| Objective This study aims to investigate the distribution of Carbapenem‐resistant Klebsiella pneumonia(CRKP) I Integron and Insertion sequence common region(ISCR) and the types of resistance gene cassettes, so as to analyze drug‐resistant mechanism and molecular epidemic features of CRKP in our hospital.Methods The 44 strains of distinct clinical isolates of CRKP were collected and identified, and the statistical questionnaire was made for the statistical analysis of clinical data of CRKP carriers. Routine drug sensitivity test was performed with Vitek 2 compact automatic microbiology analyzer,and KPC detection was conducted by revised Hodge test, and metallo‐beta‐lactamase was detected with the EDTA coordination method, and the drug‐resistant genes of the strains were tested by polymerase chain reaction method(PCR). The Enterobacterial Repetitive Intergenic Consensus(ERIC – PCR) test was conducted for the homology analysis and MLST typing of the 44 strains of CRKP. The cluster analysis was conducted in Quantity one 4.6.2 on the electrophoresis results, and tree diagrams were drawn to understand its molecular epidemiological characteristics. In the 44 strains of CRKP, PCR and genome sequencing were performed to detect the Int I/II/III, the variable regions of class I integrons, ISCR1 and the variable regions of ISCR1, and the relationship between class I integrons and ISCR1. The positive amplifications of the variable regions of class I integrons and ISCR1 were sequenced and analized.Results Clinical data indicates the average age of the CRKP carriers(chronic diseases accompanied) are 65.3±25.6 years. For the 44 strains of CRKP, 70.4% of the specimens were sputum. 22.7% strains were isolated from ICU and 29.5% were from the department of neurosurgery. All 44 CRKP strains manifested drug resistance against Carbapenem antibiotics. More than 95% were cephalosporins‐resistant. In all of the 44 strains of CRKP, KPC‐2 gene was detected, NDM‐1, SME, IMP, OXA‐48, VIM, and GES‐positive strains were not detected. For ESBLs genes: 10 strains carried SHV genes, 36 strains with TEM ‐positive, and 40 strains were CTX‐M‐9‐positive. With ERIC‐PCR typing, the 44 strains of CRKP were divided into 13 clonotypes, with the majority being type I(38.6%, 17/44).In the 44 strains of CRKP, 32 strains were carrying Int I genes with the positive rate of 72.7%, including 29 strains with variable regions(positive rate is 65.9%). The gene sequencing result showed the presence of drug‐resistant gene cassettes aad A, and Int II/III genes were not detected. 6 strains presented as ISCR1‐positive, and 1 strain had ISCR1 variable regions, of which the gene sequencing test result was drug‐resistant gene sul1. The ISCR1 in 1 strain was directly connected to the downstream of class I integrons3’ conserved domain.Conclusion(1)KPC‐2 –Producing enzymes and other resistant genes are consistently detected from clinically isolated CRKP.(2)highly homologous CRKP was manifested in the cases of our hospital. Due to its molecular cloning nature, effective measures should be taken clinically to curb its spreading.(3)Class I integrons with add A resistance gene cassettes are widely distributed in CRKP. ISCR1 with sul1 resistance gene cassettes are sparsely distributed. The fact that Class I integrons and ISCR1 are present in CRKP in tandem array might mediate horizontal transmission of resistant genes to different CRKP clone strains, and further bring about the complicated and diversified CRKP resistance capacity. |
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