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Annie Rodolakis - One of the best experts on this subject based on the ideXlab platform.
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Effectiveness of vaccine strain 1B against Tunisian field strains of Chlamydophila Abortus using mouse model
Revue De Medecine Veterinaire, 2020Co-Authors: Abdessalem Rekiki, A Bouakane, F. Bernard, S. Hammami, Annie RodolakisAbstract:SUMMARY The efficacy of vaccine strain 1B against two selected Tunisian Chlamydophila Abortus strains, isolated from an aborted (ABt35) and from a lambing ewe (MBt34), was demonstrated in a mouse model. The number of live new-born mice in vaccinated groups, challenged subsequently with the reference French C. Abortus (AB7) or either of the Tunisian isolates (ABt35 or MBt34), was not significantly different from the control pregnant group. Vaccine 1B thus provided effective protection against the selected Tunisian field C. Abortus strains. Moreover, the strain isolated from the aborted ewe (ABt35) appeared significantly more virulent than that isolated from the lambing ewe (MBt34).
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differential identification of Chlamydophila Abortus live vaccine strain 1b and c Abortus field isolates by pcr rflp
Vaccine, 2010Co-Authors: Karine Laroucau, Victoria I Siarkou, Evangelia Vretou, Annie Rodolakis, Fabien Vorimore, Konrad Sachse, Hermann Willems, Simone Magnino, Patrik M BavoilAbstract:Comparative genomic analysis of a wild-type strain of the ovine pathogen Chlamydophila Abortus and its nitrosoguanidine-induced, temperature-sensitive and virulence-attenuated live vaccine derivative identified point mutations unique to the mutant (Burall et al. [1]). Here, we evaluate the capacity of some of these mutations to either create or eliminate restriction sites using the wild-type strain C. Abortus S26/3 as a reference. Three of eight genomic sites with confirmed point mutations (CAB153, CAB636 and CAB648) were retained for analysis as each resulted in the loss of a restriction site in the genome sequence of the vaccine strain. PCR-restriction fragment length polymorphism analysis using restriction enzymes chosen to specifically target the three genomic sites was then applied to a large number of C. Abortus field isolates and reference strains. Our results indicate that the three mutations are uniquely present in the vaccine strain, and as such provide easy-to-use markers for the differential identification of the vaccine strain and wild-type isolates.
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simultaneous differential detection of Chlamydophila Abortus Chlamydophila pecorum and coxiella burnetii from aborted ruminant s clinical samples using multiplex pcr
BMC Microbiology, 2009Co-Authors: Mustapha Berri, Abdessalem Rekiki, Karim Sidi Boumedine, Annie RodolakisAbstract:Background Chlamydiosis and Q fever, two zoonosis, are important causes of ruminants' abortion around the world. They are caused respectively by strictly intracellular and Gram negative bacterium Chlamydophila Abortus (Cp. Abortus) and Coxiella burnetii (C. burnetii). Chlamydophila pecorum (Cp. pecorum) is commonly isolated from the digestive tract of clinically inconspicuous ruminants but the abortive and zoonotic impact of this bacterium is still unknown because Cp. pecorum is rarely suspected in abortion cases of small ruminants. We have developed a multiplex PCR (m-PCR) for rapid simultaneous differential detection of Cp. Abortus, Cp. pecorum and C. burnetii in clinical samples taken from infected animals.
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protection of pregnant mice against placental and splenic infection by three strains of Chlamydophila Abortus with a live 1b vaccine
Veterinary Record, 2005Co-Authors: A Bouakane, Abdessalem Rekiki, Annie RodolakisAbstract:The efficacy of a live 1B vaccine against three strains of Chlamydophila Abortus , AB16, LLG and POS, was assessed in pregnant mice in terms of the reduction in the levels of infection recorded in their placentas, fetuses and spleens. The vaccine was more effective against the AB16 strain than against the LLG and POS strains, suggesting that there are antigenic differences between the three strains.
