I continued watching the “Enhanced microbial profiling and metagenomics with nanopore sequencing” webinar. The second speaker was Steven Batinovic, a Field Application Scientist with Oxford Nanopore Technologies (ONT). The title of their session was “What you’re missing matters: delivering the future of microbial genomics with Oxford Nanopore.” Batinovic started by listing the microbiology and infectious disease applications of ONT devices. Batinovic shared examples of the use of nanopore sequencing for surveillance work. The Nanopore Only Microbial Isolate Sequencing Solution (NO-MISS) workflow can be used to generate high-quality genomes, resolve plasmids, and identify antimicrobial resistance genes (AMR). Stitching long reads together is easier than shorter reads. Batinovic described how spanning repetitive regions with long reads can help resolve bacterial genomes and any plasmids. I had not considered plasmid identification with NO-MISS workflows. Updated base calling models have improved genome accuracy, consensus assembly accuracy, and identification of modified bases. Ryan Wick’s work on bacterial genome assemblies demonstrates a consensus Q score average for a set of microbes of Q 60! The end-to-end microbial isolate WGS protocols Nanopore has include extraction methods for various organisms. The EPI2ME workflow typically uses Flye and polishes with Medaka. If a reference is provided, alignment is performed, followed by polishing. Additional options with the EPI2ME wf-bacterial-genomes include MLST and AMR detection. Gene annotation with Prokka and Salmonella serotyping are options, too. Batinovic shared the results of a bacterial outbreak analysis with v14 ONT chemistry. Susceptibility testing matched genome results. For 16S sequencing, Nanopore allows for full-length 16S sequencing. The 16S barcoding kit uses rapid attachment tags present on the primers. The EPI2mE workflow wf-16S can use Kraken2 or minimap2. Minimap2 provides resolution at the expense of compute time.
