Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, Massachusetts, USA.
PLoS Pathog. 2012;8(8):e1002828. doi: 10.1371/journal.ppat.1002828. Epub 2012 Aug 2.
A highly conserved virulence plasmid encoding a type III secretion system is shared by the three Yersinia species most pathogenic for mammals. Although factors encoded on this plasmid enhance the ability of Yersinia to thrive in their mammalian hosts, the loss of this virulence plasmid does not eliminate growth or survival in host organs. Most notably, yields of viable plasmid-deficient Yersinia pseudotuberculosis (Yptb) are indistinguishable from wild-type Yptb within mesenteric lymph nodes. To identify chromosomal virulence factors that allow for plasmid-independent survival during systemic infection of mice, we generated transposon insertions in plasmid-deficient Yptb, and screened a library having over 20,000 sequence-identified insertions. Among the previously uncharacterized loci, insertions in mrtAB, an operon encoding an ABC family transporter, had the most profound phenotype in a plasmid-deficient background. The absence of MrtAB, however, had no effect on growth in the liver and spleen of a wild type strain having an intact virulence plasmid, but caused a severe defect in colonization of the mesenteric lymph nodes. Although this result is consistent with lack of expression of the type III secretion system by Wt Yptb in the mesenteric lymph nodes, a reporter for YopE indicated that expression of the system was robust. We demonstrate that the ATPase activity of MrtB is required for growth in mice, indicating that transport activity is required for virulence. Indeed, MrtAB appears to function as an efflux pump, as the ATPase activity enhances resistance to ethidium bromide while increasing sensitivity to pyocyanin, consistent with export across the inner membrane.
三种对哺乳动物最具致病性的耶尔森氏菌共有一个高度保守的毒力质粒，该质粒编码一个 III 型分泌系统。尽管该质粒上编码的因子增强了耶尔森氏菌在其哺乳动物宿主中茁壮成长的能力，但失去这种毒力质粒并不会消除其在宿主器官中的生长或存活。最值得注意的是，缺乏可育质粒的假结核耶尔森氏菌（Yptb）的产量与野生型 Yptb 在肠系膜淋巴结中没有区别。为了鉴定允许在质粒缺失的情况下在系统性感染小鼠时生存的染色体毒力因子，我们在缺乏质粒的 Yptb 中生成转座子插入，并筛选了一个拥有超过 20000 个序列鉴定插入的文库。在以前未表征的基因座中，mrtAB 操纵子编码 ABC 家族转运蛋白的插入在缺乏质粒的背景下表现出最明显的表型。然而，MrtAB 的缺失对具有完整毒力质粒的野生型菌株在肝脏和脾脏中的生长没有影响，但导致肠系膜淋巴结定植严重缺陷。尽管这一结果与 Wt Yptb 在肠系膜淋巴结中缺乏 III 型分泌系统的表达一致，但 YopE 的报告表明该系统的表达非常强烈。我们证明了 MrtB 的 ATP 酶活性是在小鼠中生长所必需的，表明运输活性是毒力所必需的。事实上，MrtAB 似乎作为一个外排泵发挥作用，因为 ATP 酶活性增强了对溴化乙锭的抗性，同时增加了对绿脓菌素的敏感性，这与内膜的跨膜运输一致。
