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Sample records for attenuated chlamydia abortus

  1. Can Chlamydia abortus be transmitted by embryo transfer in goats?

    PubMed

    Oseikria, M; Pellerin, J L; Rodolakis, A; Vorimore, F; Laroucau, K; Bruyas, J F; Roux, C; Michaud, S; Larrat, M; Fieni, F

    2016-10-01

    The objectives of this study were to determine (i) whether Chlamydia abortus would adhere to or penetrate the intact zona pellucida (ZP-intact) of early in vivo-derived caprine embryos, after in vitro infection; and (ii) the efficacy of the International Embryo Transfer Society (IETS) washing protocol for bovine embryos. Fifty-two ZP-intact embryos (8-16 cells), obtained from 14 donors were used in this experiment. The embryos were randomly divided into 12 batches. Nine batches (ZP-intact) of five embryos were incubated in a medium containing 4 × 10(7)Chlamydia/mL of AB7 strain. After incubation for 18 hours at 37 °C in an atmosphere of 5% CO2, the embryos were washed in batches in 10 successive baths of a phosphate buffer saline and 5% fetal calf serum solution in accordance with IETS guidelines. In parallel, three batches of ZP-intact embryos were used as controls by being subjected to similar procedures but without exposure to C. abortus. The 10 wash baths were collected separately and centrifuged for 1 hour at 13,000 × g. The washed embryos and the pellets of the 10 centrifuged wash baths were frozen at -20 °C before examination for evidence of C. abortus using polymerase chain reaction. C. abortus DNA was found in all of the infected batches of ZP-intact embryos (9/9) after 10 successive washes. It was also detected in the 10th wash fluid for seven batches of embryos, whereas for the two other batches, the last positive wash bath was the eighth and the ninth, respectively. In contrast, none of the embryos or their washing fluids in the control batches were DNA positive. These results report that C. abortus adheres to and/or penetrates the ZP of in vivo caprine embryos after in vitro infection, and that the standard washing protocol recommended by the IETS for bovine embryos, failed to remove it. The persistence of these bacteria after washing makes the embryo a potential means of transmission of the bacterium during embryo transfer from

  2. Chlamydia abortus in Cows Oviducts, Occasional Event or Causal Connection?

    PubMed

    Appino, S; Vincenti, L; Rota, A; Pellegrini, S; Chieppa, M N; Cadoni, V; Pregel, P

    2015-06-01

    Fifty-seven genital tracts of regularly slaughtered culled Piedmontese cows, aged 7.4 ± 4.3 years (mean ± SD), range: 2.6-15.6 years, were grossly and microscopically examined. DNA extracted from oviducts was subjected to PCR to evaluate the presence of Chlamydia spp. The 15 PCR-positive oviducts were subjected to Sanger sequencing and showed the presence of Chamydia abortus, with an identity range between 99 and 100%. Nine of the PCR-positive samples belonged to the 24 animals with a normal macroscopic appearance of the whole genital tract (percentage of positive oviducts in normal genital tracts 9/24 = 37.5%), while six belonged to the 33 genital tracts with lesions in one or more organs (percentage of positive oviducts in pathological genital tracts 6/33 = 18.1%); of these, a single animal had salpingitis. The detection of C. abortus in bovine oviducts is of particular interest because it has never been previously investigated or reported.

  3. Identification and characterization of Chlamydia abortus isolates from yaks in Qinghai, China.

    PubMed

    Li, Zhaocai; Cao, Xiaoan; Fu, Baoquan; Chao, Yilin; Cai, Jinshan; Zhou, Jizhang

    2015-01-01

    Recently, the yak population has exhibited reproductive disorders, which are considered to be associated with Chlamydia abortus (C. abortus) in Qinghai, China. In this study, a total of 9 aborted fetuses (each from a different herd) and 126 vaginal swab samples from the 9 herds were collected and analyzed. C. abortus DNA was detected from all of the 9 aborted fetuses and 30 of the 126 vaginal swab samples (23.81%) from yak cows in the selected herds. Four C. abortus strains were isolated from embryonated egg yolk sacs inoculated with foetal organ suspensions. The isolated C. abortus strains were further identified, which showed identical restriction profiles with the C. abortus reference strain using AluI restriction enzyme in the RFLP test. Moreover, the isolated C. abortus strains and C. abortus-positive vaginal swab samples were genotyped by multiple loci variable number tandem repeat analysis and all belonged to the genotype 2 group. These findings suggested that C. abortus played a substantial role in yak abortion in Qinghai, China.

  4. High-resolution melt PCR analysis for rapid identification of Chlamydia abortus live vaccine strain 1B among C. abortus strains and field isolates.

    PubMed

    Vorimore, Fabien; Cavanna, Noémie; Vicari, Nadia; Magnino, Simone; Willems, Hermann; Rodolakis, Annie; Siarkou, Victoria I; Laroucau, Karine

    2012-09-01

    We describe a novel high-resolution melt assay that clearly differentiates Chlamydia abortus live vaccine strain 1B from field C. abortus strains and field wild-type isolates based on previously described single nucleotide polymorphisms. This modern genotyping technique is inexpensive, easy to use, and less time-consuming than PCR-RFLP.

  5. Processing of Chlamydia abortus Polymorphic Membrane Protein 18D during the Chlamydial Developmental Cycle

    PubMed Central

    Wheelhouse, Nick M.; Sait, Michelle; Aitchison, Kevin; Livingstone, Morag; Wright, Frank; McLean, Kevin; Inglis, Neil F.; Smith, David G. E.; Longbottom, David

    2012-01-01

    Background Chlamydia possess a unique family of autotransporter proteins known as the Polymorphic membrane proteins (Pmps). While the total number of pmp genes varies between Chlamydia species, all encode a single pmpD gene. In both Chlamydia trachomatis (C. trachomatis) and C. pneumoniae, the PmpD protein is proteolytically cleaved on the cell surface. The current study was carried out to determine the cleavage patterns of the PmpD protein in the animal pathogen C. abortus (termed Pmp18D). Methodology/Principal Findings Using antibodies directed against different regions of Pmp18D, proteomic techniques revealed that the mature protein was cleaved on the cell surface, resulting in a100 kDa N-terminal product and a 60 kDa carboxy-terminal protein. The N-terminal protein was further processed into 84, 76 and 73 kDa products. Clustering analysis resolved PmpD proteins into three distinct clades with C. abortus Pmp18D, being most similar to those originating from C. psittaci, C. felis and C. caviae. Conclusions/Significance This study indicates that C. abortus Pmp18D is proteolytically processed at the cell surface similar to the proteins of C. trachomatis and C. pneumoniae. However, patterns of cleavage are species-specific, with low sequence conservation of PmpD across the genus. The absence of conserved domains indicates that the function of the PmpD molecule in chlamydia remains to be elucidated. PMID:23145118

  6. Host-pathogen interactions in specific pathogen-free chickens following aerogenous infection with Chlamydia psittaci and Chlamydia abortus.

    PubMed

    Kalmar, Isabelle; Berndt, Angela; Yin, Lizi; Chiers, Koen; Sachse, Konrad; Vanrompay, Daisy

    2015-03-15

    Although Chlamydia (C.) psittaci infections are recognized as an important factor causing economic losses and impairing animal welfare in poultry production, the specific mechanisms leading to severe clinical outcomes are poorly understood. In the present study, we comparatively investigated pathology and host immune response, as well as systemic dissemination and expression of essential chlamydial genes in the course of experimental aerogeneous infection with C. psittaci and the closely related C. abortus, respectively, in specific pathogen-free chicks. Clinical signs appeared sooner and were more severe in the C. psittaci-infected group. Compared to C. abortus infection, more intense systemic dissemination of C. psittaci correlated with higher and faster infiltration of immune cells, as well as more macroscopic lesions and epithelial pathology, such as hyperplasia and erosion. In thoracic air sac tissue, mRNA expression of immunologically relevant factors, such as IFN-γ, IL-1β, IL-6, IL-17, IL-22, LITAF and iNOS was significantly stronger up-regulated in C. psittaci- than in C. abortus-infected birds between 3 and 14 days post-infection. Likewise, transcription rates of the chlamydial genes groEL, cpaf and ftsW were consistently higher in C. psittaci during the acute phase. These findings illustrate that the stronger replication of C. psittaci in its natural host also evoked a more intense immune response than in the case of C. abortus infection.

  7. Expression patterns of five polymorphic membrane proteins during the Chlamydia abortus developmental cycle

    PubMed Central

    Wheelhouse, Nick; Sait, Michelle; Wilson, Kim; Aitchison, Kevin; McLean, Kevin; Smith, David G.E.; Longbottom, David

    2012-01-01

    It has been suggested that polymorphic membrane proteins (Pmps) belonging to the Type V autotransporter protein family play an important role in the pathogenesis of Chlamydia abortus (C. abortus; formerly Chlamydophila abortus) infection. In a previous study we demonstrated the expression of all the pmps at the transcriptional level. The purpose of this study was to measure the number of Pmp positive inclusions throughout the C. abortus developmental cycle to investigate heterogeneity in expression patterns. McCoy cells were infected with C. abortus and analysed for Pmp expression over a 72 h period by fluorescent immunocytochemistry. Pmp18D could be detected at all analysed time points, and could only be accurately quantified from 36 hpi while Pmp10G positive inclusions could be visualised from 36 hpi. Expression of Pmps 13G, 16G and 17G could only be visualised later in the cycle and within less than half of visualised inclusions. These results indicate that while expression of specific Pmps is constitutive (Pmp18D), the pattern of expression of other Pmps is more variable. This suggests that different members of the Pmp family may play different roles within the developmental cycle of the organism, with some (Pmps10G and 18D) having roles throughout the cycle, while the heterogeneity of expression of others may aid in antigenic variation. PMID:22776512

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

    PubMed Central

    Vicari, Nadia; Magnino, Simone; Rodolakis, Annie; Pannekoek, Yvonne; Sachse, Konrad; Longbottom, David; Laroucau, Karine

    2015-01-01

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

  9. Comparative evaluation of the protective efficacy of two formulations of a recombinant Chlamydia abortus subunit candidate vaccine in a mouse model.

    PubMed

    Pan, Qing; Pais, Roshan; Ohandjo, Adaugo; He, Cheng; He, Qing; Omosun, Yusuf; Igietseme, J U; Eko, F O

    2015-04-01

    Chlamydia abortus (C. abortus) is the causative agent of ovine enzootic abortion (OEA) and poses a zoonotic risk to pregnant women. Current live attenuated 1B vaccines are efficacious but cause disease in vaccinated animals and inactivated vaccines are only marginally protective. We tested the ability of a new C. abortus subunit vaccine candidate based on the conserved and immunogenic polymorphic membrane protein D (Pmp18D) formulated in CpG1826+FL (Fms-like tyrosine kinase 3 Ligand; Flt3L) or Vibrio cholerae ghosts (VCG) to induce innate and cross protective immunity against genital C. abortus infection. We found that delivery of rPmp18D with VCG was more effective than with CpG+FL in up-regulating the expression of molecules critically involved in T cell activation and differentiation, including MHC II, CD40, CD80, and CD86, activation of TLRs and NLRP3 inflammasome engagement, and secretion of IL-1β and TNF-α but not IL-10 and IL-4. rVCG-Pmp18D-immunized mice elicited more robust antigen-specific IFN-γ, IgA and IgG2c antibody responses compared to CpG+FL-delivered rPmp18D. Based on the number of mice with positive vaginal cultures, length of vaginal shedding, and number of inclusion forming units recovered following challenge with the heterologous C. abortus strain B577, vaccine delivery with VCG induced superior protective immunity than delivery with a combination of CpG1826 and FL, a nasal DC-targeting adjuvant. These results demonstrate that the ability of VCG to enhance protective immunity against genital C. abortus infection is superior to that of CpG+FL adjuvants.

  10. Intratracheal infection as an efficient route for testing vaccines against Chlamydia abortus in sheep.

    PubMed

    Álvarez, D; Salinas, J; Buendía, A J; Ortega, N; del Río, L; Sánchez, J; Navarro, J A; Gallego, M C; Murcia-Belmonte, A; Cuello, F; Caro, M R

    2015-09-01

    Pregnant ewes have been widely used to test vaccines against Chlamydia abortus. However, this model entails many disadvantages such as high economic costs and long periods of pregnancy. The murine model is very useful for specific studies but cannot replace the natural host for the later stages of vaccine evaluation. Therefore, a non-pregnant model of the natural host might be useful for a vaccine trial to select the best vaccine candidates prior to use of the pregnant model. With this aim, two routes of infection were assessed in young non-pregnant sheep, namely, intranasal (IN) and intratracheal (IT). In addition, groups of non-vaccinated sheep and sheep immunised with an inactivated vaccine were established to investigate the suitability of the model for testing vaccines. After the experimental infection, isolation of the microorganism in several organs, with pathological and immunohistochemical analyses, antibody production assessment and investigation by PCR of the presence of chlamydia in the vagina or rectum were carried out. Experimental IT inoculation of C. abortus induced pneumonia in sheep during the first few days post-infection, confirming the suitability of the IT route for testing vaccines in the natural host. The course of infection and the resulting pathological signs were less severe in vaccinated sheep compared with non-vaccinated animals, demonstrating the success of vaccination. IN infection did not produce evident lesions or demonstrate the presence of chlamydial antigen in the lungs and cannot be considered an appropriate model for testing vaccines.

  11. Improving the molecular diagnosis of Chlamydia psittaci and Chlamydia abortus infection with a species-specific duplex real-time PCR.

    PubMed

    Opota, Onya; Jaton, Katia; Branley, James; Vanrompay, Daisy; Erard, Veronique; Borel, Nicole; Longbottom, David; Greub, Gilbert

    2015-10-01

    Chlamydia psittaci and Chlamydia abortus are closely related intracellular bacteria exhibiting different tissue tropism that may cause severe but distinct infection in humans. C. psittaci causes psittacosis, a respiratory zoonotic infection transmitted by birds. C. abortus is an abortigenic agent in small ruminants, which can also colonize the human placenta and lead to foetal death and miscarriage. Infections caused by C. psittaci and C. abortus are underestimated mainly due to diagnosis difficulties resulting from their strict intracellular growth. We developed a duplex real-time PCR to detect and distinguish these two bacteria in clinical samples. The first PCR (PCR1) targeted a sequence of the 16S-23S rRNA operon allowing the detection of both C. psittaci and C. abortus. The second PCR (PCR2) targeted the coding DNA sequence CPSIT_0607 unique to C. psittaci. The two PCRs showed 100 % detection for ≥ 10 DNA copies per reaction (1000 copies ml(- 1)). Using a set of 120 samples, including bacterial reference strains, clinical specimens and infected cell culture material, we monitored 100 % sensitivity and 100 % specificity for the detection of C. psittaci and C. abortus for PCR1. When PCR1 was positive, PCR2 could discriminate C. psittaci from C. abortus with a positive predictive value of 100 % and a negative predictive value of 88 %. In conclusion, this new duplex PCR represents a low-cost and time-saving method with high-throughput potential, expected to improve the routine diagnosis of psittacosis and pregnancy complication in large-scale screening programs and also during outbreaks.

  12. Chlamydiaceae Genomics Reveals Interspecies Admixture and the Recent Evolution of Chlamydia abortus Infecting Lower Mammalian Species and Humans.

    PubMed

    Joseph, Sandeep J; Marti, Hanna; Didelot, Xavier; Castillo-Ramirez, Santiago; Read, Timothy D; Dean, Deborah

    2015-11-01

    Chlamydiaceae are obligate intracellular bacteria that cause a diversity of severe infections among humans and livestock on a global scale. Identification of new species since 1989 and emergence of zoonotic infections, including abortion in women, underscore the need for genome sequencing of multiple strains of each species to advance our knowledge of evolutionary dynamics across Chlamydiaceae. Here, we genome sequenced isolates from avian, lower mammalian and human hosts. Based on core gene phylogeny, five isolates previously classified as Chlamydia abortus were identified as members of Chlamydia psittaci and Chlamydia pecorum. Chlamydia abortus is the most recently emerged species and is a highly monomorphic group that lacks the conserved virulence-associated plasmid. Low-level recombination and evidence for adaptation to the placenta echo evolutionary processes seen in recently emerged, highly virulent niche-restricted pathogens, such as Bacillus anthracis. In contrast, gene flow occurred within C. psittaci and other Chlamydiaceae species. The C. psittaci strain RTH, isolated from a red-tailed hawk (Buteo jamaicensis), is an outlying strain with admixture of C. abortus, C. psittaci, and its own population markers. An average nucleotide identity of less than 94% compared with other Chlamydiaceae species suggests that RTH belongs to a new species intermediary between C. psittaci and C. abortus. Hawks, as scavengers and predators, have extensive opportunities to acquire multiple species in their intestinal tract. This could facilitate transformation and homologous recombination with the potential for new species emergence. Our findings indicate that incubator hosts such as birds-of-prey likely promote Chlamydiaceae evolution resulting in novel pathogenic lineages. PMID:26507799

  13. Chlamydiaceae Genomics Reveals Interspecies Admixture and the Recent Evolution of Chlamydia abortus Infecting Lower Mammalian Species and Humans

    PubMed Central

    Joseph, Sandeep J.; Marti, Hanna; Didelot, Xavier; Castillo-Ramirez, Santiago; Read, Timothy D.; Dean, Deborah

    2015-01-01

    Chlamydiaceae are obligate intracellular bacteria that cause a diversity of severe infections among humans and livestock on a global scale. Identification of new species since 1989 and emergence of zoonotic infections, including abortion in women, underscore the need for genome sequencing of multiple strains of each species to advance our knowledge of evolutionary dynamics across Chlamydiaceae. Here, we genome sequenced isolates from avian, lower mammalian and human hosts. Based on core gene phylogeny, five isolates previously classified as Chlamydia abortus were identified as members of Chlamydia psittaci and Chlamydia pecorum. Chlamydia abortus is the most recently emerged species and is a highly monomorphic group that lacks the conserved virulence-associated plasmid. Low-level recombination and evidence for adaptation to the placenta echo evolutionary processes seen in recently emerged, highly virulent niche-restricted pathogens, such as Bacillus anthracis. In contrast, gene flow occurred within C. psittaci and other Chlamydiaceae species. The C. psittaci strain RTH, isolated from a red-tailed hawk (Buteo jamaicensis), is an outlying strain with admixture of C. abortus, C. psittaci, and its own population markers. An average nucleotide identity of less than 94% compared with other Chlamydiaceae species suggests that RTH belongs to a new species intermediary between C. psittaci and C. abortus. Hawks, as scavengers and predators, have extensive opportunities to acquire multiple species in their intestinal tract. This could facilitate transformation and homologous recombination with the potential for new species emergence. Our findings indicate that incubator hosts such as birds-of-prey likely promote Chlamydiaceae evolution resulting in novel pathogenic lineages. PMID:26507799

  14. Chlamydiaceae Genomics Reveals Interspecies Admixture and the Recent Evolution of Chlamydia abortus Infecting Lower Mammalian Species and Humans.

    PubMed

    Joseph, Sandeep J; Marti, Hanna; Didelot, Xavier; Castillo-Ramirez, Santiago; Read, Timothy D; Dean, Deborah

    2015-10-27

    Chlamydiaceae are obligate intracellular bacteria that cause a diversity of severe infections among humans and livestock on a global scale. Identification of new species since 1989 and emergence of zoonotic infections, including abortion in women, underscore the need for genome sequencing of multiple strains of each species to advance our knowledge of evolutionary dynamics across Chlamydiaceae. Here, we genome sequenced isolates from avian, lower mammalian and human hosts. Based on core gene phylogeny, five isolates previously classified as Chlamydia abortus were identified as members of Chlamydia psittaci and Chlamydia pecorum. Chlamydia abortus is the most recently emerged species and is a highly monomorphic group that lacks the conserved virulence-associated plasmid. Low-level recombination and evidence for adaptation to the placenta echo evolutionary processes seen in recently emerged, highly virulent niche-restricted pathogens, such as Bacillus anthracis. In contrast, gene flow occurred within C. psittaci and other Chlamydiaceae species. The C. psittaci strain RTH, isolated from a red-tailed hawk (Buteo jamaicensis), is an outlying strain with admixture of C. abortus, C. psittaci, and its own population markers. An average nucleotide identity of less than 94% compared with other Chlamydiaceae species suggests that RTH belongs to a new species intermediary between C. psittaci and C. abortus. Hawks, as scavengers and predators, have extensive opportunities to acquire multiple species in their intestinal tract. This could facilitate transformation and homologous recombination with the potential for new species emergence. Our findings indicate that incubator hosts such as birds-of-prey likely promote Chlamydiaceae evolution resulting in novel pathogenic lineages.

  15. Soluble antigens of virulent and attenuated biotypes of Brucella abortus.

    PubMed Central

    Hatten, B A; Brodeur, R D

    1978-01-01

    Several methods were used to characterize three Brucella abortus biotypes (1, 5, and 7), including the attenuated vaccine strain S-19. Chemical analysis did not reveal remarkable differences among these strains, and only minor differences were noted in elution patterns of soluble extracts subjected to column chromatography. Qualitative and quantitative differences in extract components were demonstrated, however, by polyacrylamide gel isoelectric focusing. A distinctive difference was the presence of components in extracts from one or more of the virulent biotypes that were absent in similar preparations from the attenuated strain. In addition, one component common to all virulent strains was absent in strain S-19. Results of immunodiffusion experiments employing adsorbed and unadsorbed antisera also suggested that the quantity, quality, and surface distribution of various cellular antigens differed among the biotypes studied. Images PMID:103842

  16. Prevalence and molecular identification of Chlamydia abortus in commercial dairy goat farms in a hot region in Mexico.

    PubMed

    Campos-Hernández, Eleuterio; Vázquez-Chagoyán, Juan Carlos; Salem, Abdelfattah Z M; Saltijeral-Oaxaca, Jorge Antonio; Escalante-Ochoa, Cristina; López-Heydeck, Sandra M; de Oca-Jiménez, Roberto Montes

    2014-08-01

    The aim of this study was to determine the seroprevalence and presence of Chlamydia abortus in Saanen breed female goats from commercial dairy goat farms under intensive production in the municipality of Guanajuato, Mexico. Sera were collected to determine the prevalence of anti-C. abortus IgG antibodies using recombinant enzyme-linked immunosorbent assay (rELISA) and cell culture. Polymerase chain reaction (PCR) was used to prove the presence of the pathogen in swab samples collected from the vagina and rectum of selected animals. Additionally, foetal tissue samples from a sudden abortion were collected. C. abortus prevalence in female goats of commercial milking farms sampled in Guanajuato, Mexico, was 4.87% (n = 246). Seropositive animals were found in six out of nine (66.6%) dairy goat farms sampled, and prevalence among animals in individual farms ranged between 3.44 and 13.51%. C. abortus was detected using PCR in spleen tissue from the aborted foetus. PCR-based detection, as well as isolation from vaginal and rectal swabs, was not possible in the present study. Isolation through cell culture was also unsuccessful from aborted foetal tissue samples. In conclusion, the results from rELISA and PCR show that C. abortus is present in dairy goat farms in the state of Guanajuato, Mexico.

  17. Effect of Preventive Chlamydia abortus Vaccination in Offspring Development in Sheep Challenged Experimentally

    PubMed Central

    García-Seco, Teresa; Pérez-Sancho, Marta; Salinas, Jesús; Navarro, Alejandro; Díez-Guerrier, Alberto; García, Nerea; Pozo, Pilar; Goyache, Joaquín; Domínguez, Lucas; Álvarez, Julio

    2016-01-01

    Ovine enzootic abortion, caused by Chlamydia abortus, leads to important economic losses worldwide. In addition to reproductive failures, infection may impact lamb growth during the first weeks after birth, yet this effect has not been well characterized. Vaccination can help to control the disease but variable efficacy values have been described, possibly related with factors associated with the host, the vaccine, the parameter used for efficacy determination, and the challenge conditions. In this context, we evaluated the efficacy of an inactivated standard commercial vaccine and a 1/2 diluted dose in pregnant sheep challenged with C. abortus by examining multiple indicators of vaccine effect (including incidence of reproductive failures, bacterial excretion, and evolution of weight gain of viable lambs during the first month of life). Three groups of ewes [control non-vaccinated, C (n = 18); vaccinated with standard dose, SV (n = 16); and vaccinated with 1/2 dose, DV (n = 17)], were challenged approximately 90 days post-mating and tested using direct PCR (tissue samples and vaginal swabs) and ELISA (serum) until 31 days post-reproductive outcome. There were not significant differences in the proportions of reproductive failures or bacterial shedding after birth/abortion regardless the vaccination protocol. However, a beneficial effect of vaccination on offspring growth was detected in both vaccinated groups compared with the controls, with a mean increase in weight measured at 30 days of life of 1.5 and 2.5 kg (p = 0.056) and an increase in the geometric mean of the daily gain of 8.4 and 9.7% in lambs born from DV and SV ewes compared with controls, respectively. Our results demonstrate the effect of an inactivated vaccine in the development of the offspring of C. abortus-infected ewes at a standard and a diluted dose, an interesting finding given the difficulty in achieving sufficient antigen concentration in the production of enzootic

  18. Effect of Preventive Chlamydia abortus Vaccination in Offspring Development in Sheep Challenged Experimentally.

    PubMed

    García-Seco, Teresa; Pérez-Sancho, Marta; Salinas, Jesús; Navarro, Alejandro; Díez-Guerrier, Alberto; García, Nerea; Pozo, Pilar; Goyache, Joaquín; Domínguez, Lucas; Álvarez, Julio

    2016-01-01

    Ovine enzootic abortion, caused by Chlamydia abortus, leads to important economic losses worldwide. In addition to reproductive failures, infection may impact lamb growth during the first weeks after birth, yet this effect has not been well characterized. Vaccination can help to control the disease but variable efficacy values have been described, possibly related with factors associated with the host, the vaccine, the parameter used for efficacy determination, and the challenge conditions. In this context, we evaluated the efficacy of an inactivated standard commercial vaccine and a 1/2 diluted dose in pregnant sheep challenged with C. abortus by examining multiple indicators of vaccine effect (including incidence of reproductive failures, bacterial excretion, and evolution of weight gain of viable lambs during the first month of life). Three groups of ewes [control non-vaccinated, C (n = 18); vaccinated with standard dose, SV (n = 16); and vaccinated with 1/2 dose, DV (n = 17)], were challenged approximately 90 days post-mating and tested using direct PCR (tissue samples and vaginal swabs) and ELISA (serum) until 31 days post-reproductive outcome. There were not significant differences in the proportions of reproductive failures or bacterial shedding after birth/abortion regardless the vaccination protocol. However, a beneficial effect of vaccination on offspring growth was detected in both vaccinated groups compared with the controls, with a mean increase in weight measured at 30 days of life of 1.5 and 2.5 kg (p = 0.056) and an increase in the geometric mean of the daily gain of 8.4 and 9.7% in lambs born from DV and SV ewes compared with controls, respectively. Our results demonstrate the effect of an inactivated vaccine in the development of the offspring of C. abortus-infected ewes at a standard and a diluted dose, an interesting finding given the difficulty in achieving sufficient antigen concentration in the production of enzootic

  19. Effect of Preventive Chlamydia abortus Vaccination in Offspring Development in Sheep Challenged Experimentally

    PubMed Central

    García-Seco, Teresa; Pérez-Sancho, Marta; Salinas, Jesús; Navarro, Alejandro; Díez-Guerrier, Alberto; García, Nerea; Pozo, Pilar; Goyache, Joaquín; Domínguez, Lucas; Álvarez, Julio

    2016-01-01

    Ovine enzootic abortion, caused by Chlamydia abortus, leads to important economic losses worldwide. In addition to reproductive failures, infection may impact lamb growth during the first weeks after birth, yet this effect has not been well characterized. Vaccination can help to control the disease but variable efficacy values have been described, possibly related with factors associated with the host, the vaccine, the parameter used for efficacy determination, and the challenge conditions. In this context, we evaluated the efficacy of an inactivated standard commercial vaccine and a 1/2 diluted dose in pregnant sheep challenged with C. abortus by examining multiple indicators of vaccine effect (including incidence of reproductive failures, bacterial excretion, and evolution of weight gain of viable lambs during the first month of life). Three groups of ewes [control non-vaccinated, C (n = 18); vaccinated with standard dose, SV (n = 16); and vaccinated with 1/2 dose, DV (n = 17)], were challenged approximately 90 days post-mating and tested using direct PCR (tissue samples and vaginal swabs) and ELISA (serum) until 31 days post-reproductive outcome. There were not significant differences in the proportions of reproductive failures or bacterial shedding after birth/abortion regardless the vaccination protocol. However, a beneficial effect of vaccination on offspring growth was detected in both vaccinated groups compared with the controls, with a mean increase in weight measured at 30 days of life of 1.5 and 2.5 kg (p = 0.056) and an increase in the geometric mean of the daily gain of 8.4 and 9.7% in lambs born from DV and SV ewes compared with controls, respectively. Our results demonstrate the effect of an inactivated vaccine in the development of the offspring of C. abortus-infected ewes at a standard and a diluted dose, an interesting finding given the difficulty in achieving sufficient antigen concentration in the production of enzootic

  20. Electron tomography and cryo-SEM characterization reveals novel ultrastructural features of host-parasite interaction during Chlamydia abortus infection.

    PubMed

    Wilkat, M; Herdoiza, E; Forsbach-Birk, V; Walther, P; Essig, A

    2014-08-01

    Chlamydia (C.) abortus is a widely spread pathogen among ruminants that can be transmitted to women during pregnancy leading to severe systemic infection with consecutive abortion. As a member of the Chlamydiaceae, C. abortus shares the characteristic feature of an obligate intracellular biphasic developmental cycle with two morphological forms including elementary bodies (EBs) and reticulate bodies (RBs). In contrast to other chlamydial species, C. abortus ultrastructure has not been investigated yet. To do so, samples were fixed by high-pressure freezing and processed by different electron microscopic methods. Freeze-substituted samples were analysed by transmission electron microscopy, scanning transmission electron microscopical tomography and immuno-electron microscopy, and freeze-fractured samples were analysed by cryo-scanning electron microscopy. Here, we present three ultrastructural features of C. abortus that have not been reported up to now. Firstly, the morphological evidence that C. abortus is equipped with the type three secretion system. Secondly, the accumulation and even coating of whole inclusion bodies by membrane complexes consisting of multiple closely adjacent membranes which seems to be a C. abortus specific feature. Thirdly, the formation of small vesicles in the periplasmic space of RBs in the second half of the developmental cycle. Concerning the time point of their formation and the fact that they harbour chlamydial components, these vesicles might be morphological correlates of an intermediate step during the process of redifferentiation of RBs into EBs. As this feature has also been shown for C. trachomatis and C. pneumoniae, it might be a common characteristic of the family of Chlamydiaceae.

  1. Seroprevalence and molecular characterization of Chlamydia abortus in frozen fetal and placental tissues of aborting ewes in northeastern Algeria.

    PubMed

    Hireche, Sana; Ababneh, Mustafa Mohammed Kheir; Bouaziz, Omar; Boussena, Sabrina

    2016-02-01

    Enzootic abortion of ewes is one of the most serious health problems in sheep flocks worldwide. It has a significant economic impact because abortion, decrease in milk production and weak lambs. Besides, the bacteria is zoonotic. A cross-sectional study was conducted to determine the seroprevalence and risk factors associated with Chlamydia abortus infection in 552 ewes in Constantine using a C. abortus-specific indirect ELISA kit. Chlamydial DNA was investigated in ten ovine fetuses and eight placentas using PCR- restriction fragment length polymorphism (RFLP) and DNA sequencing. The study concluded that 7.2 % of ewes were seropositive and 33.3 % of sheep flocks had at least one seropositive ewe. Adjacent farmworker visits (OR = 7.667, 95 % CI (OR) = 2.307; 27.203) was defined as a risk factor. Deliveries of weak lambs (OR = 2.920, 95 % CI (OR) = 1.022; 8.342) and septicemia in lambs (OR = 9.971, 95 % CI (OR) = 2.383; 41.713) were significantly associated with chlamydial infection. PCR-RFLP analysis revealed positive signals to C. abortus in six fetuses and four placentas. Sequencing of the omp2 gene revealed that the Algerian strain is 96 % similar with C. abortus FAS strain. C. abortus plays a major role in abortion in northeastern Algeria. Appropriate control measures must be implemented to reduce economic losses and to avoid human contamination.

  2. Profiling Antibody Responses to Infections by Chlamydia abortus Enables Identification of Potential Virulence Factors and Candidates for Serodiagnosis

    PubMed Central

    Forsbach-Birk, Vera; Foddis, Corinna; Simnacher, Ulrike; Wilkat, Max; Longbottom, David; Walder, Gernot; Benesch, Christiane; Ganter, Martin; Sachse, Konrad; Essig, Andreas

    2013-01-01

    Enzootic abortion of ewes (EAE) due to infection with the obligate intracellular pathogen Chlamydia (C.) abortus is an important zoonosis leading to considerable economic loss to agriculture worldwide. The pathogen can be transmitted to humans and may lead to serious infection in pregnant women. Knowledge about epidemiology, clinical course and transmission to humans is hampered by the lack of reliable diagnostic tools. Immunoreactive proteins, which are expressed in infected animals and humans, may serve as novel candidates for diagnostic marker proteins and represent putative virulence factors. In order to broaden the spectrum of immunogenic C. abortus proteins we applied 2D immunoblot analysis and screening of an expression library using human and animal sera. We have identified 48 immunoreactive proteins representing potential diagnostic markers and also putative virulence factors, such as CAB080 (homologue of the “macrophage infectivity potentiator”, MIP), CAB167 (homologue of the “translocated actin recruitment protein”, TARP), CAB712 (homologue of the “chlamydial protease-like activity factor”, CPAF), CAB776 (homologue of the “Polymorphic membrane protein D”, PmpD), and the “hypothetical proteins” CAB063, CAB408 and CAB821, which are predicted to be type III secreted. We selected two putative virulence factors for further characterization, i.e. CAB080 (cMIP) and CAB063, and studied their expression profiles at transcript and protein levels. Analysis of the subcellular localization of both proteins throughout the developmental cycle revealed CAB063 being the first C. abortus protein shown to be translocated to the host cell nucleus. PMID:24260366

  3. Transposon-Derived Brucella abortus Rough Mutants Are Attenuated and Exhibit Reduced Intracellular Survival

    PubMed Central

    Allen, Chris A.; Adams, L. Garry; Ficht, Thomas A.

    1998-01-01

    The O antigen of Brucella abortus has been described as a major virulence determinant based on the attenuated survival of fortuitously isolated rough variants. However, the lack of genetic definition of these mutants and the virulence of naturally occurring rough species, Brucella ovis and Brucella canis, has confused interpretation. To better characterize the role of O antigen in virulence and survival, transposon mutagenesis was used to generate B. abortus rough mutants defective in O-antigen presentation. Sequence analysis of DNA flanking the site of Tn5 insertion was used to verify insertion in genes encoding lipopolysaccharide (LPS) biosynthetic functions. Not surprisingly, each of the rough mutants was attenuated for survival in mice, but unexpected differences among the mutants were observed. In an effort to define the basis for the observed differences, the structure of the rough LPS and the sensitivity of these mutants to individual killing mechanisms were examined in vitro. All of the B. abortus rough mutants exhibited a 4- to 5-log-unit increase, compared to the smooth parental strain, in sensitivity to complement-mediated lysis. Little change was evident in the sensitivity of these organisms to hydrogen peroxide, consistent with an inability of O antigen to exclude relatively small molecules. Sensitivity to polymyxin B, which was employed as a model cationic, amphipathic peptide similar to defensins found in phagocytic cells, revealed survival differences among the rough mutants similar to those observed in the mouse. One mutant in particular exhibited hypersensitivity to polymyxin B and reduced survival in mice. This mutant was characterized by a truncated rough LPS. DNA sequence analysis of this mutant revealed a transposon interruption in the gene encoding phosphomannomutase (pmm), suggesting that this activity may be required for the synthesis of a full-length core polysaccharide in addition to O antigen. B. abortus O antigen appears to be essential

  4. Polymorphonuclear Neutrophils Are Necessary for the Recruitment of CD8+ T Cells in the Liver in a Pregnant Mouse Model of Chlamydophila abortus (Chlamydia psittaci Serotype 1) Infection

    PubMed Central

    de Oca, Roberto Montes; Buendía, Antonio J.; Del Río, Laura; Sánchez, Joaquín; Salinas, Jesús; Navarro, Jose A.

    2000-01-01

    The role of polymorphonuclear neutrophils (PMNs) in the development of the specific immune response against Chlamydophila abortus (Chlamydia psittaci serotype 1) infection was studied in a pregnant mouse model involving treatment with RB6-8C5 monoclonal antibody. PMN depletion significantly affected the immune response in the liver, in which the T-lymphocyte and F4/80+ cell populations decreased, particularly the CD8+ T-cell population. A Th1-like response, characterized by high levels of gamma interferon without detectable levels of interleukin 4 (IL-4) in serum, was observed in both depleted and nondepleted mice, although an increased production of IL-10 was detected in the depleted group. Our results suggest that PMNs play a very important role in the recruitment of other leukocyte populations to the inflammatory foci but have little influence in the polarization of the immune specific response toward a Th1-like response. PMID:10679002

  5. Polymorphonuclear neutrophils are necessary for the recruitment of CD8(+) T cells in the liver in a pregnant mouse model of Chlamydophila abortus (Chlamydia psittaci serotype 1) infection.

    PubMed

    de Oca, R M; Buendía, A J; Del Río, L; Sánchez, J; Salinas, J; Navarro, J A

    2000-03-01

    The role of polymorphonuclear neutrophils (PMNs) in the development of the specific immune response against Chlamydophila abortus (Chlamydia psittaci serotype 1) infection was studied in a pregnant mouse model involving treatment with RB6-8C5 monoclonal antibody. PMN depletion significantly affected the immune response in the liver, in which the T-lymphocyte and F4/80(+) cell populations decreased, particularly the CD8(+) T-cell population. A Th1-like response, characterized by high levels of gamma interferon without detectable levels of interleukin 4 (IL-4) in serum, was observed in both depleted and nondepleted mice, although an increased production of IL-10 was detected in the depleted group. Our results suggest that PMNs play a very important role in the recruitment of other leukocyte populations to the inflammatory foci but have little influence in the polarization of the immune specific response toward a Th1-like response.

  6. Chlamydia

    MedlinePlus

    ... you have symptoms of a chlamydia infection, your health care provider will collect a culture or perform a test called a PCR: The culture will be collected during a pelvic exam in women, or from the ... back. Your health care provider may also check you for other ...

  7. Endogenous Interleukin-12 Is Not Required for Resolution of Chlamydophila abortus (Chlamydia psittaci Serotype 1) Infection in Mice

    PubMed Central

    Del Río, Laura; Buendía, Antonio J.; Sánchez, Joaquín; Gallego, María C.; Caro, María R.; Ortega, Nieves; Seva, Juan; Pallarés, Francisco J.; Cuello, Francisco; Salinas, Jesús

    2001-01-01

    A Th1 immune response involving gamma interferon (IFN-γ) production is required to eliminate Chlamydophila abortus infections. In this study, the role of interleukin-12 (IL-12) in protecting against C. abortus infection was investigated using IL-12−/− and wild-type (WT) C57BL/6 mice to determine the role of this Th1-promoting cytokine. IL-12−/− mice were able to eliminate the C. abortus infection in a primary infection. However, there was a delay in the clearance of bacteria when IL-12−/− mice were infected with a sublethal dose of C. abortus, the delay being associated with a lower production of IFN-γ. The low level of IFN-γ was essential for survival of IL-12−/− infected mice. Both WT and IL-12−/− mice developed a Th1 immune response against C. abortus infection, since they both produced IFN-γ and immunoglobulin G2a antibody isotype. In addition, when mice were given a secondary infectious challenge with C. abortus, a protective host response which resolved the secondary infection was developed by both WT and IL-12−/− mice. The lack of IL-12 resulted in few infiltrating CD4+ T cells in the liver relative to the number in WT mice, although the number of CD8+ T cells was slightly higher. The more intense Th1 response presented by WT mice may have a pathogenic effect, as the animals showed higher morbidity after the infection. In conclusion, these results suggest that although IL-12 expedites the clearance of C. abortus infection, this cytokine is not essential for the establishment of a protective host response against the infection. PMID:11447154

  8. Mutation of purD and purF genes further attenuates Brucella abortus strain RB51.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Barate, Abhijit Kashinath; Kim, Suk; Watarai, Masahisa; Hahn, Tae-Wook

    2015-02-01

    In the present study, transposon mutagenesis was used to further attenuate Brucella abortus RB51 vaccine strain. Two purD and purF mutants were constructed, characterized and evaluated for attenuation via intracellular survival in murine macrophage-like RAW264.7 and HeLa cells, and by clearance in BALB/c mice. The purD and purF mutants showed significantly decreased intracellular survival, and complementation of these mutants with intact copies of purD or purF genes of RB51 strain was able to restore these defects. In addition, the pur mutants presented significantly lowered persistence in mice. Immunization with purD and purF mutants protected mice against a challenge with the virulent B. abortus strain 544 at a level similar to that of the parent RB51. These data suggest that genes encoding the early stages of purine biosynthesis (purD and purF) are required for intracellular survival and virulence of B. abortus.

  9. Expression of the major outer membrane protein (MOMP) of Chlamydophila abortus, Chlamydophila pecorum, and Chlamydia suis in Escherichia coli using an arabinose-inducible plasmid vector.

    PubMed

    Hoelzle, L E; Hoelzle, K; Wittenbrink, M M

    2003-10-01

    The ompA genes encoding the 40 kDa major outer membrane protein (MOMP) of Chlamydophila (Ch.) abortus, Ch. pecorum, and Chlamydia (C.) suis were cloned into the arabinose-inducible plasmid vector pBADMycHis, and recombinant MOMPs (rMOMP) from the three chlamydial species were expressed at high levels in Escherichia (E.) coli. The proteins lacking the 22 aa N-terminal signal peptide were expressed as insoluble cytoplasmic inclusion bodies which were readily purified using immobilized metal-affinity chromatography. The rMOMPs including the N-terminal signal peptide were expressed and translocated as a surface-exposed immunoaccessible protein into the outer membrane of E. coli. Transformants expressing this full-length rMOMP were significantly reduced in viability. Purified native elementary bodies (EB) and rMOMPs of the three chlamydial species purified from the E. coli cytoplasm were used for immunization of rabbits. The resulting sera were analysed for their ability to recognize homologous and heterologous rMOMP and native EB. When testing rMOMP antisera against rMOMP and EB antigens, marked cross-reactivities were detected between the three species. Using EB antisera and rMOMPs as antigens, a significant species-specific reactivity was measured.

  10. Brucella abortus mutants lacking ATP-binding cassette transporter proteins are highly attenuated in virulence and confer protective immunity against virulent B. abortus challenge in BALB/c mice.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Park, Soyeon; Park, Bo-Kyoung; Hahn, Tae-Wook

    2016-06-01

    Brucella abortus RB51 is an attenuated vaccine strain that has been most frequently used for bovine brucellosis. Although it is known to provide good protection in cattle, it still has some drawbacks including resistance to rifampicin, residual virulence and pathogenicity in humans. Thus, there has been a continuous interest on new safe and effective bovine vaccine candidates. In the present study, we have constructed unmarked mutants by deleting singly cydD and cydC genes, which encode ATP-binding cassette transporter proteins, from the chromosome of the virulent Brucella abortus isolate from Korean cow (referred to as IVK15). Both IVK15ΔcydD and ΔcydC mutants showed increased sensitivity to metal ions, hydrogen peroxide and acidic pH, which are mimic to intracellular environment during host infection. Additionally, the mutants exhibited a significant growth defect in RAW264.7 cells and greatly attenuated in mice. Vaccination of mice with either IVK15ΔcydC or IVK15ΔcydD mutant could elicit an anti-Brucella specific immunoglobulin G (IgG) and IgG subclass responses as well as enhance the secretion of interferon-gamma, and provided better protection against challenge with B. abortus strain 2308 than with the commercial B. abortus strain RB51 vaccine. Collectively, these results suggest that both IVK15ΔcydC and IVK15ΔcydD mutants could be an attenuated vaccine candidate against B. abortus. PMID:27057678

  11. Brucella abortus mutants lacking ATP-binding cassette transporter proteins are highly attenuated in virulence and confer protective immunity against virulent B. abortus challenge in BALB/c mice.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Park, Soyeon; Park, Bo-Kyoung; Hahn, Tae-Wook

    2016-06-01

    Brucella abortus RB51 is an attenuated vaccine strain that has been most frequently used for bovine brucellosis. Although it is known to provide good protection in cattle, it still has some drawbacks including resistance to rifampicin, residual virulence and pathogenicity in humans. Thus, there has been a continuous interest on new safe and effective bovine vaccine candidates. In the present study, we have constructed unmarked mutants by deleting singly cydD and cydC genes, which encode ATP-binding cassette transporter proteins, from the chromosome of the virulent Brucella abortus isolate from Korean cow (referred to as IVK15). Both IVK15ΔcydD and ΔcydC mutants showed increased sensitivity to metal ions, hydrogen peroxide and acidic pH, which are mimic to intracellular environment during host infection. Additionally, the mutants exhibited a significant growth defect in RAW264.7 cells and greatly attenuated in mice. Vaccination of mice with either IVK15ΔcydC or IVK15ΔcydD mutant could elicit an anti-Brucella specific immunoglobulin G (IgG) and IgG subclass responses as well as enhance the secretion of interferon-gamma, and provided better protection against challenge with B. abortus strain 2308 than with the commercial B. abortus strain RB51 vaccine. Collectively, these results suggest that both IVK15ΔcydC and IVK15ΔcydD mutants could be an attenuated vaccine candidate against B. abortus.

  12. Brucella abortusΔcydCΔcydD and ΔcydCΔpurD double-mutants are highly attenuated and confer long-term protective immunity against virulent Brucella abortus.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Park, Soyeon; Kim, Kiju; Hahn, Tae-Wook

    2016-01-01

    We constructed double deletion (ΔcydCΔcydD and ΔcydCΔpurD) mutants from virulent Brucella abortus biovar 1 field isolate (BA15) by deleting the genes encoding an ATP-binding cassette-type transporter (cydC and cydD genes) and a phosphoribosylamine-glycine ligase (purD). Both BA15ΔcydCΔcydD and BA15ΔcydCΔpurD double-mutants exhibited significant attenuation of virulence when assayed in murine macrophages or in BALB/c mice. Both double-mutants were readily cleared from spleens by 4 weeks post-inoculation even when inoculated at the dose of 10(8) CFU per mouse. Moreover, the inoculated mice showed no splenomegaly, which indicates that the mutants are highly attenuated. Importantly, the attenuation of in vitro and in vivo growth did not impair the ability of these mutants to confer long-term protective immunity in mice against challenge with B. abortus strain 2308. Vaccination of mice with either mutant induced humoral and cell-mediated immune responses, and provided significantly better protection than commercial B. abortus strain RB51 vaccine. These results suggest that highly attenuated BA15ΔcydCΔcydD and BA15ΔcydCΔpurD mutants can be used effectively as potential live vaccine candidates against bovine brucellosis.

  13. Experimental Infection of Richardson's Ground Squirrels (Spermophilus richardsonii) with Attenuated and Virulent Strains of Brucella abortus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Exposure of non-target species to wildlife vaccines is an important concern when evaluating a candidate vaccine for use in the field. A previous investigation of the safety of Brucella abortus strain RB51 (sRB51) in various non-target species suggested that Richardson’s ground squirrels (Spermophil...

  14. Comparative genomic analysis of Brucella abortus vaccine strain 104M reveals a set of candidate genes associated with its virulence attenuation.

    PubMed

    Yu, Dong; Hui, Yiming; Zai, Xiaodong; Xu, Junjie; Liang, Long; Wang, Bingxiang; Yue, Junjie; Li, Shanhu

    2015-01-01

    The Brucella abortus strain 104M, a spontaneously attenuated strain, has been used as a vaccine strain in humans against brucellosis for 6 decades in China. Despite many studies, the molecular mechanisms that cause the attenuation are still unclear. Here, we determined the whole-genome sequence of 104M and conducted a comprehensive comparative analysis against the whole genome sequences of the virulent strain, A13334, and other reference strains. This analysis revealed a highly similar genome structure between 104M and A13334. The further comparative genomic analysis between 104M and A13334 revealed a set of genes missing in 104M. Some of these genes were identified to be directly or indirectly associated with virulence. Similarly, a set of mutations in the virulence-related genes was also identified, which may be related to virulence alteration. This study provides a set of candidate genes associated with virulence attenuation in B.abortus vaccine strain 104M.

  15. Comparative genomic analysis of Brucella abortus vaccine strain 104M reveals a set of candidate genes associated with its virulence attenuation

    PubMed Central

    Yu, Dong; Hui, Yiming; Zai, Xiaodong; Xu, Junjie; Liang, Long; Wang, Bingxiang; Yue, Junjie; Li, Shanhu

    2015-01-01

    The Brucella abortus strain 104M, a spontaneously attenuated strain, has been used as a vaccine strain in humans against brucellosis for 6 decades in China. Despite many studies, the molecular mechanisms that cause the attenuation are still unclear. Here, we determined the whole-genome sequence of 104M and conducted a comprehensive comparative analysis against the whole genome sequences of the virulent strain, A13334, and other reference strains. This analysis revealed a highly similar genome structure between 104M and A13334. The further comparative genomic analysis between 104M and A13334 revealed a set of genes missing in 104M. Some of these genes were identified to be directly or indirectly associated with virulence. Similarly, a set of mutations in the virulence-related genes was also identified, which may be related to virulence alteration. This study provides a set of candidate genes associated with virulence attenuation in B.abortus vaccine strain 104M. PMID:26039674

  16. The Brucella abortus phosphoglycerate kinase mutant is highly attenuated and induces protection superior to that of vaccine strain 19 in immunocompromised and immunocompetent mice.

    PubMed

    Trant, Cyntia G M C; Lacerda, Thais L S; Carvalho, Natalia B; Azevedo, Vasco; Rosinha, Gracia M S; Salcedo, Suzana P; Gorvel, Jean-Pierre; Oliveira, Sergio C

    2010-05-01

    Brucella abortus is a facultative intracellular bacterial pathogen that causes abortion in domestic animals and undulant fever in humans. The mechanism of virulence of Brucella spp. is not yet fully understood. Therefore, it is crucial to identify new molecules that can function as virulence factors to better understand the host-pathogen interplay. Herein, we identified the gene encoding the phosphoglycerate kinase (PGK) of B. abortus strain 2308. To test the role of PGK in Brucella pathogenesis, a pgk deletion mutant was constructed. Replacement of the wild-type pgk by recombination was demonstrated by Southern and Western blot analyses. The B. abortus Delta pgk mutant strain exhibited extreme attenuation in bone marrow-derived macrophages and in vivo in BALB/c, C57BL/6, 129/Sv, and interferon regulatory factor-1 knockout (IRF-1 KO) mice. Additionally, at 24 h postinfection the Delta pgk mutant was not found within the same endoplasmic reticulum-derived compartment as the wild-type bacteria, but, instead, over 60% of Brucella-containing vacuoles (BCVs) retained the late endosomal/lysosomal marker LAMP1. Furthermore, the B. abortus Delta pgk deletion mutant was used as a live vaccine. Challenge experiments revealed that the Delta pgk mutant strain induced protective immunity in 129/Sv or IRF-1 KO mice that was superior to the protection conferred by commercial strain 19 or RB51. Finally, the results shown here demonstrated that Brucella PGK is critical for full bacterial virulence and that a Delta pgk mutant may serve as a potential vaccine candidate in future studies. PMID:20194591

  17. The Attenuated Brucella abortus Strain 19 Invades, Persists in, and Activates Human Dendritic Cells, and Induces the Secretion of IL-12p70 but Not IL-23

    PubMed Central

    Weinhold, Mario; Eisenblätter, Martin; Jasny, Edith; Fehlings, Michael; Finke, Antje; Gayum, Hermine; Rüschendorf, Ursula; Renner Viveros, Pablo; Moos, Verena; Allers, Kristina; Schneider, Thomas; Schaible, Ulrich E.; Schumann, Ralf R.; Mielke, Martin E.; Ignatius, Ralf

    2013-01-01

    Background Bacterial vectors have been proposed as novel vaccine strategies to induce strong cellular immunity. Attenuated strains of Brucella abortus comprise promising vector candidates since they have the potential to induce strong CD4+ and CD8+ T-cell mediated immune responses in the absence of excessive inflammation as observed with other Gram-negative bacteria. However, some Brucella strains interfere with the maturation of dendritic cells (DCs), which is essential for antigen-specific T-cell priming. In the present study, we investigated the interaction of human monocyte-derived DCs with the smooth attenuated B. abortus strain (S) 19, which has previously been employed successfully to vaccinate cattle. Methodology/Principal findings We first looked into the potential of S19 to hamper the cytokine-induced maturation of DCs; however, infected cells expressed CD25, CD40, CD80, and CD86 to a comparable extent as uninfected, cytokine-matured DCs. Furthermore, S19 activated DCs in the absence of exogeneous stimuli, enhanced the expression of HLA-ABC and HLA-DR, and was able to persist intracellularly without causing cytotoxicity. Thus, DCs provide a cellular niche for persisting brucellae in vivo as a permanent source of antigen. S19-infected DCs produced IL-12/23p40, IL-12p70, and IL-10, but not IL-23. While heat-killed bacteria also activated DCs, soluble mediators were not involved in S19-induced activation of human DCs. HEK 293 transfectants revealed cellular activation by S19 primarily through engagement of Toll-like receptor (TLR)2. Conclusions/Significance Thus, as an immunological prerequisite for vaccine efficacy, B. abortus S19 potently infects and potently activates (most likely via TLR2) human DCs to produce Th1-promoting cytokines. PMID:23805193

  18. Booster vaccination with safe, modified, live-attenuated mutants of Brucella abortus strain RB51 vaccine confers protective immunity against virulent strains of B. abortus and Brucella canis in BALB/c mice.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Kim, Kiju; Park, Bo-Kyoung; Hahn, Tae-Wook

    2015-11-01

    Brucella abortus attenuated strain RB51 vaccine (RB51) is widely used in prevention of bovine brucellosis. Although vaccination with this strain has been shown to be effective in conferring protection against bovine brucellosis, RB51 has several drawbacks, including residual virulence for animals and humans. Therefore, a safe and efficacious vaccine is needed to overcome these disadvantages. In this study, we constructed several gene deletion mutants (ΔcydC, ΔcydD and ΔpurD single mutants, and ΔcydCΔcydD and ΔcydCΔpurD double mutants) of RB51 with the aim of increasing the safety of the possible use of these mutants as vaccine candidates. The RB51ΔcydC, RB51ΔcydD, RB51ΔpurD, RB51ΔcydCΔcydD and RB51ΔcydCΔpurD mutants exhibited significant attenuation of virulence when assayed in murine macrophages in vitro or in BALB/c mice. A single intraperitoneal immunization with RB51ΔcydC, RB51ΔcydD, RB51ΔcydCΔcydD or RB51ΔcydCΔpurD mutants was rapidly cleared from mice within 3 weeks, whereas the RB51ΔpurD mutant and RB51 were detectable in spleens until 4 and 7 weeks, respectively. Vaccination with a single dose of RB51 mutants induced lower protective immunity in mice than did parental RB51. However, a booster dose of these mutants provided significant levels of protection in mice against challenge with either the virulent homologous B. abortus strain 2308 or the heterologous Brucella canis strain 26. In addition, these mutants were found to induce a mixed but T-helper-1-biased humoral and cellular immune response in immunized mice. These data suggest that immunization with a booster dose of attenuated RB51 mutants provides an attractive strategy to protect against either bovine or canine brucellosis.

  19. Comparison between Immunization Routes of Live Attenuated Salmonella Typhimurium Strains Expressing BCSP31, Omp3b, and SOD of Brucella abortus in Murine Model.

    PubMed

    Kim, Won K; Moon, Ja Y; Kim, Suk; Hur, Jin

    2016-01-01

    Live, attenuated Salmonella Typhimurium vaccine candidate expressing BCSP31, Omp3b, and SOD proteins of Brucella abortus was constructed. Thirty BALB/c mice were divided equally into three groups, Group A, were intraperitoneally (IP) inoculated with 100 μl of approximately 1.2 × 10(6) colony-forming units (CFUs)/ml of the Salmonella containing vector only in 100 μl as a control. And groups B and C mice were orally and IP immunized with approximately 1.2 × 10(9) CFU/ml of the mixture of three delivery strains in 10 μl and IP immunized with approximately 1.2 × 10(6) CFU/ml of the mixture in 100 μl, respectively. The serum IgG, TNF-α and IFN-γ concentrations in groups B (except Omp3b) and C were significantly higher than those in group A. Following challenge with B. abortus strain 544; challenge strain was detected <10(3) CFU from the spleen of all mice of group C. These results suggest that IP immunization with the mixture of the vaccine candidate can induce immune responses, and can effectively protect mice against brucellosis.

  20. Comparison between Immunization Routes of Live Attenuated Salmonella Typhimurium Strains Expressing BCSP31, Omp3b, and SOD of Brucella abortus in Murine Model

    PubMed Central

    Kim, Won K.; Moon, Ja Y.; Kim, Suk; Hur, Jin

    2016-01-01

    Live, attenuated Salmonella Typhimurium vaccine candidate expressing BCSP31, Omp3b, and SOD proteins of Brucella abortus was constructed. Thirty BALB/c mice were divided equally into three groups, Group A, were intraperitoneally (IP) inoculated with 100 μl of approximately 1.2 × 106 colony-forming units (CFUs)/ml of the Salmonella containing vector only in 100 μl as a control. And groups B and C mice were orally and IP immunized with approximately 1.2 × 109 CFU/ml of the mixture of three delivery strains in 10 μl and IP immunized with approximately 1.2 × 106 CFU/ml of the mixture in 100 μl, respectively. The serum IgG, TNF-α and IFN-γ concentrations in groups B (except Omp3b) and C were significantly higher than those in group A. Following challenge with B. abortus strain 544; challenge strain was detected <103 CFU from the spleen of all mice of group C. These results suggest that IP immunization with the mixture of the vaccine candidate can induce immune responses, and can effectively protect mice against brucellosis. PMID:27148232

  1. Chlamydia Testing

    MedlinePlus

    ... Amplification Test (NAAT); Chlamydia trachomatis Culture; Chlamydia trachomatis DNA Probe Related tests: Gonorrhea Testing , HIV Antibody and HIV Antigen , Syphilis Tests , Herpes Testing , HPV Test , Trichomonas Testing All content on Lab Tests Online has ...

  2. Evidence for Chlamydia in wild mammals of the Serengeti.

    PubMed

    Pospischil, Andreas; Kaiser, Carmen; Hofmann-Lehmann, Regina; Lutz, Hans; Hilbe, Monika; Vaughan, Lloyd; Borel, Nicole

    2012-10-01

    Only limited information is available on the presence of Chlamydiaceae in wildlife, a deficit that is particularly acute concerning mammalian wildlife in Africa. In a retrospective analysis of organ material from an earlier study on wild mammals from the Seregenti National Park, 521 samples from 54 animals of 14 mammalian species were investigated. The presence of Chlamydiaceae was analyzed using molecular methods and immunohistochemistry. Chlamydial DNA was detected by real-time polymerase chain reaction in formalin-fixed and paraffin-embedded tissues from large ruminants (African buffaloes, Syncerus caffer, n=4) and a large predator (spotted hyena, Crocuta crocuta, n=1). Microarray results revealed Chlamydia abortus in all cases, confirmed by sequencing of selected samples, and a mixed infection with Chlamydia abortus and Chlamydia pneumoniae in an African buffalo. This is the first report of Chlamydiaceae in African wildlife of the Serengeti area.

  3. Complementation of Brucella abortus RB51 with a functional wboA gene results in O-antigen synthesis and enhanced vaccine efficacy but no change in rough phenotype and attenuation.

    PubMed

    Vemulapalli, R; He, Y; Buccolo, L S; Boyle, S M; Sriranganathan, N; Schurig, G G

    2000-07-01

    Brucella abortus RB51 is a stable rough, attenuated mutant vaccine strain derived from the virulent strain 2308. Recently, we demonstrated that the wboA gene in RB51 is disrupted by an IS711 element (R. Vemulapalli, J. R. McQuiston, G. G. Schurig, N. Srirauganathan, S. M. Halling, and S. M. Boyle, Clin. Diagn. Lab. Immunol. 6:760-764, 1999). Disruption of the wboA gene in smooth, virulent B. abortus, Brucella melitensis, and Brucella suis results in rough, attenuated mutants which fail to produce the O polysaccharide (O antigen). In this study, we explored whether the wboA gene disruption is responsible for the rough phenotype of RB51. We complemented RB51 with a functional wboA gene, and the resulting strain was designated RB51WboA. Colony and Western blot analyses indicated that RB51WboA expressed the O antigen; immunoelectron microscopy revealed that the O antigen was present in the cytoplasm. Crystal violet staining, acryflavin agglutination, and polymyxin B sensitivity studies indicated that RB51WboA had rough phenotypic characteristics similar to those of RB51. Bacterial clearance studies of BALB/c mice indicated no increase in the survival ability of RB51WboA in vivo compared to that of RB51. Vaccination of mice with live RB51WboA induced antibodies to the O antigen which were predominantly of the immunoglobulin G2a (IgG2a) and IgG3 isotypes. After in vitro stimulation of splenocytes with killed bacterial cells, quantitation of gamma interferon in the culture supernatants indicated that RB51WboA immunization induced higher levels of gamma interferon than immunization with RB51. Mice vaccinated with RB51WboA were better protected against a challenge infection with the virulent strain 2308 than those vaccinated with RB51. These studies indicate that in addition to the disruption of the wboA gene there is at least one other mutation in RB51 responsible for its rough phenotype. These studies also suggest that the expressed O antigen in RB51WboA is responsible

  4. Chlamydia Infections

    MedlinePlus

    ... PID). PID can cause permanent damage to your reproductive system. This can lead to long-term pelvic pain, infertility, and ectopic pregnancy. Women who have had chlamydia infections more than once are at higher risk of serious reproductive health complications. Men often don't have health ...

  5. Chlamydia bacteriophages.

    PubMed

    Śliwa-Dominiak, Joanna; Suszyńska, Ewa; Pawlikowska, Małgorzata; Deptuła, Wiesław

    2013-11-01

    Phages are called "good viruses" due to their ability to infect and kill pathogenic bacteria. Chlamydia are small, Gram-negative (G-) microbes that can be dangerous to human and animals. In humans, these bacteria are etiological agents of diseases such as psittacosis or respiratory tract diseases, while in animals, the infection may result in enteritis in cattle and chronic bowel diseases, as well as miscarriages in sheep. The first-known representative of chlamydiaphages was Chp1. It was discovered in Chlamydia psittaci isolates. Since then, four more species of chlamydiaphages have been identified [Chp2, Chp3, φCPG1 φCPAR39 (φCpn1) and Chp4]. All of them were shown to infect Chlamydia species. This paper described all known chlamydiaphages. They were characterised in terms of origin, host range, and their molecular structure. The review concerns the characterisation of bacteriophages that infects pathogenic and dangerous bacteria with unusual, intracellular life cycles that are pathogenic. In the era of antibiotic resistance, it is difficult to cure chlamydophilosis. Those bacteriophages can be an alternative to antibiotics, but before this happens, we need to get to know chlamydiaphages better. PMID:23903989

  6. Chlamydia bacteriophages.

    PubMed

    Śliwa-Dominiak, Joanna; Suszyńska, Ewa; Pawlikowska, Małgorzata; Deptuła, Wiesław

    2013-11-01

    Phages are called "good viruses" due to their ability to infect and kill pathogenic bacteria. Chlamydia are small, Gram-negative (G-) microbes that can be dangerous to human and animals. In humans, these bacteria are etiological agents of diseases such as psittacosis or respiratory tract diseases, while in animals, the infection may result in enteritis in cattle and chronic bowel diseases, as well as miscarriages in sheep. The first-known representative of chlamydiaphages was Chp1. It was discovered in Chlamydia psittaci isolates. Since then, four more species of chlamydiaphages have been identified [Chp2, Chp3, φCPG1 φCPAR39 (φCpn1) and Chp4]. All of them were shown to infect Chlamydia species. This paper described all known chlamydiaphages. They were characterised in terms of origin, host range, and their molecular structure. The review concerns the characterisation of bacteriophages that infects pathogenic and dangerous bacteria with unusual, intracellular life cycles that are pathogenic. In the era of antibiotic resistance, it is difficult to cure chlamydophilosis. Those bacteriophages can be an alternative to antibiotics, but before this happens, we need to get to know chlamydiaphages better.

  7. Clueing in on Chlamydia.

    ERIC Educational Resources Information Center

    Gibbons, Wendy

    1991-01-01

    Chlamydia's role in female infertility is discussed. The relationship of this organism to other diseases such as leprosy and tuberculosis is explained. Conditions caused by Chlamydia such as Pelvic Inflammatory Disease (PID) are described. (KR)

  8. Detection and genotyping of Chlamydia species responsible for reproductive disorders in Algerian small ruminants.

    PubMed

    Merdja, Salah-Eddine; Khaled, Hamza; Aaziz, Rachid; Vorimore, Fabien; Bertin, Claire; Dahmani, Ali; Bouyoucef, Abdallah; Laroucau, Karine

    2015-02-01

    Chlamydiosis in small ruminants is a zoonotic disease mainly related to Chlamydia abortus. This bacterium is responsible for abortions and reproductive disorders in sheep and goats. Stillbirth and infertility, leading to important economic losses, are also associated with this pathology. In Algeria, abortion cases are frequently reported by veterinarians but, except for brucellosis which is a notifiable disease in this country, abortive diseases are in general poorly studied. In order to detect and genotype Chlamydia species in small ruminants in different areas of Algeria, a study was conducted on samples collected from females (164 blood samples and 199 vaginal swabs) between October 2011 and March 2013. Serum samples were tested with a C. abortus-specific indirect ELISA test. Fourteen samples (8.5 %), from six farms (6/20, 30 %) were tested positive. Vaginal swabs were analysed with a real-time PCR targeting all Chlamydiaceae spp. Thirty samples (15 %) were diagnosed positive in 16 farms (16/25, 64 %). Positive samples were all re-tested with a C. abortus- and a C. pecorum-specific real-time PCR. Finally, 13/30 (43.3 %) and 6/30 (20 %) were identified as C. abortus and C. pecorum, respectively. Enough concentrated C. abortus samples were genotyped by multi-loci variable number of tandem repeat (VNTR) analysis (MLVA), and all were related to the genotype [2] group which mainly includes French C. abortus isolates. C. pecorum-positive samples were genotyped by multi-locus sequence typing (MLST). Interestingly, two of them were successfully genotyped and showed identical MLST sequences to VB2, AB10, E58 and SBE, a group which includes C. pecorum isolates considered as highly pathogenic. These findings suggest a possible role of C. abortus and C. pecorum strains in the aetiology of abortion in Algerian small ruminants.

  9. A dual-targeting approach to inhibit Brucella abortus replication in human cells

    PubMed Central

    Czyż, Daniel M.; Jain-Gupta, Neeta; Shuman, Howard A.; Crosson, Sean

    2016-01-01

    Brucella abortus is an intracellular bacterial pathogen and an etiological agent of the zoonotic disease known as brucellosis. Brucellosis can be challenging to treat with conventional antibiotic therapies and, in some cases, may develop into a debilitating and life-threatening chronic illness. We used multiple independent assays of in vitro metabolism and intracellular replication to screen a library of 480 known bioactive compounds for novel B. abortus anti-infectives. Eighteen non-cytotoxic compounds specifically inhibited B. abortus replication in the intracellular niche, which suggests these molecules function by targeting host cell processes. Twenty-six compounds inhibited B. abortus metabolism in axenic culture, thirteen of which are non-cytotoxic to human host cells and attenuate B. abortus replication in the intracellular niche. The most potent non-cytotoxic inhibitors of intracellular replication reduce B. abortus metabolism in axenic culture and perturb features of mammalian cellular biology including mitochondrial function and receptor tyrosine kinase signaling. The efficacy of these molecules as inhibitors of B. abortus replication in the intracellular niche suggests “dual-target” compounds that coordinately perturb host and pathogen are promising candidates for development of improved therapeutics for intracellular infections. PMID:27767061

  10. The Number, Organization, and Size of Polymorphic Membrane Protein Coding Sequences as well as the Most Conserved Pmp Protein Differ within and across Chlamydia Species.

    PubMed

    Van Lent, Sarah; Creasy, Heather Huot; Myers, Garry S A; Vanrompay, Daisy

    2016-01-01

    Variation is a central trait of the polymorphic membrane protein (Pmp) family. The number of pmp coding sequences differs between Chlamydia species, but it is unknown whether the number of pmp coding sequences is constant within a Chlamydia species. The level of conservation of the Pmp proteins has previously only been determined for Chlamydia trachomatis. As different Pmp proteins might be indispensible for the pathogenesis of different Chlamydia species, this study investigated the conservation of Pmp proteins both within and across C. trachomatis,C. pneumoniae,C. abortus, and C. psittaci. The pmp coding sequences were annotated in 16 C. trachomatis, 6 C. pneumoniae, 2 C. abortus, and 16 C. psittaci genomes. The number and organization of polymorphic membrane coding sequences differed within and across the analyzed Chlamydia species. The length of coding sequences of pmpA,pmpB, and pmpH was conserved among all analyzed genomes, while the length of pmpE/F and pmpG, and remarkably also of the subtype pmpD, differed among the analyzed genomes. PmpD, PmpA, PmpH, and PmpA were the most conserved Pmp in C. trachomatis,C. pneumoniae,C. abortus, and C. psittaci, respectively. PmpB was the most conserved Pmp across the 4 analyzed Chlamydia species.

  11. Toll-Like Receptor 4-Linked Janus Kinase 2 Signaling Contributes to Internalization of Brucella abortus by Macrophages

    PubMed Central

    Lee, Jin Ju; Kim, Dong Hyeok; Kim, Dae Geun; Lee, Hu Jang; Min, Wongi; Rhee, Man Hee; Cho, Jae Youl; Watarai, Masahisa

    2013-01-01

    Brucella abortus is an intracellular pathogen that uses a crafty strategy to invade and proliferate within host cells, but the distinct signaling pathways associated with phagocytic mechanisms of B. abortus remain unclear. The present study was performed to test the hypothesis that Toll-like receptor 4 (TLR4)-linked signaling interacting with Janus kinase 2 (JAK2) plays an essential role in B. abortus phagocytosis by macrophages. The effects of TLR4-JAK2 signaling on B. abortus phagocytosis in murine macrophage RAW 264.7 cells were observed through an infection assay and confocal microscopy. We determined that the uptake of B. abortus was negatively affected by the dysfunction of TLR4 and JAK2. F-actin polymerization detected by flow cytometry and F-actin assay was amplified for B. abortus entry, whereas that event was attenuated by the disruption of TLR4 and JAK2. Importantly, JAK2 phosphorylation and actin skeleton reorganization were suppressed immediately after B. abortus infection in bone marrow-derived macrophages (BMDMs) from TLR4−/− mice, showing the cooperation of JAK2 with TLR4. Furthermore, small GTPase Cdc42 participated in the intermediate pathway of TLR4-JAK2 signaling on B. abortus phagocytosis. Consequently, TLR4-associated JAK2 activation in the early cellular signaling events plays a pivotal role in B. abortus-induced phagocytic processes in macrophages, implying the pathogenic significance of JAK2-mediated entry. Here, we elucidate that this specific phagocytic mechanism of B. abortus might provide achievable strategies for inhibiting B. abortus invasion. PMID:23630962

  12. The Chlamydia psittaci Genome: A Comparative Analysis of Intracellular Pathogens

    PubMed Central

    Saluz, Hans Peter

    2012-01-01

    Background Chlamydiaceae are a family of obligate intracellular pathogens causing a wide range of diseases in animals and humans, and facing unique evolutionary constraints not encountered by free-living prokaryotes. To investigate genomic aspects of infection, virulence and host preference we have sequenced Chlamydia psittaci, the pathogenic agent of ornithosis. Results A comparison of the genome of the avian Chlamydia psittaci isolate 6BC with the genomes of other chlamydial species, C. trachomatis, C. muridarum, C. pneumoniae, C. abortus, C. felis and C. caviae, revealed a high level of sequence conservation and synteny across taxa, with the major exception of the human pathogen C. trachomatis. Important differences manifest in the polymorphic membrane protein family specific for the Chlamydiae and in the highly variable chlamydial plasticity zone. We identified a number of psittaci-specific polymorphic membrane proteins of the G family that may be related to differences in host-range and/or virulence as compared to closely related Chlamydiaceae. We calculated non-synonymous to synonymous substitution rate ratios for pairs of orthologous genes to identify putative targets of adaptive evolution and predicted type III secreted effector proteins. Conclusions This study is the first detailed analysis of the Chlamydia psittaci genome sequence. It provides insights in the genome architecture of C. psittaci and proposes a number of novel candidate genes mostly of yet unknown function that may be important for pathogen-host interactions. PMID:22506068

  13. Chlamydia pecorum: fetal and placental lesions in sporadic caprine abortion.

    PubMed

    Giannitti, Federico; Anderson, Mark; Miller, Myrna; Rowe, Joan; Sverlow, Karen; Vasquez, Marce; Cantón, Germán

    2016-03-01

    Chlamydial abortion in small ruminants is usually associated with Chlamydia abortus infection. Although Chlamydia pecorum has been detected in aborted ruminants and epidemiological data suggests that C. pecorum is abortigenic in these species, published descriptions of lesions in fetuses are lacking. This work describes fetoplacental lesions in a caprine abortion with C. pecorum infection, and further supports the abortigenic role of C. pecorum in ruminants. A 16-month-old Boer goat aborted twin fetuses at ~130 days of gestation. Both fetuses (A and B) and the placenta of fetus A were submitted for postmortem examination and diagnostic workup. At autopsy, the fetuses had moderate anasarca, intermuscular edema in the hindquarters (A), and brachygnathia and palatoschisis (B). In the placenta, the cotyledons were covered by yellow fibrinosuppurative exudate that extended into the adjacent intercotyledonary areas. Histologically, there was severe suppurative and necrotizing placentitis with vasculitis (arteriolitis) and thrombosis, multifocal lymphohistiocytic and neutrophilic hepatitis (A), and fibrinosuppurative enteritis in both fetuses. Chlamydia antigen was detected in the placenta by the direct fluorescent antibody test and in fetal intestines by immunohistochemistry. Nested polymerase chain reaction of DNA extracted from formalin-fixed, paraffin-embedded sections of placenta and intestine amplified 400 bp of the Chlamydia 16S rRNA gene that was sequenced and found to be 99% identical to C. pecorum by BLAST analysis. Other known abortigenic infectious agents were ruled out by specific testing. It is concluded that C. pecorum infection is associated with fetoplacental lesions and sporadic abortion in goats. PMID:26965241

  14. Increasing diversity within Chlamydiae.

    PubMed

    Corsaro, Daniele; Valassina, Marcello; Venditti, Danielle

    2003-01-01

    In recent years, 16S ribosomal DNA analyses has allowed the recognition of new chlamydia organisms, requiring the creation of new species, genera, and families within this unique, deep lineage of prokaryotes. The trachoma and psittaci groups chlamydiae are now recognized as separate genera, Chlamydia and Chlamydophila, respectively, and biovars of each group have been elevated to the species rank. Simkania and Parachlamydia have been associated with human respiratory infections, while Waddlia seems to be implicated in abortion in bovins. DNA amplification studies targeting the 16S rDNA have revealed a richer diversity within chlamydiae, identifying new lineages from both environmental and clinical samples. Further studies will be of interest to both examine the ecology and evaluate the clinical importance of these novel chlamydiae. Herein, we provide a summary of literature and our data about the novel chlamydial lineages.

  15. Seroepidemiologic survey for Chlamydia suis in wild boar (Sus scrofa) populations in Italy.

    PubMed

    Di Francesco, Antonietta; Donati, Manuela; Morandi, Federico; Renzi, Maria; Masia, Marco Antonio; Ostanello, Fabio; Salvatore, Daniela; Cevenini, Roberto; Baldelli, Raffaella

    2011-07-01

    We used serology to estimate the prevalence of exposure to chlamydiae in Italian populations of wild boars (Sus scrofa). Sera from 173 hunter-killed wild boars harvested during the 2006-2009 hunting seasons in three Italian regions were tested for antibodies to Chlamydia suis, Chlamydophila pecorum, Chlamydophila abortus, and Chlamydophila psittaci by the microimmunofluorescence test. Antibody titers to chlamydiae ≥ 1:32 were detected in 110 of the 173 samples tested (63.6%). Specific reactivity could be assessed only in 44 sera with antibody titers to C. suis that were two- to threefold higher than antibody titers against the other chlamydial species; the other 66 sera had similar reactivity against all the chlamydia species tested. Antibody to C. suis was detected in sera from wild boar populations with rare or no known contact with domestic pigs. These results suggest that the wild boar could be a chlamydia reservoir and may acquire chlamydiae independent of contacts with the domestic pig. PMID:21719838

  16. Zinc and Chlamydia trachomatis

    SciTech Connect

    Sugarman, B.; Epps, L.R.

    1985-07-01

    Zinc was noted to have significant effects upon the infection of McCoy cells by each of two strains of Chlamydia trachomatis. With a high or low Chlamydia inoculant, the number of infected cells increased up to 200% utilizing supplemental zinc (up to 1 x 10/sup -4/ M) in the inoculation media compared with standard Chlamydia cultivation media (8 x 10/sup -6/ M zinc). Ferric chloride and calcium chloride did not effect any such changes. Higher concentrations of zinc, after 2 hr of incubation with Chlamydia, significantly decreased the number of inclusions. This direct effect of zinc on the Chlamydia remained constant after further repassage of the Chlamydia without supplemental zinc, suggesting a lethal effect of the zinc. Supplemental zinc (up to 10/sup -4/ M) may prove to be a useful addition to inoculation media to increase the yield of culturing for Chlamydia trachomatis. Similarly, topical or oral zinc preparations used by people may alter their susceptibility to Chamydia trachomatis infections.

  17. Emendation of the family Chlamydiaceae: proposal of a single genus, Chlamydia, to include all currently recognized species.

    PubMed

    Sachse, Konrad; Bavoil, Patrik M; Kaltenboeck, Bernhard; Stephens, Richard S; Kuo, Cho-Chou; Rosselló-Móra, Ramon; Horn, Matthias

    2015-03-01

    The family Chlamydiaceae (order Chlamydiales, phylum Chlamydiae) comprises important, obligate intracellular bacterial pathogens of humans and animals. Subdivision of the family into the two genera Chlamydia and Chlamydophila has been discussed controversially during the past decade. Here, we have revisited the current classification in the light of recent genomic data and in the context of the unique biological properties of these microorganisms. We conclude that neither generally used 16S rRNA sequence identity cut-off values nor parameters based on genomic similarity consistently separate the two genera. Notably, no easily recognizable phenotype such as host preference or tissue tropism is available that would support a subdivision. In addition, the genus Chlamydophila is currently not well accepted and not used by a majority of research groups in the field. Therefore, we propose the classification of all 11 currently recognized Chlamydiaceae species in a single genus, the genus Chlamydia. Finally, we provide emended descriptions of the family Chlamydiaceae, the genus Chlamydia, as well as the species Chlamydia abortus, Chlamydia caviae and Chlamydia felis.

  18. Chlamydia (For Parents)

    MedlinePlus

    ... of STDs, including chlamydia. Latex condoms provide greater protection than natural-membrane condoms. The female condom, made ... against STDs. Although birth control pills offer no protection against STDs, they may provide some protection against ...

  19. Studies on a Smooth Phage-resistant Variant of Brucella abortus

    PubMed Central

    Corbel, M. J.; Morris, J. A.

    1974-01-01

    The immunological properties of a phage-resistant variant of Brucella abortus were examined. This variant, which was smooth and identical with the phage-susceptible parent strain in all cultural, biochemical and serological properties studied, was strongly antigenic in rabbits and guineapigs, inducing high titres of antibodies and delayed hypersensitivity to Br. abortus antigens. It was also fully virulent for mice and guinea-pigs. Contrary to other reports on the properties of phage-resistant Brucella strains, the in vivo properties of this strain showed no evidence of attenuation of virulence. PMID:4209104

  20. 9 CFR 113.65 - Brucella Abortus Vaccine.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... Bacterial Vaccines § 113.65 Brucella Abortus Vaccine. Brucella Abortus Vaccine shall be prepared as a desiccated live culture bacterial vaccine from smooth colonial forms of the Brucella abortus organism... shall be incubated at 35 to 37 °C for 96 hours. If growth not typical of Brucella abortus organisms...

  1. 9 CFR 113.65 - Brucella Abortus Vaccine.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... Bacterial Vaccines § 113.65 Brucella Abortus Vaccine. Brucella Abortus Vaccine shall be prepared as a desiccated live culture bacterial vaccine from smooth colonial forms of the Brucella abortus organism... shall be incubated at 35 to 37 °C for 96 hours. If growth not typical of Brucella abortus organisms...

  2. 9 CFR 113.65 - Brucella Abortus Vaccine.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... Bacterial Vaccines § 113.65 Brucella Abortus Vaccine. Brucella Abortus Vaccine shall be prepared as a desiccated live culture bacterial vaccine from smooth colonial forms of the Brucella abortus organism... shall be incubated at 35 to 37 °C for 96 hours. If growth not typical of Brucella abortus organisms...

  3. Chlamydia infections in women

    MedlinePlus

    ... for laboratory-based detection of Chlamydia trachomatis and Neisseria gonorrhoeae , 2014. MMWR 2014;63(No. RR-2):1 - 24. Centers for Disease Control and Prevention. Sexually transmitted diseases treatment guidelines. 2010. MMWR. 2010;59(RR-12):1- ...

  4. Chlamydia - CDC Fact Sheet

    MedlinePlus

    ... can cause serious, permanent damage to a woman’s reproductive system, making it difficult or impossible for her to ... chlamydia causes no symptoms, it can damage your reproductive system. Women with symptoms may notice •• An abnormal vaginal ...

