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John E. Moore - One of the best experts on this subject based on the ideXlab platform.
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Development of a novel molecular detection method for clustered regularly interspaced short palindromic repeats (CRISPRs) in Taylorella organisms.
Journal of medical microbiology, 2015Co-Authors: Yasushi Hara, Sandrine Petry, Motoo Matsuda, T. Nakajima, Shizuko Kagawa, Marie Akamatsu, Motoki Yahiro, John E. MooreAbstract:Contagious equine metritis is a bacterial infectious disease of horses caused by Taylorella equigenitalis, a Gram-negative eubacterium. The disease has been described in several continents, including Europe, North America and Asia. A novel molecular method was developed to detect clustered regularly interspaced short palindromic repeats (CRISPRs), which were separated by non-repetitive unique spacer regions (NRUSRs) of similar length, in the Taylorella equigenitalis EQ59 strain using a primer pair, f-/r-TeCRISPR-ladder, by PCR amplification. In total, 31 Taylorella isolates (17 T. equigenitalis and 14 Taylorella asinigenitalis) were examined. The T. equigenitalis isolates came from thoroughbred and cold-blooded horses from nine countries during 1980–1996, whilst the T. asinigenitalis isolates all originated from donkey jacks in France and the USA during 1997–2006. PAGE fractionated all of the 13 CRISPRs separated by 12 NRUSRs in T. equigenitalis EQ59. Permutation examples of CRISPRs, which were separated by NRUSRs for small-sized ladders, consisting of two doublet bands were shown. Putative CRISPRs separated by NRUSRs were amplified with 14/17 (82.4 %) geographically disparate T. equigenitalis isolates using the newly designed primer pair. Approximately 82.4 % of the T. equigenitalis isolates had CRISPRs separated by NRUSRs. The CRISPR locus was also found in the French T. asinigenitalis strain MCE3. Putative CRISPRs separated by NRUSRs were detected similarly in 4/14 (28.6 %) T. asinigenitalis isolates. Overall, a more detailed understanding of the molecular biology of CRISPRs within Taylorella organisms may help elucidate the pathogenic virulence and transmission mechanisms associated with this important equine pathogen.
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Molecular identification and characterization of clustered regularly interspaced short palindromic repeat (CRISPR) gene cluster in Taylorella equigenitalis
Folia Microbiologica, 2013Co-Authors: Yasushi Hara, John E. Moore, Kyohei Hayashi, Takuya Nakajima, Shizuko Kagawa, Akihiro Tazumi, Motoo MatsudaAbstract:Clustered regularly interspaced short palindromic repeats (CRISPRs), of approximately 10,000 base pairs (bp) in length, were shown to occur in the Japanese Taylorella equigenitalis strain, EQ59. The locus was composed of the putative CRISPRs-associated with 5 ( cas5 ), RAMP csd1 , csd2 , recB , cas1 , a leader region, 13 CRISPR consensus sequence repeats (each 32 bp; 5′-TCAGCCACGTTCGCGTGGCTGTGTGTTTAAAG-3′). These were in turn separated by 12 non repetitive unique spacer regions of similar length. In addition, a leader region, a transposase/IS protein, a leader region, and cas3 were also seen. All seven putative open reading frames carry their ribosome binding sites. Promoter consensus sequences at the −35 and −10 regions and putative intrinsic ρ-independent transcription terminator regions also occurred. A possible long overlap of 170 bp in length occurred between the recB and cas1 loci. Positive reverse transcription PCR signals of cas5 , RAMP csd1 , csd2 - recB / cas1 , and cas3 were generated. A putative secondary structure of the CRISPR consensus repeats was constructed. Following this, CRISPR results of the T. equigenitalis EQ59 isolate were subsequently compared with those from the Taylorella asinigenitalis MCE3 isolate.
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Demonstration of the absence of intervening sequences (IVSs) within 16S rRNA genes of Taylorella equigenitalis and Taylorella asinigenitalis isolates.
