China is a major consumer of pork， and improving pork production and meat quality is the key research direction of pig genetic improvement. Pork quality traits are affected by multiple factors such as breed， feeding management， nutrient composition， environmental factors and post-slaughter treatment， and the regulatory mechanism is relatively complex. It is difficult for traditional evaluation methods to fully reveal the underlying genetic mechanisms affecting pork quality changes. With the development of high-throughput sequencing technology， the combination of genomic， transcriptome， proteomic， epigenome， metabolomic and microbiome technologies provides a more comprehensive perspective for in-depth understanding of the formation mechanism and genetic basis of pork quality traits. This paper focuses on reviewing the application progress of omics technology and multi-omics combined methods in pork meat quality traits， and discusses the challenges which the technology faces and its future development prospects， aiming to provide methods for the analysis of the genetic mechanism of meat quality traits and ideas for pig molecular breeding.

With the development of the pig industry， carcass traits have become not only crucial for pig breeding but also of increasing concern to consumers. However， traditional phenotypic data collection for carcass traits requires slaughtering pigs before measurement， making it impossible to directly select elite breeding individuals. Instead， selection must rely on progeny or sibling testing， leading to issues such as low accuracy of estimated breeding values， high breeding costs， and poor selection efficiency. Therefore， how to assess pig carcass traits using non-destructive measurement methods has remained a key research focus. Non-invasive techniques， such as computed tomography （CT）， magnetic resonance imaging （MRI）， dual-energy X-ray absorptiometry （DXA）， and the most commonly used ultrasound technology， can provide critical information for in vivo assessment of pig carcass traits. This， in turn， offers valuable references for pig breeding， quality control， pricing， and processing. Furthermore， with the continuous advancement of artificial intelligence， integrating modern breeding techniques， non-invasive internal detection technologies， and the Internet of Things （IoT） will optimize genetic parameter estimation for breeding selection， thereby providing core support for precision pig farming.

Canines play multiple roles in human society by virtue of their highly developed olfactory system. Understanding the physiological and anatomical basis of the canine olfactory system is crucial for unraveling the mechanisms of their exceptional olfactory capabilities， comprehending their behavior and evolution， and optimizing their applications across various fields. This review systematically summarizes the key anatomical structures and physiological functions of the canine olfactory system， covering the nasal cavity architecture， characteristics of the olfactory epithelium， olfactory receptor mechanisms， and neural pathways. It specifically elaborates on the structures of the main and accessory olfactory systems， as well as the anatomy and function of the olfactory bulb， while further exploring the connectivity mechanisms of canine olfactory pathways and their impact on cognition. Finally， the review outlines current research limitations and proposes future research directions， aiming to establish a comprehensive anatomical and physiological foundation for a deeper understanding of this core sensory system in canines.

Feed costs account for about 60% to 70% of the total cost of livestock and poultry farming， and feed efficiency（FE） traits， as a core indicator reflecting the animal's feed utilization ability， are directly related to breeding benefits and resource utilization efficiency. Improving FE is the top priority for the current livestock industry to achieve cost reduction， efficiency improvement and sustainable development. With the rapid development of sequencing technology， the application of single-omics sequencing technology in the research related to livestock and poultry FE traits has increased， generating a large amount of data related to FE traits， including genome， epigenome， transcriptome， proteome， metabolome and microbiome. These data are of great significance for studying and revealing the complex regulatory process of livestock and poultry FE traits. However， it is difficult to effectively reveal the complex mechanism of livestock and poultry FE traits by relying solely on single-omics sequencing technology. By integrating multi-omics technologies， its internal mechanism and phenotypic association can be systematically and comprehensively analyzed. Based on the current research progress， this article systematically explains the concept of FE and commonly used evaluation indicators， as well as the automated measurement equipments for livestock and poultry FE. It also focuses on reviewing the analytical methods of single-omics and integrated multi-omics and their applications in livestock and poultry FE research， in order to provide reference and ideas for in-depth research in this field.

Rumen microorganisms play a vital role in the digestive systems of ruminants， significantly affecting their lactation and health. The composition of this microbial community directly impacts lactation performance of animals. Consequently， regulating the types and quantities of these microorganisms can improve milk yield and quality. Conversely， an imbalance in the rumen microbial community can cause disease in animals. This paper reviews the diversity and composition of rumen microbial communities， focusing specifically on its impact on lactation and disease susceptibility in ruminants. The aim is to lay the groundwork for studying rumen microbial diversity and function， and to elucidate the molecular mechanisms by which these microorganisms regulate production performance and health. It also may offer valuable references for formulating more effective feeding management strategies for ruminants.

Animal gut microbiome is a complex ecosystem composed of bacteria， fungi， protozoa， bacteriophages， and other microorganisms. Through synergistic interactions， it maintains intestinal microecological balance and deeply participates in host physiological regulation， immune responses， and metabolic functions. Dysbiosis of gut microbiota not only restricts host growth and development， causes immune disorders， and metabolic diseases but also interacts with host genomes via microbial metabolites to affect epigenetic modifications （e.g.， DNA methylation）， thereby exerting profound impacts on host health. As a key epigenetic mechanism， DNA methylation shapes host physiological phenotypes by regulating gene expression and maintaining genomic stability. However， the collaborative mechanisms between DNA methylation and gut microbiota remain unclear. This paper reviews the interaction patterns between gut microbiome and DNA methylation， focusing on the regulatory pathways of microbial metabolites on host methylation modifications and the potential mechanisms by which they jointly influence animal health， growth and development， and production performance. Aiming to provide a theoretical basis and research perspective for optimizing animal production performance and regulating host health， and to promote the application of research on gut microbiota-epigenetic interactions in the livestock sector.

Trichomoniasis in cattle is a sexually transmitted disease caused by Trichomonas foetus， characterized by inducing bovine infertility， vaginitis， endometritis， abortion， and occasionally pyometra， and because of the insidious onset of the disease， farmers do not pay enough attention to it， thus causing major economic losses to the farming industry. Currently， the prevention and control of bovine trichomoniasis consists mainly of vaccination and medication， but due to the poor effectiveness of its control， the prevention and control of the disease still face serious challenges. Therefore， this review summarises and discusses the biological characteristics of T. foetus in cattle， and its adhesion and toxicity damage to host cells， the impact of reproductive system， the mechanism of immune activation and evasion， with a view to providing core theoretical support for the development of novel diagnostic reagents， targeted drug design and long-lasting vaccine development.

Brucellosis， an important zoonotic disease caused by Brucella infection， poses significant threats to animal health and public health security. Therein abortion induced by Brucella infection causes substantial losses and hazards to the livestock industry， and is currently a key and challenging issue in brucellosis research though the underlying mechanism remains unclear. Based on this， this review comprehensively summarizes the molecular mechanisms of Brucella-induced abortion from the perspective of pathogen-host interactions， mainly including： 1） The immune evasion mechanisms of Brucella， such as various immune evasion strategies mediated by lipopolysaccharide （LPS） and type IV secretion system （T4SS）； 2） The imbalance of the embryonic immune microenvironment triggered by infection， involving disruption of immune cell homeostasis， destruction of hormonal regulation homeostasis and alterations in the metabolic microenvironment； 3） The molecular mechanisms of placental tissue damage caused by Brucella infection， including impairment of trophoblast cells and dysfunction of decidual cells. Brucella induce pregnancy abnormalities through a three-stage cascade amplification effect of "Immune evasion-Microenvironment imbalance-Direct damage". This review aims to deepen the understanding of the mechanisms of brucellosis-induced abortion and provides new directions and references for the development of precise diagnostic technologies and novel vaccines against brucellosis.

Infection by Mycoplasma hyopneumoniae （Mhp） is the primary etiological agent responsible for mycoplasmal pneumonia of swine （MPS）. In cases of co-infection with other respiratory pathogens， including porcine reproductive and respiratory syndrome virus， porcine circo-virus type 2， Streptococcus suis， Glaesserella parasuis， and Actinobacillus pleuropneumoniae， the condition may progress to porcine respiratory disease complex （PRDC）， which presents with significantly greater clinical severity. This synergistic pathogenesis results in elevated mortality rates， substantially compromises swine health and productive performance， and inflicts considerable economic losses upon the swine industry. Antibiotic treatment， although capable of alleviating clinical manifestations such as coughing and dyspnea， fails to eradicate the underlying infection and may potentially promote the development of antimicrobial resistance. Accordingly， the optimization and development of vaccines targeting this pathogen have emerged as a critical research direction in the field of veterinary medicine. However， conventional vaccines continue to demonstrate persistent limitations regarding protective efficacy and acceptability of administration protocols. Consequently， the development of novel vaccine platforms with enhanced safety and efficacy profiles has become an imperative requirement. Subunit vaccines， engineered based on immunodominant components derived from the pathogen， demonstrate significant advantages including high antigen purity， superior safety profiles， and favorable stability characteristics. Owing to these attributes， they have consequently emerged as a focal research area in Mhp vaccine development both domestically and internationally. This review comprehensively examines the challenges in Mhp vaccine research， with a particular focus on recent advancements in subunit vaccine development. It provides an in-depth analysis of their application advantages， candidate antigen screening strategies， and progress in clinical implementation. By synthesizing existing research， we aim to provide theoretical insights for researchers and promote innovation in Mhp vaccine development.

Marek's disease （MD） is a highly contagious infectious disease caused by Marek's disease virus （MDV）， which is characterized by severe immunosuppression， nerve lesions and T-cell lymphoma. Currently， the main prevention and control strategy for MD relies on vaccination with artificial attenuated or natural avirulent vaccine strains such as CVI988/Rispens， 814， SB-1 and HVT Fc-126. The above vaccines can induce both cellular and humoral immune responses in the host to effectively control viral infections. MDV genome， characterized by its large capacity and stable replication， can persistently stimulate the host immune response， thereby making it an ideal live recombinant vaccine vector. By expressing protective antigens from other pathogens， recombinant MDV vaccines can confer cross-protection and prevent infections caused by multiple pathogens. In recent years， based on homologous recombination， bacterial artificial chromosomes， Fosmid libraries， and gene editing technologies such as CRISPR/Cas9， researchers have successfully constructed a series of MDV recombinant live-vector vaccines， which have shown promising applications in the prevention and control of various avian diseases. This review summarizes recent advances in the construction strategies of recombinant MDV， MDV-vectored vaccines incorporating protective antigens from diverse pathogens， and the evaluation of their immunoprotective efficacy. The aim is to provide a theoretical foundation and technical references for developing safer and more efficient multivalent vaccines.

