Branched-chain fatty acids (BCFA) have recently been reported to play a role in human gut health during early life. However, little information is available on the fecal BCFA profiles in young ruminants and whether they are associated with the development of neonatal calf diarrhea. The objectives of this study were to (1) characterize BCFA profiles in feces collected from young calves, (2) compare the fecal BCFA composition between diarrheic and nondiarrheic dairy calves, and (3) explore the potential relationships between BCFA and microbiota in the feces. A total of 32 male Holstein dairy calves (13 ± 3 d old) with the same diet management were grouped as diarrheic (n = 16) or healthy (n = 16) based on fecal score (determined by liquid fecal consistency with some solid particles); diarrhea cases were defined as fecal score ≥2 for at least 2 d. Fecal samples were collected on the seventh day after calf arrival, and the fecal BCFA and microbial profiles were assessed using gas chromatograph and amplicon sequencing, respectively. In total, 7 BCFA were detected in the feces of all dairy calves; however, the concentrations of fecal BCFA differed between diarrheic and nondiarrheic calves. Specifically, the concentrations of iso-C16:0, iso-C17:0, anteiso-C17:0, and total even-chain BCFA were significantly higher in the feces of diarrheic calves. When the associations between BCFA and bacteria were studied, the relative abundance of Eggerthella was positively correlated with the concentrations of iso-C16:0 (ρ = 0.67), iso-17:0 (ρ = 0.77), anteiso-C17:0 (ρ = 0.73), and iso-C18:0 (ρ = 0.65), whereas the relative abundance of Subdoligranulum was positively correlated with the concentrations of iso-C14:0 (ρ = 0.62), iso-C15:0 (ρ = 0.78), and anteiso-C15:0 (ρ = 0.63). Use of random forest algorithm showed that BCFA such as anteiso-C15:0, iso-C16:0, iso-C17:0, iso-C18:0, and total even-chain BCFA could be used as biomarkers to differentiate diarrheic calves from healthy ones. Our findings generated fundamental knowledge on the potential roles of BCFA in neonatal calf gut health. Follow-up studies with larger animal populations are warranted to validate the feasibility of using BCFA as indicators of health status in neonatal calves.

The objective of this study was to assess the impact of Lactobacillus plantarum, Bacillus subtilis, and a lignocellulolytic enzyme system on the nutritional value, fermentation profiles, rumen digestion, and bacterial community of fresh waxy corn stalk silage. Fresh waxy corn stalks harvested after 90 days of growth were treated with no additives (CON), compound multiple lactobacilli (ML, comprising Lactobacillus plantarum at 1.0 × 10⁵ cfu/g fresh weight and Bacillus subtilis at 1.0 × 10⁵ cfu/g fresh weight), a lignocellulolytic enzyme system (LE, at 500 g/t), and a combination of the lignocellulolytic enzyme system and multiple lactobacilli (MLLE). Throughout the fermentation process, the contents of dry matter (DM) and water-soluble carbohydrates (WSC), as well as the counts of lactic acid bacteria, yeast, and mold, continuously decreased with the extension of the fermentation time. In contrast, the concentrations of acetic acid, propionic acid, and ammonia N progressively increased over time. Notably, the pH initially decreased and then increased as fermentation progressed, while the lactic acid concentration first increased and then decreased with the extension of the fermentation time. After 45 days of fermentation, the DM contents of LE and MLLE were significantly lower than that of CON, while the WSC content of MLLE was significantly higher than that of CON. LE most effectively increased the ED of DM and neutral detergent fiber (NDF). Compared to ML and LE, MLLE showed a greater increase in the “a” value of DM and NDF. The DM content in LE and MLLE was lower than in CON, with a higher WSC content. Both ML and MLLE produced more lactic acid than CON and LE, resulting in a lower pH in these groups. Additionally, the acetic acid content and ammonia N content were lower in ML and MLLE compared to CON and LE. The counts of lactic acid bacteria, yeast, and mold declined over the fermentation period, dropping below detectable levels on the seventh and fifth days, respectively. Relative to CON, the three additives reduced the relative abundance of Weissella and Klebsiella and increased that of Lactobacillus. In conclusion, LE improved the nutritional value and rumen digestion of the silage, ML enhanced its fermentation profiles and bacterial community, and MLLE significantly improved the nutritional value, rumen digestion characteristics, fermentation profiles, and bacterial community of the fresh waxy corn stalk silage.

