ABSTRACT
Objective To explore the association of ATP-binding cassette (ABC) efflux pump gene Rv1217c-Rv1218c and the drug resistance of Mycobacterium tuberculosis.Methods A total of 34 Mycobacterium tuberculosis clinical isolates including 24 drug-resistance isolates which were resistant to riffampicin,isoniazid,streptomycin or ethambutol and resistant to at least one second-line antituberculosis drug,and 10 drug-sensitive isolates were involved in this study.The RNA of isolated strains was extracted and then reverse transcribed. Gene expression was performed by real-time polymerase chain reaction (PCR) and data was analyzed by t test and Logistic regression analysis.Results The expressions of Rv1217c in rifampicin-resistant group (2.13 ± 1.89,t =3.44,P＜0.01),isoniazid-resistant group ( 1.84 ± 1.86,t =3.16,P＜ 0.05),streptomycin-resistant group ( 1.86 ±1.96,t=2.78,P＜0.05) and ethambutol-resistant groups (3.36±2.35,t=3.04,P＜0.05) were all higher than sensitive isolates (0.42 ± 0.31).The expressions of Rv1218c in rifampicin-resistant group (2.54±1.84,t=3.82,P＜0.01),isoniazid-resistant group (2.34± 1.84,t=3.72,P＜0.01),streptomycin-resistant group (2.15±1.86,t=3.01,P＜0.01) and ethambutol-resistant groups (3.78± 1.78,t=4.22,P＜0.01 ) were all higher than sensitive isolates (0.65 ± 0.42).The expressions of Rv1217c and Rv1218c in multidrug resistant group were 2.74±2.07 and 3.33± 1.77,respectively,which were both higher than polydrug resistant group (0.79 ± 0.47 and 1.03 ± 0.79,respectively; t =2.91,P＜0.05 ; t =3.84,P＜0.01,respectively).Logistic regression analysis found that high Rv1217c expression was positively correlated with rifampicin resistance and negatively correlated with isoniazid resistance (both P＜ 0.01 ),while high Rv1218c expression was negatively correlated with rifampicin resistance and positively correlated with isoniazid resistance (both P＜0.01 ).High expressions of two genes were both positively correlated with ethambutol resistance and multidrug resistance (both P＜0.01 ) and not statistically correlated with streptomycin resistance (P＞0.05).Conclusions The expressions of ABC efflux pump gene,Rv1217c-Rv1218c,are correlated with multiple drug resistance.The overexpression may contribute to the multidrug resistance of Mycobacterium tuberculosis.
ABSTRACT
Objective To investigate the molecular epidemiology and mechanism of carbapenem resistance of Escherichia coli collected from intensive care units(ICUs)of general surgery.Methods Agardilution were carried out to confirmed the drug-susceptibility,pulsed-field gel electrophoresis(PFGE)were performed to analyze the molecular epidcmiology of carbapenem-resistance isolates.Specific PCR,DNA sequencing,conjugation experiments,plasmids extraction,plasmid transformation assays and SDS-PAGE of outer membrane proteins(OMPs)were carried to confirm genotype of carbapenemase and its transmission mechanism.Results PFGE showed the isolates belonged to 10 clonotype,and all the clinical isolates were resistant to β-lactams including imipenem and meropenem,but uncertain to aminoglycosides,specific PCR and DNA sequencing revealed that all isolates encoded carbapenem-hydrolyzing enzyme gene,KPC-2.Plasmid DNA extraction and plasmid transformation assays from some isolates comfirmed that KPC-2 encoded on a 56 kb plasmid.SDS-PAGE analysis confirmed that there are alterations in OMPs of Escherichia coli.Conclusion Escherichia coli isolates with carbapenem resistance are collected from our hospital,production of KPC-2 carbapenemase mainly contributed to reduced susceptibility of carbapenem in Escherichia coli,the alterations in OMPs may as a cofactor in high-level drug-resistance in Escherichia coli.
ABSTRACT
Objective To study molecular epidemiology and carbapenem-resistance mechanism of four Escherichia coli strains isolated from general surgery wards. Methods Antibiotic susceptibility was carried out by K-B gar diffusion and agar dilution methods. Carbapenemases were screened by three dimensional test and EDTA-Na_2-disk synergy test. Pulsed-field gel electropboresis (PFGE) was performed to analyze molecular epidemiology of isolates. Plasmid was extracted by using an alkalinelysis technique. Conjunction experiment, transformation assay, specific PCR and DNA sequencing were performed to confirm carbapenemase genotype and its transmission mechanism Results Four Escherichia coli isolates were resistant to most antimicrobials including carbapenem. PFGE showed that the four isolates belong to four different clonal strains. Specific PCR and DNA sequence analysis identified that carbapenem resistance in four clinical isolates was mediated by KPC-2 encoded on an approximately 56 000 bp plasmid, and this plasmid did not harbor aminoglycosides and fluorquinolones resistant genes. Conclusion Four Escherichia coli isolates with carbapenem resistance are obtained from our hospital, and KPC-2 plasmid is main cause of carbapenem resistance in these isolates.