Isolation, Indentification Of The Escherichia Coli Causing Edema Disease Of Swine And Cloning And Expression Of FedF Gene

Edema disease of swine(ED) was first reported by Shanks in 1938.ED is a fatal disease caused by certain serotypes of Escherichia coli ,which usually occurs in pigs shortly after weaning when various nutrional and environmental factors predispose them to the disease. ED has been found throughout the world and usually reported in our country.ED is one main disease which plaqued the swine farmery.ED is an endemic disease or a sporadically occurring disease .The incidence of the disease is about from 30% to 40% in the affected swinery. Farmers reported that, on average, mortality increased from 80% to 100% following an outbreak of ED. Average daily gain (ADG) tended to be lower on farms with the problem. The disease appears to be of economic significance and poses a challenge as far as control.Increased risk of resistance among E. coli has been associated with the use of various antimicrobials in the feed.The disease caused a great economic loss in swine farmery.ED is a serious disease mainly for post-weaned piglets that causes systemic vascular damage as a result of intestinal infection with Shiga toxin-producing Escherichia coli (SLTEC). After SLTEC colonizes in the intestine via the F18 pilus, Shiga toxin 2 variant(Stx2e) produced by theSLTEC is absorbed into the capillary vessel and then damages endothelial cells.The.*-manifestations of ED include palpebral edema, neurological impairment, lateralrecumbence, and sudden death?Some E.coli isolates recovered from pigs with diarrhea may also secrete heat-lable(LTl) or heat-liable (ST1 or STII) enterotoxins (Wittig et al,1995)Fifty seven strains Escherichia coli have been isolated from swine.Seven strains of Escherichia coli isolated from provinces having edema were identified by Gram dyeing,biochemical reaction and serotyping. These strains had pathogenicity and were able to produce Vero toxin,which could be confirmed by the following examinations. Their broth culture could lead kunming mice to death.Axenic filtrate of the broth culture could induce death of kunming mice and Vero cell. These strains were tested by PCR to dectect genes of SLT-IIe,F18,LTl and ST1. Seven strains were positive for SLT-IIe, five strains were positive for F18, but all were negative for LT1 and ST1. The results accorded with the literature having being reported. Antibiotics susceptibility test showed their resistance patterns were different. All of the seven strains could lead porcine to having edema and then to death.We selected ED3(O139) field isolated strain which lead little mouse to death in the least dose.The results of the experiment showed that ED3 field isolated strain injection had its LD50 as 2.55 X 108 CFU for little mice.On the other hand,we preparedmuiti-inactivated thalli antigen to study the immunogenicity to little mice. Its immunogenicity in pig was detected by micro-aggulutination test .The GMT was 1:8 7days post-inncculation.The GMT reached its peak 21 days post-innoculation and it was 1:128. The GMT maintained higher level as 1:64 55 days post-innoculation.The 967bp nucleotide fragment including the whole encoding regions of fedF was amplified by PCR from a field E.coli strain called Ee which responsible for the edema disease in piglets in Hubei province. The 967bp and 897 nucleotide fragment.was cloned into pMD-18T vector. The 967 nucleotides sequence including the whole encoding regions of fedF between site Sad and Hindlll compared with the strain 107/86 reported by foreigner. Result showed that the nucleotides sequence homology between Ee Sac I and site Hindlll were analyzed and the amino acids were deduced .The nucleotides sequence of whole encoding region and deduced amino acids were compared between strain Ee and 107/86 was 100% and amino acids homology was 100%.Comparsion of the fedF gene with the AF/R1 pilus .The levels of identity of the protein homologs was zero. After R.E. analysis and sequencing, the fedF gene in pMD18-T was ligated in pGEX-KG, the recombinant plasmid pGEX-KG/fedF was constructed and analysed with R.E., the protein of fedFs...