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International audience; The present study compared multiplex PCR (mPCR) and Whole Genome Sequencing (WGS) using the SCCmecFinder database to identify the Staphylococcal Cassette Chromosome (SCC) mec in five Staphylococcus aureus (SA) and nine non-aureus staphylococci (NAS) isolated from dairy cattle. mPCR identified an SCCmecIV in four SA and one NAS, but could not differentiate between SCCmecII and IV in the fifth SA, that all harbored the mecA gene and were phenotypically resistant to cefoxitin. SCCmecFinder confirmed the presence of an SCCmecIVc(2B) in four SA and of the SCCmecIVa(2B) in the fifth SA and the one NAS. Both methods also detected one untypeable SCCmec in another cefoxitin-resistant NAS harboring the mecA gene and a pseudo SCCmec in one cefoxitin-sensitive NAS harboring one mecC-related gene. No SCCmec elements were identified either in one cefoxitin-sensitive NAS harboring the mecA2 gene, or in five NAS (one resistant and four sensitive to cefoxitin) harboring the mecA1 gene. SCCmecFinder could even not identify the presence of any mecA1 gene in these five NAS, whose presence was nevertheless confirmed by ResFinder. The conclusions of this study are: (i) mPCR and WGS sequencing using SCCmecFinder are complementary methodologies to identify SCCmec; (ii) SCCmecFinder and ResFinder to a lesser extent cannot identify all mec gene allotypes; (iii) a specific classification of the SCCmec in NAS would be epidemiologically helpful; (iv) presence of a mecA gene and a complete SCCmec is linked to cefoxitin resistance, whereas presence of other mec genes and of pseudo or no SCCmec is not.

Bluetongue virus is a vector-borne pathogen affecting ruminants that has caused major epidemics in France. Reconstructing the history of bluetongue in French cattle under control strategies such as vaccination has been hampered by the high level of sub-clinical infection, incomplete case data and poor understanding of vaccine uptake over time and space. To tackle these challenges, we used three age-structured serological surveys carried out in cattle (N = 22,342) from ten administrative subdivisions called departments. We fitted catalytic models within a Bayesian MCMC framework to reconstruct the force of seroconversion from infection or vaccination, and the population-level susceptibility per semester between 2007 and 2016. In the departments of the study area, we estimated that 36% of cattle had been infected prior to vaccine rollout that became compulsory from July 2008. The last outbreak case was notified in December 2009, at which time 83% of the animals were seropositive, under the cumulative effect of vaccination and infection. The probability of seroconversion per semester dropped below 10% after 2010 when vaccination became optional. Vaccine uptake was smaller during the 2012 campaign than during the one in 2011, with strong regional contrasts. Eighty four percent of cattle were susceptible when bluetongue re-emerged in 2015. Thus, serological surveys can be used to estimate vaccine uptake and the magnitude of infection, the relative effect of which can sometimes be inferred using prior knowledge on reported incidence and vaccination dates. Keywords: Bluetongue, Epidemiology, France, Immunization, Serology

This study was funded by the Blue-Med project in the framework of the PRIMA program, an Art.185 initiative supported and funded under European Union Horizon 2020; International audience; Bluetongue virus (BTV) is the etiologic agent of a non-contagious arthropod-borne disease transmitted to wild and domestic ruminants. BTV induces a large panel of clinical manifestations ranging from asymptomatic infection to lethal hemorrhagic fever. Despite the fact that BTV has been studied extensively, we still have little understanding of the molecular determinants of BTV virulence. In our report, we have performed a comparative yeast two-hybrid (Y2H) screening approach to search direct cellular targets of the NS4 virulence factor encoded by two different serotypes of BTV: BTV8 and BTV27. This led to identifying Wilms’ tumor 1-associated protein (WTAP) as a new interactor of the BTV-NS4. In contrast to BTV8, 1, 4 and 25, NS4 proteins from BTV27 and BTV30 are unable to interact with WTAP. This interaction with WTAP is carried by a peptide of 34 amino acids (NS422−55) within its putative coil-coiled structure. Most importantly, we showed that binding to WTAP is restored with a chimeric protein where BTV27-NS4 is substituted by BTV8-NS4 in the region encompassing residue 22 to 55. We also demonstrated that WTAP silencing reduces viral titers and the expression of viral proteins, suggesting that BTV-NS4 targets a cellular function of WTAP to increase its viral replication.

International audience; Bluetongue virus serotype 8 (BTV-8) was reported for the first time in Europe in 2006, causing the largest bluetongue outbreak ever recorded. France was mostly impacted in 2007/09. Trade restrictions were implemented all along. Vaccination became available from 2008: a limited number of doses was first administered in an emergency vaccination campaign, followed by two nationwide compulsory vaccination campaigns in 2009 and 2010. France regained a disease-free status in December 2012, but BTV may have kept circulating undetected as infected herds have been reported again since August 2015. We developed a stochastic dynamic compartmental model of BTV transmission in cattle and sheep to analyze the relative importance of vector active flight and host movements in disease spread, and assess the effectiveness of control measures. We represented BTV transmission both within and between French administrative subdivisions called cantons, during the 2007/09 outbreak and until the end of 2010, when compulsory vaccination was interrupted. Within-canton transmission was vector-borne, and between canton transmission could occur through three contact networks that accounted for movements of: (i) vectors between pastures located at close distance; (ii) cattle and sheep between pastures of the same farm; (iii) traded cattle. We estimated the model parameters by approximate Bayesian computation, using data from the 2007 French outbreak. With this framework, we were able to reproduce the BTV-8 epizootic wave. Host movements between distant pastures of the same farm were found to have a major contribution to BTV spread to disease-free areas, thus raising practical questions about herd management during outbreaks. We found that cattle trade restrictions had been well complied with; without them, the whole French territory would have been infected by winter 2007. The 2008 emergency vaccination campaign had little impact on disease spread as almost half vaccine doses had likely been administered to already immune cattle. Alternatively, establishing a vaccination buffer zone would have allowed a better control of BTV in 2008: limiting its spatial expansion and decreasing the number of infected cattle and sheep. We also showed a major role of compulsory vaccination in controlling the outbreak in 2009 and 2010, though we predicted a possible low-level circulation after the last detection.

