With increasing scaled cattle cultivation, estrus accurate identification of individual has become the management focus of breeding. The performance of behavior and physiological changes, which are important markers in distinguishing estrus cows and determining the optimal insemination time, are closely related to itself follicular development. A better knowledge of the regularity and mechanism of these markers by modern technology during estrus cycle would help to improve the efficiency of automated estrus identification and become the focus in this field. Therefore, this paper reviews the research progress on estrus behavior and physiological characteristics of cow, analyzes the existing related issues in estrus identification techniques, and summarizes the recent research on identification of specific compounds of saliva, urine and milk in estrus cows, aiming to provide reference for developing newly efficient automated estrus identification technologies.

Cryopreservation of embryos is important for long-distance embryo transfer and conservation of genetic resources. However, porcine embryos are sensitive to low temperatures due to their high cytoplasmic lipid content, which increases the difficulty of their preservation. Studies have indicated that techniques such as reducing lipid content and optimizing medium composition can significantly enhance the efficiency of porcine embryo cryopreservation. Additionally, protecting the cytoskeleton and restoring mitochondrial function are also crucial strategies that contribute to this advancement. These improvements collectively foster the broader application and sustainable development of this critical technology.Therefore, this article introduces embryo cryopreservation and summaries multiple methods and measures to improve embryo freezing efficiency. Meanwhile, the writers'team will focus more on the mechanism of cryopreservation of porcine embryos by discussing the intrinsic properties of embryos and the effects of cryopreservation on transcriptome alteration to help people better understand and advance research on porcine embryo cryopreservation.

Estrus identification is an important link in cow breeding management.Timely and accurate identification of estrus cows has important significance to improve reproduction performance.However, silent estrus cows can't be identified by using pedometer or smart collar, which has become the main limiter of further improving reproductive efficiency.To this end, this paper reviews the characteristics of silent estrus cows, and analyzes the involving inducements and regulation mechanisms in previous studies.Analysis indicates that booming growth of internet of things, electromechanical, and sensing technologies provide facilities for revealing mechanism and laws of physiological and behavioral indices in estrus cows.A revelation of these mechanism and regularities will be helpful to promote the development of highly effective and accurate automated estrus identification technology, which will break through the bottleneck of silent estrus identification technology.

The glucose sensing receptor (T1R2/T1R3) in the gastrointestinal tract of ruminants can detect the presence of glucose, transmit sweet signals to the central nervous system, and promote the secretion of gastrointestinal hormones and up-regulate the expression of glucose transporters through gastrointestinal hormones. Glucose transporters mainly consist of SGLT1 and GLUT2. The former one is primarily responsible for transporting glucose into gastrointestinal epithelial cells, while the latter one basically involves in transporting glucose from gastrointestinal epithelial cells into adjacent blood vessels. Glucose sensing receptors and transporters work together to fulfill the glucose absorption process in the gastrointestinal tract. In this paper, the functional characteristics of glucose sensing receptors and transporters in the gastrointestinal tract of ruminants and their relevant regulatory mechanisms were reviewed, in order to provide insights and references for improving the utilization efficiency of glucose in ruminants.

Trypanosomes are a type of blood-borne protozoa transmitted by vectors, which can infect humans and animals and cause trypanosomiasis being responsible for huge disease burdens. The surface of trypanosomes is a dense "coat" formed by a single variable surface glycoprotein (VSG), which regularly undergoes replacement at each stage of infection to evade host immune killing. The genomes of trypanosomes contain hundreds to thousands of VSG genes and pseudogenes, and parasites employ precise expression regulation mechanisms to exclusively express one type of VSG at each specific infection stage. VSGs have good antigenicity, which is mainly determined by the immunodominant epitopes, glycosylation modifications, subdomains and their conformation located in the top lobe of the N-terminal domain (NTD). VSG-specific IgM responses play an important role in trypanosome clearance, but the functions of IgG-mediated immune lysis are very limited. Host specific IgG immune responses become stereotyped, targeting only a limited set of immunodominant epitopes in each VSG, thus leading to reduction of cross reactivity with different VSGs. This article reviews the updates of VSG in expression regulation, protein structure, immunogenicity and immune responses, providing a theoretical basis for a profound understanding of the pathogenic mechanisms of trypanosomes and the development of effective prevention and control measures against trypanosomiasis.

Flow virometry (FVM) represents a cutting-edge technological approach leveraging flow cytometry for the precise detection and analysis of individual virus particles and their unique characteristics. This innovative method offers a novel means of detecting, quantifying, and characterizing individual virus particles. FVM enables the determination of various parameters including the concentration of intact virus particles, the abundance of specific target antigens present on the virus surface, and the relative diameter of the viruses. Within the field of technique, FVM extends beyond virus analysis, allowing for the effective detection and characterization of exosomes and microvesicles originating from cells. With the continuous development and optimization of instruments, fluorescent dyes and labeling strategies, FVM has been widely used and studied. This paper summarizes the development history of FVM, as well as the common labeling methods and application fields of viruses and other microparticles, aiming to provide reference for the further application of FVM in virology, immunology, biomedics and other research fields.

Pasteurella multocida (P. multocida) can infect a wide range of animals, causing hemorrhagic septicemia or infectious pneumonia. Iron is an essential nutrient for growth, colonization, and proliferation of P. multocida during infection of the host, and competition for iron ions in the host is a critical link in the pathogenesis of this pathogen. In recent years, a series of significant research advances have progressed in the iron uptake system of P. multocida, as well as its occurrence and regulatory mechanisms. The mechanisms of iron uptake by transferrin, heme receptors and siderophore, and the mechanism of expression and regulation of the P. multocida iron uptake system are all described in this paper. Aiming to provide systematic theoretical knowledge for the study of the molecular pathogenesis of the P. multocida iron uptake system and to spark new ideas for the investigation and development of molecular target drugs and subunit vaccines of P. multocida.

A significant portion of the global population is affected by allergies to cats, with symptoms ranging from rhinitis to asthma and respiratory disorders, which have a serious impact on the lives of allergy sufferers. Fel d 1 is currently recognized as the main allergen in cats. It is a secretory globulin primarily produced in the sebaceous and salivary glands of cats, distributed onto their fur during grooming and eventually dispersed into the air. To address the challenge of cat allergies in humans, researchers have attempted various approaches to reduce the secretion of Fel d 1 or control its spread. These efforts aim to alleviate or eliminate allergic reactions in cat allergy patients, including methods such as cleaning cat fur, maintaining a clean indoor environment, breeding hypoallergenic cats, and administering vaccines to cats. This paper provides an overview of the research progress on feline allergens, management and control of cat allergies, and treatment measures.

Infectious respiratory diseases caused by animal pathogens pose a major challenge to livestock and poultry breeding and pet medical care. Infectious diseases caused by pathogens not only endanger animal health, but also affect human public health safety. At present, the development of new veterinary drugs for these diseases is facing bottlenecks. This paper focuses on summarizing the research progress of veterinary drugs for animal pathogenic infectious respiratory diseases in China, and comprehensively summarizes the characteristics of veterinary drugs for animal pathogenic infectious respiratory diseases in China, such as types, research and development time, medication rules, and veterinary drug dosage forms, in order to summarize the research and development progress of veterinary drugs for animal pathogenic infectious respiratory diseases in China. At the same time, according to the characteristics of the categories, dosage forms and clinical applications of chemical drugs and traditional Chinese medicine in animal respiratory veterinary drugs, this paper analyzes that the future research and development trend of new veterinary drugs in China needs to focus on the efficient application of existing drugs, focusing on the optimization of drug dosage forms and the development of multi-target drugs. The purpose of this paper is to provide a medication plan for the effective prevention and control of animal pathogenic infectious respiratory diseases, and to provide a reference for the future research and development direction of veterinary clinical new veterinary drugs.

Chylothorax is a condition in which a rupture or obstruction of the thoracic duct is caused by a different etiology, which subsequently leads to the entry of coeliac into the thoracic cavity. Surgical intervention is usually required when conservative treatment fails. Because thoracic duct anatomy varies widely in different cats, contrast imaging of thoracic duct branches prior to ligation thoracic duct is an essential step in surgical planning. With the gradual application of computerized tomography (CT) imaging technology in canine and feline lymphangiography, a variety of minimally invasive preoperative CT lymphangiography methods have been proposed. This article summarizes the current CT lymphangiography protocols in the diagnosis and treatment of feline chylothorax for clinical reference.

Immunochromatography is a typical rapid diagnostic technology based on antigen-antibody-specific immune reaction, using different materials as tracer markers, which is rapid, specific and inexpensive, and has wide application value for on-site detection of common viral diseases in pigs. In this paper, based on the principle of immunochromatographic technology, the current common immunochromatographic techniques were categorized according to different signal manifestation modes, colorimetric immunochromatographic techniques, fluorescent immunochromatographic techniques of the representative markers were introduced in detail, the application of immunochromatographic techniques in the rapid detection of common porcine viral diseases was reviewed, and the advantages and challenges of immunochromatographic test strips with different markers were compared, with a view to providing ideas for the development of rapid detection techniques for porcine viral diseases.

