During the estrous cycle of mammals， a large number of follicles are recruited. Typically， only one follicle is selected for dominance and ovulation in monotocous （single-offspring） species per cycle， while only a few follicles are selected for dominance in polytocous （multiple-offspring） species. The vast majority of the remaining follicles undergo atresia. In recent years， significant progress has been made in research focusing on follicular dominance selection， although the underlying mechanisms have not yet been fully elucidated. Insulin-like growth factor 1 （IGF-1） has been demonstrated to play a crucial role in follicular development. This review summarizes the roles and mechanisms of IGF-1 in mammalian follicular development， focusing on its involvement in dominant follicle selection， promotion of follicular growth and development， inhibition of granulosa cell apoptosis， and maintenance of normal reproductive performance in female mammals.

Ovarian folliculogenesis is fundamental to the reproductive activity of female mammals， encompassing the formation of primordial follicles， follicular recruitment， dominant follicle selection， ovulation of mature follicles， and the follicular atresia process that persists throughout development. Ferroptosis， a novel form of regulated cell death， plays a critical role in follicular development， particularly during atresia. This review systematically explores the molecular mechanisms of ferroptosis， emphasizing its impact on follicular atresia， primordial follicle activation， and oocyte maturation， as well as the regulatory effects of antioxidants on ferroptosis. The aim is to provide a theoretical reference for further research in this domain.

Animal mammary bioreactors utilize mammalian-specific lactoprotein gene promoters to direct the efficient and specific expression of exogenous genes in the animal mammary gland， ultimately obtaining protein expression products through the secretory function of the mammalian mammary gland. This article focuses on the application progress and research prospects of animal mammary bioreactors， providing theoretical reference value for future research and industrial development of animal mammary bioreactors. Through literature review and practical research， a brief overview is provided from the aspects of the principle， characteristics， target animal selection， application fields， challenges faced， and development prospects of animal mammary bioreactors. Different animal mammary bioreactors have their own advantages and disadvantages， and target animals can be reasonably selected according to actual needs and production practices. Compared with other traditional expression systems， mammary bioreactors are a novel expression mode with advantages such as high yield， easy purification， low cost， high biological activity， and high safety， and are widely used in improving animal milk quality， producing recombinant pharmaceutical proteins， making antibodies， synthesizing genetic engineering vaccines， and other fields. Although many influencing factors of mammary bioreactors are still unknown， there are some problems that urgently need to be solved， and challenges are also faced. However， with the rise of gene editing technologies such as CRISPR/Cas9， as long as technological innovation and research and development investment are continuously increased， their application prospects are very broad.

Tetraploid embryo technology is an important tool for livestock genetic breeding and biomedical research， and its application potential is becoming more and more prominent. In this paper， we systematically review the key technologies， bottlenecks and future development directions of tetraploid in vitro embryo production in livestock. It focuses on the induction methods （e.g.， physical， chemical， and biological methods）， optimization of in vitro culture systems， and factors affecting the developmental efficiency of tetraploid embryos of major livestock species， such as pigs， cows， and sheep. By comparing the developmental characteristics of tetraploid embryos from different species， we reveal the core limiting factors such as abnormal activation of the embryonic genome and defective placental development. Combined with the cutting-edge technologies of gene editing and embryo-like culture， the application of this echnology in the fields of efficient breeding， disease modeling and xenotransplantation is envisioned， with a view to providing theoretical references and technological guidance for related research.

Excessive fat accumulation in the ovine caudal region compromises feed efficiency and reproductive performance， consequently elevating production costs and diminishing producer incomes. Furthermore， tail docking fails to mitigate caudal adiposity； instead， it promotes ectopic fat deposition within the gluteal musculature. Consequently， the development of thin-tailed sheep populations through genome-wide selection has emerged as a critical objective for sheep breeders， and the key to addressing this challenge lies in identifying genes associated with ovine tail phenotypes. This review systematically synthesizes current knowledge on the functions of key genes （e.g. BMP2， PDGFD， and TBXT） and their epigenetic regulatory mechanisms involved in regulating sheep tail types， focusing on both adipogenesis and tail vertebrae length. Additionally， it explores the conservation value of fat-tailed sheep as a unique genetic resource. These findings provide a theoretical foundation for the genomic selection and breeding of sheep breeds with specific tail conformations.

Methionine （Met）， as an essential amino acid for ruminants， plays a critical role in production performance， reproductive efficiency， immune function， and regulation of the rumen environment. However， dietary Met is susceptible to degradation by rumen microbes， leading to insufficient absorption in the small intestine. To address this， rumen-protected methionine （RPM） is developed through coating technology or chemical modification， effectively reducing rumen degradation rates and enhancing its utilization in the small intestine. Supplementation with RPM improves ruminant production performance， optimizes rumen microbial flora， enhances nitrogen balance， boosts reproductive performance， regulates immune gene expression， and strengthens antioxidant capacity. This article systematically reviews the technological principles of RPM and its application progress in ruminant production， providing a theoretical foundation for the practical implementation of RPM.

The rapid development of large-scale and intensive livestock and poultry farming has led to a significant increase in the frequency of heat stress， which results in a series of harms to the production performance and physical health of livestock and poultry. Studies have found that the body damages of livestock and poultry caused by heat stress are closely related to endotoxins. Under heat stress conditions， the increase in peripheral blood volume leads to insufficient blood supply for the gastrointestinal tract， resulting in damages to its barrier function. Consequently， an increasing translocation of bacterial endotoxins into the blood system would lead to damages to various organs of the body. This article reviews the generation of endotoxin in livestock and poultry under heat stress conditions， its damage to the bodies of livestock and poultry under heat stress conditions， and the research progresses on the relevant body damages and mechanisms， with the aim of providing an innovative reference for the prevention and control of endotoxins hazards in livestock and poultry exposed to heat stress.

Bile acids， which are significant metabolites， are generated by the liver and conveyed to the intestine. Microorganisms in the intestine modify and process bile acids into secondary bile acids. Historically， it was believed that bile acids were engaged in the absorption and metabolism processes of lipids and fat-soluble vitamins. An increasing number of studies have shown that bile acids can act as multiple signaling metabolites， which are essential for maintaining bile acid homeostasis， enterohepatic circulation， metabolism， intestinal barrier integrity， and intestinal mucosal immune homeostasis by regulating bile acid-related receptors and interactions with the microbiota. Unique lymphocyte populations cooperate with each other to jointly maintain the stability of the intestinal immune system. Among these elements， the equilibrium between pro-inflammatory T helper 17 cells （T_H17） and anti-inflammatory regulatory T cells （T_reg） holds particular significance. It is of great importance in safeguarding the homeostasis of the intestinal immune system. This article reviews the complex relationship and latest research progress between bile acids， intestinal microorganisms and intestinal mucosal immunity from the following three aspects： the interaction between bile acids and intestinal microorganisms， bile acid-related receptors， and the balance between T_H17/T_reg cells， in order to provide a theoretical basis for revealing the physiological functions of bile acids and their application in animal production.

The intestine， as the organ responsible for food digestion and nutrient absorption within the body， its structural and functional integrity is of critical importance for health maintainance. The brush border is an essential intestinal structure that， through its highly ordered array of microvilli， greatly expands the absorptive surface area and plays a central role in mediating nutrient transport， maintaining epithelial homeostasis， and forming a physical barrier defense. A clear understanding of the brush border’s microstructure is of great value for both basic and clinical research related to intestinal function. This article systematically reviews the structural components of the brush border and the ultrastructure of microvilli， with a focus on the actin core bundles within microvilli； the cross-linking roles of bundling proteins such as Fimbrin， Villin， and Espin-1； the mechanical stabilization and dynamic regulation provided by the terminal web through proteins including Calmodulin， Spectrin， and Myosin； and the regulatory function of the intermicrovillar adhesion complex in the ordered assembly of microvilli. By offering a systematic overview of microvillar architecture and functional proteins， this work provides an important theoretical basis for deep understanding the assembly and regulatory mechanisms of small-intestinal microvilli and their targeted modulation in intestinal diseases.

Bisphenol A （BPA） is an environmental endocrine disrupting chemical which widely used in plastic production. Due to the large-scale production and use of BPA， it poses a threat to human and animal health. This article systematically sorts out the research results on the toxic effects of BPA at home and abroad in recent years， and comprehensively reviews the sources and routes of BPA exposure， its metabolic transformation in animals， as well as the progress of its toxic effects on livestock and poultry health. It will help to improve the understanding of the toxicity of BPA， and is expected to provide valuable reference materials for in-depth studies on the animal toxicity mechanism of BPA and prevention of the hazards of BPA in livestock and poultry farming.

Ferroptosis， a biologically significant form of regulated cell death， participates in the pathological processes of various metabolic diseases and toxicoses. The occurrence of ferroptosis adversely impacts animal production performance and may even lead to mortality， resulting in substantial economic losses for livestock farmers. Consequently， identifying effective functional additives to mitigate or block ferroptosis is of paramount importance. Plant extracts， defined as secondary metabolites isolated from plant tissues through standardized processes， typically contain bioactive compounds with well-documented pharmacological or regulatory functions， including alkaloids， flavonoids， polyphenols， and polysaccharides. These natural constituents exhibit regulatory effects on iron ion homeostasis， glutathione metabolism， and inhibition of lipid peroxidation， thereby counteracting ferroptosis. This review summarizes the pathogenic mechanisms of ferroptosis and the modulatory roles of plant extracts in its regulation， aiming to provide novel insights for the development of functional additives in animal husbandry.

With global population growth and rising consumption，the large-scale expansion of animal husbandry has become an irreversible trend. However， alongside ensuring food supply， the rapid development of livestock and poultry farming has also brought about severe ecological and environmental problems. Notably， the farming system has become both an important source and sink of emerging pollutants. As an emission source， the farming process continuously releases emerging pollutants such as Antibiotics， persistent organic pollutants， endocrine disruptors， microplastics， pesticides. As a sink， these pollutants accumulate in livestock through the food chain， creating a multi-level amplification effect that poses significant threats to ecosystems and human health. This paper reviewed the main types and sources of emerging pollutants in livestock farming， their occurrence characteristics and environmental behaviors， analyzed their potential impacts on the environment and human health. We further discussed the pollutants management idea of reduction at the source，interception during the process， and purification at the end. We also summarized the current research gaps，and outlined future research directions. This paper aims to provide a scientific basis for China's emerging pollutant control system construction and policy optimization，and to promote the sustainable development of the livestock industry.

Clostridium novyi is an opportunistic pathogen widely distributed in nature，primarily infects cattle and sheep，causing a range of acute diseases such as infectious necrotic hepatitis （black disease）， septicemia， and gas gangrene， which severely hindered the development of global livestock industries. This review systematically summarizes the pathogenic mechanisms and prevention and control techniques of Clostridium novyi type B， and conducts a comparative evaluation of the principles， advantages， and limitations of associated vaccines. The analysis aims to provide both theoretical foundations and methodological insights for advancing mechanistic studies and optimizing prevention practices against Clostridium novyi type B infections.

