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The objective of this experiment was to explore the effects of three-way hybridization on rumen microbes and metabolites in sheep using rumen metagenomics and metabolomics. Healthy Hu and CAH (Charolais × Australian White × Hu) male lambs of similar birth weight and age were selected for short-term fattening after intensive weaning to collect rumen fluid for sequencing. Rumen metagenomics diversity showed that Hu and CAH sheep were significantly segregated at the species, KEGG-enzyme, and CAZy-family levels. Moreover, the CAH significantly increased the ACE and Chao1 indices. Further, correlation analysis of the abundance of the top 80 revealed that the microorganisms were interrelated at the species, KEGG-enzyme, and CAZy-family levels. Overall, the microbiome significantly affected metabolites of the top five pathways, with the strongest correlation found with succinic acid. Meanwhile, species-level microbial markers significantly affected rumen differential metabolites. In addition, rumen microbial markers in Hu sheep were overall positively correlated with down-regulated metabolites and negatively correlated with up-regulated metabolites. In contrast, rumen microbial markers in CAH lambs were overall negatively correlated with down-regulated metabolites and positively correlated with up-regulated metabolites. These results suggest that three-way crossbreeding significantly affects rumen microbial community and metabolite composition, and that significant interactions exist between rumen microbes and metabolites.

The Qinghai-Tibet Plateau is a valuable genetic resource pool, and the high-altitude adaptation of Tibetan pigs is a classic example of the adaptive evolution of domestic animals. Here, we report the presence of Darwinian positive selection signatures in Tibetan pigs (TBPs) using 348 genome-wide datasets (127 whole-genome sequence datasets (WGSs) and 221 whole-genome single-nucleotide polymorphism (SNP) chip datasets). We characterized a high-confidence list of genetic signatures related response to high-altitude adaptation in Tibetan pigs, including 4,598 candidate SNPs and 131 candidate genes. Functional annotation and enrichment analysis revealed that 131 candidate genes are related to multiple systems and organs in Tibetan pigs. Notably, eight of the top ten novel genes, RALB, NBEA, LIFR, CLEC17A, PRIM2, CDH7, GK5 and FAM83B, were highlighted and associated with improved adaptive heart functions in Tibetan pigs high-altitude adaptation. Moreover, genome-wide association analysis revealed that 29 SNPs were involved in 13 candidate genes associated with at least one adaptive trait. In particular, among the top ten candidate genes, CLEC17A is related to a reduction in hemoglobin (HGB) in Tibetan pigs. Overall, our study provides a robust SNP/gene list involving genetic adaptation for Tibetan pig high-altitude adaptation, and it will be a valuable resource for future Tibetan pig studies., (© 2024. The Author(s).)

Red clover (Trifolium pratense) isoflavone was supplemented to dairy cows, and antioxidant capacity of milk was assessed. Treated cows increased the activities of antioxidant enzymes, reduced production of oxidation products, and enhanced the concentrations of vitamin E and vitamin C. Moreover, milk fatty acid profile was positive influenced by 8 g/kg red clover isoflavone, with changes in the lower saturated and higher unsaturated fatty acids. We further demonstrated the efficacy of antioxidant capacity of milk in mice, found that milk from cows feeding red clover isoflavone increased the expressions of antioxidant enzymes, and alleviated lipopolysaccharide (LPS)-stimulated tissue damage of duodenum and jejunum, which was related to upregulated metabolism pathways of carbohydrate, lipid, and amino acid, as well as downregulated inflammatory related pathways. Together, dietary supplementation of red clover isoflavone is an effective way to improve milk antioxidant capacity, providing a natural strategy for developing functional foods., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

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 (NH 3 -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&#95;7 and Succinivibrionaceae&#95;UCG&#95;001 in the STE group, whereas the relative abundance of Rikenellaceae&#95;RC9&#95;gut&#95;group significantly decreased ( p < 0.05). According to the correlation analysis between rumen microflora and VFAs, the relative abundance of Succinivibrionaceae&#95;UCG&#95;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.

A novel ratiometric fluorescent probe based on non-conjugated polymer dots (NCPDs) and gold nanocluster (AuNCs) was constructed to determine tert-butylhydroquinone (TBHQ). The probe exhibited dual emission peaks at 480 nm and 630 nm under 370 nm excitation. The fluorescence of AuNCs was quenched by TBHQ due to strong electrostatic interactions, whereas the emission of NCPDs increased. The ratio of fluorescence intensity at 480 nm to 630 nm (F 480 / F 630 ) was monitored as analytical signal response. The probe have been utilized for the detection of TBHQ with good linear relationship in the range of 0.2 to 60 μg/mL. The limit of detection (LOD) and the limit of quantitation (LOQ) were 0.048 μg/mL and 0.159 μg/L, respectively. Three levels of spiked-in TBHQ concentrations were obtained with recovery rates from 80 % to 102 %. The present study provided an effective ratiometric fluorescence method for selective screening of TBHQ in food samples., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Copy number variations (CNVs) critically influence individual genetic diversity and phenotypic traits. In this study, we employed whole-genome resequencing technology to conduct an in-depth analysis of 50 pigs from five local swine populations [Rongchang pig (RC), Wuzhishan pig (WZS), Tibetan pig (T), Yorkshire (YL) and Landrace (LR)], aiming to assess their genetic potential and explore their prospects in the field of animal model applications. We identified a total of 96,466 CNVs, which were subsequently integrated into 7112 non-redundant CNVRs, encompassing 1.3% of the swine genome. Functional enrichment analysis of the genes within these CNVRs revealed significant associations with sensory perception, energy metabolism, and neural-related pathways. Further selective scan analyses of the local pig breeds RC, T, WZS, along with YL and LR, uncovered that for the RC variety, the genes PLA2G10 and ABCA8 were found to be closely related to fat metabolism and cardiovascular health. In the T breed, the genes NCF2 and CSGALNACT1 were associated with immune response and connective tissue characteristics. As for the WZS breed, the genes PLIN4 and CPB2 were primarily linked to fat storage and anti-inflammatory responses. In summary, this research underscores the pivotal role of CNVs in fostering the diversity and adaptive evolution of pig breeds while also offering valuable insights for further exploration of the advantageous genetic traits inherent to China's local pig breeds. This facilitates the creation of experimental animal models tailored to the specific characteristics of these breeds, contributing to the advancement of livestock and biomedical research.

This experiment was conducted to investigate the effect of three-way hybrid sheep and Hu sheep on serum indicators, rumen fermentation, rumen enzyme activity, and microorganisms in sheep. Healthy and similar birth weights from three groups (Hu, n = 11; Charolais × Australian White × Hu, CAH, n = 11; Charolais × Dorper × Hu, CDH, n = 11) were selected to be fed by the ewes until 45 days of age. Subsequently, they were weaned intensively and underwent short-term fattening for 3 months along with selected male lambs fed intensively. During this period, they were fed and watered ad libitum. Blood and rumen fluid were collected and analyzed for serum indicators and rumen fluid microorganisms, enzyme activity, and VFA, respectively, at the end of the fattening period. Compared with Hu lamb, the offspring of the three-way hybrid lamb showed significant improvements in body weight, serum lactate dehydrogenase, and creatinine content. However, there was no significant effect on serum immunity and antioxidant indices. In addition, the rumen fluid volatile fatty acid (VFA) molar concentration and microcrystalline cellulose and lipase content were significantly lower in the three-way hybrid lamb compared to Hu lamb, but β-glucosidase, amylase, pepsin, and VFA molar ratio were not significantly affected. Subsequently, 16S rRNA sequencing diversity analysis revealed that three-way hybrid lamb significantly increased rumen microbial ACE and Chao1 indices compared to Hu lamb. Meanwhile, the abundance of Verrucomicrobiota and Synergistota significantly increased at the phylum level. Correlation analysis showed that Prevotella had the highest proportion, while Rikenellaceae&#95;RC9&#95;gut&#95;group correlated most closely with others genus. The microbial communities isovaleric acid molar concentration and proportion were strongly correlated. In addition, there were significant differences in correlations between microbial communities and isobutyric acid, butyric acid and valeric acid content, and their molar proportion, but they were not significantly correlated with digestive enzymes. From the functional enrichment analysis, it was found that hybrid progeny were mainly enriched in the pyruvate metabolism, microbial metabolism in diverse environments, carbon metabolism, and quorum sensing pathways. In contrast, the Hu sheep were primarily enriched in the cysteine and methionine, amino sugar and nucleotide sugar, and biosynthesis of secondary metabolite pathways. These results suggest that hybridization can play a role in regulating organismal metabolism and improve animal production performance by influencing the structure and characteristics of microbial communities.

