Relationship Between Express Changes Of Multiple Drug Resistance Related Genes And Drug Tolerance In Shigella Flexneri

Bacillary dysentery is a kind of infectious disease, which do great harm to human health and induce millions of people's death every year. Shigella flexneri is one of the causative agents of bacillary dysentery, and it's also the dominant strain in developing countries. Nowadays treating with bacillary dysentery, the only method is to use antibiotics. Although appropriate antibiotic therapy may reduce the duration of symptoms and carriage of Shigella flexneri, unreasonable use of antibiotics in clinical therapy may also increase the development of antibiotic resistant strains, finally lead the failure of clinical therapy. The characteristic multiple drug resistance in Shigella flexneri makes it impervious to antibiotics for clinical therapy, but the mechanism of resistance is not very clear.In this study, we chose to analyze the marA, acrB, tolC, tolA, tolB and tolQ genes for their roles in multiple drug resistance in Shigella flexneri 21573. The standard Shigella flexneri 21573 strain was cultured under artificial conditions with successively increasing concentrations of ciprofloxacin for 15 generations, and antibiotic sensitivity tests were done. In order to investigate whether the gene products of marA, acrB, tolC, tolA, tolB and tolQ are involved in multiple drug resistance, the expression level of the target genes in the 15 generations of Shigella flexneri 21573 were examined by real-time PCR. Furthermore, the correlation coefficients of marA and other targets genes were calculated in order to get in-depth understanding of the regulation of MarA to the AcrAB-TolC and Pal-Tol systems.Results of antibiotic sensitivity tests indicated that the induced Shigella flexneri 21573 strain acquired multiple drug resistance. And the antibiotic tolerances were developed at varying generation points, for instance, resistance to quinolones was obtained in the 1st generation, resistance to gentamicin, amikacin, chloramphenicol, tetracycline and streptomycin were obtained during the 3rd to 5th generations, and resistance to ampicillin, erythromycin, cephalosporin and novobiocin were obtained in the 13th generation.Results of real-time PCR showed that almost each target gene had two high expression peaks, the higest one was in the 13th generation and another one was around the 4th genenration, which was coincident with the gain of multiple drug resistance in Shigella flexneri 21573.Correlation analysis indicated that the express level of marA was statistically correlated with acrB (r=0.937539); in addition, it also significantly correlated with tolA and tolQ (r=0.885657, r=0.850747, respectively). However, the correlations for the express level of tolC, tolB and marA were not significant (r=0.369789, r=0.597783, respectively).The express chanange patern of marA, acrB, tolC, tolA, tolB and tolQ genes during the production of multiple drug resistance were obtained in our study. The gene products of marA, tolC and acrB were improved to be involved in multiple drug resistance in Shigella flexneri 21573, also the gene products of tolA, tolQ and tolB, three important parts of the Tol-Pal system were firstly shown to be involved in multiple drug resistance in Shigella flexneri 21573. Additionally, despite of acrB, our data also suggested that MarA might upregulate the expression of tolA and tolQ.