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Abstract Background The microbiota and metabolites in the gastrointestinal tracts of female animals at different reproductive periods are very important to the growth, development, and health of themselves and their offspring. However, the changes in the gastrointestinal microbiota and metabolites throughout reproductive period of different sheep breeds and their effects on the growth and development of offspring lambs are still unclear. Hence, this study presents an assessment of the reproductive hormone levels, immune levels, rumen microbiota, and metabolites in Hu sheep and Suffolk ewes at different reproductive periods and their effects on the growth and development of offspring lambs. Results Hu sheep and Suffolk during non-pregnancy, pregnancy, and lactation were used as the research objects to determine reproductive and immune indexes of ewes at different periods, analyze rumen microbiome and metabolome, and track the growth performance and development of offspring lambs. The results showed that the reproductive hormone and immune levels of Hu sheep and Suffolk underwent adaptive changes across different reproductive periods. Compared with non-pregnancy, the microbial energy metabolism and lipid metabolism function decreased during Hu sheep pregnancy, and energy metabolism function decreased during lactation. In Suffolk, energy metabolism, glycan biosynthesis, and metabolism function were enhanced during pregnancy, and the metabolism of cofactors and vitamins was enhanced during lactation. Prevotella increased in Suffolk during pregnancy and lactation (P < 0.05) and was positively correlated with the birth weight and body size of the lambs (P < 0.05). Moreover, the abundances of Butyrivibrio and Rikenellaceae_RC9_gut_group during pregnancy were positively correlated with the intestinal immunity of the offspring lambs (P < 0.05), thereby regulating the intestinal immunity level of the lambs. Metabolomic analysis revealed that the protein digestion, absorption, and amino acid metabolism of Hu sheep were enhanced during pregnancy, which provided amino acids for the growth and development of pregnant ewes and fetuses and was significantly correlated with the birth weight, body size, and intestinal immunity of lambs (P < 0.05). Simultaneously, there was an increase in acetate and propionate during the pregnancy and lactation period of both Hu sheep and Suffolk, providing energy for ewes during reproductive period. Moreover, the microbiota during the lactation period was significantly correlated with the milk quality and lambs daily gain (P < 0.05). Conclusions This study revealed the characteristic succession changes in the rumen microbiota and its metabolites at different reproductive periods in sheep breeds and their regulation of reproductive hormone and immune levels and identified their potential effects on the growth and development of offspring lambs. The findings provide valuable insights into the health and feeding management of different sheep breeds during the reproductive stage. Video Abstract

IntroductionResveratrol (Res), a natural plant antitoxin polyphenol, is widely used in animal husbandry due to its antioxidant and anti-inflammatory properties, and current research has focused on humans, sows, and female mice. This study aimed to analyze the effects of dietary Res supplementation in ewes on antioxidant activity, immune responses, hormone levels, rumen microbiota and metabolites across various reproductive stages (estrus, pregnancy, and lactation).MethodsTwenty-four healthy ewe lambs (Hu sheep, 2 months old) with a similar body weight (BW) (mean: 21.79 ± 2.09 kg) were selected and randomly divided into two groups: the control group (Con) and the Res group (Res). The Res group received 10 mg/kg Res (based on BW) in addition to their basal diet.ResultsRes increased the levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) in ewes at sexual maturity (p < 0.05). Additionally, Res supplementation induced significant increases in serum glutathione peroxidase (GSH-Px), IgG, FSH, and LH levels during estrus (p < 0.05); serum IgA, IgG and IgM during pregnancy and lactation (p < 0.05); and serum LH, glucose, GSH-Px, and catalase (CAT) levels during lactation (p < 0.05). Meanwhile, serum interleukin 1β (IL-1β) (p =0.005) and cholesterol levels (p = 0.041) during the lactation stage decreased following Res supplementation. Notably, colostrum IgA, IgG, and fat concentrations were significantly higher in the Res group than in the Con group (p < 0.05). Moreover, Res altered the rumen microbiota in ewes. Specifically, the relative abundance of Prevotella (p < 0.05) during pregnancy and Rikenellaceae_RC9_gut_group (p < 0.001) during lactation were significantly increased in ewes under Res treatment. The abundance of Rikenellaceae_RC9_gut_group was positively correlated with the levels of Ig A, Ig M, E2, FSH, LH, GSH-PX, and CAT. Additionally, Res altered the activity of metabolic pathways such as progesterone-mediated oocyte maturation, the estrogen signaling pathway, ovarian steroidogenesis, and the AMPK signaling pathway, and the levels of AICAR and 2-hydroxyestradiol metabolites, both during pregnancy and lactation.DiscussionThere findings show that Res can improve health, antioxidant status, and immune activity throughout the reproductive cycle in ewes by regulating rumen microorganisms and metabolites.