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molecular characterisation and ovine live vaccine 1b evaluation toward a Chlamydophila Abortus strain isolated from springbok antelope abortion
Veterinary Microbiology, 2004Co-Authors: Mustapha Berri, F. Bernard, A Lecu, F Ollivetcourtois, Annie RodolakisAbstract:Chlamydiosis is a zoonosis with a worldwide distribution. The reservoir of susceptible hosts is large and includes birds and both domestic and wild mammals. Chlamydial infection, determined serologically, seems to be widespread among wild ruminants in the Paris zoo (France). In February 2003, an abortion case was reported within the springbok (Antidorcas marsupialis) herd of the zoo. PCR assay using primers targeting the polymorph membrane protein gene (pmp) family was performed on both vaginal swab and placenta samples revealing the presence of Chlamydophila. The inoculation into chicken embryos of an infected placenta extract led to the successful isolation of a C. Abortus strain referred to as ASb1. The omp1 gene coding the major outer membrane protein (momp) and the 16S–23S rRNA spacer region of ASb1 were compared to those of various strains by restriction fragment length polymorphism (RFLP). The RFLP analysis showed that this isolate belonged to Chlamydophila Abortus species and is highly related to known domestic ruminant's strains causing abortion. The efficacy of a live vaccine 1B, based on a temperature-sensitive mutant of the ovine abortion reference strain AB7, was tested. Protection-challenge experiments in a mouse model show that the ASb1 strain led to mice abortions and that vaccination with 1B vaccine provided them with effective protection.
David Longbottom - One of the best experts on this subject based on the ideXlab platform.
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genome sequence of the Chlamydophila Abortus variant strain llg
Journal of Bacteriology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Evangelia Vretou, David Smith, Alex F Lainson, David LongbottomAbstract:Chlamydophila Abortus is a common cause of ruminant abortion. Here we report the genome sequence of strain LLG, which differs genotypically and phenotypically from the wild-type strain S26/3. Genome sequencing revealed differences between LLG and S26/3 to occur in pseudogene content, in transmembrane head/inc family proteins, and in biotin biosynthesis genes.
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identification sequencing and molecular analysis of chp4 a novel chlamydiaphage of Chlamydophila Abortus belonging to the family microviridae
Journal of General Virology, 2011Co-Authors: Michelle Sait, Nick Wheelhouse, Morag Livingstone, Rebecca Graham, Neil F Inglis, David LongbottomAbstract:Members of the family Microviridae have been identified in a number of chlamydial species infecting humans (phage CPAR39 in Chlamydophila pneumoniae), other mammals (φCPG1 in Chlamydophila caviae, Chp2 in Chlamydophila Abortus and Chp3 in Chlamydophila pecorum) and birds (Chp1 in Chlamydophila psittaci). This study describes the identification and genome sequencing of Chp4, an icosahedral, 4530 bp, ssDNA phage in C. Abortus. Chp4 is predicted to contain eight ORFs, six of which could be assigned putative functions based on sequence similarity to characterized bacteriophage. Gene order and content were highly conserved amongst chlamydiaphage, with the highest sequence variability occurring in the IN5 and INS variable regions of the VP1 major coat protein, which has been associated with host cell recognition and binding. Phylogenetic analysis of VP1 indicated that Chp4 is a member of the Chlamydiamicrovirus, and is most closely related to phage φCPG1 and CPAR39.
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inflammatory cytokine responses in a pregnant mouse model of Chlamydophila Abortus infection
Veterinary Microbiology, 2010Co-Authors: Karen P Kerr, Nick Wheelhouse, Morag Livingstone, Gary Entrican, Declan J Mckeever, Ian E Anderson, David LongbottomAbstract:Abstract Chlamydophila Abortus ( C. Abortus ) is the aetiological agent of ovine enzootic abortion (OEA). The highly elevated expression of the pro-inflammatory cytokine tumour necrosis factor-alpha (TNFα) and low-level expression of interferon-gamma (IFNγ) that are detected in C. Abortus- infected placentas have been implicated in the pathogenesis of OEA. Late-term abortions similar to those occurring in sheep have also been observed in mouse models of C. Abortus infection. Since mouse studies have contributed significantly to our understanding of the immunological responses to chlamydial infections and serve as a good model for rapidly assessing candidate vaccines for OEA, we investigated local expression of TNFα and IFNγ in infected mice. At various time points over the course of infection mice were sacrificed, serum samples obtained for serum antibody and cytokine analyses, and livers and placental tissues were removed and fixed to determine C. Abortus colonisation and cytokine expression. Immunostaining for C. Abortus was significantly greater in placenta compared to liver ( P C. Abortus infection but not placental/fetal infection in mice. In contrast to sheep, murine placental TNFα expression does not correlate with C. Abortus infection, suggesting that the immunopathogenesis of chlamydial abortion differs in these species.
