【Objective】 This study was aimed to clone SMAD homolog 4 (SMAD4) gene and explore its biological function in sheep,so as to provide a basis for studying the regulatory role of SMAD4 gene in the growth and development of hair follicles.【Method】 Using the cDNA of skin in Small-tailed Han sheep as a template,the CDS region of SMAD4 gene was amplified by PCR and cloned, and was sequenced for bioinformatics analysis.Real-time quantitative PCR was used to detect the expression of SMAD4 gene in heart,liver,spleen,kidney,skin and muscle in Small-tailed Han sheep and Xinji Fine wool sheep.【Result】 The CDS region of SMAD4 gene in Small-tailed Han sheep was successfully cloned,which contained 1 662 bp and encoded 553 amino acids.Phylogenetic tree analysis showed that the nucleotide sequence of SMAD4 gene in sheep was the closest to Capra hircus and Bos mutus,and the most distant with Gallus gallus.Bioinformatics analysis found that SMAD4 protein was a non-transmembrane protein without signal peptides,with phosphorylation and glycosylation sites,and belonged to hydrophilic proteins,mainly in nucleus.The secondary structure of SMAD4 protein was dominated by random coil,which was consistent with the tertiary structure.The results of protein interaction analysis showed that SMAD4 protein interacted with SMAD2,SMAD1,TGFBR1,TGFBR2,SMAD3 and other proteins.Real-time quantitative PCR results showed that SMAD4 gene was widely expressed in sheep.Compared with Xinji Fine wool sheep,the expression of SMAD4 gene in heart,spleen and skin of Small-tailed Han sheep were significantly increased (P＜0.05),but the expression of SMAD4 gene in lung and muscle were significantly decreased (P＜0.05).【Conclusion】 The CDS region of SMAD4 gene in sheep was successfully cloned,which was expressed in various tissues of sheep,and there were significant differences in heart,spleen,skin,lung and muscle between Small-tailed Han sheep and Xinji Fine wool sheep.The results provided a theoretical basis for further exploration of the regulation of hair follicle development by SMAD4 gene in sheep.

【Objective】 This study was aimed to developed a rapid,accurate,and sensitive visualization detection method for Cryptosporidium parvum (C. parvum). 【Method】 In this study,the recombinase polymerase amplification (RPA) was combined with the CRISPR/Cas12a system,and the results were read by the fluorescence reporter system and the lateral flow strip biosensor. The crRNA and RPA primers were designed and screened against the small subunit ribosome RNA (SSU rRNA) gene of C. parvum,and two different single-stranded DNA (ssDNA) reporters were used to present the detection results by fluorescence values and the LFS biosensor.The specificity of the RPA-CRISPR/Cas12a assay was verified by detecting C. parvum and 7 other pathogens.The sensitivity of the RPA-CRISPR/Cas12a detection method was evaluated by serially diluted recombinant plasmids and C. parvum genome,and the method was applied to clinical sample detection.【Result】 The RPA-CRISPR/Cas12a assay established in this study could detect C. parvum in samples within 90 min,and had no cross-reactivity with Escherichia coli,Giardia duodenalis,Clonorchis sinensis,Toxoplasma gondii,Fasciola hepatica,Pentatrimonas hominis,and Salmonella,the method could specifically detect C. parvum.The sensitivity test results showed that the minimum detection limit was 10 oocysts/mL.Using this method to detect 70 cattle feces samples and 50 human feces samples,the positive rate of C. parvum was 15.7% (11/70) and 6.0% (3/50) after reading the results by the fluorescent reporter system and lateral flow strips biosensor,which was completely consistent with the results of the nested PCR.【Conclusion】 The RPA-CRISPR/Cas12a assay for C. parvum had high specificity and sensitivity with intuitive results,without relying on expensive instrumentation,and had broad application value as a new strategy for more rapid and portable identification of C. parvum on-site and risk monitoring.

【Objective】 The aim of this study was to obtain the CDS sequence of frizzled 1 (FZD1) gene in sheep,determine its biological function,explore its polymorphism and clarify its expression differences in different breeds and tissues.【Method】 The heart,liver,spleen,lung,kidney,muscle and skin tissues of Small-tail Han sheep and Xinji Fine Wool sheep were collected and total RNA was extracted and reverse transcribed to synthesize cDNA.According to the predicted sequence of sheep FZD1 gene in NCBI database(accession No.:XM_027968435.2),the target gene sequence was obtained by TA cloning using the Small-tail Han sheep skin tissue cDNA as the template,and bioinformatics analysis was performed on the sequence to predict its possible encoded protein structure and function.Real-time quantitative PCR was used to detect the difference of FZD1 gene expression in different tissues of Small-tail Han sheep and Xinji Fine Wool sheep.Sequencing and high resolution melting analysis (HRM) were used for polymorphism detection to determine the possible single nucleotide polymorphisms(SNP) sites.【Result】 The CDS region of sheep FZD1 gene was successfully cloned,and the total length of the sequence was 1 941 bp,encoding 646 amino acids.The amino acid sequence of FZD1 of sheep had the highest similarity with Capra hircus (99.5%),and the lowest similarity with Homo sapiens (96.7%).The phylogenetic tree showed that FZD1 of sheep had the closest genetic relationship with Capra hircus and the farthest genetic relationship with Homo sapiens.The FZD1 protein contained 7 transmembrane domains,42 phosphorylation sites,and 2 N-glycosylation sites,mainly distributed on the cell membrane.The prediction results of the secondary structure of FZD1 protein showed that random coil,alpha helix,extended chain,and beta turn accounted for 48.30%,35.15%,15.02% and 1.55%,respectively,which was consistent with the predicted tertiary structure.The Real-time quantitative PCR results showed that the expression of FZD1 gene in the muscle and skin tissues of Small-tail Han sheep were significantly higher than that of Xinji Fine Wool sheep,while the expression in the heart and lung were significantly lower than that of Xinji Fine Wool sheep (P＜0.05).The expression of FZD1 gene were the highest in the lungs and skin of both breeds of sheep,and were significantly higher than other tissues (P＜0.05).The sequencing results and HRM validation showed that there were two SNPs (g.9443458 or g.9443458) at the 1 273 and 1 276 bp loci of the FZD1 gene.These two SNPs were in a fully linked imbalanced state,with only two haplotypes present,and both SNPs were moderately polymorphic.【Conclusion】 The total length of FZD1 gene CDS was 1 941 bp,encoding 646 amino acids.As seven transmembrane receptor proteins located on the cell membrane,FZD1 protein had 42 possible phosphorylation sites.It was speculated that the signaling of FZD1 was conducted through phosphorylation and dephosphorylation.The expression of FZD1 gene was the highest in the skin tissue of Small-tailed Han sheep,suggesting that FZD1 gene might be involved in the regulation of hair follicle development.

In recent years,with the improvement of people’s living standards,the demand for livestock products has increased year by year,leading to the rapid increase of the demand for animal husbandry,and the situation of grain shortage and livestock has become more and more serious.Agricultural and sideline products can be processed after refining a variety of products,such as crop corn straw,rice husk,cake dregs,dregs,fruit and vegetable residue,etc.,because of its rich nutrients,large yield,low price,is a high quality fermented feed.Therefore,the problem of improving the shortage of feed resources and the problem of unreasonable structure of feed resources is the focus of the development of animal husbandry in recent years.Fermentation can improve the nutritional value and palatability of agricultural and sideline products,reduce the content of antinutritional factors,increase the content of beneficial bacteria,promote the digestion and absorption of nutrients,thus improve the feed utilization of ruminant feed and reduce the feeding cost.Fermented agricultural and sideline products have the advantages of high nutritional value,low anti-nutritional factor content,and rich in probiotics,which have broad application prospects in ruminant production.The author summarizes the overview of agricultural and sideline products,the strains and fermentation process and the application of fermentation in ruminant production,aiming to provide reference for the development and application of fermented agricultural and sideline products.

【Objective】 The purpose of this experiment was to explore the application effect of earthworm hydrolysate in Muscovy duck healthy culture under high temperature environment.【Method】 A total of 360 healthy Muscovy ducks were randomly divided into 3 groups (6 replicates per each group,20 ducks per replicate).The Muscovy ducks in control group were fed the basal diet,and in test groups were fed with basal diet that supplemented with 1.5% and 2.5% earthworm hydrolysate,respectively.The test period was 55 days.The growth performance,heat shock protein 70 (HSP70) mRNA expression,hormone levels (triiodothyronine,tetraiodothyronine,corticosterone,cortisol) in serum,intestinal morphological parameters (villi height,crypt depth,villi width,villi height/crypt depth ratio and surface area of villi) and intestinal barrier-related parameters (diamine oxidase,D-lactate,lipopolysaccharide) of Muscovy ducks were measured.【Result】 Under high temperature environment,compared with the control group,① 1.5% and 2.5% earthworm hydrolysate boosted the average daily gain (ADG) of Muscovy ducks at 21-42 and 16-70 d (P＜0.01),decreased the feed conversation ratio(F/G) at 21-42 d (P＜0.05),and reduced the F/G at 16-70 d (P＜0.01).2.5% earthworm hydrolysate markedly decreased the F/G of Muscovy ducks at 42-70 d (P＜0.05).②1.5% and 2.5% earthworm hydrolysate decreased the mRNA level of HSP70 in Muscovy ducks at 42 and 70 days old (P＜0.01).③At 42 days old,1.5% earthworm hydrolysate increased the level of cortisol in Muscovy ducks (P＜0.01),and 2.5% earthworm hydrolysate decreased the levels of triiodothyronine (P＜0.05) and tetraiodothyronine (P＜0.01),and increased the level of corticosterone (P＜0.05) in Muscovy ducks.At 70 days old,1.5% earthworm solution decreased the level of corticosterone (P＜0.01),2.5% earthworm hydrolysate decreased the levels of triiodothyronine and cortisol (P＜0.05),and increased the level of tetraiodothyronine (P＜0.01) in Muscovy ducks.④1.5% earthworm hydrolysate increased the intestinal thickness and intraepithelial lymphocyte of Muscovy ducks (P＜0.01),2.5% earthworm hydrolysate increased the villi length and intestinal thickness of Muscovy ducks (P＜0.01).⑤At 42 days old,1.5% and 2.5% earthworm hydrolysate increased the activity of diamine oxidase in Muscovy ducks (P＜0.01).At 70 days old,1.5% earthworm hydrolysate decreased the serum level of LPS in Muscovy ducks (P＜0.01),and 2.5% earthworm hydrolysate decreased the activity of diamine oxidase,the levels of D-lactate and LPS in serum of Muscovy ducks (P＜0.01).【Conclusion】 Dietary earthworm hydrolysate could improve the growth performance of Muscovy ducks,increase the ability to resist heat stress.It could be used for healthy breeding of Muscovy ducks under high temperature environment.

【Objective】 To explore the effects of Astragalus polysaccharide (APS) on reproductive performance and antioxidant capacity of Yellow-feathered breeding hens during laying period.【Method】 A total of one hundred and ninety-two 21-week-old healthy Lingnan Yellow-feathered breeder hens were randomly divided into four treatment groups.Group one was the basal diet control group,and groups two to four were fed the basal diet supplemented with 100,200 and 300 mg/kg APS,respectively.Each treatment group contained six replicates with eight chickens per replicate.During the experiment,the daily feed intake,egg number and egg weight were repeatedly recorded,and fifty eggs in each experimental group were selected for incubation to determine the hatching performance.Twenty-four breeding eggs were randomly selected from each experimental group for egg quality determination,and two chickens were selected from each replicate for blood collection and sample collection,and index of reproductive organ and plasma antioxidant were determined.【Result】 Dietary APS had no significant effects on laying rate,average egg weight,daily egg weight,feed to egg ratio and unqualified egg rate of Yellow-feathered breeder hens (P＞0.05).Dietary APS supplementation could significantly affect yolk color,but had no significant effects on other egg quality indexes (P＞0.05),and yolk color in 200 mg/kg APS group was significantly higher than that in control group (P＜0.05).Dietary APS supplementation significantly decreased the percentage of abdominal fat and increased the ovarian index,and the percentage of abdominal fat in 200 mg/kg APS group was significantly lower than that in control group and 300 mg/kg APS group (P＜0.05),and the ovarian index in 100 mg/kg APS group was significantly higher than that in control group (P＜0.05).The fertilization rate of 100 mg/kg APS group was significantly higher than that of control group (P＜0.05).Dietary APS could significantly decrease the content of malondialdehyde (MDA) and increase the activity of glutathioneperoxidase (GSH-Px) in plasma,the content of MDA in 300 mg/kg APS group was significantly lower than that in control group (P＜0.05).The activity of GSH-Px in 200 and 300 mg/kg APS group was significantly higher than that of control group (P＜0.05).【Conclusion】 Dietary supplementation of APS could improve the hatching performance and egg quality of breeding eggs,and enhance the body’s antioxidant level of Yellow-feathered breeder hens. The suitable supplemental level of APS was 200 mg/kg for Yellow-feathered breeder hens during laying period.

【Objective】 The experiment was conducted to study the effect of puerarin on growth performance,blood biochemical indexes,antioxidant capacity and caecum microflora of broilers challenged with Escherichia coli.【Method】 216 healthy 1-day-old White-feathered broilers were randomly divided into 3 groups with 6 replicates in each group and 12 birds in each replicate (6 males and 6 females).The experiment lasted for 42 d.Broilers in control group (CON group) and Escherichia coli challenge group (EC group) were fed basal diet,and the broilers in Escherichia coli challenge + puerarin group (ECPR group) were fed the basal diet supplemented with 20 mg/kg puerarin.At the age of 10 day,broilers in EC and ECPR groups were fed 1 mL Escherichia coli K88 solution (1×10⁸ CFU/mL) per chicken,while chickens in CON group were fed with the same volume of sterilized medium.On the 21st day of the experiment,blood was collected from broiler chickens.After slaughter,different intestinal segments,immune organs,and cecal chyme were collected to measure the blood biochemical indicators,immune organ indexes,blood and intestinal antioxidant indicators,and cecal microbiota of broiler chickens.【Result】 ①Puerarin had no significant effect on the growth performance and immune organ indexes of broiler challenged with Escherichia coli (P＞0.05).②Compared with CON group,the content/activity of plasma albumin (ALB),alanine aminotransferase (ALT),alkaline phosphatase (ALP),high-density lipoprotein (HDL),and uric acid (UA) in EC group were significantly increased,but the content of plasma glucose (GLU) was significantly decreased (P＜0.05).Compared with EC group,the plasma ALP activity in ECPR group was significantly decreased,while the plasma contents of calcium (Ca) and phosphorus (P) were significantly increased (P＜0.05).③Compared with CON group,plasma hydrogen peroxide (H₂O₂) content in EC group was significantly increased,whereas the duodenal glutathione peroxidase (GSH-Px) and catalase (CAT) activities,jejunal GSH-Px,superoxide dismutase (SOD) and inducible nitric oxide synthase (iNOS) activities,and ileal GSH-Px activities were significantly decreased (P＜0.05).Compared with EC group,plasma malondialdehyde (MDA) and H₂O₂ contents in ECPR group were significantly decreased,but duodenal GSH-Px and CAT activities,jejunal GSH-Px and iNOS activities,and ileal GSH-Px activity in ECPR group were significantly increased (P＜0.05).④Compared with CON group,the content of Escherichia coli in the cecum in EC group was significantly increased (P＜0.05).【Conclusion】 Dietary supplemented with 20 mg/kg puerarin could effectively mitigate the adverse effects on broiler chickens challenged with Escherichia coli by regulating blood biochemical indexes and improving intestine antioxidant capacity.

【Objective】 Modern ecological pig farming techniques differ from traditional techniques.This study was aimed to analyze the bacterial composition of feces from pigs that underwent a transition to an ecological pig farming model,focus on changes in species diversity and community structure,in order to investigate the effects of modern ecological pig farming techniques on the gut microbiota of pigs.【Method】 Eight fattening pigs with similar body weights were selected from a traditional pig farm and transferred to an ecological pig farm for breeding.Fecal samples were collected every 10 days until the pigs reached the age of slaughter.16S rDNA high-throughput sequencing was performed on mixed fecal samples from each time period,and the sequencing data was analyzed using bioinformatics software.The microbiota was annotated based on operational taxonomic units (OTUs) of microbial 16S rDNA gene sequences in order to analyze the changes in species diversity and community structure.【Result】 The bacterial community structure of pig feces changed after being transferred from a traditional pig farm to an ecological pig farm,with an increase in diversity and richness during the breeding period.Proteobacteria,Bacteroidetes,etc.were the dominant phyla,uncultured bacteria,Moheibacter,etc.were dominant genera.The abundance of common pathogenic bacteria such as Elizabethkingia,Escherichia-Shigella,and Corynebacterium decreased to lower levels in feces before slaughter,but bacteria that helped with intestinal digestion,such as Lactobacillus,Treponema and Clostridium,increased in abundance.【Conclusion】 The bacterial community structure of the pig population changed significantly under modern ecological pig farming techniques,forming a unique dominant bacterial community that differs from that of common pig farms.

【Objective】 This study was aimed to investigate the effect of nicotinamide (NAM) on rumen fermentation status and lactate metabolism and its related enzyme activity in lactating dairy cows,and provide data reference for nicotinamide regulating rumen carbohydrate metabolism.【Method】 Forty Holstein dairy cows in good body condition,with similar lactation age (170 d±50 d),litter size (2.23±0.62),milk yield (36.17 kg±7.40 kg) and health,were randomly divided into control and experiment groups (NAM7,NAM11 and NAM15),with 10 dairy cows in each group.The cows in control group were fed the basal diet,while the cows in NAM7,NAM11 and NAM15 groups were supplemented with 7,11 and 15 g/d nicotinamide in the basic diet,respectively.The experiment period lasted for a total of 75 d,including 15 d for the pre-test period and 60 d for the trial period.Rumen fluid was collected before morning feeding on days 30 and 60 of the trial period,and the levels of volatile fatty acids such as acetic acid,propionic acid and butyric acid,and the concentrations of pyruvic acid,malic acid and succinic acid in rumen fluid were determined by high performance gas chromatography.The lactic acid,pyruvate kinase and lactate dehydrogenase were determined by spectrophotometer.【Result】 ①The rumen fermentation efficiency of dairy cows in three experiment groups were extremely significantly higher than that in control group (P＜0.01),among which,the rumen fermentation efficiency of dairy cows in NAM7 group was extremely significantly higher than that in NAM11 and NAM15 groups (P＜0.01).②The concentrations of rumen pyruvate,malate and succinate in NAM7 group were significantly higher and the lactic acid concentration in NAM7 group was significantly lower than that in control group (P＜0.05).③The NAD⁺ concentration in three experiment groups were significantly higher than that in control group (P＜0.05),and the NAD⁺ concentration in NAM7 group was extremely significantly higher than that in NAM11 and NAM15 groups (P＜0.01).The activities of rumen pyruvate kinase,malate dehydrogenase and succinate dehydrogenase in NAM7 group were significantly higher than that in control group (P＜0.05),and the NADH concentration and lactate dehydrogenase activity were significantly lower than that in control group (P＜0.05).【Conclusion】 Nicotinamide could improve rumen fermentation efficiency,the concentrations of NAD⁺,pyruvic acid,malic acid and succinic acid,and the activities of pyruvate kinase,malate dehydrogenase and succinate dehydrogenase in Holstein dairy cows,reduce the concentrations of lactic acid and NADH,and lactate dehydrogenase activity in rumen.The recommended nicotinamide addition rate was 7 g/d.

【Objective】 To study the effects of earthworm hydrolysate on growth performance,serum biochemical,antioxidant,and immune indices of laying hens.【Method】 A total of 432 healthy 12-weeks old Hy-line Pink laying hens with similar body weight were randomly divided into 4 groups with 6 replicates per group and 18 hens per replicate.Hens in control group were fed a basal diet,and hens in experimental groups were fed a basal diet supplemented with 1%,2% and 3% earthworm hydrolysate,respectively.The experiment lasted for 6 weeks.During the experiment period,daily feed intake was recorded daily and weighted at 13th and 18th weeks old.At 18th weeks old six laying hens from each replicate were randomly selected to collect serum for the determination of serum biochemical indices,antioxidant indices and immune indices.【Result】 Compared with the control group,①The differences in the growth performance indexes of laying hens of each test group were not significant (P＞0.05).②Diets supplemented with 1% and 3% earthworm hydrolysate significantly decreased the serum albumin/globulin (A/G) (P＜0.05), and significantly increased the level of total protein (TP) of laying hens (P＜0.05).Supplemented with 3% earthworm hydrolysate significantly increased the level of globulin (GLB) of laying hens (P＜0.05).③Diet supplemented with 1%,2%,and 3% earthworm hydrolysate extremely significantly increased the activities of serum total antioxidant capacity (T-AOC),superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of laying hens (P＜0.01).Supplemented with 1% earthworm hydrolysate significantly decreased the level of serum malondialdehyde (MDA) level of laying hens (P＜0.01).④Diet supplemented with 1% earthworm hydrolysate extremely increased the level of serum immunoglobulin G (IgG) (P＜0.01),and significantly increased the level of serum IgM of laying hens (P＜0.05).Supplemented with 1%,2%,and 3% earthworm hydrolysate significantly decreased the levels of serum interleukin-2 (IL-2) and IL-6 of laying hens (P＜0.01).Supplemented with 1% earthworm hydrolysate significantly decreased the levels of serum IL-17 of laying hens (P＜0.01).Supplemented with 1% and 2% earthworm hydrolysate significantly decreased the level of serum tumor necrosis factor α (TNF-α) of laying hens (P＜0.01).【Conclusion】 On the basis of without affecting growth performance,diet supplemented with earthworm hydrolysate could enhance antioxidant capacity and immune function of laying hens,and the optimum dosage was 1%.

Isoleucine,as one of the important essential amino acids for pigs,is a member of the branched chain amino acid.Isoleucine has the physiological functions of maintaining glucose homeostasis,regulating protein and lipid synthesis and catabolism.Dietary isoleucine at optimal level can promote the growth performance,improve the intestinal morphology and structure,promote the intestinal development of pigs by promoting the expression of intestinal nutrient transporters and regulating intestinal flora,improve immunity by promoting the proliferation of immune cells,increasing the expression of immune reactive substances and host defense peptides.It can also improve the carcass traits and pork quality.Under the background of high feed price and active promotion of diversified diets plan with lower protein and lower soybean meal,the application of isoleucine in pig nutrition has attracted much attention,the biosynthesis technology of L-isoleucine and balance of isoleucine and other amino acids especially branched chain amino acid in lower protein and lower soybean meal diets.In addition,the nutritional requirements of pigs for isoleucine in different physiological stages,and the ileal digestibable isoleucine parameters of common feed materials and unconventional feed materials and their dynamic prediction models are worthy of further study in future production and application.Thus,in this paper,the nutritional physiological effects of isoleucine and its nutritional requirements and application in pig production were reviewed,in order to provide reference for precise nutrition supply of pigs and cost reduction and efficiency increase in pig breeding in the future.

【Objective】 This experiment was conducted to optimize and establish the technology of utilizing forage triticale in dairy cows diets,and investigate the effect of replacing oat hay in diet with triticale silage on the production performance and economic efficiency of dairy cows.【Method】 Triticale (JS 3th) harvested at lactation stage was selected for making hay and silage,and then the conventional nutrient composition of triticale hay,triticale silage and oat hay were detected by chemical method,and their nutritional quality characteristics were analyzed and compared.45 Chinese Holstein lactating dairy cows were randomly divided into 3 groups,with 15 cows in each group.The dairy cows in control group were fed the total mixed rations (TMR),and the dairy cows in groups 1 and 2 were fed with the same dry matter content of triticale silage to replace 50% and 100% oat hay in TMR,respectively.The trial period was 63 days,with a pre-test period of 7 days and a main test period of 56 days.During this period,the milk yield was recorded daily,and milk samples were collected each week to determine milk quality.The economic benefits of dairy cows were calculated based on feed costs and milk production.【Result】 ①The contents of crude fat (EE),crude ash (Ash),acidic detergent fiber (ADF) and neutral detergent fiber (NDF) of triticale silage in dairy cows were significantly lower than those of triticale hay and oat hay (P＜0.05),and the silage fermentation indexes all met the quality silage standard.②The dry matter intake (DMI) of dairy cows in group 2 was significantly higher than that in control group (P＜0.05),the 4% fat correction milk (FCM) of dairy cows in groups 1 and 2 were significantly higher than that in control group (P＜0.05),and there were no significant difference of milk yield and feed efficiency in dairy cows among three groups (P＞0.05).③The milk fat of dairy cows in groups 1 and 2 were significantly higher than that in control group (P＜0.05),and the milk fat and milk protein of dairy cows in group 1 were significantly higher than that in control group (P＜0.05).④The feed costs in group 2 was higher than that in control group and group 1,and the milk income in group 1 was higher than that in control group and group 2.Compared with control group,the net incremental gain of dairy cows in groups 1 and 2 were higher by 2.90 and 1.93 RMB/(head·d),respectively.【Conclusion】 Triticale silage replacing oat hay could improve the production performance,milk quality and economic efficiency of dairy cows.Therefore,triticale silage could be used as a high-quality roughage to replace oat hay in diets,with the best results at 50% replacement of oat hay.

【Objective】 The purpose of this study was to investigate the effects of sodium humate and acidifier on growth performance and meat quality of Yellow feather broilers.【Method】 A total of 540 Yellow feather broilers at 42 days were randomly divided into 3 groups (6 replicates per group,30 birds per replicate).The broilers in control group were fed the basal diet and the broilers in test group were fed the basal diet added with 0.5% sodium humate and acidifier (0.5% group) and 1% sodium humate and acidifier (1% group),respectively.Among them,sodium humate and acidifier consisted of sodium humate,calcium citrate and calcium lactate in the ratio of 5∶4∶1.The test was conducted for 21 d.The fasting weight of broilers was weighed in replicates were used to calculate the growth performance of broilers,and two broilers with body weight close to the average weight were randomly selected for slaughter in each replicate group to determine the slaughter performance and organ index of broilers.Pectoral muscles on both sides were taken and used to determine muscle antioxidation and meat quality index.【Result】 ①Compared with the control and 1% groups,the final weight (FW),average daily gain (ADG),pectoral muscle rate,spleen index,total superoxide dismutase (T-SOD) activity and pH_{45 min} of pectoral muscle were significantly higher in 0.5% group of Yellow feather broilers,while the feed-to-weight ratio (F/G),abdominal fat rate,drip loss fraction and shear force of pectoral muscle were significantly lower (P＜0.05).②The concentration of malondialdehyde (MDA) in the pectoral muscle of Yellow feather broilers in the 0.5% group was significantly lower compared to the control group (P＜0.05).【Conclusion】 The combined supplementation of 0.5% sodium humate and acidifier in the diet could significantly increase the growth performance,slaughter performance, and antioxidant function of pectoral muscle of Yellow feather broilers, and improve meat quality.

【Objective】 The objective of this study was to investigate the distribution of fungiform and circumvallate papillae on the tongue of Sanhe cattle,the differences in the amount and morphology of taste buds on the two types of papillae.【Method】 Five 20-month-old and two 32-month-old Sanhe bulls from Hulun Buir,Inner Mongolia,China,were used in this study.The fungiform and circumvallate papillae on the tongue were recorded and counted.In addition,one of the tongue was divided into four regions:The torus linguae region,the lingual posterior region,the lingual middle region,and the apex linguae region.After sampling in different regions,the fungiform papillae,circumvallate papillae and their taste buds were observed using hematoxylin-eosin staining method.In total,1 233 complete serial tissue slices were made,and taste bud of 32 fungiform papillae and 4 circumvallate papillae were found by the serial tissue slices.【Result】 The average total length of the cattle tongue was 34.4 cm,the average width was 8.5 cm,and the average length of the torus linguae region of the tongue was 15.4 cm.The distribution of fungiform and circumvallate papillae on both sides of the tongues were similar.There were a large number of fungiform papillae,with an average of 155,mainly distributed in all areas from the apex linguae on the dorsal side of the tongue to the anterior part of the torus linguae,as well as the lateral and ventral sides of the apex linguae,showing a distribution pattern of dense anterior and sparse posterior.For the single fungiform papilla from the apex linguae to the torus linguae of the tongue,the number and density of taste buds gradually decreased,with an average of 5.1 taste buds.The number of circumvallate papillae was relatively small,with an average of 25,mainly distributed on both sides of the posterior part of the torus linguae,but the density of taste buds in a single circumvallate papilla was higher than that of the fungiform papilla,with an average number of taste buds of 524.8.Through slice observation,mushroom-shaped fungiform papillae protruded from the surface of the tongue,with wider circumvallate papillae than the fungiform papillae and a moat around them.【Conclusion】 The distribution characteristics of lingual papillae was basically the same on both sides and the density of taste buds in circumvallate papilla was higher than that of the fungiform papilla in Sanhe cattle. It could provide prospective insights and basic information for taste physiology research.

【Objective】 The differentially expressed genes of fat metabolism in 4- and 9-month-old Tan sheep were compared in this study,and the genes affecting fat metabolism in different muscle tissues of Tan sheep were excavated.【Method】 Longissimus dorsi muscle and biceps femoris muscle of 4- and 9-month-old Tan sheep were collected,the transcriptomic sequencing was performed by Illumian software,and the fat and fatty acid contents of longissimus dorsi muscle and biceps femoris muscle were analyzed.【Result】 Compared with 4-month-old Tan sheep,the fat content in biceps femoris muscle of 9-month-old Tan sheep was significantly decreased (P＜0.01),the fat content in longissimus dorsi muscle was significantly increased (P＜0.01),the unsaturated fatty acid contents such as oleic acid were increased,and the saturated fatty acid contents such as myristic acid and palmitic acid were decreased.Transcriptome sequencing results showed that a total of 893 differentially expressed genes (435 up-regulated and 458 down-regulated) were screened in biceps femoris muscle,and 809 differentially expressed genes (544 up-regulated and 265 down-regulated) were screened in longissimus dorsi muscle.GO function enrichment analysis showed that the differentially expressed genes in biceps femoris muscle of Tan sheep were significantly enriched to 327 entries,which were related to fat deposition,such as TCA cycle,respiratory chain complex,hydrogen ion transmembrane transporter activity and so on;28 terms were significantly enriched in longissimus dorsi muscle,including organic acid catabolic metabolism,fatty acid metabolism,extracellular space and so on.KEGG pathway annotation results showed that the differentially expressed genes in biceps femoris muscle of Tan sheep were significantly enriched to 25 pathways,9 of which were involved in fat metabolism,such as TCA cycle,pyruvate metabolism and so on;8 pathways were significantly enriched in longissimus dorsi muscle,2 of which were involved in fat deposition,namely,degradation of valine,leucine and isoleucine and glutathione metabolism.【Conclusion】 By screening the genes related to fat metabolism pathways,MDH2 and PPARGC1A genes in biceps femoris muscle and GSTK1 and MCCC2 genes in longissimus dorsi muscle might be related to fat metabolism in muscles of 9-month-old Tan sheep. The results provided a theoretical reference for further research on fat deposition in muscle tissue of Tan sheep at different growth stages.

Through long-term natural and artificial selection,sheep have developed a rich and diverse variety of breeds that provide humans with meat,milk,wool,fur and other living items.Selection is accompanied by the generation of selection signals,such as population differentiation,decreased genomic diversity and the presence of high-frequency alleles in regions with long-range extended haplotype purity in selected regions.For different types of selection signals,corresponding analysis methods have been derived,such as population differentiation-based analysis,genomic heterozygosity-based analysis,haplotype and linkage disequilibrium-based analysis,and allele frequency profile-based analysis.In recent years,with the development of high-throughput sequencing technology,researchers have been able to mine a large number of functional genes associated with economic traits by performing selection signal analysis in sheep,providing more genetic information for sheep molecular breeding.The author introduces the classification and further presents the advances in utilizing these methods to screen sheep functional genes,the prospects of application and research are predicated on selection signal analysis,which provides valuable information for further applied research of selection signal analysis in sheep genetic breeding.

【Objective】 The objective of this study was to understand the growth and development patterns differences between Holstein and Jersey cattle.【Method】 The study measured four body measurements,including withers height,hip height,body length,and chest girth of 96 Holstein and 225 Jersey cattle from two herds in Shanxi and Hebei provinces.Four nonlinear growth models,Logistic,Gompertz,Bertalanffy and Brody models,were used to fit growth curves for four body measurements,and the estimated values of the growth parameters A,B,and K were obtained from different growth models.The model with the largest coefficient of determination (R²) was selected as the best fitting model for each body measurements trait of the three test groups,Holstein cattle from Shanxi,Jersey cattle from Shanxi and Jersey cattle from Hebei.【Result】 The best fit model for the growth curves of body height (R²≥0.939) and neck height (R²≥0.886) of Holstein and Jersey cattle was Brody model.For body length (R²≥0.942),the Bertalanffy model and the Brody model were selected as the best fit models for Holstein and Jersey cattle,respectively.The superiority of the Brody,Bertalanffy,and Logistic models was identified for growth curve modeling of chest girth (R²≥0.930) for Holstein cattle from Shanxi,Jersey cattle from Shanxi and Jersey cattle from Hebei,respectively.At the same age,the estimated values of each body measurement for Holstein cattle from Shanxi were higher than those of Jersey cattle from Shanxi,with the differences between two breeds gradually increasing with age.The estimated value for mature withers height was 155.8 cm for Holstein cattle and 123.9 cm for Jersey cattle,mature hip height was 161.2 cm for Holstein cattle and 133.1 cm for Jersey cattle,mature body length was 168.6 cm for Holstein cattle and 147.1 cm for Jersey cattle,and mature chest girth was 210.9 cm for Holstein cattle and 186.3 cm for Jersey cattle.【Conclusion】 Holstein and Jersey cattle had different growth patterns,which provide a better understanding of growth and development in Holstein and Jersey cattle and contribute valuable insights for feeding management.

The quality of semen determines the fertilization rate and reproductive efficiency of domestic animals,and is an important factor affecting the efficiency of domestic animals production.Sperm motility refers to the percentage of sperm moving forward in a straight line in semen,which is one of the most important evaluation indicators of domestic animals semen quality.Improving sperm motility is the key to improve semen quality.Sperm motility is a complex trait regulated by multiple genes and pathways,which is a medium to high heritability trait.Therefore,genetic improvement has become an effective way to improve sperm motility.Compared with the traditional means of genetic improvement,new breeding technologies such as genome editing and molecular marker assisted selection have the characteristics of high efficiency,strong purpose and short cycle,providing new opportunities for domestic animals breeding.At present,the number of known genes affecting sperm motility of domestic animals is limited,which limits the application of new breeding technology in genetic improvement of sperm motility.Therefore,the identification of key genes regulating the sperm motility is important for domestic animals breeding.Genes mainly regulate sperm motility from two aspects of sperm structure and sperm function,including sperm structure defects,energy metabolism,ion channels,DNA integrity,semen composition and other aspects.This paper introduces the functional genes that regulate sperm motility in both sperm structure and sperm function,and summarizes their mechanism and research progress,aiming to provide reference for molecular breeding related to sperm motility.

【Objective】 The aim of this study was to investigate the genetic variation of single nucleotide polymorphism (SNP) of hormone-sensitive lipase (LIPE) gene exon 1 in Muchuan Black-bone chickens,and analyzed the correlation with body weight and intramuscular fat (IMF) content.【Method】 The blood of 106 Muchuan Black-bone chickens was collected,and the SNP site of LIPE gene exon 1 was detected by PCR amplification and direct sequencing.SPSS 21.0 was used to analyze the SNP polymorphism of LIPE gene exon 1 and its correlation with body weight and IMF content of chest muscle in Muchuan Black-bone chickens.The effect of SNP of LIPE gene exon 1 on its amino acid was analyzed by bioinformatics online tool.Real-time quantitative PCR was used to detect the relative expression of LIPE gene in different tissues and chest muscles of different genotypes of Muchuan Black-bone chickens.【Result】 A total of 3 SNP sites were detected of LIPE gene exon 1 in Muchuan Black-bone chickens.Genotypes of AA,AC and CC were found in SNP1 (c.66 A＞C).Genotypes of AA and AG were found in SNP2 (c.440 A＞G).Genotypes of GG and GA were found in SNP3 (c.462 G＞A).Chi-square fitness test showed that SNP2 and SNP3 of LIPE gene exon 1 in Muchuan Black-bone chickens were in Hardy-Weinberg equilibrium (P＞0.05),and SNP1 deviated from Hardy-Weinberg equilibrium (P＜0.05).Association analysis showed that the IMF content in chest muscle of AG genotype with SNP2 locus was significantly higher than that of AA genotype (P＜0.05),and there was no significant difference in body weight among different genotypes with each locus (P＞0.05).Amino acid sequence alignment analysis showed that SNP2 was a non-synonymous mutation,which caused lysine to become arginine.Real-time quantitative PCR results showed that the expression level of LIPE gene was higher in adipose tissue,and the mRNA expression level of LIPE gene in chest muscle of AA genotype with SNP2 locus was extremely significantly higher than that of AG genotype (P＜0.01).【Conclusion】 There are three SNP of LIPE gene exon 1 in Muchuan Black-bone chickens,among which SNP2 locus has a significant effect on IMF content in Muchuan Black-bone chickens,and it could be used as a candidate molecular genetic marker for IMF content selection in Muchuan Black-bone chickens.

Automatic estrus detection in cows is an essential component of modern livestock management.Efficient estrus detection methods can shorten the calving interval,reduce breeding costs,and increase reproductive efficiency.Traditional manual estrus detection is time-consuming and labor-intensive,and is no longer suitable for large-scale farming.Automated systems are the inevitable trend for estrus detection in the future.However,there are no mature automated identification systems for cows in slient estrus.Heart rate and respiratory rate are basic physiological indicators of cows,and the sexual excitement of estrus cows may cause changes in their heart rate and respiratory rate.Therefore,automated monitoring of heart rate and respiratory rate of estrus cows may become a new direction for estrus detection in the future.The author reviews the types of automated estrus detection systems for cows,introduces their related technical principles and achievements,compares the detection indicators and advantages and disadvantages of different estrus detection systems,and prospects the application prospects of new indicators for estrus detection monitoring in the future,providing a reference for related research.

【Objective】 This study was aimed to explore the genetic diversity of Changbaishan Chinese bee,and provide theoretical basis for its conservation and utilization.【Method】 A total of 195 broods of Changbaishan Chinese bee were collected from 7 sampling sites in the national Mesobee (Changbaishan Chinese bee) conservation area.11 microsatellite markers were used for genotyping,and genetic parameters such as polymorphism information content (PIC),expected heterozygosity (He),observed heterozygosity (Ho),inbred coefficient (Fis),differentiation coefficient (Fst) and interpopulation gene flow (Nm) were evaluated to study the genetic diversity of each population,and construct phylogenetic tree.【Result】 A total of 103 alleles were detected among 195 Changbaishan Chinese bee.Except AC306,AG005C,A028,AP049 and AP313,the other 6 microsatellite loci showed high polymorphism.The average polymorphism information content, expected heterozygosity and observed heterozygosity were 0.5080,0.5467 and 0.4662,respectively.The average inbred coefficient of Changbaishan Chinese bee was -0.0107.The differentiation coefficient of the 7 populations ranged from 0.0217 (Changbai and Fusong) to 0.3389 (Antu and Huinan).The gene flow between populations ranged from 0.7126 to 0.9785.The phylogenetic tree showed that there were three different evolutionary branches of the Changbaishan Chinese bee.【Conclusion】 The overall genetic polymorphism of Changbaishan Chinese bee was high,and the genetic diversity of different sampling sites was different.The results had important guiding significance for the future protection and utilization of Changbaishan Chinese bee.

【Objective】 This study was aimed to understand the prevalence and molecular characteristics of Enterovirus G (EV-G) in pig farms in Yunnan,so as to provide the theoretical basis for the prevention and control of EV-G in Yunnan.【Method】 RT-PCR basing on the primers from the highly conserved region of the 5'-untranslated region (UTR) of EV-G was used for the detection of EV-G-positive samples from clinical diarrhea samples of pigs collected in Yunnan from 2013 to 2022.The farm distribution,swine age composition and the type of positive samples were analyzed to determine the infection characteristics of EV-G in Yunnan.The complete sequence of EV-G VP1 gene was amplified using nested PCR specific primers.MegAlign software was used to analyze the similarity and key amino acid mutations of VP1 between Yunnan strains and the reference strains of 20 EV-G genotypes.A genetic evolutionary tree of VP1 gene was constructed by Mega 7.0 software.Additionally,the antigenic index of VP1 protein was analyzed using the Jameson-Wolf method in Protean software.【Result】 A total positive rate of 19.42% (377/1 941) for EV-G was observed in the clinical samples collected in Yunnan from 2013-2022.Among them,the EV-G infection rates in pig populations from Dali (54.54%),Yuxi (45.45%) and Honghe (43.24%) were significantly higher than that in other regions of Yunnan.The positive infection rates in suckling piglets (25.19%) and nursery pigs (24.68%) were higher than that in other pig age groups.The detection rate of positive cloacal swab (53.58%) was higher than that of intestinal contents samples (29.71%) and fecal samples (16.71%).According to the analysis of the similarity and genetic evolution of VP1 gene of EV-G showed that 17 strains from 20 Yunnan representatives belonged to EV-G1 genotype,2 strains belonged to EV-G6 genotype,and 1 strain belonged to EV-G17 genotype.VP1 protein of 17 Yunnan isolates of EV-G1 genotype exhibited significantly enhanced antigenic activity at positions 125-150 and 170-190,where multiple hydrophilic amino acid mutations were observed (P¹²⁷S,P¹⁴³S,G¹⁴⁶R and P¹⁸⁸S).【Conclusion】 Multiple genotypes such as EV-G1,EV-G6 and EV-G17 were prevalent in pig populations in Yunnan,with the EV-G1 type being the main genotype.VP1 protein of EV-G1 genotype Yunnan isolate exhibited significant antigenic activity at 125-150 and 170-190 amino acids,which might mediate the infection and epidemic of EV-G1 genotype strains in Yunnan.

【Objective】 The aim of the experiment was to study the immune protection effect of the inactivated trivalent vaccine (La Sota strain＋BX13 strain＋DC strain) of Newcastle disease,avian influenza (H9 subtype),and Avian adenovirus disease (group Ⅰ,type 4) on epidemic strains of Avian influenza virus (AIV),and provide the data for supporting the research of the inactived trivalent vaccine.【Method】 In this study,AIV was isolated and identified in samples collected from infected chickens in Shijiazhuang-Hebei,Miyun-Beijing,Hengshui-Hebei and Nanning-Guangxi during 2019-2022.The MegAlign software was used to analyze the similarity and phylogenetic relationship among the BX13 and the isolated strains from 2019 to 2022.SPF chickens were immunized with the inactivated trivalent vaccine and the serum was collected to determine the HI titer 21 d after immunization.At the same time,a challenge test was conducted using the homologous BX13 strain and the AIV isolates in this experiment to test the protective effect of the inactivated trivalent vaccine on the H9 subtype AIV epidemic strains.【Result】 Four strains of AIV were isolated from Hebei,Beijing and Guangxi from 2019 to 2022,all of which were H9 subtype AIVs,named SJZ19,MY20,HS21 and NN22 strains, respectively.The similarity among the BX13 strain and the four isolated strains from 2019 to 2022 was relatively high,ranging from 95.3% to 96.3%.BX13 strain and 4 isolates all belonged to the branch of DK-HK-Y280-97.After being immunized with the inactivated trivalent vaccine,the HI antibody titers of the H9 subtype AIV in chickens of each immunization group significantly increased,reaching 9.2-9.9 log2,while the control group showed negative antibodies.After being challenged with homologous BX13 strain,100% (5/5) of chickens in control group were positive,while 100% (10/10) of chickens in immune group were negative,with a protection rate of 100%.After being challenged with four H9 subtype AIV isolates,chickens in control group were 100% (5/5) positive,while chickens in immune group were 90% (9/10) to 100% (10/10) negative,with a protection rate of 90% to 100%.【Conclusion】 The inactivated trivalent vaccine prepared by BX13 strain had good immune protection effect on the current epidemic virus strains.

【Objective】 The purpose of this study was to identify the bacterial pathogens and its biological characteristics causing the death of calves in a large-scale cattle farm in Kashi,Xinjiang,so as to provide reference for the treatment of bacterial-associated diseases and rational drug for the area.【Method】 The pathological dissection and pathogen isolation on dead calves were conducted,single colony from liver (KS-1) and anal swab (KS-2) were selected for purification and culture.The isolates were identified by morphological observation,biochemical test,16S rRNA gene amplification together with phylogenetic group.The pathogenicity and drug resistance of the isolates were evaluated by virulence gene detection,pathogenicity test and drug sensitivity test.【Result】 After inoculation,the lung,liver,heart blood,joint fluid and mesenteric lymph nodes of dead calves showed a consistent colony growth pattern,as did anal swab samples from healthy calves.The isolated strains were identified as Escherichia coli(E.coli) by the growth morphology of the colony on selective medium,the morphological features of Gram staining microscopy,the biochemical properties and the verification of E.coli 16S rRNA.Phylogenetic analysis showed that KS-1 belonged to group A and KS-2 to group B1.KS-1 contained 8 virulence-related genes (crl,papC,hlyE,eaeA,ompA,ompC,iucD and iutA),and KS-2 contained 3 virulence-related genes (crl,hlyE and ompF).The pathogenicity test results showed that following intraperitoneal injection of 1×10⁸ CFU of the isolate,all mice in KS-1 group died within 16 h,however no mice in KS-2 group died.The results of drug sensitivity tests showed that KS-1 was sensitive to 17 drugs,including piperacillin/tazobactam,cefazolin,cefuroxime,cefuroxime,etc.,it was moderately sensitive to ampicillin/sulbactam and piperacillin,and resistant to ampicillin and cotrimoxazole.KS-2 was sensitive to piperacillin/tazobactam,cefotetan,imipenem,meropenem,amikacin and nitrofurantoin,moderately sensitive to tobramycin,and resistant to the other 14 drugs.【Conclusion】 In this study,two calf-derived E.coli strains were isolated,of which KS-1 was more virulent and carried abundant virulence genes,it was sensitive to most commonly used antibiotics,and was resistant to ampicillin and sulfamethoxazole,while KS-2 carried few virulence genes and showed multi-drug resistance.

【Objective】 This experiment aimed to produce a novel cecropin A and lysozyme fusion peptide (CecA-Lyz) using Pichia pastoris and to explore its antibacterial activity.【Method】 The gene encoding the fusion peptide CecA-Lyz was optimized according to the codon preference of Pichia pastoris and the physicochemical properties of CecA-Lyz was analyzed.The optimized CecA-Lyz gene was inserted into the plasmid pPIC9K-HSA to construct the recombinant plasmid pPIC9K-HSA-CecA-Lyz,which contained a 6×His tag and HSA DⅠ&DⅡ region at N-terminal.The recombinant plasmid was linearized using PmeⅠ and electrotransformed into the Pichia pastoris GS115 competent cell.The recombinant strain of high-level expressed CecA-Lyz protein was selected in YPD+G418 plate.Moreover,the optimum induction conditions of recombinant CecA-Lyz protein,including the methanol concentration and induction time were explored,and the antibacterial activity of recombinant CecA-Lyz was verified.【Result】 After codon optimization,the GC content of the CecA-Lyz gene was 48.9%,and the codon adaptation index (CAI) was 0.87,which was suitable for secretory expression in Pichia pastoris.The expected molecular weight of CecA-Lyz protein was 20.8 ku,with a theoretical isoelectric point of 9.62 and relatively stable physicochemical properties.Moreover,the optimized CecA-Lyz gene was successfully contructed into expression plamid and produced recombinant CecA-Lyz fusion protein in Pichia pastoris.The optimal methanol concentration for the induction of recombinant CecA-Lyz protein was 2.5% and the optimal induction time was 72 h.In addition,the recombinant fusion peptide without N-terminal HSA DⅠ&DⅡ was obtained by TEV digestion.The recombinant CecA-Lyz was initially shown to have antibacterial activity against Escherichia coli by plate counting and Oxford cup methods,including inhibition of the growth of drug-resistant Escherichia coli isolated from duck farms,with the inhibitory diameter up to 1.45 cm,but no significant inhibition against Salmonella and Staphylococcus.【Conclusion】 The recombinant CecA-Lyz fusion peptide could be stably produced in Pichia pastoris to obtain the fusion peptide without HSA tag and with the effect of inhibiting growth of Escherichia coli,which provided research ideas and references for further development of antimicrobial peptides.

【Objective】 In order to further understand the relationship between mammary microecological changes and the etiology of mastitis,the diversity and composition of microflora in milk of healthy cows,cows with subclinical mastitis and cows with clinical mastitis were compared in this study.【Method】 Firstly,PCR was used to detect the main mastitis-related pathogens in the milk of healthy cows,subclinical mastitis and clinical mastitis.Furthermore,16S rRNA high-throughput sequencing technology was applied to analyze the microbial diversity,microbiota composition and predictive function of milk from healthy cows,cows with subclinical mastitis and clinical mastitis,respectively.【Result】 Compared with the milk of healthy cows,the composition of milk microbiota in cows with subclinical mastitis and clinical mastitis had significant changes at phylum and genus levels;The abundances of Staphylococcus,Rolstonia,Brevibacterium,Ligilactobacillus and Blautia were significantly increased (P＜0.05) or extremely significantly increased (P＜0.01),and Rolstonia and Ralstonia pickettii were significantly enriched (P＜0.05) in the milk of subclinical mastitis;The relative abundances of Lactobacillus,Staphylococcus and Limosilactobacillus were significantly increased (P＜0.05) in milk of cows with clinical mastitis,and Bacteroidetes,Bacillus,Staphylococcus and Staphylococcus caprae were significantly enriched(P＜0.05).The abundances of Bacteroides,Treponema and Streptococcus in milk of cows with clinical mastitis were significantly higher than that of milk with subclinical mastitis (P＜0.05).Pseudomonas and Azospira were biomarkers of healthy milk, Rolstonia and Ralstonia pickettii were biomarkers of subclinical mastitis milk,and Staphylococcus and Staphylococcus caprae were biomarkers of clinical mastitis milk.Compared with the milk of healthy cows,the polysaccharide biosynthesis and metabolic functions of microorganisms in milk of subclinical mastitis were extremely significantly down-regulated (P＜0.01),while the carbohydrate metabolism,folding classification and degradation,replication and repair,energy metabolism,cofactor and vitamin metabolism,translation,nucleotide metabolism,cell growth and death of microorganisms in milk of clinical mastitis were extremely significantly up-regulated (P＜0.05).The functions of cell movement,metabolism,signal transduction,neurodegenerative diseases,nervous system and other functions were extremely significantly down-regulated (P＜0.01),and the functions of signal molecules and interactions were significantly down-regulated (P＜0.05).【Conclusion】 The alteration of microflora diversity in the milk of cows with subclinical mastitis and clinical mastitis were initially characterized in this study.The results showed that the diversity,species composition and function of milk microflora of cows with subclinical mastitis and clinical mastitis were significantly changed.This might be closely related to the cause of cow mastitis and the degree of mammary inflammation.

Avian pathogenic Escherichia coli (APEC) is an important pathogen that causes local and systemic infections in birds of different ages,and is also an important reservoir or source of virulence genes for human extraintestinal pathogenic Escherichia coli(ExPEC).With the increasing number of APEC-resistant strains,there is an urgent need to develop effective ways to prevent and control APEC infection,and preventing bacterial infection at the initial stage is one of the most effective measures to combat infection.In addition to mediating bacterial binding to host tissues,adhesins are the first step in the successful establishment of an initial infection.Therefore,using adhesin as a target to prepare a vaccine to prevent APEC infection will be a very promising strategy.The authors focus on the structure and functions of various adhesins,including fimbrial adhesins,afimbrial adhesins (Afa) and atypical adhesins.It also discusses their potential as candidate vaccine adhesins for APEC,some of the adhesins mentioned include type 1 fimbriae,P fimbriae,common fimbriae,Yqi fimbriae,Stg fimbriae,curli fimbriae,temperature-sensitive hemagglutinin (Tsh),AatA,FdeC,YeeJ,flagellin,lipopolysaccharide (LPS), outer membrane protein A,etc.Once successfully developed,the adhesin-based vaccines is expected to become a powerful means of preventing and controlling APEC infection.

【Objective】 This study was aimed to investigate the intestinal Coronavirus infection in dogs and cats in Beijing,and to explore the correlation between canine and feline Coronavirus infection and age,season,etc.,in order to provide reference for the prevention and treatment of canine and feline Coronavirus disease.【Method】 Specific primers were designed according to the conserved regions of each pathogen.PCR or RT-PCR were used to detect canine and feline Coronavirus from 402 canine and feline anal swabs (192 canine and 210 feline samples),and to detect the mixed infection of canine and feline Parvovirus and Canine astrovirus (CaAstV).Chi-square test was used to analyze the correlation between Canine coronavirus (CCoV) and Feline coronavirus (FCoV) infection and season.Regression statistics were used to analyze the relationship between CCoV and FCoV infection and age.According to the clinical sample information,the immunization situation of dogs and cats in the five districts of Beijing was analyzed.【Result】 In 192 canine samples,the positive rate of CCoV was 44.27% (85/192),and the mixed infection rate of CCoV and Canine parvorius (CPV) was 34.12% (29/85), no CaAstV positive samples were detected.In 192 feline samples,the positive rate of FCoV was 45.71% (96/210),and the mixed infection rate of FCoV and Feline panleukopenia virus (FPV) was 60.42% (58/96).Among 192 canine samples,the positive rate of CCoV was 19.39% (19/98) in healthy dogs and 70.21% (66/94) in diseased dogs.Among 210 feline samples,the positive rate of FCoV was 24.41% (31/127) in healthy cats and 78.31% (65/83) in sick cats.The highest detection rate of CCoV in winter was 42.35% (36/85).The detection rate of FCoV in four seasons was similar,and the detection rate in winter was relatively high (29.17% (28/85)).In different age groups,the positive rate of CCoV was 28.6%-87.8%,and the positive rate of CCoV was 87.8% in dogs aged 0 to 3 months,which was the main group of CCoV incidence.The positive rate of FCoV was 36.4% to 53.8% in different age groups,and there was no significant difference among different age groups (P>0.05).【Conclusion】 The infection rate of intestinal coronavirus in dogs and cats in Beijing was relatively higher,and it was more prevalent in winter.There was no significant difference in the detection rates of CCoV and FCoV among dogs and cats of different age groups.The coinfection of Coronavirus and Parvovirus was more common in clinical practice.

Atypical porcine pestivirus (APPV) is a new Pestivirus that has emerged in recent years and has caused significant losses to the global pig industry.APPV is considered to be the causative agent of congenital A-Ⅱ tremor in piglets,which can lead to generalized muscle tremors that interfere with feeding and ultimately death by starvation.APPV was first discovered in the United States in 2015 and has been mainly concentrated on Americas,Western Europe and East Asia.It has now been detected in several provinces and cities in China.Several methods have been developed to detect APPV infection such as PCR and serological techniques.Although some progress have been made in the study of APPV,the understanding of its origin,epidemiology and transmission dynamics is still very limited.The difficulty of isolating and culturing APPV and the lack of suitable alternative experimental animal models have limited the development of commercial vaccines,making prevention and control challenging.The author reviewed recent national and international advances in APPV evolution and typing,genome structure,pathogen isolation and culture,tissue tropism and transmission routes,pathogen detection techniques and vaccine development,with a view to fully understanding its protein function and molecular mechanism,in order to provide theoretical basis for researchers to develop effective diagnostic tools and successful control strategies.

【Objective】 The purpose of this experiment was to identify the circulating strain of Swine influenza virus (SIV) and its molecular biological characteristics in Guizhou province,and provide important reference for the etiological research and epidemiological investigation of swine influenza.【Method】 70 nasal swabs of pigs were collected from some farms in Guizhou province,identified by RT-PCR and inoculated with canine kidney epithelial cells (MDCK)for virus isolation and identification.The whole genome sequence of the strain was obtained by cloning and sequencing.The genetic recombination and key amino acid molecular characteristics of the strain were analyzed by bioinformatics online software.【Result】 The strain of H1N1 subtype SIV was isolated and named A/swine/Guizhou/ZY/2022 (H1N1).Whole genome sequencing and genetic evolution analysis showed that this strain belonged to G4 type Eurasian avian like H1N1 SIV.Bioinformatics online prediction showed that HA and NA genes of the isolated strain belonged to Eurasian avian like H1N1 branch,NS gene belonged to North American recombinant branch,and the rest of the gene fragments belonged to the 2009 pandemic H1N1 branch.The cleavage site of HA protein was PSIQSR↓GL,there were 4 potential glycosylation sites,and the receptor binding sites and antigenic sites were highly consistent with Eurasian avian like H1N1 SIV.The NA protein was structurally intact with 5 potential glycosylation sites.There were Q⁵⁹¹R and R²⁵¹K mutations in PB2 protein,N³⁷⁵S,L⁴⁷³V and S³¹N mutations in PB1 protein and M2 protein,respectively.【Conclusion】 A 3-source rearranged H1N1 subtype SIV strain was isolated from Guizhou province,which belonged to G4 type Eurasian avian like H1N1 virus,and its binding ability to human sialic acid α2,6-GAL receptor might be enhanced,suggesting to strengthen the epidemiological surveillance of swine influenza.

【Objective】 This experiment was aimed to investigate the transport characteristics of lekethromycin across Caco-2 cell monolayer model,and to provide further experimental evidence for further understanding of the absorption process of lekethromycin in vivo.【Method】 A Caco-2 cell monolayer model was established to verify the compactness of the Caco-2 cell monolayer model and to screen the model by measuring the transmembrane resistance (TEER) value and the apparent permeability Papp_(AP-BL) of the low permeability drug fluorescein sodium permeability test.The transport characteristics of lekethromycin from the apical side (AP) to the basolateral side (BL) and from the basolateral side to the apical side in the Caco-2 cell monolayer model were investigated,and the effects of three factors,drug concentration,pH and efflux transporter (P-gp,MRP2 and BCRP) inhibitors on the transportation of lekethromycin in Caco-2 monolayer model were analyzed.The drug concentrations were measured by UPLC-MS/MS,and its apparent permeability and efflux ratio were calculated.【Result】 The resistance value test and sodium fluorescein leakage test for 20 days showed that with the increase of culture time,the transmembrane resistance value of Caco-2 cell monolayer model gradually increased and reached about 400 Ω·cm² at the 20th day.The linear relationship of fluorescein sodium was good within 0.1-6 μg/mL (R²=0.991),and the calculated fluorescein sodium Papp_(AP-BL) was (1.57×10^-7±9.92×10^-9) cm/s,which was less than 0.5×10^-6 cm/s specified in the test.The Caco-2 cell monolayer model was successfully established and could be used to study the transport characteristics of lekethromycin.Lekethromycin Papp_(AP-BL) values did not change appreciably over time and were low,while Papp_(BL-AP) values increased over time,reaching a maximum level of 2.0350×10^-6 cm/s at 120 min of incubation,and intestinal efflux of lekethromycin was strong.Papp_(BL-AP) values increased with increasing concentrations of lekethromycin in a concentration-dependent manner.When the pH of the solution was 8.0,the Papp_(BL-AP) value (39.3871×10^-6 cm/s) was significantly higher than that of the solutions at pH 6.5 and 7.4,and alkaline intestinal fluid accelerated lekethromycin efflux.Verapamil,probenecid,and GF120918 inhibitors all significantly inhibited the lateral transport of lekethromycin from BL to AP,with efflux ratios greater than 1.5,and efflux proteins might be involved in transport.【Conclusion】 Lekethromycin showed poor transport and low absorption rate in the in vitro Caco-2 cell monolayer model,showing low permeability compounds.With the increase of concentration and pH,efflux was enhanced,efflux transporters P-gp,MRP2 and BCRP might be involved in the transport process of lekethromycin.

【Objective】 The aim of this study was to investigate the prevalence and antimicrobial resistance characteristics of Salmonella in a certain pigeon farm in Guangdong province.【Method】 In this study,diseased pigeons with diarrhea were collected from the pigeon farm for dissection,and Salmonella was isolated and identified from the sample.Serotype identification was performed on the strains identified as Salmonella,and the sensitivity of the isolated strains to 13 antimicrobial drugs was determined using agar dilution method.Furthermore,resistance and virulence genes were tested on the isolated strains.【Result】 A total of 27 strains of Salmonella were isolated from the samples collected from the pigeon farm,with a isolation rate of 67.50% (27/40).The serotype identification results showed that all isolated strains were Salmonella Typhimurium.The drug sensitivity test results showed that the resistance rate of the isolated strains to naphthyric acid and sulfamethoxazole was 100.00%.The resistance rate to streptomycin,tetracycline,and tigecycline was 25.93%.The resistance rate to ampicillin was 11.11% and the resistance rate to flufenicol and chloramphenicol was 3.70%.37.04% of the isolated strains showed multiple drug resistance,and resistance to tigecycline was detected.The results of drug resistance gene testing showed that a total of 5 drug resistance genes could be detected in the isolated strains,among which the detection rates of SulⅡ and tet(A) genes were relatively high,with 29.63% and 25.93%,respectively.The detection rates of bla_TEM,floR and cat1 genes were relatively low,with 11.11%,3.70% and 3.70%,respectively.The results of virulence gene testing showed that the detection rate of mogA,mgtC,bcfA,spvR,spvC,stn and sefA genes was 100.00%,and the detection rate of sseL genes was 96.30%.【Conclusion】 This study successfully isolated 27 strains of Salmonella from pigeons,with severe drug resistance and detection of tigecycline resistant strains.They also carried multiple virulence genes,which might be the main cause of diarrhea in this pigeon population.The results of this study could provide reference for the prevention and control of Salmonella in pigeon farms.

【Objective】 The aim of the present study was to determine the pathogenic bacteria responsible for neurologic signs of Jinding ducks.【Method】 Twenty-two sick ducks,which displayed neurologic signs and lesions of fibrinous pericarditis,perihepatitis and airsacculitis,were randomly selected from four 24- to 48-day-old flocks of Jinding ducks from a region with intensive egg-laying duck production in Liaoning province.Brain tissues were sampled and bacterial isolation were performed by using tryptic soy agar (TSA) and MacConkey’s agar (MCA),and identified by Gram and Wright’s staining,biochemical testing,and 16S rDNA gene sequencing.【Result】 The 19 strains of bacteria isolated from the brain tissue of Jinding duck were all Gram-negative bacteria.Among them,17 (named H1-H17) failed to grow in MCA and tested negative in the biochemical assays,whereas other 2 isolates (named H18 and H19) formed red colonies in MCA,fermented glucose,lactose,and maltose,and were methyl red and indole positive.On the basis of comparative analyses of the 16S rDNA amplicon sequences,the 17 isolates represented by H1 shared 99.1%-100% identities with reference strains of RA serotypes 1 to 19.The H18 and H19 isolates were 99.2%-99.9% identical to the selected eight reference Escherichia coli strains.Phylogenetic analysis based on the amplified 16S rDNA amplicon sequences further demonstrated that the 17 isolates represented by H1 belonged to RA and that isolates H18 and H19 belonged to Escherichia coli.【Conclusion】 In the studied cases,RA was responsible for neurologic signs in Jinding ducks,and that Escherichia coli could also cause neurologic signs in Jinding ducks.

Swine Salmonella is a zoonotic pathogen that can cause acute systemic infections in young pigs,seriously hindering the development of the breeding industry.Swine Salmonella can be transmitted to humans through the food chain,posing a potential threat to public health.Antibiotics are commonly used to control swine Salmonella in production,but the widespread use of antibiotics has led to the continuous enhancement of its resistance and the expansion of its resistance spectrum,increasing the difficulty of prevention and control.With the continuous development of bacteria detection technology and in-depth research on Salmonella,low-cost,simple and specific detection methods have been developed and applied,providing technical support for the detection of Salmonella.Vaccine immunization is one of the important means to prevent swine Salmonella.Currently,the vaccines on the market are mainly live attenuated vaccines,which have the advantages of good immunogenicity and durable protection,and inactivated vaccines and genetic engineering vaccines are also under development.In addition to antibiotics for the treatment and control of Salmonella,traditional Chinese medicine preparations and micro-ecological drugs also have good inhibitory effects on Salmonella,and are considered to be good substitute products with good development and application prospects.In this paper,the current situation of drug resistance of swine Salmonella was described,and the research progress of detection methods and control technologies of swine Salmonella was summarized,in order to provide theoretical basis and new ideas for the prevention and control of swine Salmonella.

【Objective】 This study was aimed to investigate the mechanism of action of Cangzhuxiangliansan in the treatment of damp-heat diarrhea in swine based on network pharmacology and molecular docking techniques.【Method】 The active ingredients and related targets of Cangzhuxiangliansan were obtained through the Traditional Chinese Medicine Systematic Pharmacology Database (TCMSP) and literature collection.The related targets of damp-heat diarrhea in swine were obtained through GeneCards,OMIM and DisGeNET databases.The composition-target network of Cangzhuxiangliansan was mapped by Cytoscape 3.9.1 software. Protein-protein interaction (PPI) analysis of intersecting targets was carried out by STRING database,PPI network was constructed and topology analysis was performed by Cytoscape 3.9.1 software to obtain core targets.GO function and KEGG signaling pathway enrichment analysis were carried out by DAVID database.Molecular docking simulations of key active ingredients of drugs with core potential targets of action were performed using AutoDock Vina software.【Result】 A total of 149 active ingredient targets,1 246 disease targets and 50 intersecting targets were obtained after screening.The results of the PPI network analysis showed that tumor necrosis factor (TNF),tumor antigen p53 (TP53),interleukin-6 (IL6),vascular endothelial growth factor A (VEGFA) and matrix metalloproteinase-9 (MMP9) were the key targets for drug action in the disease.GO functional analysis showed that there were 129 entries in biological processe,7 entries in cell component,and 17 entries in molecular function.The results of KEGG enrichment analysis showed that the drug exerted its therapeutic effects mainly through affecting IL17 signaling pathway,Toll-like receptor signaling pathway and TNF signaling pathway.The molecular docking results showed that the binding energy of the core active ingredient to the key target protein of the disease was ＜－5.0 kJ/mol.【Conclusion】 Cangzhu xiangliansan might act on targets such as TNF,TP53,IL6 and VEGFA through components such as quercetin,wogonin,and R-tetrahydroberberine,affecting the main inflammatory signaling pathways such as TNF and IL17 in the treatment of damp-heat diarrhea in swine.

【Objective】 The aim of this experiment was to investigate the prevalence of Escherichia coli with plasmid-mediated colistin resistance gene mcr-1 in different regions and years,and to analyze its drug resistance.【Method】 In 2017,2018,2019 and 2021,anal swabs and fecal samples of chicken were randomly collected from Kaifeng and Nanyang in Henan province and Ganzhou in Jiangxi province.The bacteria were isolated and cultured,and the suspected colonies were selected for Gram staining microscopy.Susceptibility test was used to detect the sensitivity and multiple drug resistance of the isolates to 14 common antibacterial drugs.Common clinical drug resistance genes were isolated by PCR amplification.Phylogenetic analysis of mcr-1 gene positive isolates was performed by multiplex PCR.【Result】 A total of 724 strains of Escherichia coli of chicken origin were isolated,including 364 in Henan province and 360 in Jiangxi province.The resistance rate of 724 isolates to tetracycline was the highest (93.65%),followed by flufenicol (87.98%),and amicacin and colistin were more sensitive (4.83% and 18.51%,respectively).96 isolates of 724 strains carried mcr-1 gene,and all of them showed multi-drug resistance,with 96.88% resistance rates to doxycycline and flufenicol.Among the 96 isolates carrying mcr-1 gene,27 carried other clinical drug resistance genes,among which 14 carried fosA3 gene,11 carried bla_CTX-M-1G gene,13 carried bla_CTX-M-9G gene,and 3 carried bla_NDM gene.11 isolates carried both fosA3 and β-lactamase genes.Phylogenetic analysis showed that 87 isolates carrying mcr-1 gene belonged to group A,4 isolates belonged to group B1,and 5 isolates belonged to group D.【Conclusion】 There were differences in the mcr-1 gene Escherichia coli of chickens from different years and regions.mcr-1 gene positive isolates showed obvious multi-drug resistance,and some isolates carrying mcr-1 gene also carried other clinical drug resistance genes.Continuous monitoring of the prevalence of mcr-1 gene in different regions and fields was helpful to better understand the formation and spread of bacterial resistance,and then took effective prevention and control measures.

【Objective】 The objective of this study was to analyze the biological characteristics and genomic information of a phage of Salmonella Enteritidis isolated and purified from sewage.【Method】 Using Salmonella Enteritidis as host,a phage of Salmonella Enteritidis was isolated and purified from mixed samples of lake water from South China Agricultural University and sewage from a broiler farm in Guangzhou.The host spectrum,optimal multiplicity of infection,one-step growth curve,thermal stability and pH stability of phage were detected.The genome of the bacteriophage was sequenced and bioinformatics analyzed,and the bacteriostatic ability of the bacteriophage against Salmonella in milk contaminated at different temperatures was investigated.【Result】 A phage of Salmonella Enteritidis was successfully isolated and named GD01,which could form round,transparent and clearly edged plaque on a double-layer agar plate.Under electron microscopy,the head of GD01 was icosahedral symmetry,with a diameter of 62 nm and a tail length of 224.4 nm,belonged to the Caudovirales,Siphoviridae. The host spectrum was narrow,and the optimum pH was 8.0,the optimum temperature was 37 ℃,the optimal multiplicity of infection was 0.01.The one-step growth curve showed that the latency of GD01 was 10 min,the lysis period was 110 min,and the average amount of cleavage was 50 PFU/cell.Whole genome sequencing analysis results showed that the total length of GD01 genome was 60 109 bp,the GC content was 56.22%,it was double-stranded DNA,contained 50 open reading frame (ORF),and did not contain virulence genes,drug resistance genes and tRNA genes.The results of in vitro bacteriostatic test showed that phage GD01 still had good bacteriostatic effect on Salmonella after treatment at 37 ℃ for 8 h and at 4 ℃ for 72 h.【Conclusion】 The isolated bacteriophage GD01 of Salmonella Enteritidis had strong acid-base tolerance and temperature tolerance,clear bioinformatics properties,and good antibacterial effect in vitro,which had the potential and application prospect of replacing antibiotics.

【Objective】 The aim of this study was to explore the potential mechanism of Scutellaria baicalensis Georgi extract in preventing and treating Streptococcus suis type 2 (SS2) infection.【Method】 The transcriptome data of SS2-infected cells were collected by GEO database,and differentially expressed genes were screened by GEO2R online software.The protein-protein interaction (PPI) inetwork was constructed using STRING database and Cytoscape v3.8.2 software,and topological analysis was performed.The changes of core protein,biological function and signal transduction pathway after SS2 infection were screened by GO function and KEGG pathway enrichment analysis.Liquid chromatography mass spectrometry (LC-MS) was used to analyze and identify the components of Scutellaria baicalensis Georgi extract,and Swiss ADME and SwissTargetPredication databases were used to screen the active components of Scutellaria baicalensis Georgi extract and predict its target.The mechanism of anti-SS2 infection of Scutellaria baicalensis Georgi extract was further defined by combining the disease target with the active ingredient target by network pharmacology.Molecular docking and Real-time quantitative PCR were used to verify the feasibility of the network pharmacological prediction results.【Result】 1 308 differential genes of SS2 infection were identified by two sets of gene chips provided by GEO database,and 70 core genes were screened by PPI analysis.GO functional enrichment analysis showed that significant enrichment items mainly involved cytokine-mediated signaling pathways and responses to cytokine stimulation.KEGG pathway enrichment analysis involved the TNF,PI3K-Akt,JAK-STAT and other signal transduction pathways.A total of 31 active components of Scutellaria baicalensis Georgi extract,including baicalin and baicalin,etc.,which might affect core targets such as VEGFA,TNF and PPARG,involving multiple biological functions and signaling pathways such as inflammatory response,apoptosis and promotion of angiogenesis.Molecular docking results showed that baicalin and baicalin of Scutellaria baicalensis Georgi extract had good binding energy with TNF,PPARG and VEGFA.Real-time quantitative PCR results showed that the mRNA levels of CA9,PPARG,JUN,SCD,JAK3 and VEGFA genes in PAMs cells were significantly up-regulated after SS2 infection (P＜0.05),and the mRNA expression levels of these genes were significantly decreased after the intervention of Scutellaria baicalensis Georgi extract (P＜0.05).【Conclusion】 After SS2 infection,cytokine mediated biological processes,TNF and other signaling pathways changed and the active components of Scutellaria baicalensis Georgi extract might jointly participate in apoptosis regulation,inflammatory response,angiogenesis and other biological processes through AMPK,PI3K-Akt,GFR and TNF signaling pathways,thereby preventing SS2 infection.

【Objective】 This study was aimed to evaluate the protective effects of different concentrations of epigallocatechin gallate (EGCG) on cisplatin-induced acute kidney injury in rats,and to select the best concentration of EGCG to alleviate cisplatin nephrotoxicity.【Method】 Thirty Wistar rats were randomly divided into 5 groups:Normal control group (CON),cisplatin group (CIS),low dose (20 mg/kg) EGCG＋cisplatin group (LCE),medium dose (40 mg/kg) EGCG＋cisplatin group (CE) and high dose (80 mg/kg) EGCG＋cisplatin group (HCE).Rats in CON group and CIS group were perfused with normal saline (40 mg/kg) for 28 days,and CIS group was intraperitoneally injected with CIS (7 mg/kg) on the 26th day.Rats in LCE,MCE and HCE groups were given corresponding dose of EGCG for 28 days,CIS was injected intraperitoneally on the 26th day,and serum and kidney tissues were taken on the 29th day.Firstly,the changes of physiological and biochemical indexes of rats were detected during drug administration,then the pathological changes of kidney tissue of rats in each group were detected,the protective effect of EGCG on acute kidney injury induced by CIS was preliminarily evaluated,and the best effective concentration of EGCG was selected.Then the protein expression changes of apoptosis (Bax, Bcl-2 and Caspase-3) and inflammation (IL-6 and IL-10) were detected by Western blotting to further confirm the protective effect of EGCG.【Result】 Compared with CON group,the diet,drinking water and body weight of rats in CIS group decreased significantly on the 28th day,while urea nitrogen (BUN) and creatinine (CRE) increased significantly (P＜0.05).The renal tubules were obviously dilated,with protein cast,and the renal injury was serious.Compared with CIS group,the diet,water consumption and weight of rats in MCE group were increased significantly (P＜0.05),and the contents of BUN and CRE in serum were decreased significantly (P＜0.05).The renal tubular dilatation in MCE group was significantly improved,but there was no significant change in LCE and HCE groups.Therefore,40 mg/kg EGCG was selected for the follow-up experiment.Western blotting result showed that compared with CON group,the expressions of Bax,IL-6 and Caspase-3 in CIS group were increased significantly (P＜0.05),while the expressions of Bcl-2 and IL-10 decreased significantly (P＜0.05).Compared with CIS group,the expressions of Bax,IL-6 and Caspase-3 in MCE group were decreased significantly (P＜0.05),while the expressions of Bcl-2 and IL-10 were increased significantly (P＜0.05).【Conclusion】 EGCG had a protective effect on CIS-induced acute kidney injury in rats (40 mg/kg was the best),which was related to the inhibition of renal cell apoptosis and inflammation.

Zearalenone (ZEA) is a mycotoxin produced by the fungi of Fusarium genera,which widely contaminates corn,wheat,barley,and other grains.After metabolism and conversion,ZEA can be distributed throughout the body via the bloodstream,causing damage in liver,kidney,uterus,testis and other organs through estrogenic effects,oxidative damage, cell apoptosis, etc.In recent years,the contamination of ZEA in crops not only poses serious challenges to livestock farming and feed industries but also threatens human health.A deep comprehension of the metabolism and toxic mechanism in animals is particularly essential for developing new prevention strategies to mitigate the negative impacts of ZEA.The author focuses on the absorption,distribution,metabolism and molecular toxicology of ZEA to clarify its toxic mechanism in animals.

【Objective】 The purpose of this experiment was to study and determine the degree of resistance of Staphylococcus aureus to common antibiotics and traditional Chinese medicine,and to explore the influence of traditional Chinese medicine on the expression of resistance genes in Staphylococcus aureus.【Method】 The resistance of Staphylococcus aureus SA1 and SA2 to commonly used antibiotics was detected by Kirby-Bauer method.The minimum inhibitory concentrations (MIC) of 10 traditional Chinese medicines was determined,and the 4 traditional Chinese medicines with the best effect were selected.The strains were added to BHI medium containing 3% glucose at different concentrations of 4 traditional Chinese medicine (0,1/2,1/4 and 1/8 MIC),and the effects of 4 traditional Chinese medicines on the expression of some drug resistance genes in Staphylococcus aureus were detected by Real-time quantitative PCR.【Result】 The results of drug sensitivity test showed that SA1 strain was highly resistant to ampicillin,moderately resistant to ceftazidime,and sensitive to other drugs.SA2 strain had high resistance to ampicillin,penicillin,erythromycin,clindamycin,vancomycin and oxacillin,and sensitive to other drugs.The results of MIC determination of 2 strains of Staphylococcus aureus by 10 kinds of traditional Chinese medicine decoction showed that Coptis chinensis,Cortex Phellodendri chinensis,Galla chinensis and Sanguisorba officinalis L.had the best inhibitory effect on 2 strains.Real-time quantitative PCR results showed,under different liquid concentrations, Galla chinensis significantly decreased the expression of mecA and mecB genes in SA1 strain (P＜0.01). Coptis chinensis and Cortex Phellodendri chinensis significantly decreased mecB gene expression in SA1 strain (P＜0.01). Coptis chinensis and Sanguisorba officinalis L. significantly decreased mecA gene expression in SA2 strain (P＜0.01).【Conclusion】 Traditional Chinese medicine could inhibit Staphylococcus aureus and reduce the expression level of related drug resistance genes to a certain extent,among which Coptis chinensis,Galla chinensis and Sanguisorba officinalis L.have the best effect,followed by Cortex Phellodendri chinensis.The results provided a reference for the prevention and treatment of Staphylococcus aureus with traditional Chinese medicine.

【Objective】 The experiment was aimed to investigate the effect of alliin on the expression of cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE) in liver of mice with nonalcoholic fatty liver disease (NAFLD),in order to elucidate the pathway of action of alliin to improve NAFLD and provide scientific basis for clinical drug development.【Method】 Fifty male C57/6J mice were randomly divided into five groups:Normal group,model group,sodium hydrosulfide group,allicin group and allicin＋PAG group (DL-propargylglycine,PAG),with 10 mice in each group.The normal group was fed with normal feed and the other four groups were fed with high fat feed.The allicin group was given 20 mg/kg allicin by gavage.Allicin＋PAG group was given 20 mg/kg of allicin by gavage,and at the same time,15 mg/kg PAG was injected intraperitoneally.Sodium hydrosulfide group was injected with 2 mg/kg sodium hydrosulfide by intraperitoneal injection.Normal and model groups were given equal doses of distilled water by gavage,once a day,weighed once a week.After 6 weeks of prophylactic administration,mice were euthanized to observe liver morphology and pathological changes,and the serum total cholesterol (TC),triglyceride (TG),high density lipoprotein (HDL-C),and low density lipoprotein (LDL-C) were detected.The protein expression of CBS and CSE in liver of mice were detected by Western blotting.【Result】 Compared with normal group,the body weight of mice in model group was increased significantly (P＜0.05),there was no significant change in the body weight of mice in other groups (P＞0.05).Morphological observation results showed that the liver of mice in model group were yellowish,lusterless and flowery,while the liver of mice in the other groups had a smooth surface,uniform texture and a color close to that of mice in normal group.HE staining of liver pathology showed that the slices of the normal group were normal,while the model group showed obvious fat vacuoles and lipid droplet deposition,indicating that the fatty liver model was successfully established.Compared with model group,there was a significant improvement in sodium hydrosulfide group and the allicin group.Antilipidemic effect of allicin＋PAG group was weakened compared with allicin group.TG,TC and LDL-C in serum of mice were significantly higher in model group compared with normal group (P＜0.05),and TC and TG were significantly lower in all administered groups compared with model group (P＜0.05).Compared with normal group,the protein expression of CBS and CSE of mice were significantly decreased in model group (P＜0.05),and the protein expression of CBS and CSE in mice were significantly or extremely up-regulated in sodium hydrosulfide and allicin groups after the intervention (P＜0.05 or P＜0.01).There was no significant difference in the expression of CBS and CSE in the allicin ＋ PAG group compared with model group (P＞0.05).【Conclusion】 Allicin could significantly increase the expression of CBS and CSE in liver of NAFLD mice induced by high-fat diet,improve the indicators of lipids in serum,reduce the lipid deposition in mouse hepatocytes,and play a role in protecting the liver.

【Objective】 The aim of this study was to determine the toxicity and safety of Huning powder and provide scientific basis for its clinical application in the treatment of chicken respiratory disease syndrome.【Method】 In the acute toxicity test,24 mice were randomly divided into control group and Huning powder low,medium and high doses of groups.Mice in Huning powder low,medium and high dose groups were given 2.5,5 and 10 g/kg BW Huning powder once with 0.2 mL/10 g BW,respectively,while mice in control group were given equal volume of sterilized distilled water.The toxicity and death of the mice were observed for 7 days.In the maximum dose test,20 mice were randomly divided into control group and test group.Mice in test group were given 10 g/kg BW Huning powder with 0.2 mL/10 g BW for 4 times within 24 h,mice in control group were given equal volume of sterilized distilled water.On the 15th day of the trial,the body weight was recorded and the pathological changes of main organs were observed.In the subacute toxicity test,40 mice were randomly divided into control group and Huning powder low,medium and high doses of groups (2.5,5 and 10 g/kg BW),all mice were administered continuously for 28 days,at 12 h after the last administer,mice were weighed,blood was collected,dissected and the heart,liver,spleen,lung and kidney were collected,the change of body weight and organ coefficient of mice were calculated,the contents of tumor necrosis factor-α (TNF-α),interleukin-4 (IL-4) and IL-6 in serum of mice were determined and histopathological examination was performed for the heart,liver,spleen,lung and kidney.In the bone marrow micronucleus test,50 mice were randomly divided into control group,positive control group (cyclophosphamide 40 mg/kg BW) and Huning powder low,medium and high doses of groups (2.5,5 and 10 g/kg BW),mice in each group were administered 2 times (interval 24 h),at 6 h after the last administer,mice were sacrificed and bone marrow smears were made,the rate of micronucleus and PCE/NCE value were calculated.In the sperm abnormality test,25 male mice were randomly divided into control group,positive control group (cyclophosphamide 40 mg/kg BW) and Huning powder low,medium and high doses of groups (2.5,5 and 10 g/kg BW),administered once a day for 5 days,the mice were sacrificed on day 35 after first administer and sperm smears were made,the rate of sperm deformity was calculated.【Result】 In the acute toxicity test,all mice in each group were survived without any toxic symptoms,and the LD₅₀ of Huning powder was not obtained,the maximum tolerance of Huning powder in mice was 40 g/kg BW,indicating that the Huning powder was safe and had no acute toxicity.In the subacute toxicity test,compared with male mice in control group,the weight of mice in Huning powder low dose group were significantly increased on the 7th day of administration (P＜0.05),the weight of mice in Huning powder medium dose group were extremely significantly increased on the 28th day of administration (P＜0.01),and Huning powder had no significant effect on the main organ coefficients and cytokine contents of mice in each group (P＞0.05),necropsy and pathological examination showed no obvious abnormal changes,indicating that Huning powder had no subacute toxicity.In the bone marrow micronucleus test and sperm abnormality test,Huning powder did not affect the bone marrow micronucleus rate and sperm deformity rate of mice (P＞0.05),and the PCE/NCE values were all within the normal range,indicating that Huning powder had no genetic toxicity.【Conclusion】 Huning powder at the dose 10 g/kg BW and below was safe and nontoxic.

【Objective】 The aim of this study was to investigate the effects of Sonchus oleraceus L.aqueous extract on inflammation-related factors and inflammatory signaling pathways in mouse pneumonia model induced by lipopolysaccharide (LPS),to explore the anti-inflammatory effects of Sonchus oleraceus L.aqueous extract on mouse pneumonia.【Method】 Preparation of aqueous extract of Sonchus oleraceus L.,the test mice were randomly divided into blank group,model group,positive control group,and Sonchus oleraceus L.aqueous extract low,medium and high concentrations groups,mice in blank group were not treated,mice in model group were gavaged with 0.3 mL of saline,mice in positive control group were gavaged with 0.3 mL of 5 mg/kg dexamethasone,and mice in Sonchus oleraceus L.aqueous extract low,medium and high concentrations groups were gavaged with 0.3 mL of 100,200 and 400 mg/mL Sonchus oleraceus L.aqueous extract,respectively.After being administered by gavage for 1 week,mice in blank group were injected intraperitoneally with 0.3 mL of saline,and mice in the rest of groups were injected intraperitoneally with 0.3 mL of 30 mg/kg LPS to induce the establishment of a mouse pneumonia model. The levels of interleukin-6 (IL-6),IL-10 and transforming growth factor-β (TGF-β) in serum of each group were measured by ELISA,and the expression levels of inflammatory signaling pathway (HMGB1/TLR4/NF-κB) proteins in lungs of mice in each group were measured by IHC and Western blotting.【Result】 Compared with blank group,the serum levels of IL-6 in mice were significantly up-regulated (P＜0.05),the levels of IL-10 and TGF-β in mice were up-regulated,and the distribution and expression of HMGB1,TLR4 and NF-κB proteins in lung of mice were highly significantly up-regulated in model group (P＜0.01).Compared with model group,the serum levels of pro-inflammatory factor IL-6 in mice was down-regulated,anti-inflammatory factor IL-10 was up-regulated,and anti-inflammatory factor TGF-β was highly significantly up-regulated in Sonchus oleraceus L.aqueous extract-treated groups (P＜0.01).HMGB1,TLR4 and NF-κB was abundantly distributed on the epithelial cells surrounding the alveoli in lungs.Compared with model group,the distribution of HMGB1,TLR4 and NF-κB were highly significantly down-regulated in lung of mice treated with aqueous extract of Sonchus oleraceus L.(P＜0.01),and showed a dose-dependent effect,the protein expression of HMGB1 and TLR4 were exremely significantly or down-regulation (P＜0.01),and the protein expression of NF-κB was significantly or extremely significantly down-regulated (P＜0.05 or P＜0.01).【Conclusion】 The aqueous extract of Sonchus oleraceus L.could inhibit the secretion of pro-inflammatory factor IL-6 and the expression of signaling pathway HMGB1,TLR4 and NF-κB in lung of mice with pneumonia induced by intraperitoneal injection of LPS,and promote the secretion of anti-inflammatory factor IL-10 and TGF-β in serum of mice,indicating that the aqueous extract of Sonchus oleraceus L.had anti-pneumonia effect.