This study aimed to identify circLMO7 involved in porcine skeletal muscle development and to elucidate its underlying molecular mechanisms. This study isolated and cultured skeletal muscle satellite cells （SMSCs） from the extensor digitorum longus muscle of 1-day-old Jinfen White pigs. The circular characteristics of circLMO7 were verified by Sanger sequencing， RNase R， and actinomycin D treatment. After overexpression and interference of circLMO7 （3 replicates per treatment）， qPCR， Western blot， EdU， CCK-8， cell cycle analysis， and immunofluorescence were used to detect its functions in the proliferation and differentiation of porcine skeletal muscle satellite cells. The targeting relationship between circLMO7 and miR-145-5p was verified by dual-luciferase reporter gene assay and rescue experiments， revealing that it might regulate cell proliferation and differentiation through the miR-145-5p/IGF1R/AKT signaling pathway. As a result， a new circRNA called circLMO7 was identified in the transcriptome sequencing data of the longissimus dorsi of Jinfen White pigs， and transcribed from exon 3， 10 and 11 of LMO7 gene， which was expressed in most tissues of Jinfen White pigs， especially in the lung. Moreover， the expression of circLMO7 first decreased and then increased with the differentiation of skeletal muscle satellite cells. It was found that overexpression of circLMO7 promoted proliferation and differentiation of porcine SMSCs， while interference results showed the opposite effect. In addition， the results of the dual-luciferase reporter gene and rescue test showed that circLMO7 might act as a ceRNA， regulated the expression of IGF1R through adsorption of miR-145-5p， and then mediated the AKT signaling pathway， thus promoting the proliferation and differentiation of porcine SMSCs. The results of this study indicate that circLMO7 can alleviate the inhibitory effect of miR-145-5p on the IGF1R/AKT pathway， promote the proliferation and differentiation of porcine SMSCs， and thus participate in skeletal muscle development.

This study aims to investigate the effect of miR-885-5p on the proliferation and differentiation of porcine primary myoblasts by targeting COL1A1 and explore its underlying mechanisms. High-throughput sequencing was first performed to analyze the miRNA transcriptome in muscle samples of Shaziling pigs at different ages. Subsequently， RT-qPCR was used to detect the expression levels of miR-885-5p in various tissues and primary myoblasts of the pig. An in vitro model of porcine primary myoblasts was established， and miR-885-5p was overexpressed or inhibited. The effects of miR-885-5p on cell proliferation and differentiation were evaluated through cell cycle analysis， EdU labeling， CCK-8 assays， flow cytometry and immunofluorescence experiments. Bioinformatics tools were employed to predict the target genes of miR-885-5p， followed by dual-luciferase reporter assays to verify its targeting effect on COL1A1. Furthermore， the expression of COL1A1 in various tissues and porcine primary myoblasts was examined， and COL1A1 was interfered to assess whether miR-885-5p regulates myoblast proliferation and differentiation via COL1A1. Finally， Western blot was performed to analyze the effects of miR-885-5p on muscle fiber type conversion and its relationship with the PI3K/AKT signaling pathway. The results showed： 1） The expression of miR-885-5p significantly increased with age in the longissimus dorsi and soleus muscles. In porcine primary myoblasts， miR-885-5p expression was gradually elevated during the proliferative phase and significantly decreased during the differentiation phase. Overexpression of miR-885-5p significantly promoted myoblast proliferation， enhanced cell viability， and upregulated the expression of related genes and proteins， but inhibited cell differentiation， leading to a decrease in myotube formation and lower levels of differentiation markers and proteins. Inhibition of miR-885-5p yielded the opposite effects. 2） Sequence prediction and dual-luciferase reporter assays confirmed that miR-885-5p directly targeted the 3' UTR of COL1A1， and their expressions were significantly negatively correlated in the muscle tissue and primary myoblasts of Shaziling pigs. Furthermore， interference with COL1A1 significantly promoted myoblast proliferation and inhibited differentiation. 3） Overexpression of miR-885-5p and interference with COL1A1 reduced the protein expression of MyHCI and PPARβ/δ while upregulating MyHC-IIb expression. Inhibition of miR-885-5p caused the opposite effects， indicating that miR-885-5p regulates muscle fiber type conversion by targeting COL1A1. Additionally， overexpression of miR-885-5p and interference with COL1A1 significantly decreased the phosphorylation levels of PI3K and AKT， while inhibition of miR-885-5p increased phosphorylation levels. This study suggests that miR-885-5p promotes the proliferation and inhibits the differentiation of porcine primary myoblasts by targeting COL1A1 and regulating the PI3K/AKT signaling pathway， while promoting the conversion of muscle fibers towards a glycolytic type.

This study aimed to investigate the potential of integrating Meta-GWAS and pathway annotation information through the PoPS method in genomic selection for growth traits in Large White pigs， and to evaluate its effects on prediction accuracy and heritability enrichment. Phenotypic and genotypic data from 2 857 Large White pigs raised in a national core breeding farm in southern China were used. The studied traits included average daily gain at 100 kg （ADG100） and loin muscle depth at 100 kg （LMDEP100）. Gene feature matrices were constructed based on Meta-GWAS results from PigBiobank and GO/KEGG annotation information， and PoPS scores were calculated.Using 5 different thresholds （TOP_0.1%， TOP_0.5%， TOP_1%， TOP_5%， TOP_10%）， feature genes and their associated genetic markers were selected； meanwhile， an equal number of genes were randomly sampled as a control set. These genome feature sets were incorporated into the GFBLUP model and compared with the baseline GBLUP model. The PoPS-based approach under the optimal threshold was further compared with TWAS and colocalization methods. In addition， LDAK was used to estimate heritability enrichment for the feature gene sets， providing a comprehensive evaluation of the performance of the PoPS method in genomic selection. For both ADG100 and LMDEP100， GFBLUP models outperformed GBLUP overall. In particular， when using the TOP_0.5% of feature genes as prior information， the prediction accuracy of the GFBLUP model for ADG100 increased by 10.17% compared with GBLUP， and also outperformed GFBLUP models based on TWAS and colocalization features. Moreover， genomic features selected by PoPS showed stronger heritability enrichment for target traits. Compared with GBLUP， PoPS consistently improved prediction accuracy， with the most pronounced effect observed at the TOP_0.5% threshold. Relative to TWAS and colocalization， PoPS demonstrated superior performance， while the selected genomic features explained more heritability and showed stronger enrichment effects.

This study aimed to construct a backfat thickness correction formula applicable to Qinglian Black pigs at 85-115 kg body weight， and to identify candidate genes affecting this trait through genome-wide association study （GWAS）. We measured live backfat thickness in 393 Qinglian Black pigs from the same batch， raised under identical conditions with good growth and development performance. Based on the correction model framework of the U.S. National Swine Improvement Federation （NSIF）， the nonlinear least squares estimation was used， the correction coefficient was calculated， and a backfat-thickness correction formula was constructed. Furthermore， the genome-wide association study （GWAS） was performed on corrected backfat thickness using the 66K liquid-phase chip and the population data. The Qinglian Black pig 85-115 kg backfat thickness correction formula was established by using nonlinear least squares estimation （boar correction coefficient： 28.192， R²=0.841 5； sow correction coefficient： 34.648， R²=0.725 9）. Ten significant single nucleotide polymorphisms （SNPs） associated with backfat thickness were identified， which distributed on chromosomes 1， 2， 10， and 11 （at a false discovery rate （FDR）0.01）. Gene annotation and pathway enrichment analysis were performed on genes retrieved from significant SNPs， identifying the gene MAPKAP1， GRK2 and LMX1B as potential candidate genes influencing the backfat thickness in Qinglian Black pigs. This study established a 85-115 kg backfat thickness correction system for Qinglian Black pigs and identified MAPKAP1， GRK2， and LMX1B as candidate genes for backfat thickness traits， providing theoretical and technical support for precision breeding in local pig breeds.

The study aimed to comprehensively assess the egg-production genetic potential of the Taihang chicken， a unique local breed from Hebei Province， under two distinct conservation breeding strategies and to validate the effectiveness of low-coverage whole-genome resequencing technology in constructing genomic relationship matrices and estimating genetic parameters for traits with low-heritability. A total of 956 Taihang chickens from two conservation populations were selected： the Tian-Kai （TK） population （family-based equal-contribution conservation） and the Ning-Jin （NJ） population （random retention conservation）. After quality control， 924 individuals were remained （382 from TK， 542 from NJ）. Seven key reproductive traits were recorded： age at first egg （AFE）， early egg production（EN1）， peak egg production （EN2）， late egg production （EN3）， total eggs to 300 days （EN300）， total eggs to 400 days （EN400）， and broodiness. Genomic DNA libraries were constructed using a Tn5 transposase-based protocol and sequenced on a next-generation platform， sequencing data with an average depth of 1.21× per individual was obtained. Base-Var （v0.0.1.3） was employed to identify polymorphic loci from the aligned low-coverage whole-genome resequencing data and infer allele frequencies for SNP genotyping. Genotype imputation was performed with STITCH to obtain high-density genome-wide SNP data. Based on these high-density genome-wide SNP data， a genomic relationship matrix （GRM） was constructed， and the heritability and genetic/phenotypic correlations between traits were estimated using bivariate animal model in BLUPF90 software. The results indicated that the heritabilities of egg production traits in both conservation populattions were generally low to moderate （ranging from 0.08±0.14 to 0.31±0.16）， but significant differences existed between the two populations： The TK population exhibited a significantly higher heritability for AFE （0.32±0.15） compared with NJ population （0.16±0.15）， whereas NJ population displayed a significantly higher heritability for broodiness （0.30±0.19） than TK population （0.08±0.16）， indicating that conservation strategy may shift allele-frequency distributions. Correlation analyses revealed significant negative associations between egg numbers and both AFE and broodiness， with distinct correlation patterns between the two populations. Notably， total egg production during the early laying period （up to 30 weeks） was highly correlated with total egg production up to 400 days， suggesting its utility as an early selection index. The study also confirmed that GRMs constructed from low-coverage whole-genome resequencing effectively overcome the lack of pedigree information in conservation populations and provide reliable genetic parameter estimates. In summary， first， low-coverage whole-genome resequencing technology has significant advantages for estimating genetic parameters of traits with low heritability. Second， different conservation strategies markedly influence the genetic structure and trait performance of Taihang chickens： the family-based equal- contribution strategy preserves genetic variations more favorable for egg production， while the random retention strategy retains more genetic variations associated with broodiness. Finally， early egg-production traits reliably predict overall laying performance in both populations. Consequently， for genetic improvement programs based on these two conservation populations， it is recommended to advance breeding value estimation and selection decisions to the early laying stage （approximately 30 weeks） and to tailor selection emphasis according to the genetic structure of each population， thereby achieving both protective utilization of genetic resources and efficient breeding objectives.

