Molecular Epidemiology Of Mycobacterium Tuberculosis Isolates In Yangzhou

Tuberculosis(TB)is one of the most important zoonotic diseases in the world,which seriously endangers human health and poses a serious challenge to the global public health security.In 1882,Koch,a German scientist,revealed that Mycobacterium tuberculosis(MTB)a pathogen of tuberculosis,which opened up a thorough study of tuberculosis.MTB is a small,and rod-shaped gram positive bacterium with no flagellum or spores.The presence of MTB can be detected by acid-fast stain.In clinical,it mainly infects human lungs,and patients often show severe cough,accompanied by phlegm or blood silk,body weakness,chest pain,low fever,night sweats and other symptoms.According to the WHO reports,there are about 10.4 million of new TB patients in the world during 2016,with about 1.03 million of TB/HIV double infection,and 600 thousand of MDR-TB infection,and 14 countries are facing the challenges of TB,TB/HIV and MDR-TB,including China.Therefore,the clinical diagnosis,drug treatment and surveillance of MTB have become the top priority in the prevention and control of major infectious diseases in China.In this study,MTB isolates in Yangzhou was collected,and the epidemiological investigation and drug sensitivity test of MTB were carried out.The genotype characteristics of MTB in Yangzhou were studied by MLVA molecular typing,and the potential correlation between genotypes and drug resistance,sex,age and time was analyzed.By whole genome squencing analysis,drug resistance prediction and accuracy of the predicted results were evaluated.1 Epidemiological studies of tuberculosis in YangzhouFrom the perspective of population distribution,male more likely to be infected than female in Yangzhou,and the prevalence level of themobile population is 1.5 times than that of the local population.In age distribution,most female patients were under 35 years old or above 60 years old than other age groups,showing a "double peaks" phenomenon.In terms of occupational distribution,TB patients are mainly in three occupations of farmers,retirees and householders.From the time distribution,tuberculosis in Yangzhou is mainly prevalent in the two seasons of spring and summer.311 strains of MTB and 16 strains of nontuberculous mycobacteria(NTM)were identified by PNB/TCH test and multilocus PCR from 327 clinical isolates collected in Yangzhou,including one strain of M.canettii and one strain of M.africanum subtype I.The drug sensitivity test of 176 strains of MTB showed that 76 strains were resistant to drug resistance,and the overall resistance rate was 43.18%,which was 1.08%higher than the level of the national epidemiological survey on tuberculosis.It indicated that the general drug resistance level in Yangzhou was higher and the relapse patients were more susceptible to drug resistance than those of the new patients.From the perspective of bacteriology,smear negative and culture positive strains had a high risk of drug resistance.Thus,attention should be paid to smear negative and culture positive patients with drug sensitivity test,and timely and accurate diagnosis,is urgent to prevent the emergence of drug resistance.2 MLVA study of Mycobacterium tuberculosis in Yangzhou24 MIRU-VNTR gene loci were selected to study the molecular typing of 325 MTB isolates collected in Yangzhou.11 strains of MTB were found to be mixed infection strains.Repeated analysis of VNTR gene loci of 314 single strains of MTB showed that different strains had different preferences for different VNTR loci.By calculating,the clustering rate is 11.46%,and the possible infectious risk factors of cluster strains are analyzed,which benefits the visit investigation.At the same time,the polymorphism of 24 MIRU-VNTR gene loci were analyzed.The high resolution sites were Mtub21 and QUBllb.The middle resolution loci were Mtub04,MIRU40,MIRU10,MIRU16,MIRU26,MIRU31;ETRE,QUB26,QUB4156,ETRA and MIRU31,and these 12 loci were more suitable for the study of the molecular typing of MTB in Yangzhou.Compared with the relevant international databases,the genotypes of MTB in Yangzhou has obvious aggregation and conservative.3 Whole-genome sequencing analysis of Mycobacterium tuberculosis isolates in Yangzhou94 strains of MTB were sequenced from the clinical isolates of Yangzhou.Their whole genome sequences were obtained and the drug resistance of each strain was predicted.Compared with the results of drug sensitivity test,the consistency rate was more than 70%,of which the four first-line anti-tuberculosis drugs of INH,RFP,EMB and SM were 82.56%,95.24%,86.90%and 86.90%respectively,and four second-line anti-tuberculosis drugs of CPM,AK,PAS and LVFX were 92.86%,91.67%,94.40%,73.81%respectively.It also shows that the drug resistance prediction based on the whole genome sequence has a high relibility,except for LVFX.The total genomic sequences of 94 MTB in Yangzhou were analyzed with the core SNVs database.84 strains were lineage 2 and 10 strains were lineage 4.Lineage 2 was the most prevalent in Yangzhou,and all of them belong to Beijing family.Cluster analysis of 94 MTB strains based on the whole genome sequence was consistent with the results of lineage analysis.