In livestock and poultry production,skeletal muscle development and fat deposition have an important effect on product quality. Regulating skeletal muscle development and fat deposition to improve carcass quality and meat quality of livestock and poultry is of great significance for improving the quality and efficiency of breeding industry. Proline hydroxylases(PHDs) are a class of non-heme iron-dependent dioxygenases that not only act as an important regulator of hypoxic-inducing factor signaling pathway,but also participate in many physiological functions by regulating oxygen adaptation level in cells,apoptosis,cell metabolism and cell transport. On the basis of introducing the physiological function and activity regulation of PHD,this paper summarized the regulatory role of PHDs in animal skeletal muscle development and fat deposition,and analyzed the possible mechanism of its role,in order to provide new ideas and targets for improving the quality of livestock and poultry products by regulating animal skeletal muscle development and fat deposition.

Pregnancy-associated glycoproteins (PAGs) belonging to the multigene family encoded by the aspartic protease gene, are a group of specific glycoproteins synthesized and secreted by placental trophoblast giant cells of artiodactyls, which can be detected in the maternal circulation after embryo attachment. It has the potential to serve multiple functions, including embryonic attachment, placental development, pregnancy maintenance, embryo survival, proteolytic activity, and immune regulation, which are most often used as biomarkers for early pregnancy diagnosis and assessment of embryo loss, as well as indicators of evaluation of fetal viability and monitoring placental health. In this paper, the origins and characteristics and biological functions of the PAGs family and its application in livestock production were reviewed, in order to provide reference for further research of PAGs biological functions and its rapid development in animal husbandry.

Brucellosis is a worldwide zoonotic disease caused by Brucella, which not only induces gigantic economic losses to sheep industry, but also poses significant safety hazards to public health. Brucella possesses the capability of intracellular parasitism and immune evasion, and thus the current measure of vaccination has difficulty in effectively preventing and controling Brucella infection. Cultivating new breeds of sheep resistant to brucellosis is expected to cut off the transmission route of brucellosis from the animal origin. This paper briefly describes the immune evasion mechanism of Brucella dependent on virulence factors, and systematically reviews the research progress of brucellosis resistance breeding in China and abroad, including family-based selection, genetic engineering selection and marker-assisted selection. Furthermore, candidate disease resistance genes were organized according to the development stages of inflammation and immune response. Hoping to provide a theoretical foundation for studying the pathogenic mechanism of brucellosis, identifying disease resistance genes and breeding disease resistance sheep breeds.

Both cold and heat stress can negatively affect the daily gain and feed conversion rate of cattle, which could reduce the production efficiency of the beef cattle industry. This article provides a comprehensive overview of the mechanisms underlying the physiological and biochemical responses of cattle from different regions under cold and heat stress. Additionally, the molecular mechanisms and regulatory genes associated with cold and heat stress were investigated, and practical approaches to mitigate these stressors, such as altering feeding habits and molecular breeding grounded in genome-wide association studies (GWAS), were deliberated and suggested within the beef cattle sector. The review provided the theoretical basis for a better understanding of the mechanisms of cold and heat stress in the beef cattle industry, which could be used in breeding programs to improve production efficiency under cold and heat stress in China.

Mitochondrial autophagy (mitophagy) is an evolutionarily conserved process that selectively identifies, isolates and degrades specific targets. When cells are under adverse conditions, mitophagy eliminates excess or damaged mitochondria to maintain cellular homeostasis and reduce stress on cellular operations. Mitophagy plays an important role in regulation of mammalian reproduction, mainly involving in various physiological processes such as oocyte maturation, senescence and early embryonic development. This paper reviewed the biological functions of mitochondrial autophagy, the regulation of mitochondrial autophagy and the mechanisms of mitochondrial autophagy in mammalian reproduction, providing a reference for further studies on the role of mitochondrial autophagy in mammalian reproductive development.

The proper storage of poultry semen is crucial for preserving genetic resources and ensuring the sustainable development of poultry farming. The unique structural characteristics of avian sperm make them particularly susceptible to oxidative stress during storage, resulting in imbalances in the antioxidant system and a decline in sperm vitality and fertilization ability. To maintain sperm functionality, the supplementation of antioxidants can be employed to mitigate the damage. This review aims to outline the structural features of avian sperm and the characteristics of their antioxidant system. It also summarizes the application and effects of antioxidants in the storage of poultry semen, offering more reference for the research in avian sperm storage.

Reproductive tract microbiome plays an important role in maintaining the reproductive tract health and reproductive efficiency of the host. The disruption of reproductive tract microbiome homeostasis may lead to a variety of reproductive diseases and cause huge losses to the animal husbandry economy. In recent years, some progress has been made in the research of reproductive tract microbiome of dairy cows, which is crucial to the prevention and control of postpartum reproductive diseases in dairy cows. Different physiological stages of breeding cows and postpartum reproductive tract microorganisms may affect their pregnancy ability. Therefore, this paper summarizes the composition of reproductive tract microbiota of healthy dairy cows, the changes in reproductive tract microbiota at different physiological stages and their correlation with the establishment of pregnancy in dairy cows, as well as the influence of reproductive microecology disturbance on dairy cow reproduction. The purpose of this review is to provide references for improving the reproductive efficiency of dairy cows and preventing and controlling reproductive tract diseases by microorganisms.

Copy number variation (CNV) is an important component of genomic structural variation (SV). Compared with SNP (single nucleotide polymorphism), CNV shows more complex genetic variation, which is of great significance in the study of genetic mechanism of important economic traits, disease inducements, and evolution of species. As a unique bovine species in the Qinghai-Tibet Plateau (QTP), with the rapid development of high-throughput sequencing technology and the continuous upgrading of yak reference genomes, the study of CNV at the whole genome level in yak has made important progress in recent years. Here, on the basis of describing the formation principle, function mechanism and detection method of genomic CNV, the current research status of genomic CNV in yak in recent 10 years were reviewed, and some existing problems and shortcomings were analyzed and explored. Furthermore, the future development and application trends were prospected, in order to provide reference for further yak genome research and accelerating molecular breeding process of yak genetic resources.

Bovine coronavirus is an important pathogen that can infect bovine intestinal tract, respiratory tract and cause calf diarrhea, adult winter dysentery and bovine respiratory diseases of all ages, which seriously endangers the development of cattle industry. The spike protein is the main structural protein and immunogenic protein of the virus, which plays an important role in the initial infection of the virus and the induction of protective immunity. This paper describes the structural features, biological functions and application studies of the virulence of the spike protein, as well as its immunogenicity, causes of mutation and the latest research progress of the potential dual-receptor system, with the hope of providing a scientific basis for the development of a new type of vaccine and target therapeutic drugs, so as to formulate a rapid and effective safe prevention and control countermeasures to deal with the emergence of new strains of the disease.

Pseudorabies virus (PRV) belongs to the α-herpesvirus subfamily, which can infect the host nervous system and establish latent infection in neurons, and PRV is an important pathogen threatening the global swine industry. Similar to other α-herpesviruses, the virion of PRV has a protein-rich inner membrane layer that links the DNA-containing capsids to the glycoprotein filled envelope layer to form the complete structure of the virion. Studies have shown that during the infection cycle of PRV, tegument proteins can play important biological functions in maintaining the stability of the morphological structure of the virus, promoting the replication and nuclear egress of the virus, participating in the secondary envelopment of the virus in the cytoplasm, and fighting against the host immune system by acting alone or interacting with other proteins. Therefore, this paper focuses on the recent research progress on the biological functions of PRV tegument proteins, in order to provide important references for the etiology and antiviral research of pseudorabies virus.

Bacterial outer membrane vesicles (OMVs) are vesicle-like structures released by Gram-negative bacteria during their growth. OMVs can be considered as "long-range weapons" of bacteria. OMVs play an important role in bacterial pathogenesis, which functions are to attack host tissues and help bacterial pathogens establish their biological niche, damage host cell functions, and regulate host defense. The substances secreted by bacteria into vesicles can drive special functions in the environment through combination, such as quorum sensing communication, biofilm formation, nutrition acquisition, antibiotic resistance, stress response, competition or defense against other microorganisms, environment, microbial community status, nucleic acid transfer, horizontal gene transfer, delivery of toxins and virulence factors. These characteristics play an indispensable role in the growth cycle of bacteria. This article mainly reviews the latest research progress in the structure, secretory characteristics, and regulatory pathogenicity mechanism of bacterial OMVs, providing theoretical guidance for the in-depth study of bacterial pathogenicity mechanism and the development of new antibacterial strategies.

