Influence of epinephrine reactivity to stress on meat quality in goats.

The magnitude of physiological responses to a stressor can vary among individual goats within a herd; however, whether these differences can differentially affect meat quality is not known. This study was conducted to determine the influence of the magnitude of epinephrine response (ER) to acute stress on muscle metabolome and meat quality in goats. Male Spanish goats (6 mo old) were transported for 180 min. (N = 75 goats; 25 goats/d) to impose stress. Blood samples were obtained after transport for analysis of physiological responses. Goats were slaughtered using humane procedures and samples were collected for muscle metabolomics and meat quality analyses. The data obtained from blood and muscle/meat analysis were then categorized based on epinephrine concentrations into low (LE), medium (ME), and high (HE) ER groups (n = 12/ER group). The physiological and meat quality variables were analyzed as a Completely Randomized Design in SAS, and metabolomics data were analyzed using R software. Plasma glucose concentrations were significantly high in the HE group, low in the LE group, and intermediate in the ME group (P < 0.05). However, leukocyte counts and cortisol, norepinephrine, blood urea nitrogen, and creatine concentrations were not different among the ER groups. Muscle (Longissimus dorsi) glycogen concentrations (15 min postmortem) were significantly higher (P < 0.05) in the ME and LE groups than in the HE group. However, postmortem Longissimus muscle pH and temperature (15 min and 24 h), 24 h calpastatin and desmin levels, and rib chop color (L*, a*, and b*), cooking loss, and Warner-Bratzler shear force values were unaffected by ER. Targeted metabolomics analysis of Longissimus muscle (15 min) revealed that diacyl phosphatidylcholines (C38:0; 40:6) and sphingomyelin (C20:2) were significantly different (P < 0.05) among the ER groups, with the concentrations of these metabolites being consistently high in the LE group. These differential muscle metabolite concentrations suggest that ER can influence biochemical pathways associated with cell membrane integrity and signaling. ER had a significant effect on dopamine concentrations, with the levels increasing with increasing levels of ER. The results indicate that differences in epinephrine reactivity can influence selected physiological responses and muscle metabolites; however, it does not significantly influence meat quality attributes.

Habituation to Livestock Trailer and Its Influence on Stress Responses during Transportation in Goats.

This experiment was conducted to determine the effects of habituation to livestock trailers on stress responses in goats transported for long periods. Intact male Spanish goats (12-month old; BW = 31.6 ± 0.34 kg; N = 168) were separated into two treatment (TRT) groups and maintained on two different paddocks. Concentrate supplement was fed to one group inside two livestock trailers (5.0 × 2.3 m each; habituated group, H), while the other group received the concentrate supplement, but not inside the trailers (non-habituated, NH). After 4 weeks of habituation period, goats were subjected to a 10-h transportation stress in four replicates (n = 21 goats/replicate/TRT). Blood samples were collected by a trained individual by jugular venipuncture into vacutainer tubes before loading (Preload), 20 min after loading (0 h), and at 2-h intervals thereafter (Time) for analysis of stress responses. There was a tendency for a TRT effect (p < 0.1) on tyramine and metanephrine concentrations. Phenylethylamine and 5-methoxytryptamine concentrations were significantly greater (p < 0.05) in the H group compared to the NH group. Both dopamine and 5-methoxytryptamine concentrations decreased (p < 0.05) with transportation time; however, TRT × Time interaction effects were not significant. Habituation to trailers may be beneficial in mood and energy stabilization in goats during long-distance transportation.

Differential gene expression analysis using RNA-seq in the blood of goats exposed to transportation stress.

Transportation stress causes significant changes in physiological responses in goats; however, studies exploring the transcriptome of stress are very limited. The objective of this study was to determine the differential gene expressions and related pathways in the blood samples using RNA-seq procedure in Spanish goats subjected to different durations of transportation stress. Fifty-four male Spanish goats (8-mo old; BW = 29.7 ± 2.03 kg) were randomly subjected to one of three treatments (TRT; n = 18 goats/treatment): (1) transported for 180 min, (2) transported for 30 min, or (3) held in pens (control). Blood samples were collected before and after treatment for stress hormone, metabolite, and transcriptomic analysis. RNA-seq technology was used to obtain the transcriptome profiles of blood. Analysis of physiological data using SAS showed that plasma cortisol concentrations were higher (P < 0.01) in 180 min and 30 min groups compared to the control group. Enrichment analysis of DEGs related to transportation stress through Gene Ontology and KEGG databases revealed that the differentially expressed genes related to inflammatory pathways, caspases, and apoptosis such as IL1R2, CASP14, CD14, TLR4, and MAPK14 were highly enriched in the transported group of goats compared to non-transported goats. Stress in goats leads to a sequence of events at cellular and molecular levels that causes inflammation and apoptosis.

