. Specifically, the contig N50 is 693 kbp, and the sequences of most chromosomes, the mitochondrial genome, and the 2-micron plasmid are complete. Our annotation predicts 92 genes that are not present in the reference genome of the laboratory strain S288c, over 70% of which were expressed. We predicted functions for 43 of these genes, 28 of which were previously uncharacterized and unnamed. Remarkably, many of these genes are predicted to be involved in stress tolerance and carbon metabolism and are shared with a Brazilian bioethanol production strain, even though the strains differ dramatically at most genetic loci. Lastly, the Y22-3 genome sequence provides an exceptionally high-quality resource for basic and applied research in bioenergy and genetics.« less

Genome sequence and analysis of a stress-tolerant, wild-derived strain of Saccharomyces cerevisiae used in biofuels research

DOE PAGES

McIlwain, Sean J.; Peris, Davis; Sardi, Maria; ...

2016-04-20

The genome sequences of more than 100 strains of the yeast Saccharomyces cerevisiae have been published. Unfortunately, most of these genome assemblies contain dozens to hundreds of gaps at repetitive sequences, including transposable elements, tRNAs, and subtelomeric regions, which is where novel genes generally reside. Relatively few strains have been chosen for genome sequencing based on their biofuel production potential, leaving an additional knowledge gap. Here, we describe the nearly complete genome sequence of GLBRCY22-3 (Y22-3), a strain of S. cerevisiae derived from the stress-tolerant wild strain NRRL YB-210 and subsequently engineered for xylose metabolism. After benchmarking several genome assemblymore » approaches, we developed a pipeline to integrate Pacific Biosciences (PacBio) and Illumina sequencing data and achieved one of the highest quality genome assemblies for any S. cerevisiae strain. Specifically, the contig N50 is 693 kbp, and the sequences of most chromosomes, the mitochondrial genome, and the 2-micron plasmid are complete. Our annotation predicts 92 genes that are not present in the reference genome of the laboratory strain S288c, over 70% of which were expressed. We predicted functions for 43 of these genes, 28 of which were previously uncharacterized and unnamed. Remarkably, many of these genes are predicted to be involved in stress tolerance and carbon metabolism and are shared with a Brazilian bioethanol production strain, even though the strains differ dramatically at most genetic loci. Lastly, the Y22-3 genome sequence provides an exceptionally high-quality resource for basic and applied research in bioenergy and genetics.« less

Brucella Infection in Asian Sea Otters (Enhydra lutris lutris) on Bering Island, Russia.

PubMed

Burgess, Tristan L; Johnson, Christine Kreuder; Burdin, Alexander; Gill, Verena A; Doroff, Angela M; Tuomi, Pamela; Smith, Woutrina A; Goldstein, Tracey

2017-10-01

Infection with Brucella spp., long known as a cause of abortion, infertility, and reproductive loss in domestic livestock, has increasingly been documented in marine mammals over the past two decades. We report molecular evidence of Brucella infection in Asian sea otters (Enhydra lutris lutris). Brucella DNA was detected in 3 of 78 (4%) rectal swab samples collected between 2004 and 2006 on Bering Island, Russia. These 78 animals had previously been documented to have a Brucella seroprevalence of 28%, markedly higher than the prevalence documented in sea otters (Enhydra lutris) in North America. All of the DNA sequences amplified were identical to one or more previously isolated Brucella spp. including strains from both terrestrial and marine hosts. Phylogenetic analysis of this sequence suggested that one animal was shedding Brucella spp. DNA with a sequence matching a Brucella abortus strain, whereas two animals yielded a sequence matching a group of strains including isolates classified as Brucella pinnipedialis and Brucella melitensis. Our results highlight the diversity of Brucella spp. within a single sea otter population.

The genome sequence of four isolates from the family Lichtheimiaceae.

PubMed

Chibucos, Marcus C; Etienne, Kizee A; Orvis, Joshua; Lee, Hongkyu; Daugherty, Sean; Lockhart, Shawn R; Ibrahim, Ashraf S; Bruno, Vincent M

2015-07-01

This study reports the release of draft genome sequences of two isolates of Lichtheimia corymbifera and two isolates of L. ramosa. Phylogenetic analyses indicate that the two L. corymbifera strains (CDC-B2541 and 008-049) are closely related to the previously sequenced L. corymbifera isolate (FSU 9682) while our two L. ramosa strains CDC-B5399 and CDC-B5792 cluster apart from them. These genome sequences will further the understanding of intraspecies and interspecies genetic variation within the Mucoraceae family of pathogenic fungi. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Noncytopathogenic Pestivirus Strains Generated by Nonhomologous RNA Recombination: Alterations in the NS4A/NS4B Coding Region

PubMed Central

Gallei, Andreas; Orlich, Michaela; Thiel, Heinz-Juergen; Becher, Paul

2005-01-01

Several studies have demonstrated that cytopathogenic (cp) pestivirus strains evolve from noncytopathogenic (noncp) viruses by nonhomologous RNA recombination. In addition, two recent reports showed the rapid emergence of noncp Bovine viral diarrhea virus (BVDV) after a few cell culture passages of cp BVDV strains by homologous recombination between identical duplicated viral sequences. To allow the identification of recombination sites from noncp BVDV strains that evolve from cp viruses, we constructed the cp BVDV strains CP442 and CP552. Both harbor duplicated viral sequences of different origin flanking the cellular insertion Nedd8*; the latter is a prerequisite for their cytopathogenicity. In contrast to the previous studies, isolation of noncp strains was possible only after extensive cell culture passages of CP442 and CP552. Sequence analysis of 15 isolated noncp BVDVs confirmed that all recombinant strains lack at least most of Nedd8*. Interestingly, only one strain resulted from homologous recombination while the other 14 strains were generated by nonhomologous recombination. Accordingly, our data suggest that the extent of sequence identity between participating sequences influences both frequency and mode (homologous versus nonhomologous) of RNA recombination in pestiviruses. Further analyses of the noncp recombinant strains revealed that a duplication of 14 codons in the BVDV nonstructural protein 4B (NS4B) gene does not interfere with efficient viral replication. Moreover, an insertion of viral sequences between the NS4A and NS4B genes was well tolerated. These findings thus led to the identification of two genomic loci which appear to be suited for the insertion of heterologous sequences into the genomes of pestiviruses and related viruses. PMID:16254361

Characterization of Bacteroides forsythus Strains from Cat and Dog Bite Wounds in Humans and Comparison with Monkey and Human Oral Strains

PubMed Central

Hudspeth, M. K.; Gerardo, S. Hunt; Maiden, M. F. J.; Citron, D. M.; Goldstein, E. J. C.

1999-01-01

Bacteroides forsythus strains recovered from cat and dog bite wound infections in humans (n = 3), monkey oral strains (n = 3), and the human oral ATCC 43037 type strain were characterized by using phenotypic characteristics, enzymatic tests, whole cell fatty acid analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, PCR fingerprinting, and 16S rDNA (genes coding for rRNA) sequencing. All three bite wound isolates grew on brucella agar supplemented with 5% sheep blood, vitamin K1, and hemin. These strains, unlike the ATCC strain and previously described monkey oral and human clinical strains, did not require N-acetylmuramic acid supplementation for growth as pure cultures. However, their phenotypic characteristics, except for catalase production, were similar to those of previously identified strains. PCR fingerprinting analysis showed differences in band patterns from the ATCC strain. Also, SDS-PAGE and whole cell fatty acid analysis indicated that the dog and cat bite wound strains were similar but not identical to the human B. forsythus ATCC 43037 type strain and the monkey oral strains. The rDNA sequence analysis indicated that the three bite wound isolates had 99.93% homology with each other and 98.9 and 99.22% homology with the human ATCC 43037 and monkey oral strains, respectively. These results suggest that there are host-specific variations within each group. PMID:10325363

Complete genome sequence analysis of novel human bocavirus reveals genetic recombination between human bocavirus 2 and human bocavirus 4.

PubMed

Khamrin, Pattara; Okitsu, Shoko; Ushijima, Hiroshi; Maneekarn, Niwat

2013-07-01

Epidemiological surveillance of human bocavirus (HBoV) was conducted on fecal specimens collected from hospitalized children with diarrhea in Chiang Mai, Thailand in 2011. By partial sequence analysis of VP1 gene, an unusual strain of HBoV (CMH-S011-11), was initially identified as HBoV4. The complete genome sequence of CMH-S011-11 was performed and analyzed further to clarify whether it was a recombinant strain or a new HBoV variant. Analysis of complete genome sequence revealed that the coding sequence starting from NS1, NP1 to VP1/VP2 was 4795 nucleotides long. Interestingly, the nucleotide sequence of NS1 gene of CMH-S011-11 was most closely related to the HBoV2 reference strains detected in Pakistan, which contradicted to the initial genotyping result of the partial VP1 region in the previous study. In addition, comparison of NP1 nucleotide sequence of CMH-S011-11 with those of other HBoV1-4 reference strains also revealed a high level of sequence identity with HBoV2. On the other hand, nucleotide sequence of VP1/VP2 gene of CMH-S011-11 was most closely related to those of HBoV4 reference strains detected in Nigeria. The overall full-length sequence analysis revealed that this CMH-S011-11 was grouped within HBoV4 species, but located in a separate branch from other HBoV4 prototype strains. Recombination analysis revealed that CMH-S011-11 was the result of recombination between HBoV2 and HBoV4 strains with the break point located near the start codon of VP2. Copyright © 2013 Elsevier B.V. All rights reserved.

Rhizobium favelukesii sp. nov., isolated from the root nodules of alfalfa (Medicago sativa L).

PubMed

Torres Tejerizo, Gonzalo; Rogel, Marco Antonio; Ormeño-Orrillo, Ernesto; Althabegoiti, María Julia; Nilsson, Juliet Fernanda; Niehaus, Karsten; Schlüter, Andreas; Pühler, Alfred; Del Papa, María Florencia; Lagares, Antonio; Martínez-Romero, Esperanza; Pistorio, Mariano

2016-11-01

Strains LPU83T and Or191 of the genus Rhizobium were isolated from the root nodules of alfalfa, grown in acid soils from Argentina and the USA. These two strains, which shared the same plasmid pattern, lipopolysaccharide profile, insertion-sequence fingerprint, 16S rRNA gene sequence and PCR-fingerprinting pattern, were different from reference strains representing species of the genus Rhizobium with validly published names. On the basis of previously reported data and from new DNA-DNA hybridization results, phenotypic characterization and phylogenetic analyses, strains LPU83T and Or191 can be considered to be representatives of a novel species of the genus Rhizobium, for which the name Rhizobium favelukesii sp. nov. is proposed. The type strain of this species is LPU83T (=CECT 9014T=LMG 29160T), for which an improved draft-genome sequence is available.

Native valve endocarditis due to a novel strain of Legionella.

PubMed

Pearce, Meghan M; Theodoropoulos, Nicole; Noskin, Gary A; Flaherty, John P; Stemper, Mary E; Aspeslet, Teresa; Cianciotto, Nicholas P; Reed, Kurt D

2011-09-01

Legionellae are Gram-negative bacteria which are capable of causing disease, most commonly in the form of pneumonia. We describe a case of native valve endocarditis caused by a Legionella strain which by genotypic (16S rRNA and mip gene sequencing) and phenotypic analyses is unlike previously described strains of Legionella.

Strain/species identification in metagenomes using genome-specific markers

PubMed Central

Tu, Qichao; He, Zhili; Zhou, Jizhong

2014-01-01

Shotgun metagenome sequencing has become a fast, cheap and high-throughput technology for characterizing microbial communities in complex environments and human body sites. However, accurate identification of microorganisms at the strain/species level remains extremely challenging. We present a novel k-mer-based approach, termed GSMer, that identifies genome-specific markers (GSMs) from currently sequenced microbial genomes, which were then used for strain/species-level identification in metagenomes. Using 5390 sequenced microbial genomes, 8 770 321 50-mer strain-specific and 11 736 360 species-specific GSMs were identified for 4088 strains and 2005 species (4933 strains), respectively. The GSMs were first evaluated against mock community metagenomes, recently sequenced genomes and real metagenomes from different body sites, suggesting that the identified GSMs were specific to their targeting genomes. Sensitivity evaluation against synthetic metagenomes with different coverage suggested that 50 GSMs per strain were sufficient to identify most microbial strains with ≥0.25× coverage, and 10% of selected GSMs in a database should be detected for confident positive callings. Application of GSMs identified 45 and 74 microbial strains/species significantly associated with type 2 diabetes patients and obese/lean individuals from corresponding gastrointestinal tract metagenomes, respectively. Our result agreed with previous studies but provided strain-level information. The approach can be directly applied to identify microbial strains/species from raw metagenomes, without the effort of complex data pre-processing. PMID:24523352

Investigating the Molecular Mechanisms of Organophosphate and Pyrethroid Resistance in the Fall Armyworm Spodoptera frugiperda

PubMed Central

Carvalho, Renato A.; Omoto, Celso; Field, Linda M.; Williamson, Martin S.; Bass, Chris

2013-01-01

The fall armyworm Spodoptera frugiperda is an economically important pest of small grain crops that occurs in all maize growing regions of the Americas. The intensive use of chemical pesticides for its control has led to the selection of resistant populations, however, to date, the molecular mechanisms underlying resistance have not been characterised. In this study the mechanisms involved in the resistance of two S. frugiperda strains collected in Brazil to chlorpyrifos (OP strain) or lambda-cyhalothrin (PYR strain) were investigated using molecular and genomic approaches. To examine the possible role of target-site insensitivity the genes encoding the organophosphate (acetylcholinesterase, AChE) and pyrethroid (voltage-gated sodium channel, VGSC) target-site proteins were PCR amplified. Sequencing of the S. frugiperda ace-1 gene identified several nucleotide changes in the OP strain when compared to a susceptible reference strain (SUS). These result in three amino acid substitutions, A201S, G227A and F290V, that have all been shown previously to confer organophosphate resistance in several other insect species. Sequencing of the gene encoding the VGSC in the PYR strain, identified mutations that result in three amino acid substitutions, T929I, L932F and L1014F, all of which have been shown previously to confer knockdown/super knockdown-type resistance in several arthropod species. To investigate the possible role of metabolic detoxification in the resistant phenotype of the OP and PYR stains all EST sequences available for S. frugiperda were used to design a gene-expression microarray. This was then used to compare gene expression in the resistant strains with the susceptible reference strain. Members of several gene families, previously implicated in metabolic resistance in other insects were found to be overexpressed in the resistant strains including glutathione S-transferases, cytochrome P450s and carboxylesterases. Taken together these results provide evidence that both target-site and metabolic mechanisms underlie the resistance of S. frugiperda to pyrethroids and organophosphates. PMID:23614047

Investigating the molecular mechanisms of organophosphate and pyrethroid resistance in the fall armyworm Spodoptera frugiperda.

PubMed

Carvalho, Renato A; Omoto, Celso; Field, Linda M; Williamson, Martin S; Bass, Chris

2013-01-01

The fall armyworm Spodoptera frugiperda is an economically important pest of small grain crops that occurs in all maize growing regions of the Americas. The intensive use of chemical pesticides for its control has led to the selection of resistant populations, however, to date, the molecular mechanisms underlying resistance have not been characterised. In this study the mechanisms involved in the resistance of two S. frugiperda strains collected in Brazil to chlorpyrifos (OP strain) or lambda-cyhalothrin (PYR strain) were investigated using molecular and genomic approaches. To examine the possible role of target-site insensitivity the genes encoding the organophosphate (acetylcholinesterase, AChE) and pyrethroid (voltage-gated sodium channel, VGSC) target-site proteins were PCR amplified. Sequencing of the S. frugiperda ace-1 gene identified several nucleotide changes in the OP strain when compared to a susceptible reference strain (SUS). These result in three amino acid substitutions, A201S, G227A and F290V, that have all been shown previously to confer organophosphate resistance in several other insect species. Sequencing of the gene encoding the VGSC in the PYR strain, identified mutations that result in three amino acid substitutions, T929I, L932F and L1014F, all of which have been shown previously to confer knockdown/super knockdown-type resistance in several arthropod species. To investigate the possible role of metabolic detoxification in the resistant phenotype of the OP and PYR stains all EST sequences available for S. frugiperda were used to design a gene-expression microarray. This was then used to compare gene expression in the resistant strains with the susceptible reference strain. Members of several gene families, previously implicated in metabolic resistance in other insects were found to be overexpressed in the resistant strains including glutathione S-transferases, cytochrome P450s and carboxylesterases. Taken together these results provide evidence that both target-site and metabolic mechanisms underlie the resistance of S. frugiperda to pyrethroids and organophosphates.

Nucleotide Sequences of Genes Coding for Fimbrial Proteins in a Cryptic Genospecies of Haemophilus spp. Isolated from Neonatal and Genital Tract Infections

PubMed Central

Gousset, Nathalie; Rosenau, Agnes; Sizaret, Pierre-Yves; Quentin, Roland

1999-01-01

Nineteen isolates belonging to a cryptic genospecies of Haemophilus (referred to here as genital strains) isolated from genital tract infections (6 strains) and from neonatal infections (13 strains) were studied for fimbrial genes. Sixteen strains exhibit peritrichous fimbriae observed by electron microscopy. By PCR with primers corresponding to the extreme ends of the Haemophilus influenzae type b (Hib) hifA and hifD genes and Southern blotting, a hifA-like gene (named ghfA) and a hifD-like gene (named ghfD) were identified in 6 of the 19 strains. Five of these six strains were from the genital tracts of adults, and one was from a neonate. For each gene, the nucleotide sequence was identical for the six strains. A hifE-like gene (named ghfE) was amplified from only one of the 19 genital strains of Haemophilus, but the ghfE probe gave a signal in Southern hybridization with the five other strains positive for ghfA and ghfD. Therefore, these strains may carry a ghfE-like gene. The Hib fimbrial gene cluster is located between the purE and pepN genes as previously described. For the 13 genital Haemophilus strains that lack fimbrial genes, this region corresponds to a noncoding sequence. Another major fimbrial gene designated the fimbrin gene was previously identified in a nontypeable H. influenzae strain. A fimbrin-like gene was identified for all of our 19 genital strains. This gene is similar to the ompP5 gene of many Haemophilus strains. Therefore, other, unidentified genes may explain the piliation observed in electron microscopy on genital Haemophilus strains which do not possess LKP-like fimbrial genes. Fimbrial genes were significantly associated with strains isolated from the genital tract. They may confer on the strain the ability to survive in the genital tract. PMID:9864189

Bradyrhizobium sacchari sp. nov., a legume nodulating bacterium isolated from sugarcane roots.

PubMed

de Matos, Gustavo Feitosa; Zilli, Jerri Edson; de Araújo, Jean Luiz Simões; Parma, Marcia Maria; Melo, Itamar Soares; Radl, Viviane; Baldani, José Ivo; Rouws, Luc Felicianus Marie

2017-11-01

Members of the genus Bradyrhizobium are well-known as nitrogen-fixing microsymbionts of a wide variety of leguminous species, but they have also been found in different environments, notably as endophytes in non-legumes such as sugarcane. This study presents a detailed polyphasic characterization of four Bradyrhizobium strains (type strain BR 10280 T ), previously isolated from roots of sugarcane in Brazil. 16S rRNA sequence analysis, multilocus sequence analysis (MLSA) and analysis of the 16S-23S rRNA internal transcribed spacer showed that these strains form a novel clade close to, but different from B. huanghuaihaiense strain CCBAU 23303 T . Average nucleotide identity (ANI) analyses confirmed that BR 10280 T represents a novel species. Phylogenetic analysis based on nodC gene sequences also placed the strains close to CCBAU 23303 T , but different from this latter strain, the sugarcane strains did not nodulate soybean, although they effectively nodulated Vigna unguiculata, Cajanus cajan and Macroptilium atropurpureum. Physiological traits are in agreement with the placement of the strains in the genus Bradyrhizobium as a novel species for which the name Bradyrhizobium sacchari sp. nov. is proposed.

Isolation and sequence characterization of DNA-A genome of a new begomovirus strain associated with severe leaf curling symptoms of Jatropha curcas L.

PubMed

Chauhan, Sushma; Rahman, Hifzur; Mastan, Shaik G; Pamidimarri, D V N Sudheer; Reddy, Muppala P

2018-07-20

Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, such as mosaic and leaf curling in Jatropha curcas. The molecular characterization of these viral strains will help in developing management strategies to control the disease. In this study, J. curcas that was infected with begomovirus and showed acute leaf curling symptoms were identified. DNA-A segment from pathogenic viral strain was isolated and sequenced. The sequenced genome was assembled and characterized in detail. The full-length DNA-A sequence was covered by primer walking. The genome sequence showed the general organization of DNA-A from begomovirus by the distribution of ORFs in both viral and anti-viral strands. The genome size ranged from 2844 bp-2852 bp. Three strains with minor nucleotide variations were identified, and a phylogenetic analysis was performed by comparing the DNA-A segments from other reported begomovirus isolates. The maximum sequence similarity was observed with Euphorbia yellow mosaic virus (FN435995). In the phylogenetic tree, no clustering was observed with previously reported begomovirus strains isolated from J. curcas host. The strains isolated in this study belong to new begomoviral strain that elicits symptoms of leaf curling in J. curcas. The results indicate that the probable origin of the strains is from Jatropha mosaic virus infecting J. gassypifolia. The strains isolated in this study are referred as Jatropha curcas leaf curl India virus (JCLCIV) based on the major symptoms exhibited by host J. curcas. Copyright © 2018 Elsevier B.V. All rights reserved.

Serovar distribution of a DNA sequence involved in the antigenic relationship between Leptospira and equine cornea

PubMed Central

Lucchesi, Paula MA; Parma, Alberto E; Arroyo, Guillermo H

2002-01-01

Background Horses infected with Leptospira present several clinical disorders, one of them being recurrent uveitis. A common endpoint of equine recurrent uveitis is blindness. Serovar pomona has often been incriminated, although others have also been reported. An antigenic relationship between this bacterium and equine cornea has been described in previous studies. A leptospiral DNA fragment that encodes cross-reacting epitopes was previously cloned and expressed in Escherichia coli. Results A region of that DNA fragment was subcloned and sequenced. Samples of leptospiral DNA from several sources were analysed by PCR with two primer pairs designed to amplify that region. Reference strains from serovars canicola, icterohaemorrhagiae, pomona, pyrogenes, wolffi, bataviae, sentot, hebdomadis and hardjo rendered products of the expected sizes with both pairs of primers. The specific DNA region was also amplified from isolates from Argentina belonging to serogroups Canicola and Pomona. Both L. biflexa serovar patoc and L. borgpetersenii serovar tarassovi rendered a negative result. Conclusions The DNA sequence related to the antigen mimicry with equine cornea was not exclusively found in serovar pomona as it was also detected in several strains of Leptospira belonging to different serovars. The results obtained with L. biflexa serovar patoc strain Patoc I and L. borgpetersenii serovar tarassovi strain Perepelicin suggest that this sequence is not present in these strains, which belong to different genomospecies than those which gave positive results. This is an interesting finding since L. biflexa comprises nonpathogenic strains and serovar tarassovi has not been associated clinically with equine uveitis. PMID:11869455

Genotyping of ancient Mycobacterium tuberculosis strains reveals historic genetic diversity.

PubMed

Müller, Romy; Roberts, Charlotte A; Brown, Terence A

2014-04-22

The evolutionary history of the Mycobacterium tuberculosis complex (MTBC) has previously been studied by analysis of sequence diversity in extant strains, but not addressed by direct examination of strain genotypes in archaeological remains. Here, we use ancient DNA sequencing to type 11 single nucleotide polymorphisms and two large sequence polymorphisms in the MTBC strains present in 10 archaeological samples from skeletons from Britain and Europe dating to the second-nineteenth centuries AD. The results enable us to assign the strains to groupings and lineages recognized in the extant MTBC. We show that at least during the eighteenth-nineteenth centuries AD, strains of M. tuberculosis belonging to different genetic groups were present in Britain at the same time, possibly even at a single location, and we present evidence for a mixed infection in at least one individual. Our study shows that ancient DNA typing applied to multiple samples can provide sufficiently detailed information to contribute to both archaeological and evolutionary knowledge of the history of tuberculosis.

Complete genome sequence of bacteriocin-producing Lactobacillus plantarum KLDS1.0391, a probiotic strain with gastrointestinal tract resistance and adhesion to the intestinal epithelial cells.

PubMed

Jia, Fang-Fang; Zhang, Lu-Ji; Pang, Xue-Hui; Gu, Xin-Xi; Abdelazez, Amro; Liang, Yu; Sun, Si-Rui; Meng, Xiang-Chen

2017-10-01

Lactobacillus plantarum KLDS1.0391 is a probiotic strain isolated from the traditional fermented dairy products and identified to produce bacteriocin against Gram-positive and Gram-negative bacteria. Previous studies showed that the strain has a high resistance to gastrointestinal stress and has a high adhesion ability to the intestinal epithelial cells (Caco-2). We reported the entire genome sequence of this strain, which contains a circular 2,886,607-bp chromosome and three circular plasmids. Genes, which are related to the biosynthesis of bacteriocins, the stress resistance to gastrointestinal tract environment and adhesive performance, were identified. Whole genome sequence of Lactobacillus plantarum KLDS1.0391 will be helpful for its applications in food industry. Copyright © 2017 Elsevier Inc. All rights reserved.

Phylogenetic relationship of Ornithobacterium rhinotracheale strains.

PubMed

DE Oca-Jimenez, Roberto Montes; Vega-Sanchez, Vicente; Morales-Erasto, Vladimir; Salgado-Miranda, Celene; Blackall, Patrick J; Soriano-Vargas, Edgardo

2018-04-10

The bacterium Ornithobacterium rhinotracheale is associated with respiratory disease in wild birds and poultry. In this study, the phylogenetic analysis of nine reference strains of O. rhinotracheale belonging to serovars A to I, and eight Mexican isolates belonging to serovar A, was performed. The analysis was extended to include available sequences from another 23 strains available in the public domain. The analysis showed that the 40 sequences formed six clusters, I to VI. All eight Mexican field isolates were placed in cluster I. One of the reference strains appears to present genetic diversity not previously recognized and was placed in a new genetic cluster. In conclusion, the phylogenetic analysis of O. rhinotracheale strains, based on the 16S rRNA gene, is a suitable tool for epidemiologic studies.

Native Valve Endocarditis Due to a Novel Strain of Legionella ▿

PubMed Central

Pearce, Meghan M.; Theodoropoulos, Nicole; Noskin, Gary A.; Flaherty, John P.; Stemper, Mary E.; Aspeslet, Teresa; Cianciotto, Nicholas P.; Reed, Kurt D.

2011-01-01

Legionellae are Gram-negative bacteria which are capable of causing disease, most commonly in the form of pneumonia. We describe a case of native valve endocarditis caused by a Legionella strain which by genotypic (16S rRNA and mip gene sequencing) and phenotypic analyses is unlike previously described strains of Legionella. PMID:21795511

Resequencing Pathogen Microarray (RPM) for prospective detection and identification of emergent pathogen strains and variants

NASA Astrophysics Data System (ADS)

Tibbetts, Clark; Lichanska, Agnieszka M.; Borsuk, Lisa A.; Weslowski, Brian; Morris, Leah M.; Lorence, Matthew C.; Schafer, Klaus O.; Campos, Joseph; Sene, Mohamadou; Myers, Christopher A.; Faix, Dennis; Blair, Patrick J.; Brown, Jason; Metzgar, David

2010-04-01

High-density resequencing microarrays support simultaneous detection and identification of multiple viral and bacterial pathogens. Because detection and identification using RPM is based upon multiple specimen-specific target pathogen gene sequences generated in the individual test, the test results enable both a differential diagnostic analysis and epidemiological tracking of detected pathogen strains and variants from one specimen to the next. The RPM assay enables detection and identification of pathogen sequences that share as little as 80% sequence similarity to prototype target gene sequences represented as detector tiles on the array. This capability enables the RPM to detect and identify previously unknown strains and variants of a detected pathogen, as in sentinel cases associated with an infectious disease outbreak. We illustrate this capability using assay results from testing influenza A virus vaccines configured with strains that were first defined years after the design of the RPM microarray. Results are also presented from RPM-Flu testing of three specimens independently confirmed to the positive for the 2009 Novel H1N1 outbreak strain of influenza virus.

Purification and sequence analysis of 4-methyl-5-nitrocatechol oxygenase from Burkholderia sp. strain DNT.

PubMed Central

Haigler, B E; Suen, W C; Spain, J C

1996-01-01

4-Methyl-5-nitrocatechol (MNC) is an intermediate in the degradation of 2,4-dinitrotoluene by Burkholderia sp. strain DNT. In the presence of NADPH and oxygen, MNC monooxygenase catalyzes the removal of the nitro group from MNC to form 2-hydroxy-5-methylquinone. The gene (dntB) encoding MNC monooxygenase has been previously cloned and characterized. In order to examine the properties of MNC monooxygenase and to compare it with other enzymes, we sequenced the gene encoding the MNC monooxygenase and purified the enzyme from strain DNT. dntB was localized within a 2.2-kb ApaI DNA fragment. Sequence analysis of this fragment revealed an open reading frame of 1,644 bp with an N-terminal amino acid sequence identical to that of purified MNC monooxygenase from strain DNT. Comparison of the derived amino acid sequences with those of other genes showed that DntB contains the highly conserved ADP and flavin adenine dinucleotide (FAD) binding motifs characteristic of flavoprotein hydroxylases. MNC monooxygenase was purified to homogeneity from strain DNT by anion exchange and gel filtration chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single protein with a molecular weight of 60,200, which is consistent with the size determined from the gene sequence. The native molecular weight determined by gel filtration was 65,000, which indicates that the native enzyme is a monomer. It used either NADH or NADPH as electron donors, and NADPH was the preferred cofactor. The purified enzyme contained 1 mol of FAD per mol of protein, which is also consistent with the detection of an FAD binding motif in the amino acid sequence of DntB. MNC monooxygenase has a narrow substrate specificity. MNC and 4-nitrocatechol are good substrates whereas 3-methyl-4-nitrophenol, 3-methyl-4-nitrocatechol, 4-nitrophenol, 3-nitrophenol, and 4-chlorocatechol were not. These studies suggest that MNC monooxygenase is a flavoprotein that shares some properties with previously studied nitrophenol oxygenases. PMID:8830701

Genome sequence of Prevotella intermedia SUNY aB G8-9K-3, a biofilm forming strain with drug-resistance.

PubMed

Moon, Ji-Hoi; Kim, Minjung; Lee, Jae-Hyung

Prevotella intermedia has long been known to be as the principal etiologic agent of periodontal diseases and associated with various systemic diseases. Previous studies showed that the intra-species difference exists in capacity of biofilm formation, antibiotic resistance, and serological reaction among P. intermedia strains. Here we report the genome sequence of P. intermedia SUNY aB G8-9K-3 (designated ATCC49046) that displays a relatively high antimicrobial resistant and biofilm-forming capacity. Genome sequencing information provides important clues in understanding the genetic bases of phenotypic differences among P. intermedia strains. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Sequence variations in RepMP2/3 and RepMP4 elements reveal intragenomic homologous DNA recombination events in Mycoplasma pneumoniae.

PubMed

Spuesens, Emiel B M; Oduber, Minoushka; Hoogenboezem, Theo; Sluijter, Marcel; Hartwig, Nico G; van Rossum, Annemarie M C; Vink, Cornelis

2009-07-01

The gene encoding major adhesin protein P1 of Mycoplasma pneumoniae, MPN141, contains two DNA sequence stretches, designated RepMP2/3 and RepMP4, which display variation among strains. This variation allows strains to be differentiated into two major P1 genotypes (1 and 2) and several variants. Interestingly, multiple versions of the RepMP2/3 and RepMP4 elements exist at other sites within the bacterial genome. Because these versions are closely related in sequence, but not identical, it has been hypothesized that they have the capacity to recombine with their counterparts within MPN141, and thereby serve as a source of sequence variation of the P1 protein. In order to determine the variation within the RepMP2/3 and RepMP4 elements, both within the bacterial genome and among strains, we analysed the DNA sequences of all RepMP2/3 and RepMP4 elements within the genomes of 23 M. pneumoniae strains. Our data demonstrate that: (i) recombination is likely to have occurred between two RepMP2/3 elements in four of the strains, and (ii) all previously described P1 genotypes can be explained by inter-RepMP recombination events. Moreover, the difference between the two major P1 genotypes was reflected in all RepMP elements, such that subtype 1 and 2 strains can be differentiated on the basis of sequence variation in each RepMP element. This implies that subtype 1 and subtype 2 strains represent evolutionarily diverged strain lineages. Finally, a classification scheme is proposed in which the P1 genotype of M. pneumoniae isolates can be described in a sequence-based, universal fashion.

Veillonella infantium sp. nov., an anaerobic, Gram-stain-negative coccus isolated from tongue biofilm of a Thai child.

PubMed

Mashima, Izumi; Liao, Yu-Chieh; Miyakawa, Hiroshi; Theodorea, Citra F; Thawboon, Boonyanit; Thaweboon, Sroisiri; Scannapieco, Frank A; Nakazawa, Futoshi

2018-04-01

A strain of a novel anaerobic, Gram-stain-negative coccus was isolated from the tongue biofilm of a Thai child. This strain was shown, at the phenotypic level and based on 16S rRNA gene sequencing, to be a member of the genus Veillonella. Comparative analysis of the 16S rRNA, dnaK and rpoB gene sequences indicated that phylogenetically the strain comprised a distinct novel branch within the genus Veillonella. The novel strain showed 99.8, 95.1 and 95.9 % similarity to partial 16S rRNA, dnaK and rpoB gene sequences, respectively, to the type strains of the two most closely related species, Veillonelladispar ATCC 17748 T and Veillonellatobetsuensis ATCC BAA-2400 T . The novel strain could be discriminated from previously reported species of the genus Veillonella based on partial dnaK and rpoB gene sequencing and average nucleotide identity values. The major acid end-product produced by this strain was acetic acid under anaerobic conditions in trypticase-yeast extract-haemin with 1 % (w/v) glucose or fructose medium. Lactate was fermented to acetic acid and propionic acid. Based on these observations, this strain represents a novel species, for which the name Veillonella infantium sp. nov. is proposed. The type strain is T11011-4 T (=JCM 31738 T =TSD-88 T ).

Improved High-Quality Draft Genome Sequence and Annotation of Burkholderia contaminans LMG 23361T.

PubMed

Jung, Ji Young; Ahn, Youngbeom; Kweon, Ohgew; LiPuma, John J; Hussong, David; Marasa, Bernard S; Cerniglia, Carl E

2017-04-20

Burkholderia contaminans LMG 23361 is the type strain of the species isolated from the milk of a dairy sheep with mastitis. Some pharmaceutical products contain disinfectants such as benzalkonium chloride (BZK) and previously we reported that B. contaminans LMG 23361 T possesses the ability to inactivate BZK with high biodegradation rates. Here, we report an improved high-quality draft genome sequence of this strain. Copyright © 2017 Jung et al.

Identification of three homologous latex-clearing protein (lcp) genes from the genome of Streptomyces sp. strain CFMR 7.

PubMed

Nanthini, Jayaram; Ong, Su Yean; Sudesh, Kumar

2017-09-10

Rubber materials have greatly contributed to human civilization. However, being a polymeric material does not decompose easily, it has caused huge environmental problems. On the other hand, only few bacteria are known to degrade rubber, with studies pertaining them being intensively focusing on the mechanism involved in microbial rubber degradation. The Streptomyces sp. strain CFMR 7, which was previously confirmed to possess rubber-degrading ability, was subjected to whole genome sequencing using the single molecule sequencing technology of the PacBio® RS II system. The genome was further analyzed and compared with previously reported rubber-degrading bacteria in order to identify the potential genes involved in rubber degradation. This led to the interesting discovery of three homologues of latex-clearing protein (Lcp) on the chromosome of this strain, which are probably responsible for rubber degrading activities. Genes encoding oxidoreductase α-subunit (oxiA) and oxidoreductase β-subunit (oxiB) were also found downstream of two lcp genes which are located adjacent to each other. In silico analysis reveals genes that have been identified to be involved in the microbial degradation of rubber in the Streptomyces sp. strain CFMR 7. This is the first whole genome sequence of a clear-zone-forming natural rubber- degrading Streptomyces sp., which harbours three Lcp homologous genes with the presence of oxiA and oxiB genes compared to the previously reported Gordonia polyisoprenivorans strain VH2 (with two Lcp homologous genes) and Nocardia nova SH22a (with only one Lcp gene). Copyright © 2017 Elsevier B.V. All rights reserved.

Phylogeny of the Defined Murine Microbiota: Altered Schaedler Flora

PubMed Central

Dewhirst, Floyd E.; Chien, Chih-Ching; Paster, Bruce J.; Ericson, Rebecca L.; Orcutt, Roger P.; Schauer, David B.; Fox, James G.

1999-01-01

The “altered Schaedler flora” (ASF) was developed for colonizing germfree rodents with a standardized microbiota. The purpose of this study was to identify each of the eight ASF strains by 16S rRNA sequence analysis. Three strains were previously identified as Lactobacillus acidophilus (strain ASF 360), Lactobacillus salivarius (strain ASF 361), and Bacteroides distasonis (strain ASF 519) based on phenotypic criteria. 16S rRNA analysis indicated that each of the strains differed from its presumptive identity. The 16S rRNA sequence of strain ASF 361 is essentially identical to the 16S rRNA sequences of the type strains of Lactobacillus murinis and Lactobacillus animalis (both isolated from mice), and all of these strains probably belong to a single species. Strain ASF 360 is a novel lactobacillus that clusters with L. acidophilus and Lactobacillus lactis. Strain ASF 519 falls into an unnamed genus containing [Bacteroides] distasonis, [Bacteroides] merdae, [Bacteroides] forsythus, and CDC group DF-3. This unnamed genus is in the Cytophaga-Flavobacterium-Bacteroides phylum and is most closely related to the genus Porphyromonas. The spiral-shaped strain, strain ASF 457, is in the Flexistipes phylum and exhibits sequence identity with rodent isolates of Robertson. The remaining four ASF strains, which are extremely oxygen-sensitive fusiform bacteria, group phylogenetically with the low-G+C-content gram-positive bacteria (Firmicutes, Bacillus-Clostridium group). ASF 356, ASF 492, and ASF 502 fall into Clostridium cluster XIV of Collins et al. Morphologically, ASF 492 resembles members of this cluster, Roseburia cecicola, and Eubacterium plexicaudatum. The 16S rRNA sequence of ASF 492 is identical to that of E. plexicaudatum. Since the type strain and other viable original isolates of E. plexicaudatum have been lost, strain ASF 492 is a candidate for a neotype strain. Strain ASF 500 branches deeply in the low-G+C-content gram-positive phylogenetic tree but is not closely related to any organisms whose 16S rRNA sequences are currently in the GenBank database. The 16S rRNA sequence information determined in the present study should allow rapid identification of ASF strains and should permit detailed analysis of the interactions of ASF organisms during development of intestinal disease in mice that are coinfected with a variety of pathogenic microorganisms. PMID:10427008

Complete Genomic Sequence and Comparative Analysis of the Genome Segments of Sweet Potato Chlorotic Stunt Virus in China

PubMed Central

Qin, Yanhong; Wang, Li; Zhang, Zhenchen; Qiao, Qi; Zhang, Desheng; Tian, Yuting; Wang, Shuang; Wang, Yongjiang; Yan, Zhaoling

2014-01-01

Background Sweet potato chlorotic stunt virus (family Closteroviridae, genus Crinivirus) features a large bipartite, single-stranded, positive-sense RNA genome. To date, only three complete genomic sequences of SPCSV can be accessed through GenBank. SPCSV was first detected from China in 2011, only partial genomic sequences have been determined in the country. No report on the complete genomic sequence and genome structure of Chinese SPCSV isolates or the genetic relation between isolates from China and other countries is available. Methodology/Principal Findings The complete genomic sequences of five isolates from different areas in China were characterized. This study is the first to report the complete genome sequences of SPCSV from whitefly vectors. Genome structure analysis showed that isolates of WA and EA strains from China have the same coding protein as isolates Can181-9 and m2-47, respectively. Twenty cp genes and four RNA1 partial segments were sequenced and analyzed, and the nucleotide identities of complete genomic, cp, and RNA1 partial sequences were determined. Results indicated high conservation among strains and significant differences between WA and EA strains. Genetic analysis demonstrated that, except for isolates from Guangdong Province, SPCSVs from other areas belong to the WA strain. Genome organization analysis showed that the isolates in this study lack the p22 gene. Conclusions/Significance We presented the complete genome sequences of SPCSV in China. Comparison of nucleotide identities and genome structures between these isolates and previously reported isolates showed slight differences. The nucleotide identities of different SPCSV isolates showed high conservation among strains and significant differences between strains. All nine isolates in this study lacked p22 gene. WA strains were more extensively distributed than EA strains in China. These data provide important insights into the molecular variation and genomic structure of SPCSV in China as well as genetic relationships among isolates from China and other countries. PMID:25170926

Stochastic precision analysis of 2D cardiac strain estimation in vivo

NASA Astrophysics Data System (ADS)

Bunting, E. A.; Provost, J.; Konofagou, E. E.

2014-11-01

Ultrasonic strain imaging has been applied to echocardiography and carries great potential to be used as a tool in the clinical setting. Two-dimensional (2D) strain estimation may be useful when studying the heart due to the complex, 3D deformation of the cardiac tissue. Increasing the framerate used for motion estimation, i.e. motion estimation rate (MER), has been shown to improve the precision of the strain estimation, although maintaining the spatial resolution necessary to view the entire heart structure in a single heartbeat remains challenging at high MERs. Two previously developed methods, the temporally unequispaced acquisition sequence (TUAS) and the diverging beam sequence (DBS), have been used in the past to successfully estimate in vivo axial strain at high MERs without compromising spatial resolution. In this study, a stochastic assessment of 2D strain estimation precision is performed in vivo for both sequences at varying MERs (65, 272, 544, 815 Hz for TUAS; 250, 500, 1000, 2000 Hz for DBS). 2D incremental strains were estimated during left ventricular contraction in five healthy volunteers using a normalized cross-correlation function and a least-squares strain estimator. Both sequences were shown capable of estimating 2D incremental strains in vivo. The conditional expected value of the elastographic signal-to-noise ratio (E(SNRe|ɛ)) was used to compare strain estimation precision of both sequences at multiple MERs over a wide range of clinical strain values. The results here indicate that axial strain estimation precision is much more dependent on MER than lateral strain estimation, while lateral estimation is more affected by strain magnitude. MER should be increased at least above 544 Hz to avoid suboptimal axial strain estimation. Radial and circumferential strain estimations were influenced by the axial and lateral strain in different ways. Furthermore, the TUAS and DBS were found to be of comparable precision at similar MERs.

Phylogeny of the ammonia-producing ruminal bacteria Peptostreptococcus anaerobius, Clostridium sticklandii, and Clostridium aminophilum sp. nov

NASA Technical Reports Server (NTRS)

Paster, B. J.; Russell, J. B.; Yang, C. M.; Chow, J. M.; Woese, C. R.; Tanner, R.

1993-01-01

In previous studies, gram-positive bacteria which grew rapidly with peptides or an amino acid as the sole energy source were isolated from bovine rumina. Three isolates, strains C, FT (T = type strain), and SR, were considered to be ecologically important since they produced up to 20-fold more ammonia than other ammonia-producing ruminal bacteria. On the basis of phenotypic criteria, the taxonomic position of these new isolates was uncertain. In this study, the 16S rRNA sequences of these isolates and related bacteria were determined to establish the phylogenetic positions of the organisms. The sequences of strains C, FT, and SR and reference strains of Peptostreptococcus anaerobius, Clostridium sticklandii, Clostridium coccoides, Clostridium aminovalericum, Acetomaculum ruminis, Clostridium leptum, Clostridium lituseburense, Clostridium acidiurici, and Clostridium barkeri were determined by using a modified Sanger dideoxy chain termination method. Strain C, a large coccus purported to belong to the genus Peptostreptococcus, was closely related to P. anaerobius, with a level of sequence similarity of 99.6%. Strain SR, a heat-resistant, short, rod-shaped organism, was closely related to C. sticklandii, with a level of sequence similarity of 99.9%. However, strain FT, a heat-resistant, pleomorphic, rod-shaped organism, was only distantly related to some clostridial species and P. anaerobius. On the basis of the sequence data, it was clear that strain FT warranted designation as a separate species. The closest known relative of strain FT was C. coccoides (level of similarity, only 90.6%). Additional strains that are phenotypically similar to strain FT were isolated in this study.(ABSTRACT TRUNCATED AT 250 WORDS).

ABCG-like transporter of Trypanosoma cruzi involved in benznidazole resistance: gene polymorphisms disclose inter-strain intragenic recombination in hybrid isolates.

PubMed

Franco, Jaques; Ferreira, Renata C; Ienne, Susan; Zingales, Bianca

2015-04-01

Benznidazole (BZ) is one of the two drugs for Chagas disease treatment. In a previous study we showed that the Trypanosoma cruzi ABCG-like transporter gene, named TcABCG1, is over-expressed in parasite strains naturally resistant to BZ and that the gene of TcI BZ-resistant strains exhibited several single nucleotide polymorphisms (SNPs) as compared to the gene of CL Brener BZ-susceptible strain. Here we report the sequence of TcABCG1 gene of fourteen T. cruzi strains, with diverse degrees of BZ sensitivity and belonging to different discrete typing units (DTUs) and Tcbat group. Although DTU-specific SNPs and amino acid changes were identified, no direct correlation with BZ-resistance phenotype was found. Thus, it is plausible that the transporter abundance is a determinant factor for drug resistance, as pointed out above. Sequence data were used for Bayesian phylogenies and network genealogy analysis. The network showed a high degree of reticulation suggesting genetic exchange between the parasites. TcI and TcII clades were clearly separated. Tcbat sequences were close to TcI. A fourth clade clustered TcABCG1 haplotypes of TcV, TcVI and TcIII strains, with closer proximity to TcI. Analysis of the recombination patterns indicated that hybrid strains contain haplotypes that are mosaics most likely derived by intragenic recombination of parental sequences. The data confirm that TcII and TcIII as the parentals of TcV and TcVI DTUs. Since genetic fingerprint of TcI was found in TcIII, we sustain the previously proposed "Two Hybridization model" for the origin of hybrid strains. Among the twenty best BLASTP hits in databases, orthologues of TcABCG1 transporter were found in Leishmania spp. and African trypanosomes, though their function remains undescribed. Copyright © 2015 Elsevier B.V. All rights reserved.

Genetic discovery in Xylella fastidiosa through sequence analysis of selected randomly amplified polymorphic DNAs.

PubMed

Chen, Jianchi; Civerolo, Edwin L; Jarret, Robert L; Van Sluys, Marie-Anne; de Oliveira, Mariana C

2005-02-01

Xylella fastidiosa causes many important plant diseases including Pierce's disease (PD) in grape and almond leaf scorch disease (ALSD). DNA-based methodologies, such as randomly amplified polymorphic DNA (RAPD) analysis, have been playing key roles in genetic information collection of the bacterium. This study further analyzed the nucleotide sequences of selected RAPDs from X. fastidiosa strains in conjunction with the available genome sequence databases and unveiled several previously unknown novel genetic traits. These include a sequence highly similar to those in the phage family of Podoviridae. Genome comparisons among X. fastidiosa strains suggested that the "phage" is currently active. Two other RAPDs were also related to horizontal gene transfer: one was part of a broadly distributed cryptic plasmid and the other was associated with conjugal transfer. One RAPD inferred a genomic rearrangement event among X. fastidiosa PD strains and another identified a single nucleotide polymorphism of evolutionary value.

Hepatitis a virus genotypes and strains from an endemic area of Europe, Bulgaria 2012-2014.

PubMed

Bruni, Roberto; Taffon, Stefania; Equestre, Michele; Cella, Eleonora; Lo Presti, Alessandra; Costantino, Angela; Chionne, Paola; Madonna, Elisabetta; Golkocheva-Markova, Elitsa; Bankova, Diljana; Ciccozzi, Massimo; Teoharov, Pavel; Ciccaglione, Anna Rita

2017-07-14

Hepatitis A virus (HAV) infection is endemic in Eastern European and Balkan region countries. In 2012, Bulgaria showed the highest rate (67.13 cases per 100,000) in Europe. Nevertheless, HAV genotypes and strains circulating in this country have never been described. The present study reports the molecular characterization of HAV from 105 patients from Bulgaria. Anti-HAV IgM positive serum samples collected in 2012-2014 from different towns and villages in Bulgaria were analysed by nested RT-PCR, sequencing of the VP1/2A region and phylogenetic analysis; the results were analysed together with patient and geographical data. Phylogenetic analysis revealed two main sequence groups corresponding to the IA (78/105, 74%) and IB (27/105, 26%) sub-genotypes. In the IA group, a major and a minor cluster were observed (62 and 16 sequences, respectively). Most sequences from the major cluster (44/62, 71%) belonged to either of two strains, termed "strain 1" and "strain 2", differing only for a single specific nucleotide; the remaining sequences (18/62, 29%) showed few (1 to 4) nucleotide variations respect to strain 1 and 2. Strain 2 is identical to the strain previously responsible for an outbreak in the Czech Republic in 2008 and a large multi-country European outbreak caused by contaminated mixed frozen berries in 2013. Most sequences of the IA minor cluster and the IB group were detected in large/medium centers (LMCs). Overall, sequences from the IA major cluster were more frequent in small centers (SCs), but strain 1 and strain 2 showed an opposite relative frequency in SCs and LMCs (strain 1 more frequent in SCs, strain 2 in LMCs). Genotype IA predominated in Bulgaria in 2012-2014 and phylogenetic analysis identified a major cluster of highly related or identical IA sequences, representing 59% of the analysed cases; these isolates were mostly detected in SCs, in which HAV shows higher endemicity than in LMCs. The distribution of viral sequences suggests the existence of some differences between the transmission routes in SCs and LMCs. Molecular characterization of an increased number of isolates from Bulgaria, regularly collected over time, will be useful to explore specific transmission routes and plan appropriate preventing measures.

Bradyrhizobium algeriense sp. nov., a novel species isolated from effective nodules of Retama sphaerocarpa from Northeastern Algeria.

PubMed

Ahnia, Hadjira; Bourebaba, Yasmina; Durán, David; Boulila, Farida; Palacios, José M; Rey, Luis; Ruiz-Argüeso, Tomás; Boulila, Abdelghani; Imperial, Juan

2018-04-04

We have characterized genetic, phenotypic and symbiotic properties of bacterial strains previously isolated from nitrogen-fixing nodules of Retama sphaerocarpa from Northern Algeria. Phylogenetic analyses of 16S rRNA genes and three concatenated housekeeping genes, recA, atpD and glnII, placed them in a new divergent group that is proposed to form a new Bradyrhizobium species, Bradyrhizobium algeriense sp. nov. (type strain RST89 T , LMG 27618 and CECT 8363). Based on these phylogenetic markers and on genomic identity data derived from draft genomic sequences, Bradyrhizobium valentinum LmjM3 T , Bradyrhizobium lablabi CCBAU 23086 T , Bradyrhizobium retamae Ro19 T , and Bradyrhizobium jicamae PAC68 T are the closest relatives of B. algeriense RST89 T , with sequence identities of 92-94% and Average Nucleotide Identities (ANIm) under 90%, well below the 95-96% species circumscription threshold. Likewise, a comparison of whole-cell proteomic patterns, estimated by Matrix-Assisted Laser Desorption/Ionization-Time-of-Flight (MALDI-TOF) mass spectrometric analysis, yielded almost identical spectra between B. algeriense strains but significant differences with B. valentinum, Bradyrhizobium paxllaeri, Bradyrhizobium icense, B. lablabi, B. jicamae and B. retamae. A phylogenetic tree based on symbiotic gene nodC revealed that the B. algeriense sequences cluster with sequences from the Bradyrhizobium symbiovar retamae, previously defined with B. retamae strains isolated from Retama monosperma. B. algeriense strains were able to establish effective symbioses with Retama raetam, Lupinus micranthus, Lupinus albus and Genista numidica, but not with Lupinus angustifolius or Glycine max. Copyright © 2018 Elsevier GmbH. All rights reserved.

Molecular detection of kobuviruses in European roe deer (Capreolus capreolus) in Italy.

PubMed

Di Martino, Barbara; Di Profio, Federica; Melegari, Irene; Di Felice, Elisabetta; Robetto, Serena; Guidetti, Cristina; Orusa, Riccardo; Martella, Vito; Marsilio, Fulvio

2015-08-01

Kobuvirus RNA was found in 6.6 % (13/198) of stool specimens from roe deer (Capreolus capreolus) captured during the regular hunting season. Upon sequence analysis of a fragment of the 3D gene, nine strains displayed the highest nucleotide sequence identity (91.2-97.4 %) to bovine kobuviruses previously detected in either diarrhoeic or asymptomatic calves. Interestingly, four strains were genetically related to the newly discovered caprine kobuviruses (84.2-87.6 % nucleotide identity) identified in black goats in Korea.

Isolation of a novel Orientia species (O. chuto sp. nov.) from a patient infected in Dubai.

PubMed

Izzard, Leonard; Fuller, Andrew; Blacksell, Stuart D; Paris, Daniel H; Richards, Allen L; Aukkanit, Nuntipa; Nguyen, Chelsea; Jiang, Ju; Fenwick, Stan; Day, Nicholas P J; Graves, Stephen; Stenos, John

2010-12-01

In July 2006, an Australian tourist returning from Dubai, in the United Arab Emirates (UAE), developed acute scrub typhus. Her signs and symptoms included fever, myalgia, headache, rash, and eschar. Orientia tsutsugamushi serology demonstrated a 4-fold rise in antibody titers in paired serum collections (1:512 to 1:8,192), with the sera reacting strongest against the Gilliam strain antigen. An Orientia species was isolated by the in vitro culture of the patient's acute blood taken prior to antibiotic treatment. The gene sequencing of the 16S rRNA gene (rrs), partial 56-kDa gene, and the full open reading frame 47-kDa gene was performed, and comparisons of this new Orientia sp. isolate to previously characterized strains demonstrated significant sequence diversity. The closest homology to the rrs sequence of the new Orientia sp. isolate was with three strains of O. tsutsugamushi (Ikeda, Kato, and Karp), with a nucleotide sequence similarity of 98.5%. The closest homology to the 47-kDa gene sequence was with O. tsutsugamushi strain Gilliam, with a nucleotide similarity of 82.3%, while the closest homology to the 56-kDa gene sequence was with O. tsutsugamushi strain TA686, with a nucleotide similarity of 53.1%. The molecular divergence and geographically unique origin lead us to believe that this organism should be considered a novel species. Therefore, we have proposed the name "Orientia chuto," and the prototype strain of this species is strain Dubai, named after the location in which the patient was infected.

Isolation of a Novel Orientia Species (O. chuto sp. nov.) from a Patient Infected in Dubai ▿

PubMed Central

Izzard, Leonard; Fuller, Andrew; Blacksell, Stuart D.; Paris, Daniel H.; Richards, Allen L.; Aukkanit, Nuntipa; Nguyen, Chelsea; Jiang, Ju; Fenwick, Stan; Day, Nicholas P. J.; Graves, Stephen; Stenos, John

2010-01-01

In July 2006, an Australian tourist returning from Dubai, in the United Arab Emirates (UAE), developed acute scrub typhus. Her signs and symptoms included fever, myalgia, headache, rash, and eschar. Orientia tsutsugamushi serology demonstrated a 4-fold rise in antibody titers in paired serum collections (1:512 to 1:8,192), with the sera reacting strongest against the Gilliam strain antigen. An Orientia species was isolated by the in vitro culture of the patient's acute blood taken prior to antibiotic treatment. The gene sequencing of the 16S rRNA gene (rrs), partial 56-kDa gene, and the full open reading frame 47-kDa gene was performed, and comparisons of this new Orientia sp. isolate to previously characterized strains demonstrated significant sequence diversity. The closest homology to the rrs sequence of the new Orientia sp. isolate was with three strains of O. tsutsugamushi (Ikeda, Kato, and Karp), with a nucleotide sequence similarity of 98.5%. The closest homology to the 47-kDa gene sequence was with O. tsutsugamushi strain Gilliam, with a nucleotide similarity of 82.3%, while the closest homology to the 56-kDa gene sequence was with O. tsutsugamushi strain TA686, with a nucleotide similarity of 53.1%. The molecular divergence and geographically unique origin lead us to believe that this organism should be considered a novel species. Therefore, we have proposed the name “Orientia chuto,” and the prototype strain of this species is strain Dubai, named after the location in which the patient was infected. PMID:20926708

Genome Sequences of 12 Cluster AN Arthrobacter Phages

PubMed Central

Lee-Soety, Julia Y.; Bhatt, Shantanu; Adair, Tamarah L.; Bonilla, J. Alfred; Klyczek, Karen K.; Harrison, Melinda; Garlena, Rebecca A.; Bowman, Charles A.; Russell, Daniel A.; Jacobs-Sera, Deborah

2017-01-01

ABSTRACT Twelve siphoviral phages isolated using Arthrobacter sp. strain ATCC 21022 were sequenced. The phages all have relatively small genomes, ranging from 15,319 to 15,556 bp. All 12 phages are closely related to previously described cluster AN Arthrobacter phages. PMID:29122859

Spore-forming, Desulfosporosinus-like sulphate-reducing bacteria from a shallow aquifer contaminated with gasoline.

PubMed

Robertson, W J; Franzmann, P D; Mee, B J

2000-02-01

Previous studies on the geochemistry of a shallow unconfined aquifer contaminated with hydrocarbons suggested that the degradation of some hydrocarbons was linked to bacterial sulphate reduction. There was attenuation of naphthalene, 1,3,5-trimethylbenzene (TMB), toluene, p-xylene and ethylbenzene in the groundwater with concomitant loss of sulphate. Here, the recovery of eight strains of sulphate-reducing bacteria (SRB) from the contaminated site is reported. All were straight or curved rod-shaped cells which formed endospores. Amplification and sequencing of the 16S rDNA indicated that the strains were all sulphate reducers of the Gram-positive line of descent, and were most closely related to Desulfosporosinus (previously Desulfotomaculum) orientis DSM 8344 (97-98.9% sequence similarity). The strains clustered in three phylogenetic groups based on 16S rRNA sequences. Whole cell fatty acid compositions were similar to those of D. orientis DSM 8344, and were consistent with previous studies of fatty acids in soil and groundwater from the site. Microcosms containing groundwater from this aquifer indicated a role for sulphate reduction in the degradation of [ring-UL-14C]toluene, but not for the degradation of [UL-14C]benzene which could also be degraded by the microcosms. Adding one of the strains that was isolated from the groundwater (strain T2) to sulphate-enriched microcosms increased the rate of toluene degradation four- to 10-fold but had no effect on the rate of benzene degradation. The addition of molybdate, an inhibitor of sulphate reduction, to the groundwater samples decreased the rate of toluene mineralization. There was no evidence to support the mineralization of [UL-14C]benzene, [ring-UL-14C]toluene or unlabelled m-xylene, p-xylene, ethylbenzene, TMB or naphthalene by any of the strains in pure culture. Growth of all the strains was completely inhibited by 100 micromol l-1 TMB.

Genome sequencing of ovine isolates of Mycobacterium avium subspecies paratuberculosis offers insights into host association

PubMed Central

2012-01-01

Background The genome of Mycobacterium avium subspecies paratuberculosis (MAP) is remarkably homogeneous among the genomes of bovine, human and wildlife isolates. However, previous work in our laboratories with the bovine K-10 strain has revealed substantial differences compared to sheep isolates. To systematically characterize all genomic differences that may be associated with the specific hosts, we sequenced the genomes of three U.S. sheep isolates and also obtained an optical map. Results Our analysis of one of the isolates, MAP S397, revealed a genome 4.8 Mb in size with 4,700 open reading frames (ORFs). Comparative analysis of the MAP S397 isolate showed it acquired approximately 10 large sequence regions that are shared with the human M. avium subsp. hominissuis strain 104 and lost 2 large regions that are present in the bovine strain. In addition, optical mapping defined the presence of 7 large inversions between the bovine and ovine genomes (~ 2.36 Mb). Whole-genome sequencing of 2 additional sheep strains of MAP (JTC1074 and JTC7565) further confirmed genomic homogeneity of the sheep isolates despite the presence of polymorphisms on the nucleotide level. Conclusions Comparative sequence analysis employed here provided a better understanding of the host association, evolution of members of the M. avium complex and could help in deciphering the phenotypic differences observed among sheep and cattle strains of MAP. A similar approach based on whole-genome sequencing combined with optical mapping could be employed to examine closely related pathogens. We propose an evolutionary scenario for M. avium complex strains based on these genome sequences. PMID:22409516

Candida guilliermondii and Other Species of Candida Misidentified as Candida famata: Assessment by Vitek 2, DNA Sequencing Analysis, and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry in Two Global Antifungal Surveillance Programs

PubMed Central

Woosley, Leah N.; Diekema, Daniel J.; Jones, Ronald N.; Pfaller, Michael A.

2013-01-01

Candida famata (teleomorph Debaryomyces hansenii) has been described as a medically relevant yeast, and this species has been included in many commercial identification systems that are currently used in clinical laboratories. Among 53 strains collected during the SENTRY and ARTEMIS surveillance programs and previously identified as C. famata (includes all submitted strains with this identification) by a variety of commercial methods (Vitek, MicroScan, API, and AuxaColor), DNA sequencing methods demonstrated that 19 strains were C. guilliermondii, 14 were C. parapsilosis, 5 were C. lusitaniae, 4 were C. albicans, and 3 were C. tropicalis, and five isolates belonged to other Candida species (two C. fermentati and one each C. intermedia, C. pelliculosa, and Pichia fabianni). Additionally, three misidentified C. famata strains were correctly identified as Kodomaea ohmeri, Debaryomyces nepalensis, and Debaryomyces fabryi using intergenic transcribed spacer (ITS) and/or intergenic spacer (IGS) sequencing. The Vitek 2 system identified three isolates with high confidence to be C. famata and another 15 with low confidence between C. famata and C. guilliermondii or C. parapsilosis, displaying only 56.6% agreement with DNA sequencing results. Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) results displayed 81.1% agreement with DNA sequencing. One strain each of C. metapsilosis, C. fermentati, and C. intermedia demonstrated a low score for identification (<2.0) in the MALDI Biotyper. K. ohmeri, D. nepalensis, and D. fabryi identified by DNA sequencing in this study were not in the current database for the MALDI Biotyper. These results suggest that the occurrence of C. famata in fungal infections is much lower than previously appreciated and that commercial systems do not produce accurate identifications except for the newly introduced MALDI-TOF instruments. PMID:23100350

Deletion endpoint allele-specificity in the developmentally regulated elimination of an internal sequence (IES) in Paramecium.

PubMed Central

Dubrana, K; Le Mouël, A; Amar, L

1997-01-01

Ciliated protozoa undergo thousands of site-specific DNA deletion events during the programmed development of micronuclear genomes to macronuclear genomes. Two deletion elements, W1 and W2, were identified in the Paramecium primaurelia wild-type 156 strain. Here, we report the characterization of both elements in wild-type strain 168 and show that they display variant deletion patterns when compared with those of strain 156. The W1 ( 168 ) element is defective for deletion. The W2 ( 168 ) element is excised utilizing two alternative boundaries on one side, both are different from the boundary utilized to excise the W2156 element. By crossing the 156 and 168 strains, we demonstrate that the definition of all deletion endpoints are each controlled by cis -acting determinant(s) rather than by strain-specific trans-acting factor(s). Sequence comparison of all deleted DNA segments indicates that the 5'-TA-3'terminal sequence is strictly required at their ends. Furthermore the identity of the first eight base pairs of these ends to a previously established consensus sequence correlates with the frequency of the corresponding deletion events. Our data implies the existence of an adaptive convergent evolution of these Paramecium deleted DNA segment end sequences. PMID:9171098

Population Structure and Antimicrobial Resistance Profiles of Streptococcus suis Serotype 2 Sequence Type 25 Strains.

PubMed

Athey, Taryn B T; Teatero, Sarah; Takamatsu, Daisuke; Wasserscheid, Jessica; Dewar, Ken; Gottschalk, Marcelo; Fittipaldi, Nahuel

2016-01-01

Strains of serotype 2 Streptococcus suis are responsible for swine and human infections. Different serotype 2 genetic backgrounds have been defined using multilocus sequence typing (MLST). However, little is known about the genetic diversity within each MLST sequence type (ST). Here, we used whole-genome sequencing to test the hypothesis that S. suis serotype 2 strains of the ST25 lineage are genetically heterogeneous. We evaluated 51 serotype 2 ST25 S. suis strains isolated from diseased pigs and humans in Canada, the United States of America, and Thailand. Whole-genome sequencing revealed numerous large-scale rearrangements in the ST25 genome, compared to the genomes of ST1 and ST28 S. suis strains, which result, among other changes, in disruption of a pilus island locus. We report that recombination and lateral gene transfer contribute to ST25 genetic diversity. Phylogenetic analysis identified two main and distinct Thai and North American clades grouping most strains investigated. These clades also possessed distinct patterns of antimicrobial resistance genes, which correlated with acquisition of different integrative and conjugative elements (ICEs). Some of these ICEs were found to be integrated at a recombination hot spot, previously identified as the site of integration of the 89K pathogenicity island in serotype 2 ST7 S. suis strains. Our results highlight the limitations of MLST for phylogenetic analysis of S. suis, and the importance of lateral gene transfer and recombination as drivers of diversity in this swine pathogen and zoonotic agent.

Population Structure and Antimicrobial Resistance Profiles of Streptococcus suis Serotype 2 Sequence Type 25 Strains

PubMed Central

Athey, Taryn B. T.; Teatero, Sarah; Takamatsu, Daisuke; Wasserscheid, Jessica; Dewar, Ken; Gottschalk, Marcelo; Fittipaldi, Nahuel

2016-01-01

Strains of serotype 2 Streptococcus suis are responsible for swine and human infections. Different serotype 2 genetic backgrounds have been defined using multilocus sequence typing (MLST). However, little is known about the genetic diversity within each MLST sequence type (ST). Here, we used whole-genome sequencing to test the hypothesis that S. suis serotype 2 strains of the ST25 lineage are genetically heterogeneous. We evaluated 51 serotype 2 ST25 S. suis strains isolated from diseased pigs and humans in Canada, the United States of America, and Thailand. Whole-genome sequencing revealed numerous large-scale rearrangements in the ST25 genome, compared to the genomes of ST1 and ST28 S. suis strains, which result, among other changes, in disruption of a pilus island locus. We report that recombination and lateral gene transfer contribute to ST25 genetic diversity. Phylogenetic analysis identified two main and distinct Thai and North American clades grouping most strains investigated. These clades also possessed distinct patterns of antimicrobial resistance genes, which correlated with acquisition of different integrative and conjugative elements (ICEs). Some of these ICEs were found to be integrated at a recombination hot spot, previously identified as the site of integration of the 89K pathogenicity island in serotype 2 ST7 S. suis strains. Our results highlight the limitations of MLST for phylogenetic analysis of S. suis, and the importance of lateral gene transfer and recombination as drivers of diversity in this swine pathogen and zoonotic agent. PMID:26954687

Sequence diversity of the leukotoxin (lktA) gene in caprine and ovine strains of Mannheimia haemolytica.

PubMed

Vougidou, C; Sandalakis, V; Psaroulaki, A; Petridou, E; Ekateriniadou, L

2013-04-20

Mannheimia haemolytica is the aetiological agent of pneumonic pasteurellosis in small ruminants. The primary virulence factor of the bacterium is a leukotoxin (LktA), which induces apoptosis in susceptible cells via mitochondrial targeting. It has been previously shown that certain lktA alleles are associated either with cattle or sheep. The objective of the present study was to investigate lktA sequence variation among ovine and caprine M haemolytica strains isolated from pneumonic lungs, revealing any potential adaptation for the caprine host, for which there is no available data. Furthermore, we investigated amino acid variation in the N-terminal part of the sequences and its effect on targeting mitochondria. Data analysis showed that the prevalent caprine genotype differed at a single non-synonymous site from a previously described uncommon bovine allele, whereas the ovine sequences represented new, distinct alleles. N-terminal sequence differences did not affect the mitochondrial targeting ability of the isolates; interestingly enough in one case, mitochondrial matrix targeting was indicated rather than membrane association, suggesting an alternative LktA trafficking pattern.

Complete Genome Sequence of an Avian Paramyxovirus Type 4 from North America Reveals a Shorter Genome and New Genotype

PubMed Central

Parthiban, Manoharan; Kaliyaperumal, Manimaran; Xiao, Sa; Nayak, Baibaswata; Paldurai, Anandan; Kim, Shin-Hee; Ladman, Brian S.; Preskenis, Lauren A.; Gelb, Jack; Collins, Peter L.

2013-01-01

An avian paramyxovirus type 4 (APMV-4) was isolated from a duck in Delaware in 2010. Its genome is 15,048 nucleotides (nt) long, which is shorter by 6 nt than those for all previously reported strains. Phylogenetic analysis revealed that this strain formed a separate cluster within APMV-4 strains. PMID:23405329

Recombinational hotspot specific to female meiosis in the mouse major histocompatibility complex.

PubMed

Shiroishi, T; Hanzawa, N; Sagai, T; Ishiura, M; Gojobori, T; Steinmetz, M; Moriwaki, K

1990-01-01

The wm7 haplotype of the major histocompatibility complex (MHC), derived from the Japanese wild mouse Mus musculus molossinus, enhances recombination specific to female meiosis in the K/A beta interval of the MHC. We have mapped crossover points of fifteen independent recombinants from genetic crosses of the wm7 and laboratory haplotypes. Most of them were confined to a short segment of approximately 1 kilobase (kb) of DNA between the A beta 3 and A beta 2 genes, indicating the presence of a female-specific recombinational hotspot. Its location overlaps with a sex-independent hotspot previously identified in the Mus musculus castaneus CAS3 haplotype. We have cloned and sequenced DNA fragments surrounding the hotspot from the wm7 haplotype and the corresponding regions from the hotspot-negative B10.A and C57BL/10 strains. There is no significant difference between the sequences of these three strains, or between these and the published sequences of the CAS3 and C57BL/6 strains. However, a comparison of this A beta 3/A beta 2 hotspot with a previously characterized hotspot in the E beta gene revealed that they have a very similar molecular organization. Each hotspot consists of two elements, the consensus sequence of the mouse middle repetitive MT family and the tetrameric repeated sequences, which are separated by 1 kb of DNA.

Draft Genome Sequences of Six Mycobacterium immunogenum, Strains Obtained from a Chloraminated Drinking Water Distribution System Simulator

EPA Science Inventory

We report the draft genome sequences of six Mycobacterium immunogenum isolated from a chloraminated drinking water distribution system simulator subjected to changes in operational parameters. M. immunogenum, a rapidly growing mycobacteria previously reported as the cause of hyp...

Comparative genome analysis and characterization of the Salmonella Typhimurium strain CCRJ_26 isolated from swine carcasses using whole-genome sequencing approach.

PubMed

Panzenhagen, P H N; Cabral, C C; Suffys, P N; Franco, R M; Rodrigues, D P; Conte-Junior, C A

2018-04-01

Salmonella pathogenicity relies on virulence factors many of which are clustered within the Salmonella pathogenicity islands. Salmonella also harbours mobile genetic elements such as virulence plasmids, prophage-like elements and antimicrobial resistance genes which can contribute to increase its pathogenicity. Here, we have genetically characterized a selected S. Typhimurium strain (CCRJ_26) from our previous study with Multiple Drugs Resistant profile and high-frequency PFGE clonal profile which apparently persists in the pork production centre of Rio de Janeiro State, Brazil. By whole-genome sequencing, we described the strain's genome virulent content and characterized the repertoire of bacterial plasmids, antibiotic resistance genes and prophage-like elements. Here, we have shown evidence that strain CCRJ_26 genome possible represent a virulence-associated phenotype which may be potentially virulent in human infection. Whole-genome sequencing technologies are still costly and remain underexplored for applied microbiology in Brazil. Hence, this genomic description of S. Typhimurium strain CCRJ_26 will provide help in future molecular epidemiological studies. The analysis described here reveals a quick and useful pipeline for bacterial virulence characterization using whole-genome sequencing approach. © 2018 The Society for Applied Microbiology.

Virulence Studies of Different Sequence Types and Geographical Origins of Streptococcus suis Serotype 2 in a Mouse Model of Infection

PubMed Central

Auger, Jean-Philippe; Fittipaldi, Nahuel; Benoit-Biancamano, Marie-Odile; Segura, Mariela; Gottschalk, Marcelo

2016-01-01

Multilocus sequence typing previously identified three predominant sequence types (STs) of Streptococcus suis serotype 2: ST1 strains predominate in Eurasia while North American (NA) strains are generally ST25 and ST28. However, ST25/ST28 and ST1 strains have also been isolated in Asia and NA, respectively. Using a well-standardized mouse model of infection, the virulence of strains belonging to different STs and different geographical origins was evaluated. Results demonstrated that although a certain tendency may be observed, S. suis serotype 2 virulence is difficult to predict based on ST and geographical origin alone; strains belonging to the same ST presented important differences of virulence and did not always correlate with origin. The only exception appears to be NA ST28 strains, which were generally less virulent in both systemic and central nervous system (CNS) infection models. Persistent and high levels of bacteremia accompanied by elevated CNS inflammation are required to cause meningitis. Although widely used, in vitro tests such as phagocytosis and killing assays require further standardization in order to be used as predictive tests for evaluating virulence of strains. The use of strains other than archetypal strains has increased our knowledge and understanding of the S. suis serotype 2 population dynamics. PMID:27409640

Sequence analysis of the msp4 gene of Anaplasma ovis strains

USGS Publications Warehouse

de la Fuente, J.; Atkinson, M.W.; Naranjo, V.; Fernandez de Mera, I. G.; Mangold, A.J.; Keating, K.A.; Kocan, K.M.

2007-01-01

Anaplasma ovis (Rickettsiales: Anaplasmataceae) is a tick-borne pathogen of sheep, goats and wild ruminants. The genetic diversity of A. ovis strains has not been well characterized due to the lack of sequence information. In this study, we evaluated bighorn sheep (Ovis canadensis) and mule deer (Odocoileus hemionus) from Montana for infection with A. ovis by serology and sequence analysis of the msp4 gene. Antibodies to Anaplasma spp. were detected in 37% and 39% of bighorn sheep and mule deer analyzed, respectively. Four new msp4 genotypes were identified. The A. ovis msp4 sequences identified herein were analyzed together with sequences reported previously for the characterization of the genetic diversity of A. ovis strains in comparison with other Anaplasma spp. The results of these studies demonstrated that although A. ovis msp4 genotypes may vary among geographic regions and between sheep and deer hosts, the variation observed was less than the variation observed between A. marginale and A. phagocytophilum strains. The results reported herein further confirm that A. ovis infection occurs in natural wild ruminant populations in Western United States and that bighorn sheep and mule deer may serve as wildlife reservoirs of A. ovis. ?? 2006.

16S-23S rRNA gene internal transcribed spacer sequences for analysis of the phylogenetic relationships among species of the genus Porphyromonas.

PubMed

Conrads, Georg; Citron, Diane M; Tyrrell, Kerin L; Horz, Hans-Peter; Goldstein, Ellie J C

2005-03-01

The 16S-23S rRNA gene internal transcribed spacer (ITS) regions of 11 reference strains of Porphyromonas species, together with Bacteroides distasonis and Tannerella forsythensis, were analysed to examine interspecies relationships. Compared with the phylogenetic tree generated using 16S rRNA gene sequences, the resolution of the ITS sequence-based tree was higher, but species positioning and clustering were similar with both approaches. The recent separation of Porphyromonas gulae and Porphyromonas gingivalis into distinct species was confirmed by the ITS data. In addition, analysis of the ITS sequences of 24 clinical isolates of Porphyromonas asaccharolytica plus the type strain ATCC 25260(T) divided the sequences into two clusters, of which one was alpha-fucosidase-positive (like the type strain) while the other was alpha-fucosidase-negative. The latter resembled the previously studied unusual extra-oral isolates of 'Porphyromonas endodontalis-like organisms' (PELOs) which could therefore be called 'Porphyromonas asaccharolytica-like organisms' (PALOs), based on the genetic identification. Moreover, the proposal of alpha-fucosidase-negative P. asaccharolytica strains as a new species should also be considered.

Comparison and phylogenetic analysis of the ISS gene in two predominant avian pathogenic E. coli serogroups isolated from avian colibacillosis in Iran.

PubMed

Zahraei Salehi, Taghi; Derakhshandeh, Abdollah; Tadjbakhsh, Hasan; Karimi, Vahid

2013-02-01

The ISS (increased serum survival) gene and its protein product (ISS) of avian pathogenic Escherichia coli (APEC) are important characteristics of resistance to the complement system. The aims of this study were to clone, sequence and characterize sequence diversity of the ISS gene between two predominant serogroups in Iran and among those previously deposited in Genbank. The ISS gene of 309 bp from the APEC χ1390 strain was amplified by PCR, cloned and sequenced using pTZ57R/T vector. The ISS gene from the χ1390 strain has 100% identity among different serogroups of APEC in different geographical regions throughout the world. Phylogenetic analysis shows two different phylogenic groups among the different strains. Strong association of nucleotide sequences among different E. coli strains suggests that it may be a conserved gene and could be a suitable antigen to control and detect avian pathogenic E. coli, at least in our region. Currently, our group is working on the ISS protein as candidate vaccine in SPF poultry. Copyright © 2012 Elsevier Ltd. All rights reserved.

Diagnostics for Yaws Eradication: Insights From Direct Next-Generation Sequencing of Cutaneous Strains of Treponema pallidum

PubMed Central

Marks, Michael; Fookes, Maria; Wagner, Josef; Butcher, Robert; Ghinai, Rosanna; Sokana, Oliver; Sarkodie, Yaw-Adu; Lukehart, Sheila A; Solomon, Anthony W; Mabey, David C W; Thomson, Nicholas

2018-01-01

Abstract Background Yaws-like chronic ulcers can be caused by Treponema pallidum subspecies pertenue, Haemophilus ducreyi, or other, still-undefined bacteria. To permit accurate evaluation of yaws elimination efforts, programmatic use of molecular diagnostics is required. The accuracy and sensitivity of current tools remain unclear because our understanding of T. pallidum diversity is limited by the low number of sequenced genomes. Methods We tested samples from patients with suspected yaws collected in the Solomon Islands and Ghana. All samples were from patients whose lesions had previously tested negative using the Centers for Disease Control and Prevention (CDC) diagnostic assay in widespread use. However, some of these patients had positive serological assays for yaws on blood. We used direct whole-genome sequencing to identify T. pallidum subsp pertenue strains missed by the current assay. Results From 45 Solomon Islands and 27 Ghanaian samples, 11 were positive for T. pallidum DNA using the species-wide quantitative polymerase chain reaction (PCR) assay, from which we obtained 6 previously undetected T. pallidum subsp pertenue whole-genome sequences. These show that Solomon Islands sequences represent distinct T. pallidum subsp pertenue clades. These isolates were invisible to the CDC diagnostic PCR assay, due to sequence variation in the primer binding site. Conclusions Our data double the number of published T. pallidum subsp pertenue genomes. We show that Solomon Islands strains are undetectable by the PCR used in many studies and by health ministries. This assay is therefore not adequate for the eradication program. Next-generation genome sequence data are essential for these efforts. PMID:29045605

Comprehensive Insights in the Mycobacterium avium subsp. paratuberculosis Genome Using New WGS Data of Sheep Strain JIII-386 from Germany

PubMed Central

Möbius, Petra; Hölzer, Martin; Felder, Marius; Nordsiek, Gabriele; Groth, Marco; Köhler, Heike; Reichwald, Kathrin; Platzer, Matthias; Marz, Manja

2015-01-01

Mycobacterium avium (M. a.) subsp. paratuberculosis (MAP)—the etiologic agent of Johne’s disease—affects cattle, sheep, and other ruminants worldwide. To decipher phenotypic differences among sheep and cattle strains (belonging to MAP-S [Type-I/III], respectively, MAP-C [Type-II]), comparative genome analysis needs data from diverse isolates originating from different geographic regions of the world. This study presents the so far best assembled genome of a MAP-S-strain: Sheep isolate JIII-386 from Germany. One newly sequenced cattle isolate (JII-1961, Germany), four published MAP strains of MAP-C and MAP-S from the United States and Australia, and M. a. subsp. hominissuis (MAH) strain 104 were used for assembly improvement and comparisons. All genomes were annotated by BacProt and results compared with NCBI (National Center for Biotechnology Information) annotation. Corresponding protein-coding sequences (CDSs) were detected, but also CDSs that were exclusively determined by either NCBI or BacProt. A new Shine–Dalgarno sequence motif (5′-AGCTGG-3′) was extracted. Novel CDSs including PE-PGRS family protein genes and about 80 noncoding RNAs exhibiting high sequence conservation are presented. Previously found genetic differences between MAP-types are partially revised. Four of ten assumed MAP-S-specific large sequence polymorphism regions (LSPSs) are still present in MAP-C strains; new LSPSs were identified. Independently of the regional origin of the strains, the number of individual CDSs and single nucleotide variants confirms the strong similarity of MAP-C strains and shows higher diversity among MAP-S strains. This study gives ambiguous results regarding the hypothesis that MAP-S is the evolutionary intermediate between MAH and MAP-C, but it clearly shows a higher similarity of MAP to MAH than to Mycobacterium intracellulare. PMID:26384038

[Analysis on molecular epidemiology of rubella virus in Shandong province during 2000-2007].

PubMed

Wang, Chang-Yin; Zhu, Zhen; Xu, Ai-Qiang; Xiong, Ping; Song, Li-Zhi; Xu, Qing; Feng, Lei; Xu, Wen-Bo

2010-11-01

Analyze the genetic characteristics of sixteen strains of wild-type rubella viruses derived from Vero cells, Rk13 cells or Vero/slam cells, and isolated from throat samples in Shandong province during 2000-2007. The 1107 nucleotide sequence of nucleoprotein (E1) gene of these isolates were amplified by RT-PCR, and the PCR products were directly sequenced. Comparing with the gene tree that was constructed based on the 739 gene sequences of the WHO reference strains, twelve isolated strains belonged to 1E genotype, one strain belonged to 1F genotype, three strains belonged to 2A genotype. The first strain belonged to 1E genotype was isolated in Shandong province in 2001, then genotype 1E became dominant genotype of wild rubella viruses circulated. The 1E genotype circulated from 2006-2007 was different compared with that circulated from 2001 to 2002, but no significant deviation in temporal and geographic distribution was found. The strain belonged to Genotype 1F was only isolated during 2000 to 2001. The three strains of 2A genotype of rubella viruses were similar to rubella viruses vaccine strain (BRDII). The most nucleotide mutation of rubella viruses among the sixteen strains were nonsense mutation, and the amino acid sequences were highly conservative with no change in important antigen sites. Alike the previous reports, there was the same amino acid mutation in protein E1 at the site of 338 in all of the 1E genotype rubella viruses isolated during 2001- 2007 in Shandong (Leu338 --> Phe338).

Divergence in substrate specificity by the vOTU domain of various strains of highly-pathogenic PRRSV and the implications to pathogenicity

USDA-ARS?s Scientific Manuscript database

Porcine reproductive and respiratory syndrome virus (PRRSV) is widespread with a high variation in sequence and virulence among the divergent strains and causes an economically destructive disease. A viral ovarian domain protease (vOTU) has been previously identified within the nonstructural protein...

Identification of putative plant pathogenic determinants from a draft genome sequence of an opportunistic klebsiella pneumoniae strain

USDA-ARS?s Scientific Manuscript database

Klebsiella pneumoniae has been known historically as a causal agent of bacterial pneumonia. More recently, K. pneumoniaerepresentatives have been shown to have a broad ecological distribution and are recognized nitrogen-fixers. Previously, we demonstrated the capacity of K. pneumoniae strain Kp 5-1R...

Complete genomic sequence of an infectious pancreatic necrosis virus isolated from rainbow trout (Oncorhynchus mykiss) in China.