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evidence of Chlamydophila Abortus vaccine strain 1b as a possible cause of ovine enzootic abortion
Vaccine, 2010Co-Authors: Nick Wheelhouse, Kevin Aitchison, Karine Laroucau, Jill R Thomson, David LongbottomAbstract:Chlamydophila Abortus, the agent of ovine enzootic abortion (OEA), is a major cause of lamb mortality worldwide. Disease can be controlled through the use of vaccines based on the 1B temperature-sensitive mutant strain of C. Abortus. This study investigated suspected OEA cases across Scotland for the presence of the 1B strain by analysis of recently identified unique point mutations (9). Thirty-five cases were C. Abortus-positive and 14 came from vaccinated flocks. Analysis of single nucleotide polymorphisms by PCR-RFLP and sequence analysis revealed the presence of point mutations consistent with the presence of the 1B vaccine strain in 5 of these 14 samples. Quantitative real-time PCR revealed comparable numbers of genome copies of the 1B strain in infected placentas to those present following wild-type infection. This study is the first to demonstrate the presence of the 1B vaccine strain in the placentas of OEA cases and suggests a probable causal role in the disease.
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Inflammatory cytokine responses in a pregnant mouse model of Chlamydophila Abortus infection.
Veterinary microbiology, 2010Co-Authors: Karen Kerr, Morag Livingstone, Gary Entrican, Nicholas Wheelhouse, Ian E Anderson, Declan Mckeever, David LongbottomAbstract:Chlamydophila Abortus (C. Abortus) is the aetiological agent of ovine enzootic abortion (OEA). The highly elevated expression of the pro-inflammatory cytokine tumour necrosis factor-alpha (TNFalpha) and low-level expression of interferon-gamma (IFNgamma) that are detected in C. Abortus-infected placentas have been implicated in the pathogenesis of OEA. Late-term abortions similar to those occurring in sheep have also been observed in mouse models of C. Abortus infection. Since mouse studies have contributed significantly to our understanding of the immunological responses to chlamydial infections and serve as a good model for rapidly assessing candidate vaccines for OEA, we investigated local expression of TNFalpha and IFNgamma in infected mice. At various time points over the course of infection mice were sacrificed, serum samples obtained for serum antibody and cytokine analyses, and livers and placental tissues were removed and fixed to determine C. Abortus colonisation and cytokine expression. Immunostaining for C. Abortus was significantly greater in placenta compared to liver (P
Victoria I Siarkou - One of the best experts on this subject based on the ideXlab platform.
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genome sequence of the Chlamydophila Abortus variant strain llg
Journal of Bacteriology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Evangelia Vretou, David Smith, Alex F Lainson, David LongbottomAbstract:Chlamydophila Abortus is a common cause of ruminant abortion. Here we report the genome sequence of strain LLG, which differs genotypically and phenotypically from the wild-type strain S26/3. Genome sequencing revealed differences between LLG and S26/3 to occur in pseudogene content, in transmembrane head/inc family proteins, and in biotin biosynthesis genes.
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genetic variability of Chlamydophila Abortus strains assessed by pcr rflp analysis of polymorphic membrane protein encoding genes
Veterinary Microbiology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Konrad Sachse, Simone Magnino, B K Markey, Evangelia VretouAbstract:This study used PCR-RFLP to investigate the genetic variability of pmp-encoding genes from fifty-two Chlamydophila Abortus (C. Abortus) strains originating from abortion cases from various geographical regions and host species. Six primer pairs were used to PCR-amplify DNA fragments encoding eighteen pmps. PCR products were digested using four restriction endonucleases and Bayesian methodologies were used to compare RFLP profiles and assign strains to a RFLP genotype. Strains could be assigned to 2 genotypes in the region encoding pmp18D, 3 genotypes in the regions encoding pmp1A-pmp2B, pmp3E-pmp6H and pmp11G-pmp15G, 4 genotypes in the region encoding pmp7G-pmp10G and 5 genotypes in the region encoding pmp16G-pmp17G. In all regions, the majority of strains (88.4-96.1%) had the same genotype as the reference strain S26/3. No correlation could be made between genotype, host species or geographical origin except for the two variant Greek strains, LLG and POS, which formed a discrete genotype in all pmp-encoding regions except pmp18D. Relative rates of evolution calculated for each pmp-encoding gene locus suggest that differing selective pressures and functional constraints may exist on C. Abortus polymorphic membrane proteins. These findings suggest that although intraspecies heterogeneity of pmp-encoding genes in C. Abortus is low, the sequence heterogeneity should be an important consideration when using pmps as the basis for novel diagnostics or vaccine development.
