粪便分离大肠埃希菌CRISPR系统在耐药及毒力中的作用

Effect of CRISPR system on drug resistance and virulence of Escherichia coli isolated from faecal specimen

目的探索粪便分离大肠埃希菌成簇规律间隔短回文重复序列(clustered regularly interspaced short palindromic repeats, CRISPR)在耐药及毒力中的作用。方法收集某院2018年8-12月分离自慢性腹泻患者及健康体检者粪便样本的大肠埃希菌。采用纸片扩散法检测其药物敏感性,PCR法检测并比较分析腹泻患者及健康体检者粪便样本分离大肠埃希菌的系统发育群、耐药基因及CRISPR系统。结果共收集到142株大肠埃希菌,源于63例慢性腹泻患者(疾病组)和79例健康体检者(健康组)。药敏结果显示,氨苄西林耐药率为48.0%,其余抗菌药物敏感率为73.1%~100.0%。63株源于慢性腹泻患者粪便样本分离大肠埃希菌中CRISPR系统的检出率显著低于79株源于健康体检者(3.2%vs 46.8%,χ^(2)=33.538,P<0.001)。在被检测的耐药基因中,63株疾病组中11株阳性(7株blaCTX-M-1组、3株blaCTX-M-9组和1株mcr-1),79株健康组中4株blaCTX-M-1组,未携带耐药基因菌株中CRISPR系统的检出率显著高于携带耐药基因菌株(0.0%vs 29.9%,P=0.011)。43株高毒力菌株(B2群和D群)中CRISPR系统的检出率显著高于99株低毒力菌株(A群和B1群)(44.2%vs 20.2%,χ^(2)=8.656,P=0.003)。结论粪便分离大肠埃希菌对常用抗菌药物仍保持较高敏感性。CRISPR系统可能在粪便分离大肠埃希菌毒力及耐药基因的传播方面发挥重要作用。

Objective To explore the role of clustered regularly interspaced short palindromic repeats(CRISPR) in the drug resistance and virulence of Escherichia coli isolated from faecal specimen. Methods E. coli isolates were collected from faecal specimens from patients with chronic diarrhea and healthy people in a hospital during August to December 2018. Antimicrobial susceptibility of the strains were detected using Kirby-Bauer method. PCR method was used to detect the phylogeny, resistance genes and CRISPR system of the E. coli isolates. Results A total of 142 E. coli strains were collected from 63 patients with chronic diarrhea(disease group) and 79 healthy people(health group). The results showed that the resistance rate to ampicillin was 48.0%, and susceptibility rate to other antibiotics were from 73.1% to 100.0%. The detection rate of CRISPR system in health group was significantly higher than that in disease group(46.8% vs 3.2%, χ^(2)=33.538, P<0.001). Drug-resistant gene test showed 11 positive strains in the 63 isolates from disease group(7 blaCTX-M-1 group, 3 blaCTX-M-9 group and 1 mcr-1), and 4 in the 79 isolates from health group(4 blaCTX-M-1 group). The detection rate of CRISPR system in isolates carried non drug resistant genes was significantly higher than those carried drug resistant genes(0.0% vs 29.9%, P=0.011). The detection rate of CRISPR system in 43 highly virulent strains(B2 and D) was significantly higher than that in 99 lowly virulent strains(A and B1)(44.2% vs 20.2%, χ^(2)=8.656, P=0.003). Conclusion E. coli strains isolated from faecal specimen is still highly sensitive to commonly used antibiotics. CRISPR system may play an important role in the virulence and the transmission of drug-resistant genes in E. coli isolates from faecal specimen.