The transition period for dairy cows usually refers to the 3 weeks pre-calving to the 3 weeks post-calving. During this period, dairy cows undergo metabolic and physiological adaptations because of their susceptibility to metabolic and infectious diseases. Poor feeding management under these circumstances may adversely affect the health and subsequent production performance of the cows. Owing to long-term adaptation and evolution, the rumen has become a unique ecosystem inhabited by a complex microbial community closely associated with its natural host. Dietary components are metabolized by the rumen microbiota, and volatile fatty acids and microbial protein products can be used as precursor substances for synthesizing meat and milk components. The successful transition of perinatal dairy cows includes changes in diet, physiology, and the rumen microbiota. Rumen microbial profiles have been confirmed to be heritable and repairable; however, adverse circumstances affect rumen microbial composition, host digestion and metabolism, as well as postpartum production traits of dairy cows for a certain period. Preliminary evidence indicates a close relationship between the rumen microbiota and animal performance. Therefore, changes in rumen microbes during the transition period and the intrinsic links between the microbiota and host postpartum phenotypic traits need to be better understood to optimize production performance in ruminants., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the content of this paper., (© 2024 The Authors.)

The inclusion of fibrolytic enzymes in the diet is believed to have positive effects on animal production. Hence, the objective of this study was to investigate the impact of supplementing diets with a commercial fibrolytic enzyme preparation ( Vistamax ; mixture of xylanase and cellulase) derived from Trichoderma reesei on lactational performance, digestibility, and plasma nutrient levels in high-producing dairy cows. Two dietary energy levels were considered: a normal energy diet (metabolizable energy = 2.68 Mcal/kg) and a slightly lower energy diet (metabolizable energy = 2.55 Mcal/kg). A total of 120 lactating Holstein cows (parity = 2; Days in Milk = 113 ± 23) were randomly assigned to four treatment groups using a 2 * 2 factorial arrangement. The dietary treatments consisted of: (1) normal energy diet without enzyme supplementation (NL); (2) normal energy diet with enzyme supplementation (NLE); (3) slightly lower energy diet without enzyme supplementation (SL); and (4) slightly lower energy diet with enzyme supplementation (SLE). The amount of enzyme added to the diets was determined based on previous in vitro studies and supplier recommendations. The enzyme and premix were mixed prior to the preparation of the total mixed ration, and the trial lasted for a duration of 42 days. The results indicated that the application of the fibrolytic enzyme did not have a significant effect on dry matter intake (DMI), but it did enhance the digestibility of dry matter (DM), neutral detergent fiber (NDF), potentially digestible NDF (pdNDF), organic matter (OM), milk production, milk urea nitrogen (MUN), and blood urea nitrogen (BUN). On the other hand, the slightly lower energy diet resulted in a decrease in DMI, milk production, milk protein yield, plasma free amino acids (FAA), and an increase in plasma B-hydroxybutyrate (BHBA). In conclusion, the inclusion of the fibrolytic enzyme in the diets of dairy cows led to improvements in the digestibility of DM, NDF, pdNDF, OM, milk production, and feed efficiency. Furthermore, the application of the enzyme to the slightly lower energy diet resulted in milk production levels comparable to those observed in cows fed the untreated normal energy diet., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Yang, Zhao and Lin.)

Rumen microbial urease inhibitors have been proposed for regulating nitrogen emission and improving nitrogen utilization efficiency in ruminant livestock industry. However, studies on plant-derived natural inhibitors of rumen microbial urease are limited. Urease accessory protein UreG, plays a crucial role in facilitating urease maturation, is a new target for design of urease inhibitor. The objective of this study was to select the potential effective inhibitor of rumen microbial urease from major protoberberine alkaloids in Rhizoma Coptidis by targeting UreG. Our results showed that berberine chloride and epiberberine exerted superior inhibition potential than other alkaloids based on GTPase activity study of UreG. Berberine chloride inhibition of UreG was mixed type, while inhibition kinetics type of epiberberine was uncompetitive. Furthermore, epiberberine was found to be more effective than berberine chloride in inhibiting the combination of nickel towards UreG and inducing changes in the second structure of UreG. Molecular modeling provided the rational structural basis for the higher inhibition potential of epiberberine, amino acid residues in G1 motif and G3 motif of UreG formed interactions with D ring of berberine chloride, while interacted with A ring and D ring of epiberberine. We further demonstrated the efficacy of epiberberine in the ruminal microbial fermentation with low ammonia release and urea degradation. In conclusion, our study clearly indicates that epiberberine is a promising candidate as a safe and effective inhibitor of rumen microbial urease and provides an optimal strategy and suitable feed additive for regulating nitrogen excretion in ruminants in the future. KEY POINTS: • Epiberberine is the most effective inhibitor of rumen urease from Rhizoma Coptidis. • Urease accessory protein UreG is an effective target for design of urease inhibitor. • Epiberberine may be used as natural feed additive to reducing NH 3 release in ruminants., (© 2024. The Author(s).)

Introduction: The rumen microbial community plays a crucial role in the digestion and metabolic processes of ruminants. Although sequencing-based studies have helped reveal the diversity and functions of bacteria in the rumen, their physiological and biochemical characteristics, as well as their dynamic regulation along the digestion process in the rumen, remain poorly understood. Addressing these gaps requires pure culture studies to demystify the intricate mechanisms at play. Bacteria exhibit morphological differentiation associated with different species. Based on the difference in size or shape of microorganisms, size fractionation by filters with various pore sizes can be used to separate them., Methods: In this study, we used polyvinylidene difluoride filters with pore sizes of 300, 120, 80, 40, 20, 8, 6, 2.1, and 0.6 μm. Bacterial suspensions were successively passed through these filters for the analysis of microbial population distribution using 16S rRNA gene sequences., Results: We found that bacteria from the different pore sizes were clustered into four branches (> 120 μm, 40-120 μm, 6-20 μm, 20-40 μm, and < 0.6 μm), indicating that size fractionation had effects on enriching specific groups but could not effectively separate dominant groups by cell size alone. The species of unclassified Flavobacterium, unclassified Chryseobacterium, unclassified Delftia , Methylotenera mobilis , unclassified Caulobacteraceae, unclassified Oligella , unclassified Sphingomonas , unclassified Stenotrophomonas , unclassified Shuttleworthia , unclassified Sutterella , unclassified Alphaproteobacteria, and unclassified SR1 can be efficiently enriched or separated by size fractionation., Discussion: In this study, we investigated the diversity of sorted bacteria populations in the rumen for preliminary investigations of the relationship between the size and classification of rumen bacteria that have the potential to improve our ability to isolate and culture bacteria from the rumen in the future., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Liu, Zheng, Wang and Zhao.)

The Microbiome Protocols eBook (MPB) serves as a crucial bridge, filling gaps in microbiome protocols for both wet experiments and data analysis. The first edition, launched in 2020, featured 152 meticulously curated protocols, garnering widespread acclaim. We now extend a sincere invitation to researchers to participate in the upcoming 2nd version of MPB, contributing their valuable protocols to advance microbiome research., Competing Interests: The authors declare no conflict of interest., (© 2024 The Authors. iMeta published by John Wiley & Sons Australia, Ltd on behalf of iMeta Science.)

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 (NH 3 -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&#95;Selenomonadaceae and Rikenellaceae&#95;RC9&#95;gut&#95;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 NH 3 -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 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&#95;XPB1014&#95;group , Prevotella , and Rikenellaceae&#95;RC9&#95;gut&#95;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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Sha, Liu, He, Zhao, Hu, Wang, Li, Shao, Wang, Chen, Yang and Xie.)

