Sdudy On Sika Deer Tuberculosis By Genetyping

Tuberculosis is a sort of bacterial infection between human and animal,its hosts include the Sika Deer,which is one of the most important economic animals of China.The Mycobacterium tuberculosis and M.bovis are the pathogenic bacteria of Deer tuberculosis, which also can infect human.So the sick deer,especially the open tuberculosis deer could be the main animal reservoir.So the main objective of this paper was making research on the molecular epidemiology of the deer tuberculosis in the Northeast of PRC with MIRU and Spoligotyping,cmpared the difference of this two methoeds,investigate the situation of deer tuberculosis in China's north-eastern region,and analyse the genetype characteristics.At first,we did the isolating culture and identification.The details were the comparison of different pretreatments and culture mediums,morphology characterization and PCR characterization.The result indicated that the acid treatment were better than alkali treatment on the positive rate,7H9 medium made a good appearance on isolating culture but for the subculture,L-J and sodium pyruvate medium were more compatible.After the characterization,we finally separated 120 mycobacterium tuberculosis in samples collected from the northeast of China(JiLin,LiaoNin,HeiLongJiang),incuding 92 mycobacterium tuberculosis,28 M.bovis.The MIRU was applied to the research of molecular epidemiology of the Sika Deer tuberculosis.The 12-MIRU loci were amplified with PCR which extracted from the above-mentioned mycobacterium tuberculosis,the number of copies of tandem repeats of 12 MIRU were confirmed based on the length of PCR amplification products.The discriminatory power of MIRU was assessed by Hunter-Gaston index(HGDI).We found that 120 distinct patterns were seperated by MIRU typing with 12 loci yield,the resolution was 100%,which was more higher than that used in human tuberculosis.The results of diversity analysis showed:MIRU4,MIRU16,MIRU24,MIRU26,MIRU39,MIRU40 were highly discriminative(h≥0.6),MIRU2,MIRU10,MIRU20,MIRU23,MIRU31 were moderately discriminative(0.6＞h≥0.3),and loci MIRU27 were poorly discriminative(h=0.0697),this result description MIRU has a high ablity of resolution.Strains have less relevant,genetyping cluster of the strains are not appaer.The apply of Spoligotyping on the molecular epidemiology of the Sika Deer tuberculosis in the Northeast of PRC.The DR loci were amplified by PCR which used extracted genomic DNA of MTB isolates as template.Determind genetype by probe hybridization and chemiluminescence.The result of experimence of Spoligotyping distinction 120 strains for 34-type,and 29-type has 1 strain,the other type has 4 or more strains.Comparing between MIRU and Spoligotyping,the resolution of MIRU is higher. It had a better genetype result for pathogens of Sika deer TB with 6-MIRU loci which has higher polymorphism,the time spent on experimence will be shorter.It makes MIRU becomes more quick and concise.Beacuse 43-probewas needed in experimence,the costs of Spoligotyping are higher than MIRU,for large-scale testing of Mycobacterium tuberculosis it is not enough economic So MIRU will be used as preferred method in study on the molecular epidemiology of the Sika Deer tuberculosis.Spoligotyping can use as a added.Strains in different regions have different character in the Northeast of PRC,the genetyping cluster of the strains are not appaer,it is that tha TB of sika deer in the Northeast of PRC was prevented and controled better,large-scale outbreak of TB diseases did not happen,only some sporadic cases were found in different areas,and cross-infection is minor.