PubMed

Ji, Feng; Zhao, Jing-Zhuang; Liu, Miao; Lu, Tong-Yan; Liu, Hong-Bai; Yin, Jiasheng; Xu, Li-Ming

2017-04-01

Infectious pancreatic necrosis (IPN) is a significant disease of farmed salmonids resulting in direct economic losses due to high mortality in China. However, no gene sequence of any Chinese infectious pancreatic necrosis virus (IPNV) isolates was available. In the study, moribund rainbow trout fry samples were collected during an outbreak of IPN in Yunnan province of southwest China in 2013. An IPNV was isolated and tentatively named ChRtm213. We determined the full genome sequence of the IPNV ChRtm213 and compared it with previously identified IPNV sequences worldwide. The sequences of different structural and non-structural protein genes were compared to those of other aquatic birnaviruses sequenced to date. The results indicated that the complete genome sequence of ChRtm213 strain contains a segment A (3099 nucleotides) coding a polyprotein VP2-VP4-VP3, and a segment B (2789 nucleotides) coding a RNA-dependent RNA polymerase VP1. The phylogenetic analyses showed that ChRtm213 strain fell within genogroup 1, serotype A9 (Jasper), having similarities of 96.3% (segment A) and 97.3% (segment B) with the IPNV strain AM98 from Japan. The results suggest that the Chinese IPNV isolate has relative closer relationship with Japanese IPNV strains. The sequence of ChRtm213 was the first gene sequence of IPNV isolates in China. This study provided a robust reference for diagnosis and/or control of IPNV prevalent in China.

Complete coding regions of the prototypes enterovirus B93 and C95: phylogenetic analyses of the P1 and P3 regions of EV-B and EV-C strains.

PubMed

Junttila, N; Lévêque, N; Magnius, L O; Kabue, J P; Muyembe-Tamfum, J J; Maslin, J; Lina, B; Norder, H

2015-03-01

Complete coding regions were sequenced for two new enterovirus genomes: EV-B93 previously identified by VP1 sequencing, derived from a child with acute flaccid paralysis in the Democratic Republic of Congo; and EV-C95 from a French soldier with acute gastroenteritis in Djibouti. The EV-B93 P1 had more than 30% nucleotide divergence from other EV-B types, with highest similarity to E-15 and EV-B80. The P1 nucleotide sequence of EV-C95 was most similar, 71%, to CV-A21. Complete coding regions for the new enteroviruses were compared with those of 135 EV-B and 176 EV-C strains representing all types available in GenBank. When strains from the same outbreak or strains isolated during the same year in the same geographical region were excluded, 27 of the 58 EV-B, and 16 of the 23 EV-C types were represented by more than one sequence. However, for EV-B the P3 sequences formed three clades mainly according to origin or time of isolation, irrespective of type, while for EV-C the P3 sequences segregated mainly according to disease manifestation, with most strains causing paralysis, including polioviruses, forming one clade, and strains causing respiratory illness forming another. There was no intermixing of types between these two clades, apart from two EV-C96 strains. The EV-B P3 sequences had lower inter-clade and higher intra-clade variability as compared to the EV-C sequences, which may explain why inter-clade recombinations are more frequent in EV-B. Further analysis of more isolates may shed light on the role of recombinations in the evolution of EV-B in geographical context. © 2014 Wiley Periodicals, Inc.

A Wide Variety of Clostridium perfringens Type A Food-Borne Isolates That Carry a Chromosomal cpe Gene Belong to One Multilocus Sequence Typing Cluster

PubMed Central

Xiao, Yinghua; Wagendorp, Arjen; Moezelaar, Roy; Abee, Tjakko

2012-01-01

Of 98 suspected food-borne Clostridium perfringens isolates obtained from a nationwide survey by the Food and Consumer Product Safety Authority in The Netherlands, 59 strains were identified as C. perfringens type A. Using PCR-based techniques, the cpe gene encoding enterotoxin was detected in eight isolates, showing a chromosomal location for seven isolates and a plasmid location for one isolate. Further characterization of these strains by using (GTG)5 fingerprint repetitive sequence-based PCR analysis distinguished C. perfringens from other sulfite-reducing clostridia but did not allow for differentiation between various types of C. perfringens strains. To characterize the C. perfringens strains further, multilocus sequence typing (MLST) analysis was performed on eight housekeeping genes of both enterotoxic and non-cpe isolates, and the data were combined with a previous global survey covering strains associated with food poisoning, gas gangrene, and isolates from food or healthy individuals. This revealed that the chromosomal cpe strains (food strains and isolates from food poisoning cases) belong to a distinct cluster that is significantly distant from all the other cpe plasmid-carrying and cpe-negative strains. These results suggest that different groups of C. perfringens have undergone niche specialization and that a distinct group of food isolates has specific core genome sequences. Such findings have epidemiological and evolutionary significance. Better understanding of the origin and reservoir of enterotoxic C. perfringens may allow for improved control of this organism in foods. PMID:22865060

[Phylogenetic analysis of genomes of Vibrio cholerae strains isolated on the territory of Rostov region].

PubMed

Kuleshov, K V; Markelov, M L; Dedkov, V G; Vodop'ianov, A S; Kermanov, A V; Pisanov, R V; Kruglikov, V D; Mazrukho, A B; Maleev, V V; Shipulin, G A

2013-01-01

Determination of origin of 2 Vibrio cholerae strains isolated on the territory of Rostov region by using full genome sequencing data. Toxigenic strain 2011 EL- 301 V. cholerae 01 El Tor Inaba No. 301 (ctxAB+, tcpA+) and nontoxigenic strain V. cholerae O1 Ogawa P- 18785 (ctxAB-, tcpA+) were studied. Sequencing was carried out on the MiSeq platform. Phylogenetic analysis of the genomes obtained was carried out based on comparison of conservative part of the studied and 54 previously sequenced genomes. 2011EL-301 strain genome was presented by 164 contigs with an average coverage of 100, N50 parameter was 132 kb, for strain P- 18785 - 159 contigs with a coverage of69, N50 - 83 kb. The contigs obtained for strain 2011 EL-301 were deposited in DDBJ/EMBL/GenBank databases with access code AJFN02000000, for strain P-18785 - ANHS00000000. 716 protein-coding orthologous genes were detected. Based on phylogenetic analysis strain P- 18785 belongs to PG-1 subgroup (a group of predecessor strains of the 7th pandemic). Strain 2011EL-301 belongs to groups of strains of the 7th pandemic and is included into the cluster with later isolates that are associated with cases of cholera in South Africa and cases of import of cholera to the USA from Pakistan. The data obtained allows to establish phylogenetic connections with V cholerae strains isolated earlier.

Genotyping-By-Sequencing (GBS) Detects Genetic Structure and Confirms Behavioral QTL in Tame and Aggressive Foxes (Vulpes vulpes)

PubMed Central

Johnson, Jennifer L.; Wittgenstein, Helena; Mitchell, Sharon E.; Hyma, Katie E.; Temnykh, Svetlana V.; Kharlamova, Anastasiya V.; Gulevich, Rimma G.; Vladimirova, Anastasiya V.; Fong, Hiu Wa Flora; Acland, Gregory M.; Trut, Lyudmila N.; Kukekova, Anna V.

2015-01-01

The silver fox (Vulpes vulpes) offers a novel model for studying the genetics of social behavior and animal domestication. Selection of foxes, separately, for tame and for aggressive behavior has yielded two strains with markedly different, genetically determined, behavioral phenotypes. Tame strain foxes are eager to establish human contact while foxes from the aggressive strain are aggressive and difficult to handle. These strains have been maintained as separate outbred lines for over 40 generations but their genetic structure has not been previously investigated. We applied a genotyping-by-sequencing (GBS) approach to provide insights into the genetic composition of these fox populations. Sequence analysis of EcoT22I genomic libraries of tame and aggressive foxes identified 48,294 high quality SNPs. Population structure analysis revealed genetic divergence between the two strains and more diversity in the aggressive strain than in the tame one. Significant differences in allele frequency between the strains were identified for 68 SNPs. Three of these SNPs were located on fox chromosome 14 within an interval of a previously identified behavioral QTL, further supporting the importance of this region for behavior. The GBS SNP data confirmed that significant genetic diversity has been preserved in both fox populations despite many years of selective breeding. Analysis of SNP allele frequencies in the two populations identified several regions of genetic divergence between the tame and aggressive foxes, some of which may represent targets of selection for behavior. The GBS protocol used in this study significantly expanded genomic resources for the fox, and can be adapted for SNP discovery and genotyping in other canid species. PMID:26061395

Genotyping-By-Sequencing (GBS) Detects Genetic Structure and Confirms Behavioral QTL in Tame and Aggressive Foxes (Vulpes vulpes).

PubMed

Johnson, Jennifer L; Wittgenstein, Helena; Mitchell, Sharon E; Hyma, Katie E; Temnykh, Svetlana V; Kharlamova, Anastasiya V; Gulevich, Rimma G; Vladimirova, Anastasiya V; Fong, Hiu Wa Flora; Acland, Gregory M; Trut, Lyudmila N; Kukekova, Anna V

2015-01-01

The silver fox (Vulpes vulpes) offers a novel model for studying the genetics of social behavior and animal domestication. Selection of foxes, separately, for tame and for aggressive behavior has yielded two strains with markedly different, genetically determined, behavioral phenotypes. Tame strain foxes are eager to establish human contact while foxes from the aggressive strain are aggressive and difficult to handle. These strains have been maintained as separate outbred lines for over 40 generations but their genetic structure has not been previously investigated. We applied a genotyping-by-sequencing (GBS) approach to provide insights into the genetic composition of these fox populations. Sequence analysis of EcoT22I genomic libraries of tame and aggressive foxes identified 48,294 high quality SNPs. Population structure analysis revealed genetic divergence between the two strains and more diversity in the aggressive strain than in the tame one. Significant differences in allele frequency between the strains were identified for 68 SNPs. Three of these SNPs were located on fox chromosome 14 within an interval of a previously identified behavioral QTL, further supporting the importance of this region for behavior. The GBS SNP data confirmed that significant genetic diversity has been preserved in both fox populations despite many years of selective breeding. Analysis of SNP allele frequencies in the two populations identified several regions of genetic divergence between the tame and aggressive foxes, some of which may represent targets of selection for behavior. The GBS protocol used in this study significantly expanded genomic resources for the fox, and can be adapted for SNP discovery and genotyping in other canid species.

Characterization of a Large Antibiotic Resistance Plasmid Found in Enteropathogenic Escherichia coli Strain B171 and Its Relatedness to Plasmids of Diverse E. coli and Shigella Strains.

PubMed

Hazen, Tracy H; Michalski, Jane; Nagaraj, Sushma; Okeke, Iruka N; Rasko, David A

2017-09-01

Enteropathogenic Escherichia coli (EPEC) is a leading cause of severe infantile diarrhea in developing countries. Previous research has focused on the diversity of the EPEC virulence plasmid, whereas less is known regarding the genetic content and distribution of antibiotic resistance plasmids carried by EPEC. A previous study demonstrated that in addition to the virulence plasmid, reference EPEC strain B171 harbors a second, larger plasmid that confers antibiotic resistance. To further understand the genetic diversity and dissemination of antibiotic resistance plasmids among EPEC strains, we describe the complete sequence of an antibiotic resistance plasmid from EPEC strain B171. The resistance plasmid, pB171_90, has a completed sequence length of 90,229 bp, a GC content of 54.55%, and carries protein-encoding genes involved in conjugative transfer, resistance to tetracycline ( tetA ), sulfonamides ( sulI ), and mercury, as well as several virulence-associated genes, including the transcriptional regulator hha and the putative calcium sequestration inhibitor ( csi ). In silico detection of the pB171_90 genes among 4,798 publicly available E. coli genome assemblies indicates that the unique genes of pB171_90 ( csi and traI ) are primarily restricted to genomes identified as EPEC or enterotoxigenic E. coli However, conserved regions of the pB171_90 plasmid containing genes involved in replication, stability, and antibiotic resistance were identified among diverse E. coli pathotypes. Interestingly, pB171_90 also exhibited significant similarity with a sequenced plasmid from Shigella dysenteriae type I. Our findings demonstrate the mosaic nature of EPEC antibiotic resistance plasmids and highlight the need for additional sequence-based characterization of antibiotic resistance plasmids harbored by pathogenic E. coli . Copyright © 2017 American Society for Microbiology.

Precipitation of alacranite (As8S9) by a novel As(V)-respiring anaerobe strain MPA-C3.

PubMed

Mumford, Adam C; Yee, Nathan; Young, Lily Y

2013-10-01

Strain MPA-C3 was isolated by incubating arsenic-bearing sediments under anaerobic, mesophilic conditions in minimal media with acetate as the sole source of energy and carbon, and As(V) as the sole electron acceptor. Following growth and the respiratory reduction of As(V) to As(III), a yellow precipitate formed in active cultures, while no precipitate was observed in autoclaved controls, or in uninoculated media supplemented with As(III). The precipitate was identified by X-ray diffraction as alacranite, As8 S9 , a mineral previously only identified in hydrothermal environments. Sequencing of the 16S rRNA gene indicated that strain MPA-C3 is a member of the Deferribacteres family, with relatively low (90%) identity to Denitrovibrio acetiphilus DSM 12809. The arsenate respiratory reductase gene, arrA, was sequenced, showing high homology to the arrA gene of Desulfitobacterium halfniense. In addition to As(V), strain MPA-C3 utilizes NO3(-), Se(VI), Se(IV), fumarate and Fe(III) as electron acceptors, and acetate, pyruvate, fructose and benzoate as sources of carbon and energy. Analysis of a draft genome sequence revealed multiple pathways for respiration and carbon utilization. The results of this work demonstrate that alacranite, a mineral previously thought to be formed only chemically under hydrothermal conditions, is precipitated under mesophilic conditions by the metabolically versatile strain MPA-C3. © 2013 John Wiley & Sons Ltd and Society for Applied Microbiology.

Sequence-based genotyping clarifies conflicting historical morphometric and biological data for 5 Eimeria species infecting turkeys.

PubMed

El-Sherry, S; Ogedengbe, M E; Hafeez, M A; Sayf-Al-Din, M; Gad, N; Barta, J R

2015-02-01

Unlike with Eimeria species infecting chickens, specific identification and nomenclature of Eimeria species infecting turkeys is complicated, and in the absence of molecular data, imprecise. In an attempt to reconcile contradictory data reported on oocyst morphometrics and biological descriptions of various Eimeria species infecting turkey, we established single oocyst derived lines of 5 important Eimeria species infecting turkeys, Eimeria meleagrimitis (USMN08-01 strain), Eimeria adenoeides (Guelph strain), Eimeria gallopavonis (Weybridge strain), Eimeria meleagridis (USAR97-01 strain), and Eimeria dispersa (Briston strain). Short portions (514 bp) of mitochondrial cytochrome c oxidase subunit I gene (mt COI) from each were amplified and sequenced. Comparison of these sequences showed sufficient species-specific sequence variation to recommend these short mt COI sequences as species-specific markers. Uniformity of oocyst features (dimensions and oocyst structure) of each pure line was observed. Additional morphological features of the oocysts of these species are described as useful for the microscopic differentiation of these Eimeria species. Combined molecular and morphometric data on these single species lines compared with the original species descriptions and more recent data have helped to clarify some confusing, and sometimes conflicting, features associated with these Eimeria spp. For example, these new data suggest that the KCH and KR strains of E. adenoeides reported previously represent 2 distinct species, E. adenoeides and E. meleagridis, respectively. Likewise, analysis of the Weybridge strain of E. adenoeides, which has long been used as a reference strain in various studies conducted on the pathogenicity of E. adenoeides, indicates that this coccidium is actually a strain of E. gallopavonis. We highly recommend mt COI sequence-based genotyping be incorporated into all studies using Eimeria spp. of turkeys to confirm species identifications and so that any resulting data can be associated correctly with a single named Eimeria species. © 2015 Poultry Science Association Inc.

Complete genome sequence of the Phaeobacter gallaeciensis type strain CIP 105210(T) (= DSM 26640(T) = BS107(T)).

PubMed

Frank, Oliver; Pradella, Silke; Rohde, Manfred; Scheuner, Carmen; Klenk, Hans-Peter; Göker, Markus; Petersen, Jörn

2014-06-15

Phaeobacter gallaeciensis CIP 105210(T) (= DSM 26640(T) = BS107(T)) is the type strain of the species Phaeobacter gallaeciensis. The genus Phaeobacter belongs to the marine Roseobacter group (Rhodobacteraceae, Alphaproteobacteria). Phaeobacter species are effective colonizers of marine surfaces, including frequent associations with eukaryotes. Strain BS107(T) was isolated from a rearing of the scallop Pecten maximus. Here we describe the features of this organism, together with the complete genome sequence, comprising eight circular replicons with a total of 4,448 genes. In addition to a high number of extrachromosomal replicons, the genome contains six genomic island and three putative prophage regions, as well as a hybrid between a plasmid and a circular phage. Phylogenomic analyses confirm previous results, which indicated that the originally reported P. gallaeciensis type-strain deposit DSM 17395 belongs to P. inhibens and that CIP 105210(T) (= DSM 26640(T)) is the sole genome-sequenced representative of P. gallaeciensis.

Genome-scale investigation of phenotypically distinct but nearly clonal Trichoderma strains

PubMed Central

Weld, Richard J.; Cox, Murray P.; Bradshaw, Rosie E.; McLean, Kirstin L.; Stewart, Alison; Steyaert, Johanna M.

2016-01-01

Biological control agents (BCA) are beneficial organisms that are applied to protect plants from pests. Many fungi of the genus Trichoderma are successful BCAs but the underlying mechanisms are not yet fully understood. Trichoderma cf. atroviride strain LU132 is a remarkably effective BCA compared to T. cf. atroviride strain LU140 but these strains were found to be highly similar at the DNA sequence level. This unusual combination of phenotypic variability and high DNA sequence similarity between separately isolated strains prompted us to undertake a genome comparison study in order to identify DNA polymorphisms. We further investigated if the polymorphisms had functional effects on the phenotypes. The two strains were clearly identified as individuals, exhibiting different growth rates, conidiation and metabolism. Superior pathogen control demonstrated by LU132 depended on its faster growth, which is a prerequisite for successful distribution and competition. Genome sequencing identified only one non-synonymous single nucleotide polymorphism (SNP) between the strains. Based on this SNP, we successfully designed and validated an RFLP protocol that can be used to differentiate LU132 from LU140 and other Trichoderma strains. This SNP changed the amino acid sequence of SERF, encoded by the previously undescribed single copy gene “small EDRK-rich factor” (serf). A deletion of serf in the two strains did not lead to identical phenotypes, suggesting that, in addition to the single functional SNP between the nearly clonal Trichoderma cf. atroviride strains, other non-genomic factors contribute to their phenotypic variation. This finding is significant as it shows that genomics is an extremely useful but not exhaustive tool for the study of biocontrol complexity and for strain typing. PMID:27190719

Three novel ascomycetous yeast species of the Kazachstania clade, Kazachstania saulgeensis sp. nov., Kazachstaniaserrabonitensis sp. nov. and Kazachstania australis sp. nov. Reassignment of Candida humilis to Kazachstania humilis f.a. comb. nov. and Candida pseudohumilis to Kazachstania pseudohumilis f.a. comb. nov.

PubMed

Jacques, Noémie; Sarilar, Véronique; Urien, Charlotte; Lopes, Mariana R; Morais, Camila G; Uetanabaro, Ana Paula T; Tinsley, Colin R; Rosa, Carlos A; Sicard, Delphine; Casaregola, Serge

2016-12-01

Five ascosporogenous yeast strains related to the genus Kazachstania were isolated. Two strains (CLIB 1764T and CLIB 1780) were isolated from French sourdoughs; three others (UFMG-CM-Y273T, UFMG-CM-Y451 and UFMG-CM-Y452) were from rotting wood in Brazil. The sequences of the French and Brazilian strains differed by one and three substitutions, respectively, in the D1/D2 large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS). The D1/D2 LSU rRNA sequence of these strains differed by 0.5 and 0.7 % from Kazachstania exigua, but their ITS sequences diverged by 8.1 and 8.3 %, respectively, from that of the closest described species Kazachstania barnettii. Analysis of protein coding sequences of RPB1, RPB2 and EF-1α distinguished the French from the Brazilian strains, with respectively 3.3, 6 and 11.7 % substitutions. Two novel species are described to accommodate these newly isolated strains: Kazachstania saulgeensis sp. nov. (type strain CLIB 1764T=CBS 14374T) and Kazachstania serrabonitensis sp. nov. (type strain UFMG-CM-Y273T=CLIB 1783T=CBS 14236T). Further analysis of culture collections revealed a strain previously assigned to the K. exigua species, but having 3.8 % difference (22 substitutions and 2 indels) in its ITS with respect to K. exigua. Hence, we describe a new taxon, Kazachstania australis sp. nov. (type strain CLIB 162T=CBS 2141T), to accommodate this strain. Finally, Candida humilis and Candida pseudohumilis are reassigned to the genus Kazachstania as new combinations. On the basis of sequence analysis, we also propose that Candida milleri and Kazachstania humilis comb. nov. are conspecific.

Thermophily in the Geobacteraceae: Geothermobacter ehrlichii gen. nov., sp. nov., a novel thermophilic member of the Geobacteraceae from the "Bag City" hydrothermal vent.

PubMed

Kashefi, Kazem; Holmes, Dawn E; Baross, John A; Lovley, Derek R

2003-05-01

Little is known about the microbiology of the "Bag City" hydrothermal vent, which is part of a new eruption site on the Juan de Fuca Ridge and which is notable for its accumulation of polysaccharide on the sediment surface. A pure culture, designated strain SS015, was recovered from a vent fluid sample from the Bag City site through serial dilution in liquid medium with malate as the electron donor and Fe(III) oxide as the electron acceptor and then isolation of single colonies on solid Fe(III) oxide medium. The cells were gram-negative rods, about 0.5 micro m by 1.2 to 1.5 micro m, and motile and contained c-type cytochromes. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain SS015 placed it in the family Geobacteraceae in the delta subclass of the Proteobacteria. Unlike previously described members of the Geobacteraceae, which are mesophiles, strain SS015 was a thermophile and grew at temperatures of between 35 and 65 degrees C, with an optimum temperature of 55 degrees C. Like many previously described members of the Geobacteraceae, strain SS015 grew with organic acids as the electron donors and Fe(III) or nitrate as the electron acceptor, with nitrate being reduced to ammonia. Strain SS015 was unique among the Geobacteraceae in its ability to use sugars, starch, or amino acids as electron donors for Fe(III) reduction. Under stress conditions, strain SS015 produced copious quantities of extracellular polysaccharide, providing a model for the microbial production of the polysaccharide accumulation at the Bag City site. The 16S rDNA sequence of strain SS015 was less than 94% similar to the sequences of previously described members of the Geobacteraceae; this fact, coupled with its unique physiological properties, suggests that strain SS015 represents a new genus in the family Geobacteraceae. The name Geothermobacter ehrlichii gen. nov., sp. nov., is proposed (ATCC BAA-635 and DSM 15274). Although strains of Geobacteraceae are known to be the predominant Fe(III)-reducing microorganisms in a variety of Fe(III)-reducing environments at moderate temperatures, strain SS015 represents the first described thermophilic member of the Geobacteraceae and thus extends the known environmental range of this family to hydrothermal environments.

Whole-genome comparative analysis of three phytopathogenic Xylella fastidiosa strains.

PubMed

Bhattacharyya, Anamitra; Stilwagen, Stephanie; Ivanova, Natalia; D'Souza, Mark; Bernal, Axel; Lykidis, Athanasios; Kapatral, Vinayak; Anderson, Iain; Larsen, Niels; Los, Tamara; Reznik, Gary; Selkov, Eugene; Walunas, Theresa L; Feil, Helene; Feil, William S; Purcell, Alexander; Lassez, Jean-Louis; Hawkins, Trevor L; Haselkorn, Robert; Overbeek, Ross; Predki, Paul F; Kyrpides, Nikos C

2002-09-17

Xylella fastidiosa (Xf) causes wilt disease in plants and is responsible for major economic and crop losses globally. Owing to the public importance of this phytopathogen we embarked on a comparative analysis of the complete genome of Xf pv citrus and the partial genomes of two recently sequenced strains of this species: Xf pv almond and Xf pv oleander, which cause leaf scorch in almond and oleander plants, respectively. We report a reanalysis of the previously sequenced Xf 9a5c (CVC, citrus) strain and the two "gapped" Xf genomes revealing ORFs encoding critical functions in pathogenicity and conjugative transfer. Second, a detailed whole-genome functional comparison was based on the three sequenced Xf strains, identifying the unique genes present in each strain, in addition to those shared between strains. Third, an "in silico" cellular reconstruction of these organisms was made, based on a comparison of their core functional subsystems that led to a characterization of their conjugative transfer machinery, identification of potential differences in their adhesion mechanisms, and highlighting of the absence of a classical quorum-sensing mechanism. This study demonstrates the effectiveness of comparative analysis strategies in the interpretation of genomes that are closely related.

Diversity and Evolution of Mycobacterium tuberculosis: Moving to Whole-Genome-Based Approaches

PubMed Central

Niemann, Stefan; Supply, Philip

2014-01-01

Genotyping of clinical Mycobacterium tuberculosis complex (MTBC) strains has become a standard tool for epidemiological tracing and for the investigation of the local and global strain population structure. Of special importance is the analysis of the expansion of multidrug (MDR) and extensively drug-resistant (XDR) strains. Classical genotyping and, more recently, whole-genome sequencing have revealed that the strains of the MTBC are more diverse than previously anticipated. Globally, several phylogenetic lineages can be distinguished whose geographical distribution is markedly variable. Strains of particular (sub)lineages, such as Beijing, seem to be more virulent and associated with enhanced resistance levels and fitness, likely fueling their spread in certain world regions. The upcoming generalization of whole-genome sequencing approaches will expectedly provide more comprehensive insights into the molecular and epidemiological mechanisms involved and lead to better diagnostic and therapeutic tools. PMID:25190252

Examination of the taxonomic position of Penicillium strains used in blue cheese production based on the partial sequence of β-tubulin.

PubMed

Ogawa, Yoshio; Hirose, Dai; Akiyama, Ayano; Ichinoe, Masakatsu

2014-01-01

Penicillium roqueforti is a well known starter used for blue cheese production. Two closely related species, P. carneum and P. paneum, were previously classified as varieties of P. roqueforti. Penicillium roqueforti does not produce patulin, a mycotoxin harmful for human health, whereas both P. carneum and P. paneum actively produce this toxin. From the viewpoint of food safety, it is thus important to confirm that P. carneum and P. paneum are not used for cheese production. In the present study, the taxonomic position of Penicillium strains used for blue cheese production was examined on the basis of the partial sequence of β-tubulin. Twenty-eight Penicillium strains isolated from blue cheeses were investigated. All the examined strains belonged to P. roqueforti. Therefore, the Penicillium strains used for production of the blue cheese samples examined here do not negatively impact on human health.

Genetic diversity and distribution of a distinct strain of Chili leaf curl virus and associated betasatellite infecting tomato and pepper in Oman.

PubMed

Khan, Akhtar J; Akhtar, Sohail; Al-Zaidi, Amal M; Singh, Achuit K; Briddon, Rob W

2013-10-01

Tomato and pepper are widely grown in Oman for local consumption. A countrywide survey was conducted during 2010-2011 to collect samples and assess the diversity of begomoviruses associated with leaf curl disease of tomato and pepper. A virus previously only identified on the Indian subcontinent, chili leaf curl virus (ChLCV), was found associated with tomato and pepper diseases in all vegetable grown areas of Oman. Some of the infected plant samples were also found to contain a betasatellite. A total of 19 potentially full-length begomovirus and eight betasatellite clones were sequenced. The begomovirus clones showed >96% nucleotide sequence identity, showing them to represent a single species. Comparisons to sequences available in the databases showed the highest levels of nucleotide sequence identity (88.0-91.1%) to isolates of the "Pakistan" strain of ChLCV (ChLCV-PK), indicating the virus from Oman to be a distinct strain, for which the name Oman strain (ChLCV-OM) is proposed. An analysis for recombination showed ChLCV-OM likely to have originated by recombination between ChLCV-PK (the major parent), pepper leaf curl Lahore virus and a third strain of ChLCV. The betasatellite sequences obtained were shown to have high levels of identity to isolates of tomato leaf curl betasatellite (ToLCB) previous shown to be present in Oman. For the disease in tomato Koch's postulates were satisfied by Agrobacterium-mediated inoculation of virus and betasatellites clones. This showed the symptoms induced by the virus in the presence of the betasatellite to be enhanced, although viral DNA levels were not affected. ChLCV-OM is the fourth begomovirus identified in tomato in Oman and the first in Capsicum. The significance of these findings is discussed. Copyright © 2013 Elsevier B.V. All rights reserved.

BIOCHEMICAL AND PHYLOGENETIC CHARACTERIZATION OF TWO NOVEL DEEP-SEA THERMOCOCCUS ISOLATES WITH POTENTIALLY BIOTECHNOLOGICAL APPLICATIONS

EPA Science Inventory

The partial 16S rDNA gene sequences of two thermophilic archaeal strains, TY and TYS, previously isolated from the Guaymas Basin hydrothermal vent site were determined. Lipid analyses and a comparative analysis performed with 16S rDNA sequences of similar thermophilic species sho...

Whole Genome Complete Resequencing of Bacillus subtilis Natto by Combining Long Reads with High-Quality Short Reads

PubMed Central

Kamada, Mayumi; Hase, Sumitaka; Sato, Kengo; Toyoda, Atsushi; Fujiyama, Asao; Sakakibara, Yasubumi

2014-01-01

De novo microbial genome sequencing reached a turning point with third-generation sequencing (TGS) platforms, and several microbial genomes have been improved by TGS long reads. Bacillus subtilis natto is closely related to the laboratory standard strain B. subtilis Marburg 168, and it has a function in the production of the traditional Japanese fermented food “natto.” The B. subtilis natto BEST195 genome was previously sequenced with short reads, but it included some incomplete regions. We resequenced the BEST195 genome using a PacBio RS sequencer, and we successfully obtained a complete genome sequence from one scaffold without any gaps, and we also applied Illumina MiSeq short reads to enhance quality. Compared with the previous BEST195 draft genome and Marburg 168 genome, we found that incomplete regions in the previous genome sequence were attributed to GC-bias and repetitive sequences, and we also identified some novel genes that are found only in the new genome. PMID:25329997

Stable integration and expression of heterologous genes in several lactobacilli using an integration vector constructed from the integrase and attP sequences of phage ΦAT3 isolated from Lactobacillus casei ATCC 393.

PubMed

Lin, Chao-Fen; Lo, Ta-Chun; Kuo, Yang-Cheng; Lin, Thy-Hou

2013-04-01

An integration vector capable of stably integrating and maintaining in the chromosomes of several lactobacilli over hundreds of generations has been constructed. The major integration machinery used is based on the ΦAT3 integrase (int) and attP sequences determined previously. A novel core sequence located at the 3' end of the tRNA(leu) gene is identified in Lactobacillus fermentum ATCC 14931 as the integration target by the integration vector though most of such sequences found in other lactobacilli are similar to that determined previously. Due to the lack of an appropriate attB site in Lactococcus lactis MG1363, the integration vector is found to be unable to integrate into the chromosome of the strain. However, such integration can be successfully restored by cotransforming the integration vector with a replicative one harboring both attB and erythromycin resistance sequences into the strain. Furthermore, the integration vector constructed carries a promoter region of placT from the chromosome of Lactobacillus rhamnosus TCELL-1 which is used to express green fluorescence and luminance protein genes in the lactobacilli studied.

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.

Demonstration that a Klebsiella pneumoniae subsp. pneumoniae isolated from an insect (Nezara viridula) harbors a functional plasmid-borne type IV secretion system

USDA-ARS?s Scientific Manuscript database

Previously, we reported the isolation of Klebsiella pneumoniae subspecies pneumoniae strain Kp 5-1 from a southern green stink bug (Nezara viridula) that is a significant pest of numerous economically important crops. We subsequently sequenced the strains whole genome. Here, we report the presence...

Genome Sequence of Pantoea annatis strain CFH 7-1, which is associated with a vector-borne cotton fruit disease

USDA-ARS?s Scientific Manuscript database

Pantoea ananatis is a bacterium with versatile niches that vary from pathogenic to beneficial. We present the genome of strain CFH 7-1, which was recovered from a diseased greenhouse cotton boll previously caged with a field-collected cotton fleahopper (Pseudatomoscelis seriatus). These data will ...

Simultaneous Differentiation and Typing of Entamoeba histolytica and Entamoeba dispar

PubMed Central

Zaki, Mehreen; Meelu, Parool; Sun, Wei; Clark, C. Graham

2002-01-01

Sequences corresponding to some of the polymorphic loci previously reported from Entamoeba histolytica have been detected in Entamoeba dispar. Comparison of nucleotide sequences of two loci between E. dispar strain SAW760 and E. histolytica strain HM-1:IMSS revealed significant differences in both repeat and flanking regions. The tandem repeat units varied not only in sequence but also in number and arrangement between the two species at both the loci. Using the sequences obtained, primer pairs aimed at amplifying species-specific products were designed and tested on a variety of E. histolytica and E. dispar samples. Amplification results were in complete agreement with the original species classification in all cases, and the PCR products displayed discernible size and pattern variations among the isolates. PMID:11923344

Pantoea ananatis Genetic Diversity Analysis Reveals Limited Genomic Diversity as Well as Accessory Genes Correlated with Onion Pathogenicity.

PubMed

Stice, Shaun P; Stumpf, Spencer D; Gitaitis, Ron D; Kvitko, Brian H; Dutta, Bhabesh

2018-01-01

Pantoea ananatis is a member of the family Enterobacteriaceae and an enigmatic plant pathogen with a broad host range. Although P. ananatis strains can be aggressive on onion causing foliar necrosis and onion center rot, previous genomic analysis has shown that P. ananatis lacks the primary virulence secretion systems associated with other plant pathogens. We assessed a collection of fifty P. ananatis strains collected from Georgia over three decades to determine genetic factors that correlated with onion pathogenic potential. Previous genetic analysis studies have compared strains isolated from different hosts with varying diseases potential and isolation sources. Strains varied greatly in their pathogenic potential and aggressiveness on different cultivated Allium species like onion, leek, shallot, and chive. Using multi-locus sequence analysis (MLSA) and repetitive extragenic palindrome repeat (rep)-PCR techniques, we did not observe any correlation between onion pathogenic potential and genetic diversity among strains. Whole genome sequencing and pan-genomic analysis of a sub-set of 10 strains aided in the identification of a novel series of genetic regions, likely plasmid borne, and correlating with onion pathogenicity observed on single contigs of the genetic assemblies. We named these loci Onion Virulence Regions (OVR) A-D. The OVR loci contain genes involved in redox regulation as well as pectate lyase and rhamnogalacturonase genes. Previous studies have not identified distinct genetic loci or plasmids correlating with onion foliar pathogenicity or pathogenicity on a single host pathosystem. The lack of focus on a single host system for this phytopathgenic disease necessitates the pan-genomic analysis performed in this study.

Pantoea ananatis Genetic Diversity Analysis Reveals Limited Genomic Diversity as Well as Accessory Genes Correlated with Onion Pathogenicity

PubMed Central

Stice, Shaun P.; Stumpf, Spencer D.; Gitaitis, Ron D.; Kvitko, Brian H.; Dutta, Bhabesh

2018-01-01

Pantoea ananatis is a member of the family Enterobacteriaceae and an enigmatic plant pathogen with a broad host range. Although P. ananatis strains can be aggressive on onion causing foliar necrosis and onion center rot, previous genomic analysis has shown that P. ananatis lacks the primary virulence secretion systems associated with other plant pathogens. We assessed a collection of fifty P. ananatis strains collected from Georgia over three decades to determine genetic factors that correlated with onion pathogenic potential. Previous genetic analysis studies have compared strains isolated from different hosts with varying diseases potential and isolation sources. Strains varied greatly in their pathogenic potential and aggressiveness on different cultivated Allium species like onion, leek, shallot, and chive. Using multi-locus sequence analysis (MLSA) and repetitive extragenic palindrome repeat (rep)-PCR techniques, we did not observe any correlation between onion pathogenic potential and genetic diversity among strains. Whole genome sequencing and pan-genomic analysis of a sub-set of 10 strains aided in the identification of a novel series of genetic regions, likely plasmid borne, and correlating with onion pathogenicity observed on single contigs of the genetic assemblies. We named these loci Onion Virulence Regions (OVR) A-D. The OVR loci contain genes involved in redox regulation as well as pectate lyase and rhamnogalacturonase genes. Previous studies have not identified distinct genetic loci or plasmids correlating with onion foliar pathogenicity or pathogenicity on a single host pathosystem. The lack of focus on a single host system for this phytopathgenic disease necessitates the pan-genomic analysis performed in this study. PMID:29491851

Multilocus sequence typing of Pseudomonas syringae sensu lato confirms previously described genomospecies and permits rapid identification of P. syringae pv. coriandricola and P. syringae pv. apii causing bacterial leaf spot on parsley.

PubMed

Bull, Carolee T; Clarke, Christopher R; Cai, Rongman; Vinatzer, Boris A; Jardini, Teresa M; Koike, Steven T

2011-07-01

Since 2002, severe leaf spotting on parsley (Petroselinum crispum) has occurred in Monterey County, CA. Either of two different pathovars of Pseudomonas syringae sensu lato were isolated from diseased leaves from eight distinct outbreaks and once from the same outbreak. Fragment analysis of DNA amplified between repetitive sequence polymerase chain reaction; 16S rDNA sequence analysis; and biochemical, physiological, and host range tests identified the pathogens as Pseudomonas syringae pv. apii and P. syringae pv. coriandricola. Koch's postulates were completed for the isolates from parsley, and host range tests with parsley isolates and pathotype strains demonstrated that P. syringae pv. apii and P. syringae pv. coriandricola cause leaf spot diseases on parsley, celery, and coriander or cilantro. In a multilocus sequence typing (MLST) approach, four housekeeping gene fragments were sequenced from 10 strains isolated from parsley and 56 pathotype strains of P. syringae. Allele sequences were uploaded to the Plant-Associated Microbes Database and a phylogenetic tree was built based on concatenated sequences. Tree topology directly corresponded to P. syringae genomospecies and P. syringae pv. apii was allocated appropriately to genomospecies 3. This is the first demonstration that MLST can accurately allocate new pathogens directly to P. syringae sensu lato genomospecies. According to MLST, P. syringae pv. coriandricola is a member of genomospecies 9, P. cannabina. In a blind test, both P. syringae pv. coriandricola and P. syringae pv. apii isolates from parsley were correctly identified to pathovar. In both cases, MLST described diversity within each pathovar that was previously unknown.

Diagnostics for Yaws Eradication: Insights From Direct Next-Generation Sequencing of Cutaneous Strains of Treponema pallidum.

PubMed

Marks, Michael; Fookes, Maria; Wagner, Josef; Butcher, Robert; Ghinai, Rosanna; Sokana, Oliver; Sarkodie, Yaw-Adu; Lukehart, Sheila A; Solomon, Anthony W; Mabey, David C W; Thomson, Nicholas

2018-03-05

Yaws-like chronic ulcers can be caused by Treponema pallidum subspecies pertenue, Haemophilus ducreyi, or other, still-undefined bacteria. To permit accurate evaluation of yaws elimination efforts, programmatic use of molecular diagnostics is required. The accuracy and sensitivity of current tools remain unclear because our understanding of T. pallidum diversity is limited by the low number of sequenced genomes. We tested samples from patients with suspected yaws collected in the Solomon Islands and Ghana. All samples were from patients whose lesions had previously tested negative using the Centers for Disease Control and Prevention (CDC) diagnostic assay in widespread use. However, some of these patients had positive serological assays for yaws on blood. We used direct whole-genome sequencing to identify T. pallidum subsp pertenue strains missed by the current assay. From 45 Solomon Islands and 27 Ghanaian samples, 11 were positive for T. pallidum DNA using the species-wide quantitative polymerase chain reaction (PCR) assay, from which we obtained 6 previously undetected T. pallidum subsp pertenue whole-genome sequences. These show that Solomon Islands sequences represent distinct T. pallidum subsp pertenue clades. These isolates were invisible to the CDC diagnostic PCR assay, due to sequence variation in the primer binding site. Our data double the number of published T. pallidum subsp pertenue genomes. We show that Solomon Islands strains are undetectable by the PCR used in many studies and by health ministries. This assay is therefore not adequate for the eradication program. Next-generation genome sequence data are essential for these efforts. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Delineation of the Species Haemophilus influenzae by Phenotype, Multilocus Sequence Phylogeny, and Detection of Marker Genes▿ †

PubMed Central

Nørskov-Lauritsen, Niels; Overballe, Merete D.; Kilian, Mogens

2009-01-01

To obtain more information on the much-debated definition of prokaryotic species, we investigated the borders of Haemophilus influenzae by comparative analysis of H. influenzae reference strains with closely related bacteria including strains assigned to Haemophilus haemolyticus, cryptic genospecies biotype IV, and the never formally validated species “Haemophilus intermedius”. Multilocus sequence phylogeny based on six housekeeping genes separated a cluster encompassing the type and the reference strains of H. influenzae from 31 more distantly related strains. Comparison of 16S rRNA gene sequences supported this delineation but was obscured by a conspicuously high number of polymorphic sites in many of the strains that did not belong to the core group of H. influenzae strains. The division was corroborated by the differential presence of genes encoding H. influenzae adhesion and penetration protein, fuculokinase, and Cu,Zn-superoxide dismutase, whereas immunoglobulin A1 protease activity or the presence of the iga gene was of limited discriminatory value. The existence of porphyrin-synthesizing strains (“H. intermedius”) closely related to H. influenzae was confirmed. Several chromosomally encoded hemin biosynthesis genes were identified, and sequence analysis showed these genes to represent an ancestral genotype rather than recent transfers from, e.g., Haemophilus parainfluenzae. Strains previously assigned to H. haemolyticus formed several separate lineages within a distinct but deeply branching cluster, intermingled with strains of “H. intermedius” and cryptic genospecies biotype IV. Although H. influenzae is phenotypically more homogenous than some other Haemophilus species, the genetic diversity and multicluster structure of strains traditionally associated with H. influenzae make it difficult to define the natural borders of that species. PMID:19060144

Complete genome sequence of Dehalogenimonas lykanthroporepellens type strain (BL-DC-9T) and comparison to Dehalococcoides strains

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

Siddaramappa, Shivakumara; Delano, Susana; Green, Lance D.

2012-01-01

Dehalogenimonas lykanthroporepellens is the type species of the genus Dehalogenimonas, which belongs to a deeply branching lineage within the phylum Chloroflexi. This strictly anaerobic, mesophilic, non spore forming, Gram negative staining bacterium was first isolated from chlorinated solvent contaminated groundwater at a Superfund site located near Baton Rouge, Louisiana, USA. D. lykanthroporepellens was of interest for genome sequencing for two reasons: (a) its unusual ability to couple growth with reductive dechlorination of environmentally important polychlorinated aliphatic alkanes and (b) its phylogenetic position distant from previously sequenced bacteria. The 1,686,510 bp circular chromosome of strain BL-DC-9{sup T} contains 1,720 predicted proteinmore » coding genes, 47 tRNA genes, a single large subunit rRNA (23S-5S) locus, and a single, orphan, small unit rRNA (16S) locus.« less

Genome Analysis of Listeria monocytogenes Sequence Type 8 Strains Persisting in Salmon and Poultry Processing Environments and Comparison with Related Strains

PubMed Central

Fagerlund, Annette; Langsrud, Solveig; Schirmer, Bjørn C. T.; Møretrø, Trond; Heir, Even

2016-01-01

Listeria monocytogenes is an important foodborne pathogen responsible for the disease listeriosis, and can be found throughout the environment, in many foods and in food processing facilities. The main cause of listeriosis is consumption of food contaminated from sources in food processing environments. Persistence in food processing facilities has previously been shown for the L. monocytogenes sequence type (ST) 8 subtype. In the current study, five ST8 strains were subjected to whole-genome sequencing and compared with five additionally available ST8 genomes, allowing comparison of strains from salmon, poultry and cheese industry, in addition to a human clinical isolate. Genome-wide analysis of single-nucleotide polymorphisms (SNPs) confirmed that almost identical strains were detected in a Danish salmon processing plant in 1996 and in a Norwegian salmon processing plant in 2001 and 2011. Furthermore, we show that L. monocytogenes ST8 was likely to have been transferred between two poultry processing plants as a result of relocation of processing equipment. The SNP data were used to infer the phylogeny of the ST8 strains, separating them into two main genetic groups. Within each group, the plasmid and prophage content was almost entirely conserved, but between groups, these sequences showed strong divergence. The accessory genome of the ST8 strains harbored genetic elements which could be involved in rendering the ST8 strains resilient to incoming mobile genetic elements. These included two restriction-modification loci, one of which was predicted to show phase variable recognition sequence specificity through site-specific domain shuffling. Analysis indicated that the ST8 strains harbor all important known L. monocytogenes virulence factors, and ST8 strains are commonly identified as the causative agents of invasive listeriosis. Therefore, the persistence of this L. monocytogenes subtype in food processing facilities poses a significant concern for food safety. PMID:26953695

Genotypic and phenotypic characterization of Cryptococcus flavescens, beneficial biocontrol agents for controlling Fusarium head blight of wheat

USDA-ARS?s Scientific Manuscript database

Cryptococcus flavescens strain OH182.9_3C (3C) previously displayed significant biological control activity against Fusarium head blight, a globally important disease of wheat; however, the diversity within C. flavescens has not been previously characterized. Multilocus sequence typing was performed...

Human Treponema pallidum 11q/j isolate belongs to subsp. endemicum but contains two loci with a sequence in TP0548 and TP0488 similar to subsp. pertenue and subsp. pallidum, respectively

PubMed Central

Mikalová, Lenka; Strouhal, Michal; Oppelt, Jan; Grange, Philippe Alain; Janier, Michel; Benhaddou, Nadjet; Dupin, Nicolas; Šmajs, David

2017-01-01

Background Treponema pallidum subsp. endemicum (TEN) is the causative agent of endemic syphilis (bejel). An unusual human TEN 11q/j isolate was obtained from a syphilis-like primary genital lesion from a patient that returned to France from Pakistan. Methodology/Principal findings The TEN 11q/j isolate was characterized using nested PCR followed by Sanger sequencing and/or direct Illumina sequencing. Altogether, 44 chromosomal regions were analyzed. Overall, the 11q/j isolate clustered with TEN strains Bosnia A and Iraq B as expected from previous TEN classification of the 11q/j isolate. However, the 11q/j sequence in a 505 bp-long region at the TP0488 locus was similar to Treponema pallidum subsp. pallidum (TPA) strains, but not to TEN Bosnia A and Iraq B sequences, suggesting a recombination event at this locus. Similarly, the 11q/j sequence in a 613 bp-long region at the TP0548 locus was similar to Treponema pallidum subsp. pertenue (TPE) strains, but not to TEN sequences. Conclusions/Significance A detailed analysis of two recombinant loci found in the 11q/j clinical isolate revealed that the recombination event occurred just once, in the TP0488, with the donor sequence originating from a TPA strain. Since TEN Bosnia A and Iraq B were found to contain TPA-like sequences at the TP0548 locus, the recombination at TP0548 took place in a treponeme that was an ancestor to both TEN Bosnia A and Iraq B. The sequence of 11q/j isolate in TP0548 represents an ancestral TEN sequence that is similar to yaws-causing treponemes. In addition to the importance of the 11q/j isolate for reconstruction of the TEN phylogeny, this case emphasizes the possible role of TEN strains in development of syphilis-like lesions. PMID:28263990

High-resolution community profiling of arbuscular mycorrhizal fungi.

PubMed

Schlaeppi, Klaus; Bender, S Franz; Mascher, Fabio; Russo, Giancarlo; Patrignani, Andrea; Camenzind, Tessa; Hempel, Stefan; Rillig, Matthias C; van der Heijden, Marcel G A

2016-11-01

Community analyses of arbuscular mycorrhizal fungi (AMF) using ribosomal small subunit (SSU) or internal transcribed spacer (ITS) DNA sequences often suffer from low resolution or coverage. We developed a novel sequencing based approach for a highly resolving and specific profiling of AMF communities. We took advantage of previously established AMF-specific PCR primers that amplify a c. 1.5-kb long fragment covering parts of SSU, ITS and parts of the large ribosomal subunit (LSU), and we sequenced the resulting amplicons with single molecule real-time (SMRT) sequencing. The method was applicable to soil and root samples, detected all major AMF families and successfully discriminated closely related AMF species, which would not be discernible using SSU sequences. In inoculation tests we could trace the introduced AMF inoculum at the molecular level. One of the introduced strains almost replaced the local strain(s), revealing that AMF inoculation can have a profound impact on the native community. The methodology presented offers researchers a powerful new tool for AMF community analysis because it unifies improved specificity and enhanced resolution, whereas the drawback of medium sequencing throughput appears of lesser importance for low-diversity groups such as AMF. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

Erwinia amylovora CRISPR Elements Provide New Tools for Evaluating Strain Diversity and for Microbial Source Tracking

PubMed Central

McGhee, Gayle C.; Sundin, George W.

2012-01-01

Clustered regularly interspaced short palindromic repeats (CRISPRs) comprise a family of short DNA repeat sequences that are separated by non repetitive spacer sequences and, in combination with a suite of Cas proteins, are thought to function as an adaptive immune system against invading DNA. The number of CRISPR arrays in a bacterial chromosome is variable, and the content of each array can differ in both repeat number and in the presence or absence of specific spacers. We utilized a comparative sequence analysis of CRISPR arrays of the plant pathogen Erwinia amylovora to uncover previously unknown genetic diversity in this species. A total of 85 E. amylovora strains varying in geographic isolation (North America, Europe, New Zealand, and the Middle East), host range, plasmid content, and streptomycin sensitivity/resistance were evaluated for CRISPR array number and spacer variability. From these strains, 588 unique spacers were identified in the three CRISPR arrays present in E. amylovora, and these arrays could be categorized into 20, 17, and 2 patterns types, respectively. Analysis of the relatedness of spacer content differentiated most apple and pear strains isolated in the eastern U.S. from western U.S. strains. In addition, we identified North American strains that shared CRISPR genotypes with strains isolated on other continents. E. amylovora strains from Rubus and Indian hawthorn contained mostly unique spacers compared to apple and pear strains, while strains from loquat shared 79% of spacers with apple and pear strains. Approximately 23% of the spacers matched known sequences, with 16% targeting plasmids and 5% targeting bacteriophage. The plasmid pEU30, isolated in E. amylovora strains from the western U.S., was targeted by 55 spacers. Lastly, we used spacer patterns and content to determine that streptomycin-resistant strains of E. amylovora from Michigan were low in diversity and matched corresponding streptomycin-sensitive strains from the background population. PMID:22860008

Erwinia amylovora CRISPR elements provide new tools for evaluating strain diversity and for microbial source tracking.

PubMed

McGhee, Gayle C; Sundin, George W

2012-01-01

Clustered regularly interspaced short palindromic repeats (CRISPRs) comprise a family of short DNA repeat sequences that are separated by non repetitive spacer sequences and, in combination with a suite of Cas proteins, are thought to function as an adaptive immune system against invading DNA. The number of CRISPR arrays in a bacterial chromosome is variable, and the content of each array can differ in both repeat number and in the presence or absence of specific spacers. We utilized a comparative sequence analysis of CRISPR arrays of the plant pathogen Erwinia amylovora to uncover previously unknown genetic diversity in this species. A total of 85 E. amylovora strains varying in geographic isolation (North America, Europe, New Zealand, and the Middle East), host range, plasmid content, and streptomycin sensitivity/resistance were evaluated for CRISPR array number and spacer variability. From these strains, 588 unique spacers were identified in the three CRISPR arrays present in E. amylovora, and these arrays could be categorized into 20, 17, and 2 patterns types, respectively. Analysis of the relatedness of spacer content differentiated most apple and pear strains isolated in the eastern U.S. from western U.S. strains. In addition, we identified North American strains that shared CRISPR genotypes with strains isolated on other continents. E. amylovora strains from Rubus and Indian hawthorn contained mostly unique spacers compared to apple and pear strains, while strains from loquat shared 79% of spacers with apple and pear strains. Approximately 23% of the spacers matched known sequences, with 16% targeting plasmids and 5% targeting bacteriophage. The plasmid pEU30, isolated in E. amylovora strains from the western U.S., was targeted by 55 spacers. Lastly, we used spacer patterns and content to determine that streptomycin-resistant strains of E. amylovora from Michigan were low in diversity and matched corresponding streptomycin-sensitive strains from the background population.

An axenic culture system for fruiting body formation by an edible bolete phylogenetically related to culinary-medicinal penny bun mushroom, Boletus edulis Bull.:Fr. strains from China.

PubMed

Fu, Shao Chun; Zhang, Mei Yan; Shang, Xiao Dong; Chen, Ming Jie; Tan, Qi

2011-01-01

The ability of two freshly isolated Boletus stains to fruit under axenic conditions was tested using different solid and liquid nutrient media. One strain (YNCX04) produced numerous primordia from which fruiting bodies, 12 mm and 10 mm in length, with grey, convex pilei, and yellow-white, clavate stipes developed between 15 and 30 d after inoculation of fungal mycelium onto a solid medium consisting of mineral salts, thiamine, glucose, potato, an extract of Cunninghamia lanceolata root, and agar. The other strain (YNB200) produced numerous primordia but no sporophores. Strain YNCX04 lost the ability to form fruiting bodies in axenic culture 6 mo after initial isolation but retained the ability to form primordia for up to 18 mo. Based on internal transcribed spacer sequencing data, strains YNB200 and YNCX04 formed a sub-cluster together with four previously designated Boletus edulis strains from China. Phylogenetic analysis placed the Chinese strains closer to B. aestivalis than to European and North American strains of B. edulis, although a 29-bp fragment specific to all the B. aestivalis strains was absent from all the Chinese strains. Furthermore, partial 18S rDNA sequences from strains YNB200 and YNCX04 exhibited 98% similarity with an 18S rDNA sequence from B. edulis strain Be3. Further molecular studies are indicated to more accurately establish the taxonomic positions ofF3 and F4-3, as well as the Chinese strains designated as B. edulis.

Deconvoluting simulated metagenomes: the performance of hard- and soft- clustering algorithms applied to metagenomic chromosome conformation capture (3C)

PubMed Central

DeMaere, Matthew Z.

2016-01-01

Background Chromosome conformation capture, coupled with high throughput DNA sequencing in protocols like Hi-C and 3C-seq, has been proposed as a viable means of generating data to resolve the genomes of microorganisms living in naturally occuring environments. Metagenomic Hi-C and 3C-seq datasets have begun to emerge, but the feasibility of resolving genomes when closely related organisms (strain-level diversity) are present in the sample has not yet been systematically characterised. Methods We developed a computational simulation pipeline for metagenomic 3C and Hi-C sequencing to evaluate the accuracy of genomic reconstructions at, above, and below an operationally defined species boundary. We simulated datasets and measured accuracy over a wide range of parameters. Five clustering algorithms were evaluated (2 hard, 3 soft) using an adaptation of the extended B-cubed validation measure. Results When all genomes in a sample are below 95% sequence identity, all of the tested clustering algorithms performed well. When sequence data contains genomes above 95% identity (our operational definition of strain-level diversity), a naive soft-clustering extension of the Louvain method achieves the highest performance. Discussion Previously, only hard-clustering algorithms have been applied to metagenomic 3C and Hi-C data, yet none of these perform well when strain-level diversity exists in a metagenomic sample. Our simple extension of the Louvain method performed the best in these scenarios, however, accuracy remained well below the levels observed for samples without strain-level diversity. Strain resolution is also highly dependent on the amount of available 3C sequence data, suggesting that depth of sequencing must be carefully considered during experimental design. Finally, there appears to be great scope to improve the accuracy of strain resolution through further algorithm development. PMID:27843713

Molecular analysis of an oyster-related norovirus outbreak.

PubMed

Nenonen, Nancy P; Hannoun, Charles; Olsson, Margareta B; Bergström, Tomas

2009-06-01

Contaminated raw oysters were implicated in a severe outbreak of norovirus (NoV) gastroenteritis affecting 30 restaurant guests. To define the outbreak source by using molecular methods to characterize NoV strains detected in patient and oyster samples. Molecular epidemiological studies based on nucleotide sequencing and phylogenetic analyses of patient and oyster NoV strains, and comparison to background dataset. NoV genotype (G) I.1 was detected in the one patient stool analyzed by in-house TaqMan real time RT-PCR and classical nested RT-PCR targeting NoV RNA-dependent polymerase (RdRp, 285 nt), and by nested RT-PCR targeting RdRp-capsid-poly(A)-3' (3085 nt). Patient strain showed >or=99% similarity (285 nt) with three NoV strains detected in two of five oysters examined by classical nested RT-PCR (RdRp). A third oyster tested positive for NoV GII.3. Phylogenetic analysis showed clustering of patient and oyster strains related to this outbreak with GI.1 strains from previous local outbreaks, and mussel studies. Sequence data revealed >or=99% similarity (285 nt) between NoV GI.1 strains detected in patient stool and suspect oysters, linking the contaminated oysters to the outbreak. Identification of human NoV GI and GII strains in oysters indicated contamination of human fecal origin, presumably from inappropriate storage in the harbor. Comparative long-fragment analysis of the patient strain revealed 99% similarity (3085 nt) with NoV GI.1 strains detected in previous outbreaks and environmental mussel studies from West Sweden, 87% with M87661 (Norwalk68) and 96% with L23828 (SRSV-KY-89/89/J). These results indicated considerable genomic stability of NoV GI.1 strains over time.

Complete genome sequence of Lactobacillus salivarius Ren, a probiotic strain with anti-tumor activity.

PubMed

Sun, Erna; Ren, Fazheng; Liu, Songling; Ge, Shaoyang; Zhang, Ming; Guo, Huiyuan; Jiang, Lu; Zhang, Hao; Zhao, Liang

2015-09-20

Lactobacillus salivarius Ren (LsR) (CGMCC No. 3606) is a probiotic strain that was isolated from the feces of a healthy centenarian living in Bama, Guangxi, China. Previous studies have shown that this strain decreases 4-nitroquinoline 1-oxide (4-NQO)-induced genotoxicity in vitro. It also suppresses 4-NQO-induced oral carcinogenesis and 1,2-dimethylhydrazine (DMH)-induced colorectal carcinogenesis, and therefore may be used as an adjuvant therapeutic agent for cancer. Here, we report the complete genome sequence of LsR that consists of a circular chromosome of 1751,565 bp and two plasmids (pR1, 176,951 bp; pR2, 49,848 bp). Copyright © 2015 Elsevier B.V. All rights reserved.

Tracing outbreaks of Streptococcus equi infection (strangles) in horses using sequence variation in the seM gene and pulsed-field gel electrophoresis.

PubMed

Lindahl, Susanne; Söderlund, Robert; Frosth, Sara; Pringle, John; Båverud, Viveca; Aspán, Anna

2011-11-21

Strangles is a serious respiratory disease in horses caused by Streptococcus equi subspecies equi (S. equi). Transmission of the disease occurs by direct contact with an infected horse or contaminated equipment. Genetically, S. equi strains are highly homogenous and differentiation of strains has proven difficult. However, the S. equi M-protein SeM contains a variable N-terminal region and has been proposed as a target gene to distinguish between different strains of S. equi and determine the source of an outbreak. In this study, strains of S. equi (n=60) from 32 strangles outbreaks in Sweden during 1998-2003 and 2008-2009 were genetically characterized by sequencing the SeM protein gene (seM), and by pulsed-field gel electrophoresis (PFGE). Swedish strains belonged to 10 different seM types, of which five have not previously been described. Most were identical or highly similar to allele types from strangles outbreaks in the UK. Outbreaks in 2008/2009 sharing the same seM type were associated by geographic location and/or type of usage of the horses (racing stables). Sequencing of the seM gene generally agreed with pulsed-field gel electrophoresis profiles. Our data suggest that seM sequencing as a epidemiological tool is supported by the agreement between seM and PFGE and that sequencing of the SeM protein gene is more sensitive than PFGE in discriminating strains of S. equi. Copyright © 2011 Elsevier B.V. All rights reserved.

Sensitive Next-Generation Sequencing Method Reveals Deep Genetic Diversity of HIV-1 in the Democratic Republic of the Congo.

PubMed

Rodgers, Mary A; Wilkinson, Eduan; Vallari, Ana; McArthur, Carole; Sthreshley, Larry; Brennan, Catherine A; Cloherty, Gavin; de Oliveira, Tulio

2017-03-15

As the epidemiological epicenter of the human immunodeficiency virus (HIV) pandemic, the Democratic Republic of the Congo (DRC) is a reservoir of circulating HIV strains exhibiting high levels of diversity and recombination. In this study, we characterized HIV specimens collected in two rural areas of the DRC between 2001 and 2003 to identify rare strains of HIV. The env gp41 region was sequenced and characterized for 172 HIV-positive specimens. The env sequences were predominantly subtype A (43.02%), but 7 other subtypes (33.14%), 20 circulating recombinant forms (CRFs; 11.63%), and 20 unclassified (11.63%) sequences were also found. Of the rare and unclassified subtypes, 18 specimens were selected for next-generation sequencing (NGS) by a modified HIV-switching mechanism at the 5' end of the RNA template (SMART) method to obtain full-genome sequences. NGS produced 14 new complete genomes, which included pure subtype C ( n = 2), D ( n = 1), F1 ( n = 1), H ( n = 3), and J ( n = 1) genomes. The two subtype C genomes and one of the subtype H genomes branched basal to their respective subtype branches but had no evidence of recombination. The remaining 6 genomes were complex recombinants of 2 or more subtypes, including subtypes A1, F, G, H, J, and K and unclassified fragments, including one subtype CRF25 isolate, which branched basal to all CRF25 references. Notably, all recombinant subtype H fragments branched basal to the H clade. Spatial-geographical analysis indicated that the diverse sequences identified here did not expand globally. The full-genome and subgenomic sequences identified in our study population significantly increase the documented diversity of the strains involved in the continually evolving HIV-1 pandemic. IMPORTANCE Very little is known about the ancestral HIV-1 strains that founded the global pandemic, and very few complete genome sequences are available from patients in the Congo Basin, where HIV-1 expanded early in the global pandemic. By sequencing a subgenomic fragment of the HIV-1 envelope from study participants in the DRC, we identified rare variants for complete genome sequencing. The basal branching of some of the complete genome sequences that we recovered suggests that these strains are more closely related to ancestral HIV-1 strains than to previously reported strains and is evidence that the local diversification of HIV in the DRC continues to outpace the diversity of global strains decades after the emergence of the pandemic. Copyright © 2017 Rodgers et al.

Origins of the E. coli Strain Causing an Outbreak of Hemolytic–Uremic Syndrome in Germany

PubMed Central

Rasko, David A.; Webster, Dale R.; Sahl, Jason W.; Bashir, Ali; Boisen, Nadia; Scheutz, Flemming; Paxinos, Ellen E.; Sebra, Robert; Chin, Chen-Shan; Iliopoulos, Dimitris; Klammer, Aaron; Peluso, Paul; Lee, Lawrence; Kislyuk, Andrey O.; Bullard, James; Kasarskis, Andrew; Wang, Susanna; Eid, John; Rank, David; Redman, Julia C.; Steyert, Susan R.; Frimodt-Møller, Jakob; Struve, Carsten; Petersen, Andreas M.; Krogfelt, Karen A.; Nataro, James P.; Schadt, Eric E.; Waldor, Matthew K.

2011-01-01

BACKGROUND A large outbreak of diarrhea and the hemolytic–uremic syndrome caused by an unusual serotype of Shiga-toxin–producing Escherichia coli (O104:H4) began in Germany in May 2011. As of July 22, a large number of cases of diarrhea caused by Shiga-toxin–producing E. coli have been reported — 3167 without the hemolytic–uremic syndrome (16 deaths) and 908 with the hemolytic–uremic syndrome (34 deaths) — indicating that this strain is notably more virulent than most of the Shiga-toxin–producing E. coli strains. Preliminary genetic characterization of the outbreak strain suggested that, unlike most of these strains, it should be classified within the enteroaggregative pathotype of E. coli. METHODS We used third-generation, single-molecule, real-time DNA sequencing to determine the complete genome sequence of the German outbreak strain, as well as the genome sequences of seven diarrhea-associated enteroaggregative E. coli serotype O104:H4 strains from Africa and four enteroaggregative E. coli reference strains belonging to other serotypes. Genomewide comparisons were performed with the use of these enteroaggregative E. coli genomes, as well as those of 40 previously sequenced E. coli isolates. RESULTS The enteroaggregative E. coli O104:H4 strains are closely related and form a distinct clade among E. coli and enteroaggregative E. coli strains. However, the genome of the German outbreak strain can be distinguished from those of other O104:H4 strains because it contains a prophage encoding Shiga toxin 2 and a distinct set of additional virulence and antibiotic-resistance factors. CONCLUSIONS Our findings suggest that horizontal genetic exchange allowed for the emergence of the highly virulent Shiga-toxin–producing enteroaggregative E. coli O104:H4 strain that caused the German outbreak. More broadly, these findings highlight the way in which the plasticity of bacterial genomes facilitates the emergence of new pathogens. PMID:21793740

Enhanced arbovirus surveillance with deep sequencing: Identification of novel rhabdoviruses and bunyaviruses in Australian mosquitoes.

PubMed

Coffey, Lark L; Page, Brady L; Greninger, Alexander L; Herring, Belinda L; Russell, Richard C; Doggett, Stephen L; Haniotis, John; Wang, Chunlin; Deng, Xutao; Delwart, Eric L

2014-01-05

Viral metagenomics characterizes known and identifies unknown viruses based on sequence similarities to any previously sequenced viral genomes. A metagenomics approach was used to identify virus sequences in Australian mosquitoes causing cytopathic effects in inoculated mammalian cell cultures. Sequence comparisons revealed strains of Liao Ning virus (Reovirus, Seadornavirus), previously detected only in China, livestock-infecting Stretch Lagoon virus (Reovirus, Orbivirus), two novel dimarhabdoviruses, named Beaumont and North Creek viruses, and two novel orthobunyaviruses, named Murrumbidgee and Salt Ash viruses. The novel virus proteomes diverged by ≥ 50% relative to their closest previously genetically characterized viral relatives. Deep sequencing also generated genomes of Warrego and Wallal viruses, orbiviruses linked to kangaroo blindness, whose genomes had not been fully characterized. This study highlights viral metagenomics in concert with traditional arbovirus surveillance to characterize known and new arboviruses in field-collected mosquitoes. Follow-up epidemiological studies are required to determine whether the novel viruses infect humans. © 2013 Elsevier Inc. All rights reserved.

Molecular epidemiology of drug-resistant Neisseria gonorrhoeae in Russia (Current Status, 2015).

PubMed

Kubanov, Alexey; Vorobyev, Denis; Chestkov, Aleksandr; Leinsoo, Arvo; Shaskolskiy, Boris; Dementieva, Ekaterina; Solomka, Viktoria; Plakhova, Xenia; Gryadunov, Dmitry; Deryabin, Dmitriy

2016-08-09

The widespread distribution of Neisseria gonorrhoeae strains that are resistant to previously used and clinically implemented antibiotics is a significant global public health problem. In line with WHO standards, the national Gonococcal Antimicrobial Surveillance Programme (RU-GASP) has been in existence in Russia since 2004; herein, the current status (2015) is described, including associations between N. gonorrhoeae antimicrobial susceptibility, primary genetic resistance determinants and specific strain sequence types. A total of 124 N. gonorrhoeae strains obtained from 9 regions in Russia in 2015 were examined using N. gonorrhoeae Multi-Antigen Sequence Typing (NG-MAST), an antimicrobial susceptibility test according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria and an oligonucleotide microarray for the identification of mutations in the penA, ponA, rpsJ, gyrA and parC genes responsible for penicillin G, tetracycline, and fluoroquinolone resistance. Genogroup (G) isolates were evaluated based on their porB and tbpB sequence types (STs). NG-MAST analysis showed a diversified population of N. gonorrhoeae in Russia with 58 sequence types, 35 of which were described for the first time. The STs 807, 1544, 1993, 5714, 9476 and 12531, which were typical for some Russian Federation regions and several countries of the former Soviet Union, were represented by five or more isolates. The internationally widespread ST 1407 was represented by a single strain in the present study. Division into genogroups facilitated an exploration of the associations between N. gonorrhoeae sequence type, antimicrobial resistance spectra and genetic resistance determinant contents. Preliminarily susceptible (G-807, G-12531) and resistant (G-5714, G-9476) genogroups were revealed. The variability in the most frequently observed STs and genogroups in each participating region indicated geographically restricted antimicrobial susceptibility in N. gonorrhoeae populations. Resistance or intermediate susceptibility to previously recommended antimicrobials, such as penicillin G (60.5 %), ciprofloxacin (41.1 %) and tetracycline (25 %), is common in the N. gonorrhoeae population. Based on previous reports and current data, ceftriaxone and spectinomycin should be recommended for first-line empiric antimicrobial monotherapy for gonorrhoea in Russia.

The genome sequence of E. coli W (ATCC 9637): comparative genome analysis and an improved genome-scale reconstruction of E. coli

PubMed Central

2011-01-01

Background Escherichia coli is a model prokaryote, an important pathogen, and a key organism for industrial biotechnology. E. coli W (ATCC 9637), one of four strains designated as safe for laboratory purposes, has not been sequenced. E. coli W is a fast-growing strain and is the only safe strain that can utilize sucrose as a carbon source. Lifecycle analysis has demonstrated that sucrose from sugarcane is a preferred carbon source for industrial bioprocesses. Results We have sequenced and annotated the genome of E. coli W. The chromosome is 4,900,968 bp and encodes 4,764 ORFs. Two plasmids, pRK1 (102,536 bp) and pRK2 (5,360 bp), are also present. W has unique features relative to other sequenced laboratory strains (K-12, B and Crooks): it has a larger genome and belongs to phylogroup B1 rather than A. W also grows on a much broader range of carbon sources than does K-12. A genome-scale reconstruction was developed and validated in order to interrogate metabolic properties. Conclusions The genome of W is more similar to commensal and pathogenic B1 strains than phylogroup A strains, and therefore has greater utility for comparative analyses with these strains. W should therefore be the strain of choice, or 'type strain' for group B1 comparative analyses. The genome annotation and tools created here are expected to allow further utilization and development of E. coli W as an industrial organism for sucrose-based bioprocesses. Refinements in our E. coli metabolic reconstruction allow it to more accurately define E. coli metabolism relative to previous models. PMID:21208457

Chromosomal Copy Number Variation in Saccharomyces pastorianus Is Evidence for Extensive Genome Dynamics in Industrial Lager Brewing Strains.

PubMed

van den Broek, M; Bolat, I; Nijkamp, J F; Ramos, E; Luttik, M A H; Koopman, F; Geertman, J M; de Ridder, D; Pronk, J T; Daran, J-M

2015-09-01

Lager brewing strains of Saccharomyces pastorianus are natural interspecific hybrids originating from the spontaneous hybridization of Saccharomyces cerevisiae and Saccharomyces eubayanus. Over the past 500 years, S. pastorianus has been domesticated to become one of the most important industrial microorganisms. Production of lager-type beers requires a set of essential phenotypes, including the ability to ferment maltose and maltotriose at low temperature, the production of flavors and aromas, and the ability to flocculate. Understanding of the molecular basis of complex brewing-related phenotypic traits is a prerequisite for rational strain improvement. While genome sequences have been reported, the variability and dynamics of S. pastorianus genomes have not been investigated in detail. Here, using deep sequencing and chromosome copy number analysis, we showed that S. pastorianus strain CBS1483 exhibited extensive aneuploidy. This was confirmed by quantitative PCR and by flow cytometry. As a direct consequence of this aneuploidy, a massive number of sequence variants was identified, leading to at least 1,800 additional protein variants in S. pastorianus CBS1483. Analysis of eight additional S. pastorianus strains revealed that the previously defined group I strains showed comparable karyotypes, while group II strains showed large interstrain karyotypic variability. Comparison of three strains with nearly identical genome sequences revealed substantial chromosome copy number variation, which may contribute to strain-specific phenotypic traits. The observed variability of lager yeast genomes demonstrates that systematic linking of genotype to phenotype requires a three-dimensional genome analysis encompassing physical chromosomal structures, the copy number of individual chromosomes or chromosomal regions, and the allelic variation of copies of individual genes. Copyright © 2015, van den Broek et al.

Chromosomal Copy Number Variation in Saccharomyces pastorianus Is Evidence for Extensive Genome Dynamics in Industrial Lager Brewing Strains

PubMed Central

van den Broek, M.; Bolat, I.; Nijkamp, J. F.; Ramos, E.; Luttik, M. A. H.; Koopman, F.; Geertman, J. M.; de Ridder, D.; Pronk, J. T.

2015-01-01

Lager brewing strains of Saccharomyces pastorianus are natural interspecific hybrids originating from the spontaneous hybridization of Saccharomyces cerevisiae and Saccharomyces eubayanus. Over the past 500 years, S. pastorianus has been domesticated to become one of the most important industrial microorganisms. Production of lager-type beers requires a set of essential phenotypes, including the ability to ferment maltose and maltotriose at low temperature, the production of flavors and aromas, and the ability to flocculate. Understanding of the molecular basis of complex brewing-related phenotypic traits is a prerequisite for rational strain improvement. While genome sequences have been reported, the variability and dynamics of S. pastorianus genomes have not been investigated in detail. Here, using deep sequencing and chromosome copy number analysis, we showed that S. pastorianus strain CBS1483 exhibited extensive aneuploidy. This was confirmed by quantitative PCR and by flow cytometry. As a direct consequence of this aneuploidy, a massive number of sequence variants was identified, leading to at least 1,800 additional protein variants in S. pastorianus CBS1483. Analysis of eight additional S. pastorianus strains revealed that the previously defined group I strains showed comparable karyotypes, while group II strains showed large interstrain karyotypic variability. Comparison of three strains with nearly identical genome sequences revealed substantial chromosome copy number variation, which may contribute to strain-specific phenotypic traits. The observed variability of lager yeast genomes demonstrates that systematic linking of genotype to phenotype requires a three-dimensional genome analysis encompassing physical chromosomal structures, the copy number of individual chromosomes or chromosomal regions, and the allelic variation of copies of individual genes. PMID:26150454

Singular over-representation of an octameric palindrome, HIP1, in DNA from many cyanobacteria.

PubMed

Robinson, N J; Robinson, P J; Gupta, A; Bleasby, A J; Whitton, B A; Morby, A P

1995-03-11

An octameric palindrome (5'-GCGATCGC-3') is abundant in cyanobacterial sequences within databases (GenBank/EMBL) and was designated HIP1 (highly iterated palindrome). The frequency of occurrence of all 256 octameric palindromes has now been determined in sub-databases revealing large and unique over-representation of HIP1 in cyanobacterial entries. DNA sequences from other bacteria were searched for any over-represented octameric palindromes analogous to HIP1. Only two sequences were identified, in the genomes of a thermophile and halophilic archaebacteria, although these were less abundant than HIP1 in cyanobacteria and relate to codon usage. To test the proposed widespread distribution of HIP1 in DNA from the cyanobacterium Synechococcus PCC 6301, randomly selected genomic clones were partly sequenced. HIP1 constituted 2.5% of the novel sequences, equivalent to a site on average once every 320 nucleotides. An oligonucleotide including HIP1 was also tested in PCR. Multiple products were obtained using template DNA from cyanobacterial strains in which HIP1 is abundant in known sequences, and some strains generated characteristic HIP-PCR banding patterns. However, analysis of DNA from one strain (not previously represented in databases) by random sequencing, HIP-PCR and Pvul digestion, confirms that not all cyanobacterial genomes are rich in HIP1.

West Nile virus, Anopheles flavivirus, a novel flavivirus as well as Merida-like rhabdovirus Turkey in field-collected mosquitoes from Thrace and Anatolia.

PubMed

Öncü, Ceren; Brinkmann, Annika; Günay, Filiz; Kar, Sırrı; Öter, Kerem; Sarıkaya, Yasemen; Nitsche, Andreas; Linton, Yvonne-Marie; Alten, Bülent; Ergünay, Koray

2018-01-01

Mosquitoes are involved in the transmission and maintenance of several viral diseases with significant health impact. Biosurveillance efforts have also revealed insect-specific viruses, observed to cocirculate with pathogenic strains. This report describes the findings of flavivirus and rhabdovirus screening, performed in eastern Thrace and Aegean region of Anatolia during 2016, including and expanding on locations with previously-documented virus activity. A mosquito cohort of 1545 individuals comprising 14 species were collected and screened in 108 pools via generic and specific amplification and direct metagenomics by next generation sequencing. Seven mosquito pools (6.4%) were positive in the flavivirus screening. West Nile virus lineage 1 clade 1a sequences were characterized in a pool Culex pipiens sensu lato specimens, providing the initial virus detection in Aegean region following 2010 outbreak. In an Anopheles maculipennis sensu lato pool, sequences closely-related to Anopheles flaviviruses were obtained, with similarities to several African and Australian strains of this new insect-specific flavivirus clade. In pools comprising Uranotaenia unguiculata (n=3), Cx. pipiens s.l. (n=1) and Aedes caspius (n=1) mosquitoes, sequences of a novel flavivirus, distantly-related to Flavivirus AV2011, identified previously in Spain and Turkey, were characterized. Moreover, DNA forms of the novel flavivirus were detected in two Ur. unguiculata pools. These sequences were highly-similar to the sequences amplified from viral RNA, with undisrupted reading frames, suggest the occurrence of viral DNA forms in natural conditions within mosquito hosts. Rhabdovirus screening revealed sequences of a recently-described novel virus, named the Merida-like virus Turkey (MERDLVT) in 5 Cx. pipiens s.l. pools (4.6%). Partial L and N gene sequences of MERDLVT were well-conserved among strains, with evidence for geographical clustering in phylogenetic analyses. Metagenomics provided the near-full genomic sequence in a specimen, revealing an identical genome organization and limited divergence from the prototype MERDLVT isolate. Copyright © 2017 Elsevier B.V. All rights reserved.

Microbial Characterization During the Early Habitation of the International Space Station

NASA Technical Reports Server (NTRS)

Castro, V. A.; Thrasher, A. N.; Healy, M.; Ott, C. M.; Pierson, D. L.

2004-01-01

An evaluation of the microbiota from air, water, and surface samples provided a baseline of microbial characterization onboard the International Space Station (ISS) to gain insight into bacterial and fungal contamination during the initial stages of construction and habitation. Using 16S genetic sequencing and rep-PCR, 63 bacterial strains were isolated for identification and fingerprinted for microbial tracking. Of the bacterial strains that were isolated and fingerprinted, 19 displayed similarity to each other. The use of these molecular tools allowed for the identification of bacteria not previously identified using automated biochemical analysis and provided a clear indication of the source of several ISS contaminants. Strains of Bradyrhizobium and Sphingomonas unable to be identified using sequencing were identified by comparison of rep-PCR DNA fingerprints. Distinct DNA fingerprints for several strains of Methylobacterium provided a clear indication of the source of an ISS water supply contaminant. Fungal and bacterial data acquired during monitoring do not suggest there is a current microbial hazard to the spacecraft, nor does any trend indicate a potential health risk. Previous spacecraft environmental analysis indicated that microbial contamination will increase with time and will require continued surveillance. Copyright 2004 Springer-Verlag.

Complete genome sequence of DSM 30083(T), the type strain (U5/41(T)) of Escherichia coli, and a proposal for delineating subspecies in microbial taxonomy.

PubMed

Meier-Kolthoff, Jan P; Hahnke, Richard L; Petersen, Jörn; Scheuner, Carmen; Michael, Victoria; Fiebig, Anne; Rohde, Christine; Rohde, Manfred; Fartmann, Berthold; Goodwin, Lynne A; Chertkov, Olga; Reddy, Tbk; Pati, Amrita; Ivanova, Natalia N; Markowitz, Victor; Kyrpides, Nikos C; Woyke, Tanja; Göker, Markus; Klenk, Hans-Peter

2014-01-01

Although Escherichia coli is the most widely studied bacterial model organism and often considered to be the model bacterium per se, its type strain was until now forgotten from microbial genomics. As a part of the G enomic E ncyclopedia of B acteria and A rchaea project, we here describe the features of E. coli DSM 30083(T) together with its genome sequence and annotation as well as novel aspects of its phenotype. The 5,038,133 bp containing genome sequence includes 4,762 protein-coding genes and 175 RNA genes as well as a single plasmid. Affiliation of a set of 250 genome-sequenced E. coli strains, Shigella and outgroup strains to the type strain of E. coli was investigated using digital DNA:DNA-hybridization (dDDH) similarities and differences in genomic G+C content. As in the majority of previous studies, results show Shigella spp. embedded within E. coli and in most cases forming a single subgroup of it. Phylogenomic trees also recover the proposed E. coli phylotypes as monophyla with minor exceptions and place DSM 30083(T) in phylotype B2 with E. coli S88 as its closest neighbor. The widely used lab strain K-12 is not only genomically but also physiologically strongly different from the type strain. The phylotypes do not express a uniform level of character divergence as measured using dDDH, however, thus an alternative arrangement is proposed and discussed in the context of bacterial subspecies. Analyses of the genome sequences of a large number of E. coli strains and of strains from > 100 other bacterial genera indicate a value of 79-80% dDDH as the most promising threshold for delineating subspecies, which in turn suggests the presence of five subspecies within E. coli.

Using PCR-based detection and genotyping to trace Streptococcus salivarius meningitis outbreak strain to oral flora of radiology physician assistant.

PubMed

Srinivasan, Velusamy; Gertz, Robert E; Shewmaker, Patricia L; Patrick, Sarah; Chitnis, Amit S; O'Connell, Heather; Benowitz, Isaac; Patel, Priti; Guh, Alice Y; Noble-Wang, Judith; Turabelidze, George; Beall, Bernard

2012-01-01

We recently investigated three cases of bacterial meningitis that were reported from a midwestern radiology clinic where facemasks were not worn during spinal injection of contrast agent during myelography procedures. Using pulsed field gel electrophoresis we linked a case strain of S. salivarius to an oral specimen of a radiology physician assistant (RPA). We also used a real-time PCR assay to detect S. salivarius DNA within a culture-negative cerebrospinal fluid (CSF) specimen. Here we extend this investigation through using a nested PCR/sequencing strategy to link the culture-negative CSF specimen to the case strain. We also provide validation of the real-time PCR assay used, demonstrating that it is not solely specific for Streptococcus salivarius, but is also highly sensitive for detection of the closely related oral species Streptococcus vestibularis. Through using multilocus sequence typing and 16S rDNA sequencing we further strengthen the link between the CSF case isolate and the RPA carriage isolate. We also demonstrate that the newly characterized strains from this study are distinct from previously characterized S. salivarius strains associated with carriage and meningitis.

Investigation of a Possible Link Between Vaccination and the 2010 Sheep Pox Epizootic in Morocco.

PubMed

Haegeman, A; Zro, K; Sammin, D; Vandenbussche, F; Ennaji, M M; De Clercq, K

2016-12-01

Sheep pox is endemic in most parts of Northern Africa and has the potential to cause severe economic problems. Live attenuated vaccines are used in Morocco, and in many other countries, to control the disease. Sheep pox virus (SPPV) re-appeared in 2010 causing a nodular clinical form previously not observed in Morocco. The severe clinical signs observed during the course of this outbreak and initial reports citing similarity in nucleotide sequence between the Moroccan vaccine strain and field isolates warranted a more in depth analysis of this epizootic. In this study, sequence analysis showed that isolates obtained from four provinces of eastern Morocco were identical, demonstrating that a single SPPV strain was responsible for the 2010 epizootic. In addition, the genome fragments sequenced and phylogenetic analyses undertaken as part of this study showed significant differences between field isolates and the Moroccan vaccine strain. New PCR methods were developed to differentiate between wild-type isolates and vaccine strains of SPPV. Using these methods, no trace of wild-type SPPV was found in the vaccine and no evidence was found to suggest that the vaccine strain was causing clinical disease. © 2015 Blackwell Verlag GmbH.

Molecular epidemiological analysis of paired pol/env sequences from Portuguese HIV type 1 patients.

