【Objective】 The aim of this study was to investigate the genetic basis on which the phenotypic advantage of Jinnan cattle were based and provide a reference for the subsequent utilization of germplasm resources and molecular breeding in Jinan cattle.【Method】 12 healthy purebred Jinnan cattle ear margin tissues were collected, and genomic DNA was extracted for whole genome resequencing.Compare the sequencing data and the genome data of 12 Red Angus cattle in NCBI database to the bovine reference genome (ARS-UCD1.2), and detect the SNPs loci in the population, and annotate the mutation sites using ANNOVAR software. The variant genes of Jinnan cattle and Red Angus cattle were analyzed by Venn method to screen for specific variant genes of Jinnan cattle, and conduct GO function and KEGG pathway enrichment analysis.The functional interaction of significantly enriched variant genes were analyzed by Cytoscape software.【Result】 A total of 2 621 153 222 clean reads were sequenced from the Jinnan cattle genome, with an average Q30 value of 93.5%.The base distribution was uniform and without significant bias, and the average sequencing depth of 11.19×and an average coverage rate (≥ 4×) of 92.64% were obtained.A total of 117 773 201 SNPs loci were detected.A total of 405 506 non-synonymous, stopgain/stoploss SNPs were obtained by annotating the exon region, covering 16 690 genes, and 5 289 specific variant genes were screened out in Jinnan cattle.GO and KEGG analysis showed that genes were widely enriched in mitochondrial, ribosome, organelle membrane related biological processes, as well as oxidative phosphorylation and cytochrome P450 (CYP) metabolic pathways.Mitochondrial respiratory chain protein (NDU), cytochrome C oxidase (COX), and glucuronic acid transferase (UGT) were widely enriched in the oxidative phosphorylation pathway.The CYP1A1, CYP2E1, UGT1A6, and UGT1A1 genes were enriched in multiple metabolic pathways related to cytochrome P450.The GSTM2, GSTT1, GSTT2, and GSTK1 genes were enriched in the glutathione (GST) metabolic pathway and cytochrome P450 related pathways.The sub network of mitochondrial ribosomal protein (MRP) and ribosomal protein (RP) gene families were enriched in the process of ribosomal protein formation.【Conclusion】 The cattle-specific variant genes were significantly associated with cellular energy metabolism.It was inferred that they had a potential genetic effect on the feed efficiency of Jinan cattle.The results provided direction for screening SNPs markers that significantly affect fattening traits and breeding Jinnan cattle with high feed conversion.

【Objective】 The codon usage pattern of Toll-like receptor 5 (TLR5) gene was intestigated, and the influencing factors of codon usage bias was analyzed.【Method】 The nucleotide composition and codon usage pattern of TLR5 gene complete coding (CDS) sequence in 15 different species were calculated and statistically analyzed by softwares such as CodonW, BioEdit and R, and then cluster analysis was conducted to infer species with similar biological functions of TLR5 gene based on relative synonymous codon usage (RSCU) and CDS, respectively.The possible factors responsible for the codon bias were analyzed by ENC-plot, Neutrality-plot and PR2-plot.【Result】 The GC content at the third base (GC3) of TLR5 gene CDS in 9 species was greater than 0.5, there were 14 species had the effective number of codon (ENC) values greater than 35, the AU skew (AU skew) and GC skew (GC skew) values of codons in 15 species were less than 0.The CDS of the TLR5 gene in different species had 31 high-frequency codons, of which 21 end in G or C.All of these species had four optimal codons, whose biased strongly codons were GCC, AGA, AGG and CUG.The results of RSCU hierarchical and phylogenetic clustering results were not exactly the same, but both two clustering methods suggested a similar pattern of codon usage of TLR5 gene in Sus scrofa, Capra hircus and Bubalus bubalis, with similar patterns in Gallus gallus and Anas platyrhynchos, Oncorhynchus mykiss and Carassius auratus.ENC-plot analysis found that TLR5 gene of all species were distributed far below the standard curve, and Neutrality plot showed a significant positive correlation between the average values of the first and second base (GC12) and GC3 (P<0.01), while PR2-plot showed an inconsistency usage of the third codon base.【Conclusion】 There was a bias in the codon usage pattern of TLR5 gene in different species, which was influenced by both base mutation and translational selection, of which base mutation was the main influencing factor.The findings would provide a new perspective and theoretical understanding for the subsequent biological function mining of TLR5 gene.

【Objective】 The purpose of this study was to analyze the molecular characteristics of VP2 protein of a Feline panleukopenia virus (FPV) LN3 strain from Shenyang, Liaoning province in 2022 by bioinformatics methods.【Method】 FPV colloidal gold test strips were used to detect feces of sick cats with clinical symptoms such as vomiting and diarrhea.The extracted viral DNA from the feces of the diseased cat was amplificated by PCR and sequenced.Sequences were spliced by Seqman software.The VP2 sequence of FPV LN3 strain was subjected to similarity alignment and genetic evolution analysis with FPV and Canine parvovirus (CPV) strains in NCBI.The VP2 protein of FPV LN3 strain was predicted by bioinformatics software, included physical and chemical properties, hydrophilicity and hydrophobicity, transmembrane region, glycosylation site, subcellular localization, protein secondary structure, tertiary structure, etc.【Result】 The full length of VP2 gene of FPV LN3 strain was 1 755 bp, which was composed by 584 amino acids.The FPV LN3 strain and other 15 strains registered on GenBank belong to the same major branch, with similarity ranging from 98.9% to 99.7%.FPV LN3 strain and CPV were not belong to the same branch.Bioinformatics software prediction results showed that the VP2 protein of FPV LN3 strain was hydrophilic and had no transmembrane structure.VP2 protein contained 7 potential N-glycosylation sites, 86 O-glycosylation sites and 50 phosphorylation sites.The probability of VP2 protein in cytoplasm, nucleus and mitochondria were 43.5%, 34.8% and 21.7%, respectively.The randon coil, alpha helix, extended chain and beta turn in the secondary structure of VP2 protein accounted for 61.82%, 8.90%, 24.32% and 4.97%, respectively, which was consistent with the prediction results of the tertiary structure.There were 20 antigenic epitopes of VP2 protein.【Conclusion】 VP2 was a key gene for genetic variation in FPV.The VP2 protein of FPV LN3 strain was a hydrophilic and non transmembrane stable protein, which contained 20 antigenic epitopes.The results could provide theoretical basis for further study of the function of FPV VP2 protein and the development of new vaccines.

Polylactic-co-glycolic acid (PLGA) is a polymer composed of lactic acid and glycolic acid, which has good biodegradability and biocompatibility, as a new carrier and delivery system, and is widely used in biology, medicine and other fields.PLGA has the functions of antigen display and antigen encapsulation, which can protect and encapsulate a series of substances including bioactive compounds (such as protein and peptides), nucleic acids and immunomodulatory molecules from protease-mediated mucosal surface degradation, and can also slowly release drugs or antigens to reduce the times of immunization and medication, so it has great application potential in the treatment of intestinal diseases.In addition, drugs or antigens can be coupled to the surface of PLGA particles to display antigens, which shows excellent characteristics in vaccine development and drug preparation.PLGA can also be used as an adjuvant to enhance the immune protection effect of the vaccine.The surface modification function of PLGA can be used for targeted drug delivery.Targeted delivery of therapeutic molecules to specific parts of the body can not only reduce adverse side effects, but also increase the concentration of local raw materials to improve drug efficacy.PLGA nanoparticles can be loaded with different types of drugs individually or jointly, with low immunogenicity, and they are easy to be regulated by controlled chemical synthesis.Therefore, PLGA is widely used as a non-viral gene delivery system and drug delivery platform with great application potential in biomedical field.This paper focuses on reviewing the structural characteristics and preparation methods of PLGA nanoparticles, as well as the research progress and application prospects of PLGA-based prepared nanocarrier platforms in the fields of vaccine development and drug delivery, aiming to provide a reference for the related research of PLGA nanoparticles.

【Objective】 The aim of this study was to screen the differentially expressed genes of Porcine circovirus type Ⅱ (PCV2) infected porcine alveolar macrophage cell line (3D4/2) by transcriptome sequencing, and lay a foundation for understanding the host immune cell mechanism of PCV2 infection and the development of anti-PCV2 infection drugs.【Method】 PCV2 NJ2002 strain with multiplicity of infection (MOI) equal to 1 was used to treat 3D4/2 cells, and transcriptome sequencing was performed using Illumina NovaSeq 6000 sequencing platform.DeSeq 2.0 software was used for differential expressed genes analysis.GO and KEGG function analysis and transcription factor targeting analysis were also performed for the differentially expressed genes.The genes related to immunity and apoptosis were selected for Real-time quantitative PCR verification, and the protein expression levels of intracellular phosphatidylinositol kinase (PI3K), phosphorylated intracellular phosphatidylinositol kinase (p-PI3K), protein kinase B (Akt) and phosphorylated protein kinase B (p-Akt) were detected by Western blotting.【Result】 Transcriptome sequencing results showed that compared with the control group, 713 differentially expressed genes were obtained in PCV2 infection group, of which 313 were up-regulated and 400 were down-regulated. GO function and KEGG pathway enrichment analysis showed that differentially expressed genes were related to immune response, mainly concentrated in extracellular matrix receptor interaction pathway, metabolic pathway, HIF-1 signaling pathway, viral carcinogenesis, PI3K-Akt signaling pathway.The results of Real-time quantitative PCR were consistent with those of transcriptome sequencing.Western blotting results showed that the phosphorylation levels of PI3K and Akt were significantly increased after PCV2 infected 3D4/2 cells for 24 h (P<0.05).Transcription factor targeting analysis showed that differentially expressed genes were closely related to nuclear transcription factor Y subunit β (NFYB) and ETS transcription factor (ELK4).【Conclusion】 PCV2 might affect the downstream inflammatory signaling pathway and apoptosis signaling pathway to enhance its self-replication ability through the PI3K-Akt signaling pathway.NFYB and ELK4 transcription played important roles in PCV2 infection and could be considered as targets for anti-PCV2 drugs.The results provided theoretical basis for further understanding the mechanism of PCV2 infection in host immune cells and related drug development.

