| With the unceasingly araplication of caprine cultivacation scale in our country, The sorts of caprine' s disease are also unceasingly increasing. In rencent years,a kind of chronical pneumonia in caprine is popular, which is one new disease after scale cultivationffhen the disease occure, the caprine shows cough, dyspnoea, chronic emaciaton and die in the end. It make great loss in economic and pose severely threaten to the flock of caprine cultivacation. After epidemiological investigation and pathogen inspecting, the pathogen Pseudomonas aeruginosa was idenficated..The aim of this study is on the bases of isolating and identificating the pathengon, investicating the episode status of the disease in Shandong province, and pay more attention to the study of the main virulence factor----Exotoxin A, for to lay bases for studying the pathogenesis. This studyincludes four parts.Parti:I so I at ion and identification of caprine Pseudomonas aeruginosa Using micorobioloy method, systemical micorobiology diagnosis was proceeded to one kind of new chronic pneumonia pathogen. After isolating and cultivation microscop, founding that the pathogen is one kind of G short bacilli, it haven' t spore and capsule, but have flagella, it can produce water disolved blue and green pigment. Using biochemical test and epidemiological investigation and other tests, it was identificated as pseudomonas areuginosa. With the 24hr broth culture of this bacterium separately inoculated 3-day chick(0. Iml/per) > 10-day chichen embryo(0. Iml/per) and rabbit(0. 3ml/per), all the animals die in 24h. After inoculated healthy caprine, it replicated the same clinical symptom as the naturely diseased goat.Parti I: Proper at ion of plate agglutination and sero logical investigation of caprine chronical pneumonia by Pseudomonas aeruginosa .Plate agglutination antigen was preparated with the purified pathogen. After stability test , sterile test , self-agglutination test and crossing agglutination test, It proved that the plate agglutination antigen was good. With the diagnostic antigen, from November of the year 2001 till May of 2002, 246 caprine was inspected in Tai' an, Heze, Jining> Liaocheng, leifang, Jinan, Rizhao, Laiwu of Shandong province, plate agglutination test was used to test the antibody titer. The result manifest that the rate of positivityis between 75%~100%, the mean antibody titer is 1. 61og2?. Olog2, and the mean titer of the caprine with clinical symptom is 3.001og2 . Statistical analysis suggest that excepting Tai' an is significantly higher than Heze, Weifang, Rizhao, Jinan, the mean difference in other areas isn' t significant. All the flock of caprine that was investigated lies the positive infection.PartlllrStudies on PEA PA-SEHl was grown in a dialyzed medium of tripticase soy broth enriched with 1% glycerol(v/v) and 1/20 volume of 1M monosodium glutamate, The cultures were shaken at 32C at 120 cycles per min for 22 hr. The cultural supernatant fluid was precipitated by the addition of (NH<):S04 to 30% saturation to get rid of the mixed protein and then (NH<)2S04 to 60% saturation to extract the crud exotoxin, DEAE and sephedex G-200 was used to purify the exotoxin A. Concentration was detected by spectrophotography, and purification was detected by SDS-PAGE, electrophoresis result show one strap of 66KD molecular weight, the exotoxin A was innoculated into 15g BALB/C mouse, there was no pathogenicity with 0. 085g/per. But four-fold concentrated 0. 2ml supernatant can cause the mouse die. Result suggests that Pseudomonas aeruginosa isolate PA-SD-1 can produce Exotoxin A, but only exotoxinA can' t cause the mouse die, this result indicate that it must have other virulence factor that cause the disease. But Exotoxin A was used to immune the rabbit, after several times immnunity, the antibody titer can reach 101og2, it suggests that the exotoxin A have good immunogenicity. With the Exotoxin A, ELISA method was created to inspect the anti- PEA antibody of the caprine serum, this method have high sensitivity a... |
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