  5. Delivery of Chlamydia vaccines.

    PubMed

    Igietseme, Joseph; Eko, Francis; He, Qing; Bandea, Claudiu; Lubitz, Werner; Garcia-Sastre, Adolfo; Black, Carolyn

    2005-05-01

    The plethora of ocular, genital and respiratory diseases of Chlamydia, including nongonococcal urethritis, cervicitis pelvic inflammatory disease, ectopic pregnancy, tubal factor infertility, conjunctivitis, blinding trachoma and interstitial pneumonia, and chronic diseases that may include atherosclerosis, multiple sclerosis, adult onset asthma and Alzheimer's disease, still pose a considerable public health challenge to many nations. Although antibiotics are effective against Chlamydia when effectively diagnosed, asymptomatic infections are rampart, making clinical presentation of complications often the first evidence of an infection. Consequently, the current medical opinion is that an effective prophylactic vaccine would constitute the best approach to protect the human population from the most severe consequences of these infections. Clinical and experimental studies have demonstration that Chlamydia immunity in animals and humans is mediated by T cells and a complementary antibody response, and the completion of the genome sequencing of several isolates of Chlamydia is broadening our knowledge of the immunogenic antigens with potential vaccine value. Thus, major advances have been made in defining the essential elements of a potentially effective subunit vaccine design and parameters for evaluation. However, the challenge to develop effective delivery systems and human compatible adjuvants that would boost the immune response to achieve long-lasting protective immunity remains an elusive objective in chlamydial vaccine research. In response to evolving molecular and cellular technologies and novel vaccinology approaches, considerable progress is being made in the construction of novel delivery systems, such as DNA and plasmid expression systems, viral vectors, living and nonliving bacterial delivery systems, the use of chemical adjuvants, lipoprotein constructs and the codelivery of vaccines and specific immuno-modulatory biological agonists targeting

  6. Defensin Susceptibility and Colonization in the Mouse Model of AJ100, a Polymyxin B Resistant, Brucella abortus RB51 Isolate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacterial facultative intracellular pathogens selected for sensitivity to polymyxin B have been shown to be attenuated. However, the effect on pathogenicity of increased resistance to polymyxin B has not been studied. A polymyxin resistant mutant, Brucella abortus AJ100, was characterized by comp...

  7. Longitudinal prevalence and faecal shedding of Chlamydia pecorum in sheep.

    PubMed

    Yang, Rongchang; Jacobson, Caroline; Gardner, Graham; Carmichael, Ian; Campbell, Angus J D; Ryan, Una

    2014-09-01

    The prevalence and faecal shedding of Chlamydia spp. in sheep in Australia has not been well described. Two species-specific quantitative PCRs (qPCRs) targeting the chlamydial outer membrane protein cell surface antigen gene (ompA) were validated and used to determine the prevalence and faecal shedding of C. abortus and C. pecorum from faecal samples of lambs at three sampling times (weaning, post-weaning and pre-slaughter) from eight farms in South Australia, New South Wales, Victoria and Western Australia. A total of 3412 faecal samples were collected and screened from approximately 1189 lambs across the four states. C. abortus was not detected in any of the samples screened. The overall prevalence of C. pecorum was 1027/3412 (30.1%) and median bacterial concentrations at weaning, post-weaning and pre-slaughter were 1.8 × 10(7), 1.2 × 10(7) and 9.6 × 10(5)/g faeces, respectively. A subset of C. pecorum positive samples from each farm, (n = 48) was sequenced to confirm their identity. The present study demonstrates that C. pecorum is prevalent in Australian sheep, highlighting a need for further research on the impact of this bacterium on production.

  8. Mutational Analysis of the Chlamydia muridarum Plasticity Zone

    PubMed Central

    Rajaram, Krithika; Giebel, Amanda M.; Toh, Evelyn; Hu, Shuai; Newman, Jasmine H.; Morrison, Sandra G.; Kari, Laszlo; Morrison, Richard P.

    2015-01-01

    Pathogenically diverse Chlamydia spp. can have surprisingly similar genomes. Chlamydia trachomatis isolates that cause trachoma, sexually transmitted genital tract infections (chlamydia), and invasive lymphogranuloma venereum (LGV) and the murine strain Chlamydia muridarum share 99% of their gene content. A region of high genomic diversity between Chlamydia spp. termed the plasticity zone (PZ) may encode niche-specific virulence determinants that dictate pathogenic diversity. We hypothesized that PZ genes might mediate the greater virulence and gamma interferon (IFN-γ) resistance of C. muridarum compared to C. trachomatis in the murine genital tract. To test this hypothesis, we isolated and characterized a series of C. muridarum PZ nonsense mutants. Strains with nonsense mutations in chlamydial cytotoxins, guaBA-add, and a phospholipase D homolog developed normally in cell culture. Two of the cytotoxin mutants were less cytotoxic than the wild type, suggesting that the cytotoxins may be functional. However, none of the PZ nonsense mutants exhibited increased IFN-γ sensitivity in cell culture or were profoundly attenuated in a murine genital tract infection model. Our results suggest that C. muridarum PZ genes are transcribed—and some may produce functional proteins—but are dispensable for infection of the murine genital tract. PMID:25939505

  9. Tissue-Resident T Cells as the Central Paradigm of Chlamydia Immunity

    PubMed Central

    Johnson, Raymond M.

    2016-01-01

    For almost 2 decades, results from Chlamydia pathogenesis investigations have been conceptualized using a cytokine polarization narrative. Recent viral immunity studies identifying protective tissue-resident memory T cells (Trm) suggest an alternative paradigm based on localized immune networks. As Chlamydia vaccines enter the preclinical pipeline and, in the case of an attenuated trachoma vaccine, are given to human subjects, it may be useful to ask whether cytokine polarization is the appropriate framework for understanding and evaluating vaccine efficacy. In this review, we revisit C. trachomatis pathogenesis data from mice and humans using a Trm narrative and note a comfortable concordance with the Chlamydia pathogenesis literature. PMID:26787715

  10. Tissue-Resident T Cells as the Central Paradigm of Chlamydia Immunity.

    PubMed

    Johnson, Raymond M; Brunham, Robert C

    2016-04-01

    For almost 2 decades, results from Chlamydia pathogenesis investigations have been conceptualized using a cytokine polarization narrative. Recent viral immunity studies identifying protective tissue-resident memory T cells (Trm) suggest an alternative paradigm based on localized immune networks. As Chlamydia vaccines enter the preclinical pipeline and, in the case of an attenuated trachoma vaccine, are given to human subjects, it may be useful to ask whether cytokine polarization is the appropriate framework for understanding and evaluating vaccine efficacy. In this review, we revisit C. trachomatis pathogenesis data from mice and humans using a Trm narrative and note a comfortable concordance with the Chlamydia pathogenesis literature.

  11. Identification of Chlamydiae and Mycoplasma species in ruminants with ocular infections.

    PubMed

    Gupta, S; Chahota, R; Bhardwaj, B; Malik, P; Verma, S; Sharma, M

    2015-02-01

    Infectious keratoconjunctivitis (IKC) is a highly contagious ocular inflammatory condition, which is often reported in domestic small and large ruminants. Multiple infectious aetiologies are reported to be involved, but information about the role of certain fastidious bacterial pathogens such as chlamydiae and mycoplasmas is limited in India. Hence, this study was performed to determine the role of these pathogens and their identification by molecular approach. A total of 53 samples from 31 ovine, 14 caprine and eight bovine having clinical symptoms were collected and tested using species-specific PCR tests for chlamydiae and mycoplasmas followed by nucleotide sequence analysis. The results showed 77.41, 14.29 and 25% samples were chlamydiae positive in ovine, caprine and bovine, respectively, whereas 41.93, 14.29 and 37.5% prevalence of mycoplasma infection was detected in ovine, caprine and bovines, respectively. Chlamydophila abortus, Chlamydophila psittaci, Mycoplasma arginini and Mycoplasma hyorhinis were detected from tested samples. To the best of our knowledge, this is the first time these species are identified in IKC cases from India. Coinfection of both chlamydial and mycoplasmal species was detected in eight IKC cases of ovine which suggest synergistic roles played by both chlamydiae and mycoplasma in IKC samples.

  12. Characterization and protective property of Brucella abortus cydC and looP mutants.

    PubMed

    Truong, Quang Lam; Cho, Youngjae; Barate, Abhijit Kashinath; Kim, Suk; Hahn, Tae-Wook

    2014-11-01

    Brucella abortus readily multiplies in professional or nonprofessional phagocytes in vitro and is highly virulent in mice. Isogenic mutants of B. abortus biovar 1 strain IVKB9007 lacking the ATP/GDP-binding protein motif A (P-loop) (named looP; designated here the IVKB9007 looP::Tn5 mutant) and the ATP-binding/permease protein (cydC; designated here the IVKB9007 cydC::Tn5 mutant) were identified and characterized by transposon mutagenesis using the mini-Tn5Km2 transposon. Both mutants were found to be virtually incapable of intracellular replication in both murine macrophages (RAW264.7) and the HeLa cell line, and their virulence was significantly impaired in BALB/c mice. Respective complementation of the IVKB9007 looP::Tn5 and IVKB9007 cydC::Tn5 mutants restored their ability to survive in vitro and in vivo to a level comparable with that of the wild type. These findings indicate that the cydC and looP genes play important roles in the virulence of B. abortus. In addition, intraperitoneal immunization of mice with a dose of the live IVKB9007 looP::Tn5 and IVKB9007 cydC::Tn5 mutants provided a high degree of protection against challenge with pathogenic B. abortus strain 544. Both mutants should be evaluated further as a live attenuated vaccine against bovine brucellosis for their ability to stimulate a protective immune response.

  13. Characterization of a murine model of intranasal infection suitable for testing vaccines against C. abortus.

    PubMed

    Buendía, A J; Nicolás, L; Ortega, N; Gallego, M C; Martinez, C M; Sanchez, J; Caro, M R; Navarro, J A; Salinas, J

    2007-01-15

    Mouse models have been widely used to test candidate vaccines against Chlamydophila abortus infection in mice. Although the induction of a systemic infection by endogenous or intraperitoneal inoculation is a useful tool for understanding the immune mechanism involved in the protection conferred by the vaccination, a different approach is necessary to understand other factors of the infection, such as mucosal immunity or the colonization of target organs. To test whether C. abortus intranasal model of infection in mice is a useful tool for testing vaccines in a first group of experiments mice, were infected intranasally with C. abortus to characterize the model of infection. When this model was used to test vaccines, two inactivated experimental vaccines, one of them adjuvated with QS-21 and another with aluminium hydroxide, and a live attenuated vaccine (strain 1B) were used. Non-vaccinated control mice died within the first 8 days, after displaying substantial loss of weight. Histologically, the mice showed lobar fibrinopurulent bronchointerstitial pneumonia. Prior immunization with QS-21 adjuvated vaccine or 1B vaccine presented mortality and the recipients showed a greater number of T cells in the lesions, especially CD8(+) T cells, than the control mice and mice immunized with vaccine adjuvated with aluminium hydroxide. The results confirm that the C. abortus intranasal model of infection in mice is a useful tool for testing vaccines.

  14. Chlamydia pneumoniae (TWAR).

    PubMed Central

    Kuo, C C; Jackson, L A; Campbell, L A; Grayston, J T

    1995-01-01

    Chlamydia pneumoniae (TWAR) is a recently recognized third species of the genus Chlamydia that causes acute respiratory disease. It is distinct from the other two chlamydial species that infect humans, C. trachomatis and C. psittaci, in elementary body morphology and shares less than 10% of the DNA homology with those species. The organism has a global distribution, with infection most common among children between the ages of 5 and 14 years. In children, TWAR infection is usually mild or asymptomatic, but it may be more severe in adults. Pneumonia and bronchitis are the most common clinical manifestations of infection, and TWAR is responsible for approximately 10% of cases of pneumonia and 5% of cases of bronchitis in the United States. The microimmunofluorescence serologic assay is specific for TWAR and can distinguish between recent and past infections. The organism can be isolated in cell culture; however, PCR techniques have recently facilitated its detection in tissues and clinical specimens. PMID:8665464

  15. The Brucella abortus General Stress Response System Regulates Chronic Mammalian Infection and Is Controlled by Phosphorylation and Proteolysis*

    PubMed Central

    Kim, Hye-Sook; Caswell, Clayton C.; Foreman, Robert; Roop, R. Martin; Crosson, Sean

    2013-01-01

    Brucella spp. are adept at establishing a chronic infection in mammals. We demonstrate that core components of the α-proteobacterial general stress response (GSR) system, PhyR and σE1, are required for Brucella abortus stress survival in vitro and maintenance of chronic murine infection in vivo. ΔphyR and ΔrpoE1 null mutants exhibit decreased survival under acute oxidative and acid stress but are not defective in infection of primary murine macrophages or in initial colonization of BALB/c mouse spleens. However, ΔphyR and ΔrpoE1 mutants are attenuated in spleens beginning 1 month postinfection. Thus, the B. abortus GSR system is dispensable for colonization but is required to maintain chronic infection. A genome-scale analysis of the B. abortus GSR regulon identified stress response genes previously linked to virulence and genes that affect immunomodulatory components of the cell envelope. These data support a model in which the GSR system affects both stress survival and the interface between B. abortus and the host immune system. We further demonstrate that PhyR proteolysis is a unique feature of GSR control in B. abortus. Proteolysis of PhyR provides a mechanism to avoid spurious PhyR protein interactions that inappropriately activate GSR-dependent transcription. We conclude that the B. abortus GSR system regulates acute stress adaptation and long term survival within a mammalian host and that PhyR proteolysis is a novel regulatory feature in B. abortus that ensures proper control of GSR transcription. PMID:23546883

  16. Analysis of Polymorphic Membrane Protein Expression in Cultured Cells Identifies PmpA and PmpH of Chlamydia psittaci as Candidate Factors in Pathogenesis and Immunity to Infection

    PubMed Central

    Van Lent, Sarah; De Vos, Winnok H.; Huot Creasy, Heather; Marques, Patricia X.; Ravel, Jacques; Vanrompay, Daisy; Bavoil, Patrik; Hsia, Ru-ching

    2016-01-01

    The polymorphic membrane protein (Pmp) paralogous families of Chlamydia trachomatis, Chlamydia pneumoniae and Chlamydia abortus are putative targets for Chlamydia vaccine development. To determine whether this is also the case for Pmp family members of C. psittaci, we analyzed transcription levels, protein production and localization of several Pmps of C. psittaci. Pmp expression profiles were characterized using quantitative real-time PCR (RT-qPCR), immunofluorescence (IF) and immuno-electron microscopy (IEM) under normal and stress conditions. We found that PmpA was highly produced in all inclusions as early as 12 hpi in all biological replicates. In addition, PmpA and PmpH appeared to be unusually accessible to antibody as determined by both immunofluorescence and immuno-electron microscopy. Our results suggest an important role for these Pmps in the pathogenesis of C. psittaci, and make them promising candidates in vaccine development. PMID:27631978

  17. Analysis of Polymorphic Membrane Protein Expression in Cultured Cells Identifies PmpA and PmpH of Chlamydia psittaci as Candidate Factors in Pathogenesis and Immunity to Infection.

    PubMed

    Van Lent, Sarah; De Vos, Winnok H; Huot Creasy, Heather; Marques, Patricia X; Ravel, Jacques; Vanrompay, Daisy; Bavoil, Patrik; Hsia, Ru-Ching

    2016-01-01

    The polymorphic membrane protein (Pmp) paralogous families of Chlamydia trachomatis, Chlamydia pneumoniae and Chlamydia abortus are putative targets for Chlamydia vaccine development. To determine whether this is also the case for Pmp family members of C. psittaci, we analyzed transcription levels, protein production and localization of several Pmps of C. psittaci. Pmp expression profiles were characterized using quantitative real-time PCR (RT-qPCR), immunofluorescence (IF) and immuno-electron microscopy (IEM) under normal and stress conditions. We found that PmpA was highly produced in all inclusions as early as 12 hpi in all biological replicates. In addition, PmpA and PmpH appeared to be unusually accessible to antibody as determined by both immunofluorescence and immuno-electron microscopy. Our results suggest an important role for these Pmps in the pathogenesis of C. psittaci, and make them promising candidates in vaccine development. PMID:27631978

  18. Complement fixation test to assess humoral immunity in cattle and sheep vaccinated with Brucella abortus RB51.

    PubMed

    Adone, R; Ciuchini, F

    1999-11-01

    The live attenuated Brucella abortus strain RB51 is a rifampin-resistant, lipopolysaccharide (LPS) O-chain-deficient mutant of virulent B. abortus 2308. The reduced O-chain content in RB51 prevents this bacterium from inducing antibodies detectable by the conventional serologic tests for bovine brucellosis diagnosis that mainly identify antibodies to LPS. The absence of available serologic tests for RB51 also complicates the diagnosis of possible RB51 infections in humans exposed to this strain. The purpose of this study was to evaluate the suitability of a complement fixation (CF) test performed with the rough strain B. abortus RB51, previously deprived of anticomplementary activity, in detecting anti-B. abortus RB51 antibodies in cattle and sheep experimentally vaccinated with this strain. The results of this study showed that a CF test with RB51 as the antigen is able to specifically detect antibodies following RB51 vaccination in cattle and sheep. In addition, this method could be a useful tool for detecting B. abortus RB51 infection in humans.

  19. B-Cell-Deficient Mice Show an Exacerbated Inflammatory Response in a Model of Chlamydophila abortus Infection

    PubMed Central

    Buendía, Antonio J.; Del Río, Laura; Ortega, Nieves; Sánchez, Joaquín; Gallego, María C.; Caro, María R.; Navarro, Jose A.; Cuello, Francisco; Salinas, Jesús

    2002-01-01

    The resolution of Chlamydophila abortus (Chlamydia psittaci serotype 1) infection is dependent on gamma interferon and CD8+ T cells, and classically, B cells have been considered to play a minimal role in host defense. The role of B cells in the immune response was studied by using a model of infection in mice with genetically modified immunoglobulin M transmembrane domains (μMT). In the absence of B cells, infection with C. abortus leads to an acute severe fatal disease that involves a disseminated intravascular coagulation syndrome. μMT mice displayed an increased level of proinflammatory cytokines in serum, and an increased number of neutrophils was observed in the lesions. The possible deleterious role of neutrophils in the pathogenesis of disease in μMT mice was determined by depletion of the neutrophils with the monoclonal antibody RB6-8C5. This led to an enhancement of the bacterial burden and early mortality in both μMT and wild-type mice, while necrotic lesions remained. Analysis of the presence of immunoregulatory cytokines showed significantly lower levels of transforming growth factor β in the sera of μMT mice. However, mice lacking mature B cells were able to establish a specific immune response that protected them from a secondary challenge. Taken together, these data suggest an immunomodulatory role for B cells in the early events of C. abortus primary infection that can protect mice against an exaggerated inflammatory response. PMID:12438369

  20. B-cell-deficient mice show an exacerbated inflammatory response in a model of Chlamydophila abortus infection.

    PubMed

    Buendía, Antonio J; Del Río, Laura; Ortega, Nieves; Sánchez, Joaquín; Gallego, María C; Caro, María R; Navarro, Jose A; Cuello, Francisco; Salinas, Jesús

    2002-12-01

    The resolution of Chlamydophila abortus (Chlamydia psittaci serotype 1) infection is dependent on gamma interferon and CD8(+) T cells, and classically, B cells have been considered to play a minimal role in host defense. The role of B cells in the immune response was studied by using a model of infection in mice with genetically modified immunoglobulin M transmembrane domains ( micro MT). In the absence of B cells, infection with C. abortus leads to an acute severe fatal disease that involves a disseminated intravascular coagulation syndrome. micro MT mice displayed an increased level of proinflammatory cytokines in serum, and an increased number of neutrophils was observed in the lesions. The possible deleterious role of neutrophils in the pathogenesis of disease in micro MT mice was determined by depletion of the neutrophils with the monoclonal antibody RB6-8C5. This led to an enhancement of the bacterial burden and early mortality in both micro MT and wild-type mice, while necrotic lesions remained. Analysis of the presence of immunoregulatory cytokines showed significantly lower levels of transforming growth factor beta in the sera of micro MT mice. However, mice lacking mature B cells were able to establish a specific immune response that protected them from a secondary challenge. Taken together, these data suggest an immunomodulatory role for B cells in the early events of C. abortus primary infection that can protect mice against an exaggerated inflammatory response.

  1. Chlamydia cell biology and pathogenesis

    PubMed Central

    Elwell, Cherilyn; Mirrashidi, Kathleen; Engel, Joanne

    2016-01-01

    Chlamydia spp. are important causes of human disease for which no effective vaccine exists. These obligate intracellular pathogens replicate in a specialized membrane compartment and use a large arsenal of secreted effectors to survive in the hostile intracellular environment of the host. In this Review, we summarize the progress in decoding the interactions between Chlamydia spp. and their hosts that has been made possible by recent technological advances in chlamydial proteomics and genetics. The field is now poised to decipher the molecular mechanisms that underlie the intimate interactions between Chlamydia spp. and their hosts, which will open up many exciting avenues of research for these medically important pathogens. PMID:27108705

  2. Chlamydia cell biology and pathogenesis.

    PubMed

    Elwell, Cherilyn; Mirrashidi, Kathleen; Engel, Joanne

    2016-06-01

    Chlamydia spp. are important causes of human disease for which no effective vaccine exists. These obligate intracellular pathogens replicate in a specialized membrane compartment and use a large arsenal of secreted effectors to survive in the hostile intracellular environment of the host. In this Review, we summarize the progress in decoding the interactions between Chlamydia spp. and their hosts that has been made possible by recent technological advances in chlamydial proteomics and genetics. The field is now poised to decipher the molecular mechanisms that underlie the intimate interactions between Chlamydia spp. and their hosts, which will open up many exciting avenues of research for these medically important pathogens.

  3. Brucella abortus Cyclic β-1,2-Glucan Mutants Have Reduced Virulence in Mice and Are Defective in Intracellular Replication in HeLa Cells

    PubMed Central

    Briones, Gabriel; Iñón de Iannino, Nora; Roset, Mara; Vigliocco, Ana; Paulo, Patricia Silva; Ugalde, Rodolfo A.

    2001-01-01

    Null cyclic β-1,2-glucan synthetase mutants (cgs mutants) were obtained from Brucella abortus virulent strain 2308 and from B. abortus attenuated vaccinal strain S19. Both mutants show greater sensitivity to surfactants like deoxycholic acid, sodium dodecyl sulfate, and Zwittergent than the parental strains, suggesting cell surface alterations. Although not to the same extent, both mutants display reduced virulence in mice and defective intracellular multiplication in HeLa cells. The B. abortus S19 cgs mutant was completely cleared from the spleens of mice after 4 weeks, while the 2308 mutant showed a 1.5-log reduction of the number of brucellae isolated from the spleens after 12 weeks. These results suggest that cyclic β-1,2-glucan plays an important role in the residual virulence of the attenuated B. abortus S19 strain. Although the cgs mutant was cleared from the spleens earlier than the wild-type parental strain (B. abortus S19) and produced less inflammatory response, its ability to confer protection against the virulent strain B. abortus 2308 was fully retained. Equivalent levels of induction of spleen gamma interferon mRNA and anti-lipopolysaccharide (LPS) of immunoglobulin G2a (IgG2a) subtype antibodies were observed in mice injected with B. abortus S19 or the cgs mutant. However, the titer of anti-LPS antibodies of the IgG1 subtype induced by the cgs mutant was lower than that observed with the parental S19 strain, thus suggesting that the cgs mutant induces a relatively exclusive Th1 response. PMID:11401996

  4. Increases of efficacy as vaccine against Brucella abortus infection in mice by simultaneous inoculation with avirulent smooth bvrS/bvrR and rough wbkA mutants.

    PubMed

    Grilló, María Jesús; Manterola, Lorea; de Miguel, María Jesús; Muñoz, Pilar María; Blasco, José María; Moriyón, Ignacio; López-Goñi, Ignacio

    2006-04-01

    The Brucella abortus S19 and RB51 strains are the most widely used live vaccines against bovine brucellosis. However, both can induce abortion and milk excretion, S19 vaccination interferes in serological tests, and RB51 is less effective. We have shown previously that a rough wbkAB. abortus mutant is attenuated and a better vaccine than RB51 in BALB/c mice, and that mutants in the two-component regulatory system bvrS/bvrR are markedly attenuated while keeping a smooth lipopolysaccharide (S-LPS). In this work, we tested whether simultaneous inoculation with live bvrS increases wbkA vaccine efficacy in mice. Even at high doses, the bvrS mutant was cleared much faster from spleens than the wbkA mutant. The splenic persistence of the wbkA mutant increased when inoculated along with the bvrS mutant, but also with inactivated bvrS cells or with purified B. abortus S-LPS, strongly suggesting that S-LPS in the bvrS mutant played a determinant role in the wbkA persistence. When inoculated alone, both mutants protected against virulent B. abortus but less than when inoculated simultaneously, and the protection afforded by the combination was better than that obtained with B. abortus S19. Increased protection was also obtained after simultaneous inoculation of the wbkA mutant and inactivated bvrS cells or purified S-LPS, showing again the role played by the S-LPS in the bvrS cells. In mice, the bvrS-wbkA combination induced an antibody response reduced with respect to B. abortus S19 vaccination. Thus, the simultaneous use of live bvrS and wbkA B. abortus mutants seems a promising approach to overcome the problems of the S19 andRB51 vaccines.

  5. Recent advances in Brucella abortus vaccines.

    PubMed

    Dorneles, Elaine M S; Sriranganathan, Nammalwar; Lage, Andrey P

    2015-07-08

    Brucella abortus vaccines play a central role in bovine brucellosis control/eradication programs and have been successfully used worldwide for decades. Strain 19 and RB51 are the approved B. abortus vaccines strains most commonly used to protect cattle against infection and abortion. However, due to some drawbacks shown by these vaccines much effort has been undertaken for the development of new vaccines, safer and more effective, that could also be used in other susceptible species of animals. In this paper, we present a review of the main aspects of the vaccines that have been used in the brucellosis control over the years and the current research advances in the development of new B. abortus vaccines.

  6. A rapid cycleave PCR method for distinguishing the vaccine strain Brucella abortus A19 in China.

    PubMed

    Nan, Wenlong; Zhang, Yueyong; Tan, Pengfei; Xu, Zouliang; Chen, Yuqi; Mao, Kairong; Chen, Yiping

    2016-05-01

    Brucellosis is a widespread zoonotic disease caused by Brucella spp. Immunization with attenuated vaccines has proved to be an effective method of prevention; however, it may also interfere with diagnosis. Brucella abortus strain A19, which is homologous to B. abortus strain S19, is widely used for the prevention of bovine brucellosis in China. For effective monitoring of the control of brucellosis, it is essential to distinguish A19 from field strains. Single-nucleotide polymorphism-based assays offer a new approach to such discrimination studies. In the current study, we developed a cycleave PCR assay that successfully distinguished attenuated vaccine strains A19 and S19 from 22 strains of B. abortus and 57 strains of 5 other Brucella species. The assay gave a negative reaction with 4 non-Brucella species. The minimum sensitivity of the assay, evaluated using 10-fold dilutions of chromosomal DNA, was 7.6 fg for the A19 strain and 220 fg for the single non-A19/non-S19 Brucella strain tested (B. abortus 104M). The assay was also reproducible (intra- and interassay coefficients of variation: 0.003-0.01 and 0.004-0.025, respectively). The cycleave assay gave an A19/S19-specific reaction in 3 out of 125 field serum samples, with the same 3 samples being positive in an alternative A19/S19-specific molecular assay. The cycleave assay gave a total of 102 Brucella-specific reactions (3 being the A19/S19-specific reactions), whereas an alternative Brucella-specific assay gave 92 positive reactions (all also positive in the cycleave assay). Therefore, this assay represents a simple, rapid, sensitive, and specific tool for use in brucellosis control.

  7. Killing of Brucella abortus by bovine serum.

    PubMed

    Corbeil, L B; Blau, K; Inzana, T J; Nielsen, K H; Jacobson, R H; Corbeil, R R; Winter, A J

    1988-12-01

    Studies of the serum bactericidal system in bovine brucellosis were undertaken to investigate the role of the humoral immune response in protection of cattle against the facultative intracellular parasite Brucella abortus. Fresh sera from normal control cattle, infected cattle, and cattle immunized with B. abortus cell envelopes were collected before treatment and during the course of immunization or infection. Normal fresh bovine serum or fresh agammaglobulinemic serum from colostrum-deprived calves was effective in killing smooth virulent B. abortus 2308, but rough strains RB51 (a rough mutant of strain 2308) and 45/20 were much more sensitive to serum. The difference in susceptibility to serum was shown to be correlated with differences in lipopolysaccharide chemotype, with the more resistant strain 2308 having O polysaccharide and the more susceptible strains 45/20 and RB51 lacking O side chains. By treatment of fresh serum with MgCl2 and EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid] killing was shown to occur via the classical pathway of complement activation. When antibody to B. abortus was present, killing of strain RB51 increased but killing of smooth strain 2308 decreased. The earliest antibody response in serum from infected animals did not interfere with killing. When affinity-purified bovine immunoglobulins specific for B. abortus smooth lipopolysaccharide were added to fresh normal bovine serum, immunoglobulin G1 (IgG1) and IgG2 isotypes blocked killing but IgM and IgA isotypes did not. Thus, it appears that serum from previously unexposed animals or animals early during infection can kill smooth B. abortus, an appropriate defense mechanism before the organism becomes intracellular. At later stages of infection, blocking antibodies predominate.

  8. Chlamydia Infections - Multiple Languages: MedlinePlus

    MedlinePlus

    ... Are Here: Home → Multiple Languages → All Health Topics → Chlamydia Infections URL of this page: https://medlineplus.gov/ ... V W XYZ List of All Topics All Chlamydia Infections - Multiple Languages To use the sharing features ...

  9. Immunochemical identification of Brucella abortus lipopolysaccharide epitopes.

    PubMed Central

    Rojas, N; Freer, E; Weintraub, A; Ramirez, M; Lind, S; Moreno, E

    1994-01-01

    Sera from Brucella abortus-infected and -vaccinated bovines recognized four lipopolysaccharide (LPS) determinants: two in the O-polysaccharide (A and C), one in the core oligosaccharide from rough Brucella LPS (R), and one in lipid A (LA). From 46 different hybridomas secreting monoclonal antibodies (MAbs) against various LPS moieties, 9 different specificities were identified. Two epitopes, A and C/Y, were present in the O-polysaccharide. Two epitopes were found in the core oligosaccharide (R1 and R2) of rough Brucella LPS. MAbs against R1 and R2 epitopes reacted against LPS from different rough Brucella species; however, MAbs directed to the R2 epitope also reacted against enterobacterial LPS from deep rough mutants. Three epitopes (LA1, LA2, and LA3) were located in the lipid A backbone. Different sets of MAbs recognized two epitopes in the lipid A-associated outer membrane protein (LAOmp3-1 and LAOmp3-2). LPS preparations from smooth brucellae had small amounts of rough-type LPS. Although LPS from rough brucellae did not show smooth-type LPS in western blots (immunoblots), two hybridomas generated from mice immunized with rough B. abortus produced antibodies against smooth B. abortus LPS. Results are discussed in relation to the structure and function of B. abortus LPS and to previous findings on the epitopic density of the molecule. Images PMID:7496947

  10. Brucella abortus-infected B cells induce osteoclastogenesis.

    PubMed

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2016-09-01

    Brucella abortus is an intracellular bacterium that establishes lifelong infections in livestock and humans although the mechanisms of its chronicity are poorly understood. Activated B cells have long lifespan and B. abortus infection activates B cells. Our results indicate that the direct infection of B cells with B. abortus induced matrix metalloproteinase-9 (MMP-9), receptor activator for NF κB ligand (RANKL), tumor necrosis factor (TNF)-α and interleukin (IL)-6 secretion. In addition, supernatants from B. abortus-infected B cells induced bone marrow-derived monocytes to undergo osteoclastogenesis. Using osteoprotegerin, RANKL's decoy receptor, we determined that RANKL is involved in osteoclastogenesis induced by supernatants from B. abortus-infected B cells. The results presented here shed light on how the interactions of B. abortus with B cells may have a role in the pathogenesis of brucellar osteoarticular disease.

  11. Proteomic analysis of Brucella abortus cell envelope and identification of immunogenic candidate proteins for vaccine development.

    PubMed

    Connolly, Joseph P; Comerci, Diego; Alefantis, Timothy G; Walz, Alexander; Quan, Marian; Chafin, Ryan; Grewal, Paul; Mujer, Cesar V; Ugalde, Rodolfo A; DelVecchio, Vito G

    2006-07-01

    Brucella abortus is the etiologic agent of bovine brucellosis and causes a chronic disease in humans known as undulant fever. In livestock the disease is characterized by abortion and sterility. Live, attenuated vaccines such as S19 and RB51 have been used to control the spread of the disease in animals; however, they are considered unsafe for human use and they induce abortion in pregnant cattle. For the development of a safer and equally efficacious vaccine, immunoproteomics was utilized to identify novel candidate proteins from B. abortus cell envelope (CE). A total of 163 proteins were identified using 2-DE with MALDI-TOF MS and LC-MS/MS. Some of the major protein components include outer-membrane protein (OMP) 25, OMP31, Omp2b porin, and 60 kDa chaperonin GroEL. 2-DE Western blot analyses probed with antiserum from bovine and a human patient infected with Brucella identified several new immunogenic proteins such as fumarate reductase flavoprotein subunit, F0F1-type ATP synthase alpha subunit, and cysteine synthase A. The elucidation of the immunome of B. abortus CE identified a number of candidate proteins for developing vaccines against Brucella infection in bovine and humans.

  12. Characterization and Protective Property of Brucella abortus cydC and looP Mutants

    PubMed Central

    Truong, Quang Lam; Cho, Youngjae; Barate, Abhijit Kashinath; Kim, Suk

    2014-01-01

    Brucella abortus readily multiplies in professional or nonprofessional phagocytes in vitro and is highly virulent in mice. Isogenic mutants of B. abortus biovar 1 strain IVKB9007 lacking the ATP/GDP-binding protein motif A (P-loop) (named looP; designated here the IVKB9007 looP::Tn5 mutant) and the ATP-binding/permease protein (cydC; designated here the IVKB9007 cydC::Tn5 mutant) were identified and characterized by transposon mutagenesis using the mini-Tn5Km2 transposon. Both mutants were found to be virtually incapable of intracellular replication in both murine macrophages (RAW264.7) and the HeLa cell line, and their virulence was significantly impaired in BALB/c mice. Respective complementation of the IVKB9007 looP::Tn5 and IVKB9007 cydC::Tn5 mutants restored their ability to survive in vitro and in vivo to a level comparable with that of the wild type. These findings indicate that the cydC and looP genes play important roles in the virulence of B. abortus. In addition, intraperitoneal immunization of mice with a dose of the live IVKB9007 looP::Tn5 and IVKB9007 cydC::Tn5 mutants provided a high degree of protection against challenge with pathogenic B. abortus strain 544. Both mutants should be evaluated further as a live attenuated vaccine against bovine brucellosis for their ability to stimulate a protective immune response. PMID:25253663

  13. Endometritis caused by Chlamydia trachomatis.

    PubMed Central

    Mårdh, P A; Møller, B R; Ingerselv, H J; Nüssler, E; Weström, L; Wølner-Hanssen, P

    1981-01-01

    Chlamydia trachomatis was found to be the aetiological agent of endometritis in three women with concomitant signs of salpingitis. All patients developed a significant antibody response to the organism. Chlamydia were recovered from aspirated uterine contents of two patients and darkfield examination of histological sections showed chlamydial inclusions in endometrial cells in one patient. Thus, C trachomatis can be recovered from the endometrium of patients in whom the cervical culture result is negative. In one patient curettage showed endometritis with a characteristic plasma-cell infiltration. The occurrence of chlamydial endometritis may explain why irregular bleeding is a common finding in patients with salpingitis. It also suggests a canalicular spread of chlamydia from the cervix to the Fallopian tubes. Images PMID:7237083

  14. Studies on recombinant glucokinase (r-glk) protein of Brucella abortus as a candidate vaccine molecule for brucellosis.

    PubMed

    Vrushabhendrappa; Singh, Amit Kumar; Balakrishna, Konduru; Sripathy, Murali Harishchandra; Batra, Harsh Vardhan

    2014-09-29

    Brucellosis is one of the most prevalent zoonotic diseases of worldwide distribution caused by the infection of genus Brucella. Live attenuated vaccines such as B. abortus S19, B. abortus RB51 and B. melitensis Rev1 are found most effective against brucellosis infection in animals, contriving a number of serious side effects and having chances to revert back into their active pathogenic form. In order to engineer a safe and effective vaccine candidate to be used in both animals and human, a recombinant subunit vaccine molecule comprising the truncated region of glucokinase (r-glk) gene from B. abortus S19 was cloned and expressed in Escherichia coli BL21DE3 host. Female BALB/c mice immunized with purified recombinant protein developed specific antibody titer of 1:64,000. The predominant IgG2a and IgG2b isotypes signified development of Th1 directed immune responses. In vitro cell cytotoxicity assay using anti-r-glk antibodies incubated with HeLa cells showed 81.20% and 78.5% cell viability against lethal challenge of B. abortus 544 and B. melitensis 16M, respectively. The lymphocyte proliferative assay indicated a higher splenic lymphocyte responses at 25μg/ml concentration of protein which implies the elevated development of memory immune responses. In contrast to control, the immunized group of mice intra-peritoneal (I.P.) challenged with B. abortus 544 were significantly protected with no signs of necrosis and vacuolization in their liver and spleen tissue. The elevated B-cell response associated with Th1 adopted immunity, significant in vitro cell viability as well as protection afforded in experimental animals after challenge, supplemented with histopathological analysis are suggestive of r-glk protein as a prospective candidate vaccine molecule against brucellosis.

  15. Infection of C57BL/6 mice by Trypanosoma musculi modulates host immune responses during Brucella abortus cocolonization.

    PubMed

    Lowry, Jake E; Leonhardt, Jack A; Yao, Chaoqun; Belden, E Lee; Andrews, Gerard P

    2014-01-01

    Brucellosis, which results in fetal abortions in domestic and wildlife animal populations, is of major concern in the US and throughout much of the world. The disease, caused by Brucella abortus, poses an economic threat to agriculture-based communities. A moderately efficacious live attenuated vaccine (B. abortus strain RB51) exists. However, even with vaccine use, outbreaks occur. Evidence suggests that elk (Cervus canadensis), a wild host reservoir, are the source of recent outbreaks in domestic cattle herds in Wyoming, USA. Brucella abortus establishes a chronic, persistent infection in elk. The molecular mechanisms allowing the establishment of this persistent infective state are currently unknown. A potential mechanism could be that concurrent pathogen burdens contribute to persistence. In Wyoming, elk are chronically infected with Trypanosoma cervi, which may modulate host responses in a similar manner to that documented for other trypanosomes. To identify any synergistic relationship between the two pathogens, we simulated coinfection in the well-established murine brucellosis model using Trypanosoma musculi and B. abortus S19. Groups of C57BL/6 mice (Mus musculus) were infected with either B. abortus strain 19 (S19) or T. musculi or both. Sera were collected weekly; spleens from euthanized mice were tested to determine bacterial load near the end of normal brucellosis infection. Although changes in bacterial load were observed during the later stages of brucellosis in those mice coinfected with T. musculi, the most significant finding was the suppression of gamma interferon early during the infection along with an increase in interleukin-10 secretion compared with mice infected with either pathogen alone. These results suggest that immune modulatory events occur in the mouse during coinfection and that further experiments are warranted to determine if T. cervi impacts Brucella infection in elk.

  16. Chlamydia pneumoniae and asthma

    PubMed Central

    Cook, P; Davies, P; Tunnicliffe, W; Ayres, J; Honeybourne, D; Wise, R

    1998-01-01

    BACKGROUND—This study was designed to test the association of Chlamydia pneumoniae infection with asthma in a multiracial population, after adjustments for several potential confounding variables.
METHODS—Antibodies to C pneumoniae were measured by microimmunofluorescence in 123 patients with acute asthma, 1518 control subjects admitted to the same hospital with various non-cardiovascular, non-pulmonary disorders, and 46 patients with severe chronic asthma, including some with "brittle" asthma. Acute infection or reinfection was defined by titres of IgG of ⩾512 or IgM ⩾8 or a fourfold rise in IgG, and previous infection by IgG 64-256 or IgA ⩾8. Logistic regression was used to control for likely confounders, including ethnic origin, age, sex, smoking habit, steroid medication, diabetes mellitus and social deprivation, on antibody levels.
RESULTS—Antibody titres consistent with acute C pneumoniae infection were found in 5.7% of patients with acute asthma and 5.7% of control patients, while 14.6% of patients with acute asthma and 12.7% of control patients had titres suggesting previous infection. These two groups did not differ significantly. However, titres suggesting previous infection were found in 34.8% of patients with severe chronic asthma: the difference between this group and the control group was statistically significant with an adjusted odds ratio of 3.99 (95% confidence interval 1.60 to 9.97).
CONCLUSIONS—These data raise important questions about the previously demonstrated association of C pneumoniae infection with asthma, and suggest that future studies of this association should give particular attention to the presence or absence of a history of severe chronic asthma.

 PMID:9741366

  17. WrpA Is an Atypical Flavodoxin Family Protein under Regulatory Control of the Brucella abortus General Stress Response System

    PubMed Central

    Herrou, Julien; Czyż, Daniel M.; Willett, Jonathan W.; Kim, Hye-Sook; Chhor, Gekleng; Babnigg, Gyorgy; Kim, Youngchang

    2016-01-01

    ABSTRACT The general stress response (GSR) system of the intracellular pathogen Brucella abortus controls the transcription of approximately 100 genes in response to a range of stress cues. The core genetic regulatory components of the GSR are required for B. abortus survival under nonoptimal growth conditions in vitro and for maintenance of chronic infection in an in vivo mouse model. The functions of the majority of the genes in the GSR transcriptional regulon remain undefined. bab1_1070 is among the most highly regulated genes in this regulon: its transcription is activated 20- to 30-fold by the GSR system under oxidative conditions in vitro. We have solved crystal structures of Bab1_1070 and demonstrate that it forms a homotetrameric complex that resembles those of WrbA-type NADH:quinone oxidoreductases, which are members of the flavodoxin protein family. However, B. abortus WrbA-related protein (WrpA) does not bind flavin cofactors with a high affinity and does not function as an NADH:quinone oxidoreductase in vitro. Soaking crystals with flavin mononucleotide (FMN) revealed a likely low-affinity binding site adjacent to the canonical WrbA flavin binding site. Deletion of wrpA (ΔwrpA) does not compromise cell survival under acute oxidative stress in vitro or attenuate infection in cell-based or mouse models. However, a ΔwrpA strain does elicit increased splenomegaly in a mouse model, suggesting that WrpA modulates B. abortus interaction with its mammalian host. Despite high structural homology with canonical WrbA proteins, we propose that B. abortus WrpA represents a functionally distinct member of the diverse flavodoxin family. IMPORTANCE Brucella abortus is an etiological agent of brucellosis, which is among the most common zoonotic diseases worldwide. The general stress response (GSR) regulatory system of B. abortus controls the transcription of approximately 100 genes and is required for maintenance of chronic infection in a murine model; the majority of

  18. A potent Brucella abortus 2308 Δery live vaccine allows for the differentiation between natural and vaccinated infection.

    PubMed

    Zhang, Junbo; Yin, Shuanghong; Guo, Fei; Meng, Ren; Chen, Chuangfu; Zhang, Hui; Li, Zhiqiang; Fu, Qiang; Shi, Huijun; Hu, Shengwei; Ni, Wei; Li, Tiansen; Zhang, Ke

    2014-08-01

    Brucellosis is a globally distributed zoonotic disease that causes animal and human diseases. However, the current Brucella abortus vaccines (S19 and RB51) are deficient; they can cause abortion in pregnant animals. Moreover, when the vaccine S19 is used, tests cannot differentiate natural from vaccinated infection. Therefore, a safer and more potent vaccine is needed. A Brucella abortus 2308 ery promoter mutant (Δery) was constructed to overcome these drawbacks. The growth of the Δery mutant was significantly attenuated in macrophages and mice and induced high protective immunity in mice. Moreover, Δery induced an anti-Brucella-specific IgG (immunoglobulin G) response and stimulated the expression of interferon-gamma (INF-γ) and interleukin-4 (IL-4). Furthermore, the expression of EryA antigen allowed for the serological differentiation between natural and vaccinated infection in mice. These results indicate that the Δery mutant is a potential attenuated live vaccine candidate against virulent Brucella abortus 2308 (S2308) infection.

  19. Overexpression of protective antigen as a novel approach to enhance vaccine efficacy of Brucella abortus strain RB51.

    PubMed

    Vemulapalli, R; He, Y; Cravero, S; Sriranganathan, N; Boyle, S M; Schurig, G G

    2000-06-01

    Brucella abortus strain RB51 is an attenuated rough strain that is currently being used as the official live vaccine for bovine brucellosis in the United States and several other countries. We reasoned that overexpression of a protective antigen(s) of B. abortus in strain RB51 should enhance its vaccine efficacy. To test this hypothesis, we overexpressed Cu/Zn superoxide dismutase (SOD) protein of B. abortus in strain RB51. This was accomplished by transforming strain RB51 with a broad-host-range plasmid, pBBR1MCS, containing the sodC gene along with its promoter. Strain RB51 overexpressing SOD (RB51SOD) was tested in BALB/c mice for its ability to protect against challenge infection with virulent strain 2308. Mice vaccinated with RB51SOD, but not RB51, developed antibodies and cell-mediated immune responses to Cu/Zn SOD. Strain RB51SOD vaccinated mice developed significantly (P < 0.05) more resistance to challenge than those vaccinated with strain RB51 alone. The presence of the plasmid alone in strain RB51 did not alter its vaccine efficacy. Also, overexpression of SOD did not alter the attenuation characteristic of strain RB51.

  20. SodA is a major metabolic antioxidant in Brucella abortus 2308 that plays a significant, but limited, role in the virulence of this strain in the mouse model.

    PubMed

    Martin, Daniel W; Baumgartner, John E; Gee, Jason M; Anderson, Eric S; Roop, R Martin

    2012-07-01

    The gene designated BAB1_0591 in the Brucella abortus 2308 genome sequence encodes the manganese-cofactored superoxide dismutase SodA. An isogenic sodA mutant derived from B. abortus 2308, designated JB12, displays a small colony phenotype, increased sensitivity in vitro to endogenous superoxide generators, hydrogen peroxide and exposure to acidic pH, and a lag in growth when cultured in rich and minimal media that can be rescued by the addition of all 20 amino acids to the growth medium. B. abortus JB12 exhibits significant attenuation in both cultured murine macrophages and experimentally infected mice, but this attenuation is limited to the early stages of infection. Addition of the NADPH oxidase inhibitor apocynin to infected macrophages does not alleviate the attenuation exhibited by JB12, suggesting that the basis for the attenuation of the B. abortus sodA mutant is not an increased sensitivity to exogenous superoxide generated through the oxidative burst of host phagocytes. It is possible, however, that the increased sensitivity of the B. abortus sodA mutant to acid makes it less resistant than the parental strain to killing by the low pH encountered during the early stages of the development of the brucella-containing vacuoles in macrophages. These experimental findings support the proposed role for SodA as a major cytoplasmic antioxidant in brucella. Although this enzyme provides a clear benefit to B. abortus 2308 during the early stages of infection in macrophages and mice, SodA appears to be dispensable once the brucellae have established an infection.

  1. Coombs antiglobulin test using Brucella abortus 99 as antigen to detect incomplete antibodies induced by B abortus RB51 vaccine in cattle.

    PubMed

    Ciuchini, Franco; Adone, Rosanna; Pasquali, Paolo

    2002-11-01

    This study showed that vaccination of cattle with Brucella abortus rough strain RB51 induces incomplete antibodies that can be detectable by a Coombs antiglobulin test using the B. abortus 99 smooth strain.

  2. Genetic characterization of a Tn5-disrupted glycosyltransferase gene homolog in Brucella abortus and its effect on lipopolysaccharide composition and virulence.

    PubMed

    McQuiston, J R; Vemulapalli, R; Inzana, T J; Schurig, G G; Sriranganathan, N; Fritzinger, D; Hadfield, T L; Warren, R A; Lindler, L E; Snellings, N; Hoover, D; Halling, S M; Boyle, S M

    1999-08-01

    We constructed a rough mutant of Brucella abortus 2308 by transposon (Tn5) mutagenesis. Neither whole cells nor extracted lipopolysaccharide (LPS) from this mutant, designated RA1, reacted with a Brucella O-side-chain-specific monoclonal antibody (MAb), Bru-38, indicating the absence of O-side-chain synthesis. Compositional analyses of LPS from strain RA1 showed reduced levels of quinovosamine and mannose relative to the levels in the parental, wild-type strain, 2308. We isolated DNA flanking the Tn5 insertion in strain RA1 by cloning a 25-kb XbaI genomic fragment into pGEM-3Z to create plasmid pJM6. Allelic exchange of genomic DNA in B. abortus 2308 mediated by electroporation of pJM6 produced kanamycin-resistant clones that were not reactive with MAb Bru-38. Southern blot analysis of genomic DNA from these rough clones revealed Tn5 in a 25-kb XbaI genomic fragment. A homology search with the deduced amino acid sequence of the open reading frame disrupted by Tn5 revealed limited homology with various glycosyltransferases. This B. abortus gene has been named wboA. Transformation of strain RA1 with a broad-host-range plasmid bearing the wild-type B. abortus wboA gene resulted in the restoration of O-side-chain synthesis and the smooth phenotype. B. abortus RA1 was attenuated for survival in mice. However, strain RA1 persisted in mice spleens for a longer time than the B. abortus vaccine strain RB51, but as expected, neither strain induced antibodies specific for the O side chain.

  3. Bactericidal Effect of Silver Nanoparticles on Intramacrophage Brucella abortus 544

    PubMed Central

    Alizadeh, Hamed; Salouti, Mojtaba; Shapouri, Reza

    2014-01-01

    Background: Brucellosis is an infectious disease that is caused by Brucella spp. As Brucella spp. are intramacrophage pathogens, the treatment of this infection is very difficult. On the other hand, due to the side effects of the brucellosis treatment regime, it is necessary to find new antimicrobial agents against it. Objectives: The aim of this study was to investigate the antimicrobial effect of silver nanoparticles against Brucella abortus 544 in the intramacrophage condition. Materials and Methods: The antimicrobial effect of silver nanoparticles was determined by an agar well diffusion method. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of silver nanoparticles against B. abortus 544 were determined by a broth macrodilution method. The effect of time on the antimicrobial activity of silver nanoparticles was analyzed. The effect of silver nanoparticles on the intramacrophage survival of B. abortus 544 was studied on mice peritoneal macrophages. Results: The well diffusion agar study showed that silver nanoparticles have an antimicrobial effect on B. abortus 544. The MIC and MBC of silver nanoparticles against B. abortus 544 were; 6 ppm and 8 ppm, respectively. The silver nanoparticles showed antibacterial effects within 40 minutes. The results of the macrophage culture indicated that silver nanoparticles have antibacterial activity against intramacrophage B. abortus 544, and the highest efficiency was observed at a concentration of 8-10 ppm of silver nanoparticles. Conclusions: The results showed that silver nanoparticles have an antimicrobial effect against intramacrophage B. abortus 544. PMID:25147682

  4. Protective effects of recombinant Brucella abortus Omp28 against infection with a virulent strain of Brucella abortus 544 in mice.

    PubMed

    Lim, Jeong Ju; Kim, Dong Hyeok; Lee, Jin Ju; Kim, Dae Geun; Min, Wongi; Lee, Hu Jang; Rhee, Man Hee; Kim, Suk

    2012-09-01

    The outer membrane proteins (OMPs) of Brucella (B.) abortus have been extensively studied, but their immunogenicity and protective ability against B. abortus infection are still unclear. In the present study, B. abortus Omp28, a group 3 antigen, was amplified by PCR and cloned into a maltose fusion protein expression system. Recombinant Omp28 (rOmp28) was expressed in Escherichia coli and was then purified. Immunogenicity of rOmp28 was confirmed by Western blot analysis with Brucella-positive mouse serum. Furthermore, humoral- or cell-mediated immune responses measured by the production of IgG1 or IgG2a in rOmp28-immunized mice and the ability of rOmp28 immunization to protect against B. abortus infection were evaluated in a mouse model. In the immunogenicity analysis, the mean titers of IgG1 and IgG2a produced by rOmp28-immunized mice were 20-fold higher than those of PBS-treated mice throughout the entire experimental period. Furthermore, spleen proliferation and bacterial burden in the spleen of rOmp28-immunized mice were approximately 1.5-fold lower than those of PBS-treated mice when challenged with virulent B. abortus. These findings suggest that rOmp28 from B. abortus is a good candidate for manufacturing an effective subunit vaccine against B. abortus infection in animals.

  5. Protective effects of recombinant Brucella abortus Omp28 against infection with a virulent strain of Brucella abortus 544 in mice

    PubMed Central

    Lim, Jeong Ju; Kim, Dong Hyeok; Lee, Jin Ju; Kim, Dae Geun; Min, Wongi; Lee, Hu Jang; Rhee, Man Hee

    2012-01-01

    The outer membrane proteins (OMPs) of Brucella (B.) abortus have been extensively studied, but their immunogenicity and protective ability against B. abortus infection are still unclear. In the present study, B. abortus Omp28, a group 3 antigen, was amplified by PCR and cloned into a maltose fusion protein expression system. Recombinant Omp28 (rOmp28) was expressed in Escherichia coli and was then purified. Immunogenicity of rOmp28 was confirmed by Western blot analysis with Brucella-positive mouse serum. Furthermore, humoral- or cell-mediated immune responses measured by the production of IgG1 or IgG2a in rOmp28-immunized mice and the ability of rOmp28 immunization to protect against B. abortus infection were evaluated in a mouse model. In the immunogenicity analysis, the mean titers of IgG1 and IgG2a produced by rOmp28-immunized mice were 20-fold higher than those of PBS-treated mice throughout the entire experimental period. Furthermore, spleen proliferation and bacterial burden in the spleen of rOmp28-immunized mice were approximately 1.5-fold lower than those of PBS-treated mice when challenged with virulent B. abortus. These findings suggest that rOmp28 from B. abortus is a good candidate for manufacturing an effective subunit vaccine against B. abortus infection in animals. PMID:23000585

  6. Chlamydiae and Mycoplasma infections in pulmonary MALT lymphoma

    PubMed Central

    Chanudet, E; Adam, P; Nicholson, A G; Wotherspoon, A C; Ranaldi, R; Goteri, G; Pileri, S A; Ye, H; Müller-Hermelink, H K; Du, M-Q

    2007-01-01

    Chlamydia pneumoniae, Chlamydia trachomatis and Chlamydia psittaci were detected at low frequencies (<20%) among 69 pulmonary mucosa-associated lymphoid tissue (MALT) lymphomas, 30 other lymphoproliferative disorders (LPD) and 44 non-LPD. The incidence of individual Chlamydiae was generally higher in MALT lymphoma than non-LPD, although not reaching statistical significance. Mycoplasma pneumoniae DNA was not detected. PMID:17876330

  7. Enhanced efficacy of recombinant Brucella abortus RB51 vaccines against B. melitensis infection in mice.

    PubMed

    Vemulapalli, Ramesh; Contreras, Andrea; Sanakkayala, Neelima; Sriranganathan, Nammalwar; Boyle, Stephen M; Schurig, Gerhardt G

    2004-09-01

    Brucella abortus strain RB51 is an attenuated rough strain, currently being used as the official live vaccine for bovine brucellosis in the USA and several other countries. In strain RB51, the wboA gene, encoding a glycosyltransferase required for the O-side chain synthesis, is disrupted by an IS711 element. Recently, we have demonstrated that strain RB51WboA, RB51 complemented with a functional wboA gene, remains rough but expresses low quantities of O-side chain in the cytoplasm. Mice vaccinated with strain RB51WboA develop greatly enhanced resistance against challenge with B. abortus virulent strain 2308. We have also demonstrated that overexpression of Cu/Zn superoxide dismutase (SOD) in strain RB51 (RB51SOD) significantly increases its vaccine efficacy against strain 2308 challenge. In this study, we constructed a new recombinant strain, RB51SOD/WboA, that over expresses SOD with simultaneous expression of O-side chain in the cytoplasm. We tested the vaccine potential of strains RB51SOD, RB51WboA, RB51SOD/WboA against challenge with virulent Brucella melitensis 16M and B. abortus 2308 in mice. In comparison with strain RB51, strain RB51SOD induced better protection against strain 2308, but not strain 16M, challenge. Similar to strain RB51WboA, vaccination with strain RB51SOD/WboA resulted in complete protection of the mice from infection with strain 2308. When challenged with strain 16M, mice vaccinated with either strain RB51WboA or strain RB51SOD/WboA were significantly better protected than those vaccinated with strain RB51 or RB51SOD. These results suggest that strains RB51WboA and RB51SOD/WboA are good vaccine candidates for inducing enhanced protection against B. melitensis infection.

  8. Advances in sampling and screening for chlamydia.

    PubMed

    Hocking, Jane S; Guy, Rebecca; Walker, Jennifer; Tabrizi, Sepehr N

    2013-03-01

    Chlamydia is the most commonly diagnosed bacterial sexually transmitted infection in the developed world, with diagnosis rates continuing to increase each year. As chlamydia is largely asymptomatic, screening and treatment is the main way to detect cases and reduce transmission. Recent advances in self-collected specimens and laboratory tests has made chlamydia screening easier to implement as well as possible in nonclinical settings. This review will discuss new approaches to specimen collection and how these have expanded opportunities for reaching target populations for chlamydia screening. Furthermore, it will discuss how advanced molecular microbiological methods can be used with self-collected specimens to further our knowledge of the epidemiology of chlamydia and the dynamics of transmission.

  9. The ferrous iron transporter FtrABCD is required for the virulence of Brucella abortus 2308 in mice.

    PubMed

    Elhassanny, Ahmed E M; Anderson, Eric S; Menscher, Evan A; Roop, R Martin

    2013-06-01

    Iron transport has been linked to the virulence of Brucella strains in both natural and experimental hosts. The genes designated BAB2_0837-0840 in the Brucella abortus 2308 genome sequence are predicted to encode a CupII-type ferrous iron transporter homologous to the FtrABCD transporter recently described in Bordetella. To study the role of the Brucella FtrABCD in iron transport, an isogenic ftrA mutant was constructed from B. abortus 2308. Compared with the parental strain, the B. abortus ftrA mutant displays a decreased capacity to use non-haem iron sources in vitro, a growth defect in a low iron medium that is enhanced at pH 6, and studies employing radiolabelled FeCl3 confirmed that FtrABCD transports ferrous iron. Transcription of the ftrA gene is induced in B. abortus 2308 in response to iron deprivation and exposure to acid pH, and similar to other Brucella iron acquisition genes that have been examined the iron-responsiveness of ftrA is dependent upon the iron response regulator Irr. The B. abortus ftrA mutant exhibits significant attenuation in both cultured murine macrophages and experimentally infected mice, supporting the proposition that ferrous iron is a critical iron source for these bacteria in the mammalian host.

  10. Immune response triggered by Brucella abortus following infection or vaccination.

    PubMed

    Dorneles, Elaine M S; Teixeira-Carvalho, Andréa; Araújo, Márcio S S; Sriranganathan, Nammalwar; Lage, Andrey P

    2015-07-17

    Brucella abortus live vaccines have been used successfully to control bovine brucellosis worldwide for decades. However, due to some limitations of these live vaccines, efforts are being made for the development of new safer and more effective vaccines that could also be used in other susceptible species. In this context, understanding the protective immune responses triggered by B. abortus is critical for the development of new vaccines. Such understandings will enhance our knowledge of the host/pathogen interactions and enable to develop methods to evaluate potential vaccines and innovative treatments for animals or humans. At present, almost all the knowledge regarding B. abortus specific immunological responses comes from studies in mice. Active participation of macrophages, dendritic cells, IFN-γ producing CD4(+) T-cells and cytotoxic CD8(+) T-cells are vital to overcome the infection. In this review, we discuss the characteristics of the immune responses triggered by vaccination versus infection by B. abortus, in different hosts.

  11. Antibody responses to Brucella abortus 2308 in cattle vaccinated with B. abortus RB51.

    PubMed

    Stevens, M G; Olsen, S C

    1996-03-01

    Cattle vaccinated with Brucella abortus rough strain RB51 (SRB51) produced small amounts of serum immunoglobulin G (IgG) but no IgM antibody to smooth strain 2308 (S2308) bacteria and produced no IgG or IgM antibody to S2308 lipopolysaccharide (LPS). Western immunoblot analysis revealed that antiserum from SRB51-vaccinated cattle contained IgG antibody that reacted with S2308 proteins of 84 to <20 kDa. However, antiserum from the vaccinated cattle did not contain agglutinating B. abortus antibody in the tube agglutination test for brucellosis. These results suggest that SRB51-vaccinated cattle produced no antibody to S2308 LPS, although they did produce nonagglutinating IgG antibody that reacted with S2308 bacteria and bacterial proteins of 84 to <20 kDa.

  12. Induction of antigen-specific Th1-type immune responses by gamma-irradiated recombinant Brucella abortus RB51.

    PubMed

    Sanakkayala, Neelima; Sokolovska, Anna; Gulani, Jatinder; Hogenesch, Harm; Sriranganathan, Nammalwar; Boyle, Stephen M; Schurig, Gerhardt G; Vemulapalli, Ramesh

    2005-12-01

    Brucella abortus strain RB51 is an attenuated rough mutant used as the live vaccine against bovine brucellosis in the United States and other countries. We previously reported the development of strain RB51 as a bacterial vaccine vector for inducing Th1-type immune responses against heterologous proteins. Because safety concerns may preclude the use of strain RB51-based recombinant live vaccines, we explored the ability of a gamma-irradiated recombinant RB51 strain to induce heterologous antigen-specific immune responses in BALB/c mice. Exposure of strain RB51G/LacZ expressing Escherichia coli beta-galactosidase to a minimum of 300 kilorads of gamma radiation resulted in complete loss of replicative ability. These bacteria, however, remained metabolically active and continued to synthesize beta-galactosidase. A single intraperitoneal inoculation of mice with 10(9) CFU equivalents of gamma-irradiated, but not heat-killed, RB51G/LacZ induced a beta-galactosidase-specific Th1-type immune response. Though no obvious differences were detected in immune responses to B. abortus-specific antigens, mice vaccinated with gamma-irradiated, but not heat-killed, RB51G/LacZ developed significant protection against challenge with virulent B. abortus. In vitro experiments indicated that gamma-irradiated and heat-killed RB51G/LacZ induced maturation of dendritic cells; however, stimulation with gamma-irradiated bacteria resulted in more interleukin-12 secretion. These results suggest that recombinant RB51 strains exposed to an appropriate minimum dose of gamma radiation are unable to replicate but retain their ability to stimulate Th1 immune responses against the heterologous antigens and confer protection against B. abortus challenge in mice.

  13. Genital Chlamydia trachomatis: An update

    PubMed Central

    Malhotra, Meenakshi; Sood, Seema; Mukherjee, Anjan; Muralidhar, Sumathi; Bala, Manju

    2013-01-01

    Chlamydia trachomatis is the most common cause of curable bacterial sexually transmitted infection (STI) worldwide. It manifests primarily as urethritis in males and endocervicitis in females. Untreated chlamydial infection in man can cause epididymitis and proctitis. Though most women with Chlamydia infection are asymptomatic or have minimal symptoms, some develop salpingitis, endometritis, pelvic inflammatory disease (PID), ectopic pregnancy and tubal factor infertility. It is associated with an increased risk for the transmission or acquisition of HIV and is also attributed to be a risk factor for the development of cervical carcinoma. Early diagnosis and treatment of infected individuals is required to prevent the spread of the disease and severe sequelae. Traditionally, tissue culture was considered the gold standard for the diagnosis. However, with the availability of newer diagnostic techniques particularly molecular methods which are not only highly sensitive and specific but are cost-effective also, the diagnosis has became fast and easy. The purpose of this review is to study the various aspects of genital C. trachomatis infection. Also the advances related to the clinical picture, various diagnostic modalities, prevention, treatment, drug resistance and control measures will be dealt with. PMID:24135174

  14. Molecular Genetic Analysis of Chlamydia Species.

    PubMed

    Sixt, Barbara S; Valdivia, Raphael H

    2016-09-01

    Species of Chlamydia are the etiologic agent of endemic blinding trachoma, the leading cause of bacterial sexually transmitted diseases, significant respiratory pathogens, and a zoonotic threat. Their dependence on an intracellular growth niche and their peculiar developmental cycle are major challenges to elucidating their biology and virulence traits. The last decade has seen tremendous advances in our ability to perform a molecular genetic analysis of Chlamydia species. Major achievements include the generation of large collections of mutant strains, now available for forward- and reverse-genetic applications, and the introduction of a system for plasmid-based transformation enabling complementation of mutations; expression of foreign, modified, or reporter genes; and even targeted gene disruptions. This review summarizes the current status of the molecular genetic toolbox for Chlamydia species and highlights new insights into their biology and new challenges in the nascent field of Chlamydia genetics. PMID:27607551

  15. Diverse Genetic Regulon of the Virulence-Associated Transcriptional Regulator MucR in Brucella abortus 2308

    PubMed Central

    Caswell, Clayton C.; Elhassanny, Ahmed E. M.; Planchin, Emilie E.; Roux, Christelle M.; Weeks-Gorospe, Jenni N.; Ficht, Thomas A.; Dunman, Paul M.

    2013-01-01

    The Ros-type regulator MucR is one of the few transcriptional regulators that have been linked to virulence in Brucella. Here, we show that a Brucella abortus in-frame mucR deletion strain exhibits a pronounced growth defect during in vitro cultivation and, more importantly, that the mucR mutant is attenuated in cultured macrophages and in mice. The genetic basis for the attenuation of Brucella mucR mutants has not been defined previously, but in the present study the genes regulated by MucR in B. abortus have been elucidated using microarray analysis and real-time reverse transcription-PCR (RT-PCR). In B. abortus 2308, MucR regulates a wide variety of genes whose products may function in establishing and maintaining cell envelope integrity, polysaccharide biosynthesis, iron homeostasis, genome plasticity, and transcriptional regulation. Particularly notable among the MucR-regulated genes identified is arsR6 (nolR), which encodes a transcriptional regulator previously linked to virulence in Brucella melitensis 16 M. Importantly, electrophoretic mobility shift assays (EMSAs) determined that a recombinant MucR protein binds directly to the promoter regions of several genes repressed by MucR (including arsR6 [nolR]), and in Brucella, as in other alphaproteobacteria, MucR binds to its own promoter to repress expression of the gene that encodes it. Overall, these studies have uncovered the diverse genetic regulon of MucR in Brucella, and in doing so this work has begun to define the MucR-controlled genetic circuitry whose misregulation contributes to the virulence defect of Brucella mucR mutants. PMID:23319565

  16. Overexpression of Brucella putative glycosyltransferase WbkA in B. abortus RB51 leads to production of exopolysaccharide

    PubMed Central

    Dabral, Neha; Jain-Gupta, Neeta; Seleem, Mohamed N.; Sriranganathan, Nammalwar; Vemulapalli, Ramesh

    2015-01-01

    Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis in mammals. Brucella strains containing the O-polysaccharide in their cell wall structure exhibit a smooth phenotype whereas the strains devoid of the polysaccharide show rough phenotype. B. abortus strain RB51 is a stable rough attenuated mutant which is used as a licensed live vaccine for bovine brucellosis. Previous studies have shown that the wboA gene, which encodes a glycosyltransferase required for the synthesis of O-polysaccharide, is disrupted in B. abortus RB51 by an IS711 element. Although complementation of strain RB51 with a functional wboA gene results in O-polysaccharide synthesis in the cytoplasm, it does not result in smooth phenotype. The aim of this study was to determine if overexpression of Brucella WbkA or WbkE, two additional putative glycosyltransferases essential for O-polysaccharide synthesis, in strain RB51 would result in the O-polysaccharide synthesis and smooth phenotype. Our results demonstrate that overexpression of wbkA or wbkE gene in RB51 does not result in O-polysaccharide expression as shown by Western blotting with specific antibodies. However, wbkA, but not wbkE, overexpression leads to the development of a clumping phenotype and the production of exopolysaccharide(s) containing mannose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Moreover, we found that the clumping recombinant strain displays increased adhesion to polystyrene plates. The recombinant strain was similar to strain RB51 in its attenuation characteristic and in its ability to induce protective immunity against virulent B. abortus challenge in mice. PMID:26157707

  17. Overexpression of Brucella putative glycosyltransferase WbkA in B. abortus RB51 leads to production of exopolysaccharide.

    PubMed

    Dabral, Neha; Jain-Gupta, Neeta; Seleem, Mohamed N; Sriranganathan, Nammalwar; Vemulapalli, Ramesh

    2015-01-01

    Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis in mammals. Brucella strains containing the O-polysaccharide in their cell wall structure exhibit a smooth phenotype whereas the strains devoid of the polysaccharide show rough phenotype. B. abortus strain RB51 is a stable rough attenuated mutant which is used as a licensed live vaccine for bovine brucellosis. Previous studies have shown that the wboA gene, which encodes a glycosyltransferase required for the synthesis of O-polysaccharide, is disrupted in B. abortus RB51 by an IS711 element. Although complementation of strain RB51 with a functional wboA gene results in O-polysaccharide synthesis in the cytoplasm, it does not result in smooth phenotype. The aim of this study was to determine if overexpression of Brucella WbkA or WbkE, two additional putative glycosyltransferases essential for O-polysaccharide synthesis, in strain RB51 would result in the O-polysaccharide synthesis and smooth phenotype. Our results demonstrate that overexpression of wbkA or wbkE gene in RB51 does not result in O-polysaccharide expression as shown by Western blotting with specific antibodies. However, wbkA, but not wbkE, overexpression leads to the development of a clumping phenotype and the production of exopolysaccharide(s) containing mannose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Moreover, we found that the clumping recombinant strain displays increased adhesion to polystyrene plates. The recombinant strain was similar to strain RB51 in its attenuation characteristic and in its ability to induce protective immunity against virulent B. abortus challenge in mice.

  18. Coordinated Zinc Homeostasis Is Essential for the Wild-Type Virulence of Brucella abortus

    PubMed Central

    Sheehan, Lauren M.; Budnick, James A.; Roop, R. Martin

    2015-01-01

    ABSTRACT Metal homeostasis in bacterial cells is a highly regulated process requiring intricately coordinated import and export, as well as precise sensing of intracellular metal concentrations. The uptake of zinc (Zn) has been linked to the virulence of Brucella abortus; however, the capacity of Brucella strains to sense Zn levels and subsequently coordinate Zn homeostasis has not been described. Here, we show that expression of the genes encoding the zinc uptake system ZnuABC is negatively regulated by the Zn-sensing Fur family transcriptional regulator, Zur, by direct interactions between Zur and the promoter region of znuABC. Moreover, the MerR-type regulator, ZntR, controls the expression of the gene encoding the Zn exporter ZntA by binding directly to its promoter. Deletion of zur or zntR alone did not result in increased zinc toxicity in the corresponding mutants; however, deletion of zntA led to increased sensitivity to Zn but not to other metals, such as Cu and Ni, suggesting that ZntA is a Zn-specific exporter. Strikingly, deletion of zntR resulted in significant attenuation of B. abortus in a mouse model of chronic infection, and subsequent experiments revealed that overexpression of zntA in the zntR mutant is the molecular basis for its decreased virulence. IMPORTANCE The importance of zinc uptake for Brucella pathogenesis has been demonstrated previously, but to date, there has been no description of how overall zinc homeostasis is maintained and genetically controlled in the brucellae. The present work defines the predominant zinc export system, as well as the key genetic regulators of both zinc uptake and export in Brucella abortus. Moreover, the data show the importance of precise coordination of the zinc homeostasis systems as disregulation of some elements of these systems leads to the attenuation of Brucella virulence in a mouse model. Overall, this study advances our understanding of the essential role of zinc in the pathogenesis of

  19. Prevalence of Chlamydia psittaci and Other Chlamydia Species in Wild Birds in Poland

    PubMed Central

    Krawiec, Marta; Piasecki, Tomasz

    2015-01-01

    Abstract Avian chlamydiosis is a zoonotic disease occurring in humans, poultry, and exotic birds. It has been suggested that some wild bird species play an important role as reservoirs for Chlamydia, especially Chlamydia psittaci. Whereas C. psittaci is the predominant chlamydial agent in birds, in the present study we have determined the prevalence of different species of Chlamydia among selected wild bird species in Poland using a rapid and sensitive real-time PCR method. In total, 369 free-living birds from 35 bird species and 15 orders were examined. Samples from 27 birds (7.3%) were positive for chlamydial DNA in the PCR; 22 positive samples (81.5%) belonged to C. psittaci, three to Chlamydia trachomatis (11.1%), and two (7.4%) classified only to the genus Chlamydia. Most of C. psittaci–positive samples belonged to five orders: Anseriformes, Columbiformes, Gruiformes, Phasianiformes, and Passeriformes. All C. trachomatis samples were obtained from Eurasian coots (Gruiformes). Two Chlamydia-positive samples not classified to any Chlamydia species were obtained from a common wood pigeon (Columbiformes) and a common buzzard (Accipitriformes). Detection of C. psittaci and C. trachomatis in free-living bird populations force to think on significance of birds as reservoir of varied Chlamydia species and their epidemiological importance. PMID:26501593

  20. Prevalence of Chlamydia psittaci and Other Chlamydia Species in Wild Birds in Poland.

    PubMed

    Krawiec, Marta; Piasecki, Tomasz; Wieliczko, Alina

    2015-11-01

    Avian chlamydiosis is a zoonotic disease occurring in humans, poultry, and exotic birds. It has been suggested that some wild bird species play an important role as reservoirs for Chlamydia, especially Chlamydia psittaci. Whereas C. psittaci is the predominant chlamydial agent in birds, in the present study we have determined the prevalence of different species of Chlamydia among selected wild bird species in Poland using a rapid and sensitive real-time PCR method. In total, 369 free-living birds from 35 bird species and 15 orders were examined. Samples from 27 birds (7.3%) were positive for chlamydial DNA in the PCR; 22 positive samples (81.5%) belonged to C. psittaci, three to Chlamydia trachomatis (11.1%), and two (7.4%) classified only to the genus Chlamydia. Most of C. psittaci-positive samples belonged to five orders: Anseriformes, Columbiformes, Gruiformes, Phasianiformes, and Passeriformes. All C. trachomatis samples were obtained from Eurasian coots (Gruiformes). Two Chlamydia-positive samples not classified to any Chlamydia species were obtained from a common wood pigeon (Columbiformes) and a common buzzard (Accipitriformes). Detection of C. psittaci and C. trachomatis in free-living bird populations force to think on significance of birds as reservoir of varied Chlamydia species and their epidemiological importance.

  1. Atherosclerosis, inflammation and Chlamydia pneumoniae

    PubMed Central

    Fazio, Giovanni; Giovino, Maria; Gullotti, Alessandro; Bacarella, Daniela; Novo, Giuseppina; Novo, Salvatore

    2009-01-01

    Coronary heart disease is the single most common cause of illness and death in the developed world. Coronary atherosclerosis is by far the most frequent cause of ischemic heart disease, and plaque disruption with superimposed thrombosis is the main cause of the acute coronary syndromes of unstable angina, myocardial infarction, and sudden death. Atherosclerosis is the result of a complex interaction between blood elements, disturbed flow, and vessel wall abnormality, involving several pathological processes: inflammation, with increased endothelial permeability, endothelial activation, and monocyte recruitment; growth, with smooth muscle cell proliferation, migration, and matrix synthesis; degeneration, with lipid accumulation; necrosis, possibly related to the cytotoxic effect of oxidized lipid; calcification/ossification, which may represent an active rather than a dystrophic process; and thrombosis, with platelet recruitment and fibrin formation. In this review we discuss these processes and the possible pathological effects of Chlamydia infection and the ensuing phlogosis. PMID:21160574

  2. Chlamydia pneumoniae and cardiovascular disease.

    PubMed Central

    Campbell, L. A.; Kuo, C. C.; Grayston, J. T.

    1998-01-01

    Chlamydia pneumoniae is a ubiquitous pathogen that causes acute respiratory disease. The spectrum of C. pneumoniae infection has been extended to atherosclerosis and its clinical manifestations. Seroepidemiologic studies have associated C. pneumoniae antibody with coronary artery disease, myocardial infarction, carotid artery disease, and cerebrovascular disease. The association of C. pneumoniae with atherosclerosis is corroborated by the presence of the organism in atherosclerotic lesions throughout the arterial tree and the near absence of the organism in healthy arterial tissue. C. pneumoniae has also been isolated from coronary and carotid atheromatous plaques. To determine whether chronic infection plays a role in initiation or progression of disease, intervention studies in humans have been initiated, and animal models of C. pneumoniae infection have been developed. This review summarizes the evidence for the association and potential role of C. pneumoniae in cardiovascular disease. PMID:9866733

  3. The placenta findings from an XYY abortus: a case report

    PubMed Central

    2013-01-01

    Introduction The placenta morphology from an XYY pregnancy abortus has not been reported in the medical literature. This case report consists of the first detailed documentation. The reported case is also highly unusual because the mother had two prior pregnancies with fetuses being confirmed to have Zellweger syndrome and one prior molar pregnancy. Case presentation A 43-year-old Caucasian woman presented for induction of labor secondary to diagnosis of XYY chromosomes by chorionic villus sample. Conclusions This is the first detailed observation of placenta morphology in an XYY abortus. Although not highly specific, the observation is very unique and should prompt further investigation of karyotyping of the fetus or infant because an XYY individual may be viable and grow to adulthood. The association of an XYY abortus and prior pregnancies with Zellweger syndrome and one prior molar pregnancy is also highly notable. PMID:24083480

  4. Characterization and Immunogenicity of Outer Membrane Vesicles from Brucella abortus.

    PubMed

    Kaur, Gagandeep; Singh, Satparkash; Sunil Kumar, B V; Mahajan, Kanika; Verma, Ramneek

    2016-01-01

    Bovine brucellosis is a worldwide spread zoonotic disease. The objectives of this study were characterization of outer membrane vesicles from B. abortus and to evaluate their immunogenicity in mice. For this purpose, OMVs were derived from B. abortus strain 99 using ultracentrifugation method. Isolated OMVs were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis and transmission electron microscopy which revealed spherical 20-300 nm structures rich in proteins. OMVs also showed immuno-reactivity with mice antisera in Western blot. Further, indirect ELISA showed specific and high-titer immune responses against the antigens present in OMVs suggesting their potential for a safe acellular vaccine candidate.

  5. Chlamydia

    MedlinePlus

    ... include: Bleeding between periods Burning when urinating Fever Low back pain Lower abdominal pain Nausea Pain during sex Unusual ... include: Bleeding between periods Burning when urinating Fever Low back pain Lower abdominal pain Nausea Pain during sex Unusual ...

  6. Chlamydia

    MedlinePlus

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  7. Efficacy of Brucella abortus vaccine strain RB51 compared to the reference vaccine Brucella abortus strain 19 in water buffalo.

    PubMed

    Caporale, Vincenzo; Bonfini, Barbara; Di Giannatale, Elisabetta; Di Provvido, Andrea; Forcella, Simona; Giovannini, Armando; Tittarelli, Manuela; Scacchia, Massimo

    2010-01-01

    Approximately 250,000 water buffalo (Bubalus bubalis) live in the Campania region of southern Italy where the breeding of this species is very popular. Of these animals, almost 150,000 are concentrated in the Caserta province where the prevalence of Brucella abortus in this species represents approximately 20% at herd level. The Italian brucellosis eradication programme provides a slaughter and vaccination strategy for this province. B. abortus strain RB51 (RB51) has become the official vaccine for the prevention of brucellosis in cattle in several countries. The aim of this study was to evaluate the efficacy of RB51 in water buffalo compared to the B. abortus S19 vaccine (S19). The study was performed in accordance with a protocol described in mice. Female buffalo aged five months were inoculated. Five received a RB51 dosage on two occasions that was three times greater than that approved for use in cattle and a booster after one month, five received B. abortus S19 vaccine at the standard dosage and three controls received a phosphate buffer solution. Buffalo were then challenged with a virulent B. abortus strain 544 thirty days post vaccination. Antibodies that developed in the five animals vaccinated with RB51 were not detected by the Rose Bengal test or complement fixation test (CFT) and were also tested by CFT prepared with RB51 antigen. After culling, B. abortus was cultured from the spleen, retropharyngeal and supra-mammary lymph nodes. A statistical evaluation was performed to assess the immunogenicity values obtained in buffalo vaccinated with S19, compared to those obtained in buffalo vaccinated with the RB51 vaccine and in the unvaccinated control group.

  8. Geomapping of chlamydia and gonorrhoea in Birmingham

    PubMed Central

    Shahmanesh, M.; Gayed, S.; Ashcroft, M.; Smith, R.; Roopnarainsingh, R.; Dunn, J.; Ross, J.

    2000-01-01

    Objective: To investigate if the core population hypothesis is applicable to patients with genital chlamydia infections. Design: Retrospective cross sectional study. Setting: Two genitourinary medicine (GUM) clinics in the city of Birmingham and eight adjacent clinics. Subjects: All patients with chlamydia (n = 665) or gonorrhoea (n = 584) attending between 1 October 1995 and 30 September 1996 with a postcode within the Birmingham health district. Controls were 727 patients seen in the same period with no infection. Methods: Postcodes were used to calculate population prevalence rates per 100 000 aged 15–65 in the 39 wards of the city and to estimate the socioeconomic status using the Super Profile (SP). Ethnic specific rates were also calculated. Data were obtained on gonorrhoea and chlamydia isolation from all the major laboratories of the city over the same time period. Results: GUM clinic attenders accounted for 67.6% and 82.5% of all chlamydia and gonorrhoea isolates reported by the laboratories and that were available for our epidemiological analysis. Both infections were more common in men and in black ethnic groups. However, patients with gonorrhoea only infection were more likely to be of black ethnicity than those with chlamydia only infection (p = 0.0001) and to have different SP distribution (p = 0.0001). On logistic regression age <20 years, male sex, black ethnicity, and living in neighbourhoods with SP J ("have nots") were predictive of both infections compared with controls. Overall chlamydia and gonorrhoea prevalence rates were 129 and 98.4 per 105 respectively. Corresponding rates for whites was 64.7 and 37.2 and for black ethnic groups 1105 and 1183 per 105 of each ethnic group. Eight adjacent wards accounted for 41% of the chlamydia and 66.5% of the gonorrhoea. Conclusion: In a large urban setting patients attending GUM clinics with chlamydia belong to core population groups with similar, but not identical, sociodemographic characteristics to

  9. Vaccination with live Escherichia coli expressing Brucella abortus Cu/Zn superoxide dismutase protects mice against virulent B. abortus.

