Detection and differentiation of haemosporidian parasites in chickens (Gallus gallus domesticus) using molecular diagnostic approach

Abstract

Haemosporidioses in domestic chicken (Gallus gallus domesticus) are not only widely distribute but causing high mortality and non-specific symptoms. The multiplex PCR assay developed in this study can concurrently detect and differentiate four haemosporidian parasites Leucocytozoon caulleryi, Leucocytozoon sabrazesi, Plasmodium juxtanucleare, and Plasmodium gallinaceum. Comparing to microscopic examination performed in parallel on 157 samples, multiplex PCR assay exhibited significantly higher detection rate in diagnosis of haemosporidians (p = 0.000). The results showed that, L. sabrazesi, P. juxtanucleare, and P. gallinaceum detection rate of multiplex PCR were all higher than that of microscopic examination with p = 0.002, 0.000, and 0.004, respectively. Detection limit of multiplex PCR on quadruple targets mixture was 109 copies/µl. Multiple infections were also detected more effectively by multiplex PCR. The commonness of mix infection in chicken haemosporidian parasites was further confirm as the new method have been applied to field samples from Nan province, Prachinburi province, and Chachoengsao province, in which mix infections were detected in all three sampling sites, ranging from 4% to 37%. Overall, infection rate of haemosporidian parasites in chickens from three province were 77.2%, 93.8%, and 39.5%, respectively, by using multiplex PCR. This study contributes a protocol to detect all four avian haemosporidian parasites in chicken, either single or mixed infections.