Dairy cows affected by intramammary infection, subclinical and clinical mastitis: changes in milk metabolome profiles

Immune and inflammatory responses during mammary tissue infection of dairy cows lead to an increase in SCC in milk. Metabolomics is an important branch of system biology that studies metabolites to elucidate disease etiology and characteristics, to monitor disease and develop biomarkers. Several studies were conducted to study milk metabolomic profiles. However, knowledge about the influence of only intramammary infection on milk metabolome are still lacking. The aim of this study was to investigated the quarter-milk metabolomic profile of dairy cows affected by intramammary infection, subclinical mastitis, and clinical mastitis. Multiparous Holstein-Friesian dairy cows from one farm. Animals were between 30 and 100 days in milk, not under antimicrobial treatment, and without an history of treatment in the previous 6 months. Among them 80 quarters were enrolled and equally divided in four groups: healthy (H) quarters with SCC<200.000 cells/mL and negative bacteriological milk cultures (BMC); intramammary infection (IMI) with SCC<200.000 cells/mL and positive BMC; subclinical mastitis (SCM) with SCC<200.000 cells/mL, positive BMC, and absence of clinical signs; and clinical mastitis (CM) with SCC<200.000 cells/mL, positive BMC, and presence of clinical signs. Milk metabolomics analysis were conducted by 1H-NMR. Differences in metabolites were assessed with a linear mixed-effects model. A p-value<0.05 was accepted, whereas a 0.05≤p-value≤0.10 was considered as trend to significance. The SCC were 49.200, 66.100, 1.982.800, and 11.834.700 cells/mL for H, IMI, SCM, and CM, respectively. Main pathogens were Staphylococcus aureus (24.4%) and Escherichia coli (23.3%). 45 metabolites were identified of which 34 differed significantly among groups. 12 metabolites differed only in CM compared to all other groups with greater levels in leucine, taurine, valine, acetate, formate, pyruvate, 5-dodecenoic acid, and 3-hydroxybutyrate, and lower levels in arabinose, galactose, ribose, and ascorbate. Other 18 metabolites changed from H to CM, with a progressive increase of 2-aminoadipate, alanine, dimethylamine, n-acetyl-glycine, and lactate, and decrease of creatine-phosphate, o-phosphocholine, glucose, lactose, maltose, cis-aconitate, carnitine, fumarate, phenylacetate, 2-ketobutyrate, acetoacetate, citicoline, and orotate. In conclusion, both SCM and CM showed impaired energy and lipid metabolisms, and altered systemic energy status with worse conditions during CM. Moreover, milk metabolome suggested a progressive increase in inflammation, blood-milk barrier damage, oxidative stress, and microorganism metabolism. Increased cell proliferation was suspected during IMI and SCM, with progressive substrate consumption in CM.