The rumen microbiota—a symbiont to its host and consists of critical functional substances—plays a vital role in the animal body and represents a new perspective in the study of adaptive evolution in animals. This study used Slide Viewer slicing analysis system, gas chromatography, RT-qPCR and other technologies, as well as 16S and metabolomics determination methods, to measure and analyze the microstructure of rumen epithelium, rumen fermentation parameters, rumen transport genes, rumen microbiota and metabolites in Tibetan sheep and Hu sheep. The results indicate that the rumen nipple height and cuticle thickness of Tibetan sheep are significantly greater than those of Hu sheep (p

Microbiota density plays an important role in maintaining host metabolism, immune function, and health, and age has a specific effect on the composition of intestinal microbiota. Therefore, the age-specific effects of age differences on the structure and function of the ileum microbiota in Tibetan sheep were investigated by determining the density of the ileum microbiota, the content of VFAs, and the expression levels of their transporter-related genes at different ages. The results showed that the contents of acetic acid and propionic acid in the ileum of Tibetan sheep in the 1.5-year-old group were significantly higher (p < 0.05) than those in other age groups, and that the contents of total VFAs were also significantly higher (p < 0.05) than those in other age groups. The relative densities of ileum Rf, Ra, and Fs were significantly higher in the 1.5-year-old group than in the other age groups (p < 0.05). The ileum epithelial VFAs transport-related genes AE2, MCT-4, and NHE1 had the highest expression in the 1.5-year-old group, and the expression of DRA was significantly lower in the 1.5-year-old group than in the 6-year-old group (p < 0.05). Correlation analysis showed that Cb, Sr, and Tb were significantly positively correlated with butyric acid concentration (p < 0.05) and negatively correlated with acetic acid, but the difference was not significant (p > 0.05); MCT-1, MCT-4, and AE2 were significantly positively correlated (p < 0.05) with acetic, propionic, and isobutyric acid concentrations; NHE1, NHE2, and MCT-4 were highly significantly positively correlated (p < 0.01) with Romboutsia and unclassified_Peptostreptococcaceae, while acetic acid was significantly positively correlated (p < 0.05) with NK4A214_group; Romboutsia, and unclassified_Peptostreptococcaceae were significantly positively correlated (p < 0.05). Therefore, compared with other ages, the 1.5-year-old Tibetan sheep had a stronger fermentation and metabolic capacity in the ileum under traditional grazing conditions on the plateau, which could provide more energy for Tibetan sheep during plateau acclimatization.

The experiment was conducted to investigate the potential effects of steviol glycosides on growth performance, rumen fermentation processes, and microbial diversity in Hu sheep. A single-factor design was used for the trial. Twenty healthy weaned Hu lambs, possessing comparable body weights averaging 18.31 ± 1.24 kg, were randomly allocated into two distinct groups: the control group (CON) and the experimental group (STE), with each comprising 10 lambs. The CON was fed the basal diet, and the STE was supplemented with 0.07% steviol glycosides based on the basal diet. During the experimental period, variations in body weight and feed intake were closely monitored and recorded. After feeding for 90 d, blood was collected to determine blood biochemical indices, and rumen fluid samples were gathered for an in-depth analysis of rumen fermentation parameters and microbial diversity. The outcomes revealed no statistically significant differences in growth performance or serum biochemical indices between the two groups (p > 0.05). Rumen pH in STE and CON was within the normal range. The rumen ammonia nitrogen (NH3-N) and acetic acid (AA) content of STE decreased significantly compared with CON (p < 0.05). No significant variations were observed in the levels of other volatile fatty acids (VFAs) between the two groups (p > 0.05). The rumen microbial OTUs count, as well as the Shannon, Simpson, Chao1, and Ace indices, were notably lower in the STE group compared to the CON group (p < 0.05). Additionally, at the phylum level, the relative abundance of Firmicutes, Bacteroidetes, and Proteobacteria collectively accounted for over 97% of the total phylum composition. In comparison to the CON group, the STE group exhibited an increase in the relative abundance of Proteobacteria (p < 0.05), accompanied by a significant reduction in the relative abundance of Patescibacteria and Desulfobacteria (p < 0.05). At the genus level, there was a notable increase in the relative abundance of Prevotella_7 and Succinivibrionaceae_UCG_001 in the STE group, whereas the relative abundance of Rikenellaceae_RC9_gut_group significantly decreased (p < 0.05). According to the correlation analysis between rumen microflora and VFAs, the relative abundance of Succinivibrionaceae_UCG_001 displayed a significant negative correlation with AA (p < 0.05), whereas Lactobacillus exhibited a notable positive correlation with isobutyric acid (IBA) (p < 0.05). In summary, steviol glycosides had no significant effect on the production performance and blood biochemical indexes of Hu sheep. Steviol glycosides can improve rumen fermentation parameters and rumen microflora structure of Hu sheep and have a certain effect on rumen microbial diversity and composition.

During the adaptive evolution of animals, the host and its gut microbiota co-adapt to different elevations. Currently, there are few reports on the rumen microbiota–hepato-intestinal axis of Tibetan sheep at different altitudes. Therefore, the purpose of this study was to explore the regulatory effect of rumen microorganism–volatile fatty acids (VFAs)–VFAs transporter gene interactions on the key enzymes and genes related to gluconeogenesis in Tibetan sheep. The rumen fermentation parameters, rumen microbial densities, liver gluconeogenesis activity and related genes were determined and analyzed using gas chromatography, RT-qPCR and other research methods. Correlation analysis revealed a reciprocal relationship among rumen microflora–VFAs-hepatic gluconeogenesis in Tibetan sheep at different altitudes. Among the microbiota, Ruminococcus flavefaciens (R. flavefaciens), Ruminococcus albus (R. albus), Fibrobactersuccinogenes and Ruminobacter amylophilus (R. amylophilus) were significantly correlated with propionic acid (p < 0.05), while propionic acid was significantly correlated with the transport genes monocarboxylate transporter 4 (MCT4) and anion exchanger 2 (AE2) (p < 0.05). Propionic acid was significantly correlated with key enzymes such as pyruvate carboxylase, phosphoenolpyruvic acid carboxylase and glucose (Glu) in the gluconeogenesis pathway (p < 0.05). Additionally, the expressions of these genes were significantly correlated with those of the related genes, namely, forkhead box protein O1 (FOXO1) and mitochondrial phosphoenolpyruvate carboxykinase 2 (PCK2) (p < 0.05). The results showed that rumen microbiota densities differed at different altitudes, and the metabolically produced VFA contents differed, which led to adaptive changes in the key enzyme activities of gluconeogenesis and the expressions of related genes.