    PubMed

    Oñate, A A; Vemulapalli, R; Andrews, E; Schurig, G G; Boyle, S; Folch, H

    1999-02-01

    Vaccination of mice with Escherichia coli expressing Brucella Cu/Zn superoxide dismutase (SOD) [E. coli(pBSSOD)] induced a significant level of protection against virulent Brucella abortus challenge, although this level was not as high as the one reached with B. abortus vaccine strain RB51. In addition, vaccination with E. coli(pBSSOD) induced antibodies to Cu/Zn SOD and a strong proliferative response of splenocytes when stimulated in vitro with a thioredoxin-Cu/Zn SOD fusion protein.

  10. Characterization of Brucella abortus O-polysaccharide and core lipopolysaccharide mutants and demonstration that a complete core is required for rough vaccines to be efficient against Brucella abortus and Brucella ovis in the mouse model.

    PubMed

    Monreal, D; Grilló, M J; González, D; Marín, C M; De Miguel, M J; López-Goñi, I; Blasco, J M; Cloeckaert, A; Moriyón, I

    2003-06-01

    Brucella abortus rough lipopolysaccharide (LPS) mutants were obtained by transposon insertion into two wbk genes (wbkA [putative glycosyltransferase; formerly rfbU] and per [perosamine synthetase]), into manB (pmm [phosphomannomutase; formerly rfbK]), and into an unassigned gene. Consistent with gene-predicted roles, electrophoretic analysis, 2-keto-3-manno-D-octulosonate measurements, and immunoblots with monoclonal antibodies to O-polysaccharide, outer and inner core epitopes showed no O-polysaccharide expression and no LPS core defects in the wbk mutants. The rough LPS of manB mutant lacked the outer core epitope and the gene was designated manB(core) to distinguish it from the wbk manB(O-Ag). The fourth gene (provisionally designated wa**) coded for a putative glycosyltransferase involved in inner core synthesis, but the mutant kept the outer core epitope. Differences in phage and polymyxin sensitivity, exposure or expression of outer membrane protein, core and lipid A epitopes, and lipid A acylation demonstrated that small changes in LPS core caused significant differences in B. abortus outer membrane topology. In mice, the mutants showed different degrees of attenuation and induced antibodies to rough LPS and outer membrane proteins. Core-defective mutants and strain RB51 were ineffective vaccines against B. abortus in mice. The mutants per and wbkA induced protection but less than the standard smooth vaccine S19, and controls suggested that anti O-polysaccharide antibodies accounted largely for the difference. Whereas no core-defective mutant was effective against B. ovis, S19, RB51, and the wbkA and per mutants afforded similar levels of protection. These results suggest that rough Brucella vaccines should carry a complete core for maximal effectiveness.

  11. Draft Genome Sequence of Brucella abortus Virulent Strain 544

    PubMed Central

    Singh, D. K.; Kumar, Ashok; Tiwari, A. K.; Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S.; Sridhar, Jayavel; Gunasekaran, Paramasamy

    2015-01-01

    Here, we present the draft genome sequence and annotation of Brucella abortus virulent strain 544. The genome of this strain is 3,289,405 bp long, with 57.2% G+C content. A total of 3,259 protein-coding genes and 60 RNA genes were predicted. PMID:25953161

  12. Draft Genome Sequence of Brucella abortus Virulent Strain 544.

    PubMed

    Singh, D K; Kumar, Ashok; Tiwari, A K; Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Sridhar, Jayavel; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2015-05-07

    Here, we present the draft genome sequence and annotation of Brucella abortus virulent strain 544. The genome of this strain is 3,289,405 bp long, with 57.2% G+C content. A total of 3,259 protein-coding genes and 60 RNA genes were predicted.

  13. Seroprevalence of Brucella abortus and Leptospira hardjo in cattle

    PubMed Central

    Pandian, S. Jegaveera; Ray, Pradeep Kumar; Chandran, P. C.; Kumar, Manoj

    2015-01-01

    Aim: The aim was to assess the seroprevalence of B. abortus and Leptospira hardjo in the cattle population of Bihar, this work was carried out. Materials and Methods: Randomly selected 450 cattle from nine districts of Bihar were serologically screened for antibodies against L. hardjo and B. abortus. DAS-ELISA for leptospira and AB-ELISA for brucella were carried out. Based on the results prevalence in each district and the state are reported herewith. Results and Discussion: In this study, it was found that the seroprevalence of L. hardjo was 9.11% and that of B. abortus was 12.2% in Bihar. Indigenous cattle were found to be less susceptible to leptospirosis and brucellosis even though they accounted for 83.11% of the study population. Conclusion: Although there was no acute disease, antibodies detected against L. hardjo and B. abortus in the cattle population indicated the presence of chronic and subclinical infection, which could challenge the fertility of the animals. PMID:27047076

  14. Brucella abortus RB51 in milk of vaccinated adult cattle.

    PubMed

    Miranda, Karina Leite; Poester, Fernando Padilla; Dorneles, Elaine Maria Seles; Resende, Thiago Magalhães; Vaz, Adil Knackfuss; Ferraz, Sandra Maria; Lage, Andrey Pereira

    2016-08-01

    The aim of this study was to evaluate the shedding of Brucella abortus in the milk of cows vaccinated with a full dose of RB51 during lactation. Eighteen cows, nine previously vaccinated with S19 as calves and nine non-vaccinated, were immunized subcutaneously with 1.3×10(10)CFU of B. abortus RB51, 30-60days after parturition. Milk samples from all animals were collected daily until day 7, and at weekly interval for the next 9 weeks after vaccination. To evaluate the shedding of B. abortus, milk samples were submitted for culture and PCR. No B. abortus was isolated from any sample tested. Only one sample, collected on first day after vaccination from a cow previously vaccinated, was faintly positive in the PCR. In conclusion, the public health hazard associated with milk consumption from cows vaccinated with RB51 in post-partum is very low, despite vaccination with the full dose and regardless of previous S19 vaccination.

  15. Bovine T-lymphocyte lines reactive with Brucella abortus.

    PubMed

    Smith, R; Kapatsa, J C; Rosenbaum, B A; Adams, L G

    1990-04-01

    Bovine T-cell lines reactive with Brucella abortus were established by repeated stimulation with B abortus and mitomycin C-treated autologous antigen-presenting cells. Representative results were obtained, using 33 cell lines from 14 cows. Cultures responded to the virulent laboratory strain 2308, the vaccine strain 19, and the rough mutant strain RB51 in thymidine-incorporation assays. The cells in these cultures required antigen-presenting cells for their response to B abortus. Autologous antigen-presenting cells were optimal for most lines tested, although some T-cell lines could respond to B abortus in the presence of some, but not all, allogeneic antigen-presenting cells. The cell lines expressed cell surface markers characteristics of activated bovine T cels. Of the cell lines tested for expression of cluster-determinant (CD) 4 and CD8 cell surface antigens, no cells in any cultures expressed the bovine CD8 equivalent, but all cultures included CD4+ cells in variable amounts. Some cell lines consisted of up to 50% CD2+CD4-CD8- cells. None of the cell lines tested expressed surface immunoglobulin or other bovine B-cell markers. Thus, these long-term cell lines appear to include 2 T-lymphocyte subsets: the helper/inducer subset and a second subset expressing a phenotype similar to major histocompatibility complex-unrestricted cytolytic cells in other species.

  16. Virulence of Brucella abortus isolated from cattle and water buffalo.

    PubMed

    Adesiyun, Abiodun A; Fosgate, Geoffrey T; Seebaransingh, Ravi; Brown, Gabriel; Stoute, Simone; Stewart-Johnson, Alva

    2011-01-01

    Brucellosis has been documented in domestic water buffalo (Bubalus bubalis) but published literature is limited despite the importance of this species in tropical agricultural systems. The objective of this study was to compare the virulence of Brucella abortus isolates recovered from cattle and water buffalo. Nineteen strains of B. abortus from cattle and domestic water buffalo in Trinidad were intraperitoneally inoculated into BALB/c mice. Spleens were cultured for B. abortus and histopathological severity scores were calculated based on lymphoid depletion, lymphoid necrosis, splenitis, and macrophage accumulation. A general linear model approach was used to estimate the effect of isolate source (cattle versus water buffalo) on virulence. Isolates of water buffalo origin were significantly less virulent in the mouse model based on recovered B. abortus from splenic tissues, spleen/weight ratio, and lymphoid necrosis but not overall histopathological severity scores. Further investigation of isolates recovered from water buffalo might provide the key to the development of procedures for brucellosis control in tropical environments.

  17. Identification of an IS711 element interrupting the wboA gene of Brucella abortus vaccine strain RB51 and a PCR assay to distinguish strain RB51 from other Brucella species and strains.

    PubMed

    Vemulapalli, R; McQuiston, J R; Schurig, G G; Sriranganathan, N; Halling, S M; Boyle, S M

    1999-09-01

    Brucella abortus vaccine strain RB51 is a natural stable attenuated rough mutant derived from the virulent strain 2308. The genetic mutations that are responsible for the roughness and the attenuation of strain RB51 have not been identified until now. Also, except for an assay based on pulsed-field gel electrophoresis, no other simple method to differentiate strain RB51 from its parent strain 2308 is available. In the present study, we demonstrate that the wboA gene encoding a glycosyltransferase, an enzyme essential for the synthesis of O antigen, is disrupted by an IS711 element in B. abortus vaccine strain RB51. Exploiting this feature, we developed a PCR assay that distinguishes strain RB51 from all other Brucella species and strains tested.

  18. Chlamydia Peritonitis and Ascites Mimicking Ovarian Cancer

    PubMed Central

    Macer, Matthew; Azodi, Masoud

    2016-01-01

    Background. Pelvic inflammatory disease (PID) rarely results in diffuse ascites. Severe adhesive disease secondary to PID may lead to the formation of inclusion cysts and even pelvic peritoneal nodularity due to postinflammatory scarring and cause an elevation of serum CA-125 levels. The constellation of these findings may mimic an ovarian neoplasm. Case. We report a case of a 22-year-old female who presented with multiple pelvic cysts and diffuse ascites due to Chlamydia trachomatis infection. The initial gynecologic exam did not reveal obvious evidence of PID; however, a positive Chlamydia trachomatis test, pathologic findings, and the exclusion of other etiologies facilitated the diagnosis. Conclusion. Chlamydia trachomatis and other infectious agents should be considered in the differential diagnosis of a young sexually active female with abdominal pain, ascites, and pelvic cystic masses. Thorough workup in such a population may reduce the number of more invasive procedures as well as unnecessary repeat surgical procedures. PMID:27747116

  19. Immunization of Mice with Recombinant Protein CobB or AsnC Confers Protection against Brucella abortus Infection

    PubMed Central

    Du, Xinying; Yu, Shuang; Bai, Yaoxia; Chen, Yanfen; Wang, Tongkun; Wang, Zhoujia; Yu, Yaqing; Peng, Guangneng; Huang, Kehe; Huang, Liuyu; Wang, Yufei; Chen, Zeliang

    2012-01-01

    Due to drawbacks of live attenuated vaccines, much more attention has been focused on screening of Brucella protective antigens as subunit vaccine candidates. Brucella is a facultative intracellular bacterium and cell mediated immunity plays essential roles for protection against Brucella infection. Identification of Brucella antigens that present T-cell epitopes to the host could enable development of such vaccines. In this study, 45 proven or putative pathogenesis-associated factors of Brucella were selected according to currently available data. After expressed and purified, 35 proteins were qualified for analysis of their abilities to stimulate T-cell responses in vitro. Then, an in vitro gamma interferon (IFN-γ) assay was used to identify potential T-cell antigens from B. abortus. In total, 7 individual proteins that stimulated strong IFN-γ responses in splenocytes from mice immunized with B. abortus live vaccine S19 were identified. The protective efficiencies of these 7 recombinant proteins were further evaluated. Mice given BAB1_1316 (CobB) or BAB1_1688 (AsnC) plus adjuvant could provide protection against virulent B. abortus infection, similarly with the known protective antigen Cu-Zn SOD and the license vaccine S19. In addition, CobB and AsnC could induce strong antibodies responses in BALB/c mice. Altogether, the present study showed that CobB or AsnC protein could be useful antigen candidates for the development of subunit vaccines against brucellosis with adequate immunogenicity and protection efficacy. PMID:22383953

  20. The Lipopolysaccharide Core of Brucella abortus Acts as a Shield Against Innate Immunity Recognition

    PubMed Central

    Iriarte, Maite; Manček-Keber, Mateja; Barquero-Calvo, Elías; Palacios-Chaves, Leyre; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Martirosyan, Anna; von Bargen, Kristine; Grilló, María-Jesús; Jerala, Roman; Brandenburg, Klaus; Llobet, Enrique; Bengoechea, José A.; Moreno, Edgardo

    2012-01-01

    Innate immunity recognizes bacterial molecules bearing pathogen-associated molecular patterns to launch inflammatory responses leading to the activation of adaptive immunity. However, the lipopolysaccharide (LPS) of the gram-negative bacterium Brucella lacks a marked pathogen-associated molecular pattern, and it has been postulated that this delays the development of immunity, creating a gap that is critical for the bacterium to reach the intracellular replicative niche. We found that a B. abortus mutant in the wadC gene displayed a disrupted LPS core while keeping both the LPS O-polysaccharide and lipid A. In mice, the wadC mutant induced proinflammatory responses and was attenuated. In addition, it was sensitive to killing by non-immune serum and bactericidal peptides and did not multiply in dendritic cells being targeted to lysosomal compartments. In contrast to wild type B. abortus, the wadC mutant induced dendritic cell maturation and secretion of pro-inflammatory cytokines. All these properties were reproduced by the wadC mutant purified LPS in a TLR4-dependent manner. Moreover, the core-mutated LPS displayed an increased binding to MD-2, the TLR4 co-receptor leading to subsequent increase in intracellular signaling. Here we show that Brucella escapes recognition in early stages of infection by expressing a shield against recognition by innate immunity in its LPS core and identify a novel virulence mechanism in intracellular pathogenic gram-negative bacteria. These results also encourage for an improvement in the generation of novel bacterial vaccines. PMID:22589715

  1. Mutant Brucella abortus Membrane Fusogenic Protein Induces Protection against Challenge Infection in Mice

    PubMed Central

    de Souza Filho, Job Alves; Martins, Vicente de Paulo; Campos, Priscila Carneiro; Alves-Silva, Juliana; Santos, Nathalia V.; de Oliveira, Fernanda Souza; Menezes, Gustavo B.; Azevedo, Vasco; Cravero, Silvio Lorenzo

    2015-01-01

    Brucella species can cause brucellosis, a zoonotic disease that causes serious livestock economic losses and represents a public health threat. The mechanism of virulence of Brucella spp. is not yet fully understood. Therefore, it is crucial to identify new molecules that serve as virulence factors to better understand this host-pathogen interplay. Here, we evaluated the role of the Brucella membrane fusogenic protein (Mfp) and outer membrane protein 19 (Omp19) in bacterial pathogenesis. In this study, we showed that B. abortus Δmfp::kan and Δomp19::kan deletion mutant strains have reduced persistence in vivo in C57BL/6 and interferon regulatory factor 1 (IRF-1) knockout (KO) mice. Additionally, 24 h after macrophage infection with a Δmfp::kan or Δomp19::kan strain expressing green fluorescent protein (GFP) approximately 80% or 65% of Brucella-containing vacuoles (BCVs) retained the late endosomal/lysosomal marker LAMP-1, respectively, whereas around 60% of BCVs containing wild-type S2308 were found in LAMP-1-negative compartments. B. abortus Δomp19::kan was attenuated in vivo but had a residual virulence in C57BL/6 and IRF-1 KO mice, whereas the Δmfp::kan strain had a lower virulence in these same mouse models. Furthermore, Δmfp::kan and Δomp19::kan strains were used as live vaccines. Challenge experiments revealed that in C57BL/6 and IRF-1 KO mice, the Δmfp::kan strain induced greater protection than the vaccine RB51 and protection similar that of vaccine S19. However, a Δomp19::kan strain induced protection similar to that of RB51. Thus, these results demonstrate that Brucella Mfp and Omp19 are critical for full bacterial virulence and that the Δmfp::kan mutant may serve as a potential vaccine candidate in future studies. PMID:25644010

  2. Mutant Brucella abortus membrane fusogenic protein induces protection against challenge infection in mice.

    PubMed

    de Souza Filho, Job Alves; de Paulo Martins, Vicente; Campos, Priscila Carneiro; Alves-Silva, Juliana; Santos, Nathalia V; de Oliveira, Fernanda Souza; Menezes, Gustavo B; Azevedo, Vasco; Cravero, Silvio Lorenzo; Oliveira, Sergio Costa

    2015-04-01

    Brucella species can cause brucellosis, a zoonotic disease that causes serious livestock economic losses and represents a public health threat. The mechanism of virulence of Brucella spp. is not yet fully understood. Therefore, it is crucial to identify new molecules that serve as virulence factors to better understand this host-pathogen interplay. Here, we evaluated the role of the Brucella membrane fusogenic protein (Mfp) and outer membrane protein 19 (Omp19) in bacterial pathogenesis. In this study, we showed that B. abortus Δmfp::kan and Δomp19::kan deletion mutant strains have reduced persistence in vivo in C57BL/6 and interferon regulatory factor 1 (IRF-1) knockout (KO) mice. Additionally, 24 h after macrophage infection with a Δmfp::kan or Δomp19::kan strain expressing green fluorescent protein (GFP) approximately 80% or 65% of Brucella-containing vacuoles (BCVs) retained the late endosomal/lysosomal marker LAMP-1, respectively, whereas around 60% of BCVs containing wild-type S2308 were found in LAMP-1-negative compartments. B. abortus Δomp19::kan was attenuated in vivo but had a residual virulence in C57BL/6 and IRF-1 KO mice, whereas the Δmfp::kan strain had a lower virulence in these same mouse models. Furthermore, Δmfp::kan and Δomp19::kan strains were used as live vaccines. Challenge experiments revealed that in C57BL/6 and IRF-1 KO mice, the Δmfp::kan strain induced greater protection than the vaccine RB51 and protection similar that of vaccine S19. However, a Δomp19::kan strain induced protection similar to that of RB51. Thus, these results demonstrate that Brucella Mfp and Omp19 are critical for full bacterial virulence and that the Δmfp::kan mutant may serve as a potential vaccine candidate in future studies.

  3. Controllable attenuators

    NASA Astrophysics Data System (ADS)

    Krylov, G. M.; Khoniak, E. I.; Tynynyka, A. N.; Iliushenko, V. N.; Sikolenko, S. F.

    Methods for the synthesis of controllable attenuators and their implementations are examined. In particular, attention is given to the general properties of controllable attenuators, control elements, types of controllable attenuators and methods of their analysis, and synthesis of the control characteristic of attenuators. The discussion also covers the efficiency of attenuator control, the use of transmission line segments in wide-band controllable attenuators, and attenuators with a discretely controlled transmission coefficient.

  4. Targeted Disruption of Chlamydia trachomatis Invasion by in Trans Expression of Dominant Negative Tarp Effectors

    PubMed Central

    Parrett, Christopher J.; Lenoci, Robert V.; Nguyen, Brenda; Russell, Lauren; Jewett, Travis J.

    2016-01-01

    Chlamydia trachomatis invasion of eukaryotic host cells is facilitated, in part, by the type III secreted effector protein, Tarp. The role of Tarp in chlamydiae entry of host cells is supported by molecular approaches that examined recombinant Tarp or Tarp effectors expressed within heterologous systems. A major limitation in the ability to study the contribution of Tarp to chlamydial invasion of host cells was the prior absence of genetic tools for chlamydiae. Based on our knowledge of Tarp domain structure and function along with the introduction of genetic approaches in C. trachomatis, we hypothesized that Tarp function could be disrupted in vivo by the introduction of dominant negative mutant alleles. We provide evidence that transformed C. trachomatis produced epitope tagged Tarp, which was secreted into the host cell during invasion. We examined the effects of domain specific Tarp mutations on chlamydial invasion and growth and demonstrate that C. trachomatis clones harboring engineered Tarp mutants lacking either the actin binding domain or the phosphorylation domain had reduced levels of invasion into host cells. These data provide the first in vivo evidence for the critical role of Tarp in C. trachomatis pathogenesis and indicate that chlamydial invasion of host cells can be attenuated via the introduction of engineered dominant negative type three effectors.

  5. Targeted Disruption of Chlamydia trachomatis Invasion by in Trans Expression of Dominant Negative Tarp Effectors.

    PubMed

    Parrett, Christopher J; Lenoci, Robert V; Nguyen, Brenda; Russell, Lauren; Jewett, Travis J

    2016-01-01

    Chlamydia trachomatis invasion of eukaryotic host cells is facilitated, in part, by the type III secreted effector protein, Tarp. The role of Tarp in chlamydiae entry of host cells is supported by molecular approaches that examined recombinant Tarp or Tarp effectors expressed within heterologous systems. A major limitation in the ability to study the contribution of Tarp to chlamydial invasion of host cells was the prior absence of genetic tools for chlamydiae. Based on our knowledge of Tarp domain structure and function along with the introduction of genetic approaches in C. trachomatis, we hypothesized that Tarp function could be disrupted in vivo by the introduction of dominant negative mutant alleles. We provide evidence that transformed C. trachomatis produced epitope tagged Tarp, which was secreted into the host cell during invasion. We examined the effects of domain specific Tarp mutations on chlamydial invasion and growth and demonstrate that C. trachomatis clones harboring engineered Tarp mutants lacking either the actin binding domain or the phosphorylation domain had reduced levels of invasion into host cells. These data provide the first in vivo evidence for the critical role of Tarp in C. trachomatis pathogenesis and indicate that chlamydial invasion of host cells can be attenuated via the introduction of engineered dominant negative type three effectors. PMID:27602332

  6. Targeted Disruption of Chlamydia trachomatis Invasion by in Trans Expression of Dominant Negative Tarp Effectors

    PubMed Central

    Parrett, Christopher J.; Lenoci, Robert V.; Nguyen, Brenda; Russell, Lauren; Jewett, Travis J.

    2016-01-01

    Chlamydia trachomatis invasion of eukaryotic host cells is facilitated, in part, by the type III secreted effector protein, Tarp. The role of Tarp in chlamydiae entry of host cells is supported by molecular approaches that examined recombinant Tarp or Tarp effectors expressed within heterologous systems. A major limitation in the ability to study the contribution of Tarp to chlamydial invasion of host cells was the prior absence of genetic tools for chlamydiae. Based on our knowledge of Tarp domain structure and function along with the introduction of genetic approaches in C. trachomatis, we hypothesized that Tarp function could be disrupted in vivo by the introduction of dominant negative mutant alleles. We provide evidence that transformed C. trachomatis produced epitope tagged Tarp, which was secreted into the host cell during invasion. We examined the effects of domain specific Tarp mutations on chlamydial invasion and growth and demonstrate that C. trachomatis clones harboring engineered Tarp mutants lacking either the actin binding domain or the phosphorylation domain had reduced levels of invasion into host cells. These data provide the first in vivo evidence for the critical role of Tarp in C. trachomatis pathogenesis and indicate that chlamydial invasion of host cells can be attenuated via the introduction of engineered dominant negative type three effectors. PMID:27602332

  7. The small protein CydX is required for function of cytochrome bd oxidase in Brucella abortus

    PubMed Central

    Sun, Yao-Hui; de Jong, Maarten F.; den Hartigh, Andreas B.; Roux, Christelle M.; Rolán, Hortensia G.; Tsolis, Renée M.

    2012-01-01

    A large number of hypothetical genes potentially encoding small proteins of unknown function are annotated in the Brucella abortus genome. Individual deletion of 30 of these genes identified four mutants, in BAB1_0355, BAB2_0726, BAB2_0470, and BAB2_0450 that were highly attenuated for infection. BAB2_0726, an YbgT-family protein located at the 3′ end of the cydAB genes encoding cytochrome bd ubiquinal oxidase, was designated cydX. A B. abortus cydX mutant lacked cytochrome bd oxidase activity, as shown by increased sensitivity to H2O2, decreased acid tolerance and increased resistance to killing by respiratory inhibitors. The C terminus, but not the N terminus, of CydX was located in the periplasm, suggesting that CydX is an integral cytoplasmic membrane protein. Phenotypic analysis of the cydX mutant, therefore, suggested that CydX is required for full function of cytochrome bd oxidase, possibly via regulation of its assembly or activity. PMID:22919638

  8. Polarized Cell Division of Chlamydia trachomatis

    PubMed Central

    Abdelrahman, Yasser; Ouellette, Scot P.; Belland, Robert J.; Cox, John V.

    2016-01-01

    Bacterial cell division predominantly occurs by a highly conserved process, termed binary fission, that requires the bacterial homologue of tubulin, FtsZ. Other mechanisms of bacterial cell division that are independent of FtsZ are rare. Although the obligate intracellular human pathogen Chlamydia trachomatis, the leading bacterial cause of sexually transmitted infections and trachoma, lacks FtsZ, it has been assumed to divide by binary fission. We show here that Chlamydia divides by a polarized cell division process similar to the budding process of a subset of the Planctomycetes that also lack FtsZ. Prior to cell division, the major outer-membrane protein of Chlamydia is restricted to one pole of the cell, and the nascent daughter cell emerges from this pole by an asymmetric expansion of the membrane. Components of the chlamydial cell division machinery accumulate at the site of polar growth prior to the initiation of asymmetric membrane expansion and inhibitors that disrupt the polarity of C. trachomatis prevent cell division. The polarized cell division of C. trachomatis is the result of the unipolar growth and FtsZ-independent fission of this coccoid organism. This mechanism of cell division has not been documented in other human bacterial pathogens suggesting the potential for developing Chlamydia-specific therapeutic treatments. PMID:27505160

  9. Polarized Cell Division of Chlamydia trachomatis.

    PubMed

    Abdelrahman, Yasser; Ouellette, Scot P; Belland, Robert J; Cox, John V

    2016-08-01

    Bacterial cell division predominantly occurs by a highly conserved process, termed binary fission, that requires the bacterial homologue of tubulin, FtsZ. Other mechanisms of bacterial cell division that are independent of FtsZ are rare. Although the obligate intracellular human pathogen Chlamydia trachomatis, the leading bacterial cause of sexually transmitted infections and trachoma, lacks FtsZ, it has been assumed to divide by binary fission. We show here that Chlamydia divides by a polarized cell division process similar to the budding process of a subset of the Planctomycetes that also lack FtsZ. Prior to cell division, the major outer-membrane protein of Chlamydia is restricted to one pole of the cell, and the nascent daughter cell emerges from this pole by an asymmetric expansion of the membrane. Components of the chlamydial cell division machinery accumulate at the site of polar growth prior to the initiation of asymmetric membrane expansion and inhibitors that disrupt the polarity of C. trachomatis prevent cell division. The polarized cell division of C. trachomatis is the result of the unipolar growth and FtsZ-independent fission of this coccoid organism. This mechanism of cell division has not been documented in other human bacterial pathogens suggesting the potential for developing Chlamydia-specific therapeutic treatments.

  10. New lymphogranuloma venereum Chlamydia trachomatis variant, Amsterdam.

    PubMed

    Spaargaren, Joke; Fennema, Han S A; Morré, Servaas A; de Vries, Henry J C; Coutinho, Roel A

    2005-07-01

    We retrospectively conducted a study of men who have sex with men who visited the Amsterdam, the Netherlands, sexually transmitted diseases clinic from January 2002 to December 2003 and had rectal Chlamydia trachomatis infections. We found that symptomatic (73%) as well as asymptomatic (43%) patients were infected with a new C. trachomatis LGV variant.

  11. Polarized Cell Division of Chlamydia trachomatis.

    PubMed

    Abdelrahman, Yasser; Ouellette, Scot P; Belland, Robert J; Cox, John V

    2016-08-01

    Bacterial cell division predominantly occurs by a highly conserved process, termed binary fission, that requires the bacterial homologue of tubulin, FtsZ. Other mechanisms of bacterial cell division that are independent of FtsZ are rare. Although the obligate intracellular human pathogen Chlamydia trachomatis, the leading bacterial cause of sexually transmitted infections and trachoma, lacks FtsZ, it has been assumed to divide by binary fission. We show here that Chlamydia divides by a polarized cell division process similar to the budding process of a subset of the Planctomycetes that also lack FtsZ. Prior to cell division, the major outer-membrane protein of Chlamydia is restricted to one pole of the cell, and the nascent daughter cell emerges from this pole by an asymmetric expansion of the membrane. Components of the chlamydial cell division machinery accumulate at the site of polar growth prior to the initiation of asymmetric membrane expansion and inhibitors that disrupt the polarity of C. trachomatis prevent cell division. The polarized cell division of C. trachomatis is the result of the unipolar growth and FtsZ-independent fission of this coccoid organism. This mechanism of cell division has not been documented in other human bacterial pathogens suggesting the potential for developing Chlamydia-specific therapeutic treatments. PMID:27505160

  12. Evidence for the effectiveness of a chlamydia awareness campaign: increased population rates of chlamydia testing and detection.

    PubMed

    Chen, Marcus Y; Karvelas, Maria; Sundararajan, Vijaya; Hocking, Jane S; Fairley, Christopher K

    2007-04-01

    The objective of this study was to determine the effectiveness of a statewide campaign aimed at increasing chlamydia awareness and testing among younger people. In November 2002, a narrowcast media campaign targeting men and women aged 16-29 years was launched in Victoria, Australia. This was expanded in June 2003. Data on chlamydia testing via Medicare and chlamydia notifications, before and after the campaign, were compared to determine possible effects of the campaign on population rates of chlamydia testing and detection. During the campaign, chlamydia testing rates increased significantly for both women (P=0.04) and men (P=0.04), while testing rates before and after the campaign remained relatively stable. Although testing rates increased, only 4.3% of Victorian women and 1.9% of men aged 16-30 were tested through Medicare in 2003. The increase in chlamydia testing over the study period was closely paralleled by an increase in notification rates for chlamydia, with strong correlations between the two (r=0.97, P<0.001). In conclusion, an estimated minimum of A$70 was spent on the campaign for each additional chlamydia test performed. Testing within the framework of a national chlamydia screening programme may be a more cost-effective way of increasing chlamydia testing. PMID:17509173

  13. Brucella abortus Cell Cycle and Infection Are Coordinated.

    PubMed

    De Bolle, Xavier; Crosson, Sean; Matroule, Jean-Yves; Letesson, Jean-Jacques

    2015-12-01

    Brucellae are facultative intracellular pathogens. The recent development of methods and genetically engineered strains allowed the description of cell-cycle progression of Brucella abortus, including unipolar growth and the ordered initiation of chromosomal replication. B. abortus cell-cycle progression is coordinated with intracellular trafficking in the endosomal compartments. Bacteria are first blocked at the G1 stage, growth and chromosome replication being resumed shortly before reaching the intracellular proliferation compartment. The control mechanisms of cell cycle are similar to those reported for the bacterium Caulobacter crescentus, and they are crucial for survival in the host cell. The development of single-cell analyses could also be applied to other bacterial pathogens to investigate their cell-cycle progression during infection.

  14. Characterization of Recombinant B. abortus Strain RB51SOD Toward Understanding the Uncorrelated Innate and Adaptive Immune Responses Induced by RB51SOD Compared to Its Parent Vaccine Strain RB51

    PubMed Central

    Zhu, Jianguo; Larson, Charles B.; Ramaker, Megan Ann; Quandt, Kimberly; Wendte, Jered M.; Ku, Kimberly P.; Chen, Fang; Jourdian, George W.; Vemulapalli, Ramesh; Schurig, Gerhardt G.; He, Yongqun

    2011-01-01

    Brucella abortus is a Gram-negative, facultative intracellular pathogen for several mammals, including humans. Live attenuated B. abortus strain RB51 is currently the official vaccine used against bovine brucellosis in the United States and several other countries. Overexpression of protective B. abortus antigen Cu/Zn superoxide dismutase (SOD) in a recombinant strain of RB51 (strain RB51SOD) significantly increases its vaccine efficacy against virulent B. abortus challenge in a mouse model. An attempt has been made to better understand the mechanism of the enhanced protective immunity of RB51SOD compared to its parent strain RB51. We previously reported that RB51SOD stimulated enhanced Th1 immune response. In this study, we further found that T effector cells derived from RB51SOD-immunized mice exhibited significantly higher cytotoxic T lymphocyte activity than T effector cells derived from RB51-immunized mice against virulent B. abortus-infected target cells. Meanwhile, the macrophage responses to these two strains were also studied. Compared to RB51, RB51SOD cells had a lower survival rate in macrophages and induced lower levels of macrophage apoptosis and necrosis. The decreased survival of RB51SOD cells correlates with the higher sensitivity of RB51SOD, compared to RB51, to the bactericidal action of either Polymyxin B or sodium dodecyl sulfate (SDS). Furthermore, a physical damage to the outer membrane of RB51SOD was observed by electron microscopy. Possibly due to the physical damage, overexpressed Cu/Zn SOD in RB51SOD was found to be released into the bacterial cell culture medium. Therefore, the stronger adaptive immunity induced by RB51SOD did not correlate with the low level of innate immunity induced by RB51SOD compared to RB51. This unique and apparently contradictory profile is likely associated with the differences in outer membrane integrity and Cu/Zn SOD release. PMID:22919576

  15. Characterization of recombinant B. abortus strain RB51SOD toward understanding the uncorrelated innate and adaptive immune responses induced by RB51SOD compared to its parent vaccine strain RB51.

    PubMed

    Zhu, Jianguo; Larson, Charles B; Ramaker, Megan Ann; Quandt, Kimberly; Wendte, Jered M; Ku, Kimberly P; Chen, Fang; Jourdian, George W; Vemulapalli, Ramesh; Schurig, Gerhardt G; He, Yongqun

    2011-01-01

    Brucella abortus is a Gram-negative, facultative intracellular pathogen for several mammals, including humans. Live attenuated B. abortus strain RB51 is currently the official vaccine used against bovine brucellosis in the United States and several other countries. Overexpression of protective B. abortus antigen Cu/Zn superoxide dismutase (SOD) in a recombinant strain of RB51 (strain RB51SOD) significantly increases its vaccine efficacy against virulent B. abortus challenge in a mouse model. An attempt has been made to better understand the mechanism of the enhanced protective immunity of RB51SOD compared to its parent strain RB51. We previously reported that RB51SOD stimulated enhanced Th1 immune response. In this study, we further found that T effector cells derived from RB51SOD-immunized mice exhibited significantly higher cytotoxic T lymphocyte activity than T effector cells derived from RB51-immunized mice against virulent B. abortus-infected target cells. Meanwhile, the macrophage responses to these two strains were also studied. Compared to RB51, RB51SOD cells had a lower survival rate in macrophages and induced lower levels of macrophage apoptosis and necrosis. The decreased survival of RB51SOD cells correlates with the higher sensitivity of RB51SOD, compared to RB51, to the bactericidal action of either Polymyxin B or sodium dodecyl sulfate (SDS). Furthermore, a physical damage to the outer membrane of RB51SOD was observed by electron microscopy. Possibly due to the physical damage, overexpressed Cu/Zn SOD in RB51SOD was found to be released into the bacterial cell culture medium. Therefore, the stronger adaptive immunity induced by RB51SOD did not correlate with the low level of innate immunity induced by RB51SOD compared to RB51. This unique and apparently contradictory profile is likely associated with the differences in outer membrane integrity and Cu/Zn SOD release.

  16. Brucella abortus Invasion of Osteoblasts Inhibits Bone Formation

    PubMed Central

    Scian, Romina; Barrionuevo, Paula; Fossati, Carlos A.; Giambartolomei, Guillermo H.

    2012-01-01

    Osteoarticular brucellosis is the most common presentation of the active disease in humans. Loss of bone is a serious complication of localized bacterial infection of bones or the adjacent tissue, and brucellosis proved not to be the exception. The skeleton is a dynamic organ system which is constantly remodeled. Osteoblasts are responsible for the deposition of bone matrix and are thought to facilitate the calcification and mineralization of the bone matrix, and their function could be altered under infectious conditions. In this article, we describe immune mechanisms whereby Brucella abortus may invade and replicate within osteoblasts, inducing apoptosis, inhibiting mineral and organic matrix deposition, and inducing upregulation of RANKL expression. Additionally, all of these mechanisms contributed in different ways to bone loss. These processes implicate the activation of signaling pathways (mitogen-activated protein kinases [MAPK] and caspases) involved in cytokine secretion, expression of activating molecules, and cell death of osteoblasts. In addition, considering the relevance of macrophages in intracellular Brucella survival and proinflammatory cytokine secretion in response to infection, we also investigated the role of these cells as modulators of osteoblast survival, differentiation, and function. We demonstrated that supernatants from B. abortus-infected macrophages may also mediate osteoblast apoptosis and inhibit osteoblast function in a process that is dependent on the presence of tumor necrosis factor alpha (TNF-α). These results indicate that B. abortus may directly and indirectly harm osteoblast function, contributing to the bone and joint destruction observed in patients with osteoarticular complications of brucellosis. PMID:22547546

  17. Molecular characterization of Brucella abortus chromosome II recombination.

    PubMed

    Tsoktouridis, Georgios; Merz, Christian A; Manning, Simon P; Giovagnoli-Kurtz, Renée; Williams, Leanne E; Mujer, Cesar V; Hagius, Sue; Elzer, Philip; Redkar, Rajendra J; Patra, Guy; DelVecchio, Vito G

    2003-10-01

    Large-scale genomic rearrangements including inversions, deletions, and duplications are significant in bacterial evolution. The recently completed Brucella melitensis 16M and Brucella suis 1330 genomes have facilitated the investigation of such events in the Brucella spp. Suppressive subtractive hybridization (SSH) was employed in identifying genomic differences between B. melitensis 16M and Brucella abortus 2308. Analysis of 45 SSH clones revealed several deletions on chromosomes of B. abortus and B. melitensis that encoded proteins of various metabolic pathways. A 640-kb inversion on chromosome II of B. abortus has been reported previously (S. Michaux Charachon, G. Bourg, E. Jumas Bilak, P. Guigue Talet, A. Allardet Servent, D. O'Callaghan, and M. Ramuz, J. Bacteriol. 179:3244-3249, 1997) and is further described in this study. One end of the inverted region is located on a deleted TATGC site between open reading frames BMEII0292 and BMEII0293. The other end inserted at a GTGTC site of the cyclic-di-GMP phosphodiesterase A (PDEA) gene (BMEII1009), dividing PDEA into two unequal DNA segments of 160 and 977 bp. As a consequence of inversion, the 160-bp segment that encodes the N-terminal region of PDEA was relocated at the opposite end of the inverted chromosomal region. The splitting of the PDEA gene most likely inactivated the function of this enzyme. A recombination mechanism responsible for this inversion is proposed.

  18. Synthesis of protein in host-free reticulate bodies of Chlamydia psittaci and Chlamydia trachomatis

    SciTech Connect

    Hatch, T.P.; Miceli, M.; Silverman, J.A.

    1985-06-01

    Synthesis of protein by the obligate intracellular parasitic bacteria Chlamydia psittaci (6BC) and Chlamydia trachomatis (serovar L2) isolated from host cells (host-free chlamydiae) was demonstrated for the first time. Incorporation of (/sup 35/S)methionine and (/sup 35/S)cysteine into trichloroacetic acid-precipitable material by reticulate bodies of chlamydiae persisted for 2 h and was dependent upon a exogenous source of ATP, an ATP-regenerating system, and potassium or sodium ions. Magnesium ions and amino acids stimulated synthesis; chloramphenicol, rifampin, oligomycin, and carbonyl cyanide p-trifluoromethoxyphenylhydrazone (a proton ionophore) inhibited incorporation. Ribonucleoside triphosphates (other than ATP) had little stimulatory effect. The optimum pH for host-free synthesis was between 7.0 and 7.5. The molecular weights of proteins synthesized by host-free reticulate bodies closely resembled the molecular weights of proteins synthesized by reticulate bodies in an intracellular environment, and included outer membrane proteins. Elementary bodies of chlamydiae were unable to synthesize protein even when incubated in the presence of 10 mM dithiothreitol, a reducing agent which converted the highly disulfide bond cross-linked major outer membrane protein to monomeric form.

  19. Brucella abortus DNA is a major bacterial agonist to activate the host innate immune system.

    PubMed

    Campos, Priscila Carneiro; Gomes, Marco Túlio Ribeiro; Guimarães, Gabriela; Costa Franco, Miriam Maria Silva; Marim, Fernanda Martins; Oliveira, Sergio Costa

    2014-12-01

    Immunity against Brucella abortus depends on the recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs). Signaling pathways triggered by Brucella DNA involves TLR9, AIM2 and possibly STING and MAVS. Herein, we review the advances in B. abortus DNA sensing by host innate immune receptors and the progress in this field.

  20. Draft Genome Sequence of the Live Vaccine Strain Brucella abortus 82

    PubMed Central

    Shevtsov, Alexander; Zholdybayeva, Elena; Shevtsova, Elena; Momynkulov, Dauren; Sytnik, Igor; Karibaev, Talgat; Chsherbakov, Andrey; Momynaliev, Kuvat

    2013-01-01

    Vaccination is a crucial part of the brucellosis eradication programs worldwide. A live vaccine strain of Brucella abortus 82 has been successfully used for the vaccination of cattle against brucellosis in the former Soviet republics for the last 39 years. Here, we report the genome sequence of Brucella abortus 82. PMID:24371203

  1. Simultaneous identification of antibodies to Brucella abortus and Staphylococcus aureus in milk samples by flow cytometry.

    PubMed

    Iannelli, D; D'Apice, L; Fenizia, D; Serpe, L; Cottone, C; Viscardi, M; Capparelli, R

    1998-03-01

    Two flow cytometric assays are described herein. The single cytometric test (SCT) detects antibodies to either Brucella abortus or Staphylococcus aureus in the serum or milk of a cow or water buffalo. The double cytometric test (DCT) detects both anti-B. abortus and anti-S. aureus antibodies concurrently. In the SCT, the sample to be tested is incubated in succession with the antigen (either B. abortus or S. aureus) and the proper secondary antiserum (fluorescein isothiocyanate-labelled rabbit anti-cow immunoglobulin antiserum or rabbit anti-water buffalo immunoglobulin antiserum). In the DCT, the sample to be tested is incubated first with B. abortus and S. aureus antigens and then with the secondary antiserum. The B. abortus antigen used in the DCT is covalently bound to 3-microm-diameter latex particles. The difference in size between B. abortus and S. aureus permits the establishment of whether the antibodies are directed against one, the other, or both antigens. When compared to the complement fixation test, the SCT and DCT each show a specificity and a sensitivity of 100%. The SCT has been used previously to detect anti-S. aureus antibodies. Here its use is extended to the detection of anti-B. abortus antibodies. The DCT is described here for the first time. The DCT appears to be useful for large-scale brucellosis eradication programs. It offers the possibility of using one test to identify animals that are serologically positive for both B. abortus and S. aureus.

  2. Immunization of Mice with Recombinant Brucella abortus Organic Hydroperoxide Resistance (Ohr) Protein Protects Against a Virulent Brucella abortus 544 Infection.

    PubMed

    Hop, Huynh Tan; Reyes, Alisha Wehdnesday Bernardo; Simborio, Hannah Leah Tadeja; Arayan, Lauren Togonon; Min, Won Gi; Lee, Hu Jang; Lee, Jin Ju; Chang, Hong Hee; Kim, Suk

    2016-01-01

    In this study, the Brucella abortus ohr gene coding for an organic hydroperoxide resistance protein (Ohr) was cloned into a maltose fusion protein expression system (pMAL), inserted into Escherichia coli, and purified, and its immunogenicity was evaluated by western blot analysis using Brucella-positive mouse sera. The purified recombinant Ohr (rOhr) was treated with adjuvant and injected intraperitoneally into BALB/c mice. A protective immune response analysis revealed that rOhr induced a significant increase in both the IgG1 and IgG2a titers, and IgG2a reached a higher level than IgG1 after the second and third immunizations. Additionally, immunization with rOhr induced high production of IFN-γ as well as proinflammatory cytokines such as TNF, MCP-1, IL-12p70, and IL-6, but a lesser amount of IL-10, suggesting that rOhr predominantly elicited a cell-mediated immune response. In addition, immunization with rOhr caused a significantly higher degree of protection against a virulent B. abortus infection compared with a positive control group consisting of mice immunized with maltose-binding protein. These findings showed that B. abortus rOhr was able to induce both humoral and cell-mediated immunity in mice, which suggested that this recombinant protein could be a potential vaccine candidate for animal brucellosis.

  3. Vaccination of Elk (Cervus canadensis) with Brucella abortus Strain RB51 Overexpressing Superoxide Dismutase and Glycosyltransferase Genes Does Not Induce Adequate Protection against Experimental Brucella abortus Challenge.

    PubMed

    Nol, Pauline; Olsen, Steven C; Rhyan, Jack C; Sriranganathan, Nammalwar; McCollum, Matthew P; Hennager, Steven G; Pavuk, Alana A; Sprino, Phillip J; Boyle, Stephen M; Berrier, Randall J; Salman, Mo D

    2016-01-01

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosyltransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further, work is needed for development of an effective brucellosis vaccine for use in elk. PMID:26904509

  4. Vaccination of Elk (Cervus canadensis) with Brucella abortus Strain RB51 Overexpressing Superoxide Dismutase and Glycosyltransferase Genes Does Not Induce Adequate Protection against Experimental Brucella abortus Challenge.

    PubMed

    Nol, Pauline; Olsen, Steven C; Rhyan, Jack C; Sriranganathan, Nammalwar; McCollum, Matthew P; Hennager, Steven G; Pavuk, Alana A; Sprino, Phillip J; Boyle, Stephen M; Berrier, Randall J; Salman, Mo D

    2016-01-01

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosyltransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further, work is needed for development of an effective brucellosis vaccine for use in elk.

  5. Vaccination of Elk (Cervus canadensis) with Brucella abortus Strain RB51 Overexpressing Superoxide Dismutase and Glycosyltransferase Genes Does Not Induce Adequate Protection against Experimental Brucella abortus Challenge

    PubMed Central

    Nol, Pauline; Olsen, Steven C.; Rhyan, Jack C.; Sriranganathan, Nammalwar; McCollum, Matthew P.; Hennager, Steven G.; Pavuk, Alana A.; Sprino, Phillip J.; Boyle, Stephen M.; Berrier, Randall J.; Salman, Mo D.

    2016-01-01

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area. In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the development of effective disease management strategies for wild elk herds is of utmost importance, not only for the prevention of reintroduction of brucellosis to cattle, but also for the overall health of the Greater Yellowstone Area elk populations. In two studies, we evaluated the efficacy of B. abortus strain RB51 over-expressing superoxide dismutase and glycosyltransferase for protecting elk from infection and disease caused by B. abortus after experimental infection with a virulent B. abortus strain. Our data indicate that the recombinant vaccine does not protect elk against brucellosis. Further, work is needed for development of an effective brucellosis vaccine for use in elk. PMID:26904509

  6. [Follicular conjunctivitis due to Chlamydia trachomatis].

    PubMed

    Basualdo, J A; Huarte, L; Bautista, E; Niedfeld, G; Alfonso, G; Rosso, N; Geronés, M; Galeppi, I

    2001-01-01

    During two years (1997-1999) an investigation of possible infections of chlamydial etiology in outpatients with follicular conjunctivitis was carried out, through the use of specific assays. Fifty-seven selected patients with presumptive inclusion conjunctivitis were diagnosed by means of ophthalmoscopic examination and bilateral tarsal-conjunctiva swabbing for microorganisms. The possible presence of Chlamydia trachomatis was tested by immunofluorescence microscopy and isolation in cell culture of McCoy line. Of the 57 conjunctivitis patients screened, 37 (65%) proved to be positive by cell culture (CC) and 27 (47%) by direct immunofluorescence (IFD). A good agreement between the two assays was observed, where the CC was more sensitive than IFD. Of these 37 patients with chlamydial conjunctivitis, 23 (62%) were women, with over one-third of them ranging in age from 45 to 65 years. Their clinical records revealed an evolution period of 1 to 12 months. Eighteen (78%) of these women reported previous genital pathology, while 4 (29%) of the 14 men had a history of urethritis by Chlamydia trachomatis. A high frequency of follicular conjunctivitis by Chlamydia (65%) in the screened patients was observed, without any evidence of urogenital signs and symptoms at the moment of the study.

  7. Interaction of Chlamydiae with human macrophages.

    PubMed

    Herweg, Jo-Ana; Rudel, Thomas

    2016-02-01

    The phylum Chlamydiae contains several members that are well-known human pathogens, like Chlamydia trachomatis and C. pneumoniae. Establishing a chronic bacterial infection requires the active evasion of the host immune response. A major arm of the innate immune defence is constituted by macrophages, which fight infections by removing bacteria and triggering an adaptive immune response. However, some pathogenic Chlamydia infect and survive in macrophages at least for a certain period of time. Therefore, macrophages can serve as vehicles for the dissemination of bacterial infections from the primary infection site via the urogenital or respiratory tract to distant sites in the body. The capacity to infect macrophages seems to depend on the chlamydial strain and the source of macrophages. In vitro infections of macrophages with C. trachomatis, C. psittaci and C. pneumoniae reveal low efficiency of infection and progeny formation, as well as failure to develop mature inclusions. In contrast, the emerging pathogen, Simkania negevensis, actively replicates in macrophages. Here we summarize the current knowledge of the intracellular and molecular key mechanisms of C. trachomatis, C. pneumoniae and S. negevensis infections in human macrophages. PMID:26613554

  8. Evolution, phylogeny, and molecular epidemiology of Chlamydia.

    PubMed

    Nunes, Alexandra; Gomes, João P

    2014-04-01

    The Chlamydiaceae are a family of obligate intracellular bacteria characterized by a unique biphasic developmental cycle. It encompasses the single genus Chlamydia, which involves nine species that affect a wide range of vertebral hosts, causing infections with serious impact on human health (mainly due to Chlamydia trachomatis infections) and on farming and veterinary industries. It is believed that Chlamydiales originated ∼700mya, whereas C. trachomatis likely split from the other Chlamydiaceae during the last 6mya. This corresponds to the emergence of modern human lineages, with the first descriptions of chlamydial infections as ancient as four millennia. Chlamydiaceae have undergone a massive genome reduction, on behalf of the deletional bias "use it or lose it", stabilizing at 1-1.2Mb and keeping a striking genome synteny. Their phylogeny reveals species segregation according to biological properties, with huge differences in terms of host range, tissue tropism, and disease outcomes. Genome differences rely on the occurrence of mutations in the >700 orthologous genes, as well as on events of recombination, gene loss, inversion, and paralogous expansion, affecting both a hypervariable region named the plasticity zone, and genes essentially encoding polymorphic and transmembrane head membrane proteins, type III secretion effectors and some metabolic pathways. Procedures for molecular typing are still not consensual but have allowed the knowledge of molecular epidemiology patterns for some species as well as the identification of outbreaks and emergence of successful clones for C. trachomatis. This manuscript intends to provide a comprehensive review on the evolution, phylogeny, and molecular epidemiology of Chlamydia.

  9. Post-exposure serological and bacteriological responses of water buffalo (Bubalus bubalis) to Brucella abortus biovar 1 following vaccination with Brucella abortus strain RB51.

    PubMed

    Diptee, M D; Asgarali, Z; Campbell, M; Fosgate, G; Adesiyun, A A

    2007-12-01

    Serological and bacteriological responses to Brucella abortus biovar 1 following vaccination with B. abortus strain RB51 (RB51) were evaluated in thirty domestic water buffalo (Bubalus bubalis) randomly divided into five treatment groups. Groups I to V received, respectively, the recommended dose (RD) of RB51 vaccine once, RD twice 4 weeks apart, double RD once, double RD twice 4 weeks apart, and saline once (control). Vaccination did not result in a serological response. Experimental animals released 27 weeks post initial inoculation (27 PIIW) into a brucellosis-positive herd failed to seroconvert after 29 weeks. Experimental challenge commenced at 57 PIIW. All animals received B. abortus biovar 1 intraconjunctivally at 0, 5 and 9 weeks post experimental exposure (PEEW). Serum samples collected at 4, 8 and 13 PEEW were negative. At 16 PEEW all animals received B. abortus biovar 1 subcutaneously (SC), and all seroconverted by 20 PEEW. Five of twenty-six animals were positive for Brucella infection on bacterial culture. Brucella abortus biovar 1 was isolated from three animals; B. abortus RB51 was isolated from two. Treatment group, age and sex had no effect on the isolation of Brucellae (P>0.05).

  10. Intermediate rough Brucella abortus S19Δper mutant is DIVA enable, safe to pregnant guinea pigs and confers protection to mice.

    PubMed

    Lalsiamthara, Jonathan; Gogia, Neha; Goswami, Tapas K; Singh, R K; Chaudhuri, Pallab

    2015-05-21

    Brucella abortus S19 is a smooth strain used as live vaccine against bovine brucellosis. Smooth lipopolysaccharide (LPS) is responsible for its residual virulence and serological interference. Rough mutants defective of LPS are more attenuated but confers lower level of protection. We describe a modified B. abortus S19 strain, named as S19Δper, which exhibits intermediate rough phenotype with residual O-polysaccharide (OPS). Deletion of perosamine synthetase gene resulted in substantial attenuation of S19Δper mutant without affecting immunogenic properties. It mounted strong immune response in Swiss albino mice and conferred protection similar to S19 vaccine. Immunized mice produced higher levels of IFN-γ, IgG2a and thus has immune response inclined towards Th1 cell mediated immunity. Sera from immunized animals did not show agglutination reaction with RBPT antigen and thus could serve as DIVA (Differentiating Infected from Vaccinated Animals) vaccine. S19Δper mutant displayed more susceptibility to serum complement mediated killing and sensitivity to polymyxin B. Pregnant guinea pigs injected with S19Δper mutant completed full term of pregnancy and did not cause abortion, still birth or birth of weak offspring. S19Δper mutant with intermediate rough phenotype displayed remarkable resemblance to S19 vaccine strain with improved properties of safety, immunogenicity and DIVA capability for control of bovine brucellosis.

  11. Isolation of single Chlamydia-infected cells using laser microdissection.

    PubMed

    Podgorny, Oleg V; Polina, Nadezhda F; Babenko, Vladislav V; Karpova, Irina Y; Kostryukova, Elena S; Govorun, Vadim M; Lazarev, Vassili N

    2015-02-01

    Chlamydia are obligate intracellular parasites of humans and animals that cause a wide range of acute and chronic infections. To elucidate the genetic basis of chlamydial parasitism, several approaches for making genetic modifications to Chlamydia have recently been reported. However, the lack of the available methods for the fast and effective selection of genetically modified bacteria restricts the application of genetic tools. We suggest the use of laser microdissection to isolate of single live Chlamydia-infected cells for the re-cultivation and whole-genome sequencing of single inclusion-derived Chlamydia. To visualise individual infected cells, we made use of the vital labelling of inclusions with the fluorescent Golgi-specific dye BODIPY® FL C5-ceramide. We demonstrated that single Chlamydia-infected cells isolated by laser microdissection and placed onto a host cell monolayer resulted in new cycles of infection. We also demonstrated the successful use of whole-genome sequencing to study the genomic variability of Chlamydia derived from a single inclusion. Our work provides the first evidence of the successful use of laser microdissection for the isolation of single live Chlamydia-infected cells, thus demonstrating that this method can help overcome the barriers to the fast and effective selection of Chlamydia.

  12. IL-17/il-23 and Chlamydia trachomatis.

    PubMed

    Shavlakadze, N; Gorgoshidze, B

    2010-06-01

    Ch. trachomatis is a gram negative bacteria, infecting the most organs of the uro-genital tract and harming greatly the woman's or man's reproductive field. Moreover, as this is not the limit of its destructive nature, it can form a favorable ground for developing ulcer and tumor processes. As observed, a special place is occupied by Ch. trachomatis chronization skill, which is developed irrespective the mighty humoral and cellular respond to its intrusion into the host's organism. It is also known that T (CD4) lymphocytes and their products - cytokines are directly involved into these processes. IL-17 and its regulator IL-23 are among them. For the significance of the above-mentioned processes for expiring the chlamydiosis immune-pathogenesis we have studied the problem in the patients with IL-17 and IL-23 chlamydiosis. We have investigated 56 chlamydia infected patients; 31 non-infected patients who were the carriers of a different pathology flora and 21 healthy donors. The investigation covered some impaired localities as well as the organism overall. To gain the objective we analyzed clinical -anamnesis data and carried out the appropriate instrumental, laboratorial and immunological researches. Stating the chlamydia infection was carried on with the serological and immunofluorescentical and PCR methods. The study of IL-17 and IL-23 is done by ELISA and RT-PCR methods.The findings after the statistical analyses makes us drive to the following conclusions: IL-17 occurs in almost all the patients infected with chlamydia - their organs or systemic environment compared with the patients of non-chlamydia infection (97% against 21%, P<0,05). At the same time, IL-17 has been measured by higher parameters, than IL-23. The highest parameters of IL-17 were recorded with the patients having acute chlamydiosis in the impaired localities and also with the patients having arthritis and high antiovarial antibodies, IL-23 in high digits was recorded with the patients having a

  13. Immunization with Recombinant Brucella Species Outer Membrane Protein Omp16 or Omp19 in Adjuvant Induces Specific CD4+ and CD8+ T Cells as Well as Systemic and Oral Protection against Brucella abortus Infection▿

    PubMed Central

    Pasquevich, Karina A.; Estein, Silvia M.; Samartino, Clara García; Zwerdling, Astrid; Coria, Lorena M.; Barrionuevo, Paula; Fossati, Carlos A.; Giambartolomei, Guillermo H.; Cassataro, Juliana

    2009-01-01

    Available vaccines against Brucella spp. are live attenuated Brucella strains. In order to engineer a better vaccine to be used in animals and humans, our laboratory aims to develop an innocuous subunit vaccine. Particularly, we are interested in the outer membrane proteins (OMPs) of B. abortus: Omp16 and Omp19. In this study, we assessed the use of these proteins as vaccines against Brucella in BALB/c mice. Immunization with lipidated Omp16 (L-Omp16) or L-Omp19 in incomplete Freund's adjuvant (IFA) conferred significant protection against B. abortus infection. Vaccination with unlipidated Omp16 (U-Omp16) or U-Omp19 in IFA induced a higher degree of protection than the respective lipidated versions. Moreover, the level of protection induced after U-Omp16 or U-Omp19 immunization in IFA was similar to that elicited by live B. abortus S19 immunization. Flow cytometric analysis showed that immunization with U-Omp16 or U-Omp19 induced antigen-specific CD4+ as well as CD8+ T cells producing gamma interferon. In vivo depletion of CD4+ or CD8+ T cells in mice immunized with U-Omp16 or U-Omp19 plus IFA resulted in a loss of the elicited protection, indicating that both cell types are mediating immune protection. U-Omp16 or U-Omp19 vaccination induced a T helper 1 response, systemic protection in aluminum hydroxide formulation, and oral protection with cholera toxin adjuvant against B. abortus infection. Both immunization routes exhibited a similar degree of protection to attenuated Brucella vaccines (S19 and RB51, respectively). Overall these results indicate that U-Omp16 or U-Omp19 would be a useful candidate for a subunit vaccine against human and animal brucellosis. PMID:18981242

  14. Brucella abortus Choloylglycine Hydrolase Affects Cell Envelope Composition and Host Cell Internalization

    PubMed Central

    Marchesini, María Inés; Connolly, Joseph; Delpino, María Victoria; Baldi, Pablo C.; Mujer, Cesar V.; DelVecchio, Vito G.; Comerci, Diego J.

    2011-01-01

    Choloylglycine hydrolase (CGH, E.C. 3.5.1.24) is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh) and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization. PMID:22174816

  15. Brucella abortus choloylglycine hydrolase affects cell envelope composition and host cell internalization.

    PubMed

    Marchesini, María Inés; Connolly, Joseph; Delpino, María Victoria; Baldi, Pablo C; Mujer, Cesar V; DelVecchio, Vito G; Comerci, Diego J

    2011-01-01

    Choloylglycine hydrolase (CGH, E.C. 3.5.1.24) is a conjugated bile salt hydrolase that catalyses the hydrolysis of the amide bond in conjugated bile acids. Bile salt hydrolases are expressed by gastrointestinal bacteria, and they presumably decrease the toxicity of host's conjugated bile salts. Brucella species are the causative agents of brucellosis, a disease affecting livestock and humans. CGH confers Brucella the ability to deconjugate and resist the antimicrobial action of bile salts, contributing to the establishment of a successful infection through the oral route in mice. Additionally, cgh-deletion mutant was also attenuated in intraperitoneally inoculated mice, which suggests that CGH may play a role during systemic infection other than hydrolyzing conjugated bile acids. To understand the role CGH plays in B. abortus virulence, we infected phagocytic and epithelial cells with a cgh-deletion mutant (Δcgh) and found that it is defective in the internalization process. This defect along with the increased resistance of Δcgh to the antimicrobial action of polymyxin B, prompted an analysis of the cell envelope of this mutant. Two-dimensional electrophoretic profiles of Δcgh cell envelope-associated proteins showed an altered expression of Omp2b and different members of the Omp25/31 family. These results were confirmed by Western blot analysis with monoclonal antibodies. Altogether, the results indicate that Brucella CGH not only participates in deconjugation of bile salts but also affects overall membrane composition and host cell internalization.

  16. Molecular characterization of Chlamydia pneumoniae in animals and humans from Argentina: Genetic characterization of Chlamydia pneumoniae.

    PubMed

    Frutos, María C; Monetti, Marina S; Mosmann, Jessica; Kiguen, Ana X; Venezuela, Fernando R; Ré, Viviana E; Cuffini, Cecilia G

    2016-10-01

    In this study, genetic diversity of Chlamydia pneumoniae was investigated and the relationships between sequences amplified of different sources, clinical conditions and geographical regions of central Argentina were established. Samples amplified were similar to human C. pneumoniae patterns and show the high clonality of the population.

  17. Molecular characterization of Chlamydia pneumoniae in animals and humans from Argentina: Genetic characterization of Chlamydia pneumoniae.

    PubMed

    Frutos, María C; Monetti, Marina S; Mosmann, Jessica; Kiguen, Ana X; Venezuela, Fernando R; Ré, Viviana E; Cuffini, Cecilia G

    2016-10-01

    In this study, genetic diversity of Chlamydia pneumoniae was investigated and the relationships between sequences amplified of different sources, clinical conditions and geographical regions of central Argentina were established. Samples amplified were similar to human C. pneumoniae patterns and show the high clonality of the population. PMID:27328126

  18. Chlamydia trachomatis genotypes in school adolescents, Italy.

    PubMed

    Stefanelli, Paola; Sulis, Giorgia; Renna, Giovanna; Gargiulo, Franco; Zanotti, Paola; Capelli, Michela; De Francesco, Maria Antonia; Donato, Francesco; Pecorelli, Sergio; Matteelli, Alberto

    2015-12-01

    Chlamydia trachomatis genogroups using ompA and multilocus sequence typing (MLST) were determined in consecutive isolates from school students aged 18 or older in the district of Brescia, Italy, 2012-2013. Among 40 samples, 4 ompA genovars and 18 STs were identified. Genovar E predominated (70 %) including five STs derived from ST59 (29 % of all isolates). This study, combining ompA and MLST typing of C. trachomatis school teenagers, suggests limited mixing and sexual interchange in this population.

  19. Neutralization of Chlamydia trachomatis in cell culture.

    PubMed Central

    Howard, L V

    1975-01-01

    Neutralization of Chlamydia trachomatis was assayed by the decrease in inclusion-forming units in baby hamster kidney cells grown in culture. Five percent fresh guinea pig sera increased neutralization titers of rabbit antisera 100- to 1,000-fold but had no effect when normal rabbit sera were tested. Neutralization of a type A or B trachoma isolate was strain specific. Neutralization by human eye secretions and sera also was demonstrated when guinea pig sera were included in the test. All of the six human sera tested showed strain specificity against types A or B, in agreement with typing by the fluorescent antibody technique. PMID:1091549

  20. Genetic diversity of Chlamydia among captive birds from central Argentina.

    PubMed

    Frutos, María C; Monetti, Marina S; Vaulet, Lucia Gallo; Cadario, María E; Fermepin, Marcelo Rodríguez; Ré, Viviana E; Cuffini, Cecilia G

    2015-01-01

    To study the occurrence of Chlamydia spp. and their genetic diversity, we analysed 793 cloacal swabs from 12 avian orders, including 76 genera, obtained from 80 species of asymptomatic wild and captive birds that were examined with conventional nested polymerase chain reaction and quantitative polymerase chain reaction. Chlamydia spp. were not detected in wild birds; however, four species (Chlamydia psittaci, Chlamydia pecorum, Chlamydia pneumoniae and Chlamydia gallinacea) were identified among captive birds (Passeriformes, n = 20; Psittaciformes, n = 15; Rheiformes, n = 8; Falconiformes n = 2; Piciformes n = 2; Anseriformes n = 1; Galliformes n = 1; Strigiformes n = 1). Two pathogens (C. pneumoniae and C. pecorum) were identified simultaneously in samples obtained from captive birds. Based on nucleotide-sequence variations of the ompA gene, three C. psittaci-positive samples detected were grouped into a cluster with the genotype WC derived from mammalian hosts. A single positive sample was phylogenetically related to a new strain of C. gallinacea. This report contributes to our increasing understanding of the abundance of Chlamydia in the animal kingdom. PMID:25469538

  1. Genetic diversity of Chlamydia among captive birds from central Argentina.

    PubMed

    Frutos, María C; Monetti, Marina S; Vaulet, Lucia Gallo; Cadario, María E; Fermepin, Marcelo Rodríguez; Ré, Viviana E; Cuffini, Cecilia G

    2015-01-01

    To study the occurrence of Chlamydia spp. and their genetic diversity, we analysed 793 cloacal swabs from 12 avian orders, including 76 genera, obtained from 80 species of asymptomatic wild and captive birds that were examined with conventional nested polymerase chain reaction and quantitative polymerase chain reaction. Chlamydia spp. were not detected in wild birds; however, four species (Chlamydia psittaci, Chlamydia pecorum, Chlamydia pneumoniae and Chlamydia gallinacea) were identified among captive birds (Passeriformes, n = 20; Psittaciformes, n = 15; Rheiformes, n = 8; Falconiformes n = 2; Piciformes n = 2; Anseriformes n = 1; Galliformes n = 1; Strigiformes n = 1). Two pathogens (C. pneumoniae and C. pecorum) were identified simultaneously in samples obtained from captive birds. Based on nucleotide-sequence variations of the ompA gene, three C. psittaci-positive samples detected were grouped into a cluster with the genotype WC derived from mammalian hosts. A single positive sample was phylogenetically related to a new strain of C. gallinacea. This report contributes to our increasing understanding of the abundance of Chlamydia in the animal kingdom.

  2. Interaction of chlamydiae and host cells in vitro.

    PubMed Central

    Moulder, J W

    1991-01-01

    The obligately intracellular bacteria of the genus Chlamydia, which is only remotely related to other eubacterial genera, cause many diseases of humans, nonhuman mammals, and birds. Interaction of chlamydiae with host cells in vitro has been studied as a model of infection in natural hosts and as an example of the adaptation of an organism to an unusual environment, the inside of another living cell. Among the novel adaptations made by chlamydiae have been the substitution of disulfide-bond-cross-linked polypeptides for peptidoglycans and the use of host-generated nucleotide triphosphates as sources of metabolic energy. The effect of contact between chlamydiae and host cells in culture varies from no effect at all to rapid destruction of either chlamydiae or host cells. When successful infection occurs, it is usually followed by production of large numbers of progeny and destruction of host cells. However, host cells containing chlamydiae sometimes continue to divide, with or without overt signs of infection, and chlamydiae may persist indefinitely in cell cultures. Some of the many factors that influence the outcome of chlamydia-host cell interaction are kind of chlamydiae, kind of host cells, mode of chlamydial entry, nutritional adequacy of the culture medium, presence of antimicrobial agents, and presence of immune cells and soluble immune factors. General characteristics of chlamydial multiplication in cells of their natural hosts are reproduced in established cell lines, but reproduction in vitro of the subtle differences in chlamydial behavior responsible for the individuality of the different chlamydial diseases will require better in vitro models. PMID:2030670

  3. Prevention of lethal experimental infection of C57BL/6 mice by vaccination with Brucella abortus strain RB51 expressing Neospora caninum antigens.

    PubMed

    Ramamoorthy, Sheela; Sanakkayala, Neelima; Vemulapalli, Ramesh; Duncan, Robert B; Lindsay, David S; Schurig, Gerhart S; Boyle, Stephen M; Kasimanickam, Ramanathan; Sriranganathan, Nammalwar

    2007-11-01

    Bovine abortions caused by the intracellular protozoal parasite Neospora caninum are a major concern to cattle industries worldwide. A strong Th1 immune response is required for protection against N. caninum. Brucella abortus strain RB51 is currently used as a live, attenuated vaccine against bovine brucellosis. Strain RB51 can also be used as an expression vector for heterologous protein expression. In this study, putative protective antigens of N. caninum MIC1, MIC3, GRA2, GRA6 and SRS2, were expressed individually in B. abortus strain RB51. The ability of each of the recombinant RB51 strains to induce N. caninum-specific immunity was assessed in C57BL/6 mice. Mice were immunised by two i.p. inoculations, 4 weeks apart. Five weeks after the second immunisation, spleen cells from the vaccinated mice secreted high levels of IFN-gamma and IL-10 upon in vitro stimulation with N. caninum whole cell lysate antigens. N. caninum-specific antibodies of both IgG1 and IgG2a subtypes were detected in the serum of the vaccinated mice. Mice in the vaccinated and control groups were challenged with 2 x 10(7)N. caninum tachyzoites i.p. and observed for 28 days after vaccination. All unvaccinated control mice died within 7 days. Mice in the MIC1 and GRA6 vaccine groups were completely protected while the mice in the SRS2, GRA2 and MIC3 vaccinated groups were partially protected and experienced 10-50% mortality. The non-recombinant RB51 vector control group experienced an average protection of 69%. These results suggest that expression of protective antigens of N. caninum in B. abortus strain RB51 is a novel approach towards the development of a multivalent vaccine against brucellosis and neosporosis.

  4. Impact of azithromycin resistance mutations on the virulence and fitness of Chlamydia caviae in guinea pigs.

    PubMed

    Binet, Rachel; Bowlin, Anne K; Maurelli, Anthony T; Rank, Roger G

    2010-03-01

    Azithromycin (AZM) is a major drug used in the treatment and prophylaxis of infections caused by Chlamydia, yet no significant clinical resistance has been reported for these obligate intracellular bacteria. Nevertheless, spontaneous AZM resistance (Azm(r)) arose in vitro at frequencies ranging from 3 x 10(-8) to 8 x 10(-10) for clonal isolates of Chlamydia caviae, which is a natural pathogen of guinea pigs. Sequencing of the unique 23S rRNA gene copy in 44 independent Azm(r) isolates identified single mutations at position A(2058) or A(2059) (Escherichia coli numbering system). While SP(6)AZ(1) (A(2058)C) and SP(6)AZ(2) (A(2059)C) Azm(r) mutants showed growth defects in cell culture and were less pathogenic in the guinea pig ocular infection model than in the parent SP(6), the three isogenic C. caviae isolates grew equally well in the animal. On the other hand, coinoculation of the C. caviae parent strain with one of the Azm(r) strains was detrimental for the mutant strain. This apparent lack of association between pathology and bacterial load in vivo showed that virulence of the two Azm(r) mutants of C. caviae was attenuated. While chlamydial growth in vitro reflects the ability of the bacteria to multiply in permissive cells, survival in the host is a balance between cellular multiplication and clearance by the host immune system. The obligate intracellular nature of Chlamydia may therefore limit emergence of resistance in vivo due to the strength of the immune response induced by the wild-type antibiotic-sensitive bacteria at the time of antibiotic treatment.

  5. Brucella abortus Synthesizes Phosphatidylcholine from Choline Provided by the Host

    PubMed Central

    Comerci, Diego J.; Altabe, Silvia; de Mendoza, Diego; Ugalde, Rodolfo A.

    2006-01-01

    The Brucella cell envelope is characterized by the presence of phosphatidylcholine (PC), a common phospholipid in eukaryotes that is rare in prokaryotes. Studies on the composition of Brucella abortus 2308 phospholipids revealed that the synthesis of PC depends on the presence of choline in the culture medium, suggesting that the methylation biosynthetic pathway is not functional. Phospholipid composition of pmtA and pcs mutants indicated that in Brucella, PC synthesis occurs exclusively via the phosphatidylcholine synthase pathway. Transformation of Escherichia coli with an expression vector containing the B. abortus pcs homologue was sufficient for PC synthesis upon induction with IPTG (isopropyl-β-d-thiogalactopyranoside), while no PC formation was detected when bacteria were transformed with a vector containing pmtA. These findings imply that Brucella depends on choline provided by the host cell to form PC. We could not detect any obvious associated phenotype in the PC-deficient strain under vegetative or intracellular growth conditions in macrophages. However, the pcs mutant strain displays a reproducible virulence defect in mice, which suggests that PC is necessary to sustain a chronic infection process. PMID:16484204

  6. Tracing the primordial Chlamydiae: extinct parasites of plants?

    PubMed

    Subtil, Agathe; Collingro, Astrid; Horn, Matthias

    2014-01-01

    Chlamydiae are obligate intracellular bacteria found as symbionts and pathogens in a wide range of eukaryotes, including protists, invertebrates, and vertebrates. It was recently proposed that an ancient chlamydial symbiont facilitated the establishment of primary plastids in a tripartite symbiosis with cyanobacteria and early eukaryotes. In this review, we summarize recent advances in understanding of the lifestyle and the evolutionary history of extant Chlamydiae. We reconstruct and describe key features of the ancient chlamydial symbiont. We propose that it was already adapted to an intracellular lifestyle before the emergence of Archaeplastida, and that several observations are compatible with an essential contribution of Chlamydiae to the evolution of algae and plants.

  7. Immunogenicity and protective effect of recombinant Brucella abortus Ndk (rNdk) against a virulent strain B. abortus 544 infection in BALB/c mice.

    PubMed

    Hop, Huynh Tan; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Arayan, Lauren Togonon; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2015-02-01

    In this study, we particularly evaluated the protective effect of recombinant protein encoded by Brucella abortus 544 ndk (nucleoside diphosphate kinase) gene against B. abortus infection in the BALB/c mice. Cloning and expression of B. abortus Ndk was accomplished by PCR amplification into a pMAL expression system, and purification of a recombinant Ndk (rNdk). As for the determination of IgG responses, rNdk induced vigorous IgG production, especially higher in IgG2a compared to IgG1 with titers of 5.2 and 4.8, respectively, whereas titers of these in mice immunized with MBP were 2.4 of IgG2a and 2.6 of IgG1. The analysis of cytokine has revealed that rNdk can strongly induce production of IFN-γ as well as proinflammatory cytokines (TNF, MCP1 and IL-6) but not much IL-10, suggesting rNdk elicited predominantly cell-mediated immune responses. Furthermore, the spleen proliferation and bacterial burden in the spleen of rNdk immunized mice were significantly lower than those of MBP-immunized mice against virulent B. abortus challenge (P < 0.01). Conclusionly, rNdk immunization enables to elicit both of the humoral and cellular response, ultimately enhancing protection level in experimental mice, suggesting that rNdk of B. abortus might be a useful candidate for subunit vaccine for brucellosis in animals.

  8. Amoebal host range, host-free survival and disinfection susceptibility of environmental Chlamydiae as compared to Chlamydia trachomatis.

    PubMed

    Coulon, Céline; Eterpi, Mickael; Greub, Gilbert; Collignon, Anne; McDonnell, Gerald; Thomas, Vincent

    2012-04-01

    The term 'Chlamydia-like organisms' encompasses obligate intracellular bacterial species phylogenetically close to Chlamydiaceae. Most are associated with free-living amoebae, and several could be responsible for respiratory tract infections and abortion in human and animals. Despite increasing concern about their pathogenic role, the prevalence, biodiversity and ecology of Chlamydia-related bacteria still remain largely unknown. In this study, six members of the Chlamydiales were tested, including Parachlamydia acanthamoebae (two different strains), Protochlamydia naegleriophila, Waddlia chondrophila, Criblamydia sequanensis and Chlamydia trachomatis as a reference. Intracellular growth was tested in 11 different Acanthamoeba strains, demonstrating significant differences in host susceptibilities to infection depending on strains investigated. Survival of host-free bacteria in suspension or dried onto surfaces was also explored, demonstrating that Chlamydia-like organisms present better survival capacity than C. trachomatis. Longer survival times were observed for bacteria suspended in rich culture medium, with survivors being detected after 10 weeks incubation. We also tested susceptibility of host-free Chlamydia-like organisms to several disinfection treatments. Each chemical biocide tested reduced viability of host-free Chlamydia by more than 4 logs. Conversely, all Chlamydia-like organisms tested resisted exposure at 55 °C for 10 min, while C. trachomatis was completely inactivated. PMID:22141597

  9. Biotyping and Genotyping (MLVA16) of Brucella abortus Isolated from Cattle in Brazil, 1977 to 2008

    PubMed Central

    Minharro, Sílvia; Silva Mol, Juliana P.; Dorneles, Elaine M. S.; Pauletti, Rebeca B.; Neubauer, Heinrich; Melzer, Falk; Poester, Fernando P.; Dasso, Maurício G.; Pinheiro, Elaine S.; Soares Filho, Paulo M.; Santos, Renato L.; Heinemann, Marcos B.; Lage, Andrey P.

    2013-01-01

    Brucellosis is a worldwide distributed zoonosis that causes important economic losses to animal production. In Brazil, information on the distribution of biovars and genotypes of Brucella spp. is scarce or unavailable. This study aimed (i) to biotype and genotype 137 Brazilian cattle isolates (from 1977 to 2008) of B. abortus and (ii) to analyze their distribution. B. abortus biovars 1, 2 and 3 (subgroup 3b) were confirmed and biovars 4 and 6 were first described in Brazil. Genotyping by the panel 1 revealed two groups, one clustering around genotype 40 and another around genotype 28. Panels 2A and 2B disclosed a high diversity among Brazilian B. abortus strains. Eighty-nine genotypes were found by MLVA16. MLVA16 panel 1 and 2 showed geographic clustering of some genotypes. Biotyping and MLVA16 genotyping of Brazilian B. abortus isolates were useful to better understand the epidemiology of bovine brucellosis in the region. PMID:24324670

  10. The laboratory diagnosis of Chlamydia trachomatis infections.

    PubMed

    Chernesky, Max A

    2005-01-01

    Lower genital tract infections with Chlamydia trachomatis are predominantly asymptomatic in men and women. Diagnostic technology has provided several approaches to the diagnosis of C trachomatis. Outside of cells, Chlamydia can die or degrade without optimal storage and transportation. Because some of the other assays perform better on certain specimen types, it is important for laboratories to recognize these differences and provide advice to physicians and nurses collecting patient specimens, with the objective of diagnosing lower genital tract infections to prevent transmission and upper tract damage. Most invasive specimens, such as cervical or urethral swabs, may be collected for culture, antigen or nucleic acid detection. Noninvasive samples such as first-void urine and vaginal swabs can be easily collected by the patient; these samples must be tested by more sensitive nucleic acid amplification tests. These newer investigative strategies should enable implementation of screening programs to identify and treat partners. Serology has not been particularly useful for the diagnosis of acute C trachomatis infections in adults. Presently, it appears that antibiotic-resistant C trachomatis is not a clinical problem. Laboratories providing C trachomatis diagnosis require participation in continuous quality improvement programs.

  11. Genome Sequences of 11 Brucella abortus Isolates from Persistently Infected Italian Regions.

    PubMed

    Garofolo, Giuliano; Foster, Jeffrey T; Drees, Kevin; Zilli, Katiuscia; Platone, Ilenia; Ancora, Massimo; Cammà, Cesare; De Massis, Fabrizio; Calistri, Paolo; Di Giannatale, Elisabetta

    2015-01-01

    Bovine brucellosis, typically caused by Brucella abortus, has been eradicated from much of the developed world. However, the disease remains prevalent in southern Italy, persisting as a public and livestock health concern. We report here the whole-genome sequences of 11 isolates from cattle (Bos taurus) and water buffalo (Bubalus bubalis) that are representative of the current genetic diversity of B. abortus lineages circulating in Italy. PMID:26679575

  12. Genome Sequences of 11 Brucella abortus Isolates from Persistently Infected Italian Regions

    PubMed Central

    Foster, Jeffrey T.; Drees, Kevin; Zilli, Katiuscia; Platone, Ilenia; Ancora, Massimo; Cammà, Cesare; De Massis, Fabrizio; Calistri, Paolo; Di Giannatale, Elisabetta

    2015-01-01

    Bovine brucellosis, typically caused by Brucella abortus, has been eradicated from much of the developed world. However, the disease remains prevalent in southern Italy, persisting as a public and livestock health concern. We report here the whole-genome sequences of 11 isolates from cattle (Bos taurus) and water buffalo (Bubalus bubalis) that are representative of the current genetic diversity of B. abortus lineages circulating in Italy. PMID:26679575

  13. Genome Sequences of 11 Brucella abortus Isolates from Persistently Infected Italian Regions.

    PubMed

    Garofolo, Giuliano; Foster, Jeffrey T; Drees, Kevin; Zilli, Katiuscia; Platone, Ilenia; Ancora, Massimo; Cammà, Cesare; De Massis, Fabrizio; Calistri, Paolo; Di Giannatale, Elisabetta

    2015-12-17

    Bovine brucellosis, typically caused by Brucella abortus, has been eradicated from much of the developed world. However, the disease remains prevalent in southern Italy, persisting as a public and livestock health concern. We report here the whole-genome sequences of 11 isolates from cattle (Bos taurus) and water buffalo (Bubalus bubalis) that are representative of the current genetic diversity of B. abortus lineages circulating in Italy.

