Loop-mediated isothermal amplification (LAMP) assay for the diagnosis of fasciolosis in sheep and its application under field conditions

[Background] Loop-mediated isothermal amplification (LAMP) is a very specific, efficient, and rapid gene amplification procedure in which the reaction can run at a constant temperature. In the current study we have developed a LAMP assay to improve the diagnosis of Fasciola spp. in the faeces of sheep.[Findings] After the optimisation of the LAMP assay we have shown similar results between this technique and the standard PCR using the outer primers of the LAMP reaction. In both cases the limit of detection was 10 pg; also, the diagnosis of fasciolosis was confirmed during the first week post-infection in experimental infected sheep by both techniques. In eight naturally infected sheep, the infection with F. hepatica was confirmed in all animals before a treatment with triclabendazole and on day 30 post treatment in two sheep using the LAMP assay; however, when we carried out the standard PCR with the outer primers, the results before treatment were the same but on day 30 post-treatment the infection was only confirmed in one out of the two sheep. On the other hand, the standard PCR took around 3 h to obtain a result, comparing with 1 h and 10 min for the LAMP assay.[Conclusions] The LAMP assay described here could be a good alternative to conventional diagnostic methods to detect F. hepatica in faeces since it solves the drawbacks of the standard PCR.We thank Drs Manga-González and González-Lanza (Instituto de Ganadería de Montaña, CSIC-ULE, Spain) for providing DNA samples of D. dendriticum and C. daubneyi. This study has been funded by the national project INIA (Instituto Nacional de Investigaciones Agrarias: RTA2010-00094-C03-02), Ministry of Economy and Competitiveness (Ministerio de Economia y Competitividad), and also co-funded by the European Regional Development Fund (Fondos Feder).We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).Peer reviewe