The objectives of this study were to evaluate changes in profiles of odd- and branched-chain fatty acids (OBCFA), including pentadecanoic acid (C15:0), 13-methyltetradecanoic acid (iso-C15:0), 12-methyltetradecanoic acid (anteiso-C15:0), 14-methylpentadecanoic acid (iso-C16:0), heptadecanoic acid (C17:0), 15-methylhexadecanoic acid (iso-C17:0), and 14-methylhexadecanoic acid (anteiso-C17:0) during in vitro fermentation of pure carbohydrates mixtures in the buffer-rumen fluid. The second objective was to correlate the changes in the OBCFA profile to the corresponding changes in ruminal fermentation parameters, microbial crude protein (MCP) synthesis, and bacterial populations. Five pure carbohydrates mixtures containing different cellulose: starch (C:S) ratios, i.e., 0:100, 25:75, 50:50, 75:25, and 100:0, were incubated for 6, 12, 18, and 24 h in vitro. The results showed that there was significant interaction (P < 0.05) between C:S and incubation time for changes in all OBCFA profiles, except iso-C17:0. The highest concentration of total OBCFA (3.94 mg/g dry matter; DM) was observed in the residues after 24 h of fermentation when the C:S was 0:100, while the lowest concentration of OBCFA (1.65 mg/g DM) was produced after 6 h of incubation when the C:S was 50:50. The correlation analysis revealed that the concentration of iso-C16:0 might be a potential marker for the estimation of total volatile fatty acids (ρ = 0.78) and MCP synthesis (ρ = 0.82) in the rumen. Compared to starch degrading bacteria, cellulolytic bacteria had stronger correlations with OBCFA concentrations, and the strongest correlation was found between the population of Ruminococcus flavefaciens with C15:0 concentration (ρ = 0.70). Notably, this is the first paper reporting relationship between OBCFA with rumen fermentation products and microbial protein synthesis based on fermentation of pure carbohydrates mixtures in vitro, and thus avoid confounding interference from dietary protein and fat presence in the in vivo studies. However, more in-depth experiments are needed to substantiate the current findings.

Although odd-chain fatty acids (OCFA) and branched-chain fatty acids (BCFA) are found in bovine milk and have some positive influences on human and animal health, their concentrations in bovine colostrum and transition milk have not been reported. In this study, we reported the OCFA and BCFA concentrations in colostrum and transition milk and their stability after heating or freezing treatments (or both), which are processes commonly applied in dairy calf management. Milk samples were collected from 12 Holstein dairy cows (6 primiparous and 6 multiparous) at the first milking (colostrum), fifth milking (transition milk), and ninth milking (mature milk) after calving, respectively, and were used for fatty acid analysis using gas chromatography. The sum concentration of OCFA and BCFA (termed OBCFA) was 134 mg/100 g of milk in the colostrum, which was 24% and 35% lower than that in the transition milk and mature milk, respectively. Among these fatty acids detected, C15:0 and C17:0 were the top 2 abundant fatty acids in all milk types, accounting for 20 to 25% and 21 to 24% of the total concentration of OBCFA, respectively. Additionally, anteiso-C17:0 was the most abundant BCFA, followed by iso-C17:0, anteiso-C15:0, iso-C16:0, iso-C15:0, iso-C18:0, and iso-C14:0 in 3 types of milk. Significant interactions between milk type and cow parity were observed for all OCFA and BCFA concentrations. The milk samples were also treated with heating (at 65°C for 60 min), freezing (at -20°C for 30 d), and heating and freezing (at 65°C for 60 min and then at -20°C for 30 d), and milk OCFA and BCFA concentrations were similar between these treatments. In conclusion, the OBCFA concentration was lower in colostrum, compared with transition and mature milks, and it remained stable after heating and freezing treatments.

This study examined branched-chain fatty acids (BCFAs)’ effects on oxidative stress, energy metabolism, inflammation, tight junction disruption, apoptosis, and Toll-like receptor 4/nuclear factor kappa-B (TLR4/NF-κB) signaling in lipopolysaccharide (LPS)-induced calf small intestinal epithelial cells (CSIECs). Eight groups were used: a control group, an LPS-induced group, and six BCFA treatment groups (12-methyltridecanoic acid (iso-C14:0), 13-methyltetradecanoic acid (iso-C15:0), 14-methylpentadecanoic acid (iso-C16:0), 15-methylhexadecanoic acid (iso-C17:0), 12-methyltetradecanoic acid (anteiso-C15:0), and 14-methylhexadecanoic acid (anteiso-C17:0)) with LPS. The BCFA pretreatments significantly increased CSIEC activity compared to the LPS-induced group, with iso-C14:0 showing the highest activity (89.73%). BCFA reduced Reactive Oxygen Species (ROS) generation and malondialdehyde (MDA) levels and improved the superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) activities and glutathione (GSH) levels. Iso-C16:0 optimized total antioxidant capacity (T-AOC). BCFA enhanced the mitochondrial membrane potential, Adenosine Triphosphate (ATP) enzyme activity, and ATP content, with iso-C14:0 increasing ATP by 27.01%. BCFA downregulated interleukin (IL)-1β, IL-8, tumor necrosis factor (TNF)-α, and interferon (INF)-γ gene expression, reduced IL-6 levels, and increased IL-10 expression. Myeloid differentiation factor 88 (MyD88) mRNA levels were reduced. BCFA alleviated Zonula Occludin (ZO-1), Claudin-1, and Claudin-4 decrease and increased Occludin levels. BCFA mitigated LPS-induced increases in Caspase-3 and BCL2-Associated X (BAX) mRNA levels, reduced Caspase-8 and Caspase-9 expression, and increased B-Cell Lymphoma-2 (BCL-2) mRNA levels. The Entropy Weight-TOPSIS method was adopted, and it was discovered that iso-C15:0 has the best effect. In summary, BCFA supplementation mitigated oxidative stress and enhanced mitochondrial function. BCFA inhibited TLR4/NF-κB signaling pathway overactivation, regulated inflammatory cytokine gene expression, reduced cellular apoptosis, preserved tight junction integrity, and supported barrier function.