【Objective】 This study was aimed to investigate the effect of compound Shuanghuanglian preparation on the phagocytosis and secretion function of macrophages, and provide scientific basis for the in-depth development of compound Shuanghuanglian and the rational selection of drugs for poultry and livestock.【Method】 Lonicera japonica, Scutellaria baicalensis, Forsythia suspense and Andrographis paniculata were dried and crushed to prepare extracts, and Shuanghuanglian oral solution (Lonicera japonica:Scutellaria baicalensis:Forsythia suspense=1:1:2), compound Shuanghuanglian oral solution (Lonicera japonica:Scutellaria baicalensis:Forsythia suspense:Andrographis paniculata=1:1:2:2), and Andrographis paniculata oral solution were prepared in the proportions of the three drugs, and adjusted to a pH of 7.0, with a concentration of 1 g/mL.Three drugs were applied to the monocyte-macrophage cells in the mice.RAW264.7 cell viability was determined using CCK-8 method to establish the safe concentration of three drugs.The maximum permissible concentration of the drugs was diluted into three concentrations using a two-fold dilution method.Lipopolysaccharide (LPS) and hydrocortisone succinate sodium (HCSS) were employed to simulate the states of inflammation and immunosuppression in the body.The three drugs were used to treat cells in the two aforementioned states, as well as normal cells.The macrophage Fc/C3b receptor activity and phagocytosis capacity were measured using YC and EA rosette method as well as neutral red phagocytosis.Additionally, the macrophage secretion capacity was evaluated via ELISA.【Result】 The safe concentration range of compound Shuanghuanglian, Shuanghuanglian and Andrographis paniculata were 0.625-2.5, 3.125-12.5 and 0.625-2.5 mg/mL, respectively.Different concentrations of compound Shuanghuanglian, Shuanghuanglian and Andrographis paniculata acted on macrophages, and the cell phagocytic ability was significantly higher than that of blank control group (P<0.05).Compound Shuanghuanglian had the ability to reduce the active phagocytosis of LPS macrophage RAW264.7 and enhance the low phagocytosis of HCSS macrophages RAW264.7.Different concentrations of compound Shuanghuanglian could decrease the Fc/C3b receptor activity of activated macrophages and enhance the Fc/C3b receptor activity of low.Different concentrations of compound Shuanghuanglian had the ability to promote macrophages to secrete nitric oxide (NO), tumor necrosis factor-α (TNF-α), γ-interferon (IFN-γ) and lysozyme (LZM), and inhibit the secretion of NO, TNF-α, IFN-γ and interleukin 6 (IL-6) by LPS macrophages.【Conclusion】 Compound Shuanghuanglian could enhance the immunity and anti-inflammatory ability of the body by activating the Fc/C3b receptor activity of macrophages, while inhibiting the secretion function of activated macrophages to reduce the damage caused by hyperimmune function.The results provided a reference basis for the in-depth development of the traditional Chinese veterinary medicine compound Shuanghuanglian.

【Objective】 This experiment was aimed to reveal the effect of Qilan compound on immune function of pregnant sows, provide reference for relieving immune suppression of pregnant sows, and provide data support for clinical application of Qilan compound.【Method】 The extract of Qilan compound was prepared by reflux heating and lyophilization, and its main active components were detected by phenol-sulfuric acid method, HPLC and UV spectrophotometry.25 Landrace binary sows (30-40 gestural age) were selected and divided 5 groups:Negative control group (group N), 0.1%, 0.2% and 0.4% Qilan compound groups (groups L, M and H) and positive control group (group A), 5 sows in each group.The experiment lasted for 30 days, and blood samples were collected on 1, 15 and 29 days.Serum biochemical indices (total protein, albumin, urea nitrogen, glucose, calcium, phosphorus and triglyceride), serum antioxidant function (total antioxidant capacity, glutathione peroxidase, superoxide dismutase and malondialdehyde), immunoglobulin (IgG, IgM and IgA), inflammatory factors (IL-6 and IL-1β) and PRRSV antibody levels were determined.【Result】 The contents of polysaccharide, epigoitrin and icariin in Qilan compound were 56.0%, 0.035% and 0.39%, respectively.On days 15 and 29 of gestation, compared with group N, the contents of serum albumin, IgM and IgA, total antioxidant capacity and glutathione peroxidase activity in group A were significantly increased (P<0.05).The activities of glutathione peroxidase and superoxide dismutase in groups L and H were significantly increased (P<0.05). The contents of malondialdehyde and IL-1β were significantly decreased in all groups (P<0.05). The S/P values in groups M and H were significantly decreased (P<0.05). Compared with group A, on day 15 of gestation, the levels of serum total protein, calcium, total antioxidant capacity and IgM and IgA contents in group L were significantly decreased (P<0.05). The activity of superoxide dismutase in group H was significantly increased, and the levels of serum total protein, calcium, triglyceride, IgM, IL-6 contents and S/P value were significantly decreased (P<0.05). On day 29 of gestation, serum total antioxidant capacity, serum calcium level and IgG, IgA contents in group L were significantly decreased (P<0.05). Serum calcium level, superoxide dismutase activity and IL-1β content in group M were significantly decreased (P<0.05).【Conclusion】 Qilan compound could significantly promote the immune function of pregnant sows, and it could be used for the prevention and control of immunosuppressive disease of pregnant sows.

【Objective】 This study was conducted to investigate the effects of Lactobacillus reuteri LR1 replacing antibiotics on the expression of miRNA in different intestinal segments of piglets, so as to screen potential miRNA for studying the mechanism of Lactobacillus reuteri LR1 on regulating intestinal function of piglets.【Method】 A total of 144 cross-bred (Duroc×Landrace×Yorkshire) weaned piglets with similar body weight at 21 days old were selected and randomly divided into 3 groups, with 8 replicates in each group and 6 piglets in each replicate.The piglets in control group were fed with the basic diet, the piglets in antibiotic and probiotic groups were fed with the basic diet supplemented with compound antibiotics (100 mg/kg quinoline ethanol+75 mg/kg aureomycin) and 5×10¹⁰ CFU Lactobacillus reuteri LR1, respectively.The experiment lasted for 42 days.At the end of the experiment, 1 piglet from each replicate was randomly selected and slaughtered, the jejunum tissue was collected for miRNA high-throughput sequencing and analysis.The expression of the selected miRNA in duodenum, jejunum, ileum and colon were detected by Real-time quantitative PCR.【Result】 Lactobacillus reuteri LR1 supplementation caused significant changes in miRNA expression, altering the sort of highly expressed miRNA in jejunum.The potential target genes of differentially expressed miRNA were mainly enriched in metabolic related pathways, PI3K-Akt, MAPK, Chemokine and Ras signaling pathways.Real-time quantitative PCR results showed that the expression of differential miRNA in jejunum among 3 groups had the same result with RNA-Seq.Moreover, the expression of up-regulated miRNA showed a relatively consistent trend between control and antibiotic groups, down-regulated miRNA showed a relatively consistent trend between control and probiotic groups, while the highly expressed miRNA in intestine showed a relatively consistent expression trend among 3 groups.【Conclusion】 Adding Lactobacillus reuteri LR1 in diet replacing antibiotics could cause changes in the expression of intestinal miRNA.The differentially expressed miRNA screened could provide a foundation for research on the regulating intestinal development and nutrient absorption metabolism by Lactobacillus reuteri LR1 in piglets.

【Objective】 Keel bone fracture (KBF) is one of the important welfare problems for health and production of laying hens.This study was aimed to assess responses of fear behavior, production performance and egg quality of laying hens in later laying period with or without KBF.【Method】 At 60-week-age, a total of 100 Hyline Brown laying hens and Lindian chickens were selected and housed individually in traditional battery cage.The experimental subjects were divided into keel fractured and unfractured laying hens by palpation and portable X-ray.The fear behavioral responses of laying hens were detected sequentially by tonic immobility (TI) tests, open field tests (OFT) and startle reflex tests (SRT).The production performance and egg quality of laying hens at 68 weeks of age were detemined.【Result】 In TI test, the elevated duration time of laying hens with KBF from two breeds were significantly higher than that of normal laying hens (P<0.05).In OFT test, the number of areas entered, steps and time spent in the outer zone of Lindian chickens and Hyline Brown laying hens with KBF were significantly lower than that of normal laying hens (P<0.05), and the time spent in the inner zone of laying hens with KBF were significantly higher than that of normal laying hens (P<0.05).The movement latency of fractured Hyline Brown laying hens were significantly higher than that of normal laying hens (P<0.05).In SRT test, the total score of startle reflex of two laying breeds with KBF was significantly lower than that of normal laying hens (P<0.05).Therefore, the responses of fear behavior of laying hens with KBF station were stronger.The laying rate of laying hens with KBF was significantly lower than that of normal laying hens in two varieties (P<0.05).The egg shape index of fractured Hyline Brown laying hens was significantly lower than that of normal laying hens (P<0.05), and there was no significant difference in other egg quality parameters between fractured and normal laying hens (P>0.05).【Conclusion】 KBF increased the responses of fear behavior, and reduced the welfare quality and production performance of laying hens at the later laying period.

【Objective】 This experiment was conducted to investigate the effects of nicotinamide on performance and apparent digestibility of nutrients in lactating dairy cows.【Method】 Forty healthy Holstein cows with similar lactation days, parity and milk yield in middle and late lactation were randomly divided into 4 groups with 10 cows in each group, which were divided into control group (CK group) and experimental groups (NAM7, NAM11 and NAM15 groups), respectively.Dairy cows in CK group were fed a basal diet, in NAM7, NAM11 and NAM15 groups were given 7, 11 and 15 g/d niacinamide solution on the basis of CK group, respectively.The experiment lasted for 75 days, including 15 days of prefeeding-period and 60 days of trial.Milk production was recorded every 15 days.At the same time, milk, feed and blood samples were collected on the 1, 30 and 60 days of the trial period to determine milk composition such as milk fat, milk protein and lactose, somatic cell count, and conventional nutrients such as dry matter, organic matter and crude protein in the feed, and acid insoluble ash content and niacinamide content in plasma and milk, respectively.At the last 3 days of the experiment, fecal samples were collected by rectal fecal extraction method to determine the apparent digestibility of nutrients such as dry matter, organic matter and crude protein.【Result】 The results showed as follows:①Compared with CK group, milk yield and 4% standard milk yield in NAM7, NAM11 and NAM15 groups were extremely significantly or significantly increased (P<0.01 or P<0.05), the somatic cell scores were extremely significantly decreased (P<0.01).The milk fat percentage in NAM7 group was significantly increassed (P<0.05), and milk fat, lactose and milk protein yields were extremely significantly increased (P<0.01).Milk fat yields were extremely significantly increased (P<0.01), lactose and milk protein yields were significantly increased (P<0.05) in NAM11 group, and milk fat and milk protein yields were significantly increased (P<0.05) in NAM15 group. ②Compared with CK group, the apparent digestibility of crude protein, crude fat, total energy, neutral detergent fiber and calcium in NAM7 and NAM11 groups were significantly increased (P<0.05), and the apparent digestibility of dry matter and phosphorus had an increasing trend (0.05<P<0.10).The apparent digestibility of crude protein and total energy in NAM15 group was significantly increased (P<0.05).The apparent digestibility of acid detergent fiber in NAM7 group was significantly higher than that in CK and NAM15 groups (P<0.05).③The plasma nicotinamide content in NAM15 group was extremely significantly higher than that in CK group (P<0.01).Milk nicotinamide content in NAM15 group was extremely significantly higher than that in CK, NAM7 and NAM11 groups (P<0.01), and that in NAM11 group was extremely significantly higher than that in CK and NAM7 groups (P<0.01).【Conclusion】 Niacinamide could increase milk yield and improve milk quality of lactating dairy cows.At the same time, the apparent digestibility of crude protein, crude fat, neutral detergent fiber, energy and calcium were improved, and the effect was the best in NAM7 group.It was suggested that the appropriate amount of niacinamide used in production was 7 g/d.

