Abstract：
The wide dissemination of plasmids carrying antibiotic resistance determinants among bacteria is a severe threat to global public health. Here, we characterized an extensively drug-resistant (XDR) Klebsiella pneumoniae NTU107224 by whole genome sequencing (WGS) in combination with phenotypic tests. Broth dilution method was used to determine the minimal inhibitory concentrations (MICs) of NTU107224 to 24 antibiotics. The whole genome sequence of NTU107224 was determined by Nanopore/Illumina hybrid genome sequencing. Conjugation assay was performed to determine the transferability of plasmids in NTU107224 to recipient K. pneumoniae 1706. Larvae infection model was used to determine the effect(s) of conjugative plasmid pNTU107224-1 on bacterial virulence. Among the 24 antibiotics tested, XDR K. pneumoniae NTU107224 had low MICs only for amikacin (≤1 μg/mL), polymyxin B (0.25 μg/mL), colistin (0.25 μg/mL), eravacycline (0.25 μg/mL), cefepime/zidebactam (1 μg/mL), omadacycline (4 μg/mL), and tigecycline (0.5 μg/mL). Whole genome sequencing showed that the closed NTU107224 genome comprises a 5,076,795-bp chromosome, a 301,404-bp plasmid named pNTU107224-1, and a 78,479-bp plasmid named pNTU107224-2. IncHI1B plasmid pNTU107224-1 contained three class 1 integrons accumulated various antimicrobial resistance genes (including carbapenemase genes blaVIM-1, blaIMP-23, and truncated blaOXA-256) and the blast results suggested the dissemination of IncHI1B plasmids in China. By day 7 after infection, larvae infected with K. pneumoniae 1706 and transconjugant had 70% and 15% survival rates, respectively. We found that the conjugative plasmid pNTU107224-1 is closely related to IncHI1B plasmids disseminated in China and contributes to the virulence and antibiotic resistance of pathogens.
摘要：
携带抗生素抗性决定簇的质粒在细菌中的广泛传播是对全球公共卫生的严重威胁。这里,我们通过全基因组测序(WGS)结合表型试验鉴定了广泛耐药(XDR)肺炎克雷伯菌NTU107224.肉汤稀释法用于确定NTU107224对24种抗生素的最小抑制浓度(MIC)。NTU107224的全基因组序列通过Nanopore/Illumina杂合基因组测序确定。进行缀合测定以确定NTU107224中的质粒向受体肺炎克雷伯菌1706的可转移性。幼虫感染模型用于确定接合质粒pNTU107224-1对细菌毒力的影响。在测试的24种抗生素中，XDR肺炎克雷伯菌NTU107224仅对阿米卡星(≤1μg/mL)具有低MIC，多粘菌素B（0.25μg/mL），粘菌素（0.25μg/mL），埃拉环素（0.25μg/mL），头孢吡肟/齐达巴坦（1μg/mL），奥马环素（4μg/mL），和替加环素（0.5μg/mL）。全基因组测序显示，封闭的NTU107224基因组包含一个5,076,795bp的染色体，名为pNTU107224-1的301,404-bp质粒和名为pNTU107224-2的78,479-bp质粒。IncHI1B质粒pNTU107224-1包含三个1类整合子，积累了各种抗菌抗性基因（包括碳青霉烯酶基因blaVIM-1，blaIMP-23和截短的blaOXA-256），爆炸结果表明IncHI1B质粒在中国传播。感染后第7天,感染肺炎克雷伯菌1706和转体的幼虫有70%和15%的存活率，分别。我们发现接合质粒pNTU107224-1与在中国传播的IncHI1B质粒密切相关，并有助于病原体的毒力和抗生素耐药性。