The rumen microbiota and metabolites play an important role in energy metabolism and immune regulation of the host. However, the regulatory mechanism of rumen microbiota and metabolite interactions with host on Tibetan sheep’s plateau adaptability is still unclear. We analyzed the ruminal microbiome and metabolome, host transcriptome and serum metabolome characteristics of Tibetan sheep at different ages. Biomarkers Butyrivibrio, Lachnospiraceae_XPB1014_group, Prevotella, and Rikenellaceae_RC9_gut_group were found in 4 months, 1.5 years, 3.5 years, and 6 years Tibetan sheep, respectively. The rumen microbial metabolites were mainly enriched in galactose metabolism, unsaturated fatty acid biosynthesis and fatty acid degradation pathways, and had significant correlation with microbiota. These metabolites further interact with mRNA, and are co-enriched in arginine and proline metabolism, metabolism of xenobiotics by cytochrome P450, propanoate metabolism, starch and sucrose metabolism, gap junction pathway. Meanwhile, serum metabolites also have a similar function, such as chemical carcinogenesis − reactive oxygen species, limonene and pinene degradation, and cutin, suberine and wax biosynthesis, thus participating in the regulation of the body’s immune and energy-related metabolic processes. This study systematically revealed that rumen microbiota, metabolites, mRNA and serum metabolites of Tibetan sheep were involved in the regulation of fermentation metabolic function and immune level of Tibetan sheep at different ages, which provided a new perspective for plateau adaptability research of Tibetan sheep at different ages.

Since the banning of antibiotics, the use of feed additives to improve meat quality to satisfy people's pursuit of high quality has become a research hotspot. Yeast culture (YC) is rich in proteins, mannan oligosaccharides, peptides, and yeast cell metabolites, etc., and its use as a feed additive has a positive impact on improving meat quality. So the study aimed to provide a theoretical basis for YC improving mutton flavor and quality by detecting and analyzing the effects of YC on muscle physicochemical properties, amino acids, fatty acids, flavor composition, expression of related genes, and rumen microbiota of lambs. A total of 20 crossbred F1 weaned lambs (Australian white sheep♂ × Hu sheep♀; average 23.38 ± 1.17 kg) were randomly assigned to 2 groups, the control group (CON) and the 1.0% YC supplemented group (YC) (n = 10), and were reared in separate pens. The experiment had a pre-feeding period of 10 d and a treatment period of 60 d. After the experiment, 6 lambs in each group were randomly selected for slaughtering. The results showed that dietary YC supplementation increased rumen total VFA and acetate concentrations (p

ABSTRACT Different reproductive stages of mammals involve complex biological processes, and the intestinal microbiota, as an endocrine organ or an “invisible organ,” is involved in the regulation of hormone levels, immune function, and metabolism. However, the effects of the rumen microbiota, its metabolites, and blood metabolites on the reproductive performance of ruminants remain unclear. This study revealed that the Prevotella abundance increased significantly during pregnancy (P < 0.01); the Fibrobacter abundance increased significantly during lactation (P < 0.05); and rumen microbial carbohydrate metabolism, glucose biosynthesis, and metabolic functions were significantly enriched during pregnancy (P < 0.05). Microbial metabolic profile analysis showed that the differentially abundant microbial metabolites during pregnancy and lactation were mainly enriched in the biosynthesis of ubiquinone and other terpenoid quinones, and there was a certain correlation with the microbiota. Among them, sapindoside A was increased during pregnancy, nicotinamide riboside and β-cryptoxanthin were reduced during pregnancy, and L-tryptophan was significantly increased during lactation. In addition, the volatile fatty acid levels in lactation were significantly higher than those in non-pregnancy and pregnancy (P < 0.05), and the NH3-N content during pregnancy was significantly higher than that during lactation and non-pregnancy (P < 0.05). Moreover, there were differences in the serum metabolite levels at different reproductive stages, and similar metabolites existed when comparing the rumen metabolites, which were mainly enriched in arachidonic acid metabolism, vitamin B6 metabolism, and ABC transporter protein, resulting in significantly higher serum IgA and IgM levels during lactation than during non-pregnancy and pregnancy (P < 0.05). IMPORTANCE Our study illustrates the succession of the rumen microbiota and its metabolites in Small-tailed Han sheep at different reproductive stages. Among them, Firmicutes and Prevotella, which are related to energy metabolism, increased in abundance during pregnancy, while Fibrobacter, a fiber-degrading bacterium, increased in abundance during lactation. At the same time, the microbial metabolic profile and serum metabolic profile characteristics of different reproductive stages were revealed, and some functional pathways and metabolites related to energy and immunity were found. This study provides a reference for the health management of ruminants during non-pregnancy, pregnancy, and lactation.