  14. [Detection of a clonal complex with Brucella abortus biovar 2 genotype as founder in B. abortus isolates from Argentina].

    PubMed

    Hollender, Daiana; Conde, Sandra B; Salustio, Eduardo; Samartino, Luis E

    2013-01-01

    Brucella abortus is the causative agent of bovine brucellosis, a worldwide zoonosis. Up to date, eight biovars of B. abortus have been described. In Argentina, biovar 1 is the most frequently isolated. However, biovar 2, which is more pathogenic than biovar 1, is also found. Molecular methods for subtyping isolates are necessary for allowing epidemiological surveillance and control of eradication programs. Due to the genetic homogeneity of the genus Brucella, the development of molecular typing tools has been difficult. The publication of microorganism genomes facilitates the design of this approach. The aim of this work was to employ a Multiple Locus VNTR Analysis (MLVA) scheme for strains from Argentina isolated in our laboratory. From the 56 isolates analyzed, 47 different genotypic profiles were obtained. All the strains typed as biovar 2 showed the same profile. This scheme allowed assigning each isolate to the biovar it belongs to. All the genotypes were related using the goeBURST analysis and biovar 2 was proposed as founder.

  15. Brucella abortus Invasion of Synoviocytes Inhibits Apoptosis and Induces Bone Resorption through RANKL Expression

    PubMed Central

    Scian, Romina; Barrionuevo, Paula; Rodriguez, Ana María; Arriola Benitez, Paula Constanza; García Samartino, Clara; Fossati, Carlos Alberto; Giambartolomei, Guillermo Hernán

    2013-01-01

    Arthritis is one of the most common complications of human active brucellosis, but its pathogenic mechanisms have not been completely elucidated. In this paper, we describe the role of synoviocytes in the pathogenesis of brucellar arthritis. Our results indicate that Brucella abortus infection inhibited synoviocyte apoptosis through the upregulation of antiapoptotic factors (cIAP-2, clusterin, livin, and P21/CIP/CDNK1A). In contrast, infection did not change the expression of proteins that have been involved in apoptosis induction such as Bad, Bax, cleaved procaspase 3, CytC, and TRAIL, among others; or their expression was reduced, as occurs in the case of P-p53(S15). In addition, B. abortus infection induced upregulation of adhesion molecules (CD54 and CD106), and the adhesion of monocytes and neutrophils to infected synoviocytes was significantly higher than to uninfected cells. Despite this increased adhesion, B. abortus-infected synoviocytes were able to inhibit apoptosis induced by supernatants from B. abortus-infected monocytes and neutrophils. Moreover, B. abortus infection increased soluble and membrane RANKL expression in synoviocytes that further induced monocytes to undergo osteoclastogenesis. The results presented here shed light on how the interactions of B. abortus with synovial fibroblasts may have an important role in the pathogenesis of brucellar arthritis. PMID:23509146

  16. Characterization of culture supernatant proteins from Brucella abortus and its protection effects against murine brucellosis.

    PubMed

    Lee, Jin Ju; Lim, Jeong Ju; Kim, Dae Geun; Simborio, Hannah Leah; Kim, Dong Hyeok; Reyes, Alisha Wehdnesday Bernardo; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2014-09-01

    In this study, we characterized the secreted proteins of Brucella abortus into the enriched media under the bacterial laboratory growth condition and investigated the pathogenic importance of culture supernatant (CS) proteins to B. abortus infection. CS proteins from stationary phase were concentrated and analyzed using 2D electrophoresis. In MALDI TOF/TOF analysis, more than 27 proteins including CuZn SOD, Dps, Tat, OMPs, Adh, LivF, Tuf, SucC, GroEL and DnaK were identified. Cytotoxic effects of CS proteins were found to increase in a dose-dependent manner in RAW 264.7 cells. Upon B. abortus challenge into phagocytes, however, CS proteins pre-treated cells exhibited lower bacterial uptake and intracellular replication compared to untreated cells. Immunization with CS proteins induced a strong humoral and cell mediated immune responses and exhibited significant higher degree of protection against virulence of B. abortus infection compared to mice immunized with Brucella broth protein (BBP). Taken together, these results indicate that B. abortus secreted a number of soluble immunogenic proteins under laboratory culture condition, which can promote antibody production resulted in enhancing host defense against to subsequently bacterial infection. Moreover, further analysis of CS proteins may help to understand the pathogenic mechanism of B. abortus infection and host-pathogen interaction.

  17. Inositol-Requiring Enzyme 1-Dependent Activation of AMPK Promotes Brucella abortus Intracellular Growth

    PubMed Central

    Liu, Ning; Li, Yingying; Dong, Chunyan; Xu, Xiaohan; Wei, Pan; Sun, Wanchun

    2016-01-01

    ABSTRACT AMP-activated protein kinase (AMPK) is a serine/threonine kinase that is well conserved during evolution. AMPK activation inhibits production of reactive oxygen species (ROS) in cells via suppression of NADPH oxidase. However, the role of AMPK during the process of Brucella infection remains unknown. Our data demonstrate that B. abortus infection induces AMPK activation in HeLa cells in a time-dependent manner. The known AMPK kinases LKB1, CAMKKβ, and TAK1 are not required for the activation of AMPK by B. abortus infection. Instead, this activation is dependent on the RNase activity of inositol-requiring enzyme 1 (IRE1). Moreover, we also found that B. abortus infection-induced IRE1-dependent activation of AMPK promotes B. abortus intracellular growth with peritoneal macrophages via suppression of NADPH-derived ROS production. IMPORTANCE Previous studies showed that B. abortus infection does not promote any oxidative burst regulated by NADPH oxidase. However, the underlying mechanism remains elusive. We report for the first time that AMPK activation caused by B. abortus infection plays important role in NADPH oxidase-derived ROS production. PMID:26755628

  18. Mouse Study Offers Hope for Vaccine Against Chlamydia

    MedlinePlus

    ... page: https://medlineplus.gov/news/fullstory_160004.html Mouse Study Offers Hope for Vaccine Against Chlamydia Bacteria's ... 2016 (HealthDay News) -- A new Canadian study with mice suggests there is hope for a vaccine to ...

  19. Application of DNA chip scanning technology for automatic detection of Chlamydia trachomatis and Chlamydia pneumoniae inclusions.

    PubMed

    Bogdanov, Anita; Endrész, Valeria; Urbán, Szabolcs; Lantos, Ildikó; Deák, Judit; Burián, Katalin; Önder, Kamil; Ayaydin, Ferhan; Balázs, Péter; Virok, Dezso P

    2014-01-01

    Chlamydiae are obligate intracellular bacteria that propagate in the inclusion, a specific niche inside the host cell. The standard method for counting chlamydiae is immunofluorescent staining and manual counting of chlamydial inclusions. High- or medium-throughput estimation of the reduction in chlamydial inclusions should be the basis of testing antichlamydial compounds and other drugs that positively or negatively influence chlamydial growth, yet low-throughput manual counting is the common approach. To overcome the time-consuming and subjective manual counting, we developed an automatic inclusion-counting system based on a commercially available DNA chip scanner. Fluorescently labeled inclusions are detected by the scanner, and the image is processed by ChlamyCount, a custom plug-in of the ImageJ software environment. ChlamyCount was able to measure the inclusion counts over a 1-log-unit dynamic range with a high correlation to the theoretical counts. ChlamyCount was capable of accurately determining the MICs of the novel antimicrobial compound PCC00213 and the already known antichlamydial antibiotics moxifloxacin and tetracycline. ChlamyCount was also able to measure the chlamydial growth-altering effect of drugs that influence host-bacterium interaction, such as gamma interferon, DEAE-dextran, and cycloheximide. ChlamyCount is an easily adaptable system for testing antichlamydial antimicrobials and other compounds that influence Chlamydia-host interactions. PMID:24189259

  20. Differences in Treatment of Chlamydia trachomatis by Ambulatory Care Setting.

    PubMed

    Pearson, William S; Gift, Thomas L; Leichliter, Jami S; Jenkins, Wiley D

    2015-12-01

    Chlamydia trachomatis (CT) is the most commonly reported sexually transmitted infection (STI) in the US and timely, correct treatment can reduce CT transmission and sequelae. Emergency departments (ED) are an important location for diagnosing STIs. This study compared recommended treatment of CT in EDs to treatment in physician offices. Five years of data (2006-2010) were analyzed from the National Ambulatory Medical Care Survey, and the National Hospital Ambulatory Medical Care Surveys (NHAMCS), including the Outpatient survey (NHAMCS-OPD) and Emergency Department survey (NHAMCS-ED). All visits with a CT diagnosis and those with a diagnosis of unspecified venereal disease were selected for analysis. Differences in receipt of recommended treatments were compared between visits to physician offices and emergency departments using Chi square tests and logistic regression models. During the 5 year period, approximately 3.2 million ambulatory care visits had diagnosed CT or an unspecified venereal disease. A greater proportion of visits to EDs received the recommended treatment for CT compared to visits to physician offices (66.1 vs. 44.9 %, p < .01). When controlling for patients' age, sex and race/ethnicity, those presenting to the ED with CT were more likely to receive the recommended antibiotic treatment than patients presenting to a physician's office (OR 2.16; 95 % CI 1.04-4.48). This effect was attenuated when further controlling for patients' expected source of payment. These analyses demonstrate differences in the treatment of CT by ambulatory care setting as well as opportunities for increasing use of recommended treatments for diagnosed cases of this important STI. PMID:25940936

  1. A LysR-family transcriptional regulator required for virulence in Brucella abortus is highly conserved among the α-proteobacteria.

    PubMed

    Sheehan, Lauren M; Budnick, James A; Blanchard, Catlyn; Dunman, Paul M; Caswell, Clayton C

    2015-10-01

    Small RNAs are principal elements of bacterial gene regulation and physiology. Two small RNAs in Brucella abortus, AbcR1 and AbcR2, are required for wild-type virulence. Examination of the abcR loci revealed the presence of a gene encoding a LysR-type transcriptional regulator flanking abcR2 on chromosome 1. Deletion of this lysR gene (bab1_1517) resulted in the complete loss of abcR2 expression while no difference in abcR1 expression was observed. The B. abortus bab1_1517 mutant strain was significantly attenuated in macrophages and mice, and bab1_1517 was subsequently named vtlR for virulence-associated transcriptional LysR-family regulator. Microarray analysis revealed three additional genes encoding small hypothetical proteins also under the control of VtlR. Electrophoretic mobility shift assays demonstrated that VtlR binds directly to the promoter regions of abcR2 and the three hypothetical protein-encoding genes, and DNase I footprint analysis identified the specific nucleotide sequence in these promoters that VtlR binds to and drives gene expression. Strikingly, orthologs of VtlR are encoded in a wide range of host-associated α-proteobacteria, and it is likely that the VtlR genetic system represents a common regulatory circuit critical for host-bacterium interactions.

  2. Ivermectin inhibits growth of Chlamydia trachomatis in epithelial cells.

    PubMed

    Pettengill, Matthew A; Lam, Verissa W; Ollawa, Ikechukwu; Marques-da-Silva, Camila; Ojcius, David M

    2012-01-01

    Ivermectin is currently approved for treatment of both clinical and veterinary infections by nematodes, including Onchocerca cervicalis in horses and Onchocerca volvulus in humans. However, ivermectin has never been shown to be effective against bacterial pathogens. Here we show that ivermectin also inhibits infection of epithelial cells by the bacterial pathogen, Chlamydia trachomatis, at doses that could be envisioned clinically for sexually-transmitted or ocular infections by Chlamydia. PMID:23119027

  3. Chlamydia trachomatis infection in African American women who exclusively have sex with women.

    PubMed

    Muzny, Christina A; Kapil, Richa; Austin, Erika L; Brown, LaDraka; Hook, Edward W; Geisler, William M

    2016-10-01

    Little is known about whether Chlamydia trachomatis can be sexually transmitted between women or how often it occurs in women who have sex with women (WSW). We investigated Chlamydia trachomatis prevalence and serum Chlamydia trachomatis-specific antibody responses among African American WSW who reported a lifetime history of sex only with women (exclusive WSW) (n = 21) vs. an age-matched group of women reporting sex with women and men (WSWM) (n = 42). Participants completed a survey, underwent a pelvic examination in which a cervical swab was collected for Chlamydia trachomatis nucleic acid amplification testing (NAAT), and had serum tested for anti-Chlamydia trachomatis IgG1 and IgG3 antibodies using a Chlamydia trachomatis elementary body-based ELISA. No exclusive WSW had a positive Chlamydia trachomatis NAAT vs. 5 (11.9%) WSWM having a positive Chlamydia trachomatis NAAT (p = 0.16). Compared with WSWM, WSW were significantly less likely to be Chlamydia trachomatis seropositive (7 [33.3%] vs. 29 [69%], p = 0.007). Among Chlamydia trachomatis seropositive women, all were seropositive by IgG1, and the magnitude of Chlamydia trachomatis-specific IgG1 responses did not differ in Chlamydia trachomatis-seropositive WSW vs. WSWM. In conclusion, Chlamydia trachomatis seropositivity was relatively common in exclusive African American WSW, though significantly less common than in African American WSWM.

  4. Chlamydiae interaction with the endoplasmic reticulum: contact, function and consequences.

    PubMed

    Derré, Isabelle

    2015-07-01

    Chlamydiae and chlamydiae-related organisms are obligate intracellular bacterial pathogens. They reside in a membrane-bound compartment termed the inclusion and have evolved sophisticated mechanisms to interact with cellular organelles. This review focuses on the nature, the function(s) and the consequences of chlamydiae-inclusion interaction with the endoplasmic reticulum (ER). The inclusion membrane establishes very close contact with the ER at specific sites termed ER-inclusion membrane contact sites (MCSs). These MCSs are constituted of a specific set of factors, including the C. trachomatis effector protein IncD and the host cell proteins CERT and VAPA/B. Because CERT and VAPA/B have a demonstrated role in the non-vesicular trafficking of lipids between the ER and the Golgi, it was proposed that Chlamydia establish MCSs with the ER to acquire host lipids. However, the recruitment of additional factors to ER-inclusion MCSs, such as the ER calcium sensor STIM1, may suggest additional functions unrelated to lipid acquisition. Finally, chlamydiae interaction with the ER appears to induce the ER stress response, but this response is quickly dampened by chlamydiae to promote host cell survival. PMID:25930206

  5. Significant roles played by IL-10 in Chlamydia infections.

    PubMed

    Hakimi, Hamid; Zare-Bidaki, Mohammad; Zainodini, Nahid; Assar, Shokrollah; Arababadi, Mohammad Kazemi

    2014-06-01

    Chlamydia species are obligate intracellular parasites which cause usually asymptomatic genital tract infections and also are associated with several complications. Previous studies demonstrated that immune responses to Chlamydia species are different and the diseases will be limited to some cases. Additionally, Chlamydia species are able to modulate immune responses via regulating expression of some immune system molecules including cytokines. IL-10, as the main anti-inflammatory cytokine, plays important roles in the induction of immune-tolerance against self-antigen and also immune-homeostasis after microbe elimination. Furthermore, it has been documented that ectopic expression of IL-10 is associated with several chronic infectious diseases. Therefore, it can be hypothesized that changes in the regulation of this cytokine can be associated with infection with several species of Chlamydia and their associated complications. This review collected the recent information regarding the association and relationship of IL-10 with Chlamydia infections. Another aim of this review article is to address recent data regarding the association of genetic variations (polymorphisms) of IL-10 and Chlamydia infections.

  6. Risks of Brucella abortus spillover in the Greater Yellowstone area.

    PubMed

    Schumaker, B

    2013-04-01

    Recurrent spillover of Brucella abortus from wildlife reservoirs to domestic cattle in the Greater Yellowstone Area (GYA) has prevented the United States from completely eradicating bovine brucellosis. Risks to cattle are a function of the size and location of wildlife and livestock populations, the degree and nature of spatio-temporal interactions between the various hosts, the level of disease in wildlife, and the susceptibility of livestock herds. While the brucellosis prevalence in wild, free-ranging GYA bison (Bison bison) is high, current management actions have successfully limited contact between bison and cattle. Under current management practices, the risks to cattle in the GYA are predominantly from wild elk (Cervus elaphus). Intra- and inter-species transmission events, while uncommon, are nevertheless crucial for the maintenance of brucellosis in the GYA. Future management actions should focus on decreasing elk herd densities and group sizes and on understanding the behavioural and environmental drivers that result in co-mingling that makes transmission possible.

  7. Isolation, purification, and partial characterization of Brucella abortus matrix protein.

    PubMed

    Moriyon, I; Berman, D T

    1983-01-01

    Peptidoglycan sacculi with peptidoglycan-associated proteins were prepared from cell envelopes of Brucella abortus by extraction with sodium dodecyl sulfate (SDS) at 50 degrees C. On extraction of these preparations with SDS at 100 degrees C, a protein was obtained whose removal from peptidoglycan was confirmed by electron microscopy. Incubation of the 50 degrees C SDS-extracted cell envelopes with 50 mM MgCl2 in SDS-2-beta-mercaptoethanol at 37 degrees C also extracted the protein, along with lipopolysaccharide. At temperatures below 60 degrees C, the protein did not bind SDS strongly and had an apparent molecular weight greater than 92,000 in SDS-polyacrylamide gel electrophoresis. At higher temperatures, SDS bound strongly, and the apparent molecular weight was 38,000. Urea at 5 M did not alter the electrophoretic mobility of this 38,000-molecular-weight form. Immunoelectrophoresis in detergents with antisera to cell envelopes, carbohydrate staining of SDS-polyacrylamide gels, and production of anti-lipopolysaccharide antibodies by mice immunized with the purified protein indicated that lipopolysaccharide was present in free and protein-bound forms. Sequential gel filtration in SDS-EDTA and SDS-NaCl removed most lipopolysaccharide. After further purification by preparative SDS-polyacrylamide gel electrophoresis, a gas-liquid chromatographic analysis showed residual lipid tightly associated with the protein. The results suggested that the interactions between matrix proteins and other outer membrane components are stronger in B. abortus than in Escherichia coli, which was used as a control throughout. PMID:6401696

  8. Isolation, purification, and partial characterization of Brucella abortus matrix protein.

    PubMed

    Moriyon, I; Berman, D T

    1983-01-01

    Peptidoglycan sacculi with peptidoglycan-associated proteins were prepared from cell envelopes of Brucella abortus by extraction with sodium dodecyl sulfate (SDS) at 50 degrees C. On extraction of these preparations with SDS at 100 degrees C, a protein was obtained whose removal from peptidoglycan was confirmed by electron microscopy. Incubation of the 50 degrees C SDS-extracted cell envelopes with 50 mM MgCl2 in SDS-2-beta-mercaptoethanol at 37 degrees C also extracted the protein, along with lipopolysaccharide. At temperatures below 60 degrees C, the protein did not bind SDS strongly and had an apparent molecular weight greater than 92,000 in SDS-polyacrylamide gel electrophoresis. At higher temperatures, SDS bound strongly, and the apparent molecular weight was 38,000. Urea at 5 M did not alter the electrophoretic mobility of this 38,000-molecular-weight form. Immunoelectrophoresis in detergents with antisera to cell envelopes, carbohydrate staining of SDS-polyacrylamide gels, and production of anti-lipopolysaccharide antibodies by mice immunized with the purified protein indicated that lipopolysaccharide was present in free and protein-bound forms. Sequential gel filtration in SDS-EDTA and SDS-NaCl removed most lipopolysaccharide. After further purification by preparative SDS-polyacrylamide gel electrophoresis, a gas-liquid chromatographic analysis showed residual lipid tightly associated with the protein. The results suggested that the interactions between matrix proteins and other outer membrane components are stronger in B. abortus than in Escherichia coli, which was used as a control throughout.

  9. [Current aspects of Chlamydia trachomatis infection].

    PubMed

    de Barbeyrac, Bertille

    2013-04-01

    The number of detection and diagnosis of urogenital infections with Chlamydia trachomatis is increasing among both men and women. Three-quarters involve young people between 15 and 24 years. Infection, often asymptomatic, is more common in women. It is necessary to identify it to avoid complications.The number of rectal lymphogranuloma venereum (LGV) is also growing. The affected patients are homo/bisexuel men frequently co-infected with HIV. Nucleic acid amplification tests (NAATs) are the tests of choice to the diagnosis of C. trachomatis infection regardless of the clinical situation. Most of tests simultaneously detect C. trachomatis and Neisseria gonorrhoeae. The recommended treatment regimens for a non-complicated infection to C. trachomatis is azithromycin 1g orally in a single dose or doxycyline 100 mg orally twice a day for 7 days. Doxycyclin for 21 days remains the treatment of choice for LGV. Patients should be instructed to refer their sex partners for treatment. PMID:23419460

  10. [Chlamydia infection in neonates and infants].

    PubMed

    Sarlangue, J; Castella, C

    2005-04-01

    Incidence of chlamydial infection depends on maternal colonization during pregnancy, which is different in each population. The transmission is not obligatory but when present, it occurs at birth through the genital tractus. Chlamydia trachomatis infection is the first cause of neonatal conjunctivitis, with no influence of eye lotion application at birth. C. trachomatis is also responsible for interstitial pneumonia with possible consequences on the lung function. The laboratory diagnosis relies on the identification of intracellular bacteria in patient samples by the mean of culture or PCR. Systemic antibiotherapy by macrolides is always necessary, with local application in the case of conjunctivitis. The key point is the detection of colonization of pregnant women with identified risk factors. In positive case, oral treatment of both parents is recommended.

  11. Role of Chlamydia in Multiple Sclerosis.

    PubMed

    Ivanova, M V; Kolkova, N I; Morgunova, E Yu; Pashko, Yu P; Zigangirova, N A; Zakharova, M N

    2015-09-01

    Chlamydia and antibodies to them were detected by serological, molecular biological, and culture methods in the sera and cerebrospinal fluid of patients with multiple sclerosis and in the reference groups of subjects without neurological diseases. Correlations between the agent presence in the biological fluids of patients and clinical characteristics of the disease were analyzed. C. pneumoniae were more incident in the biological liquids of patients with multiple sclerosis than in healthy volunteers. On the other hand, the incidence of the agent in the patients was not high and its presence did not correlate with the clinical manifestations. C. trachomatis was equally rare in the patients and volunteers. The studies indicated the existence of a group of patients infected by C. pneumoniae in the cohort of patients with multiple sclerosis, but the impact of this agent for the disease course remains unclear.

  12. [Chlamydia pneumoniae infections--diagnostic methods].

    PubMed

    Stepień, Ewa; Pieniazek, Piotr; Branicka, Agnieszka; Bozek, Maria

    2002-01-01

    Gram-negative bacteria Chlamydia pneumonia was found in 1989 to cause acute and chronic respiratory tract infections. This agent has been as well associated with other disease: atherogenesis and coronary heart disease. This study is aimed both at making an introduction to the issues related to C. pneumoniae diagnosis and presenting contemporary laboratory methods. Given the limitations of traditional diagnostics methods, serodiagnosis (EIA) and nucleic acids amplification (PCR, hybridisation) provide the most convincing evidence of C. pneumoniae infections. Culture and direct fluorescence antibody (DFA) may be useful in confirming these results. A variety of methods applied can provide an opportunity to detect bacteria in different clinical samples--incl. sputum, nasopharyngeal and throat swabs, bronchoalveolar lavage (BAL) and tissues from biopsy and autopsy. PMID:12184026

  13. Caspase-2-Dependent Dendritic Cell Death, Maturation, and Priming of T Cells in Response to Brucella abortus Infection

    PubMed Central

    Li, Xinna; He, Yongqun

    2012-01-01

    Smooth virulent Brucella abortus strain 2308 (S2308) causes zoonotic brucellosis in cattle and humans. Rough B. abortus strain RB51, derived from S2308, is a live attenuated cattle vaccine strain licensed in the USA and many other countries. Our previous report indicated that RB51, but not S2308, induces a caspase-2-dependent apoptotic and necrotic macrophage cell death. Dendritic cells (DCs) are professional antigen presenting cells critical for bridging innate and adaptive immune responses. In contrast to Brucella-infected macrophages, here we report that S2308 induced higher levels of apoptotic and necrotic cell death in wild type bone marrow-derived DCs (WT BMDCs) than RB51. The RB51 and S2308-induced BMDC cell death was regulated by caspase-2, indicated by the minimal cell death in RB51 and S2308-infected BMDCs isolated from caspase-2 knockout mice (Casp2KO BMDCs). More S2308 bacteria were taken up by Casp2KO BMDCs than wild type BMDCs. Higher levels of S2308 and RB51 cells were found in infected Casp2KO BMDCs compared to infected WT BMDCs at different time points. RB51-infected wild type BMDCs were mature and activated as shown by significantly up-regulated expression of CD40, CD80, CD86, MHC-I, and MHC-II. RB51 induced the production of cytokines TNF-α, IL-6, IFN-γ and IL12/IL23p40 in infected BMDCs. RB51-infected WT BMDCs also stimulated the proliferation of CD4+ and CD8+ T cells compared to uninfected WT BMDCs. However, the maturation, activation, and cytokine secretion are significantly impaired in Casp2KO BMDCs infected with RB51 or Salmonella (control). S2308-infected WT and Casp2KO BMDCs were not activated and could not induce cytokine production. These results demonstrated that virulent smooth strain S2308 induced more apoptotic and necrotic dendritic cell death than live attenuated rough vaccine strain RB51; however, RB51, but not its parent strain S2308, induced caspase-2-mediated DC maturation, cytokine production, antigen presentation, and T

  14. Exploring Chlamydia Positivity among Females on College Campuses, 2008-2010

    ERIC Educational Resources Information Center

    Habel, Melissa A.; Leichliter, Jami S.; Torrone, Elizabeth

    2016-01-01

    Objective: Describe chlamydia positivity among young women tested at college health centers by student characteristics: age, race/ethnicity, and institution type. Participants: During 2008-2010, colleges participating in a national infertility prevention program provided chlamydia testing data from females aged 18-24. Methods: Chlamydia positivity…

  15. Vaccination of elk (Cervus canadensis) with Brucella abortus strain RB51 overexpressing superoxide dismutase and glycosyltransferase genes does not induce adequate protection against experimental brucella abortus challenge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In recent years, elk (Cervus canadensis) have been implicated as the source of Brucella abortus infection for numerous cattle herds in the Greater Yellowstone Area (GYA). In the face of environmental and ecological changes on the landscape, the range of infected elk is expanding. Consequently, the d...

  16. A B lymphocyte mitogen is a Brucella abortus virulence factor required for persistent infection

    PubMed Central

    Spera, Juan Manuel; Ugalde, Juan Esteban; Mucci, Juan; Comerci, Diego J.; Ugalde, Rodolfo Augusto

    2006-01-01

    Microbial pathogens with the ability to establish chronic infections have evolved strategies to actively modulate the host immune response. Brucellosis is a disease caused by a Gram-negative intracellular pathogen that if not treated during the initial phase of the infection becomes chronic as the bacteria persist for the lifespan of the host. How this pathogen and others achieve this action is a largely unanswered question. We report here the identification of a Brucella abortus gene (prpA) directly involved in the immune modulation of the host. PrpA belongs to the proline-racemase family and elicits a B lymphocyte polyclonal activation that depends on the integrity of its proline-racemase catalytic site. Stimulation of splenocytes with PrpA also results in IL-10 secretion. Construction of a B. abortus-prpA mutant allowed us to assess the contribution of PrpA to the infection process. Mice infected with B. abortus induced an early and transient nonresponsive status of splenocytes to both Escherichia coli LPS and ConA. This phenomenon was not observed when mice were infected with a B. abortus-prpA mutant. Moreover, the B. abortus-prpA mutant had a reduced capacity to establish a chronic infection in mice. We propose that an early and transient nonresponsive immune condition of the host mediated by this B cell polyclonal activator is required for establishing a successful chronic infection by Brucella. PMID:17053080

  17. N-Formyl-Perosamine Surface Homopolysaccharides Hinder the Recognition of Brucella abortus by Mouse Neutrophils.

    PubMed

    Mora-Cartín, Ricardo; Chacón-Díaz, Carlos; Gutiérrez-Jiménez, Cristina; Gurdián-Murillo, Stephany; Lomonte, Bruno; Chaves-Olarte, Esteban; Barquero-Calvo, Elías; Moreno, Edgardo

    2016-06-01

    Brucella abortus is an intracellular pathogen of monocytes, macrophages, dendritic cells, and placental trophoblasts. This bacterium causes a chronic disease in bovines and in humans. In these hosts, the bacterium also invades neutrophils; however, it fails to replicate and just resists the killing action of these leukocytes without inducing significant activation or neutrophilia. Moreover, B. abortus causes the premature cell death of human neutrophils. In the murine model, the bacterium is found within macrophages and dendritic cells at early times of infection but seldom in neutrophils. Based on this observation, we explored the interaction of mouse neutrophils with B. abortus In contrast to human, dog, and bovine neutrophils, naive mouse neutrophils fail to recognize smooth B. abortus bacteria at early stages of infection. Murine normal serum components do not opsonize smooth Brucella strains, and neutrophil phagocytosis is achieved only after the appearance of antibodies. Alternatively, mouse normal serum is capable of opsonizing rough Brucella mutants. Despite this, neutrophils still fail to kill Brucella, and the bacterium induces cell death of murine leukocytes. In addition, mouse serum does not opsonize Yersinia enterocolitica O:9, a bacterium displaying the same surface polysaccharide antigen as smooth B. abortus Therefore, the lack of murine serum opsonization and absence of murine neutrophil recognition are specific, and the molecules responsible for the Brucella camouflage are N-formyl-perosamine surface homopolysaccharides. Although the mouse is a valuable model for understanding the immunobiology of brucellosis, direct extrapolation from one animal system to another has to be undertaken with caution. PMID:27001541

  18. Rapid and specific identification of Brucella abortus using the loop-mediated isothermal amplification (LAMP) assay.

    PubMed

    Kang, Sung-Il; Her, Moon; Kim, Ji-Yeon; Lee, Jin Ju; Lee, Kichan; Sung, So-Ra; Jung, Suk Chan

    2015-06-01

    A rapid and accurate diagnosis of brucellosis is required to reduce and prevent the spread of disease among animals and the risk of transfer to humans. In this study, a Brucella abortus-specific (Ba) LAMP assay was developed, that had six primers designed from the BruAb2_0168 region of chromosome I. The specificity of this LAMP assay was confirmed with Brucella reference strains, B. abortus vaccine strains, B. abortus isolates and phylogenetically or serologically related strains. The detection limit of target DNA was up to 20 fg/μl within 60 min. The sensitivity of the new LAMP assay was equal to or slightly higher than other PCR based assays. Moreover, this Ba-LAMP assay could specifically amplify all B. abortus biovars compared to previous PCR assays. To our knowledge, this is the first report of specific detection of B. abortus using a LAMP assay. The Ba-LAMP assay can offer a rapid, sensitive and accurate diagnosis of bovine brucellosis in the field.

  19. The role of TREM-2 in internalization and intracellular survival of Brucella abortus in murine macrophages.

    PubMed

    Wei, Pan; Lu, Qiang; Cui, Guimei; Guan, Zhenhong; Yang, Li; Sun, Changjiang; Sun, Wanchun; Peng, Qisheng

    2015-02-15

    Triggering receptor expressed on myeloid cells-2 (TREM-2) is a cell surface receptor primarily expressed on macrophages and dendritic cells. TREM-2 functions as a phagocytic receptor for bacteria as well as an inhibitor of Toll like receptors (TLR) induced inflammatory cytokines. However, the role of TREM-2 in Brucella intracellular growth remains unknown. To investigate whether TREM-2 is involved in Brucella intracellular survival, we chose bone marrow derived macrophages (BMDMs), in which TREM-2 is stably expressed, as cell model. Colony formation Units (CFUs) assay suggests that TREM-2 is involved in the internalization of Brucella abortus (B. abortus) by macrophages, while silencing of TREM-2 decreases intracellular survival of B. abortus. To further study the underlying mechanisms of TREM-2-mediated bacterial intracellular survival, we examined the activation of B. abortus-infected macrophages through determining the kinetics of activation of the three MAPKs, including ERK, JNK and p38, and measuring TNFα production in response to lipopolysaccharide (LPS) of Brucella (BrLPS) or B. abortus stimulation. Our data show that TREM-2 deficiency promotes activation of Brucella-infected macrophages. Moreover, our data also demonstrate that macrophage activation promotes killing of Brucella by enhancing nitric oxygen (NO), but not reactive oxygen species (ROS) production, macrophage apoptosis or cellular death. Taken together, these findings provide a novel interpretation of Brucella intracellular growth through inhibition of NO production produced by TREM-2-mediated activated macrophages.

  20. N-Formyl-Perosamine Surface Homopolysaccharides Hinder the Recognition of Brucella abortus by Mouse Neutrophils.

    PubMed

    Mora-Cartín, Ricardo; Chacón-Díaz, Carlos; Gutiérrez-Jiménez, Cristina; Gurdián-Murillo, Stephany; Lomonte, Bruno; Chaves-Olarte, Esteban; Barquero-Calvo, Elías; Moreno, Edgardo

    2016-06-01

    Brucella abortus is an intracellular pathogen of monocytes, macrophages, dendritic cells, and placental trophoblasts. This bacterium causes a chronic disease in bovines and in humans. In these hosts, the bacterium also invades neutrophils; however, it fails to replicate and just resists the killing action of these leukocytes without inducing significant activation or neutrophilia. Moreover, B. abortus causes the premature cell death of human neutrophils. In the murine model, the bacterium is found within macrophages and dendritic cells at early times of infection but seldom in neutrophils. Based on this observation, we explored the interaction of mouse neutrophils with B. abortus In contrast to human, dog, and bovine neutrophils, naive mouse neutrophils fail to recognize smooth B. abortus bacteria at early stages of infection. Murine normal serum components do not opsonize smooth Brucella strains, and neutrophil phagocytosis is achieved only after the appearance of antibodies. Alternatively, mouse normal serum is capable of opsonizing rough Brucella mutants. Despite this, neutrophils still fail to kill Brucella, and the bacterium induces cell death of murine leukocytes. In addition, mouse serum does not opsonize Yersinia enterocolitica O:9, a bacterium displaying the same surface polysaccharide antigen as smooth B. abortus Therefore, the lack of murine serum opsonization and absence of murine neutrophil recognition are specific, and the molecules responsible for the Brucella camouflage are N-formyl-perosamine surface homopolysaccharides. Although the mouse is a valuable model for understanding the immunobiology of brucellosis, direct extrapolation from one animal system to another has to be undertaken with caution.

  1. Brucella abortus Induces the Secretion of Proinflammatory Mediators from Glial Cells Leading to Astrocyte Apoptosis

    PubMed Central

    García Samartino, Clara; Delpino, M. Victoria; Pott Godoy, Clara; Di Genaro, María Silvia; Pasquevich, Karina A.; Zwerdling, Astrid; Barrionuevo, Paula; Mathieu, Patricia; Cassataro, Juliana; Pitossi, Fernando; Giambartolomei, Guillermo H.

    2010-01-01

    Central nervous system (CNS) invasion by bacteria of the genus Brucella results in an inflammatory disorder called neurobrucellosis. In this study we present in vivo and in vitro evidence that B. abortus and its lipoproteins activate the innate immunity of the CNS, eliciting an inflammatory response that leads to astrogliosis, a characteristic feature of neurobrucellosis. Intracranial injection of heat-killed B. abortus (HKBA) or outer membrane protein 19 (Omp19), a B. abortus lipoprotein model, induced astrogliosis in mouse striatum. Moreover, infection of astrocytes and microglia with B. abortus induced the secretion of interleukin (IL)−6, IL-1β, tumor necrosis factor (TNF)-α, macrophage chemoattractant protein−1, and KC (CXCL1). HKBA also induced these inflammatory mediators, suggesting the involvement of a structural component of the bacterium. Accordingly, Omp19 induced the same cytokine and chemokine secretion pattern. B. abortus infection induced astrocyte, but not microglia, apoptosis. Indeed, HKBA and Omp19 elicited not only astrocyte apoptosis but also proliferation, two features observed during astrogliosis. Apoptosis induced by HKBA and L-Omp19 was completely suppressed in cells of TNF receptor p55−/− mice or when the general caspase inhibitor Z-VAD-FMK was added to cultures. Hence, TNF-α signaling via TNF receptor (TNFR) 1 through the coupling of caspases determines apoptosis. Our results provide proof of the principle that Brucella lipoproteins could be key virulence factors in neurobrucellosis and that astrogliosis might contribute to neurobrucellosis pathogenesis. PMID:20093491

  2. Immunoproteomics of Brucella abortus RB51 as candidate antigens in serological diagnosis of brucellosis.

    PubMed

    Kim, Ji-Yeon; Sung, So-Ra; Lee, Kichan; Lee, Hyang-Keun; Kang, Sung-Il; Lee, Jin Ju; Jung, Suk Chan; Park, Yong Ho; Her, Moon

    2014-08-15

    The current brucellosis serodiagnostic assays are chiefly based on detecting anti-LPS (lipopolysaccharide) antibodies. However, cross-reaction with some gram-negative bacteria can occasionally induce due to similar O-polysaccharide (OPS) structure. Therefore, the aim of the present study was to identify new candidate antigens from Brucella abortus RB51, a mutant strain lacking the LPS portion, which might be valuable in brucellosis diagnosis. To detect potential antigens, immobilized pH gradients (IPG) strips with three ranges (pH 3-5.6, 4-7 and 6-11) were applied. After separating the insoluble proteins of B. abortus RB51 using two-dimensional electrophoresis (2-DE), their immunogenicity was evaluated by western blotting using four types of antisera - B. abortus, Yersinia enterocolitica O:9 and Escherichia coli O157:H7-positive, and B. abortus-negative bovine sera. Among the several immunogenic spots, the spots showing specific reactivity with only the B. abortus-positive antisera, were considered as candidate antigens. Overall, eleven immuno-reactive proteins were identified, as follows: Cu/Zn superoxide dismutase, histidinol dehydrogenase, chaperonin DnaK, chaperonin GroES, beta-ketoadipyl CoA thiolase, two-component response regulator, the cell-division protein FtsZ, aldehyde dehydrogenase, 50s ribosomal protein L10 and invasion protein B. These selected highly immunogenic protein spots might be useful as alternative antigens for brucellosis and helpful in reducing the cross-reactivity.

  3. Conditional Gene Expression in Chlamydia trachomatis Using the Tet System

    PubMed Central

    Wickstrum, Jason; Sammons, Lindsay R.; Restivo, Keasha N.; Hefty, P. Scott

    2013-01-01

    Chlamydia trachomatis is maintained through a complex bi-phasic developmental cycle that incorporates numerous processes that are poorly understood. This is reflective of the previous paucity of genetic tools available. The recent advent of a method for transforming Chlamydia has enabled the development of essential molecular tools to better study these medically important bacteria. Critical for the study of Chlamydia biology and pathogenesis, is a system for tightly controlled inducible gene expression. To accomplish this, a new shuttle vector was generated with gene expression controlled by the Tetracycline repressor and anhydryotetracycline. Evaluation of GFP expression by this system demonstrated tightly controlled gene regulation with rapid protein expression upon induction and restoration of transcription repression following inducer removal. Additionally, induction of expression could be detected relatively early during the developmental cycle and concomitant with conversion into the metabolically active form of Chlamydia. Uniform and strong GFP induction was observed during middle stages of the developmental cycle. Interestingly, variable induced GFP expression by individual organisms within shared inclusions during later stages of development suggesting metabolic diversity is affecting induction and/or expression. These observations support the strong potential of this molecular tool to enable numerous experimental analyses for a better understanding of the biology and pathogenesis of Chlamydia. PMID:24116144

  4. PPARγ-mediated increase in glucose availability sustains chronic Brucella abortus infection in alternatively activated macrophages

    PubMed Central

    Xavier, Mariana N.; Winter, Maria G.; Spees, Alanna M.; den Hartigh, Andreas B.; Nguyen, Kim; Roux, Christelle M.; Silva, Teane M. A.; Atluri, Vidya L.; Kerrinnes, Tobias; Keestra, A. Marijke; Monack, Denise M.; Luciw, Paul A.; Eigenheer, Richard A.; Bäumler, Andreas J.; Santos, Renato L.; Tsolis, Renée M.

    2013-01-01

    SUMMARY Eradication of persistent intracellular bacterial pathogens with antibiotic therapy is often slow or incomplete. However, strategies to augment antibiotics are hampered by our poor understanding of the nutritional environment that sustains chronic infection. Here we show that the intracellular pathogen Brucella abortus survives and replicates preferentially in alternatively activated macrophages (AAM), which are more abundant during chronic infection. A metabolic shift induced by peroxisome proliferator activated receptor γ (PPARγ), which increases intracellular glucose availability, is identified as a causal mechanism promoting enhanced bacterial survival in AAM. Glucose uptake was crucial for increased replication of B. abortus in AAM, and chronic infection, as inactivation of the bacterial glucose transporter gluP reduced both intracellular survival in AAM and persistence in mice. Thus, a shift in intracellular nutrient availability induced by PPARγ promotes chronic persistence of B. abortus within AAM and targeting this pathway may aid in eradicating chronic infection. PMID:23954155

  5. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants.

    PubMed

    Mol, Juliana P S; Pires, Simone F; Chapeaurouge, Alexander D; Perales, Jonas; Santos, Renato L; Andrade, Hélida M; Lage, Andrey P

    2016-01-01

    Brucella abortus is the etiological agent of bovine brucellosis, a zoonotic disease that causes significant economic losses worldwide. The differential proteomic profile of bovine chorioallantoic membrane (CAM) explants at early stages of infection with B. abortus (0.5, 2, 4, and 8 h) was determined. Analysis of CAM explants at 0.5 and 4 h showed the highest differences between uninfected and infected CAM explants, and therefore were used for the Differential Gel Electrophoresis (DIGE). A total of 103 spots were present in only one experimental group and were selected for identification by mass spectrometry (MALDI/ToF-ToF). Proteins only identified in extracts of CAM explants infected with B. abortus were related to recognition of PAMPs by TLR, production of reactive oxygen species, intracellular trafficking, and inflammation.

  6. Proteomic Profile of Brucella abortus-Infected Bovine Chorioallantoic Membrane Explants

    PubMed Central

    Mol, Juliana P. S.; Pires, Simone F.; Chapeaurouge, Alexander D.; Perales, Jonas; Santos, Renato L.; Andrade, Hélida M.; Lage, Andrey P.

    2016-01-01

    Brucella abortus is the etiological agent of bovine brucellosis, a zoonotic disease that causes significant economic losses worldwide. The differential proteomic profile of bovine chorioallantoic membrane (CAM) explants at early stages of infection with B. abortus (0.5, 2, 4, and 8 h) was determined. Analysis of CAM explants at 0.5 and 4 h showed the highest differences between uninfected and infected CAM explants, and therefore were used for the Differential Gel Electrophoresis (DIGE). A total of 103 spots were present in only one experimental group and were selected for identification by mass spectrometry (MALDI/ToF-ToF). Proteins only identified in extracts of CAM explants infected with B. abortus were related to recognition of PAMPs by TLR, production of reactive oxygen species, intracellular trafficking, and inflammation. PMID:27104343

  7. PPARγ-mediated increase in glucose availability sustains chronic Brucella abortus infection in alternatively activated macrophages.

    PubMed

    Xavier, Mariana N; Winter, Maria G; Spees, Alanna M; den Hartigh, Andreas B; Nguyen, Kim; Roux, Christelle M; Silva, Teane M A; Atluri, Vidya L; Kerrinnes, Tobias; Keestra, A Marijke; Monack, Denise M; Luciw, Paul A; Eigenheer, Richard A; Bäumler, Andreas J; Santos, Renato L; Tsolis, Renée M

    2013-08-14

    Eradication of persistent intracellular bacterial pathogens with antibiotic therapy is often slow or incomplete. However, strategies to augment antibiotics are hampered by our poor understanding of the nutritional environment that sustains chronic infection. Here we show that the intracellular pathogen Brucella abortus survives and replicates preferentially in alternatively activated macrophages (AAMs), which are more abundant during chronic infection. A metabolic shift induced by peroxisome proliferator-activated receptor γ (PPARγ), which increases intracellular glucose availability, is identified as a causal mechanism promoting enhanced bacterial survival in AAMs. Glucose uptake was crucial for increased replication of B. abortus in AAMs, and for chronic infection, as inactivation of the bacterial glucose transporter gluP reduced both intracellular survival in AAMs and persistence in mice. Thus, a shift in intracellular nutrient availability induced by PPARγ promotes chronic persistence of B. abortus within AAMs, and targeting this pathway may aid in eradicating chronic infection. PMID:23954155

  8. Genetic stability of Brucella abortus isolates from an outbreak by multiple-locus variable-number tandem repeat analysis (MLVA16)

    PubMed Central

    2014-01-01

    Background Brucellosis caused by Brucella abortus is one of the most important zoonoses in the world. Multiple-locus variable-number tandem repeat analysis (MLVA16) has been shown be a useful tool to epidemiological traceback studies in B. abortus infection. Thus, the present study aimed (i) to evaluate the genetic diversity of B. abortus isolates from a brucellosis outbreak, and (ii) to investigate the in vivo stability of the MLVA16 markers. Results Three-hundred and seventy-five clinical samples, including 275 vaginal swabs and 100 milk samples, were cultured from a brucellosis outbreak in a cattle herd, which adopted RB51 vaccination and test-and-slaughter policies. Thirty-seven B. abortus isolates were obtained, eight from milk and twenty-nine from post-partum/abortion vaginal swabs, which were submitted to biotyping and genotyping by MLVA16. Twelve B. abortus isolates obtained from vaginal swabs were identified as RB51. Twenty four isolates, seven obtained from milk samples and seventeen from vaginal swabs, were identified as B. abortus biovar 3, while one isolate from vaginal swabs was identified as B. abortus biovar 1. Three distinct genotypes were observed during the brucellosis outbreak: RB observed in all isolates identified as RB51; W observed in all B. abortus biovar 3 isolates; and Z observed in the single B. abortus biovar 1 isolate. Epidemiological and molecular data show that the B. abortus biovar 1 genotype Z strain is not related to the B. abortus biovar 3 genotype W isolates, and represents a new introduction B. abortus during the outbreak. Conclusions The results of the present study on typing of multiple clinical B. abortus isolates from the same outbreak over a sixteen month period indicate the in vivo stability of MLVA16 markers, a low genetic diversity among B. abortus isolates and the usefulness of MLVA16 for epidemiological studies of bovine brucellosis. PMID:25015840

  9. Chlamydia Serine Protease Inhibitor, targeting HtrA, as a New Treatment for Koala Chlamydia infection.

    PubMed

    Lawrence, Amba; Fraser, Tamieka; Gillett, Amber; Tyndall, Joel D A; Timms, Peter; Polkinghorne, Adam; Huston, Wilhelmina M

    2016-01-01

    The koala, an iconic marsupial native to Australia, is a threatened species in many parts of the country. One major factor in the decline is disease caused by infection with Chlamydia. Current therapeutic strategies to treat chlamydiosis in the koala are limited. This study examines the effectiveness of an inhibitor, JO146, which targets the HtrA serine protease for treatment of C. pecorum and C. pneumoniae in vitro and ex vivo with the aim of developing a novel therapeutic for koala Chlamydia infections. Clinical isolates from koalas were examined for their susceptibility to JO146. In vitro studies demonstrated that treatment with JO146 during the mid-replicative phase of C. pecorum or C. pneumoniae infections resulted in a significant loss of infectious progeny. Ex vivo primary koala tissue cultures were used to demonstrate the efficacy of JO146 and the non-toxic nature of this compound on peripheral blood mononuclear cells and primary cell lines established from koala tissues collected at necropsy. Our results suggest that inhibition of the serine protease HtrA could be a novel treatment strategy for chlamydiosis in koalas. PMID:27530689

  10. "Candidatus Mesochlamydia elodeae" (Chlamydiae: Parachlamydiaceae), a novel chlamydia parasite of free-living amoebae.

    PubMed

    Corsaro, Daniele; Müller, Karl-Dieter; Wingender, Jost; Michel, Rolf

    2013-02-01

    Vannella sp. isolated from waterweed Elodea sp. was found infected by a chlamydia-like organism. This organism behaves like a parasite, causing the death through burst of its host. Once the vannellae degenerated, the parasite was successfully kept in laboratory within a Saccamoeba sp. isolated from the same waterweed sample, which revealed in fine through electron microscopy to harbor two bacterial endosymbionts: the chlamydial parasite we introduce and another endosymbiont initially and naturally present in the host. Herein, we provide molecular-based identification of both the amoeba host and its two endosymbionts, with special focus on the chlamydia parasite. High sequence similarity values of the 18S rDNA permitted to assign the amoeba to the species Saccamoeba lacustris (Amoebozoa, Tubulinea). The bacterial endosymbiont naturally harbored by the host belonged to Sphingomonas koreensis (Alpha-Proteobacteria). The chlamydial parasite showed a strict specificity for Saccamoeba spp., being unable to infect a variety of other amoebae, including Acanthamoeba, and it was itself infected by a bacteriophage. Sequence similarity values of the 16S rDNA and phylogenetic analysis indicated that this strain is a new member of the family Parachlamydiaceae, for which we propose the name "Candidatus Mesochlamydia elodeae."

  11. Chlamydia Serine Protease Inhibitor, targeting HtrA, as a New Treatment for Koala Chlamydia infection

    PubMed Central

    Lawrence, Amba; Fraser, Tamieka; Gillett, Amber; Tyndall, Joel D. A.; Timms, Peter; Polkinghorne, Adam; Huston, Wilhelmina M.

    2016-01-01

    The koala, an iconic marsupial native to Australia, is a threatened species in many parts of the country. One major factor in the decline is disease caused by infection with Chlamydia. Current therapeutic strategies to treat chlamydiosis in the koala are limited. This study examines the effectiveness of an inhibitor, JO146, which targets the HtrA serine protease for treatment of C. pecorum and C. pneumoniae in vitro and ex vivo with the aim of developing a novel therapeutic for koala Chlamydia infections. Clinical isolates from koalas were examined for their susceptibility to JO146. In vitro studies demonstrated that treatment with JO146 during the mid-replicative phase of C. pecorum or C. pneumoniae infections resulted in a significant loss of infectious progeny. Ex vivo primary koala tissue cultures were used to demonstrate the efficacy of JO146 and the non-toxic nature of this compound on peripheral blood mononuclear cells and primary cell lines established from koala tissues collected at necropsy. Our results suggest that inhibition of the serine protease HtrA could be a novel treatment strategy for chlamydiosis in koalas. PMID:27530689

  12. Chlamydia Serine Protease Inhibitor, targeting HtrA, as a New Treatment for Koala Chlamydia infection.

    PubMed

    Lawrence, Amba; Fraser, Tamieka; Gillett, Amber; Tyndall, Joel D A; Timms, Peter; Polkinghorne, Adam; Huston, Wilhelmina M

    2016-01-01

    The koala, an iconic marsupial native to Australia, is a threatened species in many parts of the country. One major factor in the decline is disease caused by infection with Chlamydia. Current therapeutic strategies to treat chlamydiosis in the koala are limited. This study examines the effectiveness of an inhibitor, JO146, which targets the HtrA serine protease for treatment of C. pecorum and C. pneumoniae in vitro and ex vivo with the aim of developing a novel therapeutic for koala Chlamydia infections. Clinical isolates from koalas were examined for their susceptibility to JO146. In vitro studies demonstrated that treatment with JO146 during the mid-replicative phase of C. pecorum or C. pneumoniae infections resulted in a significant loss of infectious progeny. Ex vivo primary koala tissue cultures were used to demonstrate the efficacy of JO146 and the non-toxic nature of this compound on peripheral blood mononuclear cells and primary cell lines established from koala tissues collected at necropsy. Our results suggest that inhibition of the serine protease HtrA could be a novel treatment strategy for chlamydiosis in koalas.

  13. "Candidatus Mesochlamydia elodeae" (Chlamydiae: Parachlamydiaceae), a novel chlamydia parasite of free-living amoebae.

    PubMed

    Corsaro, Daniele; Müller, Karl-Dieter; Wingender, Jost; Michel, Rolf

    2013-02-01

    Vannella sp. isolated from waterweed Elodea sp. was found infected by a chlamydia-like organism. This organism behaves like a parasite, causing the death through burst of its host. Once the vannellae degenerated, the parasite was successfully kept in laboratory within a Saccamoeba sp. isolated from the same waterweed sample, which revealed in fine through electron microscopy to harbor two bacterial endosymbionts: the chlamydial parasite we introduce and another endosymbiont initially and naturally present in the host. Herein, we provide molecular-based identification of both the amoeba host and its two endosymbionts, with special focus on the chlamydia parasite. High sequence similarity values of the 18S rDNA permitted to assign the amoeba to the species Saccamoeba lacustris (Amoebozoa, Tubulinea). The bacterial endosymbiont naturally harbored by the host belonged to Sphingomonas koreensis (Alpha-Proteobacteria). The chlamydial parasite showed a strict specificity for Saccamoeba spp., being unable to infect a variety of other amoebae, including Acanthamoeba, and it was itself infected by a bacteriophage. Sequence similarity values of the 16S rDNA and phylogenetic analysis indicated that this strain is a new member of the family Parachlamydiaceae, for which we propose the name "Candidatus Mesochlamydia elodeae." PMID:23224611

  14. The genetic basis of plasmid tropism between Chlamydia trachomatis and Chlamydia muridarum.

    PubMed

    Wang, Yibing; Cutcliffe, Lesley T; Skilton, Rachel J; Ramsey, Kyle H; Thomson, Nicholas R; Clarke, Ian N

    2014-10-01

    The development of genetic transformation technology for Chlamydia trachomatis using its endogenous plasmid has recently been described. Chlamydia muridarum cannot be transformed by the C. trachomatis plasmid, indicating a barrier between chlamydial species. To determine which regions of the plasmid conferred the species specificity, we used the novel approach of transforming wild-type C. muridarum carrying the endogenous plasmid pNigg and forced recombination with the C. trachomatis vector pGFP::SW2 which carries the complete C. trachomatis plasmid (pSW2). Penicillin and chloramphenicol-resistant transformants expressing the green fluorescent protein were selected. Recovery of plasmids from these transformants showed they were recombinants. The differences between the pSW2 and pNigg allowed identification of the recombination breakpoints and showed that pGFP::SW2 had exchanged a ~ 1 kbp region with pNigg covering CDS 2. The recombinant plasmid (pSW2NiggCDS2) is maintained under antibiotic selection when transformed into plasmid-cured C. muridarum. The ability to select for recombinants in C. muridarum shows that the barrier is not at transformation, but at the level of plasmid replication or maintenance. Our studies show that CDS 2, together with adjoining sequences, is the main determinant of plasmid tropism. PMID:24700815

  15. The genetic basis of plasmid tropism between Chlamydia trachomatis and Chlamydia muridarum

    PubMed Central

    Wang, Yibing; Cutcliffe, Lesley T; Skilton, Rachel J; Ramsey, Kyle H; Thomson, Nicholas R; Clarke, Ian N

    2014-01-01

    The development of genetic transformation technology for Chlamydia trachomatis using its endogenous plasmid has recently been described. Chlamydia muridarum cannot be transformed by the C. trachomatis plasmid, indicating a barrier between chlamydial species. To determine which regions of the plasmid conferred the species specificity, we used the novel approach of transforming wild-type C. muridarum carrying the endogenous plasmid pNigg and forced recombination with the C. trachomatis vector pGFP::SW2 which carries the complete C. trachomatis plasmid (pSW2). Penicillin and chloramphenicol-resistant transformants expressing the green fluorescent protein were selected. Recovery of plasmids from these transformants showed they were recombinants. The differences between the pSW2 and pNigg allowed identification of the recombination breakpoints and showed that pGFP::SW2 had exchanged a ∼ 1 kbp region with pNigg covering CDS 2. The recombinant plasmid (pSW2NiggCDS2) is maintained under antibiotic selection when transformed into plasmid-cured C. muridarum. The ability to select for recombinants in C. muridarum shows that the barrier is not at transformation, but at the level of plasmid replication or maintenance. Our studies show that CDS 2, together with adjoining sequences, is the main determinant of plasmid tropism. PMID:24700815

  16. Testing commercial sex workers for chlamydia and gonorrhoea on outreach.

    PubMed

    Macauley, S; Creighton, S

    2009-06-01

    To assess the feasibility of testing indoor commercial sex workers (CSW) for Chlamydia trachomatis and Neisseria gonorrhoeae in an outreach setting. All CSW seen on outreach over a 6-week period were offered self-taken vulval swabs for chlamydia and gonorrhoea testing. Feasibility was assessed by all the outreach workers on a standardised proforma. Of the 93 women offered the service, 40 accepted, of whom five (12%) had not previously accessed sexual health services. The majority of women declining the service had recently attended a sexual health clinic. Three cases of chlamydia and one of gonorrhoea were diagnosed. The cost per sexually transmitted infection (STI) was pound 392.50. Most of this group of women were knowledgeable about sexual health and were already having regular check-ups, but a significant minority did not know how to access STI care. Offering STI testing on outreach was feasible and cost effective. PMID:19155241

  17. Premature apoptosis of Chlamydia-infected cells disrupts chlamydial development.

    PubMed

    Ying, Songmin; Pettengill, Matthew; Latham, E Ray; Walch, Axel; Ojcius, David M; Häcker, Georg

    2008-11-15

    The obligate intracellular development of Chlamydia suggests that the bacteria should be vulnerable to premature host cell apoptosis, but because Chlamydia-infected cells are apoptosis resistant, this has never been able to be tested. We have devised a system to circumvent the apoptotic block imposed by chlamydial infection. When the proapoptotic protein Bim(S) was experimentally induced, epithelial cells underwent apoptosis that was not blocked by chlamydial infection. Apoptosis during the developmental cycle prevented the generation of infectious bacteria and caused transcriptional changes of bacterial genes and loss of intracellular ATP. Intriguingly, although apoptosis resulted in destruction of host cell structures and of the Chlamydia inclusion, and prevented generation of elementary bodies, Bim(S) induction in the presence of a caspase inhibitor allowed differentiation into morphologically normal but noninfectious elementary bodies. These data show that chlamydial infection renders host cells apoptosis resistant at a premitochondrial step and demonstrate the consequences of premature apoptosis for development of the bacteria.

  18. [Chlamydia: from population screening to individual repeated screening].

    PubMed

    Bally, F; Quach, A

    2014-10-01

    Chlamydia trachomatis is a frequent sexually transmitted infection especially in young adults and adolescents. Its complications can impair a woman's reproductive potential. chlamydia control has several challenges. These include asymptomatic infections; a long duration of untreated infections; re-infections and partner treatments. Any person with infection is at high risk of re-infection. Repeated screening would decrease, at an individual level, the risk of complications. General practitioners, gynaecologists and centres for sexual health could participate in Chlamydia screening for asymptomatic infections, in Switzerland, the cost of the laboratory test is fixed by national tariff regulations. The cost is high and prohibitive for many, especially adolescents and young adults and needs to be lowered.

  19. A re-evaluation of the role of B cells in protective immunity to Chlamydia infection.

    PubMed

    Li, Lin-Xi; McSorley, Stephen J

    2015-04-01

    Chlamydia trachomatis is the etiological agent of the most commonly reported bacterial sexual transmitted infection (STI) in North America and Europe. The control of Chlamydia infection is hindered by the asymptomatic nature of initial infection but the consequence of untreated infection seriously threatens the reproductive health of young women. Unfortunately, there is no licensed vaccine for Chlamydia vaccine, in part due to our incomplete understanding of the immune response to Chlamydia urogenital infection. It has been well established that T cell-mediated immunity plays a dominant role in protective immunity against Chlamydia and thus the importance of B cells is somewhat underappreciated. Here, we summarize recent progress on understanding the role of B cells during Chlamydia genital tract infections and discuss how B cells and humoral immunity make an effective contribution to host defense against important intracellular pathogens, including Chlamydia.

  20. A re-evaluation of the role of B cells in protective immunity to Chlamydia infection

    PubMed Central

    Li, Lin-Xi; McSorley, Stephen J.

    2015-01-01

    Chlamydia trachomatis is the etiological agent of the most commonly reported bacterial sexual transmitted infection (STI) in North America and Europe. The control of Chlamydia infection is hindered by the asymptomatic nature of initial infection but the consequence of untreated infection seriously threatens the reproductive health of young women. Unfortunately, there is no licensed vaccine for Chlamydia vaccine, in part due to our incomplete understanding of the immune response to Chlamydia urogenital infection. It has been well established that T cell-mediated immunity plays a dominant role in protective immunity against Chlamydia and thus the importance of B cells is somewhat underappreciated. Here, we summarize recent progress on understanding the role of B cells during Chlamydia genital tract infections and discuss how B cells and humoral immunity make an effective contribution to host defense against important intracellular pathogens, including Chlamydia. PMID:25704502

  1. Characterization of betaine aldehyde dehydrogenase (BetB) as an essential virulence factor of Brucella abortus.

    PubMed

    Lee, Jin Ju; Kim, Jae Hong; Kim, Dae Geun; Kim, Dong Hyeok; Simborio, Hannah Leah; Min, Won Gi; Rhee, Man Hee; Lim, Jong Hwan; Chang, Hong Hee; Kim, Suk

    2014-01-10

    The pathogenic mechanisms of Brucellosis used to adapt to the harsh intracellular environment of the host cell are not fully understood. The present study investigated the in vitro and in vivo characteristics of B. abortus betaine aldehyde dehydrogenase (BetB) (Gene Bank ID: 006932) using a betB deletion mutant constructed from virulent B. abortus 544. In test under stress conditions, including osmotic- and acid stress-resistance, the betB mutant had a lower osmotic-resistance than B. abortus wild-type. In addition, the betB mutant showed higher internalization rates compared to the wild-type strain; however, it also displayed replication failures in HeLa cells and RAW 264.7 macrophages. During internalization, compared to the wild-type strain, the betB mutant was more adherent to the host surface and showed enhanced phosphorylation of protein kinases, two processes that promote phagocytic activity, in host cells. During intracellular trafficking, colocalization of B. abortus-containing phagosomes with LAMP-1 was elevated in betB mutant-infected cells compared to the wild-type cells. In mice, the betB mutant was predominantly cleared from spleens compared to the wild-type strain after 2 weeks post-infection, and the vaccination test with the live betB mutant showed effective protection against challenge infection with the virulent wild-type strain. These findings suggested that the B. abortus betB gene substantially affects the phagocytic pathway in human phagocytes and in host cells in mice. Furthermore, this study highlights the potential use of the B. abortus betB mutant as a live vaccine for the control of brucellosis.

  2. Diagnostic Procedures to Detect Chlamydia trachomatis Infections.

    PubMed

    Meyer, Thomas

    2016-08-05

    The intracellular life style of chlamydia and the ability to cause persistent infections with low-grade replication requires tests with high analytical sensitivity to directly detect C. trachomatis (CT) in medical samples. Nucleic acid amplification tests (NAATs) are the most sensitive assays with a specificity similar to cell culture and are considered the method of choice for CT detection. In addition, NAATs can be performed on various clinical specimens that do not depend on specific transport and storage conditions, since NAATs do not require infectious bacteria. In the case of lower genital tract infections, first void urine and vaginal swabs are the recommended specimens for testing males and females, respectively. Infections of anorectal, oropharyngeal and ocular epithelia should also be tested by NAAT analysis of corresponding mucosal swabs. In particular, anorectal infections of men who have sex with men (MSM) should include evaluation of lymphogranuloma venereum (LGV) by identification of genotypes L1, L2 or L3. Detection of CT antigens by enzyme immunoassay (EIAs) or rapid diagnostic tests (RDTs) are unsuitable due to insufficient sensitivity and specificity. Recent PCR-based RDTs, however, are non-inferior to standard NAATs, and might be used at the point-of-care. Serology finds application in the diagnostic work-up of suspected chronic CT infection but is inappropriate to diagnose acute infections.

  3. Diagnostic Procedures to Detect Chlamydia trachomatis Infections

    PubMed Central

    Meyer, Thomas

    2016-01-01

    The intracellular life style of chlamydia and the ability to cause persistent infections with low-grade replication requires tests with high analytical sensitivity to directly detect C. trachomatis (CT) in medical samples. Nucleic acid amplification tests (NAATs) are the most sensitive assays with a specificity similar to cell culture and are considered the method of choice for CT detection. In addition, NAATs can be performed on various clinical specimens that do not depend on specific transport and storage conditions, since NAATs do not require infectious bacteria. In the case of lower genital tract infections, first void urine and vaginal swabs are the recommended specimens for testing males and females, respectively. Infections of anorectal, oropharyngeal and ocular epithelia should also be tested by NAAT analysis of corresponding mucosal swabs. In particular, anorectal infections of men who have sex with men (MSM) should include evaluation of lymphogranuloma venereum (LGV) by identification of genotypes L1, L2 or L3. Detection of CT antigens by enzyme immunoassay (EIAs) or rapid diagnostic tests (RDTs) are unsuitable due to insufficient sensitivity and specificity. Recent PCR-based RDTs, however, are non-inferior to standard NAATs, and might be used at the point-of-care. Serology finds application in the diagnostic work-up of suspected chronic CT infection but is inappropriate to diagnose acute infections. PMID:27681919

  4. Diagnostic Procedures to Detect Chlamydia trachomatis Infections.

    PubMed

    Meyer, Thomas

    2016-01-01

    The intracellular life style of chlamydia and the ability to cause persistent infections with low-grade replication requires tests with high analytical sensitivity to directly detect C. trachomatis (CT) in medical samples. Nucleic acid amplification tests (NAATs) are the most sensitive assays with a specificity similar to cell culture and are considered the method of choice for CT detection. In addition, NAATs can be performed on various clinical specimens that do not depend on specific transport and storage conditions, since NAATs do not require infectious bacteria. In the case of lower genital tract infections, first void urine and vaginal swabs are the recommended specimens for testing males and females, respectively. Infections of anorectal, oropharyngeal and ocular epithelia should also be tested by NAAT analysis of corresponding mucosal swabs. In particular, anorectal infections of men who have sex with men (MSM) should include evaluation of lymphogranuloma venereum (LGV) by identification of genotypes L1, L2 or L3. Detection of CT antigens by enzyme immunoassay (EIAs) or rapid diagnostic tests (RDTs) are unsuitable due to insufficient sensitivity and specificity. Recent PCR-based RDTs, however, are non-inferior to standard NAATs, and might be used at the point-of-care. Serology finds application in the diagnostic work-up of suspected chronic CT infection but is inappropriate to diagnose acute infections. PMID:27681919

  5. Diagnostic Procedures to Detect Chlamydia trachomatis Infections

    PubMed Central

    Meyer, Thomas

    2016-01-01

    The intracellular life style of chlamydia and the ability to cause persistent infections with low-grade replication requires tests with high analytical sensitivity to directly detect C. trachomatis (CT) in medical samples. Nucleic acid amplification tests (NAATs) are the most sensitive assays with a specificity similar to cell culture and are considered the method of choice for CT detection. In addition, NAATs can be performed on various clinical specimens that do not depend on specific transport and storage conditions, since NAATs do not require infectious bacteria. In the case of lower genital tract infections, first void urine and vaginal swabs are the recommended specimens for testing males and females, respectively. Infections of anorectal, oropharyngeal and ocular epithelia should also be tested by NAAT analysis of corresponding mucosal swabs. In particular, anorectal infections of men who have sex with men (MSM) should include evaluation of lymphogranuloma venereum (LGV) by identification of genotypes L1, L2 or L3. Detection of CT antigens by enzyme immunoassay (EIAs) or rapid diagnostic tests (RDTs) are unsuitable due to insufficient sensitivity and specificity. Recent PCR-based RDTs, however, are non-inferior to standard NAATs, and might be used at the point-of-care. Serology finds application in the diagnostic work-up of suspected chronic CT infection but is inappropriate to diagnose acute infections.

  6. Tetracycline Susceptibility in Chlamydia suis Pig Isolates.

    PubMed

    Donati, Manuela; Balboni, Andrea; Laroucau, Karine; Aaziz, Rachid; Vorimore, Fabien; Borel, Nicole; Morandi, Federico; Vecchio Nepita, Edoardo; Di Francesco, Antonietta

    2016-01-01

    The aims of the present study were to assess the prevalence of Chlamydia suis in an Italian pig herd, determine the tetracycline susceptibility of C. suis isolates, and evaluate tet(C) and tetR(C) gene expression. Conjunctival swabs from 20 pigs were tested for C. suis by real-time polymerase chain reaction, and 55% (11) were positive. C. suis was then isolated from 11 conjunctival swabs resampled from the same herd. All positive samples and isolates were positive for the tet(C) resistance gene. The in vitro susceptibility to tetracycline of the C. suis isolates showed MIC values ranging from 0.5 to 4 μg/mL. Tet(C) and tetR(C) transcripts were found in all the isolates, cultured both in the absence and presence of tetracycline. This contrasts with other Gram-negative bacteria in which both genes are repressed in the absence of the drug. Further investigation into tet gene regulation in C. suis is needed.

  7. Experimental infection of goat fetuses in utero with a stable, rough mutant of Brucella abortus.

    PubMed

    Roop, R M; Jeffers, G; Bagchi, T; Walker, J; Enright, F M; Schurig, G G

    1991-09-01

    Fetuses of goats in their last trimester of pregnancy were experimentally infected with Brucella abortus strain RB51, a stable rough mutant deficient in the perosamine O-chain content of its lipopolysaccharide. RB51 maintained its rough phenotype in vivo and did not induce abortion. Infection with RB51 resulted in the production of significant levels of IgG type antibodies specific for B abortus cellular antigens distinct from the perosamine O-chain. These findings suggest that strain RB51 will be useful in the pregnant goat for studying the role of brucella antigens other than the lipopolysaccharide O-chain in the immune response to brucellosis.

  8. Murine and Bovine γδ T Cells Enhance Innate Immunity against Brucella abortus Infections

    PubMed Central

    Skyberg, Jerod A.; Thornburg, Theresa; Rollins, MaryClare; Huarte, Eduardo; Jutila, Mark A.; Pascual, David W.

    2011-01-01

    γδ T cells have been postulated to act as a first line of defense against infectious agents, particularly intracellular pathogens, representing an important link between the innate and adaptive immune responses. Human γδ T cells expand in the blood of brucellosis patients and are active against Brucella in vitro. However, the role of γδ T cells in vivo during experimental brucellosis has not been studied. Here we report TCRδ−/− mice are more susceptible to B. abortus infection than C57BL/6 mice at one week post-infection as measured by splenic colonization and splenomegaly. An increase in TCRγδ cells was observed in the spleens of B. abortus-infected C57BL/6 mice, which peaked at two weeks post-infection and occurred concomitantly with diminished brucellae. γδ T cells were the major source of IL-17 following infection and also produced IFN-γ. Depletion of γδ T cells from C57BL/6, IL-17Rα−/−, and GMCSF−/− mice enhanced susceptibility to B. abortus infection although this susceptibility was unaltered in the mutant mice; however, when γδ T cells were depleted from IFN-γ−/− mice, enhanced susceptibility was observed. Neutralization of γδ T cells in the absence of TNF-α did not further impair immunity. In the absence of TNF-α or γδ T cells, B. abortus-infected mice showed enhanced IFN-γ, suggesting that they augmented production to compensate for the loss of γδ T cells and/or TNF-α. While the protective role of γδ T cells was TNF-α-dependent, γδ T cells were not the major source of TNF-α and activation of γδ T cells following B. abortus infection was TNF-α-independent. Additionally, bovine TCRγδ cells were found to respond rapidly to B. abortus infection upon co-culture with autologous macrophages and could impair the intramacrophage replication of B. abortus via IFN-γ. Collectively, these results demonstrate γδ T cells are important for early protection to B. abortus infections. PMID:21765931

  9. Paper-based molecular diagnostic for Chlamydia trachomatis

    PubMed Central

    Linnes, Jacqueline C.; Fan, Andy; Rodriguez, Natalia M.; Lemieux, Bertrand; Kong, Huimin; Klapperich, Catherine M.

    2014-01-01

    Herein we show the development of a minimally instrumented paper-based molecular diagnostic for point of care detection of sexually transmitted infections caused by Chlamydia trachomatis. This new diagnostic platform incorporates cell lysis, isothermal nucleic acid amplification, and lateral flow visual detection using only a pressure source and heat block, eliminating the need for expensive laboratory equipment. This paper-based test can be performed in less than one hour and has a clinically relevant limit of detection that is 100x more sensitive than current rapid immunoassays used for chlamydia diagnosis. PMID:25309740

  10. Transmission of Chlamydia and genital warts during sleepwalking.

    PubMed

    Mohanty, Kailash

    2008-02-01

    A boy aged 15 years infected a girl of 13 years with Chlamydia and genital warts. The boy has been engaged in sexual activities for the last four years. There was no dispute that the boy had had sex with the girl. He was prosecuted but acquitted by the Court on the ground of 'defence of sleepwalking'. This is the first case where sexually transmitted infections like Chlamydia and genital warts have been transmitted sexually through sleepwalking. This case also raises the issues of underage sex and issues of confidentiality. PMID:18334071

  11. Transmission of Chlamydia and genital warts during sleepwalking.

    PubMed

    Mohanty, Kailash

    2008-02-01

    A boy aged 15 years infected a girl of 13 years with Chlamydia and genital warts. The boy has been engaged in sexual activities for the last four years. There was no dispute that the boy had had sex with the girl. He was prosecuted but acquitted by the Court on the ground of 'defence of sleepwalking'. This is the first case where sexually transmitted infections like Chlamydia and genital warts have been transmitted sexually through sleepwalking. This case also raises the issues of underage sex and issues of confidentiality.

  12. Chlamydia genomics: providing novel insights into chlamydial biology.

    PubMed

    Bachmann, Nathan L; Polkinghorne, Adam; Timms, Peter

    2014-08-01

    Chlamydiaceae are obligate intracellular pathogens that have successfully evolved to colonize a diverse range of hosts. There are currently 11 described species of Chlamydia, most of which have a significant impact on the health of humans or animals. Expanding chlamydial genome sequence information has revolutionized our understanding of chlamydial biology, including aspects of their unique lifecycle, host-pathogen interactions, and genetic differences between Chlamydia strains associated with different host and tissue tropisms. This review summarizes the major highlights of chlamydial genomics and reflects on the considerable impact these have had on understanding the biology of chlamydial pathogens and the changing nature of genomics tools in the 'post-genomics' era.

  13. Simultaneous subcutaneous and conjunctival administration of the influenza viral vector based Brucella abortus vaccine to pregnant heifers provides better protection against B. abortus 544 infection than the commercial B. abortus S19 vaccine.

    PubMed

    Tabynov, Kaissar; Orynbayev, Mukhit; Renukaradhya, Gourapura J; Sansyzbay, Abylai

    2016-09-30

    In this study, we explored possibility of increasing the protective efficacy of our novel influenza viral vector based B. abortus vaccine (Flu-BA) in pregnant heifers by adapting an innovative method of vaccine delivery. We administered the vaccine concurrently via the conjunctival and subcutaneous routes to pregnant heifers, and these routes were previously tested individually. The Flu-BA vaccination of pregnant heifers (n=9) against a challenge B. abortus 544 infection provided protection from abortion, infection of heifers and fetuses/calves by 88.8%, 100% and 100%, respectively (alpha=0.004-0.0007 vs. negative control; n=7). Our candidate vaccine using this delivery method provided slightly better protection than the commercial B. abortus S19 vaccine in pregnant heifers (n=8), which provided protection from abortion, infection of heifers and fetuses/calves by 87.5%, 75% and 87.5%, respectively. This improved method of the Flu-BA vaccine administration is highly recommended for the recovery of farms which has high prevalence of brucellosis.

  14. Simultaneous subcutaneous and conjunctival administration of the influenza viral vector based Brucella abortus vaccine to pregnant heifers provides better protection against B. abortus 544 infection than the commercial B. abortus S19 vaccine.

    PubMed

    Tabynov, Kaissar; Orynbayev, Mukhit; Renukaradhya, Gourapura J; Sansyzbay, Abylai

    2016-09-30

    In this study, we explored possibility of increasing the protective efficacy of our novel influenza viral vector based B. abortus vaccine (Flu-BA) in pregnant heifers by adapting an innovative method of vaccine delivery. We administered the vaccine concurrently via the conjunctival and subcutaneous routes to pregnant heifers, and these routes were previously tested individually. The Flu-BA vaccination of pregnant heifers (n=9) against a challenge B. abortus 544 infection provided protection from abortion, infection of heifers and fetuses/calves by 88.8%, 100% and 100%, respectively (alpha=0.004-0.0007 vs. negative control; n=7). Our candidate vaccine using this delivery method provided slightly better protection than the commercial B. abortus S19 vaccine in pregnant heifers (n=8), which provided protection from abortion, infection of heifers and fetuses/calves by 87.5%, 75% and 87.5%, respectively. This improved method of the Flu-BA vaccine administration is highly recommended for the recovery of farms which has high prevalence of brucellosis. PMID:27595898

  15. Toll-Like Receptor 6 Plays an Important Role in Host Innate Resistance to Brucella abortus Infection in Mice

    PubMed Central

    de Almeida, Leonardo A.; Macedo, Gilson C.; Marinho, Fábio A. V.; Gomes, Marco T. R.; Corsetti, Patrícia P.; Silva, Aristóbolo M.; Cassataro, Juliana; Giambartolomei, Guillermo H.

    2013-01-01

    Brucella abortus is recognized by several Toll-like receptor (TLR)-associated pathways triggering proinflammatory responses that affect both the nature and intensity of the immune response. Previously, we demonstrated that B. abortus-mediated dendritic cell (DC) maturation and control of infection are dependent on the adaptor molecule MyD88. However, the involvement of all TLRs in response to B. abortus infection is not completely understood. Therefore, we decided to evaluate the requirement for TLR6 in host resistance to B. abortus. Here, we demonstrated that TLR6 is an important component for triggering an innate immune response against B. abortus. An in vitro luciferase assay indicated that TLR6 cooperates with TLR2 to sense Brucella and further activates NF-κB signaling. However, in vivo analysis showed that TLR6, not TLR2, is required for the efficient control of B. abortus infection. Additionally, B. abortus-infected dendritic cells require TLR6 to induce tumor necrosis factor alpha (TNF-α) and interleukin-12 (IL-12). Furthermore, our findings demonstrated that the mitogen-activated protein kinase (MAPK) signaling pathway is impaired in TLR2, TLR6, and TLR2/6 knockout (KO) DCs when infected with B. abortus, which may account for the lower proinflammatory cytokine production observed in TLR6 KO mouse dendritic cells. In summary, the results presented here indicate that TLR6 is required to trigger innate immune responses against B. abortus in vivo and is required for the full activation of DCs to induce robust proinflammatory cytokine production. PMID:23460520

  16. The novel chlamydial adhesin CPn0473 mediates the lipid raft‐dependent uptake of Chlamydia pneumoniae

    PubMed Central

    Fechtner, Tim; Galle, Jan N.

    2016-01-01

    Summary Chlamydiae are Gram‐negative, obligate intracellular pathogens that pose a serious threat to public health worldwide. Chlamydial surface molecules are essential for host cell invasion. The first interaction with the host cell is thereby accomplished by the Outer membrane complex protein B (OmcB) binding to heparan sulfate moieties on the host cell surface, followed by the interaction of the chlamydial polymorphic membrane proteins (Pmps) with host cell receptors. Specifically, the interaction of the Pmp21 adhesin and invasin with its human interaction partner, the epidermal growth factor receptor, results in receptor activation, down‐stream signalling and finally internalization of the bacteria. Blocking both, the OmcB and Pmp21 adhesion pathways, did not completely abolish infection, suggesting the presence of additional factors relevant for host cell invasion. Here, we show that the novel surface protein CPn0473 of Chlamydia pneumoniae contributes to the binding and invasion of infectious chlamydial particles. CPn0473 is expressed late in the infection cycle and located on the infectious chlamydial cell surface. Soluble recombinant CPn0473 as well as rCPn0473‐coupled fluorescent latex beads adhere to human epithelial HEp‐2 cells. Interestingly, in classical infection blocking experiments pretreatment of HEp‐2 cells with rCPn0473 does not attenuate adhesion but promotes dose‐dependently internalization by C. pneumoniae suggesting an unusual mode of action for this adhesin. This CPn0473‐dependent promotion of infection by C. pneumoniae depends on two different domains within the protein and requires intact lipid rafts. Thus, inhibition of the interaction of CPn0473 with the host cell could provide a way to reduce the virulence of C. pneumoniae. PMID:26780295

  17. The novel chlamydial adhesin CPn0473 mediates the lipid raft-dependent uptake of Chlamydia pneumoniae.

    PubMed

    Fechtner, Tim; Galle, Jan N; Hegemann, Johannes H

    2016-08-01

    Chlamydiae are Gram-negative, obligate intracellular pathogens that pose a serious threat to public health worldwide. Chlamydial surface molecules are essential for host cell invasion. The first interaction with the host cell is thereby accomplished by the Outer membrane complex protein B (OmcB) binding to heparan sulfate moieties on the host cell surface, followed by the interaction of the chlamydial polymorphic membrane proteins (Pmps) with host cell receptors. Specifically, the interaction of the Pmp21 adhesin and invasin with its human interaction partner, the epidermal growth factor receptor, results in receptor activation, down-stream signalling and finally internalization of the bacteria. Blocking both, the OmcB and Pmp21 adhesion pathways, did not completely abolish infection, suggesting the presence of additional factors relevant for host cell invasion. Here, we show that the novel surface protein CPn0473 of Chlamydia pneumoniae contributes to the binding and invasion of infectious chlamydial particles. CPn0473 is expressed late in the infection cycle and located on the infectious chlamydial cell surface. Soluble recombinant CPn0473 as well as rCPn0473-coupled fluorescent latex beads adhere to human epithelial HEp-2 cells. Interestingly, in classical infection blocking experiments pretreatment of HEp-2 cells with rCPn0473 does not attenuate adhesion but promotes dose-dependently internalization by C. pneumoniae suggesting an unusual mode of action for this adhesin. This CPn0473-dependent promotion of infection by C. pneumoniae depends on two different domains within the protein and requires intact lipid rafts. Thus, inhibition of the interaction of CPn0473 with the host cell could provide a way to reduce the virulence of C. pneumoniae.

