Clustered regularly interspaced short palindromic repeats (CRISPR) / CRISPR-related protein (Cas) system is a heritable prokaryotic adaptive immune system against invading nucleic acids. It is an editing tool for many organisms that can generate highly specific gene double-strand breaks and repair them by non-homologous end joining (NHEJ) or homologous recombination (HR). Due to the complexity of its genetic manipulation, Mycobacterium tuberculosis genome research has many difficulties. Therefore, the emergence of the CRISPR/Cas system has a milestone significance for the research of Mycobacterium tuberculosis. In this review, we focus on the application of the CRISPR/Cas system in Mycobacterium tuberculosis and summarizes the present research progress in order to provide a reference for the selection of Mycobacterium tuberculosis research methods.

Foot-and-mouth disease (FMD) is an acute, hot, and highly contagious infectious disease caused by foot-and-mouth disease virus (FMDV) infection of cloven-hoofed animals. FMDV has seven serotypes and it spreads rapidly, which seriously affects the development of animal husbandry. FMDV is the prototype member of the Aphthovirus genus within the Picornaviridae family, and its genome encodes 4 structural proteins and 10 non-structural proteins. After infecting the host, FMDV uses its proteins to affect the host innate immune response through a variety of pathways and methods, which is beneficial to the microenvironment of FMDV replication. These strategies include FMDV involvement in autophagy, endoplasmic reticulum stress and stress granule formation of cellular processes, subverting the functions of various host proteins, such as hijacking, cleaving host proteins or interfering with the expression of host proteins, removing ubiquitin from host proteins, and inhibiting the phosphorylation of host proteins, which are also the focus of current research. Based on the existing research results, we summarized the research progress of FMDV protein in suppressing the innate immunity of host in recent years, intending to provide a reference for the research and prevention of FMDV.

Copy number variation (CNV) is an important type of genetic variation which widely exists in livestock and poultry genomes. It usually occurs in the form of deletion or duplication of large DNA sequences, which directly affects gene expression and function. CNV is closely related to many genetic diseases and phenotypic traits, and therefore has attracted close attention of animal genetics and breeding workers. The formation causes, action mechanism, detection methods, and the research progress of CNV in poultry were summarized in this article, and the difficulties and problems in current CNV studies were discussed, which aims to provide a reference for the realization of CNV as a molecular genetic marker in poultry for assisted breeding.

At present, human's demand for meat is increasing, but the environment pressure caused by the meat production of livestock is also increasingly apparent. The development of green and sustainable meat substitutes to reduce global carbon emissions and meet the increasing demand for meat has become the hotspot of social concerns. Although artificial meat belongs to food, it still lacks in-depth analysis on whether it brings challenges or opportunities to the livestock industry. In this paper, the classification and development history of artificial meat, the existing core manufacturing technology and market prospect of artificial meat are summarized, and the importance of using the diversity of livestock and poultry germplasm resources to promote cell-based meat are analyzed.

The aim of this study was to obtain the complete coding sequence (CDS) of CCAR1 gene, and to explore its subcellular localization, expression profile and its effects on cell prolification and action mechanism in pig. In this study, the cDNA from kidney tissue of 1-day-old Mashen pigs were used as the template to obtain the full-length CDS of CCAR1 gene by RT-PCR and sequencing. The cellular immunofluorescence staining was used to explore the subcellular localization of CCAR1 in PK15 cells. The temporal and spatial expression profile of CCAR1 was investigated by qRT-PCR in this study. The CCAR1 gene in PK15 cells was knocked out by using CRISPR/Cas9 gene editing technology, and the effects of CCAR1 gene on cell proliferation and expression of cell proliferation and apoptosis related genes were investigated by qRT-PCR, Western blot and CCK8 (cell counting kit 8) technologies in this experiment. The results showed that the complete CDS region of pig CCAR1 gene was 3 459 bp in length (MH301308.1). CCAR1 protein was localized in both cytoplasm and nucleus of PK15 cells. The expression profiles of CCAR1 mRNA between Large White and Mashen pigs was similar, which was expressed in all detected tissues, with the highest expression in kidney and small intestine, middle expression in spleen, liver, cerebellum and muscle, and the lowest expression in heart and subcutaneous fat. Temporal expression results showed that CCAR1 was expressed in both psoas muscle and longissimus dorsi muscle at 3 developmental stages both in Mashen and Large White pigs. The CRISPR/Cas9 gene editing system effectively reduced the expression of CCAR1. CCK8 results showed that after 48 hours of transfection, compared with the control group, the proliferation of cells in the experimental groups were extremely significantly inhibited (P<0.01). After CCAR1 gene knocked out, the expression level of Mki67, a marker of cell proliferation, was significantly decreased (P<0.05). There was no significant difference in the expression level of Caspase3 and the core protein β-catenin in Wnt pathway between control group and experimental groups, and the expression level of downstream target gene C-myc of Wnt pathway was decreased significantly(P<0.05). CCAR1 gene was expressed almost in all tissues at different developmental stages, and affected cell proliferation by regulating the expression levels of Mki67 and C-myc, and played an important role in growth and development of pig.

The aim of this study was to use bioinformatics methods to screen non-synonymous single nucleotide polymorphisms (nsSNPs) sites with potential biological functions in chicken IGF2 gene and provide a theoretical basis for carrying out marker-assisted selection to improve important economic traits of chicken. Twelve nsSNPs sites of chicken IGF2 gene were retrieved from dbSNP database. SIFT, Polyphen-2, PhD-SNP, and SNAP were used to predict the possible functional SNPs. The amino acid stability of mutation sites was analyzed by I-Mutant3.0 and Mupro methods. Multiple sequence alignment and conserved prediction of evolutionary sites were carried out on the amino acid sequence encoded by chicken IGF2 gene, and combined with MutPred2 to predict the possible functional consequences caused by mutations. Finally, Sopma was used to predict the secondary structure of IGF2 wild-type and mutant proteins, and I-TASSER was used to construct their tertiary structure. The results showed that rs740391349 (E29G), rs735633122 (T30P), rs739078786 (L31P), rs736255842 (E35G) and rs736800980 (V37G) might affect the protein function of chicken IGF2, and all the mutations reduced the protein stability of IGF2. Multiple sequence alignment and conservative analysis showed that rs740391349 (E29G), rs735633122 (T30P), and rs736255842 (E35G) were highly conserved and exposed functional residues. MutPred2 and secondary structural analysis showed that mutations at rs735633122 (T30P) and rs736255842 (E35G) both led to a decreased percentage of α-helix. Tertiary structural analysis showed that rs735633122 (T30P) and rs736255842 (E35G) could change the spatial structure of IGF2 protein. In summary, mutations at T30P and E35G seriously affect the structure of chicken IGF2 protein and may be the significant functional SNPs affecting chicken growth and body composition traits.

