Chansap Supanan, Cheukamud Werachon, Suthisintong Thitikul, Kueakhai Pornanan, Changklungmoa Narin
Research Unit for Vaccine and Diagnosis of Parasitic Diseases, Faculty of Allied Health Sciences, Burapha University, Chonburi, Thailand.
Front Vet Sci. 2025 May 22;12:1547937. doi: 10.3389/fvets.2025.1547937. eCollection 2025.
Fasciolosis is an important zoonotic disease caused by species ( spp.). spp. infection has the potential to affect the livestock economy. Furthermore, liver flukes have been found to present Triclabendazole resistance in many countries. Vaccines are used to prevent fasciolosis and are currently considered the best alternative. However, no liver fluke vaccine is commercially available at present. Cathepsin Ls (FgCatLs) are vital enzymes for the liver fluke's survival. Therefore, this study aimed to design and investigate the immune response of multi-epitope Cathepsin L (MeCatL) driven short peptide vaccine for fasciolosis using immunoinformatic tools. FgCatLs sequences were predicted Linear B cell (BCL)- and Helper T lymphocyte (HTL)-specific immunogenic Eepitopes. The selected epitopes were marked on FgCatL's alignments. Novel epitopes were constructed from three criteria, including the selection process taking non-conserved host regions, overlapping FgCatLs sequences, and the highest percent conserved residues. Novel epitopes of BCL and HTL were linked with a linker to design a short peptide. MeCatL driven short peptide presented high antigenicity, non-allergenicity, non-toxicity, and good solubility. MeCatL driven short peptide was predicted and refined the tertiary structure. The refined MeCatL driven short peptide model indicated good quality structure that was investigated by Ramachandran plot, ERRAT, and Z-score. The refined MeCatL driven short peptide model interacted with Toll-like receptor 2 (TLR-2). The lowest energy was -1222.4 kJ/mol. The levels of IgM, IgG1, and IgG2 were increased in immune simulation. MeCatL driven short peptide was synthesized and immunized in mice. IgG1 and IgG2a levels were increased after week 2, indicating IgG1 levels were dominating. MeCatL driven short peptide immunized sera can detect single proteins, including rFgCatL1, rFgCatL1G, and rFgCatL1H. In addition, MeCatL driven short peptide immunized sera was specifically detected in the cecal epithelium of NEJ and adult stages. These findings suggest that the MeCatL short peptide is a promising vaccine candidate, capable of inducing targeted immune responses, though further studies are needed to confirm its protective efficacy .
肝片吸虫病是由肝片吸虫属（Fasciola spp.）引起的一种重要的人畜共患病。Fasciola spp.感染有可能影响畜牧业经济。此外，在许多国家已发现肝片吸虫对三氯苯达唑产生耐药性。疫苗用于预防肝片吸虫病，目前被认为是最佳选择。然而，目前尚无商业化的肝片吸虫疫苗。组织蛋白酶L（FgCatLs）是肝片吸虫生存的关键酶。因此，本研究旨在利用免疫信息学工具设计并研究多表位组织蛋白酶L（MeCatL）驱动的肝片吸虫病短肽疫苗的免疫反应。预测FgCatLs序列的线性B细胞（BCL）和辅助性T淋巴细胞（HTL）特异性免疫原性表位。在FgCatL的比对中标记所选表位。根据三个标准构建新表位，包括选择非保守的宿主区域、重叠FgCatLs序列以及最高百分比的保守残基。将BCL和HTL的新表位用连接子连接以设计短肽。MeCatL驱动的短肽具有高抗原性、无致敏性、无毒性且溶解性良好。预测并优化了MeCatL驱动短肽的三级结构。通过拉氏图、ERRAT和Z分数对优化后的MeCatL驱动短肽模型的质量结构进行了研究，结果表明其结构良好。优化后的MeCatL驱动短肽模型与Toll样受体2（TLR - 2）相互作用，最低能量为 - 1222.4 kJ/mol。在免疫模拟中，IgM、IgG1和IgG2水平升高。合成MeCatL驱动的短肽并在小鼠中进行免疫。第2周后IgG1和IgG2a水平升高，表明IgG1水平占主导。MeCatL驱动的短肽免疫血清可检测包括rFgCatL1、rFgCatL1G和rFgCatL1H在内的单一蛋白质。此外，在日本分体吸虫童虫和成虫阶段的盲肠上皮中特异性检测到MeCatL驱动的短肽免疫血清。这些发现表明，MeCatL短肽是一种有前景的疫苗候选物，能够诱导靶向免疫反应，不过还需要进一步研究来证实其保护效果。