  18. Characterization of the activity and expression of arginine decarboxylase in human and animal Chlamydia pathogens.

    PubMed

    Bliven, Kimberly A; Fisher, Derek J; Maurelli, Anthony T

    2012-12-01

    Chlamydia pneumoniae encodes a functional arginine decarboxylase (ArgDC), AaxB, that activates upon self-cleavage and converts l-arginine to agmatine. In contrast, most Chlamydia trachomatis serovars carry a missense or nonsense mutation in aaxB abrogating activity. The G115R missense mutation was not predicted to impact AaxB functionality, making it unclear whether AaxB variations in other Chlamydia species also result in enzyme inactivation. To address the impact of gene polymorphism on functionality, we investigated the activity and production of the Chlamydia AaxB variants. Because ArgDC plays a critical role in the Escherichia coli acid stress response, we studied the ability of these Chlamydia variants to complement an E. coli ArgDC mutant in an acid shock assay. Active AaxB was detected in four additional species: Chlamydia caviae, Chlamydia pecorum, Chlamydia psittaci, and Chlamydia muridarum. Of the C. trachomatis serovars, only E appears to encode active enzyme. To determine when functional enzyme is present during the chlamydial developmental cycle, we utilized an anti-AaxB antibody to detect both uncleaved and cleaved enzyme throughout infection. Uncleaved enzyme production peaked around 20 h postinfection, with optimal cleavage around 44 h. While the role ArgDC plays in Chlamydia survival or virulence is unclear, our data suggest a niche-specific function.

  19. Seroprevalence and genotype of Chlamydia in pet parrots in China.

    PubMed

    Zhang, N-Z; Zhang, X-X; Zhou, D-H; Huang, S-Y; Tian, W-P; Yang, Y-C; Zhao, Q; Zhu, X-Q

    2015-01-01

    Parrots are one of the most popular pet birds in China, and can harbour Chlamydia which has significance for human and animal health. We investigated, by indirect haemagglutination assay, the seroprevalence of Chlamydia infection in four species of parrots, namely budgerigars (Melopsittacus undulatus), lovebirds (Agapornis sp.), cockatiels (Nymphicus hollandicus) and Alexandrine parakeets (Psittacula eupatria) that were collected from Weifang and Beijing cities, North China and explored the association between potential risk factors and chlamydial seropositivity. We further determined the genotype of Chlamydia in 21 fresh faecal samples based on the ompA sequence by reconstruction of phylogenetic relationships. Of the 311 parrots examined, 35·37% (95% confidence interval 30·06-40·68) were seropositive, and species, gender, age, season and geographical location were identified as risk factors. Two PCR-positive samples represented Chlamydia psittaci genotype A. The occurrence of C. psittaci genotype A in the droppings of two pet parrots in China suggests potential environmental contamination with Chlamydiaceae and may raise a public health concern. PMID:24588856

  20. Seroprevalence and genotype of Chlamydia in pet parrots in China.

    PubMed

    Zhang, N-Z; Zhang, X-X; Zhou, D-H; Huang, S-Y; Tian, W-P; Yang, Y-C; Zhao, Q; Zhu, X-Q

    2015-01-01

    Parrots are one of the most popular pet birds in China, and can harbour Chlamydia which has significance for human and animal health. We investigated, by indirect haemagglutination assay, the seroprevalence of Chlamydia infection in four species of parrots, namely budgerigars (Melopsittacus undulatus), lovebirds (Agapornis sp.), cockatiels (Nymphicus hollandicus) and Alexandrine parakeets (Psittacula eupatria) that were collected from Weifang and Beijing cities, North China and explored the association between potential risk factors and chlamydial seropositivity. We further determined the genotype of Chlamydia in 21 fresh faecal samples based on the ompA sequence by reconstruction of phylogenetic relationships. Of the 311 parrots examined, 35·37% (95% confidence interval 30·06-40·68) were seropositive, and species, gender, age, season and geographical location were identified as risk factors. Two PCR-positive samples represented Chlamydia psittaci genotype A. The occurrence of C. psittaci genotype A in the droppings of two pet parrots in China suggests potential environmental contamination with Chlamydiaceae and may raise a public health concern.

  1. 21 CFR 866.3120 - Chlamydia serological reagents.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Chlamydia serological reagents. 866.3120 Section 866.3120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3120...

  2. 21 CFR 866.3120 - Chlamydia serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Chlamydia serological reagents. 866.3120 Section 866.3120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3120...

  3. 21 CFR 866.3120 - Chlamydia serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Chlamydia serological reagents. 866.3120 Section 866.3120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3120...

  4. 21 CFR 866.3120 - Chlamydia serological reagents.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Chlamydia serological reagents. 866.3120 Section 866.3120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3120...

  5. 21 CFR 866.3120 - Chlamydia serological reagents.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Chlamydia serological reagents. 866.3120 Section 866.3120 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3120...

  6. DNA stability of Chlamydia trachomatis and Neisseria gonorrhoeae in urine.

    PubMed

    Le Guern, Rémi; Miaux, Brigitte; Pischedda, Patricia; Herwegh, Stéphanie; Courcol, René

    2016-07-01

    We evaluated the DNA stability of Chlamydia trachomatis and Neisseria gonorrhoeae in 55 urine samples. Crossing threshold (Ct) values were highly similar after 3 to 14 days at room temperature (+0.002, P = 0.99). Consequently, it does not seem necessary to transfer urine specimens into a transport medium in less than 24 hours as recommended by manufacturers. PMID:27130478

  7. Chlamydia Pneumoniae Infections and Sudden Unexpected Deaths in Denmark.

    ERIC Educational Resources Information Center

    Johannsen, Finn

    1993-01-01

    Blood samples from 38 runners on the Danish national orienteering team revealed no ongoing chlamydia pneumoniae, although 42% had an earlier infection, similar to the incidence in the general population. However, over 2% had an ongoing lyme borreliosis infection, and 18% had an earlier infection, which is a higher incidence than in the general…

  8. Chlamydia pneumoniae encodes a functional aromatic amino acid hydroxylase.

    PubMed

    Abromaitis, Stephanie; Hefty, P Scott; Stephens, Richard S

    2009-03-01

    Chlamydia pneumoniae is a community-acquired respiratory pathogen that has been associated with the development of atherosclerosis. Analysis of the C. pneumoniae genome identified a gene (Cpn1046) homologous to eukaryotic aromatic amino acid hydroxylases (AroAA-Hs). AroAA-Hs hydroxylate phenylalanine, tyrosine, and tryptophan into tyrosine, dihydroxyphenylalanine, and 5-hydroxytryptophan, respectively. Sequence analysis of Cpn1046 demonstrated that residues essential for AroAA-H enzymatic function are conserved and that a subset of Chlamydia species contain an AroAA-H homolog. The chlamydial AroAA-Hs are transcriptionally linked to a putative bacterial membrane transport protein. We determined that recombinant Cpn1046 is able to hydroxylate phenylalanine, tyrosine, and tryptophan with roughly equivalent activity for all three substrates. Cpn1046 is expressed within 24 h of infection, allowing C. pneumoniae to hydroxylate host stores of aromatic amino acids during the period of logarithmic bacterial growth. From these results we can conclude that C. pneumoniae, as well as a subset of other Chlamydia species, encode an AroAA-H that is able to use all three aromatic amino acids as substrates. The maintenance of this gene within a number of Chlamydia suggests that the enzyme may have an important role in shaping the metabolism or overall pathogenesis of these bacteria. PMID:19141112

  9. A mouse model for Chlamydia suis genital infection.

    PubMed

    Donati, Manuela; Di Paolo, Maria; Favaroni, Alison; Aldini, Rita; Di Francesco, Antonietta; Ostanello, Fabio; Biondi, Roberta; Cremonini, Eleonora; Ginocchietti, Laura; Cevenini, Roberto

    2015-02-01

    A mouse model for Chlamydia suis genital infection was developed. Ninety-nine mice were randomly divided into three groups and intravaginally inoculated with chlamydia: 45 mice (group 1) received C. suis purified elementary bodies (EBs), 27 (group 2) were inoculated with C. trachomatis genotype E EBs and 27 mice (group 3) with C. trachomatis genotype F EBs. Additionally, 10 mice were used as a negative control. At seven days post-infection (dpi) secretory anti-C. suis IgA were recovered from vaginal swabs of all C. suis inoculated mice. Chlamydia suis was isolated from 93, 84, 71 and 33% vaginal swabs at 3, 5, 7 and 12 dpi. Chlamydia trachomatis genotype E and F were isolated from 100% vaginal swabs up to 7 dpi and from 61 and 72%, respectively, at 12 dpi. Viable C. suis and C. trachomatis organisms were isolated from uterus and tubes up to 16 and 28 dpi, respectively. The results of the present study show the susceptibility of mice to intravaginal inoculation with C. suis. A more rapid course and resolution of C. suis infection, in comparison to C. trachomatis, was highlighted. The mouse model could be useful for comparative investigations involving C. suis and C. trachomatis species.

  10. Abortion and premature birth in cattle following vaccination with Brucella abortus strain RB51.

    PubMed

    Fluegel Dougherty, Amanda M; Cornish, Todd E; O'Toole, Donal; Boerger-Fields, Amy M; Henderson, Owen L; Mills, Ken W

    2013-09-01

    Brucella abortus RB51 is the vaccine strain currently licensed for immunizing cattle against brucellosis in the United States. Most cattle are vaccinated as heifer calves at 4-12 months of age. Adult cattle may be vaccinated in selected high-risk situations. Two herds of pregnant adult cattle in the brucellosis-endemic area of Wyoming were vaccinated with a standard label dose (1.0-3.4 × 10(10) organisms) of RB51. Reproductive losses in the vaccinated herds were 5.3% (herd A) and 0.6% (herd B) and included abortions, stillbirths, premature calves, and unbred cows (presumed early abortion). Brucella abortus was cultured from multiple tissues of aborted and premature calves (7/9), and from placenta. Isolates were identified as B. abortus strain RB51 by standard strain typing procedures and a species-specific polymerase chain reaction. Bronchopneumonia with intralesional bacteria and placentitis were observed microscopically. There was no evidence of involvement of other infectious or toxic causes of abortion. Producers, veterinarians, and laboratory staff should be alert to the risk of abortion when pregnant cattle are vaccinated with RB51, to potential human exposure, and to the importance of distinguishing field from vaccinal strains of B. abortus.

  11. Genetic characterization of Brucella melitensis and Brucella abortus geographical clusters in Italy.

    PubMed

    De Massis, Fabrizio; Garofolo, Giuliano; Cammà, Cesare; Ippoliti, Carla; Candeloro, Luca; Ancora, Massimo; Calistri, Paolo

    2015-01-01

    The genetic diversity of Brucella melitensis and Brucella abortus strains isolated in 199 cattle and sheep from 156 brucellosis outbreaks which occurred in 8 regions of Southern Italy in 2011, was determined using a Multiple-Locus Variable Number of Tandem Repeats Analysis approach. The existence of possible genetic clusters was verified through a hierarchical cluster analysis based on 'single link', which is closely related to the minimum spanning tree. The Hamming weighted distance matrix was adopted in the analysis. All calculations were performed using R and the additional libraries phangorn and Cluster. For a number of clusters, ranging from 2 to 15, the average silhouette width was calculated. The number of clusters adopted was identified according to the maximum average silhouette width. For B. abortus and B. melitensis, 6 and 11 genetic clusters were identified, respectively. Three out of 6 B. abortus clusters included the 96.7% of all B. abortus isolates. Clusters were clearly geographically separated, and this highlighted the known epidemiological links among them. Brucella melitensis genotypes resulted more heterogeneous; the 3 more representative genetic clusters included 79.7% of all B. melitensis isolates. A clear geographical clusterization of genotypes is recognizable only for 1 cluster, whereas the others are more widespread across Southern Italy. The genetic characterization of Brucella strains isolated from animals may be a useful tool to better understand the epidemiology and dissemination patterns of this pathogen through host populations.

  12. Genome Sequences of Brucella abortus and Brucella suis Strains Isolated from Bovine in Zimbabwe

    PubMed Central

    Ledwaba, Betty; Mafofo, Joseph

    2014-01-01

    This is a report of whole-genome sequences of a Brucella abortus strain and two Brucella suis strains isolated from bovine in Zimbabwe. These strains were selected based on their origin and data obtained when using multiplex PCR assays, then sequenced using next-generation sequencing technologies. PMID:25342680

  13. Brucella abortus Exposure during an Orthopedic Surgical Procedure—New Mexico, 2010

    PubMed Central

    Nichols, Megin; Thompson, Deborah; Carothers, Joshua T.; Klauber, Judy; Stoddard, Robyn A.; Guerra, Marta A.; Benoit, Tina J.; Traxler, Rita M.

    2015-01-01

    We describe a periprosthetic Brucella abortus infection in a case-patient undergoing hip replacement revision surgery, and the subsequent investigation of laboratory and surgical staff exposures. Although exposures are rare, it is important to have infection prevention recommendations for surgical procedures among patients with suspected or unidentified Brucella spp. infection. PMID:25026630

  14. Pyruvate kinase is necessary for Brucella abortus full virulence in BALB/c mouse.

    PubMed

    Gao, Jianpeng; Tian, Mingxing; Bao, Yanqing; Li, Peng; Liu, Jiameng; Ding, Chan; Wang, Shaohui; Li, Tao; Yu, Shengqing

    2016-08-25

    Brucellosis, caused by a facultative intracellular pathogen Brucella, is one of the most prevalent zoonosis worldwide. Host infection relies on several uncanonical virulence factors. A recent research hotpot is the links between carbon metabolism and bacterial virulence. In this study, we found that a carbon metabolism-related pyruvate kinase (Pyk) encoded by pyk gene (locus tag BAB_RS24320) was associated with Brucella virulence. Determination of bacterial growth curves and resistance to environmental stress factors showed that Pyk plays an important role in B. abortus growth, especially under the conditions of nutrition deprivation, and resistance to oxidative stress. Additionally, cell infection assay showed that Pyk is necessary for B. abortus survival and evading fusion with lysosomes within RAW264.7 cells. Moreover, animal experiments exhibited that the Pyk deletion significantly reduced B. abortus virulence in a mouse infection model. Our results elucidated the role of the Pyk in B. abortus virulence and provided information for further investigation of Brucella virulence associated carbon metabolism.

  15. Immune responses of bison and efficacy after booster vaccination with Brucella abortus strain RB51

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Thirty-one bison heifers were randomly assigned to saline (control; n=7) or single vaccination (n=24) with 1010 CFU of B. abortus strain RB51 (RB51). Some vaccinated bison were randomly selected for booster vaccination with 10**10 CFU of RB51 at 11 months after initial vaccination (n=16). When comp...

  16. Pyruvate kinase is necessary for Brucella abortus full virulence in BALB/c mouse.

    PubMed

    Gao, Jianpeng; Tian, Mingxing; Bao, Yanqing; Li, Peng; Liu, Jiameng; Ding, Chan; Wang, Shaohui; Li, Tao; Yu, Shengqing

    2016-01-01

    Brucellosis, caused by a facultative intracellular pathogen Brucella, is one of the most prevalent zoonosis worldwide. Host infection relies on several uncanonical virulence factors. A recent research hotpot is the links between carbon metabolism and bacterial virulence. In this study, we found that a carbon metabolism-related pyruvate kinase (Pyk) encoded by pyk gene (locus tag BAB_RS24320) was associated with Brucella virulence. Determination of bacterial growth curves and resistance to environmental stress factors showed that Pyk plays an important role in B. abortus growth, especially under the conditions of nutrition deprivation, and resistance to oxidative stress. Additionally, cell infection assay showed that Pyk is necessary for B. abortus survival and evading fusion with lysosomes within RAW264.7 cells. Moreover, animal experiments exhibited that the Pyk deletion significantly reduced B. abortus virulence in a mouse infection model. Our results elucidated the role of the Pyk in B. abortus virulence and provided information for further investigation of Brucella virulence associated carbon metabolism. PMID:27561260

  17. Molecular Epidemiology of Brucella abortus in Northern Ireland—1991 to 2012

    PubMed Central

    Byrne, Andrew; Mallon, Thomas; Skuce, Robin; Groussaud, Pauline; Dainty, Amanda; Graham, Judith; Jones, Kerri; Pollock, Lorraine; Whatmore, Adrian

    2015-01-01

    Background Brucellosis is the most common bacterial zoonoses worldwide. Bovine brucellosis caused by Brucella abortus has far reaching animal health and economic impacts at both the local and national levels. Alongside traditional veterinary epidemiology, the use of molecular typing has recently been applied to inform on bacterial population structure and identify epidemiologically-linked cases of infection. Multi-locus variable number tandem repeat VNTR analysis (MLVA) was used to investigate the molecular epidemiology of a well-characterised Brucella abortus epidemic in Northern Ireland involving 387 herds between 1991 and 2012. Results MLVA identified 98 unique B. abortus genotypes from disclosing isolates in the 387 herds involved in the epidemic. Clustering algorithms revealed the relatedness of many of these genotypes. Combined with epidemiological information on chronology of infection and geographic location, these genotype data helped to identify 7 clonal complexes which underpinned the outbreak over the defined period. Hyper-variability of some VNTR loci both within herds and individual animals led to detection of multiple genotypes associated with single outbreaks. However with dense sampling, these genotypes could still be associated with specific clonal complexes thereby permitting inference of epidemiological links. MLVA- based epidemiological monitoring data were congruent with an independent classical veterinary epidemiology study carried out in the same territory. Conclusions MLVA is a useful tool in ongoing disease surveillance of B. abortus outbreaks, especially when combined with accurate epidemiological information on disease tracings, geographical clustering of cases and chronology of infection. PMID:26325586

  18. Identification and partial characterisation of a new protective antigen of Brucella abortus.

    PubMed

    Cespedes, S; Andrews, E; Folch, H; Oñate, A

    2000-02-01

    Two novel Brucella abortus proteins were isolated from B. abortus strain RB51 and their immunological properties were determined. These proteins precipitated in the 40-60% saturated concentration range of ammonium sulphate and had a molecular mass of 32.2 kDa and 22.9 kDa, respectively. Both were able to induce a strong in-vitro blast transformation in lymphoid cells obtained from mice previously sensitised with a crude brucella protein extract. The protection studies showed that the 22.9-kDa protein used as a protective immunogen was as effective as the live B. abortus RB51 vaccine but the 32.2-kDa protein had a poor protective effect under similar conditions. The amino-terminal sequence of the 22.9-kDa and 32.2-kDa proteins was determined and analysed in a database. The lack of homology with other known B. abortus proteins indicated that both proteins were novel antigens.

  19. Antigenic, Immunologic and Genetic Characterization of Rough Strains B.abortus RB51, B.melitensis B115 and B.melitensis B18

    PubMed Central

    Adone, Rosanna; Muscillo, Michele; La Rosa, Giuseppina; Francia, Massimiliano; Tarantino, Michela

    2011-01-01

    The lipopolysaccharide (LPS) is considered the major virulent factor in Brucella spp. Several genes have been identified involved in the synthesis of the three LPS components: lipid A, core and O-PS. Usually, Brucella strains devoid of O-PS (rough mutants) are less virulent than the wild type and do not induce undesirable interfering antibodies. Such of them proved to be protective against brucellosis in mice. Because of these favorable features, rough strains have been considered potential brucellosis vaccines. In this study, we evaluated the antigenic, immunologic and genetic characteristics of rough strains B.abortus RB51, B.melitensis B115 and B.melitensis B18. RB51 derived from B.abortus 2308 virulent strain and B115 is a natural rough strain in which the O-PS is present in the cytoplasm. B18 is a rough rifampin-resistan mutant isolated in our laboratory. The surface antigenicity of RB51, B115 and B18 was evaluated by testing their ability to bind antibodies induced by rough or smooth Brucella strains. The antibody response induced by each strain was evaluated in rabbits. Twenty-one genes, involved in the LPS-synthesis, were sequenced and compared with the B.melitensis 16M strain. The results indicated that RB51, B115 and B18 have differences in antigenicity, immunologic and genetic properties. Particularly, in B115 a nonsense mutation was detected in wzm gene, which could explain the intracellular localization of O-PS in this strain. Complementation studies to evaluate the precise role of each mutation in affecting Brucella morphology and its virulence, could provide useful information for the assessment of new, attenuated vaccines for brucellosis. PMID:22065984

  20. Antigenic, immunologic and genetic characterization of rough strains B. abortus RB51, B. melitensis B115 and B. melitensis B18.

    PubMed

    Adone, Rosanna; Muscillo, Michele; La Rosa, Giuseppina; Francia, Massimiliano; Tarantino, Michela

    2011-01-01

    The lipopolysaccharide (LPS) is considered the major virulent factor in Brucella spp. Several genes have been identified involved in the synthesis of the three LPS components: lipid A, core and O-PS. Usually, Brucella strains devoid of O-PS (rough mutants) are less virulent than the wild type and do not induce undesirable interfering antibodies. Such of them proved to be protective against brucellosis in mice. Because of these favorable features, rough strains have been considered potential brucellosis vaccines. In this study, we evaluated the antigenic, immunologic and genetic characteristics of rough strains B. abortus RB51, B. melitensis B115 and B. melitensis B18. RB51 derived from B. abortus 2308 virulent strain and B115 is a natural rough strain in which the O-PS is present in the cytoplasm. B18 is a rough rifampin-resistan mutant isolated in our laboratory. The surface antigenicity of RB51, B115 and B18 was evaluated by testing their ability to bind antibodies induced by rough or smooth Brucella strains. The antibody response induced by each strain was evaluated in rabbits. Twenty-one genes, involved in the LPS-synthesis, were sequenced and compared with the B. melitensis 16M strain. The results indicated that RB51, B115 and B18 have differences in antigenicity, immunologic and genetic properties. Particularly, in B115 a nonsense mutation was detected in wzm gene, which could explain the intracellular localization of O-PS in this strain. Complementation studies to evaluate the precise role of each mutation in affecting Brucella morphology and its virulence, could provide useful information for the assessment of new, attenuated vaccines for brucellosis.

  1. Expression and Targeting of Secreted Proteins from Chlamydia trachomatis

    PubMed Central

    Bauler, Laura D.

    2014-01-01

    Chlamydia trachomatis is an obligate intracellular pathogen that replicates in a vacuole termed the inclusion. Many of the interactions of chlamydiae with the host cell are dependent upon bacterial protein synthesis and presumably exposure of these proteins to the cytosol. Because of the dearth of genetic tools for chlamydiae, previous studies examining secreted proteins required the use of heterologous bacterial systems. Recent advances in genetic manipulation of chlamydia now allow for transformation of the bacteria with plasmids. We describe here a shuttle vector system, pBOMB4, that permits expression of recombinant proteins under constitutive or conditional promoter control. We show that the inclusion membrane protein IncD is secreted in a type III-dependent manner from Yersinia pseudotuberculosis and also secreted from C. trachomatis in infected cells where it localizes appropriately to the inclusion membrane. IncD truncated of the first 30 amino acids containing the secretion signal is no longer secreted and is retained by the bacteria. Cytosolic exposure of secreted proteins can be confirmed by using CyaA, GSK, or microinjection assays. A protein predicted to be retained within the bacteria, NrdB is indeed localized to the chlamydia. In addition, we have shown that the chlamydial effector protein, CPAF, which is secreted into the host cell cytosol by a Sec-dependent pathway, also accesses the cytosol when expressed from this system. These assays should prove useful to assess the secretion of other chlamydial proteins that are potentially exposed to the cytosol of the host cell. PMID:24443531

  2. The Chromosome-Encoded Hypothetical Protein TC0668 Is an Upper Genital Tract Pathogenicity Factor of Chlamydia muridarum

    PubMed Central

    Conrad, Turner Allen; Gong, Siqi; Yang, Zhangsheng; Matulich, Patrick; Keck, Jonathon; Beltrami, Noah; Chen, Chaoqun; Zhou, Zhou; Dai, Jin

    2015-01-01

    We previously associated a missense mutation of the tc0668 gene of serial in vitro-passaged Chlamydia muridarum, a murine model of human urogenital C. trachomatis, with severely attenuated disease development in the upper genital tract of female mice. Since these mutants also contained a TC0237 Q117E missense mutation that enhances their in vitro infectivity, an effort was made here to isolate and characterize a tc0668 single mutant to determine its individual contribution to urogenital pathogenicity. Detailed genetic analysis of C. muridarum passages revealed a truncated variant with a G216* nonsense mutation of the 408-amino-acid TC0668 protein that does not produce a detectable product. Intracellular growth and infectivity of C. muridarum in vitro remain unaffected in the absence of TC0668. Intravaginal inoculation of the TC0668 null mutant into C3H/HeJ mice results in a typical course of lower genital tract infection but, unlike a pathogenic isogenic control, is unable to elicit significant chronic inflammation of the oviduct and fails to induce hydrosalpinx. Thus, TC0668 is demonstrated as an important chromosome-encoded urogenital pathogenicity factor of C. muridarum and the first with these characteristics to be discovered for a Chlamydia pathogen. PMID:26597987

  3. Examination of taxonomic uncertainties surrounding Brucella abortus bv. 7 by phenotypic and molecular approaches.

    PubMed

    Garin-Bastuji, Bruno; Mick, Virginie; Le Carrou, Gilles; Allix, Sebastien; Perrett, Lorraine L; Dawson, Claire E; Groussaud, Pauline; Stubberfield, Emma J; Koylass, Mark; Whatmore, Adrian M

    2014-03-01

    Brucella taxonomy is perpetually being reshuffled, at both the species and intraspecies levels. Biovar 7 of Brucella abortus was suspended from the Approved Lists of Bacterial Names Brucella classification in 1988, because of unpublished evidence that the reference strain 63/75 was a mixture of B. abortus biovars 3 and 5. To formally clarify the situation, all isolates previously identified as B. abortus bv. 7 in the AHVLA and ANSES strain collections were characterized by classical microbiological and multiple molecular approaches. Among the 14 investigated strains, including strain 63/75, only four strains, isolated in Kenya, Turkey, and Mongolia, were pure and showed a phenotypic profile in agreement with the former biovar 7, particularly agglutination with both anti-A/anti-M monospecific sera. These results were strengthened by molecular strategies. Indeed, genus- and species-specific methods allowed confirmation that the four pure strains belonged to the B. abortus species. The combination of most approaches excluded their affiliation with the recognized biovars (biovars 1 to 6 and 9), while some suggested that they were close to biovar 3.These assays were complemented by phylogenetic and/or epidemiological methods, such as multilocus sequence analysis (MLSA) and variable-number tandem repeat (VNTR) analysis. The results of this polyphasic investigation allow us to propose the reintroduction of biovar 7 into the Brucella classification, with at least three representative strains. Interestingly, the Kenyan strain, sharing the same biovar 7 phenotype, was genetically divergent from other three isolates. These discrepancies illustrate the complexity of Brucella taxonomy. This study suggests that worldwide collections could include strains misidentified as B. abortus bv. 7, and it highlights the need to verify their real taxonomic position.

  4. Genetic diversity of Brucella abortus and Brucella melitensis in Kazakhstan using MLVA-16.

    PubMed

    Shevtsov, Alexandr; Ramanculov, Erlan; Shevtsova, Elena; Kairzhanova, Alma; Tarlykov, Pavel; Filipenko, Maxim; Dymova, Maya; Abisheva, Gulzada; Jailbekova, Aygul; Kamalova, Dinara; Chsherbakov, Andrei; Tulegenov, Samat; Akhmetova, Assel; Sytnik, Igor; Karibaev, Talgat; Mukanov, Kasim

    2015-08-01

    Brucellosis is an endemic disease in Central Asia characterized by high infection rates in humans and animals. Currently, little is known about the genetic diversity of Brucella spp. circulating in the region, despite the high prevalence of brucellosis. This study aimed to analyze the genetic diversity of Brucella melitensis and Brucella abortus strains circulating in the Republic of Kazakhstan. We genotyped 128 B. melitensis and 124 B. abortus strains collected in regions with the highest prevalence of brucellosis. Genotyping was performed using multi-locus variable-number tandem-repeat analysis (MLVA). Analysis of a subset of 8 loci (MLVA-8) of 128 B. melitensis strains identified genotypes 42 (n=108), 43 (n=2), and 63 (n=19) related to the 'East Mediterranean' group. An MLVA-16 assay sorted 128 B. melitensis strains into 25 different genotypes. Excluding one variable locus, MLVA-15 of B. melitensis was distinct from strains originating in the Mediterranean region; however, 77% of them were identical to strains isolated in China. A minimum spanning tree for B. melitensis using MLVA-15 analysis clustered the local strains together with strains previously collected in China. MLVA-8 analysis of 124 B. abortus strains identified them as genotype 36, suggesting Eurasian distribution of this lineage. Complete MLVA-16 assay analysis clustered the strains into five genotypes, revealing little diversity of B. abortus when compared on the global scale. A minimum spanning tree for B. abortus obtained using MLVA-15 analysis clustered the 2 most prevalent genotypes (n=117) together with strains previously collected in China. Thus, MLVA analysis was used to characterize 252 strains of Brucella collected in Kazakhstan. The analysis revealed genetic homogeneity among the strains. Interestingly, identical MLVA-15 profiles were found in seemingly unrelated outbreaks in China, Turkey, and Kazakhstan. Further analysis is needed for better understanding of the epidemiology of

  5. 5-Lipoxygenase Negatively Regulates Th1 Response during Brucella abortus Infection in Mice

    PubMed Central

    Fahel, Júlia Silveira; de Souza, Mariana Bueno; Gomes, Marco Túlio Ribeiro; Corsetti, Patricia P.; Carvalho, Natalia B.; Marinho, Fabio A. V.; de Almeida, Leonardo A.; Caliari, Marcelo V.; Machado, Fabiana Simão

    2015-01-01

    Brucella abortus is a Gram-negative bacterium that infects humans and cattle, causing a chronic inflammatory disease known as brucellosis. A Th1-mediated immune response plays a critical role in host control of this pathogen. Recent findings indicate contrasting roles for lipid mediators in host responses against infections. 5-Lipoxygenase (5-LO) is an enzyme required for the production of the lipid mediators leukotrienes and lipoxins. To determine the involvement of 5-LO in host responses to B. abortus infection, we intraperitoneally infected wild-type and 5-LO-deficient mice and evaluated the progression of infection and concomitant expression of immune mediators. Here, we demonstrate that B. abortus induced the upregulation of 5-LO mRNA in wild-type mice. Moreover, this pathogen upregulated the production of the lipid mediators leukotriene B4 and lipoxin A4 in a 5-LO-dependent manner. 5-LO-deficient mice displayed lower bacterial burdens in the spleen and liver and less severe liver pathology, demonstrating an enhanced resistance to infection. Host resistance paralleled an increased expression of the proinflammatory mediators interleukin-12 (IL-12), gamma interferon (IFN-γ), and inducible nitric oxide synthase (iNOS) during the course of infection. Moreover, we demonstrated that 5-LO downregulated the expression of IL-12 in macrophages during B. abortus infection. Our results suggest that 5-LO has a major involvement in B. abortus infection, by functioning as a negative regulator of the protective Th1 immune responses against this pathogen. PMID:25583526

  6. Examination of Taxonomic Uncertainties Surrounding Brucella abortus bv. 7 by Phenotypic and Molecular Approaches

    PubMed Central

    Garin-Bastuji, Bruno; Le Carrou, Gilles; Allix, Sebastien; Perrett, Lorraine L.; Dawson, Claire E.; Groussaud, Pauline; Stubberfield, Emma J.; Koylass, Mark; Whatmore, Adrian M.

    2014-01-01

    Brucella taxonomy is perpetually being reshuffled, at both the species and intraspecies levels. Biovar 7 of Brucella abortus was suspended from the Approved Lists of Bacterial Names Brucella classification in 1988, because of unpublished evidence that the reference strain 63/75 was a mixture of B. abortus biovars 3 and 5. To formally clarify the situation, all isolates previously identified as B. abortus bv. 7 in the AHVLA and ANSES strain collections were characterized by classical microbiological and multiple molecular approaches. Among the 14 investigated strains, including strain 63/75, only four strains, isolated in Kenya, Turkey, and Mongolia, were pure and showed a phenotypic profile in agreement with the former biovar 7, particularly agglutination with both anti-A/anti-M monospecific sera. These results were strengthened by molecular strategies. Indeed, genus- and species-specific methods allowed confirmation that the four pure strains belonged to the B. abortus species. The combination of most approaches excluded their affiliation with the recognized biovars (biovars 1 to 6 and 9), while some suggested that they were close to biovar 3.These assays were complemented by phylogenetic and/or epidemiological methods, such as multilocus sequence analysis (MLSA) and variable-number tandem repeat (VNTR) analysis. The results of this polyphasic investigation allow us to propose the reintroduction of biovar 7 into the Brucella classification, with at least three representative strains. Interestingly, the Kenyan strain, sharing the same biovar 7 phenotype, was genetically divergent from other three isolates. These discrepancies illustrate the complexity of Brucella taxonomy. This study suggests that worldwide collections could include strains misidentified as B. abortus bv. 7, and it highlights the need to verify their real taxonomic position. PMID:24362435

  7. In Vitro Antibacterial Effects of Five Volatile Oil Extracts Against Intramacrophage Brucella Abortus 544

    PubMed Central

    Al-Mariri, Ayman; Saour, George; Hamou, Razan

    2012-01-01

    Background: Brucella abortus is a gram-negative facultative intracellular bacterium that can cause a highly contagious disease in sheep, goats, cattle and one-humped camels. It is responsible for one of the most important zoonosis in human. The aim of this study was to evaluate the role of Mentha piperita, Origanum majorana, Citrus lemon, Cinnamomum verum and Myristica fragrans essential volatile oil extracts on human macrophages infected by B. abortus 544. Methods: Essential volatile oil extracts from M. piperita, O. majorana, C. lemon, C. verum and M. fragrans were extracted. Human macrophages were cultured at a density of 2×105 cells per well in sterile 96-well microtiter plates, and infected with B. abortus 544 at a ratio of 1:100 bacteria/cell. Then essential volatile oil extracts were added at a concentration of 1%. At specified times; cells were washed, lysed with 0.1% Triton, and plated on 2YT agar to determine the number of intracellular bacteria. Results: Cinnamomum verum volatile oil at a concentration of 1% had the highest antibacterial activity against B. abortus 544 inside human macrophages. Its inhibitory effect observed from 24 h and continued till 144 h after the infection. Moreover, C. verum (0.1%) in combination with 1% concentration of M. piperita, O. majorana, C. lemon or M. fragrans volatile oil extracts produced a synergistic inhibitory effect against B. abortus 544. Conclusion: The results indicate that, among the five selected oil extracts, C. verum volatile oil applied either separately or in combination with other oil extracts had the most effective antimicrobial activity against Brucella. PMID:23115441

  8. Genetic diversity of Brucella abortus and Brucella melitensis in Kazakhstan using MLVA-16.

    PubMed

    Shevtsov, Alexandr; Ramanculov, Erlan; Shevtsova, Elena; Kairzhanova, Alma; Tarlykov, Pavel; Filipenko, Maxim; Dymova, Maya; Abisheva, Gulzada; Jailbekova, Aygul; Kamalova, Dinara; Chsherbakov, Andrei; Tulegenov, Samat; Akhmetova, Assel; Sytnik, Igor; Karibaev, Talgat; Mukanov, Kasim

    2015-08-01

    Brucellosis is an endemic disease in Central Asia characterized by high infection rates in humans and animals. Currently, little is known about the genetic diversity of Brucella spp. circulating in the region, despite the high prevalence of brucellosis. This study aimed to analyze the genetic diversity of Brucella melitensis and Brucella abortus strains circulating in the Republic of Kazakhstan. We genotyped 128 B. melitensis and 124 B. abortus strains collected in regions with the highest prevalence of brucellosis. Genotyping was performed using multi-locus variable-number tandem-repeat analysis (MLVA). Analysis of a subset of 8 loci (MLVA-8) of 128 B. melitensis strains identified genotypes 42 (n=108), 43 (n=2), and 63 (n=19) related to the 'East Mediterranean' group. An MLVA-16 assay sorted 128 B. melitensis strains into 25 different genotypes. Excluding one variable locus, MLVA-15 of B. melitensis was distinct from strains originating in the Mediterranean region; however, 77% of them were identical to strains isolated in China. A minimum spanning tree for B. melitensis using MLVA-15 analysis clustered the local strains together with strains previously collected in China. MLVA-8 analysis of 124 B. abortus strains identified them as genotype 36, suggesting Eurasian distribution of this lineage. Complete MLVA-16 assay analysis clustered the strains into five genotypes, revealing little diversity of B. abortus when compared on the global scale. A minimum spanning tree for B. abortus obtained using MLVA-15 analysis clustered the 2 most prevalent genotypes (n=117) together with strains previously collected in China. Thus, MLVA analysis was used to characterize 252 strains of Brucella collected in Kazakhstan. The analysis revealed genetic homogeneity among the strains. Interestingly, identical MLVA-15 profiles were found in seemingly unrelated outbreaks in China, Turkey, and Kazakhstan. Further analysis is needed for better understanding of the epidemiology of

  9. The presence of Chlamydia phage PhiCPG1 capsid protein VP1 genes and antibodies in patients infected with Chlamydia trachomatis.

    PubMed

    Ma, Jingyue; Liu, Yuan; Liu, Yuanjun; Li, Lingjie; Hou, Shuping; Gao, Xibo; Qi, Manli; Liu, Quanzhong

    2016-01-01

    Chlamydia phage PhiCPG1 has been found in Chlamydia caviae in a guinea pig model for inclusion conjunctivitis, raising the possibility that Chlamydia phage is also present in patients infected with C. trachomatis (Ct). In the present study, we assayed for presence of Chlamydia phage capsid protein VP1 genes and antibodies in 84 non-Ct controls and 206 Ct patients using an enzyme-linked immunoassay (ELISA), followed by verification with Western blot. None of the subjects were exposed to an antibiotic treatment or had a C. pneumoniae infection. The VP1 antibody test was positive in both, the ELISA and Western blot assay, in 4 Ct patients. PCR amplification experiments revealed presence of the VP1 gene in 5 Ct patients. The results suggest that Chlamydia phage capsid protein VP1 may exist in some Ct patients. PMID:27213552

  10. The bovine immune response to Brucella abortus I. A water soluble antigen precipitated by sera of some naturally infected cattle.

    PubMed Central

    Stemshorn, B; Nielsen, K

    1977-01-01

    Selected sera from cattle naturally infected with Brucella abortus precipitate water soluble antigens extracted by sonication from B. abortus. One of these antigens resembles antigen E (Baughn and Freeman) as it is excluded from Sephadex G-200 gels, migrates anodally when electrophoresed at pH 8.6, resists heating at 100 degrees C for ten minutes and appears to be susceptible to papain digestion. Precipitins specific for this antigen remained in sera from which all detectable Brucella agglutinating antibody had been removed by adsorption with live or heat killed B. abortus. The antigen has been extracted from smooth and rough strains of B abortus. Precipitins specific for this antigen have been detected in antisera produced against Brucella canis. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:405088

  11. Structural, functional and immunogenic insights on Cu,Zn Superoxide Dismutase pathogenic virulence factors from Neisseria meningitidis and Brucella abortus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacterial pathogens Neisseria meningitidis and Brucella abortus pose threats to human and animal health worldwide, causing meningococcal disease and brucellosis, respectively. Mortality from acute N. meningitidis infections remains high despite antibiotics, and brucellosis presents alimentary and he...

  12. Heat shock response of murine Chlamydia trachomatis.

    PubMed Central

    Engel, J N; Pollack, J; Perara, E; Ganem, D

    1990-01-01

    We have investigated the heat shock response in the mouse pneumonitis strain of Chlamydia trachomatis. The kinetics of the chlamydial heat shock response resembled that of other procaryotes: the induction was rapid, occurring over a 5- to 10-min time period, and was regulated at the level of transcription. Immunoblot analysis and immunoprecipitations with heterologous antisera to the heat shock proteins DnaK and GroEL demonstrated that the rate of synthesis, but not the absolute amount of these two proteins, increased after heat shock. Using a general screen for genes whose mRNAs are induced by heat shock, we identified and cloned two of these. DNA sequence analysis demonstrated that one of the genes is a homolog of dnaK. Further sequence analysis of the region upstream of the dnaK gene revealed that the chlamydial homolog of the grpE gene is located just adjacent to the dnaK gene. The second locus encoded three potential nonoverlapping open reading frames. One of the open reading frames was 52% homologous to the ribosomal protein S18 of Escherichia coli and thus presumably encodes the chlamydial homolog. Interestingly, this ribosomal protein is not known to be induced by heat shock in E. coli. S1 nuclease and primer extension analyses located the start site of the dnaK transcript to the last nucleotide of the grpE coding sequence, suggesting that these two genes, although tandemly arranged, are transcribed separately. No promoter sequences resembling the E. coli consensus heat shock promoter could be identified upstream of either the C. trachomatis dnaK, grpE, or S18 gene. The induction of the dnaK and S18 mRNAs by heat shock occurred at a transcriptional level; their induction could be blocked by rifampin. The mechanisms of induction for these two loci were not the same, however; they were differentially sensitive to chloramphenicol. Whereas the induction of dnaK mRNA required de novo protein synthesis, the induction of the S18 mRNA did not. Thus, C. trachomatis

  13. Naturally occurring lesions of the uterine tube in sheep and serologic evidence of exposure to Chlamydophila abortus.

    PubMed Central

    Tomlinson, L; Barker, I K; Foster, R A; McEwen, S A; Menzies, P I; Shewen, P E

    2000-01-01

    The uterine tubes from 405 ewes, collected at an abattoir, were assessed grossly and microscopically for abnormalities that correlated with serological evidence of exposure to Chlamydophila abortus. Gross lesions were found in 41 ewes and 86 had microscopic lesions. Enzyme immunoassay (EIA) of serum was used as an indication of exposure of individual ewes to C. abortus; 52 were found to be positive. Chi-squared analysis indicated no association between EIA-positive animals and lesions of the uterine tube. PMID:11041501

  14. Effects of partial deletion of the wzm and wzt genes on lipopolysaccharide synthesis and virulence of Brucella abortus S19.

    PubMed

    Wang, Xiuran; Wang, Lin; Lu, Tiancheng; Yang, Yanling; Chen, Si; Zhang, Rui; Lang, Xulong; Yan, Guangmou; Qian, Jing; Wang, Xiaoxu; Meng, Lingyi; Wang, Xinglong

    2014-06-01

    Brucellosis is a worldwide human and animal infectious disease, and the effective methods of its control are immunisation of animals by vaccination and elimination. Brucella abortus S19 is one of the popular vaccines with virulence in the control of cattle Brucellosis. In the present study, allelic exchange plasmids of wzm and wzt genes and partial knockout mutants of wzm and wzt were constructed to evaluate the resulting difference in virulence of B. abortus S19. PCR analysis revealed that the target genes were knocked out. The mutants were rough mutants and they could be differentiated from natural infection by the Rose Bengal plate and standard agglutination tests. The molecular weights of lipopolysaccharides of the Δwzm and Δwzt mutants were clustered between 25 and 40 kDa, and 30 and 35 kDa separately, and were markedly different from those in B. abortus S19. The virulence of B. abortus Δwzm and Δwzt was decreased compared with that of B. abortus S19 in mice. All these results identified that there were several differences between the wzm and wzt genes on lipopolysaccharide synthesis and on the virulence of B. abortus.

  15. Comparison of Cytokine Immune Responses to Brucella abortus and Yersinia enterocolitica Serotype O:9 Infections in BALB/c Mice

    PubMed Central

    Gu, Wenpeng; Wang, Xin; Qiu, Haiyan; Cui, Buyun; Zhao, Shiwen; Zheng, Han; Xiao, Yuchun; Liang, Junrong; Duan, Ran

    2013-01-01

    Brucella abortus and Yersinia enterocolitica serotype O:9 serologically cross-react in the immune response with the host; therefore, our aim was to compare the immune responses to these two pathogens. We selected typical B. abortus and Y. enterocolitica O:9 strains to study the cytokine immune response and the histopathological changes in livers and spleens of BALB/c mice. The data showed the cytokine responses to the two strains of pathogens were different, where the average levels of granulocyte-macrophage colony-stimulating factor (GM-CSF), gamma interferon (IFN-γ), interleukin-12 (IL-12), and tumor necrosis factor alpha (TNF-α) were higher with B. abortus infections than with Y. enterocolitica O:9 infections, especially for IFN-γ, while the IL-10 level was lower and the levels of IL-1β, IL-4, IL-5, and IL-6 were similar. The histopathological effects in the livers and spleens of the BALB/c mice with B. abortus and Y. enterocolitica O:9 infections were similar; however, the pathological changes in the liver were greater with B. abortus infections, while damage in the spleen was greater with Y. enterocolitica O:9 infections. These observations show that different cytokine responses and histopathological changes occur with B. abortus and Y. enterocolitica O:9 infections. PMID:24042115

  16. Seroprevalence and Risk Factors of Chlamydia Infection in Domestic Rabbits (Oryctolagus cuniculus) in China.

    PubMed

    Ni, Xiaoting; Qin, Siyuan; Lou, Zhilong; Ning, Hongrui; Sun, Xiaolin

    2015-01-01

    Chlamydia spp. are obligate intracellular bacteria distributed all over the world, known to cause various forms of diseases in animals and humans. In the present study, a serological survey was conducted to detect the seroprevalence and risk factors associated with rabbit chlamydiosis in northeast China, including Liaoning province, Jilin province, Heilongjiang province, and Inner Mongolia Autonomous Region. Antibodies to Chlamydia were determined by indirect hemagglutination assay (IHA). The overall seroprevalence was estimated at 17.88% in total of 800 blood samples. The Chlamydia seroprevalence varied in domestic rabbits from different factors, and genders of domestic rabbits were considered as major risk factors associated with Chlamydia infection. Our study revealed a widespread and high prevalence of Chlamydia infection in domestic rabbits in northeast China, with higher exposure risk in female domestic rabbits. These findings suggested the potential importance of domestic rabbits in the transmission of zoonotic Chlamydia infection, and thus Chlamydia should be taken into consideration in diagnosing rabbit diseases. To our knowledge, there is no report of Chlamydia infection in domestic rabbits in China and the results extend the host range for Chlamydia, which has important implications for public health and the local economy. PMID:25945336

  17. Chlamydia Species-Dependent Differences in the Growth Requirement for Lysosomes

    PubMed Central

    Ouellette, Scot P.; Dorsey, Frank C.; Moshiach, Simon; Cleveland, John L.; Carabeo, Rey A.

    2011-01-01

    Genome reduction is a hallmark of obligate intracellular pathogens such as Chlamydia, where adaptation to intracellular growth has resulted in the elimination of genes encoding biosynthetic enzymes. Accordingly, chlamydiae rely heavily on the host cell for nutrients yet their specific source is unclear. Interestingly, chlamydiae grow within a pathogen-defined vacuole that is in close apposition to lysosomes. Metabolically-labeled uninfected host cell proteins were provided as an exogenous nutrient source to chlamydiae-infected cells, and uptake and subsequent labeling of chlamydiae suggested lysosomal degradation as a source of amino acids for the pathogen. Indeed, Bafilomycin A1 (BafA1), an inhibitor of the vacuolar H+/ATPase that blocks lysosomal acidification and functions, impairs the growth of C. trachomatis and C. pneumoniae, and these effects are especially profound in C. pneumoniae. BafA1 induced the marked accumulation of material within the lysosomal lumen, which was due to the inhibition of proteolytic activities, and this response inhibits chlamydiae rather than changes in lysosomal acidification per se, as cathepsin inhibitors also inhibit the growth of chlamydiae. Finally, the addition of cycloheximide, an inhibitor of eukaryotic protein synthesis, compromises the ability of lysosomal inhibitors to block chlamydial growth, suggesting chlamydiae directly access free amino acids in the host cytosol as a preferred source of these nutrients. Thus, chlamydiae co-opt the functions of lysosomes to acquire essential amino acids. PMID:21408144

  18. Seroprevalence and Risk Factors of Chlamydia Infection in Domestic Rabbits (Oryctolagus cuniculus) in China.

    PubMed

    Ni, Xiaoting; Qin, Siyuan; Lou, Zhilong; Ning, Hongrui; Sun, Xiaolin

    2015-01-01

    Chlamydia spp. are obligate intracellular bacteria distributed all over the world, known to cause various forms of diseases in animals and humans. In the present study, a serological survey was conducted to detect the seroprevalence and risk factors associated with rabbit chlamydiosis in northeast China, including Liaoning province, Jilin province, Heilongjiang province, and Inner Mongolia Autonomous Region. Antibodies to Chlamydia were determined by indirect hemagglutination assay (IHA). The overall seroprevalence was estimated at 17.88% in total of 800 blood samples. The Chlamydia seroprevalence varied in domestic rabbits from different factors, and genders of domestic rabbits were considered as major risk factors associated with Chlamydia infection. Our study revealed a widespread and high prevalence of Chlamydia infection in domestic rabbits in northeast China, with higher exposure risk in female domestic rabbits. These findings suggested the potential importance of domestic rabbits in the transmission of zoonotic Chlamydia infection, and thus Chlamydia should be taken into consideration in diagnosing rabbit diseases. To our knowledge, there is no report of Chlamydia infection in domestic rabbits in China and the results extend the host range for Chlamydia, which has important implications for public health and the local economy.

  19. Interplay between Clathrin and Rab5 Controls the Early Phagocytic Trafficking and Intracellular Survival of Brucella abortus within HeLa cells*

    PubMed Central

    Lee, Jin Ju; Kim, Dae Geun; Kim, Dong Hyeok; Simborio, Hannah Leah; Min, Wongi; Lee, Hu Jang; Her, Moon; Jung, Suk Chan; Watarai, Masahisa; Kim, Suk

    2013-01-01

    Lipid raft-associated clathrin is essential for host-pathogen interactions during infection. Brucella abortus is an intracellular pathogen that circumvents host defenses, but little is known about the precise infection mechanisms that involve interaction with lipid raft-associated mediators. The aim of this study was to elucidate the clathrin-mediated phagocytic mechanisms of B. abortus. The clathrin dependence of B. abortus infection in HeLa cells was investigated using an infection assay and immunofluorescence microscopy. The redistribution of clathrin in the membrane and in phagosomes was investigated using sucrose gradient fractionation of lipid rafts and the isolation of B. abortus-containing vacuoles, respectively. Clathrin and dynamin were concentrated into lipid rafts during B. abortus infection, and the entry and intracellular survival of B. abortus within HeLa cells were abrogated by clathrin inhibition. Clathrin disruption decreased actin polymerization and the colocalization of B. abortus-containing vacuoles with clathrin and Rab5 but not lysosome-associated membrane protein 1 (LAMP-1). Thus, our data demonstrate that clathrin plays a fundamental role in the entry and intracellular survival of B. abortus via interaction with lipid rafts and actin rearrangement. This process facilitates the early intracellular trafficking of B. abortus to safe replicative vacuoles. PMID:23940042

  20. Immunization of BALB/c mice with Brucella abortus 2308ΔwbkA confers protection against wild-type infection

    PubMed Central

    Li, Zhi-qiang; Gui, Dan; Sun, Zhi-hua; Zhang, Jun-bo; Zhang, Wen-zhi; Guo, Fei

    2015-01-01

    Brucellosis is a zoonotic disease that causes animal and human diseases. Vaccination is a major measure for prevention of brucellosis, but it is currently not possible to distinguish vaccinated animals from those that have been naturally infected. Therefore, in this study, we constructed the Brucella (B.) abortus 2380 wbkA mutant (2308ΔwbkA) and evaluated its virulence. The survival of 2308ΔwbkA was attenuated in murine macrophage (RAW 264.7) and BALB/c mice, and it induced high protective immunity in mice. The wbkA mutant elicited an anti-Brucella-specific immunoglobulin G response and induced the secretion of gamma interferon. Antibodies to 2308ΔwbkA could be detected in sera from mice, implying the potential for use of this protein as a diagnostic antigen. The WbkA antigen would allow serological differentiation between infected and vaccinated animals. These results suggest that 2308ΔwbkA is a potential attenuated vaccine against 16M. This vaccine will be further evaluated in sheep. PMID:26040616

  1. Role of Epithelial-Mesenchyme Transition in Chlamydia Pathogenesis.

    PubMed

    Igietseme, Joseph U; Omosun, Yusuf; Stuchlik, Olga; Reed, Matthew S; Partin, James; He, Qing; Joseph, Kahaliah; Ellerson, Debra; Bollweg, Brigid; George, Zenas; Eko, Francis O; Bandea, Claudiu; Liu, Hsi; Yang, Genyan; Shieh, Wun-Ju; Pohl, Jan; Karem, Kevin; Black, Carolyn M

    2015-01-01

    Chlamydia trachomatis genital infection in women causes serious adverse reproductive complications, and is a strong co-factor for human papilloma virus (HPV)-associated cervical epithelial carcinoma. We tested the hypothesis that Chlamydia induces epithelial-mesenchyme transition (EMT) involving T cell-derived TNF-alpha signaling, caspase activation, cleavage inactivation of dicer and dysregulation of micro-RNA (miRNA) in the reproductive epithelium; the pathologic process of EMT causes fibrosis and fertility-related epithelial dysfunction, and also provides the co-factor function for HPV-related cervical epithelial carcinoma. Using a combination of microarrays, immunohistochemistry and proteomics, we showed that chlamydia altered the expression of crucial miRNAs that control EMT, fibrosis and tumorigenesis; specifically, miR-15a, miR-29b, miR-382 and MiR-429 that maintain epithelial integrity were down-regulated, while miR-9, mi-R-19a, miR-22 and miR-205 that promote EMT, fibrosis and tumorigenesis were up-regulated. Chlamydia induced EMT in vitro and in vivo, marked by the suppression of normal epithelial cell markers especially E-cadherin but up-regulation of mesenchymal markers of pathological EMT, including T-cadherin, MMP9, and fibronectin. Also, Chlamydia upregulated pro-EMT regulators, including the zinc finger E-box binding homeobox protein, ZEB1, Snail1/2, and thrombospondin1 (Thbs1), but down-regulated anti-EMT and fertility promoting proteins (i.e., the major gap junction protein connexin 43 (Cx43), Mets1, Add1Scarb1 and MARCKSL1). T cell-derived TNF-alpha signaling was required for chlamydial-induced infertility and caspase inhibitors prevented both infertility and EMT. Thus, chlamydial-induced T cell-derived TNF-alpha activated caspases that inactivated dicer, causing alteration in the expression of reproductive epithelial miRNAs and induction of EMT. EMT causes epithelial malfunction, fibrosis, infertility, and the enhancement of tumorigenesis of HPV

  2. Role of Epithelial-Mesenchyme Transition in Chlamydia Pathogenesis.

    PubMed

    Igietseme, Joseph U; Omosun, Yusuf; Stuchlik, Olga; Reed, Matthew S; Partin, James; He, Qing; Joseph, Kahaliah; Ellerson, Debra; Bollweg, Brigid; George, Zenas; Eko, Francis O; Bandea, Claudiu; Liu, Hsi; Yang, Genyan; Shieh, Wun-Ju; Pohl, Jan; Karem, Kevin; Black, Carolyn M

    2015-01-01

    Chlamydia trachomatis genital infection in women causes serious adverse reproductive complications, and is a strong co-factor for human papilloma virus (HPV)-associated cervical epithelial carcinoma. We tested the hypothesis that Chlamydia induces epithelial-mesenchyme transition (EMT) involving T cell-derived TNF-alpha signaling, caspase activation, cleavage inactivation of dicer and dysregulation of micro-RNA (miRNA) in the reproductive epithelium; the pathologic process of EMT causes fibrosis and fertility-related epithelial dysfunction, and also provides the co-factor function for HPV-related cervical epithelial carcinoma. Using a combination of microarrays, immunohistochemistry and proteomics, we showed that chlamydia altered the expression of crucial miRNAs that control EMT, fibrosis and tumorigenesis; specifically, miR-15a, miR-29b, miR-382 and MiR-429 that maintain epithelial integrity were down-regulated, while miR-9, mi-R-19a, miR-22 and miR-205 that promote EMT, fibrosis and tumorigenesis were up-regulated. Chlamydia induced EMT in vitro and in vivo, marked by the suppression of normal epithelial cell markers especially E-cadherin but up-regulation of mesenchymal markers of pathological EMT, including T-cadherin, MMP9, and fibronectin. Also, Chlamydia upregulated pro-EMT regulators, including the zinc finger E-box binding homeobox protein, ZEB1, Snail1/2, and thrombospondin1 (Thbs1), but down-regulated anti-EMT and fertility promoting proteins (i.e., the major gap junction protein connexin 43 (Cx43), Mets1, Add1Scarb1 and MARCKSL1). T cell-derived TNF-alpha signaling was required for chlamydial-induced infertility and caspase inhibitors prevented both infertility and EMT. Thus, chlamydial-induced T cell-derived TNF-alpha activated caspases that inactivated dicer, causing alteration in the expression of reproductive epithelial miRNAs and induction of EMT. EMT causes epithelial malfunction, fibrosis, infertility, and the enhancement of tumorigenesis of HPV

  3. Role of Epithelial-Mesenchyme Transition in Chlamydia Pathogenesis

    PubMed Central

    Igietseme, Joseph U.; Omosun, Yusuf; Stuchlik, Olga; Reed, Matthew S.; Partin, James; He, Qing; Joseph, Kahaliah; Ellerson, Debra; Bollweg, Brigid; George, Zenas; Eko, Francis O.; Bandea, Claudiu; Liu, Hsi; Yang, Genyan; Shieh, Wun-Ju; Pohl, Jan; Karem, Kevin; Black, Carolyn M.

    2015-01-01

    Chlamydia trachomatis genital infection in women causes serious adverse reproductive complications, and is a strong co-factor for human papilloma virus (HPV)-associated cervical epithelial carcinoma. We tested the hypothesis that Chlamydia induces epithelial-mesenchyme transition (EMT) involving T cell-derived TNF-alpha signaling, caspase activation, cleavage inactivation of dicer and dysregulation of micro-RNA (miRNA) in the reproductive epithelium; the pathologic process of EMT causes fibrosis and fertility-related epithelial dysfunction, and also provides the co-factor function for HPV-related cervical epithelial carcinoma. Using a combination of microarrays, immunohistochemistry and proteomics, we showed that chlamydia altered the expression of crucial miRNAs that control EMT, fibrosis and tumorigenesis; specifically, miR-15a, miR-29b, miR-382 and MiR-429 that maintain epithelial integrity were down-regulated, while miR-9, mi-R-19a, miR-22 and miR-205 that promote EMT, fibrosis and tumorigenesis were up-regulated. Chlamydia induced EMT in vitro and in vivo, marked by the suppression of normal epithelial cell markers especially E-cadherin but up-regulation of mesenchymal markers of pathological EMT, including T-cadherin, MMP9, and fibronectin. Also, Chlamydia upregulated pro-EMT regulators, including the zinc finger E-box binding homeobox protein, ZEB1, Snail1/2, and thrombospondin1 (Thbs1), but down-regulated anti-EMT and fertility promoting proteins (i.e., the major gap junction protein connexin 43 (Cx43), Mets1, Add1Scarb1 and MARCKSL1). T cell-derived TNF-alpha signaling was required for chlamydial-induced infertility and caspase inhibitors prevented both infertility and EMT. Thus, chlamydial-induced T cell-derived TNF-alpha activated caspases that inactivated dicer, causing alteration in the expression of reproductive epithelial miRNAs and induction of EMT. EMT causes epithelial malfunction, fibrosis, infertility, and the enhancement of tumorigenesis of HPV

  4. Live Brucella abortus rough vaccine strain RB51 stimulates enhanced innate immune response in vitro compared to rough vaccine strain RB51SOD and virulent smooth strain 2308 in murine bone marrow-derived dendritic cells.

    PubMed

    Surendran, Naveen; Hiltbold, Elizabeth M; Heid, Bettina; Sriranganathan, Nammalwar; Boyle, Stephen M; Zimmerman, Kurt L; Makris, Melissa R; Witonsky, Sharon G

    2011-01-10

    Brucella spp. are Gram-negative, coccobacillary, facultative intracellular pathogens. B. abortus strain 2308 is a pathogenic strain affecting cattle and humans. Rough B. abortus strain RB51, which lacks the O-side chain of lipopolysaccharide (LPS), is the live attenuated USDA approved vaccine for cattle in the United States. Strain RB51SOD, which overexpresses Cu-Zn superoxide dismutase (SOD), has been shown to confer better protection than strain RB51 in a murine model. Protection against brucellosis is mediated by a strong CD4+ Th(1) and CD8+ Tc(1) adaptive immune response. In order to stimulate a robust adaptive response, a solid innate immune response, including that mediated by dendritic cells, is essential. As dendritic cells (DCs) are highly susceptible to Brucella infection, it is possible that pathogenic strains could limit the innate and thereby adaptive immune response. By contrast, vaccine strains could limit or bolster the innate and subsequent adaptive immune response. Identifying how Brucella vaccines stimulate innate and adaptive immunity is critical for enhancing vaccine efficacy. The ability of rough vaccine strains RB51 and RB51SOD to stimulate DC function has not been characterized. We report that live rough vaccine strain RB51 induced significantly better (p ≤ 0.05) DC maturation and function compared to either strain RB51SOD or smooth virulent strain 2308, based on costimulatory marker expression and cytokine production.

  5. Genotyping of Brucella melitensis and Brucella abortus strains currently circulating in Xinjiang, China.

    PubMed

    Sun, Ming-Jun; Di, Dong-Dong; Li, Yan; Zhang, Zhi-Cheng; Yan, Hao; Tian, Li-Li; Jing, Zhi-Gang; Li, Jin-Ping; Jiang, Hai; Fan, Wei-Xing

    2016-10-01

    Brucellosis is a well-known zoonotic disease that can cause severe economic and healthcare losses. Xinjiang, one of the biggest livestock husbandry sectors in China, has gone through increasing incidence of brucellosis in cattle and small ruminants recently. In this paper, 50 B. melitensis strains and 9 B. abortus strains collected from across Xinjiang area (from 2010 to 2015) were genotyped using multiple locus variable-number tandem-repeat (VNTR) analysis (MLVA) and multi-locus sequence typing (MLST). Based on 8 loci (MLVA-8), 50 B. melitensis strains were classified into three genotypes. Genotypes 42 (n=38, 76%) and 63 (n=11, 22%) were part of the East Mediterranean group, and one genotype with pattern of 1-5-3-13-2-4-3-2 represents a single-locus variant from genotype 63. MLVA-16 resolved 50 B. melitensis strains into 28 genotypes, of which 15 are unique to Xinjiang and 10 are in common with those in adjacent country Kazakhstan and neighboring provinces of China. Minimum Spanning Tree (MST) analysis implies that B. melitensis strains collected from across Kazakhstan, Xinjiang and China areas may share a common origin. Nine B. abortus strains were sorted into three genotypes by MLVA-8, genotypes 36 (n=7, 77.8%), 86 (n=1, 11.1%) and a new genotype with pattern of 4-5-3-13-2-2-3-1. Each B. abortus strain showed distinct MLVA-16 genotypes, suggesting that B. abortus species may possess more genetic diversity than B. melitensis. Using MLST, most B. melitensis strains (n=49) were identified as sequence type ST8, and most B. abortus strains (n=8) were recognized as ST2. Two new sequence types, ST37 and ST38, represented by single strain from B. melitensis and B. abortus species respectively, were also detected in this study. These results could facilitate the pathogen surveillance in the forthcoming eradication programs and serve as a guide in source tracking in case of new outbreaks occur.

  6. Genotyping of Brucella melitensis and Brucella abortus strains currently circulating in Xinjiang, China.

    PubMed

    Sun, Ming-Jun; Di, Dong-Dong; Li, Yan; Zhang, Zhi-Cheng; Yan, Hao; Tian, Li-Li; Jing, Zhi-Gang; Li, Jin-Ping; Jiang, Hai; Fan, Wei-Xing

    2016-10-01

    Brucellosis is a well-known zoonotic disease that can cause severe economic and healthcare losses. Xinjiang, one of the biggest livestock husbandry sectors in China, has gone through increasing incidence of brucellosis in cattle and small ruminants recently. In this paper, 50 B. melitensis strains and 9 B. abortus strains collected from across Xinjiang area (from 2010 to 2015) were genotyped using multiple locus variable-number tandem-repeat (VNTR) analysis (MLVA) and multi-locus sequence typing (MLST). Based on 8 loci (MLVA-8), 50 B. melitensis strains were classified into three genotypes. Genotypes 42 (n=38, 76%) and 63 (n=11, 22%) were part of the East Mediterranean group, and one genotype with pattern of 1-5-3-13-2-4-3-2 represents a single-locus variant from genotype 63. MLVA-16 resolved 50 B. melitensis strains into 28 genotypes, of which 15 are unique to Xinjiang and 10 are in common with those in adjacent country Kazakhstan and neighboring provinces of China. Minimum Spanning Tree (MST) analysis implies that B. melitensis strains collected from across Kazakhstan, Xinjiang and China areas may share a common origin. Nine B. abortus strains were sorted into three genotypes by MLVA-8, genotypes 36 (n=7, 77.8%), 86 (n=1, 11.1%) and a new genotype with pattern of 4-5-3-13-2-2-3-1. Each B. abortus strain showed distinct MLVA-16 genotypes, suggesting that B. abortus species may possess more genetic diversity than B. melitensis. Using MLST, most B. melitensis strains (n=49) were identified as sequence type ST8, and most B. abortus strains (n=8) were recognized as ST2. Two new sequence types, ST37 and ST38, represented by single strain from B. melitensis and B. abortus species respectively, were also detected in this study. These results could facilitate the pathogen surveillance in the forthcoming eradication programs and serve as a guide in source tracking in case of new outbreaks occur. PMID:27521159

  7. Evolution and Conservation of Predicted Inclusion Membrane Proteins in Chlamydiae

    PubMed Central

    Lutter, Erika I.; Martens, Craig; Hackstadt, Ted

    2012-01-01

    Chlamydia spp. are obligate intracellular pathogens that replicate within a vacuole termed the inclusion. Chlamydiae extensively modify the inclusion membrane via the insertion of chlamydial inclusion membrane proteins (Incs) which decorate the cytosolic face of the inclusion. We have assessed the overall relatedness and phylogeny of Incs in order to identify potential evolutionary trends. Despite a high degree of conservation among Incs within C. trachomatis serovars, phylogenetic analysis showed that some Incs cluster according to clinical groupings suggesting that certain Incs may contribute to tissue tropism. Bioinformatic predictions identified Incs in five chlamydial species: 55 in C. trachomatis, 68 in C. felis, 92 in C. pneumoniae, 79 in C. caviae, and 54 in C. muridarum. Inc homologues were compared between chlamydial species and 23 core Incs were identified as shared among all species. Genomic expansion of Incs was identified in C. pneumoniae, C. caviae, and C. felis but not C. trachomatis or C. muridarum. PMID:22454599

  8. 2015 European guideline on the management of Chlamydia trachomatis infections.

    PubMed

    Lanjouw, E; Ouburg, S; de Vries, H J; Stary, A; Radcliffe, K; Unemo, M

    2016-04-01

    Chlamydia trachomatis infections, which most frequently are asymptomatic, are major public health concerns globally. The 2015 European C. trachomatis guideline provides: up-to-date guidance regarding broader indications for testing and treatment of C. trachomatis infections; a clearer recommendation of using exclusively-validated nucleic acid amplification tests for diagnosis; advice on (repeated) C. trachomatis testing; the recommendation of increased testing to reduce the incidence of pelvic inflammatory disease and prevent exposure to infection; and recommendations to identify, verify and report C. trachomatis variants. Improvement of access to testing, test performance, diagnostics, antimicrobial treatment and follow-up of C. trachomatis patients are crucial to control its spread. For detailed background, evidence base and discussions, see the background review for the present 2015 European guideline on the management of Chlamydia trachomatis infections (Lanjouw E, et al. Int J STD AIDS. 2015).

  9. Azithromycin versus Doxycycline for Urogenital Chlamydia trachomatis Infection

    PubMed Central

    Geisler, William M.; Uniyal, Apurva; Lee, Jeannette Y.; Lensing, Shelly Y.; Johnson, Shacondra; Perry, Raymond C.W.; Kadrnka, Carmel M.; Kerndt, Peter R.

    2016-01-01

    BACKGROUND Urogenital Chlamydia trachomatis infection remains prevalent and causes substantial reproductive morbidity. Recent studies have raised concern about the efficacy of azithromycin for the treatment of chlamydia infection. METHODS We conducted a randomized trial comparing oral azithromycin with doxycycline for the treatment of urogenital chlamydia infection among adolescents in youth correctional facilities, to evaluate the noninferiority of azithromycin (1 g in one dose) to doxycycline (100 mg twice daily for 7 days). The treatment was directly observed. The primary end point was treatment failure at 28 days after treatment initiation, with treatment failure determined on the basis of nucleic acid amplification testing, sexual history, and outer membrane protein A (OmpA) genotyping of C. trachomatis strains. RESULTS Among the 567 participants enrolled, 284 were randomly assigned to receive azithromycin, and 283 were randomly assigned to receive doxycycline. A total of 155 participants in each treatment group (65% male) made up the per-protocol population. There were no treatment failures in the doxycycline group. In the azithromycin group, treatment failure occurred in 5 participants (3.2%; 95% confidence interval, 0.4 to 7.4%). The observed difference in failure rates between the treatment groups was 3.2 percentage points, with an upper boundary of the 90% confidence interval of 5.9 percentage points, which exceeded the prespecified absolute 5-percentage-point cutoff for establishing the noninferiority of azithromycin. CONCLUSIONS In the context of a closed population receiving directly observed treatment for urogenital chlamydia infection, the efficacy of azithromycin was 97%, and the efficacy of doxycycline was 100%. The noninferiority of azithromycin was not established in this setting. (Funded by the National Institute of Allergy and Infectious Diseases; ClinicalTrials.gov number, NCT00980148.) PMID:26699167

  10. Detection of Chlamydia pneumoniae by polymerase chain reaction.

    PubMed

    Campbell, L A; Perez Melgosa, M; Hamilton, D J; Kuo, C C; Grayston, J T

    1992-02-01

    While criteria for serodiagnosis of Chlamydia pneumoniae infection are well established, isolation of the organism is often difficult. To increase detection of this organism, C. pneumoniae-specific sequences were identified to permit amplification of C. pneumoniae by polymerase chain reaction (PCR). A cloned C. pneumoniae 474-bp PstI fragment was shown by dot blot and Southern hybridization to differentiate C. pneumoniae from the other Chlamydia spp., react with all C. pneumoniae isolates tested, and not recognize DNA from normal throat flora or common respiratory tract agents. This cloned fragment was sequenced and primers for use in PCR were chosen on the bases of GenBank analysis, G + C ratio, and absence of secondary structure. All C. pneumoniae isolates tested were amplified by the HL-1-HR-1 primer pair or the HM-1-HR-1 primer pair, producing the expected 437- and 229-bp amplification products, respectively. None of the Chlamydia trachomatis serovars (B/TW-5/OT, C/TW-3/OT, D/UW-3/Cx, E/UW-5/Cx, F/UW-6/Cx, H/UW-4/Cx, I/UW-12/Ur, and L2/434/Bu), Chlamydia psittaci strains (Mn, 6BC, GPIC, FP, and OA), HeLa cells, or other organisms tested were amplified. Reaction conditions including MgCl2, oligonucleotides, and primer concentrations and temperature were optimized before application to clinical samples. Clinical specimens from patients from whom C. pneumoniae was isolated were also positive by PCR, while samples from patients with known C. trachomatis or C. psittaci infection were not amplified by PCR.

  11. Telling partners about chlamydia: how acceptable are the new technologies?

    PubMed Central

    2010-01-01

    Background Partner notification is accepted as a vital component in the control of chlamydia. However, in reality, many sexual partners of individuals diagnosed with chlamydia are never informed of their risk. The newer technologies of email and SMS have been used as a means of improving partner notification rates. This study explored the use and acceptability of different partner notification methods to help inform the development of strategies and resources to increase the number of partners notified. Methods Semi-structured telephone interviews were conducted with 40 people who were recently diagnosed with chlamydia from three sexual health centres and two general practices across three Australian jurisdictions. Results Most participants chose to contact their partners either in person (56%) or by phone (44%). Only 17% chose email or SMS. Participants viewed face-to-face as the "gold standard" in partner notification because it demonstrated caring, respect and courage. Telephone contact, while considered insensitive by some, was often valued because it was quick, convenient and less confronting. Email was often seen as less personal while SMS was generally considered the least acceptable method for telling partners. There was also concern that emails and SMS could be misunderstood, not taken seriously or shown to others. Despite these, email and SMS were seen to be appropriate and useful in some circumstances. Letters, both from the patients or from their doctor, were viewed more favourably but were seldom used. Conclusion These findings suggest that many people diagnosed with chlamydia are reluctant to use the new technologies for partner notification, except in specific circumstances, and our efforts in developing partner notification resources may best be focused on giving patients the skills and confidence for personal interaction. PMID:20211029

  12. Fluorescent labeling reliably identifies Chlamydia trachomatis in living human endometrial cells and rapidly and accurately quantifies chlamydial inclusion forming units.

    PubMed

    Vicetti Miguel, Rodolfo D; Henschel, Kevin J; Dueñas Lopez, Fiorela C; Quispe Calla, Nirk E; Cherpes, Thomas L

    2015-12-01

    Chlamydia replication requires host lipid acquisition, allowing flow cytometry to identify Chlamydia-infected cells that accumulated fluorescent Golgi-specific lipid. Herein, we describe modifications to currently available methods that allow precise differentiation between uninfected and Chlamydia trachomatis-infected human endometrial cells and rapidly and accurately quantify chlamydial inclusion forming units.

  13. Serological crossreactivity between Brucella abortus and Yersinia enterocolitica 0:9 I immunoblot analysis of the antibody response to Brucella protein antigens in bovine brucellosis.

    PubMed

    Kittelberger, R; Hilbink, F; Hansen, M F; Penrose, M; de Lisle, G W; Letesson, J J; Garin-Bastuji, B; Searson, J; Fossati, C A; Cloeckaert, A

    1995-12-01

    Sera from three groups of Brucella abortus infected cattle were examined in immunoblots with the following antigens: sodium dodecyl sulfate/mercapto ethanol (SDS/ME) extracts of two rought B. abortus strains (45/20 and RB51) and rough B. ovis, smooth lipopolysaccharides (SLPS) from B. abortus strain 99 and Y. enterocolitica 0:9, and a cytoplasmic extract from smooth B. abortus strain 19-S. The sera groups were: (1) 26 sera from animals, experimentally infected with B. abortus strain 544, which were all positive in the conventional brucellosis serological tests; (2) 152 sera from naturally infected cattle herds with varying titres in the conventional brucellosis tests, and (3) 30 sera from naturally infected cattle with varying titres in the conventional brucellosis tests and from which B. abortus was cultured. B. abortus strain 99 and Y. enterocolitica serotype 0:9 SLPS staining showed up frequently in all sera groups and correlated well with the strength in the conventional brucellosis tests, confirming the immunodominance of SLPS in B. abortus infections. Another immunodominant component of 50-80 kDa was found in the rough B. abortus 45/20 antigen preparation but not in the B. abortus RB51 and in the B. ovis cell extracts. This component was also recognised by sera from Y. enterocolitica 0:9 infected cattle and is probably a protein-lipopolysaccharide complex. Although many of the sera from B. abortus infected cattle with high titres in the conventional brucellosis tests showed complex protein staining patterns in blots, no protein bands other than the 50-80 kDa bands were found to be immunodominant.

  14. Perforin-2 restricts growth of Chlamydia trachomatis in macrophages.

    PubMed

    Fields, K A; McCormack, R; de Armas, L R; Podack, E R

    2013-08-01

    Chlamydia trachomatis is a Gram-negative obligate intracellular bacterium that preferentially infects epithelial cells. Professional phagocytes provide C. trachomatis only a limited ability to survive and are proficient killers of chlamydiae. We present evidence herein that identifies a novel host defense protein, perforin-2, that plays a significant role in the eradication of C. trachomatis during the infection of macrophages. Knockdown of perforin-2 in macrophages did not alter the invasion of host cells but did result in chlamydial growth that closely mirrored that detected in HeLa cells. C trachomatis L2, serovar B, and serovar D and C. muridarum were all equally susceptible to perforin-2-mediated killing. Interestingly, induction of perforin-2 expression in epithelial cells is blocked during productive chlamydial growth, thereby protecting chlamydiae from bactericidal attack. Ectopic expression of perforin-2 in HeLa cells, however, does result in killing. Overall, our data implicate a new innate resistance protein in the control of chlamydial infection and may help explain why the macrophage environment is hostile to chlamydial growth. PMID:23753625

  15. Cost-effectiveness of Chlamydia Vaccination Programs for Young Women

    PubMed Central

    Chesson, Harrell W.; Gift, Thomas L.; Brunham, Robert C.; Bolan, Gail

    2015-01-01

    We explored potential cost-effectiveness of a chlamydia vaccine for young women in the United States by using a compartmental heterosexual transmission model. We tracked health outcomes (acute infections and sequelae measured in quality-adjusted life-years [QALYs]) and determined incremental cost-effectiveness ratios (ICERs) over a 50-year analytic horizon. We assessed vaccination of 14-year-old girls and catch-up vaccination for 15–24-year-old women in the context of an existing chlamydia screening program and assumed 2 prevaccination prevalences of 3.2% by main analysis and 3.7% by additional analysis. Estimated ICERs of vaccinating 14-year-old girls were $35,300/QALY by main analysis and $16,200/QALY by additional analysis compared with only screening. Catch-up vaccination for 15–24-year-old women resulted in estimated ICERs of $53,200/QALY by main analysis and $26,300/QALY by additional analysis. The ICER was most sensitive to prevaccination prevalence for women, followed by cost of vaccination, duration of vaccine-conferred immunity, and vaccine efficacy. Our results suggest that a successful chlamydia vaccine could be cost-effective. PMID:25989525

  16. Characterization of interactions between inclusion membrane proteins from Chlamydia trachomatis

    PubMed Central

    Gauliard, Emilie; Ouellette, Scot P.; Rueden, Kelsey J.; Ladant, Daniel

    2015-01-01

    Chlamydiae are obligate intracellular pathogens of eukaryotes. The bacteria grow in an intracellular vesicle called an inclusion, the membrane of which is heavily modified by chlamydial proteins called Incs (Inclusion membrane proteins). Incs represent 7–10% of the genomes of Chlamydia and, given their localization at the interface between the host and the pathogen, likely play a key role in the development and pathogenesis of the bacterium. However, their functions remain largely unknown. Here, we characterized the interaction properties between various Inc proteins of C. trachomatis, using a bacterial two-hybrid (BACTH) method suitable for detecting interactions between integral membrane proteins. To validate this approach, we first examined the oligomerization properties of the well-characterized IncA protein and showed that both the cytoplasmic domain and the transmembrane region independently contribute to IncA oligomerization. We then analyzed a set of Inc proteins and identified novel interactions between these components. Two small Incs, IncF, and Ct222, were found here to interact with many other Inc proteins and may thus represent interaction nodes within the inclusion membrane. Our data suggest that the Inc proteins may assemble in the membrane of the inclusion to form specific multi-molecular complexes in an hierarchical and temporal manner. These studies will help to better define the putative functions of the Inc proteins in the infectious process of Chlamydia. PMID:25717440

  17. Immunity and vaccines against sexually transmitted Chlamydia trachomatis infection

    PubMed Central

    Howie, Sarah E. M.; Horner, Patrick J.; Horne, Andrew W.; Entrican, Gary

    2011-01-01

    Purpose of review To review recent findings on immunity and vaccine development to Chlamydia trachomatis. Recent findings There is increasing knowledge on the interactions between Chlamydia trachomatis and infected host cells. During genital infection the organism avoids generating protective immunity but immune responses to a number of chlamydial proteins have been associated with reproductive tract pathology. Various vaccine and adjuvant preparations have been tried experimentally. Information generated by proteomics and complex studies of serological and T-lymphocyte immune responses points to novel vaccine candidates. Summary Chlamydia trachomatis, an obligate intracellular bacterium, is the commonest sexually transmitted infection worldwide and is associated with reproductive pathology. To develop rational vaccines it is necessary to understand the complex life-cycle of the organism, the host immune response to infection and how these relate to disease. Infection does not prevent reinfection and antibiotic treatment prevents antibody production at a population level. It remains unclear what type of immune response would be sufficient to prevent infection and/or reinfection. Although the prevalence and demographics of infection and the severity of disease associations suggest it would be desirable, there is no vaccine currently available. A number of studies have identified novel vaccine candidates. PMID:21124214

  18. Prime-booster vaccination of cattle with an influenza viral vector Brucella abortus vaccine induces a long-term protective immune response against Brucella abortus infection.

    PubMed

    Tabynov, Kaissar; Yespembetov, Bolat; Ryskeldinova, Sholpan; Zinina, Nadezhda; Kydyrbayev, Zhailaubay; Kozhamkulov, Yerken; Inkarbekov, Dulat; Sansyzbay, Abylai

    2016-01-20

    This study analyzed the duration of the antigen-specific humoral and T-cell immune responses and protectiveness of a recently-developed influenza viral vector Brucella abortus (Flu-BA) vaccine expressing Brucella proteins Omp16 and L7/L12 and containing the adjuvant Montadine Gel01 in cattle. At 1 month post-booster vaccination (BV), both humoral (up to 3 months post-BV; GMT IgG ELISA titer 214±55 to 857±136, with a prevalence of IgG2a over IgG1 isotype antibodies) and T-cell immune responses were observed in vaccinated heifers (n=35) compared to control animals (n=35, injected with adjuvant/PBS only). A pronounced T-cell immune response was induced and maintained for 12 months post-BV, as indicated by the lymphocyte stimulation index (2.7±0.4 to 10.1±0.9 cpm) and production of IFN-γ (13.7±1.7 to 40.0±3.0 ng/ml) at 3, 6, 9, and 12 months post-BV. Prime-boost vaccination provided significant protection against B. abortus infection at 3, 6, 9 and 12 months (study duration) post-BV (7 heifers per time point; alpha=0.03-0.01 vs. control group). Between 57.1 and 71.4% of vaccinated animals showed no signs of B. abortus infection (or Brucella isolation) at 3, 6, 9 and 12 months post-BV; the severity of infection, as indicated by the index of infection (P=0.0003 to <0.0001) and rates of Brucella colonization (P=0.03 to <0.0001), was significantly lower for vaccinated diseased animals than appropriate control animals. Good protection from B. abortus infection was also observed among pregnant vaccinated heifers (alpha=0.03), as well as their fetuses and calves (alpha=0.01), for 12 months post-BV. Additionally, 71.4% of vaccinated heifers calved successfully whereas all pregnant control animals aborted (alpha=0.01). Prime-boost vaccination of cattle with Flu-BA induces an antigen-specific humoral and pronounced T cell immune response and most importantly provides good protectiveness, even in pregnant heifers, for at least 12 months post-BV.