Background: Specific pathogen-free ducks are a valuable laboratory resource for waterfowl disease research and poultry vaccine development. High throughput sequencing allows the systematic identification of structural variants in genomes. Copy number variation (CNV) can explain the variation of important duck genetic traits. Herein, the genome-wide CNVs of the three experimental duck species in China (Jinding ducks (JD), Shaoxing ducks (SX), and Fujian Shanma ducks (SM)) were characterized using resequencing to determine their genetic characteristics and selection signatures., Results: We obtained 4,810 CNV regions (CNVRs) by merging 73,012 CNVs, covering 4.2% of the duck genome. Functional analysis revealed that the shared CNVR-harbored genes were significantly enriched for 31 gene ontology terms and 16 Kyoto Encyclopedia of Genes and Genomes pathways (e.g., olfactory transduction and immune system). Based on the genome-wide fixation index for each CNVR, growth (SPAG17 and PTH1R), disease resistance (CATHL3 and DMBT1), and thermoregulation (TRPC4 and SLIT3) candidate genes were identified in strongly selected signatures specific to JD, SM, and SX, respectively., Conclusions: In conclusion, we investigated the genome-wide distribution of experimental duck CNVs, providing a reference to establish the genetic basis of different phenotypic traits, thus contributing to the management of experimental animal genetic resources., (© 2023. The Author(s).)

This study was to conducted to investigate the effect of red clover isoflavones on the health indicated by immune status and blood biochemistry in dairy cows. Sixty-eight healthy Holstein lactating cows were randomly divided into four treatments ( n  = 17 per treatment) from 5 blocks according to milk yield using a randomized complete block design. No initial differences in parity (2.13 ± 1.21), days in milk (165 ± 21 d), and milk yield (33.93 ± 3.81 kg/d) between groups. Cows were fed the basal diet supplemented with 0, 2, 4, or 8 g/kg red clover extract (RCE) in diet (dry matter based). Feeding, refusal feed weights, and milk yield were recorded three consecutive days in weeks 0, 4, 8, and 12. Blood was collected from the tail vein of the cows on the last day of weeks 4, 8 and 12, 1 h after the morning feeding, and analyzed for hormones, immunoglobulins, inflammatory markers, and markers of liver and kidney activities. The dry matter intake was significantly decreased by 3.7% in the 8 g/kg group ( P  < 0.05). The fat-corrected milk yield was significantly higher in both of the 2 and 4 g/kg groups ( P  < 0.01). Plasma estradiol and prolactin showed a quadratic effect with increasing RCE levels, with the highest in the 4 g/kg group ( P  < 0.05). Plasma tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β levels decreased linearly with increasing dietary RCE levels. Plasma IL-18 levels showed a quadratic effect with increasing dietary RCE levels, with significantly lower levels in both of the 2 and 4 g/kg groups ( P  < 0.05). Plasma immunoglobulin A and D-lactic acid levels showed a quadratic effect with increasing dietary RCE levels, with significantly higher level in the 4 g/kg group ( P  < 0.05). The liver function and kidney activity makers were similar ( P  > 0.05). These results recommend the supplementation of RCE at a level from 2 to 4 g/kg DM., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the content of this paper., (© 2024 The Authors.)

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., Competing Interests: The authors declare no conflict of interest.

The milk fat globule membrane ( MFGM ) is a complex tri-layer membrane that wraps droplets of lipids in milk. In recent years, it has attracted widespread attention due to its excellent bioactive functions and nutritional value. MFGM contains a diverse array of bioactive lipids, including cholesterol, phospholipids, and sphingolipids, which play pivotal roles in mediating the bioactivity of the MFGM . We sequentially summarize the main lipid types in the MFGM in this comprehensive review and outline the characterization methods used to employ them. In this comprehensive review, we sequentially describe the types of major lipids found in the MFGM and outline the characterization methods employed to study them. Additionally, we compare the structural disparities among glycerophospholipids, sphingolipids, and gangliosides, while introducing the formation of lipid rafts facilitated by cholesterol. The focus of this review revolves around an extensive evaluation of the current research on lipid isolates from the MFGM , as well as products containing MFGM lipids, with respect to their impact on human health. Notably, we emphasize the clinical trials encompassing a large number of participants. The summarized bioactive functions of MFGM lipids encompass the regulation of human growth and development, influence on intestinal health, inhibition of cholesterol absorption, enhancement of exercise capacity, and anticancer effects. By offering a comprehensive overview, the aim of this review is to provide valuable insights into the diverse biologically active functions exhibited by lipids in the MFGM .

Introduction: Milk fat is the most variable nutrient in milk, and recent studies have shown that rumen bacteria are closely related to milk fat. However, there is limited research on the relationship between rumen bacteria and milk fatty. Fatty acids (FAs) are an important component of milk fat and are associated with various potential benefits and risks to human health., Methods: In this experiment, forty-five healthy Holstein dairy cows with alike physiological and productive conditions were selected from medium-sized dairy farms and raised under the same feeding and management conditions. The experimental period was two weeks. During the experiment, raw milk and rumen fluid were collected, and milk components were determined. In this study, 8 high milk fat percentage (HF) dairy cows and 8 low milk fat percentage (LF) dairy cows were selected for analysis., Results: Results showed that the milk fat percentage in HF group was significantly greater than that of the dairy cows in the LF group. 16S rRNA gene sequencing showed that the rumen bacterial abundance of HF dairy cows was significantly higher than that in LF dairy cows; at the genus level, the bacterial abundances of Prevotellaceae&#95;UCG-001, Candidatus&#95;Saccharimonas, Prevotellaceae&#95;UCG-003 , Ruminococcus &#95;1, Lachnospiraceae&#95;XPB1014&#95;group , Lachnospiraceae&#95;AC2044&#95;group , probable&#95;genus&#95;10 and U29-B03 in HF group were significantly higher than those in the LF group. Spearman rank correlation analysis indicated that milk fat percentage was positively related to Prevotellaceae&#95;UCG-001, Candidatus&#95;Saccharimonas, Prevotellaceae&#95;UCG-003, Ruminococcus &#95;1, Lachnospiraceae&#95;XPB1014&#95;group , Lachnospiraceae&#95;AC2044&#95;group , probable&#95;genus&#95;10 and U29-B03 . Furthermore, Prevotellaceae&#95;UCG-001 was positively related to C14:0 iso, C15:0 iso, C18:0, Ruminococcus &#95;1 with C18:1 t9, Lachnospiraceae&#95;AC2044&#95;group with C18:1 t9 and C18:1 t11, U29-B03 with C15:0 iso., Discussion: To sum up, rumen bacteria in dairy cows are related to the variation of milk fat, and some rumen bacteria have potential effects on the deposition of certain fatty acids in raw milk., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Si, Liu, Huang, Chen, Yang, Wu, Li, Tang, Zhao, Zheng, Zhang and Wang.)

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.

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., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

An optofluidic sensor based on a Bragg grating in hollow-core fiber (HCF) is experimentally demonstrated. The grating is inscribed into the HCF by femtosecond laser illumination through a phase mask. Periodic index modulation is introduced into the silica material surrounding the hollow core, causing cladding mode resonance, and multiple reflection peaks are observed in the grating spectrum. These reflection peaks later shift to longer wavelengths when high-index liquid is infiltrated into the HCF. The new reflection peak results from the backward coupling of the liquid core mode of the waveguide, the mode field of which overlaps with the grating modulation surrounding the liquid core. The resonant wavelength of the liquid-core fiber grating increases with the index value of the infiltrating liquid, and optofluidic refractive index sensing is realized with the device. The highest refractive index sensitivity, 1117 nm/RIU, is obtained experimentally in the index range of 1.476-1.54. The infiltrated hollow-core fiber Bragg grating also exhibits high temperature sensitivity due to the high thermal-optic coefficient of the liquid, and a sensitivity of -301 pm/°C is achieved in the temperature range of 25°C to 60°C.

