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Isolation and Identification of Haemophilus Parasuis and the Typing by PCR-RFLP Analysis of the tbpA Gene

Author: LiuHui
Tutor: YangZengZuo
School: Northwest University of Science and Technology
Course: Preventive Veterinary Medicine
Keywords: Parasuis Isolation and identification of Susceptibility testing tbpA gene PCR-RFLP typing
CLC: S858.28
Type: Master's thesis
Year: 2011
Downloads: 111
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Abstract


Parasuis disease, also known as of pig leather Rather's disease (Gl the? Sser's Disease), is the main cause of nursery pigs caused by Haemophilus parasuis (Haemophilus parasuis, Hps) occurred polyserositis joint inflammation and meningitis, a bacterial disease. In recent years, the disease is widely popular in the world, showed a significant upward trend in morbidity and mortality caused huge economic losses to the pig industry in China. Disease hazards has been increasing its research is gradually increased, some provinces and cities have successfully isolated local parasuis strains and epidemiological studies, Shaanxi Province Vice pig Haemophilus prevalence and molecular characteristics of the epidemic strain rarely reported. This study identified five parasuis strains, isolated from the Guanzhong area of ??Shaanxi Province to establish a stable parasuis save, a common drug sensitivity test isolates and Vice pig the of Haemophilus tbpA gene PCR-RFLP typing method on five clinical isolates were genotyped, and provide some experimental evidence for the Shaanxi Province parasuis molecular epidemiological studies as well as the treatment of the disease, mainly The results are as follows: 1 in order to detect parasuis in Shaanxi Province farms infection, this test from the Guanzhong area of ??Shaanxi Province 14 of 30 samples suffering from multiple serositis and arthritis farms isolated five bacteria and its morphological observation, colony culture and biochemical characteristics of identification; according parasuis 16S rRNA primers were designed for PCR amplification of the isolates, the amplified products were ligated T vector and sequenced and sequence analysis, and found that the gene fragments obtained with GenBank at home and abroad parasuis 16S rRNA sequence homology between 99% to 100%, isolates parasuis. The results show that the Guanzhong area exists parasuis disease epidemic. 2. Parasuis preservation methods explored. Placed under different conditions by the strains were inoculated in a different culture medium was allowed to grow well after preservation, and spaced a certain time will be saved strains recovery observed survived to seek a more suitable parasuis preservation conditions . The results showed that the H. parasuis strains by TSB medium expanding twice after the culture was added to a final concentration of 20% glycerol by sterilization placed and stored at -70 ℃, is more suitable for the preservation method of H. parasuis. 3. Drug-sensitive paper method to detect five parasuis clinical isolates of nine drug sensitivity. The results showed that all strains of ofloxacin, tobramycin and kanamycin were 100% sensitive, have a higher rate of rifampicin sensitive (40%), cefuroxime, carbenicillin resistance rates were as high as 100% and 80%, respectively. This result suggests that 5 parasuis clinical isolates resistant than the strong, and the sensitivity of different strains of the same antibiotics are also different for the Guanzhong area parasuis disease treatment medication . 4 Redondo established by three restriction enzyme Taq Ⅰ, Ava Ⅰ, Afa Ⅰ on five sub-pigs Haemophilus clinical isolates of transferrin binding parasuis PCR-RFLP genotyping method, protein A encoding gene (tbpA) PCR-RFLP genotyping, four different RFLP genetic map, respectively, EBC, BBJ, EBJ EBJ, EAD, then four genotype at home and abroad Institute genotype analysis parasuis presence of multiple genotypes of strains in the Guanzhong area of ??Shaanxi Province, and has obvious geographical characteristics of each genotype, Shaanxi Province of pig Haemophilus disease molecular epidemiology study provides a theoretical basis. This test identified five parasuis strains, isolated from the Guanzhong area of ??Shaanxi Province and its drug susceptibility testing, but also isolates the based on tbpA gene PCR-RFLP genotyping for further The study provides some basis to carry out the separation of the region parasuis pathogen identification, molecular epidemiology and preventive treatment.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Livestock, poultry, wildlife diseases > Livestock > Pig
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