The gut microbiota is highly symbiotic with the host, and the microbiota and its metabolites are essential for regulating host health and physiological functions. Astragalus, as a feed additive, can improve animal immunity. However, the effects of Astragalus root powder on the rumen microbiota and their metabolites in lambs are not apparent. In this study, thirty healthy Hu sheep lambs with similar body weights (17.42 ± 2.02 kg) were randomly selected for the feeding experiment. Lambs were fed diets supplemented with 0.3% Astragalus root powder, and the rumen microbiota density and metabolome were measured to determine the effects of Astragalus on the health of lambs in the rumen. The results showed that the relative abundance of Butyrivibrio fibrisolvens (Bf), Ruminococcus flavefaciens (Rf), Succiniclasticum (Su), and Prevotella (Pr) in the rumen was increased in the Astragalus group (p < 0.01), and metabolic profiling showed that the metabolites, such as L-lyrosine and L-leucine, were upregulated in the Astragalus group (p < 0.01). KEGG functional annotation revealed that upregulated metabolites were mainly enriched in the pathways of amino acid metabolism, lipid metabolism, fatty acid biosynthesis, and bile secretion in the Astragalus group, and downregulated metabolites were enriched in the pathways of methane metabolism and other pathways. Correlation analysis revealed that butyric acid was positively correlated with Roseburia and Blautia (p < 0.05) and negatively correlated with Desulfovibrio (p < 0.05). Thus, by analyzing the interactions of Astragalus root powder with the density of rumen microorganisms and their metabolites in lambs, it was shown that Astragalus root powder could improve the structure of rumen microbiota and their metabolites and then participate in the regulation of amino acid metabolism, lipid metabolism, immune metabolism, and other pathways to improve the efficiency of energy absorption of the lambs.

The effects of isochlorogenic acid (ICGA) on ewes rumen environment, microbial diversity, and immunity at different physiological stages (estrus, pregnancy and lactation) were studied in this experiment. Twenty healthy female Hu lambs of 1.5 months with similar body weight (17.82 ± 0.98 kg) and body condition were selected and randomly divided into two groups: the control group (CON) and the ICGA group (ICGA). The lambs of CON were fed a basal diet, while the lambs of ICGA were supplemented with 0.1% ICGA based on the basal diet. Lambs rumen fermentation characteristics, microbial diversity and immunity at estrus, pregnancy, and lactation stages were determined and analyzed, respectively. The results showed that the rumen pH in CON increased first and then decreased as lambs grew (p < 0.05). However, it showed the opposite change in ICGA. The content of ammonia nitrogen (NH3-N) showed the highest at estrus stage in both groups, but it was significantly higher in ICGA than that in CON (p < 0.05). The Acetic acid/propionic acid (A/P) ratio at estrus stage and the volatile fatty acids (VFAs) at pregnancy stage in ICGA were significantly higher than those of the CON (p < 0.05). The 16S rDNA sequencing analysis showed that the Shannon, Chao 1 and ACE indexes of the ICGA were significantly higher than those of the CON both at estrus and lactation stages (p < 0.05), while they showed higher at the pregnancy stage in CON (p > 0.05). Principal component analysis (PCA) showed that there were significant differences in rumen microorganism structure between CON and ICGA at all physiological stages (p < 0.01). At the phylum level, compared with the CON, Firmicutes relative abundance of three physiological stages decreased (p > 0.05) while Bacteroidota increased (p > 0.05). The relative abundance of Synergistota at estrus stage and Patescibacteria at the lactation stage increased significantly too (p < 0.05). At the genus level, compared with the CON, the relative abundance of Prevotella at three stages showed the highest (p > 0.05), while the relative abundance of Succiniclasticum, unclassified_Selenomonadaceae and Rikenellaceae_RC9_gut_group showed different abundances at different physiological stages in ICGA. Compared with the CON, the lambs of the ICGA showed higher blood IgG, IgM, and TNF- α contents at three physiological stages and higher IL-6 contents at pregnancy stage (p < 0.05). Conclusion: Adding ICGA could regulate ewes rumen fermentation mode at different physiological stages by increasing rumen NH3-N at estrus, VFAs at pregnancy, and the ratio of A/P at lactation. It optimizes rumen microbial flora of different physiological stages by increasing Bacteroidota relative abundance while reducing Firmicutes relative abundance, maintaining rumen microbial homeostasis at pregnant stage, increasing the number of beneficial bacteria in later lactating and ewes blood immunoglobulins content at three physiological stages.

The intestinal microbiota of ruminants is an important factor affecting animal production and health. Research on the association mechanism between the intestinal microbiota and meat quality of ruminants will play a positive role in understanding the formation mechanism of meat quality in ruminants and improving production efficiency. In this study, the fatty acid composition and content, expression of related genes, and structural characteristics of the ileum microbiota of ewes of Tibetan sheep at different ages (4 months, 1.5 years, 3.5 years, and 6 years) were detected and analyzed. The results revealed significant differences in fatty acid composition and content in the muscle of Tibetan sheep at different ages (p < 0.05); in addition, the content of MUFAs in the longissimus dorsi muscle and leg muscle was higher. Similarly, the expressions of muscle-related genes differed among the different age groups, and the expression of the LPL, SCD, and FABP4 genes was higher in the 1.5-year-old group. The ileum microbiota diversity was higher in the 1.5-year-old group, the Romboutsia abundance ratio was significantly higher in the 1.5-year-old group (p < 0.05), and there was a significant positive correlation with oleic acid (C18:1n9c) (p < 0.05). In conclusion, the content of beneficial fatty acids in the longissimus dorsi muscle and leg muscle of Tibetan sheep was higher at 1.5 years of age, and the best slaughter age was 1.5 years. This study provides a reference for in-depth research on the mechanism of the influence of the gut microbiota on meat quality and related regulation.