International audience; No abstract available

International audience; Group B Streptococcus (GBS) is a host-generalist species, most notably causing disease in humans and cattle. However, the differential adaptation of GBS to its two main hosts, and the risk of animal to human infection remain poorly understood. Despite improvements in control measures across Europe, GBS is still one of the main causative agents of bovine mastitis in Portugal. Here, by whole-genome analysis of 150 bovine GBS isolates we discovered that a single CC61 clone is spreading throughout Portuguese herds since at least the early 1990s, having virtually replaced the previous GBS population. Mutations within an iron/manganese transporter were independently acquired by all of the CC61 isolates, underlining a key adaptive strategy to persist in the bovine host. Lateral transfer of bacteriocin production and antibiotic resistance genes also underscored the contribution of the microbial ecology and genetic pool within the bovine udder environment to the success of this clone. Compared to strains of human origin, GBS evolves twice as fast in bovines and undergoes recurrent pseudogenizations of human-adapted traits. Our work provides new insights into the potentially irreversible adaptation of GBS to the bovine environment.

Background Rift Valley fever virus (RVFV) causes a viral zoonosis, with discontinuous epizootics and sporadic epidemics, essentially in East Africa. Infection with this virus causes severe illness and abortion in sheep, goats, and cattle as well as other domestic animals. Humans can also be exposed through close contact with infectious tissues or by bites from infected mosquitoes, primarily of the Aedes and Culex genuses. Although the cycle of RVFV infection in savannah regions is well documented, its distribution in forest areas in central Africa has been poorly investigated. Methodology/Principal Findings To evaluate current circulation of RVFV among livestock and humans living in the Central African Republic (CAR), blood samples were collected from sheep, cattle, and goats and from people at risk, such as stock breeders and workers in slaughterhouses and livestock markets. The samples were tested for anti-RVFV immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies. We also sequenced the complete genomes of two local strains, one isolated in 1969 from mosquitoes and one isolated in 1985 from humans living in forested areas. The 1271 animals sampled comprised 727 cattle, 325 sheep, and 219 goats at three sites. The overall seroprevalence of anti-RVFV IgM antibodies was 1.9% and that of IgG antibodies was 8.6%. IgM antibodies were found only during the rainy season, but the frequency of IgG antibodies did not differ significantly by season. No evidence of recent RVFV infection was found in 335 people considered at risk; however, 16.7% had evidence of past infection. Comparison of the nucleotide sequences of the strains isolated in the CAR with those isolated in other African countries showed that they belonged to the East/Central African cluster. Conclusion and significance This study confirms current circulation of RVFV in CAR. Further studies are needed to determine the potential vectors involved and the virus reservoirs., Author Summary Rift valley fever virus (RVFV) is an arthropod-borne virus that causes serious illness in both animals and humans. RVFV is transmitted by direct contact with infectious tissues or by the bites of infected mosquito species of the Aedes and Culex genuses. Its distribution in tropical forests in central Africa is poorly documented. We assessed the current circulation of RVFV among livestock and humans in the Central African Republic (CAR) by detecting anti-RVFV immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies in sheep, cattle and goats and in people living in Bangui who were considered at risk. We also sequenced the complete genomes of two local strains, one isolated in 1969 from mosquitoes and one isolated in 1985 from humans living in forested areas. Sheep were the most frequently infected ruminants. IgM antibodies were found only during the rainy season; the frequency of IgG antibodies did not differ according to season. No evidence of recent RVFV infection was found in humans at risk; however, 16.7% had evidence of past infection. Phylogenetic analysis showed a perfect match of CAR strains with the East/Central African cluster. Our results confirm current circulation of RVFV in CAR. Further studies should be conducted to determine the vectors involved and the virus reservoirs.

Background Rift Valley fever (RVF) is a vector-borne disease affecting ruminants and humans. Madagascar was heavily affected by RVF in 2008–2009, with evidence of a large and heterogeneous spread of the disease. The identification of at-risk environments is essential to optimize the available resources by targeting RVF surveillance in Madagascar. Herein, the objectives of our study were: (i) to identify the environmental factors and areas favorable to RVF transmission to both cattle and human and (ii) to identify human behaviors favoring human infections in Malagasy contexts. Methodology/Principal Findings First, we characterized the environments of Malagasy communes using a Multiple Factor Analysis (MFA). Then, we analyzed cattle and human serological data collected at national level using Generalized Linear Mixed Models, with the individual serological status (cattle or human) as the response, and MFA factors, as well as other potential risk factors (cattle density, human behavior) as explanatory variables. Cattle and human seroprevalence rates were positively associated to humid environments (p, Author Summary Rift Valley fever virus (RVFV) is a pathogen that causes a vector-borne tropical disease. The disease affects ruminants and humans and severely impacts the health and economy of affected countries. Madagascar was heavily affected by Rift Valley fever (RVF) in 2008–2009, with evidence of a large and heterogeneous spread of the disease. Our study aims at identifying environmental and human-related risk factors for RVFV transmission. First, we characterized Malagasy environments according to their putative influence on RVFV mosquito density and population dynamics. Then we statistically analyzed cattle and human serological data collected at a national level with the individual serological status as response, and Malagasy environments previously characterized by climatic and landscape variables as well as other potential risk factors as explanatory variables. Our results identified humid environments of the western, north-western and eastern parts of the island as risky areas. The identification of at-risk environments is essential to focus veterinary surveillance and control of RVFV.

