宋内志贺氏菌1173质粒基因组及耐药基因转移机制

Bacterial plasmid genome analysis and the mechanism of resistance gene transfer in the clinical Shigella sonnei isolate 1173

摘要:

[目的]研究质粒介导宋内志贺氏菌1173耐药基因bla_CTX-M-64转移的机制。[方法]用双纸片协同扩散法验证1173是否产超广谱β-内酰胺酶（ESBL），用PCR扩增鉴定其携带的耐药基因，接合转移实验检验其耐药质粒是否可通过接合转移给其他细菌，并对接合子是否产ESBL和携带耐药基因进行检验，利用VITEK^® 2检测1173和接合子的耐药谱，提取质粒进行高通量基因组测序，并对质粒序列进行生物信息学分析，以研究其耐药基因转移机制。[结果]1173是产ESBL的多药耐药宋内志贺氏菌，携带的耐药基因有bla_CTX-M-64和bla_TEM，其中的bla_CTX-M-64可通过接合转移作用传递给受体菌EC600，并使接合子具有相应的耐药谱。经序列测定和生物信息学分析表明，介导bla_CTX-M-64水平转移的是ISEcp1-bla_CTX-M-64-Δorf477转座单元。[结论]质粒携带的bla_CTX-M-64介导1173对多类抗菌药物的耐药，ISEcp1-bla_CTX-M-64-Δorf477转座单元介导bla_CTX-M-64在细菌间的转移。

Abstract:

[Objective] To sequence and analyze plasmid p1173-CTXM from the clinical multidrug-resistant Shigella sonnei isolate 1173 with the aim of investigating the mechanism of bla_CTX-M-64resistance gene transfer. [Methods] A double-disk synergy test was conducted to determine if strain 1173 produced extended-spectrum beta-lactamase (ESBL), then antibiotic resistance genes were PCR-amplified and sequenced. Conjugation experiments were used to verify the transferability of plasmids carrying ESBL genes. The transconjugant was detected to certify the production of ESBL and the existence of corresponding resistance genes. Strains 1173 and 1173-CTXM-EC600 underwent testing for antimicrobial drug susceptibility. Plasmid p1173-CTXM-EC600 was sequenced by high-throughput genomic sequencing to illustrate the mechanism of resistance gene transfer. [Results] The multidrug-resistant isolate Shigella sonnei 1173 was shown to be an ESBL-producing strain and to carry the antibiotic resistance genes bla_CTX-M-64 and bla_TEM; however, only bla_CTX-M-64 was horizontally transferred to the recipient strain EC600 which endowed the corresponding antibiotic resistant profiles to EC600. Sequencing and bioinformatics analysis revealed that the resistance gene bla_CTX-M-64 was carried by the ISEcp1-bla_CTX-M-64-Δorf477 transposition unit. [Conclusion] The resistance gene bla_CTX-M-64 carried by plasmid p1173-CTXM mediated the resistance of Shigella sonnei isolate 1173 to multiple antibacterial agents. Horizontal transfer of bla_CTX-M-64 was mediated by the ISEcp1-bla_CTX-M-64-Δorf477 transposition unit.