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.
Background
Three horse mares inadvertently inseminated with semen from a Tayorella asinigenitalis‐positive Jack donkey developed severe, purulent endometritis whereas two Jenny donkeys mated ...naturally to the same Jack donkey did not develop clinical signs of infection.
Objectives
To isolate and identify the causative agent.
Study design
Case report.
Methods
Endometrial swabs from the infected mares were cultured on selective and non‐selective media under aerobic and microaerophilic conditions. Isolates were subjected to Gram staining, oxidase and catalase tests, the Monotayl Latex Agglutination test and PCR to test for both T. equigenitalis and T. asinigenitalis. In vitro antimicrobial susceptibility testing was performed and the bacterial isolate was genotyped using MLST.
Results
A new sequence type of T. asinigenitalis was confirmed.
Main limitations
A limited numbers of mares and donkeys are described.
Conclusions
This strain of T. asinigenitalis causes a severe venereal infection in mares but not in Jenny donkeys.
In Finland, Taylorella equigenitalis, the causative agent of contagious equine metritis (CEM), was first detected in 1992. The aim of this study was to genotype Finnish T. equigenitalis isolates to ...investigate the epidemiology of the infection in the Finnish horse population. A total of 34 T. equigenitalis isolates from 24 horses obtained during 1992–2021 were subjected to whole genome sequencing (WGS) and subsequent local ad hoc core genome multi-locus sequence typing (cgMLST) targeting 1259 loci. Classical MLST profiles were extracted from the whole-genome sequence data. Three novel MLST types, ST81, ST82 and ST83, and four previously described sequence types, ST16, ST17, ST50 and ST63 were detected among the isolates. cgMLST minimum spanning tree analysis using 12 allele difference as threshold, resulted in five clusters and three singletons. cgMLST clusters were congruent with the MLST-defined groups, except for the ST83 isolates which were divided into two clusters. However, the high discriminatory power cgMLST allowed differentiation between isolates of the same MLST type as each isolate had a unique core genome ST. Our study suggests that cgMLST has the prospective for being a standardised typing method for T. equigenitalis in the future, and further contributes to worldwide phylogenetic and spatio-temporal analyses needed to better understand the epidemiology of the bacterium.
•A new ad-hoc cgMLST scheme for T. equigenitalis was developed•The 34 isolates were divided into 34 cgMLST sequence types•Isolates with the same MLST type mainly clustered together in the cgMLST analysis•Isolates from 24 horses fell into 3 novel and 4 previously described MLST types•New sequence data from isolates from Finnhorses and Fjord horses are provided
Taylorella equigenitalis causes contagious equine metritis. Here we compared seven nucleic acid amplification tests for T. equigenitalis to select a rapid and reliable diagnostic method. The 95% ...detection limits of each assay varied greatly: real-time PCR had the lowest detection limit (0.77 fg/reaction); those of some of the conventional PCRs (cPCRs) were >100 fg/reaction. In experimentally infected samples, real-time PCR and semi-nested PCR showed the highest positive numbers (33 out of 42 samples), but two of the cPCRs detected only 2 and 7 positive results. Our results indicate that the use of sensitive molecular assays is important for the efficient detection of T. equigenitalis in clinical samples.
•Proficiency tests have been organized since 2017 in France for CEM detection by PCR.•Five PCR and 3 DNA extraction methods were used during these first 5 PTs.•PTs evaluated the performance of ...approved laboratories in the network in France.•Results highlight an efficient network composed of 20 approved laboratories in 2021.•Lowest Ct values were observed with direct lysis compared to column extraction.
