Development and performance of a serological enzyme-linked immunosorbent assay (ELISA) for ruminant liver fluke disease
López Corrales, Jesús
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Fasciola hepatica, also known as the liver fluke, is the causative agent of fasciolosis. It is classified by the World Health Organisation as a neglected tropical disease, affecting both humans and livestock globally. Moreover, infections caused by this parasite cause multimillion economic losses to the livestock industry. In order to minimise the effects of F. hepatica infections by applying the appropriate management and control interventions, it is essential to include the use of a reliable diagnostic method that is also effective in determining infections as early as possible with high sensitivity and specificity. Among the available methods, we show that serum-ELISA based on the detection of antibodies against cathepsin L1 peptidase has a high efficacy detecting infections at every developmental stage of the flukes in their definitive hosts. However, discovering new antigens is essential to provide more options for the development of liver fluke diagnostics and vaccines, allowing us to use one or more antigens to differentiate infected from vaccinated animals (DIVA). In this work, we evaluated the performance of an array of molecules produced by F. hepatica as antigens for indirect ELISA tests in comparison with recombinant cathepsin L1 (rFhCL1). Four antigens, namely rFhGAPDH, rFhGPI, rFhEnolase and rFhFBPA, were recombinantly produced during this study and assessed in ELISA, along with several more recombinant antigens (rFhHDM, rFhLAP, rFhPrx, rFhSOD1, rFhSOD3, rFhTrx, rFhCL2, rFhStf1 and rFhKT1), to analyse IgG responses in sera samples of sheep and cattle that were both experimentally and naturally infected. None of the studied antigens performed better than rFhCL1 because of their lower immunogenicity. In addition, some of them showed high background levels due to antibody cross-reactivity. In the case of E. coli produced antigens, this was possibly caused by E. coli infections in the animals or natural antibodies targeting E. coli proteins. In the case of P. pastoris produced antigens, the cross-reactivity possibly resulted from glycans present in the recombinant proteins. These sugar residues may be recognised by host antibodies produced after being fed with yeast-rich supplemented food. Finally, we studied a potential method to determine liver fluke infection stages, namely by calculating the ratio anti-rFhCL1/anti-rFhCB3 immune responses, with promising results. Future investigation including a larger number of samples and complementary parasitological data will be key to further prove its applicability.