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evolutionary relationships among Chlamydophila Abortus variant strains inferred by rrna secondary structure based phylogeny
PLOS ONE, 2011Co-Authors: Victoria I Siarkou, Alexandros Stamatakis, Ilias Kappas, Paul Hadweh, Karine LaroucauAbstract:The evolutionary relationships among known Chlamydophila Abortus variant strains including the LLG and POS, previously identified as being highly distinct, were investigated based on rRNA secondary structure information. PCR-amplified overlapping fragments of the 16S, 16S-23S intergenic spacer (IS), and 23S domain I rRNAs were subjected to cloning and sequencing. Secondary structure analysis revealed the presence of transitional single nucleotide variations (SNVs), two of which occurred in loops, while seven in stem regions that did not result in compensatory substitutions. Notably, only two SNVs, in 16S and 23S, occurred within evolutionary variable regions. Maximum likelihood and Bayesian phylogeny reconstructions revealed that C. Abortus strains could be regarded as representing two distinct lineages, one including the “classical” C. Abortus strains and the other the “LLG/POS variant”, with the type strain B577T possibly representing an intermediate of the two lineages. The two C. Abortus lineages shared three unique (apomorphic) characters in the 23S domain I and 16S-23S IS, but interestingly lacked synapomorphies in the 16S rRNA. The two lineages could be distinguished on the basis of eight positions; four of these comprised residues that appeared to be signature or unique for the “classical” lineage, while three were unique for the “LLG/POS variant”. The U277 (E. coli numbering) signature character, corresponding to a highly conserved residue of the 16S molecule, and the unique G681 residue, conserved in a functionally strategic region also of 16S, are the most pronounced attributes (autapomorphies) of the “classical” and the “LLG/POS variant” lineages, respectively. Both lineages were found to be descendants of a common ancestor with the Prk/Daruma C. psittaci variant. Compared with the “classical”, the “LLG/POS variant” lineage has retained more ancestral features. The current rRNA secondary structure-based analysis and phylogenetic inference reveal new insights into how these two C. Abortus lineages have differentiated during their evolution.
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differential identification of Chlamydophila Abortus live vaccine strain 1b and c Abortus field isolates by pcr rflp
Vaccine, 2010Co-Authors: Karine Laroucau, Victoria I Siarkou, Evangelia Vretou, Annie Rodolakis, Fabien Vorimore, Konrad Sachse, Hermann Willems, Simone Magnino, Patrik M BavoilAbstract:Comparative genomic analysis of a wild-type strain of the ovine pathogen Chlamydophila Abortus and its nitrosoguanidine-induced, temperature-sensitive and virulence-attenuated live vaccine derivative identified point mutations unique to the mutant (Burall et al. [1]). Here, we evaluate the capacity of some of these mutations to either create or eliminate restriction sites using the wild-type strain C. Abortus S26/3 as a reference. Three of eight genomic sites with confirmed point mutations (CAB153, CAB636 and CAB648) were retained for analysis as each resulted in the loss of a restriction site in the genome sequence of the vaccine strain. PCR-restriction fragment length polymorphism analysis using restriction enzymes chosen to specifically target the three genomic sites was then applied to a large number of C. Abortus field isolates and reference strains. Our results indicate that the three mutations are uniquely present in the vaccine strain, and as such provide easy-to-use markers for the differential identification of the vaccine strain and wild-type isolates.
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corrigendum to experimental infection of pregnant ewes with enteric and abortion source Chlamydophila Abortus vet microbiol 82 2001 285 291
Veterinary Microbiology, 2002Co-Authors: P Tsakos, Victoria I Siarkou, Evangelia Vretou, F Guscetti, E H Chowdhury, N Papaioannou, O PapadopoulosAbstract:Corrigendum to ‘‘Experimental infection of pregnant ewes with enteric and abortion-source Chlamydophila Abortus’’ [Vet. Microbiol. 82 (2001) 285–291] P. Tsakos, V. Siarkou, F. Guscetti, E.H. Chowdhury, N. Papaioannou, E. Vretou, O. Papadopoulos Faculty of Veterinary Medicine, Laboratory of Microbiology and Infectious Diseases, Aristotle University, 54 006 Thessaloniki, Greece Institute of Veterinary Pathology, University of Zurich, CH-8057 Zurich, Switzerland Faculty of Veterinary Medicine, Laboratory of Pathology, Aristotle University, 54 006 Thessaloniki, Greece Department of Biotechnology, Hellenic Pasteur Institute, 115 21 Athens, Greece
Evangelia Vretou - One of the best experts on this subject based on the ideXlab platform.
