Research Group Biomarkers for Infectious Diseases, Helmholtz Centre for Infection Research, Braunschweig, Germany.
Institute for Biochemistry, Biotechnology and Bioinformatics, Braunschweig University of Technology, Braunschweig, Germany.
PLoS Negl Trop Dis. 2019 Jan 24;13(1):e0007131. doi: 10.1371/journal.pntd.0007131. eCollection 2019 Jan.
Leptospirosis is the most common zoonotic disease worldwide. The diagnostic performance of a serological test for human leptospirosis is mainly influenced by the antigen used in the test assay. An ideal serological test should cover all serovars of pathogenic leptospires with high sensitivity and specificity and use reagents that are relatively inexpensive to produce and can be used in tropical climates. Peptide-based tests fulfil at least the latter two requirements, and ORFeome phage display has been successfully used to identify immunogenic peptides from other pathogens.
METHODOLOGY/PRINCIPAL FINDINGS: Two ORFeome phage display libraries of the entire Leptospira spp. genomes from five local strains isolated in Malaysia and seven WHO reference strains were constructed. Subsequently, 18 unique Leptospira peptides were identified in a screen using a pool of sera from patients with acute leptospirosis. Five of these were validated by titration ELISA using different pools of patient or control sera. The diagnostic performance of these five peptides was then assessed against 16 individual sera from patients with acute leptospirosis and 16 healthy donors and was compared to that of two recombinant reference proteins from L. interrogans. This analysis revealed two peptides (SIR16-D1 and SIR16-H1) from the local isolates with good accuracy for the detection of acute leptospirosis (area under the ROC curve: 0.86 and 0.78, respectively; sensitivity: 0.88 and 0.94; specificity: 0.81 and 0.69), which was close to that of the reference proteins LipL32 and Loa22 (area under the ROC curve: 0.91 and 0.80; sensitivity: 0.94 and 0.81; specificity: 0.75 and 0.75).
CONCLUSIONS/SIGNIFICANCE: This analysis lends further support for using ORFeome phage display to identify pathogen-associated immunogenic peptides, and it suggests that this technique holds promise for the development of peptide-based diagnostics for leptospirosis and, possibly, of vaccines against this pathogen.
钩端螺旋体病是全球最常见的人畜共患病。血清学检测用于人类钩端螺旋体病的诊断性能主要受检测方法中使用的抗原影响。理想的血清学检测应具有高灵敏度和特异性，涵盖所有致病性钩端螺旋体血清型，并使用相对廉价的试剂生产，可在热带气候下使用。基于肽的检测至少满足后两个要求，而 ORFeome 噬菌体展示已成功用于鉴定其他病原体的免疫原性肽。
方法/主要发现：构建了来自马来西亚五个地方株和七个世界卫生组织参考株的整个钩端螺旋体属全基因组的两个 ORFeome 噬菌体展示文库。随后，使用来自急性钩端螺旋体病患者的血清池筛选出 18 种独特的钩端螺旋体肽。其中 5 种通过使用不同患者或对照血清池的滴定 ELISA 进行验证。然后，使用来自 16 例急性钩端螺旋体病患者和 16 例健康供体的 16 份血清评估这 5 种肽的诊断性能，并与来自 L.interrogans 的两种重组参考蛋白进行比较。该分析显示，两种来自本地分离株的肽（SIR16-D1 和 SIR16-H1）具有良好的准确性，可用于检测急性钩端螺旋体病（ROC 曲线下面积：分别为 0.86 和 0.78；灵敏度：分别为 0.88 和 0.94；特异性：分别为 0.81 和 0.69），接近参考蛋白 LipL32 和 Loa22（ROC 曲线下面积：分别为 0.91 和 0.80；灵敏度：分别为 0.94 和 0.81；特异性：分别为 0.75 和 0.75）。
结论/意义：该分析进一步支持使用 ORFeome 噬菌体展示来鉴定与病原体相关的免疫原性肽，这表明该技术有望用于开发基于肽的钩端螺旋体病诊断方法，并可能开发针对该病原体的疫苗。
Zotero 插件