International audience; Bovine leptospirosis causes substantial reproductive failure in cattle, mainly due to infections with serovar (sv) Hardjo infection. Notwithstanding, other serovars from the serogroup (sg) Sejroe could also have important roles in bovine leptospirosis. The objective was to investigate genetic diversity of serogroup Sejroe isolates obtained from asymptomatic cattle in the state of Rio de Janeiro, Brazil. Urine and vaginal fluid (VF) were collected from clinically healthy cattle immediately after slaughter. Five isolates were recovered and characterized (serogrouping) as belonging to sg Sejroe. Sequencing of rrs and secY genes further identified them as Leptospira santarosai. Analysis of secY sequences indicated a high level of sequence homology to sv Guaricura strains. Based on culture and sequence data, we inferred that other members of sg Sejroe may be important in bovine leptospiral infection, particularly genotypes of L. santarosai serovar Guaricura.

OBJECTIVES In group B Streptococcus (GBS), cross-resistance to lincosamides, streptogramin A and pleuromutilins (LSAP) is mediated by the acquisition of lsa genes. Here, we characterized the diversity, mobility and ecology of lsa genes in this species. METHODS lsa variants were systematically identified by BLAST searches in the genomes of 531 GBS strains from different hosts and geographical origins. The associated phenotypes were determined by a microdilution MIC method. Acquisition of resistance genes was deduced from comparative genomics and phylogeny. Their mobility was tested by conjugation experiments. RESULTS lsa(E) and three variants of lsa(C) were identified in GBS strains. Two lsa(C) variants had not been previously reported. All four variants conferred LSAP phenotypes. lsa(E) was located in a multiresistance gene cluster of a single human strain. This gene was transferred by a high-frequency recombination-type mechanism between GBS strains. Two lsa(C) variants are carried in six unrelated human strains by two similar elements specifically integrated in the oriT site of four different classes of integrative and conjugative elements (ICEs). Strikingly, the acquisition of the resistance gene always occurred by the integration of the element into a resident ICE. The third lsa(C) variant was located at the same site in the core genome of 11 genetically distant bovine strains and was likely propagated by horizontal transfer of the corresponding chromosomal region. CONCLUSIONS lsa genes in GBS show distinct host specificities and modes of transfer. In general, their dissemination is mediated by recombination rather than by the transfer of conjugative elements.

International audience; We performed a simulation study to evaluate the performances of an anomaly detection algorithm considered in the frame of an automated surveillance system of cattle mortality. The method consisted in a combination of temporal regression and spatial cluster detection which allows identifying, for a given week, clusters of spatial units showing an excess of deaths in comparison with their own historical fluctuations. First, we simulated 1,000 outbreaks of a disease causing extra deaths in the French cattle population (about 200,000 herds and 20 million cattle) according to a model mimicking the spreading patterns of an infectious disease and injected these disease-related extra deaths in an authentic mortality dataset, spanning from January 2005 to January 2010. Second, we applied our algorithm on each of the 1,000 semi-synthetic datasets to identify clusters of spatial units showing an excess of deaths considering their own historical fluctuations. Third, we verified if the clusters identified by the algorithm did contain simulated extra deaths in order to evaluate the ability of the algorithm to identify unusual mortality clusters caused by an outbreak. Among the 1,000 simulations, the median duration of simulated outbreaks was 8 weeks, with a median number of 5,627 simulated deaths and 441 infected herds. Within the 12-week trial period, 73% of the simulated outbreaks were detected, with a median timeliness of 1 week, and a mean of 1.4 weeks. The proportion of outbreak weeks flagged by an alarm was 61% (i.e. sensitivity) whereas one in three alarms was a true alarm (i.e. positive predictive value). The performances of the detection algorithm were evaluated for alternative combination of epidemiologic parameters. The results of our study confirmed that in certain conditions automated algorithms could help identifying abnormal cattle mortality increases possibly related to unidentified health events.

Parasitological investigations on ruminal paramphistomosis were carried out in 2033 cattle over a 14-month period in three Algerian slaughterhouses to determine the prevalence and intensity of this disease. The prevalence of infection varied from different slaughterhouses (1.2% at El Khroub, 7.5% at Ferdjioua and 12.1% at Jijel). The worm burden was significantly higher in cattle from El Khroub than for those slaughtered in the other two sites (a mean of 984.1 parasites compared with 87.5 and 140.7 at Jijel and Ferdjioua, respectively). No significant correlation between prevalence, intensity of infection and age of cattle was noted. Overall, females were significantly more infected than males, whereas the sex of animals was independent of worm burdens recovered in slaughtered cattle. The climatic conditions of provinces in which the slaughterhouses were located (a more temperate climate for Jijel and semiarid conditions for the other two provinces) may partly explain variations noted in prevalence and intensity of infection. The present results will contribute to our understanding of the epidemiology of Paramphistomum daubneyi infections in north-eastern Algeria.

International audience; This work describes the first ultrastructural results on spermiogenesis and on the mature spermatozoon of Dicrocoelium hospes (Trematoda, Digenea) collected in Bos indicus from Senegal (Africa). Examination of this species was processed by TEM. Spermiogenesis follows the general pattern found in the digenean, but reveals a particularity consisting of the appearance of glycogen granules in the late spermatids within the testes. The mature spermatozoon possesses five distinct regions and presents all features found in Digenea gametes: two axonemes, mitochondria, nucleus and parallel cortical microtubules. However, several characters allow us to distinguish D. hospes from other digenetic trematodes within the Dicrocoeliidae family. In fact, we observed several structures that are absent in the other species of Dicrocoeliidae studied until now, such as: a cytoplasmic expansion, extramembranar ornamentation, spine-like bodies and two parallel mitochondria in the mature sperm. Moreover, additional particular characteristics were observed in this species in both extremities of the spermatozoon. This work produced new data on the ultrastructure of this trematode family which may be useful for phylogenetic purposes.