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genome sequence of the Chlamydophila Abortus variant strain llg
Journal of Bacteriology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Evangelia Vretou, David Smith, Alex F Lainson, David LongbottomAbstract:Chlamydophila Abortus is a common cause of ruminant abortion. Here we report the genome sequence of strain LLG, which differs genotypically and phenotypically from the wild-type strain S26/3. Genome sequencing revealed differences between LLG and S26/3 to occur in pseudogene content, in transmembrane head/inc family proteins, and in biotin biosynthesis genes.
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genetic variability of Chlamydophila Abortus strains assessed by pcr rflp analysis of polymorphic membrane protein encoding genes
Veterinary Microbiology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Konrad Sachse, Simone Magnino, B K Markey, Evangelia VretouAbstract:This study used PCR-RFLP to investigate the genetic variability of pmp-encoding genes from fifty-two Chlamydophila Abortus (C. Abortus) strains originating from abortion cases from various geographical regions and host species. Six primer pairs were used to PCR-amplify DNA fragments encoding eighteen pmps. PCR products were digested using four restriction endonucleases and Bayesian methodologies were used to compare RFLP profiles and assign strains to a RFLP genotype. Strains could be assigned to 2 genotypes in the region encoding pmp18D, 3 genotypes in the regions encoding pmp1A-pmp2B, pmp3E-pmp6H and pmp11G-pmp15G, 4 genotypes in the region encoding pmp7G-pmp10G and 5 genotypes in the region encoding pmp16G-pmp17G. In all regions, the majority of strains (88.4-96.1%) had the same genotype as the reference strain S26/3. No correlation could be made between genotype, host species or geographical origin except for the two variant Greek strains, LLG and POS, which formed a discrete genotype in all pmp-encoding regions except pmp18D. Relative rates of evolution calculated for each pmp-encoding gene locus suggest that differing selective pressures and functional constraints may exist on C. Abortus polymorphic membrane proteins. These findings suggest that although intraspecies heterogeneity of pmp-encoding genes in C. Abortus is low, the sequence heterogeneity should be an important consideration when using pmps as the basis for novel diagnostics or vaccine development.
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differential identification of Chlamydophila Abortus live vaccine strain 1b and c Abortus field isolates by pcr rflp
Vaccine, 2010Co-Authors: Karine Laroucau, Victoria I Siarkou, Evangelia Vretou, Annie Rodolakis, Fabien Vorimore, Konrad Sachse, Hermann Willems, Simone Magnino, Patrik M BavoilAbstract:Comparative genomic analysis of a wild-type strain of the ovine pathogen Chlamydophila Abortus and its nitrosoguanidine-induced, temperature-sensitive and virulence-attenuated live vaccine derivative identified point mutations unique to the mutant (Burall et al. [1]). Here, we evaluate the capacity of some of these mutations to either create or eliminate restriction sites using the wild-type strain C. Abortus S26/3 as a reference. Three of eight genomic sites with confirmed point mutations (CAB153, CAB636 and CAB648) were retained for analysis as each resulted in the loss of a restriction site in the genome sequence of the vaccine strain. PCR-restriction fragment length polymorphism analysis using restriction enzymes chosen to specifically target the three genomic sites was then applied to a large number of C. Abortus field isolates and reference strains. Our results indicate that the three mutations are uniquely present in the vaccine strain, and as such provide easy-to-use markers for the differential identification of the vaccine strain and wild-type isolates.
