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Joachim Frey - One of the best experts on this subject based on the ideXlab platform.
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vaccination of cattle with the n terminus of lppq of mycoplasma mycoides subsp mycoides results in type iii immune complex disease upon experimental infection
Infection and Immunity, 2015Co-Authors: Musa Mulongo, Hezron Wesonga, Jan Naessens, Joachim Frey, K C Smith, Christian Schnier, Declan J MckeeverAbstract:Contagious bovine pleuropneumonia (CBPP) is a serious respiratory disease of cattle caused by Mycoplasma mycoides subsp. mycoides. Current vaccines against CBPP induce short-lived immunity and can cause severe postvaccine reactions. Previous studies have identified the N terminus of the transmembrane lipoprotein Q (LppQ-N′) of M. mycoides subsp. mycoides as the major antigen and a possible virulence factor. We therefore immunized cattle with purified recombinant LppQ-N′ formulated in Freund's adjuvant and challenged them with M. mycoides subsp. mycoides. Vaccinated animals showed a strong seroconversion to LppQ, but they exhibited significantly enhanced postchallenge glomerulonephritis compared to the placebo group (P = 0.021). Glomerulonephritis was characterized by features that suggested the development of antigen-antibody immune complexes. Clinical signs and gross pathological scores did not significantly differ between vaccinated and placebo groups. These findings reveal for the first time the pathogenesis of enhanced disease as a result of antibodies against LppQ during challenge and also argue against inclusion of LppQ-N′ in a future subunit vaccine for CBPP.
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cyto adherence of mycoplasma mycoides subsp mycoides to bovine lung epithelial cells
BMC Veterinary Research, 2015Co-Authors: Martin Mwirigi, Paola Pilo, Joerg Jores, Joachim Frey, Jan NaessensAbstract:Background Mycoplasma mycoides subsp. mycoides (Mmm) is the causative agent of contagious bovine pleuropneumonia (CBPP), a respiratory disease of cattle, whereas the closely related Mycoplasma mycoides subsp. capri (Mmc) is a goat pathogen. Cyto-adherence is a crucial step in host colonization by mycoplasmas and subsequent pathogenesis. The aim of this study was to investigate the interactions between Mmm and mammalian host cells by establishing a cyto-adherence flow cytometric assay and comparing tissue and species specificity of Mmm and Mmc strains.
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the origin of the mycoplasma mycoides cluster coincides with domestication of ruminants
PLOS ONE, 2012Co-Authors: Anne Fischer, Edy M Vilei, Beth Shapiro, Cecilia Muriuki, Martin Heller, Christiane Schnee, Joachim Frey, Erik Bongcamrudloff, Joerg JoresAbstract:The 'Mycoplasma mycoides cluster' comprises the ruminant pathogens Mycoplasma mycoides subsp. mycoides the causative agent of contagious bovine pleuropneumonia (CBPP), Mycoplasma capricolum subsp. capripneumoniae the agent of contagious caprine pleuropneumonia (CCPP), Mycoplasma capricolum subsp. capricolum, Mycoplasma leachii and Mycoplasma mycoides subsp. capri. CBPP and CCPP are major livestock diseases and impact the agricultural sector especially in developing countries through reduced food-supply and international trade restrictions. In addition, these diseases are a threat to disease-free countries. We used a multilocus sequence typing (MLST) approach to gain insights into the demographic history of and phylogenetic relationships among the members of the 'M. mycoides cluster'. We collected partial sequences from seven housekeeping genes representing a total of 3,816 base pairs from 118 strains within this cluster, and five strains isolated from wild Caprinae. Strikingly, the origin of the 'M. mycoides cluster' dates to about 10,000 years ago, suggesting that the establishment and spread of the cluster coincided with livestock domestication. In addition, we show that hybridization and recombination may be important factors in the evolutionary history of the cluster.
