Metabolomic Changes in Naturally MAP-Infected Holstein–Friesian Heifers Indicate Immunologically Related Biochemical Reprogramming

Abstract

Johne’s disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP), causes weight loss, diarrhoea, and reduced milk yields in clinically infected cattle. Asymptomatic, subclinically infected cattle shed MAP bacteria but are frequently not detected by diagnostic tests. Herein, we compare the metabolite profiles of sera from subclinically infected Holstein–Friesian heifers and antibody binding to selected MAP antigens. The study used biobanked serum samples from 10 naturally MAP-infected and 10 control heifers, sampled monthly from ~ 1 to 19 months of age. Sera were assessed using flow infusion electrospray–high-resolution mass spectrometry (FIE–HRMS) on a Q Exactive hybrid quadrupole–Orbitrap mass spectrometer for high-throughput, sensitive, non-targeted metabolite fingerprinting. Partial least-squares discriminant analyses (PLS-DA) and hierarchical cluster analysis (HCA) of the data discriminated between naturally MAP-infected and control heifers. In total, 33 metabolites that differentially accumulated in naturally MAP-infected heifers compared to controls were identified. Five were significantly elevated within MAP-infected heifers throughout the study, i.e., leukotriene B4, bicyclo prostaglandin E2 (bicyclo PGE2), itaconic acid, 2-hydroxyglutaric acid and N6-acetyl-L-lysine. These findings highlight the potential of metabolomics in the identification of novel MAP diagnostic markers and particular biochemical pathways, which may provide insights into the bovine immune response to MAP.

Metabolomic changes in polyunsaturated fatty acids and eicosanoids as diagnostic biomarkers in Mycobacterium avium ssp. paratuberculosis (MAP)-inoculated Holstein–Friesian heifers

“…Our findings identified 11 metabolites in MAP-inoculated heifer calves that were also observed in our Taylor et al [ 16 ] study which focused on natural MAP infections. The prominence of metabolites from the fatty acyl class and fatty acids and conjugates subclass, alongside the eicosanoid subclass was notable in both studies (Additional file 2 ) [ 16 ]. However, there was an absence of metabolites belonging to the amino acids, peptides and analogues subclass in our current study (Additional file 2 ).…”

“…However, no omic study has assessed MAP infection associated changes in youngstock. Our previous metabolomic study identified 33 metabolites which were differentially accumulated within naturally MAP-infected heifer calves, including 5 metabolites which were elevated throughout the study [ 16 ]. In this study, we examined experimentally MAP-inoculated heifer calves to more clearly assess early changes occurring following infection and to validate the previously observed metabolites changes in an analogous study.…”

“…However, there was an absence of metabolites belonging to the amino acids, peptides and analogues subclass in our current study (Additional file 2 ). Changes to bioenergetic and uraemic pathways also seem specific to naturally infected heifer calves [ 16 ]. These changes may be attributable to differences in bacterial load, as indicated by the faecal culture results.…”

“…We have previously examined the sera metabolite profiles of naturally MAP-infected Holstein–Friesian (HF) heifer calves compared to controls between 1 and 19 months of age and identified 33 metabolites which were differentially accumulated in MAP-infected heifer calves [ 16 ]. Crucially, five of these metabolites were elevated throughout the study.…”

Early prediction of paratuberculosis health status in cattle by integrating NMR analysis and blood indices through the LASSO method

“…In details, the metabolomics studies, carried out on experimentally MAP-infected cattle using 1 H nuclear magnetic resonance (NMR) spectrometry, showed metabolomics changes related to energy shortages, increased fat metabolism, and altered protein turnover (12). Taylor et al examined the sera metabolic pro les of naturally MAP-infected Holstein-Friesian heifers and compared them to controls, nding changes in some metabolites related to biochemical reprogramming (16,17). In our previous study, we applied direct analysis in real time high resolution mass spectrometry (DART-HRMS) coupled to a midlevel data fusion approach to tease out molecular features able to chemically characterize the animal sera at the MAP preclinical stage (15).…”