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identification and characterization of inc766 an inclusion membrane protein in Chlamydophila Abortus infected cells
Microbial Pathogenesis, 2008Co-Authors: Evangelia Vretou, Evgenia Psarrou, Evangelia Katsiki, Kostantinos Vougas, George Th TsangarisAbstract:Abstract We have identified the gene product of locus 766 in the transmembrane head region (TMH/Inc-region) in the Chlamydophila Abortus genome by using mass spectrometry and a monoclonal antibody that reacted with the inclusion membrane. The identified protein at 32 kDa, termed Inc766, formed highly stable oligomers when solubilized in the absence of β-mercaptoethanol. These oligomers were resistant to SDS, to heat denaturation and to 8 M urea, but very sensitive to β-mercaptoethanol, consistent with conformations resulting from protein–protein interactions stabilized through disulphide bonds. Mass spectrometry analysis of immunoprecipitated infected cell lysates indicated that a dimer at 56 kDa was the most prominent form in solution. Cross-linking with DSP provided supporting evidence for the formation of oligomers in situ . Inc766 was expressed at 20–24 h post infection and its localization pattern in the extra-inclusion space was common in all C. Abortus strains tested. Taken together, Inc766 displays unique biochemical and cellular features not encountered in other Incs from other Chlamydiaceae species. Future studies of the particular characteristics especially the interactive properties of Inc766 should contribute to our understanding of the relationship of the different chlamydial species with their respective hosts.
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evaluation of two commercial assays for the detection of Chlamydophila Abortus antibodies
Veterinary Microbiology, 2007Co-Authors: Evangelia Vretou, F Radouani, E Psarrou, I Kritikos, E Xylouri, O ManganaAbstract:Abstract Two commercial enzyme-linked immunosorbent assays (ELISA), the CHEKIT ® -CHLAMYDIA which uses inactivated Chlamydophila psittaci antigen, and the Chlamydophila Abortus ELISA produced by the Institut Pourquier which uses a recombinant fragment of the 80–90 kDa protein, were evaluated with the objective to determine whether the new ELISAs would perform as improved alternatives to the complement fixation test (CFT) for the serological diagnosis of ovine enzootic abortion (OEA). The results were compared to those obtained by the CFT and the competitive ELISA (cELISA). The tests were assessed with a panel of 17 serum samples from specific pathogen-free (SPF) lambs experimentally infected with various subtypes of Chlamydophila pecorum , with sera from 45 C. Abortus -infected pregnant sheep and from 54 sheep free of OEA. The C. Abortus ELISA was identified as being more specific and sensitive than the other tests. The 4 assays were evaluated further with 254 sera from flocks with documented OEA, from flocks with no history of abortion and from animals after abortion of unknown cause. The C. Abortus ELISA by the Institut Pourquier identified less OEA-positive sera than the other assays though it identified correctly 9 of 10 OEA-positive flocks. The basis of the discordant results is discussed.
Nick Wheelhouse - One of the best experts on this subject based on the ideXlab platform.
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genome sequence of the Chlamydophila Abortus variant strain llg
Journal of Bacteriology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Evangelia Vretou, David Smith, Alex F Lainson, David LongbottomAbstract:Chlamydophila Abortus is a common cause of ruminant abortion. Here we report the genome sequence of strain LLG, which differs genotypically and phenotypically from the wild-type strain S26/3. Genome sequencing revealed differences between LLG and S26/3 to occur in pseudogene content, in transmembrane head/inc family proteins, and in biotin biosynthesis genes.
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genetic variability of Chlamydophila Abortus strains assessed by pcr rflp analysis of polymorphic membrane protein encoding genes
Veterinary Microbiology, 2011Co-Authors: Michelle Sait, Ewan M Clark, Nick Wheelhouse, Morag Livingstone, Lucy Spalding, Victoria I Siarkou, Konrad Sachse, Simone Magnino, B K Markey, Evangelia VretouAbstract:This study used PCR-RFLP to investigate the genetic variability of pmp-encoding genes from fifty-two Chlamydophila Abortus (C. Abortus) strains originating from abortion cases from various geographical regions and host species. Six primer pairs were used to PCR-amplify DNA fragments encoding eighteen pmps. PCR products were digested using four restriction endonucleases and Bayesian methodologies were used to compare RFLP profiles and assign strains to a RFLP genotype. Strains could be assigned to 2 genotypes in the region encoding pmp18D, 3 genotypes in the regions encoding pmp1A-pmp2B, pmp3E-pmp6H and pmp11G-pmp15G, 4 genotypes in the region encoding pmp7G-pmp10G and 5 genotypes in the region encoding pmp16G-pmp17G. In all regions, the majority of strains (88.4-96.1%) had the same genotype as the reference strain S26/3. No correlation could be made between genotype, host species or geographical origin except for the two variant Greek strains, LLG and POS, which formed a discrete genotype in all pmp-encoding regions except pmp18D. Relative rates of evolution calculated for each pmp-encoding gene locus suggest that differing selective pressures and functional constraints may exist on C. Abortus polymorphic membrane proteins. These findings suggest that although intraspecies heterogeneity of pmp-encoding genes in C. Abortus is low, the sequence heterogeneity should be an important consideration when using pmps as the basis for novel diagnostics or vaccine development.