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detection of mycoplasma mycoides subsp mycoides sc in bronchoalveolar lavage fluids of cows based on a taqman real time pcr discriminating wild type strains from an lppq mutant vaccine strain used for diva strategies
Journal of Microbiological Methods, 2010Co-Authors: Edy M Vilei, Joachim FreyAbstract:Contagious bovine pleuropneumonia (CBPP) is the most serious cattle disease in Africa, caused by Mycoplasma mycoides subsp. mycoides small-colony type (SC). CBPP control strategies currently rely on vaccination with a vaccine based on live attenuated strains of the organism. Recently, an lppQ− mutant of the existing vaccine strain T1/44 has been developed (Janis et al., 2008). This T1lppQ− mutant strain is devoid of lipoprotein LppQ, a potential virulence attribute of M. mycoides subsp. mycoides SC. It is designated as a potential live DIVA (Differentiating Infected from Vaccinated Animals) vaccine strain allowing both serological and etiological differentiation. The present paper reports on the validation of a control strategy for CBPP in cattle, whereby a TaqMan real-time PCR based on the lppQ gene has been developed for the direct detection of M. mycoides subsp. mycoides SC in ex vivo bronchoalveolar lavage fluids of cows and for the discrimination of wild type strains from the lppQ− mutant vaccine strain.
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functional and antigenic properties of glpo from mycoplasma mycoides subsp mycoides sc characterization of a flavin adenine dinucleotide binding site deletion mutant
Veterinary Research, 2009Co-Authors: Daniela F Bischof, Edy M Vilei, Joachim FreyAbstract:L-a-glycerophosphate oxidase (GlpO) plays a central role in virulence of Mycoplasma mycoides subsp. mycoides SC, a severe bacterial pathogen causing contagious bovine pleuropneumonia (CBPP). It is involved in production and translocation of toxic H2O2 into the host cell, causing inflammation and cell death. The binding site on GlpO for the cofactor flavin adenine dinucleotide (FAD) has been identified as Gly12Gly13Gly14Ile15Ile16Gly17. Recombinant GlpO lacking these six amino acids (GlpODFAD) was unable to bind FAD and was also devoid of glycerophosphate oxidase activity, in contrast to non-modified recombinant GlpO that binds FAD and is enzymatically active. Polyclonal monospecific antibodies directed against GlpODFAD, similarly to anti-GlpO antibodies, neutralised H2O2 production of M. mycoides subsp. mycoides SC grown in the presence of glycerol, as well as cytotoxicity towards embryonic calf nasal epithelial (ECaNEp) cells. The FAD-binding site of GlpO is therefore suggested as a valuable target site for the future construction of deletion mutants to yield attenuated live vaccines of M. mycoides subsp. mycoides SC necessary to efficiently combat CBPP. CBPP / L-a-glycerophosphate oxidase / FAD-binding site / targeted deletion mutant / neutralizing antibody
Edy M Vilei - One of the best experts on this subject based on the ideXlab platform.
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the origin of the mycoplasma mycoides cluster coincides with domestication of ruminants
PLOS ONE, 2012Co-Authors: Anne Fischer, Edy M Vilei, Beth Shapiro, Cecilia Muriuki, Martin Heller, Christiane Schnee, Joachim Frey, Erik Bongcamrudloff, Joerg JoresAbstract:The 'Mycoplasma mycoides cluster' comprises the ruminant pathogens Mycoplasma mycoides subsp. mycoides the causative agent of contagious bovine pleuropneumonia (CBPP), Mycoplasma capricolum subsp. capripneumoniae the agent of contagious caprine pleuropneumonia (CCPP), Mycoplasma capricolum subsp. capricolum, Mycoplasma leachii and Mycoplasma mycoides subsp. capri. CBPP and CCPP are major livestock diseases and impact the agricultural sector especially in developing countries through reduced food-supply and international trade restrictions. In addition, these diseases are a threat to disease-free countries. We used a multilocus sequence typing (MLST) approach to gain insights into the demographic history of and phylogenetic relationships among the members of the 'M. mycoides cluster'. We collected partial sequences from seven housekeeping genes representing a total of 3,816 base pairs from 118 strains within this cluster, and five strains isolated from wild Caprinae. Strikingly, the origin of the 'M. mycoides cluster' dates to about 10,000 years ago, suggesting that the establishment and spread of the cluster coincided with livestock domestication. In addition, we show that hybridization and recombination may be important factors in the evolutionary history of the cluster.
