PHOSPHOLIPASE: A TARGET TO DIAGNOSE INVASIVE CANDIDIASIS

说明(申请人提供)：免疫功能低下的患者是最易患侵袭性念珠菌病的患者之一。即使接受治疗，假丝酵母菌免疫低下患者的死亡率也估计在38-50%之间。目前可用的诊断试验，包括常规血培养，缺乏敏感性和特异性。因此，开发能够提供感染早期证据和/或补充现有培养和血清学检测的创新实验室方法势在必行。我们小组的最新发现表明，白色念珠菌分泌的磷脂酶B可能是白念珠菌病的一个新的诊断靶点。当我们在两种不同的念珠菌病小鼠模型中测试一对相同基因的白色念珠菌时，这一点很明显，并且表明caplb1突变体的毒力显著减弱。本提案中描述的实验旨在开发和评估基于Plb1的侵袭性念珠菌病诊断试验。这一建议的总体假设是Plb1可能作为诊断标志物有用。为了支持这一假设，我们已经证明了Plb1是分泌的，并且在体内宿主组织侵袭过程中可以检测到。重要的是，我们还发现，侵袭性念珠菌病患者(n=33)的血清中抗Plb1抗体水平显著高于健康对照组(P=0.042)。我们将通过追求以下具体目标来进一步检验这一假设：1)鉴定、表达和纯化Plb1p推导的氨基酸区域。这些多肽将用于评估Plb1作为特异性诊断标志物的潜力。2)检测兔血清中循环抗Plb1p抗体和Plb1p抗原。3)用人血清评价Plb1p的诊断价值。每种方法的有效性将在已建立的侵袭性和粘膜念珠菌病的实验动物模型以及培养证实的侵袭性念珠菌病患者和研究的对照中进行评估。

DESCRIPTION (provided by applicant): Immunocompromised patients are among the most susceptible individuals to develop invasive candidiasis. Mortality rates among candidemic immunocompromised patients have been estimated at between 38-50%, even with therapy. Currently available diagnostic tests including routine blood cultures lack sensitivity and specificity. Thus, development of innovative laboratory methods capable of providing early evidence of infection and/or complement current cultural and serological tests is imperative. Recent findings from our group suggest that phospholipase B, secreted by Candida albicans may be a novel diagnostic target for candidiasis. This was evident when we tested an isogenic strain pair of C. albicans differing only in their ability to secret PLB, in two different murine models of candidiasis, and showed that the virulence of caplb1 mutants is significantly attenuated. The experiments described in this proposal aim at developing and evaluating Plb1-based diagnostic tests for invasive candidiasis. The overall hypothesis of this proposal is that Plb1 could be useful as a diagnostic marker. In support of this hypothesis, we have shown that Plb1 is secreted and is detectable during host tissue invasion in vivo. Importantly, we also showed that sera from patients with invasive candidiasis (n=33) contained significantly higher levels of anti-Plb1 antibody compared to sera obtained from healthy controls (P = 0.042). We will examine this hypothesis further by pursuit of the following specific objectives: 1) Identification, expression and purification of regions specific to Plb1p deduced amino acid regions. These peptides will be used to evaluate the potential of Plb1 as a specific diagnostic marker. 2) Detection of circulating anti-Plb1p antibody and Plb1p antigen in rabbit sera. 3) Evaluations of diagnostic utility of Plb1p using human sera. The effectiveness of each approach will be evaluated in established experimental animal models of invasive and mucosal candidiasis and patients with culture-proven invasive candidiasis and controls for the study.