Ru Huanwei, Liu Xiaojia, Lin Chen, Yang Jingyan, Chen Fuzeng, Sun Ruifeng, Zhang Lu, Liu Jun
State Key Laboratory of Genetic Engineering, School of Life Science, Institute of Genetics, Fudan UniversityShanghai, China.
Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, Fudan UniversityShanghai, China.
Front Cell Infect Microbiol. 2017 Jun 8;7:239. doi: 10.3389/fcimb.2017.00239. eCollection 2017.
Comparative genome analyses have revealed a number of regions of difference (RD) among mycobacterial species. The functional consequences of most of these genome variations have not been studied. RD4, which encompasses Rv1506c-Rv1516c of () H37Rv, is absent in the closely related and Bacille Calmette-Guérin (BCG). On the other hand, we previously found that has an extended RD4 which includes a number of genes involved in the biosynthesis of lipooligosaccharides (LOSs). As such, there appears to be a gradual decay of RD4 in mycobacterial genomes in the order of , and (including BCG). To understand the potential effect of RD4 on mycobacterial virulence, in this study, we cloned the entire (Rv1501-1516c) and partial (Rv1501-1508c) RD4 into an integrating vector. These constructs were introduced to BCG and and the virulence of the RD4 knock-in strains were evaluated in the SCID mice and zebrafish infection models, respectively. BCG containing the entire RD4 exhibited similar levels of virulence to the parental strain but BCG containing partial RD4 (Rv1501-Rv1508c) was more attenuated. Similarly, zebrafish infection experiments showed that addition of partial RD4 also appeared to attenuate the virulence of . However, containing entire RD4 was more virulent than the wild type strain. Interestingly, BCG strains containing the entire or partial RD4 exhibited better protection of zebrafish against challenge than the parental BCG. Taken together, our data suggest that RD4 plays a role in mycobacterial virulence and that RD4 knock-in BCG strains confer improved protection. Our study has provided new insights into the biological function of RD4 and evolution of mycobacterial genomes.
比较基因组分析揭示了分枝杆菌属物种之间的一些差异区域（RD）。这些基因组变异中大多数的功能后果尚未得到研究。RD4包含（）H37Rv的Rv1506c - Rv1516c，在密切相关的和卡介苗（BCG）中不存在。另一方面，我们之前发现具有扩展的RD4，其中包括一些参与脂寡糖（LOS）生物合成的基因。因此，在分枝杆菌基因组中，RD4似乎按照、和（包括BCG）的顺序逐渐衰退。为了了解RD4对分枝杆菌毒力的潜在影响，在本研究中，我们将整个（Rv1501 - 1516c）和部分（Rv1501 - 1508c）RD4克隆到一个整合载体中。将这些构建体分别导入BCG和，并且在SCID小鼠和斑马鱼感染模型中评估RD4敲入菌株的毒力。含有整个RD4的BCG表现出与亲本菌株相似的毒力水平，但含有部分RD4（Rv1501 - Rv1508c）的BCG毒力更弱。同样，斑马鱼感染实验表明添加部分RD4也似乎减弱了的毒力。然而，含有整个RD4的比野生型菌株毒力更强。有趣的是，含有整个或部分RD4的BCG菌株比亲本BCG对斑马鱼抵御攻击的保护作用更好。综上所述，我们的数据表明RD4在分枝杆菌毒力中起作用，并且RD4敲入BCG菌株提供了更好的保护。我们的研究为RD4的生物学功能和分枝杆菌基因组的进化提供了新的见解。