Metabolomic exploration of the effects of habituation to livestock trailer and extended transportation in goats.

Goats raised for meat production are often transported long distances. Twelve-month-old male Spanish goats were used to determine the effects of habituation to trailers on plasma metabolomic profiles when transported for extended periods. In a split-plot design, 168 goats were separated into two treatment (TRT; whole plot) groups and maintained on two different paddocks. Concentrate supplement was fed to one group inside two livestock trailers (habituated group, H), while the other group received the same quantity of concentrate, but not inside the trailers (non-habituated, NH). Goats were subjected to a 10-h transportation stress in 4 replicates (n = 21 goats/replicate/TRT) after 4 weeks of habituation period. Blood samples were collected prior to loading, 20 min after loading (0 h), and at 2, 4, 6, 8, and 10 h of transportation (Time; subplot). A targeted quantitative metabolomics approach was employed to analyze the samples. The data were analyzed using R software and MIXED procedures in SAS. Several amino acids (alanine, serine, glycine, histidine, glutamate, trans-hydroxyproline, asparagine, threonine, methylhistidine, ornithine, proline, leucine, tryptophan) were higher (p < 0.05) in the H group compared to the NH group. Six long-chain acylcarnitines were higher (p < 0.05), while free (C0) and short-chain (C3, C5) carnitines were lower (p < 0.05) in the NH goats compared to the H goats. In general, amino acid concentrations decreased and long-chain acylcarnitine (>C10) levels increased with transportation time (p < 0.05). Butyric acid, α-ketoglutaric acid, and α-aminoadipic acid concentrations were lower (p < 0.05) and ß-hydroxybutyric acid concentrations were higher in the NH goats compared to the H goats. Plasma glucose, non-esterified fatty acid (NEFA) and urea nitrogen concentrations were significantly influenced by Time (p < 0.01). Plasma NEFA concentrations were significantly lower (p < 0.01) in the H group than the NH group. Habituation to trailers can be beneficial in enhancing stress coping abilities in goats due to higher concentrations of metabolites such as butyrate and certain amino acids that support antioxidant activities and immune function. Plasma long-chain acylcarnitines may be good indicators of stress during long-distance transportation in goats.

RNA-Seq exploration of the influence of stress on meat quality in Spanish goats.

Studies exploring the transcriptome of stress and its effects on meat quality are very limited, particularly in goats. Fifty-four male Spanish goats (8-mo old; BW = 29.7 ± 2.03 kg) were randomly subjected to one of three treatments (TRT; n = 18 goats/treatment): (1) transported for 180 min, (2) transported for 30 min, or (3) held in pens (control) to analyze the transcriptome of stress and meat quality in goats using RNA-seq technology. Blood samples were collected before and after treatment, and meat samples were collected after humane slaughter for stress hormone, meat quality (Longissimus dorsi), and transcriptomic analysis. Plasma epinephrine concentrations were higher (P < 0.01) in 180 min and 30 min groups compared to the control group; however, norepinephrine concentrations were not affected by the treatment. Muscle glycogen concentrations (15 min postmortem) were lower (P < 0.01) in both 30 min and 180 min groups compared to the control group. Calpastatin levels were higher (P < 0.01) in 180 min and 30 min groups than the control group. Warner-Bratzler shear force values of loin chops were the highest in the 180 min group (4 ± 0.15, kg), lowest in the control group (3.51 ± 0.10, kg), and intermediate in the 30 min group (3.78 ± 0.09, kg; P < 0.01) both at day 1 and day 6 aging time. Additionally, desmin levels of day 6 samples were lowest in the control group, highest in 180 min group, and intermediate in 30 min group (P < 0.05). RNA-seq results showed that a total of 10,633 genes were differentially expressed (5194 up regulated; 5439 down regulated) among all comparisons (blood and day 1 and day 6 muscle samples). Among these differentially expressed genes (DEGs), KLF9, AMPK, FOXO3, PTX3, GADD45, PTPN1, CASP7, MAPK4, HSPA12A, and JAK-STAT were probably associated with the effects of stress on skeletal muscle proteins and involved in biological process such as cellular response to corticosteroid stimulus, endoplasmic reticulum stress, insulin resistance, DNA repair, apoptosis, MAPK cascade and regulation of proteolysis. The KEGG analysis revealed that AMPK and JAK-SAT signaling pathways and autophagy were among the top 20 enriched pathways in our treatment comparisons. The results provide an understanding of the genes and pathways involved in stress responses and related changes in postmortem muscle metabolism and meat quality characteristics in goats.