International audience; As part of a general project aimed at elucidating the initiation of mucin-type O-glycosylation in helminth parasites, we have characterized a novel ppGalNAc-T (UDP-N-acetyl-D-galactosamine:polypeptide N-acetylgalactosaminyltransferase) from the cestode Echinococcus granulosus (Eg-ppGalNAc-T1). A full-length cDNA was isolated from a library of the tissue-dwelling larval stage of the parasite, and found to code for a 654-amino-acid protein containing all the structural features of ppGalNAc-Ts. Functional characterization of a recombinant protein lacking the transmembrane domain showed maximal activity at 28 degrees C, in the range 6.5-7.5 pH units and in the presence of Cu2+. In addition, it transferred GalNAc to a broad range of substrate peptides, derived from human mucins and O-glycosylated parasite proteins, including acceptors containing only serine or only threonine residues. Interestingly, the C-terminal region of Eg-ppGalNAc-T1 bears a highly unusual lectin domain, considerably longer than the one from other members of the family, and including only one of the three ricin B repeats generally present in ppGalNAc-Ts. Furthermore, a search for conserved domains within the protein C-terminus identified a fragment showing similarity to a recently defined domain, specialized in the binding of organic phosphates (CYTH). The role of the lectin domain in the determination of the substrate specificity of these enzymes suggests that Eg-ppGalNAc-T1 would be involved in the glycosylation of a special type of substrate. Analysis of the tissue distribution by in situ hybridization and immunohistochemistry revealed that this transferase is expressed in the hydatid cyst wall and the subtegumental region of larval worms. Therefore it could participate in the biosynthesis of O-glycosylated parasite proteins exposed at the interface between E. granulosus and its hosts.

In France, through exhaustive active surveillance, approximately 17.1 million adult cattle were tested for bovine spongiform encephalopathy from July 2001 through July 2007; approximately 3.6 million were >8 years of age. Our retrospective Western blot study of all 645 confirmed cases found that 7 were H-type and 6 were L-type.

International audience; Seventy-four cattle, from three farms endemic for tropical theileriosis in the north of Tunisia, were studied for tick populations from June 1991 to June 1992. Ticks were removed from cattle twice a month in the summer and every month the rest of the year. They were identified and assessed for Theileria infection. A total of 5083 Hyalomma adult ticks were collected and the major species found was H. detritum (84.3%). The activity of this species is limited between June and August with a peak in numbers observed at the end of June and the beginning of July. Amongst the 2356 Hyalomma ticks dissected, no evidence of salivary gland infection was found in either H. m. marginatum or H. a. excavatum. However, 12.4% (277/2230) of H. d. detritum dissected ticks were infected with Theileria species and amongst these, 62% had one to two sporoblasts in their salivary glands (range 1-91). The prevalence, but not the intensity, of infection was greater in females than in male ticks, and the cases of tropical theileriosis followed the peak of infected females. This suggests that female ticks have a more important role in theileriosis transmission than male ticks. A significantly lower number of adult H. detritum were collected from calves, at their first tick season, than from adult cattle. Finally, this study showed that the infestation level of cattle by H. d. detritum and the prevalence of Theileria-infection in these cattle varied between the three farms studied.

International audience; Census and disposal data provide a multipurpose source of information on cattle mortality. The retrospective analyses we conducted on the data gathered in the National Cattle Register produced relevant information for describing and modelling the cattle mortality baseline and evaluating the impact of the 2007-2008 Blue Tongue epidemic on the French cattle population. This work was conducted retrospectively but showed that monitoring cattle mortality near real time could help detecting unexpected events. We are thus currently working on a timely and automated system to monitor cadaver disposal requests received by rendering plants, thanks to a data interchange system recently implemented between the Ministry of Agriculture and the fallen stock companies. Besides technical and methodological challenges, using these data for surveillance purposes raises epidemiological questions that still need to be answered. The question remains notably as to whether an abnormal increased mortality is a sensitive and timely signal for detecting unexpected health events. It appears also very challenging to identify the most adequate surveillance scale (time, space and population) and the most adequate anomaly detection algorithms to apply when the characteristics of the signals to be detected (shape, amplitude, etc.) are not known a priori. In Human health, similar systems have not yet proven their ability to detect unexpected events earlier than classical surveillance systems currently in place, but they have already demonstrated their value for real time assessment of identified and potentially dangerous events. Combined with traditional surveillance systems, we think that monitoring routinely collected data could improve the surveillance of the animal population health. Even if not used for detection purposes, cattle mortality monitoring could be used to rapidly produce information on the impact and evolution of identified events, what would facilitate decision-making regarding management measures and improve the communication.

In bacteria, normal mutation frequencies are mostly around 10 −10 per base pair. However, there exists natural isolates, called “mutators,” that exhibit permanent mutation occurrences up to 1,000-fold greater than usual. As mutations play essential roles, particularly in the evolution of antibiotic resistance, bacteria showing elevated mutation rates could have an important responsibility in the emergence of antibiotic resistance, especially in the clinical background. In this announcement, we report the first complete genome sequence of the Salmonella enterica subsp. enterica serotype Heidelberg B182 mutator strain, isolated from bovine feces (France), which consists of a 4,750,465-bp circular chromosome (cB182_4750; GC, 52.2%) and one circular plasmid of 37,581 bp (pB182_37; GC, 42.8%).

International audience; Babesiosis is a tick-transmitted disease of mammalian hosts, caused by the intraerythrocytic protozoan parasites of the genus Babesia. Transmission of Babesia parasites from the vertebrate host to the tick is mediated by sexual stages, the gametocytes which are the only intraerythrocytic stages that survive and develop inside the vector. Very few data are available concerning these parasite stages and some markers are needed in order to refine our knowledge of Babesia life cycle inside the tick and to permit the monitoring of parasite transmission from vertebrate to vector. We previously identified some potential markers of the Babesia divergens gametocytes using an in silico post-genomic approach based on sequence identity between the available genomes of Plasmodium and Babesia spp. Here, one of the identified proteins, BdCCp2, was validated as a marker of sexual stages of B. divergens, in infected ticks challenged with antisera directed against recombinant BdCCp2 protein. The BdCCp2 protein was detected by Western blot in some infected ticks, as a discrete band of approximately 171 kDa, while no signal was detected in the laboratory-reared non-infected tick. BdCCp2 was also detected, by immunohistochemical analyses, in piriform or ovoid bodies, measuring 2.5-4.5 μm in diameter, in the gut of partially engorged ticks that were experimentally infected. This molecular marker can then be used in the future to characterize and analyze the biology of B. divergens gametocytes.