This study aimed to identify candidate genes and genetic markers associated with traits such as birth weight， weaning weight，3-month weight and other related traits in East Friesian and Hu crossbred sheep using the genome-wide association study （GWAS） methods， thereby accumulating a theoretical foundation for their selective breeding. Whole blood samples were collected from 1 554 healthy East Friesian and Hu crossbred sheep as experimental materials. Descriptive statistical analysis was performed for the traits of birth weight， weaning weight， 3-month weight， average daily gain from birth to weaning， average daily gain from weaning to 3 months， and average daily gain from birth to 3 months. Genetic variant data for individuals were obtained through whole-genome resequencing. GWAS for each body weight trait was conducted using PLINK， GEMMA， and R software， employing the model： y=W\mathrm{\alpha }+X\mathrm{\beta }+\mathrm{u}+\mathrm{\epsilon }. At the genome-wide significance threshold， a total of 48 significantly associated loci were identified （P2.64×10^-9）； under the suggestive significance threshold， 50 associated loci were detected （P5.29×10^-8）. Through annotation of the associated single nucleotide polymorphism （SNP） loci， candidate genes were screened， yielding 4 associated with birth weight， 1 with weaning weight， 3 with 3-month weight， 43 with average daily gain from birth to weaning， 3 with average daily gain from weaning to 3 months， and 1 with average daily gain from birth to 3 months. Haplotype analysis revealed that multiple significant SNPs within the target genes were located within haplotype blocks. The analysis suggests that genes such as DMRTA1， TMOD2， RPRM， NEDD9， HIVEP1， EYA2， PRDM5， and RSPH14 can be considered key candidate genes for body weight traits in East Friesian and Hu crossbred sheep. These findings provide important genetic variant loci and candidate genes for subsequent research on important economic traits in East Friesian and Hu crossbred sheep， offering crucial foundational data for further refining molecular breeding methods and accelerating breeding progress.

This study proposes the cumulative months in milk by 66 months of age （MIM66） as a new measurement for productive life in Holstein cattle. By integrating key environmental variables and a multi‑parity cumulative mechanism， we establish a predictive model for productive life， enabling accurate forecasting of the lifespan of cows within the herd. This approach provides support for conducting genetic evaluations of productive life. Based on calving， culling and DHI test records of 88 557 Holstein cows from nine large-scale dairy farms in Shandong Province， 8 015 cows aged over 66 months were selected for analysis. Using a general linear model， variables including first lactation 305-day milk yield， first lactation somatic cell score， age at first calving， and season at first calving were incorporated. A multi-parity cumulative mechanism was introduced， with a lactation length cap of 10 months per parity. Prediction models were constructed based on different age nodes from 30 to 64 months. The model performance was validated using data from 2 941 Holstein cows， with the coefficient of determination （R²） and Pearson correlation coefficient used to evaluate model fit and prediction accuracy. The results showed that the coefficient of determination （R²） of the MIM66 model significantly increased with advancing prediction age and additional variables. At 30 months of age （4 variables）， R² was 0.213； by 36 months （8 variables）， R² rose to 0.520， reaching above 0.9 by 50 months of age. External validation with 2 941 cows demonstrated that the model’s predictive accuracy increased with age： when cows reached 48 months of age， the prediction accuracy exceeded 0.75， and by 58 months it surpassed 0.85. The overall Pearson correlation between observed and predicted MIM66 values across all age points was 0.712 （P0.001）. The model was subsequently applied to predict MIM66 for 28 665 in-herd Holstein cattle， with an overall average MIM66 of 34.0 months. The results indicate that the prediction model can effectively forecast the productive life （MIM66） of Holstein cattle， aiding farms in early screening of cows with longevity， optimizing culling decisions， and laying the foundation for genetic evaluation of productive life.

This study aimed to investigate the expression pattern of circANKEF1 in subcutaneous adipose tissue of Jiaxian Red cattle at different ages， as well as the effect of circANKEF1 on the proliferation and differentiation of bovine preadipocytes. This study used dorsal subcutaneous adipose tissue and preadipocytes from 1-， 3- and 5-year old Jiaxian Red cattle （n=3） as experimental materials.circANKEF1 was identified and its cellular localization was determined by semi-quantitative PCR and quantitative real-time PCR （qRT-PCR）， and the expression pattern of circANKEF1 in subcutaneous adipose tissue of Jiaxian Red cattle at different ages was clarified.The characteristics of circANKEF1 were analyzed by RNase R resistance assay， nuclear‑cytoplasmic separation， and fluorescence in situ hybridization （FISH）.Based on circANKEF1， small interfering RNA （siRNA） and overexpression vectors were designed and transfected.Lipid droplet accumulation was observed by Oil Red O staining.The mRNA and protein expression levels of adipogenic marker genes （PPARγ， C/EBPα） and proliferation marker genes （PCNA， CDK2） were detected by qRT-PCR and Western blot （WB）， respectively.Cell viability and proliferation were measured by CCK-8 assay， EdU staining， and flow cytometry.These experiments were performed to comprehensively explore the effects of circANKEF1 on the proliferation and differentiation of bovine preadipocytes. The results showed that circANKEF1 was a circular non-coding RNA highly expressed in both the cytoplasm and nucleus， and its expression in subcutaneous adipose tissue was significantly increased with age （P0.01）. Gain-of-function assays indicated that in the in vitro differentiation model of bovine preadipocytes， overexpression of circANKEF1 significantly increased lipid droplet accumulation （P0.01）， significantly upregulated the mRNA and protein levels of adipogenic marker genes PPARγ and C/EBPα （P0.01） and proliferation marker genes PCNA and CDK2 （P0.01）， and as determined by CCK-8， EdU and flow cytometry cell cycle analysis， significantly enhanced cell viability （P0.01）， promoted cell proliferation （P0.05） and elevated the proportion of S-phase cells. In contrast， inhibition of circANKEF1 suppressed lipid droplet accumulation in adipocytes， significantly decreased the mRNA and protein levels of adipogenic marker genes PPARγ （P0.001） and C/EBPα （P0.05） as well as proliferation marker genes PCNA and CDK2 （P0.05）， and as showed by CCK-8 and EdU assays， markedly reduced cell viability （P0.05） and the number of positive cells. These results suggest that circANKEF1 can promote the proliferation and differentiation of bovine preadipocytes. These findings provide new insights for further understanding the molecular mechanisms underlying adipose development.

This study aimed to systematically evaluate the genomic prediction accuracy of 10 methods， including SVM model， Random Forest model， GBLUP_D model， GBLUP_A model， RRBLUP model， Bayes B model， PLS model， Bayes C model， Bayes A model and Bayes Lasso model for antler weight trait in sika deer， identify the optimal model， and provide new strategies for antler weight breeding in China. In this study， 383 sika deer were selected as the research population. The heritability of antler weight of sika deer was estimated by constructing genetic relationship matrix of whole genome marker loci. The prediction accuracy of 10 genomic selection models was compared by 5-folds cross-validation method. The heritability of antler weight in sika deer was 0.296 3， which was of medium heritability， indicating that the degree of genetic factor regulation of this trait is moderate and it has certain potential for genetic improvement. By using 5-fold cross-validation to compare the prediction accuracy of 10 genomic selection models， it was found that the Random Forest model had the highest prediction accuracy （0.533 1）， the dominant effect model GBLUP_D （0.500 1） was superior to most additive models， while PLS had the lowest （0.398 0）. These results suggested that the genetic regulatory mechanism of antler weight in sika deer was not dominated by a single additive effect， but might be jointly regulated by major genes， dominant effects， and complex gene interactions. Genomic selection technology effectively predicted antler weight in sika deer， and the Random Forest model achieved the highest prediction accuracy. Dominance effects play an important role in the genetic architecture of antler weight， suggesting that both additive and dominance effects should be considered in breeding programs.

The purpose of this study was to analyze the population variation analysis of sperm storage capacity and compare the metabolic characteristics of high and low sperm storage capacity groups in the late laying period of White Leghorns laying hen. A total of 1 100 90-week-old White Leghorns laying hens were selected， and the phenotypes such as fertility rate and effective fertilization days were recorded. Based on the phenotype （egg laying rate75%）， 10 individuals with high fertility rate and 10 individuals with low fertility rate were selected， which were respectively defined as high and low sperm storage capacity groups. Serum was separated to detect the levels of estradiol （E₂）， progesterone （P₄）， prostaglandin F_2α （PGF_2α） and testosterone （T）. At the same time， the differential metabolites between high and low sperm storage capacity groups were screened by serum non-targeted metabonomics， and KEGG pathway enrichment analysis was carried out. The results showed that the indexes of sperm storage capacity of hen population showed great individual variation in the late laying period， among which the coefficient of variation of intermittent fertilization intensity and the maximum number of consecutive fertile eggs were higher， which were 95.57% and 52.76% respectively. The levels of serum T and PGF2α were higher in the group with high sperm storage capacity （P0.05）. A total of 58 differential metabolites were identified， of which 25 were up-regulated and 33 were down-regulated in the group with high sperm storage capacity. KEGG enrichment analysis showed that sperm storage capacity was highly correlated with lipid metabolism pathway. At the same time， the significant increase of serum diketone gulonic acid and 24-methylene cholesterol in the group with high sperm storage capacity is closely related to its antioxidant capacity. In conclusion， high sperm storage capacity group has stronger antioxidant capacity and lipid metabolism activity， thus preventing the damage of lipid peroxidation to sperm， and enhancing the sperm storage capacity.

This research aimed to investigate the regulatory mechanism of miRNA-1576 on the physiological functions of endometrial epithelial cells in sheep. The sheep endometrial epithelial cells （EECs） used in this study were previously isolated and cultured in our laboratory. The experiment consisted of NC， miR-1576 mimic， NC-IN and miR-1576 inhibitor groups. Firstly， we detected the effect of miR-1576 on SLC7A5 expression using cell transfection and RT-qPCR. Dual-luciferase reporter assay was conducted to validate the targeting relationship between miR-1576 and SLC7A5. Finally， functional assays were performed by inhibiting and overexpressing miR-1576 in EECs in vitro. Techniques such as Western blot， CCK-8， flow cytometry， and wound healing assays were used to elucidate the role of miR-1576 in EECs. The results demonstrated the targeted regulatory relationship between miR-1576 and SLC7A5. Furthermore， inhibition of miR-1576 significantly promoted EEC proliferation， cell cycle progression， and the expression of related proteins （PCNA， c-MYC， and Cyclin D1）， as well as enhanced wound healing. In contrast， overexpression of miR-1576 significantly inhibited EEC proliferation， cell cycle progression， and the expression of related proteins （PCNA and Cyclin D1）， as well as inhibited wound healing. In summary， miR-1576 can target and regulate SLC7A5 expression， thereby inhibiting physiological processes such as proliferation， cell cycle progression， and migration in sheep endometrial epithelial cells. This study provides new insights into the regulatory mechanisms of miRNA in uterine physiological functions of prolific sheep.