This study aimed to explore the regulation mechanism of DNA methylation editing of nitric oxide synthase (NOS) 2 on Myoglobin, and study its effect on the expression level of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS). In this experiment, DNA methylation editing technology was used to methylate NOS2 in porcine kidney cell line PK-15 cells, and the DNA methylation level of the NOS2 gene promoter, the expression level of NOS2, the concentration of intracellular nitric oxide, as well as the expression level of myofibrillar-and myoglobin-related genes were detected in porcine kidney cells. The results revealed that the average DNA methylation level of the first 500 bp region of the NOS2 gene promoter was reduced, but the difference was not significant, while the methylation level of the CpG site near the gRNA2 binding site was increased. In addition, after DNA methylation editing of NOS2 gene, NOS2 gene expression and its iNOS protein expression were significantly reduced. With the decrease of NOS2 expression, nNOS and eNOS expression increased significantly, the expression level of MYHC-Ⅱb and MYHC-Ⅱx decreased significantly, and the expression of Myoglobin protein decreased significantly. In conclusion, the abnormal expression of the NOS2 gene can affect the generation of NO in cells, as well as the expression of NOS1 and NOS3, and the expression of genes related to muscle development.

The aim of this study was to analyse the genetic diversity and genetic structure of the Huixian Qingni Black pig, and to provide reference for its declaration, protection and utilization as a new genetic resource. A total of 68 pigs from two known local pig breeds (20 each of Bamei pigs and Hezuo pigs) and a new pig breed to be identified (28 of Huixian Qingni Black pigs) in Gansu Province were used for the study. "Zhongxin-1"50K SNP chip was used to detect whole-genome single nucleotide polymorphisms (SNPs), and a variety of softwares were used to analyse the genetic diversity, genetic structure, population differentiation indices and relationship of the Huixian Qingni Black pig. The results showed that:a total of 55 156 SNPs were detected, with 49 279 SNPs remaining after quality control; The effective population size (N_e), expected heterozygosity (H_e), observed heterozygosity (H_o), proportion of polymorphic markers (P_N) and nucleotide diversity (P_i) of Huixian Qingni Black pigs were higher than Hezuo pigs and Bamei pigs at 2.2, 0.370 7, 0.386 4, 0.915 7 and 0.378 6, respectively, and the linkage disequilibrium coefficients of Huixian Qingni Black pigs were lower and decayed faster; PCA showed that the three populations were clustered respectively, and HX population could be distinguished from HZ and BM populations according to PC1 (PC1>0), and the evolutionary tree analysis revealed that the Huixian Qingni Black pigs clustered into a single branch, while the Bamei pigs and the Hezuo pigs clustered into another large branch, and then gradually separated, and the population structure showed that the Huixian Qingni Black pigs showed a different evolutionary route from the Bamei pigs and the Hezuo pigs when K=3, but the bloodlines of the Huixian Qingni Black pigs were more mixed; The population differentiation indices between the Huixian Qingni Black pigs and Bamei pigs, Hezuo pigs were 0.123 6 and 0.159 8, respectively, indicating that there was a certain degree of genetic differentiation among the pig breeds; The IBS matrix and G matrix analyses showed that most of the individuals in Huixian Qingni Black pigs were distantly related to each other, while a few individuals were more closely related to each other. In this study, we analysed the genetic diversity and genetic structure of Huixian Qingni Black pigs based on "zhongxin-1"50K SNP chips data, and found that the genetic diversity of Huixian Qingni Black pigs is higher than that of Bamei pigs and Hezuo pigs, independently clustering, and there is obvious genetic differentiation with Bamei pigs and Hezuo pigs. It is preliminarily considered to be a new local pig breed in Gansu. There is risk of inbreeding between some of the individuals of Huixian Qingni Black pig, which needs to be strengthened to preserve the breed, this study provided a reference for further excavation of new genetic resources of Huixian Qingni Black pigs and rational breeding conservation and utilization.

This experiment aimed to investigate the effect for introduction of 25% Rhode Island Red chicken gene on the related indexes of Henan fighting chickens, which would provide the theoretical basis for the cross breeding of local variety birds. The same batch of 120-day-old purebred Henan fighting chickens and hybrid fighting chickens being introduced 25% Rhode Island Red gene (n=10 for each population, Half male and female) were selected. They were divided into 4 groups with 5 replicates per group and 1 chicken per replicate. After slaughtering, the following traits were determined, including slaughter performance, muscle properties, serum biochemistry, and mRNA levels of target genes (by real-time quantitative PCR) related to muscle development and glucose transport. All data were analyzed by SPSS 23.0 for Two-way ANOVA analysis. It showed that there was no significant difference in slaughter indexes between hybrid fighting chickens and purebred Henan fighting chickens (P>0.05). The diameter of pectoralis muscle fiber showed a significant population effect with hybrid fighting chickensP=0.014). The interaction effect between populations and genders was not significant for the pectoralis muscle fiber density (P=0.056). The pectoral muscle fiber density of hybrid roosters was significantly higher than that purebred birds (P<0.01). The muscle fiber density of leg muscle showed a significant population effect (P=0.002), in both male and female, hybrid chickens were significantly higher than purebred chickens. There was no difference in the expression of muscle development related genes (MyHC, MyoG, MyoD1) and glucose transport related gene GLUT12 between hybrid chicken and purebred fighting chicken by qRT-PCR, but the expression of GLUT4 gene in pectoral muscle(P=0.000)and leg muscle(P=0.023)showed significant interaction between population and gender. In leg muscles, shear forces have a significant gender effect (P=0.020), the muscle fiber density presented significant population effect (P=0.002). The expression of GLUT4 gene in pectoralis of crossbred cocks was significantly higher than that of purebred cocks (P<0.01), while where the level of GLUT4 of purebred hens was significantly higher than that of hybrid hens (P<0.05). In leg muscle tissue, the expression of GLUT4 in hybrid hens was significantly higher than that in purebred hens (P<0.01), but where there was no significant difference for GLUT4 level between hybrid cocks and purebred cocks (P>0.05). The analysis on serum biochemical indexes showed that the levels of serum GLO (P=0.004), CHO (P=0.011) and LDL-C (P=0.014) showed significant population effects, and the levels of hybrid cocks were all higher than those of Henan fighting cocks. In addition, the serum A/G ratio showed a significant interaction effect (P=0.013), the ratio of serum A/G in purebred fighting cocks was significantly higher than that in hybrid fighting cocks (P<0.05). Although the introduction of 25% Rhode Island Red gene did not affect the slaughtering index of fighting chicken, it significantly increased the blood lipid level and skeletal muscle fiber density of cocks, and changed the expression of glucose transporter GLUT4 gene in a tissue-specific and gender-specific manner.

This work aimed to screen key miRNA(s) that regulate skin melanin deposition in goats, and to explore its regulation mechanism on skin color or coat color. The 100-day-old fetal skin samples from the Youzhou dark goat (YZDG, n=3) and Chuandong white goat (CDWG, n=3), and skin samples from the healthy 2-3-years-old Dazu black goats (DBG, n=3), Inner Mongolia cashmere goat (IMCG, n=3) were collected to observe skin melanin deposition by section staining technology, and to explore differential miRNAs by miRNA-seq technology. Otherwise, B16-F10 cutaneous melanoma cells were cultured. The effect of miR-129-5p on melanin production was verified by cell transfection, qPCR, Western blot and melanin content detection. The results showed that melanin granules were obviously deposited in the hair bulb, hair shaft, and outer root sheath of DBG hair follicles and YZDG fetal goat skin, whereas they were not observed in the skin of CDWG fetal goats and the epidermis, hair follicles of IMCG. The sequencing results showed that 62 miRNAs were differentially expressed between YZDG and CDWG, of which 31 were up-regulated and 31 down-regulated in dark-skin goats. A total of 38 differentially expressed miRNAs were found between DBG and IMCG, 10 were up-regulated and 28 down-regulated in black-coated goats. The result showed miR-129-5p was significantly highly expressed in dark-skin and black-coated goats (P<0.05). After over-expression of miR-129-5p, the cell melanin deposition in the mimics group was increased by 18.9% (P<0.05), the expression of the TYR and TYRP1 genes was up-regulated by 57.3% and 16.5%, respectively (P<0.05), and the protein expression was significantly up-regulated by 49.2% and 40.2% (P<0.05); but the mRNA and protein expressions of MITF were not significantly changed (P>0.05). After inhibition of miR-129-5p, the mRNA expression of TYR gene and its protein expression level in the inhibitor group were significantly down-regulated by 38.9% and 21.1%, respectively (P<0.05). And TYRP1, MITF protein expression levels were down-regulated by 25.3% and 28.4%, respectively (P<0.05). The results indicated that the miR-129-5p was differentially expressed in different skin colors or coat colors of goats, and it could affect melanin deposition by regulating the transcript and protein abundance of melanin-related genes (such as TYR and TYRP1). These results demonstrate that miR-129-5p is an important regulator involved in the skin melanogenesis of goats.