International audience; Understanding where and how fast an infectious disease will spread during an epidemic is critical for its control. However, the task is a challenging one as numerous factors may interact and drive the spread of a disease, specifically when vector-borne diseases are involved. We advocate the use of simultaneous autoregressive models to identify environmental features that significantly impact the velocity of disease spread. We illustrate this approach by exploring several environmental factors influencing the velocity of bluetongue (BT) spread in France during the 2007-2008 epizootic wave to determine which ones were the most important drivers. We used velocities of BT spread estimated in 4,495 municipalities and tested sixteen covariates defining five thematic groups of related variables: elevation, meteorological-related variables, landscape-related variables, host availability, and vaccination. We found that ecological factors associated with vector abundance and activity (elevation and meteorological-related variables), as well as with host availability, were important drivers of the spread of the disease. Specifically, the disease spread more slowly in areas with high elevation and when heavy rainfall associated with extreme temperature events occurred one or two months prior to the first clinical case. Moreover, the density of dairy cattle was correlated negatively with the velocity of BT spread. These findings add substantially to our understanding of BT spread in a temperate climate. Finally, the approach presented in this paper can be used with other infectious diseases, and provides a powerful tool to identify environmental features driving the velocity of disease spread.

International audience; Field investigations in 361 liver fluke infected cattle- or sheep-breeding farms on acid soil were carried out during thirty years in March and April to record indicator plants in relation to the category of site colonized by the intermediate host of liver fluke, the snail Galba truncatula. Seven types of snail zones and six species of indicator plants were recorded in the 7709 positive sites studied. The most frequent habitats were located at the peripheral extremities of open drainage furrows. Juncus acutiflorus, Juncus effusus, Glyceria fluitans, and Agrostis stolonifera were the indicator plants. Plant indicators were highly efficient (38-80% of variance explained) when used with the size of site area within a hydrographical zone. The identification of positive sites for G. truncatula is a key for controlling the intermediate host on pastures through biological control with predatory molluscs. The positive sites may be detected using a two-step method including first categorization of hydrographical zones and then, within a zone, use of one or several indicator plants (frequently J. acutiflorus).

International audience; Staphylococcus aureus is an important pathogen in domestic ruminants. The main objective of this study was to determine the similarity of epidemiologically unrelated S. aureus isolates from bovine, ovine, and caprine hosts regardless the locus of isolation (nares and udder). By pulsed-field gel electrophoresis, seven major pulsotypes were identified among 153 isolates recovered from 12 different regions of France as well as from Brazil, the USA and Belgium. Typing of the accessory gene regulator (agr) and capsular (cap) serotype was carried out on all the isolates and revealed the predominance of agr I and III and of cap8 regardless the ruminant host species. Screening for methicilin-resistant S. aureus (MRSA) was carried out by disk diffusion and revealed a prevalence of only 3.2% of MRSA among the strains tested. These results suggest the existence of a host rather than tissue specificity among S. aureus isolates colonising the ruminant species and suggest a limited transmission of those isolates between large (bovine) and small (ovine-caprine) ruminants. The agr class and cap types correlated with pulsotype clusters rather than with a specific host species. Antimicrobial resistance appears not to have contributed to the predominance of any given genotypes, and MRSA prevalence appears very low in ruminant isolates.

Aims: The analysis by Denaturing Gradient Gel Electrophoresis (DGGE) of the PCR-amplified V3 region of 16S rRNA gene was previously shown to detect and differentiate a large number of human and animal mycoplasmas. In this study, we further assessed the suitability of the technique for epidemiological surveillance of mycoplasmas belonging to the ‘Mycoplasma mycoides’ cluster, a phylogenetic group that includes major ruminant pathogens. Methods and Results: The V3 region of 16S rRNA genes from approx. 50 field strains was amplified and analysed by DGGE. Detection and identification results were compared with the ones obtained by antigenic testing and sequence analysis. Conclusions: The DGGE technique is robust and valuable as a first-line test, but the patterns obtained for strains belonging to the ‘M. mycoides’ cluster were too variable within a taxon and in contrast too conserved between taxa to allow an unequivocal identification of isolates without further analysis. Significance and Impact of the Study: Issues raised by the quest for a single universal test able to detect and identify any mycoplasma in one clinical sample are thoroughly documented.

To assess the prevalence and relative importance of Mycoplasma bovis among the pathological agents implicated in bovine respiratory disease (BRD), we surveyed 135 veal calves from nine feedlots in eastern France during naturally occurring outbreaks of respiratory disease. Occurrence of respiratory pathogens, M. bovis, bovine viral diarrhoea (BVD) virus, bovine respiratory syncytial (BRS) virus and parainfluenza-3 (PI3) virus was investigated by seroconversion and isolation of bacteria and viruses from broncho-alveolar fluids. M. bovis and pathogenic respiratory bacteria were isolated in eight of the nine feedlots, and from 106 and 32, respectively, of the 135 tested animals. Seroconversion to PI3 virus occurred in four lots. BVD and BRS viruses were detected in eight and one lot, respectively. M. bovis was the most frequently isolated aetiologic agent in these BRD outbreaks, spreading early and widely throughout the affected units (60-100% rate of isolation and seroconversion). These results stress the importance of M. bovis in the BRD complex.