Abstract Background The gut microbiota play an important role in maintaining host metabolism, the immune system and health, while sex, genotype, diet and health have specific effects on the composition of the gut microbiota. Therefore, to explore the sex differences in the structure and function of rumen microbiota in Tibetan goats, herein we analyzed sex differences in rumen fermentation parameters, rumen microbiota and the expression of genes related to VFA transport in Tibetan goats. Results The results showed that the contents of acetic acid and propionic acid in the rumen of TGM (Tibetan goat male) were significantly higher than those in TGFm (Tibetan goat female) (P

Based on the serious phenomenon of Tibetan sheep “growing strong in warm seasons and losing weight in cold seasons”, this study explores the regulation of lncRNAs, microbiota, and metabolites in the cold season adaptation of Tibetan sheep from the perspective of the co-evolution of the host genome (first genome) and microbiome (second genome). RNA-seq results showed that 172 DE lncRNAs were identified in the rumen epithelium of Tibetan sheep in warm and cold seasons, of which 87 DE lncRNAs were significantly up-regulated in cold seasons. KEGG enrichment showed that target genes of up-regulated lncRNAs were significantly enriched in TNF signaling and oxidative phosphorylation pathways. LncRNA-mRNA regulatory network indicated that DE lncRNAs were involved in nutrient stress in the cold season by targeting ATP1B2, CADPS, TLR5, and UGT1A6. Correlation analysis showed some lncRNAs were significantly correlated with acetic acid, propionic acid, butyric acid, and rumen epithelial histomorphology and had a negative correlation with Butyrivibrio-2 and Succiniclasticum (p < 0.05). In addition, differential metabolites bilirubin and lncRNAs were co-enriched in the bile secretion pathway. lncRNAs played an important role in the adaptation process of Tibetan sheep in the cold season, and mediate the host to participate in nutrient absorption, energy utilization, and immune response, indicating that the host genome and microbial genome promote Tibetan sheep to adapt to nutrient stress in the cold season through co-evolution.

Plateau adaptation in animals involves genetic mechanisms as well as coevolutionary mechanisms of the microbiota and metabolome of the animal. Therefore, the characteristics of the rumen microbiome and metabolome, transcriptome, and serum metabolome of Tibetan sheep at different altitudes (4500 m, 3500 m, and 2500 m) were analyzed. The results showed that the rumen differential metabolites at 3500 m and 4500 m were mainly enriched in amino acid metabolism, lipid metabolism, and carbohydrate metabolism, and there was a significant correlation with microbiota. The differentially expressed genes and metabolites at middle and high altitudes were coenriched in asthma, arachidonic acid metabolism, and butanoate and propanoate metabolism. In addition, the serum differential metabolites at 3500 m and 4500 m were mainly enriched in amino acid metabolism, lipid metabolism, and metabolism of xenobiotics by cytochrome P450, and they were also related to microbiota. Further analysis revealed that rumen metabolites accounted for 7.65% of serum metabolites. These common metabolites were mainly enriched in metabolic pathways and were significantly correlated with host genes (p < 0.05). This study found that microbiota, metabolites, and epithelial genes were coenriched in pathways related to lipid metabolism, energy metabolism, and immune metabolism, which may be involved in the regulation of Tibetan sheep adaptation to plateau environmental changes.

Tibetan sheep are already well adapted to cold season nutrient stress on the Tibetan Plateau. Rumen, an important nutrient for metabolism and as an absorption organ in ruminants, plays a vital role in the cold stress adaptations of Tibetan sheep. Ruminal microbiota also plays an indispensable role in rumen function. In this study, combined multiomics data were utilized to comprehensively analyze the interaction mechanism between rumen epithelial miRNAs and microbiota and their metabolites in Tibetan sheep under nutrient stress in the cold season. A total of 949 miRNAs were identified in the rumen epithelium of both cold and warm seasons. A total of 62 differentially expressed (DE) miRNAs were screened using FC > 1.5 and p value < 0.01, and a total of 20,206 targeted genes were predicted by DE miRNAs. KEGG enrichment analysis revealed that DE miRNA-targeted genes were mainly enriched in axon guidance(ko04360), tight junction(ko04530), inflammatory mediator regulation of TRP channels(ko04750) and metabolism-related pathways. Correlation analysis revealed that rumen microbiota, rumen VFAs and DE miRNAs were all correlated. Further study revealed that the targeted genes of cold and warm season rumen epithelial DE miRNAs were coenriched with differential metabolites of microbiota in glycerophospholipid metabolism (ko00564), apoptosis (ko04210), inflammatory mediator regulation of TRP channels (ko04750), small cell lung cancer (ko05222), and choline metabolism in cancer (ko05231) pathways. There are several interactions between Tibetan sheep rumen epithelial miRNAs, rumen microbiota, and microbial metabolites, mainly through maintaining rumen epithelial barrier function and host homeostasis of choline and cholesterol, improving host immunity, and promoting energy metabolism pathways, thus enabling Tibetan sheep to effectively respond to cold season nutrient stress. The results also suggest that rumen microbiota have coevolved with their hosts to improve the adaptive capacity of Tibetan sheep to cold season nutrient stress, providing a new perspective for the study of cold season nutritional stress adaptation in Tibetan sheep.