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detection of mycoplasma mycoides subsp mycoides sc in bronchoalveolar lavage fluids of cows based on a taqman real time pcr discriminating wild type strains from an lppq mutant vaccine strain used for diva strategies
Journal of Microbiological Methods, 2010Co-Authors: Edy M Vilei, Joachim FreyAbstract:Contagious bovine pleuropneumonia (CBPP) is the most serious cattle disease in Africa, caused by Mycoplasma mycoides subsp. mycoides small-colony type (SC). CBPP control strategies currently rely on vaccination with a vaccine based on live attenuated strains of the organism. Recently, an lppQ− mutant of the existing vaccine strain T1/44 has been developed (Janis et al., 2008). This T1lppQ− mutant strain is devoid of lipoprotein LppQ, a potential virulence attribute of M. mycoides subsp. mycoides SC. It is designated as a potential live DIVA (Differentiating Infected from Vaccinated Animals) vaccine strain allowing both serological and etiological differentiation. The present paper reports on the validation of a control strategy for CBPP in cattle, whereby a TaqMan real-time PCR based on the lppQ gene has been developed for the direct detection of M. mycoides subsp. mycoides SC in ex vivo bronchoalveolar lavage fluids of cows and for the discrimination of wild type strains from the lppQ− mutant vaccine strain.
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functional and antigenic properties of glpo from mycoplasma mycoides subsp mycoides sc characterization of a flavin adenine dinucleotide binding site deletion mutant
Veterinary Research, 2009Co-Authors: Daniela F Bischof, Edy M Vilei, Joachim FreyAbstract:L-a-glycerophosphate oxidase (GlpO) plays a central role in virulence of Mycoplasma mycoides subsp. mycoides SC, a severe bacterial pathogen causing contagious bovine pleuropneumonia (CBPP). It is involved in production and translocation of toxic H2O2 into the host cell, causing inflammation and cell death. The binding site on GlpO for the cofactor flavin adenine dinucleotide (FAD) has been identified as Gly12Gly13Gly14Ile15Ile16Gly17. Recombinant GlpO lacking these six amino acids (GlpODFAD) was unable to bind FAD and was also devoid of glycerophosphate oxidase activity, in contrast to non-modified recombinant GlpO that binds FAD and is enzymatically active. Polyclonal monospecific antibodies directed against GlpODFAD, similarly to anti-GlpO antibodies, neutralised H2O2 production of M. mycoides subsp. mycoides SC grown in the presence of glycerol, as well as cytotoxicity towards embryonic calf nasal epithelial (ECaNEp) cells. The FAD-binding site of GlpO is therefore suggested as a valuable target site for the future construction of deletion mutants to yield attenuated live vaccines of M. mycoides subsp. mycoides SC necessary to efficiently combat CBPP. CBPP / L-a-glycerophosphate oxidase / FAD-binding site / targeted deletion mutant / neutralizing antibody
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mycoplasma leachii sp nov as a new species designation for mycoplasma sp bovine group 7 of leach and reclassification of mycoplasma mycoides subsp mycoides lc as a serovar of mycoplasma mycoides subsp capri
International Journal of Systematic and Evolutionary Microbiology, 2009Co-Authors: Lucia Mansosilvan, Edy M Vilei, Francois Thiaucourt, Konrad Sachse, Steven P. Djordjevic, Joachim FreyAbstract:The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. Mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii
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Domain analysis of lipoprotein LppQ in Mycoplasma mycoides subsp. mycoides SC
Antonie van Leeuwenhoek, 2008Co-Authors: Laetitia Bonvin-klotz, Edy M Vilei, Joachim Frey, Kathrin Kühni-boghenbor, Nadine Kapp, Michael H. StoffelAbstract:The lipoprotein LppQ is the most prominent antigen of Mycoplasma mycoides subsp. mycoides small colony type (SC) during infection of cattle. This pathogen causes contagious bovine pleuropneumonia (CBPP), a devastating disease of considerable socio-economic importance in many countries worldwide. The dominant antigenicity and high specificity for M. mycoides subsp . mycoides SC of lipoprotein LppQ have been exploited for serological diagnosis and for epidemiological investigations of CBPP. Scanning electron microscopy and immunogold labelling were used to provide ultrastructural evidence that LppQ is located to the cell membrane at the outer surface of M. mycoides subsp . mycoides SC. The selectivity and specificity of this method were demonstrated through discriminating localization of extracellular (i.e., in the zone of contact with host cells) vs. integral membrane domains of LppQ. Thus, our findings support the suggestion that the accessible N-terminal domain of LppQ is surface exposed and such surface localization may be implicated in the pathogenesis of CBPP.