International audience; Legume root architecture is characterized by the development of two de novo meristems, leading to the formation of lateral roots or symbiotic nitrogen-fixing nodules. Organogenesis involves networks of transcription factors, the encoding mRNAs of which are frequently targets of microRNA (miRNA) regulation. Most plant miRNAs, in contrast with animal miRNAs, are encoded as single entities in an miRNA precursor. In the model legume Medicago truncatula, we have identified the MtMIR166a precursor containing tandem copies of MIR166 in a single transcriptional unit. These miRNAs post-transcriptionally regulate a new family of transcription factors associated with nodule development, the class-III homeodomain-leucine zipper (HD-ZIP III) genes. In situ expression analysis revealed that these target genes are spatially co-expressed with MIR166 in vascular bundles, and in apical regions of roots and nodules. Overexpression of the tandem miRNA precursor correlated with MIR166 accumulation and the downregulation of several class-III HD-ZIP genes, indicating its functionality. MIR166 overexpression reduced the number of symbiotic nodules and lateral roots, and induced ectopic development of vascular bundles in these transgenic roots. Hence, plant polycistronic miRNA precursors, although rare, can be processed, and MIR166-mediated post-transcriptional regulation is a new regulatory pathway involved in the regulation of legume root architecture.

In gram-negative pathogens, extended-spectrum beta-lactamases (ESBLs) confer resistance to penicillins, cephalosporins (including extended-spectrum cephalosporins), and aztreonam, but not to cephamycins and carbapenems, and are inhibited by beta-lactamase inhibitors ([8][1]). Recently, cefotaximases

International audience; Between October and November 2006, a total of 278 bovine blood samples were examined, and 104 (37.4%) were positive for piroplasms by microscopy. A reverse line blot hybridisation with polymerase chain reaction detected Theileria annulata, T. buffeli, Babesia bovis and B. bigemina in cattle accounting for 48.6% of positive samples. The most frequently found species was T. buffeli, which was present in 39.2% of the samples. T. annulata was found in 48 samples (17.3%). Babesia infections were less frequently detected: B. bovis was found in 6.8% of the samples and B. bigemina in 4.3%. Mixed infections were detected in 45 samples, accounting for seven different combinations of species. Seven Ixodid tick species (Boophilus annulatus, Ixodes ricinus, Hyalomma marginatum, Hyalomma excavatum, Hyalomma detritum, Haemaphysalis punctata and Haemaphysalis sulcata) were collected from examined cattle in the 23 visited farms. I. ricinus was the dominant species (36%), mainly collected in the humid zone, while it seemed to be very rare in the semi-arid zone (where only 15 specimens were collected), whereas B. annulatus was the most commonly collected species in the sub-humid area (68.5% of ticks collected in this zone).

International audience; Infection with Shiga-toxin producing Escherichia coli (STEC) may result in the development of the haemolytic-uremic syndrome (HUS), the main cause of acute renal failure in children. While O157:H7 STEC are associated with large outbreaks of HUS, it is difficult to predict whether a non-O157:H7 isolate can be pathogenic for humans. The mucosal innate immune response plays a central role in the pathogenesis of HUS; therefore, we compared the induction of IL-8 and CCL20 in human colon epithelial cells infected with strains belonging to different serotypes, isolated from cattle or from HUS patients. No correlation was observed between strain virulence and chemokine gene expression. Rather, the genetic background of the strains seems to determine the chemokine gene expression profile. Investigating the contribution of different bacterial factors in this process, we show that the type III secretion system of O157:H7 bacteria, but not the intimate adhesion, is required to stimulate the cells. In addition, H7, H10, and H21 flagellins are potent inducers of chemokine gene expression when synthesized in large amount.

Aims: To quantify the variability of the Shiga toxin 2 (Stx2) production by a panel of stx2-positive Escherichia coli O157:H7/H7- isolates from healthy cattle before and after induction with enrofloxacin. Methods and Results: ProSpecT® ELISA was used to quantify the Stx2 production by stx2-positive E. coli O157:H7/H7- isolates in native conditions (basal level) or after induction with enrofloxacin. Whereas only 15·2% of the E. coli O157:H7/H7- strains studied displayed significant amounts of detectable Stx2 without induction, most of them were shown to be inducible, and at various levels, in presence of subinhibitory concentrations of enrofloxacin. Conclusions: We demonstrated the capability of a highly elevated proportion of stx2-positive, but constitutively Stx2-negative, E. coli O157:H7/H7- isolates from healthy cattle to produce significant levels of Shiga toxin Stx2 in presence of subtherapeutic concentrations of enrofloxacin, an antibiotic of the fluoroquinolones family only licensed for veterinary use. Significance and Impact of the Study: This study documents the risk that bovine-associated Shiga toxin producing E. coli isolates may become more frequently pathogenic to humans as a side-effect of the increasing use of veterinary fluoroquinolones in the oral treatment of food animals like cattle or poultry.

International audience; The ultrastructural study of spermiogenesis and the spermatozoon of Paramphistomum microbothrium gave evidence of some characteristics of this digenean. The intercentriolar body located between two striated roots and two centrioles exhibits a symmetric plane. The two external bands of this intercentriolar body are made up of a line of granules. The striated roots are about 3.7 microm long. During spermiogenesis, a flagellar rotation of 90 degrees is described. The spermatozoon is characterized in its anterior part by the existence of external ornamentations and a lateral expansion exhibiting a "spine-like body". Such a spine-like body has never been described before in a digenean. The region II of the spermatozoon is asymmetrical and also characterized by the absence of mitochondrion. The posterior end of this spermatozoon exhibits a nucleus and a few microtubules.