Abstract Background Tibetan pigs have exhibited unique characteristics from low‐altitudes pigs and adapted well to the Qinghai‐Tibet Plateau. Objectives The current study was undertaken to investigate the hypoxic adaptation of heart in Tibetan pigs. Methods The hearts of Tibetan pigs and Landrace pigs raised at high or low altitudes were compared using 3D casting technology, scanning electron microscopy and real‐time quantitative PCR (qRT‐PCR). Results We found that the ratio of the major axis to the minor axis and the density of the heart were significantly higher in Tibetan pigs than in Landrace pigs (p

Tibetan sheep can maintain a normal life and reproduce in harsh environments under extreme cold and lack of nutrition. However, the molecular and metabolic mechanisms underlying the adaptability of Tibetan sheep during the cold season are still unclear. Hence, we conducted a comprehensive analysis of rumen epithelial morphology, epithelial transcriptomics, microbiology and metabolomics in a Tibetan sheep model. The results showed that morphological structure of rumen epithelium of Tibetan sheep in cold season had adaptive changes. Transcriptomics analysis showed that the differential genes were primarily enriched in the PPAR signaling pathway (ko03320), legionellosis (ko05134), phagosome (ko04145), arginine and proline metabolism (ko00330), and metabolism of xenobiotics by cytochrome P450 (ko00980). Unique differential metabolites were identified in cold season, such as cynaroside A, sanguisorbin B and tryptophyl-valine, which were mainly enriched in arachidonic acid metabolism, arachidonic acid metabolism and linolenic acid metabolism pathways, and had certain correlation with microorganisms. Integrated transcriptome-metabolome-microbiome analysis showed that epithelial gene-GSTM3 expression was upregulated in the metabolism of xenobiotics by the cytochrome P450 pathway during the cold season, leading to the downregulation of some harmful metabolites; TLR5 gene expression was upregulated and CD14 gene expression was downregulated in the legionellosis pathway during the cold season. This study comprehensively described the interaction mechanism between the rumen host and microbes and their metabolites in grazing Tibetan sheep during the cold season. Rumen epithelial genes, microbiota and metabolites act together in some key pathways related to cold season adaptation.

Objective The extensive breeding of commercial chickens has led to a sharp decrease in the resources of many indigenous chickens, especially the indigenous chickens in the southeastern coastal region, which are on the verge of extinction, and the indigenous chickens in the northwestern region of China, which are also at risk. However, there are few reports on the evaluation of genetic diversity and conservation of genetic resources of indigenous chickens in remote areas in the Northwest of China. Methods In the present study, the genetic diversity and phylogenetic relationship of six indigenous chickens from different regions were studied based on variation in mitochondrial DNA control region (D-loop), and the degree of introgression from commercial breeds into these chickens was determined by the amount of haplotype sharing between indigenous and commercial breeds. Results Twenty-five polymorphic sites and 25 haplotypes were detected in 206 individuals. Principal component analysis showed that the Jingning chicken had the highest genetic diversity among the six indigenous chickens. According to the degree of introgression, the six indigenous breeds may be involved in haplotype sharing with commercial breeds, and the introgression from commercial chickens into the Haidong chicken is the most serious. Conclusion The genetic uniqueness of indigenous chickens has been eroded, so it is necessary to consider the protection of their genetic resources. Phylogenetic analysis suggests that the six indigenous chickens have two major matrilineal origins: one from Yunnan or its surrounding areas in China and the other from the Indian subcontinent.

The population genetic structure of domestic animals and the protection of genetic resources are major issues of concern in the field of global biology. In this study, the mtDNA D-loop sequences of 1163 Chinese indigenous cattle (Bos taurus and Bos indicus) were used to analyze their maternal origin and population genetic structure and to prioritize conservation evaluations based on a population genetic evaluation model. The results revealed three maternal origins or domestication events among Chinese indigenous cattle and the occurrence of secondary expansions but also the phenomenon of gene penetration. The population genetic evaluation model results showed that the genetic diversity of Huangpi cattle (HHP), Mountainous cattle (CMO), Bashan cattle (SBS) and Guanling cattle (GGL)populations was highest and that the Zaosheng cattle (GZN), Qinchuan cattle (SQB) and Pingliang cattle (GPL) populations contributed most to the richness of Chinese indigenous cattle genetic resources, indicating that these populations should be protected first. Our research provides a scientific basis and effective evaluation methods for the protection of genetic resources of indigenous livestock varieties.

The rumen is an important hallmark organ of ruminants and plays an important role in the metabolism and immune barrier of Tibetan sheep on the Plateau. However, there are few studies on rumen development and metabolism regulation in Tibetan sheep at different ages. Here, we comprehensively analyzed the immune function, fermentation function, rumen epithelial micromorphology and transcriptome profile of Tibetan sheep at different ages. The results showed that the concentration of IgG decreased and the concentration of IgM increased with age (p < 0.05), and the highest concentration of IgA was observed at 1.5 and 3.5 years of age. In terms of rumen fermentation characteristics, VFAs of 4-month-old lambs were the highest, followed by VFAs and NH3-N of Tibetan sheep at 3.5 years of age. Hematoxylin-eosin staining and transmission electron microscopy section examination of rumen epithelial tissue showed that the rumen papilla width increased with age (p < 0.001), the thickness of the stratum corneum decreased, the cells in the stratum corneum showed accelerated migration and the thickness of the rumen muscle layer increased (p < 0.001). Desmosomal junctions between the layers of rumen epithelium increased at 1.5 and 3.5 years old, forming a compact barrier structure, and the basal layer had more mitochondria involved in the regulation of energy metabolism. RNA-seq analysis revealed that a total of 1006 differentially expressed genes (DEGs) were identified at four ages. The DEGs of Tibetan sheep aged 4 months and 6 years were mainly enriched in the oxidation–reduction process and ISG15-protein conjugation pathway. The 1.5 and 3.5-year-olds were mainly enriched in skeletal muscle thin filament assembly, mesenchyme migration and the tight junction pathway. WGCNA showed that DEGs related to rumen microbiota metabolite VFAs and epithelial morphology were enriched in “Metabolism of xenobiotics by cytochrome P450, PPAR signaling pathway, Butanoate metabolism pathways” and participated in the regulation of rumen epithelial immune and fermentation metabolism functions of Tibetan sheep at different ages. This study systematically revealed the regulatory mechanism of rumen epithelial development and metabolism in the plateau adaptation of Tibetan sheep, providing a new approach for the study of plateau adaptation.