Introduction. Dairy calves, particularly pre-weaned calves have been identified as a common source of multidrug resistant (MDR) Escherichia coli . Gap statement. E. coli strains isolated from dairy calves and the location of their resistance genes (plasmid or chromosomal) have not been well characterised. Aim. To characterise the phenotypic and genotypic features as well as the population structure of antimicrobial-resistant E. coli isolated from calves located on dairy farms that feed waste-milk to their replacement calves. Methodology. Recto-anal swab enrichments from 40 dairy calves (≤ 14 days old) located on four dairy farms were examined for tetracycline, streptomycin, ciprofloxacin, and third-generation cephalosporin resistant E. coli . Whole genome sequencing was performed using both short- and long-read technologies on selected antimicrobial resistant E. coli . Results. Fifty-eight percent (23/40) of calves harboured antimicrobial resistant E. coli : 43 % (17/40) harboured tetracycline resistant, and 23 % (9/40) harboured chromosomal mediated AmpC producing E. coli . Whole genome sequencing of 27 isolates revealed five sequence types, with ST88 being the dominant ST (17/27, 63 % of the sequenced isolates) followed by ST1308 (3/27, 11 %), along with the extraintestinal pathogenic E. coli lineages ST69 (3/27, 11 %), ST10 (2/27, 7 %), and ST58 (2/27, 7 %). Additionally, 16 isolates were MDR, harbouring additional resistance genes that were not tested phenotypically. Oxford Nanopore long-read sequencing technologies enabled the location of multiple resistant gene cassettes in IncF plasmids to be determined. Conclusion. Our study identified a high incidence of tetracycline and streptomycin-resistant E. coli in dairy calves, and highlighted the presence of multidrug-resistant strains, emphasising the need for further investigation into potential associations with farm management practices.

Specific pathogen-free ducks are important high-grade laboratory animals, with a key role in research related to poultry biosecurity, production, and breeding. However, the genetic characteristics of experimental duck varieties remain poorly explored. Herein we performed whole-genome resequencing to construct a single nucleotide polymorphism genetic map of the genomes of 3 experimental duck varieties [Jinding ducks (JD), Shaoxing ducks (SX), and Fujian Shanma ducks (SM)] to determine their genetic characteristics and identify selection signatures. Subsequent analyses of population structure and genetic diversity revealed that each duck variety formed a monophyletic group, with SM showing richer genetic diversity than JD and SX. Further, on exploring shared selection signatures, we found 2 overlapping genomic regions on chromosome Z of all experimental ducks, which comprised immune response-related genes (IL7R and IL6ST). Moreover, growth and skeletal development (IGF1R and GDF5), meat quality (FoxO1), and stress resistance (HSP90B1 and Gpx8-b) candidate gene loci were identified in strongly selected signatures specific to JD, SM, and SX, respectively. Our results identified the population genetic basis of experimental ducks at the whole-genome level, providing a framework for future molecular investigations of genetic variations and phenotypic changes. We believe that such studies will eventually contribute to the management of experimental animal resources., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

A triple-mode colorimetric and fluorescent sensing scheme based on manganese dioxide nanoparticles (MnO 2 NPs) and carbon quantum dots (CQDs) were developed to determine nitrite. MnO 2 NPs can oxidize 3,3',5,5'-tetramethylbenzidine (TMB) into a blue oxidation product (TMBox), which is further oxidized into a yellow diimine derivative by nitrite. The ratio of absorbance at 652 nm to 452 nm was monitored as signal response for UV-vis detection mode. A "turn-off" CQDs fluorescence probe was also constructed for fluorescent detection mode. Smartphone tool kit was used to capture the color of sample for smartphone-based measurement. Various analytical performance under different detection modes were obtained and compared. The proposed methods were applied to food samples with satisfactory recoveries (83.3-106 %). The results were validated with AOAC standard spectrophotometric method. The current triple-mode detection were accurate, convenient, low-cost and fast for analyzing nitrite in foods and water samples on-site., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Ovaries are important endocrine organs in female animals that secrete various steroid hormones, which are involved in multiple physiological functions. Estrogen, one of the hormones secreted by ovaries, is essential for the overall maintenance of muscle growth and development. However, the molecular mechanisms that affect muscle growth and development in sheep following ovariectomy remain unclear. In this study, we identified 1662 differentially expressed mRNAs (DEGs) and 40 differentially expressed miRNAs (DEMs) in sheep that underwent ovariectomy compared with those that underwent sham surgery. A total of 178 DEG-DEM pairs were negatively correlated. GO and KEGG analysis showed that PPP1R13B was involved in the PI3K-Akt signaling pathway, which was essential for muscle development. Using in vitro experiments, we examined the effect of PPP1R13B on myoblast proliferation and found that overexpression or inhibition of PPP1R13B increased or decreased the expression of myoblast proliferation markers, respectively. PPP1R13B was identified as a functional downstream target of miR-485-5p. Our results suggested that miR-485-5p promoted myoblast proliferation by regulating proliferation factors in myoblasts by targeting PPP1R13B. Notably, exogenous estradiol supplementation to myoblasts regulated the expression of oar-miR-485-5p and PPP1R13B and promoted myoblast proliferation. These results provided new insights into the molecular mechanism by which ovaries influence muscle growth and development in sheep., Competing Interests: Declaration of competing interest The authors have no conflict of interest to declare. All authors read and approved the final manuscript., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Butyrate promotes the growth and gastrointestinal development of calves. But, the mechanisms behind its effects on signaling pathways of the gastrointestinal tract and rumen microbiome is unclear. This study aimed to reveal transcriptomic pathways of gastrointestinal epithelium and microbial community in response to butyrate supplementation in calves fed a high fiber starter. Fourteen Holstein bull calves (39.9 ± 3.7 kg, 14 d of age) were assigned to 2 groups (sodium butyrate group, SB; control group, Ctrl). The SB group received 0.5% SB supplementation. At d 51, the calves were slaughtered to obtain samples for analysis of the transcriptome of the rumen and jejunum epithelium as well as ruminal microbial metagenome. Sodium butyrate supplementation resulted in a higher performance in average daily gain and development of jejunum and rumen papillae. In both the rumen and jejunum epithelium, SB down-regulated pathways related to inflammation including NF-κB ( PPKCB, CXCL8, CXCL12 ), interleukin-17 ( IL17A, IL17B, MMP9 ), and chemokine ( CXCL12, CCL4, CCL8 ) and up-regulated immune pathways including the intestinal immune network for immunoglobulin A (IgA) production ( CD28 ). Meanwhile, in the jejunum epithelium, SB regulated pathways related to nutritional metabolism including nitrogen metabolism ( CA1, CA2, CA3 ), synthesis and degradation of ketone bodies ( HMGCS2, BDH1 , LOC100295719 ), fat digestion and absorption ( PLA2G2F, APOA1, APOA4 ), and the PPAR signaling pathway ( FABP4, FABP6, CYP4A11 ). The metagenome showed that SB greatly increased the relative abundance of Bacillus subtilis and Eubacterium limosum , activated ruminal microbial carbohydrate metabolism pathways and increased the abundance of carbohydrate hydrolysis enzymes. In conclusion, butyrate exhibited promoting effects on growth and gastrointestinal development by inhibiting inflammation, enhancing immunity and energy harvesting, and activating microbial carbohydrate metabolism. These findings provide new insights into the potential mechanisms behind the beneficial effects of butyrate in calf nutrition., Competing Interests: We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, and there is no professional or other personal interest of any nature or kind in any product, service and/or company that could be construed as influencing the content of this paper., (© 2023 The Authors. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co. Ltd.)