This study aimed to evaluate the genetic parameters of longevity traits of Holstein cows and provide insights for improving production performance and implementing balanced breeding. The culling records for 19 756 Holstein cows from 9 large-scale dairy farms in Jiangsu during year 2017 to 2023 were collected, and the longevity and elimination situation of cows was analyzed. The significance test for fixed effects including factors like farms, birth year, birth season and age at first calving was conducted using the GLM program in R software. The variance components and calculated heritability for longevity traits were estimated in BLUPF90. The breeding values for longevity traits of Holstein cattle in Jiangsu were evaluated and the genetic trend was depicted. The results showed that the average herd life of adult cows in large-scale farms in Jiangsu was 1 640.87 d, the production life was 917.15 d, and the average culling parity was 2.60. The longevity traits were found to had relatively low heritability, ranging from 0.07 to 0.15. The genetic and phenotypic correlations between these 7 longevity traits (herd life, production life, culling parity, L1-L4) exhibited moderate to high correlations, ranging from 0.27 to 0.99. Notably, the correlation between herd life and productive life was closer to 1. Moreover, the genetic trend for longevity traits was assessed through breeding values, and which had shown a deceleration in recent years, indicating a reduced rate of genetic improvement. This study systematically analyzed the genetic variation for longevity traits using data from 9 large-scale farms in Jiangsu. These findings can provide insights for the genetic evaluation and making breeding decisions for longevity traits in the future.

The study aimed to explore the effects of bta-miR-101 on bovine testicular sertoli cells' proliferation, apoptosis and secretion.In this study, testicular tissues of healthy bulls (n=3) aged 3 days and 13 months were collected to construct the expression profile of Angus cattle tissues. The differences in bta-miR-101 expression were verified between the two periods. Online tools (TargetScan, miRTarBase, miRDB, and miRWalk) were used to predict the target gene of miR-101, and KOBAS was used for enrichment analysis of GO and KEGG pathways. The mimics and inhibitors of miR-101 were transfected into bovine testicular sertoli cells. Cell proliferation, apoptosis, secretion-related gene expression, and cell proliferation coefficient were detected by RT-qPCR, Western Blotting, and CCK-8 techniques.There were 26 common target genes for bta-miR-101 predicted by 4 online tools. GO and KEGG enrichment analysis showed that these genes were mainly enriched in the nucleoplasm, protein-binding, and JAK-STAT receptor signaling pathways, which were associated with cell differentiation, cell cycle, apoptosis, and other biological processes to varying degrees. The results of RT-qPCR showed that bta-miR-101 expression was significantly higher in testis tissue at birth than at sexual maturity (P<0.05), consistent with previous laboratory sequencing results. After transfection of miR-101 mimics and inhibitors, it was found that miR-101 mimics could promote the expression of proliferation-related genes at mRNA and protein levels, significantly increase the proliferation coefficient of sertoli cells, and inhibit the expression of apoptosis genes at mRNA and protein levels (P<0.05); miR-101 inhibitors inhibited the expression of proliferation-related genes at mRNA and protein levels and promoted the expression of apoptotic genes at mRNA and protein levels (P<0.05). In terms of cell secretion, miR-101 had an effect on the transcription of secretion marker genes GDNF and BMP4, but had no significant effect on the secretion of androgen-binding proteins. In conclusion, miR-101 promoted proliferation and inhibited apoptosis of immature sertoli cells of the bovine testis, and had no significant effect on the secretion of androgen-binding proteins.

In dairy cattle breeding systems dominated by cryopreserved semen and artificial insemination, the post-freezing motility of bull sperm is of great interest. Sperm freezability is the key to the post-freezing sperm motility. Previous studies have shown significant differences in sperm freezability among different bulls. Therefore, it is necessary to explore genes and molecular markers that affect sperm freezing tolerance at the genetic level, in order to lay a foundation for improving the sperm freezability of Chinese bulls. The present study investigated the sperm proteome of Holstein bulls in the high sperm freezability group (n=9) and the low sperm freezability group (n=6). The fresh sperm motility of bulls in the high sperm freezability group was greater than 0.65 and post-freezing motility greater than 0.4, while the fresh sperm motility of bulls in the low sperm freezability group was greater than 0.65 and post-freezing motility less than 0.27. Quantitative proteomic analysis was conducted on sperm cells of 15 bulls using label-free proteomics technology followed by bioinformatics analysis. As a result, a total of 432 differentially expressed proteins were identified between the high and low freezability groups, which were mainly enriched in biological processes closely related to sperm energy supply, such as metabolic pathway and oxidative phosphorylation. The analysis of protein-protein interaction networks showed that there were strong interactions between proteins related to energy metabolism. Compared with the QTL database, it was found that 9 differentially expressed proteins overlapped with genes associated with percentage live sperms after thawing. Overall, several important candidate proteins were identified as potential biomarkers for the freezing resistance of bull sperms, such as HSPA1A, BSP3 and ACSL4. This study revealed the proteomic characteristics of bull sperms with different freezability, which provides important information for molecular breeding of frozen semen quality in Chinese bulls.

The aim of this study was to investigate the effects of Enterococcus faecium (EF) and Bacillus subtilis (BS) supplementation on growth performance, serum biochemistry, and intestinal health of brood-rearing laying hens, as well as the carryover effects of feeding during this period on the subsequent laying performance. A total of 528 1-day-old Roman pink laying hens were selected in a 2×2 experimental design. BS consisted of two supplemental amounts of 0 or 200 mg·kg^-1 (BS viable count:3×10¹⁰ CFU·g^-1). EF consisted of two additions of 0 or 200 mg·kg^-1 (EF viable count:2×10⁹ CFU·g^-1). This experiment included treatments with six replicates per treatment and 22 hens per replicate. The experimental period lasted 30 weeks. The experimental treatment was carried out at brood-rearing period from 1 to 20 weeks. The laying period was from 21 weeks to 30 weeks and fed with basal diet. The results showed that:1) There was a significant interaction between BS and EF on the feed to meat ratio (F/G) of laying hens from 1 to 20 weeks, indicating that the combination of the two probiotics increased F/G ratio (P<0.05). 2) BS supplementation tended to decrease serum ALT level at week 20 (P=0.067). There was an interaction effect of BS and EF on serum biochemistry of laying hens at 20 wk, as indicated by the reduction of serum total protein, serum high-density lipoprotein and total cholesterol content by the combination of the two bacteria (P<0.05). 3) EF supplementation tended to increase serum GH level at week 20 (P=0.078). 4) BS and EF increased the relative length of ileum (P<0.05). The addition of BS tended to increase the height of jejunal villus (P=0.051) and crypt depth (P=0.069). EF supplementation significantly increased jejunal villus height (P<0.05), and tended to increase the depth of jejunal crypt (P=0.072). 5) BS significantly increased the relative expression of Occludin mRNA in the jejunal mucosa (P<0.05), and had a tendency to increase the relative expression of Claudin-1 mRNA (P=0.081). 6) Adding BS tended to increase the average egg weight (P=0.077). In conclusion, supplementation of BS can enhance the growth performance, serum biochemical indexes and intestinal health of laying hens during the brooding period. Furthermore, supplementing with BS during this period can also improve their performance during the laying period, surpassing that of EF and a combination of two probiotics.

The main purpose of this experiment was to explore the effect of high proportion rumen bypass fat diet on feed intake and feeding behavior of Hulun Buir sheep during growth. Twelve Hulun Buir sheep with an average weight of (20±1.9) kg were randomly assigned to the control group (CON, n=6) and the high fat group (23.65% high rumen bypass fat diet, HF, n=6). And continuously record the behavioral activities of each Hulun Buir sheep during the experiments using a camera. The behaviors of the experimental sheep on the 83 d and 84d were continuously observed and recorded, including feeding, ruminating, drinking, standing, and lying. The time spent on each activity over a 24 h period had been recorded. Based on the feed intake, the rate of dry matter, fat, and crude protein intake were calculated. The results showed that dietary supplementation with high proportion of rumen bypass fat significantly decreased the concentrate feed intake, dry matter intake, crude protein intake and feeding speed (P<0.05), the feed intake times, daily rumination time, rumination time per kilogram of diet and drinking time (P<0.05) of growing Hulun Buir sheep (P<0.05), but increased the intake and feed speed of crude fat (P<0.01). The results suggested that dietary supplementation of high proportion of rumen bypass fat may reduce concentrate intake by affecting the feeding frequency of growing sheep, accompanied by a decrease in drinking behavior.