The study aimed to investigate the effect of GnIH on gonadal development in pigeons. In this study， graded follicular granulosa cells and theca cells of Mimas breeding pigeons were subjected to primary culture， followed by separate treatments with different concentrations of GnIH polypeptide or transfected them with siRNA-GnIH for interference （n=6）. For the GnIH polypeptide treatment assay， the control group （Group I） was treated with PBS at a GnIH polypeptide concentration of 0 ng·mL^-1， while the experimental groups （Groups II， III， and IV） were treated with GnIH polypeptides at concentrations of 1， 10， and 100 ng·mL^-1， respectively. For the GnIH interference assay， the control group was transfected with siRNA-NC， whereas the experimental group was transfected with siRNA-GnIH （5 μL， 20 μmoL·L^-1）. The results showed that GnIH polypeptide treatment significantly downregulated the mRNA and protein levels of GnIH and FSHR in granulosa cells （P0.05）， significantly decreased the mRNA levels of GnIHR， LHR， and StAR in granulosa cells （P0.05）， significantly inhibited the secretion of E₂ and P₄ in granulosa cells （P0.05）， and significantly upregulated the mRNA level of GnIH in theca cells （P0.05）， while the mRNA level of GnIHR in theca cells gradually decreased with increasing treatment concentration. After siRNA-GnIH interference， the mRNA and protein levels of GnIH in both granulosa and theca cells were significantly downregulated （P0.05）， while the mRNA levels of FSHR and LHR in granulosa and theca cells were significantly upregulated （P0.05）. Additionally， the protein level of FSHR in theca cells was significantly increased （P0.05）， and the mRNA and protein levels of StAR and 3β-HSD in both granulosa and theca cells were significantly upregulated （P0.05）， along with the mRNA level of CYP19A1was significantly upregulated in granulosa cells （P0.05）. Furthermore， siRNA-GnIH promoted E₂ expression in both granulosa and theca cells （P0.05）， but inhibited P₄ secretion in granulosa cells （P0.05）. These results indicated that GnIH exerts similar effects on gonadotropin receptors and steroidogenesis in granulosa and theca cells of pigeon as observed in other avian species. GnIH polypeptide significantly suppresses gonadotropin receptor expression， downregulates steroidogenic pathway factors and inhibits steroid hormone secretion， whereas GnIH interference produces the opposite effects in granulosa cells. While GnIH polypeptide shows no significant impact， GnIH interference markedly enhances gonadotropin receptor expression， steroidogenic pathway activity and steroid hormone secretion in theca cells.

This study aims to elucidate the dynamic changes in serum reproductive hormone profiles during the estrous cycle of Tibetan sows， and to investigate the mechanisms underlying synchronous estrus induction by two distinct estrus-inducing drugs through granulosa cell culture experiments. Fifteen healthy and non pregnant Tibetan sows with similar age （285±11 d） and body weight （82.84±4.72 kg） which installed head arm vein catheterization were selected for the experiment and randomly divided into three treatments：control group （CON， n=4）， altrenogest+pregnant mare serum gonadotropin （PMSG） group （ALT-P， n=6）， and Chinese medicine aphrodisiac powder group （Aph， n=5）， with one pig per replicate and raised in a single pen. The results showed that： 1） Compared with the CON group， the area under the luteinizing hormone （LH） curve of Tibetan sows in the ALT-P and Aph groups during estrus was significantly increased （P0.05）； The area under the LH curve in the Aph group was significantly higher than that in the ALT-P group （P0.05）. 2） Compared with the CON group， the ratio of progesterone （P₄） to estrogen （E₂） （P₄/E₂） in Tibetan sows in the Aph group significantly decreased at 1， 1.5， 2， and 2.5 hours after estrus （P0.01）， while the P₄/E₂ value in the ALT-P group significantly decreased at 1.5 and 2.5 hours after estrus （P0.05）； The P₄/E₂ values in the Aph group were significantly lower than those in the ALT-P group at 1 and 2 hours after estrus （P0.05）. Further isolation of primary ovarian granulosa cells from Tibetan sows for in vitro experiments， divided into CON group， ALT group， ALT-P group， and Aph group. The results showed that compared with the CON group， 1） The levels of E₂ in ovarian granulosa cells were significantly increased in both the ALT-P group and the Aph group （P0.05）； 2） The expression of estrogen receptor （ESR） proteins was significantly upregulated （P0.05）； 3） The mRNA expression of progesterone receptor， follicle stimulating hormone receptor， cytochrome P450， estrogen synthase， and acute regulatory protein of steroid hormone synthesis were significantly upregulated （P0.05）. The Aph group also significantly upregulated the mRNA expression of ESR and 3β-hydroxysteroid dehydrogenase （P0.05）. In summary， both ALT-P and Aph can promote estrus in Tibetan sows， and the effect of Aph on regulating reproductive hormone secretion is comparable to that of ALT-P； In vitro experiments have shown that ALT-P and Aph mainly promote follicular development in Tibetan sows by upregulating the expression of steroid hormone related genes in granulosa cells.

This study aims to investigate the effects of progesterone on ruminal digestion of roughage and the composition and function of rumen microbial communities. The experiment was divided into four progesterone concentration gradients， namely the control group （no progesterone added）， the P_0.5 group （0.5 ng·mL^-1 added）， the P₅ group （5 ng·mL^-1 added）， and the P₄₀ group （40 ng·mL^-1 added）. Three non-pregnant dairy cows were selected as rumen fluid donors for in vitro fermentation. During fermentation， gas was collected and the contents of hydrogen and methane were measured. After 24 h of fermentation， samples were collected to determine the dry matter disappearance rate， volatile fatty acids （VFA）， and ammonia nitrogen （NH₃-N） contents. Metagenomic techniques were used to determine the microbial structure and function in the fermentation liquid. In this study， compared with the control group， the addition of progesterone did not significantly affect the dry matter disappearance rate （P=0.501） and the NH₃-N content （P=0.056）. The P_0.5 group had significantly higher cumulative gas production， hydrogen production and methane production during fermentation （P0.05）， and a significantly higher concentration of acetate （P=0.025）. The P₄₀ group decreased gas production by rumen microbes. Metagenomic results showed that the structure of eukaryotic microbial communities differed significantly among the groups （P=0.006）. Compared with the control group， the P_0.5 group had significantly higher relative abundances of Bacteroidota and Methanobacteriota （P0.05）， and significantly affected genes related to acetate pathways and methanogenesis （P0.05）. The P₅ group significantly increased the relative abundance of certain yeasts （P0.05）. The addition of progesterone significantly altered the abundance of some carbohydrate enzyme genes. In summary， a progesterone concentration of 0.5 ng·mL^-1 significantly enhanced the metabolic activity of rumen microbes. The addition of progesterone promoted the growth of hydrogenotrophic methanogens and yeasts. Progesterone may influence the abundance and growth metabolism of rumen microbes， thereby affecting the expression levels of carbohydrate enzyme genes and consequently promoting the efficiency of rumen utilization of complex carbohydrates.

The aim of this trial was to investigate amino acid limiting sequence of corn-miscellaneous-meal-type low protein diet in medium-growing yellow-feathered broilers aged 61-85 days by using the partial amino acid deletion method. A total of 756 sixty-one-day-old female medium-growing yellow-feathered broilers （Mahuang） were randomly allotted into nine groups （six repetitions per group and 14 chicks per repetition）， which were fed a normal-protein diet and eight low-protein diets， respectively. The Control （NP） group was fed the normal-protein diet （crude protein 16%）. The low-protein control （LP） group was fed the low-protein diet （crude protein， 14%） with the amino acid content meeting the requirement of the yellow-feathered broilers. The low-protein low-lysine （LP-Lys） group， low-protein low-methionine （LP-Met） group， low-protein low-threonine （LP-Thr） group， low-protein low-isoleucine （LP-Ile） group， low-protein low-valine （LP-Val） group， low-protein low-arginine （LP-Arg） group， and low-protein low-tryptophan （LP-Trp） group were formulated by reducing the levels of true available Lys， Met， Thr， Ile， Val， Arg， and Trp by 20% from LP group， respectively； But amino acids that did not reach the 20% reduction threshold were adjusted based on the actual deficiency value. The trial duration was 25 days. The results showed that there were no significant differences in growth performance and body quality among all groups （P0.05）. Compared to NP group， LP group tended to have higher serum albumin level （P=0.077） and jejunal villus height/crypt depth （P=0.075）. Compared to LP group， LP-Met group tended to increase feed/gain （P=0.080）， LP-Thr group had significantly lower serum immunoglobulin A （P0.05）， LP-Val group had significantly higher serum immunoglobulin A （P0.05）， LP-Thr， LP-Ile， LP-Val and LP-Arg groups had significantly higher serum total antioxidant capacity and glutathione peroxidase activity （P0.05）， and LP-Met， LP-Thr， LP-Ile and LP-Val groups had significantly lower jejunal villus height/crypt depth （P0.05）. When using the optimal feed/gain as the evaluation index， the amino acid restriction order is MetLysValIleArgThrTrp， and the appropriate ratio of Lys， Met， Thr， Ile， Val， Arg and Trp is 100∶46∶64∶67∶80∶107∶17； When using the optimal jejunal villus height/crypt depth as the evaluation index， the amino acid restriction order is ThrArgMetValIleTrpLys， and the appropriate ratio of Lys， Met， Thr， Ile， Val， Arg and Trp is 100∶45∶78∶72∶86∶118∶19； When using the optimal serum immunoglobulin A level as the evaluation index， the amino acid restriction order is IleThrArgMetTrpLysVal， and the appropriate ratio of Lys， Met， Thr， Ile， Val， Arg and Trp is 100∶43∶81∶80∶63∶124∶19.

This experiment aims to evaluate the standardized ileal digestibility of amino acids （SID_AA） in fast-growing yellow-feathered male broilers （Xiangjia yellow chicken） fed with 10 different sources of rapeseed meal， while also constructing a predictive equation for SID_AA based on the chemical composition of rapeseed meal and validating its accuracy. A total of 1 056 one-day-old Xiangjia yellow chickens were selected， and the whole experiment was divided into two phases. In the first phase （14 to 18 days old）， 660 broilers with no significant body weight differences were chosen at 14 days of age and randomly assigned to 11 groups， with 6 replicates per group and 10 chickens per replicate. In the second phase （35 to 39 days old）， an additional 396 broilers with no significant body weight differences were selected at 35 days of age， also randomly divided into 11 groups with 6 replicates per group and 6 chickens per replicate. Ten groups were fed a semi-purified diet with 10 different rapeseed meals as the sole protein source， respectively， while one group was fed a nitrogen-free diet to estimate endogenous amino acid losses， using 0.4% titanium dioxide as an indicator to measure the SID_AA in fast-growing yellow-feathered male chickens at 14 to 18 days and 35 to 39 days. The results indicated that the coefficients of variation （CV） for crude fat， crude fiber， neutral detergent fiber， acid detergent fiber， crude ash， and phytic acid content of rapeseed meal from different sources all exceeded 10%. The results of animal testing indicated that： 1） At 14~18 days of age， there was no significant difference in the tryptophan SID_AA values of 10 rapeseed meals from different sources （P0.05）， however， the SID_AA values of the other 17 amino acids were extremely significant among groups （P0.001）. At the age of 35~39 days， there were significant differences in all amino acids' SID_AA values of 10 kinds of rapeseed meal from different sources， among which the SID_AA values of tryptophan were significantly different between groups （P=0.021）， and the SID_AA values of the other 17 amino acids were extremely significant among the groups （P0.001）. 2） At 14-18 days old， a rapid SID_AA prediction equation for arginine， histidine， leucine， lysine， phenylalanine， threonine， valine， alanine， cysteine， glycine， and proline in rapeseed meal of yellow feathered meat roosters was constructed. The optimal equation was： SID_Pro=-240.466+76.851 6×Lys+9.018 9×ADF-3.754 9×NDF+119.147 1×Trp-8.858 5×CF+25.770 7×Asp ［R²=0.995 3，RSE=1.707 9，P=0.014 0］； At 35-39 days old， a rapid SID_AA prediction equation was constructed for arginine， histidine， isoleucine， lysine， threonine， valine， alanine， aspartic acid， and serine in rapeseed meal of yellow feathered meat roosters. The optimal equation was SID_Ser=-1469.202+153.938 5×Lys+32.291×ADF-12.071×NDF+1 350.345 4×Trp+440.820 3×Arg-186.959 4×Phe-17.816 4×CP ［R²=0.999 9，RSE=0.056 6，P=0.004 0］. 3） SID_AA prediction model with the best accuracy for 14-18-day-old rapeseed meal of fast-growing yellow-feathered broilers was SID_Ala. The SID_AA prediction model with the best accuracy for rapeseed meal at 35-39 days old was SID_Lys. In conclusion， significant variations exist in the SID_AA of rapeseed meal among fast-growing yellow-feathered broiler males sourced from different origins and at varying ages. The predictive model developed in this study offers substantial reference value for accurately forecasting the SID_AA of rapeseed meal in broilers.

