Development of multi-epitope driven subunit vaccine against Fasciola gigantica using immunoinformatics approach Parismita Kalita a , Denzelle Lee Lyngdoh a , Aditya K. Padhi b , Harish Shukla a , Timir Tripathi a, ⁎ a Molecular and Structural Biophysics Laboratory, Department of Biochemistry, North-Eastern Hill University, Shillong 793022, India b Laboratory for Structural Bioinformatics, Field for Structural Molecular Biology, Center for Biosystems Dynamics Research, RIKEN, 1-7-22 Suehiro, Tsurumi, Yokohama, Kanagawa 230-0045, Japan abstract article info Article history: Received 12 May 2019 Received in revised form 3 July 2019 Accepted 3 July 2019 Available online 04 July 2019 Fascioliasis, a serious helminth disease of the livestock population, results from infection with the parasite Fasciola. Despite the alarming increase in drug resistance, a safe and fully effective vaccine for fascioliasis is still not available. In the present study, we employed high-throughput immunoinformatics approaches to design a multi-epitope based subunit vaccine using seven important F. gigantica proteins (cathepsin B, cathepsin L, leucyl aminopeptidase, thioredoxin glutathione reductase, fatty acid binding protein-1, saposin-like protein-2, and 14–3-3 protein epsilon). The CTL, HTL, and B-cell epitopes were selected for designing the vaccine on the basis of their immunogenic behavior and binding affinity. The engineered vaccine showed potential immunogenic ef- ficacy by elaborating the IFN-γ and humoral response. The modeled structure of the vaccine was docked with the toll-like receptor-2 immune receptor, and the molecular dynamics simulation was performed to understand the stability, interaction, and dynamics of the complex. Finally, in silico cloning of the resulting vaccine was per- formed to create the plasmid construct of vaccine for expression in an appropriate biological system. Experimen- tal evaluation of the designed vaccine construct in an animal model may result in a novel and immunogenic vaccine that may confer protection against F. gigantica infection. © 2019 Elsevier B.V. All rights reserved. Keywords: Liver flukes Immunoinformatic approaches Adjuvant Immunogenic epitopes Subunit vaccine Molecular dynamics simulation 1. Introduction Fasciola gigantica and F. hepatica are the platyhelminthic endopara- sites responsible for causing fascioliasis, a liver fluke disease. Fascioliasis is a neglected tropical disease (NTD) that mostly affects the farm ani- mals and human populations of developing and under-developed coun- tries. The World Health Organization (WHO) estimated that at least 2.4 million people are infected in N70 countries worldwide, with several million at risk [1]. Human cases occurred occasionally, but are now in- creasingly reported from Europe, America, Africa, South and South- East Asia [2]. Fascioliasis is prevalent in N150 countries worldwide, pos- ing a serious threat to both human health and livestock that attributes to mortality, reduction in milk and meat production, secondary bacterial infections, and expensive anthelmintic treatment [3,4]. This disease is principally treated with a single WHO-approved drug, Triclabendazole (TCZ), which is active against both immature and adult parasites [5]. The increasing incidence of animal as well as human fascioliasis, along with the emergence of TCZ-resistant parasite populations makes vaccination an alternative strategy for controlling the disease [6–10]. We have recently sequenced the whole genome of F. gigantica that has an assembled length of ~1.04 GB, encoding 20,858 genes in the genome [11]. When the parasite migrates through the peritoneal cavity to the liver, and finally to the bile ducts, they encounter a combination of both humoral and cellular immune responses inside the host. The para- sites tackle this condition by manipulating cellular processes via im- mune evasion and subversion [12,13]. Till now no commercial vaccine was available against fascioliasis; however, researchers have attempted to impose immunoprophylactic control in sheep and cattle using para- site antigens, such as glutathione-S-transferase (GST), cathepsin L-like cysteine proteases, fatty acid binding protein (FABP), leucine amino- peptidases (LAP), and fluke hemoglobin [14–17]. Vaccination with GST could provide ~57% protection to sheep infected with Fasciola and also in Taenia ovis infection [18]. Cysteine endopeptidases play an im- portant role in host-parasite interactions [19,20]. Cathepsin L1 and L2 show predominance in the E/S products of adult and juvenile F. hepatica flukes [19], and are known to participate in the digestion of host tissues, tissue penetration, immune evasion, and feeding [21–23]. Vaccination of cattle with these antigens resulted in 42–69% reduction in fluke burden and ~60% reduction in egg viability [24]. However, in International Journal of Biological Macromolecules 138 (2019) 224–233 ⁎ Corresponding author at: Department of Biochemistry, North-Eastern Hill University, Shillong 793022, India. E-mail address: timir.tripathi@gmail.com (T. Tripathi). https://doi.org/10.1016/j.ijbiomac.2019.07.024 0141-8130/© 2019 Elsevier B.V. All rights reserved. Contents lists available at ScienceDirect International Journal of Biological Macromolecules journal homepage: http://www.elsevier.com/locate/ijbiomac