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Abstract：
BACKGROUND: Klebsiella pneumoniae, a notorious pathogen for causing nosocomial infections has become a major cause of neonatal septicemia, leading to high morbidity and mortality worldwide. This opportunistic bacterium has become highly resistant to antibiotics due to the widespread acquisition of genes encoding a variety of enzymes such as extended-spectrum beta-lactamases (ESBLs) and carbapenemases. We collected Klebsiella pneumoniae isolates from a local tertiary care hospital from February 2019-February 2021. To gain molecular insight into the resistome, virulome, and genetic environment of significant genes of multidrug-resistant K. pneumoniae isolates, we performed the short-read whole-genome sequencing of 10 K. pneumoniae isolates recovered from adult patients, neonates, and hospital tap water samples.
RESULTS: The draft genomes of the isolates varied in size, ranging from 5.48 to 5.96 Mbp suggesting the genome plasticity of this pathogen. Various genes conferring resistance to different classes of antibiotics e.g., aminoglycosides, quinolones, sulfonamides, tetracycline, and trimethoprim were identified in all sequenced isolates. The highest resistance was observed towards carbapenems, which has been putatively linked to the presence of both class B and class D carbapenemases, blaNDM, and blaOXA, respectively. Moreover, the biocide resistance gene qacEdelta1 was found in 6/10 of the sequenced strains. The sequenced isolates exhibited a broad range of sequence types and capsular types. The significant antibiotic resistance genes (ARGs) were bracketed by a variety of mobile genetic elements (MGEs). Various spontaneous mutations in genes other than the acquired antibiotic-resistance genes were observed, which play an indirect role in making these bugs resistant to antibiotics. Loss or deficiency of outer membrane porins, combined with ESBL production, played a significant role in carbapenem resistance in our sequenced isolates. Phylogenetic analysis revealed that the study isolates exhibited evolutionary relationships with strains from China, India, and the USA suggesting a shared evolutionary history and potential dissemination of similar genes amongst the isolates of different origins.
CONCLUSIONS: This study provides valuable insight into the presence of multiple mechanisms of carbapenem resistance in K. pneumoniae strains including the acquisition of multiple antibiotic-resistance genes through mobile genetic elements. Identification of rich mobilome yielded insightful information regarding the crucial role of insertion sequences, transposons, and integrons in shaping the genome of bacteria for the transmission of various resistance-associated genes. Multi-drug resistant isolates that had the fewest resistance genes exhibited a significant number of mutations. K. pneumoniae isolate from water source displayed comparable antibiotic resistance determinants to clinical isolates and the highest number of virulence-associated genes suggesting the possible interplay of ARGs amongst bacteria from different sources.
RESULTS: The draft genomes of the isolates varied in size, ranging from 5.48 to 5.96 Mbp suggesting the genome plasticity of this pathogen. Various genes conferring resistance to different classes of antibiotics e.g., aminoglycosides, quinolones, sulfonamides, tetracycline, and trimethoprim were identified in all sequenced isolates. The highest resistance was observed towards carbapenems, which has been putatively linked to the presence of both class B and class D carbapenemases, blaNDM, and blaOXA, respectively. Moreover, the biocide resistance gene qacEdelta1 was found in 6/10 of the sequenced strains. The sequenced isolates exhibited a broad range of sequence types and capsular types. The significant antibiotic resistance genes (ARGs) were bracketed by a variety of mobile genetic elements (MGEs). Various spontaneous mutations in genes other than the acquired antibiotic-resistance genes were observed, which play an indirect role in making these bugs resistant to antibiotics. Loss or deficiency of outer membrane porins, combined with ESBL production, played a significant role in carbapenem resistance in our sequenced isolates. Phylogenetic analysis revealed that the study isolates exhibited evolutionary relationships with strains from China, India, and the USA suggesting a shared evolutionary history and potential dissemination of similar genes amongst the isolates of different origins.
CONCLUSIONS: This study provides valuable insight into the presence of multiple mechanisms of carbapenem resistance in K. pneumoniae strains including the acquisition of multiple antibiotic-resistance genes through mobile genetic elements. Identification of rich mobilome yielded insightful information regarding the crucial role of insertion sequences, transposons, and integrons in shaping the genome of bacteria for the transmission of various resistance-associated genes. Multi-drug resistant isolates that had the fewest resistance genes exhibited a significant number of mutations. K. pneumoniae isolate from water source displayed comparable antibiotic resistance determinants to clinical isolates and the highest number of virulence-associated genes suggesting the possible interplay of ARGs amongst bacteria from different sources.