This experiment was conducted to study the effects of metabolic energy （ME） restriction from 7 to 15 weeks of age and conversion to ad libitum （16 to 18 weeks of age ） on growth performance， intestinal morphology and cecal microorganisms of laying hens on onset of lay （18 weeks old）. Five hundred and forty Hy-Line Brown laying hens aged six weeks were randomly divided into 3 groups with 6 replicates in each group and 30 hens in each replicate. From 7 to 15 weeks of age， the experimental hens were fed the experimental diets with the same levels of other nutrients but different levels of ME （12.34 （control group）， 11.11 （12.34×90%， 90% ME group）， and 9.87 （12.34×80%， 80% ME group） MJ·kg^-1）. The hens in the control group were fed freely， and the hens in other experimental groups were fed quantitatively according to the feed intake of the hens in the control group. From 16 to 18 weeks of age， all the hens were fed freely with the same diets. The results showed that： 1） The body weight of laying hens at 15 weeks and 16 weeks of age in the ME restriction groups were significantly lower than that in the control group （P0.01）. The body weight of laying hens at 17 weeks of age in 80% ME group was significantly lower than that in control group and 90% ME group （P0.01）. At 18 weeks of age， there was no significant difference in body weight among the groups （P0.05）， but the body weight CV in ME restriction groups were significantly lower than that in the control group （P0.05）. As the increase of ME restriction from 7 to 15 weeks of age， the average daily feed intake and feed to gain ratio of laying hens from 7 to 18 weeks of age increased linearly， and the average daily ME intake decreased linearly（P0.01）. 2） There was no significant difference in digestive tract development indexes at 18 weeks of age between feed restriction groups and the control group （P0.05）， except that the relative thickness of the two gastric walls of hens in the feed restriction groups were significantly less than that in the control group （P0.05）. 3） The crypt depth of jejunum of hens in 80% ME group was significantly lower than that in control group and 90% ME group at 18 weeks of age （P0.05）， but there was no significant difference in the villus heigh and villus height/crypt depth among the groups （P0.05）. 4） ME restriction from 7 to 15 weeks of age significantly affected the α and β diversity index of cecal microorganisms in hens after switching to ad libitum for 3 weeks （18 weeks of age） （P0.05）. 5） Microbial composition analysis found that， the relative abundance of Actinobacteria and Actinobacteria in 80% ME group was significantly lower than that in control group （P0.01）， and the relative abundance of Euryarchaeota was significantly higher than that in control group and 90% ME group （P0.01）. 6） LEfSe analysis showed that the microbial groups that played important roles in the 90% ME group were concentrated in Atopiaceae， Coriobacteriales and Coriobacteria， the microbial groups that played important roles in the 80% ME group were Methanobacteria， Methanobacteriales and Methanobacteriaceae. 7） Microbial function prediction showed that ME restriction during rearing period affected the function of microbial flora of laying hen after conversion to ad libitum for 3 weeks. Cecal microorganisms in the ME restricted group were significantly enriched in starch and sucrose metabolism， galactose metabolism， pyrimidine metabolism， purine metabolism， cysteine and methionine metabolism， and citric acid cycle. In conclusion， ME restriction during the rearing period （7 to 15 weeks of age） and the conversion to ad libitum （16 to 18 weeks of age） had no adverse effects on the body weight and digestive tract development of laying hens at 18 weeks of age， and improved the flock uniformity. 80% ME restriction during the rearing period significantly improved the jejunal morphology of hens at 18 weeks of age， decreased harmful bacteria such as Actinobacteria and Proteobacteria in the cecum microbiota， and increased beneficial bacteria such as Firmicutes and Methanobacteria. The microbial biological functions were significantly enriched in the material metabolic pathways that release free energy.

Since 2010， the endemic outbreaks caused by G-II strains of porcine epidemic diarrhea virus （PEDV） have resulted in enormous losses to the swine industry in China and even worldwide. Safe and effective vaccination is the optimal approach to preventing and controlling this disease. This study developed a virus-like particle （VLP）-based vaccine candidate AP205-S1^AB using the truncated S1 protein S1^AB from PEDV-KB2013-130 （G-IIa strain） as targeted antigen. S1^AB protein was displayed on the surface of AP205-VLP via SpyCatcher/SpyTag specific covalent bond. The results showed that， after immunizing BALB/c mice with AP205-S1^AB， high levels of S1^AB and S1 protein-specific IgG antibodies were generated in mice sera. Further experiments demonstrated that these antibodies could neutralize PEDV in vitro and that the 50% sera neutralization titer reached 1∶48. Additionally， S1^AB protein-specific IgG antibodies were detected in intestinal mucosa after booster， and they were able to block PEDV from infecting Vero cells despite at low level. In summary， this study demonstrated that the AP205-S1^AB vaccine candidate exhibited strong immunogenicity in mice. The immunized mice sera could efficiently neutralize PEDV G-II strains， offering valuable insights for the development of PEDV vaccines.

This study aims to construct a Selenos-knockout BHK-21 cell line to elucidate the function of SelS protein in host antiviral defense and its regulatory mechanism on FMDV replication. Using the CRISPR-Cas9 system， a Selenos-knockout BHK-21 cell line was established： sgRNAs targeting the exon region of the Syrian hamster （Mesocricetus auratus） Selenos gene were designed and delivered via a lentiviral vector （lentiCRISPR v2）. Monoclonal cell lines were obtained through puromycin screening， and knockout efficiency was validated by sequencing and Western blot. Cell viability was assessed using the CCK-8 assay， and morphological analysis was performed. BHK21-SELENOS-KO and control cells （BHK21-Cas9-NC） were synchronously infected with FMDV. Viral replication dynamics were analyzed through cytopathic effect （CPE）， indirect immunofluorescence， Western blot， viral titer determination， and RT-qPCR. Differentially expressed genes were screened via transcriptomic RNA sequencing and validated by RT-qPCR， followed by signaling pathway enrichment analysis. Results showed that the Selenos-knockout BHK-21 cell line （BHK21-SELENOS-KO） was successfully constructed， leading to a complete loss of SelS protein function without significant alterations in cell proliferation or chromosomal stability. Viral infection assays revealed significantly enhanced FMDV replication in knockout cells： viral protein expression and viral yield increased， with mean fluorescence intensity 2.1 times higher than in control cells （P0.001）， alongside more pronounced CPE. Transcriptomic analysis identified 226 upregulated and 548 downregulated genes， with differentially expressed genes significantly enriched in immune-related pathways including cGMP-PKG， calcium signaling， and PI3K/Akt； RT-qPCR validation confirmed the sequencing results. In conclusion， SelS protein plays a crucial role in host antiviral immunity by restricting FMDV replication. The BHK21-SELENOS-KO cell line established in this study provides a novel tool for investigating virus-host interactions and offers a scientific basis for improving foot-and-mouth disease vaccine processes and developing antiviral drug targets.

This study aimed to investigate the differences between G2b and G2c subtypes of porcine epidemic diarrhea virus （PEDV） in biological properties， antigenicity， and cross-immunogenicity. Multiple sequence alignment revealed that G2c strains accumulated numerous stable amino acid mutations in critical functional domains of the S2 subunit， including HR1， HR2 and the cytoplasmic tail. Based on a reverse genetics system and the genome backbone of G2b strain， a chimeric recombinant virus harboring the S2 subunit of a representative G2c strain， named rAH2012/12-S2_QD， was successfully rescued and assessed its replication kinetics and genetic stability in Vero cells. Subsequently， the immunogenicity of the recombinant and parental viruses was evaluated in the mouse model， and cross-neutralization assays were performed. The results showed that rAH2012/12-S2_QD replicated efficiently in vitro and maintained genetic stability. Immunization with the recombinant virus induced serum IgG levels comparable to those elicited by the parental strains， indicating strong immunogenicity. Cross-neutralization tests revealed a marked reduction in neutralizing activity of G2b and G2c hyperimmune serum against heterologous strains， suggesting antigenic drift between the two subtypes. Notably， sera from rAH2012/12-S2_QD-immunized serum exhibited potent neutralizing activity against both G2b and G2c strains. Collectively， our findings demonstrated that key mutations in the S2 subunit contribute to altered viral replication and antigenicity of G2c strains. The further studies highlighted the pivotal role of the S2 subunit in cross-protective immunity. This study revealed the key role of the S2 gene in inducing neutralizing antibody production and provided a theoretical framework for the development of next generation of broad-spectrum vaccines.

Porcine circovirus type 2 （PCV2） infection is one of the major diseases harming China’s swine industry， with vaccination as the primary control strategy. To compare the immunization effects of different types of PCV2 vaccines， this study used the PCV2 recombinant baculovirus OKM strain to infect insect cells， purified the recombinant protein， and prepared a subunit vaccine； three commercial vaccines，a whole-virus inactivated PCV2 vaccine， an E. coli-expressed PCV2 subunit vaccine， and an imported baculovirus-expressed PCV2 subunit vaccine， were also selected. Each vaccine was administered to 21-28-day-old piglets confirmed negative for PCV2 nucleic acid and ELISA antibodies to evaluate immunoprotective efficacy. The results showed that the recombinant Cap protein prepared in this study formed typical virus-like particles （VLPs）. The prepared subunit vaccine and all three commercial vaccines induced PCV2 ELISA antibodies in piglets. At 35 days post-immunization， piglets were challenged with a virulent PCV2 strain and observed for 25 consecutive days. No abnormal clinical signs or pathological changes were observed in any immunized group after challenge. Although average daily gain （ADG） did not differ significantly among the four immunized groups， all were significantly higher than the non-immunized challenge control group. Protective efficiency in all immunized groups was not less than 80%. Viremia showed no significant differences among immunized groups and was significantly lower than that of the non-immunized challenge control group. These findings indicate that the immunization effect of the recombinant baculovirus-derived PCV2 subunit vaccine （OKM strain） is similar to that of commercial vaccines.