Influence of Excitability Rate on Physiological Responses to Stress in Goats.

This study was conducted to determine if excitability score (ES) in goats can influence their physiological responses when subjected to stress. Thirty-six intact male Spanish goats (8-mo-old) were individually weighed and scored for excitability: 1 for calm (13 goats), 2 for moderately excitable (11 goats), and 3 for highly excitable (12 goats). To impose stress, goats were assigned to one of three treatments (TRT) for 90 min: (i) isolation in an open pen with metal grill panels, (ii) isolation in a pen with side panels covered using tarp sheets, and (iii) no isolation (control). Blood samples were collected at 0, 30, 60, and 90 min of isolation and physiological data were analyzed using MIXED procedures in SAS. The data from the two isolation treatments were pooled and compared with that of the control group. Plasma cortisol and non-esterified fatty acid concentrations were the lowest in goats with ES 1 (p < 0.05). Neutrophil−lymphocyte ratios were also the lowest in goats with a calm temperament (p < 0.01). Application of full quadratic model using response surface methodology (PROC RSREG) in SAS revealed that the influence of ES on physiological stress responses over time was not the same between the TRT groups. The results indicate that physiological stress responses are greater in goats with an excitable temperament compared to goats with a calm temperament.

High-Condensed Tannin Diet and Transportation Stress in Goats: Effects on Physiological Responses, Gut Microbial Counts and Meat Quality.

Feeding condensed tannin (CT)-containing diets such as sericea lespedeza (Lespedeza cuneata) and reducing stress have been reported to improve meat quality and food safety in goats. In a completely randomized design with split-plot, thirty-six uncastrated male Spanish goats were assigned to 3 dietary treatments (n = 12/treatment): ground 'Serala' sericea lespedeza hay (SER), bermudagrass (Cynodon dactylon) hay (BG), or bermudagrass hay-dewormed goats (BG-DW; Control) at 75% of intake, with a corn-based supplementation (25%) for 8 weeks. Prior to slaughter, goats were either transported for 90 min to impose stress or held in pens. Basophil counts were lower (p < 0.01) in the SER group compared to BG or BG-DW groups suggesting a better anti-inflammatory capacity due to polyphenols in the SER diet. Compared to BG-DW group, cortisol level was higher (p < 0.05) and norepinephrine was lower (p < 0.05) in the SER group. The SER group had the lowest aerobic plate counts (APC) in both rumen and rectum (p < 0.01). Longissimus dorsi muscle initial pH was not affected by diet or stress. Feeding sericea hay to goats may have beneficial effects, such as enhanced antioxidant and anti-inflammatory properties during stress and reduced gut microbial counts, without changing meat quality characteristics.

Plasma metabolomic profiles as affected by diet and stress in Spanish goats.