PubMed

Abecasis, Ana B; Martins, Andreia; Costa, Inês; Carvalho, Ana P; Diogo, Isabel; Gomes, Perpétua; Camacho, Ricardo J

2011-07-01

The advent of new therapeutic approaches targeting env and the search for efficient anti-HIV-1 vaccines make it necessary to identify the number of recombinant forms using genomic regions that were previously not frequently sequenced. In this study, we have subtyped paired pol and env sequences from HIV-1 strains infecting 152 patients being clinically followed in Portugal. The percentage of strains in which we found discordant subtypes in pol and env was 25.7%. When the subtype in pol and env was concordant (65.1%), the most prevalent subtypes were subtype B (40.8%), followed by subtype C (17.8%) and subtype G (5.3%). The most prevalent recombinant form was CRF14_BGpol/Genv (7.2%).

Identification of a New DNA Region Specific for Members of Mycobacterium tuberculosis Complex

PubMed Central

Magdalena, Juana; Vachée, Anne; Supply, Philip; Locht, Camille

1998-01-01

The successful use of DNA amplification for the detection of tuberculous mycobacteria crucially depends on the choice of the target sequence, which ideally should be present in all tuberculous mycobacteria and absent from all other bacteria. In the present study we developed a PCR procedure based on the intergenic region (IR) separating two genes encoding a recently identified mycobacterial two-component system named SenX3-RegX3. The senX3-regX3 IR is composed of a novel type of repetitive sequence, called mycobacterial interspersed repetitive units (MIRUs). In a survey of 116 Mycobacterium tuberculosis strains characterized by different IS6110 restriction fragment length polymorphisms, 2 Mycobacterium africanum strains, 3 Mycobacterium bovis strains (including 2 BCG strains), and 1 Mycobacterium microti strain, a specific PCR fragment was amplified in all cases. This collection included M. tuberculosis strains that lack IS6110 or mtp40, two target sequences that have previously been used for the detection of M. tuberculosis. No PCR fragment was amplified when DNA from other organisms was used, giving a sensitivity of 100% and a specificity of 100% in the confidence limit of this study. The numbers of MIRUs were found to vary among strains, resulting in six different groups of strains on the basis of the size of the amplified PCR fragment. However, the vast majority of the strains (approximately 90%) fell within the same group, containing two 77-bp MIRUs followed by one 53-bp MIRU. PMID:9542912

Genetic characterization of strains of Saccharomyces uvarum from New Zealand wineries.

PubMed

Zhang, Hanyao; Richards, Keith D; Wilson, Sandra; Lee, Soon A; Sheehan, Hester; Roncoroni, Miguel; Gardner, Richard C

2015-04-01

We present a genetic characterization of 65 isolates of Saccharomyces uvarum isolated from wineries in New Zealand, along with the complete nucleotide sequence of a single sulfite-tolerant isolate. The genome of the New Zealand isolate averaged 99.85% nucleotide identity to CBS7001, the previously sequenced strain of S. uvarum. However, three genomic segments (37-87 kb) showed 10% nucleotide divergence from CBS7001 but 99% identity to Saccharomyces eubayanus. We conclude that these three segments appear to have been introgressed from that species. The nucleotide sequence of the internal transcribed spacer (ITS) region from other New Zealand isolates were also very similar to that of CBS7001, and hybrids showed complete genetic compatibility for some strains, with tetrads giving four viable progeny that showed 2:2 segregations of marker genes. Some strains showed high tolerance to sulfite, with genetic analysis indicating linkage of this trait to the transcription factor FZF1, but not to SSU1, the sulfite efflux pump that it regulates in order to confer sulfite tolerance in Saccharomyces cerevisiae. The fermentation characteristics of selected strains of S. uvarum showed exceptionally good cold fermentation characteristics, superior to the best commercially available strains of S. cerevisiae. Copyright © 2014 Elsevier Ltd. All rights reserved.

Phylogenetic analysis of the alfalfa weevil complex (Coleoptera: Curculionidae) in North America.

PubMed

Böttger, Jorge A Achata; Bundy, C Scott; Oesterle, Naomi; Hanson, Stephen F

2013-02-01

The Eastern, Western, and Egyptian strains of alfalfa weevil are pests introduced to North America on three separate occasions, now they share partially overlapping geographic ranges, covering most of the continental United States. Behavior, susceptibility to parasites, and subtle morphological differences separate the strains. The difficulty in differentiating among these strains morphologically has led to the application of molecular phylogeny approaches including restriction fragment-length polymorphism characterization and sequencing of mitochondrial genes. While valuable for strain identification, this approach cannot identify interstrain hybrids because mitochondrial markers are maternally inherited. The work reported here extends previous findings by comparing over 7 Kb of sequence from two mitochondrial and four nuclear loci to increase the resolution of molecular phylogeny for these weevils. The related clover leaf weevil, also an occasional pest of alfalfa, was included in the analysis because the molecular phylogeny of this weevil has not been examined to date. Analysis of nuclear loci indicate that the clover weevil is a distinct species. Furthermore, while the three alfalfa weevil strains are separable based on mitochondrial sequence data they cannot be separated using nuclearloci suggesting that they are all recently diverged members of the same species. These data refine the relationships among these strains and may find application in design of better control strategies.

Whole genome sequencing and phylogenetic analysis of Bluetongue virus serotype 2 strains isolated in the Americas including a novel strain from the western United States.

PubMed

Gaudreault, Natasha N; Mayo, Christie E; Jasperson, Dane C; Crossley, Beate M; Breitmeyer, Richard E; Johnson, Donna J; Ostlund, Eileen N; MacLachlan, N James; Wilson, William C

2014-07-01

Bluetongue is a potentially fatal arboviral disease of domestic and wild ruminants that is characterized by widespread edema and tissue necrosis. Bluetongue virus (BTV) serotypes 10, 11, 13, and 17 occur throughout much of the United States, whereas serotype 2 (BTV-2) was previously only detected in the southeastern United States. Since 1998, 10 other BTV serotypes have also been isolated from ruminants in the southeastern United States. In 2010, BTV-2 was identified in California for the first time, and preliminary sequence analysis indicated that the virus isolate was closely related to BTV strains circulating in the southeastern United States. In the current study, the whole genome sequence of the California strain of BTV-2 was compared with those of other BTV-2 strains in the Americas. The results of the analysis suggest co-circulation of genetically distinct viruses in the southeastern United States, and further suggest that the 2010 western isolate is closely related to southeastern strains of BTV. Although it remains uncertain as to how this novel virus was translocated to California, the findings of the current study underscore the need for ongoing surveillance of this economically important livestock disease.

Haemophilus ducreyi Cutaneous Ulcer Strains Diverged from Both Class I and Class II Genital Ulcer Strains: Implications for Epidemiological Studies.

PubMed

Gangaiah, Dharanesh; Spinola, Stanley M

2016-12-01

Haemophilus ducreyi has emerged as a major cause of cutaneous ulcers (CU) in yaws-endemic regions of the tropics in the South Pacific, South East Asia and Africa. H. ducreyi was once thought only to cause the genital ulcer (GU) disease chancroid; GU strains belong to 2 distinct classes, class I and class II. Using whole-genome sequencing of 4 CU strains from Samoa, 1 from Vanuatu and 1 from Papua New Guinea, we showed that CU strains diverged from the class I strain 35000HP and that one CU strain expressed β-lactamase. Recently, the Center for Disease Control and Prevention released the genomes of 11 additional CU strains from Vanuatu and Ghana; however, the evolutionary relationship of these CU strains to previously-characterized CU and GU strains is unknown. We performed phylogenetic analysis of 17 CU and 10 GU strains. Class I and class II GU strains formed two distinct clades. The class I strains formed two subclades, one containing 35000HP and HD183 and the other containing the remainder of the class I strains. Twelve of the CU strains formed a subclone under the class I 35000HP subclade, while 2 CU strains formed a subclone under the other class I subclade. Unexpectedly, 3 of the CU strains formed a subclone under the class II clade. Phylogenetic analysis of dsrA-hgbA-ncaA sequences yielded a tree similar to that of whole-genome phylogenetic tree. CU strains diverged from multiple lineages within both class I and class II GU strains. Multilocus sequence typing of dsrA-hgbA-ncaA could be reliably used for epidemiological investigation of CU and GU strains. As class II strains grow relatively poorly and are relatively more susceptible to vancomycin than class I strains, these findings have implications for methods to recover CU strains. Comparison of contemporary CU and GU isolates would help clarify the relationship between these entities.

Predicting Zoonotic Risk of Influenza A Viruses from Host Tropism Protein Signature Using Random Forest

PubMed Central

Eng, Christine L. P.; Tong, Joo Chuan; Tan, Tin Wee

2017-01-01

Influenza A viruses remain a significant health problem, especially when a novel subtype emerges from the avian population to cause severe outbreaks in humans. Zoonotic viruses arise from the animal population as a result of mutations and reassortments, giving rise to novel strains with the capability to evade the host species barrier and cause human infections. Despite progress in understanding interspecies transmission of influenza viruses, we are no closer to predicting zoonotic strains that can lead to an outbreak. We have previously discovered distinct host tropism protein signatures of avian, human and zoonotic influenza strains obtained from host tropism predictions on individual protein sequences. Here, we apply machine learning approaches on the signatures to build a computational model capable of predicting zoonotic strains. The zoonotic strain prediction model can classify avian, human or zoonotic strains with high accuracy, as well as providing an estimated zoonotic risk. This would therefore allow us to quickly determine if an influenza virus strain has the potential to be zoonotic using only protein sequences. The swift identification of potential zoonotic strains in the animal population using the zoonotic strain prediction model could provide us with an early indication of an imminent influenza outbreak. PMID:28587080

Predicting Zoonotic Risk of Influenza A Viruses from Host Tropism Protein Signature Using Random Forest.

PubMed

Eng, Christine L P; Tong, Joo Chuan; Tan, Tin Wee

2017-05-25

Influenza A viruses remain a significant health problem, especially when a novel subtype emerges from the avian population to cause severe outbreaks in humans. Zoonotic viruses arise from the animal population as a result of mutations and reassortments, giving rise to novel strains with the capability to evade the host species barrier and cause human infections. Despite progress in understanding interspecies transmission of influenza viruses, we are no closer to predicting zoonotic strains that can lead to an outbreak. We have previously discovered distinct host tropism protein signatures of avian, human and zoonotic influenza strains obtained from host tropism predictions on individual protein sequences. Here, we apply machine learning approaches on the signatures to build a computational model capable of predicting zoonotic strains. The zoonotic strain prediction model can classify avian, human or zoonotic strains with high accuracy, as well as providing an estimated zoonotic risk. This would therefore allow us to quickly determine if an influenza virus strain has the potential to be zoonotic using only protein sequences. The swift identification of potential zoonotic strains in the animal population using the zoonotic strain prediction model could provide us with an early indication of an imminent influenza outbreak.

Design and verification of a pangenome microarray oligonucleotide probe set for Dehalococcoides spp.

PubMed

Hug, Laura A; Salehi, Maryam; Nuin, Paulo; Tillier, Elisabeth R; Edwards, Elizabeth A

2011-08-01

Dehalococcoides spp. are an industrially relevant group of Chloroflexi bacteria capable of reductively dechlorinating contaminants in groundwater environments. Existing Dehalococcoides genomes revealed a high level of sequence identity within this group, including 98 to 100% 16S rRNA sequence identity between strains with diverse substrate specificities. Common molecular techniques for identification of microbial populations are often not applicable for distinguishing Dehalococcoides strains. Here we describe an oligonucleotide microarray probe set designed based on clustered Dehalococcoides genes from five different sources (strain DET195, CBDB1, BAV1, and VS genomes and the KB-1 metagenome). This "pangenome" probe set provides coverage of core Dehalococcoides genes as well as strain-specific genes while optimizing the potential for hybridization to closely related, previously unknown Dehalococcoides strains. The pangenome probe set was compared to probe sets designed independently for each of the five Dehalococcoides strains. The pangenome probe set demonstrated better predictability and higher detection of Dehalococcoides genes than strain-specific probe sets on nontarget strains with <99% average nucleotide identity. An in silico analysis of the expected probe hybridization against the recently released Dehalococcoides strain GT genome and additional KB-1 metagenome sequence data indicated that the pangenome probe set performs more robustly than the combined strain-specific probe sets in the detection of genes not included in the original design. The pangenome probe set represents a highly specific, universal tool for the detection and characterization of Dehalococcoides from contaminated sites. It has the potential to become a common platform for Dehalococcoides-focused research, allowing meaningful comparisons between microarray experiments regardless of the strain examined.

Candida theae sp. nov., a new anamorphic beverage-associated member of the Lodderomyces clade.

PubMed

Chang, Chin-Feng; Lin, Yu-Ching; Chen, Shan-Fu; Carvajal Barriga, Enrique Javier; Carvaja Barriga, Enrique Javier; Barahona, Patricia Portero; James, Stephen A; Bond, Christopher J; Roberts, Ian N; Lee, Ching-Fu

2012-02-01

Four strains representing a novel yeast species belonging to the genus Candida were independently isolated in Taiwan and Ecuador. Two strains (G17(T) and G31) were isolated in Taiwan, by pellet precipitation from plastic-bottled tea drinks produced in Indonesia, while two additional strains (CLQCA 10-049 and CLQCA 10-062) were recovered from ancient chicha fermentation vessels found in tombs in Quito, Ecuador. These four strains were morphologically, and phylogenetically identical to each other. No sexual reproduction was observed on common sporulation media. Large-subunit (LSU) rRNA gene sequence analysis revealed the four strains to belong to the Lodderomyces clade, closely related to members of the Candida parapsilosis species complex. The four strains, which have identical LSU D1/D2 sequences, differ from their closest phylogenetic neighbors, Candida orthopsilosis and Candida parapsilosis, by 6-9 nt substitutions, respectively. Physiologically, the four strains are similar to Candida parapsilosis, although they can be distinguished from their closest relative by the assimilation of arbutin, nitrite, and creatine. The Indonesian and Ecuadorian strain sets can also be distinguished from one another based on ITS sequencing, differing by 4 substitutions in ITS1 and 1 single nucleotide indel in ITS2. Collectively, the results indicate that the four strains represent a previously unrecognized species of Candida. The name Candida theae sp. nov. is proposed to accommodate these strains, with G-17(T) (BCRC 23242(T)=CBS 12239(T)=ATCC MYA-4746(T)) designated as the type strain. Copyright © 2011 Elsevier B.V. All rights reserved.

Isolation and whole genome sequencing of a Ruminococcus-like bacterium, associated with irritable bowel syndrome.

PubMed

Hynönen, Ulla; Rasinkangas, Pia; Satokari, Reetta; Paulin, Lars; de Vos, Willem M; Pietilä, Taija E; Kant, Ravi; Palva, Airi

2016-06-01

In our previous studies on the intestinal microbiota in irritable bowel syndrome (IBS), we identified a bacterial phylotype with higher abundance in patients suffering from diarrhea than in healthy controls. In the present work, we have isolated in pure culture strain RT94, belonging to this phylotype, determined its whole genome sequence and performed an extensive genomic analysis and phenotypical testing. This revealed strain RT94 to be a strict anaerobe apparently belonging to a novel species with only 94% similarity in the 16S rRNA gene sequence to the closest relatives Ruminococcus torques and Ruminococcus lactaris. The G + C content of strain RT94 is 45.2 mol% and the major long-chain cellular fatty acids are C16:0, C18:0 and C14:0. The isolate is metabolically versatile but not a mucus or cellulose utilizer. It produces acetate, ethanol, succinate, lactate and formate, but very little butyrate, as end products of glucose metabolism. The mechanisms underlying the association of strain RT94 with diarrhea-type IBS are discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Molecular Characterization of Lactobacillus plantarum DMDL 9010, a Strain with Efficient Nitrite Degradation Capacity

PubMed Central

Fei, Yong-tao; Liu, Dong-mei; Luo, Tong-hui; Chen, Gu; Wu, Hui; Li, Li; Yu, Yi-gang

2014-01-01

Nitrites commonly found in food, especially in fermented vegetables, are potential carcinogens. Therefore, limiting nitrites in food is critically important for food safety. A Lactobacillus strain (Lactobacillus sp. DMDL 9010) was previously isolated from fermented vegetables by our group, and is not yet fully characterized. A number of phenotypical and genotypical approaches were employed to characterize Lactobacillus sp. DMDL 9010. Its nitrite degradation capacity was compared with four other Lactobacillus strains, including Lactobacillus casei subsp. rhamnosus 719, Lactobacillus delbrueckii subsp. bulgaricu 1.83, Streptococcus thermophilus 1.204, and lactobacillus plantarum 8140, on MRS medium. Compared to these four Lactobacillus strains, Lactobacillus sp. DMDL 9010 had a significantly higher nitrite degradation capacity (P<0.001). Based on 16S rDNA sequencing and sequence comparison, Lactobacillus sp. DMDL 9010 was identified as either Lactobacillus plantarum or Lactobacillus pentosus. To further identify this strain, the flanking regions (922 bp and 806 bp upstream and downstream, respectively) of the L-lactate dehydrogenase 1 (L-ldh1) gene were amplified and sequenced. Lactobacillus sp. DMDL 9010 had 98.92 and 76.98% sequence identity in the upstream region with L. plantarum WCFS1 and L. pentosus IG1, respectively, suggesting that Lactobacillu sp. DMDL 9010 is an L. plantarum strain. It was therefore named L. plantarum DMDL 9010. Our study provides a platform for genetic engineering of L. plantarum DMDL 9010, in order to further improve its nitrite degradation capacity. PMID:25423449

Molecular characterization of Lactobacillus plantarum DMDL 9010, a strain with efficient nitrite degradation capacity.

PubMed

Fei, Yong-tao; Liu, Dong-mei; Luo, Tong-hui; Chen, Gu; Wu, Hui; Li, Li; Yu, Yi-gang

2014-01-01

Nitrites commonly found in food, especially in fermented vegetables, are potential carcinogens. Therefore, limiting nitrites in food is critically important for food safety. A Lactobacillus strain (Lactobacillus sp. DMDL 9010) was previously isolated from fermented vegetables by our group, and is not yet fully characterized. A number of phenotypical and genotypical approaches were employed to characterize Lactobacillus sp. DMDL 9010. Its nitrite degradation capacity was compared with four other Lactobacillus strains, including Lactobacillus casei subsp. rhamnosus 719, Lactobacillus delbrueckii subsp. bulgaricu 1.83, Streptococcus thermophilus 1.204, and lactobacillus plantarum 8140, on MRS medium. Compared to these four Lactobacillus strains, Lactobacillus sp. DMDL 9010 had a significantly higher nitrite degradation capacity (P<0.001). Based on 16S rDNA sequencing and sequence comparison, Lactobacillus sp. DMDL 9010 was identified as either Lactobacillus plantarum or Lactobacillus pentosus. To further identify this strain, the flanking regions (922 bp and 806 bp upstream and downstream, respectively) of the L-lactate dehydrogenase 1 (L-ldh1) gene were amplified and sequenced. Lactobacillus sp. DMDL 9010 had 98.92 and 76.98% sequence identity in the upstream region with L. plantarum WCFS1 and L. pentosus IG1, respectively, suggesting that Lactobacillu sp. DMDL 9010 is an L. plantarum strain. It was therefore named L. plantarum DMDL 9010. Our study provides a platform for genetic engineering of L. plantarum DMDL 9010, in order to further improve its nitrite degradation capacity.

MGmapper: Reference based mapping and taxonomy annotation of metagenomics sequence reads

PubMed Central

Lukjancenko, Oksana; Thomsen, Martin Christen Frølund; Maddalena Sperotto, Maria; Lund, Ole; Møller Aarestrup, Frank; Sicheritz-Pontén, Thomas

2017-01-01

An increasing amount of species and gene identification studies rely on the use of next generation sequence analysis of either single isolate or metagenomics samples. Several methods are available to perform taxonomic annotations and a previous metagenomics benchmark study has shown that a vast number of false positive species annotations are a problem unless thresholds or post-processing are applied to differentiate between correct and false annotations. MGmapper is a package to process raw next generation sequence data and perform reference based sequence assignment, followed by a post-processing analysis to produce reliable taxonomy annotation at species and strain level resolution. An in-vitro bacterial mock community sample comprised of 8 genuses, 11 species and 12 strains was previously used to benchmark metagenomics classification methods. After applying a post-processing filter, we obtained 100% correct taxonomy assignments at species and genus level. A sensitivity and precision at 75% was obtained for strain level annotations. A comparison between MGmapper and Kraken at species level, shows MGmapper assigns taxonomy at species level using 84.8% of the sequence reads, compared to 70.5% for Kraken and both methods identified all species with no false positives. Extensive read count statistics are provided in plain text and excel sheets for both rejected and accepted taxonomy annotations. The use of custom databases is possible for the command-line version of MGmapper, and the complete pipeline is freely available as a bitbucked package (https://bitbucket.org/genomicepidemiology/mgmapper). A web-version (https://cge.cbs.dtu.dk/services/MGmapper) provides the basic functionality for analysis of small fastq datasets. PMID:28467460

MGmapper: Reference based mapping and taxonomy annotation of metagenomics sequence reads.

PubMed

Petersen, Thomas Nordahl; Lukjancenko, Oksana; Thomsen, Martin Christen Frølund; Maddalena Sperotto, Maria; Lund, Ole; Møller Aarestrup, Frank; Sicheritz-Pontén, Thomas

2017-01-01

An increasing amount of species and gene identification studies rely on the use of next generation sequence analysis of either single isolate or metagenomics samples. Several methods are available to perform taxonomic annotations and a previous metagenomics benchmark study has shown that a vast number of false positive species annotations are a problem unless thresholds or post-processing are applied to differentiate between correct and false annotations. MGmapper is a package to process raw next generation sequence data and perform reference based sequence assignment, followed by a post-processing analysis to produce reliable taxonomy annotation at species and strain level resolution. An in-vitro bacterial mock community sample comprised of 8 genuses, 11 species and 12 strains was previously used to benchmark metagenomics classification methods. After applying a post-processing filter, we obtained 100% correct taxonomy assignments at species and genus level. A sensitivity and precision at 75% was obtained for strain level annotations. A comparison between MGmapper and Kraken at species level, shows MGmapper assigns taxonomy at species level using 84.8% of the sequence reads, compared to 70.5% for Kraken and both methods identified all species with no false positives. Extensive read count statistics are provided in plain text and excel sheets for both rejected and accepted taxonomy annotations. The use of custom databases is possible for the command-line version of MGmapper, and the complete pipeline is freely available as a bitbucked package (https://bitbucket.org/genomicepidemiology/mgmapper). A web-version (https://cge.cbs.dtu.dk/services/MGmapper) provides the basic functionality for analysis of small fastq datasets.

Molecular Identification of Paramecium bursaria Syngens and Studies on Geographic Distribution using Mitochondrial Cytochrome C Oxidase Subunit I (COI).

PubMed

Zagata, Patrycja; Greczek-Stachura, Magdalena; Tarcz, Sebastian; Rautian, Maria

2015-01-01

Paramecium bursaria is composed of five syngens that are morphologically indistinguishable but sexually isolated. The aim of the present study was to confirm by molecular methods (analyses of mitochondrial COI) the identification of P. bursaria syngens originating from different geographical locations. Phylograms constructed using both the neighbor-joining and maximum-likelihood methods based on a comparison of 34 sequences of P. bursaria strains and P. multimicronucleatum, P. caudatum and P.calkinsi strains used as outgroups revealed five clusters which correspond to results obtained previously by mating reaction. Our analysis shows the existence of 24 haplotypes for the COI gene sequence in the studied strains. The interspecies haplotype diversity was Hd = 0.967. We confirmed genetic differentiation between strains of P. bursaria and the occurrence of a correlation between geographical distribution and the correspondent syngen.

Expression of three mammalian cDNAs that interfere with RAS function in Saccharomyces cerevisiae.

PubMed Central

Colicelli, J; Nicolette, C; Birchmeier, C; Rodgers, L; Riggs, M; Wigler, M

1991-01-01

Saccharomyces cerevisiae strains expressing the activated RAS2Val19 gene or lacking both cAMP phosphodiesterase genes, PDE1 and PDE2, have impaired growth control and display an acute sensitivity to heat shock. We have isolated two classes of mammalian cDNAs from yeast expression libraries that suppress the heat shock-sensitive phenotype of RAS2Val19 strain. Members of the first class of cDNAs also suppress the heat shock-sensitive phenotype of pde1- pde2- strains and encode cAMP phosphodiesterases. Members of the second class fail to suppress the phenotype of pde1- pde2- strains and therefore are candidate cDNAs encoding proteins that interact with RAS proteins. We report the nucleotide sequence of three members of this class. Two of these cDNAs share considerable sequence similarity, but none are clearly similar to previously isolated genes. Images PMID:1849280

Partial 16S rRNA primary structure of five Actinomyces species: phylogenetic implications and development of an Actinomyces israelii-specific oligonucleotide probe.

PubMed

Stackebrandt, E; Charfreitag, O

1990-01-01

The intra- and intergeneric relationships of the genus Actinomyces were determined by comparing long 16S rRNA sequences, generated by reverse transcriptase. All species formed a phylogenetically coherent cluster in which Actinomyces bovis, A. viscosus, A. naeslundii, A. odontolyticus and A. israelii constituted genetically well defined species. A. israelii DSM 43322 (serotype 2) was not closely related to three other strains of this species (serotype 1) and, as judged from phylogenetic distances, could be accommodated within A. naeslundii, or represent a new species. In contrast to previous findings, members of the genus Actinomyces appear to be related to Bifidobacterium bifidum. Sequence information was used to develop an oligonucleotide probe for the A. israelii serotype 1 strains, which did not react with the serotype 2 strain or with rRNA from strains of eight Actinomyces species.

Antibiotic Resistance Markers in Burkholderia pseudomallei Strain Bp1651 Identified by Genome Sequence Analysis

PubMed Central

Sue, David; Gee, Jay E.; Elrod, Mindy G.; Hoffmaster, Alex R.; Randall, Linnell B.; Chirakul, Sunisa; Tuanyok, Apichai; Schweizer, Herbert P.; Weigel, Linda M.

2017-01-01

ABSTRACT Burkholderia pseudomallei Bp1651 is resistant to several classes of antibiotics that are usually effective for treatment of melioidosis, including tetracyclines, sulfonamides, and β-lactams such as penicillins (amoxicillin-clavulanic acid), cephalosporins (ceftazidime), and carbapenems (imipenem and meropenem). We sequenced, assembled, and annotated the Bp1651 genome and analyzed the sequence using comparative genomic analyses with susceptible strains, keyword searches of the annotation, publicly available antimicrobial resistance prediction tools, and published reports. More than 100 genes in the Bp1651 sequence were identified as potentially contributing to antimicrobial resistance. Most notably, we identified three previously uncharacterized point mutations in penA, which codes for a class A β-lactamase and was previously implicated in resistance to β-lactam antibiotics. The mutations result in amino acid changes T147A, D240G, and V261I. When individually introduced into select agent-excluded B. pseudomallei strain Bp82, D240G was found to contribute to ceftazidime resistance and T147A contributed to amoxicillin-clavulanic acid and imipenem resistance. This study provides the first evidence that mutations in penA may alter susceptibility to carbapenems in B. pseudomallei. Another mutation of interest was a point mutation affecting the dihydrofolate reductase gene folA, which likely explains the trimethoprim resistance of this strain. Bp1651 was susceptible to aminoglycosides likely because of a frameshift in the amrB gene, the transporter subunit of the AmrAB-OprA efflux pump. These findings expand the role of penA to include resistance to carbapenems and may assist in the development of molecular diagnostics that predict antimicrobial resistance and provide guidance for treatment of melioidosis. PMID:28396541

Genome Sequencing and Comparative Analysis of Saccharomyces cerevisiae Strains of the Peterhof Genetic Collection

PubMed Central

Drozdova, Polina B.; Tarasov, Oleg V.; Matveenko, Andrew G.; Radchenko, Elina A.; Sopova, Julia V.; Polev, Dmitrii E.; Inge-Vechtomov, Sergey G.; Dobrynin, Pavel V.

2016-01-01

The Peterhof genetic collection of Saccharomyces cerevisiae strains (PGC) is a large laboratory stock that has accumulated several thousands of strains for over than half a century. It originated independently of other common laboratory stocks from a distillery lineage (race XII). Several PGC strains have been extensively used in certain fields of yeast research but their genomes have not been thoroughly explored yet. Here we employed whole genome sequencing to characterize five selected PGC strains including one of the closest to the progenitor, 15V-P4, and several strains that have been used to study translation termination and prions in yeast (25-25-2V-P3982, 1B-D1606, 74-D694, and 6P-33G-D373). The genetic distance between the PGC progenitor and S288C is comparable to that between two geographically isolated populations. The PGC seems to be closer to two bakery strains than to S288C-related laboratory stocks or European wine strains. In genomes of the PGC strains, we found several loci which are absent from the S288C genome; 15V-P4 harbors a rare combination of the gene cluster characteristic for wine strains and the RTM1 cluster. We closely examined known and previously uncharacterized gene variants of particular strains and were able to establish the molecular basis for known phenotypes including phenylalanine auxotrophy, clumping behavior and galactose utilization. Finally, we made sequencing data and results of the analysis available for the yeast community. Our data widen the knowledge about genetic variation between Saccharomyces cerevisiae strains and can form the basis for planning future work in PGC-related strains and with PGC-derived alleles. PMID:27152522

Serendipitous discovery of Wolbachia genomes in multiple Drosophila species.

PubMed

Salzberg, Steven L; Dunning Hotopp, Julie C; Delcher, Arthur L; Pop, Mihai; Smith, Douglas R; Eisen, Michael B; Nelson, William C

2005-01-01

The Trace Archive is a repository for the raw, unanalyzed data generated by large-scale genome sequencing projects. The existence of this data offers scientists the possibility of discovering additional genomic sequences beyond those originally sequenced. In particular, if the source DNA for a sequencing project came from a species that was colonized by another organism, then the project may yield substantial amounts of genomic DNA, including near-complete genomes, from the symbiotic or parasitic organism. By searching the publicly available repository of DNA sequencing trace data, we discovered three new species of the bacterial endosymbiont Wolbachia pipientis in three different species of fruit fly: Drosophila ananassae, D. simulans, and D. mojavensis. We extracted all sequences with partial matches to a previously sequenced Wolbachia strain and assembled those sequences using customized software. For one of the three new species, the data recovered were sufficient to produce an assembly that covers more than 95% of the genome; for a second species the data produce the equivalent of a 'light shotgun' sampling of the genome, covering an estimated 75-80% of the genome; and for the third species the data cover approximately 6-7% of the genome. The results of this study reveal an unexpected benefit of depositing raw data in a central genome sequence repository: new species can be discovered within this data. The differences between these three new Wolbachia genomes and the previously sequenced strain revealed numerous rearrangements and insertions within each lineage and hundreds of novel genes. The three new genomes, with annotation, have been deposited in GenBank.

Whole-genome sequencing reveals that Shewanella haliotis Kim et al. 2007 can be considered a later heterotypic synonym of Shewanella algae Simidu et al. 1990.

PubMed

Szeinbaum, Nadia; Kellum, Cailin E; Glass, Jennifer B; Janda, J Michael; DiChristina, Thomas J

2018-04-01

Previously, experimental DNA-DNA hybridization (DDH) between Shewanellahaliotis JCM 14758 T and Shewanellaalgae JCM 21037 T had suggested that the two strains could be considered different species, despite minimal phenotypic differences. The recent isolation of Shewanella sp. MN-01, with 99 % 16S rRNA gene identity to S. algae and S. haliotis, revealed a potential taxonomic problem between these two species. In this study, we reassessed the nomenclature of S. haliotis and S. algae using available whole-genome sequences. The whole-genome sequence of S. haliotis JCM 14758 T and ten S. algae strains showed ≥97.7 % average nucleotide identity and >78.9 % digital DDH, clearly above the recommended species thresholds. According to the rules of priority and in view of the results obtained, S. haliotis is to be considered a later heterotypic synonym of S. algae. Because the whole-genome sequence of Shewanella sp. strain MN-01 shares >99 % ANI with S. algae JCM 14758 T , it can be confidently identified as S. algae.

Diversity of the Cronobacter Genus as Revealed by Multilocus Sequence Typing

PubMed Central

Joseph, S.; Sonbol, H.; Hariri, S.; Desai, P.; McClelland, M.

2012-01-01

Cronobacter (previously known as Enterobacter sakazakii) is a diverse bacterial genus consisting of seven species: C. sakazakii, C. malonaticus, C. turicensis, C. universalis, C. muytjensii, C. dublinensis, and C. condimenti. In this study, we have used a multilocus sequence typing (MLST) approach employing the alleles of 7 genes (atpD, fusA, glnS, gltB, gyrB, infB, and ppsA; total length, 3,036 bp) to investigate the phylogenetic relationship of 325 Cronobacter species isolates. Strains were chosen on the basis of their species, geographic and temporal distribution, source, and clinical outcome. The earliest strain was isolated from milk powder in 1950, and the earliest clinical strain was isolated in 1953. The existence of seven species was supported by MLST. Intraspecific variation ranged from low diversity in C. sakazakii to extensive diversity within some species, such as C. muytjensii and C. dublinensis, including evidence of gene conversion between species. The predominant species from clinical sources was found to be C. sakazakii. C. sakazakii sequence type 4 (ST4) was the predominant sequence type of cerebral spinal fluid isolates from cases of meningitis. PMID:22785185

Complete genome sequence and comparative analysis of Acetobacter pasteurianus 386B, a strain well-adapted to the cocoa bean fermentation ecosystem.

PubMed

Illeghems, Koen; De Vuyst, Luc; Weckx, Stefan

2013-08-01

Acetobacter pasteurianus 386B, an acetic acid bacterium originating from a spontaneous cocoa bean heap fermentation, proved to be an ideal functional starter culture for coca bean fermentations. It is able to dominate the fermentation process, thereby resisting high acetic acid concentrations and temperatures. However, the molecular mechanisms underlying its metabolic capabilities and niche adaptations are unknown. In this study, whole-genome sequencing and comparative genome analysis was used to investigate this strain's mechanisms to dominate the cocoa bean fermentation process. The genome sequence of A. pasteurianus 386B is composed of a 2.8-Mb chromosome and seven plasmids. The annotation of 2875 protein-coding sequences revealed important characteristics, including several metabolic pathways, the occurrence of strain-specific genes such as an endopolygalacturonase, and the presence of mechanisms involved in tolerance towards various stress conditions. Furthermore, the low number of transposases in the genome and the absence of complete phage genomes indicate that this strain might be more genetically stable compared with other A. pasteurianus strains, which is an important advantage for the use of this strain as a functional starter culture. Comparative genome analysis with other members of the Acetobacteraceae confirmed the functional properties of A. pasteurianus 386B, such as its thermotolerant nature and unique genetic composition. Genome analysis of A. pasteurianus 386B provided detailed insights into the underlying mechanisms of its metabolic features, niche adaptations, and tolerance towards stress conditions. Combination of these data with previous experimental knowledge enabled an integrated, global overview of the functional characteristics of this strain. This knowledge will enable improved fermentation strategies and selection of appropriate acetic acid bacteria strains as functional starter culture for cocoa bean fermentation processes.

Genome sequencing reveals loci under artificial selection that underlie disease phenotypes in the laboratory rat.

PubMed

Atanur, Santosh S; Diaz, Ana Garcia; Maratou, Klio; Sarkis, Allison; Rotival, Maxime; Game, Laurence; Tschannen, Michael R; Kaisaki, Pamela J; Otto, Georg W; Ma, Man Chun John; Keane, Thomas M; Hummel, Oliver; Saar, Kathrin; Chen, Wei; Guryev, Victor; Gopalakrishnan, Kathirvel; Garrett, Michael R; Joe, Bina; Citterio, Lorena; Bianchi, Giuseppe; McBride, Martin; Dominiczak, Anna; Adams, David J; Serikawa, Tadao; Flicek, Paul; Cuppen, Edwin; Hubner, Norbert; Petretto, Enrico; Gauguier, Dominique; Kwitek, Anne; Jacob, Howard; Aitman, Timothy J

2013-08-01

Large numbers of inbred laboratory rat strains have been developed for a range of complex disease phenotypes. To gain insights into the evolutionary pressures underlying selection for these phenotypes, we sequenced the genomes of 27 rat strains, including 11 models of hypertension, diabetes, and insulin resistance, along with their respective control strains. Altogether, we identified more than 13 million single-nucleotide variants, indels, and structural variants across these rat strains. Analysis of strain-specific selective sweeps and gene clusters implicated genes and pathways involved in cation transport, angiotensin production, and regulators of oxidative stress in the development of cardiovascular disease phenotypes in rats. Many of the rat loci that we identified overlap with previously mapped loci for related traits in humans, indicating the presence of shared pathways underlying these phenotypes in rats and humans. These data represent a step change in resources available for evolutionary analysis of complex traits in disease models. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Genome Sequencing Reveals Loci under Artificial Selection that Underlie Disease Phenotypes in the Laboratory Rat

PubMed Central

Atanur, Santosh S.; Diaz, Ana Garcia; Maratou, Klio; Sarkis, Allison; Rotival, Maxime; Game, Laurence; Tschannen, Michael R.; Kaisaki, Pamela J.; Otto, Georg W.; Ma, Man Chun John; Keane, Thomas M.; Hummel, Oliver; Saar, Kathrin; Chen, Wei; Guryev, Victor; Gopalakrishnan, Kathirvel; Garrett, Michael R.; Joe, Bina; Citterio, Lorena; Bianchi, Giuseppe; McBride, Martin; Dominiczak, Anna; Adams, David J.; Serikawa, Tadao; Flicek, Paul; Cuppen, Edwin; Hubner, Norbert; Petretto, Enrico; Gauguier, Dominique; Kwitek, Anne; Jacob, Howard; Aitman, Timothy J.

2013-01-01

Summary Large numbers of inbred laboratory rat strains have been developed for a range of complex disease phenotypes. To gain insights into the evolutionary pressures underlying selection for these phenotypes, we sequenced the genomes of 27 rat strains, including 11 models of hypertension, diabetes, and insulin resistance, along with their respective control strains. Altogether, we identified more than 13 million single-nucleotide variants, indels, and structural variants across these rat strains. Analysis of strain-specific selective sweeps and gene clusters implicated genes and pathways involved in cation transport, angiotensin production, and regulators of oxidative stress in the development of cardiovascular disease phenotypes in rats. Many of the rat loci that we identified overlap with previously mapped loci for related traits in humans, indicating the presence of shared pathways underlying these phenotypes in rats and humans. These data represent a step change in resources available for evolutionary analysis of complex traits in disease models. PaperClip PMID:23890820

Genomic characterisation of Leptospira inadai serogroup Lyme isolated from captured rat in Brazil and comparative analysis with human reference strain.

PubMed

Moreno, Luisa Z; Miraglia, Fabiana; Loureiro, Ana P; Kremer, Frederico S; Eslabao, Marcus R; Dellagostin, Odir A; Lilenbaum, Walter; Vasconcellos, Silvio A; Heinemann, Marcos B; Moreno, Andrea M

2018-03-12

Leptospira inadai is classified as a species of the Leptospira intermediate group that has been poorly studied due to its apparent insignificance to human and animal health. Nevertheless, over the last two decades the species has been described in human cases in India and in carrier animals in Ecuador. Here, we present the first identification and genomic characterisation of L. inadai serogroup Lyme isolated from captured rodent in Brazil. Even though the M34/99 strain was not pathogenic for hamsters, it was able to establish renal colonisation. The M34/99 strain presented high similarity with L. inadai serogroup Lyme human reference indicating that animal strain could also infect humans, although it does not represent high risk of severe disease. An extrachromosomal sequence was also identified in M34/99 strain and presented high identity with previously described L. inadai phage LinZ_10, suggesting that phage-like extrachromosomal sequence may be another feature of this understudied species.

Multilocus sequence typing of Chlamydia trachomatis among men who have sex with men reveals cocirculating strains not associated with specific subpopulations.

PubMed

Bom, Reinier J M; Matser, Amy; Bruisten, Sylvia M; van Rooijen, Martijn S; Heijman, Titia; Morré, Servaas A; de Vries, Henry J C; Schim van der Loeff, Maarten F

2013-09-01

Previous studies identified specific Chlamydia trachomatis strains circulating among men who have sex with men (MSM). This study investigates whether distinct C. trachomatis strains circulate among subpopulations within the MSM community. Participants were recruited at the sexually transmitted infection clinic of the Public Health Service of Amsterdam from 2008 to 2009. C. trachomatis samples were typed using multilocus sequence typing. Epidemiological and clinical data were derived from questionnaires and patient records. Typing of 277 samples from 260 MSM identified distinct C. trachomatis strains circulating concurrently over time. Men with lymphogranuloma venereum (LGV)-inducing strains were more likely to be infected with human immunodeficiency virus, more often had a history of STI, and had a higher frequency of risky sexual behavior. No such associations were found for non-LGV-inducing strains. MSM infected with heterosexual-associated strains were often younger (P = .04) and more often reported sex with women (P = .03), compared with men infected with MSM-associated strains. With the exception of LGV-inducing strains, no evidence was found that different C. trachomatis strains circulated in distinct subpopulations of MSM. This indicates that no separate transmission networks for C. trachomatis among MSM existed. However, younger MSM and bisexuals were more often infected with heterosexual-associated C. trachomatis strains.

Interaction of Extreme Halophilic Archaea With the Evaporites of the Solar Salterns Guerrero Negro Baja California, Mexico

NASA Astrophysics Data System (ADS)

Tamez, P.; Lopez-Cortés, A.

2008-12-01

Hypersaline environments have been significant reservoirs for the long-term evolution of specifically adapted microorganisms. Characterized to have higher salt concentrations (up to 35 g/L), they are worldwide distributed and have a commercial significance. Exportadora de Sal, Guerrero Negro, Mexico has a multipond salterns system designed to harvest common salt (NaCl) from sea water. To achieve this purpose, sea water is pumped through a set of shallow ponds where water evaporates and salts concentrate. Sequential precipitation of CaCO3, CaSO4 2H2O and NaCl occurs in a mineral formations call it evaporites. In the interior of those gypsum-encrusted and halite-encrusted minerals, communities of extremely salt-loving archaea prosper. Previous studies have showed the influence of Haloarchaeal cells in the formation of larger fluid inclusions than crystals formed in sterile salt solutions. S-layer envelopes and cells of Haloarcula strain SP8807 contributed to the nucleation of new crystals of NaCl. Given the significance of the scope in phylogenetic archaeal diversity research, this study had a polyphasic approach. SEM micrographs from a 21- 31% (w/v) gradient salt multipond system evaporites, gave an insight profile of the extreme halophilic archaeal communities thriving in the surface of the gypsum and halite evaporites. Halite crystals were form after 21 days of incubation in solid medium with archaeal cells. Both culture and non-culture dependent methods, Nested-PCR-DGGE analysis and sequencing of 16S rDNA amplified fragment genes from environmental samples and isolated strains were used for this purpose. We isolate three strains from Pond 9 (21.07% total salt concentration) and one strain from Cristallizer 20 (25.15% total salt concentration). 16S rDNA signaling gave 99% of similarity with Halogeometricum borinquense, sequence AF002984, two other strains were 99% of similarity with Halobacterium salinarum, sequence AJ496185 these strains shown different colony morphology. Other strain match, with 99% of similarity, with three sequences Haloarcula: Haloarcula japonica (EF645686), Haloarcula hispanica (DQ089681) and Haloarcula marismortui (X61689), differences among them were only 3 base pairs. We suggest that strain, C205030908, could correspond to Haloarcula californiae previously reported by Javor et al, (1982), from which there is not sequence known. Nested-PCR-DGGE analysis showed a pattern of 11 bands, meaning the archaeal community is more complex than we could detect by culture approach.

Detection of Low-Level Cardinium and Wolbachia Infections in Culicoides

PubMed Central

Mee, Peter T.; Weeks, Andrew R.; Walker, Peter J.; Hoffmann, Ary A.

2015-01-01

Bacterial endosymbionts have been identified as potentially useful biological control agents for a range of invertebrate vectors of disease. Previous studies of Culicoides (Diptera: Ceratopogonidae) species using conventional PCR assays have provided evidence of Wolbachia (1/33) and Cardinium (8/33) infections. Here, we screened 20 species of Culicoides for Wolbachia and Cardinium, utilizing a combination of conventional PCR and more sensitive quantitative PCR (qPCR) assays. Low levels of Cardinium DNA were detected in females of all but one of the Culicoides species screened, and low levels of Wolbachia were detected in females of 9 of the 20 Culicoides species. Sequence analysis based on partial 16S rRNA gene and gyrB sequences identified “Candidatus Cardinium hertigii” from group C, which has previously been identified in Culicoides from Japan, Israel, and the United Kingdom. Wolbachia strains detected in this study showed 98 to 99% sequence identity to Wolbachia previously detected from Culicoides based on the 16S rRNA gene, whereas a strain with a novel wsp sequence was identified in Culicoides narrabeenensis. Cardinium isolates grouped to geographical regions independent of the host Culicoides species, suggesting possible geographical barriers to Cardinium movement. Screening also identified Asaia bacteria in Culicoides. These findings point to a diversity of low-level endosymbiont infections in Culicoides, providing candidates for further characterization and highlighting the widespread occurrence of these endosymbionts in this insect group. PMID:26150447

Determination of evolutionary relationships of outbreak-associated Listeria monocytogenes strains of serotypes 1/2a and 1/2b by whole-genome sequencing

USDA-ARS?s Scientific Manuscript database

We used whole-genome sequencing to determine evolutionary relationships among 20 outbreak-associated clinical isolates of Listeria monocytogenes serotypes 1/2a and 1/2b. Isolates from 6 of 11 outbreaks fell outside the clonal groups or “epidemic clones” that have been previously associated with outb...

Structure and Evolution of Chlorate Reduction Composite Transposons

PubMed Central

Clark, Iain C.; Melnyk, Ryan A.; Engelbrektson, Anna; Coates, John D.

2013-01-01

ABSTRACT The genes for chlorate reduction in six bacterial strains were analyzed in order to gain insight into the metabolism. A newly isolated chlorate-reducing bacterium (Shewanella algae ACDC) and three previously isolated strains (Ideonella dechloratans, Pseudomonas sp. strain PK, and Dechloromarinus chlorophilus NSS) were genome sequenced and compared to published sequences (Alicycliphilus denitrificans BC plasmid pALIDE01 and Pseudomonas chloritidismutans AW-1). De novo assembly of genomes failed to join regions adjacent to genes involved in chlorate reduction, suggesting the presence of repeat regions. Using a bioinformatics approach and finishing PCRs to connect fragmented contigs, we discovered that chlorate reduction genes are flanked by insertion sequences, forming composite transposons in all four newly sequenced strains. These insertion sequences delineate regions with the potential to move horizontally and define a set of genes that may be important for chlorate reduction. In addition to core metabolic components, we have highlighted several such genes through comparative analysis and visualization. Phylogenetic analysis places chlorate reductase within a functionally diverse clade of type II dimethyl sulfoxide (DMSO) reductases, part of a larger family of enzymes with reactivity toward chlorate. Nucleotide-level forensics of regions surrounding chlorite dismutase (cld), as well as its phylogenetic clustering in a betaproteobacterial Cld clade, indicate that cld has been mobilized at least once from a perchlorate reducer to build chlorate respiration. PMID:23919996

Whole-Genome Sequences of DA and F344 Rats with Different Susceptibilities to Arthritis, Autoimmunity, Inflammation and Cancer

PubMed Central

Guo, Xiaosen; Brenner, Max; Zhang, Xuemei; Laragione, Teresina; Tai, Shuaishuai; Li, Yanhong; Bu, Junjie; Yin, Ye; Shah, Anish A.; Kwan, Kevin; Li, Yingrui; Jun, Wang; Gulko, Pércio S.

2013-01-01

DA (D-blood group of Palm and Agouti, also known as Dark Agouti) and F344 (Fischer) are two inbred rat strains with differences in several phenotypes, including susceptibility to autoimmune disease models and inflammatory responses. While these strains have been extensively studied, little information is available about the DA and F344 genomes, as only the Brown Norway (BN) and spontaneously hypertensive rat strains have been sequenced to date. Here we report the sequencing of the DA and F344 genomes using next-generation Illumina paired-end read technology and the first de novo assembly of a rat genome. DA and F344 were sequenced with an average depth of 32-fold, covered 98.9% of the BN reference genome, and included 97.97% of known rat ESTs. New sequences could be assigned to 59 million positions with previously unknown data in the BN reference genome. Differences between DA, F344, and BN included 19 million positions in novel scaffolds, 4.09 million single nucleotide polymorphisms (SNPs) (including 1.37 million new SNPs), 458,224 short insertions and deletions, and 58,174 structural variants. Genetic differences between DA, F344, and BN, including high-impact SNPs and short insertions and deletions affecting >2500 genes, are likely to account for most of the phenotypic variation between these strains. The new DA and F344 genome sequencing data should facilitate gene discovery efforts in rat models of human disease. PMID:23695301

Whole-genome sequences of DA and F344 rats with different susceptibilities to arthritis, autoimmunity, inflammation and cancer.

PubMed

Guo, Xiaosen; Brenner, Max; Zhang, Xuemei; Laragione, Teresina; Tai, Shuaishuai; Li, Yanhong; Bu, Junjie; Yin, Ye; Shah, Anish A; Kwan, Kevin; Li, Yingrui; Jun, Wang; Gulko, Pércio S

2013-08-01

DA (D-blood group of Palm and Agouti, also known as Dark Agouti) and F344 (Fischer) are two inbred rat strains with differences in several phenotypes, including susceptibility to autoimmune disease models and inflammatory responses. While these strains have been extensively studied, little information is available about the DA and F344 genomes, as only the Brown Norway (BN) and spontaneously hypertensive rat strains have been sequenced to date. Here we report the sequencing of the DA and F344 genomes using next-generation Illumina paired-end read technology and the first de novo assembly of a rat genome. DA and F344 were sequenced with an average depth of 32-fold, covered 98.9% of the BN reference genome, and included 97.97% of known rat ESTs. New sequences could be assigned to 59 million positions with previously unknown data in the BN reference genome. Differences between DA, F344, and BN included 19 million positions in novel scaffolds, 4.09 million single nucleotide polymorphisms (SNPs) (including 1.37 million new SNPs), 458,224 short insertions and deletions, and 58,174 structural variants. Genetic differences between DA, F344, and BN, including high-impact SNPs and short insertions and deletions affecting >2500 genes, are likely to account for most of the phenotypic variation between these strains. The new DA and F344 genome sequencing data should facilitate gene discovery efforts in rat models of human disease.

Genetic Diversity among 3-Chloroaniline- and Aniline-Degrading Strains of the Comamonadaceae

PubMed Central

Boon, Nico; Goris, Johan; De Vos, Paul; Verstraete, Willy; Top, Eva M.

2001-01-01

We examined the diversity of the plasmids and of the gene tdnQ, involved in the oxidative deamination of aniline, in five bacterial strains that are able to metabolize both aniline and 3-chloroaniline (3-CA). Three strains have been described and identified previously, i.e., Comamonas testosteroni I2 and Delftia acidovorans CA28 and BN3.1. Strains LME1 and B8c were isolated in this study from linuron-treated soil and from a wastewater treatment plant, respectively, and were both identified as D. acidovorans. Both Delftia and Comamonas belong to the family Comamonadaceae. All five strains possess a large plasmid of ca. 100 kb, but the plasmids from only four strains could be transferred to a recipient strain by selection on aniline or 3-CA as a sole source of carbon and/or nitrogen. Plasmid transfer experiments and Southern hybridization revealed that the plasmid of strain I2 was responsible for total aniline but not 3-CA degradation, while the plasmids of strains LME1 and B8c were responsible only for the oxidative deamination of aniline. Several transconjugant clones that had received the plasmid from strain CA28 showed different degradative capacities: all transconjugants could use aniline as a nitrogen source, while only some of the transconjugants could deaminate 3-CA. For all four plasmids, the IS1071 insertion sequence of Tn5271 was found to be located on a 1.4-kb restriction fragment, which also hybridized with the tdnQ probe. This result suggests the involvement of this insertion sequence element in the dissemination of aniline degradation genes in the environment. By use of specific primers for the tdnQ gene from Pseudomonas putida UCC22, the diversity of the PCR-amplified fragments in the five strains was examined by denaturing gradient gel electrophoresis (DGGE). With DGGE, three different clusters of the tdnQ fragment could be distinguished. Sequencing data showed that the tdnQ sequences of I2, LME1, B8c, and CA28 were very closely related, while the tdnQ sequences of BN3.1 and P. putida UCC22 were only about 83% identical to the other sequences. Northern hybridization revealed that the tdnQ gene is transcribed only in the presence of aniline and not when only 3-CA is present. PMID:11229899

Whole genome sequencing and phylogenetic analysis of Bluetongue virus serotype 2 strains isolated in the Americas including a novel strain from the western United States

USDA-ARS?s Scientific Manuscript database

Bluetongue is caused by an arbovirus which produces widespread edema and tissue necrosis in domestic and wild ruminants that can be fatal. Bluetongue virus serotypes 10, 11, 13, and 17 are typically found throughout the United States (US), while serotype 2 was previously only detected in the southea...

Study of canine parvovirus evolution: comparative analysis of full-length VP2 gene sequences from Argentina and international field strains.

PubMed

Gallo Calderón, Marina; Wilda, Maximiliano; Boado, Lorena; Keller, Leticia; Malirat, Viviana; Iglesias, Marcela; Mattion, Nora; La Torre, Jose

2012-02-01

The continuous emergence of new strains of canine parvovirus (CPV), poorly protected by current vaccination, is a concern among breeders, veterinarians, and dog owners around the world. Therefore, the understanding of the genetic variation in emerging CPV strains is crucial for the design of disease control strategies, including vaccines. In this paper, we obtained the sequences of the full-length gene encoding for the main capsid protein (VP2) of 11 canine parvovirus type 2 (CPV-2) Argentine representative field strains, selected from a total of 75 positive samples studied in our laboratory in the last 9 years. A comparative sequence analysis was performed on 9 CPV-2c, one CPV-2a, and one CPV-2b Argentine strains with respect to international strains reported in the GenBank database. In agreement with previous reports, a high degree of identity was found among CPV-2c Argentine strains (99.6-100% and 99.7-100% at nucleotide and amino acid levels, respectively). However, the appearance of a new substitution in the 440 position (T440A) in four CPV-2c Argentine strains obtained after the year 2009 gives support to the variability observed for this position located within the VP2, three-fold spike. This is the first report on the genetic characterization of the full-length VP2 gene of emerging CPV strains in South America and shows that all the Argentine CPV-2c isolates cluster together with European and North American CPV-2c strains.

Differential expression of the virulence-associated protein p57 and characterization of its duplicated gene rosa in virulent and attenuated strains of Renibacterium salmoninarum

USGS Publications Warehouse

O'Farrell, C. L.; Strom, M.S.

1999-01-01

Virulence mechanisms utilized by the salmonid fish pathogen Renibacterium salmoninarum are poorly understood. One potential virulence factor is p57 (also designated MSA for major soluble antigen), an abundant 57 kDa soluble protein that is predominately localized on the bacterial cell surface with significant levels released into the extracellular milieu. Previous studies of an attenuated strain, MT 239, indicated that it differs from virulent strains in the amount of surface-associated p57. In this report, we show overall expression of p57 in R. salmoninarum MT 239 is considerably reduced as compared to a virulent strain, ATCC 33209. The amount of cell-associated p57 is decreased while the level of p57 in the culture supernatant is nearly equivalent between the strains. To determine if lowered amount of cell-associated p57 was due to a sequence defect in p57, a genetic comparison was performed. Two copies of the gene encoding p57 (msa1 and msa2) were found in 33209 and MT 239, as well as in several other virulent isolates. Both copies from 33209 and MT 239 were cloned and sequenced and found to be identical to each other, and identical between the 2 strains. A comparison of msa1 and msa2 within each strain showed that their sequences diverge 40 base pairs 5, to the open reading frame, while sequences 3' to the open reading frame are essentially identical for at least 225 base pairs. Northern blot analysis showed no difference in steady state levels of rosa mRNA between the 2 strains. These data suggest that while cell-surface localization of p57 may be important for R. salmoninarum virulence, the differences in localization, and total p57 expression between 33209 anti MT 239 are not due to differences in rosa sequence or differences in steady state transcript levels.

Differential recognition of the ORF2 region in a complete genome sequence of porcine circovirus type 2 (PCV2) isolated from boar bone marrow in Korea.

PubMed

Kweon, Chang-Hee; Nguyen, Lien Thi Kim; Yoo, Mi-Sun; Kang, Seung-Won

2015-09-15

Porcine circovirus type 2 (PCV2) is the causative agent of post-weaning multisystemic wasting syndrome (PMWS) in swine. Here, a phylogenetic tree was constructed using PCV2 nucleotide sequences derived from the bone marrow of Korean boar and previously reported PCV2 sequences isolated from various countries. PCV2 from Korean boar bone marrow (KC188796) was classified into the group containing PCV2a-Canada and other PCV2 strain from Korea. While the ORF1 region of the PCV2 genome was highly conserved, ORF2 (the capsid protein coding region) was relatively variable. The nucleotide sequences for bone marrow-derived PCV2 were 93.4-99.0% homologous to the other reference sequences. The deduced amino acid sequences for the ORF1 and ORF2 coding regions were 97.4-99.3% and 84.5-97.4% homologous with the other reference strains, respectively, indicating that KC188796 did not differ markedly from the other PCV2 strains. Phylogenetic analysis demonstrated that bone marrow-derived PCV2 was highly similar to PCV2a from Canada and may be related to persistent PCV2 infections in swine. Copyright © 2015 Elsevier B.V. All rights reserved.

Occurrence and Phylogenetic Diversity of Sphingomonas Strains in Soils Contaminated with Polycyclic Aromatic Hydrocarbons

PubMed Central

Leys, Natalie M. E. J.; Ryngaert, Annemie; Bastiaens, Leen; Verstraete, Willy; Top, Eva M.; Springael, Dirk

2004-01-01

Bacterial strains of the genus Sphingomonas are often isolated from contaminated soils for their ability to use polycyclic aromatic hydrocarbons (PAH) as the sole source of carbon and energy. The direct detection of Sphingomonas strains in contaminated soils, either indigenous or inoculated, is, as such, of interest for bioremediation purposes. In this study, a culture-independent PCR-based detection method using specific primers targeting the Sphingomonas 16S rRNA gene combined with denaturing gradient gel electrophoresis (DGGE) was developed to assess Sphingomonas diversity in PAH-contaminated soils. PCR using the new primer pair on a set of template DNAs of different bacterial genera showed that the method was selective for bacteria belonging to the family Sphingomonadaceae. Single-band DGGE profiles were obtained for most Sphingomonas strains tested. Strains belonging to the same species had identical DGGE fingerprints, and in most cases, these fingerprints were typical for one species. Inoculated strains could be detected at a cell concentration of 104 CFU g of soil−1. The analysis of Sphingomonas population structures of several PAH-contaminated soils by the new PCR-DGGE method revealed that soils containing the highest phenanthrene concentrations showed the lowest Sphingomonas diversity. Sequence analysis of cloned PCR products amplified from soil DNA revealed new 16S rRNA gene Sphingomonas sequences significantly different from sequences from known cultivated isolates (i.e., sequences from environmental clones grouped phylogenetically with other environmental clone sequences available on the web and that possibly originated from several potential new species). In conclusion, the newly designed Sphingomonas-specific PCR-DGGE detection technique successfully analyzed the Sphingomonas communities from polluted soils at the species level and revealed different Sphingomonas members not previously detected by culture-dependent detection techniques. PMID:15066784

'Candidatus Phytoplasma palmicola', associated with a lethal yellowing-type disease of coconut (Cocos nucifera L.) in Mozambique.

PubMed

Harrison, Nigel A; Davis, Robert E; Oropeza, Carlos; Helmick, Ericka E; Narváez, María; Eden-Green, Simon; Dollet, Michel; Dickinson, Matthew

2014-06-01

In this study, the taxonomic position and group classification of the phytoplasma associated with a lethal yellowing-type disease (LYD) of coconut (Cocos nucifera L.) in Mozambique were addressed. Pairwise similarity values based on alignment of nearly full-length 16S rRNA gene sequences (1530 bp) revealed that the Mozambique coconut phytoplasma (LYDM) shared 100% identity with a comparable sequence derived from a phytoplasma strain (LDN) responsible for Awka wilt disease of coconut in Nigeria, and shared 99.0-99.6% identity with 16S rRNA gene sequences from strains associated with Cape St Paul wilt (CSPW) disease of coconut in Ghana and Côte d'Ivoire. Similarity scores further determined that the 16S rRNA gene of the LYDM phytoplasma shared <97.5% sequence identity with all previously described members of 'Candidatus Phytoplasma'. The presence of unique regions in the 16S rRNA gene sequence distinguished the LYDM phytoplasma from all currently described members of 'Candidatus Phytoplasma', justifying its recognition as the reference strain of a novel taxon, 'Candidatus Phytoplasma palmicola'. Virtual RFLP profiles of the F2n/R2 portion (1251 bp) of the 16S rRNA gene and pattern similarity coefficients delineated coconut LYDM phytoplasma strains from Mozambique as novel members of established group 16SrXXII, subgroup A (16SrXXII-A). Similarity coefficients of 0.97 were obtained for comparisons between subgroup 16SrXXII-A strains and CSPW phytoplasmas from Ghana and Côte d'Ivoire. On this basis, the CSPW phytoplasma strains were designated members of a novel subgroup, 16SrXXII-B.

Three distinct clades of cultured heterocystous cyanobacteria constitute the dominant N2-fixing members of biological soil crusts of the Colorado Plateau, USA

USGS Publications Warehouse

Yeager, C.M.; Kornosky, J.L.; Morgan, R.E.; Cain, E.C.; Garcia-Pichel, F.; Housman, D.C.; Belnap, J.; Kuske, C.R.

2007-01-01

The identity of the numerically dominant N2-fixing bacteria in biological soil crusts of the Colorado Plateau region and two outlying areas was determined using multiple approaches, to link the environmental diversity of nifH gene sequences to cultured bacterial isolates from the regions. Of the nifH sequence-types detected in soil crusts of the Colorado Plateau, 89% (421/473) were most closely related to nifH signature sequences from cyanobacteria of the order Nostocales. N2-fixing cyanobacterial strains were cultured from crusts and their morphotypes, 16S rRNA gene and nifH gene sequences were characterized. The numerically dominant diazotrophs in the Colorado Plateau crusts fell within three clades of heterocystous cyanobacteria. Two clades are well-represented by phylogenetically and morphologically coherent strains, corresponding to the descriptions of Nostoc commune and Scytonema hyalinum, which are widely recognized as important N2-fixing components of soil crusts. A third, previously-overlooked clade was represented by a phylogenetically coherent but morphologically diverse group of strains that encompass the morphogenera Tolypothrix and Spirirestis. Many of the strains in each of these groups contained at least two nifH copies that represent different clusters in the nifH environmental survey. ?? 2007 Federation of European Microbiological Societies.

Genome Structural Diversity among 31 Bordetella pertussis Isolates from Two Recent U.S. Whooping Cough Statewide Epidemics.

PubMed

Bowden, Katherine E; Weigand, Michael R; Peng, Yanhui; Cassiday, Pamela K; Sammons, Scott; Knipe, Kristen; Rowe, Lori A; Loparev, Vladimir; Sheth, Mili; Weening, Keeley; Tondella, M Lucia; Williams, Margaret M

2016-01-01

During 2010 and 2012, California and Vermont, respectively, experienced statewide epidemics of pertussis with differences seen in the demographic affected, case clinical presentation, and molecular epidemiology of the circulating strains. To overcome limitations of the current molecular typing methods for pertussis, we utilized whole-genome sequencing to gain a broader understanding of how current circulating strains are causing large epidemics. Through the use of combined next-generation sequencing technologies, this study compared de novo, single-contig genome assemblies from 31 out of 33 Bordetella pertussis isolates collected during two separate pertussis statewide epidemics and 2 resequenced vaccine strains. Final genome architecture assemblies were verified with whole-genome optical mapping. Sixteen distinct genome rearrangement profiles were observed in epidemic isolate genomes, all of which were distinct from the genome structures of the two resequenced vaccine strains. These rearrangements appear to be mediated by repetitive sequence elements, such as high-copy-number mobile genetic elements and rRNA operons. Additionally, novel and previously identified single nucleotide polymorphisms were detected in 10 virulence-related genes in the epidemic isolates. Whole-genome variation analysis identified state-specific variants, and coding regions bearing nonsynonymous mutations were classified into functional annotated orthologous groups. Comprehensive studies on whole genomes are needed to understand the resurgence of pertussis and develop novel tools to better characterize the molecular epidemiology of evolving B. pertussis populations. IMPORTANCE Pertussis, or whooping cough, is the most poorly controlled vaccine-preventable bacterial disease in the United States, which has experienced a resurgence for more than a decade. Once viewed as a monomorphic pathogen, B. pertussis strains circulating during epidemics exhibit diversity visible on a genome structural level, previously undetectable by traditional sequence analysis using short-read technologies. For the first time, we combine short- and long-read sequencing platforms with restriction optical mapping for single-contig, de novo assembly of 31 isolates to investigate two geographically and temporally independent U.S. pertussis epidemics. These complete genomes reshape our understanding of B. pertussis evolution and strengthen molecular epidemiology toward one day understanding the resurgence of pertussis.

Bioaccessible peptides released by in vitro gastrointestinal digestion of fermented goat milks.

PubMed

Moreno-Montoro, Miriam; Jauregi, Paula; Navarro-Alarcón, Miguel; Olalla-Herrera, Manuel; Giménez-Martínez, Rafael; Amigo, Lourdes; Miralles, Beatriz

2018-06-01

In this study, ultrafiltered goat milks fermented with the classical starter bacteria Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus salivarus subsp. thermophilus or with the classical starter plus the Lactobacillus plantarum C4 probiotic strain were analyzed using ultra-high performance liquid chromatography-quadrupole-time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) and/or high performance liquid chromatography-ion trap (HPLC-IT-MS/MS). Partial overlapping of the identified sequences with regard to fermentation culture was observed. Evaluation of the cleavage specificity suggested a lower proteolytic activity of the probiotic strain. Some of the potentially identified peptides had been previously reported as angiotensin-converting enzyme (ACE) inhibitory, antioxidant, and antibacterial and might account for the in vitro activity previously reported for these fermented milks. Simulated digestion of the products was conducted in the presence of a dialysis membrane to retrieve the bioaccessible peptide fraction. Some sequences with reported physiological activity resisted digestion but were found in the non-dialyzable fraction. However, new forms released by digestion, such as the antioxidant α s1 -casein 144 YFYPQL 149 , the antihypertensive α s2 -casein 90 YQKFPQY 96 , and the antibacterial α s2 -casein 165 LKKISQ 170 , were found in the dialyzable fraction of both fermented milks. Moreover, in the fermented milk including the probiotic strain, the k-casein dipeptidyl peptidase IV inhibitor (DPP-IV) 51 INNQFLPYPY 60 as well as additional ACE inhibitory or antioxidant sequences could be identified. With the aim of anticipating further biological outcomes, quantitative structure activity relationship (QSAR) analysis was applied to the bioaccessible fragments and led to potential ACE inhibitory sequences being proposed. Graphical abstract Ultrafiltered goat milks were fermented with the classical starter bacteria (St) and with St plus the L. plantarum C4 probiotic strain. Samples were analyzed using HPLC-IT-MS/MS and UPLC-Q-TOF-MS/MS. After simulated digestion and dialysis, some of the active sequences remained and new peptides with reported beneficial activities were released.

Improved detection of genetic markers of antimicrobial resistance by hybridization probe-based melting curve analysis using primers to mask proximal mutations: examples include the influenza H275Y substitution.

PubMed

Whiley, David M; Jacob, Kevin; Nakos, Jennifer; Bletchly, Cheryl; Nimmo, Graeme R; Nissen, Michael D; Sloots, Theo P

2012-06-01

Numerous real-time PCR assays have been described for detection of the influenza A H275Y alteration. However, the performance of these methods can be undermined by sequence variation in the regions flanking the codon of interest. This is a problem encountered more broadly in microbial diagnostics. In this study, we developed a modification of hybridization probe-based melting curve analysis, whereby primers are used to mask proximal mutations in the sequence targets of hybridization probes, so as to limit the potential for sequence variation to interfere with typing. The approach was applied to the H275Y alteration of the influenza A (H1N1) 2009 strain, as well as a Neisseria gonorrhoeae mutation associated with antimicrobial resistance. Assay performances were assessed using influenza A and N. gonorrhoeae strains characterized by DNA sequencing. The modified hybridization probe-based approach proved successful in limiting the effects of proximal mutations, with the results of melting curve analyses being 100% consistent with the results of DNA sequencing for all influenza A and N. gonorrhoeae strains tested. Notably, these included influenza A and N. gonorrhoeae strains exhibiting additional mutations in hybridization probe targets. Of particular interest was that the H275Y assay correctly typed influenza A strains harbouring a T822C nucleotide substitution, previously shown to interfere with H275Y typing methods. Overall our modified hybridization probe-based approach provides a simple means of circumventing problems caused by sequence variation, and offers improved detection of the influenza A H275Y alteration and potentially other resistance mechanisms.

Cutaneous Granulomas in Dolphins Caused by Novel Uncultivated Paracoccidioides brasiliensis

PubMed Central

Vilela, Raquel; Bossart, Gregory D.; St. Leger, Judy A.; Dalton, Leslie M.; Reif, John S.; Schaefer, Adam M.; McCarthy, Peter J.; Fair, Patricia A.

2016-01-01

Cutaneous granulomas in dolphins were believed to be caused by Lacazia loboi, which also causes a similar disease in humans. This hypothesis was recently challenged by reports that fungal DNA sequences from dolphins grouped this pathogen with Paracoccidioides brasiliensis. We conducted phylogenetic analysis of fungi from 6 bottlenose dolphins (Tursiops truncatus) with cutaneous granulomas and chains of yeast cells in infected tissues. Kex gene sequences of P. brasiliensis from dolphins showed 100% homology with sequences from cultivated P. brasiliensis, 73% with those of L. loboi, and 93% with those of P. lutzii. Parsimony analysis placed DNA sequences from dolphins within a cluster with human P. brasiliensis strains. This cluster was the sister taxon to P. lutzii and L. loboi. Our molecular data support previous findings and suggest that a novel uncultivated strain of P. brasiliensis restricted to cutaneous lesions in dolphins is probably the cause of lacaziosis/lobomycosis, herein referred to as paracoccidioidomycosis ceti. PMID:27869614

Cutaneous Granulomas in Dolphins Caused by Novel Uncultivated Paracoccidioides brasiliensis.

PubMed

Vilela, Raquel; Bossart, Gregory D; St Leger, Judy A; Dalton, Leslie M; Reif, John S; Schaefer, Adam M; McCarthy, Peter J; Fair, Patricia A; Mendoza, Leonel

2016-12-01

Cutaneous granulomas in dolphins were believed to be caused by Lacazia loboi, which also causes a similar disease in humans. This hypothesis was recently challenged by reports that fungal DNA sequences from dolphins grouped this pathogen with Paracoccidioides brasiliensis. We conducted phylogenetic analysis of fungi from 6 bottlenose dolphins (Tursiops truncatus) with cutaneous granulomas and chains of yeast cells in infected tissues. Kex gene sequences of P. brasiliensis from dolphins showed 100% homology with sequences from cultivated P. brasiliensis, 73% with those of L. loboi, and 93% with those of P. lutzii. Parsimony analysis placed DNA sequences from dolphins within a cluster with human P. brasiliensis strains. This cluster was the sister taxon to P. lutzii and L. loboi. Our molecular data support previous findings and suggest that a novel uncultivated strain of P. brasiliensis restricted to cutaneous lesions in dolphins is probably the cause of lacaziosis/lobomycosis, herein referred to as paracoccidioidomycosis ceti.

Actinomyces gaoshouyii sp. nov., isolated from plateau pika (Ochotona curzoniae).

PubMed

Meng, Xiangli; Wang, Yiting; Lu, Shan; Lai, Xin-He; Jin, Dong; Yang, Jing; Xu, Jianguo

2017-09-01

Two strains (pika_113T and pika_114) of a previously undescribed Actinomyces-like bacterium were recovered from the intestinal contents of plateau pika (Ochotona curzoniae) on the Tibet-Qinghai Plateau, China. Results from biochemical characterization indicated that the two strains were phenotypically homogeneous and distinct from other previously described species of the genus Actinomyces. Based on the comparison of 16S rRNA gene sequences and genome analysis, the bacteria were determined to be a hitherto unknown subline within the genus Actinomyces, being most closely related to type strains of Actinomyces denticolens and Actinomyces timonensis with a respective 97.2 and 97.1 % similarity in their 16S rRNA gene sequences. Phylogenetic analyses confirmed that pika_113T was well separated from any other recognized species of the genus Actinomyces and within the cluster with A. denticolens and A. timonensis. The genome of strain pika_113T displayed less than 42 % relatedness in DNA-DNA hybridization with all the available genomes of existing species of the genus Actinomyces in the NCBI database. Collectively, based on the phenotypic characteristics and phylogenetic analyses results, we propose the novel isolates as representatives of Actinomyces gaoshouyii sp. nov. The type strain of Actinomyces gaoshouyii is pika_113T (=CGMCC 4.7372T=DSM 104049T), with a genomic DNA G+C content of 71 mol%.

Characterization of Actinomyces Isolates from Infected Root Canals of Teeth: Description of Actinomyces radicidentis sp. nov.

PubMed Central

Collins, Matthew D.; Hoyles, Lesley; Kalfas, Sotos; Sundquist, Goran; Monsen, Tor; Nikolaitchouk, Natalia; Falsen, Enevold

2000-01-01

Two strains of a previously undescribed Actinomyces-like bacterium were recovered in pure culture from infected root canals of teeth. Analysis by biochemical testing and polyacrylamide gel electrophoresis of whole-cell proteins indicated that the strains closely resembled each other phenotypically but were distinct from previously described Actinomyces and Arcanobacterium species. Comparative 16S rRNA gene-sequencing studies showed the bacterium to be a hitherto unknown subline within a group of Actinomyces species which includes Actinomyces bovis, the type species of the genus. Based on phylogenetic and phenotypic evidence, we propose that the unknown bacterium isolated from human clinical specimens be classified as Actinomyces radicidentis sp. nov. The type strain of Actinomyces radicidentis is CCUG 36733. PMID:10970390

[Entification of the Rubella virus genotype 1H in Western Siberia].

PubMed

Seregin, S V; Babkin, I V; Petrova, I D; Iashina, L N; Malkova, E M; Petrov, V S

2011-01-01

Molecular epidemiological study of novel strain of Rubella virus isolated during the outbreak in Western Siberia in 2004 was described. Detailed phylogenetic analysis performed based upon entire SP-region, which encodes all three Rubella structural proteins (C, E2, and E1), was implemented. This analysis provides characterization of this strain and classifies it as 1H genotype, thereby correcting previous classification of this strain based upon shorter nucleotide sequence, only encoding E1 protein. Therefore, this study identified the genotype of the Rubella virus not previously detected in Western Siberia (and even entire Russian Federation), which highlights the importance of more extensive characterization of genetic variability of the Rubella virus, especially with regard to potential influence of vaccination on the Rubella virus mutagenesis.

Phaeobacterium nitratireducens gen. nov., sp. nov., a phototrophic gammaproteobacterium isolated from a mangrove forest sediment sample.

PubMed

Nupur; Tanuku, Naga Radha Srinivas; Shinichi, Takaichi; Pinnaka, Anil Kumar

2015-08-01

A novel brown-coloured, Gram-negative-staining, rod-shaped, motile, phototrophic, purple sulfur bacterium, designated strain AK40T, was isolated in pure culture from a sediment sample collected from Coringa mangrove forest, India. Strain AK40T contained bacteriochlorophyll a and carotenoids of the rhodopin series as major photosynthetic pigments. Strain AK40T was able to grow photoheterotrophically and could utilize a number of organic substrates. It was unable to grow photoautotrophically and did not utilize sulfide or thiosulfate as electron donors. Thiamine and riboflavin were required for growth. The dominant fatty acids were C12 : 0, C16 : 0, C18 : 1ω7c and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH). The polar lipid profile of strain AK40T was found to contain diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and eight unidentified lipids. Q-10 was the predominant respiratory quinone. The DNA G+C content of strain AK40T was 65.5 mol%. 16S rRNA gene sequence comparisons indicated that the isolate represented a member of the family Chromatiaceae within the class Gammaproteobacteria. 16S rRNA gene sequence analysis indicated that strain AK40T was closely related to Phaeochromatium fluminis, with 95.2% pairwise sequence similarity to the type strain; sequence similarity to strains of other species of the family was 90.8-94.8%. Based on the sequence comparison data, strain AK40T was positioned distinctly outside the group formed by the genera Phaeochromatium, Marichromatium, Halochromatium, Thiohalocapsa, Rhabdochromatium and Thiorhodovibrio. Distinct morphological, physiological and genotypic differences from previously described taxa supported the classification of this isolate as a representative of a novel species in a new genus, for which the name Phaeobacterium nitratireducens gen. nov., sp. nov. is proposed. The type strain of Phaeobacterium nitratireducens is AK40T ( = JCM 19219T = MTCC 11824T).

Volcanic Soils as Sources of Novel CO-Oxidizing Paraburkholderia and Burkholderia: Paraburkholderia hiiakae sp. nov., Paraburkholderia metrosideri sp. nov., Paraburkholderia paradisi sp. nov., Paraburkholderia peleae sp. nov., and Burkholderia alpina sp. nov. a Member of the Burkholderia cepacia Complex

PubMed Central

Weber, Carolyn F.; King, Gary M.

2017-01-01

Previous studies showed that members of the Burkholderiales were important in the succession of aerobic, molybdenum-dependent CO oxidizing-bacteria on volcanic soils. During these studies, four isolates were obtained from Kilauea Volcano (Hawai‘i, USA); one strain was isolated from Pico de Orizaba (Mexico) during a separate study. Based on 16S rRNA gene sequence similarities, the Pico de Orizaba isolate and the isolates from Kilauea Volcano were provisionally assigned to the genera Burkholderia and Paraburkholderia, respectively. Each of the isolates possessed a form I coxL gene that encoded the catalytic subunit of carbon monoxide dehydrogenase (CODH); none of the most closely related type strains possessed coxL or oxidized CO. Genome sequences for Paraburkholderia type strains facilitated an analysis of 16S rRNA gene sequence similarities and average nucleotide identities (ANI). ANI did not exceed 95% (the recommended cutoff for species differentiation) for any of the pairwise comparisons among 27 reference strains related to the new isolates. However, since the highest 16S rRNA gene sequence similarity among this set of reference strains was 98.93%, DNA-DNA hybridizations (DDH) were performed for two isolates whose 16S rRNA gene sequence similarities with their nearest phylogenetic neighbors were 98.96 and 99.11%. In both cases DDH values were <16%. Based on multiple variables, four of the isolates represent novel species within the Paraburkholderia: Paraburkholderia hiiakae sp. nov. (type strain I2T = DSM 28029T = LMG 27952T); Paraburkholderia paradisi sp. nov. (type strain WAT = DSM 28027T = LMG 27949T); Paraburkholderia peleae sp. nov. (type strain PP52-1T = DSM 28028T = LMG 27950T); and Paraburkholderia metrosideri sp. nov. (type strain DNBP6-1T = DSM 28030T = LMG 28140T). The remaining isolate represents the first CO-oxidizing member of the Burkholderia cepacia complex: Burkholderia alpina sp. nov. (type strain PO-04-17-38T = DSM 28031T = LMG 28138T). PMID:28270796

Volcanic Soils as Sources of Novel CO-Oxidizing Paraburkholderia and Burkholderia: Paraburkholderia hiiakae sp. nov., Paraburkholderia metrosideri sp. nov., Paraburkholderia paradisi sp. nov., Paraburkholderia peleae sp. nov., and Burkholderia alpina sp. nov. a Member of the Burkholderia cepacia Complex.

PubMed

Weber, Carolyn F; King, Gary M

2017-01-01

Previous studies showed that members of the Burkholderiales were important in the succession of aerobic, molybdenum-dependent CO oxidizing-bacteria on volcanic soils. During these studies, four isolates were obtained from Kilauea Volcano (Hawai'i, USA); one strain was isolated from Pico de Orizaba (Mexico) during a separate study. Based on 16S rRNA gene sequence similarities, the Pico de Orizaba isolate and the isolates from Kilauea Volcano were provisionally assigned to the genera Burkholderia and Paraburkholderia , respectively. Each of the isolates possessed a form I coxL gene that encoded the catalytic subunit of carbon monoxide dehydrogenase (CODH); none of the most closely related type strains possessed coxL or oxidized CO. Genome sequences for Paraburkholderia type strains facilitated an analysis of 16S rRNA gene sequence similarities and average nucleotide identities (ANI). ANI did not exceed 95% (the recommended cutoff for species differentiation) for any of the pairwise comparisons among 27 reference strains related to the new isolates. However, since the highest 16S rRNA gene sequence similarity among this set of reference strains was 98.93%, DNA-DNA hybridizations (DDH) were performed for two isolates whose 16S rRNA gene sequence similarities with their nearest phylogenetic neighbors were 98.96 and 99.11%. In both cases DDH values were <16%. Based on multiple variables, four of the isolates represent novel species within the Paraburkholderia : Paraburkholderia hiiakae sp. nov. (type strain I2 T = DSM 28029 T = LMG 27952 T ); Paraburkholderia paradisi sp. nov. (type strain WA T = DSM 28027 T = LMG 27949 T ); Paraburkholderia peleae sp. nov. (type strain PP52-1 T = DSM 28028 T = LMG 27950 T ); and Paraburkholderia metrosideri sp. nov. (type strain DNBP6-1 T = DSM 28030 T = LMG 28140 T ). The remaining isolate represents the first CO-oxidizing member of the Burkholderia cepacia complex: Burkholderia alpina sp. nov. (type strain PO-04-17-38 T = DSM 28031 T = LMG 28138 T ).

Application of a case-control study design to investigate genotypic signatures of HIV-1 transmission.

PubMed

Mota, Talia M; Murray, John M; Center, Rob J; Purcell, Damian F J; McCaw, James M

2012-06-25

The characterization of HIV-1 transmission strains may inform the design of an effective vaccine. Shorter variable loops with fewer predicted glycosites have been suggested as signatures enriched in envelope sequences derived during acute HIV-1 infection. Specifically, a transmission-linked lack of glycosites within the V1 and V2 loops of gp120 provides greater access to an α4β7 binding motif, which promotes the establishment of infection. Also, a histidine at position 12 in the leader sequence of Env has been described as a transmission signature that is selected against during chronic infection. The purpose of this study is to measure the association of the presence of an α4β7 binding motif, the number of N-linked glycosites, the length of the variable loops, and the prevalence of histidine at position 12 with HIV-1 transmission. A case-control study design was used to measure the prevalence of these variables between subtype B and C transmission sequences and frequency-matched randomly-selected sequences derived from chronically infected controls. Subtype B transmission strains had shorter V3 regions than chronic strains (p = 0.031); subtype C transmission strains had shorter V1 loops than chronic strains (p = 0.047); subtype B transmission strains had more V3 loop glycosites (p = 0.024) than chronic strains. Further investigation showed that these statistically significant results were unlikely to be biologically meaningful. Also, there was no difference observed in the prevalence of a histidine at position 12 among transmission strains and controls of either subtype. Although a genetic bottleneck is observed after HIV-1 transmission, our results indicate that summary characteristics of Env hypothesised to be important in transmission are not divergent between transmission and chronic strains of either subtype. The success of a transmission strain to initiate infection may be a random event from the divergent pool of donor viral sequences. The characteristics explored through this study are important, but may not function as genotypic signatures of transmission as previously described.

