| Shigella is the pathogen of shigellosis. It is estimated that shigellosis causes around 600,000 deaths per year in the world, and two-thirds of the deceased being children under 10 years of age. In China,shigellosis is still one of the important communicable disease which affect human health. The antimicrobial resistance in Shigella spp. has been becoming a seriously problem for controlling this disease. High frequencies of resistance in Shigella spp. to many of the first-line antimicrobial agents, such as TE in 1960's and quinolones in 1980's, have been reported from many parts of the world. Due to the importance of shigellosis and the common use of quinolones for treatmeat of bacterial infections, to study the antimicrobial susceptibility patterns of Shigella spp. with the particular reference of the quinolones and its possible mechanism is obviously necessary.The mutations in the gyrA gene were found both in vivo- and in vitro- resistant Shigella isolates. The mutation altering amino acid in Pare was found only in laboratory-grown fluoroquinolone-resistant Shigella flexneri mutants. In addition to the mutations in the genes encoding the target enzymes, quinolones resistance in Shigella spp.may also be associated with increased efflux of the5drugs. In order to give the instruction to use the quinolones more properly and control the resistance development, this study was planned to investigate the resistance and the genetic basis of the resistance to quinolones in Shigella spp..Method: A total of 73 clinical isolates were collected, including 54 strains of Shigella flexneri, 15 strains of Shigella sonnei, and 4 strains of Shigella dysenteriae. 30 strains of Shigella flexneri were isolated in Tonggu, Jiangxi. 24 strains of Shigella flexneri, 15 strains of Shigella sonnei, and 4 strains of Shigella dysenteriae were isolated in Zhengzhou, Henan. Susceptibility test (Kirby-Bauer) of 11 antimicrobial agents were performed. Four quinolones were Pipemidic adcid(PI), Ciprofloxacin(CIP), Ofloxacin(OFL), and Norfloxacin(NOR). Other sevenantimicrobial agents were Teracycline(TE), Chloramphenicol(C), Streptomycin(S), Sulfamethoxazole(SMX), Ampicillin(Am), Gentamicin(GM), and Cephalomin(CF). The quinolones resistance -determining regions (QRDR) of gyrA (648bp), parC (469bp) were amplified by polymerase chain reaction (PCR). Restriction fragment length polymorphism analysis (RFLP) and single strand conformational polymorphism analysis (SSCP) were applied to all PCR procucts. Cycle sequencing of several isolates was subsequently performed. PCR-RFLP analysis can detect the absence or persence of Hinf I sites,which indicating whether there is the nucleotide sequence change. PCR-SSCP analysis can detect a single nucleotide change as well as the loss or insertion of oligonucleotide. PCR-RFLP, PCR-SSCP, and sequencing are allsensitive and efficient gene detetion methods.Results:1.The resisrtance rates for Shigellaflexneri isolates to . TE, C, S, SMX, and Am varied from 70.8% to 100%, to CF was 36.7%. The resistance rates for Shigella sonnet isolates to TE, S, and SMX varied from 53.3% to 66.7%, to AM, CF were 26.7% and 13.3%. 2. The resistances rate for all the Shigella spp. to PI, CIP, NOR, and OFL were 79.5%, 60.3%, 41.1%, and 36.9%. The differences of resistance rates between PI and CIP, NOR and OFL were significant (both P<0.05). 3. The multiresistance rate for Shigella flexneri isolates from Jiangxi was 100%, for Shigella flexneri iaolates from Henan was 79.2%, for Shigella sonnei isolate from Henan was 40%. The differences among the above resistance rates were statistically significant (all P<0.05). 4. The resistance rates to 4 quinolones(in quinolone-recuced-susceptible isolates) for Shigella isolates from Jiangxi was signifcantly higher than that from Henan (60% Vs 21.9%, PO.05). 5.The mutations in the parC gene were firstly found in quinolone-resistant clinical strains of Shigella flexneri. DNA sequence analysis revealed five mutations that resulted in following amino acid changes: 12...
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