Research in veterinary science, 2011Co-Authors: A. Tazumi, Sandrine Petry, John E. Moore, S. Nakanishi, K. Hayashi, Erina Tasaki, T. Nakajima, Hitomi Ueno, Beverley C. Millar, Motoo MatsudaAbstract:Abstract A total of 57 Taylorella equigenitalis ( n = 22) and Taylorella asinigenitalis ( n = 35) isolates was shown not to carry any intervening sequences (IVSs) within 16S rRNA gene sequences. By contrast, we have already shown the genus Taylorella group to carry several kinds of IVSs within the 23S rRNA gene sequences.
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Development of a new molecular detection method for Taylorella equigenitalis.
Journal of basic microbiology, 2011Co-Authors: A. Tazumi, Sandrine Petry, John E. Moore, K. Hayashi, B. Cherie Millar, Junichi Hirayama, Motoo MatsudaAbstract:On PCR amplification of the intervening sequences (IVSs) in the central (helix 45) region within 23S rRNA gene sequences with T. equigenitalis (n = 34), as well as T. asinigenitalis (n = 35) and Bordetella (n = 11) isolates by using the primer pair of f-/r-23STis2, approximately 0.8 kb of the amplicons were generated, sequenced and analyzed. One IVS of approximately 70 bp in length was identified in all the Taylorella organisms but not Bordetella. PCR amplification was further developed for the convenient and rapid molecular detection of T. equigenitalis organisms with the IVS in the helix 45 region within the 23S rRNA genes as target by using the primer pairs (f-IVSde/r-23de). Thus, these results clearly demonstrated that PCR amplification with the primer pair (f-IVSde/r-23de) can be reliable in order to differentiate the T. equigenitalis isolates from both the T. asinigenitalis and Bordetella organisms. (© 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
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Homogeneity of the 16S rDNA sequence among geographically disparate isolates of Taylorella equigenitalis.
BMC veterinary research, 2006Co-Authors: Motoo Matsuda, A. Tazumi, S. Kagawa, Tsuyoshi Sekizuka, Ohoshi Murayama, John E. Moore, B. C. MillarAbstract:Background At present, six accessible sequences of 16S rDNA from Taylorella equigenitalis (T. equigenitalis) are available, whose sequence differences occur at a few nucleotide positions. Thus it is important to determine these sequences from additional strains in other countries, if possible, in order to clarify any anomalies regarding 16S rDNA sequence heterogeneity. Here, we clone and sequence the approximate full-length 16S rDNA from additional strains of T. equigenitalis isolated in Japan, Australia and France and compare these sequences to the existing published sequences.
Sandrine Petry - One of the best experts on this subject based on the ideXlab platform.
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Acute Endometritis due to Taylorella equigenitalis Transmission by Insemination of Cryopreserved Stallion Semen.
Journal of equine veterinary science, 2019Co-Authors: Marie Delerue, Marie-france Breuil, Fabien Duquesne, Marie-hélène Bayon-auboyer, Nadia Amenna-bernard, Sandrine PetryAbstract:Taylorella equigenitalis can be transmitted during artificial insemination. This report describes clinical T. equigenitalis transmission by cryopreserved stallion semen. T. equigenitalis isolates from a mare's vaginal discharge and semen from the same batch of the cryopreserved semen used for the insemination gave identical API ZYM, antibiotic susceptibility, and multilocus sequence typing results (ST-46); furthermore, the multilocus sequence typing lineage ST-46 is known to circulate in the country of semen collection. These results support the need for strict contagious equine metritis screening of processed semen before use for artificial insemination.
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Evaluation of MALDI-TOF MS and an expanded custom reference spectra database for the identification and differentiation of Taylorella equigenitalis and Taylorella asinigenitalis.