De novo sequencing, assembly and analysis of the genome of the laboratory strain Saccharomyces cerevisiae CEN.PK113-7D, a model for modern industrial biotechnology

PubMed Central

2012-01-01

Saccharomyces cerevisiae CEN.PK 113-7D is widely used for metabolic engineering and systems biology research in industry and academia. We sequenced, assembled, annotated and analyzed its genome. Single-nucleotide variations (SNV), insertions/deletions (indels) and differences in genome organization compared to the reference strain S. cerevisiae S288C were analyzed. In addition to a few large deletions and duplications, nearly 3000 indels were identified in the CEN.PK113-7D genome relative to S288C. These differences were overrepresented in genes whose functions are related to transcriptional regulation and chromatin remodelling. Some of these variations were caused by unstable tandem repeats, suggesting an innate evolvability of the corresponding genes. Besides a previously characterized mutation in adenylate cyclase, the CEN.PK113-7D genome sequence revealed a significant enrichment of non-synonymous mutations in genes encoding for components of the cAMP signalling pathway. Some phenotypic characteristics of the CEN.PK113-7D strains were explained by the presence of additional specific metabolic genes relative to S288C. In particular, the presence of the BIO1 and BIO6 genes correlated with a biotin prototrophy of CEN.PK113-7D. Furthermore, the copy number, chromosomal location and sequences of the MAL loci were resolved. The assembled sequence reveals that CEN.PK113-7D has a mosaic genome that combines characteristics of laboratory strains and wild-industrial strains. PMID:22448915

Phylogenetic analysis of Austrian canine distemper virus strains from clinical samples from dogs and wild carnivores.

PubMed

Benetka, V; Leschnik, M; Affenzeller, N; Möstl, K

2011-04-09

Austrian field cases of canine distemper (14 dogs, one badger [Meles meles] and one stone marten [Martes foina]) from 2002 to 2007 were investigated and the case histories were summarised briefly. Phylogenetic analysis of fusion (F) and haemagglutinin (H) gene sequences revealed different canine distemper virus (CDV) lineages circulating in Austria. The majority of CDV strains detected from 2002 to 2004 were well embedded in the European lineage. One Austrian canine sample detected in 2003, with a high similarity to Hungarian sequences from 2005 to 2006, could be assigned to the Arctic group (phocine distemper virus type 2-like). The two canine sequences from 2007 formed a clearly distinct group flanked by sequences detected previously in China and the USA on an intermediate position between the European wildlife and the Asia-1 cluster. The Austrian wildlife strains (2006 and 2007) could be assigned to the European wildlife group and were most closely related to, yet clearly different from, the 2007 canine samples. To elucidate the epidemiological role of Austrian wildlife in the transmission of the disease to dogs and vice versa, H protein residues related to receptor and host specificity (residues 530 and 549) were analysed. All samples showed the amino acids expected for their host of origin, with the exception of a canine sequence from 2007, which had an intermediate position between wildlife and canine viral strains. In the period investigated, canine strains circulating in Austria could be assigned to four different lineages reflecting both a high diversity and probably different origins of virus introduction to Austria in different years.

Paenibacillus phoenicis sp. nov., isolated from the Phoenix Lander assembly facility and a subsurface molybdenum mine.

PubMed

Benardini, James N; Vaishampayan, Parag A; Schwendner, Petra; Swanner, Elizabeth; Fukui, Youhei; Osman, Sharif; Satomi, Masakata; Venkateswaran, Kasthuri

2011-06-01

A novel Gram-positive, motile, endospore-forming, aerobic bacterium was isolated from the NASA Phoenix Lander assembly clean room that exhibits 100 % 16S rRNA gene sequence similarity to two strains isolated from a deep subsurface environment. All strains are rod-shaped, endospore-forming bacteria, whose endospores are resistant to UV radiation up to 500 J m(-2). A polyphasic taxonomic study including traditional phenotypic tests, fatty acid analysis, 16S rRNA gene sequencing and DNA-DNA hybridization analysis was performed to characterize these novel strains. The 16S rRNA gene sequencing convincingly grouped these novel strains within the genus Paenibacillus as a separate cluster from previously described species. The similarity of 16S rRNA gene sequences among the novel strains was identical but only 98.1 to 98.5 % with their nearest neighbours Paenibacillus barengoltzii ATCC BAA-1209(T) and Paenibacillus timonensis CIP 108005(T). The menaquinone MK-7 was dominant in these novel strains as shown in other species of the genus Paenibacillus. The DNA-DNA hybridization dissociation value was <45 % with the closest related species. The novel strains had DNA G+C contents of 51.9 to 52.8 mol%. Phenotypically, the novel strains can be readily differentiated from closely related species by the absence of urease and gelatinase and the production of acids from a variety of sugars including l-arabinose. The major fatty acid was anteiso-C(15 : 0) as seen in P. barengoltzii and P. timonensis whereas the proportion of C(16 : 0) was significantly different from the closely related species. Based on phylogenetic and phenotypic results, it was concluded that these strains represent a novel species of the genus Paenibacillus, for which the name Paenibacillus phoenicis sp. nov. is proposed. The type strain is 3PO2SA(T) ( = NRRL B-59348(T)  = NBRC 106274(T)).

The secreted esterase of Propionibacterium freudenreichii has a major role in cheese lipolysis.

PubMed

Abeijón Mukdsi, María Claudia; Falentin, Hélène; Maillard, Marie-Bernadette; Chuat, Victoria; Medina, Roxana Beatriz; Parayre, Sandrine; Thierry, Anne

2014-01-01

Free fatty acids are important flavor compounds in cheese. Propionibacterium freudenreichii is the main agent of their release through lipolysis in Swiss cheese. Our aim was to identify the esterase(s) involved in lipolysis by P. freudenreichii. We targeted two previously identified esterases: one secreted esterase, PF#279, and one putative cell wall-anchored esterase, PF#774. To evaluate their role in lipolysis, we constructed overexpression and knockout mutants of P. freudenreichii CIRM-BIA1(T) for each corresponding gene. The sequences of both genes were also compared in 21 wild-type strains. All strains were assessed for their lipolytic activity on milk fat. The lipolytic activity observed matched data previously reported in cheese, thus validating the relevance of the method used. The mutants overexpressing PF#279 or PF#774 released four times more fatty acids than the wild-type strain, demonstrating that both enzymes are lipolytic esterases. However, inactivation of the pf279 gene induced a 75% reduction in the lipolytic activity compared to that of the wild-type strain, whereas inactivation of the pf774 gene did not modify the phenotype. Two of the 21 wild-type strains tested did not display any detectable lipolytic activity. Interestingly, these two strains exhibited the same single-nucleotide deletion at the beginning of the pf279 gene sequence, leading to a premature stop codon, whereas they harbored a pf774 gene highly similar to that of the other strains. Taken together, these results clearly demonstrate that PF#279 is the main lipolytic esterase in P. freudenreichii and a key agent of Swiss cheese lipolysis.

The Secreted Esterase of Propionibacterium freudenreichii Has a Major Role in Cheese Lipolysis

PubMed Central

Abeijón Mukdsi, María Claudia; Falentin, Hélène; Maillard, Marie-Bernadette; Chuat, Victoria; Medina, Roxana Beatriz; Parayre, Sandrine

2014-01-01

Free fatty acids are important flavor compounds in cheese. Propionibacterium freudenreichii is the main agent of their release through lipolysis in Swiss cheese. Our aim was to identify the esterase(s) involved in lipolysis by P. freudenreichii. We targeted two previously identified esterases: one secreted esterase, PF#279, and one putative cell wall-anchored esterase, PF#774. To evaluate their role in lipolysis, we constructed overexpression and knockout mutants of P. freudenreichii CIRM-BIA1T for each corresponding gene. The sequences of both genes were also compared in 21 wild-type strains. All strains were assessed for their lipolytic activity on milk fat. The lipolytic activity observed matched data previously reported in cheese, thus validating the relevance of the method used. The mutants overexpressing PF#279 or PF#774 released four times more fatty acids than the wild-type strain, demonstrating that both enzymes are lipolytic esterases. However, inactivation of the pf279 gene induced a 75% reduction in the lipolytic activity compared to that of the wild-type strain, whereas inactivation of the pf774 gene did not modify the phenotype. Two of the 21 wild-type strains tested did not display any detectable lipolytic activity. Interestingly, these two strains exhibited the same single-nucleotide deletion at the beginning of the pf279 gene sequence, leading to a premature stop codon, whereas they harbored a pf774 gene highly similar to that of the other strains. Taken together, these results clearly demonstrate that PF#279 is the main lipolytic esterase in P. freudenreichii and a key agent of Swiss cheese lipolysis. PMID:24242250

High depth, whole-genome sequencing of cholera isolates from Haiti and the Dominican Republic.

PubMed

Sealfon, Rachel; Gire, Stephen; Ellis, Crystal; Calderwood, Stephen; Qadri, Firdausi; Hensley, Lisa; Kellis, Manolis; Ryan, Edward T; LaRocque, Regina C; Harris, Jason B; Sabeti, Pardis C

2012-09-11

Whole-genome sequencing is an important tool for understanding microbial evolution and identifying the emergence of functionally important variants over the course of epidemics. In October 2010, a severe cholera epidemic began in Haiti, with additional cases identified in the neighboring Dominican Republic. We used whole-genome approaches to sequence four Vibrio cholerae isolates from Haiti and the Dominican Republic and three additional V. cholerae isolates to a high depth of coverage (>2000x); four of the seven isolates were previously sequenced. Using these sequence data, we examined the effect of depth of coverage and sequencing platform on genome assembly and identification of sequence variants. We found that 50x coverage is sufficient to construct a whole-genome assembly and to accurately call most variants from 100 base pair paired-end sequencing reads. Phylogenetic analysis between the newly sequenced and thirty-three previously sequenced V. cholerae isolates indicates that the Haitian and Dominican Republic isolates are closest to strains from South Asia. The Haitian and Dominican Republic isolates form a tight cluster, with only four variants unique to individual isolates. These variants are located in the CTX region, the SXT region, and the core genome. Of the 126 mutations identified that separate the Haiti-Dominican Republic cluster from the V. cholerae reference strain (N16961), 73 are non-synonymous changes, and a number of these changes cluster in specific genes and pathways. Sequence variant analyses of V. cholerae isolates, including multiple isolates from the Haitian outbreak, identify coverage-specific and technology-specific effects on variant detection, and provide insight into genomic change and functional evolution during an epidemic.

Geodermatophilus tzadiensis sp. nov., a UV radiation-resistant bacterium isolated from sand of the Saharan desert.

PubMed

Montero-Calasanz, Maria del Carmen; Göker, Markus; Broughton, William J; Cattaneo, Arlette; Favet, Jocelyne; Pötter, Gabriele; Rohde, Manfred; Spröer, Cathrin; Schumann, Peter; Klenk, Hans-Peter; Gorbushina, Anna A

2013-05-01

Three novel Gram-positive, aerobic, actinobacterial strains, CF5/2(T), CF5/1 and CF7/1, were isolated in 2007 during environmental screening of arid desert soil in the Sahara desert, Chad. Results from riboprinting, MALDI-TOF protein spectra and 16S rRNA sequence analysis confirmed that all three strains belonged to the same species. Phylogenetic analysis of 16S rRNA sequences with the strains' closest relatives indicated that they represented a distinct species. The three novel strains also shared a number of physiological and biochemical characteristics distinct from previously named Geodermatophilus species. The novel strains' peptidoglycan contained meso-diaminopimelic acid; their main phospholipids were phosphatidylcholine, phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylinositol and a small amount of phosphatidylglycerol; MK-9(H4) was the dominant menaquinone. The major cellular fatty acids were the branched-chain saturated acids iso-C16:0 and iso-C15:0. Galactose was detected as diagnostic sugar. Based on these chemotaxonomic results, 16S rRNA gene sequence analysis and DNA-DNA hybridization between strain CF5/2(T) and the type strains of Geodermatophilus saharensis, Geodermatophilus arenarius, Geodermatophilus nigrescens, Geodermatophilus telluris and Geodermatophilus siccatus, the isolates CF5/2(T), CF5/1 and CF7/1 are proposed to represent a novel species, Geodermatophilus tzadiensis, with type strain CF5/2(T)=DSM 45416=MTCC 11411 and two reference strains, CF5/1 (DSM 45415) and CF7/1 (DSM 45420). Copyright © 2013 Elsevier GmbH. All rights reserved.

Genomic Epidemiology of Hypervirulent Serogroup W, ST-11 Neisseria meningitidis

PubMed Central

Mustapha, Mustapha M.; Marsh, Jane W.; Krauland, Mary G.; Fernandez, Jorge O.; de Lemos, Ana Paula S.; Dunning Hotopp, Julie C.; Wang, Xin; Mayer, Leonard W.; Lawrence, Jeffrey G.; Hiller, N. Luisa; Harrison, Lee H.

2015-01-01

Neisseria meningitidis is a leading bacterial cause of sepsis and meningitis globally with dynamic strain distribution over time. Beginning with an epidemic among Hajj pilgrims in 2000, serogroup W (W) sequence type (ST) 11 emerged as a leading cause of epidemic meningitis in the African ‘meningitis belt’ and endemic cases in South America, Europe, Middle East and China. Previous genotyping studies were unable to reliably discriminate sporadic W ST-11 strains in circulation since 1970 from the Hajj outbreak strain (Hajj clone). It is also unclear what proportion of more recent W ST-11 disease clusters are caused by direct descendants of the Hajj clone. Whole genome sequences of 270 meningococcal strains isolated from patients with invasive meningococcal disease globally from 1970 to 2013 were compared using whole genome phylogenetic and major antigen-encoding gene sequence analyses. We found that all W ST-11 strains were descendants of an ancestral strain that had undergone unique capsular switching events. The Hajj clone and its descendants were distinct from other W ST-11 strains in that they shared a common antigen gene profile and had undergone recombination involving virulence genes encoding factor H binding protein, nitric oxide reductase, and nitrite reductase. These data demonstrate that recent acquisition of a distinct antigen-encoding gene profile and variations in meningococcal virulence genes was associated with the emergence of the Hajj clone. Importantly, W ST-11 strains unrelated to the Hajj outbreak contribute a significant proportion of W ST-11 cases globally. This study helps illuminate genomic factors associated with meningococcal strain emergence and evolution. PMID:26629539

Genotyping of Leptospira directly in urine samples of cattle demonstrates a diversity of species and strains in Brazil.

PubMed

Hamond, C; Pestana, C P; Medeiros, M A; Lilenbaum, W

2016-01-01

The aim of this study was to identify Leptospira in urine samples of cattle by direct sequencing of the secY gene. The validity of this approach was assessed using ten Leptospira strains obtained from cattle in Brazil and 77 DNA samples previously extracted from cattle urine, that were positive by PCR for the genus-specific lipL32 gene of Leptospira. Direct sequencing identified 24 (31·1%) interpretable secY sequences and these were identical to those obtained from direct DNA sequencing of the urine samples from which they were recovered. Phylogenetic analyses identified four species: L. interrogans, L. borgpetersenii, L. noguchii, and L. santarosai with the most prevalent genotypes being associated with L. borgpetersenii. While direct sequencing cannot, as yet, replace culturing of leptospires, it is a valid additional tool for epidemiological studies. An unexpected finding from this study was the genetic diversity of Leptospira infecting Brazilian cattle.

Sequence analysis of infectious pancreatic necrosis virus isolated from Iranian reared rainbow trout (Oncorhynchus mykiss) in 2012.

PubMed

Dadar, Maryam; Peyghan, Rahim; Memari, Hamid Rajabi; Shapouri, Masod Reza Seifi Abad; Hasanzadeh, Reza; Goudarzi, Laleh Moazzami; Vakharia, Vikram N

2013-12-01

Infectious pancreatic necrosis virus (IPNV) is the causal agent of a highly contagious disease that affects many species of fish and shellfish. This virus causes economically significant diseases of farmed rainbow trout, Oncorhynchus mykiss (Walbaum), in Iran, which is often associated with the transmission of pathogens from European resources. In this study, moribund rainbow trout fry samples were collected during an outbreak of IPNV in three different fish farms in north and west provinces of Iran in 2012; and we investigated the full genome sequence of Iranian IPNV and compared it with previously identified IPNV sequences. The sequences of different structural and nonstructural-protein genes were compared to those of other aquatic birnaviruses sequenced to date. Our results show that the Iranian isolate falls within genogroup 5, serotype A2 strain SP, having 99% identity with the strain 1146 from Spain. These results suggest that the Iranian isolate may have originated from Europe.

An Ecotype of Neorickettsia risticii Causing Potomac Horse Fever in Canada

PubMed Central

Xiong, Qingming; Bekebrede, Hannah; Sharma, Pratibha; Arroyo, Luis G.; Baird, John D.

2016-01-01

ABSTRACT Neorickettsia (formerly Ehrlichia) risticii is an obligatory intracellular bacterium of digenetic trematodes. When a horse accidentally ingests aquatic insects containing encysted trematodes infected with N. risticii, the bacterium is transmitted from trematodes to horse cells and causes an acute and often fatal disease called Potomac horse fever (PHF). Since the discovery of N. risticii in the United States in 1984, using immunofluorescence and PCR assays, PHF has been increasingly recognized throughout North America and South America. However, so far, there exist only a few stable N. risticii culture isolates, all of which are from horses within the United States, and the strain diversity and environmental spreading and distribution of pathogenic N. risticii strains remain poorly understood. This paper reports the isolation of N. risticii from the blood of a horse with acute PHF in Ontario, Canada. Intracellular N. risticii colonies were detected in P388D1 cells after 47 days of culturing and 8 days after the addition of rapamycin. Molecular phylogenetic analysis based on amino acid sequences of major surface proteins P51 and Ssa1 showed that this isolate is distinct from any previously sequenced strains but closely related to midwestern U.S. strains. This is the first Canadian strain cultured, and a new method was developed to reactivate dormant N. risticii to improve culture isolation. IMPORTANCE Neorickettsia risticii is an environmental bacterium that lives inside flukes that are parasitic to aquatic snails, insects, and bats. When a horse accidentally ingests insects harboring flukes infected with N. risticii, the bacterium is transmitted to the horse and causes an acute and often fatal disease called Potomac horse fever. Although the disease has been increasingly recognized throughout North and South America, N. risticii has not been cultured outside the United States. This paper reports the first Canadian strain cultured and a new method to effectively culture isolate N. risticii from the horse blood sample. Molecular analysis showed that the genotype of this Canadian strain is distinct from previously sequenced strains but closely related to midwestern U.S. strains. Culture isolation of N. risticii strains would confirm the geographic presence of pathogenic N. risticii, help elucidate N. risticii strain diversity and environmental spreading and distribution, and improve diagnosis and development of vaccines for this dreadful disease. PMID:27474720

An Ecotype of Neorickettsia risticii Causing Potomac Horse Fever in Canada.

PubMed

Xiong, Qingming; Bekebrede, Hannah; Sharma, Pratibha; Arroyo, Luis G; Baird, John D; Rikihisa, Yasuko

2016-10-01

Neorickettsia (formerly Ehrlichia) risticii is an obligatory intracellular bacterium of digenetic trematodes. When a horse accidentally ingests aquatic insects containing encysted trematodes infected with N. risticii, the bacterium is transmitted from trematodes to horse cells and causes an acute and often fatal disease called Potomac horse fever (PHF). Since the discovery of N. risticii in the United States in 1984, using immunofluorescence and PCR assays, PHF has been increasingly recognized throughout North America and South America. However, so far, there exist only a few stable N. risticii culture isolates, all of which are from horses within the United States, and the strain diversity and environmental spreading and distribution of pathogenic N. risticii strains remain poorly understood. This paper reports the isolation of N. risticii from the blood of a horse with acute PHF in Ontario, Canada. Intracellular N. risticii colonies were detected in P388D1 cells after 47 days of culturing and 8 days after the addition of rapamycin. Molecular phylogenetic analysis based on amino acid sequences of major surface proteins P51 and Ssa1 showed that this isolate is distinct from any previously sequenced strains but closely related to midwestern U.S. strains. This is the first Canadian strain cultured, and a new method was developed to reactivate dormant N. risticii to improve culture isolation. Neorickettsia risticii is an environmental bacterium that lives inside flukes that are parasitic to aquatic snails, insects, and bats. When a horse accidentally ingests insects harboring flukes infected with N. risticii, the bacterium is transmitted to the horse and causes an acute and often fatal disease called Potomac horse fever. Although the disease has been increasingly recognized throughout North and South America, N. risticii has not been cultured outside the United States. This paper reports the first Canadian strain cultured and a new method to effectively culture isolate N. risticii from the horse blood sample. Molecular analysis showed that the genotype of this Canadian strain is distinct from previously sequenced strains but closely related to midwestern U.S. strains. Culture isolation of N. risticii strains would confirm the geographic presence of pathogenic N. risticii, help elucidate N. risticii strain diversity and environmental spreading and distribution, and improve diagnosis and development of vaccines for this dreadful disease. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Identification and Classification of bcl Genes and Proteins of Bacillus cereus Group Organisms and Their Application in Bacillus anthracis Detection and Fingerprinting▿ †

PubMed Central

Leski, Tomasz A.; Caswell, Clayton C.; Pawlowski, Marcin; Klinke, David J.; Bujnicki, Janusz M.; Hart, Sean J.; Lukomski, Slawomir

2009-01-01

The Bacillus cereus group includes three closely related species, B. anthracis, B. cereus, and B. thuringiensis, which form a highly homogeneous subdivision of the genus Bacillus. One of these species, B. anthracis, has been identified as one of the most probable bacterial biowarfare agents. Here, we evaluate the sequence and length polymorphisms of the Bacillus collagen-like protein bcl genes as a basis for B. anthracis detection and fingerprinting. Five genes, designated bclA to bclE, are present in B. anthracis strains. Examination of bclABCDE sequences identified polymorphisms in bclB alleles of the B. cereus group organisms. These sequence polymorphisms allowed specific detection of B. anthracis strains by PCR using both genomic DNA and purified Bacillus spores in reactions. By exploiting the length variation of the bcl alleles it was demonstrated that the combined bclABCDE PCR products generate markedly different fingerprints for the B. anthracis Ames and Sterne strains. Moreover, we predict that bclABCDE length polymorphism creates unique signatures for B. anthracis strains, which facilitates identification of strains with specificity and confidence. Thus, we present a new diagnostic concept for B. anthracis detection and fingerprinting, which can be used alone or in combination with previously established typing platforms. PMID:19767469

Comparative Genomic Analyses of Clavibacter michiganensis subsp. insidiosus and Pathogenicity on Medicago truncatula.

PubMed

Lu, You; Ishimaru, Carol A; Glazebrook, Jane; Samac, Deborah A

2018-02-01

Clavibacter michiganensis is the most economically important gram-positive bacterial plant pathogen, with subspecies that cause serious diseases of maize, wheat, tomato, potato, and alfalfa. Much less is known about pathogenesis involving gram-positive plant pathogens than is known for gram-negative bacteria. Comparative genome analyses of C. michiganensis subspecies affecting tomato, potato, and maize have provided insights on pathogenicity. In this study, we identified strains of C. michiganensis subsp. insidiosus with contrasting pathogenicity on three accessions of the model legume Medicago truncatula. We generated complete genome sequences for two strains and compared these to a previously sequenced strain and genome sequences of four other subspecies. The three C. michiganensis subsp. insidiosus strains varied in gene content due to genome rearrangements, most likely facilitated by insertion elements, and plasmid number, which varied from one to three depending on strain. The core C. michiganensis genome consisted of 1,917 genes, with 379 genes unique to C. michiganensis subsp. insidiosus. An operon for synthesis of the extracellular blue pigment indigoidine, enzymes for pectin degradation, and an operon for inositol metabolism are among the unique features. Secreted serine proteases belonging to both the pat-1 and ppa families were present but highly diverged from those in other subspecies.

Comparative genomic characterization of citrus-associated Xylella fastidiosa strains.

PubMed

da Silva, Vivian S; Shida, Cláudio S; Rodrigues, Fabiana B; Ribeiro, Diógenes C D; de Souza, Alessandra A; Coletta-Filho, Helvécio D; Machado, Marcos A; Nunes, Luiz R; de Oliveira, Regina Costa

2007-12-21

The xylem-inhabiting bacterium Xylella fastidiosa (Xf) is the causal agent of Pierce's disease (PD) in vineyards and citrus variegated chlorosis (CVC) in orange trees. Both of these economically-devastating diseases are caused by distinct strains of this complex group of microorganisms, which has motivated researchers to conduct extensive genomic sequencing projects with Xf strains. This sequence information, along with other molecular tools, have been used to estimate the evolutionary history of the group and provide clues to understand the capacity of Xf to infect different hosts, causing a variety of symptoms. Nonetheless, although significant amounts of information have been generated from Xf strains, a large proportion of these efforts has concentrated on the study of North American strains, limiting our understanding about the genomic composition of South American strains - which is particularly important for CVC-associated strains. This paper describes the first genome-wide comparison among South American Xf strains, involving 6 distinct citrus-associated bacteria. Comparative analyses performed through a microarray-based approach allowed identification and characterization of large mobile genetic elements that seem to be exclusive to South American strains. Moreover, a large-scale sequencing effort, based on Suppressive Subtraction Hybridization (SSH), identified 290 new ORFs, distributed in 135 Groups of Orthologous Elements, throughout the genomes of these bacteria. Results from microarray-based comparisons provide further evidence concerning activity of horizontally transferred elements, reinforcing their importance as major mediators in the evolution of Xf. Moreover, the microarray-based genomic profiles showed similarity between Xf strains 9a5c and Fb7, which is unexpected, given the geographical and chronological differences associated with the isolation of these microorganisms. The newly identified ORFs, obtained by SSH, represent an approximately 10% increase in our current knowledge of the South American Xf gene pool and include new putative virulence factors, as well as novel potential markers for strain identification. Surprisingly, this list of novel elements include sequences previously believed to be unique to North American strains, pointing to the necessity of revising the list of specific markers that may be used for identification of distinct Xf strains.

Characterization of Aeromonas hydrophila wound pathotypes by comparative genomic and functional analyses of virulence genes.

PubMed

Grim, Christopher J; Kozlova, Elena V; Sha, Jian; Fitts, Eric C; van Lier, Christina J; Kirtley, Michelle L; Joseph, Sandeep J; Read, Timothy D; Burd, Eileen M; Tall, Ben D; Joseph, Sam W; Horneman, Amy J; Chopra, Ashok K; Shak, Joshua R

2013-04-23

Aeromonas hydrophila has increasingly been implicated as a virulent and antibiotic-resistant etiologic agent in various human diseases. In a previously published case report, we described a subject with a polymicrobial wound infection that included a persistent and aggressive strain of A. hydrophila (E1), as well as a more antibiotic-resistant strain of A. hydrophila (E2). To better understand the differences between pathogenic and environmental strains of A. hydrophila, we conducted comparative genomic and functional analyses of virulence-associated genes of these two wound isolates (E1 and E2), the environmental type strain A. hydrophila ATCC 7966(T), and four other isolates belonging to A. aquariorum, A. veronii, A. salmonicida, and A. caviae. Full-genome sequencing of strains E1 and E2 revealed extensive differences between the two and strain ATCC 7966(T). The more persistent wound infection strain, E1, harbored coding sequences for a cytotoxic enterotoxin (Act), a type 3 secretion system (T3SS), flagella, hemolysins, and a homolog of exotoxin A found in Pseudomonas aeruginosa. Corresponding phenotypic analyses with A. hydrophila ATCC 7966(T) and SSU as reference strains demonstrated the functionality of these virulence genes, with strain E1 displaying enhanced swimming and swarming motility, lateral flagella on electron microscopy, the presence of T3SS effector AexU, and enhanced lethality in a mouse model of Aeromonas infection. By combining sequence-based analysis and functional assays, we characterized an A. hydrophila pathotype, exemplified by strain E1, that exhibited increased virulence in a mouse model of infection, likely because of encapsulation, enhanced motility, toxin secretion, and cellular toxicity. Aeromonas hydrophila is a common aquatic bacterium that has increasingly been implicated in serious human infections. While many determinants of virulence have been identified in Aeromonas, rapid identification of pathogenic versus nonpathogenic strains remains a challenge for this genus, as it is for other opportunistic pathogens. This paper demonstrates, by using whole-genome sequencing of clinical Aeromonas strains, followed by corresponding virulence assays, that comparative genomics can be used to identify a virulent subtype of A. hydrophila that is aggressive during human infection and more lethal in a mouse model of infection. This aggressive pathotype contained genes for toxin production, toxin secretion, and bacterial motility that likely enabled its pathogenicity. Our results highlight the potential of whole-genome sequencing to transform microbial diagnostics; with further advances in rapid sequencing and annotation, genomic analysis will be able to provide timely information on the identities and virulence potential of clinically isolated microorganisms.

Gene sequence variability of the three surface proteins of human respiratory syncytial virus (HRSV) in Texas.

PubMed

Tapia, Lorena I; Shaw, Chad A; Aideyan, Letisha O; Jewell, Alan M; Dawson, Brian C; Haq, Taha R; Piedra, Pedro A

2014-01-01

Human respiratory syncytial virus (HRSV) has three surface glycoproteins: small hydrophobic (SH), attachment (G) and fusion (F), encoded by three consecutive genes (SH-G-F). A 270-nt fragment of the G gene is used to genotype HRSV isolates. This study genotyped and investigated the variability of the gene and amino acid sequences of the three surface proteins of HRSV strains collected from 1987 to 2005 from one center. Sixty original clinical isolates and 5 prototype strains were analyzed. Sequences containing SH, F and G genes were generated, and multiple alignments and phylogenetic trees were analyzed. Genetic variability by protein domains comparing virus genotypes was assessed. Complete sequences of the SH-G-F genes were obtained for all 65 samples: HRSV-A = 35; HRSV-B = 30. In group A strains, genotypes GA5 and GA2 were predominant. For HRSV-B strains, the genotype GB4 was predominant from 1992 to 1994 and only genotype BA viruses were detected in 2004-2005. Different genetic variability at nucleotide level was detected between the genes, with G gene being the most variable and the highest variability detected in the 270-nt G fragment that is frequently used to genotype the virus. High variability (>10%) was also detected in the signal peptide and transmembrane domains of the F gene of HRSV A strains. Variability among the HRSV strains resulting in non-synonymous changes was detected in hypervariable domains of G protein, the signal peptide of the F protein, a not previously defined domain in the F protein, and the antigenic site Ø in the pre-fusion F. Divergent trends were observed between HRSV -A and -B groups for some functional domains. A diverse population of HRSV -A and -B genotypes circulated in Houston during an 18 year period. We hypothesize that diverse sequence variation of the surface protein genes provide HRSV strains a survival advantage in a partially immune-protected community.

Gene Sequence Variability of the Three Surface Proteins of Human Respiratory Syncytial Virus (HRSV) in Texas

PubMed Central

Tapia, Lorena I.; Shaw, Chad A.; Aideyan, Letisha O.; Jewell, Alan M.; Dawson, Brian C.; Haq, Taha R.; Piedra, Pedro A.

2014-01-01

Human respiratory syncytial virus (HRSV) has three surface glycoproteins: small hydrophobic (SH), attachment (G) and fusion (F), encoded by three consecutive genes (SH-G-F). A 270-nt fragment of the G gene is used to genotype HRSV isolates. This study genotyped and investigated the variability of the gene and amino acid sequences of the three surface proteins of HRSV strains collected from 1987 to 2005 from one center. Sixty original clinical isolates and 5 prototype strains were analyzed. Sequences containing SH, F and G genes were generated, and multiple alignments and phylogenetic trees were analyzed. Genetic variability by protein domains comparing virus genotypes was assessed. Complete sequences of the SH-G-F genes were obtained for all 65 samples: HRSV-A = 35; HRSV-B = 30. In group A strains, genotypes GA5 and GA2 were predominant. For HRSV-B strains, the genotype GB4 was predominant from 1992 to 1994 and only genotype BA viruses were detected in 2004–2005. Different genetic variability at nucleotide level was detected between the genes, with G gene being the most variable and the highest variability detected in the 270-nt G fragment that is frequently used to genotype the virus. High variability (>10%) was also detected in the signal peptide and transmembrane domains of the F gene of HRSV A strains. Variability among the HRSV strains resulting in non-synonymous changes was detected in hypervariable domains of G protein, the signal peptide of the F protein, a not previously defined domain in the F protein, and the antigenic site Ø in the pre-fusion F. Divergent trends were observed between HRSV -A and -B groups for some functional domains. A diverse population of HRSV -A and -B genotypes circulated in Houston during an 18 year period. We hypothesize that diverse sequence variation of the surface protein genes provide HRSV strains a survival advantage in a partially immune-protected community. PMID:24625544

Classification of Lactococcus lactis cell envelope proteinase based on gene sequencing, peptides formed after hydrolysis of milk, and computer modeling.

PubMed

Børsting, M W; Qvist, K B; Brockmann, E; Vindeløv, J; Pedersen, T L; Vogensen, F K; Ardö, Y

2015-01-01

Lactococcus lactis strains depend on a proteolytic system for growth in milk to release essential AA from casein. The cleavage specificities of the cell envelope proteinase (CEP) can vary between strains and environments and whether the enzyme is released or bound to the cell wall. Thirty-eight Lc. lactis strains were grouped according to their CEP AA sequences and according to identified peptides after hydrolysis of milk. Finally, AA positions in the substrate binding region were suggested by the use of a new CEP template based on Streptococcus C5a CEP. Aligning the CEP AA sequences of 38 strains of Lc. lactis showed that 21 strains, which were previously classified as group d, could be subdivided into 3 groups. Independently, similar subgroupings were found based on comparison of the Lc. lactis CEP AA sequences and based on normalized quantity of identified peptides released from αS1-casein and β-casein. A model structure of Lc. lactis CEP based on the crystal structure of Streptococcus C5a CEP was used to investigate the AA positions in the substrate-binding region. New AA positions were suggested, which could be relevant for the cleavage specificity of CEP; however, these could only explain 2 out of 3 found subgroups. The third subgroup could be explained by 1 to 5 AA positions located opposite the substrate binding region. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Sequence-based typing of Legionella pneumophila strains isolated from hospital water distribution systems as a complementary element of risk assessment of legionellosis in Poland.

PubMed

Pancer, Katarzyna

2013-01-01

Many factors affect the risk of Legionella infection, such as the design, construction and maintenance of water distribution systems, the presence of individuals who may be exposed and their vulnerability to infection, and the degree of water system colonization and properties of Legionella strains. For epidemiological investigations, two properties of the Legionella strains are usually determined: serotyping and genotyping (sequence-based typing, SBT). In Poland, data regarding legionellosis are fragmentary, despite the fact that this has been a notifiable disease since 2002. The number of reported cases is very low; moreover, the main method of diagnosis is serological examination (delayed diagnosis and cheaper methods), and only single cases of LD were confirmed by culture of bacteria. Therefore, after 10 years of mandatory reporting of the Legionella spp. infection in Poland, the real epidemiological situation is still unknown; however, risk assessment should be carried out, especially in hospitals. In the presented study, comparison of the sequence types of 111 isolated L. pneumophila strains (from hospital water systems) with those present in the EWGLI SBT data was undertaken for complex risk analysis as a complementary element. In total, strains of L. pneumophila belonging to 12 out of 19 STs determined in the presented study were previously reported to the EWGLI SBT database (ST1, ST42, ST59, ST81, ST87, ST114, ST152, ST191, ST371, ST421, ST461, ST520). Among these strains, only 7 STs were previously reported in the amount of ≥10 (mainly ST1, ST42, ST81). Analysis of EWGLI data were carried out and, proportionally, the highest percentage of hospital-acquired strains (clinical and environmental) was found for ST 81, ST421 and ST152, but the largest number was for ST1. Based on the EWGLI data and the presented results, it was found that persistent colonization of HWS of 3 hospitals by strains belonging to ST42, ST1, ST87 indicated an increased risk of legionellosis, especially ST42.

Serological and genetic examination of some nontypical Streptococcus mutans strains.

PubMed

Coykendall, A L; Bratthall, D; O'Connor, K; Dvarskas, R A

1976-09-01

Thirty-four strains of Streptococcus mutans whose antigenic or genetic positions were unclear or unknown with respect to the serological scheme of Bratthall (1970) and Perch et al. (1974), or the genetic (deoxyribonucleic acid base sequence homology) scheme of Coykendall were analyzed to clarify their relationship to previously well-characterized strains. Strain OMZ175 of the "new" serotype f was genetically homologous with strains of S. mutans subsp. mutans. Strains of the "new" serotype g were homologous with serotype d strains (S. mutans subsp. sobrinus). Strains isolated from wild rats constituted a new genetic group but carried the c antigen. Thus, strains within a "genospecies" (subspecies) of S. mutans may not always carry a unique or characteristic antigen. We suggest that the existence of multiple serotypes within subspecies represents antigenic variation and adaptations to hosts.

A genome-wide study of two-component signal transduction systems in eight newly sequenced mutans streptococci strains

PubMed Central

2012-01-01

Background Mutans streptococci are a group of gram-positive bacteria including the primary cariogenic dental pathogen Streptococcus mutans and closely related species. Two component systems (TCSs) composed of a signal sensing histidine kinase (HK) and a response regulator (RR) play key roles in pathogenicity, but have not been comparatively studied for these oral bacterial pathogens. Results HKs and RRs of 8 newly sequenced mutans streptococci strains, including S. sobrinus DSM20742, S. ratti DSM20564 and six S. mutans strains, were identified and compared to the TCSs of S. mutans UA159 and NN2025, two previously genome sequenced S. mutans strains. Ortholog analysis revealed 18 TCS clusters (HK-RR pairs), 2 orphan HKs and 2 orphan RRs, of which 8 TCS clusters were common to all 10 strains, 6 were absent in one or more strains, and the other 4 were exclusive to individual strains. Further classification of the predicted HKs and RRs revealed interesting aspects of their putative functions. While TCS complements were comparable within the six S. mutans strains, S. sobrinus DSM20742 lacked TCSs possibly involved in acid tolerance and fructan catabolism, and S. ratti DSM20564 possessed 3 unique TCSs but lacked the quorum-sensing related TCS (ComDE). Selected computational predictions were verified by PCR experiments. Conclusions Differences in the TCS repertoires of mutans streptococci strains, especially those of S. sobrinus and S. ratti in comparison to S. mutans, imply differences in their response mechanisms for survival in the dynamic oral environment. This genomic level study of TCSs should help in understanding the pathogenicity of these mutans streptococci strains. PMID:22475007

Characterization of Aeromonas hydrophila Wound Pathotypes by Comparative Genomic and Functional Analyses of Virulence Genes

PubMed Central

Grim, Christopher J.; Kozlova, Elena V.; Sha, Jian; Fitts, Eric C.; van Lier, Christina J.; Kirtley, Michelle L.; Joseph, Sandeep J.; Read, Timothy D.; Burd, Eileen M.; Tall, Ben D.; Joseph, Sam W.; Horneman, Amy J.; Chopra, Ashok K.; Shak, Joshua R.

2013-01-01

ABSTRACT Aeromonas hydrophila has increasingly been implicated as a virulent and antibiotic-resistant etiologic agent in various human diseases. In a previously published case report, we described a subject with a polymicrobial wound infection that included a persistent and aggressive strain of A. hydrophila (E1), as well as a more antibiotic-resistant strain of A. hydrophila (E2). To better understand the differences between pathogenic and environmental strains of A. hydrophila, we conducted comparative genomic and functional analyses of virulence-associated genes of these two wound isolates (E1 and E2), the environmental type strain A. hydrophila ATCC 7966T, and four other isolates belonging to A. aquariorum, A. veronii, A. salmonicida, and A. caviae. Full-genome sequencing of strains E1 and E2 revealed extensive differences between the two and strain ATCC 7966T. The more persistent wound infection strain, E1, harbored coding sequences for a cytotoxic enterotoxin (Act), a type 3 secretion system (T3SS), flagella, hemolysins, and a homolog of exotoxin A found in Pseudomonas aeruginosa. Corresponding phenotypic analyses with A. hydrophila ATCC 7966T and SSU as reference strains demonstrated the functionality of these virulence genes, with strain E1 displaying enhanced swimming and swarming motility, lateral flagella on electron microscopy, the presence of T3SS effector AexU, and enhanced lethality in a mouse model of Aeromonas infection. By combining sequence-based analysis and functional assays, we characterized an A. hydrophila pathotype, exemplified by strain E1, that exhibited increased virulence in a mouse model of infection, likely because of encapsulation, enhanced motility, toxin secretion, and cellular toxicity. PMID:23611906

Phylogenetic analysis of feline immunodeficiency virus strains from naturally infected cats in Belgium and The Netherlands.

PubMed

Roukaerts, Inge D M; Theuns, Sebastiaan; Taffin, Elien R L; Daminet, Sylvie; Nauwynck, Hans J

2015-01-22

Feline immunodeficiency virus (FIV) is a major pathogen in feline populations worldwide, with seroprevalences up to 26%. Virus strains circulating in domestic cats are subdivided into different phylogenetic clades (A-E), based on the genetic diversity of the V3-V4 region of the env gene. In this report, a phylogenetic analysis of the V3-V4 env region, and a variable region in the gag gene was made for 36 FIV strains isolated in Belgium and The Netherlands. All newly generated gag sequences clustered together with previously known clade A FIV viruses, confirming the dominance of clade A viruses in Northern Europe. The same was true for the obtained env sequences, with only one sample of an unknown env subtype. Overall, the genetic diversity of FIV strains sequenced in this report was low. This indicates a relatively recent introduction of FIV in Belgium and The Netherlands. However, the sample with an unknown env subtype indicates that new introductions of FIV from unknown origin do occur and this will likely increase genetic variability in time. Copyright © 2014 Elsevier B.V. All rights reserved.

Comparative Genome Analysis Between Aspergillus oryzae Strains Reveals Close Relationship Between Sites of Mutation Localization and Regions of Highly Divergent Genes among Aspergillus Species

PubMed Central

Umemura, Myco; Koike, Hideaki; Yamane, Noriko; Koyama, Yoshinori; Satou, Yuki; Kikuzato, Ikuya; Teruya, Morimi; Tsukahara, Masatoshi; Imada, Yumi; Wachi, Youji; Miwa, Yukino; Yano, Shuichi; Tamano, Koichi; Kawarabayasi, Yutaka; Fujimori, Kazuhiro E.; Machida, Masayuki; Hirano, Takashi

2012-01-01

Aspergillus oryzae has been utilized for over 1000 years in Japan for the production of various traditional foods, and a large number of A. oryzae strains have been isolated and/or selected for the effective fermentation of food ingredients. Characteristics of genetic alterations among the strains used are of particular interest in studies of A. oryzae. Here, we have sequenced the whole genome of an industrial fungal isolate, A. oryzae RIB326, by using a next-generation sequencing system and compared the data with those of A. oryzae RIB40, a wild-type strain sequenced in 2005. The aim of this study was to evaluate the mutation pressure on the non-syntenic blocks (NSBs) of the genome, which were previously identified through comparative genomic analysis of A. oryzae, Aspergillus fumigatus, and Aspergillus nidulans. We found that genes within the NSBs of RIB326 accumulate mutations more frequently than those within the SBs, regardless of their distance from the telomeres or of their expression level. Our findings suggest that the high mutation frequency of NSBs might contribute to maintaining the diversity of the A. oryzae genome. PMID:22912434

Comparative genome analysis between Aspergillus oryzae strains reveals close relationship between sites of mutation localization and regions of highly divergent genes among Aspergillus species.

PubMed

Umemura, Myco; Koike, Hideaki; Yamane, Noriko; Koyama, Yoshinori; Satou, Yuki; Kikuzato, Ikuya; Teruya, Morimi; Tsukahara, Masatoshi; Imada, Yumi; Wachi, Youji; Miwa, Yukino; Yano, Shuichi; Tamano, Koichi; Kawarabayasi, Yutaka; Fujimori, Kazuhiro E; Machida, Masayuki; Hirano, Takashi

2012-10-01

Aspergillus oryzae has been utilized for over 1000 years in Japan for the production of various traditional foods, and a large number of A. oryzae strains have been isolated and/or selected for the effective fermentation of food ingredients. Characteristics of genetic alterations among the strains used are of particular interest in studies of A. oryzae. Here, we have sequenced the whole genome of an industrial fungal isolate, A. oryzae RIB326, by using a next-generation sequencing system and compared the data with those of A. oryzae RIB40, a wild-type strain sequenced in 2005. The aim of this study was to evaluate the mutation pressure on the non-syntenic blocks (NSBs) of the genome, which were previously identified through comparative genomic analysis of A. oryzae, Aspergillus fumigatus, and Aspergillus nidulans. We found that genes within the NSBs of RIB326 accumulate mutations more frequently than those within the SBs, regardless of their distance from the telomeres or of their expression level. Our findings suggest that the high mutation frequency of NSBs might contribute to maintaining the diversity of the A. oryzae genome.

Genomic Analysis of the Chicken Infectious Anemia Virus in a Specific Pathogen-Free Chicken Population in China

PubMed Central

Li, Yang; Wang, Yixin; Fang, Lichun; Fu, Jiayuan; Cui, Shuai; Zhao, Yingjie; Cui, Zhizhong; Chang, Shuang; Zhao, Peng

2016-01-01

The antibody to chicken infectious anemia virus (CIAV) was positive in a specific pathogen-free (SPF) chicken population by ELISA test in our previous inspection, indicating a possible infection with CIAV. In this study, blood samples collected from the SPF chickens were used to isolate CIAV by inoculating into MSB1 cells and PCR amplification. A CIAV strain (SD1403) was isolated and successfully identified. Three overlapping genomic fragments were obtained by PCR amplification and sequencing. The full genome sequence of the SD1403 strain was obtained by aligning the sequences. The genome of the SD1403 strain was 2293 bp with a nucleotide identity of 94.8% to 98.5% when compared with 30 referred CIAV strains. The viral proteins VP2 and VP3 were highly conserved, but VP1 was not relatively conserved. Both amino acids 139 and 144 of VP1 were glutamine, which was in accord with the low pathogenic characteristics. In this study, we first reported that CIAV exists in Chinese SPF chicken populations and may be an important reason why attenuated vaccine can be contaminated with CIAV. PMID:27298822

Genomic Analysis of the Chicken Infectious Anemia Virus in a Specific Pathogen-Free Chicken Population in China.

PubMed

Li, Yang; Wang, Yixin; Fang, Lichun; Fu, Jiayuan; Cui, Shuai; Zhao, Yingjie; Cui, Zhizhong; Chang, Shuang; Zhao, Peng

2016-01-01

The antibody to chicken infectious anemia virus (CIAV) was positive in a specific pathogen-free (SPF) chicken population by ELISA test in our previous inspection, indicating a possible infection with CIAV. In this study, blood samples collected from the SPF chickens were used to isolate CIAV by inoculating into MSB1 cells and PCR amplification. A CIAV strain (SD1403) was isolated and successfully identified. Three overlapping genomic fragments were obtained by PCR amplification and sequencing. The full genome sequence of the SD1403 strain was obtained by aligning the sequences. The genome of the SD1403 strain was 2293 bp with a nucleotide identity of 94.8% to 98.5% when compared with 30 referred CIAV strains. The viral proteins VP2 and VP3 were highly conserved, but VP1 was not relatively conserved. Both amino acids 139 and 144 of VP1 were glutamine, which was in accord with the low pathogenic characteristics. In this study, we first reported that CIAV exists in Chinese SPF chicken populations and may be an important reason why attenuated vaccine can be contaminated with CIAV.

Full-Genome Sequencing as a Basis for Molecular Epidemiology Studies of Bluetongue Virus in India

PubMed Central

Maan, Sushila; Maan, Narender S.; Belaganahalli, Manjunatha N.; Rao, Pavuluri Panduranga; Singh, Karam Pal; Hemadri, Divakar; Putty, Kalyani; Kumar, Aman; Batra, Kanisht; Krishnajyothi, Yadlapati; Chandel, Bharat S.; Reddy, G. Hanmanth; Nomikou, Kyriaki; Reddy, Yella Narasimha; Attoui, Houssam; Hegde, Nagendra R.; Mertens, Peter P. C.

2015-01-01

Since 1998 there have been significant changes in the global distribution of bluetongue virus (BTV). Ten previously exotic BTV serotypes have been detected in Europe, causing severe disease outbreaks in naïve ruminant populations. Previously exotic BTV serotypes were also identified in the USA, Israel, Australia and India. BTV is transmitted by biting midges (Culicoides spp.) and changes in the distribution of vector species, climate change, increased international travel and trade are thought to have contributed to these events. Thirteen BTV serotypes have been isolated in India since first reports of the disease in the country during 1964. Efficient methods for preparation of viral dsRNA and cDNA synthesis, have facilitated full-genome sequencing of BTV strains from the region. These studies introduce a new approach for BTV characterization, based on full-genome sequencing and phylogenetic analyses, facilitating the identification of BTV serotype, topotype and reassortant strains. Phylogenetic analyses show that most of the equivalent genome-segments of Indian BTV strains are closely related, clustering within a major eastern BTV ‘topotype’. However, genome-segment 5 (Seg-5) encoding NS1, from multiple post 1982 Indian isolates, originated from a western BTV topotype. All ten genome-segments of BTV-2 isolates (IND2003/01, IND2003/02 and IND2003/03) are closely related (>99% identity) to a South African BTV-2 vaccine-strain (western topotype). Similarly BTV-10 isolates (IND2003/06; IND2005/04) show >99% identity in all genome segments, to the prototype BTV-10 (CA-8) strain from the USA. These data suggest repeated introductions of western BTV field and/or vaccine-strains into India, potentially linked to animal or vector-insect movements, or unauthorised use of ‘live’ South African or American BTV-vaccines in the country. The data presented will help improve nucleic acid based diagnostics for Indian serotypes/topotypes, as part of control strategies. PMID:26121128

Haemophilus ducreyi Cutaneous Ulcer Strains Diverged from Both Class I and Class II Genital Ulcer Strains: Implications for Epidemiological Studies

PubMed Central

Gangaiah, Dharanesh

2016-01-01

Background Haemophilus ducreyi has emerged as a major cause of cutaneous ulcers (CU) in yaws-endemic regions of the tropics in the South Pacific, South East Asia and Africa. H. ducreyi was once thought only to cause the genital ulcer (GU) disease chancroid; GU strains belong to 2 distinct classes, class I and class II. Using whole-genome sequencing of 4 CU strains from Samoa, 1 from Vanuatu and 1 from Papua New Guinea, we showed that CU strains diverged from the class I strain 35000HP and that one CU strain expressed β-lactamase. Recently, the Center for Disease Control and Prevention released the genomes of 11 additional CU strains from Vanuatu and Ghana; however, the evolutionary relationship of these CU strains to previously-characterized CU and GU strains is unknown. Methodology/Principal Findings We performed phylogenetic analysis of 17 CU and 10 GU strains. Class I and class II GU strains formed two distinct clades. The class I strains formed two subclades, one containing 35000HP and HD183 and the other containing the remainder of the class I strains. Twelve of the CU strains formed a subclone under the class I 35000HP subclade, while 2 CU strains formed a subclone under the other class I subclade. Unexpectedly, 3 of the CU strains formed a subclone under the class II clade. Phylogenetic analysis of dsrA-hgbA-ncaA sequences yielded a tree similar to that of whole-genome phylogenetic tree. Conclusions/Significance CU strains diverged from multiple lineages within both class I and class II GU strains. Multilocus sequence typing of dsrA-hgbA-ncaA could be reliably used for epidemiological investigation of CU and GU strains. As class II strains grow relatively poorly and are relatively more susceptible to vancomycin than class I strains, these findings have implications for methods to recover CU strains. Comparison of contemporary CU and GU isolates would help clarify the relationship between these entities. PMID:28027326

Surveillance of human influenza A(H3N2) virus from 1999 to 2009 in southern Italy.

PubMed

DE Donno, A; Idolo, A; Quattrocchi, M; Zizza, A; Gabutti, G; Romano, A; Grima, P; Donatelli, I; Guido, M

2014-05-01

The aim of this study was to evaluate the presence of influenza virus co-infections in humans and changes in the genetic variability of A(H3N2) virus strains in southern Italy from 1999 to 2009. A partial sequence of the haemagglutinin (HA) gene by human influenza H3N2 strains identified in oropharyngeal swabs from patients with influenza-like illness was analysed by DNA sequencing and a phylogenetic analysis was performed. During the seasons 1999-2000, 2002-2003, 2004-2005 and 2008-2009, the influenza viruses circulating belonged to subtype H3N2. However, A(H1N1) subtype virus and B type were respectively prevalent during the 2000-2001, 2006-2007, 2007-2008 and 2001-2002, 2003-2004, 2005-2006 seasons. The HA sequences appeared to be closely related to the sequence of the influenza A vaccine strain. Only the 2002-2003 season was characterized by co-circulation of two viral lineages: A/New York/55/01(H3N2)-like virus of the previous season and A/Fujian/411/02(H3N2)-like virus, a new H3 variant. In this study, over the decade analysed, no significant change was seen in the sequences of the HA gene of H3 viruses isolated.

The RNase P RNA from cyanobacteria: short tandemly repeated repetitive (STRR) sequences are present within the RNase P RNA gene in heterocyst-forming cyanobacteria.

PubMed Central

Vioque, A

1997-01-01

The RNase P RNA gene (rnpB) from 10 cyanobacteria has been characterized. These new RNAs, together with the previously available ones, provide a comprehensive data set of RNase P RNA from diverse cyanobacterial lineages. All heterocystous cyanobacteria, but none of the non-heterocystous strains analyzed, contain short tandemly repeated repetitive (STRR) sequences that increase the length of helix P12. Site-directed mutagenesis experiments indicate that the STRR sequences are not required for catalytic activity in vitro. STRR sequences seem to have recently and independently invaded the RNase P RNA genes in heterocyst-forming cyanobacteria because closely related strains contain unrelated STRR sequences. Most cyanobacteria RNase P RNAs lack the sequence GGU in the loop connecting helices P15 and P16 that has been established to interact with the 3'-end CCA in precursor tRNA substrates in other bacteria. This character is shared with plastid RNase P RNA. Helix P6 is longer than usual in most cyanobacteria as well as in plastid RNase P RNA. PMID:9254706

Beta-hemolytic Streptococcus dysgalactiae strains isolated from horses are a genetically distinct population within the Streptococcus dysgalactiae taxon.

PubMed

Pinho, Marcos D; Erol, Erdal; Ribeiro-Gonçalves, Bruno; Mendes, Catarina I; Carriço, João A; Matos, Sandra C; Preziuso, Silvia; Luebke-Becker, Antina; Wieler, Lothar H; Melo-Cristino, Jose; Ramirez, Mario

2016-08-17

The pathogenic role of beta-hemolytic Streptococcus dysgalactiae in the equine host is increasingly recognized. A collection of 108 Lancefield group C (n = 96) or L (n = 12) horse isolates recovered in the United States and in three European countries presented multilocus sequence typing (MLST) alleles, sequence types and emm types (only 56% of the isolates could be emm typed) that were, with few exceptions, distinct from those previously found in human Streptococcus dysgalactiae subsp. equisimilis. Characterization of a subset of horse isolates by multilocus sequence analysis (MLSA) and 16S rRNA gene sequence showed that most equine isolates could also be differentiated from S. dysgalactiae strains from other animal species, supporting the existence of a horse specific genomovar. Draft genome information confirms the distinctiveness of the horse genomovar and indicates the presence of potentially horse-specific virulence factors. While this genomovar represents most of the isolates recovered from horses, a smaller MLST and MLSA defined sub-population seems to be able to cause infections in horses, other animals and humans, indicating that transmission between hosts of strains belonging to this group may occur.

The Applied Development of a Tiered Multilocus Sequence Typing (MLST) Scheme for Dichelobacter nodosus.

PubMed

Blanchard, Adam M; Jolley, Keith A; Maiden, Martin C J; Coffey, Tracey J; Maboni, Grazieli; Staley, Ceri E; Bollard, Nicola J; Warry, Andrew; Emes, Richard D; Davies, Peers L; Tötemeyer, Sabine

2018-01-01

Dichelobacter nodosus ( D. nodosus ) is the causative pathogen of ovine footrot, a disease that has a significant welfare and financial impact on the global sheep industry. Previous studies into the phylogenetics of D. nodosus have focused on Australia and Scandinavia, meaning the current diversity in the United Kingdom (U.K.) population and its relationship globally, is poorly understood. Numerous epidemiological methods are available for bacterial typing; however, few account for whole genome diversity or provide the opportunity for future application of new computational techniques. Multilocus sequence typing (MLST) measures nucleotide variations within several loci with slow accumulation of variation to enable the designation of allele numbers to determine a sequence type. The usage of whole genome sequence data enables the application of MLST, but also core and whole genome MLST for higher levels of strain discrimination with a negligible increase in experimental cost. An MLST database was developed alongside a seven loci scheme using publically available whole genome data from the sequence read archive. Sequence type designation and strain discrimination was compared to previously published data to ensure reproducibility. Multiple D. nodosus isolates from U.K. farms were directly compared to populations from other countries. The U.K. isolates define new clades within the global population of D. nodosus and predominantly consist of serogroups A, B and H, however serogroups C, D, E, and I were also found. The scheme is publically available at https://pubmlst.org/dnodosus/.

Draft Genome Sequence of Roseovarius sp. A-2, an Iodide-Oxidizing Bacterium Isolated from Natural Gas Brine Water, Chiba, Japan.

PubMed

Yuliana, Tri; Nakajima, Nobuyoshi; Yamamura, Shigeki; Tomita, Masaru; Suzuki, Haruo; Amachi, Seigo

2017-01-01

Roseovarius sp. A-2 is a heterotrophic iodide (I - )-oxidizing bacterium isolated from iodide-rich natural gas brine water in Chiba, Japan. This strain oxidizes iodide to molecular iodine (I 2 ) by means of an extracellular multicopper oxidase. Here we report the draft genome sequence of strain A-2. The draft genome contained 46 tRNA genes, 1 copy of a 16S-23S-5S rRNA operon, and 4,514 protein coding DNA sequences, of which 1,207 (27%) were hypothetical proteins. The genome contained a gene encoding IoxA, a multicopper oxidase previously found to catalyze the oxidation of iodide in Iodidimonas sp. Q-1. This draft genome provides detailed insights into the metabolism and potential application of Roseovarius sp. A-2.

Draft Genome Sequence of a Tetrabromobisphenol A–Degrading Strain, Ochrobactrum sp. T, Isolated from an Electronic Waste Recycling Site

PubMed Central

Liang, Zhishu; Li, Guiying; Zhang, Guoxia; Das, Ranjit

2016-01-01

Ochrobactrum sp. T was previously isolated from a sludge sample collected from an electronic waste recycling site and characterized as a unique tetrabromobisphenol A (TBBPA)–degrading bacterium. Here, the draft genome sequence (3.9 Mb) of Ochrobactrum sp. T is reported to provide insights into its diversity and its TBBPA biodegradation mechanism in polluted environments. PMID:27445374

Genomic Characterization of Travel-Associated Dengue Viruses Isolated from the Entry-Exit Ports in Fujian Province, China, 2013-2015.

PubMed

Gao, Bo; Zhang, Jianming; Wang, Yuping; Chen, Fan; Zheng, Chaohui; Xie, Lianhui

2017-09-25

Over the past decade, indigenous dengue outbreaks have occurred occasionally in Fujian province in southeastern China because of sporadic imported dengue viruses (DENV). In this study, 3 DENV-2 and 2 DENV-4 strains were isolated from suspected febrile travelers at 2 ports of entry in Fujian between 2013-2015. Complete viral genome sequences of these new isolates were obtained with Sanger chemistry. Genomic sequence analyses revealed that these strains belonged to genotypes of 2-Cosmopolitan and 4-II. Consistent with the patients' travel information, phylogenetic analyses of the complete coding regions also indicated that most of the new isolates were genetically similar to the circulating strains in Southeast Asia rather than previous Chinese strains that were available. Therefore, phylogenetic analyses of the imported DENV demonstrated that multiple introductions of DENV emerged continuously in Fujian, and highlighted the importance of dengue surveillance at entry-exit ports in the subtropical regions of southern China.

Yeasts associated with the curculionid beetle Xyloterinus politus: Candida xyloterini sp. nov., Candida palmyrensis sp. nov. and three common ambrosia yeasts.

PubMed

Suh, Sung-Oui; Zhou, Jianlong

2010-07-01

Seven yeast strains were isolated from the body surface and galleries of Xyloterinus politus, the ambrosia beetle that attacks black oak trees. Based on rDNA sequence comparisons and other taxonomic characteristics, five of the strains were identified as members of the species Saccharomycopsis microspora, Wickerhamomyces hampshirensis and Candida mycetangii, which have been reported previously as being associated with insects. The remaining two yeast strains were proposed as representatives of two novel species, Candida xyloterini sp. nov. (type strain ATCC 62898(T)=CBS 11547(T)) and Candida palmyrensis sp. nov. (type strain ATCC 62899(T)=CBS 11546(T)). C. xyloterini sp. nov. is a close sister taxon to Ogataea dorogensis and assimilates methanol as a sole carbon source but lacks ascospores. On the other hand, C. palmyrensis sp. nov. is phylogenetically distinct from any other ambrosia yeast reported so far. The species was placed near Candida sophiae-reginae and Candida beechii based on DNA sequence analyses, but neither of these were close sister taxa to C. palmyrensis sp. nov.

Abr1, a Transposon-Like Element in the Genome of the Cultivated Mushroom Agaricus bisporus (Lange) Imbach

PubMed Central

Sonnenberg, Anton S. M.; Baars, Johan J. P.; Mikosch, Thomas S. P.; Schaap, Peter J.; Van Griensven, Leo J. L. D.

1999-01-01

A 300-bp repetitive element was found in the genome of the white button mushroom, Agaricus bisporus, and designated Abr1. It is present in ∼15 copies per haploid genome in the commercial strain Horst U1. Analysis of seven copies showed 89 to 97% sequence identity. The repeat has features typical of class II transposons (i.e., terminal inverted repeats, subterminal repeats, and a target site duplication of 7 bp). The latter shows a consensus sequence. When used as probe on Southern blots, Abr1 identifies relatively little variation within traditional and present-day commercial strains, indicating that most strains are identical or have a common origin. In contrast to these cultivars, high variation is found among field-collected strains. Furthermore, a remarkable difference in copy numbers of Abr1 was found between A. bisporus isolates with a secondarily homothallic life cycle and those with a heterothallic life cycle. Abr1 is a type II transposon not previously reported in basidiomycetes and appears to be useful for the identification of strains within the species A. bisporus. PMID:10427018

Whole genome assembly of a natto production strain Bacillus subtilis natto from very short read data.

PubMed

Nishito, Yukari; Osana, Yasunori; Hachiya, Tsuyoshi; Popendorf, Kris; Toyoda, Atsushi; Fujiyama, Asao; Itaya, Mitsuhiro; Sakakibara, Yasubumi

2010-04-16

Bacillus subtilis natto is closely related to the laboratory standard strain B. subtilis Marburg 168, and functions as a starter for the production of the traditional Japanese food "natto" made from soybeans. Although re-sequencing whole genomes of several laboratory domesticated B. subtilis 168 derivatives has already been attempted using short read sequencing data, the assembly of the whole genome sequence of a closely related strain, B. subtilis natto, from very short read data is more challenging, particularly with our aim to assemble one fully connected scaffold from short reads around 35 bp in length. We applied a comparative genome assembly method, which combines de novo assembly and reference guided assembly, to one of the B. subtilis natto strains. We successfully assembled 28 scaffolds and managed to avoid substantial fragmentation. Completion of the assembly through long PCR experiments resulted in one connected scaffold for B. subtilis natto. Based on the assembled genome sequence, our orthologous gene analysis between natto BEST195 and Marburg 168 revealed that 82.4% of 4375 predicted genes in BEST195 are one-to-one orthologous to genes in 168, with two genes in-paralog, 3.2% are deleted in 168, 14.3% are inserted in BEST195, and 5.9% of genes present in 168 are deleted in BEST195. The natto genome contains the same alleles in the promoter region of degQ and the coding region of swrAA as the wild strain, RO-FF-1. These are specific for gamma-PGA production ability, which is related to natto production. Further, the B. subtilis natto strain completely lacked a polyketide synthesis operon, disrupted the plipastatin production operon, and possesses previously unidentified transposases. The determination of the whole genome sequence of Bacillus subtilis natto provided detailed analyses of a set of genes related to natto production, demonstrating the number and locations of insertion sequences that B. subtilis natto harbors but B. subtilis 168 lacks. Multiple genome-level comparisons among five closely related Bacillus species were also carried out. The determined genome sequence of B. subtilis natto and gene annotations are available from the Natto genome browser http://natto-genome.org/.

Characterization of the Porphyromonas gingivalis conjugative transposon CTnPg1: determination of the integration site and the genes essential for conjugal transfer.

PubMed

Naito, Mariko; Sato, Keiko; Shoji, Mikio; Yukitake, Hideharu; Ogura, Yoshitoshi; Hayashi, Tetsuya; Nakayama, Koji

2011-07-01

In our previous study, extensive genomic rearrangements were found in two strains of the Gram-negative anaerobic bacterium Porphyromonas (Por.) gingivalis, and most of these rearrangements were associated with mobile genetic elements such as insertion sequences and conjugative transposons (CTns). CTnPg1, identified in Por. gingivalis strain ATCC 33277, was the first complete CTn reported for the genus Porphyromonas. In the present study, we found that CTnPg1 can be transferred from strain ATCC 33277 to another Por. gingivalis strain, W83, at a frequency of 10(-7) to 10(-6). The excision of CTnPg1 from the chromosome in a donor cell depends on an integrase (Int; PGN_0094) encoded in CTnPg1, whereas CTnPg1 excision is independent of PGN_0084 (a DNA topoisomerase I homologue; Exc) encoded within CTnPg1 and recA (PGN_1057) on the donor chromosome. Intriguingly, however, the transfer of CTnPg1 between Por. gingivalis strains requires RecA function in the recipient. Sequencing analysis of CTnPg1-integrated sites on the chromosomes of transconjugants revealed that the consensus attachment (att) sequence is a 13 bp sequence, TTTTCNNNNAAAA. We further report that CTnPg1 is able to transfer to two other bacterial species, Bacteroides thetaiotaomicron and Prevotella oralis. In addition, CTnPg1-like CTns are located in the genomes of other oral anaerobic bacteria, Porphyromonas endodontalis, Prevotella buccae and Prevotella intermedia, with the same consensus att sequence. These results suggest that CTns in the CTnPg1 family are widely distributed among oral anaerobic Gram-negative bacteria found in humans and play important roles in horizontal gene transfer among these bacteria.

Remarkably Divergent Regions Punctuate the Genome Assembly of the Caenorhabditis elegans Hawaiian Strain CB4856

PubMed Central

Thompson, Owen A.; Snoek, L. Basten; Nijveen, Harm; Sterken, Mark G.; Volkers, Rita J. M.; Brenchley, Rachel; van’t Hof, Arjen; Bevers, Roel P. J.; Cossins, Andrew R.; Yanai, Itai; Hajnal, Alex; Schmid, Tobias; Perkins, Jaryn D.; Spencer, David; Kruglyak, Leonid; Andersen, Erik C.; Moerman, Donald G.; Hillier, LaDeana W.; Kammenga, Jan E.; Waterston, Robert H.

2015-01-01

The Hawaiian strain (CB4856) of Caenorhabditis elegans is one of the most divergent from the canonical laboratory strain N2 and has been widely used in developmental, population, and evolutionary studies. To enhance the utility of the strain, we have generated a draft sequence of the CB4856 genome, exploiting a variety of resources and strategies. When compared against the N2 reference, the CB4856 genome has 327,050 single nucleotide variants (SNVs) and 79,529 insertion–deletion events that result in a total of 3.3 Mb of N2 sequence missing from CB4856 and 1.4 Mb of sequence present in CB4856 but not present in N2. As previously reported, the density of SNVs varies along the chromosomes, with the arms of chromosomes showing greater average variation than the centers. In addition, we find 61 regions totaling 2.8 Mb, distributed across all six chromosomes, which have a greatly elevated SNV density, ranging from 2 to 16% SNVs. A survey of other wild isolates show that the two alternative haplotypes for each region are widely distributed, suggesting they have been maintained by balancing selection over long evolutionary times. These divergent regions contain an abundance of genes from large rapidly evolving families encoding F-box, MATH, BATH, seven-transmembrane G-coupled receptors, and nuclear hormone receptors, suggesting that they provide selective advantages in natural environments. The draft sequence makes available a comprehensive catalog of sequence differences between the CB4856 and N2 strains that will facilitate the molecular dissection of their phenotypic differences. Our work also emphasizes the importance of going beyond simple alignment of reads to a reference genome when assessing differences between genomes. PMID:25995208

Characterization of HIV Type 1 Envelope Sequence Among Viral Isolates Circulating in the Northern Region of Colombia, South America

PubMed Central

Villarreal, José-Luis; Gutiérrez, Jaime; Palacio, Lucy; Peñuela, Martha; Hernández, Robin; Lemay, Guy

2012-01-01

Abstract To characterize human immunodeficiency virus (HIV-1) strains circulating in the Northern region of Colombia in South America, sequences of the viral envelope C2V3C3 region were obtained from patients with different high-risk practices. Close to 60% of the sequences were predicted to belong to macrophage-tropic viruses, according to the positions of acidic amino acids and putative N-linked glycosylation sites. This is in agreement with the fact that most of the patients were recently diagnosed individuals. Phylogenic analysis then allowed assignment of all 35 samples to subtype B viruses. This same subtype was found in previous studies carried out in other Colombian regions. This study thus expands previous analyses with previously missing data from the Northern region of the country. The number and the length of the sequences examined also help to provide a clearer picture of the prevailing situation of the present HIV epidemics in this country. PMID:22482735

Members of Microvirga and Bradyrhizobium genera are native endosymbiotic bacteria nodulating Lupinus luteus in Northern Tunisian soils.

PubMed

Msaddak, Abdelhakim; Rejili, Mokhtar; Durán, David; Rey, Luis; Imperial, Juan; Palacios, Jose Manuel; Ruiz-Argüeso, Tomas; Mars, Mohamed

2017-06-01

The genetic diversity of bacterial populations nodulating Lupinus luteus (yellow lupine) in Northern Tunisia was examined. Phylogenetic analyses of 43 isolates based on recA and gyrB partial sequences grouped them in three clusters, two of which belong to genus Bradyrhizobium (41 isolates) and one, remarkably, to Microvirga (2 isolates), a genus never previously described as microsymbiont of this lupine species. Representatives of the three clusters were analysed in-depth by multilocus sequence analysis of five housekeeping genes (rrs, recA, glnII, gyrB and dnaK). Surprisingly, the Bradyrhizobium cluster with the two isolates LluI4 and LluTb2 may constitute a new species defined by a separate position between Bradyrhizobium manausense and B. denitrificans. A nodC-based phylogeny identified only two groups: one formed by Bradyrhizobium strains included in the symbiovar genistearum and the other by the Microvirga strains. Symbiotic behaviour of representative isolates was tested, and among the seven legumes inoculated only a difference was observed i.e. the Bradyrhizobium strains nodulated Ornithopus compressus unlike the two strains of Microvirga. On the basis of these data, we conclude that L. luteus root nodule symbionts in Northern Tunisia are mostly strains within the B. canariense/B. lupini lineages, and the remaining strains belong to two groups not previously identified as L. luteus endosymbionts: one corresponding to a new clade of Bradyrhizobium and the other to the genus Microvirga. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Characterization of Streptomycin Resistance in Isolates of Erwinia amylovora in California.

PubMed

Förster, Helga; McGhee, Gayle C; Sundin, George W; Adaskaveg, James E

2015-10-01

In surveys from 2006 to 2014, streptomycin resistance in Erwinia amylovora from pear-growing areas in California declined from very high incidence in 2006 and 2007 to very low incidence in 2013 and 2014. The majority of resistant strains were designated as moderately resistant-low (MR-L), and were almost exclusively found in Sacramento County, whereas highly resistant (HR) strains were only recovered in Sutter-Yuba and San Joaquin counties. Resistance of HR strains was associated with a mutation in codon 43 of the chromosomal rpsL gene that results in a change from lysine to arginine, the same mutation that was originally reported for resistant strains from California in the mid-1970s. MR-L strains were found to harbor the strA-strB streptomycin resistance genes on transposon Tn5393a. This transposon lacks insertion sequence IS1133 that provides a promoter for efficient expression of strA-strB, resulting in lower minimum inhibitory concentrations of MR-L strains compared with those from other locations that harbor strA-strB on Tn5393::IS1133. In contrast to previously described plasmid-mediated resistance where Tn5393 is inserted in pEa34, or pEA29, Tn5393a in MR-L strains was located on plasmid pEU30. This plasmid was first described in E. amylovora from the western United States but was not associated with streptomycin resistance determinants previously. We hypothesize that Tn5393a was introduced into an E. amylovora strain carrying pEU30 and transposed into that plasmid. This hypothesis was supported by clustered regularly interspaced short palindromic repeat (CRISPR) sequence analysis that showed that two MR-L strains share the same CRISPR1 pattern as a streptomycin-sensitive strain. With current low resistance levels in California growing regions, streptomycin could be successfully used again, but applications per season should be limited and the antibiotic should be mixed and rotated with different modes of action.

Bradyrhizobium ottawaense sp. nov., a symbiotic nitrogen fixing bacterium from root nodules of soybeans in Canada

PubMed Central

Yu, Xiumei; Cloutier, Sylvie; Tambong, James T.

2014-01-01

Sixteen strains of symbiotic bacteria from root nodules of Glycine max grown in Ottawa, Canada, were previously characterized and placed in a novel group within the genus Bradyrhizobium. To verify their taxonomic status, these strains were further characterized using a polyphasic approach. All strains possessed identical 16S rRNA gene sequences that were 99.79 % similar to the closest relative, Bradyrhizobium liaoningense LMG 18230T. Phylogenetic analysis of concatenated atpD, glnII, recA, gyrB, rpoB and dnaK genes divided the 16 strains into three multilocus sequence types that were placed in a highly supported lineage distinct from named species of the genus Bradyrhizobium consistent with results of DNA–DNA hybridization. Based on analysis of symbiosis gene sequences (nodC and nifH), all novel strains were placed in a phylogenetic group with five species of the genus Bradyrhizobium that nodulate soybeans. The combination of phenotypic characteristics from several tests including carbon and nitrogen source utilization and antibiotic resistance could be used to differentiate representative strains from recognized species of the genus Bradyrhizobium. Novel strain OO99T elicits effective nodules on Glycine max, Glycine soja and Macroptilium atropurpureum, partially effective nodules on Desmodium canadense and Vigna unguiculata, and ineffective nodules on Amphicarpaea bracteata and Phaseolus vulgaris. Based on the data presented, we conclude that our strains represent a novel species for which the name Bradyrhizobium ottawaense sp. nov. is proposed, with OO99T ( = LMG 26739T = HAMBI 3284T) as the type strain. The DNA G+C content is 62.6 mol%. PMID:24969302

Development of a systematic feedback isolation approach for targeted strains from mixed culture systems.

PubMed

Poudel, Pramod; Tashiro, Yukihiro; Miyamoto, Hirokuni; Miyamoto, Hisashi; Okugawa, Yuki; Sakai, Kenji

2017-01-01

Elucidation of functions of bacteria in a mixed culture system (MCS) such as composting, activated sludge system is difficult, since the system is complicating with many unisolated bacteria. Here, we developed a systematic feedback isolation strategy for the isolation and rapid screening of multiple targeted strains from MCS. Six major strains (Corynebacterium sphenisci, Bacillus thermocloacae, Bacillus thermoamylovorans, Bacillus smithii, Bacillus humi, and Bacillus coagulans), which are detected by denaturing gradient gel electrophoresis (DGGE) analysis in our previous study on MCS for l-lactic acid production, were targeted for isolation. Based on information of suitable cultivation conditions (e.g., media, pH, temperature) from the literature, feedback isolation was performed to form 136 colonies. The following direct colony matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was optimised as the second screening to narrow down 20 candidate colonies from similar spectra patterns with six closest type strains. This step could distinguish bacteria at the species level with distance similarity scores ≥0.55 corresponding to 16S rRNA gene sequence similarity ≥98.2%, suggesting that this is an effective technique to minimize isolates close to targeted type strains. Analysis of 16S rRNA gene sequences indicated that two targeted strains and one strain related to the target had successfully been isolated, showing high similarities (99.5-100%) with the sequences from the DGGE bands, and that the other candidates were affiliated with three strains that were closely related to the target species. This study proposes a new method for systematic feedback isolation that may be useful for isolating targeted strains from MCS for further investigation. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Investigation of taxa of the family Pasteurellaceae isolated from Syrian and European hamsters and proposal of Mesocricetibacter intestinalis gen. nov., sp. nov. and Cricetibacter osteomyelitidis gen. nov., sp. nov.

PubMed

Christensen, H; Nicklas, W; Bisgaard, M

2014-11-01

Eleven strains from hamster of Bisgaard taxa 23 and 24, also referred to as Krause's groups 2 and 1, respectively, were investigated by a polyphasic approach including data published previously. Strains showed small, regular and circular colonies with smooth and shiny appearance, typical of members of the family Pasteurellaceae. The strains formed two monophyletic groups based on 16S rRNA gene sequence comparison to other members of the family Pasteurellaceae. Partial rpoB sequencing as well as published data on DNA-DNA hybridization showed high genotypic relationships within both groups. Menaquinone 7 (MK7) was found in strains of both groups as well as an unknown ubiquinone with shorter chain length than previously reported for any other member of the family Pasteurellaceae. A new genus with one species, Mesocricetibacter intestinalis gen. nov., sp. nov., is proposed to accommodate members of taxon 24 of Bisgaard whereas members of taxon 23 of Bisgaard are proposed to represent Cricetibacter osteomyelitidis gen. nov., sp. nov. Major fatty acids of type strains of type species of both genera are C(14:0), C(14:0) 3-OH/iso-C(16:1) I, C(16:1)ω7c and C(16:0). The two genera are clearly separated by phenotype from each other and from existing genera of the family Pasteurellaceae. The type strain of Mesocricetibacter intestinalis is HIM 933/7(T) ( =Kunstyr 246/85(T) =CCUG 28030(T) =DSM 28403(T)) while the type strain of Cricetibacter osteomyelitidis is HIM943/7(T) ( =Kunstyr 507/85(T) =CCUG 36451(T) =DSM 28404(T)). © 2014 IUMS.

First cases of European bat lyssavirus type 1 in Iberian serotine bats: Implications for the molecular epidemiology of bat rabies in Europe.

PubMed

Mingo-Casas, Patricia; Sandonís, Virginia; Obón, Elena; Berciano, José M; Vázquez-Morón, Sonia; Juste, Javier; Echevarría, Juan E

2018-04-01

Previous studies have shown that EBLV-1 strains exclusively hosted by Eptesicus isabellinus bats in the Iberian Peninsula cluster in a specific monophyletic group that is related to the EBLV-1b lineage found in the rest of Europe. More recently, enhanced passive surveillance has allowed the detection of the first EBLV-1 strains associated to Eptesicus serotinus south of the Pyrenees. The aim of this study is the reconstruction of the EBLV-1 phylogeny and phylodynamics in the Iberian Peninsula in the context of the European continent. We have sequenced 23 EBLV-1 strains detected on nine E. serotinus and 14 E. isabellinus. Phylogenetic analyses were performed on the first 400-bp-5' fragment of the Nucleoprotein (N) gene together with other 162 sequences from Europe. Besides, fragments of the variable region of the phosphoprotein (P) gene and the glycoprotein-polymerase (G-L) intergenic region were studied on Spanish samples. Phylogenies show that two of the new EBLV-1a strains from Iberian E. serotinus clustered together with French strains from the North of the Pyrenees, suggesting a recent expansion southwards of this subtype. The remaining seven Iberian strains from E. serotinus grouped, instead, within the cluster linked, so far, to E. isabellinus, indicating that spatial distribution prevails over species specificity in explaining rabies distribution and supporting interspecific transmission. The structure found within the Iberian Peninsula for EBLV-1b is in concordance with that described previously for E. isabellinus. Finally, we have found that the current EBLV-1 European strains could have emerged only 175 years ago according to our evolutionary dynamics analyses.

Mitochondrial gene sequences alone or combined with ITS region sequences provide firm molecular criteria for the classification of Lecanicillium species.

PubMed

Kouvelis, Vassili N; Sialakouma, Aphrodite; Typas, Milton A

2008-07-01

The recent revision of Verticillium sect. Prostrata led to the introduction of the genus Lecanicillium, which comprises the majority of the entomopathogenic strains. Sixty-five strains previously classified as Verticillium lecanii or Verticillium sp. from different geographical regions and hosts were examined and their phylogenetic relationships were determined using sequences from three mitochondrial (mt) genes [the small rRNA subunit (rns), the NADH dehydrogenase subunits 1 (nad1) and 3 (nad3)] and the ITS region. In general, single gene phylogenetic trees differentiated and placed the strains examined in well-supported (by BS analysis) groups of L. lecanii, L. longisporum, L. muscarium, and L. nodulosum, although in some cases a few uncertainties still remained. nad1 was the most informative single gene in phylogenetic analyses and was also found to contain group I introns with putative open reading frames (ORFs) encoding for GIY-YIG endonucleases. The combined use of mt gene sequences resolved taxonomic uncertainties arisen from ITS analysis and, alone or in combination with ITS sequences, helped in placing uncharacterised Verticillium lecanii and Verticillium sp. firmly into Lecanicillium species. Combined gene data from all the mt genes and all the mt genes and the ITS region together, were very similar. Furthermore, a relaxed correlation with host specificity -- at least for Homoptera -- was indicated for the rns and the combined mt gene sequences. Thus, the usefulness of mt gene sequences as a convenient molecular tool in phylogenetic studies of entomopathogenic fungi was demonstrated.

Genome sequence of the mud-dwelling archaeon Methanoplanus limicola type strain (DSM 2279 T), reclassification of Methanoplanus petrolearius as Methanolacinia petrolearia and emended descriptions of the genera Methanoplanus and Methanolacinia

DOE PAGES

Goker, Markus; Lu, Megan; Fiebig, Anne; ...

2014-06-15

Methanoplanus limicola Wildgruber et al. 1984 is a mesophilic methanogen that was isolated from a swamp composed of drilling waste near Naples, Italy, shortly after the Archaea were recognized as a separate domain of life. Methanoplanus is the type genus in the family Methanoplanaceae, a taxon that felt into disuse since modern 16S rRNA gene sequences-based taxonomy was established. Methanoplanus is now placed within the Methanomicrobiaceae, a family that is so far poorly characterized at the genome level. The only other type strain of the genus with a sequenced genome, Methanoplanus petrolearius SEBR 4847 T, turned out to be misclassifiedmore » and required reclassification to Methanolacinia. Both, Methanoplanus and Methanolacinia, needed taxonomic emendations due to a significant deviation of the G+C content of their genomes from previously published (pregenome-sequence era) values. Until now genome sequences were published for only four of the 33 species with validly published names in the Methanomicrobiaceae. Here we describe the features of M. limicola, together with the improved-high-quality draft genome sequence and an notation of the type strain, M3 T. The 3,200,946 bp long chromosome (permanent draft sequence) with its 3,064 protein-coding and 65 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.« less

The use of MALDI-TOF ICMS as an alternative tool for Trichophyton rubrum identification and typing.

PubMed

Pereira, Leonel; Dias, Nicolina; Santos, Cledir; Lima, Nelson

2014-01-01

In this study, the potential of matrix-assisted laser desorption/ionization time-of-flight intact cell mass spectrometry (MALDI-TOF ICMS) was investigated for the identification of clinical isolates. The isolates were analyzed at the species and strain level. Spectral identification by MALDI-TOF ICMS was performed for all strains, and compared with the results of sequencing of the internal transcribed spacers (ITS1 and ITS2), and the 5.8S rDNA region. PCR fingerprinting analysis using primers M13, (GACA)4, and (AC)10 was performed in order to assess the intra-specific variability of Trichophyton rubrum strains. The identification of strains at species level by MALDI-TOF ICMS was in agreement with the previously performed morphological and biochemical analysis. Sequence data confirmed spectral mass identification at species level. Intra-specific variability was assessed. Within the T. rubrum cluster, strains were distributed into smaller highly related sub-groups with a similarity values above 85%. MALDI-TOF ICMS was shown to be a rapid, low-cost and accurate alternative tool for the identification and strain typing of T. rubrum. Copyright © 2012 Elsevier España, S.L. All rights reserved.

Fabibacter misakiensis sp. nov., a marine bacterium isolated from coastal surface water.