Background: Ruminants can utilize urea as a dietary nitrogen source owing to their ability to recycle urea-N back to the rumen where numerous ureolytic bacteria hydrolyze urea into ammonia, which is used by numerous bacteria as their nitrogen source. Rumen ureolytic bacteria are the key microbes making ruminants the only type of animals independent of pre-formed amino acids for survival, thus having attracted much research interest. Sequencing-based studies have helped gain new insights into ruminal ureolytic bacterial diversity, but only a limited number of ureolytic bacteria have been isolated into pure cultures or studied, hindering the understanding of ureolytic bacteria with respect to their metabolism, physiology, and ecology, all of which are required to effectively improve urea-N utilization efficiency., Results: We established and used an integrated approach, which include urease gene (ureC) guided enrichment plus in situ agarose microsphere embedding and cultivation under rumen-simulating conditions, to isolate ureolytic bacteria from the rumen microbiome. We optimized the dilutions of the rumen microbiome during the enrichment, single-cell embedding, and then in situ cultivation of microsphere-embedded bacteria using dialysis bags placed in rumen fluid. Metabonomic analysis revealed that the dialysis bags had a fermentation profile very similar to the simulated rumen fermentation. In total, we isolated 404 unique strains of bacteria, of which 52 strains were selected for genomic sequencing. Genomic analyses revealed that 28 strains, which were classified into 12 species, contained urease genes. All these ureolytic bacteria represent new species ever identified in the rumen and represented the most abundant ureolytic species. Compared to all the previously isolated ruminal ureolytic species combined, the newly isolated ureolytic bacteria increased the number of genotypically and phenotypically characterized ureolytic species by 34.38% and 45.83%, respectively. These isolated strains have unique genes compared to the known ureolytic strains of the same species indicating their new metabolic functions, especially in energy and nitrogen metabolism. All the ureolytic species were ubiquitous in the rumen of six different species of ruminants and were correlated to dietary urea metabolism in the rumen and milk protein production. We discovered five different organizations of urease gene clusters among the new isolates, and they had varied approaches to hydrolyze urea. The key amino acid residues of the UreC protein that potentially plays critical regulatory roles in urease activation were also identified., Conclusions: We established an integrated methodology for the efficient isolation of ureolytic bacteria, which expanded the biological resource of crucial ureolytic bacteria from the rumen. These isolates play a vital role in the incorporation of dietary nitrogen into bacterial biomass and hence contribute to ruminant growth and productivity. Moreover, this methodology can enable efficient isolation and cultivation of other bacteria of interest in the environment and help bridge the knowledge gap between genotypes and phenotypes of uncultured bacteria. Video abstract., (© 2023. The Author(s).)

Introduction: Urea is an important non-protein nitrogen source for ruminants. In the rumen, ureolytic bacteria play critical roles in urea-nitrogen metabolism, however, a few ureolytic strains have been isolated and genomically sequenced. The purpose of this study was to isolate a novel ureolytic bacterial strain from cattle rumen and characterize its genome and function., Methods: The ureolytic bacterium was isolated using an anaerobic medium with urea and phenol red as a screening indicator from the rumen fluid of dairy cattle. The genome of isolates was sequenced, assembled, annotated, and comparatively analyzed. The pan-genome analysis was performed using IPGA and the biochemical activity was also analyzed by test kits., Results: A gram-positive ureolytic strain was isolated. Its genome had a length of 4.52 Mbp and predicted genes of 4223. The 16S rRNA gene and genome GTDB-Tk taxonomic annotation showed that it was a novel strain of Enterobacter hormaechei , and it was named E. hormaechei Z129. The pan-genome analysis showed that Z129 had the highest identity to E. hormaechei ATCC 49162 with a genome average nucleotide identity of 98.69% and possessed 238 unique genes. Strain Z129 was the first E. hormaechei strain isolated from the rumen as we know. The functional annotation of the Z129 genome showed genes related to urea metabolism, including urea transport ( urt A -urt E), nickel ion transport ( ure J, ton B, nix A, exb B, exb D, and rcn A), urease activation ( ure A-ureG) and ammonia assimilation ( gdh A, gln A, gln B, gln E, gln L, gls A, glt B, and glt D) were present. Genes involved in carbohydrate metabolism were also present, including starch hydrolysis ( amy E), cellulose hydrolysis ( cel B and bgl X), xylose transport ( xyl F -xyl H) and glycolysis ( pgi , pgk , fba A, eno , pfk A, gap , pyk , gpm L). Biochemical activity analysis showed that Z129 was positive for alkaline phosphatase, leucine arylamidase, acid phosphatase, naphthol-AS-BI-phosphohydrolase, α-glucosidase, β-glucosidase, and pyrrolidone arylaminase, and had the ability to use D-ribose, L-arabinose, and D-lactose. Urea-nitrogen hydrolysis rate of Z129 reached 55.37% at 48 h of incubation., Discussion: Therefore, the isolated novel ureolytic strain E. hormaechei Z129 had diverse nitrogen and carbon metabolisms, and is a preferred model to study the urea hydrolysis mechanism in the rumen., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zhong, Zheng, Wang and Zhao.)

Biochanin A (BCA), an isoflavone phytoestrogen, is a secondary metabolite produced mainly in leguminous plants. The objective of this study was to evaluate the effect of BCA on lactation performance, nitrogen metabolism, and the health of dairy goat. Thirty mid-lactation Saanen dairy goats were divided into three groups randomly: control, 2 g/d BCA group, and 6 g/d BCA group. After 36 days of feeding, 30 dairy goats were transferred to individual metabolic cages. Subsequently, milk yield, feed intake, total feces, and urine excretion were recorded and samples were collected continuously for 3 days. Blood and ruminal fluid samples were collected over the subsequent 4 days. Milk yield, milk protein, fat content, and the feed conversion ratio of dairy goat were significantly increased by the BCA treatment. The levels of serum 17β-estradiol, growth hormone, insulin-like growth factor 1, glutathione peroxidase activity, and total antioxidant capacity were also increased significantly by BCA, indicating that BCA enhanced the antioxidant capacity of dairy goat. Amino acid degradation was significantly inhibited, while the ammonia nitrogen content was reduced significantly by BCA. Total volatile fatty acids was significantly increased by BCA supplementation. In addition, the relative abundance of Verrucomicrobiota was decreased significantly. However, the growth of nitrogen metabolism and cellulolytic bacteria was significantly increased under BCA treatment, including Prevotella sp ., Treponema sp. , Ruminococcus flavefaciens , and Ruminobacter amylophilus . In conclusion, supplementation with BCA improved the milk production performance, nitrogen metabolism, rumen fermentation and antioxidant capacity, and regulated the rumen microbiome of dairy goat., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Xu, Li, Du, Zheng, Wang and Zhao.)

Background: Artificial insemination (AI) is an effective reproductive technique to improve the performance of cashmere goats and prevent the spread of diseases, and the quality of the semen determines the success of AI. The potential of Moringa oleifera leaf powder (MOLP) and Moringa oleifera leaf ethanolic extract (MOLE) to improve semen quality has been reported, but the underlying mechanisms remain unclear. For the purpose, 18 mature male cashmere goats were randomly assigned into three groups: the control (CON), MOLP, and MOLE groups. The CON group received distilled water orally; the MOLP group was orally treated with 200 mg/kg body weight (BW) MOLP; and the MOLE group was orally treated with 40 mg/kg BW MOLE., Results: Results showed that MOLE contained long-chain fatty acids and flavonoids. Treatment with MOLP and MOLE increased the activities of the serum catalase, superoxide dismutase, and glutathione peroxidase ( P < 0.05), enhanced the total antioxidant capacity ( P < 0.05), and reduced the serum malondialdehyde level ( P < 0.05). At the same time, MOLE increased the contents of serum gonadotropin releasing hormone and testosterone ( P < 0.05). Moreover, MOLE significantly increased sperm concentration, motility, and viability ( P < 0.05). Meanwhile, MOLE raised the Chao1 index ( P < 0.05) and altered the composition of the rumen microbiota; it also raised the relative abundance of Treponema ( P < 0.05) and Fibrobacter ( P < 0.05) and reduced the relative abundance of Prevotella ( P < 0.1). Correlation analysis revealed the genus Prevotella was significantly negatively correlated with sperm concentration, as well as sperm motility and viability. Furthermore, MOLE significantly increased the rumen levels of the steroid hormones testosterone and dehydroepiandrosterone ( P < 0.05), as well as the polyunsaturated fatty acids (PUFAs) alpha-Linolenic acid, gamma-Linolenic acid, docosapentaenoic acid, and 9-S-Hydroperoxylinoleicacid ( P < 0.05)., Conclusions: Oral MOLE supplementation can improve semen quality by increasing the antioxidant capacity and altering the rumen microbiota and metabolites of cashmere goats. Moreover, the MOLP supplementation could enhance the antioxidant capacity of cashmere goats., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Liang, Wu, Wang, Ma, Li, Zhao, Guo and Liu.)

