Genome-wide phylodynamic approach reveals the epidemic dynamics of the main Mycoplasma bovis subtype circulating in France.

Mycoplasma bovis is a major aetiological agent of bovine respiratory disease worldwide. Genome-based analyses are increasingly being used to monitor the genetic diversity and global distribution of M. bovis, complementing existing subtyping schemes based on locus sequencing. However, these analyses have so far provided limited information on the spatiotemporal and population dynamics of circulating subtypes. Here we applied a genome-wide phylodynamic approach to explore the epidemic dynamics of 88 French M. bovis strains collected between 2000 and 2019 in France and belonging to the currently dominant polC subtype 2 (st2). A strong molecular clock signal detected in the genomic data enabled robust phylodynamic inferences, which estimated that the M. bovis st2 population in France is composed of two lineages that successively emerged from independent introductions of international strains. The first lineage appeared around 2000 and supplanted the previously established antimicrobial-susceptible polC subtype 1. The second lineage, which is likely more transmissible, progressively replaced the first M. bovis st2 lineage population from 2005 onward and became predominant after 2010. Analyses also showed a brief decline in this second M. bovis st2 lineage population in around 2011, possibly due to the challenge from the concurrent emergence of M. bovis polC subtype 3 in France. Finally, we identified non-synonymous mutations in genes associated with lineages, which raises prospects for identifying new surveillance molecular markers. A genome-wide phylodynamic approach provides valuable resources for monitoring the evolution and epidemic dynamics of circulating M. bovis subtypes, and may prove critical for developing more effective surveillance systems and disease control strategies.

Molecular assessment of Theileria equi and Babesia caballi prevalence in horses and ticks on horses in southeastern France.

Equine piroplasmosis (EP) is a tick-borne disease caused by Babesia caballi and Theileria equi that is potentially emerging in non-endemic countries. We conducted a descriptive study to investigate EP prevalence and spatial distribution in an endemic region: the Camargue and the Plain of La Crau in France. In spring 2015 and 2016, we carried out sampling at stables (total n = 46) with a history of horses presenting chronic fever or weight loss. Overall, we collected blood from 632 horses, which were also inspected for ticks; these horses had been housed in the target stables for at least 1 year. We obtained 585 ticks from these horses and described land use around the stables. Real-time PCR was employed to assess T. equi and B. caballi prevalence in the horses and in the ticks found on the horses. For the horses, T. equi and B. caballi prevalence was 68.6% and 6.3%, respectively. For the ticks found on the horses, prevalence was 28.8% for T. equi and 0.85% for B. caballi. The most common tick species were, in order of frequency, Rhipicephalus bursa, R. sanguineus sl., Hyalomma marginatum, Haemaphysalis punctata, and Dermacentor sp. Horses bearing Rhipicephalus ticks occurred in wetter zones, closer to agricultural areas, permanent crops, and ditches, as well as in drier zones, in the more northern countryside. Compared to horses bearing R. bursa, horses bearing R. sanguineus sl. more frequently occurred near the Rhone River. Prevalence of T. equi in the ticks was as follows: Hyalomma marginatum (43%), Dermacentor sp. (40%), R. bursa (33%), R. sanguineus sl. (19%), and Haemaphysalis punctata (17%). In contrast, B. caballi only occurred in Dermacentor sp. (20%) and R. bursa (1%).

Questing tick abundance in urban and peri-urban parks in the French city of Lyon.