PubMed

Wong, Shu-Kuan; Park, Sanghwa; Lee, Jung-Sook; Lee, Keun Chul; Chiura, Hiroshi Xavier; Kogure, Kazuhiro; Hamasaki, Koji

2015-10-01

A slightly curved-rod-shaped, pink-pigmented, Gram-stain-negative, aerobic bacterial strain with gliding motility, designated SK-8T, was isolated from coastal surface water of Misaki, Japan. Phylogenetic trees generated using 16S rRNA gene sequences revealed that strain SK-8T belonged to the genus Fabibacter and showed 96.0 % sequence similarity to the type strain of the most closely related species, Fabibacter pacificus DY53T. The novel isolate was phenotypically and physiologically different from previously described strains. The major cellular fatty acids were iso-C15 : 1 G, iso-C15 : 0 and iso-C17 : 0 3-OH. Major polar lipids were phosphatidylethanolamine, two aminophospholipids and an unidentified phospholipid. The DNA G+C content was 39.1 mol% and MK-7 was the only predominant isoprenoid quinone. On the basis of this taxonomic study employing a polyphasic approach, it was suggested that strain SK-8T represents a novel species of the genus Fabibacter, with the newly proposed name Fabibacter misakiensis sp. nov. The type strain is SK-8T ( = NBRC 110216T = KCTC 32969T).

Co-occurrence of anaerobic bacteria in colorectal carcinomas.

PubMed

Warren, René L; Freeman, Douglas J; Pleasance, Stephen; Watson, Peter; Moore, Richard A; Cochrane, Kyla; Allen-Vercoe, Emma; Holt, Robert A

2013-05-15

Numerous cancers have been linked to microorganisms. Given that colorectal cancer is a leading cause of cancer deaths and the colon is continuously exposed to a high diversity of microbes, the relationship between gut mucosal microbiome and colorectal cancer needs to be explored. Metagenomic studies have shown an association between Fusobacterium species and colorectal carcinoma. Here, we have extended these studies with deeper sequencing of a much larger number (n = 130) of colorectal carcinoma and matched normal control tissues. We analyzed these data using co-occurrence networks in order to identify microbe-microbe and host-microbe associations specific to tumors. We confirmed tumor over-representation of Fusobacterium species and observed significant co-occurrence within individual tumors of Fusobacterium, Leptotrichia and Campylobacter species. This polymicrobial signature was associated with over-expression of numerous host genes, including the gene encoding the pro-inflammatory chemokine Interleukin-8. The tumor-associated bacteria we have identified are all Gram-negative anaerobes, recognized previously as constituents of the oral microbiome, which are capable of causing infection. We isolated a novel strain of Campylobacter showae from a colorectal tumor specimen. This strain is substantially diverged from a previously sequenced oral Campylobacter showae isolate, carries potential virulence genes, and aggregates with a previously isolated tumor strain of Fusobacterium nucleatum. A polymicrobial signature of Gram-negative anaerobic bacteria is associated with colorectal carcinoma tissue.

aes, the gene encoding the esterase B in Escherichia coli, is a powerful phylogenetic marker of the species.

PubMed

Lescat, Mathilde; Hoede, Claire; Clermont, Olivier; Garry, Louis; Darlu, Pierre; Tuffery, Pierre; Denamur, Erick; Picard, Bertrand

2009-12-29

Previous studies have established a correlation between electrophoretic polymorphism of esterase B, and virulence and phylogeny of Escherichia coli. Strains belonging to the phylogenetic group B2 are more frequently implicated in extraintestinal infections and include esterase B2 variants, whereas phylogenetic groups A, B1 and D contain less virulent strains and include esterase B1 variants. We investigated esterase B as a marker of phylogeny and/or virulence, in a thorough analysis of the esterase B-encoding gene. We identified the gene encoding esterase B as the acetyl-esterase gene (aes) using gene disruption. The analysis of aes nucleotide sequences in a panel of 78 reference strains, including the E. coli reference (ECOR) strains, demonstrated that the gene is under purifying selection. The phylogenetic tree reconstructed from aes sequences showed a strong correlation with the species phylogenetic history, based on multi-locus sequence typing using six housekeeping genes. The unambiguous distinction between variants B1 and B2 by electrophoresis was consistent with Aes amino-acid sequence analysis and protein modelling, which showed that substituted amino acids in the two esterase B variants occurred mostly at different sites on the protein surface. Studies in an experimental mouse model of septicaemia using mutant strains did not reveal a direct link between aes and extraintestinal virulence. Moreover, we did not find any genes in the chromosomal region of aes to be associated with virulence. Our findings suggest that aes does not play a direct role in the virulence of E. coli extraintestinal infection. However, this gene acts as a powerful marker of phylogeny, illustrating the extensive divergence of B2 phylogenetic group strains from the rest of the species.

Emergence of Pseudomonas aeruginosa with class 1 integron carrying blaVIM-2 and blaVIM-4 in the University Clinical Hospital of Bialystok (northeastern Poland).

PubMed

Michalska-Falkowska, Anna; Sacha, Paweł Tomasz; Grześ, Henryk; Hauschild, Tomasz; Wieczorek, Piotr; Ojdana, Dominika; Tryniszewska, Elżbieta Anna

2017-07-11

The effectiveness of carbapenems, considered as last-resort antimicrobials in severe infections, becomes compromised by bacterial resistance. The production of metallo-β-lactamases (MBLs) is the most significant threat to carbapenems activity among Pseudomonas aeruginosa. The aim of this study was to assess the presence and type of MBLs genes in carbapenem-resistant P. aeruginosa clinical strains, to identify the location of MBLs genes and to determine genetic relatedness between MBL-producers using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The first identified MBL-positive (with blaVIM genes) P. aeruginosa strains were isolated from patients hospitalized in the University Clinical Hospital of Bialystok in the period from September 2012 to December 2013. Variants of MBLs genes and variable integron regions were characterized by PCR and sequencing. PFGE was performed after digesting of bacterial genomes by XbaI enzyme. By MLST seven housekeeping genes were analyzed for the determination of sequence type (ST). Three strains carried the blaVIM-2 gene and one harbored the blaVIM-4 gene. The blaVIM genes resided within class 1 integrons. PCR mapping of integrons revealed the presence of four different cassette arrays. Genetic relatedness analysis by PFGE classified VIM-positive strains into four unrelated pulsotypes (A-D). MLST demonstrated the presence of four (ST 111, ST27, and ST17) different sequence type including one previously undescribed new type of ST 2342. Antimicrobial susceptibility testing showed that VIM-positive strains were resistant to carbapenems, cephalosporins, aminoglycosides, and quinolones, intermediate to aztreonam, and susceptible only to colistin. Integrons mapping, PFGE, and MLST results may point to different origin of these strains and independent introduction into hospitalized patients.

Description of Mycobacterium chelonae subsp. bovis subsp. nov., isolated from cattle (Bos taurus coreanae), emended description of Mycobacterium chelonae and creation of Mycobacterium chelonae subsp. chelonae subsp. nov.

PubMed

Kim, Byoung-Jun; Kim, Ga-Na; Kim, Bo-Ram; Jeon, Che Ok; Jeong, Joseph; Lee, Seon Ho; Lim, Ji-Hun; Lee, Seung-Heon; Kim, Chang Ki; Kook, Yoon-Hoh; Kim, Bum-Joon

2017-10-01

Three rapidly growing mycobacterial strains, QIA-37 T , QIA-40 and QIA-41, were isolated from the lymph nodes of three separate Korean native cattle, Hanwoo (Bos taurus coreanae). These strains were previously shown to be phylogenetically distinct but closely related to Mycobacterium chelonae ATCC 35752 T by taxonomic approaches targeting three genes (16S rRNA, hsp6 and rpoB) and were further characterized using a polyphasic approach in this study. The 16S rRNA gene sequences of all three strains showed 99.7 % sequence similarity with that of the M. chelonae type strain. A multilocus sequence typing analysis targeting 10 housekeeping genes, including hsp65 and rpoB, revealed a phylogenetic cluster of these strains with M. chelonae. DNA-DNA hybridization values of 78.2 % between QIA-37 T and M. chelonae indicated that it belongs to M. chelonae but is a novel subspecies distinct from M. chelonae. Phylogenetic analysis based on whole-genome sequences revealed a 95.44±0.06 % average nucleotide identity (ANI) value with M. chelonae, slightly higher than the 95.0 % ANI criterion for determining a novel species. In addition, distinct phenotypic characteristics such as positive growth at 37 °C, at which temperature M. chelonae does not grow, further support the taxonomic status of these strains as representatives of a novel subspecies of M. chelonae. Therefore, we propose an emended description of Mycobacterium chelonae, and descriptions of M. chelonae subsp. chelonae subsp. nov. and M. chelonae subsp. bovis subsp. nov. are presented; strains ATCC 35752 T (=CCUG 47445 T =CIP 104535 T =DSM 43804 T =JCM 6388 T =NCTC 946 T ) and QIA-37 T (=KCTC 39630 T =JCM 30986 T ) are the type strains of the two novel subspecies.

Genomic and Genetic Diversity within the Pseudomonas fluorescens Complex

PubMed Central

Garrido-Sanz, Daniel; Meier-Kolthoff, Jan P.; Göker, Markus; Martín, Marta; Rivilla, Rafael; Redondo-Nieto, Miguel

2016-01-01

The Pseudomonas fluorescens complex includes Pseudomonas strains that have been taxonomically assigned to more than fifty different species, many of which have been described as plant growth-promoting rhizobacteria (PGPR) with potential applications in biocontrol and biofertilization. So far the phylogeny of this complex has been analyzed according to phenotypic traits, 16S rDNA, MLSA and inferred by whole-genome analysis. However, since most of the type strains have not been fully sequenced and new species are frequently described, correlation between taxonomy and phylogenomic analysis is missing. In recent years, the genomes of a large number of strains have been sequenced, showing important genomic heterogeneity and providing information suitable for genomic studies that are important to understand the genomic and genetic diversity shown by strains of this complex. Based on MLSA and several whole-genome sequence-based analyses of 93 sequenced strains, we have divided the P. fluorescens complex into eight phylogenomic groups that agree with previous works based on type strains. Digital DDH (dDDH) identified 69 species and 75 subspecies within the 93 genomes. The eight groups corresponded to clustering with a threshold of 31.8% dDDH, in full agreement with our MLSA. The Average Nucleotide Identity (ANI) approach showed inconsistencies regarding the assignment to species and to the eight groups. The small core genome of 1,334 CDSs and the large pan-genome of 30,848 CDSs, show the large diversity and genetic heterogeneity of the P. fluorescens complex. However, a low number of strains were enough to explain most of the CDSs diversity at core and strain-specific genomic fractions. Finally, the identification and analysis of group-specific genome and the screening for distinctive characters revealed a phylogenomic distribution of traits among the groups that provided insights into biocontrol and bioremediation applications as well as their role as PGPR. PMID:26915094

DNA homology among diverse spiroplasma strains representing several serological groups.

PubMed

Lee, I M; Davis, R E

1980-11-01

Deoxyribonucleic acid (DNA) homology among 10 strains of spiroplasma associated with plants and insects was assessed by analysis of DNA-DNA hybrids with single strand specific S1 nuclease. Based on DNA homology, the spiroplasmas could be divided into three genetically distinct groups (designated I, II, and III), corresponding to three separate serogroups described previously. DNA sequence homology between the three groups was less than or equal to 5%. Based on DNA homology, group I could be divided into three subgroups (A, B, and C) that corresponded to three serological subgroups of serogroup I. Subgroup A contained Spiroplasma citri strains Maroc R8A2 and C 189; subgroup B contained strains AS 576 from honey bee and G 1 from flowers; subgroup C contained corn stunt spiroplasma strains I-747 and PU 8-17. There was 27-54% DNA sequence homology among these three subgroups. Group II contained strains 23-6 and 27-31 isolated from flowers of tulip tree (Liriodendron tulipifera L.). Group III contained strains SR 3 and SR 9, other isolates from flowers of tulip tree. Based on thermal denaturation, guanine plus cytosine contents of DNA from five type strains representing all groups and subgroups were estimated to be close to 26 mol% for group I strains, close to 25 mol% for group II strains, and close to 29 mol% for group III strains. The genome molecular weights of these five type strains were all estimated to bae about 10(9).

Identification of the psaA Gene, Coding for Pneumococcal Surface Adhesin A, in Viridans Group Streptococci other than Streptococcus pneumoniae

PubMed Central

Jado, Isabel; Fenoll, Asunción; Casal, Julio; Pérez, Amalia

2001-01-01

The gene encoding the pneumococcal surface adhesin A (PsaA) protein has been identified in three different viridans group streptococcal species. Comparative studies of the psaA gene identified in different pneumococcal isolates by sequencing PCR products showed a high degree of conservation among these strains. PsaA is encoded by an open reading frame of 930 bp. The analysis of this fragment in Streptococcus mitis, Streptococcus oralis, and Streptococcus anginosus strains revealed a sequence identity of 95, 94, and 90%, respectively, to the corresponding open reading frame of the previously reported Streptococcus pneumoniae serotype 6B strain. Our results confirm that psaA is present and detectable in heterologous bacterial species. The possible implications of these results for the suitability and potential use of PsaA in the identification and diagnosis of pneumococcal diseases are discussed. PMID:11527799

A cultivation-independent PCR-RFLP assay targeting oprF gene for detection and identification of Pseudomonas spp. in samples from fibrocystic pediatric patients.

PubMed

Lagares, Antonio; Agaras, Betina; Bettiol, Marisa P; Gatti, Blanca M; Valverde, Claudio

2015-07-01

Species-specific genetic markers are crucial to develop faithful and sensitive molecular methods for the detection and identification of Pseudomonas aeruginosa (Pa). We have previously set up a PCR-RFLP protocol targeting oprF, the gene encoding the genus-specific outer membrane porin F, whose strong conservation and marked sequence diversity allowed detection and differentiation of environmental isolates (Agaras et al., 2012). Here, we evaluated the ability of the PCR-RFLP assay to genotype clinical isolates previously identified as Pa by conventional microbiological methods within a collection of 62 presumptive Pa isolates from different pediatric clinical samples and different sections of the Hospital de Niños "Sor María Ludovica" from La Plata, Argentina. All isolates, but one, gave an oprF amplicon consistent with that from reference Pa strains. The sequence of the smaller-sized amplicon revealed that the isolate was in fact a mendocina Pseudomonas strain. The oprF RFLP pattern generated with TaqI or HaeIII nucleases matched those of reference Pa strains for 59 isolates (96%). The other two Pa isolates (4%) revealed a different RFLP pattern based on HaeIII digestion, although oprF sequencing confirmed that Pa identification was correct. We next tested the effectiveness of the PCR-RFLP to detect pseudomonads on clinical samples of pediatric fibrocystic patients directly without sample cultivation. The expected amplicon and its cognate RFLP profile were obtained for all samples in which Pa was previously detected by cultivation-dependent methods. Altogether, these results provide the basis for the application of the oprF PCR-RFLP protocol to directly detect and identify Pa and other non-Pa pseudomonads in fibrocystic clinical samples. Copyright © 2015 Elsevier B.V. All rights reserved.

Genetic Characterization of Atypical Enteropathogenic Escherichia coli Isolates from Ewes' Milk, Sheep Farm Environments, and Humans by Multilocus Sequence Typing and Pulsed-Field Gel Electrophoresis

PubMed Central

Otero, Verónica; Rodríguez-Calleja, José-María; Otero, Andrés; García-López, María-Luisa

2013-01-01

A collection of 81 isolates of enteropathogenic Escherichia coli (EPEC) was obtained from samples of bulk tank sheep milk (62 isolates), ovine feces (4 isolates), sheep farm environment (water, 4 isolates; air, 1 isolate), and human stool samples (9 isolates). The strains were considered atypical EPEC organisms, carrying the eae gene without harboring the pEAF plasmid. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 19 sequence types (ST) were detected, with none of them having been previously reported for atypical EPEC. The most frequent ST included 41 strains isolated from milk and human stool samples. Genetic typing by pulsed-field gel electrophoresis (PFGE) resulted in 57 patterns which grouped in 24 clusters. Comparison of strains isolated from the different samples showed phylogenetic relationships between milk and human isolates and also between milk and water isolates. The results obtained show a possible risk for humans due to the presence of atypical EPEC in ewes' milk and suggest a transmission route for this emerging pathogen through contaminated water. PMID:23872571

Genome sequence and physiological analysis of Yamadazyma laniorum f.a. sp. nov. and a reevaluation of the apocryphal xylose fermentation of its sister species, Candida tenuis.

PubMed

Haase, Max A B; Kominek, Jacek; Langdon, Quinn K; Kurtzman, Cletus P; Hittinger, Chris Todd

2017-05-01

Xylose fermentation is a rare trait that is immensely important to the cellulosic biofuel industry, and Candida tenuis is one of the few yeasts that has been reported with this trait. Here we report the isolation of two strains representing a candidate sister species to C. tenuis. Integrated analysis of genome sequence and physiology suggested the genetic basis of a number of traits, including variation between the novel species and C. tenuis in lactose metabolism due to the loss of genes encoding lactose permease and β-galactosidase in the former. Surprisingly, physiological characterization revealed that neither the type strain of C. tenuis nor this novel species fermented xylose in traditional assays. We reexamined three xylose-fermenting strains previously identified as C. tenuis and found that these strains belong to the genus Scheffersomyces and are not C. tenuis. We propose Yamadazyma laniorum f.a. sp. nov. to accommodate our new strains and designate its type strain as yHMH7 (=CBS 14780 = NRRL Y-63967T). Furthermore, we propose the transfer of Candida tenuis to the genus Yamadazyma as Yamadazyma tenuis comb. nov. This approach provides a roadmap for how integrated genome sequence and physiological analysis can yield insight into the mechanisms that generate yeast biodiversity. Published by Oxford University Press on behalf of FEMS 2017. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Identification of non-streptococcal organisms from human dental plaque grown on the Streptococcus-selective medium mitis-salivarius agar.

PubMed

Kim, Yeon-Hee; Lee, Si Young

2015-02-01

Mitis-salivarius (MS) agar has been used widely in microbial epidemiological studies because oral viridans streptococci can be selectively grown on this medium. Even though the previous findings reported the limited selecting power of MS agar for streptococcus strains, the identities of non-streptococcal strains from human oral samples which can grow on this medium are not clear yet. In this study, we identified non-streptococcal organisms grown on MS agar plates by polymerase chain reaction (PCR) amplification and sequencing of the 16S ribosomal RNA (rRNA) gene. Eighty bacterial colonies on MS plates were isolated from plaque samples, and bacterial identification was achieved with the rapid ID 32 Strep system and mini API reader. The bacterial colonies identified as non-streptococci by the API system were selected for further identification. The 16S rRNA gene was amplified by PCR and verified using DNA sequencing analysis for identification. Sequences were compared with those of reference organisms in the genome database of the National Center for Biotechnology Information using the Basic Local Alignment Search Tool (BLAST). Among the 11 isolated non-streptococcal strains on MS plates, 3 strains were identified as Actinomyces naeslundii, 7 strains were identified as Actinomyces oris and 1 strain were identified as Actinomyces sp. using Blastn. In this study, we showed that some oral Actinomyces species can grow on Streptococcus-selective MS agar plates. Copyright © 2014 Elsevier Ltd. All rights reserved.

Molecular evidence of Burkholderia pseudomallei genotypes based on geographical distribution.

PubMed

Zulkefli, Noorfatin Jihan; Mariappan, Vanitha; Vellasamy, Kumutha Malar; Chong, Chun Wie; Thong, Kwai Lin; Ponnampalavanar, Sasheela; Vadivelu, Jamuna; Teh, Cindy Shuan Ju

2016-01-01

Background. Central intermediary metabolism (CIM) in bacteria is defined as a set of metabolic biochemical reactions within a cell, which is essential for the cell to survive in response to environmental perturbations. The genes associated with CIM are commonly found in both pathogenic and non-pathogenic strains. As these genes are involved in vital metabolic processes of bacteria, we explored the efficiency of the genes in genotypic characterization of Burkholderia pseudomallei isolates, compared with the established pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) schemes. Methods. Nine previously sequenced B. pseudomallei isolates from Malaysia were characterized by PFGE, MLST and CIM genes. The isolates were later compared to the other 39 B. pseudomallei strains, retrieved from GenBank using both MLST and sequence analysis of CIM genes. UniFrac and hierachical clustering analyses were performed using the results generated by both MLST and sequence analysis of CIM genes. Results. Genetic relatedness of nine Malaysian B. pseudomallei isolates and the other 39 strains was investigated. The nine Malaysian isolates were subtyped into six PFGE profiles, four MLST profiles and five sequence types based on CIM genes alignment. All methods demonstrated the clonality of OB and CB as well as CMS and THE. However, PFGE showed less than 70% similarity between a pair of morphology variants, OS and OB. In contrast, OS was identical to the soil isolate, MARAN. To have a better understanding of the genetic diversity of B. pseudomallei worldwide, we further aligned the sequences of genes used in MLST and genes associated with CIM for the nine Malaysian isolates and 39 B. pseudomallei strains from NCBI database. Overall, based on the CIM genes, the strains were subtyped into 33 profiles where majority of the strains from Asian countries were clustered together. On the other hand, MLST resolved the isolates into 31 profiles which formed three clusters. Hierarchical clustering using UniFrac distance suggested that the isolates from Australia were genetically distinct from the Asian isolates. Nevertheless, statistical significant differences were detected between isolates from Malaysia, Thailand and Australia. Discussion. Overall, PFGE showed higher discriminative power in clustering the nine Malaysian B. pseudomallei isolates and indicated its suitability for localized epidemiological study. Compared to MLST, CIM genes showed higher resolution in distinguishing those non-related strains and better clustering of strains from different geographical regions. A closer genetic relatedness of Malaysian isolates with all Asian strains in comparison to Australian strains was observed. This finding was supported by UniFrac analysis which resulted in geographical segregation between Australia and the Asian countries.

The Evolutionary Divergence of Shiga Toxin-Producing Escherichia coli Is Reflected in Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) Spacer Composition

PubMed Central

Yin, Shuang; Jensen, Mark A.; Bai, Jiawei; DebRoy, Chitrita; Barrangou, Rodolphe

2013-01-01

The Shiga toxin-producing Escherichia coli (STEC) strains, including those of O157:H7 and the “big six” serogroups (i.e., serogroups O26, O45, O103, O111, O121, and O145), are a group of pathogens designated food adulterants in the United States. The relatively conserved nature of clustered regularly interspaced short palindromic repeats (CRISPRs) in phylogenetically related E. coli strains makes them potential subtyping markers for STEC detection, and a quantitative PCR (qPCR)-based assay was previously developed for O26:H11, O45:H2, O103:H2, O111:H8, O121:H19, O145:H28, and O157:H7 isolates. To better evaluate the sensitivity and specificity of this qPCR method, the CRISPR loci of 252 O157 and big-six STEC isolates were sequenced and analyzed along with 563 CRISPR1 and 624 CRISPR2 sequences available in GenBank. General conservation of spacer content and order was observed within each O157 and big-six serogroup, validating the qPCR method. Meanwhile, it was found that spacer deletion, the presence of an insertion sequence, and distinct alleles within a serogroup are sources of false-negative reactions. Conservation of CRISPR arrays among isolates expressing the same flagellar antigen, specifically, H7, H2, and H11, suggested that these isolates share an ancestor and provided an explanation for the false positives previously observed in the qPCR results. An analysis of spacer distribution across E. coli strains provided limited evidence for temporal spacer acquisition. Conversely, comparison of CRISPR sequences between strains along the stepwise evolution of O157:H7 from its O55:H7 ancestor revealed that, over this ∼7,000-year span, spacer deletion was the primary force generating CRISPR diversity. PMID:23851088

The evolutionary divergence of Shiga toxin-producing Escherichia coli is reflected in clustered regularly interspaced short palindromic repeat (CRISPR) spacer composition.

PubMed

Yin, Shuang; Jensen, Mark A; Bai, Jiawei; Debroy, Chitrita; Barrangou, Rodolphe; Dudley, Edward G

2013-09-01

The Shiga toxin-producing Escherichia coli (STEC) strains, including those of O157:H7 and the "big six" serogroups (i.e., serogroups O26, O45, O103, O111, O121, and O145), are a group of pathogens designated food adulterants in the United States. The relatively conserved nature of clustered regularly interspaced short palindromic repeats (CRISPRs) in phylogenetically related E. coli strains makes them potential subtyping markers for STEC detection, and a quantitative PCR (qPCR)-based assay was previously developed for O26:H11, O45:H2, O103:H2, O111:H8, O121:H19, O145:H28, and O157:H7 isolates. To better evaluate the sensitivity and specificity of this qPCR method, the CRISPR loci of 252 O157 and big-six STEC isolates were sequenced and analyzed along with 563 CRISPR1 and 624 CRISPR2 sequences available in GenBank. General conservation of spacer content and order was observed within each O157 and big-six serogroup, validating the qPCR method. Meanwhile, it was found that spacer deletion, the presence of an insertion sequence, and distinct alleles within a serogroup are sources of false-negative reactions. Conservation of CRISPR arrays among isolates expressing the same flagellar antigen, specifically, H7, H2, and H11, suggested that these isolates share an ancestor and provided an explanation for the false positives previously observed in the qPCR results. An analysis of spacer distribution across E. coli strains provided limited evidence for temporal spacer acquisition. Conversely, comparison of CRISPR sequences between strains along the stepwise evolution of O157:H7 from its O55:H7 ancestor revealed that, over this ∼7,000-year span, spacer deletion was the primary force generating CRISPR diversity.

Intra-genotypic Diversity of Archival G4P[8] Human Rotaviruses from Washington, DC

PubMed Central

McDonald, Sarah M.; Davis, Kristin; McAllen, John K.; Spiro, David J.; Patton, John T.

2011-01-01

Group A human rotaviruses (RVs) remain the most frequently detected viral agents associated with acute gastroenteritis in infants and young children. Despite their medical importance, relatively few complete genome sequences have been determined for commonly circulating G/P-type strains (i.e., G1P[8], G2P[4], G3P[8], G4P[8], and G9P[8]). In the current study, we sequenced the genomes of 11 G4P[8] isolates from stool specimens that were collected in Washington, DC during the years of 1974-1991. We found that the VP7-VP4-VP6-VP1-VP2-VP3-NSP1-NSP2-NSP3-NSP4-NSP5/6-encoding genes of all 11 G4P[8] RVs have the genotypes of G4-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1. By constructing phylogenetic trees for each gene, extensive intra-genotypic diversity was revealed among the G4P[8] RVs, and new sub-genotype gene alleles were identified. Several of these alleles are nearly identical to those of G3P[8] isolates previously sequenced from this same Washington, DC collection, strongly suggesting that the RVs underwent gene reassortment. On the other hand, we observed that some G4P[8] RVs exhibit completely different allele-based genome constellations, despite being collected during the same epidemic season; there was no evidence of gene reassortment between these strains. This observation extends our previous findings and supports the notion that stable, genetically-distinct clades of human RVs with the same G/P-type can co-circulate in a community. Interestingly, the sub-genotype gene alleles found in some of the DC RVs share a close evolutionary relationship with genes of more contemporary human strains. Thus, archival human RVs sequenced in this study might represent evolutionary precursors to modern-day strains. PMID:21712102

Complete genome sequence of Enterococcus faecium strain TX16 and comparative genomic analysis of Enterococcus faecium genomes

PubMed Central

2012-01-01

Background Enterococci are among the leading causes of hospital-acquired infections in the United States and Europe, with Enterococcus faecalis and Enterococcus faecium being the two most common species isolated from enterococcal infections. In the last decade, the proportion of enterococcal infections caused by E. faecium has steadily increased compared to other Enterococcus species. Although the underlying mechanism for the gradual replacement of E. faecalis by E. faecium in the hospital environment is not yet understood, many studies using genotyping and phylogenetic analysis have shown the emergence of a globally dispersed polyclonal subcluster of E. faecium strains in clinical environments. Systematic study of the molecular epidemiology and pathogenesis of E. faecium has been hindered by the lack of closed, complete E. faecium genomes that can be used as references. Results In this study, we report the complete genome sequence of the E. faecium strain TX16, also known as DO, which belongs to multilocus sequence type (ST) 18, and was the first E. faecium strain ever sequenced. Whole genome comparison of the TX16 genome with 21 E. faecium draft genomes confirmed that most clinical, outbreak, and hospital-associated (HA) strains (including STs 16, 17, 18, and 78), in addition to strains of non-hospital origin, group in the same clade (referred to as the HA clade) and are evolutionally considerably more closely related to each other by phylogenetic and gene content similarity analyses than to isolates in the community-associated (CA) clade with approximately a 3–4% average nucleotide sequence difference between the two clades at the core genome level. Our study also revealed that many genomic loci in the TX16 genome are unique to the HA clade. 380 ORFs in TX16 are HA-clade specific and antibiotic resistance genes are enriched in HA-clade strains. Mobile elements such as IS16 and transposons were also found almost exclusively in HA strains, as previously reported. Conclusions Our findings along with other studies show that HA clonal lineages harbor specific genetic elements as well as sequence differences in the core genome which may confer selection advantages over the more heterogeneous CA E. faecium isolates. Which of these differences are important for the success of specific E. faecium lineages in the hospital environment remain(s) to be determined. PMID:22769602

Complete Genome Sequence of Sporisorium scitamineum and Biotrophic Interaction Transcriptome with Sugarcane

PubMed Central

Benevenuto, Juliana; Peters, Leila P.; Carvalho, Giselle; Palhares, Alessandra; Quecine, Maria C.; Nunes, Filipe R. S.; Kmit, Maria C. P.; Wai, Alvan; Hausner, Georg; Aitken, Karen S.; Berkman, Paul J.; Fraser, James A.; Moolhuijzen, Paula M.; Coutinho, Luiz L.; Creste, Silvana; Vieira, Maria L. C.; Kitajima, João P.; Monteiro-Vitorello, Claudia B.

2015-01-01

Sporisorium scitamineum is a biotrophic fungus responsible for the sugarcane smut, a worldwide spread disease. This study provides the complete sequence of individual chromosomes of S. scitamineum from telomere to telomere achieved by a combination of PacBio long reads and Illumina short reads sequence data, as well as a draft sequence of a second fungal strain. Comparative analysis to previous available sequences of another strain detected few polymorphisms among the three genomes. The novel complete sequence described herein allowed us to identify and annotate extended subtelomeric regions, repetitive elements and the mitochondrial DNA sequence. The genome comprises 19,979,571 bases, 6,677 genes encoding proteins, 111 tRNAs and 3 assembled copies of rDNA, out of our estimated number of copies as 130. Chromosomal reorganizations were detected when comparing to sequences of S. reilianum, the closest smut relative, potentially influenced by repeats of transposable elements. Repetitive elements may have also directed the linkage of the two mating-type loci. The fungal transcriptome profiling from in vitro and from interaction with sugarcane at two time points (early infection and whip emergence) revealed that 13.5% of the genes were differentially expressed in planta and particular to each developmental stage. Among them are plant cell wall degrading enzymes, proteases, lipases, chitin modification and lignin degradation enzymes, sugar transporters and transcriptional factors. The fungus also modulates transcription of genes related to surviving against reactive oxygen species and other toxic metabolites produced by the plant. Previously described effectors in smut/plant interactions were detected but some new candidates are proposed. Ten genomic islands harboring some of the candidate genes unique to S. scitamineum were expressed only in planta. RNAseq data was also used to reassure gene predictions. PMID:26065709

Genetic recombination of tick-borne flaviviruses among wild-type strains.

PubMed

Norberg, Peter; Roth, Anette; Bergström, Tomas

2013-06-05

Genetic recombination has been suggested to occur in mosquito-borne flaviviruses. In contrast, tick-borne flaviviruses have been thought to evolve in a clonal manner, although recent studies suggest that recombination occurs also for these viruses. We re-analyzed the data and found that previous conclusions on wild type recombination were probably falsely drawn due to misalignments of nucleotide sequences, ambiguities in GenBank sequences, or different laboratory culture histories suggestive of recombination events in laboratory. To evaluate if reliable predictions of wild type recombination of tick-borne flaviviruses can be made, we analyzed viral strains sequenced exclusively for this study, and other flavivirus sequences retrieved from GenBank. We detected genetic signals supporting recombination between viruses within the three clades of TBEV-Eu, TBEV-Sib and TBEV-Fe, respectively. Our results suggest that the tick-borne encephalitis viruses may undergo recombination under natural conditions, but that geographic barriers restrict most recombination events to involve only closely genetically related viruses. Copyright © 2013 Elsevier Inc. All rights reserved.

Taxonomy of oxalotrophic Methylobacterium strains

NASA Astrophysics Data System (ADS)

Sahin, Nurettin; Kato, Yuko; Yilmaz, Ferah

2008-10-01

Most of the oxalotrophic bacteria are facultative methylotrophs and play important ecological roles in soil fertility and cycling of elements. This study gives a detailed picture of the taxonomy and diversity of these bacteria and provides new information about the taxonomical variability within the genus Methylobacterium. Twelve mesophilic, pink-pigmented, and facultatively methylotrophic oxalate-oxidizing strains were included in this work that had been previously isolated from the soil and some plant tissues by the potassium oxalate enrichment method. The isolates were characterized using biochemical tests, cellular lipid profiles, spectral characteristics of carotenoid pigments, G+C content of the DNA, and 16S rDNA sequencing. The taxonomic similarities among the strains were analyzed using the simple matching ( S SM) and Jaccard ( S J) coefficients, and the UPGMA clustering algorithm. The phylogenetic position of the strains was inferred by the neighbor-joining method on the basis of the 16S rDNA sequences. All isolates were Gram-negative, facultatively methylotrophic, oxidase and catalase positive, and required no growth factors. Based on the results of numerical taxonomy, the strains formed four closely related clusters sharing ≥85% similarity. Analysis of the 16S rDNA sequences demonstrated that oxalotrophic, pink-pigmented, and facultatively methylotrophic strains could be identified as members of the genus Methylobacterium. Except for M. variabile and M. aquaticum, all of the Methylobacterium type strains tested had the ability of oxalate utilization. Our results indicate that the capability of oxalate utilization seems to be an uncommon trait and could be used as a valuable taxonomic criterion for differentiation of Methylobacterium species.

Taxonomy of oxalotrophic Methylobacterium strains.

PubMed

Sahin, Nurettin; Kato, Yuko; Yilmaz, Ferah

2008-10-01

Most of the oxalotrophic bacteria are facultative methylotrophs and play important ecological roles in soil fertility and cycling of elements. This study gives a detailed picture of the taxonomy and diversity of these bacteria and provides new information about the taxonomical variability within the genus Methylobacterium. Twelve mesophilic, pink-pigmented, and facultatively methylotrophic oxalate-oxidizing strains were included in this work that had been previously isolated from the soil and some plant tissues by the potassium oxalate enrichment method. The isolates were characterized using biochemical tests, cellular lipid profiles, spectral characteristics of carotenoid pigments, G+C content of the DNA, and 16S rDNA sequencing. The taxonomic similarities among the strains were analyzed using the simple matching (S (SM)) and Jaccard (S (J)) coefficients, and the UPGMA clustering algorithm. The phylogenetic position of the strains was inferred by the neighbor-joining method on the basis of the 16S rDNA sequences. All isolates were Gram-negative, facultatively methylotrophic, oxidase and catalase positive, and required no growth factors. Based on the results of numerical taxonomy, the strains formed four closely related clusters sharing > or =85% similarity. Analysis of the 16S rDNA sequences demonstrated that oxalotrophic, pink-pigmented, and facultatively methylotrophic strains could be identified as members of the genus Methylobacterium. Except for M. variabile and M. aquaticum, all of the Methylobacterium type strains tested had the ability of oxalate utilization. Our results indicate that the capability of oxalate utilization seems to be an uncommon trait and could be used as a valuable taxonomic criterion for differentiation of Methylobacterium species.

Simplifying complex sequence information: a PCP-consensus protein binds antibodies against all four Dengue serotypes.

PubMed

Bowen, David M; Lewis, Jessica A; Lu, Wenzhe; Schein, Catherine H

2012-09-14

Designing proteins that reflect the natural variability of a pathogen is essential for developing novel vaccines and drugs. Flaviviruses, including Dengue (DENV) and West Nile (WNV), evolve rapidly and can "escape" neutralizing monoclonal antibodies by mutation. Designing antigens that represent many distinct strains is important for DENV, where infection with a strain from one of the four serotypes may lead to severe hemorrhagic disease on subsequent infection with a strain from another serotype. Here, a DENV physicochemical property (PCP)-consensus sequence was derived from 671 unique sequences from the Flavitrack database. PCP-consensus proteins for domain 3 of the envelope protein (EdomIII) were expressed from synthetic genes in Escherichia coli. The ability of the purified consensus proteins to bind polyclonal antibodies generated in response to infection with strains from each of the four DENV serotypes was determined. The initial consensus protein bound antibodies from DENV-1-3 in ELISA and Western blot assays. This sequence was altered in 3 steps to incorporate regions of maximum variability, identified as significant changes in the PCPs, characteristic of DENV-4 strains. The final protein was recognized by antibodies against all four serotypes. Two amino acids essential for efficient binding to all DENV antibodies are part of a discontinuous epitope previously defined for a neutralizing monoclonal antibody. The PCP-consensus method can significantly reduce the number of experiments required to define a multivalent antigen, which is particularly important when dealing with pathogens that must be tested at higher biosafety levels. Copyright © 2012 Elsevier Ltd. All rights reserved.

A novel prophage identified in strains from Salmonella enterica serovar Enteritidis is a phylogenetic signature of the lineage ST-1974

PubMed Central

D'Alessandro, Bruno; Pérez Escanda, Victoria; Balestrazzi, Lucía; Iriarte, Andrés; Pickard, Derek; Yim, Lucía; Chabalgoity, José Alejandro; Betancor, Laura

2018-01-01

Salmonella enterica serovar Enteritidis is a major agent of foodborne diseases worldwide. In Uruguay, this serovar was almost negligible until the mid 1990s but since then it has become the most prevalent. Previously, we characterized a collection of strains isolated from 1988 to 2005 and found that the two oldest strains were the most genetically divergent. In order to further characterize these strains, we sequenced and annotated eight genomes including those of the two oldest isolates. We report on the identification and characterization of a novel 44 kbp Salmonella prophage found exclusively in these two genomes. Sequence analysis reveals that the prophage is a mosaic, with homologous regions in different Salmonella prophages. It contains 60 coding sequences, including two genes, gogB and sseK3, involved in virulence and modulation of host immune response. Analysis of serovar Enteritidis genomes available in public databases confirmed that this prophage is absent in most of them, with the exception of a group of 154 genomes. All 154 strains carrying this prophage belong to the same sequence type (ST-1974), suggesting that its acquisition occurred in a common ancestor. We tested this by phylogenetic analysis of 203 genomes representative of the intraserovar diversity. The ST-1974 forms a distinctive monophyletic lineage, and the newly described prophage is a phylogenetic signature of this lineage that could be used as a molecular marker. The phylogenetic analysis also shows that the major ST (ST-11) is polyphyletic and might have given rise to almost all other STs, including ST-1974. PMID:29509137

Genetic analysis of a novel Xylella fastidiosa subspecies found in the southwestern United States.

PubMed

Randall, Jennifer J; Goldberg, Natalie P; Kemp, John D; Radionenko, Maxim; French, Jason M; Olsen, Mary W; Hanson, Stephen F

2009-09-01

Xylella fastidiosa, the causal agent of several scorch diseases, is associated with leaf scorch symptoms in Chitalpa tashkentensis, a common ornamental landscape plant used throughout the southwestern United States. For a number of years, many chitalpa trees in southern New Mexico and Arizona exhibited leaf scorch symptoms, and the results from a regional survey show that chitalpa trees from New Mexico, Arizona, and California are frequently infected with X. fastidiosa. Phylogenetic analysis of multiple loci was used to compare the X. fastidiosa infecting chitalpa strains from New Mexico, Arizona, and trees imported into New Mexico nurseries with previously reported X. fastidiosa strains. Loci analyzed included the 16S ribosome, 16S-23S ribosomal intergenic spacer region, gyrase-B, simple sequence repeat sequences, X. fastidiosa-specific sequences, and the virulence-associated protein (VapD). This analysis indicates that the X. fastidiosa isolates associated with infected chitalpa trees in the Southwest are a highly related group that is distinct from the four previously defined taxons X. fastidiosa subsp. fastidiosa (piercei), X. fastidiosa subsp. multiplex, X. fastidiosa subsp. sandyi, and X. fastidiosa subsp. pauca. Therefore, the classification proposed for this new subspecies is X. fastidiosa subsp. tashke.

Whole Genome Sequencing for Genomics-Guided Investigations of Escherichia coli O157:H7 Outbreaks.

PubMed

Rusconi, Brigida; Sanjar, Fatemeh; Koenig, Sara S K; Mammel, Mark K; Tarr, Phillip I; Eppinger, Mark

2016-01-01

Multi isolate whole genome sequencing (WGS) and typing for outbreak investigations has become a reality in the post-genomics era. We applied this technology to strains from Escherichia coli O157:H7 outbreaks. These include isolates from seven North America outbreaks, as well as multiple isolates from the same patient and from different infected individuals in the same household. Customized high-resolution bioinformatics sequence typing strategies were developed to assess the core genome and mobilome plasticity. Sequence typing was performed using an in-house single nucleotide polymorphism (SNP) discovery and validation pipeline. Discriminatory power becomes of particular importance for the investigation of isolates from outbreaks in which macrogenomic techniques such as pulse-field gel electrophoresis or multiple locus variable number tandem repeat analysis do not differentiate closely related organisms. We also characterized differences in the phage inventory, allowing us to identify plasticity among outbreak strains that is not detectable at the core genome level. Our comprehensive analysis of the mobilome identified multiple plasmids that have not previously been associated with this lineage. Applied phylogenomics approaches provide strong molecular evidence for exceptionally little heterogeneity of strains within outbreaks and demonstrate the value of intra-cluster comparisons, rather than basing the analysis on archetypal reference strains. Next generation sequencing and whole genome typing strategies provide the technological foundation for genomic epidemiology outbreak investigation utilizing its significantly higher sample throughput, cost efficiency, and phylogenetic relatedness accuracy. These phylogenomics approaches have major public health relevance in translating information from the sequence-based survey to support timely and informed countermeasures. Polymorphisms identified in this work offer robust phylogenetic signals that index both short- and long-term evolution and can complement currently employed typing schemes for outbreak ex- and inclusion, diagnostics, surveillance, and forensic studies.

Whole Genome Sequencing for Genomics-Guided Investigations of Escherichia coli O157:H7 Outbreaks

PubMed Central

Rusconi, Brigida; Sanjar, Fatemeh; Koenig, Sara S. K.; Mammel, Mark K.; Tarr, Phillip I.; Eppinger, Mark

2016-01-01

Multi isolate whole genome sequencing (WGS) and typing for outbreak investigations has become a reality in the post-genomics era. We applied this technology to strains from Escherichia coli O157:H7 outbreaks. These include isolates from seven North America outbreaks, as well as multiple isolates from the same patient and from different infected individuals in the same household. Customized high-resolution bioinformatics sequence typing strategies were developed to assess the core genome and mobilome plasticity. Sequence typing was performed using an in-house single nucleotide polymorphism (SNP) discovery and validation pipeline. Discriminatory power becomes of particular importance for the investigation of isolates from outbreaks in which macrogenomic techniques such as pulse-field gel electrophoresis or multiple locus variable number tandem repeat analysis do not differentiate closely related organisms. We also characterized differences in the phage inventory, allowing us to identify plasticity among outbreak strains that is not detectable at the core genome level. Our comprehensive analysis of the mobilome identified multiple plasmids that have not previously been associated with this lineage. Applied phylogenomics approaches provide strong molecular evidence for exceptionally little heterogeneity of strains within outbreaks and demonstrate the value of intra-cluster comparisons, rather than basing the analysis on archetypal reference strains. Next generation sequencing and whole genome typing strategies provide the technological foundation for genomic epidemiology outbreak investigation utilizing its significantly higher sample throughput, cost efficiency, and phylogenetic relatedness accuracy. These phylogenomics approaches have major public health relevance in translating information from the sequence-based survey to support timely and informed countermeasures. Polymorphisms identified in this work offer robust phylogenetic signals that index both short- and long-term evolution and can complement currently employed typing schemes for outbreak ex- and inclusion, diagnostics, surveillance, and forensic studies. PMID:27446025

Sphingomonas japonica sp. nov., isolated from the marine crustacean Paralithodes camtschatica.

PubMed

Romanenko, Lyudmila A; Tanaka, Naoto; Frolova, Galina M; Mikhailov, Valery V

2009-05-01

A Sphingomonas-like bacterium, strain KC7(T), was isolated from a marine crustacean specimen obtained from the Sea of Japan and subjected to a polyphasic study. Comparative 16S rRNA gene sequence analysis positioned the novel strain in the genus Sphingomonas as an independent lineage adjacent to a subclade containing Sphingomonas trueperi LMG 2142(T), Sphingomonas pituitosa EDIV(T) and Sphingomonas azotifigens NBRC 15497(T). Strain KC7(T) shared highest 16S rRNA gene sequence similarity (96.1 %) with S. trueperi LMG 2142(T), Sphingomonas dokdonensis DS-4(T) and S. azotifigens NBRC 15497(T); similarities to strains of other recognized Sphingomonas species were lower (96.0-93.9 %). The strain contained sphingoglycolipid and the predominant fatty acids were C(16 : 1), C(16 : 0) and C(18 : 1); 2-OH C(14 : 0) was the major 2-hydroxy fatty acid. Previously, these lipids have been found to be characteristic of members of the genus Sphingomonas sensu stricto. On the basis of phylogenetic analysis and physiological and biochemical characterization, strain KC7(T) represents a novel species of the genus Sphingomonas, for which the name Sphingomonas japonica sp. nov. is proposed. The type strain is KC7(T) (=KMM 3038(T) =NRIC 0738(T) =JCM 15438(T)).

Comparison of nucleotide sequences of recent and previous lineages of peste-des-petits-ruminants viruses of sheep and goats in Nigeria.

PubMed

Mantip, Samuel; Quan, Melvyn; Shamaki, David; Van Vuuren, Moritz

2016-08-31

Peste-des-petits-ruminants virus (PPRV) is a highly contagious, fatal and economically important viral disease of small ruminants that is still endemic and militates against the production of sheep and goats in endemic areas of the world. The aim of this study was to describe the viral strains within the country. This was carried out by collecting tissue and swab samples from sheep and goats in various agro-ecological zones of Nigeria. The phylogeny of archived PPRV strains or isolates and those circulating and causing recent outbreaks was determined by sequencing of the nucleoprotein (N)-gene. Twenty tissue and swab samples from apparently healthy and sick sheep and goats were collected randomly from 18 states, namely 3 states in each of the 6 agro-ecological zones visited. A total of 360 samples were collected. A total of 35 samples of 360 (9.7%) tested positive by reverse transcriptase-polymerase chain reaction, of which 25 were from oculo-nasal swabs and 10 were from tissue samples. Neighbour-joining phylogenetic analysis using Phylogenetic Analysis Using Parsimony (PAUP) identified four different lineages, that is, lineages I, II, III and IV. Interestingly, the Nigerian strains described in this study grouped in two separate major lineages, that is, lineages II and IV. Strains from Sokoto, Oyo, Plateau and Ondo states grouped according to the historical distribution of PPRV together with the Nigerian 75/1 strain of lineage II, while other strains from Sokoto, Oyo, Plateau, Akwa-Ibom, Adamawa, Kaduna, Lagos, Bauchi, Niger and Kano states grouped together with the East African and Asian strains of lineage IV. This finding confirms that both lineage II and IV strains of PPRV are circulating in Nigeria. Previously, only strains of lineage II were found to be present in the country.

Complete genome sequencing and phylogenetic analysis of dengue type 1 virus isolated from Jeddah, Saudi Arabia.

PubMed

Azhar, Esam I; Hashem, Anwar M; El-Kafrawy, Sherif A; Abol-Ela, Said; Abd-Alla, Adly M M; Sohrab, Sayed Sartaj; Farraj, Suha A; Othman, Norah A; Ben-Helaby, Huda G; Ashshi, Ahmed; Madani, Tariq A; Jamjoom, Ghazi

2015-01-16

Dengue viruses (DENVs) are mosquito-borne viruses which can cause disease ranging from mild fever to severe dengue infection. These viruses are endemic in several tropical and subtropical regions. Multiple outbreaks of DENV serotypes 1, 2 and 3 (DENV-1, DENV-2 and DENV-3) have been reported from the western region in Saudi Arabia since 1994. Strains from at least two genotypes of DENV-1 (Asia and America/Africa genotypes) have been circulating in western Saudi Arabia until 2006. However, all previous studies reported from Saudi Arabia were based on partial sequencing data of the envelope (E) gene without any reports of full genome sequences for any DENV serotypes circulating in Saudi Arabia. Here, we report the isolation and the first complete genome sequence of a DENV-1 strain (DENV-1-Jeddah-1-2011) isolated from a patient from Jeddah, Saudi Arabia in 2011. Whole genome sequence alignment and phylogenetic analysis showed high similarity between DENV-1-Jeddah-1-2011 strain and D1/H/IMTSSA/98/606 isolate (Asian genotype) reported from Djibouti in 1998. Further analysis of the full envelope gene revealed a close relationship between DENV-1-Jeddah-1-2011 strain and isolates reported between 2004-2006 from Jeddah as well as recent isolates from Somalia, suggesting the widespread of the Asian genotype in this region. These data suggest that strains belonging to the Asian genotype might have been introduced into Saudi Arabia long before 2004 most probably by African pilgrims and continued to circulate in western Saudi Arabia at least until 2011. Most importantly, these results indicate that pilgrims from dengue endemic regions can play an important role in the spread of new DENVs in Saudi Arabia and the rest of the world. Therefore, availability of complete genome sequences would serve as a reference for future epidemiological studies of DENV-1 viruses.

A new subtype (subgenotype) Ac (A3) of hepatitis B virus and recombination between genotypes A and E in Cameroon.

PubMed

Kurbanov, Fuat; Tanaka, Yasuhito; Fujiwara, Kei; Sugauchi, Fuminaka; Mbanya, Dora; Zekeng, Leopold; Ndembi, Nicaise; Ngansop, Charlotte; Kaptue, Lazare; Miura, Tomoyuki; Ido, Eiji; Hayami, Masanori; Ichimura, Hiroshi; Mizokami, Masashi

2005-07-01

Blood samples (n=544) from two different populations (Pygmies and Bantus) in Cameroon, West Africa, were analysed. Serological tests indicated that the anti-hepatitis C virus (HCV) prevalence in Bantus (20.3 %) was higher than that in Pygmies (2.3 %, P<0.0001), whereas the distribution of hepatitis B virus (HBV) serological markers was equally high in both populations: in total, 9.4, 17.3 and 86.8 % for HBsAg, anti-HBs and anti-HBc, respectively. HBV genotype A (HBV/A) and HBV/E were predominant (43.5 % each) in both populations, and HBV/D was found in a minority (13 %). The preS/S region was sequenced in nine cases (five HBV/A and four HBV/E) and the complete genome in six cases (four HBV/A and two HBV/E). Subsequent phylogenetic analysis revealed that the HBV/A strains were distinct from the subtypes (subgenotypes) described previously, Ae (A2) and Aa (A1), and in the preS/S region they clustered with previously reported sequences from Cameroon. Based on the nucleotide difference from Aa (A1) and Ae (A2), more than 4 % in the complete genome, the Cameroonian strains were suggested to represent a new subtype (subgenotype), designated HBV/Ac (A3). A high (3.9 %) nucleotide divergence in HBV/Ac (A3) strains suggested that the subtype (subgenotype) has a long natural history in the population of Cameroon. One of the HBV/Ac (A3) strains was found to be a recombinant with an HBV/E-specific sequence in the polymerase reverse transcriptase domain. Further cohort studies will be required to assess detailed epidemiological, virological and clinical characteristics of HBV/Ac (A3), as well as its recombinant form.

Toward a consistent model for strain accrual and release for the New Madrid Seismic Zone, central United States

USGS Publications Warehouse

Hough, S.E.; Page, M.

2011-01-01

At the heart of the conundrum of seismogenesis in the New Madrid Seismic Zone is the apparently substantial discrepancy between low strain rate and high recent seismic moment release. In this study we revisit the magnitudes of the four principal 1811–1812 earthquakes using intensity values determined from individual assessments from four experts. Using these values and the grid search method of Bakun and Wentworth (1997), we estimate magnitudes around 7.0 for all four events, values that are significantly lower than previously published magnitude estimates based on macroseismic intensities. We further show that the strain rate predicted from postglacial rebound is sufficient to produce a sequence with the moment release of one Mmax6.8 every 500 years, a rate that is much lower than previous estimates of late Holocene moment release. However, Mw6.8 is at the low end of the uncertainty range inferred from analysis of intensities for the largest 1811–1812 event. We show that Mw6.8 is also a reasonable value for the largest main shock given a plausible rupture scenario. One can also construct a range of consistent models that permit a somewhat higher Mmax, with a longer average recurrence rate. It is thus possible to reconcile predicted strain and seismic moment release rates with alternative models: one in which 1811–1812 sequences occur every 500 years, with the largest events being Mmax∼6.8, or one in which sequences occur, on average, less frequently, with Mmax of ∼7.0. Both models predict that the late Holocene rate of activity will continue for the next few to 10 thousand years.

Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing

PubMed Central

Tsai, Yu-Chih; Deming, Clayton; Segre, Julia A.; Kong, Heidi H.; Korlach, Jonas

2016-01-01

ABSTRACT Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant “genomes” are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT) sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation. PMID:26861018

Intricate interactions between the bloom-forming cyanobacterium Microcystis aeruginosa and foreign genetic elements, revealed by diversified clustered regularly interspaced short palindromic repeat (CRISPR) signatures.

PubMed

Kuno, Sotaro; Yoshida, Takashi; Kaneko, Takakazu; Sako, Yoshihiko

2012-08-01

Clustered regularly interspaced short palindromic repeats (CRISPR) confer sequence-dependent, adaptive resistance in prokaryotes against viruses and plasmids via incorporation of short sequences, called spacers, derived from foreign genetic elements. CRISPR loci are thus considered to provide records of past infections. To describe the host-parasite (i.e., cyanophages and plasmids) interactions involving the bloom-forming freshwater cyanobacterium Microcystis aeruginosa, we investigated CRISPR in four M. aeruginosa strains and in two previously sequenced genomes. The number of spacers in each locus was larger than the average among prokaryotes. All spacers were strain specific, except for a string of 11 spacers shared in two closely related strains, suggesting diversification of the loci. Using CRISPR repeat-based PCR, 24 CRISPR genotypes were identified in a natural cyanobacterial community. Among 995 unique spacers obtained, only 10 sequences showed similarity to M. aeruginosa phage Ma-LMM01. Of these, six spacers showed only silent or conservative nucleotide mutations compared to Ma-LMM01 sequences, suggesting a strategy by the cyanophage to avert CRISPR immunity dependent on nucleotide identity. These results imply that host-phage interactions can be divided into M. aeruginosa-cyanophage combinations rather than pandemics of population-wide infectious cyanophages. Spacer similarity also showed frequent exposure of M. aeruginosa to small cryptic plasmids that were observed only in a few strains. Thus, the diversification of CRISPR implies that M. aeruginosa has been challenged by diverse communities (almost entirely uncharacterized) of cyanophages and plasmids.

Intricate Interactions between the Bloom-Forming Cyanobacterium Microcystis aeruginosa and Foreign Genetic Elements, Revealed by Diversified Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) Signatures

PubMed Central

Kuno, Sotaro; Kaneko, Takakazu; Sako, Yoshihiko

2012-01-01

Clustered regularly interspaced short palindromic repeats (CRISPR) confer sequence-dependent, adaptive resistance in prokaryotes against viruses and plasmids via incorporation of short sequences, called spacers, derived from foreign genetic elements. CRISPR loci are thus considered to provide records of past infections. To describe the host-parasite (i.e., cyanophages and plasmids) interactions involving the bloom-forming freshwater cyanobacterium Microcystis aeruginosa, we investigated CRISPR in four M. aeruginosa strains and in two previously sequenced genomes. The number of spacers in each locus was larger than the average among prokaryotes. All spacers were strain specific, except for a string of 11 spacers shared in two closely related strains, suggesting diversification of the loci. Using CRISPR repeat-based PCR, 24 CRISPR genotypes were identified in a natural cyanobacterial community. Among 995 unique spacers obtained, only 10 sequences showed similarity to M. aeruginosa phage Ma-LMM01. Of these, six spacers showed only silent or conservative nucleotide mutations compared to Ma-LMM01 sequences, suggesting a strategy by the cyanophage to avert CRISPR immunity dependent on nucleotide identity. These results imply that host-phage interactions can be divided into M. aeruginosa-cyanophage combinations rather than pandemics of population-wide infectious cyanophages. Spacer similarity also showed frequent exposure of M. aeruginosa to small cryptic plasmids that were observed only in a few strains. Thus, the diversification of CRISPR implies that M. aeruginosa has been challenged by diverse communities (almost entirely uncharacterized) of cyanophages and plasmids. PMID:22636003

Genetic Diversity Among Botulinum Neurotoxin Producing Clostridial Strains

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

Hill, K K; Smith, T J; Helma, C H

2006-07-06

Clostridium botulinum is a taxonomic designation for many diverse anaerobic spore forming rod-shaped bacteria which have the common property of producing botulinum neurotoxins (BoNTs). The BoNTs are exoneurotoxins that can cause severe paralysis and even death in humans and various other animal species. A collection of 174 C. botulinum strains were examined by amplified fragment length polymorphism (AFLP) analysis and by sequencing of the 16S rRNA gene and BoNT genes to examine genetic diversity within this species. This collection contained representatives of each of the seven different serotypes of botulinum neurotoxins (BoNT A-G). Analysis of the16S rRNA sequences confirmed earliermore » reports of at least four distinct genomic backgrounds (Groups I-IV) each of which has independently acquired one or more BoNT serotypes through horizontal gene transfer. AFLP analysis provided higher resolution, and can be used to further subdivide the four groups into sub-groups. Sequencing of the BoNT genes from serotypes A, B and E in multiple strains confirmed significant sequence variation within each serotype. Four distinct lineages within each of the BoNT A and B serotypes, and five distinct lineages of serotype E strains were identified. The nucleotide sequences of the seven serotypes of BoNT were compared and show varying degrees of interrelatedness and recombination as has been previously noted for the NTNH gene which is linked to BoNT. These analyses contribute to the understanding of the evolution and phylogeny within this species and assist in the development of improved diagnostics and therapeutics for treatment of botulism.« less

Functional Role of Infective Viral Particles on Metal Reduction

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

Coates, John D.

2014-04-01

A proposed strategy for the remediation of uranium (U) contaminated sites was based on the immobilization of U by reducing the oxidized soluble U, U(VI), to form a reduced insoluble end product, U(IV). Previous studies identified Geobacter sp., including G. sulfurreducens and G. metallireducens, as predominant U(VI)-reducing bacteria under acetate-oxidizing and U(VI)-reducing conditions. Examination of the finished genome sequence annotation of the canonical metal reducing species Geobacter sulfurreducens strain PCA and G. metallireduceans strain GS-15 as well as the draft genome sequence of G. uraniumreducens strain Rf4 identified phage related proteins. In addition, the completed genome for Anaeromyxobacter dehalogenans andmore » the draft genome sequence of Desulfovibrio desulfuricans strain G20, two more model metal-reducing bacteria, also revealed phage related sequences. The presence of these gene sequences indicated that Geobacter spp., Anaeromyxobacter spp., and Desulfovibrio spp. are susceptible to viral infection. Furthermore, viral populations in soils and sedimentary environments in the order of 6.4×10{sup 6}–2.7×10{sup 10} VLP’s cm{sup -3} have been observed. In some cases, viral populations exceed bacterial populations in these environments suggesting that a relationship may exist between viruses and bacteria. Our preliminary screens of samples collected from the ESR FRC indicated that viral like particles were observed in significant numbers. The objective of this study was to investigate the potential functional role viruses play in metal reduction specifically Fe(III) and U(VI) reduction, the environmental parameters affecting viral infection of metal reducing bacteria, and the subsequent effects on U transport.« less

SnSAG5 is an alternative surface antigen of Sarcocystis neurona strains that is mutually exclusive to SnSAG1.

PubMed

Crowdus, Carolyn A; Marsh, Antoinette E; Saville, Willliam J; Lindsay, David S; Dubey, J P; Granstrom, David E; Howe, Daniel K

2008-11-25

Sarcocystis neurona is an obligate intracellular parasite that causes equine protozoal myeloencephalitis (EPM). Previous work has identified a gene family of paralogous surface antigens in S. neurona called SnSAGs. These surface proteins are immunogenic in their host animals, and are therefore candidate molecules for development of diagnostics and vaccines. However, SnSAG diversity exists in strains of S. neurona, including the absence of the major surface antigen gene SnSAG1. Instead, sequence for an alternative SnSAG has been revealed in two of the SnSAG1-deficient strains. Herein, we present data characterizing this new surface protein, which we have designated SnSAG5. The results indicated that the protein encoded by the SnSAG5 sequence is indeed a surface-associated molecule that has characteristics consistent with the other SAGs identified in S. neurona and related parasites. Importantly, Western blot analyses of a collection of S. neurona strains demonstrated that 6 of 13 parasite isolates express SnSAG5 as a dominant surface protein instead of SnSAG1. Conversely, SnSAG5 was not detected in SnSAG1-positive strains. One strain, which was isolated from the brain of a sea otter, did not express either SnSAG1 or SnSAG5. Genetic analysis with SnSAG5-specific primers confirmed the presence of the SnSAG5 gene in Western blot-positive strains, while also suggesting the presence of a novel SnSAG sequence in the SnSAG1-deficient, SnSAG5-deficient otter isolate. The findings provide further indication of S. neurona strain diversity, which has implications for diagnostic testing and development of vaccines against EPM as well as the population biology of Sarcocystis cycling in the opossum definitive host.

Genomic and Transcriptomic Analysis of Escherichia coli Strains Associated with Persistent and Transient Bovine Mastitis and the Role of Colanic Acid.

PubMed

Lippolis, John D; Holman, Devin B; Brunelle, Brian W; Thacker, Tyler C; Bearson, Bradley L; Reinhardt, Timothy A; Sacco, Randy E; Casey, Thomas A

2018-01-01

Escherichia coli is a leading cause of bacterial mastitis in dairy cattle. It is most often transient in nature, causing an infection that lasts 2 to 3 days. However, E. coli has been shown to cause a persistent infection in a minority of cases. Mechanisms that allow for a persistent E. coli infection are not fully understood. The goal of this work was to determine differences between E. coli strains originally isolated from dairy cattle with transient and persistent mastitis. Using RNA sequencing, we show gene expression differences in nearly 200 genes when bacteria from the two clinical phenotypes are compared. We sequenced the genomes of the E. coli strains and report genes unique to the two phenotypes. Differences in the wca operon, which encodes colanic acid, were identified by DNA as well as RNA sequencing and differentiated the two phenotypes. Previous work demonstrated that E. coli strains that cause persistent infections were more motile than those that cause transient infections. Deletion of genes in the wca operon from a persistent-infection strain resulted in a reduction of motility as measured in swimming and swarming assays. Furthermore, colanic acid has been shown to protect bacteria from complement-mediated killing. We show that transient-infection E. coli strains were more sensitive to complement-mediated killing. The deletion of genes from the wca operon caused a persistent-infection E. coli strain to become sensitive to complement-mediated killing. This work identifies important differences between E. coli strains that cause persistent and transient mammary infections in dairy cattle. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

Description of Immundisolibacter cernigliae gen. nov., sp. nov., a high-molecular-weight polycyclic aromatic hydrocarbon-degrading bacterium within the class Gammaproteobacteria, and proposal of Immundisolibacterales ord. nov. and Immundisolibacteraceae fam. nov.

PubMed

Corteselli, Elizabeth M; Aitken, Michael D; Singleton, David R

2017-04-01

The bacterial strain TR3.2T was isolated from aerobic bioreactor-treated soil from a polycyclic aromatic hydrocarbon (PAH)-contaminated site in Salisbury, NC, USA. Strain TR3.2T was identified as a member of 'Pyrene Group 2' or 'PG2', a previously uncultivated cluster of organisms associated with the degradation of high-molecular-weight PAHs by stable-isotope probing. Based on its 16S rRNA gene sequence, the strain was classified as a member of the class Gammaproteobacteria but possessed only 90.5 % gene identity to its closest described relative, Methylococcus capsulatus strain Bath. Strain TR3.2T grew on the PAHs pyrene, phenanthrene, anthracene, benz[a]anthracene and fluorene, as well as the azaarene carbazole, and could additionally metabolize a limited number of organic acids. Optimal growth occurred aerobically under mesophilic temperature, neutral pH and low salinity conditions. Strain TR3.2T was catalase and oxidase positive. Predominant fatty acids were C17 : 0 cyclo and C16 : 0. Genomic G+C content of the single chromosome was 67.79 mol% as determined by complete genome sequencing. Due to the high sequence divergence from any cultivated species and its unique physiological properties compared to its closest relatives, strain TR3.2T is proposed as a representative of a novel order, family, genus and species within the class Gammaproteobacteria, for which the name Immundisolibacter cernigliae gen. nov., sp. nov. is proposed. The associated order and family are therefore proposed as Immundisolibacteralesord. nov. and Immundisolibacteraceaefam. nov. The type strain of the species is TR3.2T (=ATCC TSD-58T=DSM 103040T).

Natural non-homologous recombination led to the emergence of a duplicated V3-NS5A region in HCV-1b strains associated with hepatocellular carcinoma.

PubMed

Le Guillou-Guillemette, Hélène; Pivert, Adeline; Bouthry, Elise; Henquell, Cécile; Petsaris, Odile; Ducancelle, Alexandra; Veillon, Pascal; Vallet, Sophie; Alain, Sophie; Thibault, Vincent; Abravanel, Florence; Rosenberg, Arielle A; André-Garnier, Elisabeth; Bour, Jean-Baptiste; Baazia, Yazid; Trimoulet, Pascale; André, Patrice; Gaudy-Graffin, Catherine; Bettinger, Dominique; Larrat, Sylvie; Signori-Schmuck, Anne; Saoudin, Hénia; Pozzetto, Bruno; Lagathu, Gisèle; Minjolle-Cha, Sophie; Stoll-Keller, Françoise; Pawlotsky, Jean-Michel; Izopet, Jacques; Payan, Christopher; Lunel-Fabiani, Françoise; Lemaire, Christophe

2017-01-01

The emergence of new strains in RNA viruses is mainly due to mutations or intra and inter-genotype homologous recombination. Non-homologous recombinations may be deleterious and are rarely detected. In previous studies, we identified HCV-1b strains bearing two tandemly repeated V3 regions in the NS5A gene without ORF disruption. This polymorphism may be associated with an unfavorable course of liver disease and possibly involved in liver carcinogenesis. Here we aimed at characterizing the origin of these mutant strains and identifying the evolutionary mechanism on which the V3 duplication relies. Direct sequencing of the entire NS5A and E1 genes was performed on 27 mutant strains. Quasispecies analyses in consecutive samples were also performed by cloning and sequencing the NS5A gene for all mutant and wild strains. We analyzed the mutant and wild-type sequence polymorphisms using Bayesian methods to infer the evolutionary history of and the molecular mechanism leading to the duplication-like event. Quasispecies were entirely composed of exclusively mutant or wild-type strains respectively. Mutant quasispecies were found to have been present since contamination and had persisted for at least 10 years. This V3 duplication-like event appears to have resulted from non-homologous recombination between HCV-1b wild-type strains around 100 years ago. The association between increased liver disease severity and these HCV-1b mutants may explain their persistence in chronically infected patients. These results emphasize the possible consequences of non-homologous recombination in the emergence and severity of new viral diseases.

Complete genome sequence of Metallosphaera cuprina, a metal sulfide-oxidizing archaeon from a hot spring.

PubMed

Liu, Li-Jun; You, Xiao-Yan; Zheng, Huajun; Wang, Shengyue; Jiang, Cheng-Ying; Liu, Shuang-Jiang

2011-07-01

The genome of the metal sulfide-oxidizing, thermoacidophilic strain Metallosphaera cuprina Ar-4 has been completely sequenced and annotated. Originally isolated from a sulfuric hot spring, strain Ar-4 grows optimally at 65°C and a pH of 3.5. The M. cuprina genome has a 1,840,348-bp circular chromosome (2,029 open reading frames [ORFs]) and is 16% smaller than the previously sequenced Metallosphaera sedula genome. Compared to the M. sedula genome, there are no counterpart genes in the M. cuprina genome for about 480 ORFs in the M. sedula genome, of which 243 ORFs are annotated as hypothetical protein genes. Still, there are 233 ORFs uniquely occurring in M. cuprina. Genome annotation supports that M. cuprina lives a facultative life on CO(2) and organics and obtains energy from oxidation of sulfidic ores and reduced inorganic sulfuric compounds.

Analysis of the VP2 protein gene of canine parvovirus strains from affected dogs in Japan.

PubMed

Soma, Takehisa; Taharaguchi, Satoshi; Ohinata, Tsuyoshi; Ishii, Hiroshi; Hara, Motonobu

2013-04-01

To clarify the evolution of canine parvovirus 2 (CPV-2) that has recently been epidemic in Japan, VP2 gene sequences at positions 3556-4166 were analyzed in 107 CPV-2 strains obtained from rectal swabs of diarrheic dogs from 2009 to 2011. CPV-2b (95 strains) was more frequently detected than CPV-2a (nine strains), while CPV-2c was not detected. Remaining three strains were identified as the original type CPV-2, which should be derived from vaccines. These findings are similar to the previous results involving Japanese strains, suggesting there has been no great change in the recent CPV-2 epidemic in Japan. This epidemic is the same as that in Taiwan. Furthermore, a 324-lle mutant, which has been reported in Korean and Chinese strains, was detected in 66.7% of CPV-2a strains. Copyright © 2012 Elsevier Ltd. All rights reserved.

Characterization of virulent West Nile virus Kunjin strain, Australia, 2011.

PubMed

Frost, Melinda J; Zhang, Jing; Edmonds, Judith H; Prow, Natalie A; Gu, Xingnian; Davis, Rodney; Hornitzky, Christine; Arzey, Kathleen E; Finlaison, Deborah; Hick, Paul; Read, Andrew; Hobson-Peters, Jody; May, Fiona J; Doggett, Stephen L; Haniotis, John; Russell, Richard C; Hall, Roy A; Khromykh, Alexander A; Kirkland, Peter D

2012-05-01

To determine the cause of an unprecedented outbreak of encephalitis among horses in New South Wales, Australia, in 2011, we performed genomic sequencing of viruses isolated from affected horses and mosquitoes. Results showed that most of the cases were caused by a variant West Nile virus (WNV) strain, WNV(NSW2011), that is most closely related to WNV Kunjin (WNV(KUN)), the indigenous WNV strain in Australia. Studies in mouse models for WNV pathogenesis showed that WNV(NSW2011) is substantially more neuroinvasive than the prototype WNV(KUN) strain. In WNV(NSW2011), this apparent increase in virulence over that of the prototype strain correlated with at least 2 known markers of WNV virulence that are not found in WNV(KUN). Additional studies are needed to determine the relationship of the WNV(NSW2011) strain to currently and previously circulating WNV(KUN) strains and to confirm the cause of the increased virulence of this emerging WNV strain.

Whole genome phylogenetic investigation of a West Nile virus strain isolated from a tick sampled from livestock in north eastern Kenya.

PubMed

Lwande, Olivia Wesula; Venter, Marietjie; Lutomiah, Joel; Michuki, George; Rumberia, Cecilia; Gakuya, Francis; Obanda, Vincent; Tigoi, Caroline; Odhiambo, Collins; Nindo, Fredrick; Symekher, Samwel; Sang, Rosemary

2014-11-28

West Nile virus (WNV) has a wide geographical distribution and has been associated to cause neurological disease in humans and horses. Mosquitoes are the traditional vectors for WNV; however, the virus has also been isolated from tick species in North Africa and Europe which could be a means of introduction and spread of the virus over long distances through migratory birds. Although WNV has been isolated in mosquitoes in Kenya, paucity of genetic and pathogenicity data exists. We previously reported the isolation of WNV from ticks collected from livestock and wildlife in Ijara District of Kenya, a hotspot for arbovirus activity. Here we report the full genome sequence and phylogenetic investigation of their origin and relation to strains from other regions. A total of 10,488 ticks were sampled from animal hosts, classified to species and processed in pools of up to eight ticks per pool. Virus screening was performed by cell culture, RT-PCR and sequencing. Phylogenetic analysis was carried out to determine the evolutionary relationships of our isolate. Among other viruses, WNV was isolated from a pool of Rhipicephalus pulchellus sampled from cattle, sequenced and submitted to GenBank (Accession number: KC243146). Comparative analysis with 27 different strains revealed that our isolate belongs to lineage 1 and clustered relatively closely to isolates from North Africa and Europe, Russia and the United States. Overall, Bayesian analysis based on nucleotide sequences showed that lineage 1 strains including the Kenyan strain had diverged 200 years ago from lineage 2 strains of southern Africa. Ijara strain collected from a tick sampled on livestock was closest to another Kenyan strain and had diverged 20 years ago from strains detected in Morocco and Europe and 30 years ago from strains identified in the USA. To our knowledge, this is the first characterized WNV strain isolated from R. pulchellus. The epidemiological role of this tick in WNV transmission and dissemination remains equivocal but presents tick verses mosquito virus transmission has been neglected. Genetic data of this strain suggest that lineage 1 strains from Africa could be dispersed through tick vectors by wild migratory birds to Europe and beyond.

Persistent natural infection of a Culex tritaeniorhynchus cell line with a novel Culex tritaeniorhynchus rhabdovirus strain.

PubMed

Gillich, Nadine; Kuwata, Ryusei; Isawa, Haruhiko; Horie, Masayuki

2015-09-01

Culex tritaeniorhynchus rhabdovirus (CTRV) is a mosquito virus that establishes persistent infection without any obvious cell death. Therefore, occult infection by CTRV can be present in mosquito cell lines. In this study, it is shown that NIID-CTR cells, which were derived from Cx. tritaeniorhynchus, are persistently infected with a novel strain of CTRV. Complete genome sequencing of the infecting strain revealed that it is genetically similar but distinct from the previously isolated CTRV strain, excluding the possibility of contamination. These findings raise the importance of further CTRV studies, such as screening of CTRV in other mosquito cell lines. © 2015 The Societies and Wiley Publishing Asia Pty Ltd.

Livestock rabies outbreaks in Shanxi province, China.

PubMed

Feng, Ye; Shi, Yanyan; Yu, Mingyang; Xu, Weidi; Gong, Wenjie; Tu, Zhongzhong; Ding, Laixi; He, Biao; Guo, Huancheng; Tu, Changchun

2016-10-01

Dogs play an important role in rabies transmission throughout the world. In addition to the severe human rabies situation in China, spillover of rabies virus from dogs in recent years has caused rabies outbreaks in sheep, cattle and pigs, showing that there is an increasing threat to other domestic animals. Two livestock rabies outbreaks were caused by dogs in Shanxi province, China from April to October in 2015, resulting in the deaths of 60 sheep, 10 cattle and one donkey. Brain samples from one infected bovine and the donkey were determined to be rabies virus (RABV) positive by fluorescent antibody test (FAT) and reverse transcription polymerase chain reaction (RT-PCR). The complete RABV N genes of the two field strains, together with those of two previously confirmed Shanxi dog strains, were amplified, sequenced and compared phylogenetically with published sequences of the N gene of RABV strains from Shanxi and surrounding provinces. All of the strains from Shanxi province grouped closely, sharing 99.6 %-100 % sequence identity, indicating the wide distribution and transmission of dog-mediated rabies in these areas. This is the first description of donkey rabies symptoms with phylogenetic analysis of RABVs in Shanxi province and surrounding regions. The result emphasizes the need for mandatory dog rabies vaccination and improved public education to eradicate dog rabies transmission.

PathogenFinder--distinguishing friend from foe using bacterial whole genome sequence data.

PubMed

Cosentino, Salvatore; Voldby Larsen, Mette; Møller Aarestrup, Frank; Lund, Ole

2013-01-01

Although the majority of bacteria are harmless or even beneficial to their host, others are highly virulent and can cause serious diseases, and even death. Due to the constantly decreasing cost of high-throughput sequencing there are now many completely sequenced genomes available from both human pathogenic and innocuous strains. The data can be used to identify gene families that correlate with pathogenicity and to develop tools to predict the pathogenicity of newly sequenced strains, investigations that previously were mainly done by means of more expensive and time consuming experimental approaches. We describe PathogenFinder (http://cge.cbs.dtu.dk/services/PathogenFinder/), a web-server for the prediction of bacterial pathogenicity by analysing the input proteome, genome, or raw reads provided by the user. The method relies on groups of proteins, created without regard to their annotated function or known involvement in pathogenicity. The method has been built to work with all taxonomic groups of bacteria and using the entire training-set, achieved an accuracy of 88.6% on an independent test-set, by correctly classifying 398 out of 449 completely sequenced bacteria. The approach here proposed is not biased on sets of genes known to be associated with pathogenicity, thus the approach could aid the discovery of novel pathogenicity factors. Furthermore the pathogenicity prediction web-server could be used to isolate the potential pathogenic features of both known and unknown strains.

Genome sequence of the pink to light reddish-pigmented Rubellimicrobium mesophilum type strain (DSM 19309(T)), a representative of the Roseobacter group isolated from soil, and emended description of the species.

PubMed

Riedel, Thomas; Spring, Stefan; Fiebig, Anne; Petersen, Jörn; Göker, Markus; Klenk, Hans-Peter

2014-06-15

Rubellimicrobium mesophilum Dastager et al. 2008 is a mesophilic and light reddish-pigmented representative of the Roseobacter group within the alphaproteobacterial family Rhodobacteraceae. Representatives of the Roseobacter group play an important role in the marine biogeochemical cycles and were found in a broad variety of marine environments associated with algal blooms, different kinds of sediments, and surfaces of invertebrates and vertebrates. Roseobacters were shown to be widely distributed, especially within the total bacterial community found in coastal waters, as well as in mixed water layers of the open ocean. Here we describe the features of R. mesophilum strain MSL-20(T) together with its genome sequence and annotation generated from a culture of DSM 19309(T). The 4,927,676 bp genome sequence consists of one chromosome and probably one extrachromosomal element. It contains 5,082 protein-coding genes and 56 RNA genes. As previously reported, the G+C content is significantly different from the actual genome sequence-based G+C content and as the type strain tests positively for oxidase, the species description is emended accordingly. The genome was sequenced as part of the activities of the Transregional Collaborative Research Centre 51 (TRR51) funded by the German Research Foundation (DFG).

The genomic sequence of ectromelia virus, the causative agent of mousepox.

PubMed

Chen, Nanhai; Danila, Maria I; Feng, Zehua; Buller, R Mark L; Wang, Chunlin; Han, Xiaosi; Lefkowitz, Elliot J; Upton, Chris

2003-12-05

Ectromelia virus is the causative agent of mousepox, an acute exanthematous disease of mouse colonies in Europe, Japan, China, and the U.S. The Moscow, Hampstead, and NIH79 strains are the most thoroughly studied with the Moscow strain being the most infectious and virulent for the mouse. In the late 1940s mousepox was proposed as a model for the study of the pathogenesis of smallpox and generalized vaccinia in humans. Studies in the last five decades from a succession of investigators have resulted in a detailed description of the virologic and pathologic disease course in genetically susceptible and resistant inbred and out-bred mice. We report the DNA sequence of the left-hand end, the predicted right-hand terminal repeat, and central regions of the genome of the Moscow strain of ectromelia virus (approximately 177,500 bp), which together with the previously sequenced right-hand end, yields a genome of 209,771 bp. We identified 175 potential genes specifying proteins of between 53 and 1924 amino acids, and 29 regions containing sequences related to genes predicted in other poxviruses, but unlikely to encode for functional proteins in ectromelia virus. The translated protein sequences were compared with the protein database for structure/function relationships, and these analyses were used to investigate poxvirus evolution and to attempt to explain at the cellular and molecular level the well-characterized features of the ectromelia virus natural life cycle.

Genomic characterization of a Helicobacter pylori isolate from a patient with gastric cancer in China

PubMed Central

2014-01-01

Background Helicobacter pylori is well known for its relationship with the occurrence of several severe gastric diseases. The mechanisms of pathogenesis triggered by H. pylori are less well known. In this study, we report the genome sequence and genomic characterizations of H. pylori strain HLJ039 that was isolated from a patient with gastric cancer in the Chinese province of Heilongjiang, where there is a high incidence of gastric cancer. To investigate potential genomic features that may be involved in pathogenesis of carcinoma, the genome was compared to three previously sequenced genomes in this area. Result We obtained 42 contigs with a total length of 1,611,192 bp and predicted 1,687 coding sequences. Compared to strains isolated from gastritis and ulcers in this area, 10 different regions were identified as being unique for HLJ039; they mainly encoded type II restriction-modification enzyme, type II m6A methylase, DNA-cytosine methyltransferase, DNA methylase, and hypothetical proteins. A unique 547-bp fragment sharing 93% identity with a hypothetical protein of Helicobacter cinaedi ATCC BAA-847 was not present in any other previous H. pylori strains. Phylogenetic analysis based on core genome single nucleotide polymorphisms shows that HLJ039 is defined as hspEAsia subgroup, which belongs to the hpEastAsia group. Conclusion DNA methylations, variations of the genomic regions involved in restriction and modification systems, are the “hot” regions that may be related to the mechanism of H. pylori-induced gastric cancer. The genome sequence will provide useful information for the deep mining of potential mechanisms related to East Asian gastric cancer. PMID:24565107

Molecular diversity, cultivation, and improved detection by fluorescent in situ hybridization of a dominant group of human gut bacteria related to Roseburia spp. or Eubacterium rectale.

PubMed

Aminov, Rustam I; Walker, Alan W; Duncan, Sylvia H; Harmsen, Hermie J M; Welling, Gjalt W; Flint, Harry J

2006-09-01

Phylogenetic analysis was used to compare 16S rRNA sequences from 19 cultured human gut strains of Roseburia and Eubacterium rectale with 356 related sequences derived from clone libraries. The cultured strains were found to represent five of the six phylotypes identified. A new oligonucleotide probe, Rrec584, and the previous group probe Rint623, when used in conjunction with a new helper oligonucleotide, each recognized an average of 7% of bacteria detected by the eubacterial probe Eub338 in feces from 10 healthy volunteers. Most of the diversity within this important group of butyrate-producing gut bacteria can apparently be retrieved through cultivation.

Molecular Diversity, Cultivation, and Improved Detection by Fluorescent In Situ Hybridization of a Dominant Group of Human Gut Bacteria Related to Roseburia spp. or Eubacterium rectale

PubMed Central

Aminov, Rustam I.; Walker, Alan W.; Duncan, Sylvia H.; Harmsen, Hermie J. M.; Welling, Gjalt W.; Flint, Harry J.

2006-01-01

Phylogenetic analysis was used to compare 16S rRNA sequences from 19 cultured human gut strains of Roseburia and Eubacterium rectale with 356 related sequences derived from clone libraries. The cultured strains were found to represent five of the six phylotypes identified. A new oligonucleotide probe, Rrec584, and the previous group probe Rint623, when used in conjunction with a new helper oligonucleotide, each recognized an average of 7% of bacteria detected by the eubacterial probe Eub338 in feces from 10 healthy volunteers. Most of the diversity within this important group of butyrate-producing gut bacteria can apparently be retrieved through cultivation. PMID:16957265

Phylogeny and S1 Gene Variation of Infectious Bronchitis Virus Detected in Broilers and Layers in Turkey.