Francois Thiaucourt - One of the best experts on this subject based on the ideXlab platform.
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highly dynamic genomic loci drive the synthesis of two types of capsular or secreted polysaccharides within the mycoplasma mycoides cluster
Applied and Environmental Microbiology, 2015Co-Authors: Clothilde Bertin, Corinne Pauroblot, Josiane Courtois, Florence Tardy, F Poumarat, Pascal Sirandpugnet, Patrice Gaurivaud, Lucia Mansosilvan, Christine Citti, Francois ThiaucourtAbstract:Mycoplasmas of the Mycoplasma mycoides cluster are all ruminant pathogens. Mycoplasma mycoides subsp. mycoides is responsible for contagious bovine pleuropneumonia and is known to produce capsular polysaccharide (CPS) and exopolysaccharide (EPS). Previous studies have strongly suggested a role for Mycoplasma mycoides subsp. mycoides polysaccharides in pathogenicity. Mycoplasma mycoides subsp. mycoides-secreted EPS was recently characterized as a β(1→6)-galactofuranose homopolymer (galactan) identical to the capsular product. Here, we extended the characterization of secreted polysaccharides to all other members of the M. mycoides cluster: M. capricolum subsp. capripneumoniae, M. capricolum subsp. capricolum, M. leachii, and M. mycoides subsp. capri (including the LC and Capri serovars). Extracted EPS was characterized by nuclear magnetic resonance, resulting in the identification of a homopolymer of β(1→2)-glucopyranose (glucan) in M. capricolum subsp. capripneumoniae and M. leachii. Monoclonal antibodies specific for this glucan and for the Mycoplasma mycoides subsp. mycoides-secreted galactan were used to detect the two polysaccharides. While M. mycoides subsp. capri strains of serovar LC produced only capsular galactan, no polysaccharide could be detected in strains of serovar Capri. All strains of M. capricolum subsp. capripneumoniae and M. leachii produced glucan CPS and EPS, whereas glucan production and localization varied among M. capricolum subsp. capricolum strains. Genes associated with polysaccharide synthesis and forming a biosynthetic pathway were predicted in all cluster members. These genes were organized in clusters within two loci representing genetic variability hot spots. Phylogenetic analysis showed that some of these genes, notably galE and glf, were acquired via horizontal gene transfer. These findings call for a reassessment of the specificity of the serological tests based on mycoplasma polysaccharides.
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mycoplasma mycoides from mycoides small colony to capri a microevolutionary perspective
BMC Genomics, 2011Co-Authors: Francois Thiaucourt, Woubit Salah, Benoit Vacherie, Anne Marie Lomenech, Valerie Barbe, Virginie Dupuy, Marc Breton, Lucia Mansosilvan, Daniel Jacob, Alain BlanchardAbstract:Background The Mycoplasma mycoides cluster consists of five species or subspecies that are ruminant pathogens. One subspecies, Mycoplasma mycoides subspecies mycoides Small Colony (MmmSC), is the causative agent of contagious bovine pleuropneumonia. Its very close relative, Mycoplasma mycoides subsp. capri (Mmc), is a more ubiquitous pathogen in small ruminants causing mastitis, arthritis, keratitis, pneumonia and septicaemia and is also found as saprophyte in the ear canal. To understand the genetics underlying these phenotypic differences, we compared the MmmSC PG1 type strain genome, which was already available, with the genome of an Mmc field strain (95010) that was sequenced in this study. We also compared the 95010 genome with the recently published genome of another Mmc strain (GM12) to evaluate Mmc strain diversity.
