Objective To detect rpoB mutations in rifampin resistant mycobacterium tuberculosis strains isolated from Shanghai, and to evaluate the implication of applicating line probe assay (LiPA). Methods A fragment (213bp) of rpoB gene of 58 Mycobacterium tuberculosis isolates was amplified and sequenced, 18 rifampin resistant strains and 10 susceptible strains were selected to detect mutation by LiPA. Results Mutations of rpoB gene in 17 strains of the 18 rifampin resistant isolates were found by LiPA, and there were no mutations in any of the 10 susceptible strains. The sensitivity of LiPA was 94.4% and the concordance with drug susceptibility of Mycobacterium tuberculosis was 96.4%. Conclusions LiPA is a useful method for the rapid detection of mutations of rpoB gene in rifampin resistant Mycobacterium tuberculosis with high sensitivity.

Objective To compare the phage based splitting assay and BACTEC 460 in the rapid detection of Mycobacterium tuberculosis ( M. tuberculosis ). Methods 30 clinical isolates of M. tuberculosis, 10 strains of non M. tuberculosis, 7 strains of non mycobacterium and 60 sputum specimens of pulmonary tuberculosis patients were detected with phage based splitting assay and BACTEC 460 system. Results All the strains of M. tuberculosis clinical isolates detected with phage based splitting assay were positive, while all of the non M. tuberculosis and non mycobacterium strains were negative. 41 sputum specimens with BACTEC 460 culture positive and 19 sputum specimens with BACTEC 460 negative were detected with phage based splitting assay, the number of positive specimens was 34 (82.9%) and 5 (26.3%) respectively.Conclusions The phage based splitting assay can detect the M. tuberculosis easily and quickly in two days with high sensitivity and specificity.

Objective To develop new method for rapid detection of mutations in rpoB gene related with resistance to rifampin of M. tuberculosis . Methods According to the sequence of wild type M. tuberculosis, five oligonucleotide probes covering the 69 bp hyper variable region of rpoB gene were designed and immobilized on nylon membrane strips. Thereafter, the target rpoB gene fragment was obtained by PCR using biotin labeled primers and thereafter the PCR product was denatured and hybridized with probes on membrane. The results of reverse dot blot hybridization were compared with the results of drug sensitivity test and sequencing. Results PCR products from 36 RFP resistant and 22 RFP susceptible isolates were detected by the assay of reverse dot blot hybridization, showing that the susceptibility and the specificity rate were 88.9% and 86.4% respectively,and the coincidence with biochemical method and sequencing is 87.9% and 89.7% respectively. Conclusions Reverse dot blot hybridization is a rapid and sensitive method to detect the rpoB gene mutations,which may be used in early detection of the resistance of M. tuberculosis to rifampin.