PubMed

Yilmaz, Huseyin; Altan, Eda; Cizmecigil, Utku Y; Gurel, Aydin; Ozturk, Gulay Yuzbasioglu; Bamac, Ozge Erdogan; Aydin, Ozge; Britton, Paul; Monne, Isabella; Cetinkaya, Burhan; Morgan, Kenton L; Faburay, Bonto; Richt, Juergen A; Turan, Nuri

2016-09-01

The avian coronavirus infectious bronchitis virus (AvCoV-IBV) is recognized as an important global pathogen because new variants are a continuous threat to the poultry industry worldwide. This study investigates the genetic origin and diversity of AvCoV-IBV by analysis of the S1 sequence derived from 49 broiler flocks and 14 layer flocks in different regions of Turkey. AvCoV-IBV RNA was detected in 41 (83.6%) broiler flocks and nine (64.2%) of the layer flocks by TaqMan real-time RT-PCR. In addition, AvCoV-IBV RNA was detected in the tracheas 27/30 (90%), lungs 31/49 (62.2%), caecal tonsils 7/22 (31.8%), and kidneys 4/49 (8.1%) of broiler flocks examined. Pathologic lesions, hemorrhages, and mononuclear infiltrations were predominantly observed in tracheas and to a lesser extent in the lungs and a few in kidneys. A phylogenetic tree based on partial S1 sequences of the detected AvCoV-IBVs (including isolates) revealed that 1) viruses detected in five broiler flocks were similar to the IBV vaccines Ma5, H120, M41; 2) viruses detected in 24 broiler flocks were similar to those previously reported from Turkey and to Israel variant-2 strains; 3) viruses detected in seven layer flocks were different from those found in any of the broiler flocks but similar to viruses previously reported from Iran, India, and China (similar to Israel variant-1 and 4/91 serotypes); and 4) that the AVCoV-IBV, Israeli variant-2 strain, found to be circulating in Turkey appears to be undergoing molecular evolution. In conclusion, genetically different AvCoV-IBV strains, including vaccine-like strains, based on their partial S1 sequence, are circulating in broiler and layer chicken flocks in Turkey and the Israeli variant-2 strain is undergoing evolution.

Complete genome sequence and the expression pattern of plasmids of the model ethanologen Zymomonas mobilis ZM4 and its xylose-utilizing derivatives 8b and 2032.

PubMed

Yang, Shihui; Vera, Jessica M; Grass, Jeff; Savvakis, Giannis; Moskvin, Oleg V; Yang, Yongfu; McIlwain, Sean J; Lyu, Yucai; Zinonos, Irene; Hebert, Alexander S; Coon, Joshua J; Bates, Donna M; Sato, Trey K; Brown, Steven D; Himmel, Michael E; Zhang, Min; Landick, Robert; Pappas, Katherine M; Zhang, Yaoping

2018-01-01

Zymomonas mobilis is a natural ethanologen being developed and deployed as an industrial biofuel producer. To date, eight Z. mobilis strains have been completely sequenced and found to contain 2-8 native plasmids. However, systematic verification of predicted Z. mobilis plasmid genes and their contribution to cell fitness has not been hitherto addressed. Moreover, the precise number and identities of plasmids in Z. mobilis model strain ZM4 have been unclear. The lack of functional information about plasmid genes in ZM4 impedes ongoing studies for this model biofuel-producing strain. In this study, we determined the complete chromosome and plasmid sequences of ZM4 and its engineered xylose-utilizing derivatives 2032 and 8b. Compared to previously published and revised ZM4 chromosome sequences, the ZM4 chromosome sequence reported here contains 65 nucleotide sequence variations as well as a 2400-bp insertion. Four plasmids were identified in all three strains, with 150 plasmid genes predicted in strain ZM4 and 2032, and 153 plasmid genes predicted in strain 8b due to the insertion of heterologous DNA for expanded substrate utilization. Plasmid genes were then annotated using Blast2GO, InterProScan, and systems biology data analyses, and most genes were found to have apparent orthologs in other organisms or identifiable conserved domains. To verify plasmid gene prediction, RNA-Seq was used to map transcripts and also compare relative gene expression under various growth conditions, including anaerobic and aerobic conditions, or growth in different concentrations of biomass hydrolysates. Overall, plasmid genes were more responsive to varying hydrolysate concentrations than to oxygen availability. Additionally, our results indicated that although all plasmids were present in low copy number (about 1-2 per cell), the copy number of some plasmids varied under specific growth conditions or due to heterologous gene insertion. The complete genome of ZM4 and two xylose-utilizing derivatives is reported in this study, with an emphasis on identifying and characterizing plasmid genes. Plasmid gene annotation, validation, expression levels at growth conditions of interest, and contribution to host fitness are reported for the first time.

Isolation and distribution of a novel iron-oxidizing crenarchaeon from acidic geothermal springs in Yellowstone National Park.

PubMed

Kozubal, M; Macur, R E; Korf, S; Taylor, W P; Ackerman, G G; Nagy, A; Inskeep, W P

2008-02-01

Novel thermophilic crenarchaea have been observed in Fe(III) oxide microbial mats of Yellowstone National Park (YNP); however, no definitive work has identified specific microorganisms responsible for the oxidation of Fe(II). The objectives of the current study were to isolate and characterize an Fe(II)-oxidizing member of the Sulfolobales observed in previous 16S rRNA gene surveys and to determine the abundance and distribution of close relatives of this organism in acidic geothermal springs containing high concentrations of dissolved Fe(II). Here we report the isolation and characterization of the novel, Fe(II)-oxidizing, thermophilic, acidophilic organism Metallosphaera sp. strain MK1 obtained from a well-characterized acid-sulfate-chloride geothermal spring in Norris Geyser Basin, YNP. Full-length 16S rRNA gene sequence analysis revealed that strain MK1 exhibits only 94.9 to 96.1% sequence similarity to other known Metallosphaera spp. and less than 89.1% similarity to known Sulfolobus spp. Strain MK1 is a facultative chemolithoautotroph with an optimum pH range of 2.0 to 3.0 and an optimum temperature range of 65 to 75 degrees C. Strain MK1 grows optimally on pyrite or Fe(II) sorbed onto ferrihydrite, exhibiting doubling times between 10 and 11 h under aerobic conditions (65 degrees C). The distribution and relative abundance of MK1-like 16S rRNA gene sequences in 14 acidic geothermal springs containing Fe(III) oxide microbial mats were evaluated. Highly related MK1-like 16S rRNA gene sequences (>99% sequence similarity) were consistently observed in Fe(III) oxide mats at temperatures ranging from 55 to 80 degrees C. Quantitative PCR using Metallosphaera-specific primers confirmed that organisms highly similar to strain MK1 comprised up to 40% of the total archaeal community at selected sites. The broad distribution of highly related MK1-like 16S rRNA gene sequences in acidic Fe(III) oxide microbial mats is consistent with the observed characteristics and growth optima of Metallosphaera-like strain MK1 and emphasizes the importance of this newly described taxon in Fe(II) chemolithotrophy in acidic high-temperature environments of YNP.

Mitogenic effect contributes to increased virulence of Streptococcus suis sequence type 7 to cause streptococcal toxic shock-like syndrome.

PubMed

Zheng, H; Ye, C; Segura, M; Gottschalk, M; Xu, J

2008-09-01

Streptococcus suis serotype 2 sequence type 7 strains emerged in 1996 and caused a streptococcal toxic shock-like syndrome in 1998 and 2005 in China. Evidence indicated that the virulence of S. suis sequence type 7 had increased, but the mechanism was unknown. The sequence type 7 strain SC84, isolated from a patient with streptococcal toxic shock-like syndrome during the Sichuan outbreak, and the sequence type 1 strain 31533, a typical highly pathogenic strain isolated from a diseased pig, were used in comparative studies. In this study we show the mechanisms underlying cytokine production differed between the two types of strains. The S. suis sequence type 7 strain SC84 possesses a stronger capacity to stimulate T cells, naive T cells and peripheral blood mononuclear cell proliferation than does S. suis sequence type 1 strain 31533. The T cell response to both strains was dependent upon the presence of antigen-presenting cells. Histo-incompatible antigen-presenting cells were sufficient to provide the accessory signals to naive T cell stimulated by the two strains, indicating that both sequence type 7 and 1 strains possess mitogens; however, the mitogenic effect was different. Therefore, we propose that the difference in the mitogenic effect of sequence type 7 strain SC84 compared with the sequence type 1 strain 31533 of S. suis may be associated with the clinical, epidemiological and microbiological difference, where the ST 7 strains have a larger mitogenic effect.

Mitogenic effect contributes to increased virulence of Streptococcus suis sequence type 7 to cause streptococcal toxic shock-like syndrome

PubMed Central

Zheng, H; Ye, C; Segura, M; Gottschalk, M; Xu, J

2008-01-01

Streptococcus suis serotype 2 sequence type 7 strains emerged in 1996 and caused a streptococcal toxic shock-like syndrome in 1998 and 2005 in China. Evidence indicated that the virulence of S. suis sequence type 7 had increased, but the mechanism was unknown. The sequence type 7 strain SC84, isolated from a patient with streptococcal toxic shock-like syndrome during the Sichuan outbreak, and the sequence type 1 strain 31533, a typical highly pathogenic strain isolated from a diseased pig, were used in comparative studies. In this study we show the mechanisms underlying cytokine production differed between the two types of strains. The S. suis sequence type 7 strain SC84 possesses a stronger capacity to stimulate T cells, naive T cells and peripheral blood mononuclear cell proliferation than does S. suis sequence type 1 strain 31533. The T cell response to both strains was dependent upon the presence of antigen-presenting cells. Histo-incompatible antigen-presenting cells were sufficient to provide the accessory signals to naive T cell stimulated by the two strains, indicating that both sequence type 7 and 1 strains possess mitogens; however, the mitogenic effect was different. Therefore, we propose that the difference in the mitogenic effect of sequence type 7 strain SC84 compared with the sequence type 1 strain 31533 of S. suis may be associated with the clinical, epidemiological and microbiological difference, where the ST 7 strains have a larger mitogenic effect. PMID:18803762

Comparative genomics provides new insights into the diversity, physiology, and sexuality of the only industrially exploited tremellomycete: Phaffia rhodozyma.

PubMed

Bellora, Nicolás; Moliné, Martín; David-Palma, Márcia; Coelho, Marco A; Hittinger, Chris Todd; Sampaio, José P; Gonçalves, Paula; Libkind, Diego

2016-11-09

The class Tremellomycete (Agaricomycotina) encompasses more than 380 fungi. Although there are a few edible Tremella spp., the only species with current biotechnological use is the astaxanthin-producing yeast Phaffia rhodozyma (Cystofilobasidiales). Besides astaxanthin, a carotenoid pigment with potent antioxidant activity and great value for aquaculture and pharmaceutical industries, P. rhodozyma possesses multiple exceptional traits of fundamental and applied interest. The aim of this study was to obtain, and analyze two new genome sequences of representative strains from the northern (CBS 7918 T , the type strain) and southern hemispheres (CRUB 1149) and compre them to a previously published genome sequence (strain CBS 6938). Photoprotection and antioxidant related genes, as well as genes involved in sexual reproduction were analyzed. Both genomes had ca. 19 Mb and 6000 protein coding genes, similar to CBS 6938. Compared to other fungal genomes P. rhodozyma strains and other Cystofilobasidiales have the highest number of intron-containing genes and highest number of introns per gene. The Patagonian strain showed 4.4 % of nucleotide sequence divergence compared to the European strains which differed from each other by only 0.073 %. All known genes related to the synthesis of astaxanthin were annotated. A hitherto unknown gene cluster potentially responsible for photoprotection (mycosporines) was found in the newly sequenced P. rhodozyma strains but was absent in the non-mycosporinogenic strain CBS 6938. A broad battery of enzymes that act as scavengers of free radical oxygen species were detected, including catalases and superoxide dismutases (SODs). Additionally, genes involved in sexual reproduction were found and annotated. A draft genome sequence of the type strain of P. rhodozyma is now available, and comparison with that of the Patagonian population suggests the latter deserves to be assigned to a distinct variety. An unexpected genetic trait regarding high occurrence of introns in P. rhodozyma and other Cystofilobasidiales was revealed. New genomic insights into fungal homothallism were also provided. The genetic basis of several additional photoprotective and antioxidant strategies were described, indicating that P. rhodozyma is one of the fungi most well-equipped to cope with environmental oxidative stress, a factor that has probably contributed to shaping its genome.

Diversity and phylogenetic relationships among Bartonella strains from Thai bats.

PubMed

McKee, Clifton D; Kosoy, Michael Y; Bai, Ying; Osikowicz, Lynn M; Franka, Richard; Gilbert, Amy T; Boonmar, Sumalee; Rupprecht, Charles E; Peruski, Leonard F

2017-01-01

Bartonellae are phylogenetically diverse, intracellular bacteria commonly found in mammals. Previous studies have demonstrated that bats have a high prevalence and diversity of Bartonella infections globally. Isolates (n = 42) were obtained from five bat species in four provinces of Thailand and analyzed using sequences of the citrate synthase gene (gltA). Sequences clustered into seven distinct genogroups; four of these genogroups displayed similarity with Bartonella spp. sequences from other bats in Southeast Asia, Africa, and Eastern Europe. Thirty of the isolates representing these seven genogroups were further characterized by sequencing four additional loci (ftsZ, nuoG, rpoB, and ITS) to clarify their evolutionary relationships with other Bartonella species and to assess patterns of diversity among strains. Among the seven genogroups, there were differences in the number of sequence variants, ranging from 1-5, and the amount of nucleotide divergence, ranging from 0.035-3.9%. Overall, these seven genogroups meet the criteria for distinction as novel Bartonella species, with sequence divergence among genogroups ranging from 6.4-15.8%. Evidence of intra- and intercontinental phylogenetic relationships and instances of homologous recombination among Bartonella genogroups in related bat species were found in Thai bats.

Complete Genome Sequence of Pigmentation Negative Yersinia Pestis strain Cadman Running head: Complete Genome Sequence of Y. pestis strain Cadman

DTIC Science & Technology

2016-10-27

Institute of Infectious Diseases, Fort Detrick, Frederick, Maryland, USA 9 10 11 Running head: Complete Genome Sequence of Y. pestis strain Cadman...1 Complete Genome Sequence of Pigmentation Negative Yersinia pestis strain Cadman 1 2 3 Sean Lovetta, Kitty Chaseb, Galina Korolevaa, Gustavo...we report the genome sequence of Yersinia pestis strain Cadman, an attenuated strain 25 lacking the pgm locus. Y. pestis is the causative agent of

Species Identification and Strain Typing of Staphylococcus agnetis and Staphylococcus hyicus Isolates from Bovine Milk by Use of a Novel Multiplex PCR Assay and Pulsed-Field Gel Electrophoresis.

PubMed

Adkins, P R F; Middleton, J R; Calcutt, M J; Stewart, G C; Fox, L K

2017-06-01

Staphylococcus hyicus and Staphylococcus agnetis are two coagulase-variable staphylococcal species that can be isolated from bovine milk and are difficult to differentiate. The objectives of this study were to characterize isolates of bovine milk origin from a collection that had previously been characterized as coagulase-positive S. hyicus based on phenotypic species identification methods and to develop a PCR-based method for differentiating S. hyicus , S. agnetis , and Staphylococcus aureus Isolates ( n = 62) were selected from a previous study in which milk samples were collected from cows on 15 dairy herds. Isolates were coagulase tested and identified to the species level using housekeeping gene sequencing. A multiplex PCR to differentiate S. hyicus , S. agnetis , and S. aureus was developed. Pulsed-field gel electrophoresis was conducted to strain type the isolates. Based on gene sequencing, 44/62 of the isolates were determined to be either S. agnetis ( n = 43) or S. hyicus ( n = 1). Overall, 88% (37/42) of coagulase-positive S. agnetis isolates were found to be coagulase positive at 4 h. The herd-level prevalence of coagulase-positive S. agnetis ranged from 0 to 2.17%. Strain typing identified 23 different strains. Six strains were identified more than once and from multiple cows within the herd. Three strains were isolated from cows at more than one time point, with 41 to 264 days between samplings. These data suggest that S. agnetis is likely more prevalent on dairy farms than S. hyicus Also, some S. agnetis isolates in this study appeared to be contagious and associated with persistent infections. Copyright © 2017 American Society for Microbiology.

NEP: web server for epitope prediction based on antibody neutralization of viral strains with diverse sequences

PubMed Central

Chuang, Gwo-Yu; Liou, David; Kwong, Peter D.; Georgiev, Ivelin S.

2014-01-01

Delineation of the antigenic site, or epitope, recognized by an antibody can provide clues about functional vulnerabilities and resistance mechanisms, and can therefore guide antibody optimization and epitope-based vaccine design. Previously, we developed an algorithm for antibody-epitope prediction based on antibody neutralization of viral strains with diverse sequences and validated the algorithm on a set of broadly neutralizing HIV-1 antibodies. Here we describe the implementation of this algorithm, NEP (Neutralization-based Epitope Prediction), as a web-based server. The users must supply as input: (i) an alignment of antigen sequences of diverse viral strains; (ii) neutralization data for the antibody of interest against the same set of antigen sequences; and (iii) (optional) a structure of the unbound antigen, for enhanced prediction accuracy. The prediction results can be downloaded or viewed interactively on the antigen structure (if supplied) from the web browser using a JSmol applet. Since neutralization experiments are typically performed as one of the first steps in the characterization of an antibody to determine its breadth and potency, the NEP server can be used to predict antibody-epitope information at no additional experimental costs. NEP can be accessed on the internet at http://exon.niaid.nih.gov/nep. PMID:24782517

Partial sequencing of sodA gene and its application to identification of Streptococcus dysgalactiae subsp. dysgalactiae isolated from farmed fish.

PubMed

Nomoto, R; Kagawa, H; Yoshida, T

2008-01-01

To investigate the difference between Lancefield group C Streptococcus dysgalactiae (GCSD) strains isolated from diseased fish and animals by sequencing and phylogenetic analysis of the sodA gene. The sodA gene of Strep. dysgalactiae strains isolated from fish and animals were amplified and its nucleotide sequences were determined. Although 100% sequence identity was observed among fish GCSD strains, the determined sequences from animal isolates showed variations against fish isolate sequences. Thus, all fish GCSD strains were clearly separated from the GCSD strains of other origin by using phylogenetic tree analysis. In addition, the original primer set was designed based on the determined sequences for specifically amplify the sodA gene of fish GCSD strains. The primer set yield amplification products from only fish GCSD strains. By sequencing analysis of the sodA gene, the genetic divergence between Strep. dysgalactiae strains isolated from fish and mammals was demonstrated. Moreover, an original oligonucletide primer set, which could simply detect the genotype of fish GCSD strains was designed. This study shows that Strep. dysgalactiae isolated from diseased fish could be distinguished from conventional GCSD strains by the difference in the sequence of the sodA gene.

Nucleotide and deduced amino acid sequence of the envelope gene of the Vasilchenko strain of TBE virus; comparison with other flaviviruses.

PubMed

Gritsun, T S; Frolova, T V; Pogodina, V V; Lashkevich, V A; Venugopal, K; Gould, E A

1993-02-01

A strain of tick-borne encephalitis virus known as Vasilchenko (Vs) exhibits relatively low virulence characteristics in monkeys, Syrian hamsters and humans. The gene encoding the envelope glycoprotein of this virus was cloned and sequenced. Alignment of the sequence with those of other known tick-borne flaviviruses and identification of the recognised amino acid genetic marker EHLPTA confirmed its identity as a member of the TBE complex. However, Vs virus was distinguishable from eastern and western tick-borne serotypes by the presence of the sequence AQQ at amino acid positions 232-234 and also by the presence of other specific amino acid substitutions which may be genetic markers for these viruses and could determine their pathogenetic characteristics. When compared with other tick-borne flaviviruses, Vs virus had 12 unique amino acid substitutions including an additional potential glycosylation site at position (315-317). The Vs virus strain shared closest nucleotide and amino acid homology (84.5% and 95.5% respectively) with western and far eastern strains of tick-borne encephalitis virus. Comparison with the far eastern serotype of tick-borne encephalitis virus, by cross-immunoelectrophoresis of Vs virions and PAGE analysis of the extracted virion proteins, revealed differences in surface charge and virus stability that may account for the different virulence characteristics of Vs virus. These results support and enlarge upon previous data obtained from molecular and serological analysis.

Phenotypic variation in Lactococcus lactis subsp. lactis isolates derived from intestinal tracts of marine and freshwater fish.

PubMed

Itoi, S; Yuasa, K; Washio, S; Abe, T; Ikuno, E; Sugita, H

2009-09-01

We compared phenotypic characteristics of Lactococcus lactis subsp. lactis derived from different sources including the intestinal tract of marine fish and freshwater fish, and cheese starter culture. In the phylogenetic analysis based on partial 16S rRNA gene nucleotide sequences (1371 bp), freshwater fish-, marine fish- and cheese starter culture-derived strains were identical to that of L. lactis subsp. lactis previously reported. Fermentation profiles determined using the API 50 CH system were similar except for fermentation of several sugars including l-arabinose, mannitol, amygdalin, saccharose, trehalose, inulin and gluconate. The strains did have distinct levels of halotolerance: marine fish-derived strains > cheese starter-derived strain > freshwater fish-derived isolate. Lactococcus lactis subsp. lactis showed extensive diversity in phenotypic adaptation to various environments. The phenotypic properties of these strains suggested that L. lactis subsp. lactis strains from fish intestine have additional functions compared with the cheese starter-derived strain that has previously described. The unique phenotypic traits of the fish intestinal tract-derived L. lactis subsp. lactis might make them useful as a probiotics in aquaculture, and contribute to the development of functional foods and novel food additives, since the strains derived from fish intestines might have additional functions such as antibacterial activity.

Genetic analysis of a human rotavirus that belongs to subgroup I but has an RNA pattern typical of subgroup II human rotaviruses.

PubMed Central

Nakagomi, O; Nakagomi, T; Hoshino, Y; Flores, J; Kapikian, A Z

1987-01-01

We have previously found (O. Nakagomi, T. Nakagomi, H. Oyamada, and T. Suto, J. Med. Virol. 17:29-34, 1985), during an epidemiological study in Japan, a novel human rotavirus that belongs to subgroup I but has a long RNA pattern typical of subgroup II human rotaviruses. From the stool specimen containing this virus, we successfully isolated in MA104 cells a rotavirus, designated AU-1, which possesses these novel characteristics. The possibility that strain AU-1 was a laboratory contaminant of an animal rotavirus previously adapted to tissue culture cells was ruled out, and the identity of the AU-1 strain was established. Genetic analysis by RNA-RNA hybridization revealed that the AU-1 strain is not a simple reassortant between subgroup I and II human rotaviruses but that it shares a high level of sequence homology only with the gene encoding VP7 (the major neutralization protein) of serotype 3 human rotaviruses. Weak homology of the genomic RNA segments was also observed between the AU-1 strain and animal rotavirus strains, including rhesus rotavirus strain RRV and bovine rotavirus strain NCDV. These results suggest that the AU-1 strain may be an animal rotavirus that infected a human. Images PMID:3038947

Genetic and Molecular Characterization of β-Lactamase-Negative Ampicillin-Resistant Haemophilus influenzae with Unusually High Resistance to Ampicillin

PubMed Central

Kaczmarek, Frank S.; Gootz, Thomas D.; Dib-Hajj, Fadia; Shang, Wenchi; Hallowell, Shawn; Cronan, Melissa

2004-01-01

Previous studies with beta-lactamase-negative, ampicillin-resistant (BLNAR) Haemophilus influenzae from Japan, France, and North America indicate that mutations in ftsI encoding PBP3 confer ampicillin MICs of 1 to 4 μg/ml. Several BLNAR strains with ampicillin MICs of 4 to 16 μg/ml recently isolated from North America were studied. Pulsed-field gel electrophoresis identified 12 unique BLNAR strains; sequencing of their ftsI transpeptidase domains identified 1 group I and 11 group II mutants, as designated previously (K. Ubukata, Y. Shibasaki, K. Yamamoto, N. Chiba, K. Hasegawa, Y. Takeuchi, K. Sunakawa, M. Inoue, and M. Konno, Antimicrob. Agents Chemother. 45:1693-1699, 2001). Geometric mean ampicillin MICs for several clinical isolates were 8 to 10.56 μg/ml. Replacement of the ftsI gene in H. influenzae Rd with the intact ftsI from several clinical isolates resulted in integrants with typical BLNAR geometric mean ampicillin MICs of 1.7 to 2.2 μg/ml. Cloning and purification of His-tagged PBP3 from three clinical BLNAR strains showed significantly reduced Bocillin binding compared to that of PBP3 from strain Rd. Based on these data, changes in PBP3 alone could not account for the high ampicillin MICs observed for these BLNAR isolates. In an effort to determine the presence of additional mechanism(s) of ampicillin resistance, sequencing of the transpeptidase regions of pbp1a, -1b, and -2 was performed. While numerous changes were observed compared to the sequences from Rd, no consistent pattern correlating with high-level ampicillin resistance was apparent. Additional analysis of the resistant BLNAR strains revealed frame shift insertions in acrR for all four high-level, ampicillin-resistant isolates. acrR was intact for all eight low-level ampicillin-resistant and four ampicillin-susceptible strains tested. A knockout of acrB made in one clinical isolate (initial mean ampicillin MIC of 10.3 μg/ml) lowered the ampicillin MIC to 3.67 μg/ml, typical for BLNAR strains. These studies illustrate that BLNAR strains with high ampicillin MICs exist that have combined resistance mechanisms in PBP3 and in the AcrAB efflux pump. PMID:15105114

Candida kuoi sp. nov., an anamorphic species of the Starmerella yeast clade that synthesizes sophorolipids.

PubMed

Kurtzman, Cletus P

2012-09-01

A novel strain of anamorphic yeast, designated strain NRRL Y-27208(T), was isolated from concentrated grape juice in Cape Province, South Africa. Analysis of nuclear large subunit rRNA gene sequences from the D1/D2 domains separated the novel isolate from strains of Starmerella bombicola and Starmerella meliponinorum, as well as from species of the genus Candida that are members of the Starmerella clade. Compared to previously described species, strain NRRL Y-27208(T) is most closely related to S. bombicola but can be separated from this species by its ability to grow on D-ribose and erythritol. Strain NRRL Y-27208(T) produced sophorolipids that have an open chain structure similar to Candida batistae, Candida riodocensis and Candida stellata, which is in contrast to the closed chain sophorolipids produced by S. bombicola and Candida apicola. The analyses showed that NRRL Y-27208(T) (= CBS 7267(T)) represents a novel species distinct from previously described species, for which the name Candida kuoi sp. nov. is proposed.

Phenotypic and genetic diversity of chlorine-resistant Methylobacterium strains isolated from various environments.

PubMed Central

Hiraishi, A; Furuhata, K; Matsumoto, A; Koike, K A; Fukuyama, M; Tabuchi, K

1995-01-01

Strains of pink-pigmented facultative methylotrophs which were isolated previously from various environments and assigned tentatively to the genus Methylobacterium were characterized in comparison with authentic strains of previously known species of this genus. Most of the isolates derived from chlorinated water supplies exhibited resistance to chlorine, whereas 29 to 40% of the isolates from air, natural aquatic environments, and clinical materials were chlorine resistant. None of the tested authentic strains of Methylobacterium species obtained from culture collections exhibited chlorine resistance. Numerical analysis of phenotypic profiles showed that the test organisms tested were separated from each other except M. organophilum and M. rhodesianum. The chlorine-resistant isolates were randomly distributed among all clusters. The 16S ribosomal DNA (rDNA) sequence-based phylogenetic analyses showed that representatives of the isolates together with known Methylobacterium species formed a line of descent distinct from that of members of related genera in the alpha-2 subclass of the Proteobacteria and were divided into three subclusters within the Methylobacterium group. These results demonstrate that there is phenotypic and genetic diversity among chlorine-resistant Methylobacterium strains within the genus. PMID:7793931

Phenotypic and genetic diversity of chlorine-resistant Methylobacterium strains isolated from various environments.

PubMed

Hiraishi, A; Furuhata, K; Matsumoto, A; Koike, K A; Fukuyama, M; Tabuchi, K

1995-06-01

Strains of pink-pigmented facultative methylotrophs which were isolated previously from various environments and assigned tentatively to the genus Methylobacterium were characterized in comparison with authentic strains of previously known species of this genus. Most of the isolates derived from chlorinated water supplies exhibited resistance to chlorine, whereas 29 to 40% of the isolates from air, natural aquatic environments, and clinical materials were chlorine resistant. None of the tested authentic strains of Methylobacterium species obtained from culture collections exhibited chlorine resistance. Numerical analysis of phenotypic profiles showed that the test organisms tested were separated from each other except M. organophilum and M. rhodesianum. The chlorine-resistant isolates were randomly distributed among all clusters. The 16S ribosomal DNA (rDNA) sequence-based phylogenetic analyses showed that representatives of the isolates together with known Methylobacterium species formed a line of descent distinct from that of members of related genera in the alpha-2 subclass of the Proteobacteria and were divided into three subclusters within the Methylobacterium group. These results demonstrate that there is phenotypic and genetic diversity among chlorine-resistant Methylobacterium strains within the genus.

Comparative Genomic Analysis for Genetic Variation in Sacbrood Virus of Apis cerana and Apis mellifera Honeybees From Different Regions of Vietnam

PubMed Central

Reddy, Kondreddy Eswar; Thu, Ha Thi; Yoo, Mi Sun; Ramya, Mummadireddy; Reddy, Bheemireddy Anjana; Lien, Nguyen Thi Kim; Trang, Nguyen Thi Phuong; Duong, Bui Thi Thuy; Lee, Hyun-Jeong; Kang, Seung-Won

2017-01-01

Abstract Sacbrood virus (SBV) is one of the most common viral infections of honeybees. The entire genome sequence for nine SBV infecting honeybees, Apis cerana and Apis mellifera, in Vietnam, namely AcSBV-Viet1, AcSBV-Viet2, AcSBV-Viet3, AmSBV-Viet4, AcSBV-Viet5, AmSBV-Viet6, AcSBV-Viet7, AcSBV-Viet8, and AcSBV-Viet9, was determined. These sequences were aligned with seven previously reported complete genome sequences of SBV from other countries, and various genomic regions were compared. The Vietnamese SBVs (VN-SBVs) shared 91–99% identity with each other, and shared 89–94% identity with strains from other countries. The open reading frames (ORFs) of the VN-SBV genomes differed greatly from those of SBVs from other countries, especially in their VP1 sequences. The AmSBV-Viet6 and AcSBV-Viet9 genome encodes 17 more amino acids within this region than the other VN-SBVs. In a phylogenetic analysis, the strains AmSBV-Viet4, AcSBV-Viet2, and AcSBV-Viet3 were clustered in group with AmSBV-UK, AmSBV-Kor21, and AmSBV-Kor19 strains. Whereas, the strains AmSBV-Viet6 and AcSBV-Viet7 clustered separately with the AcSBV strains from Korea and AcSBV-VietSBM2. And the strains AcSBV-Viet8, AcSBV-Viet1, AcSBV-Viet5, and AcSBV-Viet9 clustered with the AcSBV-India, AcSBV-Kor and AcSBV-VietSBM2. In a Simplot graph, the VN-SBVs diverged stronger in their ORF regions than in their 5′ or 3′ untranslated regions. The VN-SBVs possess genetic characteristics which are more similar to the Asian AcSBV strains than to AmSBV-UK strain. Taken together, our data indicate that host specificity, geographic distance, and viral cross-infections between different bee species may explain the genetic diversity among the VN-SBVs in A. cerana and A. mellifera and other SBV strains. PMID:29117376

Genetic heterogeneity of L-Zagreb mumps virus vaccine strain.

PubMed

Kosutic-Gulija, Tanja; Forcic, Dubravko; Santak, Maja; Ramljak, Ana; Mateljak-Lukacevic, Sanja; Mazuran, Renata

2008-07-10

The most often used mumps vaccine strains Jeryl Lynn (JL), RIT4385, Urabe-AM9, L-Zagreb and L-3 differ in immunogenicity and reactogenicity. Previous analyses showed that JL, Urabe-AM9 and L-3 are genetically heterogeneous. We identified the heterogeneity of L-Zagreb throughout the entire genome. Two major variants were defined: variant A being identical to the consensus sequence of viral seeds and vaccine(s) and variant B which differs from variant A in three nucleotide positions. The difference between viral variants in L-Zagreb strain is insufficient for distinct viral strains to be defined. We demonstrated that proportion of variants in L-Zagreb viral population depends on cell substrate used for viral replication in vitro and in vivo. L-Zagreb strain should be considered as a single strain composed of at least two variant viral genomes.

Use of Fe(III) as an electron acceptor to recover previously uncultured hyperthermophiles: isolation and characterization of Geothermobacterium ferrireducens gen. nov., sp. nov.

PubMed

Kashefi, Kazem; Holmes, Dawn E; Reysenbach, Anna-Louise; Lovley, Derek R

2002-04-01

It has recently been recognized that the ability to use Fe(III) as a terminal electron acceptor is a highly conserved characteristic in hyperthermophilic microorganisms. This suggests that it may be possible to recover as-yet-uncultured hyperthermophiles in pure culture if Fe(III) is used as an electron acceptor. As part of a study of the microbial diversity of the Obsidian Pool area in Yellowstone National Park, Wyo., hot sediment samples were used as the inoculum for enrichment cultures in media containing hydrogen as the sole electron donor and poorly crystalline Fe(III) oxide as the electron acceptor. A pure culture was recovered on solidified, Fe(III) oxide medium. The isolate, designated FW-1a, is a hyperthermophilic anaerobe that grows exclusively by coupling hydrogen oxidation to the reduction of poorly crystalline Fe(III) oxide. Organic carbon is not required for growth. Magnetite is the end product of Fe(III) oxide reduction under the culture conditions evaluated. The cells are rod shaped, about 0.5 microm by 1.0 to 1.2 microm, and motile and have a single flagellum. Strain FW-1a grows at circumneutral pH, at freshwater salinities, and at temperatures of between 65 and 100 degrees C with an optimum of 85 to 90 degrees C. To our knowledge this is the highest temperature optimum of any organism in the Bacteria. Analysis of the 16S ribosomal DNA (rDNA) sequence of strain FW-1a places it within the Bacteria, most closely related to abundant but uncultured microorganisms whose 16S rDNA sequences have been previously recovered from Obsidian Pool and a terrestrial hot spring in Iceland. While previous studies inferred that the uncultured microorganisms with these 16S rDNA sequences were sulfate-reducing organisms, the physiology of the strain FW-1a, which does not reduce sulfate, indicates that these organisms are just as likely to be Fe(III) reducers. These results further demonstrate that Fe(III) may be helpful for recovering as-yet-uncultured microorganisms from hydrothermal environments and illustrate that caution must be used in inferring the physiological characteristics of at least some thermophilic microorganisms solely from 16S rDNA sequences. Based on both its 16S rDNA sequence and physiological characteristics, strain FW-1a represents a new genus among the Bacteria. The name Geothermobacterium ferrireducens gen. nov., sp. nov., is proposed (ATCC BAA-426).

Distinguishing molecular features and clinical characteristics of a putative new rhinovirus species, human rhinovirus C (HRV C).

PubMed

McErlean, Peter; Shackelton, Laura A; Andrews, Emily; Webster, Dale R; Lambert, Stephen B; Nissen, Michael D; Sloots, Theo P; Mackay, Ian M

2008-04-02

Human rhinoviruses (HRVs) are the most frequently detected pathogens in acute respiratory tract infections (ARTIs) and yet little is known about the prevalence, recurrence, structure and clinical impact of individual members. During 2007, the complete coding sequences of six previously unknown and highly divergent HRV strains were reported. To catalogue the molecular and clinical features distinguishing the divergent HRV strains, we undertook, for the first time, in silico analyses of all available polyprotein sequences and performed retrospective reviews of the medical records of cases in which variants of the prototype strain, HRV-QPM, had been detected. Genomic analyses revealed that the six divergent strains, residing within a clade we previously called HRV A2, had the shortest polyprotein of all picornaviruses investigated. Structure-based amino acid alignments identified conserved motifs shared among members of the genus Rhinovirus as well as substantive deletions and insertions unique to the divergent strains. Deletions mostly affected regions encoding proteins traditionally involved in antigenicity and serving as HRV and HEV receptor footprints. Because the HRV A2 strains cannot yet be cultured, we created homology models of predicted HRV-QPM structural proteins. In silico comparisons confirmed that HRV-QPM was most closely related to the major group HRVs. HRV-QPM was most frequently detected in infants with expiratory wheezing or persistent cough who had been admitted to hospital and required supplemental oxygen. It was the only virus detected in 65% of positive individuals. These observations contributed to an objective clinical impact ranging from mild to severe. The divergent strains did not meet classification requirements for any existing species of the genus Rhinovirus or Enterovirus. HRV A2 strains should be partitioned into at least one new species, putatively called Human rhinovirus C, populated by members detected with high frequency, from individuals with respiratory symptoms requiring hospital admission.

The safe enterocin DD14 is a leaderless two-peptide bacteriocin with anti-Clostridium perfringens activity.

PubMed

Caly, Delphine L; Chevalier, Mickaël; Flahaut, Christophe; Cudennec, Benoit; Al Atya, Ahmed Khassaf; Chataigné, Gabrielle; D'Inca, Romain; Auclair, Eric; Drider, Djamel

2017-03-01

Enterococcus faecalis 14, a strain previously isolated from meconium, displayed activity against four Clostridium perfringens isolates when co-cultured on agar plates. The anti-Clostridium activity was ascribed to the production of enterocin DD14, which was subsequently purified. The minimum inhibitory concentration (MIC) of enterocin DD14 against one collection strain and one clinical C. perfringens strain was determined at 50 µg/mL. Furthermore, using the intestinal epithelial cell line IPEC-1, it was shown that E. faecalis 14 was not cytotoxic after 24 h of contact, and no cytotoxicity was observed when IPEC-1 cells were incubated with pure enterocin DD14 for 4 h. Enterocin DD14 was characterised using mass spectrometry and was shown to consist of two small proteins of 5200.74 Da and 5206.41 Da, respectively. The two peptides (DD14A and DD14B) have highly similar amino acid sequences and no signal peptide, which classifies enterocin DD14 as a class IIb leaderless two-peptide bacteriocin. The genes encoding DD14A and DD14B were sequenced and were shown to be 100% identical to other previously described enterocins MR10A and MR10B, in contrast to the producing strains, which are different. Consequently, the present in vitro study supports the potential of this E. faecalis 14 strain and/or its purified enterocin DD14 as putative anti-C. perfringens compounds in chickens. Copyright © 2017. Published by Elsevier B.V.

Lachancea quebecensis sp. nov., a yeast species consistently isolated from tree bark in the Canadian province of Québec.

PubMed

Freel, Kelle C; Charron, Guillaume; Leducq, Jean-Baptiste; Landry, Christian R; Schacherer, Joseph

2015-10-01

A thorough sampling of maple, oak, birch, and apple tree bark in North America yielded a set of isolates that represent a yeast species not yet formally described. The strains obtained were all isolated from the Canadian province of Québec. These four isolates have identical electrophoretic karyotypes, distinct from other species of the genus Lachancea, and are most closely related to the formally recognized species Lachancea thermotolerans according to the D1/D2 domain of the LSU rDNA gene and 5.8S–ITS region. Previous studies revealed the existence of a population of strains closely related to L. thermotolerans, with unique D1/D2 sequences and the ability to grow on melibiose, which is also true for these isolates. The sequences obtained here (for the D1/D2, and 5.8S–ITS region) are identical among the four strains, and in a phylogenetic analysis of the D1/D2 region, the strains form a distinct clade with the previously described population closely related to L. thermotolerans, composed of isolates from Japan, as well as from the provinces of Ontario and Québec in Canada. On the basis of select physiological and phylogenetic characteristics, a novel ascosporogenous yeast species, Lachancea quebecensis sp. nov., is proposed. The type strain LL11_022T ( = CBS 14138T = CLIB 1763T = UCDFST 15-106T) was isolated from maple tree bark in the Station Duchesnay, QC region of Québec, Canada. The MycoBank number is MB811749.

Halorubrum chaoviator sp. nov., a haloarchaeon isolated from sea salt in Baja California, Mexico, Western Australia and Naxos, Greece

PubMed Central

Mancinelli, Rocco L.; Landheim, Ragnhild; Sanchez-Porro, Cristina; Dornmayr-Pfaffenhuemer, Marion; Gruber, Claudia; Legat, Andrea; Ventosa, Antonio; Radax, Christian; Ihara, Kunio; White, Melisa R.; Stan-Lotter, Helga

2011-01-01

Three halophilic isolates, strains Halo-G*T, AUS-1 and Naxos II, were compared. Halo-G* was isolated from an evaporitic salt crystal from Baja California, Mexico, whereas AUS-1 and Naxos II were isolated from salt pools in Western Australia and the Greek island of Naxos, respectively. Halo-G*T had been exposed previously to conditions of outer space and survived 2 weeks on the Biopan facility. Chemotaxonomic and molecular comparisons suggested high similarity between the three strains. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the strains clustered with Halorubrum species, showing sequence similarities of 99.2–97.1 %. The DNA–DNA hybridization values of strain Halo-G*T and strains AUS-1 and Naxos II are 73 and 75 %, respectively, indicating that they constitute a single species. The DNA relatedness between strain Halo-G*T and the type strains of 13 closely related species of the genus Halorubrum ranged from 39 to 2 %, suggesting that the three isolates constitute a different genospecies. The G+C content of the DNA of the three strains was 65.5–66.5 mol%. All three strains contained C20C20 derivatives of diethers of phosphatidylglycerol, phosphatidylglyceromethylphosphate and phosphatidylglycerolsulfate, together with a sulfated glycolipid. On the basis of these results, a novel species that includes the three strains is proposed, with the name Halorubrum chaoviator sp. nov. The type strain is strain Halo-G*T (=DSM 19316T =NCIMB 14426T =ATCC BAA-1602T). PMID:19567575

The Genetic Analysis of an Acinetobacter johnsonii Clinical Strain Evidenced the Presence of Horizontal Genetic Transfer.

PubMed

Montaña, Sabrina; Schramm, Sareda T J; Traglia, German Matías; Chiem, Kevin; Parmeciano Di Noto, Gisela; Almuzara, Marisa; Barberis, Claudia; Vay, Carlos; Quiroga, Cecilia; Tolmasky, Marcelo E; Iriarte, Andrés; Ramírez, María Soledad

2016-01-01

Acinetobacter johnsonii rarely causes human infections. While most A. johnsonii isolates are susceptible to virtually all antibiotics, strains harboring a variety of β-lactamases have recently been described. An A. johnsonii Aj2199 clinical strain recovered from a hospital in Buenos Aires produces PER-2 and OXA-58. We decided to delve into its genome by obtaining the whole genome sequence of the Aj2199 strain. Genome comparison studies on Aj2199 revealed 240 unique genes and a close relation to strain WJ10621, isolated from the urine of a patient in China. Genomic analysis showed evidence of horizontal genetic transfer (HGT) events. Forty-five insertion sequences and two intact prophages were found in addition to several resistance determinants such as blaPER-2, blaOXA-58, blaTEM-1, strA, strB, ereA, sul1, aacC2 and a new variant of blaOXA-211, called blaOXA-498. In particular, blaPER-2 and blaTEM-1 are present within the typical contexts previously described in the Enterobacteriaceae family. These results suggest that A. johnsonii actively acquires exogenous DNA from other bacterial species and concomitantly becomes a reservoir of resistance genes.

Periodic re-emergence of endemic strains with strong epidemic potential-a proposed explanation for the 2004 Indonesian dengue epidemic.

PubMed

Ong, Swee Hoe; Yip, Jin Teen; Chen, Yen Liang; Liu, Wei; Harun, Syahrial; Lystiyaningsih, Erlin; Heriyanto, Bambang; Beckett, Charmagne G; Mitchell, Wayne P; Hibberd, Martin L; Suwandono, Agus; Vasudevan, Subhash G; Schreiber, Mark J

2008-03-01

Indonesia experienced a severe dengue epidemic in the first quarter of 2004 with 58,301 cases and 658 deaths reported to the WHO. All four dengue virus (DENV) serotypes were detected, with DENV-3 the predominant strain. To ascertain the molecular epidemiology of the DENV associated with the epidemic, complete genomes of 15 isolates were sequenced from patient serum collected in Jakarta during the epidemic, and two historical DENV-3 isolates from previous epidemics in 1988 and 1998 were selectively sequenced for comparative studies. Phylogenetic trees for all four serotypes indicate the viruses are endemic strains that have been circulating in Indonesia for a few decades. Whole-genome phylogeny showed the 2004 DENV-3 isolates share high similarity with those isolated in 1998 during a major epidemic in Sumatra. Together these subtype I DENV-3 strains form a Sumatran-Javan clade with demonstrated epidemic potential. No newly-acquired amino acid mutations were found while comparing genomes from the two epidemics. This suggests re-emergence of little-changed endemic strains as causative agents of the epidemic in 2004. Notably, the molecular evidence rules out change in the viral genomes as the trigger of the epidemic.

Microbial compositional changes in broiler chicken cecal contents from birds challenged with different Salmonella vaccine candidate strains.

PubMed

Park, Si Hong; Kim, Sun Ae; Rubinelli, Peter M; Roto, Stephanie M; Ricke, Steven C

2017-05-31

Previously, we constructed and characterized the vaccine efficacy of Salmonella Typhimurium mutant strains in poultry with either inducible mviN expression (P BAD -mviN) or methionine auxotrophy (ΔΔmetRmetD). The aim of the present study was to assess potential impact of these Salmonella vaccine strains on the cecal microbiota using a next generation sequencing (NGS). The cecal microbial community obtained from unvaccinated (group 1) and vaccinated chickens (group 2, vaccinated with P BAD -mviN; group 3, vaccinated with wild type; group 4, vaccinated with ΔΔmetRmetD) were subjected to microbiome sequencing analysis with an Illumina MiSeq platform. The NGS microbiome analysis of chicken ceca revealed considerable changes in microbial composition in the presence of the different vaccine strains and exhibited detectable patterns of distinctive clustering among the respective groups (the R value of unweighted PCoA plot was 0.68). The present study indicates that different S. Typhimurium vaccine strains can differentially influence the microbiota of the ceca in terms of presence but not in the relative abundance of microbiota. Copyright © 2017 Elsevier Ltd. All rights reserved.

Acquisition of virulence genes by a carrier strain gave rise to the ongoing epidemics of meningococcal disease in West Africa.

PubMed

Brynildsrud, Ola Brønstad; Eldholm, Vegard; Bohlin, Jon; Uadiale, Kennedy; Obaro, Stephen; Caugant, Dominique A

2018-05-22

In the African meningitis belt, a region of sub-Saharan Africa comprising 22 countries from Senegal in the west to Ethiopia in the east, large epidemics of serogroup A meningococcal meningitis have occurred periodically. After gradual introduction from 2010 of mass vaccination with a monovalent meningococcal A conjugate vaccine, serogroup A epidemics have been eliminated. Starting in 2013, the northwestern part of Nigeria has been affected by yearly outbreaks of meningitis caused by a novel strain of serogroup C Neisseria meningitidis (NmC). In 2015, the strain spread to the neighboring country Niger, where it caused a severe epidemic. Following a relative calm in 2016, the largest ever recorded epidemic of NmC broke out in Nigeria in 2017. Here, we describe the recent evolution of this new outbreak strain and show how the acquisition of capsule genes and virulence factors by a strain previously circulating asymptomatically in the African population led to the emergence of a virulent pathogen. This study illustrates the power of long-read whole-genome sequencing, combined with Illumina sequencing, for high-resolution epidemiological investigations. Copyright © 2018 the Author(s). Published by PNAS.

Characterization of Francisella species isolated from the cooling water of an air conditioning system.

PubMed

Gu, Quan; Li, Xunde; Qu, Pinghua; Hou, Shuiping; Li, Juntao; Atwill, Edward R; Chen, Shouyi

2015-01-01

Strains of Francisella spp. were isolated from cooling water from an air conditioning system in Guangzhou, China. These strains are Gram negative, coccobacilli, non-motile, oxidase negative, catalase negative, esterase and lipid esterase positive. In addition, these bacteria grow on cysteine-supplemented media at 20 °C to 40 °C with an optimal growth temperature of 30 °C. Analysis of 16S rRNA gene sequences revealed that these strains belong to the genus Francisella. Biochemical tests and phylogenetic and BLAST analyses of 16S rRNA, rpoB and sdhA genes indicated that one strain was very similar to Francisella philomiragia and that the other strains were identical or highly similar to the Francisella guangzhouensis sp. nov. strain 08HL01032 we previously described. Biochemical and molecular characteristics of these strains demonstrated that multiple Francisella species exist in air conditioning systems.

Rhodoluna lacicola gen. nov., sp. nov., a planktonic freshwater bacterium with stream-lined genome

PubMed Central

Schmidt, Johanna; Taipale, Sami J.; Doolittle, W. Ford; Koll, Ulrike

2014-01-01

A pure culture of an actinobacterium previously described as ‘Candidatus Rhodoluna lacicola’ strain MWH-Ta8 was established and deposited in two public culture collections. Strain MWH-Ta8T represents a free-living planktonic freshwater bacterium obtained from hypertrophic Meiliang Bay, Lake Taihu, PR China. The strain was characterized by phylogenetic and taxonomic investigations, as well as by determination of its complete genome sequence. Strain MWH-Ta8T is noticeable due to its unusually low values of cell size (0.05 µm3), genome size (1.43 Mbp), and DNA G+C content (51.5 mol%). Phylogenetic analyses based on 16S rRNA gene and RpoB sequences suggested that strain MWH-Ta8T is affiliated with the family Microbacteriaceae with Pontimonas salivibrio being its closest relative among the currently described species within this family. Strain MWH-Ta8T and the type strain of Pontimonas salivibrio shared a 16S rRNA gene sequence similarity of 94.3 %. The cell-wall peptidoglycan of strain MWH-Ta8T was of type B2β (B10), containing 2,4-diaminobutyric acid as the diamino acid. The predominant cellular fatty acids were anteiso-C15 : 0 (36.5 %), iso-C16 : 0 (16.5 %), iso-C15 : 0 (15.6 %) and iso-C14 : 0 (8.9 %), and the major (>10 %) menaquinones were MK-11 and MK-12. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and two unknown glycolipids. The combined phylogenetic, phenotypic and chemotaxonomic data clearly suggest that strain MWH-Ta8T represents a novel species of a new genus in the family Microbacteriaceae, for which the name Rhodoluna lacicola gen. nov., sp. nov. is proposed. The type strain of the type species is MWH-Ta8T ( = DSM 23834T = LMG 26932T). PMID:24984700

Rhodoluna lacicola gen. nov., sp. nov., a planktonic freshwater bacterium with stream-lined genome.

PubMed

Hahn, Martin W; Schmidt, Johanna; Taipale, Sami J; Doolittle, W Ford; Koll, Ulrike

2014-09-01

A pure culture of an actinobacterium previously described as 'Candidatus Rhodoluna lacicola' strain MWH-Ta8 was established and deposited in two public culture collections. Strain MWH-Ta8(T) represents a free-living planktonic freshwater bacterium obtained from hypertrophic Meiliang Bay, Lake Taihu, PR China. The strain was characterized by phylogenetic and taxonomic investigations, as well as by determination of its complete genome sequence. Strain MWH-Ta8(T) is noticeable due to its unusually low values of cell size (0.05 µm(3)), genome size (1.43 Mbp), and DNA G+C content (51.5 mol%). Phylogenetic analyses based on 16S rRNA gene and RpoB sequences suggested that strain MWH-Ta8(T) is affiliated with the family Microbacteriaceae with Pontimonas salivibrio being its closest relative among the currently described species within this family. Strain MWH-Ta8(T) and the type strain of Pontimonas salivibrio shared a 16S rRNA gene sequence similarity of 94.3 %. The cell-wall peptidoglycan of strain MWH-Ta8(T) was of type B2β (B10), containing 2,4-diaminobutyric acid as the diamino acid. The predominant cellular fatty acids were anteiso-C15 : 0 (36.5 %), iso-C16 : 0 (16.5 %), iso-C15 : 0 (15.6 %) and iso-C14 : 0 (8.9 %), and the major (>10 %) menaquinones were MK-11 and MK-12. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and two unknown glycolipids. The combined phylogenetic, phenotypic and chemotaxonomic data clearly suggest that strain MWH-Ta8(T) represents a novel species of a new genus in the family Microbacteriaceae, for which the name Rhodoluna lacicola gen. nov., sp. nov. is proposed. The type strain of the type species is MWH-Ta8(T) ( = DSM 23834(T) = LMG 26932(T)). © 2014 IUMS.

Analysis of the vp2 gene sequence of a new mutated mink enteritis parvovirus strain in PR China

PubMed Central

2010-01-01

Background Mink enteritis virus (MEV) causes a highly contagious viral disease of mink with a worldwide distribution. MEV has a linear, single-stranded, negative-sense DNA with a genome length of approximately 5,000 bp. The VP2 protein is the major structural protein of the parvovirus encoded by the vp2 gene. VP2 is highly antigenic and plays important roles in determining viral host ranges and tissue tropisms. This study describes the bionomics and vp2 gene analysis of a mutated strain, MEV-DL, which was isolated recently in China and outlines its homologous relationships with other selected strains registered in Genbank. Results The MEV-DL strain can infect F81 cells with cytopathic effects. Pig erythrocytes were agglutinated by the MEV-DL strain. The generation of MEV-DL in F81 cells could infect mink within three months and cause a disease that was similar to that caused by wild-type MEV. A comparative analysis of the vp2 gene nucleotide (nt) sequence of MEV-DL showed that this was more than 99% homologous with other mink enteritis parvoviruses in Genbank. However, the nucleotide residues at positions 1,065 and 1,238 in the MEV-DL strain of the vp2 gene differed from those of all the other MEV strains described previously. It is noteworthy that the mutation at the nucleotide residues position 1,238 led to Asp/Gly replacement. This may lead to structural changes. A phylogenetic tree and sequence distance table were obtained, which showed that the MEV-DL and ZYL-1 strains had the closest inheritance distance. Conclusions A new variation of the vp2 gene exists in the MEV-DL strain, which may lead to structural changes of the VP2 protein. Phylogenetic analysis showed that MEV-DL may originate from the ZYL-1 strain in DaLian. PMID:20540765

Rapid identification of causative species in patients with Old World leishmaniasis.

PubMed Central

Minodier, P; Piarroux, R; Gambarelli, F; Joblet, C; Dumon, H

1997-01-01

Conventional methods for the identification of species of Leishmania parasite causing infections have limitations. By using a DNA-based alternative, the present study tries to develop a new tool for this purpose. Thirty-three patients living in Marseilles (in the south of France) were suffering from visceral or cutaneous leishmaniasis. DNA of the parasite in clinical samples (bone marrow, peripheral blood, or skin) from these patients were amplified by PCR and were directly sequenced. The sequences observed were compared to these of 30 strains of the genus causing Old World leishmaniasis collected in Europe, Africa, or Asia. In the analysis of the sequences of the strains, two different sequence patterns for Leishmania infantum, one sequence for Leishmania donovani, one sequence for Leishmania major, two sequences for Leishmania tropica, and one sequence for Leishmania aethiopica were obtained. Four sequences were observed among the strains from the patients: one was similar to the sequence for the L. major strains, two were identical to the sequences for the L. infantum strains, and the last sequence was not observed within the strains but had a high degree of homology with the sequences of the L. infantum and L. donovani strains. The L. infantum strains from all immunocompetent patients had the same sequence. The L. infantum strains from immunodeficient patients suffering from visceral leishmaniasis had three different sequences. This fact might signify that some variants of L. infantum acquire pathogenicity exclusively in immunocompromised patients. To dispense with the sequencing step, a restriction assay with HaeIII was used. Some restriction patterns might support genetic exchanges in members of the genus Leishmania. PMID:9316906

Identifying Group-Specific Sequences for Microbial Communities Using Long k-mer Sequence Signatures

PubMed Central

Wang, Ying; Fu, Lei; Ren, Jie; Yu, Zhaoxia; Chen, Ting; Sun, Fengzhu

2018-01-01

Comparing metagenomic samples is crucial for understanding microbial communities. For different groups of microbial communities, such as human gut metagenomic samples from patients with a certain disease and healthy controls, identifying group-specific sequences offers essential information for potential biomarker discovery. A sequence that is present, or rich, in one group, but absent, or scarce, in another group is considered “group-specific” in our study. Our main purpose is to discover group-specific sequence regions between control and case groups as disease-associated markers. We developed a long k-mer (k ≥ 30 bps)-based computational pipeline to detect group-specific sequences at strain resolution free from reference sequences, sequence alignments, and metagenome-wide de novo assembly. We called our method MetaGO: Group-specific oligonucleotide analysis for metagenomic samples. An open-source pipeline on Apache Spark was developed with parallel computing. We applied MetaGO to one simulated and three real metagenomic datasets to evaluate the discriminative capability of identified group-specific markers. In the simulated dataset, 99.11% of group-specific logical 40-mers covered 98.89% disease-specific regions from the disease-associated strain. In addition, 97.90% of group-specific numerical 40-mers covered 99.61 and 96.39% of differentially abundant genome and regions between two groups, respectively. For a large-scale metagenomic liver cirrhosis (LC)-associated dataset, we identified 37,647 group-specific 40-mer features. Any one of the features can predict disease status of the training samples with the average of sensitivity and specificity higher than 0.8. The random forests classification using the top 10 group-specific features yielded a higher AUC (from ∼0.8 to ∼0.9) than that of previous studies. All group-specific 40-mers were present in LC patients, but not healthy controls. All the assembled 11 LC-specific sequences can be mapped to two strains of Veillonella parvula: UTDB1-3 and DSM2008. The experiments on the other two real datasets related to Inflammatory Bowel Disease and Type 2 Diabetes in Women consistently demonstrated that MetaGO achieved better prediction accuracy with fewer features compared to previous studies. The experiments showed that MetaGO is a powerful tool for identifying group-specific k-mers, which would be clinically applicable for disease prediction. MetaGO is available at https://github.com/VVsmileyx/MetaGO. PMID:29774017

Multi-gene phylogenetic analysis reveals that shochu-fermenting Saccharomyces cerevisiae strains form a distinct sub-clade of the Japanese sake cluster.

PubMed

Futagami, Taiki; Kadooka, Chihiro; Ando, Yoshinori; Okutsu, Kayu; Yoshizaki, Yumiko; Setoguchi, Shinji; Takamine, Kazunori; Kawai, Mikihiko; Tamaki, Hisanori

2017-10-01

Shochu is a traditional Japanese distilled spirit. The formation of the distinguishing flavour of shochu produced in individual distilleries is attributed to putative indigenous yeast strains. In this study, we performed the first (to our knowledge) phylogenetic classification of shochu strains based on nucleotide gene sequences. We performed phylogenetic classification of 21 putative indigenous shochu yeast strains isolated from 11 distilleries. All of these strains were shown or confirmed to be Saccharomyces cerevisiae, sharing species identification with 34 known S. cerevisiae strains (including commonly used shochu, sake, ale, whisky, bakery, bioethanol and laboratory yeast strains and clinical isolate) that were tested in parallel. Our analysis used five genes that reflect genome-level phylogeny for the strain-level classification. In a first step, we demonstrated that partial regions of the ZAP1, THI7, PXL1, YRR1 and GLG1 genes were sufficient to reproduce previous sub-species classifications. In a second step, these five analysed regions from each of 25 strains (four commonly used shochu strains and the 21 putative indigenous shochu strains) were concatenated and used to generate a phylogenetic tree. Further analysis revealed that the putative indigenous shochu yeast strains form a monophyletic group that includes both the shochu yeasts and a subset of the sake group strains; this cluster is a sister group to other sake yeast strains, together comprising a sake-shochu group. Differences among shochu strains were small, suggesting that it may be possible to correlate subtle phenotypic differences among shochu flavours with specific differences in genome sequences. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Correlation of genetic variability with safety of mumps vaccine Urabe AM9 strain.

PubMed

Amexis, G; Fineschi, N; Chumakov, K

2001-08-15

The Urabe AM9 strain of mumps vaccine live is known for its genetic instability and some vaccines derived from this strain were withdrawn from the market due to an excessive number of vaccine-associated parotitis and meningitis cases. To identify the molecular basis of this instability, we determined complete nucleotide sequences of several stocks of the Urabe strain used for vaccine production by different manufacturers and of two clinical isolates from cases of vaccine-associated meningitis. In contrast to previously published studies relating the Lys335 --> Glu mutation in the viral HN gene with neurovirulence of mumps virus, we could not confirm any association of this mutation with the safety of mumps vaccine. Each of the three vaccine stocks studied had its own characteristic profile of mutations that was identified by cDNA sequencing and quantitated by mutant analysis by PCR and restriction enzyme cleavage. Determination of the mutational profile of mumps vaccine lots could allow vaccine manufacturers to characterize seed viruses and monitor the consistency of vaccine production to prevent emergence of virulent revertants.

Evidence of Brucella strain ST27 in bottlenose dolphin (Tursiops truncatus) in Europe.

PubMed

Cvetnić, Željko; Duvnjak, Sanja; Đuras, Martina; Gomerčić, Tomislav; Reil, Irena; Zdelar-Tuk, Maja; Špičić, Silvio

2016-11-30

Marine mammal brucellosis has been known for more than 20 years, but recent work suggests it is more widespread than originally thought. Brucella (B.) pinnipedialis has been isolated from pinnipeds, while B. ceti strains have been associated with cetaceans. Here we report a Brucella strain isolated from multiple lymph nodes of one bottlenose dolphin (Tursiops truncatus) during routine examination of dolphin carcasses found in the Croatian part of the northern Adriatic Sea during the summer of 2015. Classical bacteriological biotyping, PCR-based techniques (single, multiplex, PCR-RFLP) and 16S rRNA DNA sequencing were used to identify Brucella spp. Multiple-locus variable number tandem repeat analysis of 16 loci and multilocus sequence typing of 9 loci were used for genotyping and species determination. The combination of bacteriological, molecular and genotyping techniques identified our strain as ST27, previously identified as a human pathogen. This report provides, to our knowledge, the first evidence of ST27 in the Adriatic Sea in particular and in European waters in general. The zoonotic nature of the strain and its presence in the Adriatic, which is inhabited by bottlenose dolphins, suggest that the strain may pose a significant threat to human health. Copyright © 2016 Elsevier B.V. All rights reserved.

Genetic characterization of epizootic hemorrhagic disease virus strains isolated from cattle in Israel

USDA-ARS?s Scientific Manuscript database

Epizootic hemorrhagic disease virus (EHDV), an Orbivirus not previously reported in Israel, was isolated from Israeli cattle during a “bluetongue like” disease outbreak in 2006. To ascertain the origin of this new virus, three isolates from the outbreak were fully sequenced and compared with availab...

Bacteriophage prevalence in the genus Azospirillum and analysis of the first genome sequence of an Azospirillum brasilense integrative phage.

PubMed

Boyer, Mickaël; Haurat, Jacqueline; Samain, Sylvie; Segurens, Béatrice; Gavory, Frédérick; González, Víctor; Mavingui, Patrick; Rohr, René; Bally, René; Wisniewski-Dyé, Florence

2008-02-01

The prevalence of bacteriophages was investigated in 24 strains of four species of plant growth-promoting rhizobacteria belonging to the genus Azospirillum. Upon induction by mitomycin C, the release of phage particles was observed in 11 strains from three species. Transmission electron microscopy revealed two distinct sizes of particles, depending on the identity of the Azospirillum species, typical of the Siphoviridae family. Pulsed-field gel electrophoresis and hybridization experiments carried out on phage-encapsidated DNAs revealed that all phages isolated from A. lipoferum and A. doebereinerae strains had a size of about 10 kb whereas all phages isolated from A. brasilense strains displayed genome sizes ranging from 62 to 65 kb. Strong DNA hybridizing signals were shown for most phages hosted by the same species whereas no homology was found between phages harbored by different species. Moreover, the complete sequence of the A. brasilense Cd bacteriophage (phiAb-Cd) genome was determined as a double-stranded DNA circular molecule of 62,337 pb that encodes 95 predicted proteins. Only 14 of the predicted proteins could be assigned functions, some of which were involved in DNA processing, phage morphogenesis, and bacterial lysis. In addition, the phiAb-Cd complete genome was mapped as a prophage on a 570-kb replicon of strain A. brasilense Cd, and a region of 27.3 kb of phiAb-Cd was found to be duplicated on the 130-kb pRhico plasmid previously sequenced from A. brasilense Sp7, the parental strain of A. brasilense Cd.

Bacteriophage Prevalence in the Genus Azospirillum and Analysis of the First Genome Sequence of an Azospirillum brasilense Integrative Phage▿

PubMed Central

Boyer, Mickaël; Haurat, Jacqueline; Samain, Sylvie; Segurens, Béatrice; Gavory, Frédérick; González, Víctor; Mavingui, Patrick; Rohr, René; Bally, René; Wisniewski-Dyé, Florence

2008-01-01

The prevalence of bacteriophages was investigated in 24 strains of four species of plant growth-promoting rhizobacteria belonging to the genus Azospirillum. Upon induction by mitomycin C, the release of phage particles was observed in 11 strains from three species. Transmission electron microscopy revealed two distinct sizes of particles, depending on the identity of the Azospirillum species, typical of the Siphoviridae family. Pulsed-field gel electrophoresis and hybridization experiments carried out on phage-encapsidated DNAs revealed that all phages isolated from A. lipoferum and A. doebereinerae strains had a size of about 10 kb whereas all phages isolated from A. brasilense strains displayed genome sizes ranging from 62 to 65 kb. Strong DNA hybridizing signals were shown for most phages hosted by the same species whereas no homology was found between phages harbored by different species. Moreover, the complete sequence of the A. brasilense Cd bacteriophage (ΦAb-Cd) genome was determined as a double-stranded DNA circular molecule of 62,337 pb that encodes 95 predicted proteins. Only 14 of the predicted proteins could be assigned functions, some of which were involved in DNA processing, phage morphogenesis, and bacterial lysis. In addition, the ΦAb-Cd complete genome was mapped as a prophage on a 570-kb replicon of strain A. brasilense Cd, and a region of 27.3 kb of ΦAb-Cd was found to be duplicated on the 130-kb pRhico plasmid previously sequenced from A. brasilense Sp7, the parental strain of A. brasilense Cd. PMID:18065619

Mycobacterium tuberculosis Whole Genome Sequences From Southern India Suggest Novel Resistance Mechanisms and the Need for Region-Specific Diagnostics.

PubMed

Manson, Abigail L; Abeel, Thomas; Galagan, James E; Sundaramurthi, Jagadish Chandrabose; Salazar, Alex; Gehrmann, Thies; Shanmugam, Siva Kumar; Palaniyandi, Kannan; Narayanan, Sujatha; Swaminathan, Soumya; Earl, Ashlee M

2017-06-01

India is home to 25% of all tuberculosis cases and the second highest number of multidrug resistant cases worldwide. However, little is known about the genetic diversity and resistance determinants of Indian Mycobacterium tuberculosis, particularly for the primary lineages found in India, lineages 1 and 3. We whole genome sequenced 223 randomly selected M. tuberculosis strains from 196 patients within the Tiruvallur and Madurai districts of Tamil Nadu in Southern India. Using comparative genomics, we examined genetic diversity, transmission patterns, and evolution of resistance. Genomic analyses revealed (11) prevalence of strains from lineages 1 and 3, (11) recent transmission of strains among patients from the same treatment centers, (11) emergence of drug resistance within patients over time, (11) resistance gained in an order typical of strains from different lineages and geographies, (11) underperformance of known resistance-conferring mutations to explain phenotypic resistance in Indian strains relative to studies focused on other geographies, and (11) the possibility that resistance arose through mutations not previously implicated in resistance, or through infections with multiple strains that confound genotype-based prediction of resistance. In addition to substantially expanding the genomic perspectives of lineages 1 and 3, sequencing and analysis of M. tuberculosis whole genomes from Southern India highlight challenges of infection control and rapid diagnosis of resistant tuberculosis using current technologies. Further studies are needed to fully explore the complement of diversity and resistance determinants within endemic M. tuberculosis populations. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Application of the major capsid protein as a marker of the phylogenetic diversity of Emiliania huxleyi viruses.

PubMed

Rowe, Janet M; Fabre, Marie-Françoise; Gobena, Daniel; Wilson, William H; Wilhelm, Steven W

2011-05-01

Studies of the Phycodnaviridae have traditionally relied on the DNA polymerase (pol) gene as a biomarker. However, recent investigations have suggested that the major capsid protein (MCP) gene may be a reliable phylogenetic biomarker. We used MCP gene amplicons gathered across the North Atlantic to assess the diversity of Emiliania huxleyi-infecting Phycodnaviridae. Nucleotide sequences were examined across >6000 km of open ocean, with comparisons between concentrates of the virus-size fraction of seawater and of lysates generated by exposing host strains to these same virus concentrates. Analyses revealed that many sequences were only sampled once, while several were over-represented. Analyses also revealed nucleotide sequences distinct from previous coastal isolates. Examination of lysed cultures revealed a new richness in phylogeny, as MCP sequences previously unrepresented within the existing collection of E. huxleyi viruses (EhV) were associated with viruses lysing cultures. Sequences were compared with previously described EhV MCP sequences from the North Sea and a Norwegian Fjord, as well as from the Gulf of Maine. Principal component analysis indicates that location-specific distinctions exist despite the presence of sequences common across these environments. Overall, this investigation provides new sequence data and an assessment on the use of the MCP gene. © 2011 Federation of European Microbiological Societies Published by Blackwell Publishing Ltd. All rights reserved.

Discovery of Escherichia coli CRISPR sequences in an undergraduate laboratory.

PubMed

Militello, Kevin T; Lazatin, Justine C

2017-05-01

Clustered regularly interspaced short palindromic repeats (CRISPRs) represent a novel type of adaptive immune system found in eubacteria and archaebacteria. CRISPRs have recently generated a lot of attention due to their unique ability to catalog foreign nucleic acids, their ability to destroy foreign nucleic acids in a mechanism that shares some similarity to RNA interference, and the ability to utilize reconstituted CRISPR systems for genome editing in numerous organisms. In order to introduce CRISPR biology into an undergraduate upper-level laboratory, a five-week set of exercises was designed to allow students to examine the CRISPR status of uncharacterized Escherichia coli strains and to allow the discovery of new repeats and spacers. Students started the project by isolating genomic DNA from E. coli and amplifying the iap CRISPR locus using the polymerase chain reaction (PCR). The PCR products were analyzed by Sanger DNA sequencing, and the sequences were examined for the presence of CRISPR repeat sequences. The regions between the repeats, the spacers, were extracted and analyzed with BLASTN searches. Overall, CRISPR loci were sequenced from several previously uncharacterized E. coli strains and one E. coli K-12 strain. Sanger DNA sequencing resulted in the discovery of 36 spacer sequences and their corresponding surrounding repeat sequences. Five of the spacers were homologous to foreign (non-E. coli) DNA. Assessment of the laboratory indicates that improvements were made in the ability of students to answer questions relating to the structure and function of CRISPRs. Future directions of the laboratory are presented and discussed. © 2016 by The International Union of Biochemistry and Molecular Biology, 45(3):262-269, 2017. © 2016 The International Union of Biochemistry and Molecular Biology.

Use of essential gene, encoding prophobilinogen deaminase from extreme psychrophilic Colwellia sp. C1, to generate temperature-sensitive strain of Francisella novicida.

PubMed

Pankowski, J A

2016-08-01

Previously, several essential genes from psychrophilic bacteria have been substituted for their homologues in mesophilic bacterial pathogens to make the latter temperature sensitive. It has been noted that an essential ligA gene from an extreme psychrophile, Colwellia sp. C1, yielded a gene product that is inactivated at 27°C, the lowest that has been observed for any psychrophilic enzyme, and hypothesized that other essential proteins of that strain would also have low inactivation temperatures. This work describes the partial sequencing of the genome of Colwellia sp. C1 strain and the identification of 24 open reading frames encoding homologues of highly conserved bacterial essential genes. The gene encoding porphobilinogen deaminase (hemC), which is involved in the pathway of haem synthesis, has been tested for its ability to convert Francisella novicida into a temperature-sensitive strain. The hybrid strain carrying the C1-derived hemC gene exhibited a temperature-sensitive phenotype with a restrictive temperature of 36°C. These results support the conclusion that Colwellia sp. C1 is a rich source of heat-labile enzymes. The issue of biosafety is often raised when it comes to work with pathogenic organisms. The main concern is caused by the risk of researchers being exposed to infectious doses of dangerous microbes. This paper analyses essential genes identified in partial genomic sequence of the psychrophilic bacterium Collwelia sp. C1. These sequences can be used as a mean of generating temperature-sensitive strains of pathogenic bacteria. Such strains are incapable of surviving at the temperature of human body. This means they could be applied as vaccines or for safer work with dangerous organisms. © 2016 The Society for Applied Microbiology.

Multiplex Amplification Refractory Mutation System Polymerase Chain Reaction (ARMS-PCR) for diagnosis of natural infection with canine distemper virus

PubMed Central

2010-01-01

Background Canine distemper virus (CDV) is present worldwide and produces a lethal systemic infection of wild and domestic Canidae. Pre-existing antibodies acquired from vaccination or previous CDV infection might interfere the interpretation of a serologic diagnosis method. In addition, due to the high similarity of nucleic acid sequences between wild-type CDV and the new vaccine strain, current PCR derived methods cannot be applied for the definite confirmation of CD infection. Hence, it is worthy of developing a simple and rapid nucleotide-based assay for differentiation of wild-type CDV which is a cause of disease from attenuated CDVs after vaccination. High frequency variations have been found in the region spanning from the 3'-untranslated region (UTR) of the matrix (M) gene to the fusion (F) gene (designated M-F UTR) in a few CDV strains. To establish a differential diagnosis assay, an amplification refractory mutation analysis was established based on the highly variable region on M-F UTR and F regions. Results Sequences of frequent polymorphisms were found scattered throughout the M-F UTR region; the identity of nucleic acid between local strains and vaccine strains ranged from 82.5% to 93.8%. A track of AAA residue located 35 nucleotides downstream from F gene start codon highly conserved in three vaccine strains were replaced with TGC in the local strains; that severed as target sequences for deign of discrimination primers. The method established in the present study successfully differentiated seven Taiwanese CDV field isolates, all belonging to the Asia-1 lineage, from vaccine strains. Conclusions The method described herein would be useful for several clinical applications, such as confirmation of nature CDV infection, evaluation of vaccination status and verification of the circulating viral genotypes. PMID:20534175

Multiplex Amplification Refractory Mutation System Polymerase Chain Reaction (ARMS-PCR) for diagnosis of natural infection with canine distemper virus.

PubMed

Chulakasian, Songkhla; Lee, Min-Shiuh; Wang, Chi-Young; Chiou, Shyan-Song; Lin, Kuan-Hsun; Lin, Fong-Yuan; Hsu, Tien-Huan; Wong, Min-Liang; Chang, Tien-Jye; Hsu, Wei-Li

2010-06-10

Canine distemper virus (CDV) is present worldwide and produces a lethal systemic infection of wild and domestic Canidae. Pre-existing antibodies acquired from vaccination or previous CDV infection might interfere the interpretation of a serologic diagnosis method. In addition, due to the high similarity of nucleic acid sequences between wild-type CDV and the new vaccine strain, current PCR derived methods cannot be applied for the definite confirmation of CD infection. Hence, it is worthy of developing a simple and rapid nucleotide-based assay for differentiation of wild-type CDV which is a cause of disease from attenuated CDVs after vaccination. High frequency variations have been found in the region spanning from the 3'-untranslated region (UTR) of the matrix (M) gene to the fusion (F) gene (designated M-F UTR) in a few CDV strains. To establish a differential diagnosis assay, an amplification refractory mutation analysis was established based on the highly variable region on M-F UTR and F regions. Sequences of frequent polymorphisms were found scattered throughout the M-F UTR region; the identity of nucleic acid between local strains and vaccine strains ranged from 82.5% to 93.8%. A track of AAA residue located 35 nucleotides downstream from F gene start codon highly conserved in three vaccine strains were replaced with TGC in the local strains; that severed as target sequences for deign of discrimination primers. The method established in the present study successfully differentiated seven Taiwanese CDV field isolates, all belonging to the Asia-1 lineage, from vaccine strains. The method described herein would be useful for several clinical applications, such as confirmation of nature CDV infection, evaluation of vaccination status and verification of the circulating viral genotypes.

Prevalence of Avian-Pathogenic Escherichia coli Strain O1 Genomic Islands among Extraintestinal and Commensal E. coli Isolates

PubMed Central

Johnson, Timothy J.; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Johnson, James R.; Logue, Catherine M.

2012-01-01

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include pathogens of humans and animals. Previously, the genome of avian-pathogenic E. coli (APEC) O1:K1:H7 strain O1, from ST95, was sequenced and compared to those of several other E. coli strains, identifying 43 genomic islands. Here, the genomic islands of APEC O1 were compared to those of other sequenced E. coli strains, and the distribution of 81 genes belonging to 12 APEC O1 genomic islands among 828 human and avian ExPEC and commensal E. coli isolates was determined. Multiple islands were highly prevalent among isolates belonging to the O1 and O18 serogroups within phylogenetic group B2, which are implicated in human neonatal meningitis. Because of the extensive genomic similarities between APEC O1 and other human ExPEC strains belonging to the ST95 phylogenetic lineage, its ability to cause disease in a rat model of sepsis and meningitis was assessed. Unlike other ST95 lineage strains, APEC O1 was unable to cause bacteremia or meningitis in the neonatal rat model and was significantly less virulent than uropathogenic E. coli (UPEC) CFT073 in a mouse sepsis model, despite carrying multiple neonatal meningitis E. coli (NMEC) virulence factors and belonging to the ST95 phylogenetic lineage. These results suggest that host adaptation or genome modifications have occurred either in APEC O1 or in highly virulent ExPEC isolates, resulting in differences in pathogenicity. Overall, the genomic islands examined provide targets for further discrimination of the different ExPEC subpathotypes, serogroups, phylogenetic types, and sequence types. PMID:22467781

Prevalence of avian-pathogenic Escherichia coli strain O1 genomic islands among extraintestinal and commensal E. coli isolates.

PubMed

Johnson, Timothy J; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Johnson, James R; Logue, Catherine M; Nolan, Lisa K

2012-06-01

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include pathogens of humans and animals. Previously, the genome of avian-pathogenic E. coli (APEC) O1:K1:H7 strain O1, from ST95, was sequenced and compared to those of several other E. coli strains, identifying 43 genomic islands. Here, the genomic islands of APEC O1 were compared to those of other sequenced E. coli strains, and the distribution of 81 genes belonging to 12 APEC O1 genomic islands among 828 human and avian ExPEC and commensal E. coli isolates was determined. Multiple islands were highly prevalent among isolates belonging to the O1 and O18 serogroups within phylogenetic group B2, which are implicated in human neonatal meningitis. Because of the extensive genomic similarities between APEC O1 and other human ExPEC strains belonging to the ST95 phylogenetic lineage, its ability to cause disease in a rat model of sepsis and meningitis was assessed. Unlike other ST95 lineage strains, APEC O1 was unable to cause bacteremia or meningitis in the neonatal rat model and was significantly less virulent than uropathogenic E. coli (UPEC) CFT073 in a mouse sepsis model, despite carrying multiple neonatal meningitis E. coli (NMEC) virulence factors and belonging to the ST95 phylogenetic lineage. These results suggest that host adaptation or genome modifications have occurred either in APEC O1 or in highly virulent ExPEC isolates, resulting in differences in pathogenicity. Overall, the genomic islands examined provide targets for further discrimination of the different ExPEC subpathotypes, serogroups, phylogenetic types, and sequence types.

Streptomyces chiangmaiensis sp. nov. and Streptomyces lannensis sp. nov., isolated from the South-East Asian stingless bee (Tetragonilla collina).

PubMed

Promnuan, Yaowanoot; Kudo, Takuji; Ohkuma, Moriya; Chantawannakul, Panuwan

2013-05-01

Two novel actinomycetes, strains TA4-1(T) and TA4-8(T,) were isolated from the South-East Asian stingless bee (Tetragonilla collina Smith 1857), collected from Chiang Mai Province, Thailand. The morphological and chemotaxonomic properties of strains TA4-1(T) and TA4-8(T) were consistent with the genus Streptomyces, i.e. the formation of aerial mycelia bearing spiral spore chains, the presence of the ll-isomer of diaminopimelic acid in cell walls, iso- and anteiso-branched fatty acids with carbon chain lengths 14-17 atoms as the major fatty acids and MK-9(H8) as the predominant menaquinone plus minor amounts of MK-9(H6) and MK-9(H10). Analysis of 16S rRNA gene sequences showed that strains TA4-1(T) and TA4-8(T) exhibited 98.8 and 98.1% sequence similarity, respectively, with Streptomyces chromofuscus NRRL B-12175(T) and 98.9% sequence similarity with each other. This study suggested that strains TA4-1(T) and TA4-8(T) were distinct from previously described species of the genus Streptomyces. In addition, the low degrees of DNA-DNA relatedness between the isolates and S. chromofuscus JCM 4354(T) warranted assigning strains TA4-1(T) and TA4-8(T) to two novel species. The names Streptomyces chiangmaiensis sp. nov. (type strain TA4-1(T)  = JCM 16577(T)  = TISTR 1981(T)) and Streptomyces lannensis sp. nov. (type strain TA4-8(T)  = JCM 16578(T)  = TISTR 1982(T)) are proposed. The species names indicate the geographical locations where the stingless bees reside.

Distribution and Phylogeny of Immunoglobulin-Binding Protein G in Shiga Toxin-Producing Escherichia coli and Its Association with Adherence Phenotypes▿

PubMed Central

Merkel, Viktor; Ohder, Barbara; Bielaszewska, Martina; Zhang, Wenlan; Fruth, Angelika; Menge, Christian; Borrmann, Erika; Middendorf, Barbara; Müthing, Johannes; Karch, Helge; Mellmann, Alexander

2010-01-01

eibG in Shiga toxin-producing Escherichia coli (STEC) O91 encodes a protein (EibG) which binds human immunoglobulins G and A and contributes to bacterial chain-like adherence to human epithelial cells. We investigated the prevalence of eibG among STEC, the phylogeny of eibG, and eibG allelic variations and their impact on the adherence phenotype. eibG was found in 15.0% of 240 eae-negative STEC strains but in none of 157 eae-positive STEC strains. The 36 eibG-positive STEC strains belonged to 14 serotypes and to eight multilocus sequence types (STs), with serotype O91:H14/H− and ST33 being the most common. Sequences of the complete eibG gene (1,527 bp in size) from eibG-positive STEC resulted in 21 different alleles with 88.11% to 100% identity to the previously reported eibG sequence; they clustered into three eibG subtypes (eibG-α, eibG-β, and eibG-γ). Strains expressing EibG-α and EibG-β displayed a mostly typical chain-like adherence pattern (CLAP), with formation of long chains on both human and bovine intestinal epithelial cells, whereas strains with EibG-γ adhered in short chains, a pattern we termed atypical CLAP. The same adherence phenotypes were displayed by E. coli BL21(DE3) clones containing the respective eibG-α, eibG-β, and eibG-γ subtypes. We propose two possible evolutionary scenarios for eibG in STEC: a clonal development of eibG in strains with the same phylogenetic background or horizontal transfer of eibG between phylogenetically unrelated STEC strains. PMID:20547747

Comparative Genomics and Identification of an Enterotoxin-Bearing Pathogenicity Island, SEPI-1/SECI-1, in Staphylococcus epidermidis Pathogenic Strains.

PubMed

Argemi, Xavier; Nanoukon, Chimène; Affolabi, Dissou; Keller, Daniel; Hansmann, Yves; Riegel, Philippe; Baba-Moussa, Lamine; Prévost, Gilles

2018-02-25

Staphylococcus epidermidis is a leading cause of nosocomial infections, majorly resistant to beta-lactam antibiotics, and may transfer several mobile genetic elements among the members of its own species, as well as to Staphylococcus aureus ; however, a genetic exchange from S. aureus to S. epidermidis remains controversial. We recently identified two pathogenic clinical strains of S. epidermidis that produce a staphylococcal enterotoxin C3-like (SEC) similar to that by S. aureus pathogenicity islands. This study aimed to determine the genetic environment of the SEC-coding sequence and to identify the mobile genetic elements. Whole-genome sequencing and annotation of the S. epidermidis strains were performed using Illumina technology and a bioinformatics pipeline for assembly, which provided evidence that the SEC-coding sequences were located in a composite pathogenicity island that was previously described in the S. epidermidis strain FRI909, called SePI-1/SeCI-1, with 83.8-89.7% nucleotide similarity. Various other plasmids were identified, particularly p_3_95 and p_4_95, which carry antibiotic resistance genes ( hsrA and dfrG , respectively), and share homologies with SAP085A and pUSA04-2-SUR11, two plasmids described in S. aureus . Eventually, one complete prophage was identified, ΦSE90, sharing 30 out of 52 coding sequences with the Acinetobacter phage vB_AbaM_IME200. Thus, the SePI-1/SeCI-1 pathogenicity island was identified in two pathogenic strains of S. epidermidis that produced a SEC enterotoxin causing septic shock. These findings suggest the existence of in vivo genetic exchange from S. aureus to S. epidermidis .