Salmonella poses a significant threat as a foodborne pathogen impacting global public health and the poultry industry. The escalating challenge of antibiotic resistance has intensified the difficulties in its control and prevention. Bacteriophage （phage） therapy， characterized by its high host specificity and intrinsic self-replication capability， has emerged as a promising novel biocontrol strategy to replace or complement antibiotics. Currently， several phage-based products targeting Salmonella are commercially deployed in livestock farming and poultry product processing. Extensive research and field trials have demonstrated their efficacy in reducing Salmonella loads within live poultry， decreasing mortality rates， enhancing production performance， and significantly reducing Salmonella contamination on poultry meat and eggs. Despite the promising potential， further field trials within commercial farm environments and robust clinical studies are imperative to evaluate clinical efficacy comprehensively， optimize application protocols， and promote standardized implementation.

Porcine epidemic diarrhea virus （PEDV） is a major pathogen causing diarrhea in piglets and has led to significant economic losses in the global swine industry since its emergence. In practice， co-infection with multiple pathogens is commonly observed alongside PEDV， complicating its prevention and control. These co-infecting factors mainly include various viruses， bacteria， parasites， and toxins， which can exacerbate clinical symptoms in infected pigs. This review systematically summarizes the factors commonly co-infecting with PEDV and their impact on its pathogenicity. The results show that porcine kobuvirus （PKV）， porcine Sapelovirus （PSV）， porcine deltacoronavirus （PDCoV）， porcine rotavirus （PRoV） and transmissible gastroenteritis virus （TGEV） are the most common porcine diarrheal viruses co-infecting with PEDV， with PKV and PSV representing emerging trends in recent years. Among bacteria， Escherichia coli， C. perfringens， and C. difficile are frequently detected in co-infections with PEDV. Additionally， parasites such as Entamoeba spp. and various toxins have also been confirmed to co-infect with PEDV. In terms of pathogenicity， PEDV co-infection can exacerbate clinical symptoms， increase PEDV load in the intestine， cause more severe intestinal barrier damage， and trigger a stronger immune response in pigs. These findings lay a foundation for future studies on the mechanisms of PEDV co-infection. Moreover， this review analyzes current prevention and control strategies and identifies existing limitations， aiming to raise awareness of PEDV co-infection among the livestock industries and to promote the implementation of scientific and sustainable control measures， thereby supporting healthier pig production， and improving overall farming efficiency.

Bovine mastitis is an inflammatory response of the mammary tissue triggered by pathogenic microorganism infections， which severely impairs dairy production performance and compromises milk product safety， leading to substantial economic losses in the livestock industry. Consequently， elucidating its pathogenesis is of critical importance for prevention and treatment strategies. Conventionally， mastitis has been attributed primarily to exogenous bacterial infections or physical trauma； however， recent studies reveal that gastrointestinal microbiota and their metabolites can systemically influence mammary health via the "gut-mammary axis，" offering novel insights into the disease’s mechanistic underpinnings. Building on prior research， this review systematically summarizes the pathogenesis of bovine mastitis and highlights the escalating challenges posed by traditional antibiotic therapies due to rising antimicrobial resistance and disruption of mammary microbiota. In parallel， herbal medicines have emerged as a promising research focus， exhibiting distinctive advantages in mastitis prevention and treatment owing to their multi-component， multi-target synergistic actions and regulatory effects on the gut-mammary axis. Starting from the inflammatory cascade and immune regulation mechanisms initiated by pathogenic infections， this article focuses on the multi-pathway mechanisms through which herbal medicines exert therapeutic effects—by targeting pathogens， modulating inflammatory responses， and regulating immune functions—thereby offering innovative perspectives for the green prevention and control of bovine mastitis.

This study aimed to investigate the regulatory role of 6-phosphofructo-2-kinase/fructose-2，6-bisphosphatase 3 （PFKFB3） in the proliferation and adipogenic differentiation of porcine intramuscular preadipocytes， screen its interacting proteins and provide clues for understanding their regulatory mechanisms. PFKFB3 was overexpressed or interfered in porcine intramuscular preadipocytes. qRT-PCR was used to detect changes in the expression of proliferation marker genes. CCK-8 and EdU assays were employed to measure changes in the number of viable cells and cells in the proliferation phase， respectively. After inducing adipogenic differentiation of porcine intramuscular preadipocytes， qRT-PCR and Oil Red O staining were used to assess its regulatory effect on adipogenic differentiation. Co-immunoprecipitation combined with liquid chromatography-tandem mass spectrometry was utilized to identify and screen proteins interacting with PFKFB3. The results showed that overexpression of PFKFB3 significantly increased the number of EdU-positive cells （P<0.01） and cell viability （P<0.01）， and significantly upregulated the expression of proliferation-related genes CDK1， CCND1， PCNA， and KI67 （P<0.05）. Conversely， interfering with PFKFB3 significantly inhibited cell proliferation （P<0.05）. Regarding adipogenic differentiation， overexpression of PFKFB3 significantly increased the number of lipid droplets （P<0.01） and triglyceride content （P<0.01）， and significantly upregulated the expression of key adipogenic genes PPARγ， CEBPα， CEBPβ， and LPL （P<0.05）. Interfering with PFKFB3 significantly inhibited adipogenic differentiation （P<0.05）. Furthermore， Flag-tag immunoprecipitation combined with mass spectrometry analysis identified 17 specific interacting proteins of PFKFB3. GO analysis indicated that these proteins were primarily involved in processes such as ATP binding and RNA binding. STRING protein-protein interaction network prediction revealed interactions between PFKFB3 and proteins including CHORDC1， ALAS1， ASCC3， and KCNK2. In summary， PFKFB3 positively regulates the proliferation and adipogenic differentiation of porcine intramuscular preadipocytes. Potential interacting proteins of PFKFB3 were preliminarily screened， providing new clues for in-depth analysis of the molecular mechanisms underlying porcine intramuscular fat deposition and improvement of pork quality.

The aim of this study was to develop different methods for screening genetic variants by integrating genomic data， phenotypic information， and functional annotations， and to compare and evaluate the prediction accuracy of 8 genomic selection （GS） methods for complex traits. Genotype-phenotype data from 542 Beijing ducks and 2 548 Duroc pigs were analyzed using three dimensions of variant selection： 1） genomic information， including linkage disequilibrium combined with principal component analysis （LD_PCA）， other principal component variants （LD_outPCA）， and Pearson correlation coefficient analysis （LD_PCC）； 2） phenotypic information， including LD combined with genome-wide association study （LD_GWAS） and EPISNP analysis （LD_EPI）； and 3） functional annotation information， using the weight distribution method （WDM）. In total， 8 different screening methods were applied to construct genotype matrices， and the prediction accuracies of different GS models were compared. The LD_PCC method， when combined with 6 GS models （GBLUP， BayesA， BayesB， BayesC， Bayes LASSO， and RRGBLUP）， achieved the highest mean prediction accuracies for duck fossil length， pig age to 100 kg， backfat thickness， and teat number， improving by approximately 4%-13.9% compared with the initial genotype matrix， with accuracies of 0.709 3， 0.440 0， 0.497 4， and 0.440 1， respectively. The WDM method achieved the highest accuracies for backfat thickness and teat number， with improvements of about 10% compared with unweighted models. In conclusion， this study demonstrates that WDM and LD_PCC can effectively enhance the prediction accuracy of GS， providing valuable insights for further exploration of variant selection methods and their application in breeding practice.

This study aimed to explore the effect of glucose on the proliferation， migratory capacity， and programmed cell apoptosis of in vitro-cultured secondary dermal papilla cells （DPCs） derived from Yanshan cashmere goats. Skin samples were collected from healthy male Yanshan cashmere goats aged about 6 months old. Secondary hair follicles were then harvested from this tissue， and the derived DPCs were subjected to a process of extraction and purification. A growth curve was established after cell counting. The purified DPCs population was then subjected to a random distribution across 5 groups， each containing 6 replicate cultures. Glucose solutions of 0 （control group）， 25， 50， 100 and 200 μg·mL^-1 were added to the culture medium， respectively. The proliferation ability of DPCs was evaluated by the CCK-8 method at 2， 12， 24 and 48 h of culture. A cell scratch assay was conducted to evaluate the migratory capacity of DPCs， while the extent of apoptosis was quantified via flow cytometry following Annexin V-FITC/PI dual staining. The transcriptional responses to glucose exposure were assessed by real-time qPCR， measuring the mRNA abundance of key proliferation markers （PCNA， CCND1） and apoptosis regulators （Bcl-2， Bax， Caspase-3， FGF21， FGF5， P53）. Furthermore， Western blot analysis was performed to determine the effect of glucose on the expression of corresponding functional proteins， including the proliferation- and anti-apoptosis-related factors （PCNA， CCND1， Bcl-2）， as well as the pro-apoptotic executors （Bax， Caspase-3）. The results demonstrated that at the 12-hour time point， both 100 and 200 μg·mL^-1 glucose treatments significantly enhanced the proliferation and migration rates of DPCs （P<0.05）， while concurrently suppressing apoptosis （P<0.05）. Further investigation revealed these concentrations markedly upregulated the mRNA expression of proliferation- and anti-apoptosis-related genes （PCNA， CCND1， Bcl-2） （P<0.05）， and downregulated transcripts of key apoptosis-associated genes （Bax， Caspase-3， FGF21， FGF5， P53） （P<0.05）. At the protein level， the 100 μg·mL^-1 treatment significantly elevated the expression of proliferation and anti-apoptotic markers （PCNA， CCND1， Bcl-2） and reduced pro-apoptotic Bax protein （P<0.05）. Conversely， the 200 μg·mL^-1 glucose group specifically induced a pronounced downregulation of the critical apoptosis executor Caspase-3 （P<0.05）. In conclusion， the pro-proliferative and anti-apoptotic effects observed in cashmere goat DPCs following treatment with 100 and 200 μg·mL^-1 glucose are mediated through a dual regulatory mechanism. This involves the coordinated transcriptional and translational upregulation of key proliferative markers （PCNA， CCND1） and the anti-apoptotic protein Bcl-2， concurrent with the suppression of specific apoptosis-related transcripts （FGF21， FGF5， P53） and both the gene and protein expression of pro-apoptotic mediators Bax and Caspase-3.