The purpose of this study was to select the key genes of mammary gland development at different physiological stages in dairy goats by weighted gene co-expression network analysis(WGCNA). GSE14008 mammary gland tissue microarray data set of dairy goats at different physiological stages (pregnancy 46 days, 70 days, 90 days, 110 days and 40 days postpartum) was downloaded from GEO database, and co-expression analysis was carried out using WGCNA package of R language. The target modules were selected by correlation analysis between the modules and physiological stages, and the hub genes were selected according to the connectivity degree. After enrichment analysis of the modules using DAVID website, the protein-protein interaction network of the modules was constructed using String website and the core genes were obtained using Cytoscape software, and finally the target genes were obtained from intersection of core genes and hub genes. A total of 8 443 genes from 18 samples were analyzed for co-expression of weighted genes. The 30 modules were obtained, and 4 target modules related to different physiological stages and 30 hub genes of each module were selected. Meanwhile, protein-protein interaction network of 4 modules and 20 core genes of each network were also obtained. Finally, 13 target genes related to mammary gland development were obtained from the 4 modules, which were UQCR, RGL2, NOTCH1, PTBP1, PPP5C, FZR1, UBE2L3, TNF, MAT2A, ITGB2, GPR18, JAK1 and CSN2 genes. The key processes of mammary gland development at different physiological stages and the genes that played the key role in these processes were elucidated by WGCNA, GO enrichment analysis, PPI network and other bioinformatics techniques, which provided a new idea and clue for the further research on the mechanism of mammary gland development.

The aim of this study was to explore whether miR-17-3p could target KCTD15 to regulate the preadipocytes differentiation of Yanbian Yellow cattle. Bioinformatics softwares were used to identify homology and predict target genes of miR-17-3p. miR-17-3p mimic or miR-17-3p inhibitor and their negative control were transfected into bovine preadipocytes to overexpress or inhibit the expression of miR-17-3p. The binding sites of miR-17-3p to KCTD15 were verified via double luciferase report system. qRT-PCR and Western blot were used to detect the effect of miR-17-3p on the expressions of PPARγ, C/EBPα and KCTD15 both at mRNA and protein levels. Bioinformatics software prediction showed that KCTD15 was the target gene of miR-17-3p, and miR-17-3p had high homology in mammals. The mRNA and protein expressions of adipogenic marker genes PPARγ and C/EBPα after transfection of miR-17-3p mimic were significantly or extremely significantly higher than those of the control group transfected with NC-mimic (P<0.05 or P<0.01). After the expression of miR-17-3p was inhibited, the expressions of PPARγ and C/EBPα were significantly or extremely significantly lower than that in the control group (P<0.05 or P<0.01). Through the dual-luciferase reporter assay verification, the overexpression of miR-17-3p extremely significantly inhibited the fluorescent activity of the luciferase reporter gene vector containing the 3'UTR fragment of KCTD15 (P<0.01). Overexpression of miR-17-3p could also significantly or extremely significantly inhibit the expression of KCTD15 mRNA and protein(P<0.05 or P<0.01). However, inhibition of miR-17-3p expression could significantly increase the expression of KCTD15 mRNA and protein(P<0.05). These results suggest that miR-17-3p may regulate the differentiation of preadipocytes in Yanbian Yellow cattle by inhibiting the expression of KCTD15.

The aim of this study was to verify the up-regulation and down-regulation of TGFβ2 and PPP3CA genes in transcriptome sequencing in monotocous group compared to polytocous group, and to explore the regulatory mechanism of TGFβ2 and PPP3CA genes on ovarian granulosa cells of Qianbei Ma goat. The monotocous and polytocous Qianbei Ma goat ewes were used as the research object. The expression levels of TGFβ2 and PPP3CA genes in the gonadal axis of monotocous and polytocous Qianbei Ma goat were detected by RT-qPCR technology; The eukaryotic expression vector of target genes were constructed. Qianbei Ma goat ovarian granular cells were cultured, the pEGFP-N3-TGFβ2 and pEGFP-N3-PPP3CA recombinant plasmids were transfected into ovarian granular cells by liposome transfection method, and the concentrations of estradiol(E2) and progesterone(P4) in the medium were detected. The effects of TGFβ2 and PPP3CA genes on the proliferation and apoptosis of ovarian granulosa cells were detected using CCK-8 and Annexin V-FITC methods. Then RT-qPCR was used to detect the effect of overexpressing TGFβ2, PPP3CA genes on TGFβ2, PPP3CA, GnRHR, FSHR, BMP4, TIMP3 genes expression at the cell level. RT-qPCR results showed that TGFβ2 and PPP3CA genes were expressed in the tested tissues, and the expression levels in the monotocous group were generally higher than those in the polytocous group; Immunofluorescence result confirmed that the cultured cells were Qianbei Ma goat ovarian granulosa cells; After the eukaryotic expression plasmids of TGFβ2 and PPP3CA genes were transfected into ovarian granulosa cells, the expressions of TGFβ2 and PPP3CA genes significantly increased while the expressions of FSHR, GnRHR, BMP4, TIMP3 genes were significantly inhibited in ovarian granulosa cells (P<0.01). The results of cell proliferation and apoptosis showed that TGFβ2 and PPP3CA genes could promote the apoptosis of granulosa cells and inhibit their proliferation. The result of enzyme-linked immunosorbent assay showed that TGFβ2 and PPP3CA genes could significantly promote the secretion of E2, but had no significant effect on the secretion of P4. The eukaryotic expression vector of TGFβ2 and PPP3CA genes were successfully constructed in this study. Fluorescence quantitative results showed that overexpression of TGFβ2 and PPP3CA genes inhibited the expression of reproduction-related genes, suggesting that TGFβ2 and PPP3CA genes had a certain effect on goat reproduction-related genes expression. These results will provide basic data for further research on the influence of TGFβ2 and PPP3CA genes on the fecundity of Qianbei Ma goat.