Diagnostic microbiology and infectious disease, 2019Co-Authors: Sandrine Petry, Fabien Duquesne, Marie-hélène Bayon-auboyer, Amandine Wilhelm, Hervé Morvan, Benoit GassilloudAbstract:Abstract Misidentification between Taylorella equigenitalis, the causative agent of contagious equine metritis (CEM), and Taylorella asinigenitalis is observed by the gold standard culture method. The performance of matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS) for Taylorella species identification was evaluated using 85 T. equigenitalis and 28 T. asinigenitalis strains selected on the basis of multilocus sequence typing data. Seven of the T. equigenitalis and 9 of the T. asinigenitalis strains were used to generate in-house reference spectra to expand the existing commercial Bruker database. Two bacterial incubation times and 3 different sample preparation procedures were compared. Overall, we demonstrated the usefulness of MALDI-TOF MS as a differential diagnostic tool for CEM; however, commercial spectra databases should be expanded with T. asinigenitalis reference spectra to achieve the expected performance. Moreover, direct spotting of 48-h colonies was not only the most efficient protocol but also the easiest to implement in a clinical setting.
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Towards European harmonisation of contagious equine metritis diagnosis through interlaboratory trials
The Veterinary record, 2018Co-Authors: Sandrine Petry, Marie-france Breuil, Fabien Duquesne, Claire LaugierAbstract:The performance of culture and PCR methods routinely used to diagnose contagious equine metritis (CEM) was evaluated and compared by two interlaboratory trials involving a total of 24 European laboratories, including 22 National Reference Laboratories for CEM. Samples were swab specimens artificially contaminated with bacteria present in the genital tract of Equidae, some with and some without Taylorella equigenitalis, the causative agent of CEM, and T asinigenitalis, responsible for possible misidentification as T equigenitalis Throughout both interlaboratory trials, PCR performed better in terms of specificity and sensitivity than the culture method, supporting the assertion that PCR should be accepted for CEM diagnosis. However, the culture performance during the second interlaboratory trial was better than during the first one, suggesting that the expertise of participants improved. This reveals the advantage of regular interlaboratory trials to constantly improve the expertise of laboratories. It also shows the need to develop new culture media that are more selective and/or better geared to the metabolism of T equigenitalis in order to improve the bacteriological diagnosis of CEM.
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Contagious equine metritis cases reported in France since 2006
The Veterinary record, 2015Co-Authors: Marie-france Breuil, Fabien Duquesne, Claire Laugier, E. Leperchois, B. Ferry, G. Collin, Sandrine PetryAbstract:CONTAGIOUS equine metritis (CEM) is a sexually transmitted disease of equids caused by Taylorella equigenitalis , a Gram-negative microaerophilic coccobacillus of the Taylorella genus. The acute form of the disease is characterised by mucopurulent vaginal discharge and variable degrees of vaginitis, endometritis and cervicitis, leading to temporary infertility (Timoney 2011). It is assumed that disease dissemination results from the transfer of carrier stallions and mares, but is also linked to artificial breeding (Schulman and others 2013). After its discovery in 1977, CEM spread rapidly worldwide, causing international concern within the breeding horse industry (Matsuda and Moore 2003; Timoney 2011). It became one of the most regulated equine diseases and is a notifiable disease of the World Organisation for Animal Health. Despite this, many countries are still considered to have endemic status for CEM within their non-Thoroughbred populations (Schulman and others 2013), probably due to several factors, for example, the absence or the shortcomings of national monitoring, surveillance and reporting programmes, and lack of knowledge about the ecology of T equigenitalis . In France, CEM was reported every year until February 2012, except in the Thoroughbred population which has been CEM free since 2006. No cases have been reported since then, although an increase in the prevalence of the disease was expected since the national regulations (with the exception of artificial breeding) were replaced in 2006 by a voluntary professional system. In this context, the aim of …
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Development of a novel molecular detection method for clustered regularly interspaced short palindromic repeats (CRISPRs) in Taylorella organisms.
