This afternoon, I came back from visiting KBase in Berkeley. I am still watching sessions from previous London Callings to learn about different topics I will discuss in the upcoming Portable Genome Sequencing course. Tonight, I found the session by David R. Greig from Public Health England in the UK. Their 2021 London Calling session was entitled “Accessing the accessory genome to better understand the diversity of Shiga toxin-producing Escherichia coli 0157:H7 during an outbreak.” They used Oxford Nanopore Technologies to study Shiga-toxin producing Escherichia coli (STEC). STEC are characterized by prophage encoded Shiga toxin. They culture and isolate STEC, extract DNA, and sequence on Illumina to characterize STEC. Grieg and team wanted to retrospectively sequence samples from outbreaks and use nanopore sequencing. DNA was extracted with the Revolution Fire Monkey kit that I had never heard of… They used a MinION Mk1B and MIN106 flow cells with the Rapid Sequencing/Barcoding Kit. They used two pipelines, starting with FAST5 files and performing read hygiene, reference-based mapping or characterization through assembly using Flye. For an outbreak associated with raw pet food, the sequencing indicated variability in pro phages. For an outbreak associated with a mud-based obstacle, samples varied in plasmid content. An outbreak associated with raw drinking milk had samples from several sources. Sequencing identified an stx2a-encoding prophage. This information provided new insights about the accessory genome of STEC. Importantly, rapid characterization of outbreak samples can help to learn about genetic factors affecting outbreak strain evolution. I learned about STEC and variability of stx2a location and plasmid content.