The objective of this study was to explore the expression differences of integrin αv in different parts of the fallopian tube at different reproduction stages of yak, and provide some important theoretical foundations for understanding the effect of integrin αv on the reproductive performance of yak. The samples of the tubal umbrella, ampulla, and isthmus of healthy female yaks (3-6 years old) during follicular, luteal and pregnancy stages were collected, the fallopian tubes of the follicular phase, the luteal phase and the pregnancy phase were divided into 9 groups according to the umbrella, ampulla and isthmus. Real-time PCR (qRT-PCR) and Western-blotting (WB) were used to detect the expression of integrin αv gene and protein. The expression of integrin αv protein was localized by immunohistochemistry. Results: 1) qRT-PCR results showed that, during the follicular phase, the expression of the integrin αv gene in the umbrella of the fallopian tube was the highest and extremely significantly higher than those in the ampulla and isthmus (P<0.01). During the luteal phase, the expression of the integrin αv gene in the isthmus of the fallopian tube was the highest and extremely significantly higher than those in the umbrella and ampulla (P<0.01). During the pregnancy phase, the expression of the integrin αv gene in the umbrella of the fallopian tube was the highest and extremely significantly higher than those in the ampulla and isthmus (P<0.01). 2) Western-blotting results showed that, during the follicular phase, the expression of integrin αv protein in the umbrella of the fallopian tube was the highest and extremely significantly higher than those in the ampulla and isthmus (P<0.01). During the luteal phase, the expression of integrin αv protein in the isthmus of the fallopian tube was the highest and extremely significantly higher than those in the umbrella and ampulla (P<0.01). During the pregnancy phase, the expression of integrin αv protein in tubal isthmus and umbrella was not significantly different, but these were extremely significantly higher than that in the ampulla (P<0.01). 3) The results of immunohistochemistry showed that integrin αv protein was positively expressed in ciliated cells, secretory cells, basal cells, muscular layers and serous glands of the fallopian tube umbrella, ampulla and isthmus. The results indicated that the expression of integrin αv in different parts of the fallopian tube at different reproduction stages of the yak were significantly different, which showed that αv might be involved in a series of reproductive processes such as fertilization and early embryonic development, the results provided basic information for the study of yak's reproductive performance.

The aim of this experiment was to study the expression pattern of taste receptor family 1 subtypes 1 (T1R1) and 3 (T1R3) during epididymal development of Congjiang Xiang pig, and to explore the possible role of these taste receptors in mammalian male reproductive function and its potential medical value. In this study, the differential expressions of T1R1 and T1R3 in epididymis at 4 key developmental periods (neonatal (15 d), peri-puberty (30 d), puberty (60 d) and sexual maturity (180 d)) of Congjiang Xiang pigs were analyzed. RT-qPCR, immunohistochemistry (IHC) and Western blot were used to detect the changes and distribution of the two taste receptors in epididymis of Congjiang Xiang pigs at different ages. The results of RT-qPCR showed that the expression of TAS1R1 and TAS1R3 mRNA increased gradually from neonatal (15 d) to sexual maturity (180 d), and there was a significant difference between each period (P<0.01). The results of Western blot showed that the expression of T1R1/T1R3 protein was the highest on the 180 d and the lowest on the 15 d. The average protein abundance of T1R1/T1R3 was as follows: 180 d > 30 d > 60 d > 15 d. The results of IHC showed that T1R1 and T1R3 proteins were distributed in the epididymis of Congjiang Xiang pigs at 4 periods, in which T1R1 protein was mainly concentrated in epithelial cell membrane, especially in basal and narrow cells, while T1R3 protein was strongly positive in stereocilia, annular vacuoles and spermatozoa. In summary, the expression of T1R1/T1R3 in the epididymis of Congjiang Xiang pigs increased gradually from 15 d to the peak of sexual maturation, which was related to the differential expression of T1R1/T1R3 in epithelial basal cells, narrow cells and stereocilia of epididymis. These special expression patterns were time related to the physiological function of epididymis, so it is speculated that T1R1/T1R3 are involved in the regulation of sperm maturation and storage in epididymis.

The study aimed to explore the effect and related molecular mechanisms of folic acid supplementation on the immunity and milking related traits of dairy cattle with subclinical mastitis, which would lay a foundation for folic acid supplementation of cows with subclinical mastitis. A total of 29 lactating cows were selected based on their parity, body weight and somatic cell count (SCC>500×10³ cells·mL^-1 for 2 consecutive months) from a Holstein cow population in a farm near Beijing, China. The 29 cows were randomly divided into feeding group (n=18) and control group (n=11). The 18 cattle in feeding group were fed with extra-coated folic acid (120 mg/500 kg per day) for two weeks, and no treatment in the control group. Then collected DHI data (including somatic cell count, daily milk yield, etc.) were analyzed. Subsequently, four individuals were randomly selected in each group to collect leukocytes, extract RNA and perform transcriptome sequencing. DESeq2 was used to perform differential expression gene analysis using RNA-Seq data. KOBAS3.0 pathway database was used for KEGG and GO functional analysis. Somatic cell counts (SCC) showed a significant decrease (P<0.05) after folic acid supplementation, however, no significant effect was observed on milking related traits. A total of 277 differentially expressed genes (P<0.05, log₂|Fold Change|>1) were detected in the comparison between the two groups, including 105 up-regulated and 172 down-regulated genes. GO analysis showed that these genes were mainly enriched in the immune-related functions terms， including inflammation, immune response, regulation of cytokine production, cell differentiation, leukocyte chemotaxis and migration, and regulation of immune system processes. In addition, KEGG pathway analysis revealed that these DEGs were involved in innate immunity and cellular immune response pathways. In conclusion, folic acid supplementation has an important impact on immune-related traits and expression level of genes related to immune pathways. The current research provides a foundation for the promotion and application of folic acid supplementation.