Background: Stevia straw is a byproduct of sugar crop stevia. It is a good feed material because of richness in nutrients and active substances (steviosides and flavonoids). However, due to improper utilization such as piling, burning and so on, it became a large amount of wasted straw resources and lead to environmental pollution., Methods: We added 0%, 0.2%, 0.4%, 0.6%, 0.8%, 1.0% and 1.5% of stevia stalk to study the effects of different stevia stalk concentrations on nutrient utilization and rumen fermentation in sheep (based on sheep diet). In vitro fermentation method was used, with 17 repetitions for each treatment. All fermentation substrate based on sheep diet with different stevia stalk concentrations were fermented for 2 h, 6 h, 12 h, 24 h and 48 h, then the gas production, dry matter degradability (DMD), crude protein degradability (CPD), neutral detergent fiber degradability (NDFD), acid detergent fiber degradability (ADFD), pH, ammonia nitrogen (NH 3 -N) and volatile fatty acids (VFAs) were determined., Results: The results showed that at different fermentation time, the change trend of gas production in each teatment was basically same, but the maximum occurred in 1.0% treatment at 48 h. The DMD, CPD, NDFD and ADFD of sheep diets increased with fermentation time increasing, especially the CPD 48h , NDFD 48h and ADFD 48h of diets in 0.8%, 1.0% and 1.5% treatments were significantly higher than those in control ( P < 0.05). The pH of fermentation substrate in each treatment remained within the normal range of 6.21∼7.25. NH 3 -N 24h-48h in 0.8%, 1.0% and 1.5% treatments were higher than that in control. At 6 h-12 h, the total acid content of 0.8% and 1.0% treatments were significantly higher than those of other treatments ( P < 0.05), it reached the highest in 1.0% treatment. According to overall evaluation, effect ranking of stevia stalk on sheep nutrient utilization was as follows: 1.0% >0.8% >1.5% >0.4% >0.6% >0.2%. Overall, 1.0% stevia stalk could promote nutrient degradation and sheep rumen fermentation., Competing Interests: The authors declare there are no competing interests., (©2023 Zhang et al.)

The beef quality significantly varies between breeds. Pingliang Red Cattle resembles Wagyu in fat deposition and flavor. To screen key factors affecting beef quality, we performed meat quality trait testing, RNA-seq, and metabolomics on the longissimus dorsi of Pingliang Red Cattle, Wagyu cross F1 generation, and Simmental cattle. The gene and metabolite expression profiles were similar between Pingliang Red Cattle and Wagyu cross F1 generation. Genes such as FASN , ACACA , PLIN1 , and FABP4 were significantly upregulated in the Pingliang Red Cattle and Wagyu cross F1 generation ( P < 0.05). Similarly, numerous metabolites, such as 3-iodo-l-tyrosine, arachidonic acid, and cis -aconitate, which may improve the beef quality such as fat deposition and tenderness, were found in higher levels in the Pingliang Red Cattle and Wagyu cross F1 generation. This study revealed differences in the transcriptional and metabolic levels between Pingliang Red Cattle and premium beef breeds, suggesting that Pingliang Red Cattle harbors the genetic potential for breeding high-grade beef cattle.

The Tibetan pig is an endemic economic animal in the plateau region of China, and has a unique adaptation mechanism to the plateau hypoxic environment. Research into microRNAs (miRNAs) involved in the mechanism underlying hypoxia adaptation of Tibetan pig is very limited. Therefore, we isolated alveolar type II epithelial (ATII) cells from the lungs of the Tibetan pig, cultured them in normoxia/hypoxia (21% O 2 ; 2% O 2 ) for 48 h, and performed high-throughput sequencing analysis. We identified a hypoxic stress-related ssc-miR-141 and predicted its target genes. The target genes of ssc-miR-141 were mainly enriched in mitogen-activated protein kinase (MAPK), autophagy-animal, and Ras signaling pathways. Further, we confirmed that PDCD4 may serve as the target gene of ssc-miR-141. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis was performed to confirm the expression levels of ssc-miR-141 and PDCD4 , and a dual-luciferase gene reporter system was used to verify the targeted linkage of ssc-miR-141 to PDCD4 . The results showed that the expression level of ssc-miR-141 in the hypoxia group was higher than that in the normoxia group, while the expression level of PDCD4 tended to show the opposite trend and significantly decreased under hypoxia. These findings suggest that ssc-miR-141 is associated with hypoxia adaptation and provide a new insight into the role of miRNAs from ATII cells of Tibetan pig in hypoxia adaptation.

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.

To study the effects of adding quinoa seed (raw grain) to the diet of the Luhua chicken on the cecal microorganism abundance and fatty acid composition of muscle, 120 49-day-old healthy female dewormed Luhua chickens (body weight 1476.21 ± 101.39 g) were randomly divided into 4 groups, with 3 replicates in each group and 10 chickens in each repetition. The control group (CK group) was fed a basal diet and the experimental groups were fed with 4% (Q4), 8% (Q8), and 12% (Q12) quinoa seed (raw grain) added to the basal diet for 75 days. After 121 days of age, the animals were slaughtered and the 16S rRNA characteristics of cecal flora, as well as composition and content of fatty acids in muscle, were determined and analyzed. The content of unsaturated fatty acids (UFAs), docosahexaenoic acid (C22:6n3; DHA) and n-3 polyunsaturated fatty acids (n-3 PUFAs) in the breast and leg muscles significantly increased in the experimental groups supplemented with quinoa seeds (p < 0.05). However, the content of saturated fatty acids (SAFs) and ratio of n-6/n-3 in breast muscle and leg muscle significantly decreased (p < 0.05). In addition, adding a certain percentage of quinoa seeds in the diet can also affect the community composition and content of microorganisms in the ceca of Luhua chickens. At the phylum level, the Proteobacteria, Actinobacteria, Synergistetes and Melainabacteria in experimental groups (Q4, Q8 and Q12) were significantly lower than those in the CK group (p < 0.05). At the genus level, Desulfovibrio, Synergistes, Olsenella, Parabacteroides, Mailhella, Sutterella and Ruminiclostridiu in group Q4 were significantly lower than those in group CK (p < 0.05) while Faecalibacterium in Q8 group, and Lawsonia and Faecalibacterium in Q12 group were significantly higher than those in the CK group (p < 0.05). Enrichment analysis of the microbial function showed that compared with the CK group, Metabolism and Enzyme Families were significantly enriched in the Q4 group (p < 0.05). Cellular Processes and Signaling were significantly enriched in the Q8 group (p < 0.05). The association analysis of fatty acids with microorganisms showed that the abundance of Faecalibacterium, Lawsonia and Meagmonas was significantly correlated with partial SFAs and UFAs (p < 0.05). In conclusion, adding quinoa seeds to diets significantly increased the content of muscle DHA, UFAs and n-3 PUFAs. The content of SAFs and the n-6/n-3 ratio were significantly reduced. Taken together, quinoa can effectively improve the cecal microbiota structure, inhibit the number of harmful bacteria and increase the number of beneficial bacteria, regulating the intestinal environment and promoting the body health of female Luhua chickens.

