Department of Veterinary Population Medicine, University of Minnesota, Saint Paul, USA.
BMC Microbiol. 2010 Oct 22;10:268. doi: 10.1186/1471-2180-10-268.
Two genotypically and microbiologically distinct strains of Mycobacterium avium subsp. paratuberculosis (MAP) exist - S and C MAP strains that primarily infect sheep and cattle, respectively. Concentration of iron in the cultivation medium has been suggested as one contributing factor for the observed microbiologic differences. We recently demonstrated that S strains have defective iron storage systems, leading us to propose that these strains might experience iron toxicity when excess iron is provided in the medium. To test this hypothesis, we carried out transcriptional and proteomic profiling of these MAP strains under iron-replete or -deplete conditions.
We first complemented M.smegmatisΔideR with IdeR of C MAP or that derived from S MAP and compared their transcription profiles using M. smegmatis mc(2)155 microarrays. In the presence of iron, sIdeR repressed expression of bfrA and MAP2073c, a ferritin domain containing protein suggesting that transcriptional control of iron storage may be defective in S strain. We next performed transcriptional and proteomic profiling of the two strain types of MAP under iron-deplete and -replete conditions. Under iron-replete conditions, C strain upregulated iron storage (BfrA), virulence associated (Esx-5 and antigen85 complex), and ribosomal proteins. In striking contrast, S strain downregulated these proteins under iron-replete conditions.. iTRAQ (isobaric tag for relative and absolute quantitation) based protein quantitation resulted in the identification of four unannotated proteins. Two of these were upregulated by a C MAP strain in response to iron supplementation. The iron-sparing response to iron limitation was unique to the C strain as evidenced by repression of non-essential iron utilization enzymes (aconitase and succinate dehydrogenase) and upregulation of proteins of essential function (iron transport, [Fe-S] cluster biogenesis and cell division).
Taken together, our study revealed that C and S strains of MAP utilize divergent metabolic pathways to accommodate in vitro iron stress. The knowledge of the metabolic pathways these divergent responses play a role in are important to 1) advance our ability to culture the two different strains of MAP efficiently, 2) aid in diagnosis and control of Johne's disease, and 3) advance our understanding of MAP virulence.
存在两种基因型和微生物学上不同的分支结核分枝杆菌亚种。 副结核分枝杆菌（MAP） - S 和 C MAP 菌株，主要分别感染绵羊和牛。 培养基中铁的浓度已被建议为观察到的微生物差异的一个促成因素。 我们最近证明 S 株具有缺陷的铁储存系统，这导致我们提出当培养基中提供过量的铁时，这些菌株可能会经历铁毒性。 为了检验这一假设，我们在铁充足或缺乏条件下对这些 MAP 菌株进行了转录组和蛋白质组谱分析。
我们首先用 C MAP 或 S MAP 衍生的 IdeR 对 M. smegmatisΔideR 进行了补充，并使用 M. smegmatis mc(2)155 微阵列比较了它们的转录谱。 在铁存在的情况下，sIdeR 抑制了 bfrA 和 MAP2073c 的表达，后者是一种含铁蛋白的含铁蛋白，这表明 S 株中铁储存的转录控制可能存在缺陷。 接下来，我们在铁缺乏和充足条件下对两种 MAP 菌株类型进行了转录组和蛋白质组谱分析。 在铁充足的条件下，C 株上调了铁储存（BfrA），毒力相关（Esx-5 和抗原 85 复合物）和核糖体蛋白。 与此形成鲜明对比的是，S 株在铁充足的条件下下调了这些蛋白质。 iTRAQ（相对和绝对定量的同重同位素标签）基于蛋白质定量导致鉴定出四个未注释的蛋白质。 其中两种在 C 株对铁补充的反应中上调。 对铁限制的铁节约反应是 C 株所特有的，表现为非必需铁利用酶（乌头酸酶和琥珀酸脱氢酶）的抑制和必需功能（铁运输，[Fe-S]簇生物发生和细胞分裂）的蛋白质的上调。
总之，我们的研究表明，C 和 S 株 MAP 利用不同的代谢途径来适应体外铁应激。 了解这些不同反应所涉及的代谢途径对于 1）提高我们有效培养两种不同 MAP 菌株的能力，2）有助于约翰氏病的诊断和控制，以及 3）提高我们对 MAP 毒力的理解非常重要。
