Ma Yibing, Pirolo Mattia, Giarratana Lily, Leth Nielsen Karen, Häussler Susanne, Guardabassi Luca
Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg, Denmark.
Department of Clinical Microbiology, Rigshospitalet, København Ø, Denmark.
Antimicrob Agents Chemother. 2025 Apr 2;69(4):e0141724. doi: 10.1128/aac.01417-24. Epub 2025 Feb 25.
acquires fosfomycin resistance through chromosomal mutations that reduce drug uptake and by drug-inactivating enzymes. However, the complete resistance mechanisms remain to be elucidated. The aim of this study was to elucidate the genetic mechanisms regulating fosfomycin susceptibility in uropathogenic (UPEC). We constructed a highly saturated transposon mutant library containing >340,000 unique Tn5 insertions in a clinical UPEC strain. We conducted transposon-directed insertion site sequencing (TraDIS) to screen for chromosomal genes whose mutations are beneficial for bacterial growth and survival in the presence of fosfomycin at 4 and 32 µg/mL. TraDIS analysis identified 67 genes including known resistance determinants ( = 13) as well as a set of novel genes modulating fosfomycin susceptibility ( = 54). These genes are involved in pyruvate metabolism, pentose phosphate pathway, nucleotide biosynthesis, DNA repair, protein translation, cellular iron homeostasis, and biotin biosynthesis. Deletion of 16 selected genes in the wild-type strain resulted in growth advantages and decreased susceptibility when exposed to fosfomycin. Notably, deletion of DNA repair genes (i.e. and ) and purine synthesis genes (i.e., and its upstream gene ) led to the most significant advantages in competitive and non-competitive growth in the presence of fosfomycin, as well as the highest increase of fosfomycin MIC (8- to 16-fold). These findings provide a genome-wide overview of genes required for maintaining fosfomycin susceptibility in , highlighting new mutations and functional pathways that may be used by UPEC to develop clinical resistance.
通过减少药物摄取的染色体突变和药物失活酶获得对磷霉素的抗性。然而，完整的抗性机制仍有待阐明。本研究的目的是阐明调节尿路致病性大肠杆菌（UPEC）中磷霉素敏感性的遗传机制。我们构建了一个高度饱和的转座子突变体文库，在一株临床UPEC菌株中包含>340,000个独特的Tn5插入。我们进行了转座子定向插入位点测序（TraDIS），以筛选其突变对在4和32μg/mL磷霉素存在下细菌生长和存活有益的染色体基因。TraDIS分析鉴定出67个基因，包括已知的抗性决定因素（n = 13）以及一组调节磷霉素敏感性的新基因（n = 54）。这些基因参与丙酮酸代谢、磷酸戊糖途径、核苷酸生物合成、DNA修复、蛋白质翻译、细胞铁稳态和生物素生物合成。在野生型菌株中缺失16个选定的基因导致在暴露于磷霉素时生长优势和敏感性降低。值得注意的是，缺失DNA修复基因（即 和 ）和嘌呤合成基因（即， 和其上游基因 ）在存在磷霉素的情况下在竞争性和非竞争性生长中导致最显著的优势，以及磷霉素MIC的最高增加（8至16倍）。这些发现提供了在UPEC中维持磷霉素敏感性所需基因的全基因组概述，突出了UPEC可能用于产生临床抗性的新突变和功能途径。