This study aimed to investigate the impact of the porcine circovirus type 2 （PCV2） ORF5 gene on the transcriptome of porcine kidney epithelial cells （PK-15）， elucidating its regulatory mechanisms at the RNA level. PK-15 cells were transfected with eukaryotic plasmid PCAGGS-HA-ORF5 to construct ORF5 overexpression model，Total RNA was extracted， and its integrity was verified. High-throughput sequencing was performed to identify differentially expressed mRNAs， circular RNAs （circRNAs）， and long non-coding RNAs （lncRNAs） between ORF5-overexpressing and control cells. The sequencing results were validated using quantitative real-time PCR （qRT-PCR）. Bioinformatics analyses， including Gene Ontology （GO） functional enrichment and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway analysis， were conducted， and an RNA regulatory network was constructed. Transcriptome sequencing revealed that ORF5 overexpression significantly altered the expression of 131 circRNAs （65 upregulated， 66 downregulated）， 155 lncRNAs （68 upregulated， 87 downregulated）， and 2 495 mRNAs （1 229 upregulated， 1 266 downregulated） in PK-15 cells. qRT-PCR validation of randomly selected differentially expressed RNAs confirmed the reliability of the sequencing data. GO analysis indicated that these RNAs were primarily associated with membrane-bound organelles and ribonucleoprotein complexes in cellular components， nucleic acid binding and transcription factor activity in molecular functions， and immune response and apoptotic regulation in biological processes. KEGG pathway analysis highlighted their involvement in TNF， NF-κB， and NOD-like receptor signaling pathways， as well as cell cycle regulation. Furthermore， an intricate interaction network was observed among differentially expressed circRNAs， lncRNAs， miRNAs， and mRNAs. These findings demonstrate that ORF5 modulates a complex RNA regulatory network in PK-15 cells， playing a crucial role in PCV2 infection. This study provides novel insights into the pathogenic mechanisms of PCV2.

This study aims to investigate the impact of the NS7 protein of the PDCoV JS2021-LX strain on PDCoV proliferation and pathogenicity. Using a bacterial artificial chromosome （BAC）-based reverse genetics system previously established in our laboratory， we generated an NS7-deficient recombinant virus （rPDCoV-ΔNS7） through CRISPR-Cas9-mediated gene editing and homologous recombination. Subsequently， the original NS7 gene fragment was replaced through homologous recombination to obtain a recombinant plasmid with NS7 gene deletion. After transfecting LLC-PK1 cells， the recombinant virus lacking the NS7 protein was rescued. The rescued rPDCoV-ΔNS7 was characterized both in vitro and in vivo using parental PDCoV and rescued wild-type virus （rPDCoV） as controls. 1） In vitro replication kinetics analysis revealed that rPDCoV-ΔNS7 exhibited significantly lower viral titers than PDCoV and rPDCoV at 12 and 24 hours post-infection （hpi） （P0.01）， but reached comparable titers at 36 hpi （P0.05）. 2） Plaque morphology analysis showed no significant differences among the three viruses. 3） In vivo， piglets infected with rPDCoV-ΔNS7 displayed similar clinical symptoms， pathological lesions， fecal virus shedding patterns， and tissue tropism compared to those infected with PDCoV or rPDCoV. 4） Quantitative analysis of viral loads in rectal swabs and various tissues confirmed no significant differences among the three groups. The results indicated that the NS7 gene deletion of the PDCoV JS2021-LX strain does not affect its replication and pathogenicity.

Porcine deltacoronavirus （PDCoV）， a newly identified enteric coronavirus that primarily targets suckling piglets， manifesting clinically as acute vomiting， profuse diarrhea， and severe dehydration. To investigate the impact of palmitoylation sites in the PDCoV spike protein on viral replication and immunogenicity. The palmitoylation sites of the S protein were predicted using CSS Palm 4.0 and verified with the palmitoylation inhibitor 2-bromopalmitate （2-BP）. Recombinant viruses with mutated palmitoylation sites were constructed using CRISPR/Cas9 technology on the rPDCoV-CHN-HN-2014 plasmid backbone. Virus growth kinetics were assessed through viral growth curves. Inactivated vaccines prepared from the viruses were used to immunize mice. Serum levels of specific antibodies were detected using virus neutralization tests and indirect ELISA. The results showed that the PDCoV S protein possessed eight palmitoylation sites. The recombinant viruses rPDCoV-S-C12S and rPDCoV-S-C1131S were successfully rescued. The growth kinetics analysis revealed that the infectivity titers of all viruses in host cells initially increased and then decreased. The titer of rPDCoV-S-C12S peaked at 24 hours post-infection （hpi）， while the titers of rPDCoV-CHN-HN-2014 and rPDCoV-S-C1131S peaked at 36 hpi. In virus neutralization tests， the neutralizing antibody titers induced by rPDCoV-CHN-HN-2014， rPDCoV-S-C12S， and rPDCoV-S-C1131S all reached 1：64. Indirect ELISA detected serum IgG antibody levels in mice， and no significant differences were observed among the different groups.​ This study provides a theoretical foundation and experimental data for further investigation into PDCoV pathogenesis and the development of gene-deleted attenuated vaccines.

To establish a rapid and efficient serological method for detecting bovine coronavirus infection （BCoV）， this study selected the conserved region of the BCoV S1 gene based on bioinformatics analysis. A truncated S1 protein was obtained through prokaryotic expression and used as a coating antigen to develop an indirect ELISA for BCoV antibody detection. The reaction conditions were optimized， and its specificity， sensitivity， repeatability， and concordance were evaluated. The established method was applied to conduct an epidemiological survey of 1 450 bovine serum samples from the Inner Mongolia Autonomous Region. The results showed that the 401-595 aa region of the BCoV S1 protein exhibited high conservation and strong immunogenicity， enabling efficient expression. The developed indirect ELISA demonstrated specific reactions with BCoV-positive sera but no cross-reactivity with sera from eight other common bovine pathogens， indicating excellent specificity. The method could detect positive sera diluted up to 1∶1 600， demonstrating high sensitivity. Both intra- and inter-assay coefficients of variation were below 10%， confirming good repeatability. The concordance rate with the neutralization test was 93.75%. The serological survey revealed an overall BCoV antibody positivity rate of 85.45% in Inner Mongolia. Regional distribution analysis showed the highest positivity rate in central regions （91.62%）. Among age groups， adult cattle had the highest positivity rate （89.36%）. Different farming models indicated higher positivity rates in large-scale farms compared to backyard farms. This study successfully established an indirect ELISA for detecting BCoV antibodies using the truncated S1 protein， which can be applied for BCoV antibody surveillance.

To understand the incidence of feline coronavirus （FCoV）， feline panleukopenia virus （FPV）， feline calicivirus （FCV）， and feline herpesvirus （FHV）， 256 suspected cases attending several animal hospitals across the country from January-December 2024 were investigated. The results showed that the positive rates of FCoV， FPV， FCV and FHV were 24.59%， 15.79%， 55.60% and 8.00%， respectively. Cats aged 0-6 months were the high-risk group for FCoV and FPV， while cats aged 7-12 months of age were more susceptible to FHV， and cats aged over one years were the main target of FCV. Regional distribution analysis shows that FCoV infection rates are distributed in a gradient pattern， with Guangdong at 30.00%， Beijing at 13.33%， Jiangsu at 10.00%， and Chongqing at 6.70%. FCV and FHV are highly concentrated in Beijing， with FCV positivity rates of 40.00% and FHV positivity rates of 75.00%. FPV showed a “double-core” distribution， with Beijing and Guangdong tied for the highest （22.20%）， followed by Chongqing and Shandong （11.10%）. Through the detection and analysis of four viruses， this study aims to provide a basis for the prevention and control of this disease.

This study aimed to isolate and characterize lytic Salmonella phages for prophylactic use in chicks.A Salmonella phage was isolated and purified from poultry intestines using the double-layer agar method. The phage was assessed through transmission electron microscopy （TEM）， host range determination， characterization of biological properties， and whole-genome sequencing analysis. Subsequently， a prophylactic experiment was conducted where 2-8-day-old chicks were orally administered the phage （1×10⁹ PFU·mL^-1）， followed by challenge with Salmonella Enteritidis （1×10⁸ CFU·mL^-1） at 9-11 days old. The efficacy of the phage in preventing Salmonella infection in chicks was evaluated by analyzing growth performance， immune organ indices， intestinal villus height-to-crypt depth ratio （VH/CD）， and intestinal inflammatory cytokine levels. vB_SEnS_SalVB showed a 20-min latent period， burst size of 196 PFU·cell^-1， and stability at pH 4-10 and 30-60 ℃. Genomic analysis revealed a 43 618 bp dsDNA genome （49.54% GC） lacking virulence or resistance genes. Prophylactic phage administration significantly （P0.05） mitigated Salmonella-induced growth impairment （P0.05）， enhanced VH/CD ratios （P0.05）， and down-regulated the mRNA expression levels of intestinal pro-inflammatory cytokines IL-6 and TNF-α （P0.05）. This study comprehensively and systematically analyzed the biological characteristics of the bacteriophage vB_SEnS_SalVB， such as key parameters like killing kinetics， host range， burst size， and environmental tolerance. This not only verified its potential as a single-strain therapeutic agent but also provided a crucial basis for future scientific compatibility with other bacteriophages.

Mycoplasma hyorhinis （Mhr） is a significant pathogen causing substantial economic losses to the pig industry. To investigate the role of lactate dehydrogenase （LDH） in the adhesion of Mhr to host cells， recombinant LDH （rLDH） was expressed， purified， and used to prepare hyperimmune rabbit serum. The surface expression of LDH on Mhr was confirmed by colony immunoblotting， flow cytometry， and Western blot. The cell-adhesion ability of rLDH was assessed using indirect immunofluorescence and microplate binding assays， and the inhibitory effect of anti-rLDH serum on Mhr binding to cell membrane proteins was evaluated. Finally， the binding capacity of rLDH to host plasminogen （Plg） and fibronectin （Fn） was examined. The results demonstrated that rLDH was successfully expressed and purified. Immunological assays confirmed that LDH is expressed on the Mhr surface. Functional assays revealed that rLDH adheres to PK-15 cells and binds to cell membrane proteins； notably， anti-rLDH antibodies significantly inhibited Mhr binding to cell membrane proteins. Far Western blot and microplate binding assays further demonstrated that rLDH binds to both Plg and Fn. These findings indicate that LDH is a surface-expressed protein of Mhr possessing multiple adhesive activities， playing a crucial role in colonization， invasion， and pathogenesis of Mhr.