The study aimed to explore the effect of structural variation (SVs) in MYH gene family on muscle growth for longissimus dorsi muscle (LDM) of Wuzhishan pigs, and to provide a theoretical basis for further understanding of the rule and molecular mechanism of muscle growth. The study subjects were Wuzhishan pigs, and Large White pigs were used as control group. Three Wuzhishan pigs (4-month-old sows, average weight was 14.2 kg) and three Large White pigs (4-month-old sows, average weight was 58.0 kg) were raised in the same environment for one week before slaughter, fed sufficient food and water, and all pigs were starved for 24 hours before slaughter. LDM of 6 pigs were collected, complete DNA was extracted, library was built and sequenced by PacBio platform, the sequencing data of each sample was about 40 Gb. The sequencing data was aligned to reference genome. Filter condition: 50 bp≤SVs≤10 kb; Total RNA was extracted from longissimus dorsi muscle of each pig, transcriptome sequencing was performed, and gene expression was calculated. The distribution position and number of SVs on chromosomes of Wuzhishan pig and Large White pigs were counted, the specific SVs of Wuzhishan pigs were screened, and the phylogenetic tree of MYH gene family was constructed, the structure domains were predicted, and the correlation between MYH gene expression and muscle fiber indicators was analyzed. The results showed as follows: 1) There were 7 specific MYH genes in Wuzhishan pigs, which were mainly distributed in chromosomes 3, 5, 6, 12 and 13, and chromosome 12 contained 3 MYH genes; 2) Phylogenetic tree showed that 7 MYH genes of Wuzhishan pigs differentiated into 2 branches, encoding non-muscle myosin and muscle myosin respectively, MYH9 and MYH11 genes, MYH1 and MYH4 genes had the highest homology; All 7 MYH genes contained the Motor domain, and 5 MYH genes contain the Myosin-N domain. The head domain of MYH gene (Myosin-N and Motor domain) was relatively conservative, and the middle and tail domain of MYH gene were different. The MYH1, MYH4 and MYH9 genes of Wuzhishan pigs contained SVs, while those of Large White pigs did not. Pearson coefficient statistics showed that the existence of SVs in MYH genes might be related to gene expression. The correlation between the expression levels of these three MYH genes and muscle fiber indexes was detected by both RNA-seq and Q-PCR. MYH1 gene expression was positively correlated with muscle fiber density and number, MYH4 gene expression was negatively correlated with muscle fiber density and number, MYH9 gene expression was negatively correlated with muscle fiber density and positively correlated with diameter. In summary, the largest number of MYH genes distributed on chromosome 12, the MYH gene family is mainly divided into two branches: non-muscle myosin and muscle myosin. The head domain is relatively conservative, and the difference between the middle and tail domain may be the main reason for the differentiation of MYH gene family. The expression levels of MYH1, MYH4 and MYH9 genes were significantly correlated with muscle fiber density, indicating that these genes might affect LDM growth through regulating the muscle fiber density. This study not only expands the content of molecular mechanism of muscle growth and development, but also provides reference for optimizing genetic breeding model related to structural variation.

The aim of this study was to explore the expression characteristics of coding and non-coding RNAs in the pituitary gland of hermaphroditic pigs, thereby providing supporting data for the analysis of the molecular mechanisms underlying pituitary dysfunction in these pigs. Three five-month-old normal sows and three hermaphroditic pigs were used for the study. Serum hormone assays and whole transcriptome sequencing of pituitary tissues were carried out to analyze and identify differentially expressed mRNAs, lncRNAs, and miRNAs in the pituitary of hermaphroditic pigs. Additionally, a competing endogenous RNA (ceRNA) network was established to elucidate the regulatory relationships of related genes in the pituitary of hermaphroditic pigs. Disrupted serum hormone secretion and abnormal pituitary function were observed in hermaphroditic pigs. Comparison with normal sows, 1 451 differentially expressed mRNAs, 277 differentially expressed lncRNAs, and 17 differentially expressed miRNAs were identified in the pituitary gland of hermaphroditic pigs, which were enriched in biological pathways, such as the MAPK signaling pathway, the progesterone-mediated oocyte maturation, and the PRL signaling pathway. Through the analysis of the ceRNA network, competitive combinations of TCONS_00175477-novel_265-CCNB3, TCONS_00134726-novel_265-ZNF366 and TCONS_00212783-novel_265-ZNF366 were identified possibly related to abnormal pituitary hormone secretion in hermaphroditic pigs. In summary, the differentially expressed mRNAs, lncRNAs and miRNAs in the pituitary gland of hermaphroditic pigs were revealed, thereby ceRNA networks were constructed, which may be involved in regulating hormone synthesis and secretion in this study. All these were aid to support the elucidation of the molecular mechanisms underlying the dysfunction of the pituitary gland in hermaphroditic pigs.

The aim of this study was to understand the molecular pathological mechanisms of subcutaneous adipose dysfunction in pigs susceptible to metabolic diseases, and to investigate the correlation between disturbances of energy metabolism in subcutaneous adipose tissue and epigenetic regulation. The study selected wild-type male Bama pigs with similar weight and good health at 6 months old, as well as transgenic pigs susceptible to metabolic diseases, and divided them into 4 groups after being induced by a high-fat high-sugar diet (HFHSD) for 3 or 12 months. The wild-type groups were named WT-3 (n=5) and WT-12 (n=5), while the transgenic groups were named TG-3 (n=8) and TG-12 (n=4). Firstly, the animals were evaluated by serum biochemistry, containing the concentrations of triglycerides, free fatty acids, leptin, and adiponectin. Then, the animals were subjected to histopathological evaluation, and molecular features and enriched signaling pathways of metabolic disorders were obtained through transcriptome sequencing. Subsequently, key genes, metabolites, and epigenetic modifications were detected using qPCR, Western blot, and ELISA. The results of serum biochemical assay and histopathological evaluation showed that both transgenic and diet induction could cause adipose tissue dysfunction, and the pathological injury of transgenic combined diet induction group was more serious. Transcriptome sequencing results showed that adipose tissue dysfunction was characterized by impaired mitochondrial oxidative phosphorylation and decreased glucose and lipid metabolism and protein synthesis in adipose tissue. qPCR results showed that the expression of mitochondria-encoded genes was significantly down-regulated in TG-12 group. WB results showed that the key genes regulating glucose and lipid metabolism, ACLY, ACSS2 and FASN, were significantly down-regulated in TG-12 group. ELISA results showed that the content of acetyl-coA, the key intermediate metabolite, was decreased. Moreover, WB verified that the reduction of histone acetylation, which has a wide regulatory role in gene expression, may be the main cause of adipose dysfunction. The use of metabolic disease susceptible pigs reveals that reduced subcutaneous adipose mitochondrial function exerts extensive gene expression inhibitory effects through down-regulation of acetyl coenzyme A content and histone acetylation levels via epigenetic modification, providing a model and reference data for the treatment of obesity-associated subcutaneous dysfunction diseases in humans.

The study aimed to investigate the potential regulatory mechanism of nesting ability of laying hens. The 12 300-day-old Chengkou mountain chickens with good body condition and the same body weight were used in this study. Among them, 3 individuals were in normal laying period, and the remaining 9 individuals were in nesting period for 10, 20 and 30 days, respectively. The ovarian tissues of laying hens in each group were collected for anatomical observation. Normal ovary (NO) and atrophic ovary (AO) of laying hens were analyzed by RNA-seq and iTRAQ sequencing technology. Differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) were screened and functional enrichment analysis was performed to identify candidate genes related to nesting in poultry. The results showed that ovary atrophy and follicle atresia occurred in nesting hens. A total of 930 DEGs were identified in ovarian tissues, of which 430 genes were up-regulated and 500 genes were down-regulated. Meanwhile, 546 DEPs were identified, of which 178 proteins were up-regulated and 368 proteins were down-regulated. Through functional annotation and enrichment analysis, it was found that these DEGs and DEPs were significantly enriched in ECM receptor interaction, adhesion and PI3K-Akt signaling pathways. Finally, 7 candidate genes for nesting ability COMP, FN1, ITGA8, THBS1, COL4A2, COL4A1 and COL1A1 were selected by combined transcriptome and proteome analysis. In summary, key candidate genes for nesting ability were identified by transcriptome and proteome analysis, and a regulatory network for ovarian development in nesting poultry was constructed. This study provides a theoretical reference for further understanding the molecular regulation mechanism of ovarian atrophy at nesting stage, enriches the candidate genes regulating poultry nesting traits, and provides theoretical basis for genetic improvement and molecular breeding research of laying hens.

This study aimed to analyze the genomic variations and population structure of Large-tailed Han (LTH) sheep to provide guidance for scientific conservation and sustainable utilization. Whole-genomic sequencing data from 170 LTH sheep (66 rams, 104 ewes) was used to investigate the genomic variations, population structure and linkage disequilibrium by GATK, Manta, Plink and other softwares. The 1 599.56 G clean data was obtained and on average, 9.409 G clean data per sheep. In the LTH sheep population, 50 276 079 SNPs and 7 240 540 InDel were found, which were mostly distributed in intergenic and intronic region. The interchromosomal translocation (CTX) was the most common type of structural variation (SV) in the genome of LTH sheep, on average, 415.82 CTX per sheep, and also mainly distributed in the intergenic region. The copy number variation (CNV) was mostly distributed in exons, 175 CNV per sheep on average. The result of principal component analysis showed that individuals were relatively scattered and poorly clustered. According to the genetic relationship, phylogenetic tree and population structure, the 170 LTH sheep were clustered into 6 families, and there was a significant difference in family size and body size. The results of cluster analysis showed that ancestry component of some individuals was single. The linkage disequilibrium analysis showed that LTH sheep population had a relatively fast attenuation rate. The selected genes during domestication were mainly related with lipids metabolism and thermogenesis. In conclusion, LTH sheep population contains 6 families and has rich genetic diversity and remarkable conservation effect. It is suggested to expand the number of small families and to prevent inbreeding in the big families to keep a balanced population size and structure and focus on the utilization of LTH sheep.

