Isolation And Identification Of Enteropathogenic E.Coli From Animals In Panjin City Of Liaoning Province And Its Drug Resistance Recombinant Anti-human TNF-α Monoclonal Antibody

Avian colibacillosis (E.coli) is a kind of local or systemic infection disease caused by Avian pathogenic e. coli (Avian pathogenic Escherichia coli, APEC). This disease is clinical common diseases, poultry can infected in different kinds and ages, and it do great harm to poultry industry. At present three kinds of drugs including quinolone, aminoglycoside and beta lactam were mainly used for preventing and controlling escherichia coli disease in the clinic. Due to abuse of antibiotics of some individual farms and farmers, drug resistance of bacteria become stronger under the selection pressure, even worse, super bacteria come into being, and this problem has become a serious public health hazard to humans. This paper aims to collect colibacillus isolated from dead birds and dead pigs from regional scale farms in panjin of Liaoning, and then analyse drug resistance of different animal-derived e. coli to the19common antibiotic used in the clinic. Studying the drug resistance mechanism in molecular level, and provide a foundation for the prevention and development of new antibiotics.1Isolation and identification of pathogenic E. coli39bacterial strain was isolated from dead birds and dead pig from some regional scale farms in panjin, and these bacterial strain comes from all over PanJin. The39bacterial strains were identified by enterobacteriaceae bacteria biochemical kit and indole test, V-P experiment, citrate salt using experiments and seven leaf glycosides hydrolysis experiments, the results showed that there are27strains of e. coli, including18isolated from chicken,5strains of duck,2strains of goose,2strains of pig, pathogenic experiment results showed that rats mortality was86.67%. Thus proves that the27strains of E. coli were all highly pathogenic e. coli.2Drug resistance analysis of27strains of pathogenic e. coli to19kinds of antibioticsAccording to CLSI Kirby Bauer method,19kinds of antibiotic were used to make drug sensitivity test of27strains of e. coli. The results showed that among the four kinds of beta lactam drugs, drug resistant rate of e. coli to the fourth generation cephalosporins was the lowest (36%). For the third generation of cephalosporins drug cefazolin, cefotaxime, ceftriaxone, the drug resistance rat was all52%. In four kinds of quinolones, drug resistance rat of E. coli to ofloxacin was the lowest(32%); to ciprofloxacin was the highest(88%); the resistance to levofloxacin and norfloxacin were all68%. In seven kinds of aminoglycoside drugs, drug resistance rat of E. coli to streptomycin was the lowest (48%); to spectinomycin was the highest (88%). among them,drug resistance rat to gentamicin and amoxicillin was84%and72%respectively; to kanamycin, amikacin, tobramycin the resistant rate was52%,56%and68%respectively.In the four other kinds of drugs,drug resistance rat of E. coli to rifampicin was the highest (96%),to furazolidone was the lowest(28%),to tetracycline was also high(92%),to chloramphenicol the drug resistance rat was72%.Two strains of pathogenic escherichia coli from chicken were all resistant to19kinds of antibiotics. This provide a basis for the scientific and medical treatment in panjin area by drug sensitivity test.3super resistant gene (NDM-1) of2chicken-derived E. coli strains resistant to19kinds of antibiotic and detection and analysis of plasmid mediated resistance geneNdm-1is a newly discovered meta1-β-lactamase, bacteria carrying NDM-1have resistance to most antibiotics, thus is called "super resistant genes".E. coli resistance genes most exist in plasmid, and plasmid can shift with higher frequency in different ways,so that the resistance can spread faster. According to references and sequences published in GenBank,4primers were designed and synthesized.NDM-1resistance gene, plasmid-mediated quinolone resistance gene qnrA, plasmid mediated-β-lactamase (cephalosporins) drug resistance gene SHV-1and plasmid mediated quinolone and aminoglycoside resistant gene aac(6’)-ib-cr. Polymerase chain reaction (PCR) was used to carry out gene amplification of two strains chicken-derived E. coli.The results showed that there is one positive aac(6’)-ib-cr in two strains of e. coli,and the others are negative. Transferring the plasmid with resistant genes aac (6’)-ib-cr into international standard strains of E. coli ATCC25922, and compared with the original receptor bacteria, we found that the drug resistance to19kinds of antibiotic of transformation of bacteria is obviously improved. Therefore, the formation of resistance of plasmid with resistant genes aac (6’)-ib-cr are closely related with aac(6’)-ib-cr.