Health organizations worldwide have advocated that treatment records be required to show that antibiotics are used prudently by veterinarians and farmers alike. In 2000, the French government passed legislation making a farm register mandatory for all farms that raise animals for food production. The farm register is a comprehensive record designed to track all animal movements, treatments, and veterinary interventions on the farm. We conducted a survey to assess the knowledge, attitudes, and behaviors of dairy farmers toward the farm register, with particular emphasis on recording of antibiotic treatments. The 43 farmers interviewed belonged to veterinary health cooperatives. All farmers correctly named an antibiotic or antibiotic preparation, yet only 2 recognized the 5 components of the farm register. Farmer attitudes toward the register were globally positive, even though they named a wide variety of constraints. Nevertheless, 72% of farmers interviewed had a permanent health record, and approximately half had recorded either the last treatment (irrespective of drug class) or the last intramammary tube administered. Results were discussed in the light of health behavior change models that are applied in human medicine. We suggest that programs that seek compliance with the farm register should focus on educational interventions and bonus incentives.

Objectives: A florfenicol-resistant Pasteurella trehalosi isolate from a calf was investigated for the presence and the location of the gene floR. Methods: The P. trehalosi isolate 13698 was investigated for its in vitro susceptibility to antimicrobial agents and its plasmid content. A 14.9 kb plasmid, designated pCCK13698, was identified by transformation into Pasteurella multocida to mediate resistance to florfenicol, chloramphenicol and sulphonamides. The plasmid was sequenced completely and analysed for its structure and organization. Results: Plasmid pCCK13698 exhibited extended similarity to plasmid pHS-Rec from Haemophilus parasuis including the region carrying the parA, repB, rec and int genes. Moreover, it revealed similarities to plasmid RSF1010 in the parts covering the mobC and repA-repC genes and to plasmid pMVSCS1 in the parts covering the sul2-catA3-strA gene cluster. Moreover, the floR gene area corresponded to that of transposon TnfloR. In addition, two complete insertion sequences were detected that were highly similar to IS1593 from Mannheimia haemolytica and IS26 from Enterobacteriaceae. Several potential recombination sites were identified that might explain the development of plasmid pCCK13698 by recombination events. Conclusions: The results of this study showed that in the bovine pathogen P. trehalosi, floR-mediated resistance to chloramphenicol and florfenicol was associated with a plasmid, which also carried functionally active genes for resistance to sulphonamides (sul2) and chloramphenicol (catA3). This is to the best of our knowledge the first report of resistance genes in P. trehalosi and only the second report of the presence of a florfenicol-resistance gene in target bacteria of the family Pasteurellaceae.

Performance indicators are variables used to calculate on a continuous basis the operational level of a surveillance network's priority activities. A 10-step process was developed to enable network coordinators to identify specific performance indicators to help them monitor their networks. The methodology was based on a listing of surveillance activities, the definition and choice of network's global priority objectives, the construction of performance indicators and monitoring tables and the implementation of the system within the network. This process was implemented for the bovine clinical digestive salmonellosis surveillance network in France (RESSAB). The process produced a list of 26 activities synthesized into 15 global objectives, from which 12 were retained as priority objectives. This made it possible to develop 12 performance indicators. Indicators retrospectively calculated for the year 2003 indicated that RESSAB was operating according to the objectives set by the network's stakeholders and its financial supporter. The methodology was implemented successfully and was not very time consuming (12 person-days) or expensive. The decision makers and actors in the network quickly appropriated the system. The methodology can thus be considered validated through this example. Nevertheless, the risks inherent in the use of performance indicators must be addressed by ensuring the relevance of the selected indicators through external assessment and by prioritizing an internal and participatory approach to avoid a misuse of the performance indicators. In addition, considering that performance indicators address only the operation of the network, relevance of surveillance procedure should be addressed by external evaluation.

International audience; The present paper describes the spermiogenesis and the ultrastructure of the spermatozoon of Fasciola gigantica, as revealed by transmission electron microscopy. Spermiogenesis in F. gigantica begins with the formation of a differentiation zone containing 2 centrioles with associated striated roots and an intercentriolar body between them. Each centriole develops a flagellum. Proximodistal fusion of these flagella with the median cytoplasmic extension gives rise to the spermatozoon. Spermiogenesis in F. gigantica is characterized by the formation of a dorsolateral cytoplasmic expansion, an external ornamentation of the cell membrane, and spinelike bodies. These 3 structures were also observed in the anterior part of the spermatozoon. Our study describes for the first time the simultaneous presence of dorsolateral cytoplasmic expansion, external ornamentation of the plasma membrane, and spinelike bodies in the spermatozoon of a trematode.

International audience; Ivermectin is a worldwide-used antiparasitic drug largely administered to cattle as a topical formulation (pour-on). The actual plasma and faecal disposition of pour-on ivermectin in cattle was documented using an original pharmacokinetic model, and taking into account the oral ingestion of the topical drug following physiological licking as a secondary route of exposure. Six pairs of monozygotic twin cattle received successively one i.v. and two pour-on administrations of ivermectin at a 3-5-month interval. For one pour-on administration, the twins were separated into an unrestrained group and a group where self- and allo-licking were prevented. Ivermectin concentrations in the plasma and faeces were determined by HPLC. Licking resulted in a high intra-and inter-individual variability of systemic exposure after topical application. By the means of pharmacokinetic modelling, we showed that 58-87% of the pour-on dose was ingested, while only 10% was absorbed percutaneously. Approximately 72% of the ingested ivermectin transited directly into the faeces, resulting in a 7-fold higher faecal excretion of the parent drug than in the non-lickers. We conclude that topical administration does not guarantee a controlled drug delivery in cattle. More importantly, the simulations revealed that non-treated cattle could get easily contaminated by allo-licking, raising the public health problem of unexpected drug residues in edible tissues.

