Genetic Characterization of phosphomannan biosynthesis in C. auris

金黄色念珠菌是一种新近出现的真菌病原菌。这种新出现的物种已经在全球范围内崛起 自它在日本首次被发现以来的过去十年里。这种生物已经成为一个公共卫生问题，因为 它的抗真菌能力和它在重症监护和疗养院设施中坚持的能力。其中一个原因是 这种持续能力的背后是许多临床分离株表现出多重耐药性，在极少数情况下， 可以抵抗摇摆不定。最近，我们对8个金黄色葡萄球菌的细胞壁进行了检测，发现金黄色葡萄球菌 甘露聚糖含有额外的甘露醇磷酸(Mα1-PO4)分枝，这是该物种所独有的。这是一个 非常令人兴奋和潜在的非常重要的新发现。金龟子的外层是甘露聚糖层 这在结构上不同于所有其他念珠菌物种，实际上，任何其他真菌病原体，表明 Mα1-PO4结构是金黄色葡萄球菌区别于其他真菌病原菌的主要原因。在这一点上，几乎没有什么是 已知控制这种独特细胞壁表型表达的基因。在白色念珠菌中 Mnt3、Mnt5和MNN4蛋白是负责磷酸甘露糖的磷酸甘露糖基转移酶 生物合成。此外，还有7种类似MNN4的蛋白质在功能上与MNN4是冗余的，但 不在磷甘露聚糖的生物合成中起主要作用。一项对念珠菌基因组数据库的调查显示 除了5个类似MNN4的基因外，Auris还含有MNT3、MNT5和MNN4基因。由于 金黄色葡萄球菌与白色念珠菌基因的相似性我们推测MNT3、MNT5和 MNN4基因在磷甘露聚糖的生物合成中保持保守的作用，而五个基因中的一个或多个 MNN4样基因已经分化，产生影响Mα1-PO4分支和/或 磷酸化。我们将通过构建缺失突变体来评估这些基因在金黄色葡萄球菌中所起的作用 要确定它们是否影响磷甘露聚糖的生物合成和M1α-PO4分支的添加，使用高 现场核磁共振。此外，我们将确定这些突变体是否增加或降低了对 人类巨噬细胞及其杀死巨噬细胞的能力。从这项工作中，我们希望开发出更好的 对金黄色葡萄球菌甘露聚糖生物合成的理解。生成的数据将有助于为潜在的诊断提供服务 以及未来的疫苗开发。

Candida auris is a recently emergent fungal pathogen. This newly emergent species has risen worldwide over the last ten years since its first recognition in Japan.This organism has become a public health problem due to its antifungal resistance and its ability to persist in critical care and nursing home facilities. One of the reasons behind this ability to persist is that many of the clinical isolates show multidrug resistance and, in rare cases, can be pan-resistant. Recently we examined the cell wall of eight C. auris isolates and discovered that C. auris mannans contain additional mannosyl phosphate (Mα1-PO4) branching that is unique to this species. This is a very exciting and potentially very important new finding. The fact that C. auris has an outer coating of mannan that is structurally distinct from all other Candida species and, indeed, any other fungal pathogen, suggests that the Mα1-PO4 structure distinguishes C. auris from other fungal pathogens. At this point, virtually nothing is known about the genes which control the expression of this unique cell wall phenotype. In C. albicans the Mnt3, Mnt5, and Mnn4 proteins are phosphmannosyltransferases responsible for phosphomannan biosynthesis. In addition there are seven Mnn4-like proteins that are functionally redundant with Mnn4 but do not play a major role in phosphomannan biosynthesis. A survey of the Candida Genome Database revealed that C. auris also contains the MNT3, MNT5, and MNN4 genes in addition to five MNN4-like genes. Due to the similarities of the genes in C. auris to those of C. albicans we hypothesize that the MNT3,MNT5 and MNN4 genes maintain a conserved role in phosphomannan biosynthesis, while one or more of the five MNN4-like genes have diverged to produce proteins that impact the Mα1-PO4 branching and/or phosphorylation. We will assess the role that these genes play via construction of deletion mutants in C. auris to determine if they impact phosphomannan biosynthesis and the addition of M1α-PO4 branching, using high field NMR. In addition we will determine if these mutants have increased or decreased attractiveness to human macrophages and their ability to kill the macropahges. From this work we expect to develop a better understanding of mannan biosynthesis in C. auris. The data generated will help to serve potential diagnostic and vaccine development in the future.