This study aimed to explore the effects of gender， age， and altitude on the hematological and blood rheology parameters of Bamei cattle， providing a basis for further understanding of the physiological adaptation mechanisms of Bamei cattle in different environments. A total of 166 healthy Bamei cattle from different altitudes in Nyingchi were selected， and 21 hematological and 13 blood rheological parameters were measured. Descriptive statistics， correlation analysis between parameters， and variance analysis based on factors such as gender， age， and altitude were performed. The results indicated that all hematological parameters of Bamei cattle were within the reference range， showing that their blood physiological indicators were in a normal state. The further variance analysis showed that gender， age， and altitude significantly affected the blood physiological indicators. Female cattle had significantly higher red blood cell volume and platelet activity indicators than male cattle （P<0.05）. Juvenile cattle showed significantly higher lymphocyte counts and platelet numbers， as well as other immune and hematopoietic activity indicators， compared to adult cattle （P<0.05）， indicating that gender and age influence blood composition and rheological performance. The effect of altitude was particularly significant. As altitude increased， oxygen-carrying indicators such as red blood cell count （RBC）， hemoglobin （HGB）， and hematocrit （HCT） significantly increased （P<0.05）， demonstrating a typical high-altitude hematopoietic adaptation trend. At the same time， plasma viscosity， aggregation indices， and whole blood flow resistance all significantly increased （P<0.05）， suggesting that blood became more viscous and less fluid in the high-altitude environment. Further correlation analysis revealed significant positive correlations between various internal indicators of the hematological and blood rheology systems. For example， in the red blood cell system， HGB was highly correlated with HCT， and mean corpuscular volume （MCV） with mean corpuscular hemoglobin （MCH）， with correlation coefficients reaching 0.98. In blood rheology， plasma viscosity and fibrinogen were strongly correlated with erythrocyte viscosity， with a correlation coefficient of 1.00. These correlation results revealed the intrinsic interconnection and mutual influence between the blood physiological indicators of Bamei cattle. The results of this study indicate that Bamei cattle possess the ability to adapt to the hypoxic environment of the plateau through enhanced erythropoiesis and the regulation of blood rheological properties. This provides important physiological evidence for a deeper understanding of the hypoxic adaptation mechanisms in plateau livestock.

The objective of this study was to estimate the genetic parameters of reproductive traits in heifers and adult cows and to elucidate the genetic correlations of these traits between heifers and adult cows as well as across different parities. This study collected a total of 84 061 reproductive records， including calving， insemination， and pregnancy diagnosis， from 37 329 Holstein cows in 6 large-scale dairy farms in Jiangsu Province between 2020 and 2024. Among these， 31 689 records were from heifers and 52 372 records were from cows. Genetic parameters were estimated using the average information restricted maximum likelihood （AI-REML） procedure implemented in the DMU software based on animal and repeatability models. The results showed that the heritability estimates of reproductive traits in heifers ranged from 0.04 to 0.52， with the highest value observed for age at first service （0.52±0.01）， while the heritability of the other traits ranged from 0.04 to 0.22. In heifers， the genetic correlations among age at first service， age at last service， and age at first calving were high， ranging from 0.73 to 0.98. In cows， the highest genetic correlations were observed between days open and the number of services and interval between first and last service， with values ranging from 0.95 to 0.96. The genetic correlations of reproductive traits between adjacent parities were relatively high， with values ranging from 0.62 to 0.68 between heifers and primiparous cows. By contrast， the correlations between heifers and multiparous cows were low （0.05-0.15）. Furthermore， phenotypic correlations across different parities were generally low， ranging from 0.01 to 0.18. In addition， the absolute genetic correlations of number of services and interval between first and last service with other traits were relatively high， ranging from 0.28 to 0.97， suggesting that these traits could serve as important indicators of reproductive performance in both heifers and cows. This study systematically elucidated the genetic parameters of reproductive traits in Holstein cattle in Jiangsu Province， providing a theoretical basis for the genetic improvement of reproductive traits in this population.

This study aimed to elucidate the phenotypic patterns and genetic architecture of fertility traits in Ningxia Holstein cattle， providing a theoretical basis for establishing a balanced breeding program in the region. We compiled 216 787 fertility records from 92 152 Holstein cattle across 15 farms in Ningxia during 2005—2024. Genetic parameter were estimated for 4 heifer traits and 9 cow traits （first， second， and third parities） using the AI-REML algorithm in DMU. Single-trait animal models were fitted to estimate heritability within parity， whereas bivariate models were used to get genetic and phenotypic correlations. Heifer heritabilities ranged from 0.097 0±0.008 to 0.500 9±0.012 9， with AFS the highest and CR_h the lowest. Heritability estimates for fertility traits in cows across the first， second， and third parities ranged from 0.007 6 （0.002 5） to 0.150 8 （0.011 6）， 0.010 1 （0.003 4） to 0.149 9 （0.010 2）， and 0.003 9 （0.005 2） to 0.137 6 （0.013 3）， respectively； differences for the same trait between parities were small and consistently lower than those observed in heifers. Genetic and phenotypic correlations among heifer fertility traits ranged from -0.845 8 to 0.835 1 and from -0.550 7 to 0.908 4， respectively； the strongest negative genetic correlation （-0.845 8） was observed between CR_h and IFL_h. Within any parity， genetic and phenotypic correlations among cow fertility traits showed consistent trends， with phenotypic correlations consistently lower than genetic correlations. As parity increased， genetic correlations strengthened among DO， IFL_c， and CI， whereas BW， CS， CE and GL exhibited weak genetic correlations with other fertility traits and can be selected independently. The same fertility trait measured in adjacent parities showed high genetic correlations， indicating that it is largely controlled by the same gene， whereas the phenotypic correlations remained low， suggesting that the trait is strongly influenced by environmental factors. Fertility traits show low-to-moderate heritability in Ningxia Holstein cattle， with greater genetic variation in heifers. CR_h and IFL_h are strongly and negatively correlated and should be treated as core traits in a heifer fertility index. DO， IFL_c， and CI can be selected jointly， and multi-parity records for cow fertility traits can be combined to increase selection accuracy. This study provides scientific basis for developing a balanced breeding programs for Holstein cattle in Ningxia and facilitates the simultaneous improvement of fertility and productive performance.

This study aimed to analyze the effects of dietary supplementation with flaxseed （Linum usitatissimum） and Schizochytrium on rumen microorganisms and meat fatty acid profiles in yaks.Twenty-one 12-month-old male healthy yaks weighing （95.00±5.20） kg were selected and divided into 3 groups according to the test requirements， with 7 replicates in each group. Supplementary feeding group （S） served as the control group， and the 2 experimental groups were fed with flaxseed （Y） and Schizochytrium （L） respectively for 210 days to analyze the effects on the microbial community structure and differential metabolites in the rumen of housed yaks， in conjunction with 16S rRNA sequencing and non-target metabolomes. Flaxseed and Schizochytrium supplementation significantly affected growth performance， muscle fatty acid composition， and rumen microbiota. Both treatment groups showed significantly lower dry matter intake （P<0.05） but improved feed conversion ratio （P<0.05）. The Schizochytrium group exhibited significantly higher muscle docosahexaenoic acid （DHA） content （0.35% vs. control 0.04%， P<0.05） and lower n-6/n-3 PUFA ratio （5.42 vs. control 36.28）. The flaxseed group showed increased total fatty acids （1.44 g vs. control 0.85 g， P<0.05）. Rumen analysis revealed significantly higher Firmicutes phylum abundance （69% vs. control 57%， P<0.05） and significantly lower Bacteroidota abundance （21% vs. control 27%，P<0.05） in the flaxseed group. The Schizochytrium group demonstrated increased Prevotella genus abundance （7.5% vs. control 3.9%， P<0.05）. α-diversity decreased in treatment groups versus control， while β-diversity differed significantly among groups. Muscle metabolomics analysis results indicated the downregulated phospholipid and acylcarnitine metabolism （P<0.05） and upregulated fatty acid derivatives in both treatment groups. Rumen microbiota correlated significantly with muscle metabolism （e.g.， Patescibacteria positively correlated with succinic acid metabolism， P<0.05）. Flaxseed and Schizochytrium promote lipid deposition in yaks by affecting rumen microbes and muscle metabolism， providing a theoretical basis for precision feeding strategies in alpine regions.

This study aimed to identify pregnancy-associated differentially methylated genes and develop an early pregnancy detection technology utilizing methylation-specific fluorescent probes. This study focused on differentially methylated regions in gene promoters and CpG islands to identify potential biomarkers for swine pregnancy. Initially， methylation-specific PCR （MSP） was employed for preliminary validation of these differentially methylated regions. Subsequently， target-specific fluorescent probes were designed to establish an early pregnancy detection method based on quantitative methylation-specific PCR （qMSP）， followed by methodological verification. Compared with empty pregnant sows， the blood of pregnant sows at 25-30 days of gestation showed high methylation of VBP1 and CpG_280， while CpG_1360 showed low methylation， indicating that these sites may be involved in pregnancy regulation. Using β-actin as an internal reference probe， methylation probes for VBP1， CpG_280， and CpG_1360 were designed to establish a qMSP detection method for preliminary application. Concordance analysis between this methylation probe-based detection and ultrasound （B-ultrasound） showed ROC curve AUC values of 0.713 8， 0.608 0， and 0.746 6 （all>0.5）， with qMSP concordance rates of 85.0%， 78.75%， and 85.0%， respectively. This study is the first to confirm that VBP1 and CpG_280 are hypermethylated while CpG_1360 is hypomethylated during early sow pregnancy. The fluorescence probe-based methylation detection technology targeting these genes offers a novel method for early pregnancy diagnosis in swine.

This study aimed to explore the impact of different photoperiods on the hypothalamic-pituitary-gonadal （HPG） axis in poultry， to reveal the mechanisms by which light regulates the follicles development and egg production performance of hen， and to provide a theoretical basis for the development of technologies to regulate the reproduction of female poultry and enhance the reproductive performance of breeding poultry. A total of 860 healthy Xinghua hens， aged 290 days and in the egg-laying period， were randomly divided into a control group and a treatment group. The experiment lasted for 61 days， with the control group maintained on a lighting schedule of 16.5L：7.5D throughout the experimental period. The lighting schedule for the treatment group was 16.5L：7.5D from day 1 to day 7， 12L：12D from day 8 to day 42， 18L：6D from day 43 to day 50， and 16.5L：7.5D from day 51 to day 61. Egg production was recorded throughout the experiment， and ovarian development， blood hormone levels， and reproductive axis factor expression were measured on days 8， 43， and 61， respectively. The results showed that under 16.5 hours of light exposure， there was no significant difference in egg production between the two groups. Shortening the light exposure to 12 hours significantly suppressed egg production， while extending it to 18 hours rapidly increased the egg production rate， which remained higher than that of the control group after restoring 16.5 hours of light exposure. Consistent with egg production， reducing light exposure significantly decreased the number of LYF and SYF in the ovaries， while extending light exposure quickly increased the number of LYF. However， changes in light exposure had no significant effect on the ovarian index or the number of LWF. Results showed that shortened photoperiod significantly reduced GnRH levels in the treatment group （P<0.05）， while extended photoperiod restored hormone levels to those comparable with the control group， though the differences were not significant （P>0.05）. Furthermore， shortened photoperiod significantly downregulated the expression of hypothalamic GnRH and TRH （P<0.01）. In contrast， extended photoperiod increased the expression of GnIH， GnRH， VIP， and TRH in the hypothalamus， though these changes were not significantly different （P>0.05）. GnRH protein expression followed a trend consistent with that of GnRH.In the pituitary gland， shortened photoperiod led to a significant reduction in the expression of GnRHR， FSH， and PRL （P<0.01）， and a moderate decrease in TSH and LH expression. Under extended photoperiod， the expression of GnRHR， TSH， FSH， and LH recovered to levels similar to the control group.In the ovaries， shortened photoperiod resulted in decreased expression of LHR， FSHR， GnRHR， and PRLR， with significant reductions in LHR and GnRHR （P<0.05 or P<0.01）. Extended photoperiod restored the expression of these factors to control levels. Protein expression levels of FSHR， LHR， and PRLR were generally consistent with their corresponding gene expression trends， though none of the differences reached statistical significance （P>0.05）.In summary， these findings demonstrate that photoperiod regulates follicular development and laying performance in Xinghua chickens by modulating the expression of key factors along the reproductive axis. Shortened photoperiod downregulates the expression of reproductive-promoting factors and gonadal receptors， thereby reducing serum reproductive hormone levels and inhibiting both follicular development and egg production. Conversely， extended photoperiod upregulates reproductive factor expression， promoting follicular development and enhancing laying performance.