The greenhouse gas emissions of ruminants account for 80% of the total greenhouse gas emissions of livestock and poultry.Methane, as a by-product of rumen microbial metabolism, not only causes serious harmness to the environment but also reduces the feed energy utilization of ruminants.Plant extracts have become one of the research hotspots in the field of methane emission reduction due to their ability to regulate rumen microbial communities.Oregano essential oil is a plant extract extracted from oregano that contains a variety of biologically active substances.With thymol and carvacrol as the main active factors, it can improve the rumen environment, change the composition of rumen microorganisms, and regulate rumen fermentation.Biological functions have broad application prospects in the field of ruminant breeding.The methane-lowering effect of oregano essential oil has been studied on a variety of animals.The results show that oregano essential oil can reduce ruminal methane emissions by regulating rumen microorganisms and their metabolism, but the research on its mechanism of action is not in-depth enough.This article takes ruminants as the subject and reviews the mechanism of oregano essential oil to improve methane emissions by regulating the abundance and composition of bacteria, archaea and protozoa in the rumen, regulating rumen metabolism, inhibiting the carbon dioxide reduction pathway and the acetate fermentation pathway, and to provide reference for the application of oregano essential oil in feed.

【Objective】 This experiment was conducted to compare the effects of grazing on chicory grassland under forest and traditional cage system on the slaughter performance, egg quality, meat quality, lipid metabolism and immune performance of Beijing-You chickens.【Method】 The 10-week-old Beijing-You hens with similar weight were randomly divided into three groups and raised for 95 days:Grazing on chicory grassland under forest group, with grazing densities of 100 hens/677 m²(T1) and 120 hens/677 m²(T2), and the traditional cage system was used as the control group (CK), the grazing groups adopted a combination of grazing and concentrate supplementation.At the end of the experiment, slaughter performance, meat and egg qualities were measured, and blood samples collected from wing vein were used for the measurement of lipid metabolism and immune performance.【Result】 Compared with the CK group, the eviscerated yield in T1 and T2 groups were significantly increased by 21.0% and 21.6% (P<0.05), breast muscle essential amino acids, breast muscle inosine monophosphate (0.05<P<0.1), yolk color and yolk weight (P<0.05) had an increasing trend.Serum lipid metabolism indexes, such as total cholesterol and triglyceride were all decreased (0.05<P<0.1).T lymphocyte conversion rate also increased numerically (0.05<P<0.1).Compared with CK group, the egg weight, yolk color and yolk weight in T1 group was increased by 25.34%, 38.3% and 53.2% (P<0.05).Compared with T2 group, the crude ash content of breast muscle, the crude protein and total amino acid contents of thigh muscle in T1 group was increased by 4.9%, 11.8% and 7.8%, respectively (P<0.05), while the crude fat content of thigh muscle decreased by 25.9% (P<0.05).And the albumen height, yolk color and weight increased by 55.5%, 27.9% and 40.6% (P<0.05).【Conclusion】 The slaughter performance, egg quality, meat quality, lipid metabolism and immune performance of Beijing-You chickens were positively affected by grazing on chicory grassland under forest, and the improvement effect of T1 group was better.

【Objective】 The purpose of this experiment was to understand the growth performance, slaughter performance and meat quality of the lambs produced by grading crossing between multiparous Suffolk sheep and Kazakh sheep under natural grazing conditions, and evaluate the improvement effect of grading crossing of multiparous Suffolk sheep on Kazakh sheep.【Method】 The study was conducted on multiparous Suffolk sheep and its F1 and F2 generations of progressive hybridization with Kazakh sheep, and Kazakh sheep at 3 months old, which had similar physical condition and body shape.They were raised under natural grazing conditions, with a pre-trial period of 10 days and a trial period of 90 days.After the experiment, 10 lambs with an average body weight close to the group were selected.Measured the body size indicators before slaughter, and measured the slaughter performance, meat quality, nutritional composition, and blood physiological and biochemical indicators after slaughter.【Result】 ① The body height, chest depth, live weight before slaughter, carcass weight, slaughter rate, meat to bone ratio, and eye muscle area of multiparous Suffolk sheep and hybrid F1 and F2 lambs were significantly higher than those of Kazakh sheep (P<0.05).The back fat thickness and tail fat weight of hybrid F1 lambs and Kazakh sheep were significantly higher than those of the multiparous Suffolk sheep and hybrid F2 lambs (P<0.05).② The water content of the longissimus dorsi muscle in hybrid F1 lambs was significantly higher than that in the multiparous Suffolk sheep, hybrid F2 lambs and Kazakh sheep (P<0.05).The content of iron, zinc, magnesium, and crude fat in the longissimus dorsi muscle of Kazakh sheep were significantly higher than that of multiparous Suffolk sheep, hybrid F1 and F2 lambs (P<0.05).③ The white blood cell count, basophil count, neutrophil count, eosinophil count, and lymphocyte count of multiparous Suffolk sheep, hybrid F1 and F2 lambs were significantly higher than those of Kazakh sheep, while the percentage of basophils, percentage of eosinophils, total platelet count, and average platelet volume were significantly lower than those of Kazakh sheep (P<0.05).④ The alkaline phosphatase activity in the serum of multiparous Suffolk sheep, hybrid F1 and F2 lambs was significantly higher than that of Kazakh sheep, and the total protein content was significantly lower than that of Kazakh sheep (P < 0.05).The serum alanine aminotransferase activity and iron content of hybrid F1 lambs were significantly lower than those of Kazakh sheep (P<0.05).The serum albumin content of multiparous Suffolk sheep was significantly higher than that of hybrid F1 and F2 lambs, and Kazakh sheep (P<0.05).【Conclusion】 Under natural grazing conditions, the meat production performance of multiparous Suffolk sheep and their progeny of grading crossing was higher than that of Kazakh sheep.With the increase of hybridization generations, the slaughtering performance and meat quality were obviously improved.

Silage is made by green feed sealing and microorganisms fermentation in the anaerobic environment.The advantage of silaging is not only to retain the original characteristics of green feed, but also to facilitate long-term storage, and rich in nutrients such as vitamins and minerals.Silage is now widely used in the production of ruminants and is an important feedstuff.Efficient utilization of silage is conducive to the high-quality development of animal husbandry.Assess the quality of silage scientifically and quickly is the key to realize silage utilization more efficient.High-quality silage plays an important role in ruminants growth, performance and health.Based on the relevant research on silage quality assessment at home and abroad, this study includes sensory analysis, chemical analysis (wet chemical method, near-infrared method, HPLC method), biological analysis (in vivo method, nylon bag method, in vitro fermentation gas production method, rusitec method) and indicator for silage quality (RFQ, GI₂₀₀₈, CSQI, and AHQI).The advantages of each analysis method in evaluating the quality of silage and the factors that affect the evaluation results are summarized.It is suggested that when evaluating the quality of silage there should be a synergy between multiple methods.The systemic and comprehensive evaluation of silage quality from the perspective of nutrient content and digestibility of silage provides suggestions and prospects for future silage quality evaluation and silage application.

【Objective】 The aim of this study was to investigate the effect of cholesterol supplementation on estrus rate, conception rate and serum biochemical indexes of Hu sheep in southern summer of China.【Method】 108 healthy Hu sheep ewes with similar body weight (33.60 kg±1.86 kg) were selected, and 30 ewes were fed with different doses of cholesterol (0.1% and 0.5%) and detect the dynamic changes of serum cholesterol content in April (non high temperature state, average highest temperature is 20.9 ℃±3.2 ℃) to select the appropriate dose of cholesterol. Another 78 Hu sheep were randomly divided into two groups, and the experimental period was 63 days (from the first day of estrus synchronization to the 30th day of mating after natural estrus), the ewes in control group were fed with basic diet, and the ewes in experimental group were fed with basic diet supplemented with cholesterol from July to September (high temperature state, average highest temperature is 31.3 ℃±2.6 ℃), and the effects of dietary cholesterol on the estrus rate, conception rate and serum biochemical indexes of Hu sheep were studied.【Result】 No matter whether 0.1% or 0.5% cholesterol was supplemented to the basic diet under non high temperature state, the cholesterol content in serum of the two groups tended to be the same after 48 h, so 0.1% cholesterol was selected for the supplementary feeding experiment in the diet of high temperature environment.Supplemental feeding of 0.1% cholesterol on the 3rd, 30th and 63rd days could significantly increase the serum cholesterol content of Hu sheep, and the natural estrus rate (89.7%) of the experimental group were higher than those of the control group (82.1%) (P>0.05), the conception rate of the experimental group(60.0%) was significantly higher than that of the control group (46.9%)(P<0.05).The content of total protein (TP), globulin (GLB), IL-1 and triglyceride (TG) in the experimental group were significantly higher than those in the control group (P<0.05), while the content of urea nitrogen was significantly decreased (P<0.05).【Conclusion】 Adding 0.1% cholesterol to the diet of Hu sheep ewes in high temperature season could enhance its estrus rate and conception rate.

【Objective】 This study explored the protective effect of resveratrol (Res) on kidney injury in rabbits induced by aflatoxin B₁ (AFB₁).【Method】 21 rabbits aged 40 days were randomly divided into 3 groups, 7 rabbits in each group, which were control group, AFB₁ group and AFB₁+Res group.The rabbits in control group was fed with basic diet, in AFB₁ group was fed with basal diet containing 0.3 mg/kg BW AFB₁, in AFB₁+Res group was fed with basal diet containing 0.3 mg/kg BW AFB₁ and 30 mg/kg BW Res.The experiment period was 21 days.At the end of the experiment, the pathological changes of kidney tissues in rabbit were observed, and the contents of creatinine (CRE), urea nitrogen (BUN) and uric acid (UA) in serum were detected.And the glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), total antioxidant capacity (T-AOC) and malonaldehyde (MDA) contents/activeties in kidney tissues were evaluated.【Result】 Compared with the control group, the pathological changes in the kidneys of rabbits in AFB₁ group included narrowed capsular space, disordered arrangement of renal tubular epithelial cells, fuzzy and exfoliated nuclei, and glycogen accumulation in the kidneys.The contents of CRE, BUN and UA in serum of rabbits in AFB₁ group were significantly increased (P<0.05).The activities of GSH-Px, SOD and T-AOC in the kidney tissues of rabbits in AFB₁ group were significantly decreased (P<0.05), and the level of MDA was significantly increased (P<0.05).Compared with AFB₁ group, for the rabbits in Res+AFB₁ group, the renal tissue structure damage in kidney was alleviated and the accumulation of glycogen was significantly reduced, the contents of CRE, BUN and UA in serum were significantly decreased, the levels of GSH-Px, SOD and T-AOC in the kidney tissue were significantly increased (P<0.05), and the level of MDA was significantly decreased (P<0.05).【Conclusion】 Adding Res in the diet could effectively alleviate the kidney damage caused by AFB₁ via improving the antioxidant capacity to regulate the level of oxidative stress of kidney in the rabbit.

【Objective】 The purpose of this study was to elucidate the genetic structure characteristics of Jinnan cattle, and explore candidate genes related to economic traits in Jinnan cattle through selection signal detection, in order to explore the selection status during the evolutionary process.【Method】 Whole genome sequencing data of Jinnan cattle and Red Angus cattle were analyzed to identify the single nucleotide polymorphism (SNP) marker of the two populations, and analyze its position and structural characteristics in the genome.Principal component analysis (PCA) and identity by state (IBS) matrix were constructed based on SNP information.A combination of population genetic differentiation index (Fst) and nucleotide diversity ratio (θ_π) was used to screen the highly selected regions in Jinan cattle genome, and quantitative trait locus (QTL), GO function and KEGG pathway enrichment were analyzed for the selected genes.【Result】 The SNPs in Jinnan cattle population were mainly distributed in the intergene region, followed by the intron region.The results of PCA and IBS analysis showed that there was no hybridization between Jinnan cattle and Red Angus cattle, and the genetic distance between individuals in Jinnan cattle was far away.By the combined analysis of Fst and θ_π, a total of 188 potential selected regions were screened.QTL analysis showed that the selection signals of Jinnan cattle were mainly related to growth, meat quality and disease resistance traits.GO function and KEGG pathway enrichment analysis showed that 11 candidate genes related to economic traits were selected strongly in Jinnan cattle, including cathepsin 1 (CATHL1), CATHL3, cathepsin antimicrobial peptide (CAMP), CATHL4, kinase inhibitor of RAS1 (KSR1), Meis homeobox 1 (MEIS1), irregular fragment polar protein 1 (DVL1), polycomb group ring finger 1 (PCGF1), RB transcriptional repressor 1 (RB1), regenerating family member 4 (REG4) and frizzled class receptor 7 (FZD7), and SNP sites were detected in the exon region of the genes.In addition to MEIS1, PCGF1 and FZD7 genes, all other genes had non-synonymous mutation sites, and KSR1 gene had stop gain mutation, and CAMP gene had stop gain and stop loss mutation.Compared with Red Angus cattle, there were 6 unique selected genes (CATHL1, CATHL4, CAMP, MEIS1, PCGF1 and PZD7 genes) in Jinnan cattle, which were mainly related to disease resistance, growth and skeletal muscle development.【Conclusion】 This study explored the genetic structure and selection signal characteristics of Jinnan cattle from the whole genome level, and screened 6 candidate genes that might be related to the economic traits of Jinnan cattle, providing theoretical reference for the subsequent research on conservation and breeding and the molecular mechanism of characteristic trait formation in Jinnan cattle.

Natural products are derived from the components of plants, animals and microorganisms, or the metabolic components and various endogenous chemical components in organisms, mainly including plant natural products, animal natural products and microbial natural products.Natural products have a regulatory effect on animal reproductive traits.For male reproduction, natural products mainly affect spermatogenesis, sperm motility and sexual activity, in which the natural products that promote male animal reproduction include icariin, lycium barbarum polysaccharide, tribulus terrestris saponins and so on, while cannabis extract, neem, and so on have a negative impact on male reproduction.Concerning female reproduction, natural products mainly affect oogenesis, follicular development and fetal growth and development.Tea polyphenols, resveratrol, genistein, forest frog oil and other natural products can increase female reproductive capacity, while citrinin, curcumin, and other natural products can decrease female prolificacy.Hormones in natural product have the greatest influence on the reproduction of female animals, which involved in various stages from oogenesis to embryo development.During pregnancy, natural products absorbed by the mother will have certain effects on fetal development.The author mainly reviews the effects of natural products on animal reproduction, in order to provide a reference for the application of natural products in the regulation of animal reproduction.

【Objective】 In this study, betaine was used as a cryoprotectant to cryopreserve Chengde Hornless goat ear fibroblast cell lines for optimizing the cryopreservation effect, so as to preserve the genetic resources of this local breed at the cellular level.【Method】 The ear fibroblast cell line of Chengde Hornless goat was established by tissue block culture method.Vitrification of fibroblasts with betaine as a cryoprotectant, different concentrations of betaine (5%, 10%, 15%, 20% and 25%) were used as the experimental group, and different concentrations of dimethyl sulfoxide (DMSO, 10% and 20%) were used as the control group.Cryopreservation of Chengde Hornless goat ear fibroblasts was performed, the cell survival rate, cell proliferative activity and the expression of B-cell lymphoma-2(Bcl-2)/Bcl-2 related X protein (Bax) genes were measured after cell resuscitation, and the toxicity of betaine was also assessed.【Result】 Chengde Hornless goat ear fibroblast cell line established in this study had good growth trend and high cell purity.The result of bacteria, fungi and mycoplasma were negative, and the growth curve was S type, which met the requirements of establishing fibroblast cell line.When the concentration of betain was 15%, the survival rate of cells was the highest, reaching 90.86%, which was extremely significantly higher than that of control group (P<0.01).The cell proliferation activity and the expression of Bcl-2/Bax genes were significantly higher than those in control group (P<0.05), and the toxicity of betain was much lower than that of DMSO (P<0.05).【Conclusion】 In this study, betaine was successfully used as a cryoprotectant to cryopreserve the ear fibroblast cell line of Chengde Hornless goat, so as to protect the genetic resources of Chengde Hornless goat.

【Objective】 The aim of this study was to analyze the fixed effects such as birth weight, teat number, herd, parity, the year of birth and birth season on the growth traits of Yorkshire pigs, and evaluate the genetic parameters, so as to provide references for the subsequent genetic improvement of this population.【Method】 Growth assay data were collected from 1 024 French Yorkshire sows, the main growth traits included average daily gain to 100 kg (ADG), age at 100 kg live weight (AGE), backfat adjusted to 100 kg (BF), and loin muscle area adjusted to 100 kg (LMA).General linear model was used to analyze fixed effects on the growth traits in Yorkshire pigs.Variance components and heritability of the growth traits were estimated by single-trait animal model in DMU software, and used multi-trait animal model to estimate the genetic correlation and phenotypic correlation.【Result】 The population mean of ADG, AGE, BF and LMA in Yorkshire pigs were 590.26 g, 168.20 d, 9.46 mm and 33.29 cm².AGE (P=0.057) and LMA (P=0.106) complied with the normality test, ADG and BF displayed approximately normal distributions.The herd, year of birth and birth season had significant effect on each growth trait in Yorkshire pigs (P<0.05 or P<0.01), the parity and birth weight had significant effect on AGE and ADG in Yorkshire pigs (P<0.05 or P<0.01).The heritability of growth traits in Yorkshire pigs ranged from 0.277 to 0.422, which were moderate to high heritability trait.The genetic correlation coefficient of each growth trait in Yorkshire pigs ranged from-0.996 to 0.346.【Conclusion】 Herd, year of birth, birth season, parity and birth weight had a significant impact on multiple growth traits in Yorkshire pigs, and the heritability of growth traits and the genetic correlation between traits could provide some guidance for the subsequent breeding of Yorkshire pig population.

Long non-coding RNA (lncRNA) is a eukaryotic transcript that is longer than 200 nucleotides and does not have the ability to code itself.It regulates the expression of genes mainly through transcriptional level, post-transcriptional level and epigenetics, and is ubiquitous in animal and plant genomes.lncRNA is a regulatory molecule that plays a key role in the growth and development, cell differentiation and disease occurrence of animals and plants.Compared with research in the medical field, the study of lncRNA in bovine ruminants is still in its infancy, especially in the regulation of economic traits related to bovine ruminants.In recent years, the rapid development of high-throughput sequencing and microarray technology has provided more efficient and rapid methods for the identification of lncRNA.Therefore, the author reviews the research findings, feature classification, post-transcriptional horizontal regulation mechanism of lncRNA and their research on muscle growth, hair follicle development and lactation traits in bovine ruminants, which lays a theoretical foundation for further research on the regulatory mechanism of lncRNA in the growth and development of bovine ruminants.

【Objective】 This study was aimed to investigate the variation of phenotype for milking flow rate traits and the correlation among the traits to identify the key indicator of milking flow rate.【Method】 A total of 2 162 cows with 1 181 628 records of milking flow rate including milk yield per milking (MMY) and 11 milking flow rate indicators:Average flow rate (AFR), milking duration (MD), peak flow rate (PFR), proportion of low flow rate (PLFR), take off flow rate (TOFR), flow during the first 0-15 s (F0-15), flow during the first 15-30 s (F15-30), flow during the first 30-60 s (F30-60), flow during the first 60-120 s (F60-120), yield during the first 2 min (Y2M), and proportion of yield during the first 2 min (PY2M) were adopted.And the normality test, regression analysis, correlation analysis and influencing factors analysis were carried out between MMY and milking flow rate indicators.【Result】 MMY and 11 milking flow rate indicators were all normal distribution.Among them, F0-15, F15-30, F30-60, F60-120, Y2M, PFR, AFR, and MD had positive regression relationship with MMY, and the variation pattern is consistent at the level of each factor.The correlation between these traits were significant with a coefficient from -0.80 to 0.95.The correlation analysis results indicated that AFR and Y2M could be used as key indicators of milk flow rate, showing a gradual increase trend with the increase of milk MMY.There were significant differences in different years, seasons, parities, lactation stages, and milking shifts (P<0.05).【Conclusion】 AFR and Y2M could be used as key indicators for evaluating milk flow rate indicators, but in actual production management, reference values for indicators need to be set based on changes in herd structure and environment.With refined management, higher reference thresholds could be gradually set for AFR and Y2M to achieve higher milk yield, thereby increasing economic benefits.

Melanin is a kind of biological pigment, which plays an important role in antioxidant, temperature regulation and radiation protection.Although it is not necessary for biological growth and development, its existence can improve the survival and competitiveness of organisms.Animal melanin is synthesized and stored by the organelle melanosomes in melanocytes.Animal melanocytes are mainly neural crest cells that migrate and differentiate along the dorsal path.Roundabout guidance receptor 2 (Robo2) gene is a member of Robo, a family of axon guidance molecules, and is the main receptor for guiding dorsal axon guidance.By introducing the formation and synthesis pathway of animal melanin, the author briefly describes the source and function of Robo2 gene, and describes the role of Robo2 gene in the direction of neuronal growth cone movement, the expression of microphthalmia associated transcription factor (MITF) and axonal development during melanin formation, and complements the mechanism of melanin synthesis, This study provides a theoretical basis for finding the major genes affecting melanin synthesis and revealing the mechanism of melanin synthesis.However, there are still many unsolved mysteries about the specific mechanism and regulatory pathway of Robo2 gene in melanin formation, which need further research to reveal.

【Objective】 From April 2023 to May 2023, yaks in multiple areas of Lhasa city, Tibet experienced severe diarrhea, bloody stools, and even death.This study was aimed to identify the pathogenic bacteria of yak diarrhea and explore their biological characteristics, so as to provide reference for the prevention and treatment of yak diarrhea disease in the region.【Method】 Fresh fecal samples of yak diarrhea were collected for anaerobic cultivation, and preliminary identification was conducted through culture characteristics and staining microscopy.The biochemical characterization verification, multiplex PCR toxin genotyping and cpe, netB, cpb2 toxin identification were performed on suspected positive strains, the genetic evolution analysis of cpa, cpb2 and 16S rRNA genes, the growth curve of some isolated strains and animal pathogenicity tests were also conducted.The in vitro drug sensitivity of isolated strains was measured using K-B method, and the multiple drug resistance results of different isolated strains were statistically analyzed.【Result】 Eight isolated bacteria were isolated and purified from the samples, which grew vigorously in anaerobic culture medium, and exhibited typical colonies similar to Clostridium perfringens on TSC medium, 5% defibrated sheep blood plate and egg yolk agar medium.Gram staining showed that they were purple bacteria, and were preliminarily identified as Clostridium perfringens, named XZ-1 to XZ-8.The biochemical test results of the isolates were consistent with those of Clostridium perfringens.The isolates all carried cpa and cpb2 genes, which were Clostridium perfringens type A.The 16S rRNA gene analysis results showed that the similarity between the 8 isolates and the reference strains of Clostridium perfringens were 97.3% to 100%, which were in the same branch, while the isolates were in the different branches with the reference strains of Enterococcus faecalis and Escherichia coli.There was some variability between cpa and cpb2 genes. The growth curve showed that the growth patterns of the three isolates(XZ-1, XZ-4 and XZ-7) were basically consistent, with 0 to 4 h being the delayed phase, 4 to 8 h being the logarithmic phase, and 8 to 22 h being the stable phase.After 22 h, the isolates began to enter the decay phase.The results of animal pathogenicity tests showed that the isolates had strong pathogenicity to Kunming mice.The isolates were all sensitive to oxacillin, erythromycin, chloramphenicol, ofloxacin, minocycline, cefoperazone, rifampicin and roxithromycin, while had varying degrees of resistance to 22 drugs such as penicillin, piperacillin, carbendazim, medimycin and ceftriaxone.The isolates had multiple drug resistance, with up to 11 strains of XZ-7 resistant, and 75.0% of strains with more than 6 types of resistance. 【Conclusion】 This study successfully isolated 8 strains of Clostridium perfringens type A from yak with severe diarrhea and bloody stools in Lhasa.The isolates had strong pathogenicity to mice and varying degrees of resistance to drugs such as penicillin, piperacillin and carbendazim.The results provided a reference basis for the prevention and control of yak diarrhea in the region.

The major histocompatibility complex (MHC), a crucial component of the chicken immune system, is primarily responsible for presenting antigenic epitopes to specific T lymphocytes and inducing immune responses.The chicken MHC expresses an MHC class Ⅰ molecule known as BF2 and an MHC class Ⅱ molecule known as BLB2.The MHC class Ⅰ molecule binds to peptides from cytoplasmic proteins, while the MHC class Ⅱ molecule binds to peptides from intracellular vesicles and extracellular regions.The polymorphism of MHC alleles is important for the spatial structure of the molecules and the characteristics of bound peptides.Due to MHC allelic differences, different haplotypes of MHC have various peptide-binding motifs.This variance further affects the number of antigenic peptides presented by MHC molecules and the strength of the activated T cell immune response, ultimately resulting in resistance or susceptibility to the same pathogen among different chicken strains.In comparison to human MHC molecules, the compact and simple chicken MHC expresses one single class Ⅰ molecule, and this nature can determine the resistance to certain pathogens in chicken. To gain a better understanding of the role of chicken MHC molecules in the process of viral infection and the mechanisms of disease resistance, it is essential to decipher their structure and conduct functional studies.The author primarily outlines the structural characteristics of chicken MHC molecules and compares the relationship between structural differences among different haplotypes of MHC molecules and disease resistance.Finally, the identification of chicken MHC peptide-binding motifs and Avian influenza virus antigenic peptides are summarized.This work lays the foundation for a deeper comprehension of the mechanism of MHC molecules present peptides to T lymphocytes and the development of T cell epitope vaccines.

【Objective】 This experiment was aimed to construct different types of novel immune castration DNA vaccines, including KISS1, neurokinin B (NKB) single expression gene vaccines, and gonadotropin releasing hormone (GnRH), KISS1 and NKB three genes co-expression gene vaccines.After injection into animals, they could stimulate immune responses and produce corresponding antibodies, thereby achieving the goal of reducing androgens by regulating the reproductive axis.【Method】 By selecting GnRH and its upstream regulatory genes KISS1 and NKB on the hypothalamic pituitary gonadal axis as targets, and introducing a balanced lethal system instead of antibiotic screening, the recombinant plasmids was constructed.The three expression plasmids were co-expressed with the target gene through the self-cleavage function of the 2A peptide.After transfecting the recombinant plasmids into 293T cells, the cell RNA was extracted to verify its normal expression in the body.Subsequently, the successfully constructed plasmids were electrically transferred to attenuated Salmonella Choleraesuis C500 competent cells to obtain recombinant live bacterial vaccines.The constructed engineered bacteria were continuously passaged 50 times in vitro, and the stability studies were conducted on strains from the 0, 2nd, 5th, 10th, 20th, 30th, 40th and 50th generations.【Result】 Recombinant plasmids pVAX-2A/KISS1-asd, pVAX-2A/S-NKB-asd and pVAX-S/GnRH-2A/S-NKB-2A/KISS1-asd were digested to obtain bands of 498, 1 146 and 2 622 bp, respectively, consistent with the target gene.After comparing the sequencing results with the target sequence, it was found that the size and direction were consistent.The recombinant plasmids were transfected into 293T cells and subjected to RT-PCR to obtain target bands of 498, 1 146 and 1 349 bp, respectively, indicating that the target gene could be transcribed normally in eukaryotic cells.After the above recombinant plasmids were electrically transferred into attenuated Salmonella Choleraesuis C500 competent cells, the extracted plasmids were digested and the band sizes were 498, 1 146 and 2 622 bp, respectively.The sequencing results were consistent with the target sequence, indicating that the recombinant plasmids were successfully introduced into attenuated Salmonella Choleraesuis C500.The D_{600 nm} values of recombinant engineering bacteria and attenuated Salmonella Choleraesuis C500 bacteria detected every 2 h were similar, and the difference was not significant.The growth curve test results showed that during the process of 50 consecutive passages in vitro, the logarithmic growth period of both engineering bacteria and attenuated Salmonella Choleraesuis C500 bacteria were around 0-10 h, with a growth plateau period of 10-14 h.After about 16 h, entering the platform phase, the growth characteristics of the engineering bacteria were consistent with those of the attenuated Salmonella Choleraesuis C500, and did not change due to carrying plasmids.At the same time, the amplified bands of the Salmonella genus marker gene invA and virulence gene crp in each generation of bacterial fluid were 580 and 599 bp, respectively.The results showed that the recombinant engineered bacteria still possessed the characteristics of Salmonella after multiple passages, and their detoxification characteristics remained unchanged.The results of plasmid digestion in each generation were 498, 1 146 and 2 622 bp, indicating that multiple passages did not affect the stability of the plasmid, and the recombinant plasmid was able to maintain normal copy function in Salmonella Choleraesuis C500.【Conclusion】 The constructed recombinant plasmids and engineered bacterial vaccines had good stability and could be expressed normally after being introduced into the body, making them suitable for research on animal immune castration.

【Objective】 The aim of this study was to screen compounds with antagonistic Senecavirus A (SVA) activity from an FDA-approved drug library containing 127 compounds and studying their pathways of action.【Method】 An in vitro active compounds screening platform for SVA was established using luciferase recombinant Senecavirus (rSVA-NLuc) combined with Fluc high-throughput screening technology.The compounds were screened from the FDA-approved drug library for their luciferase inhibitory activity at a concentration of 10 μmol/L.The inhibitory activity was further verified by Real-time quantitative RT-PCR, and the maximum non-toxic concentration was determined by a cytotoxicity assay which was measured by the released lactate dehydrogenase (LDH) from cell.According to the four main processes of the virus infection cycle, such as adsorption, endocytosis, replication, assembly and release, different cell treatment methods and Real-time quantitative RT-PCR, indirect immunofluorescence assay(IFA), Western blotting and viral titer assay (TCID₅₀) were used to study the antagonistic mechanism of the screeded compound.【Result】 Eight candidate anti-SVA active molecules were screened from the FDA drug library containing 127 molecules, and one safe and effective compound, reboxetine mesylate, was identified by Real-time quantitative RT-PCR and cytotoxicity assay.Within 36 hours of virus infection in cells, reboxetine mesylate significantly reduced the expression of SVA VP3 protein and the SVA titer (P<0.01).The treatment of golden hamster kidney cells (BSR-T7/5) with reboxetine mesylate could reduce the adsorption and entry of SVA.Real-time quantitative RT-PCR also showed that reboxetine mesylate could inhibit the assembly stage of SVA, but it had no effect on the replication and release stage of SVA.【Conclusion】 This experiment screened a compound with low cytotoxicity and excellent SVA antagonistic effect from the FDA-approved drug library:Riboxetine mesylate.This compound fighted SVA infection by inhibiting the adsorption, entry, and assembly stages of SVA.This study provided important references for the further development of anti-SVA drugs.

【Objective】 This study was to investigate the preventive and therapeutic effects of lytic phage KM104 strain on Salmonella spp.infection in chickens with different serotypes.【Method】 In this experiment the in vitro lysis spectrum and in vivo safety of KM104 strain and the attack dose and the multiplicity of infection were determined.According to the time, frequency and concentration for use of the bacteriophage KM104 strains, the prevention and therapeutic models of Salmonella spp.infection from chickens with three different serotypes in White feather laying hens were constructed.By recording the survival number and survival rate, growth performance, clinical symptoms and pathological changes of test animals and measuring bacterial excretion of anal swab, bacterial load of spleen and organ index, the prevention and therapeutic effect of KM104 was comprehensively evaluated.【Result】 The cleavage rate of the bacteriophage KM104 in five serotypes of Salmonella spp.was 80.65% (50/62), in vitro it had good bacteriostatic effect and had no obvious pathogenic effect on chickens were found.The best treatment method:The therapeutic doses for Salmonella Pullorum, Salmonella Gallinarum and Salmonella Enteritidis were 7.96×10⁷, 1.26×10⁴ and 1.68×10⁷PFU/mL, respectively, and the survival rate of chicks was the highest (90%, 80% and 90%) by continuous feeding with high titer phage KM104 for 3 days.Compared with the untreated control group, it could effectively alleviate the clinical symptoms and organ damage of experimental chicks, and the weight gain increased significantly (P<0.05), and the bacterial excretion of anal swab, the bacterial load of spleen and organ index decreased significantly (P<0.05).【Conclusion】 Bacteriophage KM104 had broad-spectrum lytic activity against pathogenic Salmonella spp.of multiple serotypes, and had no obvious pathogenic effect on chickens were found.The method of feeding KM104 with high titer for 3 days before bacteria attack had the best preventive effect on chickens from being infected with Salmonella Pullorum, Salmonella Gallinarum and Salmonella Enteritis, and could thus be used for the development of Salmonella spp. biocontrol preparations.

Coccidiosis is a parasitic protozoan disease caused by one or more species of Eimeria parasitizing in the intestinal epithelial cells of chickens.The disease quickly leads to weight loss, malnutrition, intestinal damage, etc., and has a high economic impact on the poultry industry worldwide.The life cycle of coccidia includes a switch from asexual to sexual stages and is often restricted to a single host.Current research mainly focused on cell biology, protein expression and transport mechanisms in different life stages, host cell infection, host-parasite interactions, new anticoccidial drug screens, etc.Given the diversity of research questions and the requirement of reduce and replace animal experiments, establishing efficient in vitro culture models has become essential for coccidia biological characteristics, anti-coccidia vaccines and drugs study.In this review, the author summarized the in vitro culture model of the poultry parasite coccidia and its application in drug screening, including the chicken embryo culture model, two-dimensional culture models (primary cell culture model, immortalized cell line culture model) and 3D organoid culture model.Breakthroughs in culture techniques have opened new avenues for research on the life cycle stages of coccidia and new intervention strategies and also provide a reference for in-depth understanding of the pathogenic mechanism of coccidia and the development of the anti-coccidia drugs.

【Objective】 The aim of this study was to investigate the mixed infection of Avian leukosis virus (ALV) subgroups and the genetic variation of gp85 gene in Hubei province, and provide reference for further study on the epidemiology of mixed infection of ALV.【Method】 A dead chicken suspected of ALV infection from Yichang, Hubei province was subjected to pathological autopsy, and the diseased tissue was asepsis collected and inoculated with DF-1 cells for virus isolation.The subgroups were identified by PCR, ELISA and indirect immunofluorescence assay (IFA), and the similarity and variation of gp85 gene of isolates were further analyzed.【Result】 The autopsy results showed that the kidneys and spleen of the dead chickens were swollen, with lung bleeding, and gray white tumor nodules were visible on the surface.After the tissue homogenates were inoculated into DF-1 cells, the cell supernatant was positive for P27 antigen by ELISA, and specific green fluorescence was detected in the infected cells by IFA.The specific bands of subgroups A (692 bp), B (847 bp) and J (545 bp) could be amplified simultaneously by PCR.The results of identification showed that the natural mixed strains of ALV of subgroups A, B and J were isolated from the dead chicken, which were named as HBYC2022-A, HBYC2022-B and HBYC2022-J, respectively.HBYC2022-A shared 99.6% nucleotide identity with JS-A1201 strain in Jiangsu province.HBYC2022-B shared 99.7% similarity with JS-B1204 strain.HBYC2022-J shared 100% identity with Heilongjiang strain PK19FA01, Guangxi strain GX20YL12 J and Anhui strain AHaq02.In addition, there were no special point mutations in the amino acid sequences of the gp85 highly mutated regions (hr1 and hr2).【Conclusion】 There was mixed infection of different ALV subgroups A, B, and J in chicken flocks, suggesting that the domestic poultry farms should be vigilant and the epidemiological investigation of ALV subgroups should be strengthened.

【Objective】 This study was aimed to design multi-epitope vaccine against IpaJ protein of Salmonella Pullorum and provide a new vaccine for purifying chicken dysentery.【Method】 In this study, IEDB was used to predict the major histocompatibility complex class Ⅰ (MHC Ⅰ) molecular binding epitopes of Salmonella Pullorum IpaJ protein in T lymphocytes.NetMHCIIpan 4.0 Server was used to predict T lymphocytes MHC Ⅱ molecular binding epitopes.B lymphocyte epitope was predicted by IEDB.After the antigenicity of the selected epitopes was evaluated by VaxiJen v 2.0, the qualified epitopes were concatenated into multi-epitope vaccine by flexible linker.Antigenicity, physicochemical properties, N-glycosylation sites, secondary structure and tertiary structure of the constructed multi-epitope vaccine were predicted.Molecular docking was used to evaluate the binding ability of the multi-epitope vaccine to the immune receptor.Immune simulation was used to evaluate the immune effect of the multi-epitope vaccine.Finally, the codon was optimized for cloning expression.【Result】 After screening, the constructed multi-epitope vaccine MEV-IpaJ contained 4 MHC Ⅰ, 4 MHC Ⅱ and 8 B lymphocyte epitope dominance.The relative molecular weight of the multi-epitope vaccine MEV-IpaJ was 28.18 ku, which was a stable hydrophilic protein and had good antigenicity.There were four potential N-glycoylation sites, including α-helix, extended strand, beta turn and random coil accounted for 20.38%, 19.23%, 8.08% and 52.31%, respectively.Ramachandran mapping of tertiary structure showed that the dominant region contained 89.9% of the residual base, and the residual base of the dominant region increased to 94.0% after refinement.The mapping of tertiary structure prominent epitope also proved that the multi-epitope vaccine had good immunogenicity, and molecular docking showed that the MEV-IpaJ could dock with Toll-like receptor 2 (TLR2) and TLR4 protein molecules.The results of immune simulation showed that the MEV-IpaJ had a good immune response and could improve the expression of some cytokines.Codon optimization ensured the efficient and stable expression of the MEV-IpaJ in E.coli K12 expression system.【Conclusion】 The constructed multi-epitope vaccine MEV-IpaJ could be effectively expressed and might induce strong T cell and B cell immune responses.This study provided a new method for the design of multi-epitope vaccine of Salmonella Pullorum, and provided theoretical basis and data support for the research and development of multi-epitope vaccine of Salmonella Pullorum.

【Objective】 This study was aimed to explore the morphological and biological characteristics of Enterobacter hormaechei phage, and prevent and control Enterobacter hormaechei infections.【Method】 In this study, Enterobacter hormaechei was used as host bacteria.A strain of phage was isolated from sewage and silt by double-layer agar plate method and its biological characteristics were studied.The titer of the phage, temperature and pH stability, optimal multiplicity of infection (MOI) and one-step growth curve of phage were measured.【Result】 A strain of Enterobacter hormaechei phage was successfully isolated and identified and named H2.The phage formed uniform, round and transparent plaque on the double-layer agar plate.The results of transmission electron microscopy showed that the head of phage H2 was icosahedral symmetry, with a diameter of 85 nm±2 nm and a long tail with a tail filament length of 190 nm±2 nm, was a member of Caudovirales and the Mycobacteriophages family.The results of temperature and pH stability of phage H2 showed that the titer remained stable in the range of 35-55 ℃ and pH 5.0-9.0.The best MOI was 0.0001.One-step growth curve according to the best MOI was drew, and it was measured that incubation period was about 20 min, the outbreak period was about 50 min, and the lysis amount was about 68 PFU/cell.【Conclusion】 Phage H2 had strong cracking ability, thermal stability and acid-base stability, which provided a good material for the research and development of subsequent phage preparations.

In the intestines of vertebrates, there are a large number of structurally diverse and dynamically changing microbiota, which play important roles in the physiology, metabolism, and immunity of the intestine.Under natural conditions, these microorganisms and eukaryotes (such as worms, protozoa, fungi, etc) co-exist in the vertebrate intestine.Both parasites and microbiota can significantly alter the intestinal physiology and immune environment, creating opportunities for interaction between them.The interaction between gut microbes and parasites can greatly affect the outcome of infection, which in turn has important implications for host health.For example, parasitic infections affect host-microbial interactions to promote or protect the host from bacteria.On the other hand, the flora affects the colonization, reproduction and toxicity of the parasite, causing it to develop along the survival pattern of parasitic-reciprocal symbiosis with the host.The mechanism and results of these interactions are frontier topics in the cross-research between microbiology and parasitology.This article summarizes the latest research results on the interaction between intestinal parasites and intestinal microbiota in recent years, and puts forward its own views on factors that may not be taken into account, aiming to provide reference for the prevention and control of intestinal parasitic diseases and the study of intestinal microbiota.

【Objective】 This study was aimed to construct the chaperone prsA2 gene deletion strain and its complemental strain, and explore the roles of prsA2 gene in the anchoring of virulent factor internalin B (InlB) and pathogenicity of Listeria monocytogenes.【Method】 The deletion strain ΔprsA2 and complemental strain CΔprsA2 were constructed with shuttle vector pKSV7 and pIMK2, respectively.The growth ability of wild type 10403S, ΔprsA2 and CΔprsA2 was analyzed by growth curve.The effect of prsA2 gene deletion on the anchoring of InlB was evaluated by Western blotting.The effect of prsA2 gene deletion on the adhesion and invasion ability, migration ability between host cells and pathogenicity of Listeria monocytogenes were analyzed by intestinal epithelial cell (Caco-2) adhesion and invasion test, immunofluorescence test and mouse virulence test.【Result】 PCR amplification results showed deletion strain ΔprsA2 and its complemental strain CΔprsA2 were constructed successfully.Growth curve results showed that the growth ability of wild type 10403S, ΔprsA2 and CΔprsA2 in BHI broth exhibit no significant difference (P>0.05).The results of Western blotting showed that the InlB anchor content on the surface of the bacteria ΔprsA2 was extremely significantly lower than 10403S and CΔprsA2 (P<0.01), and the InlB anchor content in the bacterial culture supernig was extremely significantly higher than 10403S and CΔprsA2 (P<0.01).The cell test results showed that the adhesion rate and invasion rate of deletion strain ΔprsA2 on Caco-2 cells were extremely significantly lower than 10403S and CΔprsA2 (P<0.01).Deletion of prsA2 gene reduced the ability to recruit actin to form an actin tail for intercellular migration of Listeria monocytogenes.The mouse virulence test showed that the bacterial load of deletion strain ΔprsA2 in liver and spleen of mice was significantly lower than that of 10403S and CΔprsA2 (P<0.05).【Conclusion】 The deletion of chaperone prsA2 gene didn't affect the growth ability of Listeria monocytogenes in BHI broth, but the InlB anchor quantity on the surface of ΔprsA2 strain was significantly decreased, and deletion of prsA2 gene weakened the adhesion and invasion ability on Caco-2 cells, the ability of intercellular migration and the pathogenicity in mice of Listeria monocytogenes.

【Objective】 This study was aimed to identify the pathogenic bacteria that causing the disease and death of breeding sheep in a sheep farm in Qiqihar city, and to explore its etiological characteristics, so as to provide scientific basis for effective prevention and control of the disease.【Method】 The lungs from dead sheep were aseptically taken, and laboratory diagnosis was conducted based on clinical symptoms.Pathogenic bacteria were isolated and cultured.Morphological observation, biochemical test, 16S rRNA PCR amplification and sequencing, kmtⅠ gene identification, capsule serotyping, virulence gene detection, drug susceptibility test, drug resistance gene detection and mouse pathogenicity test were used to identify and analyze the pathogenic characteristics of the isolated bacteria.【Result】 The pathogenic strain was Pasteurella multocida.The isolated bacteria showed round raised colonies with smooth, grayish white surface on the blood plate, and Gram staining showed that the Gram-negative brevibacterium was clustered into lines or scattered in single. Biochemical test results showed that the strain could ferment maltose, arabinose, sucrose, xylose, glucose, fructose, trehalose, mannitol, urease, sorbitol and and ONPG, and arginine acid hydrolysis and indigo test were positive.Serotyping of the capsule showed that the isolate was Pasteurella multocida capsular serotype D. The drug sensitivity test showed that the isolates were resistant to enrofloxacin, bacitracin and lomefloxacin, intermediary to 4 drugs such as neomycin, and sensitive to 16 drugs such as ampicillin.The isolates carried 14 virulence genes and 6 resistance genes.All mice died after infection with the isolated bacteria.【Conclusion】 One Pasteurella multocida serotype D strain was successfully isolated, carrying a variety of virulence genes and drug resistance genes, and was pathogenic to mice.This study provided data for exploring the biological information and etiological characteristics of the sheep origin Pasteurella multocida.

The long-term and large-scale irregular use of antibiotics in livestock and poultry production has led to the increasingly serious problem of bacterial resistance, which has brought great difficulties to the prevention and treatment of bacterial diseases in livestock and poultry, and even threatened public health security. Phage lysin is a peptidoglycan hydrolase encoded by phage, which can kill bacteria efficiently and specifically and has anti-biofilm activity.It has become a new antibacterial agent and has been included in the treatment strategy of bacterial infection, especially drug-resistant bacterial infection.In livestock and poultry production, phage lysin has been successfully used in the prevention and treatment of bacterial diseases caused by Gram-positive bacteria such as Staphylococcus aureus and Streptococcus, while the application of phage lysin on Gram-negative bacteria such as Escherichia coli and Salmonella is still in the in vitro research stage.In this paper, the structural characteristics, mechanism of action and application of phage lysin in livestock and poultry production were reviewed, and the application status and prospect of phage lysin in livestock and poultry production were explored, in order to provide reference for further application of phage lysin in prevention and control and treatment of bacterial diseases.

【Objective】 The purpose of this experiment was to obtain high-purity Monkeypox virus (MPXV) A5L protein and prepare mouse anti-A5L highly valent antiserum, which provided materials for the function study of MPXV A5L protein and the research and development of related diagnostic reagents.【Method】 The recombinant plasmid pET-28a-A5L was constructed, and the correctly sequenced recombinant plasmid was transformed into Escherichia coli BL21 (DE3) competent cells for induction expression.The expression of recombinant protein A5L was analyzed under different IPTG induction concentration, induction temperature and induction time conditions, and the optimal expression conditions were screened.The expression of recombinant protein A5L was identified by Western blotting, and the solubility of recombinant protein was analyzed by SDS-PAGE.A5L protein was purified by His-Bind nickel column affinity chromatography and immunized mice.Antibody titers were detected by ELISA.【Result】 The results of double digestion and sequencing showed that the prokaryotic expression vector pET-28a-A5L was successfully constructed.Western blotting results showed that recombinant protein A5L was successfully expressed in Escherichia coli BL21 (DE3) competent cells.A5L protein expression was the highest at 42 ℃, IPTG concentration of 1.6 mmol/L and 16 h after induction, with a size of 40 ku.SDS-PAGE analysis showed that A5L protein mainly existed in the form of inclusion bodies, and the optimal imidazole elution concentration was 50 mmol/L.ELISA results showed that the most effective value of the prepared murine anti-A5L protein antibody was 1:205 440.【Conclusion】 The recombinant plasmid pET-28a-A5L was successfully constructed and the recombinant protein A5L was successfully expressed in the prokaryotic expression system.The prepared A5L polyclonal antibody had good immunogenicity, and the results laid a foundation for further investigation of the biological function of MPXV A5L protein.

【Objective】 The aim of this experiment was to understand the genetic and evolutionary characteristics of Duck circovirus (DuCV) in Guangxi.【Method】 A total of 761 clinical samples of ducks were collected from Guangxi during 2019-2022 to detect DuCV infection by Real-time quantitative PCR.The whole genome sequence of DuCV was amplified by PCR, the similarity of nucleotide and amino acid sequences between DuCV and reference strain were analyzed by DNAStar, the DUCV recombination event was analyzed by RDP4 and SimPlot 3.5.1 software, and the genetic evolution tree was constructed by Mega 7.0 software.The amino acid variation sites of DuCV Cap protein were analyzed by BioEdit software.【Result】 The positive rate of DuCV was 22.5%, and the genome size was between 1 988-1 996 bp.The nucleotide sequence similarity of the whole genome, Rep and Cap genes between the Guangxi DuCV strain and the reference strain was 71.9%-99.6%, 83.1%-99.9% and 52.8%-99.9%, respectively.Gene recombination analysis showed that there were recombination events in DuCV-GX13-2019 and DuCV-GX14-2020.A total of 78 amino acid sites in Cap protein were mutated.Compared with DuCV-1, DuCV-2 was completely mutated at amino acids 3, 12, 15, 23, 31, 42, 56 and 64.The strains in Guangxi were DuCV-1 and DuCV-2 genotypes, DuCV-1 was the main dominant strain, and DuCV-1b subtype was the most widely prevalent.【Conclusion】 The incidence of DuCV infection in Guangxi was serious, some strains had recombination events, the mutation rate of Cap protein was high, and various subtypes of DuCV were prevalent, which provided basic data for the epidemiological investigation of DuCV in Guangxi.

【Objective】 This study was aimed to investigate the inhibitory mechanism of Lycium barbarum quercetin on β-lactamase produced by Staphylococcus aureus, so as to provide a basic basis for the clinical development of β-lactamase inhibitors.【Method】 β-lactamase positive strains were screened by penicillin inhibition zone edge test, and the sensitivity of positive strains to penicillin was detected by drug sensitivity paper method under the action of Lycium barbarum quercetin at concentrations of 32, 64 and 128 μg/mL.The changes of penicillin content under the treatment of Lycium barbarum quercetin at concentrations of 32, 64, 128 and 256 μg/mL were detected by multifunctional microplate reader, so as to evaluate the effect of Lycium barbarum quercetin on β-lactamase synthesis and β-lactamase activity under the treatment of Lycium barbarum quercetin at concentrations of 64, 128 and 256 μg/mL.The molecular docking of Lycium barbarum quercetin and BlaZ protein was performed by AutoDockTools 1.5.6 software, and the results of molecular docking were visualized by Discovery Studio 2019 Client software.【Result】 A total of 23 strains producing β-lactamase positive strains were screened from 36 strains of Staphylococcus aureus.The results of drug sensitivity paper showed that the sensitivity of positive strains to penicillin increased with the increase of Lycium barbarum quercetin concentration.The results of multifunctional microplate reader showed that Lycium barbarum quercetin could inhibit the synthesis of β-lactamase and significantly inhibit the activity of β-lactamase(P<0.05).Molecular docking results showed that Lycium barbarum quercetin affected the binding of BlaZ protein to β-lactam antibiotics by hydrogen bonding with the conserved residue Asn161 in the Ω-loop region of BlaZ protein.【Conclusion】 Lycium barbarum quercetin could reduce the synthesis of β-lactamase, and also enhance the antibacterial effect of penicillin by reducing the activity of β-lactamase, which had potential activity as a β-lactamase inhibitor.The results provided a reference for the subsequent development of new β-lactamase inhibitors.

