Lu Junwan, Zhang Runzhi, Yu Yan, Lou Hongqiang, Li Dong, Bao Qiyu, Feng Chunlin
Medical Molecular Biology Laboratory, School of Medicine, Jinhua University of Vocational Technology, Jinhua, China.
Key Laboratory of Medical Genetics of Zhejiang Province, Key Laboratory of Laboratory Medicine, Ministry of Education, School of Laboratory Medicine and Life Sciences, Institute of Biomedical Informatics, Wenzhou Medical University, Wenzhou, China.
Front Microbiol. 2025 Apr 15;16:1577167. doi: 10.3389/fmicb.2025.1577167. eCollection 2025.
Owing to the rapid emerging of multidrug-, even pandrug-resistant pathogens, and lack of new antibiotics, the older antibiotic, fosfomycin, has been reused in recent years in the clinical practice, especially for treatment of uropathogen infections. With the increased use of fosfomycin, bacterial resistance to it has also increased drastically. Elucidating the resistance mechanism to the antimicrobial has become an urgent task.
The putative fosfomycin resistance gene was cloned, and minimal inhibitory concentrations were determined by the agar dilution method. Enzyme kinetic parameters were measured by high-performance liquid chromatography. Bioinformatics analysis was applied to understand the evolutionary characteristics of FosC3.
The strain DW0021 exhibited high level resistance to several antimicrobials including kanamycin, streptomycin, chloramphenicol, florfenicol, tetracycline, and especially higher to fosfomycin (> 1,024 μg/mL), while genome annotation indicated that no function-characterized resistance gene was associated with fosfomycin resistance. A novel functional gene designated responsible for fosfomycin resistance was identified in the chromosome of DW0021. Among the function-characterized proteins, FosC3 shared the highest amino acid similarity of 58.65% with FosC2. No mobile genetic element (MGE) was found surrounding the gene. The recombinant pMD19-/DH5α displayed a MIC value of 32 μg/mL to fosfomycin, which revealed a 128-fold increase of MIC value to fosfomycin compared to the control pMD19/ DH5α (0.25 μg/mL). FosC3 was phylogenetically close to FosC2 and exhibited a and of 82,442 ± 1,475 s, 70.99 ± 4.31 μM, respectively, and a catalytic efficiency of (1.2 ± 0.3) × 10 μM·s.
In this work, a novel functional fosfomycin thiol transferase, FosC3, which shared the highest protein sequence similarity with FosC2, was identified in . The fosfomycin inactivation enzyme FosC3 could effectively inactivate fosfomycin by chemical modification. It is implied that such mechanism facilitates to respond to fosfomycin exposure, thereby enhancing survival. However, was not related with any MGE, which differs from many other fosfomycin thiol transferase genes. As a result, is not expected to be transmitted to other species through horizontal gene transfer mechanism. Our findings will contribute to the resistance mechanism of the common pathogenic .
由于多重耐药甚至泛耐药病原体的迅速出现，以及新抗生素的缺乏，古老的抗生素磷霉素近年来在临床实践中被重新使用，尤其是用于治疗尿路病原体感染。随着磷霉素使用的增加，细菌对其的耐药性也急剧上升。阐明对抗菌药物的耐药机制已成为一项紧迫任务。
克隆假定的磷霉素耐药基因，通过琼脂稀释法测定最低抑菌浓度。用高效液相色谱法测量酶动力学参数。应用生物信息学分析来了解FosC3的进化特征。
菌株DW0021对包括卡那霉素、链霉素、氯霉素、氟苯尼考、四环素在内的几种抗菌药物表现出高水平耐药，对磷霉素的耐药性尤其高（>1024μg/mL），而基因组注释表明没有功能特征明确的耐药基因与磷霉素耐药相关。在DW0021的染色体中鉴定出一个负责磷霉素耐药的新功能基因。在功能特征明确的蛋白质中，FosC3与FosC2的氨基酸相似性最高，为58.65%。在fosC3基因周围未发现移动遗传元件（MGE）。重组体pMD19-/DH5α对磷霉素的MIC值为32μg/mL，与对照pMD19/DH5α（0.25μg/mL）相比，其对磷霉素的MIC值增加了128倍。FosC3在系统发育上与FosC2接近，其Km和Vmax分别为82442±1475s、70.99±4.31μM，催化效率为（1.2±0.3）×10μM·s。
在本研究中，在[具体菌株名称未给出]中鉴定出一种新型功能性磷霉素硫醇转移酶FosC3，它与FosC2的蛋白质序列相似性最高。磷霉素失活酶FosC3可通过化学修饰有效地使磷霉素失活。这意味着这种机制有助于[具体菌株名称未给出]应对磷霉素暴露，从而提高存活率。然而，[具体菌株名称未给出]与任何MGE均无关联，这与许多其他磷霉素硫醇转移酶基因不同。因此，预计它不会通过水平基因转移机制传播到其他物种。我们的研究结果将有助于了解常见病原菌的耐药机制。
