Fascioliasis, caused by the liver flukes Fasciola gigantica and F. hepatica, is a neglected tropical disease affecting livestock and humans with substantial economic and public health consequences. In Punjab, Pakistan, F. gigantica is the predominant species infecting cattle, buffalo, sheep, and goats. Proteomic data, particularly on parasite-derived excretory–secretory proteins (ESPs), are important to elucidate host–parasite interactions, mechanisms of immune modulation, and potential biomarkers, but comprehensive proteomic information for F. gigantica from buffalo in Pakistan has been limited. The primary objective of this study was to present a detailed proteomic characterization of adult F. gigantica recovered from naturally infected buffaloes in Punjab, with specific focus on putative ESPs.
Adult flukes were collected from naturally infected buffaloes. Somatic protein extracts were initially examined by SDS–PAGE and Western blotting, which confirmed both a high abundance of parasite proteins and a broad distribution of molecular weights. Proteomic profiling used liquid chromatography–tandem mass spectrometry (LC–MS/MS) to identify proteins present in the samples.
In silico approaches were applied to predict subcellular localization and secretion. DeepLoc 2.0, SignalP, and transmembrane helix prediction (TMHMM) were employed to distinguish classical secretory proteins (with N-terminal signal peptides) from non-classical secreted proteins that lack transmembrane helices.
Mass-spectrometry analysis generated the largest proteomic dataset reported for F. gigantica to date, with a total of 4,777 proteins identified across three adult fluke samples. The authors report that these data represent a broad proteomic repertoire from somatic extracts, encompassing proteins across molecular functions and cellular compartments relevant to parasite physiology and host interaction.
Using the combined outputs from DeepLoc 2.0, SignalP, and TMHMM, the study identified a substantial set of putative ESPs, including both classical secretory proteins containing N‑terminal signal peptides and non-classical secreted proteins that lack transmembrane helices. The predicted secretome was highlighted as a key element governing parasite strategies for tissue invasion, nutrient acquisition, and modulation of host immunity.
Functional annotation and Gene Ontology (GO) enrichment analyses revealed that the identified proteins were predominantly associated with catalytic, hydrolase, and oxidoreductase activities. These molecular functions align with established roles of parasite proteins in degrading host tissues, processing nutrients, and managing redox balance during infection.
The authors report that, of the proteins linked to host-interaction roles, 95 were associated with molecular functions, 77 with biological processes, and 65 with cellular components relevant to immune regulation, stress response, metabolic adaptation, and parasite survival. Pathway enrichment further implicated protein processing and signal transduction pathways in facilitating host invasion and establishment of chronic infection.
The proteomic profile emphasizes the multifaceted role of both somatic and putative secreted proteins in orchestrating host–parasite dynamics. ESPs are presented as central mediators capable of interfering with host immunity at multiple levels, from pathogen recognition to effector responses, thereby contributing to immune suppression and the persistence of infection. The dataset provides candidate proteins that may warrant further evaluation as biomarkers for early detection, diagnostic targets, or components for future vaccine research, although functional validation beyond in silico and proteomic annotation was not reported in the source.
Raw proteomic data generated from the three adult F. gigantica samples have been deposited in the Figshare repository and are publicly accessible at the DOI cited in the article. The work was funded by the Higher Education Commission, Pakistan (HEC-NRPU 15618) and the Punjab Agriculture Research Board (Grant No. 21-67). The authors declare no competing interests. The study was received on February 16, 2026, accepted on August 4, 2026, and published on August 21, 2026 in PLOS ONE.