Journal of medical microbiology, 2015Co-Authors: Yasushi Hara, Sandrine Petry, Motoo Matsuda, T. Nakajima, Shizuko Kagawa, Marie Akamatsu, Motoki Yahiro, John E. MooreAbstract:Contagious equine metritis is a bacterial infectious disease of horses caused by Taylorella equigenitalis, a Gram-negative eubacterium. The disease has been described in several continents, including Europe, North America and Asia. A novel molecular method was developed to detect clustered regularly interspaced short palindromic repeats (CRISPRs), which were separated by non-repetitive unique spacer regions (NRUSRs) of similar length, in the Taylorella equigenitalis EQ59 strain using a primer pair, f-/r-TeCRISPR-ladder, by PCR amplification. In total, 31 Taylorella isolates (17 T. equigenitalis and 14 Taylorella asinigenitalis) were examined. The T. equigenitalis isolates came from thoroughbred and cold-blooded horses from nine countries during 1980–1996, whilst the T. asinigenitalis isolates all originated from donkey jacks in France and the USA during 1997–2006. PAGE fractionated all of the 13 CRISPRs separated by 12 NRUSRs in T. equigenitalis EQ59. Permutation examples of CRISPRs, which were separated by NRUSRs for small-sized ladders, consisting of two doublet bands were shown. Putative CRISPRs separated by NRUSRs were amplified with 14/17 (82.4 %) geographically disparate T. equigenitalis isolates using the newly designed primer pair. Approximately 82.4 % of the T. equigenitalis isolates had CRISPRs separated by NRUSRs. The CRISPR locus was also found in the French T. asinigenitalis strain MCE3. Putative CRISPRs separated by NRUSRs were detected similarly in 4/14 (28.6 %) T. asinigenitalis isolates. Overall, a more detailed understanding of the molecular biology of CRISPRs within Taylorella organisms may help elucidate the pathogenic virulence and transmission mechanisms associated with this important equine pathogen.
Motoo Matsuda - One of the best experts on this subject based on the ideXlab platform.
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Development of a novel molecular detection method for clustered regularly interspaced short palindromic repeats (CRISPRs) in Taylorella organisms.
Journal of medical microbiology, 2015Co-Authors: Yasushi Hara, Sandrine Petry, Motoo Matsuda, T. Nakajima, Shizuko Kagawa, Marie Akamatsu, Motoki Yahiro, John E. MooreAbstract:Contagious equine metritis is a bacterial infectious disease of horses caused by Taylorella equigenitalis, a Gram-negative eubacterium. The disease has been described in several continents, including Europe, North America and Asia. A novel molecular method was developed to detect clustered regularly interspaced short palindromic repeats (CRISPRs), which were separated by non-repetitive unique spacer regions (NRUSRs) of similar length, in the Taylorella equigenitalis EQ59 strain using a primer pair, f-/r-TeCRISPR-ladder, by PCR amplification. In total, 31 Taylorella isolates (17 T. equigenitalis and 14 Taylorella asinigenitalis) were examined. The T. equigenitalis isolates came from thoroughbred and cold-blooded horses from nine countries during 1980–1996, whilst the T. asinigenitalis isolates all originated from donkey jacks in France and the USA during 1997–2006. PAGE fractionated all of the 13 CRISPRs separated by 12 NRUSRs in T. equigenitalis EQ59. Permutation examples of CRISPRs, which were separated by NRUSRs for small-sized ladders, consisting of two doublet bands were shown. Putative CRISPRs separated by NRUSRs were amplified with 14/17 (82.4 %) geographically disparate T. equigenitalis isolates using the newly designed primer pair. Approximately 82.4 % of the T. equigenitalis isolates had CRISPRs separated by NRUSRs. The CRISPR locus was also found in the French T. asinigenitalis strain MCE3. Putative CRISPRs separated by NRUSRs were detected similarly in 4/14 (28.6 %) T. asinigenitalis isolates. Overall, a more detailed understanding of the molecular biology of CRISPRs within Taylorella organisms may help elucidate the pathogenic virulence and transmission mechanisms associated with this important equine pathogen.
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Development of a single multi-locus sequence typing scheme for Taylorella equigenitalis and Taylorella asinigenitalis.
