| Currently, Shigella, a member of the Enterobacteriaceae family can be categorized into four major subtypes i.e., Shigella flexneri, Shigella dysenteriae, Shigella sonnei, and Shigella boydii. Shigellos is mainly caused by Shigella, which can induce diarrhea, dysentery and can lead to the host death worsely. Shigellosis is still a global disease which can not effectively be treated with the existing drugs. In recent years, bacillary dysentery caused by Shigella bacteria has increasingly become the focus of public health issues. Unfortunately, in the last decades, a number of fluoroquinolone-resistant strains were found. In clinic, beta-lactams, macrolides and fluoroquinolone are generally used in the treatment of shigellosis and fluoroquinolone is widely used in the treatment of Shigellosis in many countries.Fluoroquinolones are presently important drugs for the treatment of shigellosis despite the quinolone-resistance has been distributed in various geographical regions. The genes of Shigella gyr A and gyr B(encoding DNA gyrase), and par C and par E(encoding topoisomerase IV) are the target of fluoroquinolones usually.Fluoroquinolones resistance is becoming a prevalent clinical issue with the threatening use of these drugs. Resistance mechanisms are currently clarified into three distinct categories. The cellular alterations associated with each mechanism are not mutually exclusive and can accumulate to create strains that exhibit a very high level of quinolone resistance.There are three mechanisms of shigella resistance to antibiotics that have been explained. First is target-mediated quinolone resistance. Quinolone resistance is most often associated with specific mutations in DNA gyrase and topoisomerase IV. Usually, IV mutation of one type II enzyme can nearly 10-fold drug resistance. Although mutations have been mapped throughout the A and B subunits of DNA gyrase and topoisomerase IV in fluoroquinolones resistant strains, the most commonly mutated amino acids are the serine and acidic residues that anchor the water-metal ion bridge. In general, mutant DNA gyrase and topoisomerase IV preserve wild-type DNA cleavage activity with no drugs. In clinic, quinolones have little ability to increase levels of enzyme-mediated DNA cleavage. And quinolones can interact with gyrase and/or topoisomerase IV to form stable enzyme-DNA-drug complexes. The complexes can protect bacterial against naturally occurring antibiotics.Second is plasmid-mediated quinolone resistance. And the resistance as high as ~250-fold has been reported. Plasmids that confer fluoroquinolones resistance typically carry additional genes that cause resistance to other drugs. Three families of genes are associated with plasmid-mediated fluoroquinolones resistance. The first are the Qnr genes, the proteins encoded by this gene that are part of the pentapeptide repeat protein family. And nearly 100 Qnr variants have been found and classified into at least five subfamilies. The Qnr family confer fluoroquinolones resistance by two different mechanisms. First is like Mcb G and Mfp A, they decrease the binding of DNA gyrase and topoisomerase IV to DNA. Thus, they can lower the number of available enzyme targets on the chromosome to protect cells from fluoroquinolones. Second they can also bind to DNA gyrase and topoisomerase IV and restrain fluoroquinolones from inserting enzyme-DNA-drug complexes formed by the enzymes. The plasmid-encoded protein associated with fluoroquinolones resistance is aac(6’)-Ib-cr. This protein contains two specific point mutations, W102 R and D179 Y that is a variant of an aminoglycoside acetyltransferase. The enzyme acetylates the unsubstituted nitrogen of the C7 piperazine ring can decrease drug activity that have been found in norfloxacin and ciprofloxacin. Only the mutant enzyme is active against quinolones and the wild-type and mutant aminoglycoside acetyltransferases are capable of acetylating other drugs. The plasmid-encoded fluoroquinolones resistance protein is comprised of efflux pumps. And three proteins of efflux pumps have been identified, Oqx AB, Qep A1, and Qep A. Whereas Qep A1 and Qep A have been reported in human bacterial infections and Oqx AB is found almost only in animal infections.The third is chromosome-mediated quinolone resistance. The outer membrane of Gram-negative bacteria poses an additional barrier that drugs must cross to enter the cell. Therefore, drug influx in Gram-negative species is enabled by protein channels called porins. When the expression of proteins is downregulated, it can lead to low-level resistance to quinolones. The concentration of fluoroquinolones in cells can be regulated by pump-mediated efflux. And, the upregulation of these efflux pumps is caused by mutations in regulatory proteins. Enhanced expression of chromosome-encoded efflux pumps also can lead to quinolone resistance.The Osp F from Shigella, Spv C from Salmonella, and Hop AI1 from the plant pathogen Pseudomonas syringae are belong to Osp F protein family. Osp F is a effector that secrete by type three secretion system(T3SS)of Shigella. All of the effectors in the Osp F protein family are phosphothreonine lyases that specifically target host MAPKs [the extracellular signal-regulated kinases(Erk) p38 in mammals and MPK3/4/6 in plants] and inactivate host signaling pathways. And we also found that the effector Osp F are involved in shigella intrinsic resistance to broad-spectrum antibiotics-fluoroquinolones resistance.Firstly to confirm the resistant of the mutant, MICs of △Osp F were determined in LB liquid medium at 37 °C supplemented with antimicrobials at different concentrations. The mutant of △Osp F exhibited increased resistant to grow on LB plate supplemented with indicated concentration of moxifloxacin, norfloxacin, ofloxacin and ciprofloxacin. Therefore, we want to know whether the mutant of △Osp F is quinolone resistance?To determine whether Osp F deficiency will enhance the survival during lethal fluoroquinolones stress, we exposed Shigella flexneri 2a 301, mutant of △Osp F and the complement cells to fluoroquinolones. When these cells were treated with various concentration of ciprofloxacin for 1 h, the survival rate of the Osp F mutant was higher than that of wild-type and complement cells. Similar phenomenon was observed when wild-type, mutant of △Osp F and the complement cells were treated with norfloxacin, ofloxacin moxifloxacin. When cells were treated hydrogen peroxide the protective effect of the Osp F deficiency seen with antimicrobials was not observed. When wild-type and Osp F mutant cells were pretreated with subinhibitory concentrations of thiourea plus 2,2’-bipyridyl to block hydroxyl radical accumulation, the lethal action of ampicillin and ciprofloxacin was reduced in both strains by a factor of 100 to several thousand, and the Osp F mediated protection was eliminated.In conclusion, the reactive oxygen species(ROS)-dependent portion of killing by fluoroquinolones is protected by the OspF deficiency. |
