Cafiso Viviana, Stracquadanio Stefano, Lo Verde Flavia, Gabriele Giacoma, Mezzatesta Maria Lina, Caio Carla, Pigola Giuseppe, Ferro Alfredo, Stefani Stefania
Department of Biomedical and Biotechnological Sciences, University of Catania, Catania, Italy.
Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy.
Front Microbiol. 2019 Jan 7;9:3195. doi: 10.3389/fmicb.2018.03195. eCollection 2018.
Even though colistin-based treatment represents the antimicrobial-regimen backbone for the management of multidrug-resistant Gram-negative infections, colistin resistance is still rare, at least as a full resistance, in (). We investigated the genomics and transcriptomics of two clinical Extensively Drug Resistance (XDR) colistin-susceptible/resistant (COL-S/R) strain-pairs in which COL-resistance was developed after exposure to colistin therapy. The molecular characterization of the strains showed that all strains belonged to PFGE-A, ST-281, OXA-23 producers, Global Clone-II, and were resistant to imipenem, meropenem, ampicillin/sulbactam, ciprofloxacin, gentamicin, amikacin, trimethoprim/sulfamethoxazole, and susceptible to tigecycline, in agreement with NGS-acquired resistome. COL-R vs. COL-S comparative genomics, mapping on ATCC 17978 and ACICU Reference Genomes, revealed a closely related genomic phylogeny, especially between strain-pair isolates, and distinctive common genomic non-synonymous SNPs (nsSNPs) in COL-R strains. Furthermore, and nsSNPs were found. Notably we recovered, for the first time, and nsSNPs previously described only in "" mutants and associated with colistin resistance in a clinical COL-R . COL-R vs. COL-S comparative transcriptomics evidenced a strain-dependent response to the colistin resistance onset highly variable among the single COL-R strains vs. their COL-S parents and merely seven common over-expressed transcripts, i.e. the PgaB lipoprotein for biofilm-matrix production, the diacylglycerol kinase for the lipid recycling in the membrane-derived oligosaccharide cycle, a membrane non-ribosomal peptide synthetase, the Lipid A phosphoethanol aminotransferase PmrC, and three hypothetical proteins. The transcript analysis of the "COL-R related genes" and the RNA-seq data confirmed over-expression responsible for a greater positive net cell-charge, and under-expression in COL-R causing a decreased LPS production, as main mechanisms of colistin resistance. Our study reports the COL-R genomic and transcriptomic signatures reflecting the interplay between several direct and indirect potential adaptations to antimicrobial pressure, including the occurrence of SNP accumulation hotspot loci in genes related to intrinsic or adaptive colistin resistance, surface adhesion proteins and porins, and over-expressed genes involved in different pathways, i.e. biofilm production, oxidative stress response, extensive drug and COL resistance.
尽管基于黏菌素的治疗是耐多药革兰氏阴性菌感染管理的抗菌治疗方案的核心，但黏菌素耐药性仍然罕见，至少作为完全耐药性，在（）中是这样。我们研究了两对临床广泛耐药（XDR）黏菌素敏感/耐药（COL-S/R）菌株的基因组学和转录组学，其中COL耐药性是在接受黏菌素治疗后产生的。菌株的分子特征表明，所有菌株均属于PFGE-A、ST-281、OXA-23产生菌、全球克隆-II，对亚胺培南、美罗培南、氨苄西林/舒巴坦、环丙沙星、庆大霉素、阿米卡星、甲氧苄啶/磺胺甲恶唑耐药，对替加环素敏感，这与通过二代测序获得的耐药组一致。COL-R与COL-S的比较基因组学，映射到ATCC 17978和ACICU参考基因组上，揭示了密切相关的基因组系统发育关系，特别是在菌株对分离株之间，以及COL-R菌株中独特的常见基因组非同义单核苷酸多态性（nsSNPs）。此外，还发现了（具体的nsSNPs）。值得注意的是，我们首次发现了（具体的nsSNPs），这些nsSNPs以前仅在“（相关突变体）”突变体中描述过，并与临床COL-R中的黏菌素耐药性相关。COL-R与COL-S的比较转录组学表明，在单个COL-R菌株与其COL-S亲本之间，对黏菌素耐药性产生的菌株依赖性反应高度可变，仅有七个共同过度表达的转录本，即用于生物膜基质产生的PgaB脂蛋白、用于膜衍生寡糖循环中脂质回收的二酰基甘油激酶、一种膜非核糖体肽合成酶、脂质A磷酸乙醇胺转移酶PmrC以及三种假设蛋白。对“COL-R相关基因”的转录分析和RNA测序数据证实，（某些基因）过度表达导致细胞净正电荷增加，以及（某些基因）在COL-R中表达不足导致脂多糖产生减少，这是黏菌素耐药性的主要机制。我们的研究报告了COL-R的基因组和转录组特征，反映了几种直接和间接潜在适应抗菌压力之间的相互作用，包括与固有或适应性黏菌素耐药性、表面粘附蛋白和孔蛋白相关基因中SNP积累热点位点的出现，以及参与不同途径（即生物膜产生、氧化应激反应、广泛耐药和COL耐药）的过度表达基因。
