Kame A. Galan-Huerta from the Autonomous University of Nuevo Leon in Mexico presented at the Nanopore Community Meeting 2022 a ten minute session entitled “Identification of resistance mutations to direct-acting antiviral agents against HCV in infected subjects in Mexico.” They spoke about adapting protocols to detect the RNA positive sense hepatitis C virus. Glan-Huerta explained that antivirals inhibit certain non-structural proteins and viral replication. They also noted that in Mexico the predominant genotype is 1B. The team adapted the Primalscheme and Josh Quick protocols to design multiplex PCR. They proceeded to amplify from serum samples. They synthesized cDNA with SuperScript IV. The had to create three pools because they had some primer dimers. They used the LSK-109 ligation sequencing kit and the native barcoding kit. The team used R9.4 flow cells in a MinION. They used a reference and Minimap2 and Medaka to assemble and polish. They then used HCV-GLUE to search for mutations related to drug resistance. They sequenced six samples and identified two mutations related to resistance in one sample. The team then created a maximum clade credibility tree and identified a clade from Mexico. The team wants to sequence more to create a phylogeographic evaluation. It was nice to learn about research from my country and applications of tools developed for other viruses. I learned about HCV-GLUE and phylogeographic analyses with this session.