The study aimed to further understand the changes in steroid hormones and their related substances at different stages of follicular development in Holstein cows， and to provide a theoretical basis for exploring the process and mechanism of follicular development in cows. In this study， based on liquid chromatography-tandem mass spectrometry （LC-MS/MS） technology， Holstein cows （n=18） were used as research subjects. Targeted steroid metabolomics analysis was performed on follicles of three sizes at different developmental stages， with exploration and group comparison conducted on small （Small follicle， SF， <5 mm， n=6）， medium （Medium follicle， MF， 5-8 mm， n=6）， and large （Large follicle， LF， >8 mm， n=6） follicles. A total of 24 steroid substances were identified. Among them， 8 differential metabolites （6 down-regulated， 2 up-regulated） were found in the comparison between SF and MF； 6 differential metabolites （2 down-regulated， 4 up-regulated） were found in the comparison between MF and LF； and 6 differential metabolites （4 down-regulated， 2 up-regulated） were found in the comparison between SF and LF. Subsequent analysis of differential metabolites based on the KEGG database revealed that in SF and MF， the endocrine resistance pathway （P=0.04） and prostate cancer pathway （P=0.03） were significantly enriched. In conclusion， this study confirmed significant changes in steroid hormones and their related substances during follicular development in Holstein cows， providing a theoretical basis for in-depth understanding of changes in the cow reproductive system.

This study aimed to explore the differences in epididymal histological characteristics and gene expression profiles of Angora rabbits in spring，summer and autumn.Twelve 1-year-old male Angora rabbits with healthy body conditions and normal reproductive functions were chosen. Epididymal tissues of 4 Angora rabbits were collected in spring （no heat stress）， summer （intense heat stress） and autumn （no heat stress），respectively. The histological differences of the epididymis in different seasons were compared through HE staining，and the transcriptome profiles of the epididymis tissues in different seasons were analyzed using RNA-Seq technology. Histological analysis revealed that， compared with spring，heat stress in summer resulted in atrophy of the epididymal duct，disorganized arrangement of epithelial cells，and absence of sperm. In contrast，the epididymal morphology recovered in autumn，resembling that of spring，and exhibited an abundance of sperm. Transcriptome analysis revealed that a total of 1 133 differentially expressed genes （DEGs） were detected in the epididymal tissue between spring and summer，910 DEGs were detected between summer and autumn，while only 36 DEGs were detected between autumn and spring. Among them，the expression trends of 535 DEGs were opposite in spring/summer and summer/autumn groups，and their expression levels recovered in autumn and were similar to those in spring，which were consistent with the histological results of the epididymis. GO enrichment analysis revealed that the DEGs in epididymal tissue between spring and summer were mainly enriched in biological processes including immune response，response to external stimulus，sperm motility，flagellated sperm motility，epithelial cell apoptotic process，and programmed cell death，and cytokine-cytokine receptor interaction pathway. DEGs in the epididymis tissue from summer and autumn were significantly enriched in biological processes including immune response，response to external stimuli，positive regulation of flagellated sperm motility，sperm motility，and negative regulation of cell death，as well as TNF，Th1 and Th2 cell differentiation，Th17 cell differentiation，and cytokine-cytokine receptor interaction pathways. Analysis of gene expression trends indicated that the highly-expressed genes in summer，including 449 differentially expressed genes with opposite expression trends between spring/summer and summer/autumn，mainly enriched in immune responses，response to external stimulus，regulation of programmed cell death and regulation of apoptotic process. The low-expression genes in summer are mainly enriched in collagen fibril organization and amino acid transport. In conclusion， heat stress during summer damaged epididymis tissue structure and changed expression profiles of genes related to reproduction，immunity and apoptosis in Angora rabbits，which recovered to normal levels in autumn. This study provided an important theoretical basis for a in-depth understanding of the molecular mechanism of“summer infertility”in rabbits.

The study evaluated the effects of folic acid （FA） on rumen fermentation parameters， antioxidant indicators and microbial population in dairy cows in vitro. Four early lactation Holstein dairy cows were used as the donor of ruminal fluid， and the total mixed ratio of dairy cows was used as the fermentation substrate. The supplementation level of FA was 0， 1.5， 3.0， 4.5， 6.0， 7.5 and 9.0 mg·kg^-1 dry matter （DM）， respectively. After incubation at 39℃ for 24 h， ruminal DM degradability （DMD）， gas production （GP）， FA content， antioxidant indicators， fermentation parameters， microbial enzyme activity and population were determined. The results were showed as follows： 1） The DMD was greater （P<0.05） for the supplementation of FA at 1.5， 3.0 and 4.5 mg·kg^-1 DM， and the 24 h GP and theoretical maximum GP were greater （P<0.05） for the supplementation of FA at 1.5， 3.0， 4.5 and 6.0 mg·kg^-1 DM compared with other groups. 2） Ruminal fluid FA content was greater （P<0.05） and malondialdehyde content was lower （P<0.05） for the groups with FA supplementation compared with the control. The activity of superoxide dismutase was greater （P<0.05） in the groups with 1.5， 3.0， 4.5 and 6.0 mg·kg^-1 DM FA supplementation compared with other groups. The activity of glutathione peroxidase was greater （P<0.05） in the groups with supplementation of FA at 3.0， 4.5， 6.0 and 7.5 mg·kg^-1 DM than in the control and 1.5 mg·kg^-1 DM FA supplementation groups. The total antioxidant capacity activity was greater （P<0.05） for the supplementation of FA at 1.5， 3.0， 4.5， 6.0 and 7.5 mg·kg^-1 DM than for the control. 3） The groups with FA addition had greater （P<0.05） total volatile fatty acids （TVFA） and acetate production compared with the control， and the production of TVFA and acetate were greater （P<0.05） for the 3.0 and 4.5 mg·kg^-1 DM FA supplementation than for 7.5 and 9.0 mg·kg^-1 DM FA supplementation. 4） The groups with FA addition had greater （P<0.05） activities of cellobiase and xylanase compared with the control， and the activity of cellobiase was greater （P<0.05） for the supplementation of FA at 3.0， 4.5 and 6.0 mg·kg^-1 DM than for the supplementation of FA 1.5 and 7.5 mg·kg^-1 DM， and the activity of xylanase was greater （P<0.05） for the 1.5， 3.0 and 4.5 mg·kg^-1 DM FA supplementation than for the 7.5 and 9.0 mg·kg^-1 DM FA supplementation. 5） The groups with FA addition had greater （P<0.05） populations of total bacteria， protozoa and R. flavefaciens compared with the control. The total bacteria population was greater （P<0.05） in the 4.5， 6.0 and 7.5 mg·kg^-1 DM FA supplementation groups than in the 1.5 and 9.0 mg·kg^-1 DM FA supplementation groups. The R. flavefaciens population was greater （P<0.05） for the 1.5， 3.0 and 4.5 mg·kg^-1 DM FA supplementation than for the 7.5 and 9.0 mg·kg^-1 DM FA supplementation. The fungi population was greater （P<0.05） in the 3.0， 4.5， 6.0 and 7.5 mg·kg^-1 DM FA groups than in the control， 1.5 and 9.0 mg·kg^-1 DM FA groups. The R. albus population was greater （P<0.05） for the groups with 1.5， 3.0， 4.5 and 6.0 mg·kg^-1 DM FA supplementation than for the control， 7.5 and 9.0 mg·kg^-1 DM FA groups. The F. succinogenes population was greater （P<0.05） in the 4.5， 6.0， 7.5 and 9.0 mg·kg^-1 DM FA groups than in the control and 1.5 mg·kg^-1 DM FA group. In conclusion， the supplementation of FA improves rumen antioxidant status， stimulate cellulolytic and amylolytic bacteria growth， and resulting in an increase in DMD and TVFA production. The proper level of ruminal FA supply is 1.5-3.0 mg·kg^-1 DM in the condition of the current study.

This experiment aimed to investigate the effects of dietary supplementation with Glycyrrhiza polysaccharides （GCPs） on the growth performance， serum indicators， apparent nutrient digestibility， rumen fermentation parameters and microbial communities in fattening beef cattle. Forty Simmental fattening bulls with good growth status and an average body weight of 617.70±18.85 kg were selected as experimental animals and randomly divided into four groups. The experimental groups received the basal diet supplemented with 0 mg·kg^-1 BW （control group， G0）， 15 mg·kg^-1 BW （low-dose， G15）， 30 mg·kg^-1 BW （medium-dose， G30）， or 45 mg·kg^-1 BW （high-dose， G45） of GCPs， respectively. The trial included a 10-day adaptation period followed by a 90-day feeding period. The body weight were measured at 0， 45，90 d of the feeding period， daily feed intake was recorded to determine average dry matter intake and feed-to-gain ratio. At 45 and 90 d of the feeding period， 10 mL of blood sample was collected from each bull to assess serum biochemical， antioxidant and immune indicators. At the end of the trial， 300 g of feces sample was collected from each bull to evaluate nutrient apparent digestibility， and 50 mL of rumen fluid was collected to analyze rumen fermentation parameters and microbial related indicators. Compared with the G0 group： 1） During the 0-45 d and 0-90 d periods， the average daily gain and average dry matter intake of the G30 and G45 groups were significantly increased （P<0.05）； in the 0-90 d period， the feed conversion ratio of the G30 and G45 groups decreased significantly （P<0.05）， while the crude fat digestibility increased significantly （P<0.05）； 2） The serum aspartate aminotransferase level and alanine aminotransferase level of the G30 group were significantly decreased （P<0.05）； at 45 d， the serum levels of superoxide dismutase and interleukin-10 in the experimental groups increased significantly （P<0.05）； 3） The acetic acid proportion of the G15 and G30 groups was significantly decreased （P<0.05）， the butyric acid proportion of the G30 group was significantly increased （P<0.05）； 4） The Chao1 richness index of the rumen microbial community in the G30 group was significantly increased （P<0.05）， along with structural changes in microbial composition. In the G30 group， the relative abundance of Bacteroidota， Pseudomonadota， Fibrobacterota， Prevotella and Fibrobacter showed a significant upward trend， while the relative abundance of Bacillota showed a significant downward trend （P<0.05）； the differential metabolic pathways of rumen microorganisms in the G15 group were mainly enriched in Synthesis and Degradation of ketonebodies， and nitrotoluene degradation. In the G30 group， these pathways were mainly linked to Amino acid metabolism， and Fatty acid biosynthesis， while the G45 group showed enrichment in C5-branched dibasic acid metabolism. In summary， dietary supplementation of Glycyrrhiza polysaccharides enhanced the growth performance， antioxidant capacity， immune function， and nutrient digestibility of Simmental fattening beef cattle， while optimizing rumen fermentation patterns and promoting beneficial rumen microorganisms. Under the conditions of this experimental study， the optimal addition amount of GCPs was 30 mg·kg^-1 BW.

