Identification Of Biological Characteristics And Evaluation Of Lipid Inhibitors Of Genes Related To Biofilm Formation Of Mycobacterium Tuberculosis Rv1904,Rv3229c And Rv3818

Mycobacterium tuberculosis,as a pathogen that can cause zoonotic diseases,has caused widespread concern all over the world,and tuberculosis is one of the main causes of human death.It has been found that the biofilm of M.tuberculosis plays an important role in its infection and survival in vitro.The viability of M.tuberculosis in biofilm is better than that of planktonic growth under the same conditions.Biofilm is recognized as the effect of drug tolerance,and its existence can reduce the prevention and control effect of anti-tuberculosis drugs.In addition,the biofilm of M.tuberculosis may be related to intracellular survival and host defense escape mechanism.Therefore,the study on the mechanism of biofilm formation can provide new ideas for the prevention and control of M.tuberculosis,and biofilm formation-related genes are also important potential targets for new anti-tuberculosis therapy.The biofilm culture of 18 strains of recombinant Mycobacterium smegmatis preserved in our laboratory was carried out.The biofilm formation was observed and the biofilm production was quantitatively compared by crystal violet staining.Rv1904,Rv3229c and Rv3818,which can significantly increase the biofilm production of M.smegmatis(P<0.001),were selected as the target genes.Rv1904 could enhance the staining effect and sliding ability of Sudan Ⅲ in the biofilm of M.smegmatis.Rv1904 can also enhance the antioxidant capacity,low temperature and high temperature tolerance and hunger tolerance of M.smegmatis(P<0.001).Rv1904 also enhanced the intracellular colonization ability of M.smegmatis and M.tuberculosis(P<0.001).RT-PCR results showed that Rv1904 upregulated the transcription level of fabG4 related to fatty acid biosynthesis pathway(P<0.001).The formation of MS_Rv1904 biofilm decreased after the addition of two thiourea drugs(TB1 and TMTU).Rv3229c can enhance the staining effect,sliding ability and colony aggregation ability of Sudan Ⅲ in the biofilm of M.smegmatis.Rv3229c could enhance the tolerance of M.smegmatis to SDS and starvation.Rv3229c also enhanced the intracellular colonization ability of M smegmatis and M.tuberculosis(P<0.001).Rv3229c can enhance the resistance of M.smegmatis to rifampicin in planktonic state and to rifampicin and isoniazid in biofilm state.RT-PCR results showed that Rv3229c also upregulated the transcription level of fabG4(P<0.001).The formation of MS_Rv3229c biofilm decreased after the addition of two thiourea drugs.Rv3818 could enhance the staining effect,sliding ability,antioxidant capacity,low temperature tolerance and hunger tolerance of Sudan Ⅲ in the biofilm of M.smegmatis.Rv3818 also enhanced the intracellular colonization ability of M.smegmatis and M.tuberculosis(P<0.001).RT-PCR results showed that Rv3818 not only upregulated the transcription level of fabG4,but also upregulated the transcriptional levels of cysW,cysT and subl in sulfur metabolism related pathways(P<0.001).The formation of MS_Rv3818 biofilm decreased after the addition of two thiourea drugs.In summary,Rv1904,Rv3229c and Rv3818 Rv1904,Rv3229c and Rv3818 can promote the biofilm formation of M.tuberculosis and M.smegmatis;thiourea drugs,isoniazid and streptomycin alone can inhibit the biofilm formation of M.smegmatis;thiourea drugs combined with anti-tuberculosis drugs have a stronger inhibitory effect on biofilm formation.