This experiment was conducted to investigate the effects of oregano essential oil on the cecal microorganisms and muscle fatty acids of Luhua chickens. One hundred and twenty 49-day-old healthy dewormed Luhua chickens were randomly divided into four groups with three replicates per group and ten chickens per replicate. The corn−quinoa and soybean meal diets were supplemented with 0 (Q8 group), 50 (QO50 group), 100 (QO100 group) and 150 mg·kg−1 (QO150 group) of oregano essential oil, respectively, and the experiment lasted for 75 days. The composition of intestinal flora was detected by Illumina sequencing of the 16S rRNA V4 region, and the composition and content of fatty acids in the muscles were analyzed by gas chromatography. The results showed that dietary oregano essential oil can effectively increase the contents of elaidic acid (C18:ln9t), polyunsaturated fatty acids (PUFAs) and n-3 polyunsaturated fatty acids (n-3 PUFAs) in breast muscle tissues. However, the fatty acid composition and PUFA content in leg muscle tissues were not significantly improved. According to a 16S rRNA high-throughput sequencing analysis, dietary oregano essential oil supplementation with a certain concentration can change the cecal microbial community composition of broilers. At the phylum level, Elusimicrobia in the QO150 group was significantly lower than that in Q8 group (p < 0.05). At the genus level, Phascolarctobacterium, Parasutterella and Bilophila in the experimental groups (QO50, QO100 and QO150) were significantly lower than those in the Q8 group (p < 0.05). An enrichment analysis of the microbial function found that the amino acid metabolism, energy metabolism, metabolism, signal transduction and genetic information processing were mainly enriched in the experimental groups, which promoted the digestion and absorption of nutrients and enhanced intestinal barrier functioning. An analysis of the association between fatty acids and microbes found that the abundance of microbiota was significantly correlated with partially saturated fatty acids (SFAs) and unsaturated fatty acids (UFAs) (p < 0.05). In conclusion, the dietary addition of oregano essential oil can effectively improve cecal microbial community composition, promote the digestion and absorption of nutrients, and enhance intestinal barrier functioning. It can significantly improve the content of some fatty acids, and there was a certain correlation between caecum microorganisms and fatty acid deposition in muscles.

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.

Cysteamine (CS), as a feed supplement, can increase the level of growth hormone (GH) in the blood, promote animal growth. However, little attention has been paid to the effects of CS on the rumen microbiome and metabolic profile in cashmere goats. This study aimed to assess the effects of rumen microbiota, metabolites, and plasma antioxidative capacity induced by CS supplementation in cashmere goats. We selected 30 Inner Mongolia white cashmere goat ewes (aged 18 months), and randomly separate the goats into three groups ( n = 10 per group) to experiment for 40 days. Oral 0 (control group, CON), 60 (low CS, LCS), or 120 mg/kg BW -1 (high CS, HCS) coated CS hydrochloride every day. Using 16S and internal transcribed spacer (ITS) rRNA gene amplicon sequencing, we identified 12 bacterial and 3 fungal genera with significant changes among the groups, respectively. We found a significant increase in rumen NH 3 -N and total volatile fatty acid (TVFA) concentrations in the LCS and HCS groups compared with the CON. With untargeted LC-MS/MS metabolomics, we screened 59 rumen differential metabolites. Among the screened metabolites, many unsaturated and saturated fatty acids increased and decreased with CS treatment, respectively. CS supplementation increased the levels of plasma total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), GH, and insulin-like growth factor-1(IGF-1). Spearman correlation analysis revealed that the abundance of U29-B03, Lactococcus , and Brochothrix were positively associated with the levels of δ2-THA, TVFA and antioxidant capacity. In conclusion, CS significantly affected rumen microbiota and fermentation parameters, and ultimately inhibited the biohydrogenation of rumen metabolites, enhanced plasma antioxidant capacity, and regulated some hormones of the GH-IGF-1 axis. This study provides an overall view into the CS application as a strategy to improve health production in cashmere goats., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wu, Liang, Wang, Zhao, Ma, Gao, Zhao, Chen and Liu.)

Understanding the signaling pathway regulatory mechanisms in type II alveolar epithelial (ATII) cells, the progenitor cells responsible for proliferating and regenerating type I alveolar epithelial (ATI) and ATII cells, in Tibetan pigs is beneficial for exploring methods of preventing and repairing cellular damage during hypoxia. We simulated a hypoxic environment (2% O 2 ) for culture ATII cells of Tibetan pigs and Landrace pigs, with cells cultured under normoxic conditions (21% O 2 ) as a control group, and performed integrated analysis of circular RNA (circRNA)-microRNA (miRNA)-messenger RNA (mRNA) regulatory axes by whole-transcriptome sequencing. Functional enrichment analysis indicated that the source genes of the differential expressed circRNAs (DEcircRNAs) were primarily involved in cell proliferation, cellular processes, and cell killing. A series of DEcircRNAs were derived from inhibitors of apoptosis proteins and led to a key autonomous effect as modulators of cell repair in Tibetan pigs under hypoxia. The significant higher expression of COL5A1 in TL groups may inhibited apoptosis of ATII cells in Tibetan pigs under lower oxygen concentration, and may lead their better survive in the hypoxia environment. In addition, a competing endogenous RNA (ceRNA) network of functional interactions was constructed that included novel&#95;circ&#95;000898-ssc-miR-199a-5p- CAV 1 and novel&#95;circ&#95;000898-ssc-miR-378- BMP 2, based on the node genes ssc-miR-199a-5p and ssc-miR-378, which may regulate multiple miRNAs and mRNAs that mediate endoplasmic reticulum (ER) stress-induced apoptosis and inflammation and attenuate hypoxia-induced injury in ATII cells under hypoxic conditions. These results broaden our knowledge of circRNAs, miRNAs, and mRNAs associated with hypoxia and provide new insights into the hypoxic response of ATII cells in Tibetan pigs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yang, Li, Yuan, Liu, Ren, Gao, Jiao, Cai and Zhao.)

Alternative splicing (AS) allows the generation of multiple transcript variants from a single gene and affects biological processes by generating protein diversity in organisms. In total, 41,642 AS events corresponding to 9,924 genes were identified, and SE is the most abundant alternatively spliced type. The analysis of functional categories demonstrates that alternatively spliced differentially expressed genes (DEGs) were enriched in the MAPK signaling pathway and hypoxia-inducible factor 1 (HIF-1) signaling pathway. Proteoglycans in cancer between the normoxic (21% O 2 , TN and LN) and hypoxic (2% O 2 , TL and LL) groups, such as SLC2A1, HK1, HK2, ENO3 , and PFKFB3 , have the potential to rapidly proliferate alveolar type II epithelial (ATII) cells by increasing the intracellular levels of glucose and quickly divert to anabolic pathways by glycolysis intermediates under hypoxia. ACADL, EHHADH , and CPT1A undergo one or two AS types with different frequencies in ATII cells between TN and TL groups (excluding alternatively spliced DEGs shared between normoxic and hypoxic groups), and a constant supply of lipids might be obtained either from the circulation or de novo synthesis for better growth of ATII cells under hypoxia condition. MCM7 and MCM3 undergo different AS types between LN and LL groups (excluding alternatively spliced DEGs shared between normoxic and hypoxic groups), which may bind to the amino-terminal PER-SIM-ARNT domain and the carboxyl terminus of HIF - 1 α to maintain their stability. Overall, AS and expression levels of candidate mRNAs between Tibetan pigs and Landrace pigs revealed by RNA-seq suggest their potential involvement in the ATII cells grown under hypoxia conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yuan, Liu, Wang, Ren, Li, Gao, Jiao, Cai, Yang and Zhao.)