Altitude is the main external environmental pressure affecting the production performance of Tibetan sheep, and the adaptive evolution of many years has formed a certain response mechanism. However, there are few reports on the response of ruminal microbiota and host genomes of Tibetan sheep to high-altitude environments. Here, we conducted an integrated analysis of volatile fatty acids (VFAs), microbial diversity (16S rRNA), epithelial morphology, and epithelial transcriptome in the rumen of Tibetan sheep at different altitudes to understand the changes in ruminal microbiota–host interaction in response to high altitude. The differences in the nutritional quality of forage at different altitudes, especially the differences in fiber content (ADF/NDF), led to changes in rumen VFAs of Tibetan sheep, in which the A/P value (acetic acid/propionic acid) was significantly decreased (p < 0.05). In addition, the concentrations of IgA and IgG in Middle-altitude (MA) and High-altitude Tibetan sheep (HA) were significantly increased (p < 0.05), while the concentrations of IgM were significantly increased in MA (p < 0.05). Morphological results showed that the width of the rumen papilla and the thickness of the basal layer increased significantly in HA Tibetan sheep (p < 0.05). The 16S rRNA analysis found that the rumen microbial diversity of Tibetan sheep gradually decreased with increasing altitude, and there were some differences in phylum- and genus-level microbes at the three altitudes. RDA analysis found that the abundance of the Rikenellaceae RC9 gut group and the Ruminococcaceae NK4A214 group increased with altitudes. Furthermore, a functional analysis of the KEGG microbial database found the “lipid metabolism” function of HA Tibetan sheep to be significantly enriched. WGCNA revealed that five gene modules were enriched in “energy production and conversion”, “lipid transport and metabolism”, and “defense mechanisms”, and cooperated with microbiota to regulate rumen fermentation and epithelial immune barrier function, so as to improve the metabolism and immune level of Tibetan sheep at high altitude.

This study explores the effects of the coevolution of the host genome (the first genome) and gut microbiome (the second genome) on nutrition stress in Tibetan sheep during the cold season. The rumen epithelial tissue of six Tibetan sheep (Oula-type) was collected as experimental samples during the cold and warm seasons and the study lasted for half a year. The cDNA library was constructed and subjected to high-throughput sequencing. The circRNAs with significant differential expression were identified through bioinformatics analysis and functional prediction, and verified by real-time quantitative PCR (qRT-PCR). The results showed that a total of 56 differentially expressed (DE) circRNAs of rumen epithelial tissue were identified using RNA-seq technology, among which 29 were significantly upregulated in the cold season. The circRNA-miRNA regulatory network showed that DE circRNAs promoted the adaptation of Tibetan sheep in the cold season by targeting miR-150 and oar-miR-370-3p. The results of correlation analysis among circRNAs, microbiota, and metabolites showed that the circRNA NC_040275.1:28680890|28683112 had a very significant positive correlation with acetate, propionate, butyrate, and total volatile fatty acid (VFA) (p < 0.01), and had a significant positive correlation with Ruminococcus-1 (p < 0.05). In addition, circRNA NC_040256.1:78451819|78454934 and metabolites were enriched in the same KEGG pathway biosynthesis of amino acids (ko01230). In conclusion, the host genome and rumen microbiome of Tibetan sheep co-encoded a certain glycoside hydrolase (β-glucosidase) and coevolved efficient VFA transport functions and amino acid anabolic processes; thus, helping Tibetan sheep adapt to nutrient stress in the cold season in high-altitude areas.