This study was conducted to investigate the effects of Bacillus subtilis （BS） and Chitosan oligosaccharides （COS） on growth performance， nutrient utilization， slaughter performance， meat quality and cecal microflora diversity of AA broilers. A total of 288 one-day-old broilers with similar body weight （46±0.64 g） were randomly divided into 6 treatment groups with 6 replicates per treatment and 8 chickens per replicate （ half male and half female ）. The control group （CON） was fed a basal diet， and the experimental groups were fed a basal diet supplemented with 5×10⁹ CFU·kg^-1Bacillus subtilis （BS1 group）， 6×10⁹ CFU·kg^-1Bacillus subtilis （BS2 group）， 125 mg·kg^-1 COS （COS group）， 5×10⁹ CFU·kg^-1Bacillus subtilis+125 mg·kg^-1 COS （MIX Ⅰ group）， 6×10⁹ CFU·kg^-1Bacillus subtilis+125 mg·kg^-1 COS （MIX Ⅱ group）. The experiment lasted for 42 days. The results were showed as follows： 1） From 1 to 21 days of age， compared with CON， ADG in MIX Ⅱ group was significantly increased （P<0.01）， and FCR in MIX Ⅱ group was significantly increased （P<0.01）. At 22-42 days of age， compared with CON， ADG in MIX Ⅰ and MIX Ⅱ groups were significantly increased （P<0.01）. From 1 to 42 days of age， compared with CON， the ADG of MIX Ⅰ and MIX Ⅱ groups were significantly increased （P<0.01）， and the FCR of MIX Ⅱ group was significantly increased （P=0.01）. 2） Compared with CON， the semi-eviscerated rate and eviscerated rate of BS2 group were significantly increased （P<0.05）. 3） Compared with CON， the crude protein utilization rate of broilers in MIX I group and COS group was significantly increased （P<0.05）. 4） Compared with CON， there was no significant difference in pH_{45 min}， pH_{24 h}， shear force， water loss rate， L^*， a^* or b^* among groups （P>0.05）. The drip loss of breast muscle in MIX Ⅰ group was significantly increased （P<0.01）， but the drip loss of leg muscle showed a downward trend， and the difference was not significant （P>0.05）. 5） Compared with CON， the Chao1， ACE and Shannon index of cecal flora in COS group increased by 0.13%， 0.52% and 1.56%， respectively， but the difference was not significant （P>0.05）. The Simpson index of cecal flora in COS group decreased by 5.2%， but the overall difference was not significant （P>0.05）. The BS2 group was significantly deviated from the other treatment groups on the PC1 axis. BS1 group， COS group， MIX Ⅰ group and MIX Ⅱ group were close to each other and tended to gather， with more overlapping parts， indicating that the composition of cecal flora was similar. At the phylum level， compared with CON， the relative abundance of Bacteroidetes and Verrucomicrobia in the cecal flora of broilers in MIX Ⅱ group increased， and the relative abundance of Proteobacteria in the cecal flora of broilers in BS1 group increased； at the genus level， compared with CON， the relative abundance of Bacteroides and Akkermansia in the cecal flora of broilers in MIX Ⅱ group increased. In summary， dietary supplementation of Bacillus subtilis or Chitosan oligosaccharides alone or their combination can promote the growth performance， improve meat quality， enhance the diversity and richness of cecal flora， and ensure the healthy growth of broilers to a certain extent.

The purpose of this study is to provide a solution for the feed utilization of table leftover food by studying the mixed solid-state fermentation technology of table leftover food and corn germ meal. The detection indicators were dry matter disappearance rate and reducing sugar content. The single factor and response surface and other experimental methods were used to optimize the process parameters of solid-state fermentation of table leftover food-corn germ meal enzyme， and the effects of cellulase addition amount and enzyme preparation X1 addition amount on the dry matter disappearancerate and sugar content of solid-state fermentation products were investigated by single factor method based on the initial fermentation conditions. The optimum addition amount of cellulase and enzyme preparation X1 and fermentation time were obtained by response surface method. Twenty-four 22-day-old Arbor Acres （AA） cockerels of similar body weight and in good health condition were randomly divided into 3 groups， which were fed the basal diet and the two experimental diets， respectively， and the metabolizable energy value was measured. The results showed that as the addition amount of cellulase increased， the content of reducing sugar and the disappearance rate of dry matter increased. When the addition amount was 10.5 g·kg^-1， the increase of dry disappearance slowed down （P>0.05）， and when the addition amount was 13.5 g·kg^-1， the increase of reducing sugar slowed. As the addition amount of enzyme preparation X1 increased， the content of reducing sugar and the disappearance rate of dry matter tended to increase as a whole. When the addition amount was 6.0 g·kg^-1， the content of reducing sugar and the disappearance rate of dry matter both showed a downward trend. With dry matter degradation rate R1 as the value， the response surface recommended the best fermentation conditions as cellulase： 10.154 g·kg^-1； Enzyme preparation X1： 90 g·kg^-1， solid fermentation time of lactic acid bacteria was 96 h. With reducing sugar R2 as the response value， the response surface recommended the fermentation conditions as cellulase： 7.842 g·kg^-1； Enzyme preparation X1： 9.0 g·kg^-1， fermentation was 96 h. The solid-state fermentation of mixing table leftover food and corn germ meal， fermenting for 72 h at 37 ℃ with moisture content of 55%， the dry matter disappearance rate increased by 19.84%， the reducing sugar content increased by 162.44·g^-1， and the metabolizable energy value increased by 17.8% compared with the control group. The results suggested that the utilization of solid-state fermentation with enzyme and bacteria not only solves the problem of difficult drying of high-moisture table leftover food but also greatly improves its nutritional value， providing a scientific basis for its feed utilization.

Getah virus （GETV） is an arbovirus that can cause reproductive failure in pregnant sows， neurological symptoms in piglets， and even death in severe cases， posing a threat to the pig industry. This study aimed to analyze the whole genome sequence and evolutionary characteristics of a GETV strain isolated from dead piglets from an infected pig farm. Metatranscriptomic sequencing was conducted to analyze pathogens in brain and lung tissues from dead piglets. Vero CCL81 cells were used to isolate GETV， and the isolated virus was identified through observation of cytopathic effect （CPE）， plaque purification， RT-PCR， indirect immunofluorescence assay （IFA）， and transmission electron microscopy. The full-length genome sequence of the isolate was determined using Sanger sequencing. The results showed that the major pathogen in the tissue samples was characterized as GETV， and the isolated strain was designated as NM2022. Its genome， excluding a poly（A） tail， has a total length of 11 689 nt. Phylogenetic analysis indicated that NM2022 belongs to the evolutionary Group III of GETV， with nucleotide homology ranging from 93.98% to 99.86%， compared to 135 of GETV strains in the GenBank database. The amino acid sequence of the E2 protein of NM2022 is highly conserved with 21 of the GETV strains， including Chinese strains HNzmd-XP2 and JS201809-2 of porcine origin， as well as the strain NMDK1813-1 of mosquito origin， except for a serine-to-alanine mutation at position 380. These findings provide a foundation for further research on the evolutionary patterns and pathogenesis mechanisms of GETV.

Establishment of a double-antigen sandwich ELISA for the detection of antibodies against classical swine fever virus （CSFV） E2 protein. The complete E2 protein sequences of CSFV and bovine viral diarrhea virus （BVDV） were aligned using BioEdit software to identify CSFV-specific epitopes. A recombinant pCDNA3.1-2×rE2-His expression vector was constructed. The 2×rE2-His protein was expressed in CHO cells， followed by thrombin cleavage to remove the His-tag. The purified His-tag-free 2×rE2 protein was used as the coating antigen， while horseradish peroxidase （HRP）-labeled 2×rE2-His protein served as the detection antigen. After systematic optimization of reaction conditions， a double-antigen sandwich ELISA method was successfully established. The specificity， sensitivity， and repeatability of the assay were evaluated， and its concordance rate with a commercial ELISA kit was compared. The optimized assay conditions were as follows： antigen coating concentration at 0.5 μg·mL^-1， serum dilution at 1∶4， and enzyme-labeled antigen dilution at 1∶40 000. The method exhibited no cross-reactivity with positive sera against other common swine pathogens， including senecavirus A （SVA）， porcine pseudorabies virus （PRV）， porcine epidemic diarrhea virus （PEDV）， foot-and-mouth disease virus （FMDV）， and porcine reproductive and respiratory syndrome virus （PRRSV）， nor with BVDV sera， which frequently causes serological cross-reactivity， demonstrating high specificity. The maximum detectable dilution ratio for antibody-positive sera reached 1∶2 048. Both intra-assay and inter-assay coefficients of variation were below 10%， indicating excellent repeatability. The concordance rate with the IDEXX CSFV Antibody Test Kit was 97.1%. To further evaluate the assay， both the established double-antigen sandwich ELISA and the IDEXX kit were used to monitor antibody kinetics in two healthy pigs immunized with CSFV C-strain vaccine combined with an E2 subunit vaccine. The developed ELISA detected antibody responses 6 days earlier than the IDEXX kit. Both methods demonstrated that antibody levels peaked at 37 days post-immunization and subsequently stabilized. In this study， a double-antigen sandwich ELISA antibody detection method for CSFV was established with high specificity， sensitivity and reproducibility， which is suitable for the clinical diagnosis and evaluation of immunization effect of swine fever.

The objective of this study was to isolate and characterize a bovine-yak origin group A rotavirus with the G8P［11］ genotype and to analyze the genetic evolutionary features of its genome. Fecal samples from diarrheic calves at a dairy farm in Sihong， Jiangsu Province， were screened for six diarrhea-associated viruses. Rotavirus （RV）-positive samples were isolated and identified using RT-PCR， indirect immunofluorescence assay （IFA）， dsRNA-PAGE， and electron microscopy. The growth kinetics and cellular tropism of the isolated strains were characterized， and their cross-species transmission potential was evaluated. Additionally， whole-genome sequencing was performed on the isolates， followed by phylogenetic tree construction to determine their G/P genotypes. A bovine rotavirus strain was successfully isolated. Inoculation onto MA104 cells induced typical cytopathic effects， including cell rounding， shrinkage， detachment， and fragmentation. The virus reacted with VP6-specific monoclonal antibodies， exhibited a 4∶2∶3∶2 genomic RNA electrophoretic pattern， and displayed characteristic wheel-shaped virions approximately 70 nm in diameter. This strain was designated JSSH2024. A one-step growth curve revealed peak viral titers （TCID₅₀） as 10^5.75·mL^-1 in MA104 cells at 18 hours post-infection. In vitro infection assays demonstrated the virus's ability to replicate in MA104， IPEC-J2， HT-29 and HCT-8 cell lines， indicating potential for cross-species transmission. Whole-genome sequencing and phylogenetic analysis identified its genotype as G8-P［11］-I2-R2-C2-M2-A13-N2-T6-E2-H3. Eight gene segments clustered closely with bovine rotaviruses， while three segments showed closer genetic affinity to yak rotaviruses， suggesting its origin as a bovine-yak reassortant virus. This study reports the first G8P［11］ bovine-yak reassortant bovine rotavirus （BoRV） in China， providing critical insights for epidemiological surveillance and vaccine development.