The purpose of this experiment was to investigate the rumen degradation characteristics of whole sugarcane of dairy cows and its effects of using it replacing alfalfa, oat hay and concentrates in dairy cows' diet on the rumen fermentation, nutrient apparent digestibility, blood biochemical indexes and production performance of dairy cows, in order to open up new ways to replace the increasing prices feedstuff such as alfalfa, oat hay and concentrate in dairy cows' diets with whole sugarcane. In experiment one, the in-situ test, namely the rumen nylon bag test was used to investigate the rumen degradation characteristics of various nutrients of whole sugarcane, leymus chinensis and alfalfa. Experiment two was in vivo test，repeat 3 ⅹ 3 Latin square design was conducted and Holstein dairy cows with similar body weight, parities and milk yield and permanent rumen fistula were chosen to investigate the effect of using whole sugarcane replacing 25% or 50% of imported alfalfa from the Unites States in dairy cows' diet on nutrient digestibility, rumen fermentation characteristics and blood biochemical indexes of dairy cows. In experiment three, 60 Holstein dairy cows with similar parity, milk yield and milk in lactation were selected to perform production test, which aimed to explore the effects of using whole sugarcane replacing 30% alfalfa hay, 50% oat hay or 10% concentrate on the production performance, blood biochemical indexes of dairy cows and the economic benefits of dairy farms. The results showed that：1) The effective degradation rates of dry matter (DM) and acid detergent fiber (ADF) of whole sugarcane were significantly higher than leymus chinensis (P<0.05), and the effective degradation rates of neutral detergent fiber (NDF) and organic matter (OM) were significantly higher than leymus chinensis and alfalfa (P<0.05). 2) There was no significant effect on rumen pH, NH₃-N (except 25% group at the 4 h time point) and volatile fatty acid (VFA) concentrations of dairy cows when using 25% and 50% whole sugarcane to replace imported alfalfa from the Unites States in the diet. There was also no significant effect on DMI and digestibility of DM, CP, OM and ADF, while the digestibility of NDF in the 50% group was significantly increased (P<0.05). In 50% group, the content of serum urea nitrogen in 2 and 8 h was significantly lower than 25% group and control group (P<0.05) and there was no significant difference in the blood content of β-hydroxybutyric acid, rumen VFA and total VFA in each group at different time points. 3) After using 50%, 30% and 10% of whole sugarcane to replace oat hay, alfalfa hay and concentrate, respectively, no significant difference was detected in blood glucose, urea nitrogen, β-hydroxybutyric acid, the esterification of fatty acid (NEFA), and milk yield. Besides, combined with milk yield and feed cost, the economic benefits of each cow in the substitution groups increased by 3.28, 7.48 and 1.62 yuan per day. respectively, compared with control group. It can be seen that the whole sugarcane is a kind of high-quality roughage that can be used by dairy cows with high digestibility, and using it to replace 25% or 50% of imported alfalfa from the Unites States in dairy cows' diet does not affect the rumen fermentation characteristics and rumen apparent digestibility of nutrients of dairy cows. And using whole sugarcane replacing 30% alfalfa hay, 50% oat hay or 10% concentrate will not affect the blood biochemical indicators and production performance of dairy cows, and will also increase the economic benefits of the dairy farm.

The aim of this study was to investigate the effects of different ammonia concentration on production performance, immunity and antioxidant capacity of beef cattle. In this study, 16 healthy Qinchuan cows with an initial weight of (220±5) kg were randomly divided into 4 groups with 4 replicates per group and 1 cattle per replicate.Treated with ammonia at concentrations of<5 (control), (15±3), (30±3) and (45±3) mg·m^-3, respectively. The advanced experiment period was 10 days and the formal period was 30 days. During the trial period, the production performance was recorded(ADG,ADFI and F/G), and serum samples were collected from the jugular vein at day 1, 15, 30. Then the biochemical indicators, immune globulin,cytokines and antioxidant enzymes activities were measured.The results showed as follows:1) Compared with the control group, average daily gain (ADG) significantly decreased when ammonia concentration was 30 mg·m^-3, and average daily feed intake (ADFI) was significantly decreased in all ammonia treatment groups (P<0.05), and feed∶gain (F/G) increased significantly when ammonia concentration was 15 and 30 mg· m^-3 (P<0.05). 2) The content of serum CRE、BUN、ALT、AST and LDH were significantly increased when the ammonia concentration was 45 mg·m^-3 (P<0.05), and the An content increased significantly in all ammonia treatment groups compared with the control group, indicating that ammonia exposure caused damage to liver and kidney function of cattle. 3) The immunoglobulin A (IgA) level was significantly reduced in each ammonia treatment group (P<0.05), the content of IgM was significantly decreased when the ammonia concentration was 30 and 45 mg· m^-3 (P<0.05). However, there was no significant difference in IgG content among the groups (P>0.05). Ammonia exposure significantly increased the levels of IL-6 at 30 and 45 mg·m^-3， and IL-4 content was significantly increased at 45 mg· m^-3 (P<0.05), IFN-γ content was significantly reduced at 30 and 45 mg· m^-3 (P<0.05), thus inducing an inflammatory response of beef cattle. 4) Ammonia exposure significantly decreased T-AOC activity at 30 and 45 mg· m^-3 and GSH-Px activity was significantly decreased at 45 mg· m^-3 (P<0.05)， MDA content significantly increased at 30 mg· m^-3 (P<0.05);No significant difference was observed in the serum SOD and CAT activities in each group (P>0.05). In summary, this study suggested that excessive ammonia exposure reduced growth performance, impaired immunity and antioxidant capacity of beef cattle.

