| Marine environment has gradually become an important "repository" of resistant bacteria and resistance genes.Bacterial resistance can spread in the marine environment through horizontal gene transfer.The beach is an important place to entertain,and it is also the place where microbes spread and gather,and the microbial diseases spread through recreational water have become one of the most important issues of public concern.Marine Environment Fecal Contamination Indicator-Escherichia coli can swallow through the seawater to cause swallowing the fatal intestinal disease.In order to prevent and slow the spread of bacterial resistance,we must fully understand the bacterial resistance of the generation and dissemination mechanism.Therefore,this study was conducted to study the distribution characteristics and drug resistance status of Escherichia coli in the beach,and the multidrug resistance of Escherichia coli mediated by integron-gene cassette system was analyzed.Plasmid conjugation experiments revealed a plasmid-mediated level of transmission of E.coli resistance genes.The results are as follows: 1.A total of 373 strains were obtained in seawater and sands samples of Fujiazhuang Beach in Dalian.The Escherichia coli was obtained by biochemical identification in seawater samples was 69 and in sands samples was 13.K-b paper diffusion method susceptibility test was used to determine its resistance to 10 kinds of common antibiotics.The results showed that the detection rate of drug-resistant Escherichia coli in seawater was 37.7% and was resistant to all antibiotic except aztreonam.The resistance rate of tetracycline(24.6%),trimethoprim(24.6%)and trimethoprim/sulfamethoxazolethe(23.2%)was higher than other antibiotics.The resistance rate of gentamicin was 10.1%.And the resistance against streptomycin,ciprofloxacin,cephalothin,chloramphenicol and levofloxacin was lower than 10%.The multiple resistance rate of Escherichia coli in seawater was 57.7%,and the multiple resistance spectrum of Escherichia coli showed that it was resistant to up to 8 antibiotics.The resistance to ciprofloxacin was only 37.5% in the wet sand isolate,and the resistance to trimethoprim was only 40% in the dry sand isolate.2.Based on the results of the above-mentioned Escherichia coli drug resistance phenotype,the tetracycline,?-lactam,sulfonamides and quinolone resistance genes were selected and the resistance gene of Escherichia coli genome and plasmid DNA in seawater and beach was determined by PCR Detection.The results showed that the detection rate of tetracycline-resistant gene in E.coli genomic DNA was 88.2%,the detection rate of tetA gene was 64.7%,the detection rate of tetB gene was 41.2%,the detection of tetracycline-resistant gene in the plasmid The rate of tetA gene(29.4%)was lower than that of tetB gene(41.2%).The detection rates of ?-lactam resistance genes in genomic DNA and plasmid DNA were 80% and 20%,respectively.The detection rates of sulfonamide-resistant genes in genome and plasmid DNA were 50% and 66.7%,respectively.The qnrS resistance gene of quinolones was detected only on plasmid DNA,and the remaining genes were not detected.While the E.coli isolated from sands only detected in the plasmid DNA sulfonamide resistance gene sul1,the remaining drug resistance genes were not detected.3.In order to study the multidrug resistance of seawater Escherichia coli,the integraining enzyme gene of 15 strains of multiple resistant bacteria was detected,9 genomic DNA type ?integrase was positive,and 3 strains were amplified in the integrase positive isolates 11 type plasmid DNA type integrase positive,in the integrase?-positive isolates were amplified 3 class ? integrons,multi-resistant strains of class I integrin detection rate is higher The The results of the sequencing of the integron variable region showed that 8 different gene cassettes were the genes encoding the sulfonamide resistance gene dihydrofolate reductase(dfr16/A12/A17,dhfr12/17),encoding the aminoglycoside resistance gene Nucleoside transferase gene(aadA2 / 5),and encoding the forest amide nucleotide transferase gene(linF).The results suggest that integrins may mediate the multidrug resistance of E.coli.4.A strain containing a sulfonamide-resistant gene on the plasmid was selected from the isolates of Escherichia coli and subjected to a bonding experiment with the E.coli C600,and the bonding time was measured.The results showed that six of the 12 strains of Escherichia coli containing the resistant sulfonamides resistant gene were successfully joined,the rate of fusion was 50%,the time of the grafting was 2~7h,the sulfonamide resistance gene was detected in the adapter,but resistant phenotype did not show.The above results indicate that the environmental strain resistance gene can be transferred by plasmid binding in the same species strain. |
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