The study aimed to investigate the structural characteristics and genetic distribution of the ATG16L2 gene promoter region in Hainan Black goat, providing a theoretical basis for further exploring the expression regulation mechanism and function of this gene. This study focused on 200 Hainan Black goats, constructing a DNA pooling strategy. The polymorphism of the ATG16L2 gene promoter region in Hainan Black goats was initially screened using Sanger sequencing. PCR-RFLP technology was applied for genotyping the 200 Hainan Black goats. Linkage disequilibrium analysis was performed on the identified SNP loci to construct haplotypes. Bioinformatics methods were utilized to analyze the impact of SNP loci on the expression of the ATG16L2 gene in Hainan Black goats. The promoter region of the ATG16L2 gene in Hainan Black goats contained 3 linked single nucleotide polymorphisms (SNPs): SNP1 (g.30667970T > C), SNP2 (g.30668540T > C), and SNP3 (g.30668664C > T). Both SNP1 and SNP2 exhibited moderate levels of polymorphism, whereas SNP3 showed low levels. Notably, all three SNPs conformed to the principles of Hardy-Weinberg equilibrium(P > 0.05). The haplotype analysis revealed that the frequencies of haplotype H1, H2, H3, and H4 were 0.321, 0.304, 0.271, and 0.097, respectively, with H1 (CGC) being the predominant haplotype. The bioinformatics analysis results showed that there were 3 promoters, 4 CpG island regions, 2 repetitive elements LINE2 (—1 989-—1 826 bp, —562-—426 bp), hAT-Charlie (—1 804-—1 511 bp), and 5 CCAAT-Box, 13 CAAT-Box, 10 CGCG-Box, 11 GATA-Box and 2 TATA-Box of the ATG16L2 gene in Hainan Black goat. The integrated prediction from various online tools suggests that the above SNPs may affect the expression regulation of the ATG16L2 gene in Hainan Black goats by influencing the changes in transcription factors in the gene's promoter region. In this study, 3 SNP loci were identified in the promoter sequence of the ATG16L2 gene in Hainan Black goats. Among these, SNP1 and SNP2 exhibited moderate polymorphism, while SNP3 showed low polymorphism. It is predicted that these SNPs may influence transcription factor binding, thereby regulating gene expression. This provides a theoretical basis for further exploration of the function and regulatory mechanisms of the ATG16L2 gene.

The study aimed to understand the population structure and genetic differences of Ganzhou Muscovy duck. The blood samples of 20 Fujian Muscovy ducks (FJ), 20 Guizhou Tianzhu Muscovy ducks (TZ), 20 Northwest Jiangxi Muscovy ducks (GXB) and 40 Ganzhou Muscovy ducks (GZ) were collected for whole genome resequencing with a depth of 10×. Subsequently, sequence data of 15 Peking ducks (PK), 15 Mallard ducks (MD), 10 Cherry Valley ducks (CV) and 2 French Muscovy ducks (FF) were downloaded from the NCBI database. Principal component analysis, phylogenetic tree construction, heterozygosity analysis, population differentiation analysis, gene flow analysis and population linkage disequilibrium analysis were carried out using the sequence data of the above 8 populations. The results showed that the French Muscovy duck, Tianzhu Muscovy duck and Northwest Jiangxi Muscovy duck could not be completely distinguished according to the second and third principal components in the principal component analysis, and there might be hybridization among them. In phylogenetic tree analysis, the populations of Ganzhou Muscovy duck and other breeds of ducks were obviously clustered into different branches, which showed obvious genetic uniqueness and were different from other populations. In the analysis of heterozygosity, the observed heterozygosity of all populations except French Muscovy duck was lower than expected heterozygosity, indicating that the genetic diversity of 7 populations including Ganzhou Muscovy duck was decreased. According to the analysis of population differentiation, the degree of differentiation between Ganzhou Muscovy duck and other populations, from high to low, was as follows: Beijing duck, Cherry Valley duck, Mallard duck, French Muscovy duck, Fujian Muscovy duck, Tianzhu Muscovy duck, and Northwest Jiangxi Muscovy duck. In gene flow analysis, there was no single gene transfer between Ganzhou Muscovy duck and French Muscovy duck, Tianzhu Muscovy duck, and Fujian Muscovy duck, and there was also not much gene exchange with other populations. In the analysis of population genetic structure, when K=3, Ganzhou Muscovy duck was identified as an independent population, showing its unique genetic background. In the analysis of linkage disequilibrium, Ganzhou Muscovy duck showed the lowest degree of linkage disequilibrium, indicating that it had low inbreeding degree, higher genetic diversity and stronger environmental adaptability. In conclusion, the Ganzhou Muscovy duck is distinct from the other 7 duck populations and can be regarded as an independent genetic resource.

This study aimed to investigate the effect of the SREBP1 gene on the differentiation of Jinnan cattle precursor adipocytes and to explore the regulatory mechanism of this gene in the differentiation of Jinnan cattle precursor adipocytes by RNA-Seq technology. We transfected Jinnan cattle preadipocytes with an overexpression plasmid of SREBP1 gene and siRNA, and after differentiation induced by oleic acid, oil red O staining was used to observe the accumulation of lipid droplets, and triglyceride (TG) content was tested to explore the influence of SREBP1 gene on their differentiation. RT-qPCR and Western blotting (WB) were used to detect changes at mRNA and protein levels of related genes in preadipocytes overexpressing the SREBP1 gene. RNA-Seq was used to detect adipocytes that interfered with the SREBP1 gene, screened differentially expressed genes (DEGs), performed enrichment analysis of the KEGG pathway, and verified the differentially expressed genes. The potential target genes of the SREBP1 gene involved in regulating the differentiation of precursor adipocytes of Jinnan cattle were explored. Three replicates were set up for each treatment. The results showed as follows: 1) Compared with the control group, overexpression of SREBP1 gene could promote the accumulation of lipid droplets in precursor adipocytes and significantly increase the intracellular triglyceride content (P < 0.01), knockdown of SREBP1 gene could inhibit lipid droplet formation. 2) FABP4 mRNA expression significantly increased after overexpression of the SREBP1 gene (P < 0.05), the expression of FABP7 and LPL were significantly increased (P < 0.01), WB results showed that FABP4 protein expression level increased obviously; 3) A total of 227 DEGs were detected by RNA-Seq, including 67 up-regulated genes and 160 down-regulated genes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that differentially expressed genes were enriched into PPAR signaling pathway and unsaturated fatty acid biosynthesis pathway, the mRNA expression of differentially expressed genes FABP4, LPL, and FABP7 decreased significantly after interfering with SREBP1 gene (P < 0.01), the protein expression level of FABP4 also decreased obviously. In conclusion, the SREBP1 gene can promote the differentiation of precursor adipocytes of Jinnan cattle, and overexpression of this gene can promote intracellular TG accumulation, which is most likely achieved through the regulation of the PPAR signaling pathway and unsaturated fatty acid biosynthesis pathway related to lipid differentiation. The results of this study provide a theoretical reference for exploring the regulatory mechanism of the SREBP1 gene in the differentiation of bovine precursor adipocytes.

The aim of this study was to explore the role and related mechanism of ACSBG2 gene, a member of acyl-CoA synthetase family, in the response of goose liver to changes in nutritional status. In this study, 10-day-old healthy Landes geese (n=8) were used in the fasting/refeeding model, and 65-day-old Landes geese (n=6) were used in the overfeeding model. Firstly, these animal models were used to determine the response of ACSBG2 mRNA expression in goose liver and pectoral muscle to changes in nutritional status. Secondly, the effects of nutrition-related factors (glucose, insulin, sodium oleate, palmitic acid) on the mRNA expression level of ACSBG2 in goose primary hepatocytes and pectoral myocytes were determined. Then, ACSBG2 gene was overexpressed in goose primary hepatocytes and transcriptome sequencing analysis was performed to identify the downstream genes and pathways regulated by ACSBG2, which was followed by determining the mRNA expression level of ACSBG2 downstream genes in vitro and in animal models. The results showed that: 1) The expression of ACSBG2 in the liver was inhibited by fasting (P < 0.01) and induced by refeeding and overfeeding (P < 0.001). The expression of ACSBG2 in pectoral muscle was induced by refeeding (P < 0.001) and overfeeding (P < 0.01). 2) The expression of ACSBG2 in goose primary hepatocytes was inhibited by sodium oleate (P < 0.05) and palmitic acid (P < 0.001), but induced by glucose (P < 0.01) and insulin (P < 0.001). The expression of ACSBG2 in goose primary pectoral cells was inhibited by glucose (P < 0.05) and palmitic acid (P < 0.05), but induced by sodium oleate (P < 0.001). 3) The differentially expressed genes affected by ACSBG2 overexpression were mainly enriched in steroid hormone synthesis and cell adhesion-related pathways. 4) ACSBG2 may mediate the effect of changes in nutritional status on STX1A gene expression in vitro and animal models. In summary, ACSBG2 gene mainly regulates glycolipid metabolism and the expression of inflammation-related factors through steroid hormone synthesis and cell adhesion-related pathways, thereby mediating the effects of nutritional/energy changes.

This study aimed to explore the regulatory effect of differentially expressed microRNAs on Npm2 (nucleoplasmin 2) in pig oocytes. The GV-stage oocytes were collected from slaughterhouse ovaries, cultured in vitro. The expression levels of screened miRNA in GV and MⅡstage oocytes were detected using qPCR. The dual-luciferase reporter experiment was performed to verify the binding site of predicted miRNA with target gene Npm2. Further, miRNAs mimics/inhibitors were added to in vitro maturation medium to evaluate the effect of miRNA on the developmental ability of oocytes and parthenogenetic activated embryos. After the optimal concentration of miRNAs mimics/inhibitors were added, peanut agglutinin staining test was used to detect cortical granule distribution and Npm2 mRNA expression level was detected using qPCR. The results showed that, compared with GV stage oocytes, the expression levels of miR-150 and miR-7138-5p were significantly higher (P < 0.05) and the expression levels of miR-296-3p and miR-423-3p were significantly lower in MⅡ oocytes (P < 0.01), which were consistent with the previous sequencing results. There was no significant difference in the expression levels of miR-32 (P>0.05). The double luciferase assay showed that miR-32, miR-7138-5p, miR-296-3p and miR-150 had binding sites with Npm2, while the fluorescence intensity was significantly down-regulated (P < 0.05). There was no binding site between miR-296-5p, miR-423-3p and Npm2, as the fluorescence intensity was comparable (P>0.05). The optimal concentration of miRNAs mimics/inhibitors was determined. After addition of miR-32 inhibitor, miR-296-5p mimics, miR-296-5p inhibitor, miR-423-3p mimics, miR-423-3p inhibitor, miR-7138 inhibitor, miR-150 mimics and miR-150 inhibitor, cortical particle fluorescence intensity was significantly decreased (P < 0.01). The cortical particle fluorescence intensity was significantly decreased after the addition of miR-296-3p inhibitor and miR-7138-5p mimics (P < 0.05). The expression of Npm2 mRNA in MⅡ oocytes was significantly higher than that in GV oocytes, and after the addition of the optimal concentration of miR-150, miR-296-3p, miR-296-5p, miR-7138-5p mimics and/or inhibitors, the expression of Npm2 in MⅡ oocytes had significant differences compared with the GV oocytes (P < 0.05). There was no significant difference in the expression of Npm2 mRNA after miR-32 and miR-423-3p mimics/inhibitors in MⅡ oocytes compared with control group (P>0.05). These findings indicated that the screened miRNAs may affect oocyte maturation and early embryo development in vitro by regulating Npm2 expression.