【Objective】 The study was aimed to decipher the mechanism of mulberry leaf (ML) and Eucommia ulmoides (EU) against infectious bursal disease (IBD) based on network pharmacology, molecular docking and molecular dynamics simulations methods, and provide new insights for development of veterinary drugs against immunosuppression and IBD.【Method】 The active ingredients and the corresponding targets of ML and EU were retrieved from the Traditional Chinese Medicine Systems Pharmacology (TCMSP) and SwissTargetPrediction database, and IBD-related targets were collected through GeneCards and Gene Expression Omnibus (GEO) database.Furthermore, the component-target network diagram was constructed using Cytoscape 3.7.2.Then, the intersectional targets between the active ingredients and disease were obtained and the protein-protein interaction (PPI) network was constructed by STRING database and Cytoscape 3.7.2.Additionally, GO function and KEGG pathway enrichment were performed based DAVID database.Finally, Molecular docking was performed using AutoDock 1.5.6 and PyMOL software and molecular dynamics simulations were performed using Gromacs 2020.6 software.【Result】 A total of 53 active ingredients of ML and EU, 423 corresponding targets, 5 949 IBD-related targets and 153 common drug-disease targets were screened for analysis.PPI showed that the key targets mainly include interleukin-6 (IL6), catenin beta-1 (CTNNB1), proto-oncogene tyrosine-protein kinase Src (SRC), IL1B, vascular endothelial growth factor A (VEGFA).The results of GO function and KEGG signaling pathway showed that ML and EU exert effects against IBD through response to IL18, interferon-gamma receptor binding, neuroactive ligand-receptor interaction and calcium signaling pathway.Finally, the molecular docking and molecular dynamics simulations results showed that core active ingredients and key targets had a good binding, and bound stably.【Conclusion】 ML and EU might played a role on IL6, CTNNB1, SRC, IL1B, VEGFA and other main targets through the key active ingredients, such as quercetin, dehydrodieugenol, icosa-11, 14, 17-trienoic acid methyl ester, kaempferol and beta-sitosterol, and exert effects against infectious bursal disease through response to IL18, calcium signaling pathway and neuroactive ligand-receptor interactions.

【Objective】 The experiment aims to analyze the drug resistance, the drug resistance mechanism, multilocus sequence typing(MLST), the virulence genes, the affinity and Inc-type plasmids of extended-spectrum β-lactamases (ESBLs) producing Escherichia coli of avian origin in Shandong province, so as to provide reference for the rational use of antibiotics in avian clinical prevention and treatment of such bacterial diseases.【Method】 38 strains of ESBLs producing E.coli, isolated from avian farms in Shandong during April 2021 to July 2021, were recovered.The antimicrobial susceptibility test of these isolates against 16 kinds of antimicrobials was determined by the broth microdilution method.The drug resistance, the virulence genes, MLST and Inc-type plasmids were detected by whole genome sequencing.Parsnp was used to analyze the phylogenetic relationship among these isolates.【Result】 The antimicrobial susceptibility test revealed that all strains were resistant to ampicillin, ceftiofur and ceftazidime.The resistance to sulfisoxazole (97.4%), trimethoprim/sulfamethoxazole (94.7%), tetracycline (89.5%), florfenicol (86.8%), spectinomycin (84.2%) was serious. Analysis results of whole genome sequencing showed that 34.2% of 38 strains of ESBLs producing E.coli carried bla_CTX-M-55 in bla_CTX-M type, 22.1% carried bla_NDM-5 and 10.5% carried bla_NDM-4 in bla_NDM type.ESBLs producing E.coli isolates carried a large number of resistance genes that mediate commonly used antimicrobials in clinical practice, and adhesion, invasion, serum resistance, iron transport virulence genes.MLST results revealed that ST10 (13.2%) was the most prevalent ST among ESBLs producing E.coli isolates, followed by ST616 (10.5%) and ST746 (10.5%).Phylogenetic tree analysis showed that ST10, ST616 and ST746 had the same origin, respectively.Inc-type plasmids results revealed that the carrier rate of IncFIB (AP001918) was the highest (68.4%), followed by IncHI2 (42.1%).【Conclusion】 The situation of ESBLs producing E.coli isolates remains very serious, bla_CTX-M mainly accounted for the resistance to extended-spectrum β-lactams antimicrobials.Moreover, this type of strain carried a large number of virulence genes, and it was urgent to strengthen monitoring.

【Objective】 The aim of this study was to develop a transdermal formulation of rifaximin with convenient administration and good skin penetration for the treatment of intestinal diseases in livestock and poultry, and preliminarily evaluate its formulation and efficacy.【Method】 The concentration of rifaximin was detected by ultraviolet spectrophotometer.The orthogonal design method was used in the transdermal preparation test.The in vitro transdermal ability of the preparation was investigated by isolated pig skin, and the prescription components were screened by the cumulative permeation amount (Q) per unit area of 12 h.The storage stability test was carried out at 40 and 60 ℃.A mouse model of Salmonella Typhimurium infection was established, and the prepared transdermal or gavage preparation were used to treat the infection model and evaluate the therapeutic effect by comparing the survival rate, organ bacterial load and histopathological changes.【Result】 Rifaximin transdermal preparation was as follows:The system contained rifaximin 2%, azone 4%, menthol 2%, acetone glycerol 20%, dimethyl sulfoxide 30%, and anhydrous ethanol to 100%.The average cumulative permeation amount per unit area of the formulation for 12 h was 132.82 μg/cm².After placing the preparation at 60 and 40 ℃ for 10 d, the drug content was kept at 89.78% and 93.53% of the initial value respectively, which indicated its good stability.Compared with infection without treatment group, the organ bacterial load in each organ of transdermal group and gavage group were significantly reduced (P<0.05).After 3 d of administration, the survival rate of rifaximin transdermal treatment group (91.67%) was higher than that of gavage treatment group (83.33%).The pathological changes of cecum were obvious before and after treatment.Compared with infection without treatment group, the edema of the intestinal submucosa, the infiltration of inflammatory cells in the submucosa, intestinal crypts and intestinal epithelium were reduced, the number of goblet cells in the intestinal villi was increased, the integrity of the intestinal epithelium was significantly restored, and the lesions were alleviated in the two treatment groups.【Conclusion】 The transdermal preparation of rifaximin had the advantages of simple preparation process, good stability and better therapeutic effect through skin.

Animal derived foods containing veterinary drugs, microbial toxins, preservatives, and drug residues during food processing can cause significant toxic effects and allergic reactions, posing a threat to human health.Molecular imprinting technology mainly induces templates to form specific recognition sites in polymers, which has advantages such as predictability, recognition, and practicality, and has played an important role in the field of veterinary drug residue detection.The robustness, high affinity, specificity, and low production cost of polymer receptors prepared using molecular imprinting technology make them an alternative to natural receptors and toxic compounds.By combining molecular imprinting technology with sensor technology, the advantages and characteristics of both can be fully utilized.The progress of polymer science and nanotechnology can help improve the performance of molecular imprinted polymer sensors, and the applicable fields of molecular imprinted sensors are further expanded.In recent years, many high-quality literature has reported on the research of molecularly imprinted polymer sensors in the fields of biomolecules, abused drugs, and explosives, promoting the application of this technology in the fields of life sciences and hazardous chemicals.The author focuses on the principles of molecular imprinting technology and molecular imprinting sensors, and introduces the latest achievements of sensor technology based on molecular imprinted polymers, as well as research in the field of veterinary drug residue detection applications.

【Objective】 This study was aimed to explore the relieving effect of traditional Chinese medicine on liver damage caused by zearalenone (ZEA) in animals.【Method】 63 8-week-old Kunming female mice were randomly divided into 7 groups, with 9 mice in each group.In control group, physiological saline was administered orally once in the morning and once in the afternoon;ZEA group, 20 mg/kg ZEA was gavaged in the morning and physiological saline was gavaged in the afternoon.Quercetin, kaempferol, rutin, and compound ingredients groups were traditional Chinese medicine treatment groups.In the morning, 20 mg/kg ZEA was administered by gavage, and in the afternoon, 50 mg/kg quercetin, 50 mg/kg kaempferol, 100 mg/kg rutin, and 100 mg/kg compound ingredients (V_(quercetin):V_(kaempferol):V_(rutin)=1:2:2) were administered by gavage, respectively.Positive control group, 20 mg/kg ZEA were administered by gavage in the morning and 100 mg/kg vitamin E was administered by gavage in the afternoon.Each animal was gavaged with 0.2 mL each time for 7 consecutive days.The mice were euthanized on the 8th day to detect body weight, liver index, serum liver function, antioxidant enzyme activity, and liver inflammation indicators.【Result】 The weight development results showed that compared with control group, ZEA group mice had lower body weight and higher liver weight (P<0.05).Compared with ZEA group, the weight of rutin and compound groups were increased (P<0.05).Compared with control group, the serum liver function test results showed that the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), acid phosphatase (ACP), lactate dehydrogenase (LDH) in ZEA group were increased (P<0.05).Compared with ZEA group, the activity of various enzymes in traditional Chinese medicine and vitamin E treatment groups were decreased (P<0.05).The results of liver oxidative stress test showed that compared with control group, the activity of superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) content, and total antioxidant capacity (T-AOC) in liver of ZEA group were decreased (P<0.05), while the content of malondialdehyde (MDA) and LDH were increased (P<0.05).Compared with ZEA group, the activity of SOD, CAT, GSH content and T-AOC in traditional Chinese medicine and vitamin E treatment groups were increased (P<0.05), while the content of MDA and LDH were decreased (P<0.05).Inflammatory factor detection results showed that compared with control group, the content of tumor necrosis factor-α(TNF-α), interleukin-1 β (IL-1β) and IL-6 in ZEA group were increased and the content of IL-10 was decreased (P<0.05). Compared with ZEA group, TNF-α, IL-1β and IL-6 contents of traditional Chinese medicine ingredient and vitamin E treatment groups were decreased (P<0.05), while the IL-10 content were increased in quercetin and vitamin E treatment groups (P<0.05).【Conclusion】 All three traditional Chinese medicine ingredients quercetin, kaempferol, rutin and vitamin E could alleviate the liver damage caused by ZEA in mice, and the effect of kaempferol was the best.