The virulence genotype profile and presence of a pathogenicity island(s) (PAI) were studied in 18 strains of F165-positive Escherichia coli originally isolated from diseased calves or piglets. On the basis of their adhesion phenotypes and genotypes, these extraintestinal pathogenic strains were classified into three groups. The F165 fimbrial complex consists of at least two serologically and genetically distinct fimbriae: F165 1 and F165 2 . F165 1 is encoded by the foo operon ( pap -like), and F165 2 is encoded by fot ( sfa related). Strains in group 1 were foo and fot positive, strains in group 2 were foo and afa positive, and strains in group 3 were foo positive only. The strains were tested for the presence of virulence genes found mainly in extraintestinal pathogenic E. coli (ExPEC) strains. Although all the strains were positive for the papA variant encoding F11 fimbriae incD , traT , and papC , the prevalence of virulence genes commonly found in PAIs associated with ExPEC strains was highly variable, with strains of group 2 harboring most of the virulence genes tested. papG allele III was detected in all strains in group 1 and in one strain in group 3. All other strains were negative for the known alleles encoding PapG adhesins. The association of virulence genes with tRNA genes was characterized in these strains by using pulsed-field gel electrophoresis and DNA hybridization. The insertion site of the foo operon was found at the pheU tRNA locus in 16 of the 18 strains and at the selC tRNA locus in the other 2 strains. Furthermore, 8 of the 18 strains harbored a high-pathogenicity island which was inserted in either the asnT or the asnV / U tRNA locus. These results suggest the presence of one or more PAIs in septicemic strains from animals and the association of the foo operon with at least one of these islands. F165-positive strains share certain virulence traits with ExPEC, and most of them are pathogenic in piglets, as tested in experimental infections.

International audience; Pour-on formulations of endectocides are extensively used to treat and control systemic parasitic diseases in cattle, worldwide. The purpose of the present study was to investigate the influence of the natural licking behaviour of cattle on the plasma and faecal disposition of topically administered ivermectin. Twelve Holstein cattle were given one single intravenous (i.v.) (200 microg/kg) and topical (500 microg/kg) administration of ivermectin at a 5-month interval. For the pour-on administration, the animals were allocated into two groups (n=6): one control group (lickers) and one group where licking was prevented (non-lickers). Ivermectin plasma (total) clearance (270+/-57.4 ml/kg/day) was very homogeneous among the 12 cattle. In contrast, major differences between lickers and non-lickers were observed following pour-on administration. Prevention of licking resulted in an extended terminal plasma half-life (363+/-16.2 vs. 154+/-7.4 h in lickers) and in a lower and less variable systemic availability of ivermectin (19+/-4.9 vs. 33+/-18.5% in lickers). More importantly, nearly 70% of the pour-on dose was recovered as parent drug in the faeces of lickers vs. only 6.6% in non-lickers. Altogether, these results are consistent with an oral rather than percutaneous absorption of topical ivermectin in control animals, the non-systemically available fraction of ingested ivermectin providing a major contribution (80%) to the drug faecal output. The consequences of licking on the disposition of pour-on ivermectin are discussed in terms of environment, given the known ecotoxicity of this drug, and of cross-contamination. Animals licking themselves and each other could result in unexpected residues in edible tissues of untreated animals and in possible subtherapeutic drug concentrations, a factor in drug resistance. According to the Precautionary Principle, these considerations elicit concern over the use of topical drug formulations in cattle.

International audience; Babesia divergens of bovine origin was isolated, for the first time in Tunisia, from a cow of a local breed (Bos taurus) from a locality in the north-west of the country. Gerbils (Meriones unguiculatus) were used for the isolation of the parasite which was inoculated into a splenectomised Friesian calf that developed a babesiosis, thus confirming the presence of the parasite.

Rift Valley fever is an acute, zoonotic viral disease of domestic ruminants, caused by a phlebovirus (Bunyaviridae family). A large outbreak occurred in Madagascar in 2008–2009. The goal of the present study was to evaluate the point prevalence of antibodies against Rift Valley Fever Virus (RVFV) in cattle in the Anjozorobe district, located in the wet and temperate highland region of Madagascar and yet heavily affected by the disease, and analyse environmental and trade factors potentially linked to RVFV transmission. A serological study was performed in 2009 in 894 bovines. For each bovine, the following variables were recorded: age, location of the night pen, minimum distance from the pen to the nearest water point and the forest, nearest water point type, and herd replacement practices. The serological data were analyzed using a generalized linear mixed model. The overall anti-RVFV IgG seroprevalence rate was 28% [CI95% 25–31]. Age was statistically linked to prevalence (p = 10−4), being consistent with a recurrent RVFV circulation. Distance from the night pen to the nearest water point was a protective factor (p = 5.10−3), which would be compatible with a substantial part of the virus transmission being carried out by nocturnal mosquito vectors. However, water point type did not influence the risk of infection: several mosquito species are probably involved. Cattle belonging to owners who purchase animals to renew the herd were significantly more likely to have seroconverted than others (p = 0.04): cattle trade may contribute to the introduction of the virus in this area. The minimum distance of the night pen to the forest was not linked to the prevalence. This is the first evidence of a recurrent transmission of RVFV in such an ecosystem that associates a wet, temperate climate, high altitude, paddy fields, and vicinity to a dense rain forest. Persistence mechanisms need to be further investigated., Author Summary Rift Valley fever (RVF) is a viral disease of domestic ruminants, which may affect humans. The RVF virus (RVFV) may be transmitted either by mosquitoes or through direct contact with vireamic body fluids or products. Until now, this disease had been described in arid, hot and irrigated or tropical areas. Performed in the year following the 2008–2009 RVFV outbreak in Madagascar, this study demonstrates for the first time a regular and intense transmission of this disease in a temperate and mountainous region.The area chosen as a pilot project shows that cattle are regularly and heavily affected in the highlands of Madagascar. Statistical analyses suggest that (i) a substantial part of the transmission is due to mosquito vectors; (ii) many mosquito species such as Culex and Anopheles, are probably involved in the transmission; (iii) cattle trade, by a regular introduction of the virus via herds coming from infected areas of the island, may explain the recurrence of the disease in this region. Further investigations are needed to understand the mechanisms of transmission of the disease, and design and implement appropriate surveillance and control measures in this area.