Veterinary Microbiology, 2013Co-Authors: Fabien Duquesne, Laurent Hébert, Marie-france Breuil, Claire Laugier, Motoo Matsuda, Sandrine PetryAbstract:Abstract We describe here the development of a multilocus sequence typing (MLST) scheme for Taylorella equigenitalis , the causative agent of contagious equine metritis (CEM), and Taylorella asinigenitalis , a nonpathogenic bacterium. MLST was performed on a set of 163 strains collected in several countries over 35 years (1977–2012). The MLST data were analyzed using START2, MEGA 5.05 and eBURST, and can be accessed at http://pubmlst.org/Taylorella/ . Our results revealed a clonal population with 39 sequence types (ST) and no common ST between the two Taylorella species. The eBURST analysis grouped the 27 T. equigenitalis STs into four clonal complexes (CC1–4) and five unlinked STs. The 12 T. asinigenitalis STs were grouped into three clonal complexes (CC5–7) and five unlinked STs, among which CC1 (68.1% of the 113 T. equigenitalis ) and CC5 (58.0% of the 50 T. asinigenitalis ) were dominants. The CC1, still in circulation in France, contains isolates from the first CEM outbreaks that simultaneously emerged in several countries in the late 1970s. The emergence in different countries (e.g. France, Japan, and United Arab Emirates) of STs without any genetic relationship to CC1 suggests the existence of a natural worldwide reservoir that remains to be identified. T. asinigenitalis appears to behave same way since the American, Swedish and French isolates have unrelated STs. This first Taylorella sp. MLST is a powerful tool for further epidemiological investigations and population biology studies of the Taylorella genus.
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Molecular identification and characterization of clustered regularly interspaced short palindromic repeat (CRISPR) gene cluster in Taylorella equigenitalis
Folia Microbiologica, 2013Co-Authors: Yasushi Hara, John E. Moore, Kyohei Hayashi, Takuya Nakajima, Shizuko Kagawa, Akihiro Tazumi, Motoo MatsudaAbstract:Clustered regularly interspaced short palindromic repeats (CRISPRs), of approximately 10,000 base pairs (bp) in length, were shown to occur in the Japanese Taylorella equigenitalis strain, EQ59. The locus was composed of the putative CRISPRs-associated with 5 ( cas5 ), RAMP csd1 , csd2 , recB , cas1 , a leader region, 13 CRISPR consensus sequence repeats (each 32 bp; 5′-TCAGCCACGTTCGCGTGGCTGTGTGTTTAAAG-3′). These were in turn separated by 12 non repetitive unique spacer regions of similar length. In addition, a leader region, a transposase/IS protein, a leader region, and cas3 were also seen. All seven putative open reading frames carry their ribosome binding sites. Promoter consensus sequences at the −35 and −10 regions and putative intrinsic ρ-independent transcription terminator regions also occurred. A possible long overlap of 170 bp in length occurred between the recB and cas1 loci. Positive reverse transcription PCR signals of cas5 , RAMP csd1 , csd2 - recB / cas1 , and cas3 were generated. A putative secondary structure of the CRISPR consensus repeats was constructed. Following this, CRISPR results of the T. equigenitalis EQ59 isolate were subsequently compared with those from the Taylorella asinigenitalis MCE3 isolate.
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Demonstration of the absence of intervening sequences (IVSs) within 16S rRNA genes of Taylorella equigenitalis and Taylorella asinigenitalis isolates.
Research in veterinary science, 2011Co-Authors: A. Tazumi, Sandrine Petry, John E. Moore, S. Nakanishi, K. Hayashi, Erina Tasaki, T. Nakajima, Hitomi Ueno, Beverley C. Millar, Motoo MatsudaAbstract:Abstract A total of 57 Taylorella equigenitalis ( n = 22) and Taylorella asinigenitalis ( n = 35) isolates was shown not to carry any intervening sequences (IVSs) within 16S rRNA gene sequences. By contrast, we have already shown the genus Taylorella group to carry several kinds of IVSs within the 23S rRNA gene sequences.
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Development of a new molecular detection method for Taylorella equigenitalis.