This study aimed to prokaryotically express the lumpy skin disease virus （LSDV） ORF132 protein and evaluate its immunogenicity. The ORF132 gene was expressed in E. coli BL21（DE3）， purified， and emulsified with Freund's adjuvant. Mice were immunized with 100 µg of the protein per dose. The results demonstrated that the ORF132 protein elicited high titers of antigen-specific total IgG and a significantly elevated level of IFN-γ secretion compared to the PBS control group. Furthermore， it induced significant lymphocyte proliferation and a high neutralizing antibody titer. These findings indicate that the ORF132 protein is an immunodominant antigen capable of inducing robust humoral and cellular immune responses in mice， providing a promising candidate for subsequent multi-antigen combination studies and bovine trials.

This study aimed to construct a safe and effective Newcastle disease virus （NDV） pseudovirus for the quality control of NDV nucleic acid detection. The P gene of NDV was amplified by RT-PCR and linked to the lentiviral expression vector PLV-Puro to construct Puro-NDV/P. Puro-NDV/P plasmids and two helper plasmids were extracted， and a total of 293T cells were transfected， and pseudovirus particles containing NDV P genes were successfully constructed. The constructed pseudovirus particles were properly diluted and lyophilized to prepare RNA-positive quality controls， and calibrated， uniformity and stability tested. A Taqman fluorescence quantitative PCR detection method was established by successfully constructing NDV P gene pseudovirus. The results showed that 15 positive quality control products were randomly selected for fluorescence RT-PCR detection， and the Ct value coefficient of variation （CV） was 0.90%， which was good uniformity. The copy number of the prepared positive quality control products was 6.58×10² copies·μL^-1. The quality controls were stored at 4 ℃， 25 ℃ for short-term storage （1 week）， and -20 ℃ for a long time （6 months）， and the Ct value was detected by fluorescence RT-PCR without significant change， and the stability was good. A real-time fluorescence RT-PCR detection method based on NDV P gene pseudovirus was established. The NDV pseudovirus prepared in this test has the advantages of good uniformity and stability， and can be used for the research and development， evaluation and quality control of NDV nucleic acid detection kits， providing a reliable tool for the standardized detection of NDV.

Phage therapy is a novel antimicrobial treatment， in order to provide new theoretical and technical support for the prevention and treatment of E. coli infections， in this study， E. coli E6 and a broad-spectrum E. coli phage named vB_EcoM_E6 were isolated from the feces of diarrheic goats， and the potency， pH stability， thermal stability， lysis spectrum， and one-step growth curves of the phage were measured by observing the phage spots and biological properties， performing whole genome sequence analysis and evaluating the phage by animal model tests. The results showed that the lysis rate of this phage in the study was as high as 70.6%， and it was relatively stable at 50 ℃ and pH 3-10 range， and the transmission electron microscopy observation results indicated that this phage belonged to the myotail phage. Genome-wide analysis showed that the phage had a total length of 137 974 bp， a CG content of 39.13%， and 247 coding sequences （CDS）. The results of the mouse model test showed that the phage had a good protective effect against E. coli infection. This study provides a good material for further research on E. coli antimicrobial agents.

This study investigated the epidemiology of Salmonella in a large-scale broiler slaughterhouse in Guangdong， China， and characterized the genomic features and potential transmission risks of the predominant serovar S.Infantis， providing evidence for targeted control. Comprehensive sampling was performed across all processing stages of the slaughterhouse， yielding Salmonella isolates that were tested for resistance to 12 antimicrobials using the agar dilution method. Thirty-four representative isolates were subjected to whole-genome sequencing （WGS） for analyses of serotype， multilocus sequence typing （MLST）， plasmid content， and resistance determinants， followed by construction of a single nucleotide polymorphism （SNP）-based phylogeny. In addition， 10 389 publicly available S.Infantis genomes were retrieved from NCBI to clarify the potential transmission risks using whole-genome multilocus sequence typing （cgMLST） and SNP pairwise distance matrix analysis. A total of 96 isolates were recovered， with an overall prevalence of 55.49% （96/173），and the highest prevalence observed in the evisceration stage （79.25%）. All isolates exhibited multidrug resistance （MDR） with highly similar resistance profiles， showing 100% resistance to ampicillin， ceftiofur， nalidixic acid， florfenicol， and sulfamethoxazole， and over 90% resistance to chloramphenicol and tetracycline. WGS revealed S.Infantis as the predominant serovar （32/34）， dominated by ST32 （30/32）. All S.Infantis isolates harbored the large IncFIB-type pESI megaplasmid， carrying diverse resistance， virulence， metal， and disinfectant resistance genes. Comparative genomic analyses indicated that the slaughterhouse isolates clustered closely with 15 global S.Infantis strains， 12 of which originated from human infections. These findings demonstrate extensive cross-contamination of Salmonella across slaughterhouse stages， with MDR S.Infantis strains harboring a wide repertoire of resistance and virulence determinants. The close phylogenetic relationship with international human-derived isolates underscores their strong transmission potential and public health threat. Enhanced monitoring and targeted control of MDR S.Infantis， particularly within poultry slaughterhouses， should be prioritized to protect food safety and public health.

Nuclear factor of activated T cells 1 （NFAT1）， as a key transcription factor regulating T cell function， may participate in the pathogenesis of malaria by mediating the T cell exhaustion. This study aims to investigate the regulatory role of NFAT1 in the progression of Plasmodium berghei ANKA infection. By inhibiting NFAT1 activity and evaluating its effects on the expression of immune checkpoint molecules—T cell immunoglobulin and mucin domain-containing protein 3 （Tim-3）， cytotoxic T lymphocyte-associated antigen-4 （CTLA-4）， and programmed cell death-1 （PD-1）—in peripheral blood and spleen of Plasmodium-infected mice， this study further explore the potential therapeutic value of targeting NFAT1 for malaria treatment. Firstly， a C57BL/6 mouse infection model was established， serum lactate levels and peripheral blood NFAT1 protein expression （Western blot） were dynamically monitored at different time points （day 0， 4 and 7）. Flow cytometry was used to detect the expression levels of Tim-3， PD-1， and CTLA-4 on CD8⁺ T cells and NK cells in peripheral blood and spleen at different time points （day 0， 4， 6 and 8）. An intervention model was subsequently constructed， followed by sample collection at 48 hours post-intervention for immunophenotypic analysis. The results indicated that the serum lactate level in the infection group was significantly higher than that in the control group at the 4th （P<0.01） and the 7th （P<0.001）. The expression of NFAT1 protein was significantly upregulated at 7 days after infection （P<0.01）. Flow cytometry analysis showed that the proportions of Tim-3⁺， CTLA-4⁺， and PD-1⁺ subsets of CD8⁺ T cells/NK cells in the spleen and peripheral blood of mice were significantly increased at 6-8 days after infection （P<0.01）. After NFAT1 inhibitor intervention， the proportion of Tim-3⁺ CD8⁺ T/NK cells in the peripheral blood was significantly decreased （P<0.01）， and the proportions of CTLA-4⁺， Tim-3⁺ CD8⁺ T cells and Tim-3⁺， PD-1⁺ NK cells in the spleen were significantly reduced （P<0.01）. Notably， the proportion of CTLA-4⁺ NK cells in the peripheral blood was abnormally elevated after inhibitor treatment （P<0.001）.In conclusion， NFAT1 inhibition can partially down-regulate the expression of immune checkpoint molecules in CD8⁺ T/NK cells， but it is accompanied by the compensatory overexpression of CTLA-4 in NK cells， which may weaken its therapeutic effect. The research suggests that NFAT1-targeted therapy needs to be optimized in combination with the dynamic regulation mechanism of immune checkpoints， providing a new research direction for malaria immunotherapy strategies.

This study aimed to evaluate the application of metatranscriptomic sequencing technology for the identification of a co-infection event involving European and North American genotypes of PRRSV. A PRRSV-positive serum sample was collected from a swine farm in Guangxi Zhuang Autonomous Region. The complete genome sequence of the virus was obtained using metatranscriptomic sequencing， and further subjected to phylogenetic and recombination analysis. Complete genome sequences of both PRRSV1-GXHZ-202408 （15 076 bp） and PRRSV2-GXHZ-202408 （14 989 bp） were successfully recovered. Phylogenetic analysis revealed that PRRSV1-GXHZ-202408 formed a novel subclade with its closest relative SC-2020-1 （GenBank： MW115431； 90.45% nucleotide identity）， while PRRSV2-GXHZ-202408 clustered within the NADC30-like group （Lineage 1.8）， sharing 88.63% nucleotide identity with the prototype NADC30 strain （MH500776）. Notably， recombination events were identified in both strains： PRRSV1-GXHZ-202408 originated from an SC-2020-1 backbone with recombination breakpoints in NSP1-NSP2 （650-2 050 nt） and ORF6 （14 131-14 570 nt） involving LV4.2.1 （AY588319） and HKEU16 （EU076704）， whereas PRRSV2-GXHZ-202408 arose from recombination between NADC30 （MH500776， major parent） and JXA1 （EF112445， minor parent） in the NSP3-NSP9 region （5 331-8 081 nt）.In this study， metatranscriptomic sequencing was employed to not only definitively identify co-circulation of PRRSV1 and PRRSV2 in the swine farm， but also to obtain complete viral genomes and precisely characterize the genetic features of circulating strains. This approach clearly analyzed the epidemic dynamics of PRRSV in the swine farm， demonstrating that metatranscriptomic sequencing serves as a powerful tool for precise prevention and control of PRRS.

