Zhang Jing-rui, Li Gui-lian, Zhao Xiu-qin, Wan Kang-lin, Lü Jian-xin
Wenzhou Medical College School of Laboratory Medicine, Key of Laboratory Medicine, Wenzhou 325035, China.
Zhonghua Liu Xing Bing Xue Za Zhi. 2013 Apr;34(4):379-84.
To detect the changes on the expression of putative drug efflux genes caused by isoniazid-inducement in single resistance to the isoniazid Mycobacterium tuberculosis (M. tuberculosis) clinical isolates, for exploring the putative efflux genes which causing M. tuberculosis isoniazid resistance as well as the mechanism related to high expression of the putative efflux genes.
We selected 35 M. tuberculosis clinical isolates which were only resistant to isoniazid as well as 10 sensitive M. tuberculosis clinical isolates and using H37Rv as control. Each strain was cultured in 7H9 liquid medium without isoniazid and with subinhibitory isoniazid concentration (1/4 MIC) induction. After RNA extraction and reverse transcription, real-time PCR was conducted to assess the expression changes of 27 putative drug efflux pump genes with formula 2(-ΔΔCT) to calculate the expression of each putative drug efflux pump genes.
Of the 27 putative genes, 13 of them were expressed at high level. High expression of Rv1258c gene had the maximum number of 6 strains, followed by high expression of Rv0849 and Rv2265 which both had 5 strains. Fourteen strains (40.00%) out of the 35 strains had high expression pump genes. Six strains (17.14%) had only one highly expressed putative efflux pump gene. Eight strains (22.86%) had two or more highly expressed putative efflux pump genes, including two, four, five, seven genes that highly expressed in 4, 2, 1, 1 strains respectively. For the 27 putative genes, ten sensitive strains and H37Rv did not show highly expressed genes.
Rv1258c, Rv2265, Rv0849, etc. genes might be the putative efflux pumps genes of M. tuberculosis resistant to isoniazid. Isoniazid might serve as the inducer of M. tuberculosis part putative efflux pump genes, inducing activation and causing high expression of these putative efflux pump genes.
检测异烟肼诱导对异烟肼单耐药的结核分枝杆菌临床分离株中假定药物外排基因表达的变化，以探索导致结核分枝杆菌异烟肼耐药的假定外排基因以及与假定外排基因高表达相关的机制。
我们选取了35株仅对异烟肼耐药的结核分枝杆菌临床分离株以及10株敏感的结核分枝杆菌临床分离株，并以H37Rv作为对照。将每株菌分别在不含异烟肼和含亚抑菌浓度异烟肼（1/4 MIC）诱导的7H9液体培养基中培养。提取RNA并逆转录后，进行实时PCR以评估27个假定药物外排泵基因的表达变化，采用公式2(-ΔΔCT)计算每个假定药物外排泵基因的表达量。
在27个假定基因中，有13个高表达。Rv1258c基因高表达的菌株数量最多，为6株，其次是Rv0849和Rv2265基因高表达，均为5株。35株菌株中有14株（40.00%）存在高表达泵基因。6株（17.14%）仅有一种高表达的假定外排泵基因。8株（22.86%）有两种或更多种高表达的假定外排泵基因，其中分别有4、2、1、1株菌高表达2、4、5、7个基因。对于27个假定基因，10株敏感菌株和H37Rv未显示高表达基因。
Rv1258c、Rv2265、Rv0849等基因可能是结核分枝杆菌对异烟肼耐药的假定外排泵基因。异烟肼可能作为结核分枝杆菌部分假定外排泵基因的诱导剂，诱导激活并导致这些假定外排泵基因的高表达。