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mycoplasma leachii sp nov as a new species designation for mycoplasma sp bovine group 7 of leach and reclassification of mycoplasma mycoides subsp mycoides lc as a serovar of mycoplasma mycoides subsp capri
International Journal of Systematic and Evolutionary Microbiology, 2009Co-Authors: Lucia Mansosilvan, Edy M Vilei, Francois Thiaucourt, Konrad Sachse, Steven P. Djordjevic, Joachim FreyAbstract:The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. Mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii
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real time polymerase chain reaction assays for the detection of members of the mycoplasma mycoides cluster
New Zealand Veterinary Journal, 2008Co-Authors: J Fitzmaurice, Lucia Mansosilvan, Francois Thiaucourt, M Sewell, W L Mcdonald, J S OkeefeAbstract:Abstract AIM: To develop real-time PCR assays for the detection and differentiation of members of the Mycoplasma mycoides cluster. METHODS: Five real-time PCR assays were designed to allow differentiation of members of the M. mycoides cluster: an assay for detection of the M. mycoides subspecies, viz M. mycoides subsp mycoides large colony (MmmLC), M. mycoides subsp capri (Mmc), and M. mycoides subsp mycoides small colony (MmmSC); one for the detection of the M. capricolum subspecies, viz M. capricolum subsp capricolum (Mcc), M. capricolum subsp capripneumoniae (Mccp), and Mycoplasma sp bovine group 7 (BG7); and three for the specifi c detection of MmmSC, Mccp, and BG7. A panel of 74 Mycoplasma isolates from various geographical origins and a panel of 21 other bacterial isolates were used to evaluate the sensitivity and specificity of the assays. RESULTS: The assays displayed 100% analytical sensitivity in detecting all target Mycoplasma isolates. The analytical detection limit for the assays to detect th...
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real time polymerase chain reaction assays for the detection of memebers of the mycoplasma mycoides cluster
New Zealand Veterinary Journal, 2008Co-Authors: J Fitzmaurice, Lucia Mansosilvan, Francois Thiaucourt, M Sewell, W L Mcdonald, J S OkeefeAbstract:AIM: To develop real-time PCR assays for the detection and differentiation of members of the Mycoplasma mycoides cluster. METHODS: Five real-time PCR assays were designed to al-low differentiation of members of the M mycoides cluster: an assay for detection of the M mycoides subspecies, viz M mycoides subsp mycoides large colony (MmmLC), M mycoides subsp capri (Mmc), and M mycoides subsp mycoides small colony (MmmSC); one for the detection of the M capricolum subspecies, viz M capricolum subsp capricolum (Mcc), M capricolum subsp capripneumoniae (Mccp), and Mycoplasma sp bovine group 7 (BG7); and three for the specific detection of MmmSC, Mccp, and BG7. A panel of 74 Mycoplasma isolates from various geographical origins and a panel of 21 other bacterial isolates were used to evaluate the sensitivity and specificity of the as-says. RESULTS: The assays displayed 100% analytical sensitivity in detecting all target Mycoplasma isolates. The analytical detection limit for the assays to detect the M mycoides subspecies, M capricolum subspecies, and MmmSC was determined to be 100 fg of genomic DNA, while the Mccp and BG7 assays had a detection limit of 100 fg and 10 fg of genomic DNA, respectively. The M mycoides subspecies assay had a detection limit of 103 (SD 102) cfu/ml milk, 104 (SD 104) cfu per swab, and 103 (SD 103) cfu/g lung in inoculated samples. The assays displayed 100% specificity when applied to non-target bacterial isolates and to 110 culture-negative milk samples. CONCLUSIONS: The assays were highly sensitive and specific, and provide accurate detection and differentiation of the members of the M mycoides cluster. (Resume d'auteur)
Lucia Mansosilvan - One of the best experts on this subject based on the ideXlab platform.
