I watched the webinar tonight facilitated by Anna Maria Niewiadomska, Global Market Segment Manager in Public Health at Oxford Nanopore Technologies. They first spoke about using ONT for microbiology and infectious disease work. The turnaround time with the rapid library prep was one feature that Niewiadomska highlighted, as well as the real-time analysis and read-length ability. Niewiadomska highlighted bacterial genome sequencing (with the ONT NO-MISS protocol), full-length 16S, viral amplification, and metagenomics. The webinar focused on viral sequencing. Both targeted and untargeted pathogen sequencing approaches for viral whole genome sequencing have advantages and disadvantages. For example, Niewiadomska explained that an RNA virus is reverse transcribed to cDNA, and then tiled PCR amplification is used. The EPI2ME application has workflows that can be used for viral analysis. Niewiadomska summarized the ONT sequencing solutions available on one slide with a table with targeted and untargeted library prep and bioinformatics solutions. The first speaker was Daniel M. Maloney from NHS Lothian, who presented “RSVamplicon sequencing reveals global and local lineage dynamics.” They described RSV as a single-stranded RNA virus with two subtypes: RSV A and RSV B. The types are ~85% similar. RSV season starts around when the flu starts. Maloney mentioned that there has been renewed interest in RSB vaccination and surveillance. A primer set similar to that used for the ARTIC COVID system was used. A series of overlapping primer pairs was designed, and two pools were created. Maloney shared initial results indicating good coverage of the virus. Maloney noted that they designed primer sets for RSV A and RSV B, and the primer sets can be multiplexed but with a reduction of coverage. An EPI2ME workflow was developed for analysis. The research team obtained samples from several countries for testing. Maloney shared phylogenetic trees, comparing the sequencing results of viral genomes placed into phylogenetic trees. There is mixing of genomes and no clear clustering by geographic location. Interestingly, the RSV B genomes show more dominant genotypes at time points, whereas the RSV A viral genomes are more mixed at a given time. Maloney and team sequenced over 500 samples of RSV during the ’23-’24 season in Scotland. They examined the phylogenetic trees from closely related samples and patients to try to detect nosocomial transmission. They didn’t find any! However, Maloney mentioned one case of potential transmission of RSV B in a cluster of cases at Guy’s and St. Thomas’ Trust. Maloney went from developing an assay to improving knowledge of the circulating RSV genomes!
