Validation of a High-Throughput Microfluidic Real-Time PCR for the Detection of Vector-Borne Agents in Wild Birds from the Brazilian Pantanal.

Despite numerous studies on haemosporidians in wild birds from Brazil, the presence of other vector-borne agents (VBA) such as spp., spp., and Onchocercidae filariids in avian hosts remains largely unknown. The low occurrence of these VBAs might be due to the low sensitivity of traditional molecular techniques. The microfluidic real-time PCR assay, known for its high sensitivity, has emerged as a promising method to detect and study the occurrence and diversity of VBAs in both arthropod vectors and vertebrate hosts. To validate previously and standardize newly designed microfluidic real-time PCR protocols, selected positive avian blood DNA samples for spp., spp., haemosporidians, and filariids were used. The molecular occurrence rates for the selected VBAs were 18.2% for spp., 0.36% for spp., 6.2% for spp., 4.7% for spp., and 6.5% for Onchocercidae filariids. The spp. B sequence detected in a clustered with , whereas the spp. sequence detected in a clustered with . While Onchocercidae filariid 1 sequences were detected in specimens of , and grouped with spp., one sequence detected in was ancestral to the clade comprising spp. and spp. High-throughput microfluidic real-time PCR assay can be used for screening VBAs in avian hosts from South America, but new primers/probe sets should be designed for VBA genotypes present in Brazil.