From October 2017 to May 2019, 67 samples of typical goslings gout were collected from Guangdong, Sichuan, Hebei, Shandong, Anhui, Liaoning provinces and other vast goose-raising areas in China. The results showed that goose astrovirus was detected in all samples and about 94.03% of the samples were infected by two different kinds of goose astrovirus. The pathogenicity of goose astrovirus FLX variant strain (named SCCD) which could stably proliferate in goose embryos and kill 10-day-old goose embryos was more virulent than goose astrovirus FLX strain. The new type of goose astrovirus strain (named SDPD) could't stably proliferate in goose embryos and SPF chicken embryos. The whole genome nucleotides homology of goose astrovirus FLX strain with goose astrovirus SCCD strain and the new type goose astrovirus SDPD strain were 89.7% and 58.1%, the amino acids homology of ORF1b gene were 98.4% and 61.0%, the amino acids homology of ORF2 gene were 81.0% and 42.5%, respectively. The newly isolated goose astrovirus strains were inoculated into 1-day-old healthy goslings, the results showed that although the goose astrovirus SCCD strain and the new type goose astrovirus SDPD strain could cause the death of goslings and the changes of hepatitis, kidney swelling and weight loss, none of the goslings showed typical gout characteristics (urate deposition in the organs of the body), however, 44% of goslings were characterized with typical gout which was consistent with the symptoms of natural disease after coinfected by the two goose astrovirus strains. Therefore, the pathogenic agents of goslings gout may be two different kinds of goose astrovirus, namely the new type of goose astrovirus and the variant of goose astrovirus FLX.

Pseudorabies (PR) is caused by the Pseudorabies virus (PRV). It is an acute and hot highly contagious disease infecting livestock and a wide range of wild animals. In order to investigate the relationship between latent infection of Pseudorabies virus and sow production performance，this study collected production parameters of first-parity sows with wild virus gE positive and negtive in a Pseudorabies positive stable intensive farm, including total litter size, healthy litter size, weak litter size, stillbirths, mummified fetus, litter weight, number of weaning live, number of weaning qualified and weaning weight. And compared the production performance of PRV gE antibody negative and positive sows in the same intensive pig farm. The study showed that each PRV gE antibody negative sow could produce 11.96 live piglets per parity. Additionally, PRV gE antibody negative sow could provide more alive, weaning and weaning qualified piglets per parity than infection sows, which were 0.63, 0.18 and 0.28, respectively. Although the average birth weight and average weaning weight of piglets produced by PRV gE antibody positive sows were higher than those produced by negative sows, the weaning qualified rate of antibody negative sows was higher than that of antibody positive sows, indicating that the weaning live piglets produced by antibody negative sows had higher uniformity. In summary, the production performance of PRV gE antibody positive sows was lower than that of the negative sows. Eradication of PR can bring higher profit to the pig farm. Pig farm should actively eradicate the PR.

The present study aimed to isolate phages that could lyse Proteus mirabilis from sewage in Daqing, Heilongjiang, and analyze their biological characteristics and effects on biofilm and provide the basic data for phage therapy as well as prevention. Phages were isolated and purified from sewage by double plate culture method, and their plaque morphology were observed. Morphological characteristics of purified phages were observed by transmission electron microscopy. The optimal multiplicity of infection (MOI), one-step growth curve, thermal stability, pH, nucleic acids analysis and the inhibition and removal effect of biofilm were determined. Three lytic of bacteriophages of Proteus mirabilis were successfully isolated and purified, and named as vB_PmM_S, vB_PmM_W and vB_PmM_X. The phage plaques were circular, clear and transparent with smooth edge. The electron microscope observation showed that the phage particles, those head was twenty-side, head length was about (30±3) nm. The optical MOI was 0.001, the incubation period was 20 min, the burst period was 35 min, and the burst size was 70 PFU·cell^-1. The phages could withstand the temperature range from 10 to 60 ℃ and keep stable titer under pH 5.0-8.0. The phage vB_PmM_S can be stored at 4 ℃ in SM solution for a short time, and stored at -80 ℃ in glycerin (30%) or DMSO (5%) for a long time. The mixture of three bacteriophages can inhibit the biofilm formation and remove the biofilm. The results laid the foundation for the treatment of Proteus mirabilis and urinary tract infections.

This study was conducted to obtain the Clostridium perfringens β toxin (CPB) derivative and to evaluate its virulence and immunoprotection. Based on the known sequence, four amino acid mutations, R212E, L268G, Y266A and W275A, were introduced into the gene of Clostridium perfringens CPB. Meanwhile, genes of Th cell and N-terminal of flagellin were added to 5' of the CPB gene, respectively. Then the gene GTF_NCPB_m4 was optimized and synthesized, and was subsequently cloned into the prokaryotic expression vector pET-30a (＋) for expression and purification to get the recombinant protein, rTF_NCPB_m4. Reactivity of rTF_NCPB_m4with antiserum of Clostridium perfringens type C crude toxins was detected by Western blot. Meanwhile, the toxicity of rTF_NCPB_m4 to mice was evaluated. According to the method prescribed in Chinese Veterinary Pharmacopoeia (2015), rabbits were immunized with rTF_NCPB_m4 to prepare antiserum and detect the neutralizing titer against Clostridium perfringens type C crude toxins. Results showed that rTF_NCPB_m4 was presented predominantly in an insoluble form (inclusion bodies), and it could react with the antiserum of Clostridium perfringens type C crude toxins. rTF_NCPB_m4 with an injection volume of 50 μg was still not fatal to mice. Sera from rabbits immunized with rTF_NCPB_m4can neutralize 10-20 mouse minimum lethal doses (MLD) of Clostridium perfringens type C crude toxins after twice immunization. Moreover, rabbits immunized with rTF_NCPB_m4 can fully resist 1 rabbit MLD of Clostridium perfringens type C crude toxins challenge, whereas all of the rabbits died (4/4) in the control groups. These data suggest that rTF_NCPB_m4 is a potential vaccine candidate for the subunit vaccine of Clostridium perfringens type C.