The study aimed to define first estrus expression traits after calving (FETs) of dairy cows based on information recorded by collars and to estimate their genetic parameters and relationships with other routine fertility traits. Data were collected from 35 519 Holstein cows across 8 farms in Ningxia from 2020 to 2023, comprising 272 589 811 high-throughput estrus records and 649 274 reproductive records. Three estrus expression traits were defined, including the duration of first estrus expression (DFE), the interval from calving to first estrus expression (ICE) and the mean of heat indicators of first estrus expression after calving (HIE). The GLM in SAS 9.2 was used to analyze the effects of various non-genetic factors on FETs, and the DMUAI module of DMU were used to estimate the genetic parameters of FETs by employing single-trait and multi-trait model. The averages for DFE, ICE, and HIE were 45.87 h, 80.25 au and 44.43 d, respectively. Parity, age at first service, day-night, farm, and year-season of calving had significant effects on all the 3 traits (P < 0.05). DFE was a trait with medium heritability (h²=0.139), while both ICE and HIE were low heritable traits (h² < 0.1). Medium to high genetic correlations were observed between FETs (0.179 to 0.722) and between FETs and fertility traits (0.146-0.766). The collars-based information enables accurate quantification and monitoring of estrus expression in dairy cows, addressing the limitations of traditional recording methods. The FETs obtained from collars-based information are heritable and combining them with fertility traits can provide a new insight to improve genetic selection of estrus efficiency and fertility performance.

The study aimed to screen genes related to the deviation of follicular development and dominant follicle selection in cattle. In this study, 6 healthy 10-month-old Hayford young cows were divided into two groups on average after synchronization. The first group collected the first and second largest follicles before the deviation of the first follicle development wave. The second group collected the first and second largest follicles after the deviation of the first follicle development wave. RNA of follicular granulosa cells was extracted for transcriptome sequencing, and the sequencing results were compared with the reference genome. The PDF1-VS-PDF2 group was screened for differentially expressed genes affecting follicular development. ODF1-VS-ODF2 group, ODF1-VS-PDF1 group and ODF1-VS-PDF2 group screened DEGs that affected dominant follicle selection, and performed GO and KEGG enrichment analysis and PPI analysis to screen key genes. The accuracy of the screened genes was verified by RT-qPCR and Western blotting. The results showed that 220 DEGs were found in the PDF1-VS-PDF2 group, 179 of which were up-regulated and 41 were down-regulated. GO and KEGG analysis showed that PI3K-Akt signaling pathway and TGF-β signaling pathway were related to follicular development. PPI analysis showed that MYC, BRCA1, EZH2, ARID1A and SMARCA4 were hub genes. A total of 184 DEGs were found in ODF1-VS-ODF2 group, 93 were up-regulated and 91 were down-regulated. GO and KEGG analysis showed that TGF-β signaling pathway, PI3K-Akt signaling pathway, Jak-STAT signaling pathway, TNF signaling pathway, mTOR signaling pathway and cell adhesion molecules were related to follicular development. PPI analysis showed that POLR2A, FOS, HIF1A, KIT and SOCS3 were hub genes. A total of 837 DEGs were found in ODF1-VS-PDF1 group and ODF1-VS-PDF2 group, of which 360 were up-regulated and 477 were down-regulated. GO and KEGG analysis showed that mTOR signaling pathway, TNF signaling pathway, TGF-β signaling pathway, PI3K-Akt signaling pathway, progestin (P₄)-mediated oocyte maturation pathway were related to follicular development. PPI analysis showed that HIF1A, RPS9, COL1A2, PIK3R1, COL4A1, ITSN1, GNB1, RPL3, ESPL1, CUL7 were the hub genes. RT-qPCR results showed that the expression of BRCA1, ARID1A and EZH2 in PDF1 was higher than that in PDF2, and the expression of POLR2A in ODF1 was higher than that in ODF2. Western blotting results showed that the expression of BRCA1 and EZH2 in PDF1 was higher than that in PDF2. In this study, MYC, BRCA1, EZH2, ARID1A and SMARCA4 may play a role in the deviation of bovine follicle development. POLR2A, FOS, HIF1A, KIT, SOCS3, RPS9, COL1A2, PIK3R1, COL4A1, ITSN1, GNB1, RPL3, ESPL1 and CUL7 may play a role in the selection process of bovine DF. The results of this study laid a foundation for exploring the theory of regulation of bovine follicle development deviation and dominant follicle selection gene.

This study aimed to analyze the differences in proteins in the saliva of buffaloes at different estrous stages using data-independent acquisition (DIA) proteomics. Fifteen Nili-Ravi buffaloes of similar body condition (average weight of 650-750 kg), in normal oestrus and in good health condition with 2-5 lactations were injected with chloroprostenol on day 0 of the experiment. Saliva samples were collected during proestrus (day 1), estrus (day 3), and metestrus (day 6) before morning feeding (08:00-11:00 am) under the same management conditions and subjected to data-independent acquisition proteomics analysis. Saliva samples were grouped identically for each estrous period, with every 3 cows divided into one group and a total of 5 groups. The results showed that a total of 1 982 proteins were quantified in the saliva of buffaloes at different oestrus stages. Additionally, 59 differentially abundant proteins were identified in the estrus vs proestrus comparison group, and 38 differentially abundant proteins were identified in the estrus vs metestrus comparison group. Functional analyses revealed that proteins such as CBL, SOD1, SPP1, and ARPC1B might play roles in cellular regulation, antioxidant functions, and maintenance of intercellular interactions. KEGG pathway analyses revealed that during proestrus, estrus and metestrus buffalo salivary differential abundance proteins were enriched into the mTOR signalling pathway, progesterone-mediated oocyte maturation and other pathways. In conclusion, significant differentially abundant proteins were present in the saliva of buffaloes at different estrous stages. Additionally, 4 differential proteins associated with reproduction, CBL, ARPC1B, SOD1 and SPP1, were obtained by comparing different oestrus phases. These differential proteins in saliva may serve as key proteins for estrus biomarkers in buffaloes, potentially providing a reference for the development of estrus detection diagnostic kits.

This experiment was conducted to evaluate the effects of metabolizable energy (ME) intake during rearing on the growth and development of reproductive organ, hormone level and gene expression in ovary of laying hens. A total of 720 6-week-old Hyline-Brown laying hens were allocated equally to three groups with six replicates of 40 hens each, and were fed one of three diets that were nutritionally equal with the exception of ME levels. From 6 to 17 weeks of age, the crude protein (CP) levels in diet were 17.5% (6-12 weeks) and 15.5% (12-17 weeks), respectively, and the ME levels in diet were 12.34, 11.11 (12.34×90%) and 9.87(12.34×80%) MJ ·kg^-1 (control group, also called ad libitum feeding group, 90% energy-restricted feeding group, and 80% energy-restricted feeding group), respectively. The hens in control group were fed ad libitum, and the daily amount of feed in experimental groups was restricted to the absolute quantity of the diet consumed by hens in control group. The results showed as follows: 1) Body weight, body slope length and shank circumference of hens at 17 weeks of age decreased linearly (P < 0.001) with increasing levels of energy restriction. 2) The serum UN and GLU content decreased linearly (P=0.040 and 0.044, respectively), while the plasma oestradiol concentrations increased linearly (P=0.026). 3)The ovary stroma of hens in the ad libitum feeding group (ALF17W) and 80% energy-restricted feeding group (ERF17W) with significant differences (P < 0.05) in plasma oestradiol concentrations were used to screen the novel mRNA implemented by the RNA-seq. The proportion of pure reads matching to chicken reference genome was more than 94.61%, and the content of Q20 and Q30 was more than 97.38% and 92.87%, respectively. A total of 1 299 differential genes were screened in ALF17W and ERF17W, of which 961 were down-regulated and 338 were up-regulated in ERF17W. The GO functional enrichment analysis found that differentially expressed mRNAs were involved in 48 GO terms such as cell proliferation, development, and reproduction. The KEGG pathway were significantly enriched in 25 pathways, among these pathways, the cAMP signaling pathway, estrogen signaling pathway, steroid hormone biosynthesis and ovarian steroidogenesis pathway were related to energy metabolism or reproduction. The screened cAMP response element-binding protein (CREB) and steroid-producing acute regulatory protein (StAR) are enriched in the cAMP signaling pathway, and progesterone receptor (PGR), metabotropic glutamate receptor 1 (GRM1) and cytoskeletal keratin (KRT18) are enriched in the estrogen signaling pathway, they may be the potential target genes of ME intake regulating estrogen production in laying hens. The qRT-PCR results showed that the expression trends of 10 randomly selected differentially expressed genes were consistent with the RNA-Seq results. The above results showed that the body weight, serum UN and GLU decreased linearly with the decrease of ME intake during the rearing period, but plasma oestradiol level increased linearly. It is suggested that ME intake during the rearing period may regulate the expression of StAR, CREB1, KRT18, PGR and GRM1 genes in ovarian tissues, and act on the cAMP and estrogen signaling pathways to regulate the production of estrogen in laying hens.