The study was conducted to determine the effects of betaine on the performance and antioxidant function of late-phase laying hens. A total of 108 Hy-Line variety Brown laying hens with 500-day-old were selected and randomly divided into 3 groups, with 3 replicates in each group and 12 hens in each replicate. The birds in control group were fed with the basal diet, and the experimental groups were supplemented with 1 000 and 3 000 mg·kg^-1 betaine on the basal diet respectively for 42 days. Samples of eggs, serum, ovary and granulosa cells of small yellow follicles at the end of the experiment were collected. The results showed that:1) Compared with the control group, betaine supplementation linearly increased the laying rate of layers, average egg weight (P_L<0.01) and protein height, yolk color, yolk weight in the late laying period (P_L<0.05). 2) Compared with the control group, the addition of betaine to the diet resulted in a significant linear increase in the level of Prog in serum (P_L<0.01), and dietary betaine linearly and quadratic increased the level of FSH and LH in serum (P_L<0.01; P_Q<0.05). 3)The addition of betaine to the diet linearly increased the activity of SOD, GSH-Px in the serum, and the level of T-AOC,the activity of SOD, CAT, also linearly decreased the content of MDA in ovary(P_L<0.05). At the same time, dietary betaine linearly and quadratic increased the level of T-AOC and the activity of CAT in serum, decreased the content of MDA (P_L<0.01; P_Q<0.01). The addition of betaine to the diet linearly increased the mRNA expression levels of SOD, HO-1 (P_L<0.01),quadratic increased the mRNA expression level of Nrf2 (P_Q<0.01), linearly and quadratic increased the mRNA expression level of NQO1 (P_L<0.01; P_Q<0.01) and significantly increased the mRNA expression level of CAT (P<0.05). Collectively, the addition of betaine to the diet can increase the level of hormone in serum and the activity of antioxidant enzymes in serum and ovary, improve its antioxidant capacity active by promoting the expression of antioxidant related genes in small yellow follicle granulosa cells and thereby improve the production performance of late-phase laying hens. Moreover, the addition effect of high doses is better than that of low doses.

Lipophagy is a new form of autophagy, which can selectively recognize and degrade lipid droplets, and plays an important role in maintaining intracellular lipid homeostasis. The aim of this study was to investigate the effect of starvation induced lipophagy on the size of lipid droplets in bovine mammary epithelial cells. The cells were treated with 100 μmol·L^-1 linoleic acid for 24 h to establish a lipid accumulation cell model, and then the cells were starved for 0, 6, 12, 24, and 48 h, respectively. Oil red O staining was used to observe the changes in the size and number of intracellular lipid droplets with starvation time. Immunofluorescence was used to observe the co-localization of lipid droplets and autophagy protein LC3. Western blot was used to detect the expression of autophagy-related proteins LC3 and P62. The results showed that the diameter of lipid droplets significantly increased from 1.57 μm to 2.12 μm, the number of lipid droplets per cell significantly decreased from 39 to 24·cell^-1, the proportion of large lipid droplets significantly increased, and the proportion of small lipid droplets significantly decreased with the extension of starvation treatment time (P<0.05), immunofluorescence showed that LC3 co-localized with lipid droplets, and transmission electron microscopy showed that lipid droplets were encapsulated in autolysosomes. After 24 hours of starvation, the ratio of LC3Ⅱ/LC3Ⅰ protein increased by 1 fold, and the expression of P62 protein decreased significantly (P<0.05). These results suggest that lipophagy plays a regulatory role in the size of lipid droplets in the bovine mammary epithelium.

This study was conducted to explore the effects of rumen-protected methionine (RPM) on meat quality, volatile flavor substances and fatty acid composition of yak semitendinosus. Twenty-four Maiwa male yaks (4 years old; body weight:252.79±15.95 kg) were randomly divided into 4 groups:1) basal diet (CON), 2) basal diet + 5 g·d^-1 RPM (RPM1), 3) basal diet + 10 g·d^-1 RPM (RPM2), and 4) basal diet + 15 g·d^-1 RPM (RPM3). Yaks were slaughtered after feeding 70 days. The results showed that the fat content in the RPM3 group was significantly higher than those in RPM1 and RPM2 groups (P<0.05). The crude protein content in the RPM1, RPM2 and RPM3 groups were significantly higher than that in the CON group (P<0.05). A total of 42 volatile flavor compounds were detected in yak semitendinosus samples. According to the relative odor activity value, there were 9, 11, 9 and 10 key volatile flavor compounds in the CON, RPM1, RPM2 and RPM3 groups, respectively. The key volatile flavor compounds provided yak semitendinosus with unique meat flavor and fat flavor. Nonanal was the main volatile flavor compound in the CON, RPM1 and RPM2 groups, while octenol was the main volatile flavor compound in RPM3 group. The content of aldehydes in the RPM1 group was the highest among the treatments, and the types of alcohols and hydrocarbons in the RPM2 group were the most among the treatments. A total of 16 fatty acids were detected in yak semitendinosus samples. The total saturated fatty acids in the CON group were significantly lower than those in the RPM1 and RPM2 groups (P<0.05), and palmitic acid in the RPM1 and RPM2 groups were significantly higher than those in other groups (P<0.05), docosanoic acid in the RPM1 group was significantly higher than those in the CON and RPM3 groups (P<0.05). The total monounsaturated fatty acids in RPM1 group were significantly higher than those in RPM2 and RPM3 groups (P<0.05), and the myristicoleic acid in RPM1 group was significantly lower than those in other groups (P<0.05), heptadecenoic acid in CON group was significantly higher than those in other groups (P<0.05), tetracosenoic acid in the RPM1 group was significantly higher than that in the RPM2 group (P<0.05). There was no significant difference in total polyunsaturated fatty acids among the groups (P>0.05). Inconclusion, the flavor richness volatile flavor substances content of yak semitendinosus in the 10 g·d^-1 RPM group were the highest. The fatty acid content in the 5 g·d^-1 RPM group was the highest, and the distribution was reasonable.

To shorten the detection cycle of subgroup J avian leukosis virus (ALV-J) and further accelerate the process of avian leukosis decontamination, this study combined SYBR Green Ⅰ qPCR and pooling sample assay to establish a blood nucleic acid screening technique (ALV-J-B-qPCR) for rapid screening of ALV-J in low prevalence. Based on the sequences of ALV-J pol and env genes in GenBank, the specific primers for ALV-J were designed and the reaction conditions were optimized to establish the ALV-J SYBR Green Ⅰ qPCR assay. Anticoagulant DNA, blood cell DNA, peripheral blood lymphocyte DNA, peripheral blood lymphocyte cDNA and plasma cDNA were respectively prepared to perform PCR detection of ALV-J, and the best assay templates were screened. Further, the accuracy of the three mixing methods of extracting DNA from mixed blood by red blood cell cracking method, the mixed method with 200 μL total volume of blood mixed sample and the mixed method with 10 μL total volume of blood mixed sample were compared, and the best mixing method was selected. The ALV-J-B-qPCR was established by combining the above qPCR method, assay template and mixing method. Simulated screening tests with expected prevalence scenarios of 1%-2% and 4%-5% were performed using ALV-J-B-qPCR respectively, and compared with the virus isolation method. Specificity, sensitivity and reproducibility tests of the qPCR method showed that the method specifically amplifies only ALV-J and has a minimum detection limit of 1×10² copies·μL^-1 for standard plasmids, with intra-and inter-batch coefficients of variation <1%. Results on 90 clinical samples showed that the detection rate of ALV-J by this qPCR method (15.6%) was higher than that of p27 antigen ELISA and PCR (12.2%). In the assay template screening test, anticoagulated blood DNA best met the requirements of high assay accuracy, operational complexity and low cost, and was the best assay template. In the mixing method screening test, the mixing method with a total mixing volume of 10 μL (mixing size <12) had the highest detection accuracy and was the best mixing method. In a simulated screening with a sample size of 400 and an expected prevalence of 4%-5%, the detection rate of ALV-J-B-qPCR was 6.25%, which was 2.00% higher than that of the virus isolation method; in a simulated screening with a sample size of 400 and an expected prevalence of 1%-2%, the detection rate of ALV-J-B-qPCR was 2.25%, which was 0.75% higher than that of the virus isolation method. The ALV-J-B-qPCR technique established in this study has the advantages of high sensitivity, rapid detection and cost saving, which provides a new idea and method to accelerate the ALV purification process in breeding poultry farms.

