Institute of Molecular Biology & Biotechnology (IMBB), University of Lahore, Lahore 54590, Pakistan.
Institute of Microbiology, University of Veterinary and Animal Sciences, Lahore 54000, Pakistan.
Medicina (Kaunas). 2024 Jul 2;60(7):1086. doi: 10.3390/medicina60071086.
(), particularly carbapenem-resistant (CRAB), represents a grave concern in healthcare settings and is associated with high mortality. This study aimed to conduct molecular, mutational, and phylogenetic analyses of specific genes in CRAB and evaluate the synergistic effects of selected antimicrobial combinations. : Phenotypic characterization was performed on six CRAB strains by using the Modified Hodge Test (MHT) and IMP-EDTA Double-Disc Synergy Test (IMP-EDTA DDST). Carbapenemase- and metallo-beta-lactamase-encoding genes were amplified by using Polymerase Chain Reaction. Phylogenetic analysis using the MEGA 11 tool was used to determine the evolutionary relatedness of these genes. Mutational analysis was performed by using I-Mutant, MUPro, and PHD-SNP bioinformatics tools to predict mutations in the carbapenemase-encoding genes. Microdilution checkerboard titration assessed the synergistic effects of antimicrobial combinations (azithromycin-meropenem, rifampicin-meropenem, meropenem-colistin, and azithromycin-colistin) on these CRAB isolates. : The phenotypic characterization of six CRAB isolates revealed positive results for MHT and IMP-EDTA DDST. The molecular characterization revealed that carbapenemase- and MBL-encoding genes were present in all isolates with varying frequencies, including blaOXA-51 (100%) and blaIMP (0%). The sequence analysis revealed high evolutionary relatedness to sequences in the NCBI database. The mutational analysis identified 16 mutations, of which 1 mutation (P116L) in the blaOXA-58 gene predicted a change in the protein product, potentially contributing to carbapenem resistance. The checkerboard titration method did not reveal any synergism among the tested antimicrobial combinations against CRAB. : This study's findings underscore the significant challenges posed by CRAB isolates harboring multiple resistant genes in treatment. This highlights the urgent need for novel antimicrobial agents, a crucial step towards reducing mortality rates not only in Pakistan but also globally.
（碳青霉烯类耐药肠杆菌科细菌，CRAB）在医疗环境中是一个严重的问题，与高死亡率有关。本研究旨在对 CRAB 中特定基因进行分子、突变和系统发育分析，并评估选定抗菌组合的协同作用。：使用改良霍格试验（MHT）和 IMP-EDTA 双碟协同试验（IMP-EDTA DDST）对 6 株 CRAB 菌株进行表型特征分析。使用聚合酶链反应扩增碳青霉烯酶和金属β-内酰胺酶编码基因。使用 MEGA 11 工具进行系统发育分析，以确定这些基因的进化关系。使用 I-Mutant、MUPro 和 PHD-SNP 生物信息学工具进行突变分析，以预测碳青霉烯酶编码基因中的突变。微量稀释棋盘滴定法评估抗菌组合（阿奇霉素-美罗培南、利福平-美罗培南、美罗培南-黏菌素和阿奇霉素-黏菌素）对这些 CRAB 分离株的协同作用。：对 6 株 CRAB 分离株的表型特征分析显示 MHT 和 IMP-EDTA DDST 呈阳性结果。分子特征分析显示，所有分离株均存在不同频率的碳青霉烯酶和 MBL 编码基因，包括 blaOXA-51（100%）和 blaIMP（0%）。序列分析显示与 NCBI 数据库中的序列具有高度进化关系。突变分析确定了 16 个突变，其中 blaOXA-58 基因中的 1 个突变（P116L）预测蛋白产物发生变化，可能导致碳青霉烯类耐药。棋盘滴定法未发现测试的抗菌组合对 CRAB 有任何协同作用。：本研究结果强调了携带多种耐药基因的 CRAB 分离株在治疗中带来的重大挑战。这突显了迫切需要新型抗菌药物的必要性，这是降低死亡率的关键一步，不仅在巴基斯坦，而且在全球范围内。