Journal of basic microbiology, 2011Co-Authors: A. Tazumi, Sandrine Petry, John E. Moore, K. Hayashi, B. Cherie Millar, Junichi Hirayama, Motoo MatsudaAbstract:On PCR amplification of the intervening sequences (IVSs) in the central (helix 45) region within 23S rRNA gene sequences with T. equigenitalis (n = 34), as well as T. asinigenitalis (n = 35) and Bordetella (n = 11) isolates by using the primer pair of f-/r-23STis2, approximately 0.8 kb of the amplicons were generated, sequenced and analyzed. One IVS of approximately 70 bp in length was identified in all the Taylorella organisms but not Bordetella. PCR amplification was further developed for the convenient and rapid molecular detection of T. equigenitalis organisms with the IVS in the helix 45 region within the 23S rRNA genes as target by using the primer pairs (f-IVSde/r-23de). Thus, these results clearly demonstrated that PCR amplification with the primer pair (f-IVSde/r-23de) can be reliable in order to differentiate the T. equigenitalis isolates from both the T. asinigenitalis and Bordetella organisms. (© 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
Fabien Duquesne - One of the best experts on this subject based on the ideXlab platform.
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Overview of spatio-temporal distribution inferred by multi-locus sequence typing of Taylorella equigenitalis isolated worldwide from 1977 to 2018 in equidae.
Veterinary microbiology, 2020Co-Authors: Fabien Duquesne, Falk Melzer, Marie-france Breuil, Aurélie Merlin, Iratxe Pérez-cobo, Kamil Sedlák, Gudrun Overesch, David Fretin, Wojciech Iwaniak, Ulrich WerneryAbstract:Abstract The accurate identification of Taylorella equigenitalis strains is essential to improve worldwide prevention and control strategies for contagious equine metritis (CEM). This study compared 367 worldwide equine strains using multilocus sequence typing according to the geographical origin, isolation year and equine breed. The strains were divided into 49 sequence types (STs), including 10 described for the first time. Three major and three minor clonal complexes (CCs), and 11 singletons, were identified. The genetic heterogeneity was low (0.13 STs/strain) despite the wide diversity of geographical origins (n = 16), isolation years (1977 to 2018) and equine breeds (n = 18). It was highest outside Europe and in the 1977-1997 period; current major STs and CCs already existed before 1998. Previous data associated the major CC1 with the first CEM outbreaks in 1977-1978 in the United Kingdom, Australia and the United States, and revealed its circulation in France. Our study confirms its circulation in France over a longer period of time (1992-2018) and its distribution in Spain and Germany but not throughout Europe. In addition to CC1, relationships between non-European and European countries were observed only through ST4, ST17 and ST30. Within Europe, several STs emerged with cross-border circulation, in particular ST16 and ST46 from the major complexes CC2 and CC8. These results constitute a baseline for monitoring the spread of CEM outbreaks. A retrospective analysis of a higher number of strains isolated worldwide between 1977 and the early 2000s would be helpful to obtain an exhaustive picture of the original CEM situation.
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Acute Endometritis due to Taylorella equigenitalis Transmission by Insemination of Cryopreserved Stallion Semen.
Journal of equine veterinary science, 2019Co-Authors: Marie Delerue, Marie-france Breuil, Fabien Duquesne, Marie-hélène Bayon-auboyer, Nadia Amenna-bernard, Sandrine PetryAbstract:Taylorella equigenitalis can be transmitted during artificial insemination. This report describes clinical T. equigenitalis transmission by cryopreserved stallion semen. T. equigenitalis isolates from a mare's vaginal discharge and semen from the same batch of the cryopreserved semen used for the insemination gave identical API ZYM, antibiotic susceptibility, and multilocus sequence typing results (ST-46); furthermore, the multilocus sequence typing lineage ST-46 is known to circulate in the country of semen collection. These results support the need for strict contagious equine metritis screening of processed semen before use for artificial insemination.
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Evaluation of MALDI-TOF MS and an expanded custom reference spectra database for the identification and differentiation of Taylorella equigenitalis and Taylorella asinigenitalis.