  19. Prime-booster vaccination of cattle with an influenza viral vector Brucella abortus vaccine induces a long-term protective immune response against Brucella abortus infection.

    PubMed

    Tabynov, Kaissar; Yespembetov, Bolat; Ryskeldinova, Sholpan; Zinina, Nadezhda; Kydyrbayev, Zhailaubay; Kozhamkulov, Yerken; Inkarbekov, Dulat; Sansyzbay, Abylai

    2016-01-20

    This study analyzed the duration of the antigen-specific humoral and T-cell immune responses and protectiveness of a recently-developed influenza viral vector Brucella abortus (Flu-BA) vaccine expressing Brucella proteins Omp16 and L7/L12 and containing the adjuvant Montadine Gel01 in cattle. At 1 month post-booster vaccination (BV), both humoral (up to 3 months post-BV; GMT IgG ELISA titer 214±55 to 857±136, with a prevalence of IgG2a over IgG1 isotype antibodies) and T-cell immune responses were observed in vaccinated heifers (n=35) compared to control animals (n=35, injected with adjuvant/PBS only). A pronounced T-cell immune response was induced and maintained for 12 months post-BV, as indicated by the lymphocyte stimulation index (2.7±0.4 to 10.1±0.9 cpm) and production of IFN-γ (13.7±1.7 to 40.0±3.0 ng/ml) at 3, 6, 9, and 12 months post-BV. Prime-boost vaccination provided significant protection against B. abortus infection at 3, 6, 9 and 12 months (study duration) post-BV (7 heifers per time point; alpha=0.03-0.01 vs. control group). Between 57.1 and 71.4% of vaccinated animals showed no signs of B. abortus infection (or Brucella isolation) at 3, 6, 9 and 12 months post-BV; the severity of infection, as indicated by the index of infection (P=0.0003 to <0.0001) and rates of Brucella colonization (P=0.03 to <0.0001), was significantly lower for vaccinated diseased animals than appropriate control animals. Good protection from B. abortus infection was also observed among pregnant vaccinated heifers (alpha=0.03), as well as their fetuses and calves (alpha=0.01), for 12 months post-BV. Additionally, 71.4% of vaccinated heifers calved successfully whereas all pregnant control animals aborted (alpha=0.01). Prime-boost vaccination of cattle with Flu-BA induces an antigen-specific humoral and pronounced T cell immune response and most importantly provides good protectiveness, even in pregnant heifers, for at least 12 months post-BV. PMID:26709638

  20. Vector Development for the Expression of Foreign Proteins in the Vaccine Strain Brucella abortus S19

    PubMed Central

    Comerci, Diego J.; Pollevick, Guido D.; Vigliocco, Ana M.; Frasch, Alberto C. C.; Ugalde, Rodolfo A.

    1998-01-01

    A vector for the expression of foreign antigens in the vaccine strain Brucella abortus S19 was developed by using a DNA fragment containing the regulatory sequences and the signal peptide of the Brucella bcsp31 gene. This fragment was cloned in broad-host-range plasmid pBBR4MCS, resulting in plasmid pBEV. As a reporter protein, a repetitive antigen of Trypanosoma cruzi was used. The recombinant fusion protein is stably expressed and secreted into the Brucella periplasmic space, inducing a good antibody response against the T. cruzi antigen. The expression of the repetitive antigen in Brucella neither altered its growth pattern nor generated a toxic or lethal effect during experimental infection. The application of this strategy for the generation of live recombinant vaccines and the tagging of B. abortus S19 vaccine is discussed. This is the first time that a recombinant protein has been expressed in the periplasm of brucellae. PMID:9673273

  1. Infection of cattle in Kenya with Brucella abortus biovar 3 and Brucella melitensis biovar 1 genotypes.

    PubMed

    Muendo, Esther N; Mbatha, Peter M; Macharia, Joseph; Abdoel, Theresia H; Janszen, Paul V; Pastoor, Rob; Smits, Henk L

    2012-01-01

    Brucella melitensis biovar 1 was isolated from bovine milk samples from a herd in central Kenya, and Brucella abortus biovar 3 was isolated from aborted fetus materials and vaginal discharge fluids from cattle in central and eastern provinces of Kenya. All infections including those with B. melitensis were in cattle with reproductive problems kept in mixed herds indicating that cross infection occurs from small ruminants. Multiple-locus variable-number tandem repeat analysis genotyping revealed a close molecular homology of the B. melitensis isolates with an isolate from Israel and a close homology of the B. abortus isolates with an isolate from Uganda indicating that these genotypes have a wide geographic distribution. Infection of cattle with B. melitensis may complicate the control of brucellosis in this country.

  2. A case of unusual septic knee arthritis with Brucella abortus after arthroscopic meniscus surgery.

    PubMed

    Lee, Keun Hwa; Kang, Hyunseong; Kim, Taejung; Choi, Sungwook

    2016-01-01

    We present a 51-year-old male patient with Brucella abortus septic arthritis in the right knee following arthroscopic meniscus surgery. He had eaten a traditional dish of raw minced cattle conceptus (bovine fetus) that was prepared after the cow was slaughtered. Despite treatment with empirical antibiotics and debridement of the postoperative surgical wound, the infection persisted without improvement. Polymerase chain reaction sequencing identified Brucella abortus from tissue samples obtained from the patient. After confirmation of the diagnosis of brucellar infection, antibiotics were replaced with doxycycline and rifampin, which were used for 4 months. In patients with a non-specific arthralgia who eat raw meat or live close to animals, it is important to consider the possibility of septic arthritis due to infection with Brucella spp.

  3. Comparative proteome analysis of laboratory grown Brucella abortus 2308 and Brucella melitensis 16M.

    PubMed

    Eschenbrenner, Michel; Horn, Troy A; Wagner, Mary Ann; Mujer, Cesar V; Miller-Scandle, Tabbi L; DelVecchio, Vito G

    2006-07-01

    Brucella species are pathogenic agents that cause brucellosis, a debilitating zoonotic disease that affects a large variety of domesticated animals and humans. Brucella melitensis and Brucella abortus are considered major health threats because of their highly infectious nature and worldwide occurrence. The availability of the annotated genomes for these two species has allowed a comparative proteomics study of laboratory grown B. melitensis 16M and B. abortus 2308 by two-dimensional (2-D) gel electrophoresis and peptide mass fingerprinting. Computer-assisted analysis of the different 2-D gel images of strains 16M and 2308 revealed significant quantitative and qualitative differences in their protein expression patterns. Proteins involved in membrane transport, particularly the high affinity amino acids binding proteins, and those involved in Sec-dependent secretion systems related to type IV and type V secretion systems, were differentially expressed. Differential expression of these proteins may be responsible for conferring specific host preference in the two strains 2308 and 16M.

  4. A case of unusual septic knee arthritis with Brucella abortus after arthroscopic meniscus surgery.

    PubMed

    Lee, Keun Hwa; Kang, Hyunseong; Kim, Taejung; Choi, Sungwook

    2016-01-01

    We present a 51-year-old male patient with Brucella abortus septic arthritis in the right knee following arthroscopic meniscus surgery. He had eaten a traditional dish of raw minced cattle conceptus (bovine fetus) that was prepared after the cow was slaughtered. Despite treatment with empirical antibiotics and debridement of the postoperative surgical wound, the infection persisted without improvement. Polymerase chain reaction sequencing identified Brucella abortus from tissue samples obtained from the patient. After confirmation of the diagnosis of brucellar infection, antibiotics were replaced with doxycycline and rifampin, which were used for 4 months. In patients with a non-specific arthralgia who eat raw meat or live close to animals, it is important to consider the possibility of septic arthritis due to infection with Brucella spp. PMID:27130400

  5. Immune responses and protection against infection and abortion in cattle experimentally vaccinated with mutant strains of Brucella abortus.

    PubMed

    Cheville, N F; Stevens, M G; Jensen, A E; Tatum, F M; Halling, S M

    1993-10-01

    Twenty-four 10-month-old Polled Hereford heifers were inoculated SC with live cells of one of the following strains of Brucella abortus: S19 delta 31K (n = 4), S19 delta SOD (n = 4), RB51 (n = 4), and strain 19 (n = 6); controls (n = 6) were given saline solution. Heifers given the deletion mutants S19 delta 31K and S19 delta SOD, and those given strain 19 developed antibody responses to B abortus and cutaneous reactions to brucellin. Heifers given strain RB51 did not develop antibodies that reacted in the standard tube agglutination test, but sera reacted in tests, using an antibody dot-blot assay containing RB51 antigen. The S19 delta 31K and S19 delta SOD strains of B abortus isolated from lymph node tissue after vaccination did not differ genetically from the master stock strain. All heifers were bred naturally at 16 to 17 months of age, and were challenge-exposed intraconjunctivally with virulent B abortus strain 2308 during the fifth month of pregnancy. All vaccinated heifers were protected (ie, none aborted and none had B abortus isolated from their tissues after parturition). Calves born from vaccinated dams were free of B abortus. Antibody responses in heifers after challenge exposure were an indicator of immunity. All 5 control heifers (nonvaccinated) developed serum antibodies after challenge exposure; 3 aborted, and 1 delivered a small, weak calf at 8.5 months of gestation. Thus live mutant strains of B abortus can induce protective immunity when given at 10 months of age, and strain RB51 is a strong candidate for further testing.

  6. Brucella abortus strain RB51 as a vector for heterologous protein expression and induction of specific Th1 type immune responses.

    PubMed

    Vemulapalli, R; He, Y; Boyle, S M; Sriranganathan, N; Schurig, G G

    2000-06-01

    Brucella abortus strain RB51 is a stable, rough, attenuated mutant widely used as a live vaccine for bovine brucellosis. Our ultimate goal is to develop strain RB51 as a preferential vector for the delivery of protective antigens of other intracellular pathogens to which the induction of a strong Th1 type of immune response is needed for effective protection. As a first step in that direction, we studied the expression of a foreign reporter protein, beta-galactosidase of Escherichia coli, and the 65-kDa heat shock protein (HSP65) of Mycobacterium bovis in strain RB51. We cloned the promoter sequences of Brucella sodC and groE genes in pBBR1MCS to generate plasmids pBBSODpro and pBBgroE, respectively. The genes for beta-galactosidase (lacZ) and HSP65 were cloned in these plasmids and used to transform strain RB51. An enzyme assay in the recombinant RB51 strains indicated that the level of beta-galactosidase expression is higher under the groE promoter than under the sodC promoter. In strain RB51 containing pBBgroE/lacZ, but not pBBSODpro/lacZ, increased levels of beta-galactosidase expression were observed after subjecting the bacteria to heat shock or following internalization into macrophage-like J774A.1 cells. Mice vaccinated with either of the beta-galactosidase-expressing recombinant RB51 strains developed specific antibodies of predominantly the immunoglobulin G2a (IgG2a) isotype, and in vitro stimulation of their splenocytes with beta-galactosidase induced the secretion of gamma interferon (IFN-gamma), but not interleukin-4 (IL-4). A Th1 type of immune response to HSP65, as indicated by the presence of specific serum IgG2a, but not IgG1, antibodies, and IFN-gamma, but not IL-4, secretion by the specific-antigen-stimulated splenocytes, was also detected in mice vaccinated with strain RB51 containing pBBgroE/hsp65. Studies with mice indicated that expression of beta-galactosidase or HSP65 did not alter either the attenuation characteristics of strain RB51 or

  7. Brucella abortus Induces the Premature Death of Human Neutrophils through the Action of Its Lipopolysaccharide.

    PubMed

    Barquero-Calvo, Elías; Mora-Cartín, Ricardo; Arce-Gorvel, Vilma; de Diego, Juana L; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Guzmán-Verri, Caterina; Buret, Andre G; Gorvel, Jean-Pierre; Moreno, Edgardo

    2015-05-01

    Most bacterial infections induce the activation of polymorphonuclear neutrophils (PMNs), enhance their microbicidal function, and promote the survival of these leukocytes for protracted periods of time. Brucella abortus is a stealthy pathogen that evades innate immunity, barely activates PMNs, and resists the killing mechanisms of these phagocytes. Intriguing clinical signs observed during brucellosis are the low numbers of Brucella infected PMNs in the target organs and neutropenia in a proportion of the patients; features that deserve further attention. Here we demonstrate that B. abortus prematurely kills human PMNs in a dose-dependent and cell-specific manner. Death of PMNs is concomitant with the intracellular Brucella lipopolysaccharide (Br-LPS) release within vacuoles. This molecule and its lipid A reproduce the premature cell death of PMNs, a phenomenon associated to the low production of proinflammatory cytokines. Blocking of CD14 but not TLR4 prevents the Br-LPS-induced cell death. The PMNs cell death departs from necrosis, NETosis and classical apoptosis. The mechanism of PMN cell death is linked to the activation of NADPH-oxidase and a modest but steadily increase of ROS mediators. These effectors generate DNA damage, recruitments of check point kinase 1, caspases 5 and to minor extent of caspase 4, RIP1 and Ca++ release. The production of IL-1β by PMNs was barely stimulated by B. abortus infection or Br-LPS treatment. Likewise, inhibition of caspase 1 did not hamper the Br-LPS induced PMN cell death, suggesting that the inflammasome pathway was not involved. Although activation of caspases 8 and 9 was observed, they did not seem to participate in the initial triggering mechanisms, since inhibition of these caspases scarcely blocked PMN cell death. These findings suggest a mechanism for neutropenia in chronic brucellosis and reveal a novel Brucella-host cross-talk through which B. abortus is able to hinder the innate function of PMN.

  8. Draft Genome Sequences of Two Brucella abortus Strains Isolated from Cattle and Pig.

    PubMed

    Sharma, Narinder Singh; Sunita, Thakhur; Arora, A K; Mudit, Chandra; Kaur, Paviter; Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2015-01-01

    We report the draft genome sequences of two Brucella abortus strains LMN1 and LMN2 isolated from cattle and pig. The LMN1 and LMN2 have the genome size of 3,395,952 bp and 3,334,792 bp, respectively. In addition to the conserved genes of Brucella, few novel regions showing similarity to the phages were identified in both strains.

  9. Draft Genome Sequences of Two Brucella abortus Strains Isolated from Cattle and Pig

    PubMed Central

    Sharma, Narinder Singh; Sunita, Thakhur; Arora, A K; Mudit, Chandra; Kaur, Paviter; Sankarasubramanian, Jagadesan; Vishnu, Udayakumar S; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2015-01-01

    We report the draft genome sequences of two Brucella abortus strains LMN1 and LMN2 isolated from cattle and pig. The LMN1 and LMN2 have the genome size of 3,395,952 bp and 3,334,792 bp, respectively. In addition to the conserved genes of Brucella, few novel regions showing similarity to the phages were identified in both strains. PMID:26816552

  10. Effects of gamma radiation and azathioprine on Brucella abortus infection in BALB/c mice

    SciTech Connect

    Elzer, P.H.; Rowe, G.E.; Enright, F.M.; Winter, A.J. )

    1991-06-01

    Sublethal irradiation of BALB/c mice 4 hours prior to inoculation with 5 {times} 10(4) virulent Brucella abortus, caused significant (P less than 0.01) reductions in bacterial numbers in comparison with numbers in unirradiated controls. Numbers of brucellae in the spleen were significantly lower by 5 days after inoculation and decreased thereafter, so that at 2 and 3 weeks after inoculation, there were up to 1,000-fold fewer organisms in the spleen of irradiated mice. The number of brucellae in the spleen increased in irradiated mice thereafter. The course of events in the liver was similar, but developed more slowly, and peak differences in bacterial numbers were about 1 log less. These phenomena were not attributable to differences in implantation of brucellae in the liver or spleen, nor to an abnormal distribution of organisms in other organs of irradiated mice. Irradiation of mice during the plateau phase of infection also resulted in significant (P less than 0.05) reductions in bacterial counts in the spleen during the succeeding 4 weeks. Macrophage activation in the spleen, measured by a Listeria monocytogenes-killing assay, was significantly (P less than 0.01) increased by irradiation alone at 1 week after inoculation and at that time was significantly (P less than 0.01) greater in B abortus-infected, irradiated mice than in B abortus-infected controls. Histologic, cytologic, and immunologic studies revealed that the decrease in numbers of organisms between 1 and 2 weeks after inoculation in irradiated mice occurred at a time when their immune response to B abortus was suppressed and when numbers of neutrophils and monocytes infiltrating the spleen were significantly (P less than 0.01) diminished.

  11. Evaluation of transmission of Brucella abortus strain 19 in bison by intravaginal, intrauterine, and intraconjunctival inoculation.

    PubMed

    Uhrig, Samantha R; Nol, Pauline; McCollum, Matt; Salman, Mo; Rhyan, Jack C

    2013-07-01

    Bovine brucellosis, caused by the bacterium Brucella abortus, is endemic in bison (Bison bison) and elk (Cervus elaphus nelsoni) populations in the area of Yellowstone National Park, USA. Two strategies have been proposed to reduce the risk of transmission of disease in bison: remote vaccination with the vaccine RB51, and the use of immunocontraception of bison to decrease shedding of organisms from infected females. The frequent occurrence of venereal transmission in bison would complicate either of these strategies, requiring vaccination of males as well as females, and rendering immunocontraception less effective in reducing transmission of B. abortus. To address the question of venereal transmission, we inoculated each of 18 bison cows with 4.5 × 10(8) colony-forming units of B. abortus strain 19, as a surrogate of field strain, by three routes: intraconjunctival (IC), intravaginal (VI), and intracervical/intrauterine (AI). Bison semen was mixed with strain 19 inoculum for the latter route. Bison were monitored by serology and culture for 12 wk, at which time they were euthanized and specimens collected for culture. All IC-inoculated animals seroconverted on multiple tests and one was culture positive at 12 wk postexposure. Seven of eight VI bison developed suspect or positive serologic tests and four were positive at one or more time points. Weak transient serologic responses (suspect) were seen in four of five AI bison. Results showed that IC inoculation with strain 19 was a suitable surrogate for field strain to demonstrate exposure to the B. abortus. The seroconversion of four of eight VI bison indicated exposure of the immune system to the agent and the need for further studies on venereal transmission in bison.

  12. Overview of the activity of a Brucella abortus preparation, Bru-Pel.

    PubMed

    Youngner, J S; Feingold, D S; Keleti, G

    1978-11-01

    The properties of a nonviable, aqueous ether-extracted Brucela abortus preparation, Bru-Pel, are described. In addition to inducing a "virus-type" interferon response and protecting mice against challenge with otherwise lethal doses of Semliki Forest virus, Bru-Pel is demonstrated to have potent antitumor properties in mice. These antitumor effects appear to be mediated by an increase in nonspecific resistance similar to that seen with other experimental antitumor agents.

  13. Brucella abortus Induces the Premature Death of Human Neutrophils through the Action of Its Lipopolysaccharide

    PubMed Central

    Barquero-Calvo, Elías; Mora-Cartín, Ricardo; Arce-Gorvel, Vilma; de Diego, Juana L.; Chacón-Díaz, Carlos; Chaves-Olarte, Esteban; Guzmán-Verri, Caterina; Buret, Andre G.; Gorvel, Jean-Pierre; Moreno, Edgardo

    2015-01-01

    Most bacterial infections induce the activation of polymorphonuclear neutrophils (PMNs), enhance their microbicidal function, and promote the survival of these leukocytes for protracted periods of time. Brucella abortus is a stealthy pathogen that evades innate immunity, barely activates PMNs, and resists the killing mechanisms of these phagocytes. Intriguing clinical signs observed during brucellosis are the low numbers of Brucella infected PMNs in the target organs and neutropenia in a proportion of the patients; features that deserve further attention. Here we demonstrate that B. abortus prematurely kills human PMNs in a dose-dependent and cell-specific manner. Death of PMNs is concomitant with the intracellular Brucella lipopolysaccharide (Br-LPS) release within vacuoles. This molecule and its lipid A reproduce the premature cell death of PMNs, a phenomenon associated to the low production of proinflammatory cytokines. Blocking of CD14 but not TLR4 prevents the Br-LPS-induced cell death. The PMNs cell death departs from necrosis, NETosis and classical apoptosis. The mechanism of PMN cell death is linked to the activation of NADPH-oxidase and a modest but steadily increase of ROS mediators. These effectors generate DNA damage, recruitments of check point kinase 1, caspases 5 and to minor extent of caspase 4, RIP1 and Ca++ release. The production of IL-1β by PMNs was barely stimulated by B. abortus infection or Br-LPS treatment. Likewise, inhibition of caspase 1 did not hamper the Br-LPS induced PMN cell death, suggesting that the inflammasome pathway was not involved. Although activation of caspases 8 and 9 was observed, they did not seem to participate in the initial triggering mechanisms, since inhibition of these caspases scarcely blocked PMN cell death. These findings suggest a mechanism for neutropenia in chronic brucellosis and reveal a novel Brucella-host cross-talk through which B. abortus is able to hinder the innate function of PMN. PMID:25946018

  14. Comparison of Biological and Immunological Characterization of Lipopolysaccharides From Brucella abortus RB51 and S19

    PubMed Central

    Kianmehr, Zahra; Kaboudanian Ardestani, Sussan; Soleimanjahi, Hoorieh; Fotouhi, Fatemeh; Alamian, Saeed; Ahmadian, Shahin

    2015-01-01

    Background: Brucella abortus RB51 is a rough stable mutant strain, which has been widely used as a live vaccine for prevention of brucellosis in cattle instead of B. abortus strain S19. B. abortus lipopolysaccharide (LPS) has unique properties in comparison to other bacterial LPS. Objectives: In the current study, two types of LPS, smooth (S-LPS) and rough (R-LPS) were purified from B. abortus S19 and RB51, respectively. The aim of this study was to evaluate biological and immunological properties of purified LPS as an immunogenical determinant. Materials and Methods: Primarily, S19 and RB51 LPS were extracted and purified by two different modifications of the phenol water method. The final purity of LPS was determined by chemical analysis (2-keto-3-deoxyoctonate (KDO), glycan, phosphate and protein content) and different staining methods, following sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). C57BL/6 mice were immunized subcutaneously three times at biweekly intervals with the same amount of purified LPSs. The humoral immunity was evaluated by measuring specific IgG levels and also different cytokine levels, such as IFN-γ, TNF-α, IL-4 and IL-10, were determined for assessing T-cell immune response. Results: Biochemical analysis data and SDS-PAGE profile showed that the chemical nature of S19 LPS is different from RB51 LPS. Both S and R-LPS induce an immune response. T-cell immune response induced by both S and R-LPS had almost the same pattern whereas S19 LPS elicited humoral immunity, which was higher than RB51 LPS. Conclusions: Purified LPS can be considered as a safe adjuvant and can be used as a component in prophylactic and therapeutic vaccines targeting infectious disease, cancer and allergies. PMID:26862376

  15. Biological properties of RB51; a stable rough strain of Brucella abortus.

    PubMed

    Schurig, G G; Roop, R M; Bagchi, T; Boyle, S; Buhrman, D; Sriranganathan, N

    1991-07-01

    A rifampin-resistant mutant of Brucella abortus, designated RB51, was derived by repeated passage of strain 2308 on Trypticase soy supplemented with 1.5% agar and varying concentrations rifampin or penicillin. The RB51 colonies absorbed crystal violet and RB51 cell suspensions autoagglutinated, indicating a rough type colonial morphology for this strain. No O-chain component was detected in lipopolysaccharide (LPS) extracted from RB51 on SDS-PAGE gels stained with silver. Western blot analysis with the monoclonal antibody BRU 38, which is specific for the perosamine homopolymer O-chain of smooth Brucella LPS, indicated that the LPS of RB51 is highly deficient in O-chain when compared with the parenteral smooth strain 2308 or rough strain 45/20. Biochemically, RB51 resembles parental strain 2308 in its ability to utilize erythritol. Intraperitoneal inoculation of RB51 into mice results in a splenic colonization which is cleared within four weeks post infection. RB51 does not revert to smooth colony morphology upon passage in vivo (mice) or in vitro. Mice infected with RB51 produce antibodies against B. abortus antigens including class 2 and 3 outer membrane proteins but not against the O-chain. Furthermore, rabbits, goats and cattle hyperimmunized with sonicates of RB51 develop antibodies to B. abortus cellular antigens but do not develop antibodies specific for the O-chain. Immunization of mice with 1 x 10(8) viable RB51 organisms confers significant protection against challenge with virulent B. abortus strain 2308.

  16. Link between geographical origin and occurrence of Brucella abortus biovars in cow and water buffalo herds.

    PubMed

    Borriello, Giorgia; Peletto, Simone; Lucibelli, Maria G; Acutis, Pier L; Ercolini, Danilo; Galiero, Giorgio

    2013-02-01

    Sixty-three Brucella isolates from water buffaloes and cattle slaughtered within the Italian national plan for brucellosis control were characterized by multiple-locus variable-number tandem repeat analysis (MLVA). Genotyping indicated a strong influence of geographic origin on the Brucella abortus biovar distribution in areas where brucellosis is endemic and highlighted the importance of rigorous management procedures aimed at avoiding inter- and intraherd spreading of pathogens.

  17. Chlamydia pecorum Infection in Free-ranging Koalas ( Phascolarctos cinereus ) on French Island, Victoria, Australia.

    PubMed

    Legione, Alistair R; Amery-Gale, Jemima; Lynch, Michael; Haynes, Leesa; Gilkerson, James R; Sansom, Fiona M; Devlin, Joanne M

    2016-04-28

    We detected Chlamydia pecorum in two koalas ( Phascolarctos cinereus ) from a closed island population in Victoria, Australia, previously free of Chlamydia infection. The ompA and multilocus sequence type were most closely related to published isolates of livestock rather than koala origin, suggesting potential cross-species transmission of C. pecorum . PMID:26981690

  18. Mechanism of T cell mediated protection in newborn mice against a Chlamydia infection

    PubMed Central

    Pal, Sukumar; de la Maza, Luis M.

    2013-01-01

    To determine the immune components needed for protection of newborn mice against Chlamydia muridarum, animals born to Chlamydia-immunized and to sham-immunized dams were infected intranasally with C. muridarum at 2 post-natal days. T-cells isolated from immunized or sham-immunized adult mice were adoptively transferred to newborn mice at the time of infection. Also, to establish what cytokines are involved in protection, IFN-γ TNF-α, IL-10, and IL-12 were passively transferred to newborn mice. To assess the Chlamydia burden in the lungs mice were euthanized at 12 post-natal days. When T-cells from immunized adult mice were transferred, mice born to and fed by immunized dams were significantly protected as evidenced by the reduced number of Chlamydia isolated from the lungs compared to mice born to and fed by sham-immunized dams. Transfer of IFN-γ and TNF-α also significantly reduced the number of Chlamydia in the lungs of mice born to immunized dams. Transfer of IL-10 or IL-12 did not result in a significant reduction of Chlamydia. In vitro T-cell proliferation data suggest that neonatal antigen presenting cells can present Chlamydia antigens to adult T-cells. In conclusion, maternal antibodies and Chlamydia specific T-cells or Th1 cytokines are required for protection of neonates against this pathogen. PMID:23644176

  19. Mechanism of T-cell mediated protection in newborn mice against a Chlamydia infection.

    PubMed

    Pal, Sukumar; de la Maza, Luis M

    2013-01-01

    To determine the immune components needed for protection of newborn mice against Chlamydia muridarum, animals born to Chlamydia-immunized and to sham-immunized dams were infected intranasally with C. muridarum at 2 post-natal days. T-cells isolated from immunized or sham-immunized adult mice were adoptively transferred to newborn mice at the time of infection. Also, to establish what cytokines are involved in protection, IFN-γ, TNF-α, IL-10, and IL-12 were passively transferred to newborn mice. To assess the Chlamydia burden in the lungs mice were euthanized at 12 post-natal days. When T-cells from immunized adult mice were transferred, mice born to and fed by immunized dams were significantly protected as evidenced by the reduced number of Chlamydia isolated from the lungs compared to mice born to and fed by sham-immunized dams. Transfer of IFN-γ and TNF-α also significantly reduced the number of Chlamydia in the lungs of mice born to immunized dams. Transfer of IL-10 or IL-12 did not result in a significant reduction of Chlamydia. In vitro T-cell proliferation data suggest that neonatal antigen presenting cells can present Chlamydia antigens to adult T-cells. In conclusion, maternal antibodies and Chlamydia specific T-cells or Th1 cytokines are required for protection of neonates against this pathogen.

  20. Changes in chlamydia control activities in Europe between 2007 and 2012: a cross-national survey

    PubMed Central

    Sfetcu, Otilia; van der Sande, Marianne A.; Andersen, Berit; Herrmann, Björn; Ward, Helen; Götz, Hannelore M.; Uusküla, Anneli; Woodhall, Sarah C.; Redmond, Shelagh M.; Amato-Gauci, Andrew J.; Low, Nicola; van Bergen, Jan E.

    2016-01-01

    Background: In 2012, the levels of chlamydia control activities including primary prevention, effective case management with partner management and surveillance were assessed in 2012 across countries in the European Union and European Economic Area (EU/EEA), on initiative of the European Centre for Disease Control (ECDC) survey, and the findings were compared with those from a similar survey in 2007. Methods: Experts in the 30 EU/EEA countries were invited to respond to an online questionnaire; 28 countries responded, of which 25 participated in both the 2007 and 2012 surveys. Analyses focused on 13 indicators of chlamydia prevention and control activities; countries were assigned to one of five categories of chlamydia control. Results: In 2012, more countries than in 2007 reported availability of national chlamydia case management guidelines (80% vs. 68%), opportunistic chlamydia testing (68% vs. 44%) and consistent use of nucleic acid amplification tests (64% vs. 36%). The number of countries reporting having a national sexually transmitted infection control strategy or a surveillance system for chlamydia did not change notably. In 2012, most countries (18/25, 72%) had implemented primary prevention activities and case management guidelines addressing partner management, compared with 44% (11/25) of countries in 2007. Conclusion: Overall, chlamydia control activities in EU/EEA countries strengthened between 2007 and 2012. Several countries still need to develop essential chlamydia control activities, whereas others may strengthen implementation and monitoring of existing activities. PMID:26498953

  1. Two more species of Chlamydia-does it make a difference?

    PubMed

    Sachse, Konrad; Laroucau, Karine

    2015-02-01

    The recent description of Chlamydia (C.) avium and C. gallinacea as new species of the reunited genus Chlamydia can be expected to have implications on the perception of avian chlamydiosis. We discuss possible effects on epidemiology, diagnosis and our understanding of aetiopathogenesis resulting from this discovery.

  2. High seroprevalence of Chlamydia infection in sows in Hunan province, subtropical China.

    PubMed

    Zhang, Xiao-Xuan; Li, Run-Cheng; Liu, Guo-Hua; Cong, Wei; Song, Hui-Qun; Yu, Xing-Long; Zhu, Xing-Quan

    2014-04-01

    Chlamydia spp. are Gram-negative obligate intracellular bacteria, which are responsible for significant public health problems in humans and have major economic impact on animals. In the present study, the seroprevalence of Chlamydia infection in sows in Hunan province, subtropical China, was examined using indirect hemagglutination assay (IHA). Antibodies to Chlamydia were detected in 747 of 1,191 (62.7%, 95% CI 60-65.5) serum samples (IHA titer ≥ 1:16). The Chlamydia seroprevalence ranged from 35% (95% CI 25.7-44.4) to 77.1% (95% CI 69.1-85.2) among different regions in Hunan province, and the differences were statistically significant (P < 0.01). In addition, the seroprevalence of Chlamydia infection in sows was higher in summer (75.7%, 95% CI 71.3-80) and spring (63.2%, 95% CI 57.5-68.8) than in autumn (56.9%, 95% CI 51.5-62.3) and winter (48.6%, 95% CI 42-55.3), and the differences were statistically significant (P < 0.01). The results of the present investigation indicated the high seroprevalence of Chlamydia infection in sows in Hunan province, subtropical China, which poses a potential risk for human infection with Chlamydia in this province. This is the first report of Chlamydia seroprevalence in sows over the last two decades in Hunan province, subtropical China.

  3. Male sex predominance in Chlamydia trachomatis sexually acquired reactive arthritis: are women more protected by anti-chlamydia antibodies?

    PubMed Central

    Bas, S; Scieux, C; Vischer, T

    2001-01-01

    OBJECTIVE—To determine whether the humoral anti-chlamydia antibody response might be related to the ineffective bacterial elimination seen in patients with Chlamydia trachomatis reactive arthritis, particularly in men, who have a higher prevalence of the disease than women.
METHODS—The number and specificity of the antibody responses to 27 different C trachomatis antigens were determined by western blots in serum samples from patients with C trachomatis urogenital infection, with and without reactive arthritis, with a special regard to the sex of the patients.
RESULTS—Patients with reactive arthritis had antibodies to significantly fewer chlamydia antigens than those with urethritis only. Antibodies from men recognised significantly fewer antigens than antibodies from women. The IgA class antibodies were slightly more relevant than those of the IgG class for differentiation of patients with reactive arthritis from those with uncomplicated genitourinary infection.
CONCLUSIONS—In patients with acute C trachomatis infection the development of reactive arthritis may be related, particularly in men, to a deficient humoral response, to antigens which perhaps play a part in the clearance of the bacteria. Men who cannot generate antibodies to a large number of antigens may be less able to contain the local infection, allowing a wide systemic dissemination of the organisms to the joints.

 PMID:11350850

  4. The prevalence and clinical significance of Chlamydia infection in island and mainland populations of Victorian koalas (Phascolarctos cinereus).

    PubMed

    Patterson, Jade L S; Lynch, Michael; Anderson, Garry A; Noormohammadi, Amir H; Legione, Alistair; Gilkerson, James R; Devlin, Joanne M

    2015-04-01

    Chlamydia infection is known to impact the health of koalas (Phascolarctos cinereus) in New South Wales (NSW) and Queensland, but the clinical significance of Chlamydia infections in Victorian koalas is not well described. We examined the prevalence of Chlamydia infection and assessed associated health parameters in two Victorian koala populations known to be Chlamydia positive. The same testing regimen was applied to a third Victorian population in which Chlamydia had not been detected. We examined 288 koalas and collected samples from the urogenital sinus and conjunctival sacs. Detection and differentiation of Chlamydia species utilized real-time PCR and high-resolution melting curve analysis. Chlamydia pecorum was detected in two populations (prevalences: 25% and 41%, respectively) but only from urogenital sinus swabs. Chlamydia was not detected in the third population. Chlamydia pneumoniae was not detected. Chlamydia pecorum infection was positively associated with wet bottom (indicating chronic urinary tract disease) in one Chlamydia-positive population and with abnormal urogenital ultrasound findings in the other Chlamydia-positive population. The prevalence of wet bottom was similar in all populations (including the Chlamydia-free population), suggesting there is another significant cause (or causes) of wet bottom in Victorian koalas. Ocular disease was not observed. This is the largest study of Chlamydia infection in Victorian koalas, and the results suggest the potential for epidemiologic differences related to Chlamydia infections between Victorian koalas and koalas in Queensland and NSW and also between geographically distinct Victorian populations. Further studies to investigate the genotypes of C. pecorum present in Victorian koalas and to identify additional causes of wet bottom in koalas are indicated. PMID:25588005

  5. The prevalence and clinical significance of Chlamydia infection in island and mainland populations of Victorian koalas (Phascolarctos cinereus).

    PubMed

    Patterson, Jade L S; Lynch, Michael; Anderson, Garry A; Noormohammadi, Amir H; Legione, Alistair; Gilkerson, James R; Devlin, Joanne M

    2015-04-01

    Chlamydia infection is known to impact the health of koalas (Phascolarctos cinereus) in New South Wales (NSW) and Queensland, but the clinical significance of Chlamydia infections in Victorian koalas is not well described. We examined the prevalence of Chlamydia infection and assessed associated health parameters in two Victorian koala populations known to be Chlamydia positive. The same testing regimen was applied to a third Victorian population in which Chlamydia had not been detected. We examined 288 koalas and collected samples from the urogenital sinus and conjunctival sacs. Detection and differentiation of Chlamydia species utilized real-time PCR and high-resolution melting curve analysis. Chlamydia pecorum was detected in two populations (prevalences: 25% and 41%, respectively) but only from urogenital sinus swabs. Chlamydia was not detected in the third population. Chlamydia pneumoniae was not detected. Chlamydia pecorum infection was positively associated with wet bottom (indicating chronic urinary tract disease) in one Chlamydia-positive population and with abnormal urogenital ultrasound findings in the other Chlamydia-positive population. The prevalence of wet bottom was similar in all populations (including the Chlamydia-free population), suggesting there is another significant cause (or causes) of wet bottom in Victorian koalas. Ocular disease was not observed. This is the largest study of Chlamydia infection in Victorian koalas, and the results suggest the potential for epidemiologic differences related to Chlamydia infections between Victorian koalas and koalas in Queensland and NSW and also between geographically distinct Victorian populations. Further studies to investigate the genotypes of C. pecorum present in Victorian koalas and to identify additional causes of wet bottom in koalas are indicated.

  6. Intracellularly induced cyclophilins play an important role in stress adaptation and virulence of Brucella abortus.

    PubMed

    Roset, Mara S; García Fernández, Lucía; DelVecchio, Vito G; Briones, Gabriel

    2013-02-01

    Brucella is an intracellular bacterial pathogen that causes the worldwide zoonotic disease brucellosis. Brucella virulence relies on its ability to transition to an intracellular lifestyle within host cells. Thus, this pathogen must sense its intracellular localization and then reprogram gene expression for survival within the host cell. A comparative proteomic investigation was performed to identify differentially expressed proteins potentially relevant for Brucella intracellular adaptation. Two proteins identified as cyclophilins (CypA and CypB) were overexpressed in the intracellular environment of the host cell in comparison to laboratory-grown Brucella. To define the potential role of cyclophilins in Brucella virulence, a double-deletion mutant was constructed and its resulting phenotype was characterized. The Brucella abortus ΔcypAB mutant displayed increased sensitivity to environmental stressors, such as oxidative stress, pH, and detergents. In addition, the B. abortus ΔcypAB mutant strain had a reduced growth rate at lower temperature, a phenotype associated with defective expression of cyclophilins in other microorganisms. The B. abortus ΔcypAB mutant also displays reduced virulence in BALB/c mice and defective intracellular survival in HeLa cells. These findings suggest that cyclophilins are important for Brucella virulence and survival in the host cells.

  7. Intracellularly Induced Cyclophilins Play an Important Role in Stress Adaptation and Virulence of Brucella abortus

    PubMed Central

    García Fernández, Lucía; DelVecchio, Vito G.; Briones, Gabriel

    2013-01-01

    Brucella is an intracellular bacterial pathogen that causes the worldwide zoonotic disease brucellosis. Brucella virulence relies on its ability to transition to an intracellular lifestyle within host cells. Thus, this pathogen must sense its intracellular localization and then reprogram gene expression for survival within the host cell. A comparative proteomic investigation was performed to identify differentially expressed proteins potentially relevant for Brucella intracellular adaptation. Two proteins identified as cyclophilins (CypA and CypB) were overexpressed in the intracellular environment of the host cell in comparison to laboratory-grown Brucella. To define the potential role of cyclophilins in Brucella virulence, a double-deletion mutant was constructed and its resulting phenotype was characterized. The Brucella abortus ΔcypAB mutant displayed increased sensitivity to environmental stressors, such as oxidative stress, pH, and detergents. In addition, the B. abortus ΔcypAB mutant strain had a reduced growth rate at lower temperature, a phenotype associated with defective expression of cyclophilins in other microorganisms. The B. abortus ΔcypAB mutant also displays reduced virulence in BALB/c mice and defective intracellular survival in HeLa cells. These findings suggest that cyclophilins are important for Brucella virulence and survival in the host cells. PMID:23230297

  8. Detection of antibodies against Chlamydophila abortus in Costa Rican sheep flocks

    PubMed Central

    Villagra-Blanco, R.; Dolz, G.; Montero-Caballero, D.; Romero-Zúñiga, J.J.

    2015-01-01

    A total of 359 sheep samples from 15 flocks were analyzed for the presence of antibodies against Chlamydophila abortus using a commercial Enzyme linked Immunosorbent Assay (ELISA). Antibodies were detected in 19 (5.29%) sheep from 12 (80%) flocks. Seropositive animals were found in all analyzed regions (Central, Chorotega, Atlantic Huetar, North Huetar and Central Pacific) determining prevalence between 0.28% and 4.4%, and intra-flock positivity between 3.7% and 25.0%. The survey revealed two risk factors associated with seropositivity; introducing animals (males and females), embryos, or semen from other farms or from abroad without any sanitary certification, and flocks not having quarantine areas or separated boxes for diseased animals. No clinical signs of disease were observed in positive seroreactors. C. abortus seems to be present in Costa Rica in a very low prevalence in sheep flocks. Further studies, to isolate the bacteria are required. Finally, implementation of control measures to prevent the spread of C. abortus is recommended. PMID:26623377

  9. Survival of smooth, rough and transposon mutant strains of Brucella abortus in bovine mammary macrophages.

    PubMed

    Price, R E; Templeton, J W; Adams, L G

    1990-12-01

    Transposon mutants offer a unique way to evaluate the role of lipopolysaccharide (LPS) by producing a theoretical single-gene difference between the original strain and the transposon mutant strain. Comparative survival of Brucella abortus smooth strain 2308, rough RB51, smooth strain 19, and two transposon mutant strains (rough strain 2308::Tn5 Lac Z [m106] and rough strain 19::Tn5 Lac Z [m3], was tested in restrictive bovine mammary macrophages that were able to effectively reduce the percentage of intracellular bacterial survival and permissive bovine mammary macrophages that were unable to control the intracellular replication of B. abortus. The theoretical single-gene difference between strain 19 and strain 19::Tn5 lac Z [m3] and between smooth virulent strain 2308 and rough transposon mutant 2308::Tn5 lacZ [m106] is likely related to differences in LPS content or structure. Significant (P less than 0.05) reduction in the survival of rough strain 19::Tn5 Lac Z [m3] with no significant reduction in the rough transposon mutant strain 2308::Tn5 lacZ [m106] indicated that at least one factor other than LPS contributes to the intracellular survival of B. abortus in bovine macrophages.

  10. The role of innate immune signals in immunity to Brucella abortus

    PubMed Central

    Gomes, Marco Túlio R.; Campos, Priscila C.; de Almeida, Leonardo A.; Oliveira, Fernanda S.; Costa, Miriam Maria S.; Marim, Fernanda M.; Pereira, Guilherme S. M.; Oliveira, Sergio C.

    2012-01-01

    Innate immunity serves as the first line of defense against infectious agents such as intracellular bacteria. The innate immune platform includes Toll-like receptors (TLRs), retinoid acid-inducible gene-I-like receptors and other cytosolic nucleic acid sensors, nucleotide-binding and oligomerization domain-like receptors, adaptors, kinases and other signaling molecules that are required to elicit effective responses against different pathogens. Our research group has been using the Gram-negative bacteria Brucella abortus as a model of pathogen. We have demonstrated that B. abortus triggers MAPK and NF-κB signaling pathways in macrophages in a MyD88 and IRAK-4-dependent manner. Furthermore, we claimed that so far TLR9 is the most important single TLR during Brucella infection. The identification of host receptors that recognize pathogen-derived nucleic acids has revealed an essential role for nucleic acid sensing in the triggering of immunity to intracellular pathogens. Besides TLRs, herein we describe recent advances in NOD1, NOD2, and type I IFN receptors in innate immune pathways during B. abortus infection. PMID:23112959

  11. The Nramp1AA genotype confers susceptibility to Brucella abortus in water buffalo.

    PubMed

    Capparelli, Rosanna; Borriello, Giorgia; Marabelli, Romano; Roperto, Sante; Roperto, Franco; Iannelli, Domenico

    2007-02-01

    The 3' untranslated region of the water buffalo Nramp1 (natural resistance-associated macrophage protein 1) gene contains two alleles (Nramp1A and Nramp1B), as detected by the denaturing gradient gel electrophoresis (DGGE) technique. The Nramp1BB genotype is associated with resistance of water buffalo to the intracellular pathogen Brucella abortus. This article provides evidence that the Nramp1AA genotype is associated with susceptibility to the same pathogen. Susceptibility or resistance of water buffalo to B. abortus was established by agglutination, complement fixation, and skin tests. The Nramp1 genotype was established by DGGE analysis. The association between the Nramp1AA genotype and susceptibility to B. abortus was demonstrated in two independent population samples (152 cases and 281 controls; 87 cases and 124 controls, respectively). Macrophages from Nramp1AA subjects displayed a lower Nramp1 mRNA level when compared with macrophages from Nramp1BB subjects. Also, monocytes and macrophages from Nramp1AA subjects displayed a higher number of viable intracellular bacteria in comparison with monocytes and macrophages from Nramp1BB animals, providing biological significance to the results from association studies.

  12. Mannose-binding lectin haplotypes influence Brucella abortus infection in the water buffalo (Bubalus bubalis).

    PubMed

    Capparelli, R; Parlato, M; Amoroso, M G; Roperto, S; Marabelli, R; Roperto, F; Iannelli, D

    2008-04-01

    A case-control study established that the haplotype pair HYA/HYA at the MBL (mannose binding lectin) locus of water buffalo is associated with resistance to Brucella abortus infection (P < 10(-7)) and the haplotype pairs LYD/LYD with susceptibility to the same pathogen (P < 10(-7)). The subjects included in the present study were tested twice-at a 1-month interval-for the presence of anti-B. abortus antibodies in the serum by agglutination, complement fixation and flow cytometry. Cases (335 subjects) included animals consistently positive to all these tests; controls (335 subjects) comprised animals exposed yet negative by the same tests. The serum from genetically resistant subjects displayed in vitro significantly higher antibacterial activity compared to the serum from genetically susceptible subjects, lending biological significance to the results from the association study. Inhibition of the antibacterial activity following heat treatment of the serum, addition of specific MBL inhibitors (EDTA, mannose, N-acetyl-D: -glucosamine) or anti-human MBL antiserum provide convincing evidence that the antibacterial activity present in the serum results from the interaction between MBL and B. abortus. A replication study (comprising 100 cases and 100 controls) confirmed the results from the original study.

  13. Cloning and sequencing of yajC and secD homologs of Brucella abortus and demonstration of immune responses to YajC in mice vaccinated with B. abortus RB51.

    PubMed

    Vemulapalli, R; Duncan, A J; Boyle, S M; Sriranganathan, N; Toth, T E; Schurig, G G

    1998-12-01

    To identify Brucella antigens that are potentially involved in stimulating a protective cell-mediated immune response, a gene library of Brucella abortus 2308 was screened for the expression of antigens reacting with immunoglobulin G2a antibodies from BALB/c mice vaccinated with B. abortus RB51. One selected positive clone (clone MCB68) contained an insert of 2.6 kb; nucleotide sequence analysis of this insert revealed two open reading frames (ORFs). The deduced amino acid sequences of the first and second ORFs had significant similarities with the YajC and SecD proteins, respectively, of several bacterial species. Both the YajC and SecD proteins were expressed in Escherichia coli as fusion proteins with maltose binding protein (MBP). In Western blots, sera from mice vaccinated with B. abortus RB51 recognized YajC but not SecD. Further Western blot analysis with purified recombinant YajC protein indicated that mice inoculated with B. abortus 19 or 2308 or B. melitensis RM1 also produced antibodies to YajC. In response to in vitro stimulation with recombinant MBP-YajC fusion protein, splenocytes from mice vaccinated with B. abortus RB51 were able to proliferate and produce gamma interferon but not interleukin-4. This study demonstrates, for the first time, the involvement of YajC protein in an immune response to an infectious agent.

  14. What's in a word: the use, misuse, and abuse of the word "persistence" in Chlamydia biology.

    PubMed

    Bavoil, Patrik M

    2014-01-01

    The word persistence was used by Chlamydia researchers almost as soon as Chlamydia research was born to reflect the propensity of chlamydiae to cause inapparent infection in their hosts, from birds to humans. More recently, the term persistence has been used, misused, and sometimes abused amidst in vitro and in vivo studies that aim to mimick the ability of chlamydiae to emerge from the presumed inapparent state into clinically detectable infection and disease. Here, I have attempted to provide a global perspective on the state of research on chlamydial persistence, revisiting old observations that may warrant a new look, critically evaluating more recent observations and their shortcomings, and including recent developments that may help redefine chlamydiae as pathogens-or not-of both animals and humans.

  15. Inhibitory effect of red ginseng acidic polysaccharide from Korean red ginseng on phagocytic activity and intracellular replication of Brucella abortus in RAW 264.7 cells.

    PubMed

    Reyes, Alisha Wehdnesday Bernardo; Simborio, Hannah Leah Tadeja; Hop, Huynh Tan; Arayan, Lauren Togonon; Min, Won Gi; Lee, Hu Jang; Rhee, Man Hee; Chang, Hong Hee; Kim, Suk

    2016-09-30

    Korean red ginseng (KRG) has long been used in traditional Korean and Oriental medicine. However, the anti-bacterial mechanism and therapeutic efficiency of KGR for intracellular Brucella infection are still unclear. In this study, the bactericidal activity of Korean red ginseng acidic polysaccharide (RGAP) on Brucella (B.) abortus and its cytotoxic effects on RAW 264.7 cells were evaluated. In addition, B. abortus internalization and intracellular replication in macrophages were investigated after RGAP treatment. RGAP-incubated cells displayed a marked reduction in the adherence, internalization and intracellular growth of B. abortus in macrophages. Furthermore, decreased F-actin fluorescence was observed relative to untreated B. abortus-infected cells. Western blot analysis of intracellular signaling proteins revealed reduced ERK, JNK and p38α phosphorylation levels in B. abortus-infected RGAP-treated cells compared to the control. Moreover, elevated co-localization of B. abortus-containing phagosomes with lysosome-associated membrane protein 1 (LAMP-1) were observed in RGAP-treated cells compared with the control. Overall, the results of this study suggest that RGAP can disrupt phagocytic activity of B. abortus via suppression of mitogen-activated protein kinases (MAPKs) signaling proteins ERK, JNK and p38 levels and inhibit intracellular replication of B. abortus by enhancing phagolysosome fusion, which may provide an alternative control of brucellosis.

  16. Inhibitory effect of red ginseng acidic polysaccharide from Korean red ginseng on phagocytic activity and intracellular replication of Brucella abortus in RAW 264.7 cells

    PubMed Central

    Bernardo Reyes, Alisha Wehdnesday; Simborio, Hannah Leah Tadeja; Hop, Huynh Tan; Arayan, Lauren Togonon; Min, Won Gi; Lee, Hu Jang; Rhee, Man Hee; Chang, Hong Hee

    2016-01-01

    Korean red ginseng (KRG) has long been used in traditional Korean and Oriental medicine. However, the anti-bacterial mechanism and therapeutic efficiency of KGR for intracellular Brucella infection are still unclear. In this study, the bactericidal activity of Korean red ginseng acidic polysaccharide (RGAP) on Brucella (B.) abortus and its cytotoxic effects on RAW 264.7 cells were evaluated. In addition, B. abortus internalization and intracellular replication in macrophages were investigated after RGAP treatment. RGAP-incubated cells displayed a marked reduction in the adherence, internalization and intracellular growth of B. abortus in macrophages. Furthermore, decreased F-actin fluorescence was observed relative to untreated B. abortus-infected cells. Western blot analysis of intracellular signaling proteins revealed reduced ERK, JNK and p38α phosphorylation levels in B. abortus-infected RGAP-treated cells compared to the control. Moreover, elevated co-localization of B. abortus-containing phagosomes with lysosome-associated membrane protein 1 (LAMP-1) were observed in RGAP-treated cells compared with the control. Overall, the results of this study suggest that RGAP can disrupt phagocytic activity of B. abortus via suppression of mitogen-activated protein kinases (MAPKs) signaling proteins ERK, JNK and p38 levels and inhibit intracellular replication of B. abortus by enhancing phagolysosome fusion, which may provide an alternative control of brucellosis. PMID:26726017

  17. Evidence of a conserved role for Chlamydia HtrA in the replication phase of the chlamydial developmental cycle.

    PubMed

    Patel, Pooja; De Boer, Leonore; Timms, Peter; Huston, Wilhelmina May

    2014-08-01

    Identification of the HtrA inhibitor JO146 previously enabled us to demonstrate an essential function for HtrA during the mid-replicative phase of the Chlamydia trachomatis developmental cycle. Here we extend our investigations to other members of the Chlamydia genus. C. trachomatis isolates with distinct replicative phase growth kinetics showed significant loss of viable infectious progeny after HtrA was inhibited during the replicative phase. Mid-replicative phase addition of JO146 was also significantly detrimental to Chlamydia pecorum, Chlamydia suis and Chlamydia cavie. These data combined indicate that HtrA has a conserved critical role during the replicative phase of the chlamydial developmental cycle.

  18. Caspase-2 mediates a Brucella abortus RB51-induced hybrid cell death having features of apoptosis and pyroptosis

    PubMed Central

    Bronner, Denise N.; O'Riordan, Mary X. D.; He, Yongqun

    2013-01-01

    Programmed cell death (PCD) can play a crucial role in tuning the immune response to microbial infection. Although PCD can occur in different forms, all are mediated by a family of proteases called caspases. Caspase-2 is the most conserved caspase, however, its function in cell death is ill-defined. Previously we demonstrated that live attenuated cattle vaccine strain Brucella abortus RB51 induces caspase-2-mediated and caspase-1-independent PCD of infected macrophages. We also discovered that rough attenuated B. suis strain VTRS1 induces a caspase-2-mediated and caspase-1-independent proinflammatory cell death in infected macrophages, which was tentatively coined “caspase-2-mediated pyroptosis”. However, the mechanism of caspase-2-mediated cell death pathway remained unclear. In this study, we found that caspase-2 mediated proinflammatory cell death of RB51-infected macrophages and regulated many genes in different PCD pathways. We show that the activation of proapoptotic caspases-3 and -8 was dependent upon caspase-2. Caspase-2 regulated mitochondrial cytochrome c release and TNFα production, both of which are known to activate caspase-3 and caspase-8, respectively. In addition to TNFα, RB51-induced caspase-1 and IL-1β production was also driven by caspase-2-mediated mitochondrial dysfunction. Interestingly, pore formation, a phenomenon commonly associated with caspase-1-mediated pyroptosis, occurred; however, unlike its role in S. typhimurium-induced pyroptosis, pore formation did not contribute to RB51-induced proinflammatory cell death. Our data suggest that caspase-2 acts as an initiator caspase that mediates a novel RB51-induced hybrid cell death that simulates but differs from typical non-proinflammatory apoptosis and caspase-1-mediated proinflammatory pyroptosis. The initiator role of the caspase-2-mediated cell death was also conserved in cellular stress-induced cell death of macrophages treated with etoposide, naphthalene, or anti-Fas. Caspase-2

  19. Brucella abortus RB51 and hot saline extract from Brucella ovis as antigens in a complement fixation test used To detect sheep vaccinated with Brucella abortus RB51.

    PubMed

    Adone, R; Ciuchini, F

    2001-01-01

    The efficacy of Brucella abortus RB51 and hot saline extract (HSE) from Brucella ovis as antigens in complement fixation (CF) tests was comparatively evaluated in detecting immune responses of sheep vaccinated with B. abortus strain RB51. For this study, four 5-month-old sheep were vaccinated subcutaneously with 5 x 10(9) CFU of RB51, and two sheep received saline. Serum samples collected at different times after vaccination were tested for the presence of antibodies to RB51 by a CF test with RB51 as antigen, previously deprived of anticomplementary activity, and with HSE antigen, which already used as the official antigen to detect B. ovis-infected sheep. The results showed that vaccinated sheep developed antibodies which reacted weakly against HSE antigen and these antibodies were detectable for 30 days after vaccination. However, antibodies to RB51 could be detected for a longer period after vaccination by using homologous RB51 antigen in CF tests. In fact, high titers were still present at 110 days postvaccination with RB51 antigen. Sera from sheep naturally infected with B. ovis also reacted to RB51 but gave lower titers than those detected by HSE antigen. As expected, all sera from RB51-vaccinated sheep remained negative when tested with standard S-type Brucella standard antigens.

  20. DC attenuation meter

    DOEpatents

    Hargrove, Douglas L.

    2004-09-14

    A portable, hand-held meter used to measure direct current (DC) attenuation in low impedance electrical signal cables and signal attenuators. A DC voltage is applied to the signal input of the cable and feedback to the control circuit through the signal cable and attenuators. The control circuit adjusts the applied voltage to the cable until the feedback voltage equals the reference voltage. The "units" of applied voltage required at the cable input is the system attenuation value of the cable and attenuators, which makes this meter unique. The meter may be used to calibrate data signal cables, attenuators, and cable-attenuator assemblies.

  1. Chlamydia psittaci: update on an underestimated zoonotic agent.

    PubMed

    Knittler, Michael R; Sachse, Konrad

    2015-02-01

    Chlamydia (C.) psittaci is an economically relevant pathogen in poultry and pet birds, where it causes psittacosis/ornithosis, and also a human pathogen causing atypical pneumonia after zoonotic transmission. Despite its well-documented prevalence, the agent has received less attention by researchers than other Chlamydia spp. in the last decades. In the present paper, we review recently published data on C. psittaci infection and attempt to single out characteristic features distinguishing it from related chlamydial agents. It is remarkable that C. psittaci is particularly efficient in disseminating in the host organism causing systemic disease, which occasionally can take a fulminant course. At the cellular level, the pathogen's broad host cell spectrum (from epithelial cells to macrophages), its rapid entry and fast replication, proficient use of intracellular transport routes to mitochondria and the Golgi apparatus, the pronounced physical association of chlamydial inclusions with energy-providing cell compartments, as well as the subversive regulation of host cell survival during productive and persistent states facilitate the characteristic efficient growth and successful host-to-host spread of C. psittaci. At the molecular level, the pathogen was shown to upregulate essential chlamydial genes when facing the host immune response. We hypothesize that this capacity, in concert with expression of specific effectors of the type III secretion system and efficient suppression of selected host defense signals, contributes to successful establishment of the infection in the host. Concerning the immunology of host-pathogen interactions, C. psittaci has been shown to distinguish itself by coping more efficiently than other chlamydiae with pro-inflammatory mediators during early host response, which can, to some extent, explain the effective evasion and adaptation strategies of this bacterium. We conclude that thorough analysis of the large number of whole

  2. Prevalence of Chlamydia trachomatis infection in pregnant patients.

    PubMed Central

    Much, D H; Yeh, S Y

    1991-01-01

    Chlamydia is a sexually transmitted disease of epidemic proportions, infecting an estimated 4 million people a year. It results not only in infertility and ectopic pregnancy but also in infant morbidity and mortality. Ectopic pregnancy is responsible for 11 percent of maternal deaths. About 60 percent of infected women can transmit the bacteria at birth to their infants. Early detection and treatment of chlamydia in both men and women, especially prenatal women, is critical. Chlamydia trachomatis infection of the cervix was found in 8.1 percent of a group of 1,004 pregnant women at a hospital prenatal clinic by means of a direct fluorescent antibody test. The prevalence of C. trachomatis was only 0.7 percent in 277 pregnant women receiving prenatal care from private practitioners. All patients between 27 and 30 weeks gestation who tested positive were treated with oral erythromycin. Their partners were treated with tetracycline. The outcome of pregnancy in patients treated for chlamydial infection was compared with a control group of noninfected mothers from the same population. The frequency of premature rupture of the membranes, prematurity, and low Apgar scores among the treated women were not significantly different from those in the control group. There was a significant difference, however, between the two groups in the incidence of low mean birth weight infants and the presence of meconium. Children can acquire a chlamydial infection at birth from contact with infected cervico-vaginal secretions. If not detected and treated, these infected infants may develop conjunctivitis, bronchiolitis, and pneumonia. It is suggested, therefore, that all patients at prenatal clinics be screened for chlamydial cervicitis.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1910182

  3. Evaluation of bison (Bison bison) semen from Yellowstone National Park, Montana, USA, bulls for Brucella abortus shedding.

    PubMed

    Frey, Rebecca K; Clarke, P Ryan; McCollum, Matt P; Nol, Pauline; Johnson, Kammy R; Thompson, Brent D; Ramsey, Jennifer M; Anderson, Neil J; Rhyan, Jack C

    2013-07-01

    To determine if bison (Bison bison) bulls from Yellowstone National Park (YNP), Montana, USA, shed an infective dose of Brucella abortus in semen, 50 YNP bulls were captured on public lands in Montana during the winter and early spring (April-May) of 2010 and 2011. The bulls were immobilized, and blood and semen samples were collected for serology and Brucella culture. Thirty-five bulls (70%) were antibody-positive, and B. abortus was cultured from semen in three (9%) of the 35 antibody-positive or suspect bulls, though not at concentrations considered an infective dose. Eight bulls (six antibody-positive, two negative) had palpable lesions of the testes, epididymides, or seminal vesicles consistent with B. abortus infection. Breeding soundness exams and semen analysis suggested that antibody-positive bulls were more likely to have nonviable ejaculate (8/35; 23%) than bulls without detectable antibody (2/15; 13%).

  4. [Cough syncope caused by a possible Chlamydia pneumoniae pneumonia].

    PubMed

    Cinotti, R; Moubarak, G; Gervais, R; Mabo, P

    2009-09-01

    We report the case of a 61-year-old man who presented with coughing fits followed by sinus pauses and syncope. Cardiac and neurological diagnostic work-up was negative and the patient was considered to have cough syncope. As this occurred within the context of febrile pneumonia, an infectious disease was suspected but diagnostic work-up only revealed an increase of antibodies against Chlamydia pneumoniae. The responsibility of this agent is discussed. Clinical recovery was obtained with the prescription of antitussive medication.

  5. Detection of novel Chlamydiae and Legionellales from human nasal samples of healthy volunteers.

    PubMed

    Corsaro, Daniele; Venditti, Danielle

    2015-07-01

    Chlamydiae are intracellular bacterial parasites of eukaryotes, ranging from amoebae to humans. They comprise many novel members and are investigated as emerging pathogens. Environmental studies highlighted similarities between the ecologies of chlamydiae and legionellae, both groups being important agents of respiratory infections. Herein, we analyzed nasal samples from healthy persons, searching for the presence of amoebae, chlamydiae and legionellae. From a total of 25 samples, we recovered by PCR eight samples positive to chlamydiae and six samples positive to legionellae. Among these samples, four were positive to both organisms. The sequencing of 16S rDNAs allowed to identify (i) among Chlamydiae: Parachlamydia acanthamoebae, Chlamydophila psittaci, Chlamydophila felis, and members of Rhabdochlamydiaceae, Simkaniaceae and E6 lineage and (ii) among Legionellaceae: Legionella longbeachae, Legionella bozemanii and Legionella impletisoli. Unexpectedly, we also recovered Diplorickettsia sp. Amoebae collected from nasal mucosae, Acanthamoeba and Vermamoeba, were endosymbiont-free, and chlamydiae revealed refractory to amoeba coculture. This study shows common exposure to chlamydiae and legionellae and suggests open air activities like gardening as a probable additional source of infection.

  6. Prevalence of cervical Chlamydia trachomatis infection in sexually active adolescents from Salvador, Brazil.

    PubMed

    Machado, Márcia Sacramento Cunha; Costa e Silva, Bruno Fernando Borges da; Gomes, Igor Logetto Caetité; Santana, Iuri Usêda; Grassi, Maria Fernanda Rios

    2012-01-01

    The incidence of sexually transmitted diseases among adolescents is increasing worldwide. Genital Chlamydia trachomatis infection is one of the most prevalent sexually transmitted diseases in young women, and undetected disease is highly associated with long-term complications in women. Our goal was to determine the prevalence of cervical Chlamydia trachomatis infection in a sexually active population of female adolescents from Salvador, Brazil, and to describe their socio-demographic, behavioral, and clinical characteristics. 100 sexually active adolescents (10-19 years) were included in this study, between 2008 and 2010. Endocervical samples were obtained during gynecological examination. Inhouse polymerase chain reaction of cervical specimens was used for Chlamydia trachomatis detection. The overall prevalence of cervical Chlamydia trachomatis infection was 31% (95% CI 22-40). There were no statistically significant differences in the age at first sexual intercourse, number of sexual partners, and frequency of condom use between Chlamydia infected and uninfected adolescents. The prevalence of cervical Chlamydia trachomatis infection among adolescents from Salvador was the highest in Brazil up to the present date. These results demonstrate an urgent need for continued and comprehensive prevention strategies along with proper screening for Chlamydia in high-risk populations in order to decrease the rates of infection.

  7. Evaluation of five immunoassays for detection of Chlamydia psittaci in cloacal and conjunctival specimens from turkeys.

    PubMed Central

    Vanrompay, D; Van Nerom, A; Ducatelle, R; Haesebrouck, F

    1994-01-01

    Five commercially available immunoassays were evaluated for the detection of Chlamydia psittaci in cloacal and conjunctival swabs from industrially raised turkeys: IMAGEN (DAKO Diagnostics, Ely, Cambridgeshire, United Kingdom), Chlamydia CEL-VET IF (Cellabs, Brookvale, Australia), IDEIA (DAKO Diagnostics), CELISA (Cellabs), and CLEARVIEW (Unipath, Bedford, United Kingdom). Results were compared with isolation in Buffalo Green Monkey cells as a reference method. For the conjunctival samples, the sensitivities of the IMAGEN test, the Chlamydia CEL-VET IF test, the IDEIA, the CELISA, and the CLEARVIEW test were found to be 100, 66, 0, 0, and 0%, respectively, as compared to the reference test. Also for the conjunctival samples, the specificities of the IMAGEN test, the Chlamydia CEL-VET IF test, and the IDEIA were found to be 100, 11, and 92.8%, respectively. For the cloacal specimens, the sensitivities of the IMAGEN test, the Chlamydia CEL-VET IF test, the IDEIA, the CELISA, and the CLEARVIEW test were found to be 100, 93.3, 26.6, 0, and 53.3%, respectively. Also for the cloacal specimens, the specificities of the IMAGEN test, the Chlamydia CEL-VET IF test, the IDEIA, and the CLEARVIEW test were found to be 92, 12, 100, and 88%, respectively. The IMAGEN test was the most sensitive and specific direct chlamydia antigen detection test for cloacal and conjunctival samples from turkeys. PMID:8077391

  8. Detection of novel Chlamydiae and Legionellales from human nasal samples of healthy volunteers.

    PubMed

    Corsaro, Daniele; Venditti, Danielle

    2015-07-01

    Chlamydiae are intracellular bacterial parasites of eukaryotes, ranging from amoebae to humans. They comprise many novel members and are investigated as emerging pathogens. Environmental studies highlighted similarities between the ecologies of chlamydiae and legionellae, both groups being important agents of respiratory infections. Herein, we analyzed nasal samples from healthy persons, searching for the presence of amoebae, chlamydiae and legionellae. From a total of 25 samples, we recovered by PCR eight samples positive to chlamydiae and six samples positive to legionellae. Among these samples, four were positive to both organisms. The sequencing of 16S rDNAs allowed to identify (i) among Chlamydiae: Parachlamydia acanthamoebae, Chlamydophila psittaci, Chlamydophila felis, and members of Rhabdochlamydiaceae, Simkaniaceae and E6 lineage and (ii) among Legionellaceae: Legionella longbeachae, Legionella bozemanii and Legionella impletisoli. Unexpectedly, we also recovered Diplorickettsia sp. Amoebae collected from nasal mucosae, Acanthamoeba and Vermamoeba, were endosymbiont-free, and chlamydiae revealed refractory to amoeba coculture. This study shows common exposure to chlamydiae and legionellae and suggests open air activities like gardening as a probable additional source of infection. PMID:25697709

  9. Assessment of Chlamydia trachomatis prevalence by PCR and LCR in women presenting for termination of pregnancy

    PubMed Central

    Garland, S.; Tabrizi, S.; Hallo, J.; Chen, S.

    2000-01-01

    Objectives: To determine the prevalence of Chlamydia trachomatis in a patient population presenting for legal termination of pregnancy by polymerase chain reaction (PCR) and ligase chain reaction (LCR), from first catch urine and self administered tampons, and comparing with the traditionally collected endocervical swab tested by both PCR and culture. Methods: Consecutive women attending for legal termination of pregnancy were screened for chlamydia by patient collected first catch urine and tampon, and physician collected endocervical swab. Results: Of 1175 patients with complete samples, there were 33 (2.8%) in whom chlamydia was detected by two or more assays from one or more sample site. Chlamydia was detected equally well by both PCR and LCR in first catch urine (p = 0.25), tampon (p = 0.5), and endocervical swab (p = 0.5). However, both PCR and LCR were significantly better than culture of an endocervical swab (p = 0.0005) for detection of C trachomatis. Conclusion: The prevalence of chlamydia in patients presenting for termination of pregnancy was 2.8%. A simple efficient way of performing screening for chlamydia for women presenting for termination of pregnancy is by first catch urine or tampon, which can be tested by the highly sensitive amplification assays, PCR or LCR. Key Words: polymerase chain reaction; ligase chain reaction; Chlamydia trachomatis; pregnancy PMID:10961192

  10. Chlamydia screening and positivity in juvenile detention centers, United States, 2009-2011.

    PubMed

    Satterwhite, Catherine Lindsey; Newman, Daniel; Collins, Dayne; Torrone, Elizabeth

    2014-01-01

    An estimated 2.9 million new chlamydia infections occur in the United States each year. Among women, chlamydia can lead to serious adverse outcomes, including pelvic inflammatory disease and infertility. Chlamydia prevalence is highest among females aged 15-19 years. Despite long-standing recommendations directed at young, sexually active females, screening remains sub-optimal. Juvenile detention centers (JDCs) are uniquely situated to screen and treat high-risk adolescents. From 2009-2011, performance measure data on chlamydia screening coverage (proportion of eligible females screened) and positivity (proportion of females tested who were positive) were available from 126 geographically-dispersed JDCs in the United States. These facilities reported screening 55.2% of females entering the facilities (149,923), with a facility-specific median of 66.4% (range: 0-100%). Almost half (44.4%) of facilities had screening coverage levels of 75-100%. This screening resulted in the detection of 12,305 chlamydial infections, for an overall positivity of 14.7% (facility-specific median = 14.9%, range: 0-36.9%). In linear regression analysis, chlamydia positivity was inversely associated with screening coverage: as coverage increased, positivity decreased. The burden of chlamydia in JDCs is substantial; facilities should continue to deliver recommended chlamydia screening and treatment to females and identify mechanisms to increase coverage.

  11. Comparing two definitions of ethnicity for identifying young persons at risk for chlamydia.

    PubMed

    Haasnoot, A; Koedijk, F D H; Op De Coul, E L M; Götz, H M; van der Sande, M A B; Van Den Broek, I V F

    2012-05-01

    Ethnic disparities in chlamydia infections in The Netherlands were assessed, in order to compare two definitions of ethnicity: ethnicity based on country of birth and self-defined ethnicity. Chlamydia positivity in persons aged 16-29 years was investigated using data from the first round of the Chlamydia Screening Implementation (CSI, 2008-2009) and surveillance data from STI centres (2009). Logistic regression modelling showed that being an immigrant was associated with chlamydia positivity in both CSI [adjusted odds ratio (aOR) 2·3, 95% confidence interval (CI) 2·0-2·6] and STI centres (aOR 1·4, 95% CI 1·3-1·5). In both settings, 60% of immigrants defined themselves as Dutch. Despite the difference, classification by self-defined ethnicity resulted in similar associations between (non-Dutch) ethnicity and chlamydia positivity. However, ethnicity based on country of birth explained variation in chlamydia positivity better, and is objective and constant over time and therefore more useful for identifying young persons at higher risk for chlamydia infection.

  12. Comprehensive Identification of Immunodominant Proteins of Brucella abortus and Brucella melitensis Using Antibodies in the Sera from Naturally Infected Hosts.

    PubMed

    Wareth, Gamal; Eravci, Murat; Weise, Christoph; Roesler, Uwe; Melzer, Falk; Sprague, Lisa D; Neubauer, Heinrich; Murugaiyan, Jayaseelan

    2016-04-30

    Brucellosis is a debilitating zoonotic disease that affects humans and animals. The diagnosis of brucellosis is challenging, as accurate species level identification is not possible with any of the currently available serology-based diagnostic methods. The present study aimed at identifying Brucella (B.) species-specific proteins from the closely related species B. abortus and B. melitensis using sera collected from naturally infected host species. Unlike earlier reported investigations with either laboratory-grown species or vaccine strains, in the present study, field strains were utilized for analysis. The label-free quantitative proteomic analysis of the naturally isolated strains of these two closely related species revealed 402 differentially expressed proteins, among which 63 and 103 proteins were found exclusively in the whole cell extracts of B. abortus and B. melitensis field strains, respectively. The sera from four different naturally infected host species, i.e., cattle, buffalo, sheep, and goat were applied to identify the immune-binding protein spots present in the whole protein extracts from the isolated B. abortus and B. melitensis field strains and resolved on two-dimensional gel electrophoresis. Comprehensive analysis revealed that 25 proteins of B. abortus and 20 proteins of B. melitensis were distinctly immunoreactive. Dihydrodipicolinate synthase, glyceraldehyde-3-phosphate dehydrogenase and lactate/malate dehydrogenase from B. abortus, amino acid ABC transporter substrate-binding protein from B. melitensis and fumarylacetoacetate hydrolase from both species were reactive with the sera of all the tested naturally infected host species. The identified proteins could be used for the design of serological assays capable of detecting pan-Brucella, B. abortus- and B. melitensis-specific antibodies.

  13. Comprehensive Identification of Immunodominant Proteins of Brucella abortus and Brucella melitensis Using Antibodies in the Sera from Naturally Infected Hosts

    PubMed Central

    Wareth, Gamal; Eravci, Murat; Weise, Christoph; Roesler, Uwe; Melzer, Falk; Sprague, Lisa D.; Neubauer, Heinrich; Murugaiyan, Jayaseelan

    2016-01-01

    Brucellosis is a debilitating zoonotic disease that affects humans and animals. The diagnosis of brucellosis is challenging, as accurate species level identification is not possible with any of the currently available serology-based diagnostic methods. The present study aimed at identifying Brucella (B.) species-specific proteins from the closely related species B. abortus and B. melitensis using sera collected from naturally infected host species. Unlike earlier reported investigations with either laboratory-grown species or vaccine strains, in the present study, field strains were utilized for analysis. The label-free quantitative proteomic analysis of the naturally isolated strains of these two closely related species revealed 402 differentially expressed proteins, among which 63 and 103 proteins were found exclusively in the whole cell extracts of B. abortus and B. melitensis field strains, respectively. The sera from four different naturally infected host species, i.e., cattle, buffalo, sheep, and goat were applied to identify the immune-binding protein spots present in the whole protein extracts from the isolated B. abortus and B. melitensis field strains and resolved on two-dimensional gel electrophoresis. Comprehensive analysis revealed that 25 proteins of B. abortus and 20 proteins of B. melitensis were distinctly immunoreactive. Dihydrodipicolinate synthase, glyceraldehyde-3-phosphate dehydrogenase and lactate/malate dehydrogenase from B. abortus, amino acid ABC transporter substrate-binding protein from B. melitensis and fumarylacetoacetate hydrolase from both species were reactive with the sera of all the tested naturally infected host species. The identified proteins could be used for the design of serological assays capable of detecting pan-Brucella, B. abortus- and B. melitensis-specific antibodies. PMID:27144565

  14. A cohort study of Chlamydia trachomatis treatment failure in women: a study protocol

    PubMed Central

    2013-01-01

    Background Chlamydia trachomatis is the most commonly diagnosed bacterial sexually transmitted infection in the developed world and diagnosis rates have increased dramatically over the last decade. Repeat infections of chlamydia are very common and may represent re-infection from an untreated partner or treatment failure. The aim of this cohort study is to estimate the proportion of women infected with chlamydia who experience treatment failure after treatment with 1 gram azithromycin. Methods/design This cohort study will follow women diagnosed with chlamydia for up to 56 days post treatment. Women will provide weekly genital specimens for further assay. The primary outcome is the proportion of women who are classified as having treatment failure 28, 42 or 56 days after recruitment. Comprehensive sexual behavior data collection and the detection of Y chromosome DNA and high discriminatory chlamydial genotyping will be used to differentiate between chlamydia re-infection and treatment failure. Azithromycin levels in high-vaginal specimens will be measured using a validated liquid chromatography – tandem mass spectrometry method to assess whether poor azithromycin absorption could be a cause of treatment failure. Chlamydia culture and minimal inhibitory concentrations will be performed to further characterize the chlamydia infections. Discussion Distinguishing between treatment failure and re-infection is important in order to refine treatment recommendations and focus infection control mechanisms. If a large proportion of repeat chlamydia infections are due to antibiotic treatment failure, then international recommendations on chlamydia treatment may need to be re-evaluated. If most are re-infections, then strategies to expedite partner treatment are necessary. PMID:23957327

  15. DNA microarray-based detection of multiple pathogens: Mycoplasma spp. and Chlamydia spp.

    PubMed

    Schnee, Christiane; Sachse, Konrad

    2015-01-01

    Rapid detection of slow-growing or non-culturable microorganisms, such as Mycoplasma spp. and Chlamydia spp., is still a challenge to diagnosticians in the veterinary field. In addition, as epidemiological evidence on the frequency of mixed infections involving two and more bacterial species has been emerging, detection methods allowing simultaneous identification of different pathogens are required. In the present chapter, we describe DNA microarray-based procedures for the detection of 83 Mollicutes species (Mycoplasma assay) and 11 Chlamydia spp. (Chlamydia assay). The assays are suitable for use in a routine diagnostic environment, as well as in microbiological research.

  16. Mechanism of Asp24 Upregulation in Brucella abortus Rough Mutant with a Disrupted O-Antigen Export System and Effect of Asp24 in Bacterial Intracellular Survival

    PubMed Central

    Tian, Mingxing; Qu, Jing; Han, Xiangan; Ding, Chan; Wang, Shaohui; Peng, Daxin

    2014-01-01

    We previously showed that Brucella abortus rough mutant strain 2308 ΔATP (called the ΔrfbE mutant in this study) exhibits reduced intracellular survival in RAW264.7 cells and attenuated persistence in BALB/c mice. In this study, we performed microarray analysis to detect genes with differential expression between the ΔrfbE mutant and wild-type strain S2308. Interestingly, acid shock protein 24 gene (asp24) expression was significantly upregulated in the ΔrfbE mutant compared to S2308, as confirmed by quantitative reverse transcription-PCR (qRT-PCR) and Western blotting. Further studies using additional strains indicated that the upregulation of asp24 occurred only in rough mutants with disrupted O-antigen export system components, including the ATP-binding protein gene rfbE (bab1_0542) and the permease gene rfbD (bab1_0543), while the ΔwboA rough mutant (which lacks an O-antigen synthesis-related glycosyltransferase) and the RB51 strain (a vaccine strain with the rough phenotype) showed no significant changes in asp24 expression compared to S2308. In addition, abolishing the intracellular O-antigen synthesis of the ΔrfbE mutant by deleting the wboA gene (thereby creating the ΔrfbE ΔwboA double-knockout strain) recovered asp24 expression. These results indicated that asp24 upregulation is associated with intracellular O-antigen synthesis and accumulation but not with the bacterial rough phenotype. Further studies indicated that asp24 upregulation in the ΔrfbE mutant was associated neither with bacterial adherence and invasion nor with cellular necrosis on RAW264.7 macrophages. However, proper expression of the asp24 gene favors intracellular survival of Brucella in RAW264.7 cells and HeLa cells during an infection. This study reveals a novel mechanism for asp24 upregulation in B. abortus mutants. PMID:24752516

  17. Mechanism of Asp24 upregulation in Brucella abortus rough mutant with a disrupted O-antigen export system and effect of Asp24 in bacterial intracellular survival.

    PubMed

    Tian, Mingxing; Qu, Jing; Han, Xiangan; Ding, Chan; Wang, Shaohui; Peng, Daxin; Yu, Shengqing

    2014-07-01

    We previously showed that Brucella abortus rough mutant strain 2308 ΔATP (called the ΔrfbE mutant in this study) exhibits reduced intracellular survival in RAW264.7 cells and attenuated persistence in BALB/c mice. In this study, we performed microarray analysis to detect genes with differential expression between the ΔrfbE mutant and wild-type strain S2308. Interestingly, acid shock protein 24 gene (asp24) expression was significantly upregulated in the ΔrfbE mutant compared to S2308, as confirmed by quantitative reverse transcription-PCR (qRT-PCR) and Western blotting. Further studies using additional strains indicated that the upregulation of asp24 occurred only in rough mutants with disrupted O-antigen export system components, including the ATP-binding protein gene rfbE (bab1_0542) and the permease gene rfbD (bab1_0543), while the ΔwboA rough mutant (which lacks an O-antigen synthesis-related glycosyltransferase) and the RB51 strain (a vaccine strain with the rough phenotype) showed no significant changes in asp24 expression compared to S2308. In addition, abolishing the intracellular O-antigen synthesis of the ΔrfbE mutant by deleting the wboA gene (thereby creating the ΔrfbE ΔwboA double-knockout strain) recovered asp24 expression. These results indicated that asp24 upregulation is associated with intracellular O-antigen synthesis and accumulation but not with the bacterial rough phenotype. Further studies indicated that asp24 upregulation in the ΔrfbE mutant was associated neither with bacterial adherence and invasion nor with cellular necrosis on RAW264.7 macrophages. However, proper expression of the asp24 gene favors intracellular survival of Brucella in RAW264.7 cells and HeLa cells during an infection. This study reveals a novel mechanism for asp24 upregulation in B. abortus mutants.

