Optimized methods for the targeted surveillance of extended-spectrum beta-lactamase-producing <i>Escherichia coli</i> in human stool

Gallichan, Sarah, Forrest, Sally, Picton-Barlow, Esther, McKeown, Claudia, Moore, Maria, Heinz, Eva ORCID: https://orcid.org/0000-0003-4413-3756, Feasey, Nicholas ORCID: https://orcid.org/0000-0003-4041-1405, Lewis, Joseph ORCID: https://orcid.org/0000-0002-3837-5188 and Graf, Fabrice (2024) 'Optimized methods for the targeted surveillance of extended-spectrum beta-lactamase-producing <i>Escherichia coli</i> in human stool'. Microbiology Spectrum. (In Press)

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Abstract

Understanding transmission pathways of important opportunistic, drug-resistant pathogens, such as extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli, is essential to implementing targeted prevention strategies to interrupt transmission and reduce the number of infections. To link transmission of ESBL-producing E. coli (ESBL-EC) between two sources, single-nucleotide resolution of E. coli strains, as well as E. coli diversity within and between samples, is required. However, the microbiological methods to best track these pathogens are unclear. Here, we compared different steps in the microbiological workflow to determine the impact different pre-enrichment broths, pre-enrichment incubation times, selection in pre-enrichment, selective plating, and DNA extraction methods had on recovering ESBL-EC from human stool samples, with the aim to acquire high-quality DNA for sequencing and genomic epidemiology. We demonstrate that using a 4-h pre-enrichment in Buffered Peptone Water, plating on cefotaxime-supplemented MacConkey agar and extracting DNA using Lucigen MasterPure DNA Purification kit improves the recovery of ESBL-EC from human stool and produced high-quality DNA for whole-genome sequencing. We conclude that our optimized workflow can be applied for single-nucleotide variant analysis of an ESBL-EC from stool.

Item Type:	Article
Subjects:	WC Communicable Diseases > Infection. Bacterial Infections > Enteric Infections > WC 290 Escherichia coli infections
Faculty: Department:	Biological Sciences > Vector Biology Department Clinical Sciences & International Health > Clinical Sciences Department
Digital Object Identifer (DOI):	https://doi.org/10.1128/spectrum.01058-24
SWORD Depositor:	JISC Pubrouter
Depositing User:	JISC Pubrouter
Date Deposited:	17 Dec 2024 08:25
Last Modified:	17 Dec 2024 08:25
URI:	https://archive.lstmed.ac.uk/id/eprint/25713

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