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highly dynamic genomic loci drive the synthesis of two types of capsular or secreted polysaccharides within the mycoplasma mycoides cluster
Applied and Environmental Microbiology, 2015Co-Authors: Clothilde Bertin, Corinne Pauroblot, Josiane Courtois, Florence Tardy, F Poumarat, Pascal Sirandpugnet, Patrice Gaurivaud, Lucia Mansosilvan, Christine Citti, Francois ThiaucourtAbstract:Mycoplasmas of the Mycoplasma mycoides cluster are all ruminant pathogens. Mycoplasma mycoides subsp. mycoides is responsible for contagious bovine pleuropneumonia and is known to produce capsular polysaccharide (CPS) and exopolysaccharide (EPS). Previous studies have strongly suggested a role for Mycoplasma mycoides subsp. mycoides polysaccharides in pathogenicity. Mycoplasma mycoides subsp. mycoides-secreted EPS was recently characterized as a β(1→6)-galactofuranose homopolymer (galactan) identical to the capsular product. Here, we extended the characterization of secreted polysaccharides to all other members of the M. mycoides cluster: M. capricolum subsp. capripneumoniae, M. capricolum subsp. capricolum, M. leachii, and M. mycoides subsp. capri (including the LC and Capri serovars). Extracted EPS was characterized by nuclear magnetic resonance, resulting in the identification of a homopolymer of β(1→2)-glucopyranose (glucan) in M. capricolum subsp. capripneumoniae and M. leachii. Monoclonal antibodies specific for this glucan and for the Mycoplasma mycoides subsp. mycoides-secreted galactan were used to detect the two polysaccharides. While M. mycoides subsp. capri strains of serovar LC produced only capsular galactan, no polysaccharide could be detected in strains of serovar Capri. All strains of M. capricolum subsp. capripneumoniae and M. leachii produced glucan CPS and EPS, whereas glucan production and localization varied among M. capricolum subsp. capricolum strains. Genes associated with polysaccharide synthesis and forming a biosynthetic pathway were predicted in all cluster members. These genes were organized in clusters within two loci representing genetic variability hot spots. Phylogenetic analysis showed that some of these genes, notably galE and glf, were acquired via horizontal gene transfer. These findings call for a reassessment of the specificity of the serological tests based on mycoplasma polysaccharides.
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mycoplasma mycoides from mycoides small colony to capri a microevolutionary perspective
BMC Genomics, 2011Co-Authors: Francois Thiaucourt, Woubit Salah, Benoit Vacherie, Anne Marie Lomenech, Valerie Barbe, Virginie Dupuy, Marc Breton, Lucia Mansosilvan, Daniel Jacob, Alain BlanchardAbstract:Background The Mycoplasma mycoides cluster consists of five species or subspecies that are ruminant pathogens. One subspecies, Mycoplasma mycoides subspecies mycoides Small Colony (MmmSC), is the causative agent of contagious bovine pleuropneumonia. Its very close relative, Mycoplasma mycoides subsp. capri (Mmc), is a more ubiquitous pathogen in small ruminants causing mastitis, arthritis, keratitis, pneumonia and septicaemia and is also found as saprophyte in the ear canal. To understand the genetics underlying these phenotypic differences, we compared the MmmSC PG1 type strain genome, which was already available, with the genome of an Mmc field strain (95010) that was sequenced in this study. We also compared the 95010 genome with the recently published genome of another Mmc strain (GM12) to evaluate Mmc strain diversity.
