Non-targeted multimodal metabolomics data from ovine rumen fluid fractions

Abstract

In nematodes that invade the gastro-intestinal tract of the ruminant, the process of larval exsheathment marks the transition from the free-living to the parasitic stages of these parasites. To investigate larval exsheathment, a closed in vitro system that effectively reproduces the two basic components of an anaerobic rumen environment (CO2 and 39 °C) was developed to trigger exsheathment in one of the most pathogenic and model gastrointestinal parasitic nematode, Haemonchus contortus (barber‘s pole worm). This project applied the basic principles of dialysis and is the first reported use of all currently available non-specific metabolomics methodologies to identify the rumen-derived trigger compounds associated with larval development in H. contortus. Herein, we report an multimodal untargeted metabolomics and lipidomics analyses based on 20 kDa (n = 10), 8-10 kDa (n = 10) and 100 Da (n = 10) molecular weight cut-off dialyzed rumen fluid fractions, all of which demonstrated positive effects on exsheathment induction in vitro under rumen conditions, as compared to control infective third-stage larva (L3) in saline (n = 10). Metabolite identification and multivariate analyses resulted in a catalogue of over one thousand structurally annotated compounds or metabolites that form three distinct cluster groups and warrant further investigation. The multimodal metabolomics analyses revealed significant alterations between the three different rumen fluid fractions in L-lactic acid, pyruvate metabolism, gluconeogenesis, the Warburg effect, plant-pathogen interaction, plant hormone signal transduction, and biosynthesis of amino acids metabolism. Lysophosphatidylglycerol, diglyceride, phosphatidylcholine, triglyceride and a sterol were the major lipid classes identified by the non-targeted lipidomics method applied. A combination of univariate and multivariate statistical analyses revealed a panel of metabolites directly responsible for larval exsheathment and associated with amino acids as well as purine and pyruvate metabolism KEGG pathways. Among these were compounds such as methionine, leucine, alanine, phenylalanine, xanthine and hypoxanthine, that were significant in the control group and identified using two separate metabolomics streams. This study presents an in vitro system suitable for numerous functional omics’ experimentation applications and has enhanced our understanding of the distinct molecular and metabolic pathways associated with larval exsheathment. The identified metabolites may serve as metabolic signatures or novel metabolic biomarkers for pre-diagnostic purposes of haemonchosis. Alternatively, our findings can be used to develop next-generation biomimicry agents to cause infective larvae of parasitic nematodes to exsheath outside the host while on pasture for experimental or control purposes.