Contagious equine metritis (CEM) detection by PCR is recognized by the European Union according to Commission Implementing Regulation (EU) No 846/2014, and real-time PCR is now recommended by the World Organisation for Animal Health Terrestrial Manual at the same level as the culture method. The present study highlights the creation of an efficient network of approved laboratories in France in 2017 for CEM detection by real-time PCR. The network currently consists of 20 laboratories. A first proficiency test (PT) was organized by the national reference laboratory for CEM in 2017 to evaluate the performance of the early network, followed by annual proficiency tests organized for ongoing periodic assessment of network performance. Results of the 5 PTs organized from 2017 to 2021 are presented, during which 5 real-time PCRs and 3 DNA extraction methods were used. Overall, 99.20% of the qualitative data corresponded to expected results and the R-squared of global DNA amplification calculated for each PT varied from 0.728 to 0.899. DNA extraction is also an important step in the analytical process, and results were more favorable with direct lysis compared to column extraction. Focusing on the most commonly used PCR (PCR 1: 86.4% of results) showed lowest cycle threshold values with direct lysis compared to column and magnetic bead extractions, and with magnetic bead extraction compared to column extraction, but neither of these differences were statistically significant.
Taylorella equigenitalis has long been recognised as a causative agent of contagious equine metritis, but practitioners may be less familiar with Taylorella asinigenitalis, which has been identified ...more recently. Here, Abel Dorrego, Consuelo Serres and Fatima Cruz‐Lopez of the Universidad Complutense de Madrid describe T asinigenitalis and report the findings of a survey they carried out in donkeys in Spain.
Taylorella equigenitalis causes contagious equine metritis. Here we compared seven nucleic acid amplification tests for T. equigenitalis to select a rapid and reliable diagnostic method. The 95% ...detection limits of each assay varied greatly: real-time PCR had the lowest detection limit (0.77 fg/reaction); those of some of the conventional PCRs (cPCRs) were >100 fg/reaction. In experimentally infected samples, real-time PCR and semi-nested PCR showed the highest positive numbers (33 out of 42 samples), but two of the cPCRs detected only 2 and 7 positive results. Our results indicate that the use of sensitive molecular assays is important for the efficient detection of T. equigenitalis in clinical samples.
We report herein the use of crude extracts obtained from samples of Taylorella equigenitalis–infected horses for the purpose of multi-locus sequence typing (MLST). Samples (n = 36) were collected ...from horses in South Africa from 1996 to 2017: 34 from genital swabs (stored at −20°C for 2–3 y) and 2 from cryopreserved raw semen aliquots (stored at −70°C for 18 y) prior to assay. The MLST assay showed a single sequence type (ST), designated ST4, that supported a point introduction and thus a common source for the South African outbreak of contagious equine metritis.
Taylorella asinigenitalis is a non-pathogenic bacteria isolated from the genital tract of donkeys but also a cause of metritis and vaginal discharge in mares. It is closely related to Taylorella ...equigenitalis, the cause of Contagious Equine Metritis (CEM) in horses, and has been present in different countries in Europe since 1995. Up to date, there are no studies on the prevalence of T. asinigenitalis in the equine or asinine populations in Spain; this is the first report of the presence of T. asinigenitalis in donkeys (Equus asinus) from different breeds in three regions of Spain. A total of 106 healthy animals of three different Spanish donkey breeds: Andaluza (26), Majorera (12) and Zamorano-Leonés (68) were sampled between June and July 2017 and a real-time PCR was used to detect T. asinigenitalis in all samples. A total of 39/221 (17,65 %) samples from 22/106 (20,75 %) animals yielded a positive result and were further characterized by MLST; an allelic profile and Sequence Type (ST) could be assigned to 11 of the 39 positive samples, resulting in four novel STs and no clonal complexes within the PubMLST database. There were statistically significant differences in the percentage of positive animals by breed and sex, and also in the variability of STs between farms. Breeding management would have an influence on the percentage of positives in a farm; artificial insemination and separating jacks from jennies should be implemented. Further studies to detect and characterize T. asinigenitalis in donkeys and horses from Spain would be required to obtain a broader epidemiological picture in this country.
•This is the first report of Taylorella asinigenitalis in donkeys residing in Spain.•There were differences in T. asinigenitalis positivity rates by premises and sex.•The breeding management would affect the percentage of positives in a premises.•A higher bacterial load was found in males compared to females.•All the STs assigned in our study were new and related to STs found in France.
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