Improving nitrofurantoin resistance prediction in Escherichia coli from whole-genome sequence by integrating NfsA/B enzyme assays.

Autor:	Dulyayangkul P; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom.; Laboratory of Biotechnology, Chulabhorn Research Institute, Bangkok, Thailand., Sealey JE; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom., Lee WWY; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom., Satapoomin N; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom., Reding C; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom., Heesom KJ; University of Bristol Proteomics Facility, Bristol, United Kingdom., Williams PB; University Hospitals Bristol and Weston NHS Foundation Trust, Bristol, United Kingdom., Avison MB; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom.
Jazyk:	angličtina
Zdroj:	Antimicrobial agents and chemotherapy [Antimicrob Agents Chemother] 2024 Jul 09; Vol. 68 (7), pp. e0024224. Date of Electronic Publication: 2024 May 20.
DOI:	10.1128/aac.00242-24
Abstrakt:	Nitrofurantoin resistance in Escherichia coli is primarily caused by mutations damaging two enzymes, NfsA and NfsB. Studies based on small isolate collections with defined nitrofurantoin MICs have found significant random genetic drift in nfsA and nfsB , making it extremely difficult to predict nitrofurantoin resistance from whole-genome sequence (WGS) where both genes are not obviously disrupted by nonsense or frameshift mutations or insertional inactivation. Here, we report a WGS survey of 200 oqxAB -negative E. coli from community urine samples, of which 34 were nitrofurantoin resistant. We characterized individual non-synonymous mutations seen in nfsA and nfsB among this collection using complementation cloning and NfsA/B enzyme assays in cell extracts. We definitively identified R203C, H11Y, W212R, A112E, and A112T in NfsA and R121C, Q142H, F84S, P163H, W46R, K57E, and V191G in NfsB as amino acid substitutions that reduce enzyme activity sufficiently to cause resistance. In contrast, E58D, I117T, K141E, L157F, A172S, G187D, and A188V in NfsA and G66D, M75I, V93A, and A174E in NfsB are functionally silent in this context. We identified that 9/166 (5.4%) nitrofurantoin-susceptible isolates were "pre-resistant," defined as having loss of function mutations in nfsA or nfsB . Finally, using NfsA/B enzyme assays and proteomics, we demonstrated that 9/34 (26.5%) ribE wild-type nitrofurantoin-resistant isolates also carried functionally wild-type nfsB or nfsB / nfsA . In these cases, NfsA/B activity was reduced through downregulated gene expression. Our biological understanding of nitrofurantoin resistance is greatly improved by this analysis but is still insufficient to allow its reliable prediction from WGS data. Competing Interests: The authors declare no conflict of interest.
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu