【Objective】 The purpose of this study was to explore the key role of miRNA-mRNA interaction network in the regulation of estrus in gilts for explaining the genetic mechanism of miRNA-mRNA interaction in the estrus activity of gilts. 【Method】 The hypothalamus,pituitary and ovary of gilts in estrus and anestrus were selected as the study objects.The concentrations of follicle-stimulating hormone (FSH),progesterone (P₄),estradiol (E₂), and other reproductive hormones in serum were measured.After sRNA-Seq analysis,the differentially expressed miRNA were screened out by bioinformatics software,GO function and KEGG pathway enrichment analysis of differentially expressed miRNA target genes were performed using R language,and 4 differentially expressed miRNAs were verified by Real-time quantitative PCR. 【Result】 The concentration of reproductive hormone in serum was consistent with the physiological cycle of gilts.A total of 742 known miRNAs and 229 novel miRNAs in gilts were detected during estrus and anestrus,57 differentially expressed miRNAs (24 up-regulated and 33 down-regulated) were found in hypothalamus of gilts during estrus and anestrus,71 differentially expressed miRNAs (44 up-regulated and 27 down-regulated) in pituitary were found,and 140 differentially expressed miRNAs (63 up-regulated and 77 down-regulated) in ovary were found. KEGG pathway enrichment analysis showed that differentially expressed miRNA target genes in hypothalamus of gilts were mainly involved in proteoglycan in cancer,NOD-like receptor signaling pathway,Toll-like receptor signaling pathway,and cytokine receptor interaction.The differentially expressed miRNA target genes in pituitary were mainly involved in glycine, serine and threonine metabolism,GnRH secretion,cell adhesion molecules,metabolic signaling pathway.The differentially expressed miRNA target genes in ovary were mainly involved in chemokine signaling pathway,lysosome,ovarian sterogenesis, cholesterol metabolism,PPAR signaling pathway,and ECM-receptor interaction.In the target gene regulatory network related to reproduction,the key miRNA that might be mediated by hypothalamus-pituitary-ovary gonad axis to regulate the estrus activity of gilts were screened:miR-6240Z,ssc-miR-34a,ssc-miR-143-3P,ssc-miR-127,ssc-miR-21-5P,and ssc-miR-381-3p.Four differentially expressed miRNAs in ovary of gilts were verified by Real-time quantitative PCR,and the expression trends were consistent with the sequencing results. 【Conclusion】 In this study,miRNA expression profiles of hypothalamus-pituitary-ovary gonad axis of gilts in estrus and anestrus were successfully constructed,and the differentially expressed miRNA was verified.miRNAs that were significantly related to the estrus activity of gilts were screened,which provided theoretical basis for the analysis of the estrus mechanism of gilts.

【Objective】 gga-miR-1574-5p is a miRNA associated with the cellular nutrient deprivation response.This study was aimed to analyze the function of gga-miR-1574-5p and identify its targeting relationship with G3BP2 gene,so as to provide a scientific basis for further analysis of the biological function of gga-miR-1574-5p. 【Method】 The mature sequence of gga-miR-1574-5p was obtained by online tool miRBase.Target genes of gga-miR-1574-5p were predicted using TargetScan database for GO function and KEGG pathway enrichment analysis,and the binding sites of G3BP2 gene and gga-miR-1574-5p were predicted.Dual luciferase reporter gene assay was used to verify the targeting relationship between gga-miR-1574-5p and G3BP2.Real-time quantitative PCR was used to detect the effect of gga-miR-1574-5p overexpression on G3BP2 gene expression.The effect of gga-miR-1574-5p on the expression of G3BP2 protein in chicken macrophages was detected by Western blotting. 【Result】 GO function enrichment results showed that the potential target genes of gga-miR-1574-5p were mainly enriched in cellular components such as nucleus and plasma membrane,biological processes such as regulation of transcription by RNA polymerase Ⅱ and positive regulation of transcription by RNA polymerase Ⅱ,and molecular functions such as protein binding and ATP binding.KEGG pathway enrichment results showed that the target genes were mainly concentrated in oocyte meiosis,TGF-beta signaling pathway,mitophagy-animal pathway,etc.A binding site between gga-miR-1574-5p seed region and the 3'-untranslated region (3'-UTR) of G3BP2 gene in Gallus gallus was detected in TargetScan database.Compared with G3BP2-3'-UTR wild-type plasmid and NC-mimics co-transfection group,the dual luciferase activity of G3BP2-3'-UTR wild-type plasmid and gga-miR-1574-5p mimics co-transfection group was extremely significantly decreased (P＜0.01).Real-time quantitative PCR results showed that compared with NC-mimics group,transfected with 20 nmol/L gga-miR-1574-5p mimics had no significant effect on the expression of G3BP2 gene (P＞0.05),but the expression of G3BP2 gene in 30 and 50 nmol/L gga-miR-1574-5p mimics groups were significantly or extremely significantly decreased (P＜0.05 or P＜0.01).Western blotting results showed that the expression of G3BP2 protein in 20,30 and 50 nmol/L gga-miR-1574-5p mimics groups were extremely significantly lower than that in NC-mimics group (P＜0.01). 【Conclusion】 gga-miR-1574-5p had a targeting relationship with G3BP2 gene,and gga-miR-1574-5p could inhibit the expression of G3BP2 gene and protein in chicken macrophages by specifically binding to G3BP2-3'-UTR.

【Objective】 The aim of this study was to explore the promoter sequence characteristics and protein expression of LIN28A gene in Duolang sheep,so as to lay a foundation for further study of its transcriptional regulation mechanism at puberty stage. 【Method】 Primers were designed for PCR amplification,cloning sequencing,and sequence alignment based on the LIN28A gene sequence of sheep in NCBI (accession No.:NC_056055.1),and the promoter sequence of LIN28A gene in Duolang sheep was obtained,which was analyzed by online analysis software.The hypothalamus,pituitary,and ovarian tissues of Duolang sheep during juvenile,prepuberty,puberty,and post-puberty periods were collected,and the expression of LIN28A protein in hypothalamic-pituitary-ovarian (HPO) axis was detected by Western blotting. 【Result】 The length of promoter sequence of LIN28A gene in Duolang sheep was 2 089 bp,and the GC content was 56.28%.The promoter sequence of LIN28A gene covered a total of 65 transcription factor binding sites,such as MEIS1,STAT3,and COMP1,3 TATA-boxes,1 CpG island,and 9 potential transcription start sites.LIN28A protein was expressed in all three tissues of Duolang sheep,with the highest expression in ovarian,followed by hypothalamus,and the lowest expression in pituitary.The expression of LIN28A protein in hypothalamus of Duolang sheep in prepuberty was significantly higher than that in juvenile and puberty (P＜0.05).The expression of LIN28A protein in pituitary of Duolang sheep in juvenile and prepuberty was significantly higher than that in puberty and post-puberty (P＜0.05).The expression of LIN28A protein in ovarian of Duolang sheep showed a stable trend before and after of puberty (P＞0.05).During the same period,there was no significant difference in the expression of LIN28A protein in hypothalamus,pituitary and ovarian of Duolang sheep (P＞0.05). 【Conclusion】 The promoter sequence of LIN28A gene in Duolang sheep had multiple transcription factor binding sites associated with puberty,providing experimental materials for transcription factor validation and promoter activity detection in the future.The expression of LIN28A protein in HPO axis of Duolang sheep was significantly different,providing a scientific basis for further study of the function and regulatory mechanism of LIN28A gene.

【Objective】 This study aimed to investigate the functional role of the duck Rho GDP dissociation inhibitor β (ARHGDIB) gene and further elucidate the molecular mechanisms underlying subcutaneous fat deposition in ducks. 【Method】 The ARHGDIB gene was cloned from the liver tissue cDNA of 28-day-old Wuqin-10 meat ducks,and was verified to be correct by agarose gel electrophoresis and sequencing.The obtained sequences were compared and bioinformatics analyzed using online software.The relative expression of ARHGDIB gene in thymus,lung,spleen,heart,duodenum,liver,kidney,abdominal fat,pectoralis muscles,leg muscle,and subcutaneous fat of ducks was detected by Real-time quantitative PCR. 【Result】 The full length of ARHGDIB gene in ducks was 1 409 bp,with a predicted coding region of 693 bp spanning 7 exons,encoding 230 amino acids.Leucine was the most abundant amino acid (10% of the total composition).Nucleotide sequence similarity of the ARHGDIB gene in ducks with that of Anser cygnoides,Bos taurus,Canis lupus familiaris,Capra hircus,Danio rerio,Equus caballus,Felis catust,Gallus gallus,Homo sapiens,Macaca fascicularis,Mus musculus,Ovis aries,Pan troglodytes,Papio anubis,Sus scrofa,Columba livia were 82.56%,50.53%,58.75%,54.08%,24.94%,56.30%,35.31%,81.79%,54.10%,54.90%,49.44%,47.25%,56.22%,55.99%,55.17%,and 82.64%,respectively.The phylogenetic tree analysis showed that ducks were most closely related to Anser cygnoides,followed by Columba livia.The ARHGDIB protein in ducks was a stable hydrophilic protein lacking transmembrane domains.It contained 24 phosphorylation sites and 2 N-glycosylation sites.Subcellular localization prediction suggested that the protein was predominantly localized to the mitochondria.The secondary structure of the ARHGDIB protein in ducks comprises α-helices,extended chain,β-turns,and random coils,with respective proportions of 25.22%,23.91%,4.35% and 46.52%.The predicted tertiary structure aligned with the secondary structure.Real-time quantitative PCR results demonstrated that the ARHGDIB gene was ubiquitously expressed across all examined tissues of ducks,with the highest expression observed in the thymus,which was significantly higher than in other tissues (P＜0.05).Moderately high expression was detected in the spleen,heart,and lung,while subcutaneous fat showed the lowest expression,significantly lower than all other tissues (P＜0.05). 【Conclusion】 The coding region of ARHGDIB gene in ducks was 693 bp long and encodes 230 amino acids.ARHGDIB protein in ducks was hydrophilic and predominantly localized to mitochondria.The ARHGDIB gene was expressed in various tissues of ducks,with the highest levels in thymus and the lowest level in subcutaneous fat.These findings provided a theoretical foundation for further investigation of ARHGDIB gene function in ducks,and establish a basis for elucidating its regulatory role in lipid metabolism and supporting targeted duck breeding programs.

【Objective】 This study was aimed to investigate the effect of 1,8-cineole on antioxidant indexes and alterations in metabolic components of eggs,and antioxidant signaling pathways in laying hens,thereby offering a scientific foundation for the judicious incorporation of 1,8-cineole into the dietary of laying hens. 【Method】 320 Hy-Line Brown laying hens,all 46 weeks old with similar egg production and body weights,were randomly divided into 2 groups,with 10 replicates in each group and 16 chickens in each replicate.Laying hens in control group were fed with basic feed,while hens in experimental group were given the same diet with an addition of 40 mg/kg 1,8-cineole.After a 7-day pre-trial,a 56-day trial followed,during which daily measurements of production performance and egg quality were taken.Blood,liver tissues,and eggs were collected on the 7th,28th,and 56th days of the trial.Eggs were divided into two groups and stored at 4 ℃ for 30 and 40 days,respectively.Albumen,yolk,and serum were analyzed for total antioxidant capacity (T-AOC),total superoxide dismutase (T-SOD),glutathione (GSH) activity,and malondialdehyde (MDA) content.High-resolution non-targeted metabolomics analysis was employed to identify the types and relative contents of metabolic products present in egg yolks.Concurrently,Real-time quantitative PCR was utilized to assess the expression of nuclear factor erythroid 2-related factor 2 (Nrf2),NADPH quinone oxidoreductase 1 (NQO1),small Maf proteins (sMaf),and heme oxygenase-1 (HO-1) genes in liver of laying hens. 【Result】 Compared with control group,①During the experimental period of 29-56 days,the egg laying rate of hens in experimental group was significantly increased (P＜0.05),while the soft broken egg rate was significantly decreased (P＜0.05).There were no significant changes in average egg weight,average daily feed intake,and mortality rate (P＞0.05).Laying hens in experimental group exhibited a statistically significant increase in Haugh unit of eggs on the 28th and 56th days (P＜0.05).No significant differences were observed in yolk ratio,egg shape index,and eggshell thickness (P＞0.05).②On the 7th,28th,and 56th days,the activities of T-AOC and SOD in both albumen and yolk,as well as GSH activity in yolk,were significantly increased (P＜0.05),while MDA content in albumen was significantly decreased (P＜0.05).③The metabolite profiles in yolk showed significant alterations,with 365 up-regulated and 218 down-regulated metabolites under positive ion mode,and 105 up-regulated and 89 down-regulated metabolites under negative ion mode.Notably,such as choline and taurine expression were significantly increased (P＜0.05).④The T-AOC,and the activities of GSH and SOD in serum of laying eggs were significantly increased (P＜0.05).⑤The expression of Nrf2,ssMaf,and NQO1 genes in liver of laying hens were significantly increased (P＜0.05). 【Conclusion】 Under the conditions of this experiment,dietary supplementation with 1,8-cineole could enhance the antioxidant capacity of both eggs and laying hens,potentially contributing to an extended shelf life of eggs.

【Objective】 The aim of this study was to analyze the molecular mechanisms related to the effects of perinatal nutritional supplementation and early weaning on yak’s maternal nutrient metabolism and reproduction through transcriptomics. 【Method】 18 healthy late-pregnant yaks with (233.9±18.3) kg weights,2-4 litters,and similar due dates were selected and randomly divided into three groups (n=6)：graze feeding group (GF),nutritional supplementation group (SF),and nutritional supplementation and early weaning group (SW).The yaks in GF,SF,and SW groups were naturally grazed from 30 days before delivery to 90 days after delivery,and after daily grazing,the yaks in SF and SW groups were supplemented with fully mixed feed (TMR).The calves in GF and SF groups were breastfed and naturally weaned after delivery,while calves in SW group were weaned early at 60 days postpartum.Whole blood samples were collected from the yaks in GF,SF,and SW groups at 15 days before(-15 d),30 and 90 days after parturition(30 and 60 d) for transcriptomic analysis. 【Result】 ① After data filtering，the whole blood cDNA library sequencing of yak showed,the average value of each group in Q20 ＞ 95%;the average value of each group in Q30 ＞ 90%,indicating high quality of transcriptome information.The reference gene comparison statistics showed that the average mapped ratio of each group was greater than 70% standard,indicating good comparison results.② Transcriptome sequencing revealed that a total of 884,443 and 354 significantly differentially expressed genes (DEGs) were identified in SF and GF groups at -15,30 and 90 d,respectively.Additionally,502 and 160 significantly DEGs were identified in SW group compared with the GF and SF groups at 90 d,respectively.③The GO analysis results showed that the three groups were enriched to 1 464-4 638,308-891 and 253-586 significantly different terms (P＜0.05) for biological process (BP),molecular function (MF),and cellular component (CC) in the two-by-two comparisons of -15,30 and 90 d,respectively.DEGs were mainly enriched in terms related to nutrient metabolism functions such as positive regulation of response to nutrient levels,sterol-transporting ATPase activity,chylomicron,and lipoprotein particle receptor activity.④ The KEGG enrichment analyses results showed that DEGs were significantly enriched in 16 pathways in SF group compared with GF group at -15 d.Pathways related to nutrient metabolism and reproduction included protein digestion and absorption,gastric acid secretion,salivary secretion,and regulation of lipolysis in adipocytes.At 30 d,23 pathways were significantly enriched in SF vs GF group.Pathways related to nutrient metabolism and reproduction included relaxin signaling pathway,arachidonic acid metabolism,ovarian steroidogenesis,protein digestion and absorption,regulation of lipolysis in adipocytes,and prolactin signaling pathway.At 90 d,17 significantly enriched pathways were found in SF vs GF group,included insulin secretion,arginine biosynthesis,C5-branched dibasic acid metabolism,and gastric acid secretion related to nutritional metabolism and reproduction.Additionally,10 significantly enriched pathways were found in SW vs GF group.Pathways related to nutritional metabolism and reproduction included arginine biosynthesis,nitrogen metabolism,and C5-branched dibasic acid metabolism.There was 4 significantly enriched pathways in SW vs SF group. 【Conclusion】 Nutritional supplementation treatment during the perinatal period promoted nutrient digestion and absorption,lipid metabolism,and attenuated the catabolism of protein-amino acids in body stores through the secretion of salivary and gastric acid in yak dams.It also enhanced the maintenance of pregnancy,lactation activity,and postpartum recovery through the synthesis,secretion,and action pathways of reproduction-related hormones,such as epinephrine,norepinephrine,prostaglandin E2,relaxin and prolactin.

【Objective】 The aim of this study was to investigate the effects of different fermentation bedding materials on the growth performance,digestive enzyme activities,serum biochemical indexes and immune indexes of Patridge Shank chickens. 【Method】 Four hundred healthy 1-day-old Patridge Shank chickens (equal sex ratio) with similar body weights were randomly allocated to four groups:Control group (concrete floor feeding with pure chicken manure generated during the feeding process as bedding materials,without additional bedding litter),group A (wheat straw bedding litter),group B (sand bedding litter),and group C (wheat straw and sand with a volume ratio of 1∶1). No feces was removed in any group,each group was sprayed with activated and diluted bacterial solution (bacterial solution∶water=1∶20) and turned over every 7 days.The pre-feeding period was 7 days,the experimental period was 21 days.The temperature and pH of bedding were observed regularly during the experiment period and at the end,organic matter,total nitrogen content and carbon-nitrogen (C/N) ratio of the fermentation bedding materials were determined by four division method for each group,and two chickens were randomly selected from each replicate for blood sampling,slaughtering,and collecting pancreas and duodenum to determine serum biochemical indices and intestinal digestive enzyme activities. 【Result】 The pH of the bedding in each group showed an overall increasing trend with the prolongation of the rearing time of broilers, and eventually stabilized between 7.88 and 8.03, and the pH of bedding litter in experimental groups were relatively higher than that of control group throughout the whole period.The litter temperature of all experimental groups was higher than that of control group,with group A having the highest temperature and fastest warming up.Compared with control group,the moisture content of bedding litter was significantly higher in group A (P＜0.05),while the moisture and organic matter contents were significantly lower in groups B and C (P＜0.05),and the total nitrogen content of each experimental group was lower (P＜0.05),and C/N of each experimental group was higher (P＜0.05).Compared with control group,the final weight and average daily weight gain of chickens in groups A,B and C were significantly higher (P＜0.05),and there were no significant differences in average daily feed intake and feed to gain ratio (P＞0.05).Compared with control group,the activity of pancreatic trypin and lipase in pancreas of chickens were significantly increased in experimental groups (P＜0.05),the activity of α-amylase in pancreas and the activity of lipase in duodenum were enhanced in group B (P＜0.05),and the activities of lipase and α-amylase in duodenum were significantly enhanced in group C (P＜0.05).Compared with control group,the serum total protein content (TP) of chickens in group B was significantly higher (P＜0.05),the serum TP， albumin (ALB) and glucose (GLU) contents of chickens in group C were significantly higher (P＜0.05),while blood urea nitrogen (BUN) content of chickens in experimental groups were significantly lower (P＜0.05).The TP content in the serum of chickens in group C was significantly higher than that of group B (P＜0.05),and the ALB content was significantly higher than that of groups A and B (P＜0.05).Compared with control group,IgA,IgM and IgG concentration in the serum of chickens were significantly higher in group C (P＜0.05). 【Conclusion】 The bio-bed system with sand and straw litters could improve the growth performance of Patridge Shank chickens to a certain extent,enhance the activity of intestinal digestive enzymes,promote body metabolism，and improve its immune function,especially the bio-bed of mixture litters with 50% sand and 50% wheat straw was more effective.

China has abundant crop straw resources,and if it can be converted into ruminant feed to make full use of it,not only reduce the problems of resource wastage and environmental pollution,but also effectively alleviate the predicament caused by the shortage of roughage resources in China,and decrease the cost of breeding.However,straw’s high cellulose content and complex structural characteristics have become barriers to its utilization in animal production.Currently,there are various treatment methods (including physical,chemical and biological) that can improve the feeding value of crop straw,but the actual feed utilization is still restricted by many factors.This review focuses on the national major agricultural leading technology—Feed utilization technology of straw with "wall breaking and bacteria-enzyme" synergism treatment.This technology combines wall-breaking technology with bacteria and enzyme co-fermentation technology,which opens up a new way for the efficient feed utilization of crop straw.This review describes the principle,process and characteristics of feed utilization technology of straw with "wall breaking and bacteria-enzyme" synergism treatment,and its application in ruminant production,and clarifies the challenges and future development direction of the efficient use of straw resources,aiming to provide technical guidance for the feed utilization of crop straw resources and promote the sustainable development of the animal husbandry industry.

【Objective】 This study aimed to isolate and screen a duck-origin probiotic strain,investigate its biological characteristics,and select candidate strains for meat duck production,providing reference for developing novel probiotic preparations. 【Method】 Bacterial strain was isolated and purified from duck cecal contents.Identification was performed through calcium dissolution zone assay,Gram staining,biochemical tests,and 16S rDNA sequencing.Growth characteristics,acid tolerance,bile salt tolerance,free amino acid production,and organic acid content were determined.Antimicrobial efficacy against Escherichia coli,Riemerella anatipestifer,Salmonella spp.,Staphylococcus aureus,Streptococcus suis,and Avibacterium paragallinarum were evaluated through inhibition rate and apoptosis rate assays.Scanning electron microscopy characterized antibacterial morphology.Duck-derived Lactiplantibacillus plantarum extracellular proteins were isolated by ultrafiltration,with antibacterial components analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS).Safety assessment was conducted in meat ducks. 【Result】 A Gram-positive,non-spore-forming bacillus was isolated.and it was determined to be Lactobacillus genus through biochemical identification tests.Then,it was confirmed to be Lactiplantibacillus plantarum through 16S rDNA detection.Compared with Lactiplantibacillus plantarum from pickle (pickle-LP),this Lactiplantibacillus plantarum from duck (duck-LP) could quickly enter the logarithmic phase and plateau of growth,and had a better acid and bile salt resistance.It could produce more free amino acids,lactic acid,and acetic acid (P＜0.05).It had a significant inhibitory effect on Escherichia coli and Avibacterium paragallinarum at a 1∶1 addition ratio (P＜0.05).At a 1∶4 addition ratio,there was a significant inhibitory effect on the growth of Escherichia coli,Riemerella anatipestifer,and Salmonella spp.(P＜0.05).At a 1∶9 addition ratio,there was a significant inhibitory effect on Escherichia coli and Riemerella anatipestifer (P＜0.05).At a 1∶4 addition ratio,the apoptosis rates of Escherichia coli,Riemerella anatipestifer, Streptococcus suis,and Avibacterium paragallinarum were significantly increased (P＜0.05). Through LC-MS/MS,a total of 10 proteins were detected,including two antimicrobial components:Peptidoglycan-binding protein and acetate kinase.The safety test on meat ducks showed that the bacterium was safe and had no toxic side effects on meat ducks. 【Conclusion】 This experiment isolated and screened a strain of duck-LP,which had excellent growth characteristics,acid/bile tolerance,amino acid/organic acid production,and broad-spectrum antimicrobial activity.Its antibacterial mechanism might be through synergistic effects of antimicrobial proteins and organic acids,disrupting cell wall/membrane integrity.The strain showed potential as a safe probiotic feed additive for meat ducks.

【Objective】 This study aimed to investigate the effects of proline supplementation on growth performance,serum antioxidant and immune indices,and cytokine levels in weaned foals,so as to provide a reference for the healthy growth and development in foals. 【Method】 A total of 28 healthy and 5 months of age weaned foals with similarly weight were randomly assigned to four groups.Foals in control group were fed standard diet,and foals in trial groups Ⅰ,Ⅱ,and Ⅲ were supplemented with 20,40,and 60 mg/kg BW proline,respectively.The pre-feeding period was 7 d,and the experimental period was 60 d.The growth performance,antioxidant index and immune index,and cytokine contents in serum of foals were determined every 30 days. 【Result】 Compared with control group,①With the increase of proline supplementation,there was a significant growth trend of body weight in foals,the total weight gain and average daily gain of foals in group Ⅲ were significantly increased (P＜0.05).On days 0 and 60,proline supplementation exhibited an upward trend in body height,body length,chest girth,and tube girth of foals (P＞0.05).From days 31 to 60,there was a significant increase in body height growth of foals in group Ⅲ (P＜0.05).②On days 30 and 60,the activities of superoxide dismutase (SOD),glutathione peroxidase (GSH-Px),and catalase (CAT),and total antioxidant capacity (T-AOC) in serum of foals in each experimental group were extremely significantly increased (P＜0.01),while the levels of diamine oxidase (DAO) and malondialdehyde (MDA) in serum were extremely significantly decreased (P＜0.01).③On days 30,the concentrations of immunoglobulin A (IgA) and IgG in serum of foals in groups Ⅱ and Ⅲ,and IgM concentration in group Ⅲ,were extremely significantly increased (P＜0.01).On days 60,the IgM concentration of foal serum in group Ⅱ was extremely significantly increased (P＜0.01),and the concentrations of IgA and IgG in group Ⅰ were significantly increased (P＜0.05).④On days 30 and 60，the contents of tumor necrosis factor-α (TNF-α),interferon-γ (IFN-γ),interleukin-1β (IL-1β),IL-6,and IL-8 in serum of foals in each experimental group were extremely significantly decreased (P＜0.01). 【Conclusion】 Under the conditions of this experiment,proline supplementation showed a trend of improving the growth performance in weaned foals,enhancing the antioxidant capacity and immune indices,and reducing the contents of cytokines in serum.The best effect was observed at a dosage of 60 mg/kg BW.

【Objective】 This study aimed to investigate the effects of dietary net energy levels on colonic microbiota composition and short-chain fatty acid profiles in Tunchang pigs. 【Method】 Forty healthy Tunchang pigs,with an average body weight of 10.00 kg±0.86 kg and similar body condition,were randomly divided into four dietary treatment groups (JC1,JC2,JC3,and JC4),with 5 replicates per group and 2 pigs per replicate.The four treatment groups were provided experimental diets with net energy levels of 10.65,10.15,9.65,and 9.15 MJ/kg,respectively.The experiment was terminated when the pigs reached an average body weight of 25.00 kg,with a total experimental period of 42 days.At the end of the trial,one pig was selected from each replicate to collect cecal contents for gut microbiota analysis and short-chain fatty acid composition analysis. 【Result】 ①Compared with JC3 group,the final body weight and average daily gain were significantly lower (P＜0.05),the feed-to-gain ratio was significant higher (P＜0.05) of Tunchang pigs in JC1,JC2 and JC4 groups.②Analysis of the diversity of intestinal flora showed that there were significant differences in both alpha diversity and beta diversity among the groups treated with different net energy levels of rations (P＜0.05).At the phylum level of colonic microbiota composition,the relative abundance of Spirochaetes was significantly lower in JC1,JC3 and JC4 groups compared with JC2 group (P＜0.05),while the relative abundance of Actinobacteria was significantly lower in JC2 and JC3 groups compared with JC4 group (P＜0.05).At the genus level,the relative abundance of Roseburia and Coprococcus was significantly lower in JC1,JC2 and JC4 groups compared with JC3 group (P＜0.05),and the relative abundance of Ruminococcus was also significantly lower in JC1 and JC2 groups compared with JC3 group (P＜0.05).In addition,the relative abundance of Treponema in JC2 group was significantly higher than that in the other three groups (P＜0.05).③Metabolomic analysis of short-chain fatty acids in colon contents showed that the content of caproic acid was significantly higher in JC2 group than that in JC3 group (P＜0.05),and extremely significantly higher than JC4 group (P＜0.01).And butyric acid content had a tendency to be elevated in JC3 group compared with JC1,JC2 and JC4 groups (0.05＜P＜0.10). 【Conclusion】 Compared to the recommended net energy level of 10.15 MJ/kg (JC2), reducing the dietary net energy level to 9.65 MJ/kg could improve the composition of short-chain fatty acids by increasing the richness of the intestinal microbiota and the relative abundance of beneficial bacteria,thereby promoting the growth performance of Tunchang pigs.

【Objective】 The experiment aimed to evaluate the application effects of phased malic acid (MA) diet in broiler production. 【Method】 A total of 480 one-day-old Cobb male broilers were randomly divided into 4 treatments,with 8 replicates per treatment.The broilers in control group were fed a corn-soybean based diet throughout the entire period.The broilers in experimental groups were fed experimental diets supplemented with 0.4%,0.6%,and 0.8% MA from 1 to 14 days of age,and then were fed the basal diet from 15 to 42 days of age.This experiment lasted for 42 days.Growth performance was recorded during the rearing trial.After the trial,the broilers were slaughtered,and carcass traits were assessed.Additionally,samples of chicken breast,blood,and intestinal tissue were collected to evaluate meat quality,blood biochemistry,intestinal antioxidant capacity,and cytokine levels. 【Result】 Compared with the control group,① The addition of 0.4% MA in the diet during the early growth phase increased the average daily gain (ADG) of broilers from 1 to 14 days of age,reduced the feed conversion ratio,and increased the average body weight at 14 days of age (P＜0.05).And it decreased the drip loss and cooking loss of muscle,as well as the serum low-density lipoprotein cholesterol levels (P＜0.05).② The addition of 0.6% MA in the diet during the early growth phase increased ADG of broilers from 1 to 14 days of age (P＜0.05),reduced muscle shear force (P＜0.05),it also decreased serum alanine aminotransferase activity and total protein levels (P＜0.05),and reduced the content of interleukin-1β (IL-1β) in jejunal mucosa (P＜0.05).③ The addition of 0.8% MA in the diet during the early growth phase decreased the abdominal fat percentage of broilers at 42 days of age,significantly increased the activity of intestinal superoxide dismutase,and significantly decreased the activity of intestinal catalase (P＜0.05),and also reduced the content of IL-1β,IL-2,and tumor necrosis factor in the jejunal mucosa (P＜0.05). 【Conclusion】 The addition of 0.4% to 0.6% MA in the diet during the early growth phase could improve the growth performance of broilers from 1 to 14 days of age,enhance the meat quality of broilers at 42 days of age,and strengthen their intestinal antioxidant capacity and immune levels.However,the addition of 0.8% MA in the diet during the early growth phase had an adversely affect on the intestinal antioxidant capacity and immune performance of 42 days of age broilers.

【Objective】 The aim of this study was to investigate the effects of different dietary lipid levels on growth,body composition,liver biochemical parameters,digestive capacity,and related antioxidant indices in hybrid snakehead (Channa argus♂ × Channa maculate♀) larvae,and determine the optimal dietary lipid level for hybrid snakehead larvae. 【Method】 825 experimental fish with an initial body weight of (299.87±0.15) mg were randomly divided into 5 groups,each with 3 replicates and 55 fish per replicate.They were fed 5 different diets with lipid levels of 11.22%,13.22%,16.25%,19.28%,and 22.54%,respectively,and designated as L10,L13,L16,L19,and L22 groups.The fish were reared in net cages (0.50 m×0.30 m×0.50 m) for 28 days.At the end of the experiment,the weight and number of experimental fish in each cage were counted,and growth performance was calculated.Fifteen fish were randomly selected from each cage for body composition analysis.The entire intestine and liver were collected for the determination of biochemical parameters,digestive enzyme activity,and antioxidant enzyme activity. 【Result】 ①The fish in L19 group exhibited significantly higher final body weight,weight gain rate,and specific growth rate than L10 and L22 groups (P＜0.05),and their feed conversion ratio was significantly lower than L10,L13,and L22 groups (P＜0.05).The protein efficiency ratio of fish in L19 and L16 groups were significantly higher than that of L10 and L13 groups (P＜0.05).There were no significant differences in survival rate,condition factor,and viscerosomatic index among all groups(P＞0.05).②Experimental fish in L19 and L22 groups had significantly higher whole-body crude lipid content compared to L10,L13,and L16 groups (P＜0.05),and there were no significant differences in whole-body moisture,crude protein,and crude ash contents among all groups (P＞0.05).③In the livers of the experimental fish,the triglyceride content in L19 and L22 groups was significantly higher than L10,L13,and L16 groups (P＜0.05),the total cholesterol content in L19 group was significantly higher than L10 group (P＜0.05),and the activities of alanine transaminase and aspartate transaminase had no significant differences among all groups (P＞0.05).④The trypsin activity in the liver of fish in L10 group was significantly higher than that in L16,L19,and L22 groups (P＜0.05).In the intestines of fish,the trypsin activity in L19 and L22 groups was significantly lower than that in L10 group (P＜0.05),and the lipase activity in L22 group was significantly higher than L10 and L13 groups (P＜0.05).⑤In the livers of the experimental fish,the glutathione peroxidase activity in L19 group was significantly higher than L10 group (P＜0.05),and the catalase activity was significantly higher than L10 and L22 groups (P＜0.05).The malondialdehyde content in L22 group was significantly higher than that in all other groups (P＜0.05). 【Conclusion】 An appropriate dietary lipid level could enhance the growth performance,increase body lipid content,and improve the antioxidant capacity of hybrid snakehead larval.Based on the comprehensive evaluation of growth performance and antioxidant capacity,the recommended dietary lipid requirement was 17.71%-19.39%.

【Objective】 The research aimed to study the effects of dietary quercetin supplementation on egg quality and lipid metabolism of laying ducks at late phase of production. 【Method】 A total of 288 healthy Longyan laying ducks at 63 weeks of age with similar egg production rate were randomly divided into 4 groups,with 6 replicates of 12 ducks each.The ducks in control group were fed a corn-soybean meal basal diet,and the ducks in other treatments were supplemented with 100,200 and 400 mg/kg quercetin,respectively.The pre-trial period was 2 weeks,and the trial period was 9 weeks.During the pre-trial period,all ducks were fed the same basal diet.At the end of weeks 3,6,and 9,six duck eggs were selected from each replicate to determine egg quality.At the end of week 9,two ducks from each replicate were sampled via the wing vein to determine plasma lipid metabolism indexes.After slaughtered,the development of the liver and reproductive organs was assessed,and the liver was collected to analyze lipid metabolism indexes. 【Result】 ①Compared with the control group,dietary supplementation with 400 mg/kg quercetin significantly increased the egg-laying rate of ducks during the late laying period (weeks 4-6) (P＜0.05),exhibiting linear and quadratic responses with increasing dietary quercetin levels (P＜0.05).However,dietary quercetin supplementation had no significant effect on average daily feed intake,daily egg weight,feed to egg ratio,or egg breakage rate (P＞0.05).②Compared with the control group, at the end of the week 6, the addition of 100 and 400 mg/kg of quercetin significantly increased eggshell thickness in duck eggs (P＜0.05); the addition of 400 mg/kg quercetin group significantly improved the average eggshell weight, eggshell proportion and toughness at weeks 6 and 9, and throughout the entire late laying period (P＜0.05). At week 9, the dietary addition of 200 mg/kg of quercetin significantly improved eggshell strength, eggshell proportion and toughness (P＜0.05), and all these indicators showed an upward trend with increasing levels of quercetin added (P＜0.05).③With increasing dietary quercetin levels,ovarian stroma weight in laying ducks exhibited a quadratic response,initially decreasing and subsequently increasing (P＜0.05).Compared with the control group,dietary supplementation with 100 mg/kg quercetin significantly decreased ovarian stroma weight in ducks during the late laying period (P＜0.05).④Plasma LDL-C and hepatic TG contents decreased linearly and quadratically with increasing quercetin levels (P＜0.05),and plasma LDL-C content of ducks in the 200 and 400 mg/kg quercetin-added groups were decreased (P＜0.05),and hepatic TG contents of ducks in the 400 mg/kg quercetin-added group were decreased (P＜0.05) compared with the control group.⑤Based on eggshell toughness,eggshell ratio and breaking strength,the optimal supplemental levels of dietary quercetin were obtained by fitting the equation as 199.5,284.3,and 284.9 mg/kg,respectively. 【Conclusion】 Dietary quercetin addition could improve eggshell mechanical properties and lipid metabolism of laying ducks at late phase of production.Under the conditions of this experiment,the recommended quercetin supplementation level in the diet of late-phase laying ducks was 200-285 mg/kg.

【Objective】 This study aimed to investigate the effects of dietary fiber levels and digesta collection periods on ileal digesta digestive enzyme activities,nutrient flow,and short-chain fatty acid (SCFA) composition and content in growing pigs. 【Method】 A single-factor completely randomized design was employed,with 18 pigs fitted with T-cannulas at the terminal ileum randomly assigned to three treatments (6 pigs per treatment).The treatments included a control diet,a 10% alfalfa fiber diet,and a 20% alfalfa fiber diet.Ileal digesta samples were collected over two 10-day periods to determine activities of amylase,lipase,trypsin,and chymotrypsin,analyze nutrient flow,and measure SCFA content. 【Result】 ①The ileal digesta pH in growing pigs increased significantly with elevated dietary fiber levels,and the differences between groups were extremely significant (P＜0.01).The activities of amylase,trypsin,and chymotrypsin in the ileal digesta of growing pigs in high fiber group were significantly lower than those in other dietary groups (P＜0.05).Collection periods had no significant effects on ileal pH or the four digestive enzymes (P＞0.05).②Compared with the control group,the total carbohydrate and acid detergent fiber flow in the ileum of growing pigs in both experimental groups increased significantly (P＜0.01),while the crude fat flow decreased significantly (P＜0.01).Collection periods did not affect nutrient or energy flow in the ileum (P＞0.05).③The contents of acetic acid,valeric acid,and total SCFAs in the ileal digesta of growing pigs in the high fiber group were significantly higher than those in the other treatment groups (P＜0.05).As the collection period of the experiment extended,the concentrations of propionic acid,valeric acid,and total SCFAs in ileal digesta significantly increased (P＜0.01). 【Conclusion】 High-fiber diets reduced ileal digestive enzyme activities but increased ileal flows of carbohydrates,fiber,and total SCFAs in growing pigs.Collection periods did not influence digestive enzyme activities or nutrient flow,but extended fiber intake duration enhanced ileal fermentation capacity.

【Objective】 The aim of this experiment was to study the effects of different energy and protein levels of diets on the egg-laying performance,egg quality and hatching performance of Snowy Mountain chicken breeder hens,and to clarify the appropriate energy and protein levels of diets during the laying period. 【Method】 3 150 healthy Snowy Mountain chicken breeder hens with similar body weight at 22 weeks of age were selected and randomly divided into 9 groups according to a 3×3 two-factor experimental design with 3 energy levels (11.09,11.51,and 11.92 MJ/kg) and 3 protein levels (14.50%,15.50%,and 16.50%),with 7 replicates in each group and 50 hens per replicate.The trial period was 14 weeks.The number of broken eggs,unqualified eggs and qualified eggs were counted during the test period,and at the end of the test,the relevant indexes of egg quality were measured, and the eggs were hatched,and hatching performance were determeared. 【Result】 ①Dietary energy and protein levels had a significant effect on the egg breaking rate of breeding eggs,at 22 to 25 weeks of age,it was decreased with the increase of energy level,and showed a trend of increasing and then decreasing with the increase of protein level,and at 26 to 35 weeks of age,it was increased with the increase of energy level,and decreased with the increase of protein level.There was no significant interaction between dietary energy and protein levels on indexes of egg-laying performance of Snowy Mountain chicken breeder hens (P＞0.05).② Dietary energy level had no significant effect on the indicators related to the quality of breeding eggs (P＞0.05).Dietary protein level had a significant effect on the egg shape index and egg shell thickness of breeding eggs (P＜0.05),and the egg shape index of the low protein group was significantly higher than that of the medium protein group (P＜0.05),and the egg shell thickness showed an increasing trend with the increase of protein level.There was no significant interaction between dietary energy and protein levels on indices of Snowy Mountain chicken breeding egg quality (P＞0.05).③Dietary energy level had a significant effect on the hatchability of incubated eggs,fledgling rate and fertilized egg hatchability of breeder hens (P＜0.05),and the hatchability of incubated eggs,fledgling rate of the medium energy group was significantly higher than those of the low and high energy groups (P＜0.05).Dietary protein level had a significant effect on the embryo death rate and fertilized egg hatchability of breeder hens (P＜0.05),and the embryo death rate of the high protein group was significantly higher than those of the medium and low protein groups (P＜0.05),and the fertilized egg hatching rate in the medium protein group was significantly higher than that in the low protein group (P＜0.05).There was a significant interaction between dietary energy and protein levels on the hatchability of incubated eggs (P＜0.05). 【Conclusion】 Considering all the indexes of laying performance,egg quality and hatching performance of Snowy Mountain chicken breeder hens,it was recommended that the appropriate energy level of Snowy Mountain chicken breeder hens during laying period should be 11.51 MJ/kg,and the protein level should be 16.50%.

【Objective】 The purpose of this experiment was to investigate how long-term cold exposure affect ewes’ weight changes,serum biochemical indicators,antioxidant indicators,non-specific immune indicators,as well as fetal development,and oxidative damage to the liver of ewes and fetal sheep in late pregnancy.Meanwhile,the regulatory role of N-carbamylglutamate (NCG) was also investigated. 【Method】 24 healthy Mongolian sheep in late pregnancy,aged 3-4 years,with similar body weight (48.53 kg± 3.64 kg),were randomly divided into 3 groups,with 8 replicates in each group and 1 sheep in each replicate.The control group (group C) was raised in a warm house (average temperature 3.60 ℃),while the two experimental groups (groups L and LN) were out of the house (average temperature -12.20 ℃).In addition,each sheep in LN group was orally administered 10 mL of distilled water and 2.5 g of NCG suspension per day,while each sheep in groups C and L was orally administered 10 mL of distilled water per day.The pre-feeding period was 10 days,and the normal test period was 62 days.During the experiment,the weight of the ewes was recorded on the 1 st,30th,and 60th day.At the beginning and end of the trial period,the jugular vein blood of ewes was collected to measure serum biochemical indicators,antioxidant indicators,immune indicators,and hormone levels in the hypothalamic pituitary adrenal axis (HPA) and hypothalamic pituitary thyroid axis (HPT).At the end of the experiment,the experimental sheep were slaughtered and sampled to measure the fetal sheep’s body weight,organ index,as well as the oxidative damage indicators of 8-hydroxydeoxyguanosine (8-OHdG),4-hydroxynonaenoic acid (4-HNE),advanced oxidative protein products (AOPP),3-nitrotyrosine (3-NT) content,and superoxide dismutase (SOD) activity in the liver of ewe and fetal sheep. 【Result】 ①The average daily weight gain (ADG) of ewes in group LN was significantly higher than group L in the later and whole stages of the experiment (P＜0.05),while there was no significant difference between groups L and C in the early,late,and entire stages of the experiment (P＞0.05).Fetal sheep weight in group L was significantly lower than groups C and LN (P＜0.05),while there was no significant difference between groups LN and C (P＞0.05). Except for the lungs,the visceral organ weight of fetal sheep in group L was significantly lower than groups C and LN (P＜0.05).②At the end of the experiment,the serum NEFA content of ewes in group LN was significantly higher than groups C and L (P＜0.05),and the serum β-HB content of ewes in group L was significantly lower than groups C and LN.③The serum T₄ content of ewes in group L was significantly lower than group C (P＜0.05),while the group LN was between them,with no significant difference (P＞0.05).④For hepatic oxidative damage indicators,the content of 8-OHdG of ewes in group L was significantly higher than group C (P＜0.05),while group LN was higher than group C and lower than group L (P＞0.05).The SOD activity of ewes in group L was significantly lower than groups C and LN (P＜0.05),and group LN was lower than group C (P＞0.05).There was no significant difference in the oxidative damage indicators of fetal sheep liver among the three groups (P>0.05). 【Conclusion】 Cold exposure in late pregnancy of ewes reduced their ADG,abnormal serum metabolism,and oxidative damage to the liver.Supplementing NCG could improve the adverse effects of cold exposure on ewes and fetal sheep.

【Objective】 This study was aimed to analyze mRNA and long non-coding RNA (lncRNA) in cashmere goat skin induced by exogenous melatonin to participate in the regulation of alternative splicing (AS) of cashmere growth at post transcriptional level,so as to provide a reference for exploring the effect of AS regulation on melatonin promoting cashmere growth in goats. 【Method】 Six Hanshan White cashmere goats were selected and randomly assigned to control and implant groups with three replicates in each group.Take the natural year as the experiment period,skin samples were collected monthly for RNA-Seq analysis.UsedBowtie,TopHat,Cufflinks,Cuffmerge,Cuffdiff,Cuffcompare,CPAT and CPC software were used to analyze differentially expressed mRNA (DE-mRNA) and differentially expressed lncRNA (DE-lncRNA),with rMATS software to identify differential alternative splicing events (DASE) of mRNA and lncRNA in cashmere goat skin transcriptome. 【Result】 The number of DE-mRNA (FDR≤0.05) between the two groups from January to December was 23,93,43,652,1 178,194,353,357,435,36,55 and 52,respectively.The number of DE-lncRNA (FDR≤0.05) between the two groups from January to December was 15,20,10,40,191,50,63,33,79,7,9 and 5,respectively.The number of DASE between the two groups involving mRNA and lncRNA was 715 and 569,respectively.Skipping exon (SE) and mutually exclusive exons (MXE) were the main alternative splicing ways of mRNA.SE,retained intron (RI) and alternative 3'-splice site (A3SS) were the main AS ways of lncRNA.The top 3 chromosomes of DASE number in mRNA were Chr1,Chr5 and Chr10/Chr11 (juxtaposition),and the top 3 chromosomes of DASE in lncRNA were Chr1,Chr19 and Chr10,respectively.Two or more splicing isoforms of mRNA and lncRNA were found in cashmere goat skin.A total of 443 DAS-mRNA and 271 DAS-lncRNA were identified.The top 3 months of DAS-mRNA distribution were October,June and May,and the top 3 months of DAS-lncRNA distribution were May，April and August,respectively.A total of 67 DE-mRNA and 7 DE-lncRNA underwent DASE.The AS regulatory network induced by exogenous melatonin，which composed of 334 DE-mRNA and 8 DE-lncRNA associated with mRNA and lncRNA underwent AS of Pearson’s correlation coefficient (PCC) ranking in top 10 (PCC＞0.800). 【Conclusion】 There were abundant ASE in cashmere goat skin,except for SE,mRNA preferred the alternative splicing way of MXE,while lncRNA preferred RI.Compared with mRNA,the distribution of DASE in lncRNA on chromosomes was specific.When induced by exogenous melatonin,the cashmere fast-growing and the early growth period were main periods for mRNA,while the early growth and the growth period were the main periods for lncRNA occurrence of AS.The potential interaction network between DE-mRNA and DE-lncRNA involved in the transcriptome of cashmere goat skin undergoing AS was constructed,DE-mRNA and DE-lncRNA participated in regulating cashmere growth through AS at the post transcriptional level,providing a reference for further exploring the effect of AS regulation on melatonin promoting cashmere growth.

【Objective】 The aim of this study was to explore the effects of supplemental light on blood endocrine hormone concentration and circadian rhythm change in late-gestation Holstein breeding dairy cows,so as to provide a theoretical basis for large-scale and standardized light management of late-gestation Holstein breeding dairy cows under supplemental light. 【Method】 Twenty-four Holstein breeding dairy cows with similar body weights (600 kg±5 kg) in one months before parturition were selected and randomly divided into a natural light group and a supplemental light group using a randomized experimental design.Dairy cows in natural light group received natural light for 10-11 h.Based on natural light,Dairy cows in supplemental light group were supplemented with fluorescent light at 04:00-08:00 and 16:00-20:00 every day,so that the light received 16 h.During the experiment,cows had free access to food and water,and the experimental period was 63 d.Five cows in each group were selected for caudal root blood sampling at six time points (03:00,07:00,11:00,15:00,19:00,and 23:00) on the 63rd day of the experiment.Indicators such as melatonin (MT),growth hormone (GH),insulin-like growth factor-Ⅰ(IGF-Ⅰ),leptin (LEP),prolactin (PRL),and cortisol (COR) in serum were measured.At the same time,a cosine mathematical model was used to fit the changes in hormone concentrations during the day and night to observe the circadian rhythm changes of each hormone. 【Result】 ①At each sampling time point,compared with natural light group,there were no significant differences of the concentrations in various serum hormones of dairy cows in supplemental light group (P＞0.05),but the mean concentrations of MT,IGF-Ⅰ,PRL,and COR in serum of dairy cows in the supplemental light group were higher than that in natural light group,whereas the mean concentrations of GH and LEP were lower than that in natural light group.②The circadian changes of MT,GH,LEP,PRL,and COR concentration in serum of dairy cows in both groups did not show a cosine oscillation rhythm curve change (P＞0.05).IGF-Ⅰ concentration in serum of dairy cows in natural light group had circadian rhythmic expression (R²＞0.5 and P＜0.05).The concentrations of MT，GH,IGF-Ⅰ,LEP,PRL,and COR in serum of dairy cows in natural light group exhibited superior goodness-of-fit in circadian rhythm patterns compared with supplemental light group. 【Conclusion】 Supplemental light exerted effects on the rhythmicity of MT,PRL,and COR concentrations in serum of late-gestation Holstein breeding dairy cows,and altered the circadian rhythmic expression of IGF-Ⅰsecretion.Therefore,it was recommended to appropriately increase the light to improve the circadian rhythm of late-gestation Holstein breeding dairy cows,the physiological state and production performance of dairy cows could be improved by regulating the biological clock and hormone secretion.

【Objective】 The aim of this experiment was to obtain the complete mitochondrial genome of Rhabdias bufonis and analyze its characteristics. 【Method】 The mitochondrial genes of Rhabdias bufonis were amplified by PCR,and the whole mitochondrial genome of Rhabdias bufonis was obtained by sequencing,annotation,assembly and spliceover,and the sequence analysis was performed.Using Camallanus lacustris as an outgroup,based on the amino acid series of 12 protein-coding genes (PCGs) in mitochondria.The phylogenetic tree was constructed by Neighbor-Joining (NJ) and Bayesian inference (BI) method for evolutionary analysis. 【Result】 The mitochondrial genome of Rhabdias bufonis was a circular structure with a total length of 14 208 bp,consisting of 12 PCGs,22 tRNAs,2 rRNAs and 2 non-coding control regions.The base composition of the mitochondrial genome，AT content was 75%,GC content was 25%,AT-skew was 0.26,GC-skew was -0.36,showing obvious AT bias.PCGs used ATG and GTG as start codons and TGA,TAA and incomplete TA- as stop codon.Secondary structure analysis of tRNAs showed that the trnS2 and trnH genes lacked dihydroouracil (DHU) arm,and the rest were typical clover structures.Phylogenetic tree analysis showed that Tylenchina was divided into two large clades,Cephaloboidea and Tylenchoidea formed clade Ⅰ,Strongyloidoidea,Steinernematoidea,Aphelenchoidea and Panagrolaimoidea formed clade Ⅱ.In clade Ⅱ,Steinernematidae and Rhabdiasidae gathered together to form a branch and grouped to another sisters branch formed by Aphelenchoidiae and Panagrolaimidae.Howerver,Strongyloididae independently formed another large branch.In Rhabdiasidae,Rhabdias bufonis from forest frog in this study and the Rhabdias bufonis (OR725306) from toad gathered together,and it was more closely to two strains of Rhabdias kafunata.The amphibian-parasitic Rhabdias sp.and the insect-associated Steinernema sp.clustered together,which exhibited more closer to the plant-parasitic Aphelenchoididae than to the mammalian-parasitic Strongyloididae. 【Conclusion】 In this study,the complete mitochondrial genome sequence of Rhabdias bufonis from forest frog was obtained. The phylogenetic relationship between Steinernematidae and Rhabdiasidae was obviously close to that of Strongyloididae,supporting the validity of Steinernematoidea as an independent taxonomic order,and Steinernematidae should be included in Steinernematoidea.The results provided basic information for the taxonomy,population genetics and phylogeny of Rhabdiasidae.

Frozen sperm of animals has great advantages such as long-term storage,long-distance transport,and more safety and efficiency.So,it has been broadly applicated in the fields of animal genetic resources preservation,endangered species protection,and cross-regional breeding of livestock and poultry.Sperm,as a highly differentiated cell,undergoes varying degrees of structural damage during the freezing process.These damages affect the integrity,fluidity,and selective permeability of sperm membrane,as well as alter the composition and function of membrane lipids,proteins,RNA,and other molecules.Consequently,these changes directly impact the post-thaw viability of sperm.Additionally,the extent of sperm cryotolerance varies greatly due to factors such as species,variety,season,and nutrition.However,due to the lack of clear understanding of basic problems such as sperm cryodamage,cryotolerance,and its influencing factors,the present freezing technology cannot produce frozen sperm that has high enough quality and can be applied in farm except bull.This has posed significant challenges to the preservation of these animal genetic resources via frozen semen,as well as to genetic improvement and the development of new breeds through artificial insemination using frozen semen.Therefore,enhancing the further basic research on the pertinent issues and developing an efficient freezing system to minimize freezing-induced damage and improve post-thaw sperm viability would have a positive impact on the promotion and application of the technology.The authors summarize the recent research progress on the mechanisms of sperm cryodamage and differential cryotolerance in animals,so as to provide reference for the in-depth development of related research.

【Objective】 The heat shock protein family A member 5 (HSPA5) of Wuyi Black pig was analyzed through bioinformatics predictions to provide insights on its biological functions. 【Method】 The coding sequence of HSPA5 gene was amplified and sequenced,similarity analysis was performed afterward,then a phylogenetic tree was constructed.The physicochemical properties,structural features,and functional roles of the HSPA5 protein were predicted using bioinformatics software,and its expression in different tissues of Wuyi Black pig were finally measured. 【Result】 The coding sequence of HSPA5 gene in Wuyi Black pig was 1 965 bp,encoding 654 amino acids.The results of similarity and phylogenetic tree analyses showed that the amino acid sequence of HSPA5 in Wuyi Black pig had the highest similarity (100%) with the transcript variant 2 of HSPA5 in Duroc pig and that in mice.It had the closest genetic relationship with Mus musculus and the most distant genetic relationship with Larimichthys crocea.The HSPA5 protein was predicted as an acidic hydrophilic protein containing a signal peptide sequence,with the highest concentration observed in the cytoplasm.It was mainly composed of alpha helix conformation with conserved motifs within two overlapping functional domains.It contained 56 phosphorylation sites,76 O-glycosylation sites and 24 chemical interaction sites,as well as 26 antigenic determinants,11 T-cell epitopes,and 13 B-cell epitopes.It was closely related to interactions with heat shock protein member 90 (HSP90) and activating transcription factor 6 (ATF6).The Real-time quantitative PCR results showed that the expression of HSPA5 gene in liver and uterus of Wuyi Black pig was significantly higher than that in other tissues (P＜0.05). 【Conclusion】 The HSPA5 gene of Wuyi Black pig had been successfully cloned,and had the closest genetic relationship with that of Mus musculus.It was predominantly localized in the cytoplasm,featuring phosphorylation and O-glycosylation modification sites,it also interacted with proteins related to the unfolded protein response pathway.It had the highest expression in liver of Wuyi Black pig.These characteristics could provide valuable references for further studies on the biological functions of HSPA5 protein.

【Objective】 This study was aimed to systematically compare the lactation performance differences between Fleckvieh F1 and Montbéliarde F1 cows under the feeding conditions in Northern China. 【Method】 Dairy herd improvement (DHI) records from 2021 to 2024 were collected and obtained from a large-scale dairy farm in Yantai,Shandong province.The analysis focused on F1 cows from Chilean Simmental dams crossed with Fleckvieh and Montbéliarde sires.A mixed linear model was constructed using the SAS 9.4 software (Mixed procedure) to analyze the effects of breed,parity,measurement season,lactation stage and their interactions on lactation traits in Fleckvieh F1 and Montbéliarde F1 cows. 【Result】 Parity,season and lactation stage were found to significantly affect milk yield,milk fat percentage,milk protein percentage,and somatic cell score (SCS) in Fleckvieh F1 and Montbéliarde F1 cows (P＜0.05).With the increase of parity,the milk yield,milk fat percentage and milk protein percentage were significantly increased across the first three parities in Fleckvieh F1 and Montbéliarde F1 cows (P＜0.05).The milk yield and SCS were the highest in summer,while milk fat percentage and milk protein percentage were the highest in winter.With the increase of lactation days,the milk fat percentage,milk protein percentage and SCS initially declined and then increased in Fleckvieh F1 and Montbéliarde F1 cows.In the first lactation,Montbéliarde F1 cows exhibited significantly higher milk protein percentage than Fleckvieh F1 cows (P<0.05),with the difference becoming even more pronounced in the second lactation (P<0.01).Seasonal comparisons revealed that Montbéliarde F1 cows had significantly higher milk protein percentages only during winter (P<0.01).Regarding lactation stages,although Montbéliarde F1 cows demonstrated markedly higher milk protein percentage in early lactation (P<0.01),their values were significantly lower than those of Fleckvieh F1 cows in late lactation (P<0.05).Notably,the Montbéliarde F1 cows exhibited consistently lower SCS than the Fleckvieh F1 cows across all parities,seasons,and lactation stages.The results suggested that Montbéliarde F1 cows exhibited superior lactation performance and enhanced resistance to mastitis compared with Fleckvieh F1 cows. 【Conclusion】 Under the feeding conditions in Northern China,parity,season and lactation stage had varying effects on the lactation performance in Fleckvieh F1 and Montbéliarde F1 cows.The results provided valuable insights for breed improvement and production management decisions on dairy farms.

【Objective】 Fatty acid binding protein (FABP) is an important functional protein that plays a key role in regulating lipid metabolism.Especially,FABP4 is considered the main carrier protein responsible for fatty acid transport in mammalian cells,playing a crucial regulatory role in the synthesis and decomposition of lipids in the body.Therefore,exploring the expression patterns of FABP4 gene in sheep mammary glands will be significant in revealing the importance of this gene in regulating milk fat formation,providing a theoretical basis for subsequent research on sheep lactation. 【Method】 The mammary gland and nipple tissues of Small-tailed Han sheep during the empty period and at 120 days of gestation were selected as research subjects,and the structure of the mammary gland and nipple tissues before and after pregnancy were observed using the hematoxylin-eosin staining method (HE).The distribution of FABP4 protein in mammary gland and nipple tissues before and after pregnancy was observed using immunohistochemistry,and the expression of FABP4 gene and protein in mammary gland and nipple tissues before and after pregnancy were detected using Real-time quantitative PCR and Western blotting,respectively. 【Result】 FABP4 gene was positively expressed in mammary gland and nipple tissues of Small-tailed Han sheep during the empty period and pregnancy.The expression of FABP4 gene in mammary gland and nipple of Small-tailed Han sheep during the empty period was significantly or extremely significantly higher than that during the pregnancy (P＜0.05 or P＜0.01).The expression of FABP4 gene in nipple during the empty period was extremely significantly higher than that in mammary gland (P＜0.01).The expression of FABP4 protein in mammary gland and nipple of Small-tailed Han sheep during the empty period was extremely significantly higher than that during the pregnancy (P＜0.01).The expression of FABP4 protein in nipple during the empty period and pregnancy was significantly higher than that in mammary gland (P＜0.05). 【Conclusion】 The expression of FABP4 gene in mammary gland and nipple tissues of Small-tailed Han sheep during pregnancy was significantly reduced,indicating that this gene played an important role in breast development and was involved in the periodic development of breast.

Artificial insemination technology plays a significant role in livestock genetic improvement and reproductive management,while semen cryopreservation facilitates the full utilization of superior sires’ reproductive potential and reduces production costs in the breeding industry.With the popularization of artificial insemination technology,the requirements for the quality and time of cryopreservation of ruminant semen have been also increasing.However,sperms in semen are susceptible to ice crystal damage and oxidative stress damage during the freezing-thawing process,which affects sperm motility and fertilization ability.As an important substance to reduce oxidative damage and improve semen quality,exogenous antioxidants have attracted extensive attention in the cryopreservation of animal semen.At present,numerous studies have demonstrated the beneficial effects of exogenous antioxidants in ruminant semen cryopreservation,highlighting their substantial potential for improving frozen semen quality.To this end,this article systematically reviews the effects of ultra-low temperature on oxidative damage in sperm and how antioxidants protect sperm from oxidative stress by scavenging reactive oxygen species (ROS).It comprehensively discusses the application of both enzymatic and non-enzymatic antioxidants in ruminant semen cryopreservation,including specific antioxidant types,optimal concentrations,and their precise effects on sperm motility parameters.In addition,the potential application value of novel antioxidants (e.g.,kaempferol,rutin,astaxanthin) in the freezing of semen from ruminants are also explored.However,the optimal concentration,application mode and mechanism of action of antioxidants still require further investigation and refinement.By systematically analyzing the efficacy of various antioxidant types and concentrations in ruminant semen cryopreservation,this review aims to provide theoretical foundations and practical references for advancing and optimizing ruminant frozen semen technologies.

【Objective】 This study aimed to investigate the effects of different fixed effects on semen traits,estimate the genetic parameters of semen traits in Duroc boars,and evaluate the genetic and phenotypic correlations among semen traits,providing a reference for the selection and production management of breeding boars. 【Method】 Semen-related traits of Duroc boars were assessed using a computer-assisted sperm analysis (CASA) system and flow cytometry.ANOVA and significance tests were conducted to examine the fixed effects on semen traits using R software,including semen collection method,semen collection season,and semen collection month.The genetic parameters of semen traits were estimated using single-trait and multi-trait animal models,applying the PBLUP method in ASReml software. 【Result】 The fixed effects of semen collection method,semen collection season,and semen collection month had varying impacts on each semen trait.The single-trait animal model estimation revealed that most semen traits in Duroc boars had low heritability,except semen density,which had medium heritability.Heritability estimates for each trait ranged from 0.11 (mitochondrial membrane potential) to 0.29 (semen density).Multi-trait animal model estimation showed that the heritability estimates for each trait ranged from 0.10 (abnormal sperm percentage) to 0.30 (semen density).In terms of genetic correlations,a strong negative correlation was observed between reactive oxygen species (ROS) level and acrosome integrity rate (r=-0.570),and between ROS level and mitochondrial membrane potential (r=-0.730).A strong positive correlation was found between semen density and sperm motility (r=0.860),as well as between mitochondrial membrane potential and acrosome integrity rate (r=0.930).In terms of phenotypic correlation,there was a moderate correlation between semen density and sperm motility (r=0.320),as well as between mitochondrial membrane potential and acrosome integrity rate (r=0.380). 【Conclusion】 Each fixed effect affected semen traits in Duroc boars,which were low to medium heritability,and there was a genetic correlation among semen traits,which could be selected to improve the semen quality of boars.

【Objective】 The purpose of this experiment was to understand the current epidemic status of Porcine pseudorabies virus (PRV) in China,the genetic status of the epidemic strain,the codon usage bias of PRV gE gene and the influencing factors. 【Method】 In this study,a phylogenetic tree was constructed using Mega 7.0 software for 106 PRV gE gene sequences isolated in China from 2019 to 2023.Launch DnaSP 6.0 software was used for gene selection pressure analysis and gene flow analysis.CodonW 1.4.2 and SPSS 27.0.1 software were used for codon analysis. 【Result】 The results of genetic evolution analysis showed that the PRV strains prevalent in China were genotype Ⅱ,which could be further divided into genotypes 2.1 and 2.2.Gene flow analysis showed that the nucleotide diversity (Pi) of genotypes 2.1 and 2.2 in genotype Ⅱ was 0.006 and 0.002,respectively.The coefficient of genetic differentiation (Fst) was 0.381,the synonymous substitution rate (Ks) and the quantitative index of gene flow (Z) were 1.353 and 6.637,respectively.Gene selection pressure analysis showed that the non-synonymous mutation to synonymous mutation ratio (dN/dS) of genotypes 2.1 and 2.2 was 1.808 and 1.362,respectively.The results of codon base analysis showed that the content of GC3 was more than 50% in the two subtypes of genotype Ⅱ,and the ENC values of different gene coding regions of genotypes 2.1 and 2.2 was 30.04 and 30.06,respectively.Codon usage bias analysis showed that the correlation coefficients of genotypes 2.1 and 2.2 was 0.115 and 0.173,respectively.Relative synonymous codon usage (RSCU) analysis showed that the third codon of high expression in genotypes 2.1 and 2.2 was G/C base,and the third codon of low expression was mainly A/U base.A total of 25 codons with RSCU ＞1 were used as preferred codons,of which 13 had high GC content and 16 ended with base C. 【Conclusion】 Genotype Ⅱ was the dominant PRV strain in China,which had a unique base composition,high GC content and relatively low AU content,showing a low preference.Codon usage was influenced by both mutation pressure and natural selection,in which natural selection played a dominant role.The results of this study provided scientific basis and technical support for the study of the epidemic trend of porcine pseudorabies in China and the formulation of prevention and control strategies.

【Objective】 The purpose of this test was to screen the single chain antibody fragment (scFv) against the non-structural protein NS1 of Porcine parvovirus (PPV), which laid a foundation for the further study of PPV. 【Method】 The expression of PPV NS1 protein was induced by prokaryotic expression system and purified,and the recombinant NS1 protein was used for animal immunization，and serum titer level was detected.Total RNA was extracted from mouse spleen and reverse-transcribed into cDNA,which was used as template to amplify the sequences of variable domain of heavy chains (VH) and variable domain of light chains (VL).scFv gene was amplified by overlap PCR,and was connected to pSEXRTL2 phage vector and electrotransformed into E.coli XLⅠ-Blue competent cells.A murine scFv antibody library against PPV NS1 was constructed,and its quality and storage capacity were measured.Phage display technology was used to screen scFv specific to NS1 protein,and the affinity between the antigen and the positive phage clone was detected by ELISA and Western blotting. 【Result】 In this experiment,soluble PPV NS1 recombinant protein was successfully expressed and purified.The capacity of the murine anti-NS1 scFv library was 2.7×10⁷ CFU/mL,and the positive rate of the library was 87.5%.After four rounds of "adsorption-elution-enrichment" phage screening,two scFv strains with high affinity for PPV NS1 protein were obtained. 【Conclusion】 In this study,a murine scFv phage library against PPV NS1 protein was successfully constructed by prokaryotic expression and purification of recombinant protein NS1.scFv that could bind specifically to PPV NS1 recombinant protein was obtained by phage display technique.The results provided the basis for further research and development of anti-PPV drugs and diagnostic reagents.

【Objective】 The purpose of this experiment was to study the molecular characteristics,reactogenicity and expression of methionine aminopeptidase Ⅱ (MAP Ⅱ) in different stages of Echinococcus granulosus development,and provide a vaccine candidate antigen with protective effect on the definitive host. 【Method】 Bioinformatics analysis of MAP Ⅱ protein was carried out,including phylogenetic tree construction,physical and chemical properties analysis,and prediction of signal peptide,transmembrane region,antigen epitopes,phosphorylation and glycosylation sites,and secondary and tertiary structures.The prokaryotic system was used to express MAP Ⅱ protein and the antigenicity of the recombinant protein was verified.At the same time,the transcript levels of different developmental stages of Echinococcus granulosus were analyzed. 【Result】 The MAP Ⅱ protein of Echinococcus granulosus was distantly related to that of the definitive host dog and the intermediate host sheep.MAPⅡ was composed of 453 amino acids with a molecular weight of 51 023.75 u.The instability coefficient was 36.6,indicating Map Ⅱ was a stable protein.The average hydrophobicity index was －0.609 and the hydrophilicity was good.There was no signal peptide and transmembrane region in MAP Ⅱ protein.Map Ⅱ protein had a good B cell linear epitope,with a total of 38 phosphorylation sites and 1 N-glycosylation site.The secondary structure of MAPⅡ protein was dominated by alpha helix and random coil,accounting for 33.55% and 51.88%,respectively.The tertiary structure prediction showed that it was a monomeric protein,and the GMQE value was 0.85.The MAP Ⅱ recombinant protein expressed by prokaryotic system could bind to the positive serum of dogs infected with Echinococcus granulosus,and had good antigenicity.MAP Ⅱ protein was expressed in both the protoscoleces stage and the adult stage,and the expression in the adult stage was significantly higher than that in the protoscoleces stage (P＜0.05). 【Conclusion 】 Map Ⅱ protein had stable structure and dominant antigen epitopes.The immunogenicity of MAP Ⅱ recombinant protein was good.It played an important role in different developmental stages of Echinococcus granulosus,especially in adult stages,and could be used as a vaccine candidate antigen for definitive host of Echinococcus granulosus.

【Objective】 The purpose of this study was to investigate the effect of Porcine deltacoronavirus (PDCoV) on the intestinal extracellular matrix (ECM) and its dynamic regulators in weaned piglets,in order to alleviate the infection of porcine pathogens and improve the intestinal health of piglets. 【Method】 Sixteen healthy 8-day-old piglets (Duroc×Landerace×Yorkshire) were selected for the experiment and randomly divided into control and PDCoV groups,8 piglets in each group with the control group receiving 20 mL of DMEM by oral gavage and the PDCoV group receiving 20 mL of PDCoV cell culture solution (10⁸ TCID₅₀/mL) by oral gavage,and another administration was given at an interval of 8 h.All piglets were slaughtered on the third day after challenge,and the middle jejunum and distal ileum samples were collected for the assay of ECM-related indices such as intestinal collagen,fibronectin (FN),integrin (ITG) and their related regulatory factors. 【Result】 Compared with control group,the mRNA expressions of interleukin-1β (IL-1β),IL-6 and IL-8 gene in jejunum of piglets in PDCoV group were significantly increased (P＜0.05).Masson staining showed that PDCoV increased jejunal collagen deposition.Compared with control group,the mRNA expressions of FN1 gene in jejunum and ITGA1 gene in ileum were significantly reduced in PDCoV group (P＜0.05).In jejunum,the mRNA expressions of ECM regulators matrix metalloproteinase-2 (MMP2),MMP9,plasminogen activator (PAs),transport and Golgi tissue 2 (TANGO2) gene in PDCoV group were significantly lower than those in control group (P＜0.05),and the mRNA expressions of endoplasmic reticulum channel protein (SEC6A1) and phosphodiesterase 4δ (PDE4D) gene were significantly higher than those in control group (P＜0.05).Compared with control group,the mRNA expressions of MMP2,PAs and SEC6A1 gene in ileum of piglets in PDCoV group were significantly decreased (P＜0.05),and the mRNA expression of MMP9 gene was significantly increased (P＜0.05).As shown by Spearman correlation analysis,the regulatory factors MMP2, TANGO2 and PAs gene were significantly negatively correlated with proinflammatory factors IL-1β and IL-8,and were significantly positively correlated with TGF-β gene (P＜0.05).SEC6A1 and PDE4D gene were significantly positive correlation with IL-1,IL-6 and IL-8 gene (P＜0.05),and PDE4D gene was significantly negative correlation with TGF-β gene (P＜0.05). 【Conclusion】 PDCoV increased the deposition of collagens by regulating the expression of ECM regulatory factors and decreasing the expression of fibronectin and integrins,leading to the dynamic changes of intestinal ECM and disrupting the intestinal homeostasis in weaned piglets.

【Objective】 This study was conducted to express GRA1 protein of Toxoplasma gondii in prokaryotic expression system,and develop an indirect ELISA (iELISA) method for rapid detection of Toxoplasma gondii antibody,and provide material for serological investigation of Toxoplasma gondii. 【Method】 The recombinant plasmid pET-28a-GRA1 was constructed,the GRA1 protein was expressed by Escherichia coli expression system,and the protein was purified with His nickel column.The immunogenicity of the protein was identified by Western blotting.The purified GRA1 protein was coated on the solid-phase carrier as antigen,and the optimal antigen coating concentration and serum dilution were determined by checkerboard titration method.On this basis,the antigen coating conditions,blocking conditions,serum dilution to be tested and enzyme-labeled secondary antibody dilution were optimized.The cutoff value of the established method was determined by measuring the D_{450 nm} value of negative sera,and the specificity,sensitivity and reproducibility tests were carried out.At the same time,the coincidence rate between the method and the modified agglutination test (MAT) for the detection of actual samples was determined. 【Result】 The results of SDS-PAGE showed that the GRA1 protein was expressed in soluble form,and the relative molecular weight of the protein was 25 ku.Western blotting result showed that the GRA1 protein reacted specifically with the porcine positive serum of Toxoplasma gondii.The optimized ELISA reaction conditions were:The optimal antigen encapsulation concentration was 5 μg/mL,with a serum dilution of 1∶100,antigen encapsulation was performed at 37 ℃ for 2 h,and the optimal containment condition was 5% skimmed milk powder at 37 ℃ for 2 h,and the action time of the serum to be tested was 60 min,the dilution of enzyme labeled secondary antibody was 1∶20 000,and the action time of TMB was 15 min.The cutoff value of this detection method was 0.277.This method had good sensitivity,positive results could still be detected when the serum was diluted at a ratio of 1∶6 400.The coefficients of variation of intra- and inter-assay repeatability were both less than 10%,indicating good repeatability.It had no cross-reaction with positive sera of Taenia solium,Trichinella spiralis,Porcine pseudorabies virus or Porcine reproductive and respiratory syndrome virus,and had good specificity.One hundred clinical porcine sera were tested using this method and the MAT method,and the total coincidence rate was 94.0%. 【Conclusion】 In this study,a rapid and effective indirect ELISA method for detecting antibodies of Toxoplasma gondii in pigs was established,which was helpful for the diagnosis and prevention of Toxoplasma gondii infection in pigs.

【Objective】 The experiment aimed to screen out a bacteriocin of lactic acid bacteria agaist Pasteurella multocida,complete the whole genome sequencing of the isolated lactic acid bacteria,and analyze the physicochemical properties and bactericidal mechanism of its bacteriocin. 【Method】 Healthy cat feces were inoculated in MRS broth medium and cultured at 37 ℃ for 12 h to isolate bacteria.With Pasteurella multocida as the indicator bacteria,the antibacterial activity of the isolated lactic acid bacteria was determined by Oxford cup agar diffusion method and the isolated lactic acid bacteria were identified by amplifying 16S rRNA gene.The whole genome sequence was determined and annotated in the KEGG database.The biological characteristics of the antibacterial substances produced by the isolated bacteria were determined using protease sensitivity,thermal stability and acid-base stability tests.The antibacterial properties of antibacterial substances were explored by using scanning electron microscopy technology. 【Result】 One strain of lactic acid bacteria producing antibacterial substances was screened out.Morphological observation showed that the surface of the colony was smooth and the morphology was small.Gram staining showed positive cocci.16S rRNA sequencing analysis showed that the similarity between the isolated strain and Weisseria confusa was 97%,and it was identified as Weisseria confusa,named JLP1.The results of whole genome sequencing showed that the genome size of Weisseria confusa JLP1 was 2 350 892 bp,with an effective average GC content of 44.94% and 2 020 coding genes.There were a total of 1 051 genes annotated in the KEGG database,which were the three major functions of environmental information processing,metabolism,and genetic information processing systems,with a total of 49 metabolic pathways.The results of acid excretion and hydrogen peroxide excretion test showed that the antibacterial substance of Weisseria confusa JLP1 was not organic acid and hydrogen peroxide,and the antibacterial activity decreased significantly after proteinase K treatment,so it was presumed that the antibacterial substance was bacteriocin.The antibacterial activity of bacteriocin produced by Weisseria confusa JLP1 was stable within 40 to 80 ℃,and did not lose activity after 10 minutes of treatment at 100 ℃,which had good thermal stability.It had good antibacterial activity within pH 3.0-8.0 and a wide range of acid-base tolerance.The scanning results of the electron microscope showed that the bacteriocin produced by Weisseria confusa JLP1 attacked the cell membrane of Pasteurella multocida,forming pores and causing the cell surface to shrink. 【Conclusion】 Weisseria confusa JLP1 had many metabolism and amino acid biosynthesis genes,and the bacteriocin produced by the strain had stable physicochemical properties and good antibacterial effect,providing a beneficial reference for preventing Pasteurella multocida infection.

【Objective】 This study aimed to solve the limitations of Litsea cubeba essential oil(LCEO) including poor aqueous solubility,high volatility and irritation, and improve its solubility and stability,thereby achieve the purpose of sustained release and strengthen the antibacterial efficacy in vitro. 【Method】 In this study,the β-cyclodextrin(β-CD) inclusion complex of LCEO was prepared through a saturated aqueous solution method with β-CD as coating material,and the optimal preparation conditions were systematically optimized via orthogonal design experiments.The structural and morphological properties of the inclusion compound were analyzed by Fourier transform infrared spectroscopy(FT-IR),inverted fluorescence microscope and scanning electron microscope(SEM).Furthermore,the stability test,dissolution test in vitro and antibacterial test against Escherichia coli,Salmonella,Staphylococcus aureus and Streptococcus suis in vitro were conducted to a comprehensive assessment. 【Result】 The optimal preparation process of clathrate was as follows:The ratio of β-CD to LCEO was 3∶1,the inclusion temperature was 40 ℃,the inclusion time was 4 h,and the volume ratio of anhydrous ethanol to LCEO was 1∶1.Under these conditions,the average yield of clathrate was 79.33% and the average inclusion rate was 84.40%.FT-IR analysis showed that the infrared spectral characteristic peaks of LCEO were hidden in the inclusion complexes,and the surface morphology and structure of β-CD and LCEO-β-CD inclusion complexes were significantly different under inverted fluorescence microscopy and SEM.The results of light and high temperature stability tests showed that under the condition of light intensity of (4 500±500) lx and 60 ℃,the performance of the clathrate was stable within 10 days,while the content of LCEO in the physical mixture decreased significantly,indicating that the clathrate had good light and thermal stability.The results of in vitro dissolution test showed that all the LCEO in the physical mixture of LCEO/β-CD were dissolved within 120 min,while the cumulative release of LCEO-β-CD clathrate was 84.36% within 240 min.The results of kinetic model showed that the release of clathrate in simulated intestinal fluid was about 2.7 times that in simulated gastric fluid,The results showed that the inclusion compound of LCEO-β-CD inclusion complexes had sustained release and enteric solubility.The results of in vitro antibacterial test showed that the MIC of LCEO against Escherichia coli,Salmonella,Staphylococcus aureus and Streptococcus suis were 1.56,1.56,0.78 and 0.39 μL/mL,respectively,while the MIC corresponding to LCEO-β-CD inclusion compound were 0.78,0.78,0.39 and 0.19 μL/mL,respectively.The results showed that clathrate could improve the bacteriostatic effect on Escherichia coli,Salmonella,Staphylococcus aureus and Streptococcus suis,and the bacteriostatic curve showed that clathrate could prolong the bacteriostatic time. 【Conclusion】 The LCEO-β-CD inclusion complexes prepared in this study effectively overcome the poor water solubility and strong volatility of LCEO.It had sustained-release and enteric-coated effects,which could enhance the antibacterial effect and prolong the antibacterial time in vitro.

【Objective】 This study was conducted to understand the main pathogenic bacteria causing abortion in Yili horses,and provide scientific basis for their clinical guidance of medication. 【Method】 In this experiment,35 tissue samples of heart,liver,spleen,lung,kidney,umbilical cord and amniotic membrane from 5 spontaneously aborted foals were collected for bacterial enrichment and culture,purification,PCR identification and sequencing,and the isolated strains were tested for drug resistance and pathogenicity. 【Result】 Through bacterial isolation and culture,morphological observation,6 strains were isolated from the tissue of the aborted foal.One strain was a large grayish-white mucus colony on the agar plate,easy to stretch when picked,Gram-negative,short,thick,rod-shaped,individually or in pairs arranged bacilli,suspected to be Klebsiella sp.One strain was glossy surface,with an untidy edge of semi-transparent or opaque,Gram-negative,short,bluntly rounded at the ends,suspected to be Bacillus sp. One stain was stered into clusters of cocci,suspected to be Staphylococcus spp. One stain was round,white or creamy-white colony with smooth surfaces,Gram-negative,and long rods with bluntly rounded ends,suspected to be Escherichia coli.One strain was greyish-white round colony with a small bulge and β-hemolysis ring on a sheep blood agar plate,Gram-positive cocci,in pairs or in long chains,suspected to be Streptococcus spp.One strain was a round colony with a smooth surface and purple color,with neat edges,in the Salmonella chromogenic medium,and was Gram-negative,and short bacilli with bluntly rounded ends,suspected to be Salmonella.The results of PCR amplification of 16S rRNA gene of six isolates showed that the strains all showed a single band at 1 540 bp.BLAST alignment of 16S rRNA gene sequencing results showed that one isolate had more than 95% similarity to Klebsiella sp. published in GenBank, and five isolates had more than 97% similarity to Citrobacter freundii, Staphylococcus lentus, Escherichia coli, Streptococcus equi subsp. zooepidemicus and Salmonella published in GenBank, respectively.The virulence gene detection results of each strain were consistent with the expected results.The six strains of isolated pathogenic bacteria were finally identified as Klebsiella sp., Citrobacter freundii,Staphylococcus lentus,Escherichia coli, Streptococcus equi subsp.zooepidemicus and Salmonella,and named 23-KLB-16S,23-NM-16S,23-HM-16S,23-DC-16S,23-ML-16S and 23-SM-16S，respectively.The similarity and genetic evolution results showed that the isolates 23-KLB-16S and Klebsiella sp.,23-NM-16S and Citrobacter freundii,23-HM-16S and Staphylococcus lentus,23-DC-16S and Escherichia coli,23-ML-16S and Streptococcus equi subsp.zooepidemicus,23-SM-16S and Salmonella reference strains were similar and closely related.The results of drug sensitivity showed that the isolates were multi-drug resistant,generally more sensitive to tetracyclines and fluoroquinolones,and resistant to lincosamide.The results of the mouse attack test showed that the LD₅₀ of the above six pathogenic bacteria were 1.54×10²,2.59×10⁷,3.62×10⁷,7.02×10⁷,6.08×10⁶ and 8.15×10⁵ CFU/mouse,respectively,with Klebsiella sp.,Streptococcus equi subsp.zooepidemicus,and Salmonella being more pathogenic.The autopsy results of mice showed varying degrees of bruising and enlargement of liver and spleen,and hemorrhage in lung. 【Conclusion】 Six main pathogenic strains,including Klebsiella sp.and Citrobacter freundii,were isolated from tissue samples of aborted foals,and mixed infections were found.All six strains were multidrug-resistant and generally sensitive to tetracyclines and fluoroquinolones antibiotics,but resistant to lincosamides.Among them,Klebsiella sp.,Streptococcus equi subsp. zooepidemicus and Salmonella had strong pathogenicity.

In recent years,the use of antimicrobials has faced serious challenges,and the continuous increase of multi-drug-resistant microorganisms has severely limited the use of antimicrobials.The mechanism of bacterial resistance is complex and varied,and the development and production of new antimicrobials is time-consuming and costly.Therefore,it is particularly important to explore alternative therapies to treat drug-resistant bacterial infections.Antimicrobial peptides (AMPs),as a kind of unique and diverse small molecular peptides,have the characteristics of broad antibacterial spectrum,antiviral,not easy to produce drug resistance,immune regulation,etc.,and have shown great potential in the treatment of drug-resistant bacterial diseases.However,AMPs faces many challenges in clinical application,such as high toxicity to host cells and easy degradation by proteases.Researchers are actively exploring a variety of strategies to optimize the structural design of AMPs in order to improve their biological activity while reducing adverse reactions.In this paper,the origin,structural properties and mechanism of AMPs are described.The recent breakthroughs in improving the activity of AMPs are discussed,including amino acid residue replacement,recombinant fragment design,heterozygous peptide construction,incorporation of unnatural amino acids and other chemical modifications.At the same time,the potential value of AMPs combined with conventional antibiotics,lysozyme and nanomaterials was analyzed，in order to provide a theoretical basis for the development and application of AMPs in the fields of medical health,agriculture and food safety.

【Objective】 The purpose of this study was to observe the effect of Platycodon grandiflorum ultrafine powder on the diversity of intestinal flora in mice,and provide evidence for clinical application and safety of Platycodon grandiflorum ultrafine powder. 【Method】 Twenty-four male Kunming mice were randomly divided into control group and high (3.0 g/kg),medium (1.5 g/kg) and low (0.3 g/kg) dose groups of Platycodon grandiflorum ultrafine powder，6 mice in each group.Mice in each dose groups of Platycodon grandiflorum ultrafine powder were gavaged with the corresponding dose of Platycodon grandiflorum ultrafine powder dissolved in water,and mice in control group were gavaged with equal volume of purified water,the medication was administered continuously for 14 days.The body weight and feed consumption of mice in each group were recorded daily.On the 15th day of the experiment,mice in each group were sacrificed for autopsy and histopathological observation.Meanwhile,the feces of mice were aseptically collected to amplify the V3-V4 regions of the 16S rRNA gene of the fecal microbiota.High-throughput sequencing was used to analyze Alpha and Beta diversity of the microbiota and the species composition at phylum and genus levels.And microbial function prediction and analysis were conducted. 【Result】 ①The body weight gain of mice in medium and high dose groups were significantly lower than those in control group (P＜0.05).The average feed consumption of mice in medium dose group of Platycodon grandiflorum superfine powder was lower than that in control group.②Compared with control group,no pathological abnormalities were found in stomach,duodenum,jejunum and ileum of mice in each dose group of Platycodon grandiflorum superfine powder.③The Alpha diversity index in intestinal flora of mice in each dose group of Platycodon grandiflorum superfine powder was higher than that in control group(P＞0.05),and increased with the increase of the dosage.Beta diversity analysis of intestinal flora showed that low and high doses of Platycodon grandiflorum superfine powder had no significant effect on the structure of intestinal flora in mice,while medium doses of Platycodon grandiflorum superfine powder could significantly change the structure of intestinal flora in mice (P＜0.05).④Compared with control group,at phylum level,the contents of Firmicutes and Cyanobacteria in the intestinal tract of mice in medium dose group of Platycodon grandiflorum ultrafine powder were significantly increased (P＜0.05),while the abundance of Bacteroidetes,Proteobacteria and Planctomycetes was significantly decreased (P＜0.05).At genus level,the abundance of Prevotella,Parabacteroides and rc4_4 in the intestinal tract of mice in medium dose group was significantly increased (P＜0.05),while the abundance of Kaistobacter and Pseudomonas etc.was significantly decreased (P＜0.05).⑤Microbial function prediction analysis found that the intestinal microbial genome functions of mice in medium dose group of Platycodon grandiflorum superfine powder were mainly enriched in amino acid biosynthesis and cofactors,prosthetic groups,electronic carriers and vitamin biosynthesis and nucleotide biosynthesis.Compared with control group,there were 11 significantly downregulated differential metabolic pathways in medium dose group of Platycodon grandiflorum superfine powder,mainly involving the degradation of fluorobenzoates,polycyclic aromatic hydrocarbons and yeast meiosis. 【Conclusion】 Platycodon grandiflorum ultrafine powder was able to increase the diversity of intestinal flora in mice,and 1.5 g/kg Platycodon grandiflorum ultrafine powder could significantly change the structure of intestinal flora.The results of this experiment provided data support for the scientific dosage and safety of Platycodon grandiflorum ultrafine powder’s clinical use,and also support for future pharmacodynamic studies related to Platycodon grandiflorum ultrafine powder and intestinal flora.

【Objective】 This study aimed to investigate the antibacterial effect of isobavachalcone (IBC),a natural plant-derived flavonoid compound,and its combination with six commonly used antibiotics on methicillin-resistant Staphylococcus aureus (MRSA),to observe whether isobavachalcone had antibacterial synergistic effect on commonly used antibiotics,and provide reference for isobavachalcone as a potential antibacterial drug or antibacterial synergist. 【Method】 Minimal inhibitory concentrations (MIC) of six antibiotics (polymyxin B,enrofloxacin,oxytetracycline,florfenicol,ampicillin and amoxicillin) against MRSA T144 were determined by broth microdilution method.The chessboard method combined with drug sensitivity test was used to determine the effect of drug combination.The fractional inhibitory concentrations index (FICI) was used to determine their synergistic effect,and the antibacterial drugs with synergistic effect were selected.Staphylococcus aureus,Bacillus cereus,Escherichia coli and Salmonella were used as test strains to explore whether isobavachalcone combined with the screened antibacterial could have antibacterial synergistic effect on common clinical Gram-positive and Gram-negative bacteria.SYTO9-PI staining was performed to evaluate the effect of isobavachalcone on the cell membrane permeability of the tested strains.The content of antibacterials in the bacteria was determined according to the change of PI fluorescence intensity. 【Result】 The results of drug sensitivity test showed that polymyxin B,oxytetracycline,ciprofloxacin,florfenicol,ampicillin and amoxicillin all showed inhibitory effects on MRSA T144,and the MIC was 8-64 μg/mL.The results of combined drug sensitivity test showed that the combination of isobavachalcone and polymyxin B had a synergistic effect on MRSA T144 (FICI＝0.5).When isobavachalcone was combined with oxytetracycline,florfenicol,ampicillin and amoxicillin respectively,it had additive effect on MRSA T144 (FICI was 0.5625-1).There was no correlation between isobavachalcone and ciprofloxacin (FICI＝2).When isobavachalcone and polymyxin B were combined,they showed a synergistic effect on MRSA T144 (FICI＝0.5).The results of combined drug sensitivity test showed that when isobavachalcone and polymyxin B were used in combination,there was a synergistic effect on Staphylococcus aureus and Bacillus cereus (FICI was 0.258-0.375).It showed a partial synergistic effect on Escherichia coli and Salmonella (FICI was 0.531-0.563).The results of cell membrane permeability test further showed that the combined treatment group could significantly increase the number of bacteria MRSA T144,Staphylococcus aureus,Bacillus cereus,Escherichia coli and Salmonella with changed membrane permeability compared with polymyxin B and isobavachalcone alone (P＜0.05).The results of polymyxin B content in bacteria showed that compared with polymyxin B alone,the concentration of polymyxin B in Escherichia coli and MRSA T144 was significantly increased when polymyxin B was combined with isobavachalcone (P＜0.05). 【Conclusion】 Isobavachalcone combined with polymyxin B had certain antibacterial synergistic effect on MRSA T144,Staphylococcus aureus,Bacillus cereus,Escherichia coli and Salmonella,isobavachalcone could change the membrane permeability of the tested strains,thereby reducing the dose of polymyxin B.This study could provide new ideas for clinical treatment of bacterial infection,and also provide a reference for broadening the use of polymyxin B in animal husbandry in the future.

The immune system consists of immune organs,immune cells and immunologically active substances,which can recognize and eliminate pathogens and abnormal cells.The intestinal flora is involved in the body’s digestion,absorption and metabolism,and the regulation of the immune system and nervous system.Intestinal flora is a complex community composed of all microorganisms in the intestinal tract.During the long-term co-evolutionary process,there is a dynamic interplay between the intestinal flora and the host immune system,which affects the host’s intestinal health and immune function,and dysregulation of the intestinal flora can cause inflammatory reactions and diseases in the host.At the same time,intestinal flora can metabolize protein,dietary fiber,primary bile acids,linoleic acids and carbohydrates,and produce a large number of metabolites,such as short-chain fatty acids,bile acids and indole，which affect the interaction between intestinal epithelial cells and immune cells,including host metabolism,intestinal barrier and immune function.At present,there are still some shortcomings in the study of intestinal flora in livestock and poultry,such as incomplete understanding of intestinal flora diversity,unclear function and cooperation mechanism,limitations in research methods and lack of regulatory means.In this paper,the effects of intestinal flora and its metabolites on the immune system of livestock and poultry were systematically reviewed in order to better understand the complex interaction between intestinal flora and immune system of livestock and poultry,and provide theoretical basis for developing new strategies and methods for the prevention and treatment of related diseases.

【Objective】 The experiment aimed to study the bacteriostatic effect of commonly used antibiotics and 24 Chinese herbal medicines on Aeromonas hydrophila and Bacillus cereus,as well as the bacteriostatic effect of the combination of drugs,so as to select effective antibacterial formulas for production practice and treat bacterial infections. 【Method】 Several antibiotics with strong inhibitory effect on Aeromonas hydrophila and Bacillus cereus were screened by drug sensitive disk method,and the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of these antibiotics and 24 Chinese herbal medicines against the above two pathogens were determined.According to the MIC and MBC of Chinese herbal medicine,the effective Chinese herbal medicine was selected to be combined with antibiotics.The fractional inhibitory concentrations index (FICI) of the combined drugs against Aeromonas hydrophila and Bacillus cereus was determined by the broth chessboard dilution method.The combined effect of drugs was judged,and the effective combination of Chinese and Western medicine was selected.One group of Chinese and Western medicine was made into a combined prescription for in vivo test. 【Result】 The antibacterial effect of antibiotics and Chinese herbal medicine on Aeromonas hydrophila showed that the antibiotic enrofloxacin had the best effect (MIC was 0.39 mg/mL),and the Chinese herbal medicine Scutellaria baicalensis Georgi had the best effect (MIC was 7.81 mg/mL).The results of combined medication showed that the FICI of Scutellaria baicalensis Georgi and enrofloxacin was 0.127,showing a synergistic effect,and the FICI of Pericarpium granati and enrofloxacin was 0.562,showing an additive effect.The antibacterial effect of antibiotics and Chinese herbal medicine on Bacillus cereus showed that the antibiotic ceftazidime had the best effect (MIC was 0.78 mg/mL),and the Chinese herbal medicine Coptis chinensis Franch.and Scutellaria baicalensis Georgi had the best effect (MIC both were 7.81 mg/mL).The results of combined medication showed that the FICI of Coptis chinensis Franch.and ceftazidime was 0.5,showing a synergistic effect.The in vivo test results of the combined medication showed that Pericarpium granati and enrofloxacin combined medication group had complete liver structure,closely arranged hepatocytes,at the same time,the intestinal tissues were clearly defined,and the intestinal mucosa was intact,and had a good protective effect on the intestinal mucosa,effectively alleviating the damage caused by Aeromonas hydrophila to the liver and intestine of Micropterus salmoides. 【Conclusion】 In vitro test,Scutellaria baicalensis Georgi combined with enrofloxacin and Pericarpium granati combined with enrofloxacin had inhibitory effect on Aeromonas hydrophila.Coptis chinensis Franch.combined with ceftazidime had strong antibacterial effect on Bacillus cereus.The combination of Pericarpium granati and enrofloxacin had a good therapeutic effect on Aeromonas hydrophila infected Micropterus salmoides.

【Objective】 The purpose of this experiment was to prepare probiotic fermented compound dandelion powder and evaluate its clinical safety. 【Method】 The probiotic properties of Saccharomyces cerevisiae and Bacillus subtilis were detected through artificial gastrointestinal fluid tolerance tests.The probiotic fermented compound dandelion powder was made by mixing Saccharomyces cerevisiae and Bacillus subtilis with 13 Chinese herbs，such as dandelion,Pulsatilla chinensis and hawthorn through solid-state fermentation.The quality of fermentation was evaluated by measuring the pH,odor,color and texture of the fermented products.The safety of fermented products was detected through acute toxicity test in mice.Thirty Kunming mice were randomly divided into 3 groups,with 10 mice in each group,half male and half female.Mice in test groups Ⅰ and Ⅱ were gavaged with compound dandelion powder preparation and probiotic fermented compound dandelion powder preparation at a dose of 5 000 mg/kg BW，respectively.Mice in control group were gavaged with an equal volume of normal saline，continuously for 7 days.Record the clinical symptoms and death situations of mice every day.After the experiment,the mice were weighed,blood was collected from the eyeballs,and the biochemical and immune indicators of the blood were detected.After post-mortem examination of mice,internal organs were collected,organ coefficients were calculated and tissue sections were made to observe pathological changes. 【Result】 The survival rates of Saccharomyces cerevisiae and Bacillus subtilis in artificial gastric fluid were 39.91% and 63.72%，and in artificial intestinal fluid were 47.61% and 22.04%,respectively.During the fermentation process of probiotic fermented compound dandelion powder,the pH showed a downward trend with the extension of time,reaching the lowest pH at 60 h.Sensory properties observation showed that the color of the fermented product had deepened,presenting a yellowish-brown color,soft texture,and emitting a rich sour aroma.No spoilage or deterioration had occurred.The results of the acute toxicity test in mice showed that none of the mice died during the test period.The clinical manifestations and histopathological examinations of the mice in experimental group showed no significant changes compared with those in control group.The liver index of mice in test group Ⅰ was significantly lower than that of control group (P＜0.05).The heart index of mice in test group Ⅱ was significantly lower than that of control group and test group Ⅰ (P＜0.05).The coefficients of other organs showed no significant differences among the groups (P＞0.05).The ALP activity in the serum of mice in test group Ⅰ was significantly higher than that in control group (P＜0.05),while there were no significant differences in other blood biochemical indicators and immune indicators among the groups (P＞0.05). 【Conclusion】 The probiotic fermented compound dandelion powder had good fermentation quality,was non-toxic to mice and had good safety.