  18. Typing Discrepancy Between Phenotypic and Molecular Characterization Revealing an Emerging Biovar 9 Variant of Smooth Phage-Resistant B. abortus Strain 8416 in China

    PubMed Central

    Kang, Yao-Xia; Li, Xu-Ming; Piao, Dong-Ri; Tian, Guo-Zhong; Jiang, Hai; Jia, En-Hou; Lin, Liang; Cui, Bu-Yun; Chang, Yung-Fu; Guo, Xiao-Kui; Zhu, Yong-Zhang

    2015-01-01

    A newly isolated smooth colony morphology phage-resistant strain 8416 isolated from a 45-year-old cattle farm cleaner with clinical features of brucellosis in China was reported. The most unusual phenotype was its resistance to two Brucella phages Tbilisi and Weybridge, but sensitive to Berkeley 2, a pattern similar to that of Brucella melitensis biovar 1. VITEK 2 biochemical identification system found that both strain 8416 and B. melitensis strains shared positive ILATk, but negative in other B. abortus strains. However, routine biochemical and phenotypic characteristics of strain 8416 were most similar to that of B. abortus biovar 9 except CO2 requirement. In addition, multiple PCR molecular typing assays including AMOS-PCR, B. abortus special PCR (B-ab PCR) and a novel sub-biovar typing PCR, indicated that strain 8416 may belong to either biovar 3b or 9 of B. abortus. Surprisingly, further MLVA typing results showed that strain 8416 was most closely related to B. abortus biovar 3 in the Brucella MLVA database, primarily differing in 4 out of 16 screened loci. Therefore, due to the unusual discrepancy between phenotypic (biochemical reactions and particular phage lysis profile) and molecular typing characteristics, strain 8416 could not be exactly classified to any of the existing B. abortus biovars and might be a new variant of B. abortus biovar 9. The present study also indicates that the present phage typing scheme for Brucella sp. is subject to variation and the routine Brucella biovar typing needs further studies. PMID:26696984

  19. The Effector Protein BPE005 from Brucella abortus Induces Collagen Deposition and Matrix Metalloproteinase 9 Downmodulation via Transforming Growth Factor β1 in Hepatic Stellate Cells.

    PubMed

    Arriola Benitez, Paula Constanza; Rey Serantes, Diego; Herrmann, Claudia Karina; Pesce Viglietti, Ayelén Ivana; Vanzulli, Silvia; Giambartolomei, Guillermo Hernán; Comerci, Diego José; Delpino, María Victoria

    2015-12-14

    The liver is frequently affected in patients with active brucellosis. In the present study, we identified a virulence factor involved in the modulation of hepatic stellate cell function and consequent fibrosis during Brucella abortus infection. This study assessed the role of BPE005 protein from B. abortus in the fibrotic phenotype induced on hepatic stellate cells during B. abortus infection in vitro and in vivo. We demonstrated that the fibrotic phenotype induced by B. abortus on hepatic stellate (LX-2) cells was dependent on BPE005, a protein associated with the type IV secretion system (T4SS) VirB from B. abortus. Our results indicated that B. abortus inhibits matrix metalloproteinase 9 (MMP-9) secretion through the activity of the BPE005-secreted protein and induces concomitant collagen deposition by LX-2 cells. BPE005 is a small protein containing a cyclic nucleotide monophosphate binding domain (cNMP) that modulates the LX-2 cell phenotype through a mechanism that is dependent on the cyclic AMP (cAMP)/protein kinase A (PKA) signaling pathway. Altogether, these results indicate that B. abortus tilts LX-2 cells to a profibrogenic phenotype employing a functional T4SS and the secreted BPE005 protein through a mechanism that involves the cAMP and PKA signaling pathway.

  20. The Effector Protein BPE005 from Brucella abortus Induces Collagen Deposition and Matrix Metalloproteinase 9 Downmodulation via Transforming Growth Factor β1 in Hepatic Stellate Cells

    PubMed Central

    Arriola Benitez, Paula Constanza; Rey Serantes, Diego; Herrmann, Claudia Karina; Pesce Viglietti, Ayelén Ivana; Vanzulli, Silvia; Giambartolomei, Guillermo Hernán; Comerci, Diego José

    2015-01-01

    The liver is frequently affected in patients with active brucellosis. In the present study, we identified a virulence factor involved in the modulation of hepatic stellate cell function and consequent fibrosis during Brucella abortus infection. This study assessed the role of BPE005 protein from B. abortus in the fibrotic phenotype induced on hepatic stellate cells during B. abortus infection in vitro and in vivo. We demonstrated that the fibrotic phenotype induced by B. abortus on hepatic stellate (LX-2) cells was dependent on BPE005, a protein associated with the type IV secretion system (T4SS) VirB from B. abortus. Our results indicated that B. abortus inhibits matrix metalloproteinase 9 (MMP-9) secretion through the activity of the BPE005-secreted protein and induces concomitant collagen deposition by LX-2 cells. BPE005 is a small protein containing a cyclic nucleotide monophosphate binding domain (cNMP) that modulates the LX-2 cell phenotype through a mechanism that is dependent on the cyclic AMP (cAMP)/protein kinase A (PKA) signaling pathway. Altogether, these results indicate that B. abortus tilts LX-2 cells to a profibrogenic phenotype employing a functional T4SS and the secreted BPE005 protein through a mechanism that involves the cAMP and PKA signaling pathway. PMID:26667834

  1. Recombinant bovine interleukin 2 enhances immunity and protection induced by Brucella abortus vaccines in cattle.

    PubMed

    Wyckoff, John H; Howland, Jeri L; Scott, Catherine M O'Connell; Smith, Robert A; Confer, Anthony W

    2005-11-30

    Augmentation of immunization of cattle Brucella abortus S19 or a B. abortus soluble protein extract (SPEBA) vaccine through administration of recombinant bovine IL 2 (rBoIL 2) was evaluated. Seventy-five heifers were divided among 6 groups that were treated with the following: Group 1, no treatment; Group 2, rBoIL 2 (1microg/kg) on day 0; Group 3, SPEBA (2 mg) on day 0 and week 9; Group 4, SPEBA + rBoIL 2 on day 0, SPEBA on week 9; Group 5, S19 (10(7) CFU) on day 0 and week 9; Group 6, S19 + rBoIL 2 on day 0, S19 only on week 9. Approximately, 6 months after vaccination, cattle were bred by natural service, and at mid-gestation pregnant cattle were challenged intraconjunctivally with 9.1 x 10(5) CFU of virulent B. abortus S2308. Pre- and post-challenge antibody responses were measured by an enzyme-linked immunosorbent assay, a particle concentration fluorescence assay, and the card test. Lymphoproliferation (LP) responses to gamma-irradiated B. abortus and SPEBA antigens were measured in peripheral blood mononuclear cells. After vaccination, antibody responses to B. abortus elevated rapidly in SPEBA- and S19-vaccinates with and without rBoIL 2, however, these responses were significantly (P < 0.05) higher in vaccinates which also received rBoIL 2. Antibody levels for all vaccinated groups had returned to those of negative control groups by the challenge date with the exception of the SPEBA/rBoIL 2 group. In general, LP responses were higher in vaccinated or rBoIL 2-treated cattle than for unvaccinated controls. Challenge of 48 pregnant heifers resulted in abortions in 4/9 of Group 1, 0/9 of Group 2, 4/8 of Group 3, 2/9 of Group 4, 1/7 of Group 5, and 0/6 of Group 6 cattle. Treatment with rBoIL 2 alone (Group 2) provided significant (P < 0.05) protection from infection, abortions and induction of sero-positive status compared to untreated (Group 1) cattle. Co-administration of rBoIL 2 with S19 resulted in significant (P < 0.05) augmentation in onset, duration and

  2. Lateral flow-based antibody testing for Chlamydia trachomatis.

    PubMed

    Gwyn, Sarah; Mitchell, Alexandria; Dean, Deborah; Mkocha, Harran; Handali, Sukwan; Martin, Diana L

    2016-08-01

    We describe here a lateral flow-based assay (LFA) for the detection of antibodies against immunodominant antigen Pgp3 from Chlamydia trachomatis, the causative agent of urogenital chlamydia infection and ocular trachoma. Optimal signal detection was achieved when the gold-conjugate and test line contained Pgp3, creating a dual sandwich capture assay. The LFA yielded positive signals with serum and whole blood but not with eluted dried blood spots. For serum, the agreement of the LFA with the non-reference multiplex assay was 96%, the specificity using nonendemic pediatric sera was 100%, and the inter-rater agreement was κ=0.961. For whole blood, the agreement of LFA with multiplex was 81.5%, the specificity was 100%, and the inter-rater agreement was κ=0.940. The LFA was tested in a field environment and yielded similar results to those from laboratory-based testing. These data show the successful development of a lateral flow assay for detection of antibodies against Pgp3 with reliable use in field settings, which would make antibody-based testing for trachoma surveillance highly practical, especially after cessation of trachoma elimination programs. PMID:27208400

  3. Antibody-mediated modulation of arthritis induced by Chlamydia.

    PubMed Central

    Rank, R. G.; Ramsey, K. H.; Hough, A. J.

    1988-01-01

    The purpose of this investigation was to determine the role of the humoral immune response in the production of arthritis in mice immunized with the chlamydial agent of mouse pneumonitis (MoPn) (Chlamydia trachomatis biovar). Mice were made B cell deficient (BCD) by treatment with rabbit antiserum to murine IgM. Control mice included animals treated similarly with normal rabbit serum or phosphate-buffered saline. Male mice were immunized with MoPn inactivated with ultraviolet irradiation while female mice were immunized by genital tract infection with viable chlamydiae. Arthritis was elicited in all mice by intra-articular inoculation of inactivated MoPn. When knee joints were examined for pathologic changes at varying times after challenge, a marked enhancement of the arthritis was observed in both male and female BCD mice when compared with controls at all time points. These data indicated that the humoral immune response is not essential for the production of arthritic disease in this model but may have some role in the modulation of the process in immunologically intact animals. Persistence of chlamydial antigen in joint tissue of BCD mice suggested that antibody may play a role in the elimination of antigen, thus decreasing the stimulus for the development of cell-mediated immunologic injury. Regulatory role for T suppressor cells cannot be ruled out however, because B cell deficient mice have been shown to lack certain T suppressor cell subsets. Images Figure 3 Figure 4 Figure 6 Figure 7 Figure 9 PMID:3400779

  4. Comparison between ICT and PCR for diagnosis of Chlamydia trachomatis.

    PubMed

    Khan, E R; Hossain, M A; Paul, S K; Mahmud, C; Hasan, M M; Rahman, M M; Nahar, K; Kubayashi, N

    2012-04-01

    Chlamydia trachomatis is an obligate intracellular gram-negative bacterium which is the most prevalent cause of bacterial sexually transmitted infections (STI). The present study was carried to diagnose genital Chlamydia trachomatis infection among women of reproductive age, attending Mymensingh Medical College Hospital, during July 2009 to June 2010 by Immunochromatographic test (ICT) and Polymerase chain reaction (PCR). A total of 70 females were included in this study. Out of 70 cases 56 were symptomatic and 14 asymptomatic. Endocervical swabs were collected from each of the cases and examined by Immunochromatographic test (ICT) for antigen detection and Polymerase chain reaction (PCR) for detection of endogenous plasmid-based nucleic acid. A total 29(41.4%) of the cases were found positive for C. trachomatis either by ICT or PCR. Of the 56 symptomatic cases, 19(33.9%) were found ICT positive and 17(30.4%) were PCR positive. Among 14 asymptomatic females, 2(14.3%) were ICT positive and none were PCR positive. Though PCR is highly sensitive but a total of twelve cases were found ICT positive but PCR negative. It may be due to presence of plasmid deficient strain of C trachomatis which could be amplified by ompA based (Chromosomal gene) multiplex PCR.

  5. Prevalence of genital mycoplasmas, ureaplasmas and chlamydia in pregnancy.

    PubMed

    Govender, S; Theron, G B; Odendaal, H J; Chalkley, L J

    2009-11-01

    The study was designed to determine the prevalence of genital mycoplasmas, ureaplasmas and Chlamydia on women attending their first prenatal visit, in conjunction with pre-term labour or HIV status. For pre-term labour (2003), 199 women were monitored for pre-term delivery (<37 weeks); for colonisation and HIV (2005), 219 women were screened. Microbial detection was performed on DNA extracted from endocervical swabs employing PCR techniques. Colonisation was seen to be highest in the 14-20 year age group from 2003. In women aged > or = 21 years, co-colonisation was 13%, although there was a shift from co-colonisation with Mycoplasma hominis and Ureaplasma urealyticum in 2003, to other dual/triple combinations in 2005. Overall, major trends from both collection periods were that the prevalence of U. urealyticum tended to be higher in women > or = 26 years, while the prevalence of Chlamydia trachomatis and M. hominis lower. No association was evident between colonisation with M. hominis, U. urealyticum, Ureaplasma parvum and labour outcome. HIV status had no effect on the prevalence/co-colonisation of M. hominis, U. urealyticum or C. trachomatis. The importance of genital mycoplasmas, ureaplasmas and C. trachomatis in long-term aetiologies requires further investigations, certainly in relation to syndromic management regimens that fail to reduce colonisation rates. PMID:19821660

  6. Candidate vaginal microbicides with activity against Chlamydia trachomatis and Neisseriagonorrhoeae.

    PubMed

    Chu, Hencelyn; Slepenkin, Anatoly; Elofsson, Mikael; Keyser, Pia; de la Maza, Luis M; Peterson, Ellena M

    2010-08-01

    Vaginal microbicides with activity towards organisms that cause sexually transmitted infections have been proposed as a strategy to reduce transmission. Small-molecule inhibitors of Chlamydia trachomatis serovar D belonging to the class of salicylidene acylhydrazides (INPs) have been shown to work through a mechanism that involves iron restriction. Expanding on this work, ten INPs were tested against a lymphogranuloma venereum strain of C. trachomatis (serovar L2), Neisseria gonorrhoeae, and hydrogen peroxide-producing Lactobacillus crispatus and Lactobacillus jensenii. Seven INPs had minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations of <50 microM towards C. trachomatis L2. Three INPs had a MIC <12.5 microM against N. gonorrhoeae. Inhibition was reversed by iron, holo-transferrin and holo-lactoferrin but not by the iron-poor forms of these compounds. The compounds exhibited no bactericidal activity toward Lactobacillus. The INPs were not cytotoxic to HeLa 229 cells. When INP 0341 was tested in a mouse model of a Chlamydia vaginal infection there was a significant reduction in the number of mice shedding C. trachomatis up to 4 days after infection (P<0.01). In summary, select INPs are promising vaginal microbicide candidates as they inhibit the growth of two common sexually transmitted organisms in vitro, are active in a mouse model against C. trachomatis, are not cytotoxic and do not inhibit organisms that compose the normal vaginal flora. PMID:20605703

  7. MLVA16 typing of Portuguese human and animal Brucella melitensis and Brucella abortus isolates.

    PubMed

    Ferreira, Ana Cristina; Chambel, Lélia; Tenreiro, Tania; Cardoso, Regina; Flor, Lídia; Dias, Isabel Travassos; Pacheco, Teresa; Garin-Bastuji, Bruno; Le Flèche, Philippe; Vergnaud, Gilles; Tenreiro, Rogério; de Sá, Maria Inácia Corrêa

    2012-01-01

    To investigate the epidemiological relationship of isolates from different Portuguese geographical regions and to assess the diversity among isolates, the MLVA16(Orsay) assay (panels 1, 2A and 2B) was performed with a collection of 126 Brucella melitensis (46 human and 80 animal isolates) and 157 B. abortus field isolates, seven vaccine strains and the representative reference strains of each species. The MLVA16(Orsay) showed a similar high discriminatory power (HGDI 0.972 and 0.902) for both species but panel 1 and 2A markers displayed higher diversity (HGDI 0.693) in B. abortus compared to B. melitensis isolates (HGDI 0.342). The B. melitensis population belong to the "Americas" (17%) and "East Mediterranean" (83%) groups. No isolate belonged to the "West Mediterranean" group. Eighty-five percent of the human isolates (39 in 46) fit in the "East-Mediterranean" group where a single lineage known as MLVA11 genotype 116 is responsible for the vast majority of Brucella infections in humans. B. abortus isolates formed a consistent group with bv1 and bv3 isolates in different clusters. Four MLVA11 genotypes were observed for the first time in isolates from S. Jorge and Terceira islands from Azores. From the collection of isolates analysed in this study we conclude that MLVA16(Orsay) provided a clear view of Brucella spp. population, confirming epidemiological linkage in outbreak investigations. In particular, it suggests recent and ongoing colonisation of Portugal with one B. melitensis lineage usually associated with East Mediterranean countries.

  8. MLVA16 Typing of Portuguese Human and Animal Brucella melitensis and Brucella abortus Isolates

    PubMed Central

    Ferreira, Ana Cristina; Chambel, Lélia; Tenreiro, Tania; Cardoso, Regina; Flor, Lídia; Dias, Isabel Travassos; Pacheco, Teresa; Garin-Bastuji, Bruno; Le Flèche, Philippe; Vergnaud, Gilles; Tenreiro, Rogério; de Sá, Maria Inácia Corrêa

    2012-01-01

    To investigate the epidemiological relationship of isolates from different Portuguese geographical regions and to assess the diversity among isolates, the MLVA16Orsay assay (panels 1, 2A and 2B) was performed with a collection of 126 Brucella melitensis (46 human and 80 animal isolates) and 157 B. abortus field isolates, seven vaccine strains and the representative reference strains of each species. The MLVA16Orsay showed a similar high discriminatory power (HGDI 0.972 and 0.902) for both species but panel 1 and 2A markers displayed higher diversity (HGDI 0.693) in B. abortus compared to B. melitensis isolates (HGDI 0.342). The B. melitensis population belong to the “Americas” (17%) and “East Mediterranean” (83%) groups. No isolate belonged to the “West Mediterranean” group. Eighty-five percent of the human isolates (39 in 46) fit in the “East-Mediterranean” group where a single lineage known as MLVA11 genotype 116 is responsible for the vast majority of Brucella infections in humans. B. abortus isolates formed a consistent group with bv1 and bv3 isolates in different clusters. Four MLVA11 genotypes were observed for the first time in isolates from S. Jorge and Terceira islands from Azores. From the collection of isolates analysed in this study we conclude that MLVA16Orsay provided a clear view of Brucella spp. population, confirming epidemiological linkage in outbreak investigations. In particular, it suggests recent and ongoing colonisation of Portugal with one B. melitensis lineage usually associated with East Mediterranean countries. PMID:22905141

  9. Molecular structure of the Brucella abortus metalloprotein RicA, a Rab2-binding virulence effector.

    PubMed

    Herrou, Julien; Crosson, Sean

    2013-12-17

    The Gram-negative intracellular pathogen Brucella abortus is the causative agent of brucellosis, which is among the most common zoonoses globally. The B. abortus RicA protein binds the host-expressed guanosine nucleotide-binding protein, Rab2, and modulates B. abortus infection biology. We have solved the first X-ray crystal structure of RicA to 2.7 Å resolution and have quantified the affinity of RicA binding to human Rab2 in its GDP-bound and nucleotide-free forms. RicA adopts a classic γ-carbonic anhydrase (γ-CA) fold containing a left-handed β-helix followed by a C-terminal α-helix. Two homotrimers of RicA occupy the crystallographic asymmetric unit. Though no zinc was included in the purification or crystallization buffers, zinc is contained within the RicA crystals, as demonstrated by X-ray fluorescence spectroscopy. Electron density for a Zn(2+) ion coordinated by three histidine residues is evident in the putative active site of RicA. However, purified RicA preparations do not exhibit carbonic anhydrase activity, suggesting that Zn(2+) may not be the physiologically relevant metal cofactor or that RicA is not a bona fide carbonic anhydrase enzyme. Isothermal titration calorimetry (ITC) measurements of purified RicA binding to purified human Rab2 and GDP-Rab2 revealed similar equilibrium affinities (Kd ≈ 35 and 40 μM, respectively). This study thus defines RicA as a Zn(2+)-binding γ-carbonic anhydrase-like protein that binds the human membrane fusion/trafficking protein Rab2 with low micromolar affinity in vitro. These results support a model in which γ-CA family proteins may evolve unique cellular functions while retaining many of the structural hallmarks of archetypal γ-CA enzymes.

  10. Different resistance patterns of reference and field strains of Brucella abortus.

    PubMed

    Miranda, Karina L; Dorneles, Elaine M S; Poester, Fernando P; Martins Filho, Paulo S; Pauletti, Rebeca B; Lage, Andrey P

    2015-03-01

    The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO 2 . Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL) was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL). All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 ( B. abortus biovar 3) all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75-0.80 (10 μg/mL). These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus.

  11. Molecular structure of the Brucella abortus metalloprotein RicA, a Rab2-binding virulence effector

    PubMed Central

    Herrou, Julien; Crosson, Sean

    2014-01-01

    The Gram-negative intracellular pathogen Brucella abortus is the causative agent of brucellosis, which is among the most common zoonoses globally. The B. abortus RicA protein binds the host-expressed guanosine nucleotide-binding protein, Rab2, and modulates B. abortus infection biology. We have solved the first X-ray crystal structure of RicA to 2.7 Å resolution, and have quantified the affinity of RicA binding to human Rab2 in its GDP bound and nucleotide free forms. RicA adopts a classic γ-carbonic anhydrase (γ-CA) fold containing a left-handed β-helix followed by a C-terminal α-helix. Two homotrimers of RicA occupy the crystallographic asymmetric unit. Though no zinc was included in the purification or crystallization buffers, zinc is contained within the RicA crystals as demonstrated by X-ray fluorescence spectroscopy. Electron density for a Zn2+ ion coordinated by three histidine residues is evident in the putative active site of RicA. However, purified RicA preparations do not exhibit carbonic anhydrase activity, suggesting that Zn2+ may not be the physiologically relevant metal cofactor, or that RicA is not a bona fide carbonic anhydrase enzyme. Isothermal titration calorimetry (ITC) measurements of purified RicA binding to purified human Rab2 and GDP-Rab2 revealed similar equilibrium affinities (KD ≈ 35 µM and 40 µM, respectively). This study thus defines RicA as a Zn2+ binding, γ-carbonic anhydrase-like protein that binds the human membrane fusion/trafficking protein, Rab2, with low micromolar affinity in vitro. These results support a model in which γ-CA family proteins may evolve unique cellular functions while retaining many of the structural hallmarks of archetypal γ-CA enzymes. PMID:24251537

  12. Different resistance patterns of reference and field strains of Brucella abortus

    PubMed Central

    Miranda, Karina L.; Dorneles, Elaine M. S.; Poester, Fernando P.; Martins, Paulo S.; Pauletti, Rebeca B.; Lage, Andrey P.

    2015-01-01

    The aim of this study was to evaluate the growth of the B. abortus reference strains and field isolates on media containing different inhibitor agents. Reference strains were seeded on tryptose agar containing: i-erythritol (1.0 mg/mL), fuchsin (20 μg/mL and 80 μg/mL), thionin (2.5 μg/mL and 10 μg/mL), rifampicin (200 μg/mL) and safranin O (200 μg/mL). Field isolates were tested only on media containing i-erythritol, rifampicin and thionin. Furthermore, each suspension was also inoculated on tryptose agar incubated in air, to test its ability to grow without CO 2 . Sensitivity to fuchsin was similar among reference strains evaluated. Growth of S19, 544 and 2308 but not RB51 were inhibited on media containing rifampicin. Medium with safranin O showed no inhibition for RB51, 544 and 2308, but it partially inhibited the S19 growth as well as medium containing i-erythritol. Treatment/control growth ratio for 2308 on tryptose agar containing thionin (2.5 μg/mL) was approximatelly 1.0, whereas S19 and RB51 showed 0.85 and 0.89 ratios, respectively. Growth of 544, S19 and RB51 but not 2308 was completely inhibited on medium with thionin (10 μg/mL). All field strains grew on medium containing i-erythritol, but were completelly inhibited by rifampicin. With exception of A1 ( B. abortus biovar 3) all field isolates grew on medium with thionin, although some strains showed a treatment/control growth ratio of 0.75–0.80 (10 μg/mL). These results showed that tryptose agar with thionin, i-erythritol or rifampicin could be useful for differentiating vaccine, challenge and field strains of B. abortus. PMID:26221116

  13. Proinflammatory Response of Human Trophoblastic Cells to Brucella abortus Infection and upon Interactions with Infected Phagocytes.

    PubMed

    Fernández, Andrea G; Ferrero, Mariana C; Hielpos, M Soledad; Fossati, Carlos A; Baldi, Pablo C

    2016-02-01

    Trophoblasts are targets of infection by Brucella spp. but their role in the pathophysiology of pregnancy complications of brucellosis is unknown. Here we show that Brucella abortus invades and replicates in the human trophoblastic cell line Swan-71 and that the intracellular survival of the bacterium depends on a functional virB operon. The infection elicited significant increments of interleukin 8 (IL8), monocyte chemotactic protein 1 (MCP-1), and IL6 secretion, but levels of IL1beta and tumor necrosis factor-alpha (TNF-alpha) did not vary significantly. Such proinflammatory response was not modified by the absence of the Brucella TIR domain-containing proteins BtpA and BtpB. The stimulation of Swan-71 cells with conditioned medium (CM) from B. abortus-infected human monocytes (THP-1 cells) or macrophages induced a significant increase of IL8, MCP-1 and IL6 as compared to stimulation with CM from non-infected cells. Similar results were obtained when stimulation was performed with CM from infected neutrophils. Neutralization studies showed that IL1beta and/or TNF-alpha mediated the stimulating effects of CM from infected phagocytes. Reciprocally, stimulation of monocytes and neutrophils with CM from Brucella-infected trophoblasts increased IL8 and/or IL6 secretion. These results suggest that human trophoblasts may provide a local inflammatory environment during B. abortus infections either through a direct response to the pathogen or through interactions with monocytes/macrophages or neutrophils, potentially contributing to the pregnancy complications of brucellosis.

  14. Proinflammatory Response of Human Trophoblastic Cells to Brucella abortus Infection and upon Interactions with Infected Phagocytes.

    PubMed

    Fernández, Andrea G; Ferrero, Mariana C; Hielpos, M Soledad; Fossati, Carlos A; Baldi, Pablo C

    2016-02-01

    Trophoblasts are targets of infection by Brucella spp. but their role in the pathophysiology of pregnancy complications of brucellosis is unknown. Here we show that Brucella abortus invades and replicates in the human trophoblastic cell line Swan-71 and that the intracellular survival of the bacterium depends on a functional virB operon. The infection elicited significant increments of interleukin 8 (IL8), monocyte chemotactic protein 1 (MCP-1), and IL6 secretion, but levels of IL1beta and tumor necrosis factor-alpha (TNF-alpha) did not vary significantly. Such proinflammatory response was not modified by the absence of the Brucella TIR domain-containing proteins BtpA and BtpB. The stimulation of Swan-71 cells with conditioned medium (CM) from B. abortus-infected human monocytes (THP-1 cells) or macrophages induced a significant increase of IL8, MCP-1 and IL6 as compared to stimulation with CM from non-infected cells. Similar results were obtained when stimulation was performed with CM from infected neutrophils. Neutralization studies showed that IL1beta and/or TNF-alpha mediated the stimulating effects of CM from infected phagocytes. Reciprocally, stimulation of monocytes and neutrophils with CM from Brucella-infected trophoblasts increased IL8 and/or IL6 secretion. These results suggest that human trophoblasts may provide a local inflammatory environment during B. abortus infections either through a direct response to the pathogen or through interactions with monocytes/macrophages or neutrophils, potentially contributing to the pregnancy complications of brucellosis. PMID:26792938

  15. Pressure surge attenuator

    DOEpatents

    Christie, Alan M.; Snyder, Kurt I.

    1985-01-01

    A pressure surge attenuation system for pipes having a fluted region opposite crushable metal foam. As adapted for nuclear reactor vessels and heads, crushable metal foam is disposed to attenuate pressure surges.

  16. Deletion in the gene BruAb2_0168 of Brucella abortus strains: diagnostic challenges

    PubMed Central

    Dean, A S; Schelling, E; Bonfoh, B; Kulo, A E; Boukaya, G A; Pilo, P; Raoult, D

    2014-01-01

    Three Brucella abortus strains were isolated from joint hygromas from cows in northern Togo. Two deletions in the 5′ side of the gene BruAb2_0168 were identified. As this gene is used for species identification, these deletions have consequences for diagnostic procedures. Multiple locus variable number of tandem repeat (VNTR) analysis was therefore performed for species identification. The strains showed unique VNTR profiles, providing some of the first genotypic data from West Africa. More molecular and epidemiological data are needed from the region, in order to better understand transmission patterns and develop suitable diagnostic assays. PMID:24450581

  17. Successful Medical Treatment of Prosthetic Mitral Valve Endocarditis Caused by Brucella abortus

    PubMed Central

    Lee, Seung-Ah; Shin, Hyo-Sun; Lee, Hee-Sun; Choi, Hong-Mi; Kim, Hyung-Kwan

    2014-01-01

    Although Brucella endocarditis is a rare complication of human brucellosis, it is the main cause of the mortality in this disease. Traditionally, the therapeutic approach to endocarditis caused by Brucella species requires a combination of antimicrobial therapy and valve replacement surgery. In the literature, only a few cases of mitral prosthetic valve endocarditis caused by Brucella species have been successfully treated without reoperation. We present a case of a 42-year-old man with a prosthetic mitral valve infected by Brucella abortus who was cured solely by medical treatment. PMID:25469149

  18. Dibenzocyclooctadiene lignans from Schisandra spp. selectively inhibit the growth of the intracellular bacteria Chlamydia pneumoniae and Chlamydia trachomatis.

    PubMed

    Hakala, Elina; Hanski, Leena; Uvell, Hanna; Yrjönen, Teijo; Vuorela, Heikki; Elofsson, Mikael; Vuorela, Pia Maarit

    2015-10-01

    Lignans from Schisandra chinensis berries show various pharmacological activities, of which their antioxidative and cytoprotective properties are among the most studied ones. Here, the first report on antibacterial properties of six dibenzocyclooctadiene lignans found in Schisandra spp. is presented. The activity was shown on two related intracellular Gram-negative bacteria Chlamydia pneumoniae and Chlamydia trachomatis upon their infection in human epithelial cells. All six lignans inhibited C. pneumoniae inclusion formation and infectious progeny production. Schisandrin B inhibited C. pneumoniae inclusion formation even when administered 8 h post infection, indicating a target that occurs relatively late within the infection cycle. Upon infection, lignan-pretreated C. pneumoniae elementary bodies had impaired inclusion formation capacity. The presence and substitution pattern of methylenedioxy, methoxy and hydroxyl groups of the lignans had a profound impact on the antichlamydial activity. In addition our data suggest that the antichlamydial activity is not caused only by the antioxidative properties of the lignans. None of the compounds showed inhibition on seven other bacteria, suggesting a degree of selectivity of the antibacterial effect. Taken together, the data presented support a role of the studied lignans as interesting antichlamydial lead compounds.

  19. Chlamydia Infection Across Host Species Boundaries Promotes Distinct Sets of Transcribed Anti-Apoptotic Factors.

    PubMed

    Messinger, Joshua E; Nelton, Emmalin; Feeney, Colleen; Gondek, David C

    2015-01-01

    Chlamydiae, obligate intracellular bacteria, cause significant human and veterinary associated diseases. Having emerged an estimated 700-million years ago, these bacteria have twice adapted to humans as a host species, causing sexually transmitted infection (C. trachomatis) and respiratory associated disease (C. pneumoniae). The principle mechanism of host cell defense against these intracellular bacteria is the induction of cell death via apoptosis. However, in the "arms race" of co-evolution, Chlamydiae have developed mechanisms to promote cell viability and inhibit cell death. Herein we examine the impact of Chlamydiae infection across multiple host species on transcription of anti-apoptotic genes. We found mostly distinct patterns of gene expression (Mcl1 and cIAPs) elicited by each pathogen-host pair indicating Chlamydiae infection across host species boundaries does not induce a universally shared host response. Understanding species specific host-pathogen interactions is paramount to deciphering how potential pathogens become emerging diseases. PMID:26779446

  20. Causality of Chlamydiae in Arthritis and Spondyloarthritis: a Plea for Increased Translational Research.

    PubMed

    Zeidler, Henning; Hudson, Alan P

    2016-02-01

    Current molecular genetic understanding of the metabolically active persistent infection state of Chlamydia trachomatis and Chlamydia pneumoniae in the synovium in patients with arthritis and spondyloarthritis favors a causal relationship. Here, we examine how adequately the accepted criteria for that etiologic relationship are fulfilled, emphasizing the situation in which these microorganisms cannot be cultivated by standard or other means. We suggest that this unusual situation of causality by chlamydiae in rheumatic disease requires establishment of a consensus regarding microorganism-specific terminology as well as the development of new diagnostic and classification criteria. Recent studies demonstrate the value of molecular testing for diagnosis of reactive arthritis, undifferentiated spondyloarthritis, and undifferentiated arthritis caused by C. trachomatis and C. pneumoniae in clinical practice. Data regarding combination antibiotic therapy is consistent with the causative role of chlamydiae for these diseases. Observations of multiple intra-articular coinfections require more research to understand the implications and to respond to them.

  1. Detection of Chlamydiae from freshwater environments by PCR, amoeba coculture and mixed coculture.

    PubMed

    Corsaro, Daniele; Venditti, Danielle

    2009-10-01

    Water systems have been shown to be a potential source of Chlamydiae, intracellular symbionts of eukaryotes. However, their diversity is likely underestimated, and natural hosts remain undetermined in many cases. In this study, we combined PCR-based and cultivation approaches to search for chlamydiae in different freshwaters, including natural ponds, garden pots and fountains. From a total of 40 samples, we recovered 16 phylotypes, clustered into nine species-level taxa belonging to the Parachlamydiaceae, Simkaniaceae, Rhabdochlamydiaceae, cvE6 lineage and a novel lineage. Parachlamydiaceae (four species) were recovered by amoeba coculture, while the other chlamydiae were maintained in mixed eukaryotic cultures. This study confirms the widespread occurrence of novel chlamydiae in the environment and enlarges our knowledge of their biodiversity in freshwater habitats.

  2. Chlamydia Infection Across Host Species Boundaries Promotes Distinct Sets of Transcribed Anti-Apoptotic Factors

    PubMed Central

    Messinger, Joshua E.; Nelton, Emmalin; Feeney, Colleen; Gondek, David C.

    2015-01-01

    Chlamydiae, obligate intracellular bacteria, cause significant human and veterinary associated diseases. Having emerged an estimated 700-million years ago, these bacteria have twice adapted to humans as a host species, causing sexually transmitted infection (C. trachomatis) and respiratory associated disease (C. pneumoniae). The principle mechanism of host cell defense against these intracellular bacteria is the induction of cell death via apoptosis. However, in the “arms race” of co-evolution, Chlamydiae have developed mechanisms to promote cell viability and inhibit cell death. Herein we examine the impact of Chlamydiae infection across multiple host species on transcription of anti-apoptotic genes. We found mostly distinct patterns of gene expression (Mcl1 and cIAPs) elicited by each pathogen-host pair indicating Chlamydiae infection across host species boundaries does not induce a universally shared host response. Understanding species specific host-pathogen interactions is paramount to deciphering how potential pathogens become emerging diseases. PMID:26779446

  3. Asymptomatic natural Chlamydia pecorum infection reduces growth rates in calves by up to 48 percent

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Intracellular Chlamydia (C.) bacteria cause in cattle some acute but rare diseases such as abortion, sporadic bovine encephalomyelitis, kerato-conjunctivitis, pneumonia, enteritis and polyarthritis. Much more frequent, essentially ubiquitous worldwide, are low-level, asymptomatic chlamydial infecti...

  4. Persistent chlamydiae: from cell culture to a paradigm for chlamydial pathogenesis.

    PubMed Central

    Beatty, W L; Morrison, R P; Byrne, G I

    1994-01-01

    Chlamydiae are medically important bacteria responsible for a wide range of human infections and diseases. Repeated episodes of infection promote chronic inflammation associated with detrimental immune system-mediated pathologic changes. However, the true nature of chlamydial pathogenesis may encompass repeated infection superimposed upon persistent infection, which would allow for heightened immune reactivity. During the course of chlamydial infection, numerous host elaborated factors with inhibitory or modifying effects may cause alterations in the chlamydia-host cell relationship such that the organism is maintained in a nonproductive stage of growth. Abnormal or persistent chlamydiae have been recognized under a variety of cell culture systems. The numerous factors associated with altered growth suggest an innate flexibility in the developmental cycle of chlamydiae. This review evaluates in vitro studies of chlamydial persistence and correlates these model systems to features of natural chlamydial disease. Images PMID:7854252

  5. Intramuscular Immunisation with Chlamydial Proteins Induces Chlamydia trachomatis Specific Ocular Antibodies

    PubMed Central

    Badamchi-Zadeh, Alexander; McKay, Paul F.; Holland, Martin J.; Paes, Wayne; Brzozowski, Andrzej; Lacey, Charles; Follmann, Frank; Tregoning, John S.; Shattock, Robin J.

    2015-01-01

    Background Ocular infection with Chlamydia trachomatis can cause trachoma, which is the leading cause of blindness due to infection worldwide. Despite the large-scale implementation of trachoma control programmes in the majority of countries where trachoma is endemic, there remains a need for a vaccine. Since C. trachomatis infects the conjunctival epithelium and stimulates an immune response in the associated lymphoid tissue, vaccine regimens that enhance local antibody responses could be advantageous. In experimental infections of non-human primates (NHPs), antibody specificity to C. trachomatis antigens was found to change over the course of ocular infection. The appearance of major outer membrane protein (MOMP) specific antibodies correlated with a reduction in ocular chlamydial burden, while subsequent generation of antibodies specific for PmpD and Pgp3 correlated with C. trachomatis eradication. Methods We used a range of heterologous prime-boost vaccinations with DNA, Adenovirus, modified vaccinia Ankara (MVA) and protein vaccines based on the major outer membrane protein (MOMP) as an antigen, and investigated the effect of vaccine route, antigen and regimen on the induction of anti-chlamydial antibodies detectable in the ocular lavage fluid of mice. Results Three intramuscular vaccinations with recombinant protein adjuvanted with MF59 induced significantly greater levels of anti-MOMP ocular antibodies than the other regimens tested. Intranasal delivery of vaccines induced less IgG antibody in the eye than intramuscular delivery. The inclusion of the antigens PmpD and Pgp3, singly or in combination, induced ocular antigen-specific IgG antibodies, although the anti-PmpD antibody response was consistently lower and attenuated by combination with other antigens. Conclusions If translatable to NHPs and/or humans, this investigation of the murine C. trachomatis specific ocular antibody response following vaccination provides a potential mouse model for the rapid

  6. TLR2 and TLR4 signaling pathways are required for recombinant Brucella abortus BCSP31-induced cytokine production, functional upregulation of mouse macrophages, and the Th1 immune response in vivo and in vitro.

    PubMed

    Li, Jia-Yun; Liu, Yuan; Gao, Xiao-Xue; Gao, Xiang; Cai, Hong

    2014-09-01

    Brucella abortus is a zoonotic Gram-negative pathogen that causes brucelosis in ruminants and humans. Toll-like receptors (TLRs) recognize Brucella abortus and initiate antigen-presenting cell activities that affect both innate and adaptive immunity. In this study, we focused on recombinant Brucella cell-surface protein 31 (rBCSP31) to determine its effects on mouse macrophages. Our results demonstrated that rBCSP31 induced TNF-α, IL-6 and IL-12p40 production, which depended on the activation of mitogen-activated protein kinases (MAPKs) by stimulating the rapid phosphorylation of p38 and JNK and the activation of transcription factor NF-κB in macrophages. In addition, continuous exposure (>24 h) of RAW264.7 cells to rBCSP31 significantly enhanced IFN-γ-induced expression of MHC-II and the ability to present rBCSP31 peptide to CD4(+) T cells. Furthermore, we found that rBCSP31 could interact with both TLR2 and TLR4. The rBCSP31-induced cytokine production by macrophages from TLR2(-/-) and TLR4(-/-) mice was lower than that from C57BL/6 macrophages, and the activation of NF-κB and MAPKs was attenuated in macrophages from TLR2(-/-) and TLR4(-/-) mice. In addition, CD4(+) T cells from C57BL/6 mice immunized with rBCSP31 produced higher levels of IFN-γ and IL-2 compared with CD4(+) T cells from TLR2(-/-) and TLR4(-/-) mice. Macrophages from immunized C57BL/6 mice produced higher levels of IL-12p40 than those from TLR2(-/-) and TLR4(-/-) mice. Furthermore, immunization with rBCSP31 provided better protection in C57BL/6 mice than in TLR2(-/-) and TLR4(-/-) mice after B. abortus 2308 challenge. These results indicate that rBCSP31 is a TLR2 and TLR4 agonist that induces cytokine production, upregulates macrophage function and induces the Th1 immune response.

  7. TLR2 and TLR4 signaling pathways are required for recombinant Brucella abortus BCSP31-induced cytokine production, functional upregulation of mouse macrophages, and the Th1 immune response in vivo and in vitro

    PubMed Central

    Li, Jia-Yun; Liu, Yuan; Gao, Xiao-Xue; Gao, Xiang; Cai, Hong

    2014-01-01

    Brucella abortus is a zoonotic Gram-negative pathogen that causes brucelosis in ruminants and humans. Toll-like receptors (TLRs) recognize Brucella abortus and initiate antigen-presenting cell activities that affect both innate and adaptive immunity. In this study, we focused on recombinant Brucella cell-surface protein 31 (rBCSP31) to determine its effects on mouse macrophages. Our results demonstrated that rBCSP31 induced TNF-α, IL-6 and IL-12p40 production, which depended on the activation of mitogen-activated protein kinases (MAPKs) by stimulating the rapid phosphorylation of p38 and JNK and the activation of transcription factor NF-κB in macrophages. In addition, continuous exposure (>24 h) of RAW264.7 cells to rBCSP31 significantly enhanced IFN-γ-induced expression of MHC-II and the ability to present rBCSP31 peptide to CD4+ T cells. Furthermore, we found that rBCSP31 could interact with both TLR2 and TLR4. The rBCSP31-induced cytokine production by macrophages from TLR2−/− and TLR4−/− mice was lower than that from C57BL/6 macrophages, and the activation of NF-κB and MAPKs was attenuated in macrophages from TLR2−/− and TLR4−/− mice. In addition, CD4+ T cells from C57BL/6 mice immunized with rBCSP31 produced higher levels of IFN-γ and IL-2 compared with CD4+ T cells from TLR2−/− and TLR4−/− mice. Macrophages from immunized C57BL/6 mice produced higher levels of IL-12p40 than those from TLR2−/− and TLR4−/− mice. Furthermore, immunization with rBCSP31 provided better protection in C57BL/6 mice than in TLR2−/− and TLR4−/− mice after B. abortus 2308 challenge. These results indicate that rBCSP31 is a TLR2 and TLR4 agonist that induces cytokine production, upregulates macrophage function and induces the Th1 immune response. PMID:24769793

  8. Performance evaluation of a new rapid urine test for chlamydia in men: prospective cohort study

    PubMed Central

    Nadala, Elpidio-Cesar; Goh, Beng T; Magbanua, Jose-Paolo; Barber, Penelope; Swain, Alison; Alexander, Sarah; Laitila, Vivian; Michel, Claude-Edouard; Mahilum-Tapay, Lourdes; Ushiro-Lumb, Ines; Ison, Catherine

    2009-01-01

    Objective To evaluate the performance of a rapid test for chlamydia with first void male urine samples as a potential tool for diagnosis and screening of chlamydial infection in men. Design Evaluation of test performance in prospective cohort study. Settings A young people’s sexual health centre (site 1) and a genitourinary medicine clinic (site 2) in the United Kingdom. Participants 1211 men aged 16-73 attending either of the two sites. Main outcome measures Sensitivity, specificity, positive predictive value, and negative predictive value of the Chlamydia Rapid Test versus polymerase chain reaction assay. Relation between the visual signal of the Chlamydia Rapid Test and organism load. Results Detection rates for Chlamydia trachomatis infection with polymerase chain reaction were 4.4% (20/454) at site 1 and 11.9% (90/757) at site 2. Compared with polymerase chain reaction assay, the resolved sensitivity, specificity, positive predictive value, and negative predictive value of the Chlamydia Rapid Test was 82.6% (90/109), 98.5% (1085/1102), 84.1% (90/107), and 98.3% (1085/1104), respectively. The organism load in first void urine samples that were positive for chlamydia ranged from 7.28×102 to 6.93×106 plasmids/ml and correlated significantly with the visual signal of the Chlamydia Rapid Test (r=0.7897, P<0.001). Conclusions The performance of the new Chlamydia Rapid Test with first void male urine samples indicates that it would be an effective diagnostic tool for chlamydial infection in men. The availability of test results within an hour allows for immediate treatment and contact tracing, potentially reducing the risks of persistent infection and onward transmission. The test could also provide a simple and reliable alternative to nucleic acid amplification assays for testing of male urine in chlamydial screening programmes in high prevalence settings. PMID:19638650

  9. [Improvement of health care for patients with upper respiratory tract diseases associated with chlamydia infection].

    PubMed

    Kapustina, T A; Markina, A N; Parilova, O V

    2012-01-01

    At present the issues in regard to Chlamydia infection are not only limited by urogenital system. By the way optimal organization and non-urogenital chlamydiosis treatment strategy (with respiratory tract involvement in particular) have not been worked out yet and require immediate solutions. Due to new knowledge on respiratory chlamidiosis the authors discuss scientific background for future development of complex measures and main directions of health care support strategy for patients with upper respiratory associated with Chlamydia infection.

  10. Chlamydia trachomatis genital tract infection of antibody-deficient gene knockout mice.

    PubMed Central

    Su, H; Feilzer, K; Caldwell, H D; Morrison, R P

    1997-01-01

    The importance of antibody-mediated immunity in primary and secondary Chlamydia trachomatis genital tract infections was examined by using a definitive model of B-cell deficiency, the microMT/microMT gene knockout mouse. Vaginally infected B-cell-deficient microMT/microMT mice developed a self-limiting primary infection that was indistinguishable from infection of control C57BL/6 mice. Sera and vaginal secretions from infected mice were analyzed for anti-Chlamydia antibodies. C57BL/6 mice produced high-titered serum anti-Chlamydia immunoglobulin G2a (IgG2a), IgG2b, and IgA antibodies, and vaginal washes contained predominately anti-Chlamydia IgA. Serum and vaginal washes from infected B-cell-deficient mice were negative for anti-Chlamydia antibody. T-cell proliferation and delayed-type hypersensitivity assays were used as measures of Chlamydia-specific cell-mediated immunity and were found to be comparable for C57BL/6 and B-cell-deficient mice. Seventy days following primary infection, mice were rechallenged to assess acquired immunity. B-cell-deficient mice which lack anti-Chlamydia antibodies were more susceptible to reinfection than immunocompetent C57BL/6 mice. However, acquired immune resistance was evident in both strains of mice and characterized by decreased shedding of chlamydiae and an infection of shorter duration. Thus, this study demonstrates that cell-mediated immune responses alone were capable of resolving chlamydial infection; however, in the absence of specific antibody, mice were more susceptible to reinfection. Therefore, these data suggest that both humoral and cell-mediated immune responses were important mediators of immune protection in this model, though cell-mediated immune responses appear to play a more dominant role. PMID:9169723

  11. Tracer attenuation in groundwater

    NASA Astrophysics Data System (ADS)

    Cvetkovic, Vladimir

    2011-12-01

    The self-purifying capacity of aquifers strongly depends on the attenuation of waterborne contaminants, i.e., irreversible loss of contaminant mass on a given scale as a result of coupled transport and transformation processes. A general formulation of tracer attenuation in groundwater is presented. Basic sensitivities of attenuation to macrodispersion and retention are illustrated for a few typical retention mechanisms. Tracer recovery is suggested as an experimental proxy for attenuation. Unique experimental data of tracer recovery in crystalline rock compare favorably with the theoretical model that is based on diffusion-controlled retention. Non-Fickian hydrodynamic transport has potentially a large impact on field-scale attenuation of dissolved contaminants.

  12. A Live Vaccine from Brucella abortus Strain 82 for Control of Cattle Brucellosis in the Russian Federation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During the first half of the 20th century, widespread regulatory efforts to control cattle brucellosis (Brucella abortus) in the Union of Soviet Socialist Republics were essentially nonexistent, and control was limited to selective test and slaughter of serologic agglutination reactors. By the 1950...

  13. Comparison of abortion and infection after experimental challenge of pregnant bison and cattle with Brucella abortus strain 2308

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A comparative study was conducted using data from naive bison (n=45) and cattle (n=46) from 8 and 6 studies, respectively, in which a standardized Brucella abortus strain 2308 experimental challenge was administered. The incidence of abortion, fetal infection, uterine or mammary infection, or infec...

  14. Dextran sulfate sodium upregulates MAPK signaling for the uptake and subsequent intracellular survival of Brucella abortus in murine macrophages.

    PubMed

    Reyes, Alisha Wehdnesday Bernardo; Arayan, Lauren Togonon; Simborio, Hannah Leah Tadeja; Hop, Huynh Tan; Min, WonGi; Lee, Hu Jang; Kim, Dong Hee; Chang, Hong Hee; Kim, Suk

    2016-02-01

    Brucellosis is one of the major zoonoses worldwide that inflicts important health problems in animal and human. Here, we demonstrated that dextran sulfate sodium (DSS) significantly increased adhesion of Brucella (B.) abortus in murine macrophages compared to untreated cells. Even without infection, Brucella uptake into macrophages increased and F-actin reorganization was induced compared with untreated cells. Furthermore, DSS increased the phosphorylation of MAPKs (ERK1/2 and p38α) in Brucella-infected, DSS-treated cells compared with the control cells. Lastly, DSS markedly increased the intracellular survival of Brucella abortus in macrophages by up to 48 h. These results suggest that DSS enhanced the adhesion and phagocytosis of B. abortus into murine macrophages by stimulating the MAPK signaling proteins phospho-ERK1/2 and p38α and that DSS increased the intracellular survival of B. abortus by inhibiting colocalization of Brucella-containing vacuoles (BCVs) with the late endosome marker LAMP-1. This study emphasizes the enhancement of the phagocytic and intracellular modulatory effects of DSS, which may suppress the innate immune system and contribute to prolonged Brucella survival and chronic infection.

  15. Draft Genome Sequence of the Intermediate Rough Vaccine Strain Brucella abortus S19Δper Mutant.

    PubMed

    Chaudhuri, Pallab; Goswami T, Tapas K; Lalsiamthara, Jonathan; Kaur, Gurpreet; Vishnu, Udayakumar S; Sankarasubramanian, Jagadesan; Gunasekaran, Paramasamy; Rajendhran, Jeyaprakash

    2015-11-12

    Here, we report the genome sequence of the intermediate rough vaccine strain mutant, Brucella abortus S19Δper. The length of the draft genome was 3,271,238 bp, with 57.2% G+C content. A total of 3,204 protein-coding genes and 56 RNA genes were predicted.

  16. The Brucella abortus virulence regulator, LovhK, is a sensor kinase in the general stress response signaling pathway

    PubMed Central

    Kim, Hye-Sook; Willett, Jonathan W.; Jain-Gupta, Neeta; Fiebig, Aretha; Crosson, Sean

    2014-01-01

    Summary In the intracellular pathogen Brucella abortus, the general stress response (GSR) signaling system determines survival under acute stress conditions in vitro, and is required for long-term residence in a mammalian host. To date, the identity of the Brucella sensor kinase(s) that function to perceive stress and directly activate GSR signaling have remained undefined. We demonstrate that the flavin-binding sensor histidine kinase, LovhK (bab2_0652), functions as a primary B. abortus GSR sensor. LovhK efficiently and specifically phosphorylates the central GSR regulator, PhyR, and activates transcription of a set of genes that closely overlaps the known B. abortus GSR regulon. Deletion of lovhK severely compromises cell survival under defined oxidative and acid stress conditions. We further show that lovhK is required for cell survival during the early phase of mammalian cell infection and for establishment of long-term residence in a mouse infection model. Finally, we present evidence that particular regions of primary structure within the two N-terminal PAS domains of LovhK have distinct sensory roles under specific environmental conditions. This study elucidates new molecular components of a conserved signaling pathway that regulates B. abortus stress physiology and infection biology. PMID:25257300

  17. Draft Genome Sequence of the Intermediate Rough Vaccine Strain Brucella abortus S19Δper Mutant

    PubMed Central

    Chaudhuri, Pallab; Goswami T, Tapas K.; Lalsiamthara, Jonathan; Kaur, Gurpreet; Vishnu, Udayakumar S.; Sankarasubramanian, Jagadesan; Gunasekaran, Paramasamy

    2015-01-01

    Here, we report the genome sequence of the intermediate rough vaccine strain mutant, Brucella abortus S19Δper. The length of the draft genome was 3,271,238 bp, with 57.2% G+C content. A total of 3,204 protein-coding genes and 56 RNA genes were predicted. PMID:26564050

  18. Development of a dual vaccine for prevention of Brucella abortus infection and Escherichia coli O157:H7 intestinal colonization.

    PubMed

    Iannino, Florencia; Herrmann, Claudia K; Roset, Mara S; Briones, Gabriel

    2015-05-01

    Zoonoses that affect human and animal health have an important economic impact. In the study now presented, a bivalent vaccine has been developed that has the potential for preventing the transmission from cattle to humans of two bacterial pathogens: Brucella abortus and Shiga toxin-producing Escherichia coli (STEC). A 66kDa chimeric antigen, composed by EspA, Intimin, Tir, and H7 flagellin (EITH7) from STEC, was constructed and expressed in B. abortus Δpgm vaccine strain (BabΔpgm). Mice orally immunized with BabΔpgm(EITH7) elicited an immune response with the induction of anti-EITH7 antibodies (IgA) that clears an intestinal infection of E. coli O157:H7 three times faster (t=4 days) than mice immunized with BabΔpgm carrier strain (t=12 days). As expected, mice immunized with BabΔpgm(EITH7) strain also elicited a protective immune response against B. abortus infection. A Brucella-based vaccine platform is described capable of eliciting a combined protective immune response against two bacterial pathogens with diverse lifestyles-the intracellular pathogen B. abortus and the intestinal extracellular pathogen STEC.

  19. A live vaccine from Brucella abortus strain 82 for control of cattle brucellosis in the Russian Federation.

    PubMed

    Ivanov, Arkady V; Salmakov, Konstantin M; Olsen, Steven C; Plumb, Glenn E

    2011-06-01

    During the first half of the twentieth century, widespread regulatory efforts to control cattle brucellosis due to Brucella abortus in the Union of Soviet Socialist Republics were essentially non-existent, and control was limited to selective test and slaughter of serologic agglutination reactors. By the 1950s, 2-3 million cattle were being vaccinated annually with the strain 19 vaccine, but because this vaccine induced strong, long-term titers on agglutination tests that interfered with identification of cattle infected with field strains of B. abortus, its use in cattle was discontinued in 1970. Soviet scientists then began a comprehensive program of research to identify vaccines with high immunogenicity, weak responses on agglutination tests and low pathogenicity in humans, as a foundation for widespread control of cattle brucellosis. While several new vaccines that induced weak or no responses on serologic agglutination tests were identified by experiments in guinea pigs and cattle, a large body of experimental and field studies suggested that the smooth-rough strain SR82 vaccine combined the desired weak agglutination test responses with comparatively higher efficacy against brucellosis. In 1974, prior to widespread use of strain SR82 vaccine, over 5300 cattle farms across the Russian Federation were known to be infected with B. abortus. By January 2008, only 68 cattle farms in 18 regions were known to be infected with B. abortus, and strain SR82 continues to be the most widely and successfully used vaccine in many regions of the Russian Federation.

  20. Efficacy of dart or booster vaccination with strain RB51 in protecting bison against experimental Brucella abortus challenge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vaccination is an effective tool for reducing the prevalence of brucellosis in natural hosts. In this study, we characterized the efficacy of the Brucella abortus strain RB51 (RB51) vaccine in bison when delivered by single intramuscular vaccination (Hand RB51), single pneumatic dart delivery (Dart ...

  1. Protective role of antibodies induced by Brucella melitensis B115 against B. melitensis and Brucella abortus infections in mice.

    PubMed

    Adone, Rosanna; Francia, Massimiliano; Pistoia, Claudia; Petrucci, Paola; Pesciaroli, Michele; Pasquali, Paolo

    2012-06-01

    It has been demonstrated that antibodies specific for O-PS antigen of Brucella smooth strains are involved in the protective immunity of brucellosis. Since the rough strain Brucella melitensis B115 was able to protect mice against wild Brucella strains brucellosis despite the lack of anti-OPS antibodies, in this study we evaluated the biological significance of antibodies induced by this strain, directed to antigens other than O-PS, passively tranferred to untreated mice prior to infection with Brucella abortus 2308 and B. melitensis 16M virulent strains. The protective ability of specific antisera collected from mice vaccinated with B. melitensis B115, B. abortus RB51 and B. abortus S19 strains was compared. The results indicated that antibodies induced by B115 were able to confer a satisfactory protection, especially against B. abortus 2308, similar to that conferred by the antiserum S19, while the RB51 antiserum was ineffective. These findings suggest that antibodies induced by B115 could act as opsonins as well as antibodies anti-O-PS, thus triggering more efficient internalization and degradation of bacteria within phagocytes. This is the first study assessing the efficacy of antibodies directed to antigens other than O-PS in the course of brucellosis infection.

  2. [Multiplication of Brucella abortus and production of nitric oxide in two macrophage cell lines of different origin].

    PubMed

    Serafino, J; Conde, S; Zabal, O; Samartino, L

    2007-01-01

    Brucella abortus is a bacterium which causes abortions and infertility in cattle and undulant fever in humans. It multiplies intracellularly, evading the mechanisms of cellular death. Nitric oxide (NO) is important in the regulation of the immune response. In the present work, we studied the ability of three B. abortus strains to survive intracellularly in two macrophage cell lines. The bacterial multiplication in both cell lines was determined at two different times in UFC/ ml units. Moreover the inoculated cells were also observed under light-field and fluorescence microscopy stained with Giemsa and acridine orange, respectively. The stain of both cellular lines showed similar results with respect to the UFC/ml determination. The presence of B. abortus was confirmed by electronic microscopy. In both macrophage cell lines inoculated with the rough strain RB51, the multiplication diminished and the level of NO was higher, compared with cells inoculated with smooth strains (S19 and 2308). These results suggest that the absence of O-chain of LPS probably affects the intracellular growth of B. abortus.

  3. Prevalence and characterization of Chlamydia DNA in zoo animals in Japan.

    PubMed

    Kabeya, Hidenori; Sato, Shingo; Maruyama, Soichi

    2015-09-01

    Because many people visit zoos, prevention of zoonoses is important from the standpoint of public health. This study examined the prevalence of Chlamydia among zoo animals in Japan by PCR and characterized these bacteria by performing phylogenetic analyses of the sequences of the variable domain (VD) 2 and VD4 regions of the ompA gene, which encodes the Chlamydia major outer membrane protein. Fecal samples were collected from 1150 zoo animals in five zoos and examined for Chlamydia DNA. Chlamydia psittaci DNA was found in 3.9% of mammals, 7.2% of birds and 8.1% of reptiles. The prevalence of Chlamydia pneumoniae DNA was significantly higher in reptiles (5.8%) than in mammals (0.3%) and birds (0.3%). Phylogenetic analysis of the ompA VD2 region from 18 samples showed that nine were in three different clusters containing C. psittaci strains, six were in a cluster containing C. pneumoniae strains and three each formed a distinct branch. Furthermore, phylogenetic analysis of the ompA VD4 region showed that C. pneumoniae DNAs from reptiles were close to those from human patients. The C. pneumoniae DNAs from the European glass lizard, Emerald tree boa, and Panther chameleon were classified in clusters that were distinct from other strains, suggesting that these reptiles had each been infected with a specific C. pneumoniae genotype. This study showed that diverse Chlamydia strains have been prevalent among a variety of zoo animals. PMID:26215334

  4. Do the factors associated with successful contact tracing of patients with gonorrhoea and Chlamydia differ?

    PubMed Central

    Ross, J. D.; Sukthankar, A.; Radcliffe, K. W.; Andre, J.

    1999-01-01

    OBJECTIVE: To assess and compare factors which may be associated with successful contact tracing in patients with gonorrhoea and chlamydia. STUDY DESIGN: Prospective observational study of patients attending a genitourinary medicine clinic with a diagnosis of gonorrhoea or chlamydia. Multivariate analysis model including demographic, socioeconomic, and behavioural variables. RESULTS: The attendance of at least one sexual contact was associated with naming more contacts for patients with gonorrhoea (OR 1.44, 95% CI 1.04-2.01). A history of gonorrhoea was associated with successful contact tracing for patients with chlamydia (OR 1.46, 95% CI 1.12-1.9). Successful contact tracing, as defined by at least one confirmed contact attendance after the index case, was not associated with age, sex, sexual orientation, history of chlamydia, use of condoms, marital status, ethnicity, or socioeconomic status for either gonorrhoea or chlamydia. CONCLUSIONS: Differences in the composition of the core groups infected with gonorrhoea and chlamydia are not explained by differences in contact tracing success. In the clinic setting studied, the outcome of contact tracing was not associated with a variety of demographic, socioeconomic, and behaviour factors. 


 PMID:10448364

  5. Structural characterization of muropeptides from Chlamydia trachomatis peptidoglycan by mass spectrometry resolves “chlamydial anomaly”

    PubMed Central

    Packiam, Mathanraj; Weinrick, Brian; Jacobs, William R.; Maurelli, Anthony T.

    2015-01-01

    The “chlamydial anomaly,” first coined by James Moulder, describes the inability of researchers to detect or purify peptidoglycan (PG) from pathogenic Chlamydiae despite genetic and biochemical evidence and antibiotic susceptibility data that suggest its existence. We recently detected PG in Chlamydia trachomatis by a new metabolic cell wall labeling method, however efforts to purify PG from pathogenic Chlamydiae have remained unsuccessful. Pathogenic chlamydial species are known to activate nucleotide-binding oligomerization domain-containing protein 2 (NOD2) innate immune receptors by as yet uncharacterized ligands, which are presumed to be PG fragments (muramyl di- and tripeptides). We used the NOD2-dependent activation of NF-κB by C. trachomatis-infected cell lysates as a biomarker for the presence of PG fragments within specific lysate fractions. We designed a new method of muropeptide isolation consisting of a double filtration step coupled with reverse-phase HPLC fractionation of Chlamydia-infected HeLa cell lysates. Fractions that displayed NOD2 activity were analyzed by electrospray ionization mass spectrometry, confirming the presence of muramyl di- and tripeptides in Chlamydia-infected cell lysate fractions. Moreover, the mass spectrometry data of large muropeptide fragments provided evidence that transpeptidation and transglycosylation reactions occur in pathogenic Chlamydiae. These results reveal the composition of chlamydial PG and disprove the “glycanless peptidoglycan” hypothesis. PMID:26290580

  6. Pathogenic Potential of Novel Chlamydiae and Diagnostic Approaches to Infections Due to These Obligate Intracellular Bacteria

    PubMed Central

    Corsaro, Daniele; Greub, Gilbert

    2006-01-01

    Novel chlamydiae are newly recognized members of the phylum Chlamydiales that are only distantly related to the classic Chlamydiaceae, i.e., Chlamydia and Chlamydophila species. They also exibit an obligate biphasic intracellular life cycle within eukaryote host cells. Some of these new chlamydiae are currently considered potential emerging human and/or animal pathogens. Parachlamydia acanthamoebae and Simkania negevensis are both emerging respiratory human pathogens, Waddlia chondrophila could be a novel abortigenic bovine agent, and Piscichlamydia salmonis has recently been identified as an agent of the gill epitheliocystis in the Atlantic salmon. Fritschea spp. and Rhabdochlamydia spp. seem to be confined to arthropods, but some evidence for human exposure exists. In this review, we first summarize the data supporting a pathogenic potential of the novel chlamydiae for humans and other vertebrates and the interactions that most of these chlamydiae have with free-living amoebae. We then review the diagnostic approaches to infections potentially due to the novel chlamydiae, especially focusing on the currently available PCR-based protocols, mammalian cell culture, the amoebal coculture system, and serology. PMID:16614250

  7. Pathogenic potential of novel Chlamydiae and diagnostic approaches to infections due to these obligate intracellular bacteria.

    PubMed

    Corsaro, Daniele; Greub, Gilbert

    2006-04-01

    Novel chlamydiae are newly recognized members of the phylum Chlamydiales that are only distantly related to the classic Chlamydiaceae, i.e., Chlamydia and Chlamydophila species. They also exhibit an obligate biphasic intracellular life cycle within eukaryote host cells. Some of these new chlamydiae are currently considered potential emerging human and/or animal pathogens. Parachlamydia acanthamoebae and Simkania negevensis are both emerging respiratory human pathogens, Waddlia chondrophila could be a novel abortigenic bovine agent, and Piscichlamydia salmonis has recently been identified as an agent of the gill epitheliocystis in the Atlantic salmon. Fritschea spp. and Rhabdochlamydia spp. seem to be confined to arthropods, but some evidence for human exposure exists. In this review, we first summarize the data supporting a pathogenic potential of the novel chlamydiae for humans and other vertebrates and the interactions that most of these chlamydiae have with free-living amoebae. We then review the diagnostic approaches to infections potentially due to the novel chlamydiae, especially focusing on the currently available PCR-based protocols, mammalian cell culture, the amoebal coculture system, and serology.

  8. Abortion and subsequent excretion of chlamydiae from the reproductive tract of sheep during estrus.

    PubMed Central

    Papp, J R; Shewen, P E; Gartley, C J

    1994-01-01

    Chlamydia psittaci serovar 1 infection in pregnant sheep typically causes abortion or the birth of weak lambs. Eight sheep that experienced chlamydia-induced abortion during their first pregnancy were successfully rebred yearly for the past 2 years. Chlamydia-specific lipopolysaccharide was detectable for approximately 3 weeks in vaginal swabs taken from the experimentally infected sheep following abortion. There was no evidence of chlamydiae in vaginal, placental, or neonatal samples obtained immediately after each subsequent successful pregnancy. Sera collected from the experimentally infected sheep had persistent, high antibody levels to C. psittaci, suggesting continued exposure of the immune system to the organism. Examination of vaginal specimens obtained during various stages of the estrus cycle revealed detectable levels of chlamydiae only when the animal was near ovulation. Chlamydiae were not detected in swabs from sheep that did not experience abortion. Enhanced chlamydial excretion during the periovulation period of sheep may provide sufficient stimulation of the immune system to account for the persistent antibody response. Furthermore, the association between estrus and chlamydial shedding has important implications for transmission of infection to other ewes during breeding. PMID:8063395

  9. Prevalence and characterization of Chlamydia DNA in zoo animals in Japan.

    PubMed

    Kabeya, Hidenori; Sato, Shingo; Maruyama, Soichi

    2015-09-01

    Because many people visit zoos, prevention of zoonoses is important from the standpoint of public health. This study examined the prevalence of Chlamydia among zoo animals in Japan by PCR and characterized these bacteria by performing phylogenetic analyses of the sequences of the variable domain (VD) 2 and VD4 regions of the ompA gene, which encodes the Chlamydia major outer membrane protein. Fecal samples were collected from 1150 zoo animals in five zoos and examined for Chlamydia DNA. Chlamydia psittaci DNA was found in 3.9% of mammals, 7.2% of birds and 8.1% of reptiles. The prevalence of Chlamydia pneumoniae DNA was significantly higher in reptiles (5.8%) than in mammals (0.3%) and birds (0.3%). Phylogenetic analysis of the ompA VD2 region from 18 samples showed that nine were in three different clusters containing C. psittaci strains, six were in a cluster containing C. pneumoniae strains and three each formed a distinct branch. Furthermore, phylogenetic analysis of the ompA VD4 region showed that C. pneumoniae DNAs from reptiles were close to those from human patients. The C. pneumoniae DNAs from the European glass lizard, Emerald tree boa, and Panther chameleon were classified in clusters that were distinct from other strains, suggesting that these reptiles had each been infected with a specific C. pneumoniae genotype. This study showed that diverse Chlamydia strains have been prevalent among a variety of zoo animals.

  10. Using Fluorescent Proteins to Visualize and Quantitate Chlamydia Vacuole Growth Dynamics in Living Cells.

    PubMed

    Zuck, Meghan; Feng, Caroline; Hybiske, Kevin

    2015-10-13

    The obligate intracellular bacterium Chlamydia elicits a great burden on global public health. C. trachomatis is the leading bacterial cause of sexually transmitted infection and also the primary cause of preventable blindness in the world. An essential determinant for successful infection of host cells by Chlamydia is the bacterium's ability to manipulate host cell signaling from within a novel, vacuolar compartment called the inclusion. From within the inclusion, Chlamydia acquire nutrients required for their 2-3 day developmental growth, and they additionally secrete a panel of effector proteins onto the cytosolic face of the vacuole membrane and into the host cytosol. Gaps in our understanding of Chlamydia biology, however, present significant challenges for visualizing and analyzing this intracellular compartment. Recently, a reverse-imaging strategy for visualizing the inclusion using GFP expressing host cells was described. This approach rationally exploits the intrinsic impermeability of the inclusion membrane to large molecules such as GFP. In this work, we describe how GFP- or mCherry-expressing host cells are generated for subsequent visualization of chlamydial inclusions. Furthermore, this method is shown to effectively substitute for costly antibody-based enumeration methods, can be used in tandem with other fluorescent labels, such as GFP-expressing Chlamydia, and can be exploited to derive key quantitative data about inclusion membrane growth from a range of Chlamydia species and strains.

  11. Brucella abortus induces TNF-α-dependent astroglial MMP-9 secretion through mitogen-activated protein kinases

    PubMed Central

    2013-01-01

    Background Central nervous system (CNS) invasion by bacteria of the genus Brucella results in an inflammatory disorder called neurobrucellosis. We have recently demonstrated that B. abortus infects microglia and astrocytes, eliciting the production of a variety of pro-inflammatory cytokines which contribute to CNS damage. Matrix metalloproteinases (MMP) have been implicated in inflammatory tissue destruction in a range of pathological situations in the CNS. Increased MMP secretion is induced by pro-inflammatory cytokines in a variety of CNS diseases characterized by tissue-destructive pathology. Methods In this study, the molecular mechanisms that regulate MMP secretion from Brucella-infected astrocytes in vitro were investigated. MMP-9 was evaluated in culture supernatants by ELISA, zymography and gelatinolytic activity. Involvement of mitogen-activated protein kinases (MAPK) signaling pathways was evaluated by Western blot and using specific inhibitors. The role of TNF-α was evaluated by ELISA and by assays with neutralizing antibodies. Results B. abortus infection induced the secretion of MMP-9 from murine astrocytes in a dose-dependent fashion. The phenomenon was independent of bacterial viability and was recapitulated by L-Omp19, a B. abortus lipoprotein model, but not its LPS. B. abortus and L-Omp19 readily activated p38 and Erk1/2 MAPK, thus enlisting these pathways among the kinase pathways that the bacteria may address as they invade astrocytes. Inhibition of p38 or Erk1/2 significantly diminished MMP-9 secretion, and totally abrogated production of this MMP when both MAPK pathways were inhibited simultaneously. A concomitant abrogation of B. abortus- and L-Omp19-induced TNF-α production was observed when p38 and Erk1/2 pathways were inhibited, indicating that TNF-α could be implicated in MMP-9 secretion. MMP-9 secretion induced by B. abortus or L-Omp19 was completely abrogated when experiments were conducted in the presence of a TNF-α neutralizing

  12. Brucella abortus depends on pyruvate phosphate dikinase and malic enzyme but not on Fbp and GlpX fructose-1,6-bisphosphatases for full virulence in laboratory models.

    PubMed

    Zúñiga-Ripa, Amaia; Barbier, Thibault; Conde-Álvarez, Raquel; Martínez-Gómez, Estrella; Palacios-Chaves, Leyre; Gil-Ramírez, Yolanda; Grilló, María Jesús; Letesson, Jean-Jacques; Iriarte, Maite; Moriyón, Ignacio

    2014-08-15

    The brucellae are the etiological agents of brucellosis, a worldwide-distributed zoonosis. These bacteria are facultative intracellular parasites and thus are able to adjust their metabolism to the extra- and intracellular environments encountered during an infectious cycle. However, this aspect of Brucella biology is imperfectly understood, and the nutrients available in the intracellular niche are unknown. Here, we investigated the central pathways of C metabolism used by Brucella abortus by deleting the putative fructose-1,6-bisphosphatase (fbp and glpX), phosphoenolpyruvate carboxykinase (pckA), pyruvate phosphate dikinase (ppdK), and malic enzyme (mae) genes. In gluconeogenic but not in rich media, growth of ΔppdK and Δmae mutants was severely impaired and growth of the double Δfbp-ΔglpX mutant was reduced. In macrophages, only the ΔppdK and Δmae mutants showed reduced multiplication, and studies with the ΔppdK mutant confirmed that it reached the replicative niche. Similarly, only the ΔppdK and Δmae mutants were attenuated in mice, the former being cleared by week 10 and the latter persisting longer than 12 weeks. We also investigated the glyoxylate cycle. Although aceA (isocitrate lyase) promoter activity was enhanced in rich medium, aceA disruption had no effect in vitro or on multiplication in macrophages or mouse spleens. The results suggest that B. abortus grows intracellularly using a limited supply of 6-C (and 5-C) sugars that is compensated by glutamate and possibly other amino acids entering the Krebs cycle without a critical role of the glyoxylate shunt.

  13. Brucella abortus Depends on Pyruvate Phosphate Dikinase and Malic Enzyme but Not on Fbp and GlpX Fructose-1,6-Bisphosphatases for Full Virulence in Laboratory Models

    PubMed Central

    Zúñiga-Ripa, Amaia; Barbier, Thibault; Conde-Álvarez, Raquel; Martínez-Gómez, Estrella; Palacios-Chaves, Leyre; Gil-Ramírez, Yolanda; Grilló, María Jesús; Letesson, Jean-Jacques; Iriarte, Maite

    2014-01-01

    The brucellae are the etiological agents of brucellosis, a worldwide-distributed zoonosis. These bacteria are facultative intracellular parasites and thus are able to adjust their metabolism to the extra- and intracellular environments encountered during an infectious cycle. However, this aspect of Brucella biology is imperfectly understood, and the nutrients available in the intracellular niche are unknown. Here, we investigated the central pathways of C metabolism used by Brucella abortus by deleting the putative fructose-1,6-bisphosphatase (fbp and glpX), phosphoenolpyruvate carboxykinase (pckA), pyruvate phosphate dikinase (ppdK), and malic enzyme (mae) genes. In gluconeogenic but not in rich media, growth of ΔppdK and Δmae mutants was severely impaired and growth of the double Δfbp-ΔglpX mutant was reduced. In macrophages, only the ΔppdK and Δmae mutants showed reduced multiplication, and studies with the ΔppdK mutant confirmed that it reached the replicative niche. Similarly, only the ΔppdK and Δmae mutants were attenuated in mice, the former being cleared by week 10 and the latter persisting longer than 12 weeks. We also investigated the glyoxylate cycle. Although aceA (isocitrate lyase) promoter activity was enhanced in rich medium, aceA disruption had no effect in vitro or on multiplication in macrophages or mouse spleens. The results suggest that B. abortus grows intracellularly using a limited supply of 6-C (and 5-C) sugars that is compensated by glutamate and possibly other amino acids entering the Krebs cycle without a critical role of the glyoxylate shunt. PMID:24936050

  14. Identification of a protective protein from stationary-phase exoproteome of Brucella abortus.

    PubMed

    Jain, Shikha; Kumar, Subodh; Dohre, Sudhir; Afley, Prachiti; Sengupta, Nabonita; Alam, Syed I

    2014-02-01

    Brucellosis is a worldwide zoonotic disease. No Brucella vaccine is available for use in humans, and existing animal vaccines have limitations. To search the putative vaccine candidates, we studied the exoproteome of Brucella abortus NCTC 10093 using 2-DE-MS approach. Twenty-six proteins were identified using MALDI-TOF/TOF tandem mass spectrometry. Outer membrane protein 25, d-galactose periplasmic-binding protein, oligopeptide ABC transporter protein and isopropylmalate synthase were found to be the most abundant proteins. Most proteins (6, 23%) were predicted to be involved in amino acid transport and metabolism followed by carbohydrate transport and metabolism (4, 15%). Outer membrane protein 25, Omp2b porin and one hypothetical protein were predicted as outer membrane proteins. In addition, Omp28, Omp31 and one ribosomal protein (L9) were also identified. The ribosomal protein L9 was produced as a recombinant protein and was studied in mouse model for vaccine potential. It was found to be immunogenic in terms of generating serum antibody response and release of IFN-γ from mice spleen cells. Recombinant L9-immunized mice were protected against challenge with virulent B. abortus strain 544, suggesting usefulness of ribosomal protein L9 as a good vaccine candidate against brucellosis.

  15. An evaluation of ELISA using recombinant Brucella abortus bacterioferritin (Bfr) for bovine brucellosis.

    PubMed

    Hop, Huynh Tan; Arayan, Lauren Togonon; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Min, WonGi; Lee, Hu Jang; Lee, Jin Ju; Chang, Hong Hee; Kim, Suk

    2016-04-01

    To date, detection of antibodies against the lipopolysaccharide portion is the backbone of most serodiagnostic methods for brucellosis screening. However this pose a risk for false positive reactions related to other pathogens especially that of Yersinia enterocolitica O:9 which has the most prominent cross reactivity with Brucella spp. In this study, cloning and expression of Brucella abortus bacterioferritin (Bfr) was accomplished by PCR amplification into an expression vector system, and purification of a recombinant B. abortus Bfr (rBfr). The immunogenicity of rBfr was confirmed by Western blot with Brucella-positive bovine serum. To determine whether rBfr has a potential benefit for use in the serodiagnosis of bovine brucellosis, rBfr-based ELISA was performed. Interestingly, rBfr was able to detect anti-Brucella antibodies in positive sera in a dependent manner of TAT values but did not show an immunoreaction with negative samples. Particularly, average OD492 values at the lowest, medium and highest TAT titer levels were 1.4, 2.2 and 2.6-fold increase compared with the cutoff value, respectively. The accuracy, specificity and sensitivity of rBfr showed 89.09%, 93.6% and 85.33%, respectively. These findings suggest that rBfr might be a good candidate for serological diagnosis development of bovine brucellosis.

  16. G1-arrested newborn cells are the predominant infectious form of the pathogen Brucella abortus

    PubMed Central

    Deghelt, Michaël; Mullier, Caroline; Sternon, Jean-François; Francis, Nayla; Laloux, Géraldine; Dotreppe, Delphine; Van der Henst, Charles; Jacobs-Wagner, Christine; Letesson, Jean-Jacques; De Bolle, Xavier

    2014-01-01

    Several intracellular pathogens, such as Brucella abortus, display a biphasic infection process starting with a non-proliferative stage of unclear nature. Here, we study the cell cycle of B. abortus at the single-cell level, in culture and during infection of HeLa cells and macrophages. The localization of segregation and replication loci of the two bacterial chromosomes indicates that, immediately after being engulfed by host-cell endocytic vacuoles, most bacterial cells are newborn. These bacterial cells do not initiate DNA replication for the next 4 to 6 h, indicating a G1 arrest. Moreover, growth is completely stopped during that time, reflecting a global cell cycle block. Growth and DNA replication resume later, although bacteria still reside within endosomal-like compartments. We hypothesize that the predominance of G1-arrested bacteria in the infectious population, and the bacterial cell cycle arrest following internalization, may constitute a widespread strategy among intracellular pathogens to colonize new proliferation niches. PMID:25006695

  17. Myeloid decidual dendritic cells and immunoregulation of pregnancy: defective responsiveness to Coxiella burnetii and Brucella abortus

    PubMed Central

    Gorvel, Laurent; Ben Amara, Amira; Ka, Mignane B.; Textoris, Julien; Gorvel, Jean-Pierre; Mege, Jean-Louis

    2014-01-01

    Dendritic cells (DCs) are a component of the placental immune system, but their role in pregnancy is still poorly understood. Decidual DCs (dDCs) were selected from at-term pregnancy on the basis of CD14 and CD11c expression. A phenotypic analysis revealed that dDCs are characterized by the expression of monocyte-derived DC (moDCs) markers and specific markers such as HLA-G and its ligand ILT4. As demonstrated by whole-genome microarray, dDCs expressed a specific gene program markedly distinct from that of moDCs; it included estrogen- and progesterone-regulated genes and genes encoding immunoregulatory cytokines, which is consistent with the context of foeto-maternal tolerance. A functional analysis of dDCs showed that they were unable to mature in response to bacterial ligands such as lipopolysaccharide or peptidoglycan, as assessed by the expression of HLA-DR, CD80, CD83, and CD86. When dDCs were incubated with bacteria known for their placenta tropism, Coxiella burnetii and Brucella abortus, they were also unable to mature and to produce inflammatory cytokines. It is likely that the defective maturation of dDCs and their inability to produce inflammatory cytokines is related to the spontaneous release of IL-10 by these cells. Taken together, these results suggest that dDCs exhibit an immunoregulatory program, which may favor the pathogenicity of C. burnetii or B. abortus. PMID:25566514

  18. Effect of entF deletion on iron acquisition and erythritol metabolism by Brucella abortus 2308.

    PubMed

    Jain, Neeta; Rodriguez, Araceli C; Kimsawatde, Gade; Seleem, Mohamed N; Boyle, Stephen M; Sriranganathan, Nammalwar

    2011-03-01

    Brucella abortus has been shown to produce two siderophores: 2,3-dihydroxybenzoic acid (2,3-DHBA) and brucebactin. Previous studies on Brucella have shown that 2,3-DHBA is associated with erythritol utilization and virulence in pregnant ruminants. The biosynthetic pathway and role of brucebactin are not known and the only gene shown to be involved so far is entF. Using cre-lox methodology, an entF mutant was created in wild-type B. abortus 2308. Compared with the wild-type strain, the ΔentF strain showed significant growth inhibition in iron minimal media that became exacerbated in the presence of an iron chelator. For the first time, we have demonstrated the death of the ΔentF strain under iron-limiting conditions in the presence of erythritol. Addition of FeCl(3) restored the growth of the ΔentF strain, suggesting a significant role in iron acquisition. Further, complementation of the ΔentF strain using a plasmid containing an entF gene suggested the absence of any polar effects. In contrast, there was no significant difference in survival and growth between the ΔentF and wild-type strains grown in the murine macrophage cell line J774A.1, suggesting that an alternate iron acquisition pathway is present in Brucella when grown intracellulary.