The aim of this study was to establish a model of oxidative damage induced by diquat in broilers and to preliminarily reveal its molecular mechanism. A total of 160 1-day-old broilers were randomly divided into four groups: control group (C), low-dose group (T1), medium-dose group (T2) and high-dose group (T3), 4 replicates per group, 10 replicates each. Five chickens were sampled in each replicate at the 16^th(the first stage), and the remaining five chickens were sampled at 37^th (the second stage) day of age, broilers were weighed, the average body weight of each group was calculated, and diquat was injected intraperitoneally. The T1, T2 and T3 groups were intraperitoneally injected with 5, 10, and 20 mg·kg^-1 of diquat, respectively, and the control group was injected with the corresponding dose of normal saline. The day and the fifth day after the two injections, two broilers were randomly taken from each replicate and blood collection and sampling were conducted. The results showed that after two injections of diquat, compared with the control group, 1) The broilers in the experimental groups were depressed and drank less water and eaten less, 2 broilers in the T3 group died at 6 h after each injection, and the body weight of broilers in the experimental groups were extremely significant reduced at 72 h and 96 h after each injection (P < 0.01); 2) The serum activity of diamine oxidase (DAO), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the T2 and T3 groups were extremely significant increased (P < 0.01); 3) The serum activity of glutathione peroxidase (GSH-Px) in the T2 and T3 groups were extremely significant decreased (P < 0.01), and the serum levels of malondialdehyde (MDA) were extremely significant increased (P < 0.01). The serum activity of superoxide dismutase (SOD) in the T3 group after the second infection was significantly decreased (P < 0.05); 4) The serum contents of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in each experimental group were extremely significant increased (P < 0.01); 5) In the experimental groups, inflammatory cell infiltration was present, some hepatocytes were vacuolated, cytoplasmic was obvious laxity, and the villi loss of each intestinal segment was serious; Histological scores showed that the liver and small intestine scores in both the T2 and T3 groups were extremely significant higher than control group(P < 0.01), and except the histological score of duodenum at the second stage, there was no significant difference between the T1 group and the control group; 6) The mRNA expressions of ZO-1 and OCLDN in the liver and jejunum of broilers in each experimental group were extremely significant decreased(P < 0.01); 7) The mRNA expression of HO-1 in the liver of T2 group was significantly lower than T1 group (P < 0.05); The mRNA expressions of Nrf2 and HO-1 in jejunum of T2 and T3 groups were extremely significant lower than T1 group(P < 0.01). In summary, this study successfully established a model of oxidative damage induced by diquat in broilers, and the injection dose of 10 mg·kg^-1 was the most appropriate to establish the oxidative damage model, and it was found that diquat may induce the production of inflammation-related factors and inhibit the expression of antioxidant enzymes through the Nrf2/HO-1 signaling pathway, causing intestinal barrier damage in broilers, thereby causing inflammatory response in broilers.

The study evaluated the effects of coated folic acid (CFA) and coated cobalamin (CCA) on the lactation performance, nutrient digestion, rumen fermentation and hepatic lipid content of dairy cows during perinatal period. Forty-eight Holstein dairy cows during perinatal period with similar parity, previous 305-d milk yield and body weight were assigned in a 2×2 factorial and randomized block design to 4 groups: cows in the control group were fed a basal diet, and those in other 3 groups were fed basal diets supplemented with CFA 6.75 g·d^-1, CCA 0.6 g·d^-1, and CFA 6.75 g·d^-1+CCA 0.6 g·d^-1, respectively. The study started at 21 days before the excepted calving date and ended at 21 days after calving. During the postpartum 21 days, body weight change (BWC) and lactation performance were determined, and samples of blood, rumen fluid and liver tissue were collected. The results showed as follows: 1) CFA addition significantly decreased the BWC (P < 0.05) and significantly increased feed efficiency (P < 0.05). The yields of actual milk, 4% fat-corrected milk (FCM), milk fat and milk protein significantly increased (P < 0.05) with CFA or CCA addition. 2) The apparent digestibility of dry matter (DM) and neutral detergent fiber (NDF) increased significantly (P < 0.05) with CFA addition. The apparent digestibility of DM, organic matter (OM), crude protein (CP), NDF and acid detergent fiber (ADF) significantly increased with CCA addition (P < 0.05). 3) Rumen pH was not significantly changed (P>0.05), but total volatile fatty acids (TVFA) concentration increased (P < 0.05) with CFA or CCA addition. Rumen acetate concentration and acetate to propionate ratio increased significantly (P < 0.05) with CFA addition. Rumen propionate concentration increased significantly (P < 0.05), while acetate to propionate ratio decreased significantly (P < 0.05) with CCA addition. 4) Blood nonesterified fatty acids, β-hydroxybutyrate (BHB) and homocysteine (Hcy) contents decreased (P < 0.05) with CFA addition. Blood glucose content was not significantly changed (P>0.05), but folate (FA) content increased significantly (P < 0.05) with CFA addition. Blood glucose and cobalamin (CA) contents increased significantly (P < 0.05) with CCA addition. 5) Hepatic FA content increased significantly (P < 0.05), and total lipid and triglyceride (TG) contents decreased significantly (P < 0.05) with CFA addition. Hepatic CA content increased significantly (P < 0.05), but total lipid and TG contents were not significantly changed (P>0.05) with CCA addition. 6) When compared with CFA or CCA addition alone, the FCM yield, NDF digestibility and rumen TVFA concentration were greater (P < 0.05), and blood Hcy and hepatic TG contents were lower for cows receiving CFA and CCA together. In conclusion, addition of CFA and CCA together is more effective in improving lactation performance and reducing hepatic lipid content compared with CFA or CCA addition alone.

This study aims to investigate the effects of co-treatment of sodium butyrate and indole-3-propionic acid on the expression of tight junction-related proteins and inflammatory factors in Caco-2 cells. Caco-2 cells were administrated with 0.4 mmol·L^-1 sodium butyrate and 0.1 mmol·L^-1 indole-3-propionic acid each and together for 24 h; Then the transmembrane resistance (TEER) value, expression of tight junction related proteins, gene expression and secretion level of inflammatory cytokines were measured. The results showed that, compared with control group, the co-treatment of sodium butyrate and indole-3-propionic acid significantly increased the TEER value in Caco-2 cells (P < 0.01), while no significant changes were observed in sodium butyrate or indole-3-propionic group (P>0.05); In addition, the results by qPCR and Western blot showed that the relative expression of Claudin-2 mRNA was significantly down-regulated in the co-treatment group compared with the control group (P < 0.01); Compared with sodium butyrate group, the relative mRNA expression of Claudin-1 was significantly up-regulated in the co-treatment group (P < 0.05), and the protein expression of Claudin-1, ZO-1 and Occludin was highly significantly increased in the co-treatment group(P < 0.001); Compared with the indole-3-propionic acid group, the relative mRNA expression of ZO-1 was significantly up-regulated in the co-treatment group (P < 0.05), and the protein expression of Claudin-1 and ZO-1 was highly significantly increased (P < 0.001). For inflammatory cytokines, the relative mRNA expression levels of IL-6, IL-1β, IL-8 and TNF-α were significantly down-regulated in the co-treatment group (P < 0.05); Meanwhile, the results of ELISA showed that the co-treatment group highly significantly suppressed the secretion of IL-6, IL-1β, IL-8 and TNF-α in the cell supernatant compared with the control group (P < 0.001). In conclusion, compared with sodium butyrate or indole-3-propionic acid, the co-treatment of sodium butyrate and indole-3-propionic acid increased the TEER value, promoted the expression of tight junction proteins, suppressed the secretion of pro-inflammatory cytokines, and reduced intestinal permeability. Overall, these results suggested that the co-treatment of sodium butyrate and indole-3-propionic acid has a stronger role in enhancing intestinal epithelial cell barrier function and inhibiting inflammatory response.

This experiment was conducted to investigate the variability of digestive environment in different sites of the small intestine of pigs and its correlation with dietary standardized ileal digestibility of amino acids, and to provide a reference for the development of rapid evaluation of nutritive of amino acid for pig diets. In this experiment, 18 Duroc×Landrace×Yorkshire barrows with an initial body weight of (21.6±3.5)kg were randomly divided into 3 groups with 6 pigs in each group. A T-cannula was surgically equipped at the end of duodenum, the middle of the jejunum-ileum and the end of the ileum, respectively. Each group of pigs was allotted to a replicated 3×3 Latin square design with 3 diets (3 kinds of semi-homozygous diets with corn, soybean meal and wheat bran as the only protein source, respectively) and 3 phases. In each phase, there was a 5-d adaption to experimental diet, followed by a 3-d determination of digesta passage time, a 2-d collection of digesta, and a 2-d of recovery period. The pH, ion concentration, digestive enzyme activity and the passage time of digesta in each sites of small intestine were determined. The effects of intestinal sites and diets on the digestive environment were analyzed, and the simple correlation analysis was conducted between the standardized ileal digestibility of amino acid in diets and the digestive environment parameters in small intestine. The results showed that: 1) Intestinal sites and diets had significantly interactions (P < 0.05) on the K⁺ and Cl^- concentrations and chymotrypsin activity of intestinal fluid. The intestinal sites had extremely significant effects (P < 0.01) on pH, amylase and trypsin activities and Na⁺ concentration, and diets had significant effects (P < 0.05) on pH, trypsin activity and Na⁺ concentration. 2) Taking the average time of appearance and disappearance of digesta indicator as the passage time, the passage time of corn diet in duodenum, middle jejunum-ileum and terminal ileum was significantly longer (P < 0.05) than that of wheat bran diet, but there was no significantly difference (P>0.05) compared to soybean meal diet in duodenum. 3) Pigs fed the corn diet had no significant correlation (|r|≤0.68, P>0.10) between the pH, Na⁺ concentration, K⁺ concentration, passage time of digesta, digestive enzyme activities and the standardized ileal digestibility of amino acids. Pigs fed the soybean meal diet showed significantly negative correlation (|r|=0.86, P < 0.05) between the amylase activity and standardized ileal digestibility of glutamate. Feeding pigs with wheat bran diet, there was a significantly positive correlation (|r|≥0.87, P < 0.05) between Na⁺ concentration and proline, passage time of digesta and standardized ileal digestibility of methionine, histidine, arginine, aspartic acid, glutamic acid, glycine and alanine. In summary, the intestinal sites and diets of growing pigs have a significant influence and interaction on the digestive environment of small intestine. The correlation between pH, ion concentration and digestive enzyme activity and the standardized ileal digestibility of amino acids is weak, but the passage time of digesta affects protein digestion in wheat bran diet.

The aim of this study was to elucidate the differences in gut microecology of pigs induced by breast milk and formula. In this study, 12 healthy 1-day-old female Rongchang piglets with similar body weight were randomly divided into two groups (BF: breast-fed; FF: formula-fed), with 6 piglets in each group. After weaning at 30 days of age, piglets were continued feeding until 210 days of age. Body weight and serum biochemical indicators were determined, and the composition of cecal microbiota and metabolites were detected by 16S rRNA sequencing and LC-MS technology. The results showed that the average daily weight gain, serum alanine aminotransferase, glutamyl aminotransferase, alkaline phosphatase, albumin, total cholesterol, high-density lipoprotein, low-density lipoprotein levels, and albumin/globulin ratio were significantly higher (P < 0.05), while creatinine levels and aspartate transaminase/glutamic pyruvic transaminase ratio were significantly lower in BF pigs compared with FF pigs (P < 0.05). Regarding gut microbiota, compared to FF pigs, BF pigs had significantly higher chao1, ace, and simpson indices as well as significantly higher Actinobacteria, Bacteroidetes, Lactobacillus, Clostrium_XlVa, Streptococcus, Oscillibacter and Megasphaera (P < 0.05) in relative abundance, while the shannon index as well as the relative abundance of Clostridium_sensu_stricto, Turicibacter, Terrisporobacter and Mogibacterium were significantly lower (P < 0.05). Moreover, compared with FF pigs, the differential microbiota with down-regulated abundance were enriched in amino acid metabolism, cofactor and vitamin metabolism, lipid metabolism, and nucleotide metabolism in BF pigs (P < 0.05), and the differential microbiota with up-regulated abundance were enriched in carbohydrate metabolism, glycan biosynthesis and metabolism, and xenobiotic degradation and metabolism in BF pigs (P < 0.05). Differential metabolites were mainly involved in amino acid metabolism, such as histidine metabolism, arginine and proline metabolism, and tryptophan metabolism, followed by pyrimidine metabolism and prolactin signaling pathways (P < 0.05). In addition, most microbiota with genus level abundance top 8 showed significant correlation with specific metabolites (P < 0.05). In summary, the breast milk has a positive effect on increasing the abundance of beneficial bacteria and promoting amino acid metabolism. The results will provide possible explanations for some of the risks and benefits associated with piglet lactation practices, and will support theoretical guidance for the development of piglet formula.

The aim of this study was to construct multi-epitope recombinant plasmids of Eimeria tenella EtAMA1, EtAMA2, EtMIC3, EtMIC4, EtMIC13, EtROP5, and EtSAG1 antigens and verify their expression in DF-1 cells and chickens through various assisted delivery methods, we employed pcDNA3.1(+) as the eukaryotic expression vector. The eukaryotic expression plasmid pcDNA3.1-ET Seven was constructed through bioinformatics analysis, obtaining seven epitopes and reverse transcriptional amplification of gene fragments of seven antigen epitope regions in E. tenella mRNA templates at different time points. The company synthesized pcDNA3.1(+) as the eukaryotic expression vector, and constructed the eukaryotic expression plasmid pcDNA3.1-ET seven. Subsequently, the constructed positive plasmid, pcDNA3.1-ET Seven, was transfected into DF-1 cells. RT-PCR and Western blot analysis was employed to detect the expression of the ET Seven recombinant gene in DF-1 cells. The cytokine levels of pcDNA3.1-ET seven in chickens were detected by calcium phosphate nanoparticles as adjuvant plasmid delivery adjuvant, and specific IgG antibody levels were detected by indirect ELISA. The eukaryotic expression plasmid pcDNA3.1-ET Seven was successfully constructed, and the specific expression of ET Seven was detected through Western blot analysis and RT-PCR. After the plasmid was immunized with calcium phosphate nano-adjuvant, the detected cytokines IL-2, IL-6, IL-10, IL-12, IFN-γ and CD-40 were all expressed at high levels seven days after three times of immunization, and the expression levels of CD-40 and IL-6 were the highest. The pcDNA3.1-ET Seven recombinant plasmid, when conjugated with calcium phosphate adjuvant, was administered intramuscularly, resulting in a positive detection of specific IgG antibodies through indirect ELISA. The results showed that the pcDNA3.1-ET seven recombinant plasmid was successfully constructed and had good immunogenicity in chickens. This finding provides novel insights and technical support for vaccine prevention and control of chicken coccidiosis.

The aim is to study the effect of GreA protein on the biological characteristics and pathogenicity of Salmonella Enteritidis. Based on the parent strain ATCC13076, ΔgreA and ΔgreB single gene deletion strains and ΔgreAΔgreB double gene deletion strains were constructed by λ-Red recombinant system. Their growth and swimming motility, resistance to stressful environment, biofilm formation ability, and pathogenicity to cells and mice of the parents and the deletion strains were detected. RNA-seq was used to detect differentially expressed genes (DEGs). GreA protein negatively regulated the H₂O₂ resistance, the growth in vitro and biofilm formation ability of Salmonella Enteritidis, and positively regulated the motor ability, cell invasion and intracellular proliferation ability of Salmonella Enteritidis and its colonizing ability in mouse organs, but have no significant effect on resistance of Salmonella Enteritidis to high temperature and acid environment. GreA protein affects the expression of 107 genes, including positive regulation of inv operon, fim operon and flg operon, which are mainly related to bacterial motility and cell invasion; and negative regulation of cps operon, cbp operon and leu operon, which are related to amino acid synthesis and metabolism. GreA protein affects the environmental adaptability and pathogenicity of Salmonella Enteritidis by regulating the expression of metabolic pathways related to flagellar synthesis and cell invasion.

Adhesion of host small intestinal epithelial cells through fungal hairs and flagella is an indispensable and important link in pathogenic processes of Salmonella Enterison. This study aims to analyze the competitive blocking effect of the hair recombinant protein FimA-sefA expressed by prokaryotes on the adhesion of Salmonella Enteritis to intestinal epithelial cells (IPEC-J2). FimA and sefA gene fragments were cloned into pET28a (+), resulting in the construction of a recombinant plasmid pET28a-FimA-sefA, which was then transformed into the strain BL21 (DE3). The recombinant protein FimA-sefA was purified using nickel-column affinity chromatography (Ni-NTA). The adhesion of enteritis to IPEC-J2 cells was analyzed using indirect immunofluorescence (IFA) and plate counting. The molecular weight of the expressed recombinant protein FimA-sefA was 36 ku; the recombinant protein FimA-sefA, recombinant bacteria, and Salmonella Enteritis can adhere to IPEC-J2 cells. Recombinant protein FimA-sefA and recombinant bacteria can competitively block the adhesion of Salmonella Enteritis to IPEC-J2 cells, and the blocking effect of recombinant bacteria was stronger than that of the recombinant protein. We believe that the recombinant protein FimA-sefA can competitively block the adhesion of Salmonella Enteritis to IPEC-J2 cells to a certain extent, providing a new strategy for preventing and reducing enteritis infection.

The NF-κB pathway is an important target for regulation of host immune function by viruses and plays an important role in the pathogenesis of viral infections. It has shown that the ankyrin protein encoded by some poxviruses regulates the NF-κB pathway in a variety of ways, but how the ankyrin protein encoded by goat poxvirus (GTPV) regulates the NF-κB pathway remains to be explored. In this paper, we aim to investigate the effects of the ankyrin protein ORF140 encoded by GTPV on the NF-κB pathway. The effects of ankyrin protein ORF140 on the NF-κB pathway were detected using dual luciferase reporter system, its localization in HEK293T cells was observed by indirect immunofluorescence and its effects on expression of the factors associated with the activated NF-κB pathway at different time points by Western blot. Results showed that ORF140 was able to inhibit the NF-κB pathway. It was localized in the cytoplasm and did not undergo nuclear translocation even when the NF-κB pathway was activated. ORF140 was able to inhibit the nuclear translocation of NF-κB-p65 and the degradation of p-IκBα. GTPV ORF140 localizes in the cytoplasm and downregulates NF-κB transcriptional activity by inhibiting the nuclear translocation of NF-κB-p65 and the degradation of p-IκBα.

To provide a candidate strain for the development of an efficient and safe ORFV genetic engineering vaccine, the deleted ORF112 gene recombinant ORFV strain was constructed using the homologous recombination technique. Taking the ORF 112 gene as the targeted gene, the left and right homology arms and the enhanced green fluorescent protein gene (EGFP) expression frame were in-fused by the overlapping PCR method, which was connected with the pUC19T vector to construct the pUC19T-ORFV-ΔORF112-EGFP transfer vector. Subsequently, the pUC19T-ORFV-ΔORF112-EGFP transfer vector was transfected into the Vero cells to cause the homologous recombination events with the wild-type ORFV genome. The positive ORFV-ΔORF112-EGFP strain was screened and purified by limited dilution and picking a single-cell clone method. The genetic stability and replication ability of the deleted ORF112 ORFV recombinant strain were further investigated. The positive recombinant ORFV strain was purified using limited dilution and picking a single-cell clone method. After the PCR verification and sequencing identification, the recombinant ORFV-ΔORF112-EGFP strain was successfully obtained. The expression of EGFP in the recombinant virus genome was stable, with no attenuating characteristics during the serial passages. The replication and growth characteristics of this recombinant ORFV stain are the same as wild-type ORFV. The ORFV-ΔORF112-EGFP recombinant strain has been successfully constructed, purified, and screened, which maintains good genetic stability and replication ability, providing the candidate strain for the development of a genetic engineering ORFV vaccine.