With the increasingly serious problem of bacterial drug resistance,bacterial diseases in veterinary clinical practice have become more intractable,and the development of novel anti-infective drugs is extremely urgent.The anti-virulence strategy targeting bacterial virulence factors has become a promising antibacterial approach to avoid the selective pressure-induced resistance associated with conventional antibiotics.The flavonoid epigallocatechin gallate (EGCG), with its unique structures such as polyhydroxy,esterification modification,and stereoconfiguration,can bind to bacterial virulence factors and has inhibitory effects on the virulence factors of various bacteria. It is an ideal anti-virulence drug.The author reviewed the anti-virulence strategies of EGCG,summarized the physicochemical properties and pharmacological effects of EGCG,and clarified that this compound mainly inhibits bacterial virulence through direct binding to virulence factors,hindering the expression of virulence genes,preventing the secretion of virulence factors,and indirect anti-virulence action pathways.The inhibitory effects and mechanisms of EGCG on different types of virulence factors (endotoxin,hemolysin,enterotoxin,enzymes,biofilms and virulence regulatory systems) were further summarized and explored.From the perspective of clinical treatment,the application prospects and problems faced by the development of EGCG into an anti-virulence drug were discussed,aiming to provide a theoretical basis for the research of novel anti-virulent drugs in the veterinary field and promote the healthy and sustainable development of animal husbandry through anti-virulence strategies.

【Objective】 The experiment aimed to explore the mechanism of Shuanghuanglian (SHL) in regulating cross-endothelial lymphocytes against H9N2 subtype Avian influenza virus (AIV) infection,and provide new ideas and theoretical basis for the research and development of traditional Chinese medicines against Influenza virus. 【Method】 The drug toxicity of SHL-containing medium with different concentrations and action time on rat pulmonary microvascular endothelial cells (rPMVECs) was detected by MTT assay,and appropriate drug concentrations and time were screened. The effect of SHL on the replication dynamics of the H9N2 subtype avian influenza virus (H9N2 subtype AIV) in rPMVECs was investigated.The various indicators of rat peripheral lymphocyte proliferation capacity,including lymphocyte concentration,FBS concentration,and incubation time were optimized.On this basis,a Transwell co-culture system was established between rPMVECs and lymphocytes.rPMVECs were inoculated into the upper chamber and divided into six groups:Virus group,Particle group,Virus+SHL group,Particle+SHL group,EC group,and Blank group,and lymphocytes were inoculated.After incubation of H9N2 subtype AIV with cross-endothelial lymphocytes and subsequent inoculation on MDCK cells,the antiviral effect of SHL was evaluated by assessing the number of dead MDCK cells and the amount of live virus in the supernatant. 【Result】 No significant cytopathic effects were observed in rPMVECs co-cultured with ≤100 μg/mL concentration of SHL for 24 h.SHL exhibited strong inhibitory effects on H9N2 AIV proliferation when co-cultured with virus-infected rPMVECs at a concentration of 100 μg/mL for 24 h.The number of dead MDCK cells and the amount of live virus in the Particle group were significantly lower than those in Virus group (P＜0.05).The number of dead MDCK cells and the amount of live virus in the Virus+SHL group were significantly lower than those in EC group and Virus group(P＜0.05). 【Conclusion】 SHL played a crucial role in protecting the host from H9N2 subtype AIV by maintaining the integrity of endothelial cells and activating their recruitment of lymphocytes to exert immune effects,thereby reducing viral load and exerting antiviral effects.

Ketosis is a nutritional metabolic disease caused by the imbalance of glucose and lipid metabolism in perinatal dairy cows.It is mainly manifested by the accumulation of ketone bodies and the increase of free fatty acids,accompanied by abnormal concentration of blood sugar,insulin and glucagon.The disease seriously endangers the health and production performance of dairy cows and causes huge losses to the dairy farming industry.The core link of ketosis is the disorder of glucose and lipid metabolism.As an important endogenous homeostasis regulation system in mammals,the circadian clock can regulate the circadian rhythm of glucose and lipid metabolism by regulating the secretion of hormones related to glucose and lipid metabolism,the expression of rate-limiting enzyme genes and nuclear receptor genes.This paper reviewed the connection between ketosis and glycolipid metabolism disorders in dairy cows,the role of the circadian clock in the regulation of circadian rhythm homeostasis of lipid and carbohydrate metabolism in mammals,the circadian rhythm of metabolic physiology regulated by circadian clock in dairy cows,and the effects of ketosis on the circadian rhythm of glucose and lipid metabolism in dairy cows,in order to understand the pathogenesis of ketosis in dairy cows more comprehensively from the perspective of chronobiology,and provide new ideas for exploring accurate and efficient prevention and treatment strategy of ketosis.

【Objective】 This study was conducted to screen and isolate lactic acid bacteria from the cecum samples of healthy Zi geese as a candidate strain for the development of probiotics for geese. 【Method】 The isolated bacteria were isolated and identified by morphological observation,physiological and biochemical tests and 16S rDNA sequence analysis.The growth characteristics and pH curve,acid and bile salt resistance,heat resistance,osmotic pressure resistance,self-agglutination ability and surface hydrophobicity,antibacterial activity,drug sensitivity and safety were studied. 【Result】 A strain of Gram-positive bacteria was isolated from the cecum of healthy Zi geese.It was a small round colony with moist and smooth colony morphology on MRS medium,and was named DT021.The biochemical test results showed that it could ferment glucose,maltose,lactose,cellobiose,sucrose,raffinose,mannitol and esculin,the catalase and nitrate reduction tests were negative,and it was preliminarily determined as lactic acid bacteria.16S rDNA gene sequence analysis showed that the similarity between the isolated strain and 4 reference strains of Lactobacillus plantarum in NCBI was all 100%,and the isolated strain was identified as Lactobacillus plantarum.The growth curve and pH curve showed that the bacteria entered the logarithmic phase at 2 h,entered the stable phase at 16 h,pH decreased to below 4 at 18 h,and reached the lowest value of 3.82 at 32 h.The survival rates the isolated strain in MRS broth at pH 3.0 and 2.0 were 45.0% and 38.9%,respectively.The survival rates in MRS broth with 0.2% and 0.3% bile salt were 15.5% and 9.0%,respectively.After 50 ℃ water bath treatment,the survival rate was more than 50%,and the survival rate could reach 95.4% after 10% sodium chloride content broth treatment.The results of self-aggregation ability and surface hydrophobicity showed that the isolated strain was a moderately agglutinating and moderately hydrophobic strain.In addition,the isolated strain showed good antibacterial activity against five goose-derived pathogenic bacteria including Escherichia coli,Staphylococcus aureus,Salmonella,Proteus mirabilis,and Aerococcus viridans,and the diameters of the inhibition zone were all more than 13.5 mm.The results of drug test showed that the isolated strain was resistant to 6 drugs such as amikacin and norfloxacin,and was sensitive to β-lactam drugs.After 2 weeks of intragastric administration of the isolated bacteria liquid,there was no death or lesion in mice,indicating that the isolated strain had no toxic effect on mice. 【Conclusion】 Lactobacillus plantarum DT021 isolated in this study had good probiotic properties,and was safe and non-toxic side effects.It had potential development value in the field of goose probiotic preparations.

【Objective】 The purpose of this experiment was to investigate the in vitro antibacterial effect of berberine combined with antimicrobial drugs on Escherichia coli of animal origin. 【Method】 The minimum inhibitory concentration (MIC) of berberine and antimicrobial drugs (enrofloxacin,ciprofloxacin,doxycycline,flufenicol and colistin) against 18 strains of Escherichia coli of animal origin were determined by microbroth dilution method.The fractional inhibitory concentration index (FICI) of the combination of berberine and antimicrobial drugs was determined by checkerboard dilution method.For FICI≤0.50 drug combination,the time-bactericidal curve was determined to analyze its bacteriostatic dynamics. 【Result】 The resistance rates of 18 strains of Escherichia coli to enrofloxacin,ciprofloxacin,doxycycline,florfenicol,and colistin was 55.56%,55.56%,50.00%,88.89% and 27.78%,respectively.The FICI of berberine combined with enrofloxacin ranged from 0.31 to 2.00,and the synergistic rate was 33.33%.The FICI of berberine combined with ciprofloxacin ranged from 0.31 to 2.00,and the synergistic rate was 27.78%.The FICI of berberine combined with doxycycline ranged from 0.50 to 1.00,and the synergistic rate was 27.78%.The FICI of berberine combined with florfenicol ranged from 0.63 to 0.75,and the synergistic rate was 0.The FICI of berberine combined with colistin ranged from 0.31 to 1.00,and the synergistic rate was 66.67%.The time-sterilization curve showed that 1/2 MIC berberine combined with 1/2 MIC colistin against strains E01 and E06,1/2 MIC berberine combined with 1/2 MIC enrofloxacin against strain E-05,and 1/2 MIC berberine combined with 1/2 MIC doxycycline against strain E-03,the concentration of bacterial solution decreased by more than 2 lg CFU/mL compared with the monotherapy effect at 24 h,showing obvious synergistic effect. 【Conclusion】 The combination of berberine and antimicrobial drugs exhibited varying degrees of synergistic or additive effects on most strains of Escherichia coli,which provided a reference for enhancing the efficacy of antimicrobial drugs in veterinary clinical practice.

The emergence of antibiotic resistance represents a significant challenge to public health,undermining the considerable progress made over decades in combating bacterial infections.The dearth of novel and efficacious antimicrobial agents,coupled with the horizontal transfer of antibiotic resistance genes (ARGs) within the ecosystem,has served to exacerbate the dissemination of resistance.Among these mechanisms,plasmid-mediated conjugation represents the primary means by which ARGs disseminate among humans,animals,and the environment.The conjugative transfer of ARGs is frequently driven by antibiotic pressure.Moreover,certain non-antibiotic pharmaceuticals have also been observed to facilitate the horizontal transfer of ARGs.However,the role of natural compounds in non-antibiotic drugs,particularly plant extracts,in the horizontal transfer of ARGs has been undervalued.Accordingly,this review initially delineated the principal route of horizontal transfer of ARGs,namely conjugation.Thereafter,it delineated the utilization of plant extracts in the livestock and poultry industry and explored the role of plant extracts in promoting or inhibiting conjugation,and discussed the potential and challenges of applying these conjugation inhibitors in clinical trials,so as to guide the clinical development of more reasonable dosage regimens and drug residue standards.

【Objective】 This study aimed to investigate the anti-inflammatory and antibacterial effects of platelet rich plasma (PRP) on infectious endometritis in donkeys. 【Method】 Healthy donkey endometrium was selected as explants for in vitro culture,and 1.82×10⁷ CFU/mL Streptococcus equi subsp. zooepidemicus (S. zooepidimicus) was added to induce an infection-based endometritis explant model.The experiment was divided into 7 groups,which were mock control group (MCG),model group (MG),positive drug control group (PDCG),25% PRP and 50% PRP groups,MIX1 and MIX2 group.After the experiment,the culture medium and explants of each group were collected,and the concentration of interleukin-1 (IL-1) and IL-6 were detected in the culture medium,and the explants were examined by histology.The antibacterial and anti-inflammatory effects of PRP were evaluated by comparing bacterial concentrations in culture media,histopathological inflammatory changes in explants,and the levels of inflammatory factors. 【Result】 The PRP platelet concentration was 4.69×10¹¹/L,the platelet-derived growth factor (PDGF) concentration was 32.01 ng/mL,and the transforming growth factor β1 (TGF-β1) concentration was 49.37 ng/mL.The bacterial concentration in culture medium of MG group was extremely significantly higher than that of MCG group (P<0.01),and the bacterial concentrations in culture medium of 25% PRP,50% PRP,MIX1 and MIX2 groups were extremely significantly lower than that of MG group (P<0.01).In MCG group,the endometrium structure of the explants was intact,and few polymorphonuclear leukocytes were distributed.In MG group,the endometrial structure was severely damaged and polymorphonuclear leukocytes were infiltrated.Compared with MG group,PRP treated groups had more complete epithelial and glandular structures and less infiltration area of polymorphonuclear leukocytes.The concentration of IL-1 and IL-6 in culture medium of MG group was extremely significantly higher than that of MCG group (P<0.01).The concentrations of IL-1 and IL-6 in culture medium treated with PRP were extremely significantly lower than those in MG and PDCG groups (P<0.01). 【Conclusion】 In this study,an explants model of donkey infective endometritis induced by S. zooepidimicus was successfully constructed.PRP could significantly inhibit the growth of bacteria in explants,down-regulate the expression of inflammatory factors IL-1 and IL-6,and alleviate tissue inflammation in the explants.