The aim of the study is to identify the candidate biomarkers of heat stress (HS) in the urine of lactating dairy goats through the application of proton Nuclear Magnetic Resonance (1H NMR)-based metabolomic analysis. Dairy does (n = 16) in mid-lactation were submitted to thermal neutral (TN; indoors; 15 to 20°C; 40 to 45% humidity) or HS (climatic chamber; 37°C day, 30°C night; 40% humidity) conditions according to a crossover design (2 periods of 21 days). Thermophysiological traits and lactational performances were recorded and milk composition analyzed during each period. Urine samples were collected at day 15 of each period for 1H NMR spectroscopy analysis. Principal component analysis (PCA) and partial least square-discriminant analysis (PLS-DA) assessment with cross validation were used to identify the goat urinary metabolome from the Human Metabolome Data Base. HS increased rectal temperature (1.2°C), respiratory rate (3.5-fold) and water intake (74%), but decreased feed intake (35%) and body weight (5%) of the lactating does. No differences were detected in milk yield, but HS decreased the milk contents of fat (9%), protein (16%) and lactose (5%). Metabolomics allowed separating TN and HS urinary clusters by PLS-DA. Most discriminating metabolites were hippurate and other phenylalanine (Phe) derivative compounds, which increased in HS vs. TN does. The greater excretion of these gut-derived toxic compounds indicated that HS induced a harmful gastrointestinal microbiota overgrowth, which should have sequestered aromatic amino acids for their metabolism and decreased the synthesis of neurotransmitters and thyroid hormones, with a negative impact on milk yield and composition. In conclusion, HS markedly changed the thermophysiological traits and lactational performances of dairy goats, which were translated into their urinary metabolomic profile through the presence of gut-derived toxic compounds. Hippurate and other Phe-derivative compounds are suggested as urinary biomarkers to detect heat-stressed dairy animals in practice.
Caseous lymphadenitis is an infectious disease of almost all animals, particularly small ruminants that are caused by Corynebacterium pseudotuberculosis. The organism causes the formation of suppurative abscesses in superficial and visceral lymph nodes and in visceral organs. This current study was designed to elucidate the clinicopathological responses and PCR detection of the aetiological agent in the vital organs of goats challenged with C. pseudotuberculosis and its immunogenic mycolic acid extract. A total of twelve clinically healthy crossbred Boer female goats were divided into three groups: A, B, and C (four goats per group). Group A was inoculated intradermally with 2 ml of sterile phosphate buffered saline (PBS) pH 7 as a control group. Group B was inoculated intradermally with 2 ml of mycolic acid extract (1 g/ml), while group C was inoculated intradermally with 2 ml of 10⁹ colony-forming unit (cfu) of live C. pseudotuberculosis. The experimental animals were observed for clinical responses for 90 days post-inoculation and the clinical signs were scored according to the severity. The clinical signs observed in this study were temperature, heart rate, respiratory rate, rumen motility, enlargement of lymph nodes, and body condition score. The experimental animals were euthanised and tissue samples from different anatomical regions of the vital organs were collected in 10% buffered formalin, processed, sectioned, and stained with H&E. Results of both C. pseudotuberculosis and mycolic acid treated groups indicated a significant difference (p rate in group C showed a significant increase between week 1 (93.35 ± 0.42 bpm) and week 11 (86.52 ± 1.32 bpm), and the mean heart rate of group B showed a significant increase (p rate (p rate of group B only increased at week 1 (32.98 ± 1.30 bpm) and week 2 (31.87 ± 0.48 bpm). Both groups C & B showed significant decreases in rumen motility and body condition score as compared to the control. The histopathological changes were significant in group C, this was shown by mild to severe haemorrhage, congestion, degeneration and necrosis, oedema, infiltration with inflammatory cells mainly lymphocytes and macrophages, while group B was less affected and showed mild to moderate haemorrhage, congestion, degeneration and necrosis, infiltration of inflammatory cells and oedema as compared to the control group. This study concluded that C. pseudotuberculosis caused typical CLA disease with a short incubation period in the experiment. While the mycolic acid extracted from C. pseudotuberculosis caused mild clinical signs, no abscess formation, and negative PCR result. Moreover, evidence of mild to moderate histopathological changes in vital organs was also observed.