Assessment of virus and Leptospira carriage in bats in France.

With over 1,400 species worldwide, bats represent the second largest order of mammals after rodents, and are known to host major zoonotic pathogens. Here, we estimate the presence of pathogens in autochthonous bat populations. First, we set out to check our samples for PCR amplification efficiency by assessing the occurrence of inhibited PCR reactions from different types of bat samples with amplifying the housekeeping gene ß-actin. Second, we investigated the presence of five targeted pathogens in a French bat population using PCR. We targeted viral RNA of Canine distemper virus, Alphacoronavirus, Lyssavirus, Rotavirus and bacterial Leptospira DNA. To do so, we screened for these viruses in bat faecal samples as well as in oropharyngeal swab samples. The presence of Leptospira was assessed in urine, kidney, lung and faecal samples. Results showed a frequency of inhibited reactions ranging from 5 to 60% of samples, varying according to the sample itself and also suspected to vary according to sampling method and the storage buffer solution used, demonstrating the importance of the sampling and storage on the probability of obtaining negative PCR results. For pathogen assessment, rotavirus and alphacoronavirus RNA were detected in Myotis myotis, Myotis daubentonii, Myotis emarginatus and Rhinolophus ferrumequinum bats. Rotaviruses were also detected in Barbastella barbastellus. The presence of alphacoronavirus also varied seasonally, with higher frequencies in late summer and October, suggesting that juveniles potentially play an important role in the dynamics of these viruses. Leptospira DNA was detected in M. myotis and M. daubentonii colonies. The 16S rRNA sequences obtained from Leptospira positive samples showed 100% genetic identity with L. borgpetersenii. Neither canine distemper virus nor lyssavirus RNA were detected in any of the tested samples. This study is the first to show the presence of Leptospira in autochthonous French bats in addition to coronavirus and rotavirus RNA previously reported in European autochthonous bats.

Genetic identification of bat species for pathogen surveillance across France.

With more than 1400 chiropteran species identified to date, bats comprise one-fifth of all mammalian species worldwide. Many studies have associated viral zoonoses with 45 different species of bats in the EU, which cluster within 5 families of bats. For example, the Serotine bats are infected by European Bat 1 Lyssavirus throughout Europe while Myotis bats are shown infected by coronavirus, herpesvirus and paramyxovirus. Correct host species identification is important to increase our knowledge of the ecology and evolutionary pattern of bat viruses in the EU. Bat species identification is commonly determined using morphological keys. Morphological determination of bat species from bat carcasses can be limited in some cases, due to the state of decomposition or nearly indistinguishable morphological features in juvenile bats and can lead to misidentifications. The overall objective of our study was to identify insectivorous bat species using molecular biology tools with the amplification of the partial cytochrome b gene of mitochondrial DNA. Two types of samples were tested in this study, bat wing punches and bat faeces. A total of 163 bat wing punches representing 22 species, and 31 faecal pellets representing 7 species were included in the study. From the 163 bat wing punches tested, a total of 159 were genetically identified from amplification of the partial cyt b gene. All 31 faecal pellets were genetically identified based on the cyt b gene. A comparison between morphological and genetic determination showed 21 misidentifications from the 163 wing punches, representing ~12.5% of misidentifications of morphological determination compared with the genetic method, across 11 species. In addition, genetic determination allowed the identification of 24 out of 25 morphologically non-determined bat samples. Our findings demonstrate the importance of a genetic approach as an efficient and reliable method to identify bat species precisely.