The Molecular Epidemiological Survey And Establishment Of Diagnosis Target Cluster System Platform Of Mycobacterium Tuberculosis Based On Bioinformatics

The resurgence of tuberculosis in developing countries has become amajor public health issue of global concern. The high rate of missingreport, the spread and the regional expansion of tuberculosis are closelyassociated with the limitation of its detection level. The currentsituation of tuberculosis detection is not optimistic, which is still stuck inthe comprehensive analysis such as specimen smear, culture ofMycobacterium tuberculosis (needs4-6weeks), x-ray, CT screening, ESR,PPD testing, etc. However, the detection rate of TB was only32%byusing these traditional diagnostic methods. Gene detection technique andits application on Mycobacterium detection is still in the stage ofdevelopment, which lack of high sensitivity and specific targets. In addition, the genotypes and phenotypes of Mycobacterium tuberculosisare extremely complex and volatile, the virulence genes which has beenidentified and the genes which has been used in detection cannot meet thenew requirements of early diagnosis of diseases. Therefore, the selectionof TB diagnostic targets by high-throughput screening is needed urgently,it can settle molecular foundation for the establishment of a sensitive,specific and rapid detection technology. Then realize the early prevention,early detection, and early treatment of TB.In this article, we make an investigation on molecularepidemiology of tuberculosis in Jilin province firstly, then use dynamicsimulation of signal transduction networks based on genome-wide data ofTB, regulate the high-throughput annotation of central node molecule andSignal transduction protein coupling, then realize the high-throughputscreening of TB essential gene cluster and diagnostic target spectrum,provide molecular basis for early diagnosis of mycobacterium tuberculosis. Finally, large-scale verification was made onmycobacterium tuberculosis clinical strains in Jilin province.The specific content of this study are as follows:1. The research classified the21clinical strains of Mycobacteriumtuberculosis through17VNTR sites (12MIRU sites and5ETR sites).5ETR-VNTR sites only classified the strains into6genotypes, and HGDIis0.7381; the combination of12MIRU-MIRU sites and12MIRU sitesand5ETR sites classified the strains into8genotypes, HGDI are all0.8619. By VNTR sites allele diversity (h) analysis, we found6siteswhich has better classification effects in Jilin province, they are: MIRU26,40,10,27,31,39. The11strains of the21strains in this study wereBeijing strains, which accounts for52%of all strains, this illustrated thatBeijing genotype is the prevalent genotype in Jilin province. We drawevolutionary trees by the12MIRU sites and found3strains were similarto the TB strains form other regions, which is14%of total strains. 2. This study used dynamic simulation of signal transductionnetworks based on genome-wide data of TB, screened out857possibleessential genes; then found out55genes which has not been proven butmost likely to be essential genes by comment analysis of signalingpathway number and gene functions as well as the verification ofbiblio-metrological method, finally, the696genes verified bybiblio-metrological method and the55genes screened by pathwayenrichment method, and we made annotations to essential genes andrelated signal transduction networks of TB by Operon high throughputscreening database and gene sequencing rules of microbial, together withthe prediction on the corresponding functions of essential genes and theclassification of functions of essential genes, and eventually establishedthe diagnostic target cluster platform of TB.3. PCR is a simple, rapid, sensitive and specific gene diagnosistechnology, but there are still some problems, in the process of DNA amplification, because the specificity of diagnostic target is not thatstrong, false-positive and false-negative results may occur, therefore,therefore, the method needs to be further improved. If diagnostic targetsare essential genes which are highly conserved and generally not changed,the stability of the diagnostic targets can be greatly improved; same time,we need to be screened the genes only exist in Mycobacteriumtuberculosis and does not exist in other microorganisms, the specificity ofthese genes are very sharp as a diagnostic target. Finally, by usingGenome-wide high-throughput scanning technology, we screened12candidate genes through diagnostic target cluster platform of TB, andthen by Biblio-metrics method and genetic stability checking, Rv1513,Rv1974and Rv3738were screened out as diagnostic targets oftuberculosis. We verified15clinical strains of TB by PCR and comparedthe diseases which have similar early symptoms with TB, such as asthma,pneumonia and lung cancer and selected3genes as new high-specific diagnostic targets of tuberculosis.