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Dietary C18 unsaturated fatty acids are extensively biohydrogenated by rumen microbiota into 18:0 and trans-18:1 isomers. Usually, the biohydrogenation (BH) pathways produce mostly 18:0 and trans-11 18:1 as the main intermediate. However, when fed lowforage diets that decrease the rumen pH, the main BH intermediate often is the trans-10 18:1 isomer. This change in BH, called the trans-10 shift, is associated with milk fat depression, but despite that, little is known about the triggers of the trans-10 shift and its ecological role in rumen microbiota. Microbiota stressed by low rumen pH may benefit from increased trans-18:1 availability favoring the production of the trans-18:1 isomer with a slower rate of BH. Thus, we hypothesize that at low rumen pH conditions, the BH of trans-10 18:1 will be slower than that of trans-11 18:1. We tested that with an in vitro experiment using batch incubation in Hungate tubes, replicated in 5 wk. In each run, 24 tubes were allocated at 6 treatments and 4 incubation times (0, 6, 24 and 48 h). The treatments resulted from the combination of 2 buffering solution pH (6.74 vs. 5.85) and 3 substrates: 1) Control (60 mg feed DM) 2) T10 (Control plus 1.2 mg of trans- 10 18:1) and 3) T11 (Control plus 1.2 mg of trans-11 18:1). Volatile fatty acids (VFA) and long-chain fatty acids were analyzed by gas chromatography. Data were analyzed using the 2 × 3 factorial arrangement for each incubation time. Low pH resulted in less 30 to 37% DM disappearance than those with high pH at all incubation times (P < 0.05). The decrease in VFA with low pH was less expressive (≈17%) and only evident at 24 and 48 h incubation times. The low pH also decreased (P < 0.05) the BH of 18:1n-9 (less ≈47 to 53%), 18:2n-6 (less 30 to 34%) and 18:3n-3 (less 20 to 22%) at 24h and 48h incubation times, respectively. The disappearance of trans-11 18:1 was 14.6% at 6 h, 29.0% at 24 h and 32.8% at 48 h and was not affected by pH. The disappearance of trans-10 18:1 was dependent on pH. At 6 h and high pH the disappearance of both was similar, but at low pH the disappearance of trans-10 18:1 was much less (9.8%) than that of trans-11 18:1. However, at 48 h and low pH, the disappearance was similar between isomers and the disappearance of trans-10 18:1 was much greater (46.6%) than trans- 11 18:1. In summary, the BH of trans-10 18:1 is less at low pH than at high pH, and this could in part explain its abundance in some practical ruminant feeding conditions. Acknowledgements: Fundação para a Ciência e a Tecnologia (FCT) funding through PTDC/ CAL-ZOO/29654/2017(RumOmics), PTDC/CALZOO/ 4515/2021(Gene2Rumen), UIDB/00276/2020 (CIISA), and LA/P/0059/2020 (AL4Animals) projects. [ABSTRACT FROM AUTHOR]

The trans-10 shift is a recognized deviation from the normal ruminal biohydrogenation (BH) pathways of the dietary polyunsaturated fatty acids (FA) in the rumen. This phenomenon leads to the formation of the harmful t10-18:1, which is detrimental to the health of the animal, instead of the beneficial t11-18:1, which has positive effects. The phenomenon known as the t10-shift has been observed in animals consuming diets that are high in starch but low in forage. Although consuming such similar diets, there exists a significant variability in t10-shift expression among animals. The reasons be- hind the t10-shift are not well understood. With the aim of finding biomarkers associated with t10-shift, this study utilized multiple levels of information, including lipidomics, metabolomics, and proteomics of ruminal contents. Rumen contents from male lambs (n = 16) distributed by two diets with distinct forage: concentrate ratios (20:80 and 80:20) were collected after 42 d on ad libitum feed. FA from rumen contents were prepared and analyzed by gas chromatography, metaproteomics analyzes were performed from rumen bacterial pellets using LC-MS/MS-based label-free quantification, and metabolomics analyzes were performed from rumen fluid supernatants using IMS-TOF technology. Multivariate analyses and correlations were performed using the Metaboanalyst R package. Several fatty acids, proteins, and metabolites showed a strong correlation with the proportion of t10-18:1 in rumen contents. Among fatty acids, 17:0, 14:0, a-15:0, and t15-18:1 were positively correlated, while t11-18:1, t11,c13-CLA and c9,t11-CLA showed negative correlation. The metabolites, thiamine aldehyde, mevinolinic acid, 2-hydroxy-2H-benzo[h] chromene-2-carboxylate, or biotin were positively correlated with t10-18:1, as well as proteins associated with gluconeogenesis, stress response or amino acid metabolism. This work provides, for the first time, a multi-level integrated view of t10-shift phenomena in sheep. Fundação para a Ciência e a Tecnologia (FCT) provided financial support through the research pro- jects PTDC/CAL-ZOO/29654/2017 (RumOmics), PTDC/CAL-ZOO/4515/2021 (Gene2Rumen), UIDB/00276/2020 (CIISA), UIDB/04129/2020 (LEAF), and LA/P/0059/2020 (AL4Animals) [ABSTRACT FROM AUTHOR]