Diagnostic microbiology and infectious disease, 2019Co-Authors: Sandrine Petry, Fabien Duquesne, Marie-hélène Bayon-auboyer, Amandine Wilhelm, Hervé Morvan, Benoit GassilloudAbstract:Abstract Misidentification between Taylorella equigenitalis, the causative agent of contagious equine metritis (CEM), and Taylorella asinigenitalis is observed by the gold standard culture method. The performance of matrix-assisted laser desorption ionization–time-of-flight mass spectrometry (MALDI-TOF MS) for Taylorella species identification was evaluated using 85 T. equigenitalis and 28 T. asinigenitalis strains selected on the basis of multilocus sequence typing data. Seven of the T. equigenitalis and 9 of the T. asinigenitalis strains were used to generate in-house reference spectra to expand the existing commercial Bruker database. Two bacterial incubation times and 3 different sample preparation procedures were compared. Overall, we demonstrated the usefulness of MALDI-TOF MS as a differential diagnostic tool for CEM; however, commercial spectra databases should be expanded with T. asinigenitalis reference spectra to achieve the expected performance. Moreover, direct spotting of 48-h colonies was not only the most efficient protocol but also the easiest to implement in a clinical setting.
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Towards European harmonisation of contagious equine metritis diagnosis through interlaboratory trials
The Veterinary record, 2018Co-Authors: Sandrine Petry, Marie-france Breuil, Fabien Duquesne, Claire LaugierAbstract:The performance of culture and PCR methods routinely used to diagnose contagious equine metritis (CEM) was evaluated and compared by two interlaboratory trials involving a total of 24 European laboratories, including 22 National Reference Laboratories for CEM. Samples were swab specimens artificially contaminated with bacteria present in the genital tract of Equidae, some with and some without Taylorella equigenitalis, the causative agent of CEM, and T asinigenitalis, responsible for possible misidentification as T equigenitalis Throughout both interlaboratory trials, PCR performed better in terms of specificity and sensitivity than the culture method, supporting the assertion that PCR should be accepted for CEM diagnosis. However, the culture performance during the second interlaboratory trial was better than during the first one, suggesting that the expertise of participants improved. This reveals the advantage of regular interlaboratory trials to constantly improve the expertise of laboratories. It also shows the need to develop new culture media that are more selective and/or better geared to the metabolism of T equigenitalis in order to improve the bacteriological diagnosis of CEM.
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Contagious equine metritis cases reported in France since 2006
The Veterinary record, 2015Co-Authors: Marie-france Breuil, Fabien Duquesne, Claire Laugier, E. Leperchois, B. Ferry, G. Collin, Sandrine PetryAbstract:CONTAGIOUS equine metritis (CEM) is a sexually transmitted disease of equids caused by Taylorella equigenitalis , a Gram-negative microaerophilic coccobacillus of the Taylorella genus. The acute form of the disease is characterised by mucopurulent vaginal discharge and variable degrees of vaginitis, endometritis and cervicitis, leading to temporary infertility (Timoney 2011). It is assumed that disease dissemination results from the transfer of carrier stallions and mares, but is also linked to artificial breeding (Schulman and others 2013). After its discovery in 1977, CEM spread rapidly worldwide, causing international concern within the breeding horse industry (Matsuda and Moore 2003; Timoney 2011). It became one of the most regulated equine diseases and is a notifiable disease of the World Organisation for Animal Health. Despite this, many countries are still considered to have endemic status for CEM within their non-Thoroughbred populations (Schulman and others 2013), probably due to several factors, for example, the absence or the shortcomings of national monitoring, surveillance and reporting programmes, and lack of knowledge about the ecology of T equigenitalis . In France, CEM was reported every year until February 2012, except in the Thoroughbred population which has been CEM free since 2006. No cases have been reported since then, although an increase in the prevalence of the disease was expected since the national regulations (with the exception of artificial breeding) were replaced in 2006 by a voluntary professional system. In this context, the aim of …