Cattle-yak, a hybrid offspring of yak (Bos grunniens) and cattle (Bos taurus), inhabit the Qinghai-Tibet Plateau at an altitude of more than 3,000 m and obtain nutrients predominantly through grazing on natural pastures. Severe shortages of pasture in the cold season leads to reductions in the weight and disease resistance of grazing cattle-yak, which then affects their production performance. This study aimed to investigate the effect of supplementary feeding during the cold season on the rumen microbial community of cattle-yak. Six cattle-yak (bulls) were randomly divided into two groups—“grazing + supplementary feeding” (G+S) (n = 3) and grazing (G) (n = 3)—and rumen microbial community structure (based on 16S rRNA sequencing), volatile fatty acids (VFAs), and ruminal epithelial sodium ion-dependent glucose transporter 1 (SGLT1) expression were assessed. There were significant differences in the flora of the two groups at various taxonomic classification levels. For example, Bacteroidetes, Rikenellaceae, and Rikenellaceae_RC9_gut_group were significantly higher in the G+S group than in the G group (P < 0.05), while Firmicutes and Christensenellaceae_R-7_group were significantly lower in the G+S group than in the G group (P < 0.05). Kyoto Encyclopedia of Genes and Genomes (KEGG) and Clusters of Orthologous Groups (COG) analyses revealed that functions related to carbohydrate metabolism and energy production were significantly enriched in the G+S group (P < 0.05). In addition, the concentration of total VFAs, along with concentrations of acetate, propionate, and butyrate, were significantly higher in the G+S group than in the G group (P < 0.05). Furthermore, SGLT1 expression in ruminal epithelial tissue was significantly lower in the G+S group (P < 0.01). Supplementary feeding of cattle-yak after grazing in the cold season altered the microbial community structure and VFA contents in the rumen of the animals, and decreased ruminal epithelial SGLT1 expression. This indicated that supplementary feeding after grazing aids rumen function, improves adaptability of cattle-yak to the harsh environment of the Qinghai-Tibet Plateau, and enhances ability of the animals to overwinter.

As one of the important ruminants of the Qinghai-Tibet Plateau, Tibetan sheep are able to reproduce and maintain their population in this harsh environment of extreme cold and low oxygen. However, the adaptive mechanism of Tibetan sheep when nutrients are scarce in the cold season of the Plateau environment is unclear. We conducted comparative analysis rumen fermentation parameters, rumen microbes, and expression of host genes related to nutrient absorption and rumen epithelial barrier function in cold and warm season Tibetan sheep. We found that concentrations of the volatile fatty acids (VFAs) acetate, propionate and butyrate of Tibetan sheep in the cold season were significantly higher than in the warm season (P < 0.05). Microbial 16S rRNA gene analysis revealed significant differences in rumen microbiota between the cold and warm seasons, and the abundance of microbial in the cold season was significantly higher than that in the warm season (P < 0.05), and the lack of nutrients in the cold season led to a significant reduction in the expression of SGLT1, Claudin-4, and ZO-1 genes in the rumen epithelium. Correlation analysis revealed significant associations of some rumen microorganisms with the fermentation product acetate and the rumen epithelial genes SGLT1, Claudin-4, and ZO-1.

As an important ruminant on the Qinghai-Tibet Plateau, Tibetan sheep can maintain their population reproduction rate in the harsh high-altitude environment of low temperature and low oxygen, which relies on their special plateau adaptations mechanism that they have formed for a long time. Microbiomes (known as “second genomes”) are closely related to the nutrient absorption, adaptability, and health of the host. In this study, rumen fermentation characteristics, the microbiota, and rumen epithelial gene expression of Tibetan sheep in various months were analyzed. The results show that the rumen fermentation characteristics of Tibetan sheep differed in different months. The total SCFAs (short-chain fatty acids), acetate, propionate, and butyrate concentrations were highest in October and lowest in June. The CL (cellulase) activity was highest in February, while the ACX (acid xylanase) activity was highest in April. In addition, the diversity and abundance of rumen microbes differed in different months. Bacteroidetes (53.4%) and Firmicutes (27.4%) were the dominant phyla. Prevotella_1 and Rikenellaceae_RC9_gut_group were the dominant genera. The abundance of Prevotella_1 was highest in June (27.8%) and lowest in December (17.8%). In addition, the expression of CLAUDIN4 (Claudin-4) and ZO1 (Zonula occludens 1) was significantly higher in April than in August and December, while the expression of SGLT1 (Sodium glucose linked transporter 1) was highest in August. Correlation analysis showed that there were interactions among rumen fermentation characteristics, the microbiota, and host gene expression, mainly by adjusting the amino acid metabolism pathway and energy metabolism pathway to improve energy utilization. At the same time, we adjusted the balance of the rumen “core microbiota” to promote the development of rumen and maintain the homeostasis of rumen environment, which makes Tibetan sheep better able to adapt to the harsh environment in different periods of the Qinghai-Tibet Plateau.

Aim: The meat of Tibetan sheep has a unique flavor, delicious taste, and superior nutritional value. However, the change of grass will lead to a change in meat quality. This study aimed to explore the potential regulatory mechanisms of microbial metabolites with respect to meat quality traits of Tibetan sheep under nutrient stress in the cold season. Methods and results: We determined and analyzed the longissimus dorsi quality, fatty acid composition, expression of genes, and rumen microbial metabolites of Tibetan sheep in cold and warm seasons. The shear force was decreased (P < .05), the meat color a∗24 h value was increased (P < .05), and the contents of crude fat (EE) and protein (CP) were decreased in the cold season. Polyunsaturated fatty acids (PUFAs)- linoleic acid and docosahexaenoic acid increased significantly in the cold season (P < .05). The expressions of meat quality genes MC4R, CAPN1, H-FABP, and LPL were significantly higher in the warm season (P < .05), and the CAST gene was significantly expressed in the cold season (P < .01). The different microbial metabolites of Tibetan sheep in the cold and warm seasons were mainly involved in amino acid metabolism, lipid metabolism, and digestive system pathway, and there was some correlation between microbiota and meat quality traits. There are similarities between microbial metabolites enriched in the lipid metabolism pathway and muscle metabolites. Conclusion: Under nutritional stress in the cold season, the muscle tenderness of Tibetan sheep was improved, and the fat deposition capacity was weakened, but the levels of beneficial fatty acids were higher than those in the warm season, which was more conducive to healthy eating. [ABSTRACT FROM AUTHOR]