Implémentation d'une méthode de détection du virus de la diarrhée virale bovine au sein de la fondation de la promotion des productions andines Proinpa (Bolivie)

An animal infected by the Bovine Viral Diarrhea Virus will have diarrheas, which can lead to milk production reduction, reproduction problems (and particularly abortion problems) and general weakness. This all leads to an economic loss. This is why the virus detection and the carriers, or Persistently Infected (PI) animals, eradication are important. If the virus presence was previously suspected in Bolivia, since 2009 it is certain that there is BVDV infection and the prevalence is high in comparison with others areas. Furthermore, this virus can also infect « New Worlds camelids » which are the alpacas, lamas, vicunas and guanacos species living in South-America. The aim of this study was to implement a BVDV detection method for the Foundation of Andeans Productions Promotion PROINPA in Bolivia. This objective was first met by a prevalence evaluation of the virus using ELISA anti-antibodies analysis. Then, a screening of the tested herds containing animals with a positive serology was done by real-time PCR and finally the PI animals were detected by individual real-time PCR analysis or by two ELISA anti-antigens analysis’s in a 3 week's interval. There was no problem from an infrastructure viewpoint; the laboratories have all the necessary material and equipment and the labor was qualified for both analyses. But the logistic part was more difficult, because the molecular kit has to be preserved at minus twenty degrees. And this kit took 10 days to arrive from Belgium. Because none of the molecular analysis’s succeeded, we concluded that the kit had been damaged during the travel and had been degraded. The kit conservation conditions seemed to be a problem and that is why the elaboration of a new lyophilized kit was tried in Belgium. The lyophilisation of the BVDV sequences, the IPC and EPC was easy but wasn’t possible for the Master MIX (containing the Taq Polymerase (Taq)) and the Reverse Transcriptase (RT). The « Illustra Ready-To-GoTM RT-PCR Beads », from GE Healthcare, are beads made of Taq and RT that can be stored at room temperature. The combination of a lyophilized part of the LSI kit (BVDV sequences, EPC and IPC) with those beads will lead to a molecular BVDV detection kit that can be stored at room temperature. However, this combination doesn’t seem to work. This can be due to a lot of different things like the incompatibility between the primers size and design and the Taq, the temperature activities of the Taq and the RT, the salt concentration, etc. In conclusion, different experiments are required to finalize a qPCR kit resistant to long trip conditions.