This study aimed to investigate the inhibitory effect of pyrogallic acid on Actinobacillus pleuropneumoniae （APP） and its synergistic mechanism with antibiotics. The minimal inhibitory concentration （MIC） and minimal bactericidal concentration （MBC） of pyrogallic acid and commonly used antibiotics against APP were determined using the broth microdilution method. The fractional inhibitory concentration index （FICI） of drug combinations was calculated by the checkerboard dilution method. Time-kill curves were constructed using plate colony counting and spectrophotometry. Bacterial ultrastructural changes were observed by scanning electron microscopy （SEM） and transmission electron microscopy （TEM）. Biofilm inhibition was assessed by crystal violet staining. Infected cell protection rates were detected using AM/PI double staining. Potential action targets were analyzed by adding bacterial components. The expression of virulence and biofilm-related genes was analyzed by RT-qPCR. The results showed that APP was resistant to sulfisoxazole and clindamycin. The MIC and MBC of pyrogallic acid alone were both 256 μg·mL^-1， exhibiting dose-dependent antibacterial activity. Pyrogallic acid demonstrated synergistic effects in combination with sulfisoxazole， clindamycin， and doxycycline. Electron microscopy revealed damage to the bacterial cell wall/membrane. Treatment with 128 μg·mL^-1 pyrogallol reduced biofilm formation by 26.4% （P0.01） and alleviated infection-induced cell damage. The MIC of pyrogallic acid increased 4-fold after the addition of peptidoglycan or phosphatidylethanolamine， suggesting these as potential targets. The expression of virulence and biofilm-related genes was significantly downregulated （P0.05）. Pyrogallic acid exerts synergistic antibacterial effects by disrupting bacterial structure， inhibiting biofilm formation， and downregulating virulence gene expression， indicating its potential as an antibiotic synergist for drug-resistant APP infections.

This study aims to evaluate the immunoprotective efficacy of recombinant Eimeria media protein rEm-GAPDH and recombinant Eimeria intestinalis protein rEi-14-3-3 against Eimeria media infection in rabbits， providing a basis for developing recombinant subunit vaccines against E. media. Fifty-four 35-day-old coccidian-free young rabbits were randomly divided into six groups： blank control （PBS）， non-immunized challenge control （PBS）， Trx-His-S tag challenge control （100 μg Trx-His-S tag protein+1 mg Quil-A）， rEi-14-3-3 immunized group （100 μg rEi-14-3-3 protein+1 mg Quil-A）， rEm-GAPDH immunized group （100 μg rEm-GAPDH protein+1 mg Quil-A）， and rEm-GAPDH+rEi-14-3-3mixed immunized group （100 μg each protein+1 mg Quil-A）. All groups were immunized subcutaneously. A booster immunization was administered at the same dose 14 days after the primary immunization. Fourteen days post-booster， all groups except the blank control were orally challenged with 5×10⁴ sporulated E. media oocysts per rabbit. Clinical symptoms were monitored， and blood samples and body weights were collected periodically. Ten days post-infection， rabbits were euthanized for intestinal lesion scoring， oocyst output per gram of feces （OPG）， relative weight gain， specific IgG antibodies， and cytokine levels. Compared to challenge controls， all immunized groups exhibited milder clinical symptoms and intestinal lesions. The oocyst reduction rates for the rEi-14-3-3， rEm-GAPDH， and mixed immunized groups were 64.49%， 49.06% and 70.68%， respectively. Relative weight gains were significantly higher than in the non-immunized challenge group （P0.05）， with Anticoccidial Index （ACI） values of 163.47， 150.24 and 171.68， respectively. Immunized groups showed significantly elevated levels of specific IgG antibodies and cytokines （IFN-γ， IL-2 and IL-10） compared to controls （P0.05）. In conclusion， rEm-GAPDH and rEi-14-3-3 reduce weight loss and oocyst shedding， induce host cellular and humoral immune responses， and demonstrate immunoprotective potential as candidate antigens for recombinant subunit vaccines against E. media.

To clarify the species composition， dominant populations， and distribution patterns of Dipteran flies in cattle farms in north-central Zhejiang， an investigation was carried out from July to November 2024. Six farms were sampled： three large-scale cattle farms （500 head） and three small-scale cattle farms （200 head） in Hangzhou， Zhoushan， Ningbo， and Jinhua， across 4 regions. Flies were collected simultaneously with commercial yellow sticky fly papers （500 cm²） and fly traps spaced 20 meters apart for 72 hours. Specimens were identified morphologically and by mitochondrial COI sequencing followed by NCBI BLAST； a phylogenetic tree was constructed in MEGA 11 using Tabanidae as the out-group， and data were processed in Excel 2016 and GraphPad Prism 7.0.​ The results showed that： 1） A total of 1 969 flies were captured from the 6 cattle farms， belonging to 33 species， 16 genera， and 8 families. Musca was the dominant genus （65.52%）， followed by Stomoxys （13.86%）， Atherigona （7.72%）， Chrysomya （6.86%）， Hydrotaea （1.93%）， Lucilia （1.63%）， and Sarcophaga （1.37%）. Among these genera， Musca domestica， Stomoxys calcitrans （stable fly）， Atherigona orientalis， Chrysomya megacephala， Lucilia cuprina， Hydrotaea chalcogaster， and Sarcophaga （Boettcherisca） peregrina were the dominant species. Stomoxys calcitrans was a widespread species across the entire study area， being the absolute dominant species in the beef cattle farms in Jinhua （97.3%） and Zhoushan （68.6%）. Musca domestica was the dominant species of the Musca genus in 5 of the 6 cattle farms （except the farm in Chun'an， Hangzhou）， with the highest proportions in Qiantang， Hangzhou （91.7%） and Yuyao， Ningbo （80.2%）.​ Distinct region-specific distribution characteristics were observed： The cattle farm in Chun'an， Hangzhou （located in mountainous and hilly areas） had the richest diversity of Sarcophagidae species. Atherigona orientalis reached 19.7% in Yinzhou of Ningbo， acting as an indicator species. whereas Tephritidae were only detected in the Qiantang， Hangzhou farm （1.1%）.​ Molecular phylogenetic analysis revealed that， within the Muscidae family， all genera except Hydrotaea clustered into a single clade， while Hydrotaea showed a closer phylogenetic relationship with Tabanidae. The Lucilia and Hemipyrellia formed a subclade， and Chrysomya （Calliphoridae） and Sarcophaga （Sarcophagidae） formed another subclade； these two subclades further merged into a large clade. Individuals of the same species clustered stably into independent clades with high bootstrap values.​ Hydrotaea leucostoma， Sarcophaganathani（Boettcherisca chianshanensis）， and Helophilus hybridus are reported for the first time in Zhejiang Province. This study fills a knowledge gap in basic surveys of flies in north-central Zhejiang cattle farms and provides baseline data for targeted fly control and risk assessment of pathogen transmission in the region.​

As an interferon-stimulated gene， IRAV has significant antiviral activity. However， the lack of commercial-specific antibodies against porcine IRAV severely limits the research on the regulatory role of IRAV in porcine related diseases. To address this issue， in this study， recombinant IRAV protein was expressed through the Escherichia coli expression system and its monoclonal antibody was prepared. The porcine IRAV gene was amplified by PCR to construct the recombinant prokaryotic expression plasmid pET-28a（+）-IRAV. After inducing the expression of recombinant IRAV protein with IPTG and purifying it， BALB/c mice were immunized. The spleen cells of immunized mice were fused with SP2/0 myeloma cells to prepare monoclonal antibodies， and the antibody titer of ascites was determined by indirect ELISA. The specificity of the antibody was verified by IFA and Western blot. Antibody subtypes were identified using the monoclonal antibody subtype identification kit. The results indicated that the porcine recombinant IRAV protein was successfully expressed， and five specific monoclonal cell lines were obtained， named： 8H10D2H6， 5F2D12D6， 6A9C3B2， 6F9C4B7， 1C8A3G5. The ascites titers were all measured to be between 1∶500 000 and 1∶2 500 000. Five monoclonal antibodies were verified by IFA and Western blot， confirming that they could specifically bind to IRAV proteins expressed by prokaryotic and eukaryotic expression systems. After subtype identification， the monoclonal antibody strain 8H10D2H6 belongs to the IgG2a heavy chain category， while the monoclonal antibodies strains 5F2D12D6， 6A9C3B2， 6F9C4B7 and 1C8A3G5 belong to the IgG2b heavy chain category， and the light chains are all Kappa type. Using this monoclonal antibody as a tool， Western blot and IFA experiments preliminarily demonstrated that porcine IRAV can inhibit the proliferation of PRRSV. In summary， this study successfully prepared monoclonal antibodies against porcine IRAV， which can serve as an excellent immunological tool in the research of IRAV against PRRSV. This provides a scientific basis for subsequent studies on the regulatory role of IRAV proteins in important swine diseases.

This study aimed to investigate the effects and underlying mechanisms of naringin （Nar） on benzalkonium chloride （BAC）-induced injury in feline limbal stem cells （FLSCs）， and to provide experimental evidence supporting the potential use of Nar as an adjunctive ocular surface protectant. Both in vitro experiments using FLSCs and an in vivo feline corneal epithelial injury model were employed. By using CCK-8 method， DCFH-DA probe， malondialdehyde （MDA）， superoxide dismutase （SOD） and lactate dehydrogenase （LDH） assay kits， JC-1 staining， Hoechst 33342/PI double staining and transmission electron microscopy observation， real-time fluorescence quantitative PCR （qPCR） and protein immunoblotting （Western blot） method， as well as constructing a feline corneal epithelial mechanical injury model， the effects of BAC on FLSCs proliferation， injury， oxidative stress and apoptosis， and the intervention effect and mechanism of Nar were explored. BAC decreased FLSC viability in a concentration-dependent manner and significantly increased cellular ROS levels， MDA content， and LDH release， while significantly decreasing SOD activity （P0.05）. BAC also led to a significant reduction in mitochondrial membrane potential and increased cell death （P0.05）. Nar mitigated BAC-induced mitochondrial swelling， cristae loss， nuclear pyknosis， and apoptotic body formation. Nar significantly upregulated the mRNA levels of antioxidant stress-related genes Nrf2， Ho-1， and Nqo1 in BAC-treated cells （P0.05）， downregulated the expression of pro-apoptotic proteins Bax， Bad， Caspase-3， and AIF （P0.05）， and upregulated anti-apoptotic proteins Bcl-2 and Bcl-xL （P0.05）. In vivo， BAC significantly delayed corneal re-epithelialization in Felines （P0.05）， while intraocular pressure and tear film breakup time showed no significant differences among groups （P0.05）. Nar significantly accelerated epithelial repair in BAC-treated eyes. Naringin significantly alleviates BAC-induced oxidative stress and mitochondria-dependent apoptosis in FLSCs. Topical administration of Nar can effectively mitigate the delayed corneal epithelial regeneration caused by 0.01% BAC， without altering intraocular pressure or tear film stability. These findings suggest that Nar has potential as an adjunctive agent for protecting the ocular surface in veterinary ophthalmology.

