Rudder, S. | bioRxiv Science

bioRxiv2023

Uropathogenic Escherichia coli population structure and antimicrobial susceptibility in Norfolk, UK

SynopsisO_ST_ABSBackgroundC_ST_ABSHalf of all women have experienced a urinary tract infection (UTI) in their lifetime and this remains a persistent issue in rural counties like Norfolk, UK. In alignment globally, Uropathogenic E. coli (UPEC) are the main etiological agent for UTIs in Norfolk and are increasingly difficult to treat due to multi-drug resistance (MDR). ObjectiveWe set out to identify which clonal groups and resistance genes are disseminating in the community and hospitals in Norfolk, the first study of its kind for UPEC in this region. MethodsWe collected 217 clinical E. coli isolates causing UTIs in the community and hospital from the Clinical Microbiology laboratory at Norfolk and Norwich University Hospital. These were whole genome sequenced using the Illumina and MinION platforms for in silico multi-locus sequence typing and antibiotic resistance determinant detection. ResultsThe isolates were composed of 74 sequence types (STs); 8 lineages represented 57% of this population: ST73, ST12, ST69, ST131, ST404, ST95, ST127, and ST1193. Importantly, primary UTI screening deemed 8% of isolates to be MDR, with high rates of resistance to ampicillin (52.1%) and trimethoprim (36.2%) in hospitals. Of concern is the probable clonal expansion of MDR groups ST131 and ST1193 in hospitals and community settings with chromosomally encoded blaCTX-M-15, blaOXA-1, and aac(6)-Ib-cr5. ConclusionsThe burden of reported UTIs in Norfolk is largely caused by non-MDR isolates. The UPEC population is continually evolving, and monitoring samples with consideration of sources will help reduce burden of disease.

microbiology↗

bioRxiv2020

CoronaHiT: large scale multiplexing of SARS-CoV-2 genomes using Nanopore sequencing

The COVID-19 pandemic has spread to almost every country in the world since it started in China in late 2019. Controlling the pandemic requires a multifaceted approach including whole genome sequencing to support public health interventions at local and national levels. One of the most widely used methods for sequencing is the ARTIC protocol, a tiling PCR approach followed by Oxford Nanopore sequencing (ONT) of up to 96 samples at a time. There is a need, however, for a flexible, platform agnostic, method that can provide multiple throughput options depending on changing requirements as the pandemic peaks and troughs. Here we present CoronaHiT, a method capable of multiplexing up to 96 small genomes on a single MinION flowcell or >384 genomes on Illumina NextSeq, using transposase mediated addition of adapters and PCR based addition of barcodes to ARTIC PCR products. We demonstrate the method by sequencing 95 and 59 SARS-CoV-2 genomes for routine and rapid outbreak response runs, respectively, on Nanopore and Illumina platforms and compare to the standard ARTIC LoCost nanopore method. Of the 154 samples sequenced using the three approaches, genomes with [≥] 90% coverage (GISAID criteria) were generated for 64.3% of samples for ARTIC LoCost, 71.4% for CoronaHiT-ONT, and 76.6% for CoronaHiT-Illumina and have almost identical clustering on a maximum likelihood tree. In conclusion, we demonstrate that CoronaHiT can multiplex up to 96 SARS-CoV-2 genomes per MinION flowcell and that Illumina sequencing can be performed on the same libraries, which will allow significantly higher throughput. CoronaHiT provides increased coverage for higher Ct samples, thereby increasing the number of high quality genomes that pass the GISAID QC threshold. This protocol will aid the rapid expansion of SARS-CoV-2 genome sequencing globally, to help control the pandemic.

genomics↗