Comparative Genomics and Identification of an Enterotoxin-Bearing Pathogenicity Island, SEPI-1/SECI-1, in Staphylococcus epidermidis Pathogenic Strains

PubMed Central

Nanoukon, Chimène; Affolabi, Dissou; Keller, Daniel; Hansmann, Yves; Riegel, Philippe; Baba-Moussa, Lamine; Prévost, Gilles

2018-01-01

Staphylococcus epidermidis is a leading cause of nosocomial infections, majorly resistant to beta-lactam antibiotics, and may transfer several mobile genetic elements among the members of its own species, as well as to Staphylococcus aureus; however, a genetic exchange from S. aureus to S. epidermidis remains controversial. We recently identified two pathogenic clinical strains of S. epidermidis that produce a staphylococcal enterotoxin C3-like (SEC) similar to that by S. aureus pathogenicity islands. This study aimed to determine the genetic environment of the SEC-coding sequence and to identify the mobile genetic elements. Whole-genome sequencing and annotation of the S. epidermidis strains were performed using Illumina technology and a bioinformatics pipeline for assembly, which provided evidence that the SEC-coding sequences were located in a composite pathogenicity island that was previously described in the S. epidermidis strain FRI909, called SePI-1/SeCI-1, with 83.8–89.7% nucleotide similarity. Various other plasmids were identified, particularly p_3_95 and p_4_95, which carry antibiotic resistance genes (hsrA and dfrG, respectively), and share homologies with SAP085A and pUSA04-2-SUR11, two plasmids described in S. aureus. Eventually, one complete prophage was identified, ΦSE90, sharing 30 out of 52 coding sequences with the Acinetobacter phage vB_AbaM_IME200. Thus, the SePI-1/SeCI-1 pathogenicity island was identified in two pathogenic strains of S. epidermidis that produced a SEC enterotoxin causing septic shock. These findings suggest the existence of in vivo genetic exchange from S. aureus to S. epidermidis. PMID:29495323

Xylella taiwanensis sp. nov., causing pear leaf scorch disease.

PubMed

Su, C-C; Deng, W-L; Jan, F-J; Chang, C-J; Huang, H; Shih, H-T; Chen, J

2016-11-01

A Gram-stain-negative, nutritionally fastidious bacterium (PLS229T) causing pear leaf scorch was identified in Taiwan and previously grouped into Xylella fastidiosa. Yet, significant variations between PLS229T and Xylellafastidiosa were noted. In this study, PLS229T was evaluated phenotypically and genotypically against representative strains of Xylellafastidiosa, including strains of the currently known subspecies of Xylellafastidiosa, Xylella fastidiosa subsp. multiplex and 'Xylella fastidiosasubsp.pauca'. Because of the difficulty of in vitro culture characterization, emphases were made to utilize the available whole-genome sequence information. The average nucleotide identity (ANI) values, an alternative for DNA-DNA hybridization relatedness, between PLS229T and Xylellafastidiosa were 83.4-83.9 %, significantly lower than the bacterial species threshold of 95 %. In contrast, sequence similarity of 16S rRNA genes was greater than 98 %, higher than the 97 % threshold to justify if two bacterial strains belong to different species. The uniqueness of PLS229T was also evident by observing only about 87 % similarity in the sequence of the 16S-23S internal transcribed spacer (ITS) between PLS229T and strains of Xylellafastidiosa, discovering significant single nucleotide polymorphisms at 18 randomly selected housekeeping gene loci, observing a distinct fatty acid profile for PLS229T compared with Xylellafastidiosa, and PLS229T having different observable phenotypes, such as different susceptibility to antibiotics. A phylogenetic tree derived from 16S rRNA gene sequences showed a distinct PLS229T phyletic lineage positioning it between Xylellafastidiosa and members of the genus Xanthomonas. On the basis of these data, a novel species, Xylella taiwanensis sp. nov. is proposed. The type strain is PLS229T (=BCRC 80915T=JCM 31187T).

Phylogenomic analysis of the species of the Mycobacterium tuberculosis complex demonstrates that Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and Mycobacterium pinnipedii are later heterotypic synonyms of Mycobacterium tuberculosis.

PubMed

Riojas, Marco A; McGough, Katya J; Rider-Riojas, Cristin J; Rastogi, Nalin; Hazbón, Manzour Hernando

2018-01-01

The species within the Mycobacterium tuberculosis Complex (MTBC) have undergone numerous taxonomic and nomenclatural changes, leaving the true structure of the MTBC in doubt. We used next-generation sequencing (NGS), digital DNA-DNA hybridization (dDDH), and average nucleotide identity (ANI) to investigate the relationship between these species. The type strains of Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and Mycobacterium pinnipedii were sequenced via NGS. Pairwise dDDH and ANI comparisons between these, previously sequenced MTBC type strain genomes (including 'Mycobacterium canettii', 'Mycobacterium mungi' and 'Mycobacterium orygis') and M. tuberculosis H37Rv T were performed. Further, all available genome sequences in GenBank for species in or putatively in the MTBC were compared to H37Rv T . Pairwise results indicated that all of the type strains of the species are extremely closely related to each other (dDDH: 91.2-99.2 %, ANI: 99.21-99.92 %), greatly exceeding the respective species delineation thresholds, thus indicating that they belong to the same species. Results from the GenBank genomes indicate that all the strains examined are within the circumscription of H37Rv T (dDDH: 83.5-100 %). We, therefore, formally propose a union of the species of the MTBC as M. tuberculosis. M. africanum, M. bovis, M. caprae, M. microti and M. pinnipedii are reclassified as later heterotypic synonyms of M. tuberculosis. 'M. canettii', 'M. mungi', and 'M. orygis' are classified as strains of the species M. tuberculosis. We further recommend use of the infrasubspecific term 'variant' ('var.') and infrasubspecific designations that generally retain the historical nomenclature associated with the groups or otherwise convey such characteristics, e.g. M. tuberculosis var. bovis.

More Easily Cultivated Than Identified: Classical Isolation With Molecular Identification of Vaginal Bacteria

PubMed Central

Srinivasan, Sujatha; Munch, Matthew M.; Sizova, Maria V.; Fiedler, Tina L.; Kohler, Christina M.; Hoffman, Noah G.; Liu, Congzhou; Agnew, Kathy J.; Marrazzo, Jeanne M.; Epstein, Slava S.; Fredricks, David N.

2016-01-01

Background. Women with bacterial vaginosis (BV) have complex communities of anaerobic bacteria. There are no cultivated isolates of several bacteria identified using molecular methods and associated with BV. It is unclear whether this is due to the inability to adequately propagate these bacteria or to correctly identify them in culture. Methods. Vaginal fluid from 15 women was plated on 6 different media using classical cultivation approaches. Individual isolates were identified by 16S ribosomal RNA (rRNA) gene sequencing and compared with validly described species. Bacterial community profiles in vaginal samples were determined using broad-range 16S rRNA gene polymerase chain reaction and pyrosequencing. Results. We isolated and identified 101 distinct bacterial strains spanning 6 phyla including (1) novel strains with <98% 16S rRNA sequence identity to validly described species, (2) closely related species within a genus, (3) bacteria previously isolated from body sites other than the vagina, and (4) known bacteria formerly isolated from the vagina. Pyrosequencing showed that novel strains Peptoniphilaceae DNF01163 and Prevotellaceae DNF00733 were prevalent in women with BV. Conclusions. We isolated a diverse set of novel and clinically significant anaerobes from the human vagina using conventional approaches with systematic molecular identification. Several previously “uncultivated” bacteria are amenable to conventional cultivation. PMID:27449870

A Unique Capsule Locus in the Newly Designated Actinobacillus pleuropneumoniae Serovar 16 and Development of a Diagnostic PCR Assay.

PubMed

Bossé, Janine T; Li, Yanwen; Sárközi, Rita; Gottschalk, Marcelo; Angen, Øystein; Nedbalcova, Katerina; Rycroft, Andrew N; Fodor, László; Langford, Paul R

2017-03-01

Actinobacillus pleuropneumoniae causes pleuropneumonia, an economically significant lung disease of pigs. Recently, isolates of A. pleuropneumoniae that were serologically distinct from the previously characterized 15 serovars were described, and a proposal was put forward that they comprised a new serovar, serovar 16. Here we used whole-genome sequencing of the proposed serovar 16 reference strain A-85/14 to confirm the presence of a unique capsular polysaccharide biosynthetic locus. For molecular diagnostics, primers were designed from the capsule locus of strain A-85/14, and a PCR was formulated that differentiated serovar 16 isolates from all 15 known serovars and other common respiratory pathogenic/commensal bacteria of pigs. Analysis of the capsule locus of strain A-85/14 combined with the previous serological data show the existence of a sixteenth serovar-designated serovar 16-of A. pleuropneumoniae . Copyright © 2017 Bossé et al.

Erwinia amylovora Novel Plasmid pEI70: Complete Sequence, Biogeography, and Role in Aggressiveness in the Fire Blight Phytopathogen

PubMed Central

Llop, Pablo; Cabrefiga, Jordi; Smits, Theo H. M.; Dreo, Tanja; Barbé, Silvia; Pulawska, Joanna; Bultreys, Alain; Blom, Jochen; Duffy, Brion; Montesinos, Emilio; López, María M.

2011-01-01

Comparative genomics of several strains of Erwinia amylovora, a plant pathogenic bacterium causal agent of fire blight disease, revealed that its diversity is primarily attributable to the flexible genome comprised of plasmids. We recently identified and sequenced in full a novel 65.8 kb plasmid, called pEI70. Annotation revealed a lack of known virulence-related genes, but found evidence for a unique integrative conjugative element related to that of other plant and human pathogens. Comparative analyses using BLASTN showed that pEI70 is almost entirely included in plasmid pEB102 from E. billingiae, an epiphytic Erwinia of pome fruits, with sequence identities superior to 98%. A duplex PCR assay was developed to survey the prevalence of plasmid pEI70 and also that of pEA29, which had previously been described in several E. amylovora strains. Plasmid pEI70 was found widely dispersed across Europe with frequencies of 5–92%, but it was absent in E. amylovora analyzed populations from outside of Europe. Restriction analysis and hybridization demonstrated that this plasmid was identical in at least 13 strains. Curing E. amylovora strains of pEI70 reduced their aggressiveness on pear, and introducing pEI70 into low-aggressiveness strains lacking this plasmid increased symptoms development in this host. Discovery of this novel plasmid offers new insights into the biogeography, evolution and virulence determinants in E. amylovora. PMID:22174857

Erwinia amylovora novel plasmid pEI70: complete sequence, biogeography, and role in aggressiveness in the fire blight phytopathogen.

PubMed

Llop, Pablo; Cabrefiga, Jordi; Smits, Theo H M; Dreo, Tanja; Barbé, Silvia; Pulawska, Joanna; Bultreys, Alain; Blom, Jochen; Duffy, Brion; Montesinos, Emilio; López, María M

2011-01-01

Comparative genomics of several strains of Erwinia amylovora, a plant pathogenic bacterium causal agent of fire blight disease, revealed that its diversity is primarily attributable to the flexible genome comprised of plasmids. We recently identified and sequenced in full a novel 65.8 kb plasmid, called pEI70. Annotation revealed a lack of known virulence-related genes, but found evidence for a unique integrative conjugative element related to that of other plant and human pathogens. Comparative analyses using BLASTN showed that pEI70 is almost entirely included in plasmid pEB102 from E. billingiae, an epiphytic Erwinia of pome fruits, with sequence identities superior to 98%. A duplex PCR assay was developed to survey the prevalence of plasmid pEI70 and also that of pEA29, which had previously been described in several E. amylovora strains. Plasmid pEI70 was found widely dispersed across Europe with frequencies of 5-92%, but it was absent in E. amylovora analyzed populations from outside of Europe. Restriction analysis and hybridization demonstrated that this plasmid was identical in at least 13 strains. Curing E. amylovora strains of pEI70 reduced their aggressiveness on pear, and introducing pEI70 into low-aggressiveness strains lacking this plasmid increased symptoms development in this host. Discovery of this novel plasmid offers new insights into the biogeography, evolution and virulence determinants in E. amylovora.

Genetic analysis among environmental strains of Balamuthia mandrillaris recovered from an artificial lagoon and from soil in Sonora, Mexico.

PubMed

Lares-Jiménez, Luis Fernando; Booton, Gregory C; Lares-Villa, Fernando; Velázquez-Contreras, Carlos Arturo; Fuerst, Paul A

2014-11-01

Since the first report of Balamuthia mandrillaris as a causative agent of granulomatous amoebic encephalitis in humans, the environmental niche of this amoeba was assumed to be restricted to soil and dust. A single isolation from water was recently made independently by us from Northern Mexico. Now we report the isolation of 8 new strains of B. mandrillaris from Mexico. This continues the pattern of an excess of isolates from North America, compared to other parts of the world. All of the new isolates are environmental isolates, 7 from water samples and one from soil. The identity of each isolate was confirmed by PCR and by examining the sequences of the mitochondrial 16S-like rRNA gene. Success in amplification was determined using comparisons of amplifications of DNA from the strain CDC: V039 and the water strain (ITSON-BM1) as positive controls. The DNA sequences of the new isolates were compared to older strains from clinical cases using phylogenetic analysis, showing very high sequence similarity. The similarity among the new isolates and with previous clinical and environmental isolates of B. mandrillaris was also examined using biochemical and immunological studies. High homogeneity of total protein products, and similarity in antigenic moiety among the eight new isolates and two controls was found. Taken together, the molecular and biochemical studies indicate very low levels of genetic variation within B. mandrillaris. Copyright © 2014 Elsevier Inc. All rights reserved.

Polyphasic characterization and genetic relatedness of low-virulence and virulent Listeria monocytogenes isolates

PubMed Central

2012-01-01

Background Currently, food regulatory authorities consider all Listeria monocytogenes isolates as equally virulent. However, an increasing number of studies demonstrate extensive variations in virulence and pathogenicity of L. monocytogenes strains. Up to now, there is no comprehensive overview of the population genetic structure of L. monocytogenes taking into account virulence level. We have previously demonstrated that different low-virulence strains exhibit the same mutations in virulence genes suggesting that they could have common evolutionary pathways. New low-virulence strains were identified and assigned to phenotypic and genotypic Groups using cluster analysis. Pulsed-field gel electrophoresis, virulence gene sequencing and multi-locus sequence typing analyses were performed to study the genetic relatedness and the population structure between the studied low-virulence isolates and virulent strains. Results These methods showed that low-virulence strains are widely distributed in the two major lineages, but some are also clustered according to their genetic mutations. These analyses showed that low-virulence strains initially grouped according to their lineage, then to their serotypes and after which, they lost their virulence suggesting a relatively recent emergence. Conclusions Loss of virulence in lineage II strains was related to point mutation in a few virulence genes (prfA, inlA, inlB, plcA). These strains thus form a tightly clustered, monophyletic group with limited diversity. In contrast, low-virulence strains of lineage I were more dispersed among the virulence strains and the origin of their loss of virulence has not been identified yet, even if some strains exhibited different mutations in prfA or inlA. PMID:23267677

The 14alpha-Demethylasse(CYP51A1) Gene is Overexpressed in Venturia inaequalis Strains Resistant to Myclobutanil.

PubMed

Schnabel, G; Jones, A L

2001-01-01

ABSTRACT We identified the cytochrome P450 sterol 14alpha-demethylase (CYP51A1) gene from Venturia inaequalis and optional insertions located upstream from CYP51A1 and evaluated their potential role in conferring resistance to the sterol demethylation-inhibitor (DMI) fungicide my-clobutanil. The CYP51A1 gene was completely sequenced from one my-clobutanil sensitive (S) and two myclobutanil-resistant (R) strains. No nucleotide variation was found when the three sequences were aligned. Allele-specific polymerase chain reaction (PCR) analysis indicated that a previously described single base pair mutation that correlated with resistance to DMI fungicides in strains of other filamentous fungi was absent in 19 S and 32 R strains of V. inaequalis from Michigan and elsewhere. The sequencing results and PCR analyses suggest that resistance in these strains was not due to a mutation in the sterol demethylase target site for DMI fungicides. Expression of CYP51A1 was determined for strains from an orchard that had never been sprayed with DMI fungicides (baseline orchard), and the data provided a reference for evaluating the expression of strains collected from a research orchard and from three commercial Michigan apple orchards with a long history of DMI use and a high frequency of R strains. Overexpression of CYP51A1 was significantly higher in 9 of 11 R strains from the research orchard than in S strains from the baseline orchard. The high expression was correlated with the presence of a 553-bp insertion located upstream of CYP51A1. Overexpression of the CYP51A1 gene was also detected in eight of eight, five of nine, and nine of nine R strains from three commercial orchards, but the insertion was not detected in the majority of these strains. The results suggest that overexpression of the target-site CYP51A1 gene is an important mechanism of resistance in some field resistant strains of V. inaequalis, but other mechanisms of resistance also appear to exist.

Genome sequencing and analysis of a type A Clostridium perfringens isolate from a case of bovine clostridial abomasitis.

PubMed

Nowell, Victoria J; Kropinski, Andrew M; Songer, J Glenn; MacInnes, Janet I; Parreira, Valeria R; Prescott, John F

2012-01-01

Clostridium perfringens is a common inhabitant of the avian and mammalian gastrointestinal tracts and can behave commensally or pathogenically. Some enteric diseases caused by type A C. perfringens, including bovine clostridial abomasitis, remain poorly understood. To investigate the potential basis of virulence in strains causing this disease, we sequenced the genome of a type A C. perfringens isolate (strain F262) from a case of bovine clostridial abomasitis. The ∼3.34 Mbp chromosome of C. perfringens F262 is predicted to contain 3163 protein-coding genes, 76 tRNA genes, and an integrated plasmid sequence, Cfrag (∼18 kb). In addition, sequences of two complete circular plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), and two incomplete plasmid fragments, pF262A (48.5 kb) and pF262B (50.0 kb), were identified. Comparison of the chromosome sequence of C. perfringens F262 to complete C. perfringens chromosomes, plasmids and phages revealed 261 unique genes. No novel toxin genes related to previously described clostridial toxins were identified: 60% of the 261 unique genes were hypothetical proteins. There was a two base pair deletion in virS, a gene reported to encode the main sensor kinase involved in virulence gene activation. Despite this frameshift mutation, C. perfringens F262 expressed perfringolysin O, alpha-toxin and the beta2-toxin, suggesting that another regulation system might contribute to the pathogenicity of this strain. Two complete plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), unique to this strain of C. perfringens were identified.

Genome Sequencing and Analysis of a Type A Clostridium perfringens Isolate from a Case of Bovine Clostridial Abomasitis

PubMed Central

Nowell, Victoria J.; Kropinski, Andrew M.; Songer, J. Glenn; MacInnes, Janet I.; Parreira, Valeria R.; Prescott, John F.

2012-01-01

Clostridium perfringens is a common inhabitant of the avian and mammalian gastrointestinal tracts and can behave commensally or pathogenically. Some enteric diseases caused by type A C. perfringens, including bovine clostridial abomasitis, remain poorly understood. To investigate the potential basis of virulence in strains causing this disease, we sequenced the genome of a type A C. perfringens isolate (strain F262) from a case of bovine clostridial abomasitis. The ∼3.34 Mbp chromosome of C. perfringens F262 is predicted to contain 3163 protein-coding genes, 76 tRNA genes, and an integrated plasmid sequence, Cfrag (∼18 kb). In addition, sequences of two complete circular plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), and two incomplete plasmid fragments, pF262A (48.5 kb) and pF262B (50.0 kb), were identified. Comparison of the chromosome sequence of C. perfringens F262 to complete C. perfringens chromosomes, plasmids and phages revealed 261 unique genes. No novel toxin genes related to previously described clostridial toxins were identified: 60% of the 261 unique genes were hypothetical proteins. There was a two base pair deletion in virS, a gene reported to encode the main sensor kinase involved in virulence gene activation. Despite this frameshift mutation, C. perfringens F262 expressed perfringolysin O, alpha-toxin and the beta2-toxin, suggesting that another regulation system might contribute to the pathogenicity of this strain. Two complete plasmids, pF262C (4.8 kb) and pF262D (9.1 kb), unique to this strain of C. perfringens were identified. PMID:22412860

DNA-Sequence Based Typing of the Cronobacter Genus Using MLST, CRISPR-cas Array and Capsular Profiling

PubMed Central

Ogrodzki, Pauline; Forsythe, Stephen J.

2017-01-01

The Cronobacter genus is composed of seven species, within which a number of pathovars have been described. The most notable infections by Cronobacter spp. are of infants through the consumption of contaminated infant formula. The description of the genus has greatly improved in recent years through DNA sequencing techniques, and this has led to a robust means of identification. However some species are highly clonal and this limits the ability to discriminate between unrelated strains by some methods of genotyping. This article updates the application of three genotyping methods across the Cronobacter genus. The three genotyping methods were multilocus sequence typing (MLST), capsular profiling of the K-antigen and colanic acid (CA) biosynthesis regions, and CRISPR-cas array profiling. A total of 1654 MLST profiled and 286 whole genome sequenced strains, available by open access at the PubMLST Cronobacter database, were used this analysis. The predominance of C. sakazakii and C. malonaticus in clinical infections was confirmed. The majority of clinical strains being in the C. sakazakii clonal complexes (CC) 1 and 4, sequence types (ST) 8 and 12 and C. malonaticus ST7. The capsular profile K2:CA2, previously proposed as being strongly associated with C. sakazakii and C. malonaticus isolates from severe neonatal infections, was also found in C. turicensis, C. dublinensis and C. universalis. The majority of CRISPR-cas types across the genus was the I-E (Ecoli) type. Some strains of C. dublinensis and C. muytjensii encoded the I-F (Ypseudo) type, and others lacked the cas gene loci. The significance of the expanding profiling will be of benefit to researchers as well as governmental and industrial risk assessors. PMID:29033918

Identification, Characterization, and Recombinant Expression of Epidermicin NI01, a Novel Unmodified Bacteriocin Produced by Staphylococcus epidermidis That Displays Potent Activity against Staphylococci

PubMed Central

Sandiford, Stephanie

2012-01-01

We describe the discovery, purification, characterization, and expression of an antimicrobial peptide, epidermicin NI01, which is an unmodified bacteriocin produced by Staphylococcus epidermidis strain 224. It is a highly cationic, hydrophobic, plasmid-encoded peptide that exhibits potent antimicrobial activity toward a wide range of pathogenic Gram-positive bacteria including methicillin-resistant Staphylococcus aureus (MRSA), enterococci, and biofilm-forming S. epidermidis strains. Purification of the peptide was achieved using a combination of hydrophobic interaction, cation exchange, and high-performance liquid chromatography (HPLC). Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) analysis yielded a molecular mass of 6,074 Da, and partial sequence data of the peptide were elucidated using a combination of tandem mass spectrometry (MS/MS) and de novo sequencing. The draft genome sequence of the producing strain was obtained using 454 pyrosequencing technology, thus enabling the identification of the structural gene using the de novo peptide sequence data previously obtained. Epidermicin NI01 contains 51 residues with four tryptophan and nine lysine residues, and the sequence showed approximately 50% identity to peptides lacticin Z, lacticin Q, and aureocin A53, all of which belong to a new family of unmodified type II-like bacteriocins. The peptide is active in the nanomolar range against S. epidermidis, MRSA isolates, and vancomycin-resistant enterococci. Other unique features displayed by epidermicin include a high degree of protease stability and the ability to retain antimicrobial activity over a pH range of 2 to 10, and exposure to the peptide does not result in development of resistance in susceptible isolates. In this study we also show the structural gene alone can be cloned into Escherichia coli strain BL21(DE3), and expression yields active peptide. PMID:22155816

Tenacibaculum aestuarii sp. nov., isolated from a tidal flat sediment in Korea.

PubMed

Jung, Seo-Youn; Oh, Tae-Kwang; Yoon, Jung-Hoon

2006-07-01

A novel Tenacibaculum-like bacterial strain, SMK-4(T), was isolated from a tidal flat sediment in Korea. Strain SMK-4(T) was Gram-negative, pale yellow-pigmented and rod-shaped. It grew optimally at 30-37 degrees C and in the presence of 2-3 % (w/v) NaCl. It contained MK-6 as the predominant menaquinone and iso-C(15 : 0), iso-C(16 : 0) 3-OH and C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH as the major fatty acids (>10 % of total fatty acids). The DNA G+C content was 33.6 mol%. Phylogenetic trees based on 16S rRNA gene sequences showed that strain SMK-4(T) fell within the evolutionary radiation encompassed by the genus Tenacibaculum. Strain SMK-4(T) exhibited 16S rRNA gene sequence similarity levels of 95.2-98.6 % with respect to the type strains of recognized Tenacibaculum species. DNA-DNA relatedness levels and differential phenotypic properties made it possible to categorize strain SMK-4(T) as a species that is separate from previously described Tenacibaculum species. On the basis of phenotypic properties and phylogenetic and genetic distinctiveness, strain SMK-4(T) (=KCTC 12569(T)=JCM 13491(T)) should be classified as a novel Tenacibaculum species, for which the name Tenacibaculum aestuarii sp. nov. is proposed.

Discrimination of probiotic Lactobacillus strains for poultry by repetitive sequenced-based PCR fingerprinting.

PubMed

Lee, Chin Mei; Sieo, Chin Chin; Cheah, Yoke-Kqueen; Abdullah, Norhani; Ho, Yin Wan

2012-02-01

Four repetitive element sequence-based polymerase chain reaction (rep-PCR) methods, namely repetitive extragenic palindromic PCR (REP-PCR), enterobacterial repetitive intergenic consensus PCR (ERIC-PCR), polytrinucleotide (GTG)₅ -PCR and BOX-PCR, were evaluated for the molecular differentiation of 12 probiotic Lactobacillus strains previously isolated from the gastrointestinal tract of chickens and used as a multistrain probiotic. This study represents the first analysis of the comparative efficacy of these four rep-PCR methods and their combination (composite rep-PCR) in the molecular typing of Lactobacillus strains based on a discriminatory index (D). Species-specific and strain-specific profiles were observed from rep-PCR. From the numerical analysis of composite rep-PCR, BOX-PCR, (GTG)₅ -PCR, REP-PCR and ERIC-PCR, D values of 0.9118, 0.9044, 0.8897, 0.8750 and 0.8529 respectively were obtained. Composite rep-PCR analysis was the most discriminative method, with eight Lactobacillus strains, namely L. brevis ATCC 14869(T) , L. reuteri C 10, L. reuteri ATCC 23272(T) , L. gallinarum ATCC 33199(T) , L. salivarius ATCC 11741(T) , L. salivarius I 24, L. panis JCM 11053(T) and L. panis C 17, being differentiated at the strain level. Composite rep-PCR analysis is potentially a useful fingerprinting method to discriminate probiotic Lactobacillus strains isolated from the gastrointestinal tract of chickens. Copyright © 2011 Society of Chemical Industry.

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

Siegel, A.

Previous work had demonstrated the presence of a unique low-molecular-weight RNA component (LMC) in extracts of tobacco mosaic virus (TMV) infected tissue. Enough of this component has been isolated during the past year to ascertain that it has a molecular weight of 250,000 daltons and that it acts as an in vitro messenger for the synthesis of TMV capsid protein. Thus, we conclude that at least one monocistronic messenger RNA for a virion coded product is generated during TMV infection. Strains of TMV were classified according to nucleotide sequence homology of their RNAs. The strains fall into groups by themore » test employed. No differences were observed between strains within a group, whereas no homology was detected between groups. Using this information, it was possible, in part, to relate differences in capsid protein amino acid sequences to the degree of nomology of their nucleotide coding sequences. A study was initiated into the Pot Y virus group infection mechanism. In contrast to TMV infection, it was determined that for both tobacco etch and potato virus Y that: viral RNA synthesis is inhibited by actinomycin B and synthesis by virus-related proteins is inhibited by chloramphenicol.« less

Characterization of regionally associated feline immunodeficiency virus (FIV) in bobcats (Lynx rufus).

PubMed

Lagana, Danielle M; Lee, Justin S; Lewis, Jesse S; Bevins, Sarah N; Carver, Scott; Sweanor, Linda L; McBride, Roy; McBride, Caleb; Crooks, Kevin R; VandeWoude, Sue

2013-07-01

Feline immunodeficiency virus (FIV) classically infects felid species with highly divergent species-specific FIVs. However, recent studies have detected an FIV strain infecting both bobcats (Lynx rufus) and pumas (Puma concolor) in California and Florida. To further investigate this observation, we evaluated FIV from bobcats in Florida (n=25) and Colorado (n=80) between 2008 and 2011. Partial viral sequences from five Florida bobcats cluster with previously published sequences from Florida panthers. We did not detect FIV in Colorado bobcats.

Genome sequence of a novel H14N7 subtype influenza A virus isolated from a blue-winged teal (Anas discors) harvested in Texas, USA

USGS Publications Warehouse

Ramey, Andy M.; Reeves, Andrew; Poulson, Rebecca L.; Carter, Deborah L.; Davis-Fields, Nicholas; Stallknecht, David E.

2016-01-01

We report here the complete genome sequence of a novel H14N7 subtype influenza A virus (IAV) isolated from a blue-winged teal (Anas discors) harvested in Texas, USA. The genomic characteristics of this IAV strain with a previously undetected subtype combination suggest recent viral evolution within the New World wild-bird IAV reservoir.                   

Rapid identification of oral Actinomyces species cultivated from subgingival biofilm by MALDI-TOF-MS

PubMed Central

Stingu, Catalina S.; Borgmann, Toralf; Rodloff, Arne C.; Vielkind, Paul; Jentsch, Holger; Schellenberger, Wolfgang; Eschrich, Klaus

2015-01-01

Background Actinomyces are a common part of the residential flora of the human intestinal tract, genitourinary system and skin. Isolation and identification of Actinomyces by conventional methods is often difficult and time consuming. In recent years, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has become a rapid and simple method to identify bacteria. Objective The present study evaluated a new in-house algorithm using MALDI-TOF-MS for rapid identification of different species of oral Actinomyces cultivated from subgingival biofilm. Design Eleven reference strains and 674 clinical strains were used in this study. All the strains were preliminarily identified using biochemical methods and then subjected to MALDI-TOF-MS analysis using both similarity-based analysis and classification methods (support vector machine [SVM]). The genotype of the reference strains and of 232 clinical strains was identified by sequence analysis of the 16S ribosomal RNA (rRNA). Results The sequence analysis of the 16S rRNA gene of all references strains confirmed their previous identification. The MALDI-TOF-MS spectra obtained from the reference strains and the other clinical strains undoubtedly identified as Actinomyces by 16S rRNA sequencing were used to create the mass spectra reference database. Already a visual inspection of the mass spectra of different species reveals both similarities and differences. However, the differences between them are not large enough to allow a reliable differentiation by similarity analysis. Therefore, classification methods were applied as an alternative approach for differentiation and identification of Actinomyces at the species level. A cross-validation of the reference database representing 14 Actinomyces species yielded correct results for all species which were represented by more than two strains in the database. Conclusions Our results suggest that a combination of MALDI-TOF-MS with powerful classification algorithms, such as SVMs, provide a useful tool for the differentiation and identification of oral Actinomyces. PMID:25597306

Genetic Characterization of Circulating 2015 A(H1N1)pdm09 Influenza Viruses from Eastern India

PubMed Central

Mukherjee, Anupam; Nayak, Mukti Kant; Dutta, Shanta; Panda, Samiran; Satpathi, Biswa Ranjan; Chawla-Sarkar, Mamta

2016-01-01

In 2015, the swine derived A(H1N1)pdm09 pandemic strain outbreak became widespread throughout the different states of India. The reported cases and deaths in 2015 surpassed the previous years with more than 39000 laboratory confirmed cases and a death toll of more than 2500 people. There are relatively limited complete genetic sequences available for this virus from Asian countries. In this study, we describe the full genome analysis of influenza 2015 A(H1N1)pdm09 viruses isolated from West Bengal between January through December 2015. The phylogenetic analysis of the haemagglutinin sequence revealed clustering with globally circulating strains of genogroup 6B. This was further confirmed by the constructed concatenated tree using all eight complete gene segments of Kolkata A(H1N1)pdm09 isolates with the other strains from different timeline and lineages. A study from Massachusetts Institute of Technology (MIT) in 2015 reported novel mutations T200A and D225N in haemagglutinin gene of a 2014 Indian strain (A/India/6427/2014). However, in all the pandemic strains of 2014–2015 reported from India, so far including A(H1N1)pdm09 strains from Kolkata, D225N mutation was not observed, though the T200A mutation was found to be conserved. Neuraminidase gene of the analyzed strains did not show any oseltamivir resistant mutation H275Y suggesting continuation of Tamiflu® as drug of choice. The amino acid sequences of the all gene segments from 2015 A(H1N1)pdm09 isolates identified several new mutations compared to the 2009 A(H1N1)pdm09 strains, which may have contributed towards enhanced virulence, compared to 2009 A(H1N1)pdm09 strains. PMID:27997573

Genetic Characterization of Circulating 2015 A(H1N1)pdm09 Influenza Viruses from Eastern India.

PubMed

Mukherjee, Anupam; Nayak, Mukti Kant; Dutta, Shanta; Panda, Samiran; Satpathi, Biswa Ranjan; Chawla-Sarkar, Mamta

2016-01-01

In 2015, the swine derived A(H1N1)pdm09 pandemic strain outbreak became widespread throughout the different states of India. The reported cases and deaths in 2015 surpassed the previous years with more than 39000 laboratory confirmed cases and a death toll of more than 2500 people. There are relatively limited complete genetic sequences available for this virus from Asian countries. In this study, we describe the full genome analysis of influenza 2015 A(H1N1)pdm09 viruses isolated from West Bengal between January through December 2015. The phylogenetic analysis of the haemagglutinin sequence revealed clustering with globally circulating strains of genogroup 6B. This was further confirmed by the constructed concatenated tree using all eight complete gene segments of Kolkata A(H1N1)pdm09 isolates with the other strains from different timeline and lineages. A study from Massachusetts Institute of Technology (MIT) in 2015 reported novel mutations T200A and D225N in haemagglutinin gene of a 2014 Indian strain (A/India/6427/2014). However, in all the pandemic strains of 2014-2015 reported from India, so far including A(H1N1)pdm09 strains from Kolkata, D225N mutation was not observed, though the T200A mutation was found to be conserved. Neuraminidase gene of the analyzed strains did not show any oseltamivir resistant mutation H275Y suggesting continuation of Tamiflu® as drug of choice. The amino acid sequences of the all gene segments from 2015 A(H1N1)pdm09 isolates identified several new mutations compared to the 2009 A(H1N1)pdm09 strains, which may have contributed towards enhanced virulence, compared to 2009 A(H1N1)pdm09 strains.

Characteristics of emerging human-pathogenic Escherichia coli O26:H11 strains isolated in France between 2010 and 2013 and carrying the stx2d gene only.

PubMed

Delannoy, Sabine; Mariani-Kurkdjian, Patricia; Bonacorsi, Stephane; Liguori, Sandrine; Fach, Patrick

2015-02-01

Strains of Escherichia coli O26:H11 that were positive for stx2 alone (n = 23), which were not epidemiologically related or part of an outbreak, were isolated from pediatric patients in France between 2010 and 2013. We were interested in comparing these strains with the new highly virulent stx2a-positive E. coli O26 clone sequence type 29 (ST29) that has emerged recently in Europe, and we tested them by multilocus sequence typing (MLST), stx2 subtyping, clustered regularly interspaced short palindromic repeat (CRISPR) sequencing, and plasmid (ehxA, katP, espP, and etpD) and chromosomal (Z2098, espK, and espV) virulence gene profiling. We showed that 16 of the 23 strains appeared to correspond to this new clone, but the characteristics of 12 strains differed significantly from the previously described characteristics, with negative results for both plasmid and chromosomal genetic markers. These 12 strains exhibited a ST29 genotype and related CRISPR arrays (CRISPR2a alleles 67 or 71), suggesting that they evolved in a common environment. This finding was corroborated by the presence of stx2d in 7 of the 12 ST29 strains. This is the first time that E. coli O26:H11 carrying stx2d has been isolated from humans. This is additional evidence of the continuing evolution of virulent Shiga toxin-producing E. coli (STEC) O26 strains. A new O26:H11 CRISPR PCR assay, SP_O26_E, has been developed for detection of these 12 particular ST29 strains of E. coli O26:H11. This test is useful to better characterize the stx2-positive O26:H11 clinical isolates, which are associated with severe clinical outcomes such as bloody diarrhea and hemolytic uremic syndrome. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Comparative genomics provides new insights into the diversity, physiology, and sexuality of the only industrially exploited tremellomycete: Phaffia rhodozyma

DOE PAGES

Bellora, Nicolas; Moline, Martin; David-Palma, Marcia; ...

2016-11-09

The class Tremellomycete (Agaricomycotina) encompasses more than 380 fungi. Although there are a few edible Tremella spp., the only species with current biotechnological use is the astaxanthin-producing yeast Phaffia rhodozyma (Cystofilobasidiales). Besides astaxanthin, a carotenoid pigment with potent antioxidant activity and great value for aquaculture and pharmaceutical industries, P. rhodozyma possesses multiple exceptional traits of fundamental and applied interest. The aim of this study was to obtain, and analyze two new genome sequences of representative strains from the northern (CBS 7918 T, the type strain) and southern hemispheres (CRUB 1149) and compre them to a previously published genome sequence (strainmore » CBS 6938). Furthermore, photoprotection and antioxidant related genes, as well as genes involved in sexual reproduction were analyzed.« less

Comparative genomics provides new insights into the diversity, physiology, and sexuality of the only industrially exploited tremellomycete: Phaffia rhodozyma

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

Bellora, Nicolas; Moline, Martin; David-Palma, Marcia

The class Tremellomycete (Agaricomycotina) encompasses more than 380 fungi. Although there are a few edible Tremella spp., the only species with current biotechnological use is the astaxanthin-producing yeast Phaffia rhodozyma (Cystofilobasidiales). Besides astaxanthin, a carotenoid pigment with potent antioxidant activity and great value for aquaculture and pharmaceutical industries, P. rhodozyma possesses multiple exceptional traits of fundamental and applied interest. The aim of this study was to obtain, and analyze two new genome sequences of representative strains from the northern (CBS 7918 T, the type strain) and southern hemispheres (CRUB 1149) and compre them to a previously published genome sequence (strainmore » CBS 6938). Furthermore, photoprotection and antioxidant related genes, as well as genes involved in sexual reproduction were analyzed.« less

Isolation and genetic characterization of Aurantimonas and Methylobacterium strains from stems of hypernodulated soybeans.

PubMed

Anda, Mizue; Ikeda, Seishi; Eda, Shima; Okubo, Takashi; Sato, Shusei; Tabata, Satoshi; Mitsui, Hisayuki; Minamisawa, Kiwamu

2011-01-01

The aims of this study were to isolate Aurantimonas and Methylobacterium strains that responded to soybean nodulation phenotypes and nitrogen fertilization rates in a previous culture-independent analysis (Ikeda et al. ISME J. 4:315-326, 2010). Two strategies were adopted for isolation from enriched bacterial cells prepared from stems of field-grown, hypernodulated soybeans: PCR-assisted isolation for Aurantimonas and selective cultivation for Methylobacterium. Thirteen of 768 isolates cultivated on Nutrient Agar medium were identified as Aurantimonas by colony PCR specific for Aurantimonas and 16S rRNA gene sequencing. Meanwhile, among 187 isolates on methanol-containing agar media, 126 were identified by 16S rRNA gene sequences as Methylobacterium. A clustering analysis (>99% identity) of the 16S rRNA gene sequences for the combined datasets of the present and previous studies revealed 4 and 8 operational taxonomic units (OTUs) for Aurantimonas and Methylobacterium, respectively, and showed the successful isolation of target bacteria for these two groups. ERIC- and BOX-PCR showed the genomic uniformity of the target isolates. In addition, phylogenetic analyses of Aurantimonas revealed a phyllosphere-specific cluster in the genus. The isolates obtained in the present study will be useful for revealing unknown legume-microbe interactions in relation to the autoregulation of nodulation.

Conserved hypothetical protein Rv1977 in Mycobacterium tuberculosis strains contains sequence polymorphisms and might be involved in ongoing immune evasion.

PubMed

Jiang, Yi; Liu, Haican; Wang, Xuezhi; Li, Guilian; Qiu, Yan; Dou, Xiangfeng; Wan, Kanglin

2015-01-01

Host immune pressure and associated parasite immune evasion are key features of host-pathogen co-evolution. A previous study showed that human T cell epitopes of Mycobacterium tuberculosis are evolutionarily hyperconserved and thus it was deduced that M. tuberculosis lacks antigenic variation and immune evasion. Here, we selected 151 clinical Mycobacterium tuberculosis isolates from China, amplified gene encoding Rv1977 and compared the sequences. The results showed that Rv1977, a conserved hypothetical protein, is not conserved in M. tuberculosis strains and there are polymorphisms existed in the protein. Some mutations, especially one frameshift mutation, occurred in the antigen Rv1977, which is uncommon in M.tb strains and may lead to the protein function altering. Mutations and deletion in the gene all affect one of three T cell epitopes and the changed T cell epitope contained more than one variable position, which may suggest ongoing immune evasion.

RECOVIR Software for Identifying Viruses

NASA Technical Reports Server (NTRS)

Chakravarty, Sugoto; Fox, George E.; Zhu, Dianhui

2013-01-01

Most single-stranded RNA (ssRNA) viruses mutate rapidly to generate a large number of strains with highly divergent capsid sequences. Determining the capsid residues or nucleotides that uniquely characterize these strains is critical in understanding the strain diversity of these viruses. RECOVIR (an acronym for "recognize viruses") software predicts the strains of some ssRNA viruses from their limited sequence data. Novel phylogenetic-tree-based databases of protein or nucleic acid residues that uniquely characterize these virus strains are created. Strains of input virus sequences (partial or complete) are predicted through residue-wise comparisons with the databases. RECOVIR uses unique characterizing residues to identify automatically strains of partial or complete capsid sequences of picorna and caliciviruses, two of the most highly diverse ssRNA virus families. Partition-wise comparisons of the database residues with the corresponding residues of more than 300 complete and partial sequences of these viruses resulted in correct strain identification for all of these sequences. This study shows the feasibility of creating databases of hitherto unknown residues uniquely characterizing the capsid sequences of two of the most highly divergent ssRNA virus families. These databases enable automated strain identification from partial or complete capsid sequences of these human and animal pathogens.

Genetic heterogeneity of L-Zagreb mumps virus vaccine strain

PubMed Central

Kosutic-Gulija, Tanja; Forcic, Dubravko; Šantak, Maja; Ramljak, Ana; Mateljak-Lukacevic, Sanja; Mazuran, Renata

2008-01-01

Background The most often used mumps vaccine strains Jeryl Lynn (JL), RIT4385, Urabe-AM9, L-Zagreb and L-3 differ in immunogenicity and reactogenicity. Previous analyses showed that JL, Urabe-AM9 and L-3 are genetically heterogeneous. Results We identified the heterogeneity of L-Zagreb throughout the entire genome. Two major variants were defined: variant A being identical to the consensus sequence of viral seeds and vaccine(s) and variant B which differs from variant A in three nucleotide positions. The difference between viral variants in L-Zagreb strain is insufficient for distinct viral strains to be defined. We demonstrated that proportion of variants in L-Zagreb viral population depends on cell substrate used for viral replication in vitro and in vivo. Conclusion L-Zagreb strain should be considered as a single strain composed of at least two variant viral genomes. PMID:18616793

Methanohalophilus zhilinae sp. nov., an alkaliphilic, halophilic, methylotrophic methanogen

NASA Technical Reports Server (NTRS)

Mathrani, I. M.; Boone, D. R.; Mah, R. A.; Fox, G. E.; Lau, P. P.

1988-01-01

Methanohalophilus zhilinae, a new alkaliphilic, halophilic, methylotrophic species of methanogenic bacteria, is described. Strain WeN5T (T = type strain) from Bosa Lake of the Wadi el Natrun in Egypt was designated the type strain and was further characterized. This strain was nonmotile, able to catabolize dimethylsulfide, and able to grow in medium with a methyl group-containing substrate (such as methanol or trimethylamine) as the sole organic compound added. Sulfide (21 mM) inhibited cultures growing on trimethylamine. The antibiotic susceptibility pattern of strain WeN5T was typical of the pattern for archaeobacteria, and the guanine-plus-cytosine content of the deoxyribonucleic acid was 38 mol%. Characterization of the 16S ribosomal ribonucleic acid sequence indicated that strain WeN5T is phylogenetically distinct from members of previously described genera other than Methanohalophilus and supported the partition of halophilic methanogens into their own genus.

Partial gene sequences for the A subunit of methyl-coenzyme M reductase (mcrI) as a phylogenetic tool for the family Methanosarcinaceae

NASA Technical Reports Server (NTRS)

Springer, E.; Sachs, M. S.; Woese, C. R.; Boone, D. R.

1995-01-01

Representatives of the family Methanosarcinaceae were analyzed phylogenetically by comparing partial sequences of their methyl-coenzyme M reductase (mcrI) genes. A 490-bp fragment from the A subunit of the gene was selected, amplified by the PCR, cloned, and sequenced for each of 25 strains belonging to the Methanosarcinaceae. The sequences obtained were aligned with the corresponding portions of five previously published sequences, and all of the sequences were compared to determine phylogenetic distances by Fitch distance matrix methods. We prepared analogous trees based on 16S rRNA sequences; these trees corresponded closely to the mcrI trees, although the mcrI sequences of pairs of organisms had 3.01 +/- 0.541 times more changes than the respective pairs of 16S rRNA sequences, suggesting that the mcrI fragment evolved about three times more rapidly than the 16S rRNA gene. The qualitative similarity of the mcrI and 16S rRNA trees suggests that transfer of genetic information between dissimilar organisms has not significantly affected these sequences, although we found inconsistencies between some mcrI distances that we measured and and previously published DNA reassociation data. It is unlikely that multiple mcrI isogenes were present in the organisms that we examined, because we found no major discrepancies in multiple determinations of mcrI sequences from the same organism. Our primers for the PCR also match analogous sites in the previously published mcrII sequences, but all of the sequences that we obtained from members of the Methanosarcinaceae were more closely related to mcrI sequences than to mcrII sequences, suggesting that members of the Methanosarcinaceae do not have distinct mcrII genes.

Clonal deleted latent membrane protein 1 variants of Epstein-Barr virus are predominant in European extranodal NK/T lymphomas and disappear during successful treatment.

PubMed

Halabi, Mohamad Adnan; Jaccard, Arnaud; Moulinas, Rémi; Bahri, Racha; Al Mouhammad, Hazar; Mammari, Nour; Feuillard, Jean; Ranger-Rogez, Sylvie

2016-08-15

Extranodal natural killer/T-cell lymphomas (NK/TL), rare in Europe, are Epstein-Barr virus (EBV) associated lymphomas with poor outcomes. Here, we determined the virus type and analyzed the EBV latent membrane protein-1 (LMP1) gene sequence in NK/TL from French patients. Six clones of viral LMP1 were sequenced by Sanger technology in blood from 13 patients before treatment with an l-asparaginase based regimen and, for 8 of them, throughout the treatment. Blood LMP1 sequences from 21 patients without any known malignancy were tested as controls. EBV Type A was identified for 11/13 patients and for all controls. Before treatment, a clonal LMP1 gene containing a 30 bp deletion (del30) was found in 46.1% of NK/TL and only in 4.8% of controls. Treatment was less effective in these patients who died more rapidly than the others. Patients with a deleted strain evolving toward a wild-type strain during treatment reached complete remission. The LMP1 gene was sequenced by highly sensitive next-generation sequencing technology in five NK/TL nasopharyngeal biopsies, two of them originating from the previous patients. Del30 was present in 100% of the biopsies; two viruses at least coexisted in three biopsies. These results suggest that del30 may be associated with poor prognosis NK/TL and that strain evolution could be used as a potential marker to monitor treatment. © 2016 UICC.

Draft Genome Sequences for Two Metal-Reducing Pelosinus fermentans Strains Isolated from a Cr(VI)-Contaminated Site and for Type Strain R7

PubMed Central

Brown, Steven D.; Podar, Mircea; Klingeman, Dawn M.; Johnson, Courtney M.; Yang, Zamin K.; Utturkar, Sagar M.; Land, Miriam L.; Mosher, Jennifer J.; Hurt, Richard A.; Phelps, Tommy J.; Palumbo, Anthony V.; Arkin, Adam P.; Hazen, Terry C.

2012-01-01

Pelosinus fermentans 16S rRNA gene sequences have been reported from diverse geographical sites since the recent isolation of the type strain. We present the genome sequence of the P. fermentans type strain R7 (DSM 17108) and genome sequences for two new strains with different abilities to reduce iron, chromate, and uranium. PMID:22933770

Identification of 88 regulatory small RNAs in the TIGR4 strain of the human pathogen Streptococcus pneumoniae

PubMed Central

Acebo, Paloma; Martin-Galiano, Antonio J.; Navarro, Sara; Zaballos, Ángel; Amblar, Mónica

2012-01-01

Streptococcus pneumoniae is the main etiological agent of community-acquired pneumonia and a major cause of mortality and morbidity among children and the elderly. Genome sequencing of several pneumococcal strains revealed valuable information about the potential proteins and genetic diversity of this prevalent human pathogen. However, little is known about its transcriptional regulation and its small regulatory noncoding RNAs. In this study, we performed deep sequencing of the S. pneumoniae TIGR4 strain RNome to identify small regulatory RNA candidates expressed in this pathogen. We discovered 1047 potential small RNAs including intragenic, 5′- and/or 3′-overlapping RNAs and 88 small RNAs encoded in intergenic regions. With this approach, we recovered many of the previously identified intergenic small RNAs and identified 68 novel candidates, most of which are conserved in both sequence and genomic context in other S. pneumoniae strains. We confirmed the independent expression of 17 intergenic small RNAs and predicted putative mRNA targets for six of them using bioinformatics tools. Preliminary results suggest that one of these six is a key player in the regulation of competence development. This study is the biggest catalog of small noncoding RNAs reported to date in S. pneumoniae and provides a highly complete view of the small RNA network in this pathogen. PMID:22274957

Comparative genomic analysis identified a mutation related to enhanced heterologous protein production in the filamentous fungus Aspergillus oryzae.

PubMed

Jin, Feng-Jie; Katayama, Takuya; Maruyama, Jun-Ichi; Kitamoto, Katsuhiko

2016-11-01

Genomic mapping of mutations using next-generation sequencing technologies has facilitated the identification of genes contributing to fundamental biological processes, including human diseases. However, few studies have used this approach to identify mutations contributing to heterologous protein production in industrial strains of filamentous fungi, such as Aspergillus oryzae. In a screening of A. oryzae strains that hyper-produce human lysozyme (HLY), we previously isolated an AUT1 mutant that showed higher production of various heterologous proteins; however, the underlying factors contributing to the increased heterologous protein production remained unclear. Here, using a comparative genomic approach performed with whole-genome sequences, we attempted to identify the genes responsible for the high-level production of heterologous proteins in the AUT1 mutant. The comparative sequence analysis led to the detection of a gene (AO090120000003), designated autA, which was predicted to encode an unknown cytoplasmic protein containing an alpha/beta-hydrolase fold domain. Mutation or deletion of autA was associated with higher production levels of HLY. Specifically, the HLY yields of the autA mutant and deletion strains were twofold higher than that of the control strain during the early stages of cultivation. Taken together, these results indicate that combining classical mutagenesis approaches with comparative genomic analysis facilitates the identification of novel genes involved in heterologous protein production in filamentous fungi.

Identification of Group B Streptococcal Sip Protein, Which Elicits Cross-Protective Immunity

PubMed Central

Brodeur, Bernard R.; Boyer, Martine; Charlebois, Isabelle; Hamel, Josée; Couture, France; Rioux, Clément R.; Martin, Denis

2000-01-01

A protein of group B streptococci (GBS), named Sip for surface immunogenic protein, which is distinct from previously described surface proteins, was identified after immunological screening of a genomic library. Immunoblots using a Sip-specific monoclonal antibody indicated that a protein band with an approximate molecular mass of 53 kDa which did not vary in size was present in every GBS strain tested. Representatives of all nine GBS serotypes were included in the panel of strains. Cloning and sequencing of the sip gene revealed an open reading frame of 1,305 nucleotides coding for a polypeptide of 434 amino acid residues, with a calculated pI of 6.84 and molecular mass of 45.5 kDa. Comparison of the nucleotide sequences from six different strains confirmed with 98% identity that the sip gene is highly conserved among GBS isolates. N-terminal amino acid sequencing also indicated the presence of a 25-amino-acid signal peptide which is cleaved in the mature protein. More importantly, immunization with the recombinant Sip protein efficiently protected CD-1 mice against deadly challenges with six GBS strains of serotypes Ia/c, Ib, II/R, III, V, and VI. The data presented in this study suggest that this highly conserved protein induces cross-protective immunity against GBS infections and emphasize its potential as a universal vaccine candidate. PMID:10992461

Sequence analysis of the PIP5K locus in Eimeria maxima provides further evidence for eimerian genome plasticity and segmental organization.

PubMed

Song, B K; Pan, M Z; Lau, Y L; Wan, K L

2014-07-29

Commercial flocks infected by Eimeria species parasites, including Eimeria maxima, have an increased risk of developing clinical or subclinical coccidiosis; an intestinal enteritis associated with increased mortality rates in poultry. Currently, infection control is largely based on chemotherapy or live vaccines; however, drug resistance is common and vaccines are relatively expensive. The development of new cost-effective intervention measures will benefit from unraveling the complex genetic mechanisms that underlie host-parasite interactions, including the identification and characterization of genes encoding proteins such as phosphatidylinositol 4-phosphate 5-kinase (PIP5K). We previously identified a PIP5K coding sequence within the E. maxima genome. In this study, we analyzed two bacterial artificial chromosome clones presenting a ~145-kb E. maxima (Weybridge strain) genomic region spanning the PIP5K gene locus. Sequence analysis revealed that ~95% of the simple sequence repeats detected were located within regions comparable to the previously described feature-rich segments of the Eimeria tenella genome. Comparative sequence analysis with the orthologous E. maxima (Houghton strain) region revealed a moderate level of conserved synteny. Unique segmental organizations and telomere-like repeats were also observed in both genomes. A number of incomplete transposable elements were detected and further scrutiny of these elements in both orthologous segments revealed interesting nesting events, which may play a role in facilitating genome plasticity in E. maxima. The current analysis provides more detailed information about the genome organization of E. maxima and may help to reveal genotypic differences that are important for expression of traits related to pathogenicity and virulence.

Classification of "Pseudomonas azotocolligans" Anderson 1955, 132, in the genus Sphingomonas as Sphingomonas trueperi sp. nov.

PubMed

Kämpfer, P; Denner, E B; Meyer, S; Moore, E R; Busse, H J

1997-04-01

"Pseudomonas azotocolligans" ATCC 12417T (T = type strain), which was described as a diazotrophic bacterium, was reinvestigated to clarify its taxonomic position. 16S ribosomal DNA sequence comparisons demonstrated that this strain clusters phylogenetically with species of the genus Sphingomonas and represents a new species. The results of investigations of the fatty acid patterns, polar lipid profiles, and quinone system supported this placement. The substrate utilization profile and biochemical characteristics displayed no obvious similarity to the characteristics of any previously described species of the genus Sphingomonas. The new name Sphingomonas trueperi is proposed on the basis of these results and previously published data for the G + C content of the genomic DNA and the polyamine pattern.

Aquimarina salinaria sp. nov., a novel algicidal bacterium isolated from a saltpan.

PubMed

Chen, Wen-Ming; Sheu, Fu-Sian; Sheu, Shih-Yi

2012-02-01

A bacterial strain designated antisso-27(T), previously isolated from saltpan in Taiwan while screening for bacteria for algicidal activity, was characterized using the polyphasic taxonomic approach. Strain antisso-27(T) was Gram-negative, aerobic, brownish yellow colored, rod-shaped, non-flagellated and non-gliding. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain antisso-27(T) belonged to the genus Aquimarina within the family Flavobacteriaceae with relatively low sequence similarities of 94.0-96.6% to other valid Aquimarina spp. It contained iso-C(17:0) 3-OH, iso-C(15:0), iso-C(16:0), iso-C(15:1) and iso-C(15:0) 3-OH as the main fatty acids and contained a menaquinone with six isoprene units (MK-6) as the major isoprenoid quinone. Major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, an uncharacterized aminolipid and five uncharacterized phospholipids. Strain antisso-27(T) employed direct mode of algicidal lysis to Chlorella vulgaris strain 211-31; nevertheless, it released an algicidal substance against M. aeruginosa strain MTY01. This is the first study that the Aquimarina species possesses both direct and indirect algicidal activities. On the basis of the phylogenetic and phenotypic data, strain antisso-27(T) should be classified as representing a novel species, for which the name A. salinaria sp. nov. is proposed. The type strain is A. salinaria antisso-27(T) (= BCRC 80080(T) = LMG 25375(T)).

Whole-Genome Characterization and Strain Comparison of VT2f-Producing Escherichia coli Causing Hemolytic Uremic Syndrome

PubMed Central

Michelacci, Valeria; Bondì, Roslen; Gigliucci, Federica; Franz, Eelco; Badouei, Mahdi Askari; Schlager, Sabine; Minelli, Fabio; Tozzoli, Rosangela; Caprioli, Alfredo; Morabito, Stefano

2016-01-01

Verotoxigenic Escherichia coli infections in humans cause disease ranging from uncomplicated intestinal illnesses to bloody diarrhea and systemic sequelae, such as hemolytic uremic syndrome (HUS). Previous research indicated that pigeons may be a reservoir for a population of verotoxigenic E. coli producing the VT2f variant. We used whole-genome sequencing to characterize a set of VT2f-producing E. coli strains from human patients with diarrhea or HUS and from healthy pigeons. We describe a phage conveying the vtx2f genes and provide evidence that the strains causing milder diarrheal disease may be transmitted to humans from pigeons. The strains causing HUS could derive from VT2f phage acquisition by E. coli strains with a virulence genes asset resembling that of typical HUS-associated verotoxigenic E. coli. PMID:27584691

Strain improvement of chymosin-producing strains of Aspergillus niger var. awamori using parasexual recombination.

PubMed

Bodie, E A; Armstrong, G L; Dunn-Coleman, N S

1994-05-01

Parasexual recombination was used to obtain improved chymosin-producing strains and to perform genetic analysis on existing strains. Chlorate resistance was used to select for a variety of spontaneous nitrate assimilation pathway mutations in strains previously improved for chymosin production using classical strain improvement methods including mutation and screening, and selection for 2-deoxyglucose resistance (dgr). Diploids of these improved strains were generated via parasexual recombination and were isolated on selective media by complementation of nitrate assimilation mutations. A preliminary genetic analysis of diploid and haploid segregants indicated that the dgr trait, resulting in overexpression of chymosin, was recessive. Also, mutations in two different dgr genes resulted in an increased level of chymosin production. When these mutations were combined via parasexual recombination, the resulting haploid segregants produced about 15% more chymosin than either parental strain. CHEF gel electrophoresis was used to determine the chromosomal location of the integrated chymosin DNA sequences, and to verify diploidy in one case where the chromosome composition of two haploid parents differed.

Pseudomonas syringae pv. actinidiae (PSA) Isolates from Recent Bacterial Canker of Kiwifruit Outbreaks Belong to the Same Genetic Lineage

PubMed Central

Taratufolo, Maria C.; Cai, Rongman; Almeida, Nalvo F.; Goodman, Tokia; Guttman, David S.; Vinatzer, Boris A.; Balestra, Giorgio M.

2012-01-01

Intercontinental spread of emerging plant diseases is one of the most serious threats to world agriculture. One emerging disease is bacterial canker of kiwi fruit (Actinidia deliciosa and A. chinensis) caused by Pseudomonas syringae pv. actinidiae (PSA). The disease first occurred in China and Japan in the 1980s and in Korea and Italy in the 1990s. A more severe form of the disease broke out in Italy in 2008 and in additional countries in 2010 and 2011 threatening the viability of the global kiwi fruit industry. To start investigating the source and routes of international transmission of PSA, genomes of strains from China (the country of origin of the genus Actinidia), Japan, Korea, Italy and Portugal have been sequenced. Strains from China, Italy, and Portugal have been found to belong to the same clonal lineage with only 6 single nucleotide polymorphisms (SNPs) in 3,453,192 bp and one genomic island distinguishing the Chinese strains from the European strains. Not more than two SNPs distinguish each of the Italian and Portuguese strains from each other. The Japanese and Korean strains belong to a separate genetic lineage as previously reported. Analysis of additional European isolates and of New Zealand isolates exploiting genome-derived markers showed that these strains belong to the same lineage as the Italian and Chinese strains. Interestingly, the analyzed New Zealand strains are identical to European strains at the tested SNP loci but test positive for the genomic island present in the sequenced Chinese strains and negative for the genomic island present in the European strains. Results are interpreted in regard to the possible direction of movement of the pathogen between countries and suggest a possible Chinese origin of the European and New Zealand outbreaks. PMID:22590555

Phylogenetic relationships between some members of the genera Neisseria, Acinetobacter, Moraxella, and Kingella based on partial 16S ribosomal DNA sequence analysis.

PubMed

Enright, M C; Carter, P E; MacLean, I A; McKenzie, H

1994-07-01

We obtained 16S ribosomal DNA (rDNA) sequence data for strains belonging to 11 species of Proteobacteria, including the type strains of Kingella kingae, Neisseria lactamica, Neisseria meningitidis, Moraxella lacunata subsp. lacunata, [Neisseria] ovis, Moraxella catarrhalis, Moraxella osloensis, [Moraxella] phenylpyruvica, and Acinetobacter lwoffii, as well as strains of Neisseria subflava and Acinetobacter calcoaceticus. The data in a distance matrix constructed by comparing the sequences supported the proposal that the genera Acinetobacter and Moraxella and [N.] ovis should be excluded from the family Neisseriaceae. Our results are consistent with hybridization data which suggest that these excluded taxa should be part of a new family, the Moraxellaceae. The strains that we studied can be divided into the following five groups: (i) M. lacunata subsp. lacunata, [N.] ovis, and M. catarrhalis; (ii) M. osloensis; (iii) [M.] phenylpyruvica; (iv) A. calcoaceticus and A. lwoffii; and (v) N. meningitidis, N. subflava, N. lactamica, and K. kingae. We agree with the previous proposal that [N.] ovis should be renamed Moraxella ovis, as this organism is closely related to Moraxella species and not to Neisseria species. The generically misnamed taxon [M.] phenylpyruvica belongs to the proposed family Moraxellaceae, but it is sufficiently different to warrant exclusion from the genus Moraxella. Further work needs to be done to investigate genetically similar species, such as Psychrobacter immobilis, before the true generic position of this organism can be determined. Automated 16S rDNA sequencing with the PCR allows workers to accurately determine phylogenetic relationships between groups of organisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Long-term excretion of vaccine-derived poliovirus by a healthy child.

PubMed

Martín, Javier; Odoom, Kofi; Tuite, Gráinne; Dunn, Glynis; Hopewell, Nicola; Cooper, Gill; Fitzharris, Catherine; Butler, Karina; Hall, William W; Minor, Philip D

2004-12-01

A child was found to be excreting type 1 vaccine-derived poliovirus (VDPV) with a 1.1% sequence drift from Sabin type 1 vaccine strain in the VP1 coding region 6 months after he was immunized with oral live polio vaccine. Seventeen type 1 poliovirus isolates were recovered from stools taken from this child during the following 4 months. Contrary to expectation, the child was not deficient in humoral immunity and showed high levels of serum neutralization against poliovirus. Selected virus isolates were characterized in terms of their antigenic properties, virulence in transgenic mice, sensitivity for growth at high temperatures, and differences in nucleotide sequence from the Sabin type 1 strain. The VDPV isolates showed mutations at key nucleotide positions that correlated with the observed reversion to biological properties typical of wild polioviruses. A number of capsid mutations mapped at known antigenic sites leading to changes in the viral antigenic structure. Estimates of sequence evolution based on the accumulation of nucleotide changes in the VP1 coding region detected a "defective" molecular clock running at an apparent faster speed of 2.05% nucleotide changes per year versus 1% shown in previous studies. Remarkably, when compared to several type 1 VDPV strains of different origins, isolates from this child showed a much higher proportion of nonsynonymous versus synonymous nucleotide changes in the capsid coding region. This anomaly could explain the high VP1 sequence drift found and the ability of these virus strains to replicate in the gut for a longer period than expected.

New Hepatitis E Virus Genotype in Camels, the Middle East

PubMed Central

Lau, Susanna K.P.; Teng, Jade L.L.; Tsang, Alan K. L.; Joseph, Marina; Wong, Emily Y.M.; Tang, Ying; Sivakumar, Saritha; Xie, Jun; Bai, Ru; Wernery, Renate; Wernery, Ulrich; Yuen, Kwok-Yung

2014-01-01

In a molecular epidemiology study of hepatitis E virus (HEV) in dromedaries in Dubai, United Arab Emirates, HEV was detected in fecal samples from 3 camels. Complete genome sequencing of 2 strains showed >20% overall nucleotide difference to known HEVs. Comparative genomic and phylogenetic analyses revealed a previously unrecognized HEV genotype. PMID:24856611

Introductions of West Nile Virus Strains to Mexico

PubMed Central

Deardorff, Eleanor; Estrada-Franco, José G.; Brault, Aaron C.; Navarro-Lopez, Roberto; Campomanes-Cortes, Arturo; Paz-Ramirez, Pedro; Solis-Hernandez, Mario; Ramey, Wanichaya N.; Davis, C. Todd; Beasley, David W.C.; Tesh, Robert B.; Barrett, Alan D.T.

2006-01-01

Complete genome sequencing of 22 West Nile virus isolates suggested 2 independent introductions into Mexico. A previously identified mouse-attenuated glycosylation variant was introduced into southern Mexico through the southeastern United States, while a common US genotype appears to have been introduced incrementally into northern Mexico through the southwestern United States. PMID:16494762

Seismic activity and faulting associated with a large underground nuclear explosion

USGS Publications Warehouse

Hamilton, R.M.; McKeown, F.A.; Healy, J.H.

1969-01-01

The 1.1-megaton nuclear test Benham caused movement on previously mapped faults and was followed by a sequence of small earthquakes. These effects were confined to a zone extending not more than 13 kilometers from ground zero; they are apparently related to the release of natural tectonic strain.

RNA sequencing of an nsdC mutant reveals global regulation of secondary metabolic gene clusters in Aspergillus flavus

USDA-ARS?s Scientific Manuscript database

The zinc finger transcription factor nsdC is required for both sexual development and aflatoxin production in the saprophytic fungus Aspergillus flavus. While previous work with an nsdC knockout mutant was conducted in Aspergillus nidulans and A. flavus strain 3357, here we demonstrate perturbations...

PHYLOGENETIC AFFILIATION OF WATER DISTRIBUTION SYSTEM BACTERIAL ISOLATES USING 16S RDNA SEQUENCE ANALYSIS

EPA Science Inventory

In a previously described study, only 15% of the bacterial strains isolated from a water distribution system (WDS) grown on R2A agar were identifiable using fatty acid methyl esthers (FAME) profiling. The lack of success was attributed to the use of fatty acid databases of bacter...

Analysis of differentially expressed genes in two immunologically distinct strains of Eimeria maxima using suppression subtractive hybridization and dot-blot hybridization

PubMed Central

2014-01-01

Background It is well known that different Eimeria maxima strains exhibit significant antigenic variation. However, the genetic basis of these phenotypes remains unclear. Methods Total RNA and mRNA were isolated from unsporulated oocysts of E. maxima strains SH and NT, which were found to have significant differences in immunogenicity in our previous research. Two subtractive cDNA libraries were constructed using suppression subtractive hybridization (SSH) and specific genes were further analyzed by dot-blot hybridization and qRT-PCR analysis. Results A total of 561 clones were selected from both cDNA libraries and the length of the inserted fragments was 0.25–1.0 kb. Dot-blot hybridization revealed a total of 86 differentially expressed clones (63 from strain SH and 23 from strain NT). Nucleotide sequencing analysis of these clones revealed ten specific contigs (six from strain SH and four from strain NT). Further analysis found that six contigs from strain SH and three from strain NT shared significant identities with previously reported proteins, and one contig was presumed to be novel. The specific differentially expressed genes were finally verified by RT-PCR and qRT-PCR analyses. Conclusions The data presented here suggest that specific genes identified between the two strains may be important molecules in the immunogenicity of E. maxima that may present potential new drug targets or vaccine candidates for coccidiosis. PMID:24894832

Molecular characterization of amino acid deletion in VP1 (1D) protein and novel amino acid substitutions in 3D polymerase protein of foot and mouth disease virus subtype A/Iran87.

PubMed

Esmaelizad, Majid; Jelokhani-Niaraki, Saber; Hashemnejad, Khadije; Kamalzadeh, Morteza; Lotfi, Mohsen

2011-12-01

The nucleotide sequence of the VP1 (1D) and partial 3D polymerase (3D(pol)) coding regions of the foot and mouth disease virus (FMDV) vaccine strain A/Iran87, a highly passaged isolate (~150 passages), was determined and aligned with previously published FMDV serotype A sequences. Overall analysis of the amino acid substitutions revealed that the partial 3D(pol) coding region contained four amino acid alterations. Amino acid sequence comparison of the VP1 coding region of the field isolates revealed deletions in the highly passaged Iranian isolate (A/Iran87). The prominent G-H loop of the FMDV VP1 protein contains the conserved arginine-glycine-aspartic acid (RGD) tripeptide, which is a well-known ligand for a specific cell surface integrin. Despite losing the RGD sequence of the VP1 protein and an Asp(26)→Glu substitution in a beta sheet located within a small groove of the 3D(pol) protein, the virus grew in BHK 21 suspension cell cultures. Since this strain has been used as a vaccine strain, it may be inferred that the RGD deletion has no critical role in virus attachment to the cell during the initiation of infection. It is probable that this FMDV subtype can utilize other pathways for cell attachment.

Mutations Affecting Expression of the rosy Locus in Drosophila melanogaster

PubMed Central

Lee, Chong Sung; Curtis, Daniel; McCarron, Margaret; Love, Carol; Gray, Mark; Bender, Welcome; Chovnick, Arthur

1987-01-01

The rosy locus in Drosophila melanogaster codes for the enzyme xanthine dehydrogenase (XDH). Previous studies defined a "control element" near the 5' end of the gene, where variant sites affected the amount of rosy mRNA and protein produced. We have determined the DNA sequence of this region from both genomic and cDNA clones, and from the ry+10 underproducer strain. This variant strain had many sequence differences, so that the site of the regulatory change could not be fixed. A mutagenesis was also undertaken to isolate new regulatory mutations. We induced 376 new mutations with 1-ethyl-1-nitrosourea (ENU) and screened them to isolate those that reduced the amount of XDH protein produced, but did not change the properties of the enzyme. Genetic mapping was used to find mutations located near the 5' end of the gene. DNA from each of seven mutants was cloned and sequenced through the 5' region. Mutant base changes were identified in all seven; they appear to affect splicing and translation of the rosy mRNA. In a related study (T. P. Keith et al. 1987), the genomic and cDNA sequences are extended through the 3' end of the gene; the combined sequences define the processing pattern of the rosy transcript and predict the amino acid sequence of XDH. PMID:3036645

Survey of endosymbionts in the Diaphorina citri metagenome and assembly of a Wolbachia wDi draft genome.

PubMed

Saha, Surya; Hunter, Wayne B; Reese, Justin; Morgan, J Kent; Marutani-Hert, Mizuri; Huang, Hong; Lindeberg, Magdalen

2012-01-01

Diaphorina citri (Hemiptera: Psyllidae), the Asian citrus psyllid, is the insect vector of Ca. Liberibacter asiaticus, the causal agent of citrus greening disease. Sequencing of the D. citri metagenome has been initiated to gain better understanding of the biology of this organism and the potential roles of its bacterial endosymbionts. To corroborate candidate endosymbionts previously identified by rDNA amplification, raw reads from the D. citri metagenome sequence were mapped to reference genome sequences. Results of the read mapping provided the most support for Wolbachia and an enteric bacterium most similar to Salmonella. Wolbachia-derived reads were extracted using the complete genome sequences for four Wolbachia strains. Reads were assembled into a draft genome sequence, and the annotation assessed for the presence of features potentially involved in host interaction. Genome alignment with the complete sequences reveals membership of Wolbachia wDi in supergroup B, further supported by phylogenetic analysis of FtsZ. FtsZ and Wsp phylogenies additionally indicate that the Wolbachia strain in the Florida D. citri isolate falls into a sub-clade of supergroup B, distinct from Wolbachia present in Chinese D. citri isolates, supporting the hypothesis that the D. citri introduced into Florida did not originate from China.

Survey of Endosymbionts in the Diaphorina citri Metagenome and Assembly of a Wolbachia wDi Draft Genome

PubMed Central

Saha, Surya; Hunter, Wayne B.; Reese, Justin; Morgan, J. Kent; Marutani-Hert, Mizuri; Huang, Hong; Lindeberg, Magdalen

2012-01-01

Diaphorina citri (Hemiptera: Psyllidae), the Asian citrus psyllid, is the insect vector of Ca. Liberibacter asiaticus, the causal agent of citrus greening disease. Sequencing of the D. citri metagenome has been initiated to gain better understanding of the biology of this organism and the potential roles of its bacterial endosymbionts. To corroborate candidate endosymbionts previously identified by rDNA amplification, raw reads from the D. citri metagenome sequence were mapped to reference genome sequences. Results of the read mapping provided the most support for Wolbachia and an enteric bacterium most similar to Salmonella. Wolbachia-derived reads were extracted using the complete genome sequences for four Wolbachia strains. Reads were assembled into a draft genome sequence, and the annotation assessed for the presence of features potentially involved in host interaction. Genome alignment with the complete sequences reveals membership of Wolbachia wDi in supergroup B, further supported by phylogenetic analysis of FtsZ. FtsZ and Wsp phylogenies additionally indicate that the Wolbachia strain in the Florida D. citri isolate falls into a sub-clade of supergroup B, distinct from Wolbachia present in Chinese D. citri isolates, supporting the hypothesis that the D. citri introduced into Florida did not originate from China. PMID:23166822

The bias of experimental design, including strain background, in the determination of critical Streptococcus suis serotype 2 virulence factors

PubMed Central

Auger, Jean-Philippe; Chuzeville, Sarah; Roy, David; Mathieu-Denoncourt, Annabelle; Xu, Jianguo; Grenier, Daniel

2017-01-01

Streptococcus suis serotype 2 is an important porcine bacterial pathogen and emerging zoonotic agent mainly responsible for sudden death, septic shock, and meningitis. However, serotype 2 strains are genotypically and phenotypically heterogeneous. Though a multitude of virulence factors have been described for S. suis serotype 2, the lack of a clear definition regarding which ones are truly “critical” has created inconsistencies that have only recently been highlighted. Herein, the involvement of two factors previously described as being critical for S. suis serotype 2 virulence, whether the dipeptidyl peptidase IV and autolysin, were evaluated with regards to different ascribed functions using prototype strains belonging to important sequence types. Results demonstrate a lack of reproducibility with previously published data. In fact, the role of the dipeptidyl peptidase IV and autolysin as critical virulence factors could not be confirmed. Though certain in vitro functions may be ascribed to these factors, their roles are not unique for S. suis, probably due to compensation by other factors. As such, variations and discrepancies in experimental design, including in vitro assays, cell lines, and animal models, are an important source of differences between results. Moreover, the use of different sequence types in this study demonstrates that the role attributed to a virulence factor may vary according to the S. suis serotype 2 strain background. Consequently, it is necessary to establish standard experimental designs according to the experiment and purpose in order to facilitate comparison between laboratories. Alongside, studies should include strains of diverse origins in order to prevent erroneous and biased conclusions that could affect future studies. PMID:28753679

Fine-structure mapping of the complex locus Odc-rs5 relative to Igk and distal loci.

PubMed

Richards-Smith, B A; Elliott, R W

1992-01-01

Odc-rs5 was previously identified as a complex locus closely linked to the Igk complex on mouse Chromosome (Chr) 6 and comprising at least five copies of a sequence related to the mRNA encoding ornithine decarboxylase (ODC) in the genomes of mice of some inbred strains and at least seven copies in others (Richards-Smith and Elliott, Mammalian Genome 2: 215, 1992). In the present study, Odc-rs5 was shown to be composed of at least seven copies of the ODC sequence in both the Odc-rs5a and Odc-rs5b haplotypes. Based upon the distribution of DNA restriction fragments (RFs) that had previously been associated with Odc-rs5a or Odc-rs5b among 42 mice of inbred laboratory strains having various haplotypes at Igk and in mice of two congenic strains [B6.PL-Ly-2a, Ly-3a(75NS)/Cy and B6.PL-Ly-2a,Ly-3a(85NS)/Cy] and a backcross-derived stock (NAK) known to be recombinant within Igk, a fine structure map of Odc-rs5 was deduced relative to Igk and more distal loci. Odc-rs5-derived RFs were located to three distinct regions within and/or distal to Igk and to a fourth site between (Ly-3, Ly-2) and Raf-1. Additionally, DNAs from 19 mice of inbred strains and random-bred stocks derived from wild progenitors trapped at various locations were analyzed and found to exhibit an unexpected variety of combinations of RFs associated with the two Odc-rs5 haplotypes most frequently observed among inbred laboratory strains of mice.

SEROPREVALENCE AND MOLECULAR CHARACTERIZATION OF FERLAVIRUS IN CAPTIVE VIPERS OF COSTA RICA.

PubMed

Solis, Cristina; Arguedas, Randall; Baldi, Mario; Piche, Martha; Jimenez, Carlos

2017-06-01

Ferlaviruses (FV, previously referred to as ophidian paramyxoviruses, OPMV), are enveloped viruses with a negative-strand RNA genome, affecting snakes in captivity worldwide. Infection is characterized by respiratory and nervous clinical signs and carries high mortality rates, but no specific treatment or vaccine is currently available. Costa Rica has 16 species of vipers, found in captivity in collections essential for antivenom production, reintroduction, and public education. FV circulation in these populations was previously unknown, and the risk of introducing the viruses into naïve collections or free-ranging populations exists if the virus's presence is confirmed. The objective of this study was to determine seroprevalence and FV shedding in 150 samples from captive vipers in nine collections across Costa Rica. A hemagglutination inhibition (HI) assay was performed to determine the antibody titer against two Ferlavirus strains, Bush viper virus (BV) and Neotropical virus (NT), and reverse-transcriptase polymerase chain reaction (RT-PCR) and sequencing to determine virus secretion in cloacal swabs. Ferlavirus strains were replicated in Vero cells, and chicken anti-FV polyclonal antibodies were produced and used as a positive control serum for the HI. Results demonstrate that seroprevalence of anti-FV antibodies in viper serum was 26.6% (n = 40) for the BV strain and 30% (n = 45) for the NT strain in the population tested. Furthermore, molecular characterization of FV group A was possible by sequencing the virus recovered from three cloacal swabs, demonstrating circulation of FV in one collection. This study demonstrates for the first time serological evidence of FV exposure and infection in vipers in captivity in Costa Rica, and suggests cross reactivity between antibodies against both strains. Appropriate biosafety measures could prevent the spread of FV between and within collections of reptiles in the country.

AN IMPORTED CASE OF ACUTE MELIOIDOSIS CAUSED BY ST881 BURKHOLDERIA PSEUDOMALLEI.

PubMed

Zong, Zhiyong; Wang, Xiaohui; Deng, Yiyun

2016-03-01

A previously healthy Chinese male working in Malaysia returned to China with high fever. A blood culture showed Burkholderia pseudomallei strain WCBP1. This isolate was sequenced, showing type, ST881, which appears to be present in Malaysia. WCP1 had unusual susceptibility to aminoglycosides and habored the Yersinia-like fimbrial gene cluster for virulence. The patient's condition deteriorated rapidly but he recovered after receiving meropenem and intensive care support. Melioidosis is a potential problem among Chinese imigrant workers with strains new to China being identified.

Analysis of the genomic sequences and metabolites of Serratia surfactantfaciens sp. nov. YD25T that simultaneously produces prodigiosin and serrawettin W2.

PubMed

Su, Chun; Xiang, Zhaoju; Liu, Yibo; Zhao, Xinqing; Sun, Yan; Li, Zhi; Li, Lijun; Chang, Fan; Chen, Tianjun; Wen, Xinrong; Zhou, Yidan; Zhao, Furong

2016-11-03

Gram-negative bacteria of the genus Serratia are potential producers of many useful secondary metabolites, such as prodigiosin and serrawettins, which have potential applications in environmental bioremediation or in the pharmaceutical industry. Several Serratia strains produce prodigiosin and serrawettin W1 as the main bioactive compounds, and the biosynthetic pathways are co-regulated by quorum sensing (QS). In contrast, the Serratia strain, which can simultaneously produce prodigiosin and serrawettin W2, has not been reported. This study focused on analyzing the genomic sequence of Serratia sp. strain YD25 T isolated from rhizosphere soil under continuously planted burley tobacco collected from Yongding, Fujian province, China, which is unique in producing both prodigiosin and serrawettin W2. A hybrid polyketide synthases (PKS)-non-ribosomal peptide synthetases (NRPS) gene cluster putatively involved in biosynthesis of antimicrobial serrawettin W2 was identified in the genome of YD25 T , and its biosynthesis pathway was proposed. We found potent antimicrobial activity of serrawettin W2 purified from YD25 T against various pathogenic bacteria and fungi as well as antitumor activity against Hela cells. Subsequently, comparative genomic analyses were performed among a total of 133 Serratia species. The prodigiosin biosynthesis gene cluster in YD25 T belongs to the type I pig cluster, which is the main form of pig-encoding genes existing in most of the pigmented Serratia species. In addition, a complete autoinducer-2 (AI-2) system (including luxS, lsrBACDEF, lsrGK, and lsrR) as a conserved bacterial operator is found in the genome of Serratia sp. strain YD25 T . Phylogenetic analysis based on concatenated Lsr and LuxS proteins revealed that YD25 T formed an independent branch and was clearly distant from the strains that solely produce either prodigiosin or serrawettin W2. The Fe (III) ion reduction assay confirmed that strain YD25 T could produce an AI-2 signal molecule. Phylogenetic analysis using the genomic sequence of YD25 T combined with phylogenetic and phenotypic analyses support this strain as a member of a novel and previously uncharacterized Serratia species. Genomic sequence and metabolite analysis of Serratia surfactantfaciens YD25 T indicate that this strain can be further explored for the production of useful metabolites. Unveiling the genomic sequence of S. surfactantfaciens YD25 T benefits the usage of this unique strain as a model system for studying the biosynthesis regulation of both prodigiosin and serrawettin W2 by the QS system.

Analysis of Multilocus Sequence Typing and Virulence Characterization of Listeria monocytogenes Isolates from Chinese Retail Ready-to-Eat Food

PubMed Central

Wu, Shi; Wu, Qingping; Zhang, Jumei; Chen, Moutong; Guo, Weipeng

2016-01-01

Eighty Listeria monocytogenes isolates were obtained from Chinese retail ready-to-eat (RTE) food and were previously characterized with serotyping and antibiotic susceptibility tests. The aim of this study was to characterize the subtype and virulence potential of these L. monocytogenes isolates by multilocus sequence typing (MLST), virulence-associate genes, epidemic clones (ECs), and sequence analysis of the important virulence factor: internalin A (inlA). The result of MLST revealed that these L. monocytogenes isolates belonged to 14 different sequence types (STs). With the exception of four new STs (ST804, ST805, ST806, and ST807), all other STs observed in this study have been associated with human listeriosis and outbreaks to varying extents. Six virulence-associate genes (inlA, inlB, inlC, inlJ, hly, and llsX) were selected and their presence was investigated using PCR. All strains carried inlA, inlB, inlC, inlJ, and hly, whereas 38.8% (31/80) of strains harbored the listeriolysin S genes (llsX). A multiplex PCR assay was used to evaluate the presence of markers specific to epidemic clones of L. monocytogenes and identified 26.3% (21/80) of ECI in the 4b-4d-4e strains. Further study of inlA sequencing revealed that most strains contained the full-length InlA required for host cell invasion, whereas three mutations lead to premature stop codons (PMSC) within a novel PMSCs at position 326 (GAA → TAA). MLST and inlA sequence analysis results were concordant, and different virulence potentials within isolates were observed. These findings suggest that L. monocytogenes isolates from RTE food in China could be virulent and be capable of causing human illness. Furthermore, the STs and virulence profiles of L. monocytogenes isolates have significant implications for epidemiological and public health studies of this pathogen. PMID:26909076

Analysis of Multilocus Sequence Typing and Virulence Characterization of Listeria monocytogenes Isolates from Chinese Retail Ready-to-Eat Food.