BACKGROUND: In Europe, ticks are responsible for the transmission of several pathogens of medical importance, including bacteria of the Borrelia burgdorferi (s.l.) complex, the agents of Lyme borreliosis. In France, the Auvergne Rhône-Alpes region is considered a hot spot for human tick-borne pathogen infections, with an estimated annual rate of 156 cases of Lyme borreliosis per 100,000 inhabitants. Although several studies have assessed the abundance of ticks in rural areas, little consideration has been given thus far to urban green spaces in France. METHODS: This study aimed to estimate tick abundance in three parks, two urban (U1, U2) and one peri-urban (PU), in and around the city of Lyon (France). A forest in a rural area was used as a control (C). Tick sampling campaigns were performed in each site in April, May, June, July, and October 2019 using the dragging method. One hundred transects of 10 m2 each were randomly chosen in each park in places frequented by humans. The sampling sessions were carried out under semi-controlled abiotic conditions. Ticks were stored in 70% ethanol and identified to species and developmental stage under a light microscope using morphological keys. RESULTS: A total of seven ticks (nymphs and adults) were collected in the two urban parks (six in U1 and one in U2), while 499 ticks were sampled in the peri-urban park. Of the 506 ticks collected, 504 were identified as Ixodes ricinus, one as Dermacentor marginatus, and one as Ixodes frontalis. In the peri-urban park, ticks were mainly collected under the forest cover and at forest edges. Tick density under forest cover was 7.1 times higher in the control site than in the peri-urban park throughout the survey period. CONCLUSIONS: This study confirmed the presence of ticks in all of the parks surveyed, although their occurrence in the urban parks was very rare compared to the peri-urban park and the control site. These results should serve as a basis for the implementation of preventive measures.

Swab cloths as a tool for revealing environmental contamination by Q fever in ruminant farms.

Q fever is a zoonotic abortive disease of ruminants mostly transmitted by inhalation of aerosols contaminated by Coxiella burnetii. Clusters of cases or even epidemics regularly occur in humans but, to date, there is no consensus about the best way to carry out outbreak investigations in order to identify potential farms at risk. Although environmental samples might be useful during such investigations, there are few baseline data on the presence of C. burnetii in the environment of ruminant farms. We thus investigated dust samples from cattle, sheep and goat farm buildings in order to (a) estimate C. burnetii detection frequency and bacterial loads in the environment, and (b) determine whether this environmental contamination is associated with series of abortions attributed to Q fever. We considered 113 herds with a recent abortive episode potentially related (n = 60) or not (n = 53) to C. burnetii. Dust was sampled using a swab cloth and tested by a quantitative PCR method targeting the IS1111 gene. Coxiella burnetii DNA was detected on 9 of 50 cattle farms, 13 of 19 goat farms and 30 of 40 sheep farms. On 16 cloths, bacterial loads were higher than 108 genome equivalents, levels as high as in infectious materials such as placentas and aborted foetuses. Overall, the probability of detecting C. burnetii DNA was higher on small ruminant farms than cattle farms, in herds suspected of Q fever and in large herds. We conclude that swab cloths are a putative indicator of contamination of ruminant farms by C. burnetii.

Molecular epidemiology of Coxiella burnetii in French livestock reveals the existence of three main genotype clusters and suggests species-specific associations as well as regional stability.

Q fever is a worldwide zoonosis caused by the bacterium Coxiella burnetii. In domestic ruminants, Q fever main clinical manifestations are abortions. Although the clinical signs may differ between ruminant species, C. burnetii's genetic diversity remains understudied in enzootic areas. Here, we focused on France, where Q fever is enzootic, with the aims to (a) identify potential associations between C. burnetii genotypes and ruminant host species; (b) assess the distribution of C. burnetii genotypes both within French farms and across France's major livestock-farming regions; and (c) suggest a subset of markers for future genotypic studies. We used DNA samples collected between 2006 and 2015 from 301 females (160 cows, 76 ewes, 65 goats) aborted of Q fever within 7 different farming regions. C. burnetii diversity was determined using a multiple-locus variable-number of tandem repeat analysis (MLVA) considering 17 markers. Using a phylogenetic approach, we identified 3 main genotypic clusters divided into 12 sub-clusters. These clusters were significantly associated with ruminant species: almost all the cattle genotypes were found in a "cattle-specific" cluster whereas small ruminants genotypes essentially grouped into the two other clusters. The clusters also proved stable over space and time, some genotypes being more specifically observed in certain farming regions. We also observed some within-farm diversity but this diversity was restricted to a same genotypic cluster. Finally, we identified 6 MLVA markers that maximized the representativeness of the diversity described. Overall, we highlighted that molecular epidemiology is a relevant approach to assess C. burnetii's genetic diversity and to reveal the existence of species-specific associations and regional stability. These results will be valuable in the field to trace genotype circulation among ruminants and from ruminants to humans. Ultimately, the potential links between genotypes and virulence traits need to be investigated to adapt control measures in livestock farms.