This work evaluated the effects of dietary inclusion of microalga Chlorella vulgaris on post-weaning piglet growth performance and physiology. Noncastrated male piglets [n = 72; Piétrain × (Large White x Landrace), with 7.4±1.03 kg body weight (BW); 28 d] were blocked by BW and randomly assigned to 24 pens (3 animals/pen, n = 8) and each pen to one of three experimental diets, fed ad libitum: (C) Control diet (wheat/corn/soybean meal-based), (CV2.5) 2.5% C. vulgaris inclusion and (CV5) 5% C. vulgaris inclusion. The trial lasted 5 wk, with BW and feed intake determined weekly. Total tract apparent digestibility (TTAD) was determined using chromic oxide as external marker. Following wk 5 (67 d), one piglet per pen (median BW) was slaughtered and visceral organs, gastrointestinal contents and Longissimus lumborum muscle were sampled. Untargeted 1H NMR metabolomics study characterized hepatic polar metabolome following CHCl3:MetOH:H2O extraction. Multivariate analysis (PCA, PLS-DA) and identification of important metabolites enabled pathway analysis. Intramuscular fatty acid composition was determined by gas-liquid chromatography after direct transesterification to methylesters and extraction. Growth performance indicators, TTAD, digesta viscosity and pH were determined and analyzed by ANOVA with values reported as average ± SEM and significance at P ≤ 0.05. Diet did not affect average daily BW gain (566 ± 1.6 g/d), average daily feed intake (711 ± 1.8 g/d), nor feed conversion ratio (1.28 ± 0.078). Viscosity of intestinal contents in upper small intestine (3.7 ± 0.19 mPa.s) and ileum (4.8 ± 0.26 mPa.s), as well as pH of stomach (4.1 ± 0.18), upper small intestine (5.7 ± 0.08), ileum (6.6 ± 0.07), cecum (5.7 ± 0.10) and colon contents (6.1 ± 0.08) were unaffected by diets, within each sampling site. Minor negative effects of microalgae inclusion were seen on digestibility of all feed fractions (1 to 9% difference, compared with Control), namely DM, OM, CP, CF, Ash, NDF and DE. In the muscle, animals fed 5% C. vulgaris presented greater concentrations of omega-3 polyunsaturated fatty acids (PUFA, 1.68 ± 0.112 % vs. 1.39 % of total FA) and no effect in omega-6 PUFA concentration, thus a greater ratio of Omega 3 to Omega 6 (0.052 ± 0.022 vs. 0.045), when compared with Control. Multivariate analysis of liver metabolomics showed significant clustering, clearly separating both microalgaefed groups from Control. The altered metabolic profile suggests activation of antioxidant pathways, as well as carbohydrate and amino acid metabolism with dietary inclusion of C. vulgaris. Particularly, levels of antioxidant metabolic markers as glutathione, sarcosine, betaine and their precursor amino acids point to an enhanced GSH-antioxidant system activation. Overall, results indicate that C. vulgaris inclusion in postweaning diets up to 5% does not penalize growth performance and can effectively be used in piglet nutrition. Furthermore, hepatic metabolomics suggest important bioactivity imparted by C. vulgaris dietary inclusion, potentially linking enhanced antioxidant systems in the liver with stable integration of PUFA in muscle, which may improve piglet health and pork quality. [ABSTRACT FROM AUTHOR]

Protected Designation of Origin (PDO) products are gaining importance due to their high quality and increased consumer concerns about sustainability. Arouquesa PDO beef is a local product with valued organoleptic traits that contributes to local heritage and economy in rural regions of Northern Portugal. Proteomic analysis provides insights on important metabolic pathways that can be related to the production system and be used to differentiate PDO beef from beef produced under standard systems such as those used with crossbreds. The aim of this work is to compare the proteome profile of Arouquesa PDO beef with beef from crossbred animals produced under standard conditions. Muscle samples (n = 10) were collected from the abattoir from male weaners slaughtered at 8.5 mo of age: five Arouquesa breed and five Limousine (n = 2) or Charolais (n = 3) crossbreds. Samples were analyzed using 2D-DIGE and spots showing differential accumulation (P-value < 0.05 and fold-change ± 1.5) were selected for identification using in-gel tryptic digest and LC-MS/MS identification. Meat quality parameters (pH, crude protein and total lipids) were also determined. Animal body weight was 62% heavier (P < 0.05) in crossbred animals (214 ± 5.47 kg). Arouquesa beef had reduced crude protein (17.52 ± 1.39 vs. 20.46 ± 1.44 g/100 g meat) and consequently a greater value of total lipids (8.39 ± 1.86 vs. 3.62 ± 2.44 g/100 g meat). Of the 34 protein spots showing differential accumulation, 13 had greater abundances in Arouquesa beef, whereas 21 were increased in the crossbred animals. Some of the up-regulated proteins in the Arouquesa beef are involved in glycolysis (GADPH, GPI, PGM1, TPI, ENO3) or in muscle structure (MYL1, MYLPF). The down-regulated proteins were mostly related to muscle structure (MYLPF, MYL6B, TNNT1), oxygen transport (HBA, HBB) or heat shock proteins (HSPB1). We also found for some proteins several differentially regulated proteoforms between breeds (MYLPF and TPI1), not all regulated in the same way. The proteins involved in glycolysis can indicate a decreased glycolytic potential of Arouquesa muscle leading to reduced glycogen degradation during post-mortem. The proteins related to muscle structure are generally associated with greater growth performances which was not expected as Arouquesa animals had lighter final body weights. However, MYL1 is also associated to increased tenderness as expectable in Arouquesa beef due to the production system based on early slaughter. The greater abundance of proteins related to oxygen metabolism in crossbred animals suggests a greater oxidative capacity which may be related to muscle fiber composition. In conclusion, the proteome profile of Arouquesa breed points to better beef quality traits compared with commercial beef; thus, contributing to increase product valorization. We found different proteins in the two groups related to specific metabolisms showing a breed-specific response allowing the improvement of the PDO products traceability. [ABSTRACT FROM AUTHOR]