Trimethoprim is a synthetic antibacterial agent that targets folate biosynthesis by competitively binding to the di-hydrofolate reductase enzyme (DHFR). Trimethoprim is often administered synergistically with sulfonamide, another chemotherapeutic agent targeting the di-hydropteroate synthase (DHPS) enzyme in the same pathway. Clinical resistance to both drugs is widespread and mediated by enzyme variants capable of performing their biological function without binding to these drugs. These mutant enzymes were assumed to have arisen after the discovery of these synthetic drugs, but recent work has shown that genes conferring resistance to sulfonamide were present in the bacterial pangenome millions of years ago. Here, we apply phylogenetics and comparative genomics methods to study the largest family of mobile trimethoprim-resistance genes (dfrA). We show that most of the dfrA genes identified to date map to two large clades that likely arose from independent mobilization events. In contrast to sulfonamide resistance (sul) genes, we find evidence of recurrent mobilization in dfrA genes. Phylogenetic evidence allows us to identify novel dfrA genes in the emerging pathogen Acinetobacter baumannii , and we confirm their resistance phenotype in vitro. We also identify a cluster of dfrA homologues in cryptic plasmid and phage genomes, but we show that these enzymes do not confer resistance to trimethoprim. Our methods also allow us to pinpoint the chromosomal origin of previously reported dfrA genes, and we show that many of these ancient chromosomal genes also confer resistance to trimethoprim. Our work reveals that trimethoprim resistance predated the clinical use of this chemotherapeutic agent, but that novel mutations have likely also arisen and become mobilized following its widespread use within and outside the clinic. Hence, this work confirms that resistance to novel drugs may already be present in the bacterial pangenome, and stresses the importance of rapid mobilization as a fundamental element in the emergence and global spread of resistance determinants.

中文翻译：

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探索二氢叶酸还原酶基因的起源和动员以及甲氧苄啶临床耐药性的出现。


甲氧苄啶是一种合成抗菌剂，通过与二氢叶酸还原酶 (DHFR) 竞争性结合来靶向叶酸生物合成。甲氧苄啶通常与磺酰胺协同给药，磺酰胺是另一种针对同一途径中的二氢蝶酸合酶 (DHPS) 的化疗药物。对这两种药物的临床耐药性很普遍，并且是由能够在不与这些药物结合的情况下发挥其生物学功能的酶变体介导的。这些突变酶被认为是在这些合成药物发现后出现的，但最近的研究表明，赋予磺酰胺抗性的基因在数百万年前就存在于细菌全基因组中。在这里，我们应用系统发育学和比较基因组学方法来研究最大的移动甲氧苄啶抗性基因家族（ dfrA ）。我们表明，迄今为止鉴定的大多数dfrA基因都映射到两个可能由独立动员事件产生的大进化枝。与磺酰胺抗性（ sul ）基因相反，我们发现dfrA基因中反复动员的证据。系统发育证据使我们能够在新出现的病原体鲍曼不动杆菌中鉴定出新的dfrA基因，并在体外确认它们的耐药表型。我们还在隐性质粒和噬菌体基因组中鉴定了一组dfrA同源物，但我们表明这些酶不会赋予甲氧苄啶抗性。 我们的方法还使我们能够查明先前报道的dfrA基因的染色体起源，并且我们表明许多这些古老的染色体基因也赋予甲氧苄啶抗性。我们的工作表明，甲氧苄啶耐药性早于这种化疗药物的临床使用，但随着其在临床内外的广泛使用，新的突变也可能出现并被动员起来。因此，这项工作证实了细菌全基因组中可能已经存在对新药的耐药性，并强调了快速动员作为耐药性决定因素的出现和全球传播的基本要素的重要性。