In this study, we aimed to determine the tissue distribution of clinical isolates of Haemophilus parasuis (HPS) to explore the epidemic distribution and correlation of its serotypes and genotypes, and to provide scientific basis for the effective prevention and control of Glässer's disease. According to the 89 strains of HPS isolated from the clinic, the serotypes of HPS were identified by PCR, and the number of HPS strains in different parts of the isolates was counted. Sequence type (ST) identification analysis, site polymorphism analysis, BURST cluster analysis and UPGMA phylogenetic tree cluster analysis were performed by using the multilocus sequence typing method. Nine serotypes (1, 2, 4, 5, 7, 11, 12, 13, and 14) and undetermined serotypes (NT) were identified from 89 HPS clinical isolates. Serotypes 4,13,7 and 5 were the predominant serotypes, accounting for 28.09%, 22.47%, 13.48% and 10.11%, respectively, and sixty-four strains of HPS were isolated from the lung tissues. ST267, ST268, ST387 and ST365 were dominant genotypes, accounting for 26.97%, 21.35%, 8.99% and 5.62%, respectively, there were 3-13 alleles at each locus, and the polymorphisms ranged from 3(g3pd) to 71(6pgd). The BURST analysis showed that the 89 HPS were divided into 2 single ST types and 11 clonal groups (CC). The phylogenetic tree of UPGMA had 4 branches, the dominant ST type was found in 2, 3 and 4 branches, which was corresponded to the HPS isolates with virulent serotype. The epidemic serotypes and genotypes of HPS are diversified and have obvious genetic heterogeneity. At the same time, ST types and serotypes have some crossover and are related to HPS clinical pathogenicity.

This study aimed to investigate the real microbial composition, antibiotic resistance genes and parasites in feces of giant pandas. Fresh feces of six health giant pandas were collected. The microbial composition and functions, the classes and relative abundance of antibiotic resistance genes (ARGs), and parasites composition in the feces of giant pandas were analyzed by transcriptome sequencing, while the correlations between predominant bacteria and ARGs were determined. The results showed that the giant pandas contained different types of microbiomes, including bacteria, fungi and virus, most of which were bacteria. The main bacterial phyla in giant pandas consisted of Firmicutes and Proteobacteria, and the main fungal phyla consisted of Basidiomycota, Ascomycota and Mucoromycota. No significant difference of microbiomes in abundance between male and female giant pandas was observed (P>0.05). By analyzing the functions of differentially expressed Unigenes, it was found that microbiomes are mainly involved in the carbohydrates and amino acids metabolism. A total of 25 antibiotic classes and 304 ARGs were identified, and the class of efflux pump had the most related genes and highest relative abundance. Besides, a total of 126 parasites species, including nematodes, tapeworms, and trematodes from 63 genera have been found, while nematodes were the dominant species. Correlation analysis revealed that the genera Streptococcus, Escherichia, Enterococcus and Turicibacter significantly and positively correlated with the antibiotic classes of efflux pumps, quinolones, tetracyclines, glycopeptides, polypeptides and sulfonamides. This study revealed the composition of microbiomes, antibiotic resistance genes and parasite species in the feces of giant pandas through transcriptome level, which is of great significance for the prevention and controls microbial diseases and parasitic diseases in giant pandas.

The purpose of this experiment was to screen and identify the outer membrane proteins of Riemerella anatipestifer (R. anatipestifer) interacting with duck C4b-binding protein (C4BP). The preserved R. anatipestifer was resuscitated, then the outer membrane proteins of R. anatipestifer were extracted and His pull-down and LC-MS/MS were conducted by using duck C4BPα as the bait protein, and the candidate outer membrane proteins that might interact with duck C4BP were screened out. The candidate proteins were cloned, prokaryotic expression was conducted and the polyclonal antibodies were prepared by immunizing the mice. Far-western blot was conducted to verify the outer membrane proteins of R. anatipestifer interacting with duck C4BP. The clone and prokaryotic expression of functional domains of candidate proteins and C4BPα were conducted, and Far-western blot was used to identify the interaction sites between candidate proteins and C4BP. The deposition of complement C3b, C4b and C4BP on the surface of R. anatipestifer were detected by ELISA to verify the function of the candidate proteins. The results showed that a total of 3 outer membrane proteins of R.anatipestifer interacting with duck C4BP were screened out by His pull-down and LC-MS/MS, namely ECE-1, SODs and Omp62. The polyclonal antibodies of 3 outer membrane proteins were successfully prepared, the titers of 3 polyclonal antibodies were more than 1∶6 400 by ELISA, and the result of Western blot showed that 3 polyclonal antibodies could specifically react with corresponding recombinant proteins. The results of Far-western blot showed that only ECE-1 could interact with C4BP, and only the full length of ECE-1 could interact with duck C4BP, and the interaction region between duck C4BP and ECE-1 was located in SCR 2 and SCR 3 of C4BPα. Anti-ECE-1 antibody could significantly increase the C3b and C4b deposition on the surface of R. anatipestifer using 3.125% normal duck serum (NDS, P<0.05), while anti-ECE-1 antibody could significantly decrease the deposition of C4BP on the surface of R. anatipestifer using 6.25% NDS (P<0.05). The study successfully screened out and identified one outer membrane protein (ECE-1) of R. anatipestifer interacting with duck C4BP, which provide a basis to further study the mechanism of R. anatipestifer immune escape.