摘要：
背景：肺炎克雷伯菌，引起医院感染的臭名昭著的病原体已成为新生儿败血症的主要原因，导致世界范围内的高发病率和高死亡率。由于广泛获得编码多种酶如超广谱β-内酰胺酶(ESBLs)和碳青霉烯酶的基因,这种机会细菌对抗生素具有高度抗性。我们从2019年2月至2021年2月收集了当地三级医院的肺炎克雷伯菌分离株。为了获得对耐药组的分子洞察，病毒组，和多重耐药肺炎克雷伯菌分离株重要基因的遗传环境，我们对从成年患者中回收的10株肺炎克雷伯菌进行了短读全基因组测序，新生儿,还有医院的自来水样本.
结果：分离株的基因组草案大小不同，范围从5.48到5.96Mbp，表明该病原体的基因组可塑性。各种基因赋予对不同类别抗生素的抗性，例如，氨基糖苷类,喹诺酮类药物,磺胺类药物,四环素,和甲氧苄啶在所有测序的分离株中被鉴定。对碳青霉烯类抗生素的耐药性最高，这与B类和D类碳青霉烯酶的存在有关，blaNDM,和blaOXA,分别。此外，在6/10的测序菌株中发现了抗微生物剂基因qacEdelta1。测序的分离株表现出广泛的序列类型和囊膜类型。显著的抗生素抗性基因(ARGs)被各种可移动遗传元件(MGEs)所包围。观察到除获得性抗生素抗性基因以外的各种基因自发突变，在使这些细菌对抗生素产生抗药性方面发挥了间接作用。外膜孔蛋白丢失或缺乏，结合ESBL生产，在我们测序的分离株中，碳青霉烯耐药性发挥了重要作用。系统发育分析表明，研究分离株与中国菌株表现出进化关系，印度,和美国提出了共同的进化史和相似基因在不同起源的分离株中的潜在传播。
结论：这项研究为肺炎克雷伯菌菌株存在碳青霉烯类耐药的多种机制提供了有价值的见解，包括通过可移动的遗传元件获得多种抗生素耐药基因。丰富的动员体的鉴定产生了有关插入序列的关键作用的有见地的信息，转座子,和整合子塑造细菌基因组，以传播各种抗性相关基因。具有最少抗性基因的多重抗性分离株表现出大量突变。来自水源的肺炎克雷伯菌分离株显示出与临床分离株相当的抗生素抗性决定因素和最高数量的毒力相关基因，表明不同来源的细菌之间可能存在ARGs的相互作用。
结果：分离株的基因组草案大小不同，范围从5.48到5.96Mbp，表明该病原体的基因组可塑性。各种基因赋予对不同类别抗生素的抗性，例如，氨基糖苷类,喹诺酮类药物,磺胺类药物,四环素,和甲氧苄啶在所有测序的分离株中被鉴定。对碳青霉烯类抗生素的耐药性最高，这与B类和D类碳青霉烯酶的存在有关，blaNDM,和blaOXA,分别。此外，在6/10的测序菌株中发现了抗微生物剂基因qacEdelta1。测序的分离株表现出广泛的序列类型和囊膜类型。显著的抗生素抗性基因(ARGs)被各种可移动遗传元件(MGEs)所包围。观察到除获得性抗生素抗性基因以外的各种基因自发突变，在使这些细菌对抗生素产生抗药性方面发挥了间接作用。外膜孔蛋白丢失或缺乏，结合ESBL生产，在我们测序的分离株中，碳青霉烯耐药性发挥了重要作用。系统发育分析表明，研究分离株与中国菌株表现出进化关系，印度,和美国提出了共同的进化史和相似基因在不同起源的分离株中的潜在传播。
结论：这项研究为肺炎克雷伯菌菌株存在碳青霉烯类耐药的多种机制提供了有价值的见解，包括通过可移动的遗传元件获得多种抗生素耐药基因。丰富的动员体的鉴定产生了有关插入序列的关键作用的有见地的信息，转座子,和整合子塑造细菌基因组，以传播各种抗性相关基因。具有最少抗性基因的多重抗性分离株表现出大量突变。来自水源的肺炎克雷伯菌分离株显示出与临床分离株相当的抗生素抗性决定因素和最高数量的毒力相关基因，表明不同来源的细菌之间可能存在ARGs的相互作用。