At present， the remodeling of host metabolism by viral infection has been studied in a variety of viruses. As a porcine intestinal pathogenic coronavirus， the interaction between porcine epidemic diarrhea virus （PEDV） and host metabolic system has not been reported. Studies have shown that angiotensin-converting enzyme 2 （ACE2） is involved in the regulation of glucose metabolism in the pathophysiological state of the body and plays a role in anti-PEDV infection. In order to explore the regulatory effect of PEDV infection on intestinal glucose metabolism in piglets and the possible role of ACE2， the relationship between ACE2， PEDV infection and intestinal glucose metabolism in piglets was clarified. In this experiment， healthy and PED piglets were used as the research objects. By collecting blood， duodenum， jejunum and ileum tissue samples were isolated and intercepted for the following experiments： 1） The expression of PEDV M gene and N gene and protein in each segment of small intestine was detected. 2） Detection of glucose and lactic acid in blood； western blot was used to detect the expression of glucose transporter SGLT-1 and GLUT-2 protein. 3） RT-qPCR was used to detect the gene transcription of key enzymes in glycolysis and TCA cycle pathway in each intestinal segment； 4） ELISA and Western blot were used to analyze the content of Ang1-7， AngII and the protein expression of ACE2 and MasR in each intestinal segment； 5） Spearman method was used to analyze the correlation between ACE2 and PEDV infection and glucose metabolism-related enzyme expression changes.The results showed that ： 1） There was M gene expression in the jejunum tissue of diarrhea piglets， and a positive band appeared at about 552 bp. PEDV-N protein was expressed in duodenum， jejunum and ileum of piglets， with the highest expression in jejunum （P<0.01）. 2） Compared with the normal healthy group， the contents of glucose （P<0.05） and lactic acid （P<0.01） in the blood of PEDV-infected piglets were significantly or extremely significantly increased； the protein expressions of SGLT-1 and GLUT-2 in duodenum and jejunum were significantly or extremely significantly up-regulated （P<0.05 & P<0.01）. 3） The results demonstrated that the transcriptional levels of key glycolytic enzymes （HK2， PFK， PKM， and LDHA） were markedly elevated （P<0.01） in the small intestinal tissues of PEDV-infected piglets， whereas those of key TCA cycle enzymes （CS， OGDH， and IDH） were significantly suppressed （P<0.05 & P<0.01）.； 4） In the small intestinal tissues of PEDV-infected piglets， the protein expressions of ACE2 and MasR were significantly down-regulated （P<0.01）. In conjunction with these changes， the level of Ang1-7 was markedly decreased （P<0.01）， whereas that of AngII was considerably increased （P<0.01）； 5） PEDV infection was positively correlated with SGLT-1， GLUT-2， HK2， PKM and PFK in the small intestine of piglets， and negatively correlated with ACE2-Ang1-7-MasR axis. The results suggest that PEDV infection in intestinal tissues activates glycolysis and suppresses the TCA cycle， concomitant with a weakened ACE2-mediated ACE2-Ang1-7-MasR pathway. This metabolic rewiring manifests as a Warburg effect， characterized by increased glucose uptake and lactate production. Furthermore， this PEDV-induced remodeling of intestinal glucose metabolism is negatively correlated with ACE2 expression.

The study characterizes the pathogenicity and antimicrobial resistance profile of atypical enteropathogenic Escherichia coli （aEPEC） strain ZX185 isolated from sheep meat in Xinjiang， and evaluates its potential for foodborne transmission. Antimicrobial susceptibility was determined using the microbroth dilution method， and whole-genome sequencing was performed on the Illumina HiSeq platform. Bioinformatics analyses were conducted to investigate genomic features， including virulence and resistance determinants. A phylogenetic tree was constructed using core-genome single nucleotide polymorphisms （cgSNPs） to compare virulence gene content across closely related strains. The results indicated that strain ZX185 belongs to serotype O85：H4 and sequence type ST642. Although it was phenotypically susceptible to all 28 tested antibiotics， its genome harbored 43 antimicrobial resistance genes and an IncFIB plasmid replicon， suggesting potential for multidrug resistance. Additionally， this strain carries 91 virulence genes （including adhesion factors， type Ⅲ/Ⅵ secretion system effector proteins， etc.） and exhibits notable acid tolerance， with a survival rate exceeding 11% after 3 hours of treatment in simulated gastric fluid at pH≥3.5， indicating a strong potential for intestinal colonization and pathogenicity. Although cgSNP-based phylogenetic analysis indicated a close genetic relationship （26 SNPs apart） between strain ZX185 and the human-source U.S. strain MOD1-EC6173 （O85：H4， ST642）， ZX185 carries 35 additional virulence genes missing in MOD1-EC6173， predominantly associated with adhesion， T3SS/T6SS， and toxin synthesis. These findings indicate that although this sheep-derived aEPECstrain ZX185 shows phenotypic susceptibility， it possesses multidrug resistance genes and strong virulence traits， and exhibits high genomic similarity to a human-derived strain， indicating potential cross-host transmission risk. This study provides critical molecular insights into the pathogenesis of aEPEC， the evolution of antimicrobial resistance， and the foodborne transmission risk， supporting improved surveillance and control strategies.

This experiment aims to study the immune-enhancing effects of peptide gCK-16 combined with the attenuated vaccine of infectious bursal disease （IBD） and to evaluate the protective effects of the vaccination on SPF chickens（14-day-old）. To investigate the toxicity of the peptide gCK-16， the immune effect of vaccine and the levels of cytokines （IFN-γ， IL-4 and IL-6） in serum， the cells were stimulated by the different concentrations of peptide gCK-16 in vitro， and then the chickens were immunized with the different concentrations of peptide gCK-6 in combination with attenuated vaccine of IBD. Meanwhile， the challenge protection of infectious bursal disease virus （IBDV） was performed by detecting the survival rate， the bursa-weight/body-weight index （BBIX）， the splenic lymphocyte viability， the level of immunoglobulin （IgM and IgA）， the level of immunoglobulin （IgY） in the serum， the histopathological changes and the expression level of apoptosis-related proteins in the bursa. The results show that peptide gCK-16 has no obvious toxicity both in vivo and in vitro， and peptide gCK-16 combined with the attenuated vaccine of IBD can significantly increase the antibody titer of IBDV in the serum of chickens immunized with the attenuated vaccine of IBD and the level of cytokines （IFN-γ， IL-4 and IL-6） in the serum. Moreover， the peptide gCK-16 significantly promoted the stimulation index of LPS， ConA and IBDV to PBMCs. Immunization and challenge experiments also showed that the peptide gCK-16 significantly increased the survival rate and BBIX of chickens infected with IBDV， and reduced the pathological changes of bursa tissues， while the expression levels of pro-apoptotic proteins Bax， Caspase-3， Caspase-9 were significantly decreased， and the expression level of anti-apoptotic protein Bcl-2 was significantly increased. In conclusion， peptide gCK-16 can significantly enhance the humoral and cellular immune responses of the attenuated vaccine of IBD， and achieved an effective immune protection in chickens infected with IBDV， which provides theoretical reference and data support for the application of this peptide as an immunopotentiator of the attenuated vaccine of IBD.

The aim of this study was to investigate the effect of Bacillus subtilis （KC） in combination with different lactic acid bacterias on the quality and active effects of fermented Astragalus. In which the different lactic acid bacterias inludes Lactobacillus acidophilus （SS）， Lactobacillus rohitaensis （LY）， Lactobacillus plantarum （ZW）， and Streptococcus alactolyticus （FGM）. Four kinds of fermented Astragalus were prepared by the mixed probiotics （SS+KC， LY+KC， ZW+KC， and FGM+KC）， respectively. The negative control group （UNF） was the same preparation process with fermented Astragalus， but no probiotics. The quality of 4 fermented products was evaluated by measuring general physicochemical indicators （pH， ethanol content）， characteristic physicochemical indicators （total acid， organic acid， crude polysaccharide content）， total flavonoids， total saponins， total polyphenols， aflatoxin B₁ （AFB₁）， and deoxynivalenol （DON） content. HD11 cells were stimulated with different concentrations of 4 fermented products and UNF samples， respectively， and the cell proliferation activity was determined by the CCK8 method， and the phagocytic ability and cytokine expression were determined by immunofluorescence and ELISA methods. The results showed that the general physicochemical indicators and characteristic physicochemical indicators of LY+KC group， FGM+KC group， and ZW+KC group all met the requirements of plant enzyme （QB/T5323-2018）. Compared with UNF group， the total saponin content of LY+KC group was significantly reduced （P<0.05）， and the total flavonoid and total saponin contents of FGM+KC group and ZW+KC group were significantly increased （P<0.05）. Compared with UNF group， the AFB₁ and DON contents of FGM+KC group， ZW+KC group， and LY+KC group were significantly reduced （P<0.05）. At the concentration of 10 µg·mL^-1， four kinds of fermented Astragalus could promote the viability of HD11 cells， which was determined as the subsequent working concentration. After treated with ZW+KC and FGM+KC fermented Astragalus， the phagocytosis capacity and the expression of pro-inflammatory factors （IL-1β and IL-12） in HD11 cells were significantly higher than those in CON， UNF， SS+KC and LY+KC groups （P<0.05）. In conclusion， the selection of probitics is closely related to the quality and activity of fermented Astragalus. Among them， Astragalus membranaceus fermented by Bacillus subtilis combined with Lactobacillus plantarum or Streptococcus alactolyticus FGM has better quality， which can increase the content of total flavonoids and total saponins， degrade AFB₁ and DON， and improve the function of HD11 cells.

This study aimed to investigate the mechanism of Ziziphi Spinosae Semen in inhibiting LPS-induced bovine mastitis using network pharmacology， molecular docking and in vitro experiments. The core components and potential targets of Ziziphi Spinosae Semen were screened through TCMSP， Swiss Target Prediction and Pharm Mapper databases， while bovine mastitis-related targets were obtained from DisGeNET， OMIM and GeneCards databases. Venny， STRING database and Cytoscape 3.8.0 software were used for topological analysis to construct compound-target and disease-target interaction networks. Hub targets for treating bovine mastitis were screened and subjected to GO functional and KEGG pathway enrichment analyses. Molecular docking and visualization were performed between hub targets and active components using AutoDock. An LPS-induced inflammatory model was established in bMECs， with CCK-8 assay， qRT-PCR， Western blot and ELISA used to analyze the effects of Ziziphi Spinosae Semen on inflammatory factors （IL-1β， IL-6， TNF-α） and hub targets at mRNA and protein levels. Results showed that： 9 bioactive components of Ziziphi Spinosae Semen and 235 potential therapeutic targets for bovine mastitis were identified； PPI analysis revealed 4 hub targets： epidermal growth factor receptor （EGFR）， estrogen receptor 1 （ESR1）， peroxisome proliferator-activated receptor gamma （PPARG）， and phosphatidylinositol-4，5-bisphosphate 3-kinase catalytic subunit alpha （PIK3CA）； GO and KEGG enrichment analyses indicated predominant involvement in biological processes including positive regulation of cytosolic ion concentration， receptor complex formation， and steroid binding， with significant associations to pathways such as central carbon metabolism in cancer， endocrine resistance， insulin resistance， and EGFR tyrosine kinase inhibitor resistance； Molecular docking demonstrated high binding affinity between ESR1 and the key components of Ziziphi Spinosae Semen， betulinic acid and daucosterol. In vitro experiments confirmed that Ziziphi Spinosae Semen bioactive components significantly suppressed the LPS-induced upregulation of IL-1β， IL-6， TNF-α， and ESR1、PI3KCA、PPARG mRNA and ESR1 protein levels in bMECs. In conclusion， Ziziphi Spinosae Semen exerts therapeutic effects against bovine mastitis primarily through bioactive components like betulinic acid by modulating key targets including ESR1. This study provides a scientific foundation for developing Ziziphi Spinosae Semen based therapeutic strategies for bovine mastitis intervention.

