Study On The Genotypes Of ESBLs And Analysis Of ERIC-PCR In Shigella Isolates

Objectives:Bacillary dysentery caused by Shigella species is one of the most important cause of acute infectious diarrhea.Following widespread ESBLs and selection ressure of antibiotics,extended-spectrumβ-lactamases(ESBLs)-producing Shigella isolated increases gradually.In order to explore the epidemical characteristics of ESBLs-producing Shigella spp,the topic choosed isolated shigella selected from three rank 3-grade A hospitals in Tianjin as the research target,studying changed serogroup and drug-resistant characteristics of Shigella species;analysising antibiotic resistance,transmissible R-plasmid and mainly prevalent genotypes of ESBLs-producing Shigella isolated;investigating molecular epidemiology circumstance of shigella strains from Tianjin.Methods:136 Shigella isolates were collected from entero-out-patient clinical of three hospitals in Tianjin from June in 2009 to September in 2010. Identify different and superiority serotype of 136 Shigella spp.Antimicrobial susceptibility test was done by Kirby-Bauer (K-B) test,and compared the drug resistance of these strains from different serotype and source to different antibiotics.Phenotypic confirmatory test and three-dimensional extract test were used for identification of ESBLs and AmpCβ-lactamases separately. The conjugation experiment was performed in the confirmed ESBLs-producing strains in order to identify containing the transmissible R-plasmid, antibiotic resistance was compared between clinical strains and transconjugants. Polymerase chain reaction (PCR) was performed to detect the genotypes of confirmed ESBLs-producing strains.Some PCR positive products were sequenced and compared for homology on genbank. Molecular epidemiology analysis of producing-ESBLs shigella isolates was determined enterobacterial repetitive intergenic consensus PCR(ERIC-PCR).Results: 1.The study included 136 isolates of Shigella spp.(99 Shigella sonnei,34 Shigella flexneri, two Shigella boydii and one Shigella dysenteriae).Shigella sonnei is the predominant serotype in Tianjin.For S. sonnei and S flexneri,Imipnem was the most sensitive antibiotics,which sensitivity were both 100%,The resistance rates to ampicillin,piperacillin,tetracycline,streptomycin and Sulfamethoxazole-Trimetho prin were higher 75%, The resistance rates of quinolones resistance(norfloxacin, levofloxacin) is lower 3%.The difference of the two serogroups's partial resistance rates were statistically significant (P<0.05).The resistance rate of S flexneri to third,four Generation Cephalosporins and quinolones(cefotaxime, cefepime, ceftri-axome),aztreonam and Chloramphenicol were higher than S sonnei,but the others are undifferential. The resistance rate of strains contained the same serotype from different age groups were uniform.2.20 strains were positive for ESBLs phenotypic confirmatory test,the positive rate was 14.7%. All strains were not detected to produce AmpC enzyme.Eleven TEM-1 type genes,sixteen CTX-M-14 group type,four CTX-M-15 group type were founded by PCR in 20 ESBLs-producing strains.SHV,CTX-M-2 group,CTX-M-8 group and OXA group were negative.The general isolating rate of CTX-M type ESBLs was 100%. The resistance rate of ESBLs to Sulfamethoxazole-Trimethoprin, tetracycline and were significant(70%～90%);the rate to Gentamicin streptomycin and Chloramphenicol were between 30%～50%,while the rate to Cefepime, Amikacin and quinolones were lower 30%, All of the ESBLs-producing strains were sensitive to Imipnem. The transconjugant test of 16 ESBLs-producing isolates were positive,and these conjugations were resistant toβ-lactamases and partial aminoglycosides. The transmission of CTX-M producing strains was mainly mediated by plasmids.The Profile of these plasmids showed:most conjugations only contained one transferred natural plasmid. ERIC-PCR demonstrated that the strains with CTX-M-14 ESBLs genes seemed aggregate and prevail tendency.Conclusions:The main serotype sonnei of 136 Shigella isolates was Shigella. The difference of the two serogroups's partial resistance rates were statistically significant. The resistance rate of strains contained the same serotype from different age groups were uniform. the positive rate of 20 ESBLs- producing Shigella isolated was 14.7%. The CTX-M producing strains was primary genotype. ESBLs was the mian cause of leading to their multiplex resistance to mostβ-lactams.The mechanism to resiatance was mainly mediated by plasmids. CTX-M-14 ESBLs of Shigella strains was the predominant genotype in Tianjin.