Table_5_Immunoproteomic Approach of Extracellular Antigens From Paracoccidioides Species Reveals Exclusive B-Cell Epitopes.XLSX

Fungi of the Paracoccidioides genus are the etiological agents of paracoccidioidomycosis (PCM), a systemic mycosis restricted to the countries of Latin America. Currently, the Paracoccidioides complex is represented by Paracoccidioides lutzii, Paracoccidioides americana, Paracoccidioides brasiliensis, Paracoccidioides restrepiensis, and Paracoccidioides venezuelensis. Even with advances in techniques used for diagnosing fungal diseases, high rates of false-positive results for PCM are still presented. Additionally, there is no efficient antigen that can be used to follow up the efficiency of patient treatment. The immunoproteomic is considered a powerful tool for the identification of antigens. In addition, antigens are molecules recognized by the immune system, which make them excellent targets for diagnostic testing of diseases caused by microorganisms. In this vein, we investigated which antigens are secreted by species representing Paracoccidioides complex to increase the spectrum of molecules that could be used for future diagnostic tests, patient follow-up, or PCM therapy. To identify the profile of antigens secreted by Paracoccidioides spp., immunoproteomic approaches were used combining immunoprecipitation, followed by antigen identification by nanoUPLC-MSE-based proteomics. Consequently, it was possible to verify differences in the exoantigen profiles present among the studied species. Through a mass spectrometry approach, it was possible to identify 79 exoantigens in Paracoccidioides species. Using bioinformatics tools, two unique exoantigens in P. lutzii species were identified, as well as 44 epitopes exclusive to the Paracoccidioides complex and 12 unique antigenic sequences that can differentiate between Paracoccidioides species. Therefore, these results demonstrate that Paracoccidioides species have a range of B-cell epitopes exclusive to the complex as well as specific to each Paracoccidioides species. In addition, these analyses allowed us the identification of excellent biomarker candidates for epidemiology screening, diagnosis, patient follow-up, as well as new candidates for PCM therapy.

副球孢子菌属（Paracoccidioides）真菌是副球孢子菌病（paracoccidioidomycosis, PCM）的病原菌，该系统性真菌病仅流行于拉丁美洲各国。目前，副球孢子菌复合群包含卢茨副球孢子菌（Paracoccidioides lutzii）、美洲副球孢子菌（Paracoccidioides americana）、巴西副球孢子菌（Paracoccidioides brasiliensis）、雷斯特雷帕副球孢子菌（Paracoccidioides restrepiensis）以及委内瑞拉副球孢子菌（Paracoccidioides venezuelensis）。尽管真菌疾病诊断技术已取得进展，但副球孢子菌病仍存在较高的假阳性检出率。此外，目前尚无有效的抗原可用于监测患者的治疗效果。免疫蛋白质组学被认为是鉴定抗原的有力工具；而抗原作为可被免疫系统识别的分子，是微生物感染性疾病诊断检测的优质靶标。基于此，本研究旨在探究副球孢子菌复合群各代表物种分泌的抗原种类，以拓展可用于未来诊断检测、患者随访或副球孢子菌病治疗的分子谱范围。为鉴定副球孢子菌属物种分泌的抗原谱，本研究采用免疫蛋白质组学方法：先通过免疫沉淀富集抗原，再结合基于纳米超高效液相色谱-MSE（nanoUPLC-MSE）的蛋白质组学技术进行抗原鉴定。最终，本研究验证了所研究物种间外分泌抗原谱的差异。通过质谱分析，共在副球孢子菌属物种中鉴定出79种外分泌抗原。借助生物信息学工具，本研究鉴定出卢茨副球孢子菌特有的2种外分泌抗原，同时发现副球孢子菌复合群共有的44个表位，以及12条可区分不同副球孢子菌物种的独特抗原序列。综上，副球孢子菌属物种既存在该复合群共有的B细胞表位，也存在各物种特异性的表位。此外，本研究的分析结果鉴定出了适用于流行病学筛查、诊断及患者随访的优质候选生物标志物，同时也为副球孢子菌病治疗提供了新的候选靶点。