PubMed

Wu, Shi; Wu, Qingping; Zhang, Jumei; Chen, Moutong; Guo, Weipeng

2016-01-01

Eighty Listeria monocytogenes isolates were obtained from Chinese retail ready-to-eat (RTE) food and were previously characterized with serotyping and antibiotic susceptibility tests. The aim of this study was to characterize the subtype and virulence potential of these L. monocytogenes isolates by multilocus sequence typing (MLST), virulence-associate genes, epidemic clones (ECs), and sequence analysis of the important virulence factor: internalin A (inlA). The result of MLST revealed that these L. monocytogenes isolates belonged to 14 different sequence types (STs). With the exception of four new STs (ST804, ST805, ST806, and ST807), all other STs observed in this study have been associated with human listeriosis and outbreaks to varying extents. Six virulence-associate genes (inlA, inlB, inlC, inlJ, hly, and llsX) were selected and their presence was investigated using PCR. All strains carried inlA, inlB, inlC, inlJ, and hly, whereas 38.8% (31/80) of strains harbored the listeriolysin S genes (llsX). A multiplex PCR assay was used to evaluate the presence of markers specific to epidemic clones of L. monocytogenes and identified 26.3% (21/80) of ECI in the 4b-4d-4e strains. Further study of inlA sequencing revealed that most strains contained the full-length InlA required for host cell invasion, whereas three mutations lead to premature stop codons (PMSC) within a novel PMSCs at position 326 (GAA → TAA). MLST and inlA sequence analysis results were concordant, and different virulence potentials within isolates were observed. These findings suggest that L. monocytogenes isolates from RTE food in China could be virulent and be capable of causing human illness. Furthermore, the STs and virulence profiles of L. monocytogenes isolates have significant implications for epidemiological and public health studies of this pathogen.

Lactobacillus strain diversity based on partial hsp60 gene sequences and design of PCR-restriction fragment length polymorphism assays for species identification and differentiation.

PubMed

Blaiotta, Giuseppe; Fusco, Vincenzina; Ercolini, Danilo; Aponte, Maria; Pepe, Olimpia; Villani, Francesco

2008-01-01

A phylogenetic tree showing diversities among 116 partial (499-bp) Lactobacillus hsp60 (groEL, encoding a 60-kDa heat shock protein) nucleotide sequences was obtained and compared to those previously described for 16S rRNA and tuf gene sequences. The topology of the tree produced in this study showed a Lactobacillus species distribution similar, but not identical, to those previously reported. However, according to the most recent systematic studies, a clear differentiation of 43 single-species clusters was detected/identified among the sequences analyzed. The slightly higher variability of the hsp60 nucleotide sequences than of the 16S rRNA sequences offers better opportunities to design or develop molecular assays allowing identification and differentiation of either distant or very closely related Lactobacillus species. Therefore, our results suggest that hsp60 can be considered an excellent molecular marker for inferring the taxonomy and phylogeny of members of the genus Lactobacillus and that the chosen primers can be used in a simple PCR procedure allowing the direct sequencing of the hsp60 fragments. Moreover, in this study we performed a computer-aided restriction endonuclease analysis of all 499-bp hsp60 partial sequences and we showed that the PCR-restriction fragment length polymorphism (RFLP) patterns obtainable by using both endonucleases AluI and TacI (in separate reactions) can allow identification and differentiation of all 43 Lactobacillus species considered, with the exception of the pair L. plantarum/L. pentosus. However, the latter species can be differentiated by further analysis with Sau3AI or MseI. The hsp60 PCR-RFLP approach was efficiently applied to identify and to differentiate a total of 110 wild Lactobacillus strains (including closely related species, such as L. casei and L. rhamnosus or L. plantarum and L. pentosus) isolated from cheese and dry-fermented sausages.

Lactobacillus Strain Diversity Based on Partial hsp60 Gene Sequences and Design of PCR-Restriction Fragment Length Polymorphism Assays for Species Identification and Differentiation▿ †

PubMed Central

Blaiotta, Giuseppe; Fusco, Vincenzina; Ercolini, Danilo; Aponte, Maria; Pepe, Olimpia; Villani, Francesco

2008-01-01

A phylogenetic tree showing diversities among 116 partial (499-bp) Lactobacillus hsp60 (groEL, encoding a 60-kDa heat shock protein) nucleotide sequences was obtained and compared to those previously described for 16S rRNA and tuf gene sequences. The topology of the tree produced in this study showed a Lactobacillus species distribution similar, but not identical, to those previously reported. However, according to the most recent systematic studies, a clear differentiation of 43 single-species clusters was detected/identified among the sequences analyzed. The slightly higher variability of the hsp60 nucleotide sequences than of the 16S rRNA sequences offers better opportunities to design or develop molecular assays allowing identification and differentiation of either distant or very closely related Lactobacillus species. Therefore, our results suggest that hsp60 can be considered an excellent molecular marker for inferring the taxonomy and phylogeny of members of the genus Lactobacillus and that the chosen primers can be used in a simple PCR procedure allowing the direct sequencing of the hsp60 fragments. Moreover, in this study we performed a computer-aided restriction endonuclease analysis of all 499-bp hsp60 partial sequences and we showed that the PCR-restriction fragment length polymorphism (RFLP) patterns obtainable by using both endonucleases AluI and TacI (in separate reactions) can allow identification and differentiation of all 43 Lactobacillus species considered, with the exception of the pair L. plantarum/L. pentosus. However, the latter species can be differentiated by further analysis with Sau3AI or MseI. The hsp60 PCR-RFLP approach was efficiently applied to identify and to differentiate a total of 110 wild Lactobacillus strains (including closely related species, such as L. casei and L. rhamnosus or L. plantarum and L. pentosus) isolated from cheese and dry-fermented sausages. PMID:17993558

Complete Genome Sequences of Newcastle Disease Virus Strains Circulating in Chicken Populations of Indonesia

PubMed Central

Xiao, Sa; Paldurai, Anandan; Nayak, Baibaswata; Samuel, Arthur; Bharoto, Eny E.; Prajitno, Teguh Y.; Collins, Peter L.

2012-01-01

Eight highly virulent Newcastle disease virus (NDV) strains were isolated from vaccinated commercial chickens in Indonesia during outbreaks in 2009 and 2010. The complete genome sequences of two NDV strains and the sequences of the surface protein genes (F and HN) of six other strains were determined. Phylogenetic analysis classified them into two new subgroups of genotype VII in the class II cluster that were genetically distinct from vaccine strains. This is the first report of complete genome sequences of NDV strains isolated from chickens in Indonesia. PMID:22532534

Gluconacetobacter medellinensis sp. nov., cellulose- and non-cellulose-producing acetic acid bacteria isolated from vinegar.

PubMed

Castro, Cristina; Cleenwerck, Ilse; Trcek, Janja; Zuluaga, Robin; De Vos, Paul; Caro, Gloria; Aguirre, Ricardo; Putaux, Jean-Luc; Gañán, Piedad

2013-03-01

The phylogenetic position of a cellulose-producing acetic acid bacterium, strain ID13488, isolated from commercially available Colombian homemade fruit vinegar, was investigated. Analyses using nearly complete 16S rRNA gene sequences, nearly complete 16S-23S rRNA gene internal transcribed spacer (ITS) sequences, as well as concatenated partial sequences of the housekeeping genes dnaK, groEL and rpoB, allocated the micro-organism to the genus Gluconacetobacter, and more precisely to the Gluconacetobacter xylinus group. Moreover, the data suggested that the micro-organism belongs to a novel species in this genus, together with LMG 1693(T), a non-cellulose-producing strain isolated from vinegar by Kondo and previously classified as a strain of Gluconacetobacter xylinus. DNA-DNA hybridizations confirmed this finding, revealing a DNA-DNA relatedness value of 81 % between strains ID13488 and LMG 1693(T), and values <70 % between strain LMG 1693(T) and the type strains of the closest phylogenetic neighbours. Additionally, the classification of strains ID13488 and LMG 1693(T) into a single novel species was supported by amplified fragment length polymorphism (AFLP) and (GTG)5-PCR DNA fingerprinting data, as well as by phenotypic data. Strains ID13488 and LMG 1693(T) could be differentiated from closely related species of the genus Gluconacetobacter by their ability to produce 2- and 5-keto-d-gluconic acid from d-glucose, their ability to produce acid from sucrose, but not from 1-propanol, and their ability to grow on 3 % ethanol in the absence of acetic acid and on ethanol, d-ribose, d-xylose, sucrose, sorbitol, d-mannitol and d-gluconate as carbon sources. The DNA G+C content of strains ID13488 and LMG 1693(T) was 58.0 and 60.7 mol%, respectively. The major ubiquinone of LMG 1693(T) was Q-10. Taken together these data indicate that strains ID13488 and LMG 1693(T) represent a novel species of the genus Gluconacetobacter for which the name Gluconacetobacter medellinensis sp. nov. is proposed. The type strain is LMG 1693(T) ( = NBRC 3288(T) = Kondo 51(T)).

Genetic Diversity of the Hepatitis B Virus Strains in Cuba: Absence of West-African Genotypes despite the Transatlantic Slave Trade

PubMed Central

Rodríguez Lay, Licel A.; Corredor, Marité B.; Villalba, Maria C.; Frómeta, Susel S.; Wong, Meilin S.; Valdes, Lidunka; Samada, Marcia; Sausy, Aurélie; Hübschen, Judith M.; Muller, Claude P.

2015-01-01

Cuba is an HBsAg low-prevalence country with a high coverage of anti-hepatitis B vaccine. Its population is essentially the result of the population mix of Spanish descendants and former African slaves. Information about genetic characteristics of hepatitis B virus (HBV) strains circulating in the country is scarce. The HBV genotypes/subgenotypes, serotypes, mixed infections, and S gene mutations of 172 Cuban HBsAg and HBV-DNA positive patients were determined by direct sequencing and phylogenetic analysis. Phylogenetic analysis of HBV S gene sequences showed a predominance of genotype A (92.4%), subgenotype A2 (84.9%) and A1 (7.6%). Genotype D (7.0%) and subgenotype C1 (0.6%) were also detected but typical (sub)genotypes of contemporary West-Africa (E, A3) were conspicuously absent. All genotype A, D, and C strains exhibited sequence characteristics of the adw2, ayw2, and adrq serotypes, respectively. Thirty-three (19.1%) patients showed single, double, or multiple point mutations inside the Major Hydrophilic domain associated with vaccine escape; eighteen (10.5%) patients had mutations in the T-cell epitope (amino acids 28-51), and there were another 111 point mutations downstream of the S gene. One patient had an HBV A1/A2 mixed infection. This first genetic study of Cuban HBV viruses revealed only strains that were interspersed with strains from particularly Europe, America, and Asia. The absence of genotype E supports previous hypotheses about an only recent introduction of this genotype into the general population in Africa. The presence of well-known vaccine escape (3.5%) and viral resistance mutants (2.9%) warrants strain surveillance to guide vaccination and treatment strategies. PMID:25978398

Comparison of multilocus sequence typing and pulsed-field gel electrophoresis for Salmonella spp. identification in surface water

NASA Astrophysics Data System (ADS)

Kuo, Chun Wei; Hao Huang, Kuan; Hsu, Bing Mu; Tsai, Hsien Lung; Tseng, Shao Feng; Kao, Po Min; Shen, Shu Min; Chou Chiu, Yi; Chen, Jung Sheng

2013-04-01

Salmonella is one of the most important pathogens of waterborne diseases with outbreaks from contaminated water reported worldwide. In addition, Salmonella spp. can survive for long periods in aquatic environments. To realize genotypes and serovars of Salmonella in aquatic environments, we isolated the Salmonella strains by selective culture plates to identify the serovars of Salmonella by serological assay, and identify the genotypes by Multilocus sequence typing (MLST) based on the sequence data from University College Cork (UCC), respectively. The results show that 36 stream water samples (30.1%) and 18 drinking water samples (23.3%) were confirmed the existence of Salmonella using culture method combined PCR specific invA gene amplification. In this study, 24 cultured isolates of Salmonella from water samples were classified to fifteen Salmonella enterica serovars. In addition, we construct phylogenetic analysis using phylogenetic tree and Minimum spanning tree (MST) method to analyze the relationship of clinical, environmental, and geographical data. Phylogenetic tree showed that four main clusters and our strains can be distributed in all. The genotypes of isolates from stream water are more biodiversity while comparing the Salmonella strains genotypes from drinking water sources. According to MST data, we can found the positive correlation between serovars and genotypes of Salmonella. Previous studies revealed that the result of Pulsed field gel electrophoresis (PFGE) method can predict the serovars of Salmonella strain. Hence, we used the MLST data combined phylogenetic analysis to identify the serovars of Salmonella strain and achieved effectiveness. While using the geographical data combined phylogenetic analysis, the result showed that the dominant strains were existed in whole stream area in rainy season. Keywords: Salmonella spp., MLST, phylogenetic analysis, PFGE

Isolation and Distribution of a Novel Iron-Oxidizing Crenarchaeon from Acidic Geothermal Springs in Yellowstone National Park▿ †

PubMed Central

Kozubal, M.; Macur, R. E.; Korf, S.; Taylor, W. P.; Ackerman, G. G.; Nagy, A.; Inskeep, W. P.

2008-01-01

Novel thermophilic crenarchaea have been observed in Fe(III) oxide microbial mats of Yellowstone National Park (YNP); however, no definitive work has identified specific microorganisms responsible for the oxidation of Fe(II). The objectives of the current study were to isolate and characterize an Fe(II)-oxidizing member of the Sulfolobales observed in previous 16S rRNA gene surveys and to determine the abundance and distribution of close relatives of this organism in acidic geothermal springs containing high concentrations of dissolved Fe(II). Here we report the isolation and characterization of the novel, Fe(II)-oxidizing, thermophilic, acidophilic organism Metallosphaera sp. strain MK1 obtained from a well-characterized acid-sulfate-chloride geothermal spring in Norris Geyser Basin, YNP. Full-length 16S rRNA gene sequence analysis revealed that strain MK1 exhibits only 94.9 to 96.1% sequence similarity to other known Metallosphaera spp. and less than 89.1% similarity to known Sulfolobus spp. Strain MK1 is a facultative chemolithoautotroph with an optimum pH range of 2.0 to 3.0 and an optimum temperature range of 65 to 75°C. Strain MK1 grows optimally on pyrite or Fe(II) sorbed onto ferrihydrite, exhibiting doubling times between 10 and 11 h under aerobic conditions (65°C). The distribution and relative abundance of MK1-like 16S rRNA gene sequences in 14 acidic geothermal springs containing Fe(III) oxide microbial mats were evaluated. Highly related MK1-like 16S rRNA gene sequences (>99% sequence similarity) were consistently observed in Fe(III) oxide mats at temperatures ranging from 55 to 80°C. Quantitative PCR using Metallosphaera-specific primers confirmed that organisms highly similar to strain MK1 comprised up to 40% of the total archaeal community at selected sites. The broad distribution of highly related MK1-like 16S rRNA gene sequences in acidic Fe(III) oxide microbial mats is consistent with the observed characteristics and growth optima of Metallosphaera-like strain MK1 and emphasizes the importance of this newly described taxon in Fe(II) chemolithotrophy in acidic high-temperature environments of YNP. PMID:18083851

Complete genome sequence and the expression pattern of plasmids of the model ethanologen Zymomonas mobilis ZM4 and its xylose-utilizing derivatives 8b and 2032

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

Yang, Shihui; Vera, Jessica M.; Grass, Jeff

Zymomonas mobilis is a natural ethanologen being developed and deployed as an industrial biofuel producer. To date, eight Z. mobilis strains have been completely sequenced and found to contain 2-8 native plasmids. However, systematic verification of predicted Z. mobilis plasmid genes and their contribution to cell fitness has not been hitherto addressed. Moreover, the precise number and identities of plasmids in Z. mobilis model strain ZM4 have been unclear. The lack of functional information about plasmid genes in ZM4 impedes ongoing studies for this model biofuel-producing strain. In this study, we determined the complete chromosome and plasmid sequences of ZM4more » and its engineered xylose-utilizing derivatives 2032 and 8b. Compared to previously published and revised ZM4 chromosome sequences, the ZM4 chromosome sequence reported here contains 65 nucleotide sequence variations as well as a 2400-bp insertion. Four plasmids were identified in all three strains, with 150 plasmid genes predicted in strain ZM4 and 2032, and 153 plasmid genes predicted in strain 8b due to the insertion of heterologous DNA for expanded substrate utilization. Plasmid genes were then annotated using Blast2GO, InterProScan, and systems biology data analyses, and most genes were found to have apparent orthologs in other organisms or identifiable conserved domains. To verify plasmid gene prediction, RNA-Seq was used to map transcripts and also compare relative gene expression under various growth conditions, including anaerobic and aerobic conditions, or growth in different concentrations of biomass hydrolysates. Overall, plasmid genes were more responsive to varying hydrolysate concentrations than to oxygen availability. Additionally, our results indicated that although all plasmids were present in low copy number (about 1-2 per cell), the copy number of some plasmids varied under specific growth conditions or due to heterologous gene insertion. The complete genome of ZM4 and two xylose-utilizing derivatives is reported in this study, with an emphasis on identifying and characterizing plasmid genes. Furthermore, plasmid gene annotation, validation, expression levels at growth conditions of interest, and contribution to host fitness are reported for the first time.« less

Complete genome sequence and the expression pattern of plasmids of the model ethanologen Zymomonas mobilis ZM4 and its xylose-utilizing derivatives 8b and 2032

DOE PAGES

Yang, Shihui; Vera, Jessica M.; Grass, Jeff; ...

2018-05-02

Zymomonas mobilis is a natural ethanologen being developed and deployed as an industrial biofuel producer. To date, eight Z. mobilis strains have been completely sequenced and found to contain 2-8 native plasmids. However, systematic verification of predicted Z. mobilis plasmid genes and their contribution to cell fitness has not been hitherto addressed. Moreover, the precise number and identities of plasmids in Z. mobilis model strain ZM4 have been unclear. The lack of functional information about plasmid genes in ZM4 impedes ongoing studies for this model biofuel-producing strain. In this study, we determined the complete chromosome and plasmid sequences of ZM4more » and its engineered xylose-utilizing derivatives 2032 and 8b. Compared to previously published and revised ZM4 chromosome sequences, the ZM4 chromosome sequence reported here contains 65 nucleotide sequence variations as well as a 2400-bp insertion. Four plasmids were identified in all three strains, with 150 plasmid genes predicted in strain ZM4 and 2032, and 153 plasmid genes predicted in strain 8b due to the insertion of heterologous DNA for expanded substrate utilization. Plasmid genes were then annotated using Blast2GO, InterProScan, and systems biology data analyses, and most genes were found to have apparent orthologs in other organisms or identifiable conserved domains. To verify plasmid gene prediction, RNA-Seq was used to map transcripts and also compare relative gene expression under various growth conditions, including anaerobic and aerobic conditions, or growth in different concentrations of biomass hydrolysates. Overall, plasmid genes were more responsive to varying hydrolysate concentrations than to oxygen availability. Additionally, our results indicated that although all plasmids were present in low copy number (about 1-2 per cell), the copy number of some plasmids varied under specific growth conditions or due to heterologous gene insertion. The complete genome of ZM4 and two xylose-utilizing derivatives is reported in this study, with an emphasis on identifying and characterizing plasmid genes. Furthermore, plasmid gene annotation, validation, expression levels at growth conditions of interest, and contribution to host fitness are reported for the first time.« less

Serratia nematodiphila sp. nov., associated symbiotically with the entomopathogenic nematode Heterorhabditidoides chongmingensis (Rhabditida: Rhabditidae).

PubMed

Zhang, Chong-Xing; Yang, Shou-Yun; Xu, Ming-Xu; Sun, Jie; Liu, Huan; Liu, Jing-Rui; Liu, Hui; Kan, Fei; Sun, Jing; Lai, Ren; Zhang, Ke-Yun

2009-07-01

A novel red-pigmented, Gram-negative, motile, fluorescent, rod-shaped strain, DZ0503SBS1(T), with a single lateral flagellum, was isolated from the intestine of the nematode Heterorhabditidoides chongmingensis. Comparative 16S rRNA gene sequence analysis indicated that the strain is a member of the genus Serratia, sharing highest sequence similarities with Serratia marcescens subsp. sakuensis JCM 11315(T) (99.8 %), S. marcescens subsp. marcescens DSM 30121(T) (99.5 %) and Serratia ureilytica LMG 22860(T) (98.3 %). Similarities between the rpoB gene sequence of strain DZ0503SBS1(T) and those of S. marcescens subsp. sakuensis JCM 11315(T), S. marcescens subsp. marcescens DSM 30121(T) and S. ureilytica LMG 22860(T) were 98.0, 97.4 and 98.3 %, respectively. DNA-DNA hybridization values of strain DZ0503SBS1(T) with S. marcescens subsp. sakuensis JCM 11315(T), S. marcescens subsp. marcescens DSM 30121(T) and S. ureilytica LMG 22860(T) were 68.2, 65.1 and 53.0 %, respectively. The major isoprenoid quinone of strain DZ0503SBS1(T) was Q-8 and the predominant fatty acids were C(16 : 0) (34.76 %), cyclo-C(17 : 0) (20.03 %) and cyclo-C(19 : 0)omega8c (17.24 %). The cyclo-C(19 : 0)omega8c content (17.24 %) was significantly different from those found in S. marcescens subsp. sakuensis JCM 11315(T) and S. marcescens subsp. marcescens DSM 30121(T). Some characteristics of strain DZ0503SBS1(T), i.e. fluorescence and its symbiotic association with nematodes, have not been reported previously in any species of the genus Serratia. Phenotypic and biochemical characteristics and molecular data show that strain DZ0503SBS1(T) represents a novel species, for which the name Serratia nematodiphila sp. nov. is proposed; the type strain is DZ0503SBS1(T) (=KCTC 22130(T) =CGMCC 1.6853(T)).

Genomic analysis and temperature-dependent transcriptome profiles of the rhizosphere originating strain Pseudomonas aeruginosa M18

PubMed Central

2011-01-01

Background Our previously published reports have described an effective biocontrol agent named Pseudomonas sp. M18 as its 16S rDNA sequence and several regulator genes share homologous sequences with those of P. aeruginosa, but there are several unusual phenotypic features. This study aims to explore its strain specific genomic features and gene expression patterns at different temperatures. Results The complete M18 genome is composed of a single chromosome of 6,327,754 base pairs containing 5684 open reading frames. Seven genomic islands, including two novel prophages and five specific non-phage islands were identified besides the conserved P. aeruginosa core genome. Each prophage contains a putative chitinase coding gene, and the prophage II contains a capB gene encoding a putative cold stress protein. The non-phage genomic islands contain genes responsible for pyoluteorin biosynthesis, environmental substance degradation and type I and III restriction-modification systems. Compared with other P. aeruginosa strains, the fewest number (3) of insertion sequences and the most number (3) of clustered regularly interspaced short palindromic repeats in M18 genome may contribute to the relative genome stability. Although the M18 genome is most closely related to that of P. aeruginosa strain LESB58, the strain M18 is more susceptible to several antimicrobial agents and easier to be erased in a mouse acute lung infection model than the strain LESB58. The whole M18 transcriptomic analysis indicated that 10.6% of the expressed genes are temperature-dependent, with 22 genes up-regulated at 28°C in three non-phage genomic islands and one prophage but none at 37°C. Conclusions The P. aeruginosa strain M18 has evolved its specific genomic structures and temperature dependent expression patterns to meet the requirement of its fitness and competitiveness under selective pressures imposed on the strain in rhizosphere niche. PMID:21884571

Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:H− Isolates from Czech Patients with Novel Plasmid Composition Not Previously Seen in German Isolates

PubMed Central

Bauwens, Andreas; Marejková, Monika; Middendorf-Bauchart, Barbara; Prager, Rita; Kossow, Annelene; Zhang, Wenlan; Karch, Helge

2017-01-01

ABSTRACT Sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H− strains, first identified in Germany, have emerged as important pathogens throughout Europe. Besides chromosomally encoded Shiga toxin 2a (the major virulence factor), several putative virulence loci, including the hly, etp, and sfp operons, encoding EHEC hemolysin, type II secretion system proteins, and Sfp fimbriae, respectively, are located on the 121-kb plasmid pSFO157 in German strains. Here we report novel SF EHEC O157:H− strains isolated from patients in the Czech Republic. These strains share the core genomes and chromosomal virulence loci encoding toxins (stx2a and the cdtV-ABC operon) and adhesins (eae-γ, efa1, lpfAO157OI-141, and lpfAO157OI-154) with German strains but differ essentially in their plasmids. In contrast to all previously detected SF EHEC O157:H− strains, the Czech strains carry two plasmids, of 79 kb and 86 kb. The 79-kb plasmid harbors the sfp operon, but neither of the plasmids contains the hly and etp operons. Sequence analyses demonstrated that the 79-kb plasmid (pSFO157 258/98-1) evolved from pSFO157 of German strains by deletion of a 41,534-bp region via homologous recombination, resulting in loss of the hly and etp operons. The 86-kb plasmid (pSFO157 258/98-2) displays 98% sequence similarity to a 92.7-kb plasmid of an extraintestinal pathogenic E. coli bloodstream isolate. Our finding of this novel plasmid composition in SF EHEC O157:H− strains extends the evolutionary history of EHEC O157 plasmids. Moreover, the unique molecular plasmid characteristics permit the identification of such strains, thereby facilitating further investigations of their geographic distribution, clinical significance, and epidemiology. IMPORTANCE Since their first identification in Germany in 1989, sorbitol-fermenting enterohemorrhagic Escherichia coli O157:H− (nonmotile) strains have emerged as important causes of the life-threatening disease hemolytic-uremic syndrome in Europe. They account for 10 to 20% of sporadic cases of this disease and have caused several large outbreaks. The strains isolated throughout Europe share conserved chromosomal and plasmid characteristics. Here we identified novel sorbitol-fermenting enterohemorrhagic E. coli O157:H− patient isolates in the Czech Republic which differ from all such strains reported previously by their unique plasmid characteristics, including plasmid number, composition of plasmid-carried virulence genes, and plasmid origins. Our findings contribute substantially to understanding the evolution of E. coli O157 strains and their plasmids. In practical terms, they enable the identification of strains with these novel plasmid characteristics in patient stool samples and thus the investigation of their roles as human pathogens in other geographic areas. PMID:28970221

Sorbitol-Fermenting Enterohemorrhagic Escherichia coli O157:H- Isolates from Czech Patients with Novel Plasmid Composition Not Previously Seen in German Isolates.

PubMed

Bauwens, Andreas; Marejková, Monika; Middendorf-Bauchart, Barbara; Prager, Rita; Kossow, Annelene; Zhang, Wenlan; Karch, Helge; Mellmann, Alexander; Bielaszewska, Martina

2017-12-01

Sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H - strains, first identified in Germany, have emerged as important pathogens throughout Europe. Besides chromosomally encoded Shiga toxin 2a (the major virulence factor), several putative virulence loci, including the hly , etp , and sfp operons, encoding EHEC hemolysin, type II secretion system proteins, and Sfp fimbriae, respectively, are located on the 121-kb plasmid pSFO157 in German strains. Here we report novel SF EHEC O157:H - strains isolated from patients in the Czech Republic. These strains share the core genomes and chromosomal virulence loci encoding toxins ( stx 2a and the cdtV -ABC operon) and adhesins ( eae -γ, efa1 , lpfA O157OI-141 , and lpfA O157OI-154 ) with German strains but differ essentially in their plasmids. In contrast to all previously detected SF EHEC O157:H - strains, the Czech strains carry two plasmids, of 79 kb and 86 kb. The 79-kb plasmid harbors the sfp operon, but neither of the plasmids contains the hly and etp operons. Sequence analyses demonstrated that the 79-kb plasmid (pSFO157 258/98-1) evolved from pSFO157 of German strains by deletion of a 41,534-bp region via homologous recombination, resulting in loss of the hly and etp operons. The 86-kb plasmid (pSFO157 258/98-2) displays 98% sequence similarity to a 92.7-kb plasmid of an extraintestinal pathogenic E. coli bloodstream isolate. Our finding of this novel plasmid composition in SF EHEC O157:H - strains extends the evolutionary history of EHEC O157 plasmids. Moreover, the unique molecular plasmid characteristics permit the identification of such strains, thereby facilitating further investigations of their geographic distribution, clinical significance, and epidemiology. IMPORTANCE Since their first identification in Germany in 1989, sorbitol-fermenting enterohemorrhagic Escherichia coli O157:H - (nonmotile) strains have emerged as important causes of the life-threatening disease hemolytic-uremic syndrome in Europe. They account for 10 to 20% of sporadic cases of this disease and have caused several large outbreaks. The strains isolated throughout Europe share conserved chromosomal and plasmid characteristics. Here we identified novel sorbitol-fermenting enterohemorrhagic E. coli O157:H - patient isolates in the Czech Republic which differ from all such strains reported previously by their unique plasmid characteristics, including plasmid number, composition of plasmid-carried virulence genes, and plasmid origins. Our findings contribute substantially to understanding the evolution of E. coli O157 strains and their plasmids. In practical terms, they enable the identification of strains with these novel plasmid characteristics in patient stool samples and thus the investigation of their roles as human pathogens in other geographic areas. Copyright © 2017 American Society for Microbiology.

Complete Genome Analysis of an Enterovirus EV-B83 Isolated in China.

PubMed

Tang, Jingjing; Li, Qiongfen; Tian, Bingjun; Zhang, Jie; Li, Kai; Ding, Zhengrong; Lu, Lin

2016-07-12

Enterovirus B83 (EV-B83) is a recently identified member of enterovirus species B. It is a rarely reported serotype and up to date, only the complete genome sequence of the prototype strain from the United States is available. In this study, we describe the complete genomic characterization of an EV-B83 strain 246/YN/CHN/08HC isolated from a healthy child living in border region of Yunnan Province, China in 2008. Compared with the prototype strain, it had 79.6% similarity in the complete genome and 78.9% similarity in the VP1 coding region, reflecting the great genetic divergence among them. VP1-coding region alignment revealed it had 77.2-91.3% with other EV-B83 sequences available in GenBank. Similarity plot analysis revealed it had higher identity with several other EV-B serotypes than the EV-B83 prototype strain in the P2 and P3 coding region, suggesting multiple recombination events might have occurred. The great genetic divergence with previously isolated strains and the extremely rare isolation suggest this serotype has circulated at a low epidemic strength for many years. This is the first report of complete genome of EV-B83 in China.

Human polyomavirus JC variants in Papua New Guinea and Guam reflect ancient population settlement and viral evolution.

PubMed

Ryschkewitsch, C F; Friedlaender, J S; Mgone, C S; Jobes, D V; Agostini, H T; Chima, S C; Alpers, M P; Koki, G; Yanagihara, R; Stoner, G L

2000-07-01

The peopling of the Pacific was a complex sequence of events that is best reconstructed by reconciling insights from various disciplines. Here we analyze the human polyomavirus JC (JCV) in Highlanders of Papua New Guinea (PNG), in Austronesian-speaking Tolai people on the island of New Britain, and in nearby non-Austronesian-speaking Baining people. We also characterize JCV from the Chamorro of Guam, a Micronesian population. All JCV strains from PNG and Guam fall within the broad Asian group previously defined in the VP1 gene as Type 2 or Type 7, but the PNG strains were distinct from both genotypes. Among the Chamorro JCV samples, 8 strains (Guam-1) were like the Type 7 strains found in Southeast Asia, while nine strains (Guam-2) were distinct from both the mainland strains and most PNG strains. We identified three JCV variants within Papua New Guinea (PNG-1, PNG-2 and PNG-3), but none of the Southeast Asian (Type 7) strains. PNG-1 strains were present in all three populations (Highlanders and the Baining and Tolai of New Britain), but PNG-2 strains were restricted to the Highlanders. Their relative lack of DNA sequence variation suggests that they arose comparatively recently. The single PNG-3 strain, identified in an Austronesian-speaking Tolai individual, was closely related to the Chamorro variants (Guam-2), consistent with a common Austronesian ancestor. In PNG-2 variants a complex regulatory region mutation inserts a duplication into a nearby deletion, a change reminiscent of those seen in the brains of progressive multifocal leukoencephalopathy patients. This is the first instance of a complex JCV rearrangement circulating in a human population.

Histophilus somni IbpA Fic cytotoxin is conserved in disease strains and most carrier strains from cattle, sheep and bison.

PubMed

Zekarias, B; O'Toole, D; Lehmann, J; Corbeil, L B

2011-04-21

Histophilus somni causes bovine pneumonia, septicemia, myocarditis, thrombotic meningoencephalitis and arthritis, as well as a genital or upper respiratory carrier state in normal animals. However, differences in virulence factors among strains are not well studied. The surface and secreted immunoglobulin binding protein A (IbpA) Fic motif of H. somni causes bovine alveolar type 2 (BAT2) cells to retract, allowing virulent bacteria to cross the alveolar monolayer. Because H. somni IbpA is an important virulence factor, its presence was evaluated in different strains from cattle, sheep and bison to define whether there are syndrome specific markers and whether antigenic/molecular/functional conservation occurs. A few preputial carrier strains lacked IbpA by Western blotting but all other tested disease or carrier strains were IbpA positive. These positive strains had either both IbpA DR1/Fic and IbpA DR2/Fic or only IbpA DR2/Fic by PCR. IbpA Fic mediated cytotoxicity for BAT2 cells and sequence analysis of IbpA DR2/Fic from selected strains revealed conservation of sequence and function in disease and IbpA positive carrier strains. Passive protection of mice against H. somni septicemia with antibody to IbpA DR2/Fic, along with previous data, indicates that the IbpA DR1/Fic and/or DR2/Fic domains are candidate vaccine antigens for protection against many strains of H. somni. Since IbpA DR2/Fic is conserved in most carrier strains, they may be virulent if introduced to susceptible animals at susceptible sites. Conservation of the protective IbpA antigen in all disease isolates tested is encouraging for development of protective vaccines and diagnostic assays. Copyright © 2010 Elsevier B.V. All rights reserved.

Taxonomic study of aromatic-degrading bacteria from deep-terrestrial-subsurface sediments and description of Sphingomonas aromaticivorans sp. nov., Sphingomonas subterranea sp. nov., and Sphingomonas stygia sp. nov.

PubMed

Balkwill, D L; Drake, G R; Reeves, R H; Fredrickson, J K; White, D C; Ringelberg, D B; Chandler, D P; Romine, M F; Kennedy, D W; Spadoni, C M

1997-01-01

Phylogenetic analyses of 16S rRNA gene sequences by distance matrix and parsimony methods indicated that six strains of bacteria isolated from deep saturated Atlantic coastal plain sediments were closely related to the genus Sphingomonas. Five of the strains clustered with, but were distinct from, Sphingomonas capsulata, whereas the sixth strain was most closely related to Blastobacter natatorius. The five strains that clustered with S. capsulata, all of which could degrade aromatic compounds, were gram-negative, non-spore-forming, non-motile, rod-shaped organisms that produced small, yellow colonies on complex media. Their G + C contents ranged from 60.0 to 65.4 mol%, and the predominant isoprenoid quinone was ubiquinone Q-10. All of the strains were aerobic and catalase positive. Indole, urease, and arginine dihydrolase were not produced. Gelatin was not liquified, and glucose was not fermented. Sphingolipids were present in all strains; 2OH14:0 was the major hydroxy fatty acid, and 18:1 was a major constituent of cellular lipids. Acid was produced oxidatively from pentoses, hexoses, and disaccharides, but not from polyalcohols and indole. All of these characteristics indicate that the five aromatic-degrading strains should be placed in the genus Sphingomonas as currently defined. Phylogenetic analysis of 16S rRNA gene sequences, DNA-DNA reassociation values, BOX-PCR genomic fingerprinting, differences in cellular lipid composition, and differences in physiological traits all indicated that the five strains represent three previously undescribed Sphingomonas species. Therefore, we propose the following new species: Sphingomonas aromaticivorans (type strain, SMCC F199), Sphingomonas subterranea (type strain, SMCC B0478), and Sphingomonas stygia (type strain, SMCC B0712).

RNA Sequencing Reveals Differences between the Global Transcriptomes of Salmonella enterica Serovar Enteritidis Strains with High and Low Pathogenicities

PubMed Central

2014-01-01

Salmonella enterica serovar Enteritidis is one of the important causes of bacterial food-borne gastroenteritis worldwide. Field strains of S. Enteritidis are relatively genetically homogeneous; however, they show extensive phenotypic diversity and differences in virulence potential. RNA sequencing (RNA-Seq) was used to characterize differences in the global transcriptome between several genetically similar but phenotypically diverse poultry-associated field strains of S. Enteritidis grown in laboratory medium at avian body temperature (42°C). These S. Enteritidis strains were previously characterized as high-pathogenicity (HP; n = 3) and low-pathogenicity (LP; n = 3) strains based on both in vitro and in vivo virulence assays. Using the negative binomial distribution-based statistical tools edgeR and DESeq, 252 genes were identified as differentially expressed in LP strains compared with their expression in the HP strains (P < 0.05). A majority of genes (235, or 93.2%) showed significantly reduced expression, whereas a few genes (17, or 6.8%) showed increased expression in all LP strains compared with HP strains. LP strains showed a unique transcriptional profile that is characterized by significantly reduced expression of several transcriptional regulators and reduced expression of genes involved in virulence (e.g., Salmonella pathogenicity island 1 [SPI-1], SPI-5, and fimbrial and motility genes) and protection against osmotic, oxidative, and other stresses, such as iron-limiting conditions commonly encountered within the host. Several functionally uncharacterized genes also showed reduced expression. This study provides a first concise view of the global transcriptional differences between field strains of S. Enteritidis with various levels of pathogenicity, providing the basis for future functional characterization of several genes with potential roles in virulence or stress regulation of S. Enteritidis. PMID:24271167

Legionella pneumophila pangenome reveals strain-specific virulence factors.

PubMed

D'Auria, Giuseppe; Jiménez-Hernández, Nuria; Peris-Bondia, Francesc; Moya, Andrés; Latorre, Amparo

2010-03-17

Legionella pneumophila subsp. pneumophila is a gram-negative gamma-Proteobacterium and the causative agent of Legionnaires' disease, a form of epidemic pneumonia. It has a water-related life cycle. In industrialized cities L. pneumophila is commonly encountered in refrigeration towers and water pipes. Infection is always via infected aerosols to humans. Although many efforts have been made to eradicate Legionella from buildings, it still contaminates the water systems. The town of Alcoy (Valencian Region, Spain) has had recurrent outbreaks since 1999. The strain "Alcoy 2300/99" is a particularly persistent and recurrent strain that was isolated during one of the most significant outbreaks between the years 1999-2000. We have sequenced the genome of the particularly persistent L. pneumophila strain Alcoy 2300/99 and have compared it with four previously sequenced strains known as Philadelphia (USA), Lens (France), Paris (France) and Corby (England).Pangenome analysis facilitated the identification of strain-specific features, as well as some that are shared by two or more strains. We identified: (1) three islands related to anti-drug resistance systems; (2) a system for transport and secretion of heavy metals; (3) three systems related to DNA transfer; (4) two CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) systems, known to provide resistance against phage infections, one similar in the Lens and Alcoy strains, and another specific to the Paris strain; and (5) seven islands of phage-related proteins, five of which seem to be strain-specific and two shared. The dispensable genome disclosed by the pangenomic analysis seems to be a reservoir of new traits that have mainly been acquired by horizontal gene transfer and could confer evolutionary advantages over strains lacking them.

Phylogenetic placement of two previously described intranuclear bacteria from the ciliate Paramecium bursaria (Protozoa, Ciliophora): 'Holospora acuminata' and 'Holospora curviuscula'.

PubMed

Rautian, Maria S; Wackerow-Kouzova, Natalia D

2013-05-01

'Holospora acuminata' infects micronuclei of Paramecium bursaria (Protozoa, Ciliophora), whereas 'Holospora curviuscula' infects the macronucleus in other clones of the same host species. Because these micro-organisms have not been cultivated, their description has been based only on some morphological properties and host and nuclear specificities. One16S rRNA gene sequence of 'H. curviuscula' is present in databases. The systematic position of the representative strain of 'H. curviuscula', strain MC-3, was determined in this study. Moreover, for the first time, two strains of 'H. acuminata', KBN10-1 and AC61-10, were investigated. Phylogenetic analysis indicated that all three strains belonged to the genus Holospora, family Holosporaceae, order Rickettsiales within the Alphaproteobacteria.

Mutation of I176R in the E coding region weakens Japanese encephalitis virus neurovirulence, but not its growth rate in BHK-21 cells.

PubMed

Zhou, Yuyong; Wu, Rui; Zhao, Qin; Chang, Yung-Fu; Wen, Xintian; Feng, Yao; Huang, Xiaobo; Wen, Yiping; Yan, Qigui; Huang, Yong; Ma, Xiaoping; Han, Xinfeng; Cao, Sanjie

2018-05-01

Previously, we isolated the Japanese encephalitis virus (JEV) strain SCYA201201. In this study, we passed the SCYA201201 strain in Syrian baby hamster kidney (BHK-21) cells 120 times to obtain the SCYA201201-0901 strain, which exhibited an attenuated phenotype in mice. Comparison of SCYA201201-0901 amino acid sequences with those of other JEV strains revealed a single mutation, I176R, in the E coding region. Using reverse genetic technology, we provide evidence that this single E-I176R mutation does not affect virus growth in BHK-21 cells but significantly decreases JEV neurovirulence in mice. This study provides critical information for understanding the molecular mechanism of JEV attenuation.

Anaerostipes caccae gen. nov., sp. nov., a new saccharolytic, acetate-utilising, butyrate-producing bacterium from human faeces.

PubMed

Schwiertz, Andreas; Hold, Georgina L; Duncan, Sylvia H; Gruhl, Barbel; Collins, Matthew D; Lawson, Paul A; Flint, Harry J; Blaut, Michael

2002-04-01

Two strains of a previously undescribed Eubacterium-like bacterium were isolated from human faeces. The strains are Gram-variable, obligately anaerobic, catalase negative, asporogenous rod-shaped cells which produced acetate, butyrate and lactate as the end products of glucose metabolism. The two isolates displayed 99.9% 16S rRNA gene sequence similarity to each other and treeing analysis demonstrated the faecal isolates are far removed from Eubacterium sensu stricto and that they represent a new subline within the Clostridium coccoides group of organisms. Based on phenotypic and phylogenetic criteria, it is proposed that the two strains from faeces be classified as a new genus and species, Anaerostipes caccae. The type strain of Anaerostipes caccae is NCIMB 13811T (= DSM 14662T).

Involvement of the pagR gene of pXO2 in anthrax pathogenesis

PubMed Central

Liang, Xudong; Zhang, Enmin; Zhang, Huijuan; Wei, Jianchun; Li, Wei; Zhu, Jin; Wang, Bingxiang; Dong, Shulin

2016-01-01

Anthrax is a disease caused by Bacillus anthracis. Specifically, the anthrax toxins and capsules encoded by the pXO1 and pXO2 plasmids, respectively, are the major virulence factors. We previously reported that the pXO1 plasmid was retained in the attenuated strain of B. anthracis vaccine strains even after subculturing at high temperatures. In the present study, we reinvestigate the attenuation mechanism of Pasteur II. Sequencing of pXO1 and pXO2 from Pasteur II strain revealed mutations in these plasmids as compared to the reference sequences. Two deletions on these plasmids, one each on pXO1 and pXO2, were confirmed to be unique to the Pasteur II strain as compared to the wild-type strains. Gene replacement with homologous recombination revealed that the mutation in the promoter region of the pagR gene on pXO2, but not the mutation on pXO1, contributes to lethal levels of toxin production. This result was further confirmed by RT-PCR, western blot, and animal toxicity assays. Taken together, our results signify that the attenuation of the Pasteur II vaccine strain is caused by a mutation in the pagR gene on its pXO2 plasmid. Moreover, these data suggest that pXO2 plasmid encoded proteins are involved in the virulence of B. anthracis. PMID:27363681

Xanthomonas prunicola sp. nov., a novel pathogen that affects nectarine (Prunus persica var. nectarina) trees.

PubMed

López, María M; Lopez-Soriano, Pablo; Garita-Cambronero, Jerson; Beltrán, Carmen; Taghouti, Geraldine; Portier, Perrine; Cubero, Jaime; Fischer-Le Saux, Marion; Marco-Noales, Ester

2018-06-01

Three isolates obtained from symptomatic nectarine trees (Prunus persica var. nectarina) cultivated in Murcia, Spain, which showed yellow and mucoid colonies similar to Xanthomonas arboricola pv. pruni, were negative after serological and real-time PCR analyses for this pathogen. For that reason, these isolates were characterized following a polyphasic approach that included both phenotypic and genomic methods. By sequence analysis of the 16S rRNA gene, these novel strains were identified as members of the genus Xanthomonas, and by multilocus sequence analysis (MLSA) they were clustered together in a distinct group that showed similarity values below 95 % with the rest of the species of this genus. Whole-genome comparisons of the average nucleotide identity (ANI) of genomes of the strains showed less than 91 % average nucleotide identity with all other species of the genus Xanthomonas. Additionally, phenotypic characterization based on API 20 NE, API 50 CH and BIOLOG tests differentiated the strains from the species of the genus Xanthomonas described previously. Moreover, the three strains were confirmed to be pathogenic on peach (Prunus persica), causing necrotic lesions on leaves. On the basis of these results, the novel strains represent a novel species of the genus Xanthomonas, for which the name Xanthomonas prunicola is proposed. The type strain is CFBP 8353 (=CECT 9404=IVIA 3287.1).

Hypervariability generated by natural selection in an extracellular complement-inhibiting protein of serotype M1 strains of group A Streptococcus.

PubMed

Stockbauer, K E; Grigsby, D; Pan, X; Fu, Y X; Mejia, L M; Cravioto, A; Musser, J M

1998-03-17

In many countries, M1 strains of the human pathogenic bacterium group A Streptococcus are the most common serotype recovered from patients with invasive disease episodes. Strains of this serotype express an extracellular protein that inhibits complement [streptococcal inhibitor of complement (Sic)] and is therefore believed to be a virulence factor. Comparative sequence analysis of the 915-bp sic gene in 165 M1 organisms recovered from diverse localities and infection types identified 62 alleles. Inasmuch as multilocus enzyme electrophoresis and pulsed-field gel electrophoresis previously showed that most M1 organisms represent a distinct streptococcal clone, the extent of sic gene polymorphism was unexpected. The level of polymorphism greatly exceeds that recorded for all other genes examined in serotype M1 strains. All insertions and deletions are in frame, and virtually all nucleotide substitutions alter the amino acid sequence of the Sic protein. These molecular features indicate that structural change in Sic is mediated by natural selection. Study of 70 strains recovered from two temporally distinct epidemics of streptococcal infections in the former East Germany found little sharing of Sic variants among strains recovered in the different time periods. Taken together, the data indicate that sic is a uniquely variable gene and provide insight into a potential molecular mechanism contributing to fluctuations in streptococcal disease frequency and severity.

TALE-Like Effectors Are an Ancestral Feature of the Ralstonia solanacearum Species Complex and Converge in DNA Targeting Specificity.

PubMed

Schandry, Niklas; de Lange, Orlando; Prior, Philippe; Lahaye, Thomas

2016-01-01

Ralstonia solanacearum, a species complex of bacterial plant pathogens divided into four monophyletic phylotypes, causes plant diseases in tropical climates around the world. Some strains exhibit a broad host range on solanaceous hosts, while others are highly host-specific as for example some banana-pathogenic strains. Previous studies showed that transcription activator-like (TAL) effectors from Ralstonia, termed RipTALs, are capable of activating reporter genes in planta, if these are preceded by a matching effector binding element (EBE). RipTALs target DNA via their central repeat domain (CRD), where one repeat pairs with one DNA-base of the given EBE. The repeat variable diresidue dictates base repeat specificity in a predictable fashion, known as the TALE code. In this work, we analyze RipTALs across all phylotypes of the Ralstonia solanacearum species complex. We find that RipTALs are prevalent in phylotypes I and IV but absent from most phylotype III and II strains (10/12, 8/14, 1/24, and 1/5 strains contained a RipTAL, respectively). RipTALs originating from strains of the same phylotype show high levels of sequence similarity (>98%) in the N-terminal and C-terminal regions, while RipTALs isolated from different phylotypes show 47-91% sequence similarity in those regions, giving rise to four RipTAL classes. We show that, despite sequence divergence, the base preference for guanine, mediated by the N-terminal region, is conserved across RipTALs of all classes. Using the number and order of repeats found in the CRD, we functionally sub-classify RipTALs, introduce a new simple nomenclature, and predict matching EBEs for all seven distinct RipTALs identified. We experimentally study RipTAL EBEs and uncover that some RipTALs are able to target the EBEs of other RipTALs, referred to as cross-reactivity. In particular, RipTALs from strains with a broad host range on solanaceous hosts cross-react on each other's EBEs. Investigation of sequence divergence between RipTAL repeats allows for a reconstruction of repeat array biogenesis, for example through slipped strand mispairing or gene conversion. Using these studies we show how RipTALs of broad host range strains evolved convergently toward a shared target sequence. Finally, we discuss the differences between TALE-likes of plant pathogens in the context of disease ecology.

Phenotypic and genotypic discrepancy of Streptococcus pneumoniae strains isolated from Asian countries.

PubMed

Ko, Kwan Soo; Oh, Won Sup; Peck, Kyong Ran; Lee, Jang Ho; Lee, Nam Yong; Song, Jae-Hoon

2005-07-01

Non-typeable isolates of Streptococcus pneumoniae collected from Asian countries were characterized by optochin susceptibility test, bile solubility test, multilocus sequence typing of housekeeping genes, amplification of virulence-related genes, 16S rDNA-RsaI digestion, and 16S rDNA sequencing. Six of 54 non-typeable pneumococcal isolates showed divergence of gene sequences of recP and xpt from typical pneumococcal strains. Of these six atypical pneumococcal strains, two showed different results in optochin susceptibility or bile solubility test from typical pneumococcal strains. All six isolates showed high sequence dissimilarities of multilocus sequence typing, 16S rDNA sequences, and lytA sequences from typical S. pneumoniae strains. Data from this study suggest that classic tests such as optochin susceptibility and bile solubility tests may lead to incorrect identification of S. pneumoniae. These atypical strains may belong to different bacterial species from S. pneumoniae.

Multiple mobile promoter regions for the rare carbapenem resistance gene of Bacteroides fragilis.

PubMed

Podglajen, I; Breuil, J; Rohaut, A; Monsempes, C; Collatz, E

2001-06-01

Two novel insertion sequences (IS), IS1187 and IS1188, are described upstream from the carbapenem resistance gene cfiA in strains of Bacteroides fragilis. Mapping, with the RACE procedure, of transcription start sites of cfiA in these and two other previously reported IS showed that transcription of this rarely encountered gene is initiated close to a variety of B. fragilis consensus promoter sequences, as recently defined (D. P. Bayley, E. R. Rocha, and C. J. Smith, FEMS Microbiol. Lett. 193:149-154, 2000). In the cases of IS1186 and IS1188, these sequences overlap with putative Esigma(70) promoter sequences, while in IS942 and IS1187 such sequences can be observed either upstream or downstream of the B. fragilis promoters.

Veterinary Fusarioses within the United States

PubMed Central

Sutton, Deanna A.; Wiederhold, Nathan; Robert, Vincent A. R. G.; Crous, Pedro W.; Geiser, David M.

2016-01-01

Multilocus DNA sequence data were used to assess the genetic diversity and evolutionary relationships of 67 Fusarium strains from veterinary sources, most of which were from the United States. Molecular phylogenetic analyses revealed that the strains comprised 23 phylogenetically distinct species, all but two of which were previously known to infect humans, distributed among eight species complexes. The majority of the veterinary isolates (47/67 = 70.1%) were nested within the Fusarium solani species complex (FSSC), and these included 8 phylospecies and 33 unique 3-locus sequence types (STs). Three of the FSSC species (Fusarium falciforme, Fusarium keratoplasticum, and Fusarium sp. FSSC 12) accounted for four-fifths of the veterinary strains (38/47) and STs (27/33) within this clade. Most of the F. falciforme strains (12/15) were recovered from equine keratitis infections; however, strains of F. keratoplasticum and Fusarium sp. FSSC 12 were mostly (25/27) isolated from marine vertebrates and invertebrates. Our sampling suggests that the Fusarium incarnatum-equiseti species complex (FIESC), with eight mycoses-associated species, may represent the second most important clade of veterinary relevance within Fusarium. Six of the multilocus STs within the FSSC (3+4-eee, 1-b, 12-a, 12-b, 12-f, and 12-h) and one each within the FIESC (1-a) and the Fusarium oxysporum species complex (ST-33) were widespread geographically, including three STs with transoceanic disjunctions. In conclusion, fusaria associated with veterinary mycoses are phylogenetically diverse and typically can only be identified to the species level using DNA sequence data from portions of one or more informative genes. PMID:27605713

A novel amino acid substitution in a voltage-gated sodium channel is associated with knockdown resistance to permethrin in Aedes aegypti.

PubMed

Chang, Cheng; Shen, Wen-Kai; Wang, Tzu-Ting; Lin, Ying-Hsi; Hsu, Err-Lieh; Dai, Shu-Mei

2009-04-01

To identify pertinent mutations associated with knockdown resistance to permethrin, the entire coding sequence of the voltage-gated sodium channel gene Aa-para was sequenced and analyzed from a Per-R strain with 190-fold resistance to permethrin and two susceptible strains of Aedes aegypti. The longest transcript, a 6441bp open reading frame, encodes 2147 amino acid residues with an estimated molecular mass of 241kDa. A total of 33 exons were found in the Aa-para gene over 293kb of genomic DNA. Three previously unreported optional exons were identified. The first two exons, m and n, were located within the intracellular domain I/II, and the third, f', was found within the II/III linkers. The two mutually exclusive exons, d and l, were the only alternative exons in all the cDNA clones sequenced in this study. The most distinct finding was a novel amino acid substitution mutation, D1794Y, located within the extracellular linker between IVS5 and IVS6, which is concurrent with the known V1023G mutation in Aa-para of the Per-R strain. The high frequency and coexistence of the two mutations in the Per-R strain suggest that they might exert a synergistic effect to provide the knockdown resistance to permethrin. Furthermore, both cDNA and genomic DNA data from the same individual mosquitoes have demonstrated that RNA editing was not involved in amino acid substitutions of the Per-R strain.

Veterinary Fusarioses within the United States.

PubMed

O'Donnell, Kerry; Sutton, Deanna A; Wiederhold, Nathan; Robert, Vincent A R G; Crous, Pedro W; Geiser, David M

2016-11-01

Multilocus DNA sequence data were used to assess the genetic diversity and evolutionary relationships of 67 Fusarium strains from veterinary sources, most of which were from the United States. Molecular phylogenetic analyses revealed that the strains comprised 23 phylogenetically distinct species, all but two of which were previously known to infect humans, distributed among eight species complexes. The majority of the veterinary isolates (47/67 = 70.1%) were nested within the Fusarium solani species complex (FSSC), and these included 8 phylospecies and 33 unique 3-locus sequence types (STs). Three of the FSSC species (Fusarium falciforme, Fusarium keratoplasticum, and Fusarium sp. FSSC 12) accounted for four-fifths of the veterinary strains (38/47) and STs (27/33) within this clade. Most of the F. falciforme strains (12/15) were recovered from equine keratitis infections; however, strains of F. keratoplasticum and Fusarium sp. FSSC 12 were mostly (25/27) isolated from marine vertebrates and invertebrates. Our sampling suggests that the Fusarium incarnatum-equiseti species complex (FIESC), with eight mycoses-associated species, may represent the second most important clade of veterinary relevance within Fusarium Six of the multilocus STs within the FSSC (3+4-eee, 1-b, 12-a, 12-b, 12-f, and 12-h) and one each within the FIESC (1-a) and the Fusarium oxysporum species complex (ST-33) were widespread geographically, including three STs with transoceanic disjunctions. In conclusion, fusaria associated with veterinary mycoses are phylogenetically diverse and typically can only be identified to the species level using DNA sequence data from portions of one or more informative genes. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Comparative Genomic Analysis for Genetic Variation in Sacbrood Virus of Apis cerana and Apis mellifera Honeybees From Different Regions of Vietnam.

PubMed

Reddy, Kondreddy Eswar; Thu, Ha Thi; Yoo, Mi Sun; Ramya, Mummadireddy; Reddy, Bheemireddy Anjana; Lien, Nguyen Thi Kim; Trang, Nguyen Thi Phuong; Duong, Bui Thi Thuy; Lee, Hyun-Jeong; Kang, Seung-Won; Quyen, Dong Van

2017-09-01

Sacbrood virus (SBV) is one of the most common viral infections of honeybees. The entire genome sequence for nine SBV infecting honeybees, Apis cerana and Apis mellifera, in Vietnam, namely AcSBV-Viet1, AcSBV-Viet2, AcSBV-Viet3, AmSBV-Viet4, AcSBV-Viet5, AmSBV-Viet6, AcSBV-Viet7, AcSBV-Viet8, and AcSBV-Viet9, was determined. These sequences were aligned with seven previously reported complete genome sequences of SBV from other countries, and various genomic regions were compared. The Vietnamese SBVs (VN-SBVs) shared 91-99% identity with each other, and shared 89-94% identity with strains from other countries. The open reading frames (ORFs) of the VN-SBV genomes differed greatly from those of SBVs from other countries, especially in their VP1 sequences. The AmSBV-Viet6 and AcSBV-Viet9 genome encodes 17 more amino acids within this region than the other VN-SBVs. In a phylogenetic analysis, the strains AmSBV-Viet4, AcSBV-Viet2, and AcSBV-Viet3 were clustered in group with AmSBV-UK, AmSBV-Kor21, and AmSBV-Kor19 strains. Whereas, the strains AmSBV-Viet6 and AcSBV-Viet7 clustered separately with the AcSBV strains from Korea and AcSBV-VietSBM2. And the strains AcSBV-Viet8, AcSBV-Viet1, AcSBV-Viet5, and AcSBV-Viet9 clustered with the AcSBV-India, AcSBV-Kor and AcSBV-VietSBM2. In a Simplot graph, the VN-SBVs diverged stronger in their ORF regions than in their 5' or 3' untranslated regions. The VN-SBVs possess genetic characteristics which are more similar to the Asian AcSBV strains than to AmSBV-UK strain. Taken together, our data indicate that host specificity, geographic distance, and viral cross-infections between different bee species may explain the genetic diversity among the VN-SBVs in A. cerana and A. mellifera and other SBV strains. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America.

Tetrahymena australis (Protozoa, Ciliophora): A Well-Known But "Non-Existing" Taxon - Consideration of Its Identification, Definition and Systematic Position.

PubMed

Liu, Mingjian; Fan, Xinpeng; Gao, Feng; Gao, Shan; Yu, Yuhe; Warren, Alan; Huang, Jie

2016-11-01

A cryptic species of the Tetrahymena pyriformis complex, Tetrahymena australis, has been known for a long time but never properly diagnosed based on taxonomic methods. The species name is thus invalid according to the International Code of Zoological Nomenclature. Recently, a population isolated from a freshwater lake in Wuhan, China was investigated using live observations, silver staining methods and gene sequence data. This organism can be separated from other described species of the T. pyriformis complex by its relatively small body size, the number of somatic kineties and differences in sequences of two genes, namely the small subunit ribosomal RNA (SSU rRNA) and the mitochondrial cytochrome c oxidase subunit I (cox1). We compared the SSU rRNA gene sequences of all available Tetrahymena species to reveal the nucleotide differences within this genus. The sequence of the Wuhan population is identical to two sequences of a previously isolated strain of T. australis (ATCC #30831). Phylogenetic analyses indicate that these three sequences (X56167, M98015, KT334373) cluster with Tetrahymena shanghaiensis (EF070256) in a polytomy. However, sequence divergence of the cox1 gene between the Wuhan population and another strain of T. australis (ATCC #30271) is 1.4%, suggesting that these may represent different subspecies. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

Enteroaggregative Escherichia coli is the predominant diarrheagenic E. coli pathotype among irrigation water and food sources in South Africa.

PubMed

Aijuka, Matthew; Santiago, Araceli E; Girón, Jorge A; Nataro, James P; Buys, Elna M

2018-08-02

Diarrheagenic E. coli (DEC) has been implicated in foodborne outbreaks worldwide and have been associated with childhood stunting in the absence of diarrhoea. Infection is extraordinarily common, but the routes of transmission have not been determined. Therefore, determining the most prevalent pathotypes in food and environmental sources may help provide better guidance to various stakeholders in ensuring food safety and public health and advancing understanding of the epidemiology of enteric disease. We characterized 205 E. coli strains previously isolated from producer distributor bulk milk (PDBM)(118), irrigation water (48), irrigated lettuce (29) and street vendor coleslaw (10) in South Africa. Enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroaggregative E. coli (EAEC) and diffusely adherent E. coli (DAEC) were sought. We used PCR and partial gene sequencing for all 205 strains while 46 out of 205 that showed poor resolution were subsequently characterized using cell adherence (HeLa cells). PCR and partial gene sequencing of aatA and/or aaiC genes confirmed EAEC (2%, 5 out of 205) as the only pathotype. Phylogenetic analysis of sequenced EAEC strains with E. coli strains in GenBank showing ≥80% nucleotide sequence similarity based on possession of aaiC and aatA generated distinct clusters of strains separated predominantly based on their source of isolation (food source or human stool) suggesting a potential role of virulence genes in source tracking. EAEC 24%, 11 out of 46 strains (PDBM = 15%, irrigation water = 7%, irrigated lettuce = 2%) was similarly the predominant pathotype followed by strains showing invasiveness to HeLa cells, 4%, 2 out of 46 (PDBM = 2%, irrigated lettuce = 2%), among stains characterized using cell adherence. Therefore, EAEC may be the leading cause of DEC associated food and water-borne enteric infection in South Africa. Additionally, solely using molecular based methods targeting virulence gene determinants may underestimate prevalence, especially among heterogeneous pathogens such as EAEC. Copyright © 2018 Elsevier B.V. All rights reserved.

Listeria booriae sp. nov. and Listeria newyorkensis sp. nov., from food processing environments in the USA.

PubMed

Weller, Daniel; Andrus, Alexis; Wiedmann, Martin; den Bakker, Henk C

2015-01-01

Sampling of seafood and dairy processing facilities in the north-eastern USA produced 18 isolates of Listeria spp. that could not be identified at the species-level using traditional phenotypic and genotypic identification methods. Results of phenotypic and genotypic analyses suggested that the isolates represent two novel species with an average nucleotide blast identity of less than 92% with previously described species of the genus Listeria. Phylogenetic analyses based on whole genome sequences, 16S rRNA gene and sigB gene sequences confirmed that the isolates represented by type strain FSL M6-0635(T) and FSL A5-0209 cluster phylogenetically with Listeria cornellensis. Phylogenetic analyses also showed that the isolates represented by type strain FSL A5-0281(T) cluster phylogenetically with Listeria riparia. The name Listeria booriae sp. nov. is proposed for the species represented by type strain FSL A5-0281(T) ( =DSM 28860(T) =LMG 28311(T)), and the name Listeria newyorkensis sp. nov. is proposed for the species represented by type strain FSL M6-0635(T) ( =DSM 28861(T) =LMG 28310(T)). Phenotypic and genotypic analyses suggest that neither species is pathogenic. © 2015 IUMS.

Identification of chitinolytic bacteria isolated from shrimp pond sediment and characterization of their chitinase encoding gene

NASA Astrophysics Data System (ADS)

Triwijayani, A. U.; Puspita, I. D.; Murwantoko; Ustadi

2018-03-01

Chitinolytic bacteria are a group of bacteria owning enzymes that able to hydrolyze chitin. Previously, we isolated chitinolytic bacteria from shrimp pond sediment in Bantul, Yogyakarta, and obtained five isolates showing high chitinolytic index named as isolate PT1, PT2, PT5, PT6 and PB2. The aims of this study were to identify chitinolytic bacteria isolated from shrimp pond sediment and to characterize the chitinase encoding gene from each isolate. The molecular technique was performed by amplification of 16S rDNA, amplification of chitinase encoding gene and sequence analysis. Two chitinolytic bacteria of PT1 and PT2 were similar to Aeromonas bivalvium strain D15, PT5 to Pseudomonas stutzeri strain BD-2.2.1, PT6 to Serratia marcescens strain FZSF02 and PB2 to Streptomyces misionensis strain OsiRt-1. The comparison of chitinase encoding gene between three isolates with those in Gen Bank shows that PT1 had similar sequences with the chi1 gene in Aeromonas sp. 17m, PT2 with chi1 gene in A. caviae (CB101) and PT6 with chiB gene in S. Marcescens (BJL200).

Genome sequences of two closely related strains of Escherichia coli K-12 GM4792.

PubMed

Zhang, Yan-Cong; Zhang, Yan; Zhu, Bi-Ru; Zhang, Bo-Wen; Ni, Chuan; Zhang, Da-Yong; Huang, Ying; Pang, Erli; Lin, Kui

2015-01-01

Escherichia coli lab strains K-12 GM4792 Lac(+) and GM4792 Lac(-) carry opposite lactose markers, which are useful for distinguishing evolved lines as they produce different colored colonies. The two closely related strains are chosen as ancestors for our ongoing studies of experimental evolution. Here, we describe the genome sequences, annotation, and features of GM4792 Lac(+) and GM4792 Lac(-). GM4792 Lac(+) has a 4,622,342-bp long chromosome with 4,061 protein-coding genes and 83 RNA genes. Similarly, the genome of GM4792 Lac(-) consists of a 4,621,656-bp chromosome containing 4,043 protein-coding genes and 74 RNA genes. Genome comparison analysis reveals that the differences between GM4792 Lac(+) and GM4792 Lac(-) are minimal and limited to only the targeted lac region. Moreover, a previous study on competitive experimentation indicates the two strains are identical or nearly identical in survivability except for lactose utilization in a nitrogen-limited environment. Therefore, at both a genetic and a phenotypic level, GM4792 Lac(+) and GM4792 Lac(-), with opposite neutral markers, are ideal systems for future experimental evolution studies.

Genomic Variability of O Islands Encoding Tellurite Resistance in Enterohemorrhagic Escherichia coli O157:H7 Isolates

PubMed Central

Taylor, Diane E.; Rooker, Michelle; Keelan, Monika; Ng, Lai-King; Martin, Irene; Perna, Nicole T.; Burland, N. T. Valerie; Blattner, Fredrick R.

2002-01-01

Strains of Escherichia coli causing enterohemorrhagic colitis belonging to the O157:H7 lineage are reported to be highly related. Fifteen strains of E. coli O157:H7 and 1 strain of E. coli O46:H− (nonflagellated) were examined for the presence of potassium tellurite resistance (Ter). Ter genes comprising terABCDEF were shown previously to be part of a pathogenicity island also containing integrase, phage, and urease genes. PCR analysis, both conventional and light cycler based, demonstrated that about one-half of the Ter E. coli O157:H7 strains (6 of 15), including the Sakai strain, which has been sequenced, carried a single copy of the Ter genes. Five of the strains, including EDL933, which has also been sequenced, contained two copies. Three other O157:H7 strains and the O46:H− strain did not contain the Ter genes. In strains containing two copies, the Ter genes were associated with the serW and serX tRNA genes. Five O157:H7 strains resembled the O157 Sakai strain whose sequence contained one copy, close to serX, whereas in one isolate the single copy was associated with serW. There was no correlation between Ter and the ability to produce Shiga toxin ST1 or ST2. The Ter MIC for most strains, containing either one or two copies, was 1,024 μg/ml, although for a few the MIC was intermediate, 64 to 128 μg/ml, which could be increased to 512 μg/ml by pregrowth of strains in subinhibitory concentrations of potassium tellurite. Reverse transcriptase PCR analysis confirmed that in most strains Ter was constitutive but that in the rest it was inducible and involved induction of terB and terC genes. Only the terB, -C, -D, and -E genes are required for Ter. The considerable degree of homology between the ter genes on IncH12 plasmid R478, which originated in Serratia marcescens, and pTE53, from an E. coli clinical isolate, suggests that the pathogenicity island was acquired from a plasmid. This work demonstrates diversity among E. coli O157:H7 isolates, at least as far as the presence of Ter genes is concerned. PMID:12169592

Natural Variant of Collagen-Like Protein A in Serotype M3 Group A Streptococcus Increases Adherence and Decreases Invasive Potential

PubMed Central

Jewell, Brittany E.; Versalovic, Erika M.; Olsen, Randall J.; Bachert, Beth A.; Lukomski, Slawomir; Musser, James M.

2015-01-01

Group A Streptococcus (GAS) predominantly exists as a colonizer of the human oropharynx that occasionally breaches epithelial barriers to cause invasive diseases. Despite the frequency of GAS carriage, few investigations into the contributory molecular mechanisms exist. To this end, we identified a naturally occurring polymorphism in the gene encoding the streptococcal collagen-like protein A (SclA) in GAS carrier strains. All previously sequenced invasive serotype M3 GAS possess a premature stop codon in the sclA gene truncating the protein. The carrier polymorphism is predicted to restore SclA function and was infrequently identified by targeted DNA sequencing in invasive strains of the same serotype. We demonstrate that a strain with the carrier sclA allele expressed a full-length SclA protein, while the strain with the invasive sclA allele expressed a truncated variant. An isoallelic mutant invasive strain with the carrier sclA allele exhibited decreased virulence in a mouse model of invasive disease and decreased multiplication in human blood. Further, the isoallelic invasive strain with the carrier sclA allele persisted in the mouse nasopharynx and had increased adherence to cultured epithelial cells. Repair of the premature stop codon in the invasive sclA allele restored the ability to bind the extracellular matrix proteins laminin and cellular fibronectin. These data demonstrate that a mutation in GAS carrier strains increases adherence and decreases virulence and suggest selection against increased adherence in GAS invasive isolates. PMID:25561712

Emergence of Novel Human Norovirus GII.17 Strains Correlates With Changes in Blockade Antibody Epitopes.

PubMed

Lindesmith, Lisa C; Kocher, Jacob F; Donaldson, Eric F; Debbink, Kari; Mallory, Michael L; Swann, Excel W; Brewer-Jensen, Paul D; Baric, Ralph S

2017-12-05

Human norovirus is a significant public health burden, with >30 genotypes causing endemic levels of disease and strains from the GII.4 genotype causing serial pandemics as the virus evolves new ligand binding and antigenicity features. During 2014-2015, genotype GII.17 cluster IIIb strains emerged as the leading cause of norovirus infection in select global locations. Comparison of capsid sequences indicates that GII.17 is evolving at previously defined GII.4 antibody epitopes. Antigenicity of virus-like particles (VLPs) representative of clusters I, II, and IIIb GII.17 strains were compared by a surrogate neutralization assay based on antibody blockade of ligand binding. Sera from mice immunized with a single GII.17 VLP identified antigenic shifts between each cluster of GII.17 strains. Ligand binding of GII.17 cluster IIIb VLP was blocked only by antisera from mice immunized with cluster IIIb VLPs. Exchange of residues 393-396 from GII.17.2015 into GII.17.1978 ablated ligand binding and altered antigenicity, defining an important varying epitope in GII.17. The capsid sequence changes in GII.17 strains result in loss of blockade antibody binding, indicating that viral evolution, specifically at residues 393-396, may have contributed to the emergence of cluster IIIb strains and the persistence of GII.17 in human populations. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Sequence comparison of phoR, gyrB, groEL, and cheA genes as phylogenetic markers for distinguishing Bacillus amyloliquefaciens and B. subtilis and for identifying Bacillus strain B29.

PubMed

Yu, C; Jin, J; Meng, L-Q; Xia, H-H; Yuan, H-F; Wang, J; Yu, D-S; Zhao, X-Y; Sha, C-Q

2017-05-20

Given the close genetic relationship between Bacillus amyloliquefaciens and B. subtilis, distinguishing the two solely based on their physiological and biochemical characteristics and 16S rRNA sequences is difficult. Molecular identification was used to discover suitable genes for distinguishing the two bacteria, and to identify the bio-controlling strain B29, due to molecular identification has been paid more and more attention. The similarity of four genes, cheA, gyrB, groEL and phoR, of the two species was compared by the software BLASTN and MAGA, and phylogenetic tree was constructed. The B29 strain was re-identified by using the screened genes. The similarities of the four genes, gyrB, groEL, cheA and phoR, of the two species were 93-95%, 82-84%, 76-78% and 76-77%, respectively. The homologies of the four genes of the strain B29 and the strains of B. amyloliquefaciens strains were more than 95%. We determined how well the phoR and cheA genes could be used to differentiate B. amyloliquefacien and B. subtilis. The previously isolated biological control strain B29, initially classified as B. subtilis, was re-classified as B. amyloliquefaciens. Our data indicate that other than the phoR gene, the cheA gene might be a useful phylogenetic marker for differentiating B. subtilis and B. amyloliquefaciens.

Cultured representatives of two major phylogroups of human colonic Faecalibacterium prausnitzii can utilize pectin, uronic acids, and host-derived substrates for growth.

PubMed

Lopez-Siles, Mireia; Khan, Tanweer M; Duncan, Sylvia H; Harmsen, Hermie J M; Garcia-Gil, L Jesús; Flint, Harry J

2012-01-01

Faecalibacterium prausnitzii is one of the most abundant commensal bacteria in the healthy human large intestine, but information on genetic diversity and substrate utilization is limited. Here, we examine the phylogeny, phenotypic characteristics, and influence of gut environmental factors on growth of F. prausnitzii strains isolated from healthy subjects. Phylogenetic analysis based on the 16S rRNA sequences indicated that the cultured strains were representative of F. prausnitzii sequences detected by direct analysis of fecal DNA and separated the available isolates into two phylogroups. Most F. prausnitzii strains tested grew well under anaerobic conditions on apple pectin. Furthermore, F. prausnitzii strains competed successfully in coculture with two other abundant pectin-utilizing species, Bacteroides thetaiotaomicron and Eubacterium eligens, with apple pectin as substrate, suggesting that this species makes a contribution to pectin fermentation in the colon. Many F. prausnitzii isolates were able to utilize uronic acids for growth, an ability previously thought to be confined to Bacteroides spp. among human colonic anaerobes. Most strains grew on N-acetylglucosamine, demonstrating an ability to utilize host-derived substrates. All strains tested were bile sensitive, showing at least 80% growth inhibition in the presence of 0.5 μg/ml bile salts, while inhibition at mildly acidic pH was strain dependent. These attributes help to explain the abundance of F. prausnitzii in the colonic community but also suggest factors in the gut environment that may limit its distribution.

Arsenite as an electron donor for anoxygenic photosynthesis: Description of three strains of Ectothiorhodospria from Mono Lake, California, and Big Soda Lake, Nevada

USGS Publications Warehouse

McCann, Shelley; Boren, Alison; Hernandez-Maldonado, Jaime; Stoneburner, Brendon; Saltikov, Chad W; Stolz, John F.; Oremland, Ronald S.

2017-01-01

Three novel strains of photosynthetic bacteria from the family Ectothiorhodospiraceae were isolated from soda lakes of the Great Basin Desert, USA by employing arsenite (As(III)) as the sole electron donor in the enrichment/isolation process. Strain PHS-1 was previously isolated from a hot spring in Mono Lake, while strain MLW-1 was obtained from Mono Lake sediment, and strain BSL-9 was isolated from Big Soda Lake. Strains PHS-1, MLW-1, and BSL-9 were all capable of As(III)-dependent growth via anoxygenic photosynthesis and contained homologs of arxA, but displayed different phenotypes. Comparisons were made with three related species: Ectothiorhodospira shaposhnikovii DSM 2111, Ectothiorhodospira shaposhnikovii DSM 243T, and Halorhodospira halophila DSM 244. All three type cultures oxidized arsenite to arsenate but did not grow with As(III) as the sole electron donor. DNA–DNA hybridization indicated that strain PHS-1 belongs to the same species as Ect. shaposhnikovii DSM 2111 (81.1% sequence similarity), distinct from Ect. shaposhnikovii DSM 243T (58.1% sequence similarity). These results suggest that the capacity for light-driven As(III) oxidation is a common phenomenon among purple photosynthetic bacteria in soda lakes. However, the use of As(III) as a sole electron donor to sustain growth via anoxygenic photosynthesis is confined to novel isolates that were screened for by this selective cultivation criterion.

Cultured Representatives of Two Major Phylogroups of Human Colonic Faecalibacterium prausnitzii Can Utilize Pectin, Uronic Acids, and Host-Derived Substrates for Growth

PubMed Central

Lopez-Siles, Mireia; Khan, Tanweer M.; Duncan, Sylvia H.; Harmsen, Hermie J. M.; Garcia-Gil, L. Jesús

2012-01-01

Faecalibacterium prausnitzii is one of the most abundant commensal bacteria in the healthy human large intestine, but information on genetic diversity and substrate utilization is limited. Here, we examine the phylogeny, phenotypic characteristics, and influence of gut environmental factors on growth of F. prausnitzii strains isolated from healthy subjects. Phylogenetic analysis based on the 16S rRNA sequences indicated that the cultured strains were representative of F. prausnitzii sequences detected by direct analysis of fecal DNA and separated the available isolates into two phylogroups. Most F. prausnitzii strains tested grew well under anaerobic conditions on apple pectin. Furthermore, F. prausnitzii strains competed successfully in coculture with two other abundant pectin-utilizing species, Bacteroides thetaiotaomicron and Eubacterium eligens, with apple pectin as substrate, suggesting that this species makes a contribution to pectin fermentation in the colon. Many F. prausnitzii isolates were able to utilize uronic acids for growth, an ability previously thought to be confined to Bacteroides spp. among human colonic anaerobes. Most strains grew on N-acetylglucosamine, demonstrating an ability to utilize host-derived substrates. All strains tested were bile sensitive, showing at least 80% growth inhibition in the presence of 0.5 μg/ml bile salts, while inhibition at mildly acidic pH was strain dependent. These attributes help to explain the abundance of F. prausnitzii in the colonic community but also suggest factors in the gut environment that may limit its distribution. PMID:22101049

Genomics-enabled analysis of the emergent disease cotton bacterial blight

PubMed Central

Phillips, Anne Z.; Burke, Jillian; Bunn, J. Imani; Allen, Tom W.; Wheeler, Terry

2017-01-01

Cotton bacterial blight (CBB), an important disease of (Gossypium hirsutum) in the early 20th century, had been controlled by resistant germplasm for over half a century. Recently, CBB re-emerged as an agronomic problem in the United States. Here, we report analysis of cotton variety planting statistics that indicate a steady increase in the percentage of susceptible cotton varieties grown each year since 2009. Phylogenetic analysis revealed that strains from the current outbreak cluster with race 18 Xanthomonas citri pv. malvacearum (Xcm) strains. Illumina based draft genomes were generated for thirteen Xcm isolates and analyzed along with 4 previously published Xcm genomes. These genomes encode 24 conserved and nine variable type three effectors. Strains in the race 18 clade contain 3 to 5 more effectors than other Xcm strains. SMRT sequencing of two geographically and temporally diverse strains of Xcm yielded circular chromosomes and accompanying plasmids. These genomes encode eight and thirteen distinct transcription activator-like effector genes. RNA-sequencing revealed 52 genes induced within two cotton cultivars by both tested Xcm strains. This gene list includes a homeologous pair of genes, with homology to the known susceptibility gene, MLO. In contrast, the two strains of Xcm induce different clade III SWEET sugar transporters. Subsequent genome wide analysis revealed patterns in the overall expression of homeologous gene pairs in cotton after inoculation by Xcm. These data reveal important insights into the Xcm-G. hirsutum disease complex and strategies for future development of resistant cultivars. PMID:28910288

Complete Genome Sequence of a CTX-M-15-Producing Escherichia coli Strain from the H30Rx Subclone of Sequence Type 131 from a Patient with Recurrent Urinary Tract Infections, Closely Related to a Lethal Urosepsis Isolate from the Patient’s Sister

PubMed Central

Johnson, Timothy J.; Liu, Cindy M.; Sokurenko, Evgeni; Kisiela, Dagmara I.; Paul, Sandip; Andersen, Paal; Johnson, James R.; Price, Lance B.

2016-01-01

We report here the complete genome sequence, including five plasmid sequences, of Escherichia coli sequence type 131 (ST131) strain JJ1887. The strain was isolated in 2007 in the United States from a patient with recurrent cystitis, whose caregiver sister died from urosepsis caused by a nearly identical strain. PMID:27174264

Molecular Tracking, through Processing, of Campylobacter Strains Colonizing Broiler Flocks▿

PubMed Central

Elvers, Karen T.; Morris, Victoria K.; Newell, Diane G.; Allen, Vivien M.

2011-01-01

Many of the poultry flocks produced in the United Kingdom are colonized with Campylobacter, and the intensive nature of poultry processing usually results in contaminated carcasses. In this study, a previously reported molecular oligonucleotide probe method was used to track a specific flock-colonizing strain(s) on broiler carcasses during processing in two United Kingdom commercial poultry processing plants. Five Campylobacter-positive flocks were sampled at four points along the processing line, postbleed, postpluck, prechill, and postchill, and two Campylobacter-negative flocks processed immediately after positive flocks were sampled prechill. flaA was sequenced from Campylobacter strains isolated from these flocks, and strain-specific probes were synthesized. Skin and cecal samples were plated onto selective agar to give individual colonies, which were transferred onto membranes. These were then hybridized with the strain- and genus-specific probes. For all the 5 positive flocks, there was a significant reduction in campylobacters postbleed compared to postpluck but no subsequent fall on sampling pre- and postchill, and the strain(s) predominating on the carcasses throughout processing came from the flock being processed. This indicates that strains from the abattoir environment were not a significant cause of carcass contamination in flocks with well-established campylobacter colonization. However, negative flocks that were preceded by positive flocks were contaminated by strains that did not generally originate from the predominating strains recovered from the ceca of the previous positive flocks. This suggests that the abattoir environment has a significant role in the contamination of carcasses from negative but not fully colonized flocks. PMID:21705532

More Easily Cultivated Than Identified: Classical Isolation With Molecular Identification of Vaginal Bacteria.

PubMed

Srinivasan, Sujatha; Munch, Matthew M; Sizova, Maria V; Fiedler, Tina L; Kohler, Christina M; Hoffman, Noah G; Liu, Congzhou; Agnew, Kathy J; Marrazzo, Jeanne M; Epstein, Slava S; Fredricks, David N

2016-08-15

Women with bacterial vaginosis (BV) have complex communities of anaerobic bacteria. There are no cultivated isolates of several bacteria identified using molecular methods and associated with BV. It is unclear whether this is due to the inability to adequately propagate these bacteria or to correctly identify them in culture. Vaginal fluid from 15 women was plated on 6 different media using classical cultivation approaches. Individual isolates were identified by 16S ribosomal RNA (rRNA) gene sequencing and compared with validly described species. Bacterial community profiles in vaginal samples were determined using broad-range 16S rRNA gene polymerase chain reaction and pyrosequencing. We isolated and identified 101 distinct bacterial strains spanning 6 phyla including (1) novel strains with <98% 16S rRNA sequence identity to validly described species, (2) closely related species within a genus, (3) bacteria previously isolated from body sites other than the vagina, and (4) known bacteria formerly isolated from the vagina. Pyrosequencing showed that novel strains Peptoniphilaceae DNF01163 and Prevotellaceae DNF00733 were prevalent in women with BV. We isolated a diverse set of novel and clinically significant anaerobes from the human vagina using conventional approaches with systematic molecular identification. Several previously "uncultivated" bacteria are amenable to conventional cultivation. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.