To study the clinicopathology and histopathology of African swine fever (ASF), and to explore the internal relationship between pathological changes and disease occurrence and development and its pathological mechanism, 13 Landrace pigs with bodyweight about 20 kg were intramuscular injected with African swine fever virus (ASFV), strain Pig/HLJ/18 at a dose of 10²HAD₅₀·mL^-1. During the experiment, all the dead pigs were systematically dissected and sampled, paraffin sections were produced, and haematoxylin-eosin staining was performed. Clinicopathological evaluation standards for acute ASF were established, then pathological lesions (classification variables) were expressed by counting frequency and percentage, and the lesion degree (continuous variables) was graded and scored according to different pathological changes of various tissues and organs. The results showed that all infected pigs were in line with the clinical characteristics of ASF, including acute, febrile and highly infectious, with a 100% incidence rate and 100% mortality. The dead pigs showed typical characteristics of septicemia, cadavers prone to corruption, blood clotting adverse or hemolysis, rigor mortis incomplete. The main pathological lesions were hemorrhagic necrotizing lymphadenitis, acute inflammatory splenomegaly (septic spleen), cerebral edema, pulmonary edema and lung consolidation et al. The spleen and lymphonodus are the target organs attacked by ASFV, with the most significant lesions, the earliest occurrence time, the longest duration and the highest frequency. The most prominent pathological changes are blood circulation disorders, including multiple pathological manifestations such as edema, hyperemia, congestion, hemorrhage, infarction, disseminated intravascular coagulation (DIC), and the most important characteristics are hemorrhagic lesions. The inflammatory reaction of lymphocytic exudation caused by ASFV runs through the whole process, especially in the middle and later stages of the course. The results suggest that the main pathological process of acute African swine fever is a typical immune/inflammatory cascade reaction and severe systemic blood circulation disorder, which resulted in the high incidence rate and high mortality rate of acute ASF.

The purpose of this research was to investigate the effect of copper poisoning on the expression of inflammatory factors and cell proliferation in renal tissues of rat. In this study, 32 healthy rats aged 20 days were selected and randomly divided into 4 groups, 8 in each group. The rats were fed as follows: The copper concentration in the control group was 15 mg·kg^-1; high-copper group I: 30 mg·kg^-1; high-copper group Ⅱ: 60 mg·kg^-1; high-copper group Ⅲ: 120 mg·kg^-1. After continuously feeding for 6 months, the mRNA and their proteins expression of Ki-67, PCNA, IL-1β, IL-2, IL-6, IL-18, NF-κB, TNF-α in renal tissues were detected. The results demonstrated that the relative expressions of Ki-67，PCNA mRNA and proteins in renal tissues were significantly decreased with the increase of copper content in feed (P<0.05). Meanwhile, the mRNA expression levels of inflammatory factors including IL-1β, IL-2, IL-6, IL-18, NF-κB, TNF-α in renal tissues were significantly increased (P<0.05). Besides, the relative expression of IL-1β protein was up-regulated in a dose dependent manner, and the relative expression levels of IL-6 and IL-18 proteins increased first and then decreased. In summary, the finding suggested that high copper stimulation inhibited the cell proliferation of rat kidney tissue, promoted the expression of inflammatory factors, and caused inflammatory damage.

A method for the detection of curcumin in pig plasma by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was established firstly and the pharmacokinetics of curcumin solid dispersion and curcumin premix in piglets were studied. Sixteen healthy piglets (Yorkshire×Changbai), seven-week aged, half male and half female, were randomly divided into two groups receiving curcumin solid dispersant and curcumin premix orally at the dose of 100 mg·kg^-1, respectively. Then plasma samples were collected at different time points, and the blood concentration of curcumin was determined by HPLC-MS/MS. The WinNonlin 5.2.1 software was used to analyze and calculate the pharmacokinetic parameters. The pharmacokinetic parameters of curcumin solid dispersion and curcumin premix were as follows: the area under the curve (AUC) was (104.53±38.67) and (37.82±11.48) h·ng·mL^-1, time to peak concentration (T_max) was (3.25±0.38) and (2.31±0.37) h, peak concentration (C_max) was (26.65±9.65) and (9.55±2.75) ng·mL^-1, respectively, elimination half-life time (t_1/2β) was (3.55±2.17) and (6.93±0.86) h, mean residence time (MRT) was (5.23±0.53) and (4.26±0.47) h. The statistical analysis showed significant differentce (P<0.01) between curcumin solid dispersion and premix in parameters, the T_max of curcumin solid dispersion was delayed significantly, the C_max was increased obviously and the AUC was improved after the piglets were given curcumin solid dispersion. Compared with curcumin premix, the relative bioavailability of curcumin solid dispersion was 280.39%. The results showed that curcumin solid dispersion could improve the dissolution and absorption of curcumin in the intestinal tract and improve the relative bioavailability of curcumin, which provided a scientific basis for the development and clinical application of curcumin solid dispersions in the future.

This study aimed to investigate the effect of arsenic trioxide (ATO) on oxidative stress and methionine sulfoxide reductase (Msrs) expression in chicken liver. A total of 32 one-day-old chicks were divided into control group (saline solution), low-dose group (1 mg·kg^-1 ATO solution), medium-dose group (3 mg·kg^-1 ATO solution) and high-dose group (9 mg·kg^-1 ATO solution). Chickens were treated by gavage with ATO solution of corresponding concentration once a day for 5 weeks. The liver coefficient, histopathological sections, MDA level, SOD activity, Msrs gene and protein expression levels of liver were analyzed. The results showed that compared with the control group, the liver coefficient of the medium- and high-dose groups were significantly increased (P<0.01), and the liver congestion and vesicular degeneration were found in the medium- and high-dose groups. Meanwhile, the experimental groups had significantly increased MDA levels (P<0.01) and decreased SOD activity (P<0.01) as compared with the control group; MsrA, MsrB1, MsrB3 gene mRNA expression levels in the ATO treatment groups were significantly increased compared with the control group (P<0.01 or P<0.001); However, the protein levels of MsrA were reduced significantly (P<0.01). The results indicated that ATO induced oxidative stress in the liver of chickens, and at the same time promoted the expression of Msrs in liver to clear ROS and reduce liver oxidative damage.