In the study, we found that one type of tumors in yellow-feather chicken that look like a white, stiff, irregular shaped mass, growing up at the position between comb and eyelid. In order to identify the pathogenetic characteristics and causative agent of the disease, we have made a series of diagnoses and studies of the disease. Research has found that about 2% sick chicken in the laying hens, they have no other pathological changes other than the tumors. The histopathological diagnosis the pathological tissue was Hemangioendothelioma. RT-PCR tests on tumor tissue showed that the sick chickens were infected with the subgroup K avian leukosis virus (ALV-K). We have tried to isolate the viruses from 32 blood samples, 6 tumor tissues and brain tissues of infected chicks. Finally, we successfully isolated 4 strains of ALV-K from brain tissues. The 4 strains of ALV-K were named as GXJL01-GXJL04. To better understand the molecular characterization of GXJL01-GXJL04. On the other hand, we have isolated another 7 strains of ALV-K from 5 large-scale chicken farms in South China, amplifying and sequencing their env genome. The phylogenetic tree of 11 strains of ALV-K was drawn by MEGA-X software based on gp85 amino acid sequence, indicating that GXJL01-GXJL04 were most similar to the JS15SG01 and fowl glioma-inducing virus (FGV) such as Km5844 and Sp53, and they were distantly related to the genetic evolution of the other 7 ALV-K isolates in this study. What's more, in the analysis of gp85 amino acid point mutation, we have found that some amino acid sites are the same as fowl glioma-inducing virus and JS15SG01, while they are different from other 7 strains of ALV-K from south China. In the study, the tumors of the chicken was firstly diagnosed as Hemangioendothelioma, the associated pathogen was ALV-K, and the isolates GXJL01-GXJL04 were highly homologous to neurotropic strains like FGV, JS15SG01, but were different from the other circulating ALV-K in South China.

The avian influenza virus (AIV) of subtype H3 infects a wide range of hosts and is capable of cross-species transmission. In addition to causing disease in poultry, it also causes many cases of human infection, which is of great public health importance. Therefore, it is necessary to establish a rapid and sensitive method for the detection of H3 subtype AIV. In this study, the H3 subtype AIV HA gene sequence published by GenBank in recent years was used to design a pair of specific primers and TaqMan probes in the conserved region. By optimising the reaction conditions and drawing the standard curve using cRNA standard and viral nucleic acid standard as templates, a fluorescence quantitative RT-PCR detection method for H3 subtype AIV was established and its specificity, sensitivity and repeatability were evaluated. The method was further validated using laboratory-challenged chicken tissue samples and clinical swab samples. The results showed that the method was highly specific and did not cross-react with other subtypes of AIV and common avian pathogens. The lower detection limits of the method were 1.0×10² copies·μL^-1 and 10² EID₅₀·0.1 mL^-1, respectively, and the sensitivity was 10-fold higher than that of conventional RT-PCR. The coefficient of variation within and between groups was less than 1.5% and the repeatability was good. The results of the animal challenge clinical test and clinical swab samples showed that the sensitivity of the method was higher than that of the conventional RT-PCR method, and the concordance rate was 100%, which could be used for clinical detection. In conclusion, the fluorescent quantitative RT-PCR detection method for H3 subtype AIV established in this study has the characteristics of specific, rapid and sensitive, which provides certain technical support for the rapid diagnosis, monitoring, prevention and control of H3 subtype AIV.

Infectious bovine rhinotracheitis virus (IBRV) is a bovine respiratory virus that has a serious impact on the global cattle industry. Based on an epidemiological investigation of IBRV in 1-month-old calves from four large-scale Angus beef cattle breeding farms in the southern Xinjiang region, the impact of IBRV infection on the nasal microbiota of Angus calves was explored. One-month old Angus calves with respiratory symptoms (mainly including fever, cough, and difficulty breathing) were clinically investigated. Nasal swabs were collected from the calves for IBRV PCR detection. Based on the PCR detection results, 10 pure IBRV positive calves (Group P) and 10 healthy calves (Group N) with negative IBRV and no other respiratory virus infection were randomly selected, Use 16S rRNA gene sequencing technology, select V3 and V4 variable regions, and use Illumina platform to perform Paired end sequencing on the DNA fragments of nasal microbiota, analyze the composition structure and function prediction of the two groups of nasal microbiota. The results showed that 922 calves had symptoms of respiratory diseases, and the incidence rate was 8.2% (922/11 215); Among them, 98 died, with a case fatality rate of 10.6% (98/922). The positive rate of IBRV in the sample test was 22.0% (50/227). Compared with the number of taxonomic units of nasal flora of calves in group N, calves in group P showed a highly significant increase at the level of phylum, class, order, and family (P<0.01), and a trend towards an increase at the level of genus (P=0.056). In Alpha diversity, there was a significant difference in the coverage and evenness of the two groups of microbiota (P<0.05), while in Beta diversity, there was a significant difference in the structure of the nasal microbiota between Group P and Group N (P<0.05). The difference between bacteria and phyla showed that the abundance of Bacillota in group P was significantly lower than that in group N (P<0.01), and the abundance of Chloroflexota, Acidobacteriota, Cyanobacteria, Gemmatimonadota was significantly higher than that in group N (P<0.01); The abundance of Planococcus and Salinicoccus in group P were significantly lower than that in group N (P<0.05), and the abundance of Halomonas and Lactobacillus were significantly higher than that in group N (P<0.05). There are 9 metabolic pathway changes in the MetaCyc metabolic pathway in group P calves, and 7 metabolic pathway changes in the KEGG metabolic pathway abundance prediction, mainly related to participating in body synthesis, inflammatory response markers, and affecting body growth and development. In addition, there are differences in cellular metabolism, material transportation, decomposition and synthesis, and potential functions of diseases. This study preliminarily revealed a close correlation between IBRV infection and changes in the composition, structure, and function of nasal microbiota in cattle with respiratory symptom diseases, providing a theoretical reference for further exploring the pathogenesis of IBRV infection in calves.

In order to provide important experimental materials for the in-depth study of the biological functions of the M protein encoded by the ORF6 gene of porcine reproductive and respiratory syndrome virus (PRRSV), in this study, we firstly constructed a recombinant lentiviral plasmid overexpressing the ORF6 gene of PRRSV using the lentiviral packaging system. The recombinant lentiviral plasmid overexpressing PRRSV ORF6 gene was constructed using the lentiviral packaging system, and the plasmid, together with the helper plasmid, was co-transfected into HEK293T cells to obtain recombinant lentivirus; after that the recombinant lentiviruses were infected into MARC-145 cells, and screened using puromycin in combination with the finite-dilution method, and after three consecutive rounds of screening, a MARC-145^ORF6 cell line was established that expresses the M protein of PRRSV; and The effect of overexpression of PRRSV M protein on the growth of MARC-145 cells was assessed using CCK-8 assay. RT-PCR, protein immunoblotting (Western blot) and indirect immunofluorescence (IFA) were used to assess the passaging stability of the MARC-145^ORF6 cell line and to identify the subcellular localization of the M protein, and RT-qPCR was further used to assess the effect of overexpression of the M protein on interferon and related regulatory genes in MARC-145 cells; in addition, the effect of PRRSV viral titres in MARC-145^ORF6 cell line, MARC-145^Flag cell line and MARC-145 cells and plotted multi-step growth curves to compare the differences. The results of CCK-8 assay showed that there was no significant effect on the viability of MARC-145 cells after overexpression of PRRSV M protein; the results of RT-qPCR, Western blot and IFA showed that the MARC-145^ORF6 cell line was able to express the M protein of PRRSV and was stable during passaging. And the expression of PRRSV ORF6 gene and M protein were stable. In addition, stable expression of PRRSV M protein significantly down-regulated type Ⅰ interferon and its related regulatory genes in the cell line; the multi-step growth curve showed that the MARC-145^ORF6 cell line promoted the proliferation of PRRSV and increased its viral titer. In conclusion, the MARC-145^ORF6 cell line, which can stably express PRRSV M protein, was constructed in this study, and was found to significantly down-regulate the level of type Ⅰ interferon and promote PRRSV replication. The MARC-145^ORF6 cell line constructed in this study will provide an important biological material for the in-depth study of M protein function.

To establish a blocking ELISA method for the detection of porcine circovirus type 3 (PCV3) antibodies, in this study, a hybridoma cell line 2E6, which secretes a good blocking antibody, was obtained by immunization of BALB/c mice with the prokaryotic expression of PCV3 Cap recombinant protein. The recombinant Cap protein was used as the coating antigen, and the horseradish peroxidase (HRP)-labeled 2E6 monoclonal antibody was used as the detection antibody, and the conditions were continuously optimized to finally establish a blocking ELISA method for detecting PCV3 antibody. The established blocking ELISA method was used to test 50 clinical negative sera, and the threshold value of blocking rate (PI) was calculated to determine the cut-off of the method. When PI ≤ 28.30%, the result was determined as negative; when PI ≥ 35.05%, the result was determined as positive; when 28.30%<35.05%, the result was determined as suspicious. The suspicious results should be tested again and the still suspicious results will be judged as positive. Specificity tests showed no cross-reactivity with positive sera for porcine circovirus type 2 (PCV2), pseudorabies virus (PRV), porcine reproductive and respiratory syndrome virus (PRRSV) and swine fever virus (CSFV). The sensitivity test showed that the detection potency could reach 1:128. The repetition test showed that the intra- and inter-batch coefficients of variation were less than 10%. The conformity test showed that the method was highly consistent with the gold standard immunoperoxidase monolayer test (IPMA) for PCV detection with a Kappa value of 0.9. In conclusion, the blocking ELISA method established in this study has good specificity and high compliance rate, which can be used for later detection of PCV3 antibodies and provides technical support for epidemiological investigation and clinical diagnosis of PCV3.

The aim of this study was to develop a combined vaccine against porcine circovirus type 2 (PCV2) and Glaesserella parasuis (GPS), and to evaluate its immune protective effect in mice. We selected SH0165, a type 5 strain of GPS, as the vaccine strain. The G. parasuis ghosts were generated by chemically-induced lysis and the method is based on minimum inhibitory concentration (MIC) of sodium hydroxide (NaOH). Then, the ORF2 gene of PCV2 was cloned into the pEGFP-N1 vector. We developed a PCV2-GPS combined vaccine by using G. parasuis ghosts as vehicle for pEGFP-ORF2. Furthermore, four-week-old BALB/c mice were divided into commercial vaccine group, PCV2-GPS combined vaccine group, G. parasuis vaccine group, pEGFP-ORF2 group, PBS group, and blank group. The IgG antibody level against PCV2 and GPS, neutralization antibody against PCV2, PCV2 viral load of organs in mice, serum bactericidal activities, challenge protection efficacy, and level of cytokines (IFN-γ, IL-12) in serum were measured after immunization. The results showed that PCV2-GPS combined vaccine elicited significantly higher levels of IgG antibody response of PCV2 and GPS, neutralization antibody titer of PCV2, bactericidal activities and higher levels of IFN-γ than those of the control group. After PCV2 challenge, PCV2-GPS combined vaccine-immunized mice showed lower viral load in lymph nodes and lungs than the control group, and after challenge infection with GPS, immune protection efficacy of mice immunized was 70% (7/10). In conclusion, the PCV2-GPS combined vaccine significantly activated both humoral and cell-mediated immune responses, and inhibited the proliferation of PCV2 in mice, achieving a better protective effect in mice infected with GPS. The results provide biological materials and experimental data for the development of new, safe and efficient combined vaccines against porcine circovirus type 2 and Glaesserella parasuis.

The study was design to understand the molecular typing, pathogenicity and biological characteristics of Streptococcus suis meningitidis type 2 from swine. In this study, a strain of Streptococcus suis PX0923 was isolated from the brain tissue of diseased pigs, and its Serotype and molecular typing were identified by electron microscopy, 16S rRNA gene sequencing, multiplex PCR, PCR-RFLP and multiple site sequence analysis (MLST), the pathogenicity and drug sensitivity of the isolated strain were analyzed by virulence gene, biofilm formation ability, drug resistance gene detection, mouse infection test, antibiotic and Chinese medicine susceptibility test. The results showed that the isolated strain PX0923 belongs to Gram-positive bacteria. Under electron microscopy, continuous arrangement of oval shaped mycelium can be seen in chains. In terms of 16S rRNA gene sequence analysis and phylogenetic tree construction, the isolated strain was identified as S. suis type 2. MLST (multilocus sequence typing) analysis confirmed that it is type ST7 and merged with type ST1. The isolated strain PX0923 carried seven virulence genes, epf, mrp, sly, gapdh, fbps, orf2 and sao, and showed α hemolytic activity. After artificial infection of PX0923, it was found that the infected mice presented meningitis and other symptoms similar to the natural diseases, such as bleeding in the lung, spleen and liver of the infected mice. The LD₅₀ of the isolated strain was 1.08×10⁹ CFU per mice, these findings suggest that PX0923 may cause meningitis in infected animals. PX0923 carrys resistant genes aadA1 and bla-TEM. PX0923 is resistant to 12 drugs such as Gentamicin, kanamycin and tetracycline, and sensitive to traditional Chinese medicine scorpion and panax notoginseng. In summary, Streptococcus suis PX0923 from pigs was isolated as serotype 2, and MLST was classified as ST7, and carries a variety of virulence genes and drug resistance genes. Artificial infection of mice can cause multiple tissue damage such as meningitis in mice. It is sensitive to various antibiotics and traditional Chinese medicine scorpion, Panax notoginseng. The research results provide a reference basis for the prevention and treatment of streptococci and vaccine research. The results of this study provide a molecular biological basis for the epidemic and source analysis of S. suis, and provide a reference for the prevention and control of S. suis Meningitidis type 2 infection

The aim of this paper was to analyze the differences between the genome of Avibacterium paragallinella (Apg) of goose origin and chicken. In this study, three goose Apg and four chicken Apg isolates were sequenced and their genomic components were analyzed. The results of whole gene re-sequence showed that the genome fragment size of 7 isolates was 2.567 832-2.607 127 Mb, and the GC content was between 40.80% and 40.93%. The results of SNPs homology analysis showed that three strains isolated from geese and three strains of C-AP3 from chicken were clustered on the small branch of p4chr1 strain, and the other three strains from chicken were distributed on the branches of other domestic strains. In the gene homology analysis experiment, there are 76 specific genes of goose isolate and 37 specific genes of chicken isolate. The screening results of adaptive genes in geese showed that 79 genes were co-clustered, including 22 genes encoding known proteins, and the rest encoding putative proteins. In the screening test of chicken adaptive genes, 39 genes were clustered, including 10 encoding known proteins and 29 encoding putative genes. Through comparative genomics, found 118 genes related to Apg host adaptation, which laid a foundation for further elucidation of the molecular biological mechanism of Apg cross-species infection.

Corynebacterium pseudotuberculosis (Cp) infection highly impairs the production performance of livestocks, especially for the small ruminants (sheep/goats). This experiment was conducted to screen the essential oils against Cp, and to evaluate its antibacterial mechanism. Bergamot essential oil (BEO) was screened from four kinds of essential oils by K-B disk diffusion method, and the minimal inhibitory concentration (MIC) of BEO against different strains of Cp was determined by resazurin based broth dilution method. The effect of BEO on Cp infection of J774A. 1 macrophages were detected through in vitro experiments. The bacterial load in visceral organs, antimicrobial peptide genes expression in spleens and survival rates were tested to evaluate the protective effects of BEO on Cp-infected Kunming mice in vivo. Then the antibacterial mechanism of BEO was explored via the changes of cell morphology, cellular nucleotide leakage and the expression of virulence genes of Cp. The MIC value of BEO against Cp is 0.78-1.56 μL·mL^-1. The treatment of BEO significantly reduced the bacterial loads in kidney, liver and ascites of Cp-infected mice (P<0.01), up-regulated the mRNA expression levels of antibacterial peptides genes including Cramp, Bpifal 1, mBD2, mBD3 and mBD4 in spleens (P<0.01), decreased infection rate of Cp in J774A.1 macrophages and reduced mortality of Cp-infected mice (P<0.05). Scanning electron microscope analysis showed that the cell membrane was destroyed, the nucleotide was significantly leaked in Cp treated with BEO, and the mRNA expression level of virulence gene phospholipase D (pld) was significantly decreased in BEO treated Cp (P<0.01). BEO possesses strong antibacterial activity against Cp, and alleviates the infectious degree of Cp in mice, the mechanism of which may be related to the disruption of membrane integrity and inhibition of pld mRNA expression of Cp treated with BEO.

The aim of this study was to investigate the potential of canine adenovirus type 2 (CAV-2) as an oncolytic virus and to further explore the value of sperm adhesion factor (SPAM1) synergizing with CAV-2 to remodel the tumor microenvironment in tumor immunotherapy. Based on the P15-CAV-2 reverse genetic manipulation platform, the RED/ET homologous recombination system was used to construct an intermediate vector carrying the exogenous gene of SPAM1, and then the intermediate vector was used to deletion the E3 region of the CAV-2 backbone vector and replace the expression cassette of the exogenous gene of SPAM1. Then the synthetic primers was used to knockdown the kmccdB reverse screening expression cassette to construct the P15A-CAV-2-mCMV-SPAM1-SV40 polyA infectious clone plasmid and rescue the recombinant lysogenic viruses in the MDCK-E1A cell line, then the recombinant viruses' in vitro tumor lysis effect was validated. The results of sequencing and digestion showed that, a recombinant lyssavirus strain stably expressing the exogenous gene SPAM1 had been successfully constructed and rescued, and the IFA assay proved that the recombinant strain could stably express the exogenous gene at a high level, and the exogenous protein had the characteristics of diffusely distributed in the intercellular region. The recombinant lysovirus with the deletion of the E3 region expressing the exogenous gene SPAM1 was found to have a strong killing effect on the canine cancer cell line A72 by light microscopy and CCK8 assay. In conclusion, the current study successfully constructed a recombinant CAV-2 strain with a good tumor lysis effect, which lays the foundation for subsequent application in pet tumor therapy.

