The purpose of the study was to characterize the resistome, virulome, mobilome and Clustered Regularly Interspaced Short Palindromic Repeats-associated (CRISPR-Cas) system of extended-spectrum β-lactamase producing Klebsiella pneumoniae (ESBL-KP) clinical isolates and to determine their phylogenetic relatedness. The isolates were from Algeria, isolated at the University Hospital Establishment of Oran, between 2011 and 2012. ESBL-KP isolates (n = 193) were screened for several antibiotic resistance genes (ARGs) using qPCR followed by Pulsed-Field Gel Electrophoresis (PFGE). Representative isolates were selected from PFGE clusters and subjected to whole-genome sequencing (WGS). Genomic characterization of the WGS data by studying prophages, CRISPR-Cas systems, Multi-Locus Sequence Typing (MLST), serotype, ARGs, virulence genes, plasmid replicons, and their pMLST. Phylogenetic and comparative genomic were done using core genome MLST and SNP-Based analysis. Generally, the ESBL-KP isolates were polyclonal. The whole genome sequences of nineteen isolates were taken of main PFGE clusters. Sixteen sequence types (ST) were found including high-risk clones ST14, ST23, ST37, and ST147. Serotypes K1 (n = 1), K2 (n = 2), K3 (n = 1), K31 (n = 1), K62 (n = 1), and K151 (n = 1) are associated with hyper-virulence. CRISPR-Cas system was found in 47.4%, typed I-E and I-E*. About ARGs, from 193 ESBL-KP, the majority of strains were multidrug-resistant, the CTX-M-1 enzyme was predominant (99%) and the prevalence of plasmid-mediated quinolone resistance (PMQR) genes was high with aac(6')-lb-cr (72.5%) and qnr's (65.8%). From 19 sequenced isolates we identified ESBL, AmpC, and carbapenemase genes: blaCTX-M-15 (n = 19), blaOXA-48 (n = 1), blaCMY-2 (n = 2), and blaCMY-16 (n = 2), as well as non-ESBL genes: qnrB1 (n = 12), qnrS1 (n = 1) and armA (n = 2). We found IncF, IncN, IncL/M, IncA/C2, and Col replicon types, at least once per isolate. This study is the first to report qnrS in ESBL-KP in Algeria. Our analysis shows the concerning co-existence of virulence and resistance genes and would support that genomic surveillance should be a high priority in the hospital environment.

中文翻译：

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从阿尔及利亚奥兰大学医院中分离的产超广谱 β-内酰胺酶肺炎克雷伯菌的高风险克隆（2011-2012）。

本研究的目的是表征产超广谱 β-内酰胺酶肺炎克雷伯菌 (ESBL-KP) 临床分离株的抗性组、病毒组、移动组和成簇规则间隔短回文重复相关 (CRISPR-Cas) 系统系统发育相关性。分离株来自阿尔及利亚，于 2011 年至 2012 年在奥兰大学医院建立。使用 qPCR 和脉冲场凝胶电泳 (PFGE) 筛选 ESBL-KP 分离株（n = 193）的几种抗生素抗性基因 (ARG) ）。从 PFGE 簇中选择代表性分离株并进行全基因组测序 (WGS)。通过研究原噬菌体、CRISPR-Cas 系统、多位点序列分型 (MLST)、血清型、ARG、毒力基因、质粒复制子、和他们的 pMLST。使用核心基因组 MLST 和基于 SNP 的分析进行系统发育和比较基因组。通常，ESBL-KP 分离物是多克隆的。19 个分离株的全基因组序列取自主要的 PFGE 簇。发现了 16 种序列类型 (ST)，包括高风险克隆 ST14、ST23、ST37 和 ST147。血清型 K1 (n = 1)、K2 (n = 2)、K3 (n = 1)、K31 (n = 1)、K62 (n = 1) 和 K151 (n = 1) 与超毒力相关。47.4% 的 CRISPR-Cas 系统被发现，类型为 IE 和 IE*。关于 ARG，从 193 ESBL-KP 开始，大多数菌株具有多重耐药性，CTX-M-1 酶占主导地位（99%），质粒介导的喹诺酮耐药（PMQR）基因的流行率很高，aac（6 ')-lb-cr (72.5%) 和 qnr (65.8%)。我们从 19 个测序的分离株中鉴定出 ESBL、AmpC 和碳青霉烯酶基因：blaCTX-M-15 (n = 19)、blaOXA-48 (n = 1)、blaCMY-2 (n = 2) 和 blaCMY-16 (n = 2)，以及非 ESBL 基因：qnrB1 (n = 12)、qnrS1 (n = 1) 和 armA (n = 2)。我们发现了 IncF、IncN、IncL/M、IncA/C2 和 Col 复制子类型，每个分离株至少一次。本研究首次报道了阿尔及利亚 ESBL-KP 中的 qnrS。我们的分析显示了毒力和抗性基因的相关共存，并支持基因组监测应成为医院环境中的高度优先事项。