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mycoplasma leachii sp nov as a new species designation for mycoplasma sp bovine group 7 of leach and reclassification of mycoplasma mycoides subsp mycoides lc as a serovar of mycoplasma mycoides subsp capri
International Journal of Systematic and Evolutionary Microbiology, 2009Co-Authors: Lucia Mansosilvan, Edy M Vilei, Francois Thiaucourt, Konrad Sachse, Steven P. Djordjevic, Joachim FreyAbstract:The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. Mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii
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real time polymerase chain reaction assays for the detection of members of the mycoplasma mycoides cluster
New Zealand Veterinary Journal, 2008Co-Authors: J Fitzmaurice, Lucia Mansosilvan, Francois Thiaucourt, M Sewell, W L Mcdonald, J S OkeefeAbstract:Abstract AIM: To develop real-time PCR assays for the detection and differentiation of members of the Mycoplasma mycoides cluster. METHODS: Five real-time PCR assays were designed to allow differentiation of members of the M. mycoides cluster: an assay for detection of the M. mycoides subspecies, viz M. mycoides subsp mycoides large colony (MmmLC), M. mycoides subsp capri (Mmc), and M. mycoides subsp mycoides small colony (MmmSC); one for the detection of the M. capricolum subspecies, viz M. capricolum subsp capricolum (Mcc), M. capricolum subsp capripneumoniae (Mccp), and Mycoplasma sp bovine group 7 (BG7); and three for the specifi c detection of MmmSC, Mccp, and BG7. A panel of 74 Mycoplasma isolates from various geographical origins and a panel of 21 other bacterial isolates were used to evaluate the sensitivity and specificity of the assays. RESULTS: The assays displayed 100% analytical sensitivity in detecting all target Mycoplasma isolates. The analytical detection limit for the assays to detect th...
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real time polymerase chain reaction assays for the detection of memebers of the mycoplasma mycoides cluster
New Zealand Veterinary Journal, 2008Co-Authors: J Fitzmaurice, Lucia Mansosilvan, Francois Thiaucourt, M Sewell, W L Mcdonald, J S OkeefeAbstract:AIM: To develop real-time PCR assays for the detection and differentiation of members of the Mycoplasma mycoides cluster. METHODS: Five real-time PCR assays were designed to al-low differentiation of members of the M mycoides cluster: an assay for detection of the M mycoides subspecies, viz M mycoides subsp mycoides large colony (MmmLC), M mycoides subsp capri (Mmc), and M mycoides subsp mycoides small colony (MmmSC); one for the detection of the M capricolum subspecies, viz M capricolum subsp capricolum (Mcc), M capricolum subsp capripneumoniae (Mccp), and Mycoplasma sp bovine group 7 (BG7); and three for the specific detection of MmmSC, Mccp, and BG7. A panel of 74 Mycoplasma isolates from various geographical origins and a panel of 21 other bacterial isolates were used to evaluate the sensitivity and specificity of the as-says. RESULTS: The assays displayed 100% analytical sensitivity in detecting all target Mycoplasma isolates. The analytical detection limit for the assays to detect the M mycoides subspecies, M capricolum subspecies, and MmmSC was determined to be 100 fg of genomic DNA, while the Mccp and BG7 assays had a detection limit of 100 fg and 10 fg of genomic DNA, respectively. The M mycoides subspecies assay had a detection limit of 103 (SD 102) cfu/ml milk, 104 (SD 104) cfu per swab, and 103 (SD 103) cfu/g lung in inoculated samples. The assays displayed 100% specificity when applied to non-target bacterial isolates and to 110 culture-negative milk samples. CONCLUSIONS: The assays were highly sensitive and specific, and provide accurate detection and differentiation of the members of the M mycoides cluster. (Resume d'auteur)
Anja Persson - One of the best experts on this subject based on the ideXlab platform.