Foot-and-mouth disease virus (FMDV) serotype Asia 1 is still circulating in Asia countries around China, which will be a long-term threat to animal husbandry. The aim of this study was to screen porcine monoclonal neutralizing antibodies against Asia1 FMDV using single B-cell antibody technology, and to identify the antigenic sites recognized by neutralizing antibodies. FMDV Asia1/JS/05 inactivated whole virus particles were used as the bait antigen to sort antigen-specific B cells in PBMCs of immunized porcine by flow cytometry. The sequences of variable regions of the heavy and light chain of the IgG were amplified by nested PCR. The expression plasmids of the IgG heavy and light chain were constructed respectively and co-transfected into CHO-S cells to express the whole IgG antibodies. The reactivity and neutralization ability of expressed antibodies were verified by indirect ELISA, indirect immunofluorescent assay (IFA) and viral neutralizing test (VNT). The epitopes as well as the critical amino acids recognized by the neutralized antibody were identified using Western blot and neutralizing antibody escape mutants screening. Five Asia 1 FMDV-specific antibodies were obtained, of which PD3 and PD7 showed neutralizing ability. These two neutralizing antibodies recognized the same epitope and the key amino acids residue was D 72 of the VP2 protein. In this study, neutralizing antibody against Asia 1 FMDV from pig were obtained for the first time, which provides further information of the protective antigen sites of Asia1 FMDV, and make a basis for the design of molecular vaccine and the development of new tools for diagnosis of FMD Asia 1.

In order to develop FMD vaccine candidate strain that could effectively prevent and control recently epidemic strains belonging to three topotypes of serotype O FMDV, we constructed recombinant full-length clone containing the structural protein genes belonging to three topotypes of type O FMDV by gene replacement based on FMDV reverse genetic manipulation technique. The recombinant FMDV was rescued after transfection the full-length plasmid into BSR/T7 cells expressing T7 RNA polymerase. The effect of gene reconstruction on the biological characteristics of the recombinant virus was analyzed, pigs and cattle were vaccinated with the vaccines prepared from the recombinant virus and parental virus, the potential of the recombinant virus as a vaccine candidate strain for type O FMD was preliminarily studied by in vitro neutralization test. The results showed that the replacement of G-H loop gene of structural protein VP1 of FMD epidemic strain did not significantly affect the plaque phenotype and replication ability of the recombinant virus, but different G-H has obvious effect on production the cross-neutralizing antibody in pigs, and has little effect on production the cross-neutralizing antibody in cattle, indicating that FMDV G-H loop is the immune dominance epitope for pigs. This study will provide important reference for the design of FMDV vaccine in future.

African swine fever is an acute, highly contagious and deadly infectious disease caused by the African swine fever virus, which has had a huge impact on the pig breeding industry. The lack of vaccines and effective therapeutic drugs has brought great challenges to the prevention and control of African swine fever. We tried to design a fusion protein to improve the immunogenicity of African swine fever virus p30. CRM197, a non-virulent mutant of diphtheria toxin, has been successfully used as a carrier protein in polysaccharide-protein conjugate vaccines to enhance the immunogenicity of polysaccharides. In this study, a fusion expression plasmid of p30 and CRM197 catalytic domain (A fragment) was constructed, and the immunogenicity of the purified recombinant protein and p30 alone were evaluated using mice. After immunizing mice with the fusion protein p30-CRM197(A) purified by prokaryotic expression and affinity chromatography, p30 alone could excite a high level of specific IgG against p30 in a shorter period of time, and maintain a high level of lymphocyte immune activity two months after the initial epidemic. The results of this study showed that the A fragment of CRM197 alone could significantly enhance the immunogenicity of prokaryotic expression p30 protein as a protein carrier, and it could be used as a candidate intramolecular adjuvant for African swine fever subunit vaccine.

This study aimed to construct a precise gene-edited cell lines for the ALV-J receptor molecule chNHE1. In DF-1 cells, key amino acids V33 and W38 facilitating ALV-J entry into host cells via chNHE1 were mutated and deleted, respectively. Meanwhile, the codon encoding amino acids 34-37 of chNHE1 were synonymously replaced. Forty-eight monoclonal cell lines were obtained through flow cytometry sorting, and PCR identification and sequencing analysis revealed that 14 of these cell lines had chNHE1 mutations in V33, W38 deletions, and synonymous substitution of codons at amino acids 34-37, with a gene editing efficiency of 29%. In order to verify the genetic stability and proliferation of the chNHE1 gene-edited cell lines, sequencing analysis was performed on the cells lines that were continuously passaged for 25 generations, and the results showed no revertant mutations in the chNHE1 gene. Further cell count analysis showed no revertant mutations in the chNHE1 gene. Further cell count analysis showed that the proliferation level of chNHE1 gene-edited cell lines was unaffected. In order to verify the anti ALV-J infection ability of the chNHE1 gene-edited cell line, ALV-J fluorescent reporter virus (ALV-J-GFP) and ALV-J prototype strain (HPRS-103) were used for virus challenge experiment. The fluorescence observation and flow cytometry analysis results showed that the chNHE1 gene-edited cell line completely resisted the infection of ALV-J fluorescent strain; Furthermore, the results of indirect immunofluorescence assays, PCR amplification assays, and virus titer determination assays showed that the chNHE1 gene-edited cell line completely resisted the infection of HPRS-103 strain. This study utilized a fluorescence labeled CRISPR/Cas9 system combined with flow cytometry sorting to successfully construct a chNHE1 gene-edited cell line, which showed complete resistance to ALV-J infection, and exhibited good genetic stability and proliferation activity. This provides theoretical support and gene editing targets for establishing new technologies against ALV-J infections.

Thiazolidinedione (TZD) is an oral anti-diabetic drug for the treatment of insulin resistance. Abuse or improper use of TZD has adverse effects on animal and human health. This study explores effects of TZD on the growth and metabolism of chicks and underlying mechanisms of this process. Eighty chicks at the age of 6-day-old were divided into a control group (20 females, 20 males) and an experimental group (20 females, 20 males). Chicks in the control group were administered without TZD, chicks in the experimental group were daily administrated with 25 mg·(kg·d)^-1 TZD by gavage for 14 days, and the growth of chicks was measured. The biochemical kits were used to detect the levels of adenosine triphosphate (ATP) and adiponectin, the concentrations of growth and metabolism-related hormones, and the function of mitochondria. The mRNA and protein expressions of genes which are associated with cell metabolism and proliferation were detected by RT-PCR and western blotting. Results showed that TZD treatment significantly decreased the average daily gain of chicks (P < 0.05). Serum ATP, insulin (INS), growth hormone (GH), and insulin-like growth factor 1 (IGF1) levels were reduced in TZD-treated group (P < 0.05), while adiponectin and insulin-like growth factor binding protein 2 levels was improved (P < 0.05). TZD also decreased ATP levels and mitochondrial enzyme activities in the liver, kidney and muscle of chicks. Moreover, mRNA and protein expressions of adiponectin and its receptors, AMP-activated protein kinase α2 (AMPKα2), p21 and p27 were enhanced in TZD-treated chicks (P < 0.05), whereas mRNA and protein expressions of INS and its receptor, IGF1 and its receptor, phosphatidylinositol-3-kinase (PI3K), AKT, mammalian target of rapamycin (mTOR), Cyclin-dependent kinase 2 (CDK2) and Cyclin E1 were inhibited (P < 0.05). The growth metabolism-related hormones and mitochondrial function in TZD-treated chicks were down-regulated by the adiponectin-mediated AMPK signaling pathway, which inhibited its downstream PI3K/AKT/mTOR, and further led to increased expressions of p21/p27 and decreased expressions of CDK2/Cyclin E1, this inhibiting the cell proliferation in growth and metabolic tissues of chicks. Our findings suggest that TZD has an adverse effect on chicken growth through regulating the adiponectin-mediated AMPK signaling pathway, this provides a certain theoretical basis for avoiding the abuse or improper use of TZD in clinical practice and animal production.

The purpose of the study was to evaluate the clinical efficacy of the compound oxyclozanide suspension through the deworming testing, and to provide reference basis for its clinical use. Seventy-five Qinghai Huanhu yaks were divided into 5 groups randomly, with 15 yaks in each group, which were compound oxyclozanide suspension low dose (0.125 mL·kg^-1), medium dose (0.250 mL·kg^-1), high dose (0.500 mL·kg^-1), control group (0.250 mL·kg^-1) and blank control group. The test cycle was 28 days. The feces were collected on the first day and on days 7, 14, 21, and 28 after administration, and the egg counts (eggs per gram, EPG) of Fasciola hepatica and digestive tract nematodes is measured, and the precision of deworming and the rate of egg conversion were calculated. The blood routine, blood biochemical indicators value were measured before and on day 28 of the administration. The precision deworming rates of low, medium, and high doses of compound oxyclozanide suspension against yak Fasciola hepatica were 97.5%, 100.0%, and 100.0% respectively, and the egg negative conversion rates were 80%, 100% and 100% respectively. The recision deworming rate of the imported medicine against yak Fasciola hepatica was 99.4%, and the egg negative conversion rate was 93.3%. The precision deworming rate of low, medium and high doses of compound oxyclozanide suspension against yak digestive tract nematodes was 99.2%, 100.0% and 100.0%, and the egg transfer rate was 80%, 100% and 100%, respectively, while the precision deworming rate of control imported drug against yak digestive tract nematodes was 99.9%, and the egg transfer rate was 93.3%.The results of this yak test showed that the compound oxyclozanide suspension had a significant treatment effect on the Fasciola hepatica insects and digestive tract nematodes. There is no significant difference in the clinical effects between the high and medium dose groups. Therefore, the recommended dose for clinical application was 0.250 mL·kg^-1 in the medium dose group.