This study aimed to investigate the mechanism of Banlian Zijin San （BLZJS） in preventing and treating heat stress in broilers using network pharmacology， molecular docking， and animal experiments. Active components and targets of BLZJS were screened using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）. Heat stress-related disease targets were identified via the GeneCards database. Venny and Cytoscape 3.7.2 software were used to map BLZJS-heat stress intersection targets and construct a "drug-component-disease-target" network. The STRING database established a protein-protein interaction （PPI） network. GO functional enrichment and KEGG pathway enrichment analyses of targets were performed using DAVID and Metascape databases. The results of network pharmacology showed that 122 targets were intersected between Banlian Zijinsan and heat stress， and the lipid and atherosclerosis and cancer pathways were the ones with higher number of enriched genes and significance， and the molecular docking results showed that the components of Banlian Zijinsan with higher number of potential targets bound better to the core targets of heat stress diseases；For the animal experiments， 300 1-day-old broiler chickens were randomly divided into 5 groups. Except for the control group， all other groups were subjected to heat stress treatment at an ambient temperature of 37±2 ℃ for 42 days. Three treatment groups were added with 0.5%， 1%， and 2% of traditional Chinese medicine compound to their basal diet， while the model group and control group were fed with basal diet. During the experiment， Banlian Zijin San could significantly reduce the levels of IL-6 and TNF-α in the serum of broiler chickens on days 14， 28， and 42 of heat stress （P<0.05）， significantly increased the level of IL-10 （P<0.05）， significantly reduced the levels of HSP70， HSP90， IL-6， and TNF-α in liver tissue （P<0.05）， and significantly increased the level of IL-10 （P<0.05）. Mild liver damage occurred in the model group on day 14， but not in BLZJS groups. Liver damage worsened in the model group on days 28 and 42， while BLZJS groups showed no significant damage.BLZJS alleviates heat stress-induced impairments in broiler growth performance and liver function. Its multiple active components likely regulate key targets （IL-6， IL-10， TNF-α， HSP70） and signaling pathways to mitigate heat stress.

The purpose of this study was to determine the circadian variation and differences of blood glucose and lipid metabolism， immunity， oxidation and antioxidation related indexes in postpartum healthy dairy cows and subclinical ketosis dairy cows， and to evaluate the effects of ketosis on the circadian physiological homeostasis of energy metabolism， immunity and oxidative stress in dairy cows from the perspective of the circadian clock. The blood samples of postpartum healthy group （postpartum 3-21 d， blood BHBA<1.2 mmol·L^-1， n=10） and subclinical ketosis group （postpartum 3-21 d， 1.2 mmol·L^-1<blood BHBA<3.0 mmol·L^-1， n=10） were collected at different time points （09：00， 17：00 and 01：00）. The blood ketone blood glucose meter and cholesterol determination kit were used to determine the diurnal changes of glucose and lipid metabolism indexes of dairy cows in each group. The white blood cells in the blood were isolated and total RNA was extracted. The circadian changes of inflammation-related genes and circadian clock genes in white blood cells were detected by real-time fluorescence quantitative PCR. The changes of blood oxidative stress indexes were detected by ELISA kit. The results showed that compared with the healthy group， the blood β-hydroxybutyrate （BHBA） content of dairy cows in the subclinical ketosis group was significantly increased （P<0.000 1）， the blood glucose concentration （P<0.01） and the plasma cholesterol content （P<0.000 1） were significantly decreased， and the blood glucose content （P<0.05） of dairy cows in the subclinical ketosis group exhibited significant difference between day and night. The changes of hydrogen peroxide （H₂O₂） （P<0.05）， catalase （CAT） （P<0.05）， superoxide dismutase （SOD） （P<0.05） and β-N-acetylglucosaminidase （NAG） （P<0.05） in plasma of healthy cows had significant diurnal differences. The changes of plasma H₂O₂ （P<0.05） and CAT （P<0.01） in subclinical ketosis group were significantly different between day and night. Compared with the healthy group， the contents of H₂O₂ （P<0.01） and malondialdehyde （MDA） （P<0.000 1）， and the activities of glutathione peroxidase （GSH-Px） （P<0.05） and NAG （P<0.000 1） in the plasma of dairy cows in the subclinical ketosis group were significantly increased. The qPCR results showed that the mRNA expression of circadian clock genes BMAL1 （P<0.01）， RORα （P<0.05） and PER2 （P<0.01） in white blood cells of healthy cows had significant day-night differences. The mRNA expression of circadian clock genes BMAL1 （P<0.05）， NR1D1 （P<0.05）， RORα （P<0.01） and PER2 （P<0.05） in white blood cells of subclinical ketosis cows had significant day-night differences. Compared with the healthy group， the circadian expression patterns of BMAL1 （P<0.05）， RORα （P<0.01） and PER2 （P<0.01） mRNA in blood leukocytes of subclinical ketosis cows were significantly changed. The mRNA expression of inflammatory cytokines interleukin-6 （IL-6） （P<0.000 1）， interleukin-8 （IL-8） （P<0.05）， interleukin-10 （IL-10） （P<0.05）， interleukin-1β （IL-1β） （P<0.01）， and chemokine ligand 5 （CCL5） （P<0.01） in the blood of healthy cows showed significant circadian differences. The mRNA expression of IL-6 （P<0.01）， IL-10 （P<0.01） and IL-1β （P<0.000 1） in blood leukocytes of dairy cows with subclinical ketosis showed significant difference between day and night. Compared with the healthy group， the circadian expression patterns of IL-6， IL-10 and IL-1β mRNA in the blood leukocytes of dairy cows in the subclinical ketosis group changed， and the circadian differences in the expression of IL-8 and CCL5 mRNA disappeared. In summary， there are circadian differences in some blood glucose and lipid metabolism indexes， oxidative stress indexes， blood leukocyte circadian clock genes and inflammatory factor gene expression between healthy cows and subclinical ketosis cows. However， the circadian changes of some blood glucose and lipid metabolism indexes， oxidative stress indexes， blood leukocyte circadian clock genes and inflammatory factor gene expression in subclinical ketosis cows are significantly changed. The steady-state of circadian changes of plasma oxidation and antioxidant system is abnormal， and the circadian changes of circadian clock genes and inflammatory related gene expression in blood leukocytes are abnormal. The decrease of body resistance associated with ketosis may be related to the abnormal circadian rhythm of inflammatory factor expression in peripheral blood leukocytes of dairy cows. The above research preliminarily analyzed the characteristic changes of blood indexes of ketosis cows from the perspective of circadian clock， which provided research basis and theoretical basis for the accurate prevention and control and accurate diagnosis of ketosis in dairy cows.

The aim of this study was to understand the genomic characteristics and pathogenicity of epidemic porcine reproductive and respiratory syndrome virus （PRRSV） strains in China， and to explore their recombination and related variations. Pathological samples were collected， processed， and subjected to three consecutive blind passages in Marc-145 and PAM cells. Plaque purification was performed post-indirect IFA. Complete genomic sequences of the isolated strains were sequenced and analyzed. Piglet infection experiment was conducted for systematic assessment of the strains’ pathogenicity. In this study， six PRRSV type 2 strains were isolated and named WH-2021A， GZ-2021W， SX-2022M， HB-2021A， H-1， and C-1. WH-2021A， GZ-2021W， and SX-2022M belong to NADC30-like strains of L1.8.GZ-2021W is a recombinant strain with NADC30 as the main parent and highly pathogenic strain HuN4 as the secondary parent. SX-2022M is a recombinant strain with NADC30 as the main parent and classical strain CH-1a as the secondary parent. HB-2021A belongs to the L3.5 QYYZ branch， while H-1 and C-1 belong to the L8.7 highly pathogenic strain and the L5.1 classical strain， respectively. Piglets infection experiments showed that WH-2021A had the strongest pathogenicity， and the mortality rate of piglets was 100%， followed by H-1， with a mortality rate of 80%. The two recombinant strains of GZ-2021W and SX-2022M could cause a series of clinical lesions， but no piglets died. Pathological examination showed that the lesions of lung， lymph nodes and tonsils in the infected group were obvious. In this study， 6 epidemic PRRSV strains were isolated， and their genomic characteristics and pathogenicity were explored， which provides early warning of PRRS immune prevention， as well as critical data support for the recombinantion and evolutionary analysis of RRRSV-2.

This study aimed to prepare a positive control for RT-qPCR detection of Porcine Reproductive and Respiratory Syndrome Virus genotype 2 （PRRSV-2） by utilizing the property of MS2 phage capsid protein to self-assemble and encapsulate exogenous nucleic acids into armored RNA， and to investigate its stability and RNase resistance. The specific target gene （conserved region of ORF7） of PRRSV-2 was inserted downstream of the PAC site in the pET28a（+）/CP-A recombinant plasmid to construct a recombinant vector. Through prokaryotic expression， ammonium sulfate precipitation， and gel filtration chromatography purification， uniform MS2 virus-like particles （VLPs） with a diameter of 20-28 nm were obtained. Benzonase nuclease digestion and RT-qPCR were used to verify its thermal stability and RNase resistance. The recombinant vector containing MS2 phage capsid protein， PAC site， and PRRSV-2 target gene was successfully constructed. After purification， armored RNA encapsulating the target gene was formed， which was resistant to RNase degradation and stable for 7 days at 37 ℃ under sterile conditions. This armored RNA can serve as a positive control for PRRSV-2 RT-qPCR detection. Compared with traditional synthetic nucleic acids， its advantages include： 1） simulating the nucleic acid state of real viruses and monitoring the entire detection process； 2） strong RNase resistance； 3） intra- and inter-batch coefficients of variation （CV） of <2% among 3 batches； 4） stable storage at 37 ℃ for 7 days. It provides a reliable quality control tool for PRRSV-2 detection.

This study aimed to establish a rapid and effective indirect ELISA method for detecting Listeria monocytogenes infection in samples. The InlB protein was expressed and purified in vitro using a prokaryotic expression system. The optimal parameters for the indirect ELISA method were determined through optimization， an indirect ELISA method was established. Soluble expression of InlB protein was achieved using the pCold II-10 × His vector. Optimal conditions determined by a grid-screen method were： InlB protein coating concentration of 4 μg·mL^-1， blocking with 10% skim milk powder at room temperature for 2 hours， optimal serum dilution of 1∶400， incubation at 37 ℃ for 1 hour， the optimal dilution for the secondary antibody was 1∶2 500， with incubation at 37 ℃ in the dark for 8 min. The positive/negative cutoff value was determined to be 0.37. The indirect ELISA method established in this study reacted only with L. monocytogenes-positive sera， maintaining positive detection at a serum dilution of 1∶800. The coefficient of variation （CV） for intra-batch replicates ranged from 2.79% to 6.64%， while the CV for inter-batch replicates ranged from 2.61% to 7.75%. Compared with a commercial kit， the overall agreement rate was 95.83%. The indirect ELISA method established in this study for detecting L. monocytogenes-infected samples exhibits high specificity， sensitivity， and reproducibility. It can be used for rapid detection of L.monocytogenes-infected samples， providing a reference for monitoring listeriosis.