This experiment was conducted to explore the mechanism of Gegen Qinlian decoction in treating swine diarrhea. The chemical composition and targets of Gegen Qinlian decoction were searched by the analysis of the traditional Chinese medicine system pharmacology platform (TCMSP). The software of Cytoscape 3.7.1 was used to construct and analyze compound-targets networks, protein-protein interaction (PPI) network and key targets-signaling pathway network. The results showed that 60 active compounds including daidzein, puerarin, oroxindin and formononetin were obtained after screening. The compounds acted on 206 targets, among which TP53, TNF, IL-6, IL-8, Jun and STAT3 were the key targets. These key targets involve in PI3K-Akt signaling pathway, Viral carcinogenesis, and so on. Based on the results of network pharmacological analysis, Gegen Qinlian decoction may regulate diarrhea and fever through TP53, TNF, IL-6, IL-8, Jun, STAT3 and other targets. The paper provides theoretical basis and clues for the further study of Gegen Qinlian decoction in the treatment of swine diarrhea.

The objective of the current study was to explore the anti-inflammatory mechanism of Shenfukang granule on chicken kidney swelling. Ninety healthy chickens were divided into blank group (n=15) and test group (n=75), the test group was replicated by the chicken kidney swelling model through feeding high-calcium and high-protein diet. After successful modeling, the test group was randomly divided into model group, positive drug group, Shenfukang high, medium and low dose group (n=15), Shenfukang granule 2.0, 1.0 and 0.5 g·L^-1, positive drug group, drinking water to Canglan oral liquid 1.0 mL·L^-1, The drinking water of the model group was given 1.0 mL·L^-1 of normal saline for 2 times·d^-1 and 5 days. Subpteral venous blood was collected before modeling, after modeling and 2 days after the last administration to isolate the serum. The content changes of serum IL-1, TNF-α, MCP-1 and TGF-β1 content were detected by ELISA. RT-PCR was used to detect renal IL-1R, IκBα, NF-κB mRNA transcription. Results were as follows: Compared with the blank group, the serum contents of IL-1, TNF-1, MCP-1 and TGF-β1 were significantly increased in the model group (P<0.05), the transcription of IL-1R, NF-κBp65 mRNA in kidney was significantly increased (P<0.01), and IκBα mRNA in kidney was significantly decreased (P<0.01). After drinking water treatment, the serum IL-1, TNF-α, MCP-1 and TGF-β1 contents of Shenfukang granule were significantly decreased in the high and low dose group and the positive drug group (P<0.01). The results of the current study suggested that Shenfukang granule played an anti-inflammatory role in inhibiting the IL-1R/NF-κB signaling pathway by reducing the contents of IL-1, TNF-α, MCP-1 and TGF-β1 in serum of kidney swelling chicken.

To explore the biological function of NADH oxidase NOX2 from Mycoplasma bovis (Mb), according to the nox2 gene sequence of Mb strain Hubei (GenBank.CP002513.1), the primers were designed and the nox2 gene of Mb strain Lintao was amplified by PCR. Based on sequencing and gene optimization, the prokaryotic expression vector pET-nox2 was constructed, and was expressed in Escherichia coli Rosetta (DE3). Subsequently, the analysis of the enzymatic activity and the immunogenicity of recombinant proteins rMbNOX2 were completed. And then the subcellular localization of NOX2, complement-dependent bactericidal activity of anti-rMbNOX2 serum, and as well as inhibition effect of anti-rMbNOX2 serum to Mb adhering to host cells was determined. The results showed that the CDS sequence of the nox2 gene of Mb Lintao strain was 1 350 bp, and showed 99.93% homology with nox2 gene of all Mb except Mb JF4278 strain in GenBank. The result of SDS-PAGE displayed the optimized nox2 was successfully expressed in E. coli. The recombinant protein rMbNOX2 was about 67 ku. Enzyme activity analysis showed that the purified rMbNOX2 had good enzymatic activity. The results of ELISA and Western blot showed that the rMbNOX2 has excellent immunogenicity, and the Mb NOX2 distribute both in the cell membrane and cytoplasm, but it's more distributed in the cytoplasm. Complement-dependent mycoplasmacidal assay and adherence inhibition assay confirmed anti-rMbNOX2 serum has distinct complement-dependent mycoplasmacidal activity and can also effectively inhibit the adherence of Mb to host cells. The results of this study lay a foundation for further study on the biological function of Mb NOX2.

The objective of the present study was to investigate the biofilm formation, antimicrobial resistance, virulence genes, and agr genotypes of Staphylococcus aureus (S. aureus) isolated from bovine mastitis cases, and to analyze the correlations between agr genotypes and virulence genes. Biofilm formation, antimicrobial resistance, and virulence genes for 336 strains of S. aureus were detected by microtiter plate method, disk diffusion method, and PCR respectively, and the agr typing of tested strains was determined by multiplex PCR. The results showed that all 336 strains of S. aureus from bovine mastitis were biofilm producers, among which 52.1% and 47.9% of isolates tested were moderate (++) and strong (+++) biofilm producers, respectively. Antimicrobial susceptibility testing demonstrated that S. aureus strains were highly resistant to penicillin, with a resistance rate of 91.7%, followed by erythromycin (89.6%), kanamycin (72.9%), clindamycin (66.7%) and gentamicin (60.4%). However, all isolates were sensitive to nitrofurantoin and linezolid. PCR results showed that the prevalence of fnbA gene was the highest (99.7%), followed by icaD (98.2%), icaA (89.6%), clfA (86.0%), cna (56.0%), and bap (14.6%) genes. Moreover, the sea, seb, sec and tst genes were found in 26.5%, 8.3%, 6.8% and 8.3% of the isolates, respectively. The agr typing results showed that S. aureus strains belonging to agr Ⅰ was predominant in our study, accounting for 77.1% of the isolates, and the frequencies of agr Ⅱ, agr Ⅲ and agr Ⅳ genotypes were 14.0%, 4.8% and 2.1%, respectively. Statistical analysis indicated that the strains of S. aureus belonging to agr Ⅰ genotype have the potential to carry more virulence genes, while no toxin genes could be found in any of the strains belonging to agr Ⅳ. The results revealed high antimicrobial resistance to common antimicrobial agents in S. aureus isolated from bovine mastitis milk samples. Moreover, agr Ⅰ was the predominant genotype with diverse toxin genes in S. aureus from bovine mastitis, and the potential hazard should be of concern.