This study was conducted to investigate the effects of irisin on Escherichia coli (E. coli)-induced tissue damage in mice, and bacterial clearance and inflammation induced by macrophages. The mouse model was established by intraperitoneal infection with E. coli at a median lethal dose. The protective effect of irisin was evaluated by body temperature, clinical score, survival rate, degree of organ and tissue damage, bacterial load and inflammation level of tissue. E. coli was incubated with different concentrations of irisin in vitro, and the bacterial solution OD_{600 nm} was detected by microplate reader to evaluate the antibacterial effect of irisin. After pretreatment of mouse mononuclear macrophages cells (RAW264.7) with irisin, macrophages were infected with E. coli, and cell lysates and mRNA were collected to evaluate the ability of macrophages to engulf and clear bacteria and the level of M1 polarization. Results showed that irisin could significantly improve the survival rate of bacterially infected mice, reduce the clinical score, promote body temperature recovery, and reduce the liver, lung and kidney damage (P < 0.000 1), decreased the bacterial load of heart (P < 0.05), liver (P < 0.01), spleen (P < 0.01), lung (P < 0.01), kidney (P>0.05), and reduced IL-1β(P < 0.000 1) and IL-6 (P < 0.001) mRNA levels in liver and interleukin (IL)-1β (P < 0.05) and IL-6 (P < 0.01) mRNA levels in kidney. Irisin did not inhibit the growth of E. coli in vitro, but promoted RAW264.7 phagocytosis (P < 0.001) and clearance of E. coli, and also reduced the inflammatory marker IL-1β(P < 0.000 1), IL-6 (P < 0.000 1) and iNOS (P < 0.001) mRNA levels. Irisin can promote the phagocytosis and clearance of bacteria by macrophages, reduce the pro-inflammatory response of macrophages induced by bacterial infection, and alleviate tissue damage in mice.

The rise of antibiotic resistance has brought renewed attention to older drugs like polymyxins, making it important to control the spread of resistance genes. In this study, we focused on colistin-resistant Escherichia coli strains from a pig farm in Hunan, China. Out of 240 samples, 172 E. coli strains were isolated and cultured, PCR was used to identify mcr-1 positive E. coli strains. These strains were tested by ten different antibiotics to check for their drug resistance, and whole-genome sequencing (WGS) was applied to analyze their genetic background. All the nine mcr-1 positive E. coli strains exhibit high levels of multidrug resistance. Four distinct MLST types (ST10, ST196, ST5229, ST46) and 4 serotypes (O83:H5, O16:H7, O16:H51, O9:H4) in the nine strains were identified. Further bioinformatics analyses showed that the ISApl1 element, which is commonly associated with mcr-1 transmission, was not observed in the present study, while the type Ⅳ secretion system genes were detected in all the nine mcr-1 positive E. coli strains. Plasmid conjugation experiments and the presence of polymorphic MLST haplotypes indicated a vertical plasmid-mediated mcr-1 transmission possibility. Notably, our study for the first time uncovered a pig origin IncI2(Delta) plasmid carrying mcr-1 gene in E. coli which is normally presented in the human clinical E. coli strains.

We aimed to investigate the inhibitory activity of Folium Isatidis aqueous extract on the in vitro proliferation of infectious bovine rhinotracheitis virus (IBRV), and to provide new reference data for the clinical treatment of IBRV infection. The cytotoxic effects of Folium Isatidis aqueous extract on Madin-Darby bovine kidney (MDBK) cells were assessed using cell counting kit-8 (CCK-8) and MTT assays, alongside the cytopathic effect (CPE) method, to establish the maximum safe concentration (MSC). The MSC was subsequently diluted in a three-step, two-fold serial dilution to evaluate the extract's inhibitory action on IBRV through both MTT and CCK-8 assays across various administration modes. Metrics such as cell viability, drug efficacy, 50% cytotoxic concentration (CC₅₀), and 50% effective concentration (EC₅₀) were calculated, employing the Therapeutic Index (TI) as a principal evaluative criterion. The inhibitory effect of Folium Isatidis aqueous extract on the proliferation of IBRV was further confirmed by viral titration, real-time quantitative PCR, and immunofluorescence assays. The findings indicate that the MSC of Folium Isatidis aqueous extract on MDBK cells is 0.8 μg·μL^―1, with a CC₅₀ of 1.937 μg·μL^―1. The antiviral efficacy enhances with increasing concentrations within the MSC range. The extract exhibited significant inhibitory effects on IBRV across different administration modes, pre action of medicine and virus, inoculate virus first and then add medicine, add medicine first and then inoculate virus, with EC₅₀ values of 0.292 8, 0.350 1, and 0.416 1 μg·μL^―1, respectively, and corresponding TIs of 6.615 4, 5.532 7, and 4.655 1. Notably, the maximal viral suppression was observed at 36 hours post-infection when the extract was pre-mixed with the virus, as evidenced by real-time fluorescence quantitative PCR and immunofluorescence staining. The aqueous extract of Folium Isatidis demonstrates potent anti-IBRV activity, providing a scientific foundation for its clinical application and further development in veterinary medicine. This study contributes valuable insights into the antiviral properties of Folium Isatidis, underscoring its potential as a therapeutic agent against IBRV infections.

The effect of naringin (NAR) on the prevention and treatment of lung epithelial barrier damage induced by Actinobacillus pleuropneumoniae (APP) infection in mice was investigated. Forty male Kunming mice between 6 and 8 weeks of age were randomly divided into five groups, namely, the control group, the model group, the NAR low-dose group, the NAR medium-dose group and the NAR high-dose group. The drug was administered by gavage continuously for 8 d. The blank and model groups were given saline. On d 7, mice were infected with APP (1×10⁸ CFU·mL^-1, 100 μL per mouse) by tracheal intubation, and the blank group was infused with an equal volume of sterile saline, and the mice in each group were executed at 48 h after modelling, sampled and evaluated for the preventive effect of NAR. X-ray images showed that NAR reduced the APP load in the alveolar lavage fluid of mice and alleviated the pathological lung injury; moreover, NAR intervention reversed the APP-induced decrease in the expression of ZO-1, Occludin-1, and Claudin-1 tight junction proteins in the lung tissues of mice, thus promoting the repair of the lung epithelial barrier. To further explore the mechanism of NAR promoting lung epithelial barrier repair, it was found that NAR not only reduced the secretion of pro-inflammatory factors IL-6 and TNF-α, but also increased the expression of anti-inflammatory factor IL-10, and increased the expression of antioxidant enzymes SOD2 and Hmox1 in mouse lung tissues, suggesting that NAR enhances the anti-inflammatory and antioxidant capacity of the organism. In addition, NAR can inhibit the TGF-β1/Smad2/3 pathway, reduce APP-induced proliferation of lung fibrotic tissue; analysis of key proteins related to apoptosis and Tunel fluorescent labelling revealed that NAR could reduce apoptosis in a dose-dependent manner. In conclusion, NAR can repair the lung epithelial barrier damage in mice through anti-inflammatory, antioxidant, anti-fibrotic and anti-apoptotic pathways, thus effectively reversing APP-induced inflammatory lung injury and exerting the efficacy of preventing and controlling APP.

The study aimed to establish the HD11 cell line with a stable expression of the marco gene through the lentivirus expression system, providing a crucial tool for investigating the antiviral immune function of the marco gene. Initially, the marco gene coding sequence was amplified by PCR using HD11 cells as a template and cloned into the lentiviral vector to generate the recombinant lentiviral plasmid pLV3-CMV-MCS-Puro-marco. Subsequently, the recombinant plasmid pLV3-CMV-MCS-Puro-marco was co-transfected with helper plasmids pLP1, pLP2, and pMD2.G into 293T cells using a tetra-plasmid lentivirus packaging system, resulting in the construction of recombinant lentivirus carrying the chicken marco gene. The successfully packaged recombinant lentivirus was then used to infect HD11 cells, followed by puromycin (2 μg·mL^-1) selection for 72 h. The stable expression of the chicken marco gene in the HD11 cell line was confirmed by RT-qPCR analysis. This study successfully established a stable HD11 cell line expressing the chicken marco gene, providing a crucial basis for further exploration of the marco gene's function.

The pathogenic microorganisms can be sensed and recognized by innate immune system once invaded the animals. And the sensing of conserved structural components called pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) of the hosts activates a series of signaling events, leading to induction of downstream antiviral effector proteins including type Ⅰ interferons (IFNs) and proinflammatory cytokines to clear the invaders. The nucleotidyltransferase GMP-AMP (cGAMP) synthase (cGAS) is ubiquitously expressed in various types of cells and tissues, and is still the most crucial cytosolic DNA sensor till now. However, the transcription and expression of cGAS are almost undetectable in porcine kidney cell lines PK-15, which play central roles in the study of swine viral disease. To construct the PK-15 cell lines stably expressing cGAS, the cGAS gene was inserted into the genome of the target cells using lentivirus packaging system, following by the identification of gene transcription, expression, and distribution. Finally, the DNA viruses pseudorabies virus (PRV) and porcine parvovirus (PPV) were used as model viruses to confirm the role of cGAS in innate antiviral immunity. The results showed that cGAS is successfully integrated into the genome of the PK-15 cells, and can be detected in the recombinant cell lines PK-15-cGAS as early as generation 3. The infection of PRV and PPV induced markedly higher levels of interferon-related genes, and consistently the replication of PRV and PPV was dramatically inhibited in PK-15-cGAS cells. In summary, the PK-15 cell lines stably expressing cGAS were prepared successfully, which will help to explore the mechanisms of viral infection as well as immune evasion.