Abstract
Anthelmintic resistance in parasitic nematodes, particularly in Haemonchus contortus, poses a significant threat to livestock health and productivity. Since resistance mechanisms have not yet been fully elucidated, the present study employed metabolomic and proteomic analyses of H. contortus adults (females and males separately) from drug-susceptible (ISE) and benzimidazole-resistant (IRE) strains, with a focus on resistance-specific differences. Using nuclear magnetic resonance (NMR)-based metabolomics and mass spectrometry-based proteomics, significant alterations in metabolic and protein expression profiles associated with resistance were identified. Resistant adults exhibited reduced levels of multiple amino acids and tricarboxylic acid cycle intermediates, alongside elevated levels of trehalose, myo-inositol, NAD(+), and glycerophosphocholine, suggesting enhanced stress resistance and altered energy metabolism. Proteomic analysis revealed over 3000 proteins, with substantial sex-related differences. Moderate resistance-related differences in protein expression were observed, indicating adaptive biochemical pathways supporting resistance. While some resistance-associated changes in the metabolome and proteome were observed in both sexes, several were distinctly sex-specific. This finding provides the first evidence of such sex-dependent strategies in H. contortus, reinforcing the notion that anthelmintic resistance is a multifaceted and highly complex phenomenon, with many aspects yet to be fully understood. In addition, certain nematode-specific enzymes upregulated in the IRE strain - such as cysteine synthases and transthyretin-like proteins - might be considered as potential targets for future anthelmintic development.