  19. Enzymatic Activity Analysis and Catalytic Essential Residues Identification of Brucella abortus Malate Dehydrogenase

    PubMed Central

    Han, Xiangan; Tong, Yongliang; Tian, Mingxing; Zhang, Yuxi; Sun, Xiaoqing; Wang, Shaohui; Qiu, Xusheng; Ding, Chan; Yu, Shengqing

    2014-01-01

    Malate dehydrogenase (MDH) plays important metabolic roles in bacteria. In this study, the recombinant MDH protein (His-MDH) of Brucella abortus was purified and its ability to catalyze the conversion of oxaloacetate (OAA) to L-malate (hereon referred to as MDH activity) was analyzed. Michaelis Constant (Km) and Maximum Reaction Velocity (Vmax) of the reaction were determined to be 6.45 × 10−3 M and 0.87 mM L−1 min−1, respectively. In vitro studies showed that His-MDH exhibited maximal MDH activity in pH 6.0 reaction buffer at 40°C. The enzymatic activity was 100%, 60%, and 40% inhibited by Cu2+, Zn2+, and Pb2+, respectively. In addition, six amino acids in the MDH were mutated to investigate their roles in the enzymatic activity. The results showed that the substitutions of amino acids Arg 89, Asp 149, Arg 152, His 176, or Thr 231 almost abolished the activity of His-MDH. The present study will help to understand MDH's roles in B. abortus metabolism. PMID:24895685

  20. Induction of specific cytotoxic lymphocytes in mice vaccinated with Brucella abortus RB51.

    PubMed

    He, Y; Vemulapalli, R; Zeytun, A; Schurig, G G

    2001-09-01

    A safe, more sensitive, nonradioactive, neutral red uptake assay was adopted to replace the traditional 51Cr release assay for detection of Brucella-specific cytotoxic T lymphocyte (CTL) activity. Our studies indicated that Brucella abortus strain RB51 vaccination of mice induced specific CTLs against both strain RB51- and strain 2308-infected J774.A1 macrophages but not against Listeria monocytogenes-infected J774.A1 cells. The antigen-specific cytotoxic activity was exerted by T lymphocytes but not by NK cells. CD3+ CD4+ T cells secreted the highest level of gamma interferon (IFN-gamma) and were able to exert a low but significant level of specific lysis of Brucella-infected macrophages. They also exerted a low level of nonspecific lysis of noninfected macrophages. In contrast, CD3+ CD8+ T cells secreted low levels of IFN-gamma but demonstrated high levels of specific lysis of Brucella-infected macrophages with no nonspecific lysis. These findings indicate that B. abortus strain RB51 vaccination of mice induces specific CTLs and suggest that CD3+ CD4+ and CD3+ CD8+ T cells play a synergistic role in the anti-Brucella activity.

  1. Epizootic abortion related to infections by Chlamydophila abortus and Chlamydophila pecorum in water buffalo (Bubalus bubalis).

    PubMed

    Greco, G; Corrente, M; Buonavoglia, D; Campanile, G; Di Palo, R; Martella, V; Bellacicco, A L; D'Abramo, M; Buonavoglia, C

    2008-06-01

    Water buffalo (Bubalus bubalis) are affected by high rates of embryonic mortality and abortion related to infectious diseases and non-infectious factors. A number of viral and bacterial infections have been associated with reproductive failure, but there is limited information on the role of chlamydial infections. In order to investigate the presence and the role of Chlamydiaceae in water buffalo a retrospective study was performed in a herd with a history of reproductive failure. During an 11-month period, the pregnant heifers suffered an abortion rate of 36.8% between the 3rd and 7th month of pregnancy. Antibodies to Chlamydiaceae were detected in 57% of the aborted cows, and in 0% of the overtly healthy cows used as control. By a nested-PCR assay, three of 14 vaginal swabs from aborted animals tested positive for Chlamydophila agents and, additionally, three out of seven aborted fetuses tested positive for Chlamydophila spp., with two being co-infections by Cp. abortus and Cp. pecorum and one being characterised as Cp. abortus. Sequence analysis of the amplicons confirmed the results of the nested-PCR. The presence of anti-Chlamydiaceae antibodies in more than half of the aborting animals (P<0.002) and the detection of Chlamydophila agents in several fetal organs and in the vaginal swabs are consistent with the history of abortions observed in the herd and suggest an abortifacient role by Chlamydophila spp. in water buffalo (B. Bubalis) herds.

  2. Entry of the lymphogranuloma venereum strain of Chlamydia trachomatis into host cells involves cholesterol-rich membrane domains.

    PubMed

    Jutras, Isabelle; Abrami, Laurence; Dautry-Varsat, Alice

    2003-01-01

    Chlamydiae are bacterial pathogens which develop strictly inside the epithelial cells of their hosts. The mechanism used by chlamydiae to enter cells is not well characterized; however, it is thought to consist of a receptor-mediated process. In addition, the formation of clathrin-coated pits appears to be dispensable for chlamydiae to be internalized by host cells. Clathrin-independent endocytosis has recently been shown to occur through cholesterol-rich lipid microdomains, which are characterized by detergent insolubility. In the present study, we investigated whether these lipid domains play a role in Chlamydia trachomatis serovar L2 internalization by host cells. Our results show that after binding to HeLa cells, chlamydiae are associated with detergent-resistant lipid microdomains (DRMs), which can be isolated by fractionation of infected HeLa cells and flotation on a sucrose gradient. After internalization by HeLa cells, chlamydiae were still found in DRMs. In addition, extraction of plasma membrane cholesterol inhibited infection of HeLa cells by C. trachomatis. Many of the proteins associated with DRMs are glycosylphosphatidylinositol (GPI)-anchored proteins; however, our results could not identify a role for GPI-anchored proteins in the entry process. The same results were obtained for Chlamydia psittaci strain GPIC. We propose that cholesterol-rich domains participate in the entry of chlamydiae into host cells. Chlamydia binding to cholesterol-rich domains may lead to coalescence of the bacterial cells, which could trigger internalization by host cells.

  3. Incidence and persistence of carcinogenic genital human papillomavirus infections in young women with or without Chlamydia trachomatis co-infection.

    PubMed

    Vriend, Henrike J; Bogaards, Johannes A; van Bergen, Jan E A M; Brink, Antoinette A T P; van den Broek, Ingrid V F; Hoebe, Christian J P A; King, Audrey J; van der Sande, Marianne A B; Wolffs, Petra F G; de Melker, Hester E

    2015-10-01

    We assessed whether infection with chlamydia increases the incidence of carcinogenic human papillomavirus (HPV) infections and if HPV persistence is affected by chlamydia co-infection. For 1982 women (16-29 years-old) participating in two consecutive rounds of a chlamydia screening implementation trial, swabs were polymerase chain reaction tested to detect chlamydia and 14 carcinogenic HPV genotypes. HPV type-specific incidence and persistence rates were stratified for chlamydia positivity at follow-up. Associations were assessed by multilevel logistic regression analyses with correction for sexual risk factors. HPV type-specific incidence ranged from 1.4% to 8.9% and persistence from 22.7% to 59.4% after a median follow-up of 11 months (interquartile range: 11-12). Differences in 1-year HPV persistence rates between chlamydia -infected and noninfected women were less distinct than differences in HPV incidence rates (pooled adjusted odds ratios of 1.17 [95% CI: 0.69-1.96] and 1.84 [95% CI: 1.36-2.47], respectively). The effect of chlamydia co-infection on HPV-infection risk did not significantly differ by HPV genotype. In conclusion, infection with chlamydia increases the risk of infection by carcinogenic HPV types and may enhance persistence of some HPV types. Although these findings could reflect residual confounding through unobserved risk factors, our results do give reason to explore more fully the association between chlamydia and HPV type-specific acquisition and persistence.

  4. Cell mediated immune response after challenge in Omp25 liposome immunized mice contributes to protection against virulent Brucella abortus 544.

    PubMed

    Goel, Divya; Rajendran, Vinoth; Ghosh, Prahlad C; Bhatnagar, Rakesh

    2013-02-01

    Brucellosis is a disease affecting various domestic and wild life species, and is caused by a bacterium Brucella. Keeping in view the serious economic and medical consequences of brucellosis, efforts have been made to prevent the infection through the use of vaccines. Cell-mediated immune responses [CMI] involving interferon gamma and cytotoxic CD4(+) and CD8(+) T cells are required for removal of intracellular Brucella. Omp25 has been reported to be involved in virulence of Brucella melitensis, Brucella abortus and Brucella ovis. In our previous study, we have shown the protective efficacy of recombinant Omp25, when administered intradermally. In this study, the recombinant Omp25 was formulated in PC-PE liposomes and PLGA microparticles, to enhance the protective immunity generated by it. Significant protection was seen with prime and booster liposome immunization in Balb/c mice against virulent B. abortus 544 as it was comparable to B. abortus S-19 vaccine strain. However, microparticle prime and booster immunization failed to give better protection when compared to B. abortus S-19 vaccine strain. This difference can be attributed to the stimulation of cell mediated immune response in PC-PE liposome immunized mice even after challenge which converted to cytotoxicity seen in CD4(+) and CD8(+) enriched lymphocytes. However, in PLGA microparticle immunized mice, cell mediated immunity was not generated after challenge as observed by decreased cytotoxicity of CD4(+) and CD8(+) enriched lymphocytes. Our study emphasizes on the importance of liposome encapsulating Omp25 immunization in conferring protection against B. abortus 544 challenge in Balb/c mice with a single dose immunization regimen.

  5. Bacterial survival, lymph node pathology, and serological responses of bison (Bison bison) vaccinated with Brucella abortus strain RB51 or strain 19.

    PubMed

    Olsen, S C; Cheville, N F; Kunkle, R A; Palmer, M V; Jensen, A E

    1997-01-01

    From August 1993 to June 1994, 3 month-old bison (Bison bison) were vaccinated with Brucella abortus strain RB51 (SRB51, n = 6), strain 19 (S19, n = 3), or with saline (n = 1) and serologic responses and persistence of vaccine strains within lymph nodes were monitored. Bison vaccinated with S19 had granulomatous lymphadenitis and greater peak numbers of B. abortus than those vaccinated with SRB51. Bison vaccinated with RB51 had similar histological lesions and B. abortus were still present in lymph nodes at 16 weeks. Although antibodies against RB51 were produced, standard tube agglutination test responses of RB51-vaccinates remained negative. The histological lesions of B. abortus infections in bison were similar to those observed in cattle, but bison did not clear SRB51 as rapidly as cattle.

  6. Chlamydia pneumoniae Promotes Dysfunction of Pancreatic Beta Cells

    PubMed Central

    Rodriguez, Annette R.; Plascencia-Villa, Germán; Witt, Colleen M.; Yu, Jieh-Juen; José-Yacamán, Miguel; Chambers, James P.; Perry, George; Guentzel, M. Neal; Arulanandam, Bernard P.

    2015-01-01

    The human pathogen Chlamydia pneumoniae has been implicated in chronic inflammatory diseases including type 2 diabetes. Therefore, we designed a study to evaluate pancreatic beta cells and mast cells during chlamydial infection. Our study revealed that C. pneumoniae infected mast cells significantly (p< 0.005) decreased beta cell ATP and insulin production, in contrast to uninfected mast cells co-cultured with beta cells. Infected mast cells exhibited pyknotic nuclei and active caspase-3 and caspase-1 expression. Additionally, ex vivo analyses of tissues collected from C. pneumoniae infected mice showed increased interleukin-1β production in splenocytes and pancreatic tissues as was observed with in vitro mast cell-beta cell co-cultures during C. pneumoniae infection. Notably, infected mast cells promoted beta cell destruction. Our findings reveal the negative effect of C. pneumoniae on mast cells, and the consequential impact on pancreatic beta cell function and viability. PMID:25863744

  7. Chlamydia trachomatis causing neonatal conjunctivitis in a tertiary care center.

    PubMed

    Kakar, S; Bhalla, P; Maria, A; Rana, M; Chawla, R; Mathur, N B

    2010-01-01

    Chlamydia trachomatis is considered a major aetiological agent of conjunctivitis in newborns. The objective of the present study was to determine the aetiology of neonatal conjunctivitis and clinico-epidemiological correlates of chlamydial ophthalmia neonatorum. Fifty-eight newborns with signs and symptoms of conjunctivitis were studied. Conjunctival specimens were subjected to Gram staining, routine bacteriological culture, culture for Neisseria gonorrhoeae and direct fluorescent antibody (DFA) staining for diagnosis of C. trachomatis infection. C. trachomatis was detected in 18 (31%) neonates. Findings suggest that since C. trachomatis is the most common cause of neonatal conjunctivitis, routine screening and treatment of genital C. trachomatis infection in pregnant women and early diagnosis and treatment of neonatal Chlamydial conjunctivitis may be considered for its prevention and control.

  8. Chlamydia trachomatis infection in "sine causa" recurrent abortion.

    PubMed

    Olliaro, P; Regazzetti, A; Gorini, G; Milano, F; Marchetti, A; Rondanelli, E G

    One hundred and one women suffering from "sine causa" recurrent abortion were screened for Chlamydia trachomatis (C.T.) infection by using direct examination, cultural and serological procedures. In this series, C.T. infection did not appear to be related to increased risk of recurrent abortion. The culture-positive and serology-positive rates (14.85% and 34.65%, respectively) did not differ from other unselected populations. Neither time from last abortion nor type of abortion were significantly related to C.T. infection. Nonetheless, the women who underwent examination within one year from last abortion and had a culture-positive partner as well, were more likely to present with a C.T.-positive culture.

  9. Use of polymerase chain reaction for detection of Chlamydia trachomatis.

    PubMed

    Ostergaard, L; Birkelund, S; Christiansen, G

    1990-06-01

    A polymerase chain reaction (PCR) assay was developed for detection of Chlamydia trachomatis DNA. From the published sequence of the common C. trachomatis plasmid, two primer sets were selected. Detection of amplified sequences was done by agarose gel electrophoresis of cleaved or uncleaved amplified sequences, Southern hybridization, or dot blot analysis. The PCR assay was optimized and, after 40 cycles of amplification with primer set II, demonstrated a sensitivity of 10(-17) g of DNA, which corresponds to the detection of one copy of the plasmid. Because of the high sensitivity, we developed a closed system in which airborne contamination was minimized. Analysis of 228 clinical samples tested by cell culture, IDEIA enzyme immunosorbent assay (Medico-Nobel, Boots-Celltech Ltd., Berkshire, United Kingdom), and PCR showed a sensitivity of 100%, a specificity of 93% when PCR was compared with cell culture, and a corrected specificity of 99% when PCR was compared with cell culture or IDEIA.

  10. Quinolones for the Treatment of Neisseria Gonorrhoeae and Chlamydia Trachomatis

    PubMed Central

    1993-01-01

    The most commonly sexually transmitted bacteria are Neisseria gonorrhoeae and Chlamydia trachomatis. The quinolones ofloxacin and ciprofloxacin have been shown to have activity against both of these bacteria in vitro and in vivo. Ofloxacin is particularly well suited for the treatment of N. gonorrhoeae and C. trachomatis cervical infection, which can be considered the earliest manifestation of pelvic inflammatory disease (PID). Not only can ofloxacin be effectively used as a single agent, it is also useful in treating urinary tract infections caused by Enterobacteriaceae. Although it has moderate activity against anaerobes in general, ofloxacin does have activity against the anaerobes commonly isolated from female patients with soft tissue pelvic infections. Thus, ofloxacin has the potential for being utilized to treat early salpingitis. PMID:18475328

  11. The Relationship between Chlamydia trachomatis Genital Infection and Spontaneous Abortion

    PubMed Central

    Ahmadi, Amjad; Khodabandehloo, Mazaher; Ramazanzadeh, Rashid; Farhadifar, Fariba; Roshani, Daem; Ghaderi, Ebrahim; Farhangi, Niloofar

    2016-01-01

    Background: Chlamydia trachomatis is the etiology of most of sexually transmitted diseases. Colonization of C. trachomatis in the genital tract during early gestation has been associated with preterm birth, and preterm premature rupture of the membranes. The role of C. trachomatis on spontaneous abortion has not yet been proved completely. The aim of this study was to evaluate the frequency of C. trachomatis infection among pregnant women and its association with spontaneous abortion. Methods: This case-control study was conducted from August 2012 until January 2013. Totally, 218 women were included; 109 women with spontaneous abortion with gestation age between 10–20 weeks (cases), and 109 women with normal pregnancy with gestation age between 20–30 weeks (controls) in Sanandaj, Iran. DNA was extracted from endocervical swabs and a PCR test was conducted for detection of C. trachomatis infection in women using specific primers. Independent T-test and Chi-square were used for comparison of quantitative and qualitative variables, respectively, and p<0.05 was considered significant. Results: The total prevalence of C. trachomatis infection was 38(17.43%) in endocervical swabs of women. However, the number of cases with C. trachomatis infections was 25 out of 109(22.9%) in the case group and 13 out of 109(11.9%) in control group, respectively. Association between chlamydia infection and spontaneous abortion was statistically significant (OR=2.198, CI 95%: 1.058–4.56). Conclusion: Our study showed that C. trachomatis infection was associated with spontaneous abortion. Thus, screening and treatment of pregnant women may prevent this adverse pregnancy outcome. PMID:27141466

  12. TLR9 is required for MAPK/NF-κB activation but does not cooperate with TLR2 or TLR6 to induce host resistance to Brucella abortus.

    PubMed

    Gomes, Marco Túlio; Campos, Priscila Carneiro; Pereira, Guilherme de Sousa; Bartholomeu, Daniella Castanheira; Splitter, Gary; Oliveira, Sergio Costa

    2016-05-01

    Brucella abortus is a Gram-negative intracellular bacterial pathogen that causes a zoonosis of worldwide occurrence, leading to undulant fever in humans and abortion in domestic animals. B. abortus is recognized by several pattern-recognition receptors triggering pathways during the host innate immune response. Therefore, here, we determined the cooperative role of TLR9 with TLR2 or TLR6 receptors in sensing Brucella Furthermore, we deciphered the host innate immune response against B. abortus or its DNA, emphasizing the role of TLR9-MAPK/NF-κB signaling pathways in the production of proinflammatory cytokines. TLR9 is required for the initial host control of B. abortus, but this TLR was dispensable after 6 wk of infection. The susceptibility of TLR9(-/-)-infected animals to Brucella paralleled with lower levels of IFN-γ produced by mouse splenocytes stimulated with this pathogen compared with wild-type cells. However, no apparent cooperative interplay was observed between TLR2-TLR9 or TLR6-TLR9 receptors to control infection. Moreover, B. abortus or its DNA induced activation of MAPK/NF-κB pathways and production of IL-12 and TNF-α by macrophages partially dependent on TLR9 but completely dependent on MyD88. In addition, B. abortus-derived CpG oligonucleotides required TLR9 to promote IL-12 and TNF-α production by macrophages. By confocal microscopy, we demonstrated that TLR9 redistributed and colocalized with lysosomal-associated membrane protein-1 upon Brucella infection. Thus, B. abortus induced TLR9 traffic, leading to cell signaling activation and IL-12 and TNF-α production. Although TLR9 recognized Brucella CpG motifs, our results suggest a new pathway of B. abortus DNA-activating macrophages independent of TLR9.

  13. A Homologue of an Operon Required for DNA Transfer in Agrobacterium Is Required in Brucella abortus for Virulence and Intracellular Multiplication

    PubMed Central

    Sieira, Rodrigo; Comerci, Diego J.; Sánchez, Daniel O.; Ugalde, Rodolfo A.

    2000-01-01

    As part of a Brucella abortus 2308 genome project carried out in our laboratory, we identified, cloned, and sequenced a genomic DNA fragment containing a locus (virB) highly homologous to bacterial type IV secretion systems. The B. abortus virB locus is a collinear arrangement of 13 open reading frames (ORFs). Between virB1 and virB2 and downstream of ORF12, two degenerated, palindromic repeat sequences characteristic of Brucella intergenic regions were found. Gene reporter studies demonstrated that the B. abortus virB locus constitutes an operon transcribed from virB1 which is turned on during the stationary phase of growth. A B. abortus polar virB1 mutant failed to replicate in HeLa cells, indicating that the virB operon plays a critical role in intracellular multiplication. Mutants with polar and nonpolar mutations introduced in virB10 showed different behaviors in mice and in the HeLa cell infection assay, suggesting that virB10 per se is necessary for the correct function of this type IV secretion apparatus. Mouse infection assays demonstrated that the virB operon constitutes a major determinant of B. abortus virulence. It is suggested that putative effector molecules secreted by this type IV secretion system determine routing of B. abortus to an endoplasmic reticulum-related replication compartment. PMID:10940027

  14. The effects of red ginseng saponin fraction-A (RGSF-A) on phagocytosis and intracellular signaling in Brucella abortus infected RAW 264.7 cells.

    PubMed

    Arayan, Lauren Togonon; Simborio, Hannah Leah; Reyes, Alisha Wehdnesday Bernardo; Hop, Huynh Tan; Min, WonGi; Lee, Hu Jang; Rhee, Man Hee; Chang, Hong Hee; Kim, Suk

    2015-06-01

    This study indicated that RGSF-A caused a marked reduction in the adherence, internalization and intracellular growth of Brucella abortus in RGSF-A-treated cells. Furthermore, a decline in the intensity of F-actin fluorescence was observed in RGSF-A-treated cells compared with untreated B. abortus-infected cells. In addition, an evaluation of phagocytic signaling proteins by Western blot analysis revealed an apparent reduction of ERK and p38α phosphorylation levels in B. abortus-infected RGSF-A-treated cells compared with the control. Upon intracellular trafficking of the pathogen, a higher number of B. abortus-containing phagosomes colocalized with LAMP-1 in RGSF-A-treated cells compared with control cells. These results strongly suggest that inhibition of B. abortus uptake could be mediated by suppression in the activation of MAPKs signaling proteins phospho-ERK 1/2, and p38 levels. On the other hand, inhibition of intracellular replication results from the enhancement of phagolysosome fusion in host macrophages. This study highlights the phagocytic and intracellular modulating effect of RGSF-A and its potential as an alternative remedy to control B. abortus infection.

  15. Brucella abortus S19 and RB51 vaccine immunogenicity test: Evaluation of three mice (BALB/c, Swiss and CD-1) and two challenge strains (544 and 2308).

    PubMed

    Miranda, Karina Leite; Dorneles, Elaine Maria Seles; Pauletti, Rebeca Barbosa; Poester, Fernando Padilla; Lage, Andrey Pereira

    2015-01-15

    The aim of the present study was to evaluate the use of different mouse strains (BALB/c, Swiss and CD-1) and different challenge strains (Brucella abortus 544 and 2308) in the study of B. abortus vaccine (S19 and RB51) immunogenicity test in the murine model. No significant difference in B. abortus vaccine potency assay was found with the use of B. abortus 544 or B. abortus 2308 as challenge strain. Results of variance analysis showed an interaction between treatment and mouse strain; therefore these parameters could not be compared separately. When CD-1 groups were compared, those vaccinated showed significantly lower counts than non-vaccinated ones (P<0.05), independently of the vaccine received (S19 or RB51). Similar results were observed on BALB/c groups. However, in Swiss mouse groups, S19 was more protective than RB51 (P<0.05), which showed protection when compared to the non-vaccinated group (P<0.05). In summary, data from the present study showed that CD-1, BALB/c and Swiss mice strains, as well as both challenge strains, B. abortus strains 544 and 2308, can be used in immunogenicity tests of S19 and RB51 vaccines.

  16. Adrenal steroids modulate the immune response during Brucella abortus infection by a mechanism that depends on the regulation of cytokine production.

    PubMed

    Gentilini, María Virginia; Velásquez, Lis Noelia; Barrionuevo, Paula; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2015-05-01

    Human brucellosis is a protean disease with a diversity of clinical signs and symptoms resulting from infection with Brucella species. Recent reports suggest a cross-regulation between adrenal steroids (cortisol and dehydroepiandrosterone [DHEA]) and the immune system. Monocytes and macrophages are the main replication niche for Brucella. Therefore, we investigated the role of adrenal hormones on the modulation of the immune response mediated by macrophages in B. abortus infection. Cortisol treatment during B. abortus infection significantly inhibits cytokine, chemokine, and MMP-9 secretion. In contrast, DHEA treatment had no effect. However, DHEA treatment increases the expression of costimulatory molecules (CD40, CD86), the adhesion molecule CD54, and major histocompatibility complex class I (MHC-I) and MHC-II expression on the surface of B. abortus-infected monocytes. It is known that B. abortus infection inhibits MHC-I and MHC-II expression induced by gamma interferon (IFN-γ) treatment. DHEA reverses B. abortus downmodulation of the MHC-I and -II expression induced by IFN-γ. Taken together, our data indicate that DHEA immune intervention may positively affect monocyte activity during B. abortus infection.

  17. Pheno- and genotyping of Brucella abortus biovar 5 isolated from a water buffalo (Bubalus bubalis) fetus: First case reported in the Americas.

    PubMed

    Martínez, Diana; Thompson, Carolina; Draghi, Graciela; Canavesio, Vilma; Jacobo, Roberto; Zimmer, Patricia; Elena, Sebastián; Nicola, Ana M; de Echaide, Susana Torioni

    2014-09-17

    An isolate of Brucella spp. from an aborted water buffalo (Bubalus bubalis) fetus was characterized based on its pheno- and genotype. The phenotype was defined by carbon dioxide requirement, hydrogen sulfide production, sensitivity to thionin and basic fuchsin and agglutination with Brucella A and M monospecific antisera. The genotype was based on the amplification of the following genes: bcsp31, omp2ab, and eri and the species-specific localization of the insertion sequence IS711 in the Brucella chromosome via B. abortus-B. melitensis-B. ovis-B. suis (AMOS)-PCR. Unexpectedly, the isolate showed a phenotype different from B. abortus bv 1, the most prevalent strain in cattle in Argentina, and from vaccine strain 19, currently used in bovines and water buffaloes. Genotyping supported the phenotypic results, as the analysis of the omp2ab gene sequence showed an identical pattern to either B. abortus bv 5 or B. melitensis. Finally, the AMOS PCR generated a 1700-bp fragment from the isolate, different than those amplified from B. abortus bv 1 (498bp) and B. melitensis (731bp), confirming the presence of B. abortus bv 5. The OIE/FAO Reference Laboratory for Brucellosis confirmed this typing. This is the first report of B. abortus bv 5 from a water buffalo in the Americas.

  18. Adrenal Steroids Modulate the Immune Response during Brucella abortus Infection by a Mechanism That Depends on the Regulation of Cytokine Production

    PubMed Central

    Gentilini, María Virginia; Velásquez, Lis Noelia; Barrionuevo, Paula; Arriola Benitez, Paula Constanza; Giambartolomei, Guillermo Hernán

    2015-01-01

    Human brucellosis is a protean disease with a diversity of clinical signs and symptoms resulting from infection with Brucella species. Recent reports suggest a cross-regulation between adrenal steroids (cortisol and dehydroepiandrosterone [DHEA]) and the immune system. Monocytes and macrophages are the main replication niche for Brucella. Therefore, we investigated the role of adrenal hormones on the modulation of the immune response mediated by macrophages in B. abortus infection. Cortisol treatment during B. abortus infection significantly inhibits cytokine, chemokine, and MMP-9 secretion. In contrast, DHEA treatment had no effect. However, DHEA treatment increases the expression of costimulatory molecules (CD40, CD86), the adhesion molecule CD54, and major histocompatibility complex class I (MHC-I) and MHC-II expression on the surface of B. abortus-infected monocytes. It is known that B. abortus infection inhibits MHC-I and MHC-II expression induced by gamma interferon (IFN-γ) treatment. DHEA reverses B. abortus downmodulation of the MHC-I and -II expression induced by IFN-γ. Taken together, our data indicate that DHEA immune intervention may positively affect monocyte activity during B. abortus infection. PMID:25733519

  19. Comparative study on responses of cattle and water buffalo (Bubalus bubalis) to experimental inoculation of Brucella abortus biovar 1 by the intraconjunctival route--a preliminary report.

    PubMed

    Adesiyun, Abiodun A; Fosgate, Geoff T; Persad, Anil; Campbell, Mervyn; Seebaransingh, Ravi; Stewart-Johnson, Alva

    2010-12-01

    The preliminary study was conducted to assess the virulence of a strain of Brucella abortus (1969D) and to compare the susceptibility of water buffalo and cattle calves to infection by the intraconjunctival route. Seven of each cattle and water buffalo (Bubalus bubalis) calves aged 3-6 months were inoculated intraconjunctivally with counts ranging from 1.5 × 10(7) to 1.7 × 10(10) colony forming units of B. abortus. Animals were monitored over an 8-week period for clinical manifestations and serological and hematological evidence of infection. At slaughter, eight lymph nodes from each animal were sampled for bacteriological and histopathological assessments. Lymph nodes from three water buffalo (43%) and five cattle (71%) yielded B. abortus (P=0.048). Parotid/prescapular lymph nodes were most sensitive in detecting B. abortus. Our data suggest that B. abortus strain 1969D may be used as challenge strain, and water buffalo appeared to have a lower susceptibility to B. abortus infection than cattle.

  20. Brucella abortus Invasion of Osteocytes Modulates Connexin 43 and Integrin Expression and Induces Osteoclastogenesis via Receptor Activator of NF-κB Ligand and Tumor Necrosis Factor Alpha Secretion.

    PubMed

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Gentilini, María Virginia; Velásquez, Lis Noelia; Fossati, Carlos Alberto; Giambartolomei, Guillermo Hernán; Delpino, María Victoria

    2015-10-12

    Osteoarticular brucellosis is the most common localization of human active disease. Osteocytes are the most abundant cells of bone. They secrete factors that regulate the differentiation of both osteoblasts and osteoclasts during bone remodeling. The aim of this study is to determine if Brucella abortus infection modifies osteocyte function. Our results indicate that B. abortus infection induced matrix metalloproteinase 2 (MMP-2), receptor activator for NF-κB ligand (RANKL), proinflammatory cytokines, and keratinocyte chemoattractant (KC) secretion by osteocytes. In addition, supernatants from B. abortus-infected osteocytes induced bone marrow-derived monocytes (BMM) to undergo osteoclastogenesis. Using neutralizing antibodies against tumor necrosis factor alpha (TNF-α) or osteoprotegerin (OPG), RANKL's decoy receptor, we determined that TNF-α and RANKL are involved in osteoclastogenesis induced by supernatants from B. abortus-infected osteocytes. Connexin 43 (Cx43) and the integrins E11/gp38, integrin-α, integrin-β, and CD44 are involved in cell-cell interactions necessary for osteocyte survival. B. abortus infection inhibited the expression of Cx43 but did not modify the expression of integrins. Yet the expression of both Cx43 and integrins was inhibited by supernatants from B. abortus-infected macrophages. B. abortus infection was not capable of inducing osteocyte apoptosis. However, supernatants from B. abortus-infected macrophages induced osteocyte apoptosis in a dose-dependent manner. Taken together, our results indicate that B. abortus infection could alter osteocyte function, contributing to bone damage.

  1. Brucella abortus Invasion of Osteocytes Modulates Connexin 43 and Integrin Expression and Induces Osteoclastogenesis via Receptor Activator of NF-κB Ligand and Tumor Necrosis Factor Alpha Secretion

    PubMed Central

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Gentilini, María Virginia; Velásquez, Lis Noelia; Fossati, Carlos Alberto; Giambartolomei, Guillermo Hernán

    2015-01-01

    Osteoarticular brucellosis is the most common localization of human active disease. Osteocytes are the most abundant cells of bone. They secrete factors that regulate the differentiation of both osteoblasts and osteoclasts during bone remodeling. The aim of this study is to determine if Brucella abortus infection modifies osteocyte function. Our results indicate that B. abortus infection induced matrix metalloproteinase 2 (MMP-2), receptor activator for NF-κB ligand (RANKL), proinflammatory cytokines, and keratinocyte chemoattractant (KC) secretion by osteocytes. In addition, supernatants from B. abortus-infected osteocytes induced bone marrow-derived monocytes (BMM) to undergo osteoclastogenesis. Using neutralizing antibodies against tumor necrosis factor alpha (TNF-α) or osteoprotegerin (OPG), RANKL's decoy receptor, we determined that TNF-α and RANKL are involved in osteoclastogenesis induced by supernatants from B. abortus-infected osteocytes. Connexin 43 (Cx43) and the integrins E11/gp38, integrin-α, integrin-β, and CD44 are involved in cell-cell interactions necessary for osteocyte survival. B. abortus infection inhibited the expression of Cx43 but did not modify the expression of integrins. Yet the expression of both Cx43 and integrins was inhibited by supernatants from B. abortus-infected macrophages. B. abortus infection was not capable of inducing osteocyte apoptosis. However, supernatants from B. abortus-infected macrophages induced osteocyte apoptosis in a dose-dependent manner. Taken together, our results indicate that B. abortus infection could alter osteocyte function, contributing to bone damage. PMID:26459511

  2. Variable laser attenuator

    DOEpatents

    Foltyn, S.R.

    1987-05-29

    The disclosure relates to low loss, high power variable attenuators comprising one or more transmissive and/or reflective multilayer dielectric filters. The attenuator is particularly suitable to use with unpolarized lasers such as excimer lasers. Beam attenuation is a function of beam polarization and the angle of incidence between the beam and the filter and is controlled by adjusting the angle of incidence the beam makes to the filter or filters. Filters are selected in accordance with beam wavelength. 9 figs.

  3. Variable laser attenuator

    DOEpatents

    Foltyn, Stephen R.

    1988-01-01

    The disclosure relates to low loss, high power variable attenuators comprng one or more transmissive and/or reflective multilayer dielectric filters. The attenuator is particularly suitable to use with unpolarized lasers such as excimer lasers. Beam attenuation is a function of beam polarization and the angle of incidence between the beam and the filter and is controlled by adjusting the angle of incidence the beam makes to the filter or filters. Filters are selected in accordance with beam wavelength.

  4. Brucella abortus as a potential vaccine candidate: induction of interleukin-12 secretion and enhanced B7.1 and B7.2 and intercellular adhesion molecule 1 surface expression in elutriated human monocytes stimulated by heat-inactivated B. abortus.

    PubMed Central

    Zaitseva, M; Golding, H; Manischewitz, J; Webb, D; Golding, B

    1996-01-01

    Development of a vaccine which is capable of generating a strong cellular immune response associated with gamma interferon (IFN-gamma) production and cytotoxic T-cell development requires that the immunogen be capable of inducing the secretion of interleukin-12 (IL-12), which is a pivotal factor for the differentiation of Th1 or Tc1 cells. We have previously shown that the heat-inactivated gram-negative bacterium Brucella abortus can induce IFN-gamma secretion by T cells. In the present study, we demonstrate that B. abortus and lipopolysaccharide (LPS) from B. abortus can induce IL-12 p40 mRNA expression and protein secretion by human elutriated monocytes (99% pure). p40 mRNA was detected within 4 h, and p40 protein could be measured at 24 h. This induction was abrogated by anti-CD14 monoclonal antibody, suggesting that monocytes recognize B. abortus via their receptor for LPS. The biological activity of IL-12 secreted by B. abortus-stimulated monocytes was demonstrated by its ability to upregulate IFN-gamma mRNA expression in T cells separated from monocytes and B. abortus by a transwell membrane. The B. abortus-induced IL-12 also enhanced NK cytolytic activity against K562 target cells. B. abortus was shown to rapidly increase the expression of the costimulatory molecules B7.1 and B7.2 and intercellular adhesion molecule 1 on human monocytes. Together, these data indicate that B. abortus can directly activate human monocytes and provide the cytokine milieu which would direct the immune response towards Th1-Tc1 differentiation. PMID:8757841

  5. Extragenital Infections Caused by Chlamydia trachomatis and Neisseria gonorrhoeae: A Review of the Literature.

    PubMed

    Chan, Philip A; Robinette, Ashley; Montgomery, Madeline; Almonte, Alexi; Cu-Uvin, Susan; Lonks, John R; Chapin, Kimberle C; Kojic, Erna M; Hardy, Erica J

    2016-01-01

    In the United States, sexually transmitted diseases due to Chlamydia trachomatis and Neisseria gonorrhoeae continue to be a major public health burden. Screening of extragenital sites including the oropharynx and rectum is an emerging practice based on recent studies highlighting the prevalence of infection at these sites. We reviewed studies reporting the prevalence of extragenital infections in women, men who have sex with men (MSM), and men who have sex only with women (MSW), including distribution by anatomical site. Among women, prevalence was found to be 0.6-35.8% for rectal gonorrhea (median reported prevalence 1.9%), 0-29.6% for pharyngeal gonorrhea (median 2.1%), 2.0-77.3% for rectal chlamydia (median 8.7%), and 0.2-3.2% for pharyngeal chlamydia (median 1.7%). Among MSM, prevalence was found to be 0.2-24.0% for rectal gonorrhea (median 5.9%), 0.5-16.5% for pharyngeal gonorrhea (median 4.6%), 2.1-23.0% for rectal chlamydia (median 8.9%), and 0-3.6% for pharyngeal chlamydia (median 1.7%). Among MSW, the prevalence was found to be 0-5.7% for rectal gonorrhea (median 3.4%), 0.4-15.5% for pharyngeal gonorrhea (median 2.2%), 0-11.8% for rectal chlamydia (median 7.7%), and 0-22.0% for pharyngeal chlamydia (median 1.6%). Extragenital infections are often asymptomatic and found in the absence of reported risk behaviors, such as receptive anal and oral intercourse. We discuss current clinical recommendations and future directions for research. PMID:27366021

  6. Defining Species-Specific Immunodominant B Cell Epitopes for Molecular Serology of Chlamydia Species

    PubMed Central

    Rahman, K. Shamsur; Chowdhury, Erfan U.; Poudel, Anil; Ruettger, Anke; Sachse, Konrad

    2015-01-01

    Urgently needed species-specific enzyme-linked immunosorbent assays (ELISAs) for the detection of antibodies against Chlamydia spp. have been elusive due to high cross-reactivity of chlamydial antigens. To identify Chlamydia species-specific B cell epitopes for such assays, we ranked the potential epitopes of immunodominant chlamydial proteins that are polymorphic among all Chlamydia species. High-scoring peptides were synthesized with N-terminal biotin, followed by a serine-glycine-serine-glycine spacer, immobilized onto streptavidin-coated microtiter plates, and tested with mono-specific mouse hyperimmune sera against each Chlamydia species in chemiluminescent ELISAs. For each of nine Chlamydia species, three to nine dominant polymorphic B cell epitope regions were identified on OmpA, CT618, PmpD, IncA, CT529, CT442, IncG, Omp2, TarP, and IncE proteins. Peptides corresponding to 16- to 40-amino-acid species-specific sequences of these epitopes reacted highly and with absolute specificity with homologous, but not heterologous, Chlamydia monospecies-specific sera. Host-independent reactivity of such epitopes was confirmed by testing of six C. pecorum-specific peptides from five proteins with C. pecorum-reactive sera from cattle, the natural host of C. pecorum. The probability of cross-reactivity of peptide antigens from closely related chlamydial species or strains correlated with percent sequence identity and declined to zero at <50% sequence identity. Thus, phylograms of B cell epitope regions predict the specificity of peptide antigens for rational use in the genus-, species-, or serovar-specific molecular serology of Chlamydia spp. We anticipate that these peptide antigens will improve chlamydial serology by providing easily accessible assays to nonspecialist laboratories. Our approach also lends itself to the identification of relevant epitopes of other microbial pathogens. PMID:25761461

  7. Extragenital Infections Caused by Chlamydia trachomatis and Neisseria gonorrhoeae: A Review of the Literature

    PubMed Central

    Chan, Philip A.; Montgomery, Madeline; Almonte, Alexi; Lonks, John R.; Chapin, Kimberle C.; Kojic, Erna M.; Hardy, Erica J.

    2016-01-01

    In the United States, sexually transmitted diseases due to Chlamydia trachomatis and Neisseria gonorrhoeae continue to be a major public health burden. Screening of extragenital sites including the oropharynx and rectum is an emerging practice based on recent studies highlighting the prevalence of infection at these sites. We reviewed studies reporting the prevalence of extragenital infections in women, men who have sex with men (MSM), and men who have sex only with women (MSW), including distribution by anatomical site. Among women, prevalence was found to be 0.6–35.8% for rectal gonorrhea (median reported prevalence 1.9%), 0–29.6% for pharyngeal gonorrhea (median 2.1%), 2.0–77.3% for rectal chlamydia (median 8.7%), and 0.2–3.2% for pharyngeal chlamydia (median 1.7%). Among MSM, prevalence was found to be 0.2–24.0% for rectal gonorrhea (median 5.9%), 0.5–16.5% for pharyngeal gonorrhea (median 4.6%), 2.1–23.0% for rectal chlamydia (median 8.9%), and 0–3.6% for pharyngeal chlamydia (median 1.7%). Among MSW, the prevalence was found to be 0–5.7% for rectal gonorrhea (median 3.4%), 0.4–15.5% for pharyngeal gonorrhea (median 2.2%), 0–11.8% for rectal chlamydia (median 7.7%), and 0–22.0% for pharyngeal chlamydia (median 1.6%). Extragenital infections are often asymptomatic and found in the absence of reported risk behaviors, such as receptive anal and oral intercourse. We discuss current clinical recommendations and future directions for research. PMID:27366021

  8. Defining species-specific immunodominant B cell epitopes for molecular serology of Chlamydia species.

    PubMed

    Rahman, K Shamsur; Chowdhury, Erfan U; Poudel, Anil; Ruettger, Anke; Sachse, Konrad; Kaltenboeck, Bernhard

    2015-05-01

    Urgently needed species-specific enzyme-linked immunosorbent assays (ELISAs) for the detection of antibodies against Chlamydia spp. have been elusive due to high cross-reactivity of chlamydial antigens. To identify Chlamydia species-specific B cell epitopes for such assays, we ranked the potential epitopes of immunodominant chlamydial proteins that are polymorphic among all Chlamydia species. High-scoring peptides were synthesized with N-terminal biotin, followed by a serine-glycine-serine-glycine spacer, immobilized onto streptavidin-coated microtiter plates, and tested with mono-specific mouse hyperimmune sera against each Chlamydia species in chemiluminescent ELISAs. For each of nine Chlamydia species, three to nine dominant polymorphic B cell epitope regions were identified on OmpA, CT618, PmpD, IncA, CT529, CT442, IncG, Omp2, TarP, and IncE proteins. Peptides corresponding to 16- to 40-amino-acid species-specific sequences of these epitopes reacted highly and with absolute specificity with homologous, but not heterologous, Chlamydia monospecies-specific sera. Host-independent reactivity of such epitopes was confirmed by testing of six C. pecorum-specific peptides from five proteins with C. pecorum-reactive sera from cattle, the natural host of C. pecorum. The probability of cross-reactivity of peptide antigens from closely related chlamydial species or strains correlated with percent sequence identity and declined to zero at <50% sequence identity. Thus, phylograms of B cell epitope regions predict the specificity of peptide antigens for rational use in the genus-, species-, or serovar-specific molecular serology of Chlamydia spp. We anticipate that these peptide antigens will improve chlamydial serology by providing easily accessible assays to nonspecialist laboratories. Our approach also lends itself to the identification of relevant epitopes of other microbial pathogens.

  9. Disentangling screening and diagnostic Chlamydia test positivity among females testing at title x-funded and adolescent health clinics, san francisco 2009.

    PubMed

    Stephens, Sally C; Snell, Ameera; Liska, Sally; Rauch, Leah; Philip, Susan S; Bernstein, Kyle T

    2011-07-01

    By using a reason-for-test code, we compared positivity for female chlamydia and gonorrhea. At family planning clinics, there were no statistically significant differences in screening versus diagnostic positivity for either chlamydia or gonorrhea among women. However, at adolescent health clinics, diagnostic positivity was higher than screening positivity for chlamydia and gonorrhea.

  10. Twenty years of research into Chlamydia-like organisms: a revolution in our understanding of the biology and pathogenicity of members of the phylum Chlamydiae.

    PubMed

    Taylor-Brown, Alyce; Vaughan, Lloyd; Greub, Gilbert; Timms, Peter; Polkinghorne, Adam

    2015-02-01

    Chlamydiae are obligate intracellular bacteria that share a unique but remarkably conserved biphasic developmental cycle that relies on a eukaryotic host cell for survival. Although the phylum was originally thought to only contain one family, the Chlamydiaceae, a total of nine families are now recognized. These so-called Chlamydia-like organisms (CLOs) are also referred to as 'environmental chlamydiae', as many were initially isolated from environmental sources. However, these organisms are also emerging pathogens, as many, such as Parachlamydia sp., Simkania sp. and Waddlia sp., have been associated with human disease, and others, such as Piscichlamydia sp. and Parilichlamydia sp., have been documented in association with diseases in animals. Their strict intracellular nature and the requirement for cell culture have been a confounding factor in characterizing the biology and pathogenicity of CLOs. Nevertheless, the genomes of seven CLO species have now been sequenced, providing new information on their potential ability to adapt to a wide range of hosts. As new isolation and diagnostic methods advance, we are able to further explore the richness of this phylum with further research likely to help define the true pathogenic potential of the CLOs while also providing insight into the origins of the 'traditional' chlamydiae.

  11. The chlamydia knowledge, awareness and testing practices of Australian general practitioners and practice nurses: survey findings from the Australian Chlamydia Control Effectiveness Pilot (ACCEPt)

    PubMed Central

    2013-01-01

    Background ACCEPt, a large cluster randomized control trial, aims to determine if annual testing for 16 to 29 year olds in general practice can reduce chlamydia prevalence. ACCEPt is the first trial investigating the potential role of practice nurses (PN) in chlamydia testing. To inform the design of the ACCEPt intervention, we aimed to determine the chlamydia knowledge, attitudes, and testing practices of participating general practitioners (GPs) and PNs. Methods GPs and PNs from 143 clinics recruited from 52 areas in 4 Australian states were asked to complete a survey at time of recruitment. Responses of PNs and GPs were compared using conditional logistic regression to account for possible intra cluster correlation within clinics. Results Of the PNs and GPs enrolled in ACCEPt, 81% and 72% completed the questionnaire respectively. Less than a third of PNs (23%) and GPs (32%) correctly identified the two age groups with highest infection rates in women and only 16% vs 17% the correct age groups in men. More PNs than GPs would offer testing opportunistically to asymptomatic patients aged ≤25 years; women having a pap smear (84% vs 55%, P<0.01); antenatal checkup (83% vs 44%, P<0.01) and Aboriginal men with a sore throat (79% vs 33%, P<0.01), but also to patients outside of the guideline age group at the time of the survey; 26 year old males presenting for a medical check (78% vs 30%, P = <0.01) and 33 year old females presenting for a pill prescription (83% vs 55%, P<0.01). More PNs than GPs knew that retesting was recommended after chlamydia treatment (93% vs 87%, P=0.027); and the recommended timeframe was 3 months (66% vs 26%, P<0.01). A high proportion of PNs (90%) agreed that they could conduct chlamydia testing in general practice, with 79% wanting greater involvement and 89% further training. Conclusions Our survey reveals gaps in chlamydia knowledge and management among GPs and PNs that may be contributing to low testing rates in general practice. The

  12. Protective immune-response of aluminium hydroxide gel adjuvanted phage lysate of Brucella abortus S19 in mice against direct virulent challenge with B. abortus 544.

    PubMed

    Jain, Lata; Rawat, Mayank; Prajapati, Awadhesh; Tiwari, Ashok Kumar; Kumar, Bablu; Chaturvedi, V K; Saxena, H M; Ramakrishnan, Sarvanan; Kumar, Jatin; Kerketta, Priscilla

    2015-09-01

    The prophylactic efficacies of plain and alum adsorbed lysate were evaluated by direct virulent challenge in mice model. A recently isolated brucellaphage 'ϕLd' was used for generation of lysates. Twenty four h incubated Brucella abortus S19 broth cultures standardized to contain approximately 10(8) CFU/ml were found suitable for generation of lysates. Three lysate batches produced through separate cycles did not show any significant variation with respect to protein and polysaccharide contents, endotoxin level and phage counts, indicating that compositionally stable lysate preparations can be generated through an optimized production process. Three polypeptides of ∼16, 19 and 23 kDa could be identified as immuno-dominant antigens of the lysate which induced both humoral and cell-mediated immune responses in a dose dependent manner. Results of efficacy evaluation trial confirmed dose-dependent protective potencies of lysate preparation. The lysate with an antigenic dose of 0.52 μg protein and 60 μg CHO adsorbed on aluminium gel (0.1 percent aluminium concentration) exhibited the highest protective potency which was greater than that induced by standard S19 vaccine. Phage lysate methodology provides a very viable option through which an improved immunizing preparation with all desirable traits can be developed against brucellosis, and integrated with immunization programmes in a more efficient manner.

  13. Incident and recurrent Chlamydia trachomatis and Neisseria gonorrhoeae infections, active component, U.S. Armed Forces, 2010-2014.

    PubMed

    Owings, Alfred J; Clark, Leslie L; Rohrbeck, Patricia

    2016-02-01

    Chlamydia trachomatis and Neisseria gonorrhoeae infections impose a significant clinical and public health burden on the Military Health System. Repeat infections contribute significantly to that burden. This report summarizes rates and relative risks of true incident (i.e., initial or "first time ever") and recurrent (i.e., repeat) chlamydia and gonorrhea infections among active component members between 1 January 2010 and 31 December 2014. During the surveillance period, a total of 66,396 initial chlamydia and 9,138 initial gonorrhea cases were diagnosed. Annual crude rates of initial chlamydia infections increased by 23%. Crude rates of initial gonorrhea infections remained stable overall, but female rates decreased by 28.3% over the period. Among the incident cohorts, 11,699 cases of repeat chlamydia, and 1,138 cases of repeat gonorrhea were diagnosed over the period, accounting for 15.0% and 11.1% of overall cohort chlamydia and gonorrhea infections, respectively. The Army branch, those aged 17-19 years, females, non-Hispanic black service members, junior enlisted ranks, and single/never-married service members had the highest crude rates of initial chlamydia and gonorrhea infection, and (single/never-married service members excepted) highest adjusted relative risk of repeat chlamydia infection. PMID:26930148

  14. Chlamydia Induces Anchorage Independence in 3T3 Cells and Detrimental Cytological Defects in an Infection Model

    PubMed Central

    Knowlton, Andrea E.; Fowler, Larry J.; Patel, Rahul K.; Wallet, Shannon M.; Grieshaber, Scott S.

    2013-01-01

    Chlamydia are Gram negative, obligate intracellular bacterial organisms with different species causing a multitude of infections in both humans and animals. Chlamydia trachomatis is the causative agent of the sexually transmitted infection (STI) Chlamydia, the most commonly acquired bacterial STI in the United States. Chlamydial infections have also been epidemiologically linked to cervical cancer in women co-infected with the human papillomavirus (HPV). We have previously shown chlamydial infection results in centrosome amplification and multipolar spindle formation leading to chromosomal instability. Many studies indicate that centrosome abnormalities, spindle defects, and chromosome segregation errors can lead to cell transformation. We hypothesize that the presence of these defects within infected dividing cells identifies a possible mechanism for Chlamydia as a cofactor in cervical cancer formation. Here we demonstrate that infection with Chlamydia trachomatis is able to transform 3T3 cells in soft agar resulting in anchorage independence and increased colony formation. Additionally, we show for the first time Chlamydia infects actively replicating cells in vivo. Infection of mice with Chlamydia results in significantly increased cell proliferation within the cervix, and in evidence of cervical dysplasia. Confocal examination of these infected tissues also revealed elements of chlamydial induced chromosome instability. These results contribute to a growing body of data implicating a role for Chlamydia in cervical cancer development and suggest a possible molecular mechanism for this effect. PMID:23308295

  15. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection.

    PubMed

    Hielpos, M Soledad; Ferrero, Mariana C; Fernández, Andrea G; Bonetto, Josefina; Giambartolomei, Guillermo H; Fossati, Carlos A; Baldi, Pablo C

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site.

  16. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection.

    PubMed

    Hielpos, M Soledad; Ferrero, Mariana C; Fernández, Andrea G; Bonetto, Josefina; Giambartolomei, Guillermo H; Fossati, Carlos A; Baldi, Pablo C

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site. PMID:26448160

  17. CCL20 and Beta-Defensin 2 Production by Human Lung Epithelial Cells and Macrophages in Response to Brucella abortus Infection

    PubMed Central

    Fernández, Andrea G.; Bonetto, Josefina; Giambartolomei, Guillermo H.; Fossati, Carlos A.; Baldi, Pablo C.

    2015-01-01

    Both CCL20 and human β-defensin 2 (hBD2) interact with the same membrane receptor and display chemotactic and antimicrobial activities. They are produced by airway epithelia in response to infectious agents and proinflammatory cytokines. Whereas Brucella spp. can infect humans through inhalation, their ability to induce CCL20 and hBD2 in lung cells is unknown. Here we show that B. abortus induces CCL20 expression in human alveolar (A549) or bronchial (Calu-6) epithelial cell lines, primary alveolar epithelial cells, primary human monocytes, monocyte-derived macrophages and the monocytic cell line THP-1. CCL20 expression was mainly mediated by JNK1/2 and NF-kB in both Calu-6 and THP-1 cells. CCL20 secretion was markedly induced in A549, Calu-6 and THP-1 cells by heat-killed B. abortus or a model Brucella lipoprotein (L-Omp19) but not by the B. abortus lipopolysaccharide (LPS). Accordingly, CCL20 production by B. abortus-infected cells was strongly TLR2-dependent. Whereas hBD2 expression was not induced by B. abortus infection, it was significantly induced in A549 cells by conditioned media from B. abortus-infected THP-1 monocytes (CMB). A similar inducing effect was observed on CCL20 secretion. Experiments using blocking agents revealed that IL-1β, but not TNF-α, was involved in the induction of hBD2 and CCL20 secretion by CMB. In the in vitro antimicrobial assay, the lethal dose (LD) 50 of CCL20 for B. abortus (>50 μg/ml) was markedly higher than that against E. coli (1.5 μg/ml) or a B. abortus mutant lacking the O polysaccharide in its LPS (8.7 ug/ml). hBD2 did not kill any of the B. abortus strains at the tested concentrations. These results show that human lung epithelial cells secrete CCL20 and hBD2 in response to B. abortus and/or to cytokines produced by infected monocytes. Whereas these molecules do not seem to exert antimicrobial activity against this pathogen, they could recruit immune cells to the infection site. PMID:26448160

  18. Epidemiology of Chlamydophila caviae-like Chlamydia isolated from urethra and uterine cervix.

    PubMed

    Murao, Wataru; Wada, Koichiro; Matsumoto, Akira; Fujiwara, Michihisa; Fukushi, Hideto; Kishimoto, Toshio; Monden, Koichi; Kariyama, Reiko; Kumon, Hiromi

    2010-02-01

    In 2000, chlamydial strains OK133 and OK135 were isolated from 2 female patients with cervicitis. These strains were unresponsive to commercially available PCR and LCR test kits for the diagnosis of Chlamydia trachomatis infection, and their phenotypic characteristics were very similar. The OK135 nucleotide sequence in MOMP-VD2 gene closely resembled that of Chlamydophila caviae GPIC. A similar strain was isolated in 2003 from a male patient OKM2 with urethritis, from which the strain SC10-6 was cloned by the plaque purification method. The nucleotide sequence of the entire MOMP gene of SC10-6 was exactly the same as that of OK135. Thus, the strains OK135 and SC10-6, together with OK133, have been called C. caviae-like Chlamydia. We designed primers for nested PCR assay, the product of which showed a single-band 311-bp fragment, to detect C. caviae-like Chlamydia. Of swab specimens obtained from 202 patients from 2003 to 2006 (119 male and 83 female patients), 18 specimens (8.9%) from 14 male and 4 female patients were positive, suggesting that C. caviae-like Chlamydia infection is rather common. Thus far, it has not been determined whether C. caviae-like Chlamydia is pathogenic for humans.

  19. A Coming of Age Story: Chlamydia in the Post-Genetic Era

    PubMed Central

    Hooppaw, Anna J.

    2015-01-01

    Chlamydia spp. are ubiquitous, obligate, intracellular Gram-negative bacterial pathogens that undergo a unique biphasic developmental cycle transitioning between the infectious, extracellular elementary body and the replicative, intracellular reticulate body. The primary Chlamydia species associated with human disease are C. trachomatis, which is the leading cause of both reportable bacterial sexually transmitted infections and preventable blindness, and C. pneumoniae, which infects the respiratory tract and is associated with cardiovascular disease. Collectively, these pathogens are a significant source of morbidity and pose a substantial financial burden on the global economy. Past efforts to elucidate virulence mechanisms of these unique and important pathogens were largely hindered by an absence of genetic methods. Watershed studies in 2011 and 2012 demonstrated that forward and reverse genetic approaches were feasible with Chlamydia and that shuttle vectors could be selected and maintained within the bacterium. While these breakthroughs have led to a steady expansion of the chlamydial genetic tool kit, there are still roads left to be traveled. This minireview provides a synopsis of the currently available genetic methods for Chlamydia along with a comparison to the methods used in other obligate intracellular bacteria. Limitations and advantages of these techniques will be discussed with an eye toward the methods still needed, and how the current state of the art for genetics in obligate intracellular bacteria could direct future technological advances for Chlamydia. PMID:26667838

  20. A Coming of Age Story: Chlamydia in the Post-Genetic Era.

    PubMed

    Hooppaw, Anna J; Fisher, Derek J

    2015-12-14

    Chlamydia spp. are ubiquitous, obligate, intracellular Gram-negative bacterial pathogens that undergo a unique biphasic developmental cycle transitioning between the infectious, extracellular elementary body and the replicative, intracellular reticulate body. The primary Chlamydia species associated with human disease are C. trachomatis, which is the leading cause of both reportable bacterial sexually transmitted infections and preventable blindness, and C. pneumoniae, which infects the respiratory tract and is associated with cardiovascular disease. Collectively, these pathogens are a significant source of morbidity and pose a substantial financial burden on the global economy. Past efforts to elucidate virulence mechanisms of these unique and important pathogens were largely hindered by an absence of genetic methods. Watershed studies in 2011 and 2012 demonstrated that forward and reverse genetic approaches were feasible with Chlamydia and that shuttle vectors could be selected and maintained within the bacterium. While these breakthroughs have led to a steady expansion of the chlamydial genetic tool kit, there are still roads left to be traveled. This minireview provides a synopsis of the currently available genetic methods for Chlamydia along with a comparison to the methods used in other obligate intracellular bacteria. Limitations and advantages of these techniques will be discussed with an eye toward the methods still needed, and how the current state of the art for genetics in obligate intracellular bacteria could direct future technological advances for Chlamydia.

  1. Lactobacillus crispatus inhibits the infectivity of Chlamydia trachomatis elementary bodies, in vitro study

    PubMed Central

    Nardini, Paola; Ñahui Palomino, Rogers Alberto; Parolin, Carola; Laghi, Luca; Foschi, Claudio; Cevenini, Roberto; Vitali, Beatrice; Marangoni, Antonella

    2016-01-01

    Lactobacillus species dominate the vaginal microbiota of healthy reproductive-age women and protect the genitourinary tract from the attack of several infectious agents. Chlamydia trachomatis, a leading cause of sexually transmitted disease worldwide, can induce severe sequelae, i.e. pelvic inflammatory disease, infertility and ectopic pregnancy. In the present study we investigated the interference of Lactobacillus crispatus, L. gasseri and L. vaginalis, known to be dominant species in the vaginal microbiome, with the infection process of C. trachomatis. Lactobacilli exerted a strong inhibitory effect on Chlamydia infectivity mainly through the action of secreted metabolites in a concentration/pH dependent mode. Short contact times were the most effective in the inhibition, suggesting a protective role of lactobacilli in the early steps of Chlamydia infection. The best anti-Chlamydia profile was shown by L. crispatus species. In order to delineate metabolic profiles related to anti-Chlamydia activity, Lactobacillus supernatants were analysed by 1H-NMR. Production of lactate and acidification of the vaginal environment seemed to be crucial for the activity, in addition to the consumption of the carbonate source represented by glucose. The main conclusion of this study is that high concentrations of L. crispatus inhibit infectivity of C. trachomatis in vitro. PMID:27354249

  2. Improved Plaque Assay Identifies a Novel Anti-Chlamydia Ceramide Derivative with Altered Intracellular Localization

    PubMed Central

    Banhart, Sebastian; Saied, Essa M.; Martini, Andrea; Koch, Sophia; Aeberhard, Lukas; Madela, Kazimierz; Arenz, Christoph

    2014-01-01

    Chlamydia trachomatis is a medically important human pathogen causing different diseases, including trachoma, the leading cause of preventable blindness in developing countries, and sexually transmitted infections that can lead to infertility and ectopic pregnancies. There is no vaccine against C. trachomatis at present. Broad-spectrum antibiotics are used as standard therapy to treat the infection but have unwanted side effects, such as inducing persistent or recurring infections and affecting the host microbiome, necessitating the development of novel anti-Chlamydia therapies. Here, we describe the establishment of a robust, fast, and simple plaque assay using liquid overlay medium (LOM) for the identification of anti-Chlamydia compounds. Using the LOM plaque assay, we identified nitrobenzoxadiazole (NBD)-labeled 1-O-methyl-ceramide-C16 as a compound that efficiently inhibits C. trachomatis replication without affecting the viability of the host cell. Further detailed analyses indicate that 1-O-methyl-NBD-ceramide-C16 acts outside the inclusion. Thereby, 1-O-methyl-NBD-ceramide-C16 represents a lead compound for the development of novel anti-Chlamydia drugs and furthermore constitutes an agent to illuminate sphingolipid trafficking pathways in Chlamydia infections. PMID:25001308

  3. Seroprevalence of Chlamydia infection in pigs in Jiangxi province, South-Eastern China.

    PubMed

    Jiang, H H; Huang, S Y; Zhang, W B; Zhao, L; Xu, C M; Deng, S Z; Zhu, X Q

    2013-12-01

    Chlamydia are Gram-negative obligate bacteria that cause a wide range of diseases in humans and animals. To assess the risk of zoonosis posed by pigs, a total of 920 serum samples were collected from pigs in 11 administrative cities in Jiangxi province, south-eastern China, and the seroprevalence of Chlamydia antibodies was investigated by an indirect haemagglutination assay. The pathogen-specific antibodies were detected in 539 (58.59 %) pigs with seroprevalence ranging from 33.33 % (Jingdezhen) to 90.91 % (Pingxiang) among different cities (P<0.05). The highest prevalence was found in pregnant sows (80.89 %, 127/157), followed by breeding boars (79.37 %, 50/63), suckling sows (77.01 %, 67/87), fattening pigs (69.32 %, 61/88) and non-pregnant sows (62.5 %, 180/288). Piglets had the lowest prevalence of 22.78 % (54/237). The seroprevalence of Chlamydia infection among different categories of pigs was also significantly different (P<0.05). These results indicate that Chlamydia is highly prevalent in pigs in Jiangxi province and our results indicate that the presence of Chlamydia exposure in pigs may pose a potential threat to human health.

  4. Evaluation of Brucella abortus strain RB51 and strain 19 in pronghorn antelope.

    PubMed

    Elzer, Philip H; Smith, J; Roffe, T; Kreeger, T; Edwards, J; Davis, D

    2002-10-01

    Free-roaming elk and bison in the Greater Yellowstone Area remain the only wildlife reservoirs for Brucella abortus in the United States, and the large number of animals and a lack of holding facilities make it unreasonable to individually vaccinate each animal. Therefore, oral delivery is being proposed as a possible option to vaccinate these wild ungulates. One of the main problems associated with oral vaccination is the potential exposure of nontarget species to the vaccines. The purpose of this study was to determine the effects of two Brucella vaccines, strain 19 (S19) and the rough strain RB51 (SRB51), in pregnant pronghorn antelope. We conclude that S19 and SRB51 rarely colonize maternal and fetal tissues of pregnant pronghorn and were not associated with fetal death. Oral delivery of either vaccine at this dose appears to be nonhazardous to pregnant pronghorn. PMID:12381572

  5. Brucella abortus Strain 2308 Wisconsin Genome: Importance of the Definition of Reference Strains

    PubMed Central

    Suárez-Esquivel, Marcela; Ruiz-Villalobos, Nazareth; Castillo-Zeledón, Amanda; Jiménez-Rojas, César; Roop II, R. Martin; Comerci, Diego J.; Barquero-Calvo, Elías; Chacón-Díaz, Carlos; Caswell, Clayton C.; Baker, Kate S.; Chaves-Olarte, Esteban; Thomson, Nicholas R.; Moreno, Edgardo; Letesson, Jean J.; De Bolle, Xavier; Guzmán-Verri, Caterina

    2016-01-01

    Brucellosis is a bacterial infectious disease affecting a wide range of mammals and a neglected zoonosis caused by species of the genetically homogenous genus Brucella. As in most studies on bacterial diseases, research in brucellosis is carried out by using reference strains as canonical models to understand the mechanisms underlying host pathogen interactions. We performed whole genome sequencing analysis of the reference strain B. abortus 2308 routinely used in our laboratory, including manual curated annotation accessible as an editable version through a link at https://en.wikipedia.org/wiki/Brucella#Genomics. Comparison of this genome with two publically available 2308 genomes showed significant differences, particularly indels related to insertional elements, suggesting variability related to the transposition of these elements within the same strain. Considering the outcome of high resolution genomic techniques in the bacteriology field, the conventional concept of strain definition needs to be revised. PMID:27746773

  6. Evaluation of Brucella abortus strain RB51 and strain 19 in pronghorn antelope

    USGS Publications Warehouse

    Elzer, P.H.; Smith, J.; Roffe, T.; Kreeger, T.; Edwards, J.; Davis, D.

    2002-01-01

    Free-roaming elk and bison in the Greater Yellowstone Area remain the only wildlife reservoirs for Brucella abortus in the United States, and the large number of animals and a lack of holding facilities make it unreasonable to individually vaccinate each animal. Therefore, oral delivery is being proposed as a possible option to vaccinate these wild ungulates. One of the main problems associated with oral vaccination is the potential exposure of nontarget species to the vaccines. The purpose of this study was to determine the effects of two Brucella vaccines, strain 19 (S19) and the rough strain RB51 (SRB51), in pregnant pronghorn antelope. We conclude that S19 and SRB51 rarely colonize maternal and fetal tissues of pregnant pronghorn and were not associated with fetal death. Oral delivery of either vaccine at this dose appears to be nonhazardous to pregnant pronghorn.

  7. An ecological perspective on the changing face of Brucella abortus in the western United States

    USGS Publications Warehouse

    Cross, Paul C.; Maichak, Eric J.; Brennan, Angela; Scurlock, Brandon M.; Henningsen, John C.; Luikart, Gordon

    2013-01-01

    After a hiatus during the 1990s, outbreaks of Brucella abortus in cattle are occurring more frequently in some of the western states of the United States, namely, Montana, Wyoming and Idaho. This increase is coincident with increasing brucellosis seroprevalence in elk (Cervus elaphus), which is correlated with elk density. Vaccines are a seductive solution, but their use in wildlife systems remains limited by logistical, financial, and scientific constraints. Cattle vaccination is ongoing in the region. Livestock regulations, however, tend to be based on serological tests that test for previous exposure and available vaccines do not protect against seroconversion. The authors review recent ecological studies of brucellosis, with particular emphasis on the Greater Yellowstone Area, and highlight the management options and implications of this work, including the potential utility of habitat modifications and targeted hunts, as well as scavengers and predators. Finally, the authors discuss future research directions that will help us to understand and manage brucellosis in wildlife.

  8. Crystal structure of cyclic nucleotide-binding-like protein from Brucella abortus.

    PubMed

    He, Zheng; Gao, Yuan; Dong, Jing; Ke, Yuehua; Li, Xuemei; Chen, Zeliang; Zhang, Xuejun C

    2015-12-25

    The cyclic nucleotide-binding (CNB)-like protein (CNB-L) from Brucella abortus shares sequence homology with CNB domain-containing proteins. We determined the crystal structure of CNB-L at 2.0 Å resolution in the absence of its C-terminal helix and nucleotide. The 3D structure of CNB-L is in a two-fold symmetric form. Each protomer shows high structure similarity to that of cGMP-binding domain-containing proteins, and likely mimics their nucleotide-free conformation. A key residue, Glu17, mediates the dimerization and prevents binding of cNMP to the canonical ligand-pocket. The structurally observed dimer of CNB-L is stable in solution, and thus is likely to be biologically relevant.

  9. First report of orchitis in man caused by Brucella abortus biovar 1 in Ecuador.

    PubMed

    Ron-Román, Jorge; Saegerman, Claude; Minda-Aluisa, Elizabeth; Benítez-Ortíz, Washington; Brandt, Jef; Douce, Richard

    2012-09-01

    We present a 44-year-old man from a rural community in northern Ecuador who worked on a cattle farm where he was involved with primary veterinary care, including assistance during births (or calving) and placenta retention and artificial insemination, with minimal precautions. In September of 2009, quite abruptly, he developed asthenia and hypersomnia without any apparent cause or symptoms like fever, chills, or night sweats. On November 14, 2009, he suffered from pain and edema in the right testicle that coincided with pain in the abdomen. Clinical, serological, and bacteriological investigations confirmed the first case of unilateral orchitis in man in Ecuador caused by Brucella abortus biovar 1. Because brucellosis is a neglected disease, special attention should be given to it in the training of medical and veterinary students. PMID:22826490

  10. First Report of Orchitis in Man Caused by Brucella abortus Biovar 1 in Ecuador

    PubMed Central

    Ron-Román, Jorge; Saegerman, Claude; Minda-Aluisa, Elizabeth; Benítez-Ortíz, Washington; Brandt, Jef; Douce, Richard

    2012-01-01

    We present a 44-year-old man from a rural community in northern Ecuador who worked on a cattle farm where he was involved with primary veterinary care, including assistance during births (or calving) and placenta retention and artificial insemination, with minimal precautions. In September of 2009, quite abruptly, he developed asthenia and hypersomnia without any apparent cause or symptoms like fever, chills, or night sweats. On November 14, 2009, he suffered from pain and edema in the right testicle that coincided with pain in the abdomen. Clinical, serological, and bacteriological investigations confirmed the first case of unilateral orchitis in man in Ecuador caused by Brucella abortus biovar 1. Because brucellosis is a neglected disease, special attention should be given to it in the training of medical and veterinary students. PMID:22826490

  11. Properties of a cell-wall-defective variant of Brucella abortus of bovine origin.

    PubMed Central

    Corbel, M. J.; Scott, A. C.; Ross, H. M.

    1980-01-01

    The properties of an atypical Brucella strain isolated from lymph node tissue of a cow slaughtered as a brucellosis reactor were examined. The organism was Gram negative and highly pleomorphic, existing as cocci, coccobacilli, rods, branched and irregular forms which stained with fluorescent antibody conjugates prepared against rough and smooth Brucella abortus strains. It produced lecithinase and required at least 15% v/v equine or bovine serum for growth. It did not need supplementary CO2 for growth, produced H2S and was inhibited by brucella dyes and partially by i-erythritol. Growth inhibition or lysis was produced by brucella-phages. The organism was not pathogenic for guinea-pigs or mice but evoked antibodies mainly to rough Brucella antigens. Images Fig. 1 Fig. 2 Fig. 3 Fig. 1 Fig. 2 Fig. 1 Fig. 1 Fig. 2 PMID:6820027

  12. On the inactivation of Brucella abortus in naturally contaminated milk by commercial pasteurisation procedures.

    PubMed

    Van den Heever, L W; Katz, K W; Te Brugge, L A

    1982-12-01

    Following concern about the recent publication indicating the possible ability of Brucella abortus to survive commercial pasteurisation of naturally contaminated milk, such milk was subjected to biological (BT), serological and bacteriological tests. The raw milk was Brucella Milk Ring Test positive and biotype I was isolated from 4/5 BT guinea pigs, the one tested being seropositive to the Rose Bengal Test, and with serum agglutination and complement fixation titres of 160 and 36 international units respectively. After batch (63 degrees C/30 min) and high temperature, short time (72 degrees C/15 sec) pasteurisation, all 10 BT guinea pigs were bacteriologically and serologically negative, indicating that officially approved methods of commercial pasteurisation rendered naturally Brucella-contaminated raw milk safe for consumption.

  13. Crystal structure of cyclic nucleotide-binding-like protein from Brucella abortus.

    PubMed

    He, Zheng; Gao, Yuan; Dong, Jing; Ke, Yuehua; Li, Xuemei; Chen, Zeliang; Zhang, Xuejun C

    2015-12-25

    The cyclic nucleotide-binding (CNB)-like protein (CNB-L) from Brucella abortus shares sequence homology with CNB domain-containing proteins. We determined the crystal structure of CNB-L at 2.0 Å resolution in the absence of its C-terminal helix and nucleotide. The 3D structure of CNB-L is in a two-fold symmetric form. Each protomer shows high structure similarity to that of cGMP-binding domain-containing proteins, and likely mimics their nucleotide-free conformation. A key residue, Glu17, mediates the dimerization and prevents binding of cNMP to the canonical ligand-pocket. The structurally observed dimer of CNB-L is stable in solution, and thus is likely to be biologically relevant. PMID:26549229

  14. A "dipstick" enzyme immunoassay for detection of antibody to Brucella abortus in cattle sera.

    PubMed

    Nielsen, K; Ballinger, R; Stiller, J; Rosenbaum, B

    1985-07-01

    An enzyme immunoassay that utilizes antigen bound to a matrix which can be removed from the substrate to stop development is described. The assay which is performed in glass or plastic disposable tubes uses Gel-Bond film strips for attachment of antigen. The only equipment requirements are a rotary shaker and a spectrophotometer (optional). The antigen coated strips are passed through a series of tubes containing test serum, wash solution, antibody-enzyme conjugate, wash solution and substrate-chromogen taking about 45 minutes to perform. In testing sera with or without antibody to Brucella abortus a very high correlation existed between same day tests and tests performed over several days as well as with data on the same sera obtained by an enzyme immunoassay in a microtiter format.

  15. An ecological perspective on Brucella abortus in the western United States.

    PubMed

    Cross, P C; Maichak, E J; Brennan, A; Scurlock, B M; Henningsen, J; Luikart, G

    2013-04-01

    After a hiatus during the 1990s, outbreaks of Brucella abortus in cattle are occurring more frequently in some of the western states of the United States, namely, Montana, Wyoming and Idaho. This increase is coincidentwith increasing brucellosis seroprevalence in elk (Cervus elaphus), which is correlated with elk density. Vaccines are a seductive solution, but their use in wildlife systems remains limited by logistical, financial, and scientific constraints. Cattle vaccination is ongoing in the region. Livestock regulations, however, tend to be based on serological tests that test for previous exposure and available vaccines do not protect against seroconversion. The authors review recent ecological studies of brucellosis, with particular emphasis on the Greater Yellowstone Area, and highlight the management options and implications of this work, including the potential utility of habitat modifications and targeted hunts, as well as scavengers and predators. Finally, the authors discuss future research directions that will help us to understand and manage brucellosis in wildlife. PMID:23837367

  16. New point of care Chlamydia Rapid Test—bridging the gap between diagnosis and treatment: performance evaluation study

    PubMed Central

    2007-01-01

    Objective To evaluate the performance of a new Chlamydia Rapid Test with vaginal swab specimens as a potential tool for chlamydia diagnosis and screening. Design Performance evaluation study. Settings A young people’s sexual health centre (site 1) and two genitourinary medicine clinics (sites 2 and 3) in the United Kingdom. Participants 1349 women aged between 16 and 54 attending one of the three clinics. Main outcome measures Sensitivity, specificity, positive predictive value, and negative predictive value of the Chlamydia Rapid Test versus polymerase chain reaction and strand displacement amplification assays; correlation between the Chlamydia Rapid Test visual signal and organism load; acceptability to participants of self collected vaginal swabs as the specimen type for Chlamydia testing. Results Polymerase chain reaction positivity rates for Chlamydia trachomatis infection were 8.4% (56/663) at site 1, 9.4% (36/385) at site 2, and 6.0% (18/301) at site 3. Compared with polymerase chain reaction assay, the resolved sensitivity, specificity, positive predictive value, and negative predictive value of the Chlamydia Rapid Test were 83.5% (91/109), 98.9% (1224/1238), 86.7% (91/105), and 98.6% (1224/1242). Compared with strand displacement amplification assay, sensitivity and specificity of the Chlamydia Rapid Test were 81.6% (40/49) and 98.3% (578/588). Organism load of self collected vaginal swabs ranged from 5.97×102 to 1.09×109 Chlamydia plasmids per swab, which correlated well with the Chlamydia Rapid Test’s visual signal (r=0.6435, P<0.0001). Most (95.9%) surveyed participants felt comfortable about collecting their own swabs. Conclusions The performance of the Chlamydia Rapid Test with self collected vaginal swabs indicates that it would be an effective same day diagnostic and screening tool for Chlamydia infection in women. The availability of Chlamydia Rapid Test results within 30 minutes allows for immediate treatment and contact tracing

  17. What's in a word: the use, misuse, and abuse of the word “persistence” in Chlamydia biology

    PubMed Central

    Bavoil, Patrik M.

    2014-01-01

    The word persistence was used by Chlamydia researchers almost as soon as Chlamydia research was born to reflect the propensity of chlamydiae to cause inapparent infection in their hosts, from birds to humans. More recently, the term persistence has been used, misused, and sometimes abused amidst in vitro and in vivo studies that aim to mimick the ability of chlamydiae to emerge from the presumed inapparent state into clinically detectable infection and disease. Here, I have attempted to provide a global perspective on the state of research on chlamydial persistence, revisiting old observations that may warrant a new look, critically evaluating more recent observations and their shortcomings, and including recent developments that may help redefine chlamydiae as pathogens—or not—of both animals and humans. PMID:24624366

  18. Brief Report: Gonorrhea and Chlamydia Testing Increasing but Still Lagging in HIV Clinics in the United States.

    PubMed

    Berry, Stephen A; Ghanem, Khalil G; Mathews, William Christopher; Korthuis, Philip Todd; Yehia, Baligh R; Agwu, Allison L; Lehmann, Christoph U; Moore, Richard D; Allen, Sara L; Gebo, Kelly A

    2015-11-01

    Screening persons living with HIV for gonorrhea and chlamydia has been recommended since 2003. We compared annual gonorrhea/chlamydia testing to syphilis and lipid testing among 19,368 adults (41% men who have sex with men, 30% heterosexual men, and 29% women) engaged in HIV care. In 2004, 22%, 62%, and 70% of all patients were tested for gonorrhea/chlamydia, syphilis, and lipid levels, respectively. Despite increasing steadily [odds ratio per year (95% confidence interval): 1.14 (1.13 to 1.15)], gonorrhea/chlamydia testing in 2010 remained lower than syphilis and lipid testing (39%, 77%, 76%, respectively). Interventions to improve gonorrhea/chlamydia screening are needed. A more targeted screening approach may be warranted.

  19. Evaluation of Brucella abortus Phosphoglucomutase (pgm) Mutant as a New Live Rough-Phenotype Vaccine

    PubMed Central

    Ugalde, Juan Esteban; Comerci, Diego José; Leguizamón, M. Susana; Ugalde, Rodolfo Augusto

    2003-01-01

    Brucella abortus S19 is the vaccine most frequently used against bovine brucellosis. Although it induces good protection levels, it cannot be administered to pregnant cattle, revaccination is not advised due to interference in the discrimination between infected and vaccinated animals during immune-screening procedures, and the vaccine is virulent for humans. Due to these reasons, there is a continuous search for new bovine vaccine candidates that may confer protection levels comparable to those conferred by S19 but without its disadvantages. A previous study characterized the phenotype associated with the phosphoglucomutase (pgm) gene disruption in Brucella abortus S2308, as well as the possible role for the smooth lipopolysaccharide (LPS) in virulence and intracellular multiplication in HeLa cells (J. E. Ugalde, C. Czibener, M. F. Feldman, and R. A. Ugalde, Infect. Immun. 68:5716-5723, 2000). In this report, we analyze the protection, proliferative response, and cytokine production induced in BALB/c mice by a Δpgm deletion strain. We show that this strain synthesizes O antigen with a size of approximately 45 kDa but is rough. This is due to the fact that the Δpgm strain is unable to assemble the O side chain in the complete LPS. Vaccination with the Δpgm strain induced protection levels comparable to those induced by S19 and generated a proliferative splenocyte response and a cytokine profile typical of a Th1 response. On the other hand, we were unable to detect a specific anti-O-antigen antibody response by using the fluorescence polarization assay. In view of these results, the possibility that the Δpgm mutant could be used as a vaccination strain is discussed. PMID:14573645

  20. Efficacy of single calfhood vaccination of elk with Brucella abortus strain 19

    USGS Publications Warehouse

    Roffe, T.J.; Jones, L.C.; Coffin, K.; Drew, M.L.; Sweeney, Steven J.; Hagius, S.D.; Elzer, P.H.; Davis, D.

    2004-01-01

    Brucellosis has been eradicated from cattle in the states of Wyoming, Montana, and Idaho, USA. However, free-ranging elk (Cervus elaphus) that use feedgrounds in the Greater Yellowstone Area (GYA) and bison (Bison bison) in Yellowstone and Grand Teton national parks still have high seroprevalence to the disease and have caused loss of brucellosis-free status in Wyoming. Management tools to control or eliminate the disease are limited; however, wildlife vaccination is among the methods currently used by wildlife managers in Wyoming. We conducted a controlled challenge study of single calfhood vaccination. Elk calves, caught in January and February of 1999 and 2000 and acclimated to captivity for 3 weeks, were randomly assigned to control or vaccinate groups. The vaccinate groups received Brucetta abortus vaccine strain 19 (S19) by hand-delivered intramuscular injection. Calves were raised to adulthood and bred at either 2.5 or 3.5 years of age for 2000 and 1999 captures, respectively. Eighty-nine (44 controls, 45 vaccinates) pregnant elk entered the challenge portion of the study. We challenged elk at mid-gestation with pathogenic B. abortus strain 2308 by intraconjunctival instillation. Abortion occurred in significantly more (P = 0.002) controls (42; 93%) than vaccinates (32; 71%), and vaccine protected 25% of the vaccinate group. We used Brucella culture of fetus/calf tissues to determine the efficacy of vaccination for preventing infection, and we found that the number of infected fetuses/calves did not differ between controls and vaccinates (P = 0.14). Based on these data, single calfhood vaccination with S19 has low efficacy, will likely have only little to moderate effect on Brucella prevalence in elk, and is unlikely to eradicate the disease in wildlife of the GYA.