Development of a loop-mediated isothermal amplification assay for detection of Austropeplea tomentosa from environmental water samples

Abstract

Abstract Lymnaeid snails are key intermediate hosts for the development and survival of Fasciola spp., the causative agent of Fascioliasis which are economically important parasites infecting humans and livestock globally. The current control method for treating Fascioliasis is heavily reliant on anthelmintic drugs, particularly Triclabendazole (TCBZ) which has resulted in drug-resistant parasites and poses significant risk as there are no long-term efficacious alternatives available. Sustainable control measures at the farm level could include both parasite and snail control will play an important role in Fasciola spp. control and reduce the reliance on anthelmintic drugs. Implementation of such sustainable control measures requires effective identification of snails on the property however Lymnaeid snails are small and difficult to physically locate. Snail identification using an environmental DNA approach is a recent approach in which physically locating snails are not required. Austropeplea tomentosa, is the primary intermediate snail host for F. hepatica transmission in South-East Australia and we present an in-field loop-mediated isothermal amplification and water filtering method for the detection of A. tomentosa eDNA from water samples to improve current surveillance methods. This methodology is highly sensitive with a detection limit of 5 × 10− 6 ng/μL, detected in < 20 minutes, with cumulative sample preparation and amplification time under 1 hour. This proposed workflow could assist in monitoring areas to determine the risk of Fascioliasis infection and implement strategies to manage snail populations to ultimately reduce the risk of infection for humans and livestock.

Keywords

• Fasciola spp.
• Snail
• Molecular detection
• DNA diagnostics
• LAMP
• Environmental sampling