Astragalus polysaccharides have been shown to significantly modulate the expression levels of interferons and interleukin， yet the underlying mechanisms regulating these immune response pathways remain unclear. Recent studies indicate that extracellular vesicles （EVs） regulate multiple immune response pathways through the bioactive substances they carry. This study aims to investigate whether fermented Astragalus participates in the mRNA transcription of interferons and interleukin-2 genes by modulating EVs and their bioactive substances in chicken peripheral blood， thereby providing a theoretical foundation for understanding the antiviral effects of fermented Astragalus and developing novel antiviral drugs. Through the administration of fermented Astragalus， changes in the quantity of EVs in chicken peripheral blood were observed； the effects of EVs on the mRNA transcription of IFN-α， IFN-β， and IL-2 in chicken macrophages were assessed； differential miRNAs in EVs were analyzed to identify key regulatory miRNAs； and the consistency between target genes regulated by key miRNAs and the mRNA expression profile in chicken peripheral blood was further validated. The results demonstrated that the quantity of EVs in chicken peripheral blood significantly altered after the administration of fermented Astragalus， and these EVs markedly promoted the transcription of IFN-α， IFN-β， and IL-2 in chicken macrophages. Differential miRNA analysis revealed that miRNA-1388a is a crucial miRNA facilitating the transcription of IFN-α， IFN-β， and IL-2； further studies indicated that the target genes regulated by miRNA-1338a-3p were highly consistent with the mRNA expression profile in chicken peripheral blood. This study is the first to confirm that fermented Astragalus modulates the transcription of interferons and interleukin-2 genes through the regulation of chicken EVs and their mediated miRNAs. This discovery provides new insights into the antiviral mechanisms of fermented Astragalus and establishes a theoretical basis for the development of novel antiviral drugs.

This study aimed to analyze the effects of Dihydromyricetin-Chitosan Nanoparticles （DHM/HACC-NPs） on milk metabolites in dairy cows using non-targeted metabolomics. Twenty-four periparturient dairy cows with similar initial conditions （milk yield， parity， and body weight） were selected and randomly divided into a control group （CON， n=12） and a treatment group （TRT， n=12）. During the prepartum period， cows were administered 5 mL of saline （CON group） or 5 mL of DHM/HACC-NPs （TRT group） via mammary duct injection for 7 consecutive days. Milk samples were collected before the injection period （CON1， TRT1） and on the seventh day of treatment （CON7， TRT7） for metabolomic analysis. The results showed significant differences in metabolites between the CON and TRT groups. Both PCA and PLS-DA analyses indicated a clear separation between the two groups with good intra-group repeatability. A total of 118 significantly different metabolites （SDMs） were identified between the CON7 and TRT7 groups. Among these， 80 metabolites， including 8，11-Eicosadiynoic acid， LysoPC［18：1（9Z）］， and LysoPC［18：2（9Z，12Z）］， were significantly upregulated. Conversely， 38 metabolites， such as 2-Dodecylbenzenesulfonic acid， Bis（2-ethylhexyl） phthalate， and Tryptophyl-arginine， were significantly downregulated. The key differential metabolites were identified as 8，11-Eicosadiynoic acid， LysoPC［18：1（9Z）］， 2-Dodecylbenzenesulfonic acid， and Bis （2-ethylhexyl） phthalate. KEGG pathway enrichment analysis revealed that the SDMs were significantly enriched in choline metabolism， glycerophospholipid metabolism， and oxidative phosphorylation pathways. In conclusion， DHM/HACC-NPs influence key differential metabolites and the metabolic state of the mammary gland in periparturient dairy cows. This is achieved by upregulating lysophosphatidylcholines （LysoPCs） and downregulating oxidative stress-related metabolites like 2-Dodecylbenzenesulfonic acid， while also regulating lipid metabolism and maintaining mammary energy metabolism balance. These findings provide a basis for the application of DHM/HACC-NPs in the mammary health management of periparturient dairy cows.

This study aimed to explore the effect of bta-miRNA-31 and its target gene RASA1 （Ras GTPase-activating protein 1） on collagen degradation in the maternal placental tissue of dairy cows， thereby clarifying the regulatory role and mechanism by which bta-miRNA-31 contributes to retained fetal membranes （RFM）. Dairy cows were classified as RFM or non-retained fetal membranes （NRFM） based on whether fetal membranes were expelled within 12 h postpartum （n=6 per group）. The expression of bta-miRNA-31 in the maternal placental tissues was quantified by stem-loop quantitative real-time PCR. of cows in the RFM and NRFM groups. The mRNA and protein levels of RASA1， MMP-2， MMP-9， COL-Ⅲ， and COL-Ⅳ were determined by qRT-PCR and Western blot. Meanwhile， bioinformatics analysis was used to predict the target genes of bta-miRNA-31， and a dual-luciferase assay was conducted to verify its targeting relationship with RASA1. After overexpressing and silencing bta-miRNA-31 respectively， stem-loop quantitative real-time PCR was used to detect the expression changes of bta-miRNA-31 in bovine endometrial epithelial cells； qRT-PCR and Western blot were used to detect the changes in the mRNA and protein levels of RASA1， MMP-2， MMP-9， COL-Ⅲ， and COL-Ⅳ in the cells. Immunofluorescence technology was used to detect the expression changes of COL-Ⅳ in cells of each group. In RFM cows， the mRNA expression of bta-miRNA-31 in maternal placental tissues was significantly increased （P0.01）， while the mRNA and protein levels of RASA1， MMP-2， and MMP-9 were markedly decreased （P0.01）， and the mRNA and protein lvels of COL-Ⅲ and COL-Ⅳ were highly increased （P0.01）. Bioinformatics analysis showed that bta-miRNA-31 had 39 potential target genes， and the dual-luciferase assay confirmed that RASA1 had a targeting relationship with bta-miRNA-31. In vitro， overexpression of bta-miRNA-31 significantly reduced RASA1 as well as MMP-2 and MMP-9 at both mRNA and protein levels （P0.01） while increasing COL-Ⅲ and COL-Ⅳ （P0.01）； the opposite changes were observed upon bta-miRNA-31 inhibition （P0.01）. Immunofluorescence results were concordant， showing markedly increased COL-Ⅳ in the bta-miRNA-31 mimic group and decreased COL-Ⅳ in the inhibitor group （both P0.01） relative to controls. Elevated bta-miRNA-31 contributes to RFM by targeting and downregulating RASA1， thereby suppressing MMP expression， reducing collagen degradation， and impairing extracellular matrix remodeling.

The study aimed to reduce the genotyping cost of sika deer and obtain high-density genotyping data， construct a genotype reference panel using sika deer resequencing data and verified the genotype imputation effect through software， with the aim of providing a cost-effective genotyping strategy for sika deer genomic group in the experiment. A total of 298 sika deer aged 24 months and above were selected as the reference population. Genotyping data were obtained using 5x resequencing technology to construct a genotype reference panel. At the same time， 45 sika deer were sequenced and typed using 40K liquid-phase chips， and genotype imputation verification was carried out with the help of Beagle software. Imputation concordance rate reached 92.05% at a DR² threshold of 0.95， indicating that high DR² values significantly enhanced imputation performance of sika deer； when the minor allele frequency （MAF） 0.1， the imputation consistency of the locus could reach 95.13%， while when MAF≥0.3， the imputation consistency of the locus dropped to 57.9%. This suggested that the genetic diversity of the reference panel needed to be improved； Furthermore， the difference in the imputation consistency among different populations was less than 2%， indicating that the genetic structure of the reference panel was relatively stable； however， the imputing effect of sex chromosomes was 1%-2% lower than that of autosomes， and this difference suggested that the imputation method for sex chromosomes needed to be optimized； ultimately， expanding the reference population size by 6 times only resulted in a 0.27% increase in the imputing consistency. This further highlighted that enhancing the priority of genetic diversity over sample size. The reference panel of the sika deer genotype constructed in this study can effectively improve the imputation effect of the genotype. Among them， high DR² values， rich genetic diversity， and stable genetic structure are the key influencing factors. These findings could provide important references for reducing the breeding cost of sika deer.

This study aimed to investigate the genetic diversity and key mutations of Waterfowl Parvovirus （WPV） and to establish a rapid nucleic acid detection method for the early diagnosis of clinical samples and virus control. A WPV strain was isolated via Muscovy duck embryos. Based on whole genome analysis， a SYBR Green I fluorescent quantitative PCR detection method was established. Genetic analysis indicated that the isolated strain was most closely related to the Anhui DY16 strain， and several key amino acid mutations were identified in its NS and VP proteins. The established fluorescent quantitative PCR method demonstrated high sensitivity and strong specificity. In summary， a WPV strain was successfully isolated in this study， and the established rapid nucleic acid detection method is suitable for early clinical diagnosis and large-scale screening.

This study aimed to investigate the effect of bovine adipose-derived mesenchymalstem cells （bAD-MSCs） on the pyroptosis model of bovine endometrial epithelial cells （BEECs） and its related mechanism. First， BEECs were pretreated with 1 µmol·L^-1 LPS for 12 h and then stimulated with 10 µmol·L^-1 Nigericin sodium salt （Nig） for 2 h to establish a BEECs pyroptosis model. MSCs and BEECs were co-cultured for 48 h， and the expression levels of pyroptosis-related factors NLRP3， Caspase-1 and GSDMD， as well as pro-inflammatory factors TNF-α， IL-6 and IL-1β were detected by qPCR to evaluate the effect of bAD-MSCs on Nig-induced BEECs pyroptosis. Subsequently， the expression of pyroptosis-related proteins NLRP3， Caspase-1， Cleaved Caspase-1， GSDMD and Cleaved GSDMD was analyzed by Western Blot. Finally， the secretion levels of pro-inflammatory factors TNF-α， IL-6 and IL-1β were further verified by ELISA kits to comprehensively confirm the regulatory effect of bAD-MSCs on the BEECs pyroptosis model. The qPCR results showed that the expression of pyroptosis-related factors NLRP3， Caspase-1 and GSDMD and pro-inflammatory factors TNF-α， IL-6 and IL-1β in the BEECs pyroptosis model co-cultured with MSCs was significantly inhibited. The Western blot results showed that the expression of pyroptosis-related proteins NLRP3， Caspase-1， Cleaved Caspase-1， GSDMD and Cleaved GSDMD in the BEECs co-cultured with bAD-MSCs was significantly lower than that in the pyroptosis model group （P0.05）. The ELISA results showed that the expression of pro-inflammatory factors TNF-α， IL-6 and IL-1β in the BEECs co-cultured with bAD-MSCs was significantly lower than that in the pyroptosis model group. MSCs can inhibit the expression of pro-inflammatory factors TNF-α， IL-6 and IL-1β and pyroptosis-related factors NLRP3， Caspase-1， Cleaved Caspase-1， GSDMD and Cleaved GSDMD in bovine endometrial epithelial cells， and inhibit the occurrence of bovine endometritis， providing important reference value for the prevention and treatment of bovine endometritis in clinical practice