S. W. Ricketts - One of the best experts on this subject based on the ideXlab platform.
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an investigation into the suitability of a commercial real time pcr assay to screen for Taylorella equigenitalis in routine prebreeding equine genital swabs
Equine Veterinary Journal, 2009Co-Authors: Jennifer C. Ousey, L. Palmer, R. Cash, K. J. Grimes, A. P. Fletcher, A. Barrelet, A. K. Foote, F. M. Manning, S. W. RickettsAbstract:Summary Reasons for performing study: Standard bacteriological methods for identifying Taylorella equigenitalis in cervical smears are time consuming. Therefore, a more rapid real-time PCR assay was evaluated for its suitability in screening swabs. Objective: To compare the results of a commercially available real-time PCR assay with routine microbiological culture for the identification of T. equigenitalis, the causative organism of contagious equine metritis, in equine genital swab samples, under ‘field trial’ conditions. Materials and methods: Routine prebreeding genital swabs (n = 2072) collected from Thoroughbred mares and stallions during 2009 were examined together with stored T. equigenitalis positive material. Swabs were cultured for T. equigenitalis using standard microbiological techniques. Bacterial lysates were isolated from the swabs and examined for the presence of a 16S DNA fragment of T. equigenitalis, using a commercial multiplex real-time PCR assay system. Results: There was complete concordance between positive and negative results obtained by the 2 methods. Real-time PCR also detected T. equigenitalis DNA from swabs that were negative using standard microbiological culture after 6 months' storage at +4°C but from which T. equigenitalis had been isolated following collection. The sensitivities of realtime PCR and bacterial culture were both 10−3 (equivalent to 3 colony-forming units). Conclusion and clinical relevance: Routine bacterial culture of T. equigenitalis requires an incubation period of not less than 7 days before a conclusive negative result can be obtained, whereas bacterial extraction and real-time PCR assay can be completed in less than 6 h. The commercially-available PCR assay tested provided a rapid and reliable method for the identification of T. equigenitalis from equine genital swabs and could be usefully employed for the screening of mares and stallions for preseason Horserace Betting Levy Board (HBLB) Code of Practice and in other situations such as for bloodstock sales screening requirements, overcoming the current delays imposed by bacterial culture requirements. Its use could be quality assured by the existing HBLB biannual testing scheme for designated laboratories.
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An investigation into the suitability of a commercial real‐time PCR assay to screen for Taylorella equigenitalis in routine prebreeding equine genital swabs
Equine veterinary journal, 2009Co-Authors: Jennifer C. Ousey, L. Palmer, R. Cash, K. J. Grimes, A. P. Fletcher, A. Barrelet, A. K. Foote, F. M. Manning, S. W. RickettsAbstract:Summary Reasons for performing study: Standard bacteriological methods for identifying Taylorella equigenitalis in cervical smears are time consuming. Therefore, a more rapid real-time PCR assay was evaluated for its suitability in screening swabs. Objective: To compare the results of a commercially available real-time PCR assay with routine microbiological culture for the identification of T. equigenitalis, the causative organism of contagious equine metritis, in equine genital swab samples, under ‘field trial’ conditions. Materials and methods: Routine prebreeding genital swabs (n = 2072) collected from Thoroughbred mares and stallions during 2009 were examined together with stored T. equigenitalis positive material. Swabs were cultured for T. equigenitalis using standard microbiological techniques. Bacterial lysates were isolated from the swabs and examined for the presence of a 16S DNA fragment of T. equigenitalis, using a commercial multiplex real-time PCR assay system. Results: There was complete concordance between positive and negative results obtained by the 2 methods. Real-time PCR also detected T. equigenitalis DNA from swabs that were negative using standard microbiological culture after 6 months' storage at +4°C but from which T. equigenitalis had been isolated following collection. The sensitivities of realtime PCR and bacterial culture were both 10−3 (equivalent to 3 colony-forming units). Conclusion and clinical relevance: Routine bacterial culture of T. equigenitalis requires an incubation period of not less than 7 days before a conclusive negative result can be obtained, whereas bacterial extraction and real-time PCR assay can be completed in less than 6 h. The commercially-available PCR assay tested provided a rapid and reliable method for the identification of T. equigenitalis from equine genital swabs and could be usefully employed for the screening of mares and stallions for preseason Horserace Betting Levy Board (HBLB) Code of Practice and in other situations such as for bloodstock sales screening requirements, overcoming the current delays imposed by bacterial culture requirements. Its use could be quality assured by the existing HBLB biannual testing scheme for designated laboratories.