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identification sequencing and molecular analysis of chp4 a novel chlamydiaphage of Chlamydophila Abortus belonging to the family microviridae
Journal of General Virology, 2011Co-Authors: Michelle Sait, Nick Wheelhouse, Morag Livingstone, Rebecca Graham, Neil F Inglis, David LongbottomAbstract:Members of the family Microviridae have been identified in a number of chlamydial species infecting humans (phage CPAR39 in Chlamydophila pneumoniae), other mammals (φCPG1 in Chlamydophila caviae, Chp2 in Chlamydophila Abortus and Chp3 in Chlamydophila pecorum) and birds (Chp1 in Chlamydophila psittaci). This study describes the identification and genome sequencing of Chp4, an icosahedral, 4530 bp, ssDNA phage in C. Abortus. Chp4 is predicted to contain eight ORFs, six of which could be assigned putative functions based on sequence similarity to characterized bacteriophage. Gene order and content were highly conserved amongst chlamydiaphage, with the highest sequence variability occurring in the IN5 and INS variable regions of the VP1 major coat protein, which has been associated with host cell recognition and binding. Phylogenetic analysis of VP1 indicated that Chp4 is a member of the Chlamydiamicrovirus, and is most closely related to phage φCPG1 and CPAR39.
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inflammatory cytokine responses in a pregnant mouse model of Chlamydophila Abortus infection
Veterinary Microbiology, 2010Co-Authors: Karen P Kerr, Nick Wheelhouse, Morag Livingstone, Gary Entrican, Declan J Mckeever, Ian E Anderson, David LongbottomAbstract:Abstract Chlamydophila Abortus ( C. Abortus ) is the aetiological agent of ovine enzootic abortion (OEA). The highly elevated expression of the pro-inflammatory cytokine tumour necrosis factor-alpha (TNFα) and low-level expression of interferon-gamma (IFNγ) that are detected in C. Abortus- infected placentas have been implicated in the pathogenesis of OEA. Late-term abortions similar to those occurring in sheep have also been observed in mouse models of C. Abortus infection. Since mouse studies have contributed significantly to our understanding of the immunological responses to chlamydial infections and serve as a good model for rapidly assessing candidate vaccines for OEA, we investigated local expression of TNFα and IFNγ in infected mice. At various time points over the course of infection mice were sacrificed, serum samples obtained for serum antibody and cytokine analyses, and livers and placental tissues were removed and fixed to determine C. Abortus colonisation and cytokine expression. Immunostaining for C. Abortus was significantly greater in placenta compared to liver ( P C. Abortus infection but not placental/fetal infection in mice. In contrast to sheep, murine placental TNFα expression does not correlate with C. Abortus infection, suggesting that the immunopathogenesis of chlamydial abortion differs in these species.
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evidence of Chlamydophila Abortus vaccine strain 1b as a possible cause of ovine enzootic abortion
Vaccine, 2010Co-Authors: Nick Wheelhouse, Kevin Aitchison, Karine Laroucau, Jill R Thomson, David LongbottomAbstract:Chlamydophila Abortus, the agent of ovine enzootic abortion (OEA), is a major cause of lamb mortality worldwide. Disease can be controlled through the use of vaccines based on the 1B temperature-sensitive mutant strain of C. Abortus. This study investigated suspected OEA cases across Scotland for the presence of the 1B strain by analysis of recently identified unique point mutations (9). Thirty-five cases were C. Abortus-positive and 14 came from vaccinated flocks. Analysis of single nucleotide polymorphisms by PCR-RFLP and sequence analysis revealed the presence of point mutations consistent with the presence of the 1B vaccine strain in 5 of these 14 samples. Quantitative real-time PCR revealed comparable numbers of genome copies of the 1B strain in infected placentas to those present following wild-type infection. This study is the first to demonstrate the presence of the 1B vaccine strain in the placentas of OEA cases and suggests a probable causal role in the disease.