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protein specific analysis of humoral immune responses in a clinical trial for vaccines against contagious bovine pleuropneumonia
Clinical and Vaccine Immunology, 2010Co-Authors: Carl Hamsten, Massimo Scacchia, Roger D Ayling, Otto J B Huebschle, Georgina Tjipurazaire, Laura Mcauliffe, Anja PerssonAbstract:Specific humoral immune responses in a clinical trial on cattle for vaccines against contagious bovine pleuropneumonia (CBPP) were investigated. The trial included a subunit vaccine consisting of five recombinant putative variable surface proteins of the infectious agent Mycoplasma mycoides subsp. mycoides small colony type (M. mycoides SC) compared to the currently approved attenuated vaccine strain T1/44 and untreated controls. Humoral immune responses to 65 individual recombinant surface proteins of M. mycoides SC were monitored by a recently developed bead-based array assay. Responses to the subunit vaccine components were found to be weak. Animals vaccinated with this vaccine were not protected and had CBPP lesions similar to those of the untreated controls. In correlating protein-specific humoral responses to T1/44-induced immunity, five proteins associated with a protective immune response were identified by statistical evaluation, namely, MSC_1046 (LppQ), MSC_0271, MSC_0136, MSC_0079, and MSC_0431. These five proteins may be important candidates in the development of a novel subunit vaccine against CBPP.
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recombinant surface proteomics as a tool to analyze humoral immune responses in bovines infected by mycoplasma mycoides subsp mycoides small colony type
Molecular & Cellular Proteomics, 2009Co-Authors: Carl Hamsten, John B March, Maja Neiman, Jochen M Schwenk, Marica Hamsten, Anja PerssonAbstract:A systematic approach to characterize the surface proteome of Mycoplasma mycoides subspecies mycoides small colony type (M. mycoides SC), the causative agent of contagious bovine pleuropneumonia (CBPP) in cattle, is presented. Humoral immune responses in 242 CBPP-affected cattle and controls were monitored against one-third of the surface proteins of M. mycoides SC in a high throughput magnetic bead-based assay. Initially, 64 surface proteins were selected from the genome sequence of M. mycoides SC and expressed as recombinant proteins in Escherichia coli. Binding of antibodies to each individual protein could then be analyzed simultaneously in minute sample volumes with the Luminex suspension array technology. The assay was optimized on Namibian CBPP-positive sera and Swedish negative controls to allow detection and 20-fold mean signal separation between CBPP-positive and -negative sera. Signals were proven to be protein-specific by inhibition experiments, and results agreed with Western blot experiments. The potential of the assay to monitor IgG, IgM, and IgA responses over time was shown in a proof-of-concept study with 116 sera from eight animals in a CBPP vaccine study. In conclusion, a toolbox with recombinant proteins and a flexible suspension array assay that allows multiplex analysis of humoral immune responses to M. mycoides SC has been created.
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diagnosis of contagious bovine pleuropneumonia by pcr laser induced fluorescence and pcr restriction endonuclease analysis based on the 16s rrna genes of mycoplasma mycoides subsp mycoides sc
Journal of Clinical Microbiology, 1999Co-Authors: Anja Persson, Bertil Pettersson, Goran Bolske, Karlerik JohanssonAbstract:As contagious bovine pleuropneumonia (CBPP) is spreading fast in many African countries, there is an increasing demand for rapid and sensitive diagnostic methods that can be used to confirm the initial diagnosis based on clinical symptoms or pathological findings. Two PCR-based diagnostic systems for identification of the infectious agent, Mycoplasma mycoides subsp. mycoides SC (M. mycoides SC), in various samples are presented. Both systems involve group-specific amplification of the two 16S rRNA genes from mycoplasmas of the M. mycoides cluster. The laser-induced fluorescence assay is based on a unique sequence length difference between the two 16S rRNA genes in M. mycoides SC. This region was amplified by PCR, and the products were separated by polyacrylamide gel electrophoresis in a DNA sequencer. The resulting electropherogram showed two peaks for strains of M. mycoides SC and one peak for all other members of the M. mycoides cluster. The second system was based on restriction endonuclease analysis and agarose gel electrophoresis. Restriction of amplicons from a region containing a polymorphism, which is found in M. mycoides SC only, resulted in an extra band on the agarose gel because an AluI site is lacking in the rrnA operon. Specimens from cows with postmortem signs of CBPP were analyzed with the two PCR systems. M. mycoides SC was clearly identified in pleural fluid and lung tissue, and the methods were found to be robust and rapid. The results were in agreement with those obtained by conventional diagnostic techniques.
