Tilapia lake virus segment 1 PCR amplification and nanopore sequencing

Rapid and accurate diagnosis of TiLV is critical to facilitate timely response to prevent its widespread and deploy of adequate control measures. Currently, most laboratories use PCR to amplify partial pathogen genomic regions combined with sequencing of PCR amplicon(s) using conventional Sanger sequencing services for confirmatory diagnosis. The main limitation of this approach is time constraint (approx. 5-7 days) when a Sanger platform is available. Here, we report an innovative approach using specific-PCR targeting TiLV partial segment 1 region, coupled with Minion, a unique portable sequencing device from Oxford Nanopore Technologies (ONT). Using clinical samples from infected fish, we applied this approach for the rapid confirmation of TiLV in less than 12 h. The obtained sequences match near perfectly to Illumina and Sanger-verified references (99.83-100 % identity), making it suitable for rapid diagnostic of TiLV. We were also able to identify one single nucleotide polymorphism in one of the isolate. Taken together, this study suggests that PCR-MinION is a promising platform to deploy in regional aquatic animal health diagnostic laboratories in low and medium income countries, for fast and accurate confirmation and "genotyping" of emerging infectious pathogens from field samples within a single day.