Pseudomonas spp. are common microorganisms from cold-storage raw milk, and protease secreted by Pseudomonas spp. can cause the deterioration of stored milk. However, analyses of Pseudomonas spp. diversity and proteolytic activity in raw milk from different regions of China have not been extensively examined. With this aim, the diversity and proteolytic activity of Pseudomonas isolated from 25 raw cow milk samples from Inner Mongolia, Heilongjiang, Gansu, Henan, Anhui, Jiangsu, Chongqing and Hunan of China in different seasons were evaluated by PCR targeting 16S rDNA and rpoD, as well as TNBS method, respectively. A total of 116 Pseudomonas isolates from 25 raw cow milk samples were identified at the species level, including P. fluorescens, P. veronii, P. psychrophila, P. lundensis, P. lactis, P. azotoformans, P. granadensis, P. lurida, P. rhizosphaerae, P. rhodesiae and P. extremorientalis. P. fluorescens accounted for 75.8% of the total. Of all 116 Pseudomonas isolates, 68.9% of them displayed proteolytic activity at 4 °C, 81.9% at 10 °C and 85.3% at 25 °C, respectively. The aprX gene encoded a secreted and heat-resistant metalloprotease that was present in 60.3% of the Pseudomonas isolates tested. The proteases showed residual activity ranged from 73 ± 4% to 84 ± 7% residual activity after the heat treatment at 72 °C for 15 s and 62 ± 3% to 74 ± 2% after the heat treatment at 132 °C for 4 s. This is the first report to compare Pseudomonas spp. diversity and proteolytic activity at species levels in raw milk from different regions of China. The results of this study provide valuable data about the diversity and spoilage potential of Pseudomonas species in raw milk and the thermal resistance of the proteases. Therefore, these findings provide a reference for the importance to prevent Pseudomonas spp. contamination of raw cow milk to ensure the quality and safety of milk and dairy products., Competing Interests: Declaration of competing interest All authors have approved the submission and none declare any conflict of interest in the work performed or in the submission of the manuscript., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Increasing studies have demonstrated potential disproportionate functional and ecological contributions of rare taxa in a microbial community. However, the study of the microbial rare biosphere is hampered by their inherent scarcity and the deficiency of currently available techniques. Sample-wise cross contaminations might be introduced by sample index misassignment in the most widely used metabarcoding amplicon sequencing approach. Although downstream bioinformatic quality control and clustering or denoising algorithms could remove sequencing errors and non-biological artifact reads, no algorithm could eliminate high quality reads from sample-wise cross contaminations introduced by index misassignment, making it difficult to distinguish between bona fide rare taxa and potential false positives in metabarcoding studies., Results: We thoroughly evaluated the rate of index misassignment of the widely used NovaSeq 6000 and DNBSEQ-G400 sequencing platforms using both commercial and customized mock communities, and observed significant lower (0.08% vs. 5.68%) fraction of potential false positive reads for DNBSEQ-G400 as compared to NovaSeq 6000. Significant batch effects could be caused by stochastically introduced false positive or false negative rare taxa. These false detections could also lead to inflated alpha diversity of relatively simple microbial communities and underestimated that of complex ones. Further test using a set of cow rumen samples reported differential rare taxa by different sequencing platforms. Correlation analysis of the rare taxa detected by each sequencing platform demonstrated that the rare taxa identified by DNBSEQ-G400 platform had a much higher possibility to be correlated with the physiochemical properties of rumen fluid as compared to NovaSeq 6000 platform. Community assembly mechanism and microbial network correlation analysis indicated that false positive or negative rare taxa detection could lead to biased community assembly mechanism and identification of fake keystone species of the community., Conclusions: We highly suggest proper positive/negative/blank controls, technical replicate settings, and proper sequencing platform selection in future amplicon studies, especially when the microbial rare biosphere would be focused., (© 2022. The Author(s).)

The objective of this study was to determine the effect of whole flaxseed and ground flaxseed supplementation on the composition of fatty acids in plasma and milk, particularly the content of omega-3 polyunsaturated fatty acids (n-3 PUFAs). Thirty Holstein dairy cows were randomly assigned to three treatment groups. Cows were fed a total mixed ration without flaxseed (CK), 1500 g of whole flaxseed (WF), and 1500 g of ground flaxseed (GF) supplementation. There were no differences observed in dry matter intake, milk yield, energy-corrected milk, and 4% fat-corrected milk (p > 0.05). Compared with the CK group, the contents of α-linolenic acid (ALA), eicosatrienoic acid, and eicosapentaenoic acid increased in the plasma and milk WF and GF groups, and the content of docosahexaenoic acid and total n-3 PUFA was higher in GF than the other groups (p < 0.001). The ALA yield increased to 232% and 360% in WF and GF, respectively, compared to the CK group. Compared with the WF group, GF supplementation resulted in an increased milk ALA/ALA intake ratio (p < 0.001). Flaxseed supplementation increased the activity of GSH-Px and decreased the concentration of MDA in milk (p < 0.001). Plasma parameters did not differ among the treatments (p > 0.05). This result indicated that compared with the WF group, GF supplementation in the diet showed higher efficiency in increasing the total n-3 PUFA levels and the milk ALA/ALA intake ratio, and decreased the ratio of n-6 PUFAs to n-3 PUFAs in milk.

We compared the differences in testicular morphology and vascular anatomy between plateau-type Tibetan sheep and low-altitude Small-tailed Han sheep to explore any adaptations of Tibetan sheep to high altitudes and hypoxia factors. Testicular artery architecture specimens of plateau-type Tibetan sheep and lowland Small-tailed Han sheep were used to compare the physical parameters of the testis, testicular artery architecture, arterial diameter, testicular leaflets and the characteristics of epididymal head arterioles. The results showed that the weight, major axis, minor axis, overall length, caput major axis and caput minor axis in Tibetan sheep were larger than those of Small-tailed Han sheep; however, the caput major axis of epididymal head in Tibetan sheep was smaller than that of Small-tailed Han sheep. Compared with Small-tailed Han sheep, the testicular knot-like arteries of Tibetan sheep had denser spiral distribution and more small arteries, and the diameters of centripetal arteries, knot-like arteries, centrifugal arteries and epididymal microarterioles were also thicker. The 'spindle' indentation on the surface of the microarteriole of the head of the testicular leaflets and the epididymis in plateau Tibetan sheep was relatively shallow. The results showed that the physical testicular characteristics, testicular artery construction and arteriole surface structure of Tibetan sheep were adapted for the plateau environment., (© 2022 Wiley-VCH GmbH.)

Background: Milk microRNA (miRNA) with bioactivity is beneficial for human health. However, the effect of heat treatment on miRNA in milk is still not clear. In this study, the miRNAs in raw (RM), pasteurized (PM) and ultra-high-temperature (UHT) milk (UM) from the same batch were extracted, sequenced and analyzed., Results: The results showed that there was a significant difference in miRNAs between RM and UM, but not between RM and PM. The total read counts of milk miRNAs were significantly decreased by heat treatment, with the least counts in UM (P < 0.05). The average length and GC percentage of miRNAs were significantly reduced by heat treatment (P < 0.05), while there was no significant difference in these terms between RM and PM. The content of miRNAs was verified by qPCR, finding that miR-17-5p, miR-25, miR-27b and miR-9-5p were significantly reduced in UM (P < 0.05) but not significantly affected in PM (except miR-27b). In addition, the targeting gene ontology enrichment functions of the different presented miRNAs were mostly enriched in biological process, cellular component and molecular function. The top 20 enriched miRNAs with different levels in heat-treated milk were identified by the Kyoto Encyclopedia of Genes and Genomes enrichment analysis. Interestingly, most of the functions of these miRNA targeting genes are involved in cancer or inflammation activity., Conclusion: This study revealed that the bioactive miRNA in RM was lost after UHT treatment but not in pasteurized treatment. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

Milk is rich in fat, protein, minerals, vitamins, peptides, immunologically active substances, and other nutrients, and it plays an important role in satisfying human nutrition and health. However, dairy product safety incidents caused by microbial contamination have occurred. We found that the total bacterial numbers in the pasteurized product were low and far below the limit requirements of the food safety standards of the European Union, the United States, and China. At the genus level, the primary microbial groups found in milk samples were Acinetobacter, Macrococcus, Pseudomonas , and Lactococcus , while in the equipment rinse water and air samples there was contamination by Stenotrophomonas and Acinetobacter . The Source Tracker model analysis indicated that the microorganisms in the final milk products were significantly related to the contamination in product tanks and raw milk. Therefore, it is the hope that this work can provide guidance to pinpoint contamination problems using the proper quality control sampling at specific stages in the pasteurization process., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Du, Meng, Wu, Yang, Liu, Zheng, Zhang, Zhao and Wang.)