The effects of high-condensed tannin (CT) diet combined with preslaughter stress have not been studied at the metabolome level in goats. This study was conducted to determine the effects of feeding sericea lespedeza (SL; Lespedeza cuneata), a high-CT legume, and transportation stress on plasma metabolome in goats. Uncastrated male Spanish goats (age = 8 months; BW = 26.0 ± 0.48 kg) were either fed ground 'Serala' SL hay (SER), bermudagrass (Cynodon dactylon) hay (BG), or bermudagrass hay-dewormed goats (BG-DW; Control) at 75% of intake, with a corn-based supplementation (25%) for 8 weeks (n = 12/Diet). At the end of the trial, goats were subjected to one of two stress treatments (ST): transported for 90 min to impose stress (TS) or held in pens (NTS) before slaughtering, in two replicates. Live and carcass weights, and blood samples were collected at 0, 30, 60 and 90 min of transportation or holding time (Time). The data were analyzed using MIXED procedures in SAS and metabolomics data were analyzed using the R software. When measured after ST, SER group had the lowest body weight (P < 0.05) among the three diet groups. Carcass weights were high in the BG-DW, low in SER, and intermediate in BG group. Plasma creatine concentrations decreased over Time (P < 0.01) in the TS goats in all diet groups. Meat crude protein percentages were higher (P < 0.05) in SER (22.5 ± 0.22) and BG-DW (22.3 ± 0.22) groups compared to the BG group (21.6 ± 0.22). At the metabolome level, SER group had the lowest (P < 0.05) glycine, alanine, threonine, taurine, trans-hydroxyproline, methionine, and histidine concentrations and highest (P < 0.01) lysine and citrulline concentrations among the Diet groups. Butyric acid, concentration was higher (P < 0.05) in the SER group compared to BG group. Eight medium- and long-chained acylcarnitines were higher (P < 0.05) in the BG-DW group than SER or BG groups. In general, amino acid levels decreased and acylcarnitine increased with Time (P < 0.05) in all groups. Sericea diet can be beneficial in enhancing stress coping abilities in goats due to elevated butyrate, lysine, and citrulline levels; however, SER resulted in lower energy level in goats compared to BG or BG-DW groups. Fatty acid metabolism is the main energy pathway in all groups during prolonged stress. Inclusion of certain varieties of SL in the diet must be carefully controlled to prevent possible negative effect.

Social isolation of goats: significance of visual contact with conspecifics on behavioral and physiological responses.

Social isolation can increase distress in goats, particularly when they cannot maintain visual contact with conspecifics. This experiment was conducted to determine the behavioral and physiological responses in goats during isolation with or without visual contact with conspecifics. Male Spanish goats (uncastrated, 8 mo old, average weight 29.4 ± 0.59 kg) were randomly assigned to a control (CO) group with no isolation or to one of four isolation treatment (TRT) pens (1.5 × 1.5 m) with: 1) open grill panels but with no visual contact with conspecifics (IO), 2) covered grill to prevent visual contact (IC), 3) open grill with visual contact (IV), or 4) covered grill with a 30 × 30 cm window to allow visual contact (IW), for 90 min of social isolation (n = 12 goats per TRT). Blood samples were collected at 0, 30, 60, and 90 min (Time) from isolated and control goats. The experiment was repeated 1 wk later using the same animals, with each goat being subjected to the same isolation TRT the second time to study the effect of prior exposure to isolation. Friedman's two-way analysis of variance by Ranks test in SAS showed that the median frequency of vocalization (rank score) in goats was high in the IO group, low in the IV and IW groups, and intermediate in the IC group (P < 0.01). The vocalization rank score was also higher (P < 0.01) during the first 30 min of isolation in goats. The median frequency of visual contact was higher in the IW group than in the IV group (P < 0.01). The frequency of climbing behavior was high in the IC and IO groups, low in the IV group, and intermediate in the IW group (P < 0.01). Repeated measures analysis using general linear models procedures in SAS revealed that plasma cortisol and glucose concentrations tended (P < 0.1) to be the highest in the IO group than in CO, IC, IV, and IW groups. Cortisol levels were also higher (Time; P < 0.05) at 0 and 90 min compared with 30 and 60 min. Norepinephrine concentrations decreased (P < 0.05) with Time, and plasma nonesterified fatty acid (NEFA) levels were affected by TRT × Time interaction (P < 0.01). Overall, epinephrine, norepinephrine, glucose, and NEFA concentrations were lower (P < 0.01) and cortisol concentrations and lymphocyte counts were higher (P < 0.01) when goats were exposed to isolation the second time. The results showed that goats with no visual contact with conspecifics during social isolation had greater physiological stress responses and spent more time vocalizing or trying to escape the pen, which may indicate distress.