The present study aimed to investigate the role of angiotensin converting enzyme 2 (ACE2) in the infection of piglets with porcine epidemic diarrhea virus (PEDV). We investigated the expression of ACE2 before and after PEDV infection in piglets by transcriptomic analysis. Then the changes of ACE2 expression before and after PEDV infection were verified by using Porcine Intestinal Epithelial Cells (IPEC-J2) model. The changes of ACE2 mRNA and protein expression levels after PEDV infection in IPEC-J2 were detected by RT-qPCR and Western blot. And the PEDV replication levels were detected by RT-qPCR, Western blot, and TCID₅₀ after overexpression and inhibition of ACE2 expression. The results showed that the mRNA and protein expression levels of ACE2 were significantly downregulated after PEDV infection with IPEC-J2, which was consistent with the transcriptomic results. PEDV infection significantly increased in the ACE2 overexpression group and significantly decreased in the PEDV infection in the ACE2 suppression group. This study validated the role of ACE2 in the process of PEDV infection at the cellular level. PEDV infection was able to decrease ACE2 expression, overexpression of ACE2 in IPEC-J2 cells increased PEDV replication levels, and inhibition of ACE2 expression decreased PEDV replication levels.

The aim of this study was to establish a method for the isolation and in vitro culture of porcine primary peritoneal mesothelial cells (PMC) and preliminarily apply it to an in vitro infection model of Mycoplasma hyorhinis (Mhr). Porcine omentum was digested with 0.1% type Ⅰ collagenase to isolate primary porcine PMC. The cells were identified by morphology and immunological assays. The PMC cells were infected with Mhr. Thereafter, the morphological changes were observed and the cell activity damage was detected using the lactate dehydrogenase (LDH) method. Inverted phase contrast microscope observation showed that the isolated cells were pulled reticular at the early stage, and could grow up to fusion state 5-8 days later, with uniform size and polygonal slab stone-like appearance. Immunofluorescence analysis showed that the cells were positive for vimentin and cytokeratin-18, but negative for factor VIII-related antigen and CD45. Microvilli on the cell surface were observed under scanning electron microscope. It was confirmed that the isolated and cultured cells were PMC, and the purity was more than 95%. After Mhr infection, significant shrinking of the cells could be observed under light microscopy, and LDH release was significantly higher in the infected group of cells compared with the control group of cells. This study successfully established the method to isolate and culture the primary porcine PMC cells, and was initially used for Mhr infection in vitro, which could provide an in vitro cell model for studying the interaction mechanism between pathogens polyserositis-causing such as Mhr and the host.

The aims of this paper were to construct a mouse model of dust mite induced atopic dermatitis (AD) and a mouse model of asthma. In this experiment, the main allergens Der p 1, Der f 1, and Blo t 5 of the three main dust mite species (Dermatophagoides pteronyssinus, Dermatophagoides farinae, and Blomia tropicalis) were mixed in equal ratios to induce an AD mouse model and an asthma mouse model, respectively. The degree of sensitization in AD mice was also assessed by the levels of relevant inflammatory cytokines (IFN-γ, IL-4, IL-10, IL-17A) in skin and lung tissues, and total IgE serum levels. IgE levels, ear prick test, and hematoxylin-eosin staining of lung tissue (HE) were used to assess the asthma mouse model. The results showed that the skin of the sensitized area in the AD model showed crusting after breakage, and the changes of cytokine levels in the skin and lung tissues indicated that the in vivo response of the mice was in favor of Th2 response, and the IgE level was significantly higher than that of the normal group; The total serum IgE level was significantly increased in the mice of the asthma model group. The body temperature of the model mice decreased significantly after allergen excitation and did not recover the initial temperature after 90 min. In the ear prick test, the antigen attack increased the leakage of ear dye in the model mice, and the HE staining of lung tissue shows increased inflammatory cell infiltration in the model group of mice. This experiment successfully induced dust mite AD mouse model and dust mite asthma mouse model.

Mycoplasma bovis is one of the important pathogens causing bovine respiratory disease syndrome (BRD), which has caused immeasurable economic losses to the cattle industry. Mycoplasma bovis is resistant to most antibiotics, and there is no good drug to treat it at present. Therefore, vaccination is the most effective prevention and control method. The lack of a small animal evaluation model is one of the important constraints for the development of Mycoplasma bovis vaccines. The small animal challenge model was established to provide a certain research basis for the follow-up vaccine research and preparation, and reduce the scientific research cost of follow-up experiments. In order to establish a small animal model of Mycoplasma bovis infection, two-month-old Japanese big-eared white rabbits were used in this study and divided into 4 groups:1) Mycoplasma bovis strain HB0801 was challenged alone, 2) Mycoplasma bovis strain HB0801 was challenged with dexamethasone, 3) Mycoplasma bovis strain HB0801 was challenged with KLH and thioglycollate medium. 4) Blank control group. The excretion of Mycoplasma bovis was detected by PCR, and the level of antibody in serum was detected. The lungs were taken to make pathological sections after 31 days. The results showed that both the detection rate of M. bovis and the antibody against M. bovis were the highest in the group challenged with M. bovis HB0801 strain after injection of dexamethasone. The detection rate of M. bovis was 42.11%, and 91.67% of the animals tested positive for M. bovis. It was significantly higher than other groups. Except for the blank control group, the other challenge groups showed alveolar wall thickening, macrophage infiltration,and different degrees of fibrinous exudation in the lungs, which were consistent with the pathological characteristics of interstitial pneumonia caused by Mycoplasma bovis. In this study, the infection and evaluation model of Mycoplasma bovis HB0801 strain challenge in Japanese big-eared white rabbits was established, and the best modeling effect was achieved when the immunosuppression was induced by dexamethasone.

This experiment was conducted to study the effect of the Chinese herbal medicine Radix dichroa powder (RDP) on the anticoccidial effect and immune function of chick infected with Eimeria tenella at different developmental stages. Six major groups were set up in the trial, namely, 1 day before the attack, the day of the attack, day 2, day 3, day 4 and day 5 dosing time design groups, and each dosing time point included three treatment groups, namely group I (healthy control group) and group II (infected control group), fed the basal diet, and group III (RDP recommended dose group), fed at 0.1 g·kg^-1 in the basal diet and dosed continuously for 4 days, with 10 chicks in each group. At 16 days of age, 5×10⁴ sporulated oocysts of Eimeria tenella were administered orally to each chick in group II and group III. The results showed that compared with group I, the average daily gain (ADG) of group III was not significantly different (P> 0.05) in the early stage of coccidia development (1 day before and on the day of attack) and significantly decreased (P<0.05) in the late stage of development, while group II had significant decrease (P<0.05) in ADG throughout the period of coccidia development, and the decrease was greater than that of group III. Compared with group Ⅰ, the average daily feed intake (ADFI) decreased in both group Ⅱ and group Ⅲ, with the most significant decrease in group Ⅱ and improvement in group Ⅲ. Compared with group II, RDP significantly (P<0.05) reduced the number of fecal oocysts per gram (OPG) in infected chicks. Coccidia infection damaged the integrity of the cecum mucosa and thickened the cecum muscle membrane. The RDPof Chinese medicine reduced the mucosal damage caused by coccidial infection. Compared with group II, the addition of RDP in the diet reduced the cecum lesion score (P<0.05). The anticoccidial index (ACI) of group III were198.83, 190.23 and 165.49 on the 1st day before, the day of and the 2nd day of the attack, respectively, and 150.53, 155.00 and 146.18 on the 3rd, 4th and 5th day of the attack, respectively. The above results indicated that RDP had good anticoccidial effect when administered in the early stage of coccidial development, however as the coccidia developed in the body later, the therapeutic effect of RDP weakened. Compared with group II, RDP increased the levels of immunoglobulins IgA, IgM and IgG in blood, and improved the index of spleen and bursal. The results suggested that RDP improved the growth performance of infected chicks, significantly reduced the OPG and cecum lesion scores of infected chickens, and enhanced the immune function of infected chicks.

This study aimed to explore the immunomodulatory mechanism of Astragalus polysaccharide (APS) on HD11 in chicken macrophages by analyzing the effects of APS on the transcriptomes and metabolomes of HD11. A blank control group (CONT group) and an APS treatment group (A50 group) were set up, in which HD11 cells of the A50 group was treated with complete culture medium contain 50 μg·mL^-1 APS for 12 h; and HD11 cells of the CONT group were cultured in complete culture medium for 12 h. Metabolomics and transcriptomic techniques were used to analyze the differential genes and differential metabolites in CONT and A50 treated cells, and the combined analysis of the two omics was performed. Results:A total of 845 differential genes were detected in the transcriptome analysis between the A50 treatment group and the CONT group, including 415 up-regulated genes and 430 down-regulated genes. Differentially expressed genes were mainly concentrated in cytokine-cytokine receptor interaction, toll-like receptor signaling pathway, phagosome and AGE-RAGE signaling pathways, among which cytokine-cytokine receptor interaction and toll-like receptor signaling pathways were the most significant. In the metabolome analysis and comparison between the A50 treatment group and the CONT group, a total of 89 differential metabolites were screened, of which 70 were down-regulated and 19 were up-regulated. In the combined analysis of transcription and metabolism of the A50 treatment group and the CONT group, the top ten trusted pathways were purine metabolism, carbon metabolism, pentose phosphate pathway, glutathione metabolism, histidine metabolism, amino sugar and nucleotide sugar metabolism, α-homolenic acid metabolism, ABC transporter, iron death and tyrosine metabolism. APS may exert immunomodulatory effects on HD11 cells through the metabolic reprogramming of toll-like receptor signaling pathway, glycolysis and pentose phosphate pathway.

The aim of this study was to investigate the protective effect and mechanism of action of hesperidin (HDN) on high-fat feeding-induced hepatic oxidative stress injury in mice. In this study, 18 male C57BL/6 (body weight 20-23 g) mice were randomly divided into a control group, a high-fat diet (HFD) group, and a high-fat diet + hesperidin (HFD+HDN) group (300 mg·kg^-1) of six mice each. The mice in the control group were fed with basal diet (10% fat, 70% carbohydrate, 20% protein); mice in the HFD group were fed with high-fat diet (60% fat, 20% carbohydrate, 20% protein); and mice in the HFD+HDN group were fed with high-fat diet while HDN was administered by gavage at 300 mg·kg^-1 per day. After 16 weeks, mice were anaesthetised by intraperitoneal injection of 3% sodium pentobarbital (60 mg·kg^-1) and then blood was collected from the eyeballs. After blood collection, mice were decapitated and dissected for liver tissues. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), which are indicators of hepatic function, were detected using kits in the blood; The levels of malondialdehyde (MDA), activities of total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px), and the level of total antioxidant capacity (T-AOC), which are indicators of oxidative stress, were detected using the kits in the liver tissues;The liver tissues were also collected for transcriptomic sequencing to screen differentially expressed genes between the HFD and HFD+HDN group for KEGG pathway enrichment analysis, with P<0.05 as the threshold for significant enrichment, and the most significant signalling pathway affected by HDN intervention was screened accordingly. Genes with significant changes from the screened signalling pathway were selected to validate the transcriptomics results by qRT-PCR and protein immunoblotting methods, the relative content of mtDNA and the relative expression of the mitochondrial outer membrane protein(TOMM20), as well as the liver ATP content were detected. The results showed that, compared to the HFD group, the HDN intervention improved high-fat feeding-induced liver damage, significantly reduced ALT and AST activity in the blood of mice (P<0.01); Significantly reduced liver MDA levels in mice (P<0.01); Significantly increased T-AOC, T-SOD and GSH-Px levels (P<0.05); Compared to the HFD group, the HFD+HDN group of which 889 genes were up-regulated (P<0.05) and 405 genes were down-regulated (P<0.05); KEGG enrichment analysis screened the oxidative phosphorylation pathway as the most significantly up-regulated signaling pathway (P<0.000 1), Compared with the HFD group, the relative mRNA expression of Cox8b, Cox6a2, Gm10231, mt-Atp8, mt-Nd4l, Gm11237, Ndufb8, and Ndufb10 were significantly up-regulated in the liver tissues after the HDN intervention (P<0.05), and the relative mRNA expression of Atp6v0d2, Cox6c2 were significantly down-regulated (P<0.05). Protein expression of Cox6a2, Ndufb8, Ndufb10 were significantly increased (P<0.05), and the expression of Atp6v0d2d protein expression were significantly decreased (P<0.01), which was consistent with the transcriptomics results; And the relative expression, relative mtDNA content, and ATP content of TOMM20 were significantly higher after HDN intervention (P<0.05) compared with that of the HFD group. These results indicated that HDN reduces high-fat feeding-induced oxidative stress in mouse liver by modulating the oxidative phosphorylation pathway.

The present study was designed to investigate the therapeutic effect of lytic bacteriophage S13-21 on broilers infected with Salmonella enteritidis, as well as its effect on growth performance, antioxidant capacity, and cytokine and immunoglobulin content. Total of 72 1-day-old AA broilers were randomly distributed into 4 groups with 6 replicates per group and 3 chickens per replicate, namely blank control group (C), Salmonella infection group (S), phage treatment group (S+P) and antibiotic treatment group (S+A). The broilers in groups S, S+P and S+A were infected with Salmonella enteritidis by intraperitoneal injection, and the challenge dose was 200 μL per broiler (1.0×10⁹ CFU·mL^-1). Six hours later, the chickens in groups S+P and S+A were treated by intraperitoneal injection with phage dose of 1 mL per broiler (1.0×10¹² PFU·mL^-1) and antibiotic dose of 200 μL per broiler (gentamicin sulfate, 800 IU per broiler). The treatment lasted for 7 days with an interval of 12 hours. On the fifth day after treatment, broilers were weighed by repetition, and 6 chickens were randomly selected from each group for slaughter. The results showed that:1) At the end of the experiment, the survival rate of broilers in group S was only 59%, compared with group C, the body weight was significantly decreased (P<0.01), and spleen weight increased but the difference was not significant (P>0.05). The activities of T-SOD and GSH-Px in serum and other tissues were extremely significantly decreased (P<0.01), MDA and immunoglobulin content were extremely significantly increased (P<0.01). 2) Compared with group S, the survival rate of broilers in group S+P was increased by more than 25%, and the bacterial load of Salmonella in ileum and cecum were extremely significantly decreased (P<0.01). The contents of antioxidant indexes, cytokines and immunoglobulin in serum and other tissues (except some indexes in ileum) were significantly different from those in S group (P<0.05). In conclusion, Salmonella enteritidis can damage the intestinal mucosal barrier of broilers, resulting in growth retardation, tissue damage and systemic inflammatory response. Phage therapy can specifically eliminate Salmonella enteritidis infection, significantly improve the antioxidant capacity and intestinal mucosal barrier, relieve oxidative stress and inflammatory response, and has a strong protective effect. To a certain extent, it can improve the survival rate and restore the growth of chickens.

In this study, we successfully established mouse and canine diabetes models under the combined effect of high-fat diet (HFD) feeding and streptozotocin (STZ) injecting, aiming to investigate the effect and mechanism of Mitoquionl Mesylate (MitoQ) pretreatment of adipose-derived mesenchymal stem cells (ADMSCs) in enhancing the therapeutic effect of diabetic animal models. At the cellular level, ADMSCs in normal culture were pretreated with MitoQ at a concentration of 1 μmol·L^-1 in α-MEM+ cell culture medium, and the morphological changes, growth, proliferation, migration and antioxidant capacity of MitoQ-ADMSCs and ADMSCs were examined respectively. At the animal level, forty-eight 8-week old Kun-Ming(KM) male mice and 12 Chinese Field canines were selected and randomly divided into 4 groups:normal control group (NC group), diabetes model group (Diabetes group), ADMSCs treatment group alone (ADMSCs group) and combined treatment group of MitoQ pretreated ADMSCs (MitoQ-ADMSCs group). Diabetes models were made in mice and canines except for the NC group, the animals in the ADMSCs and MitoQ-ADMSCs groups were transplanted with 2×10⁶ cells each mice, and 1×10⁷ cells each canines once a week, for 3 consecutive weeks. Changes in clinical indicators were continuously monitored during modeling and treatment, and samples were collected 1 week after treatment and analyzed for serological levels, histological levels, oxidative stress levels and serum metabolomic changes in canines. The results showed that MitoQ treatment of ADMSCs did not affect their cell morphology, but could promote their growth, proliferation and migration ability, as well as enhance their antioxidant and anti-aging abilities. The treatment of ADMSCs and After transplantation of ADMSCs and MitoQ-ADMSCs, it was found that MitoQ significantly promoted the ability of ADMSCs to lower blood glucose, and improved pancreatic β-cell function and the body's blood glucose regulation ability. Staining of pancreatic and liver tissues revealed that ADMSCs could reduce tissue damage, promote insulin secretion, and reduce impaired hepatic glycogen synthesis and fibrosis levels, however the effects were limited, while MitoQ was able to enhance these effects more significantly. Detection of indicators related to oxidative stress in pancreatic tissues showed that MitoQ treatment of ADMSCs could effectively enhance antioxidant capacity of the body and reduce oxidative damage, thus enhancing the therapeutic effect. Metabolomic analysis of canine serum revealed that MitoQ treatment of ADMSCs significantly promoted the expression of antioxidant-related metabolites in the body. The results revealed that MitoQ could improve the antioxidant capacity of ADMSCs and accelerate the repair of tissue damage resulted from diabetes, thus beneficial for the treating of metabolism diseases in mice and canines.