Recently, wooden breast (WB) and white striping (WS), the newly reported myopathies attracted wide attention of global poultry industry for their increasing incidences and negative effects on meat quality and broiler performance.WS is characterized by the occurrence of white fat striations parallel to muscle fibers, while WB is mainly featured with obvious hardening of breast meat fillets. These two myopathies share similar histological lesions and often appear together. They can adversely affect breast fillet’s appearance, nutrition value, ability for further processes and consumption intention, which result in huge economic loss to the industry. In recent years,most of the studies on WS and WB were reported by international research groups, few by Chinese research teams.In this review,we summarized the research progress of WS and WB from the aspect of pathological features,impacts on meat quality and nutrition value, the underlying causes and possible detailed mechanism in order to provide reference and theoretical basis for future studies.

Kisspeptin is the coding product of the Kiss1 gene, which can bind to its G protein-coupled receptor GPR54 and activate the PLC/PKC/MAPK signaling pathway, thereby playing an important role in inhibiting tumor metastasis, regulating animal reproduction and initiating the estrus. Kisspeptin/GPR54 is not only expressed in hypothalamus but also widely expressed in pituitary and gonads, which is involved in regulating reproduction of animals. We summarizes the latest researches about kisspeptin/GPR54 regulating reproduction and puberty initiation of animals in the hypothalamic-pituitary-gonadal axis. More importantly, the location and distribution of kisspeptin/GPR54 and its possible direct regulation on gametogenesis were highlighted in the gonads in this article. Besides, some problems faced in kisspeptin/GPR54 related research and its future research directions were concluded, which will better help the research and application of kisspentin/GPR54 in animal reproduction.

The intestine is the primary organ responsible for digestion and absorption of nutrients and is frequently subjected to external environmental stimulations leading to the development of inflammatory bowel disease (IBD), which can cause serious harm to intestinal health in animals. Dietary amino acids play important roles in promoting intestinal development and maintaining intestinal health and exert diverse effects through multiple signaling pathways on the prevention and treatment of IBD, including affecting the physiological activities of intestinal epithelial cells, improving intestinal barrier function, reducing intestinal oxidative damage, regulating the production of inflammatory cytokines, and promoting the expression of endogenous antimicrobial peptides, and involved in main signaling pathways including AMP-activated kinase (AMPK), mammalian target of rapamycin (mTOR), mitogen-activated protein kinase (MAPK), Toll-like receptors (TLRs), nucleotide binding oligomerization domain (NOD)/nuclear factor kappa-B (NF-κB). In this review, basic characteristics of IBD, effects and involved signaling pathways of the amino acids on IBD, and effects and applications of amino acids in maintaining intestinal health in livestock and poultry production were reviewed, so as to provide effective clues and strategies for dietary nutrients in the prevention and treatment of IBD.

Currently, corona virus disease 2019 (COVID-19) caused by SARS-CoV-2 is still raging worldwide, which attracts great concern about coronavirus originated from animal. Porcine deltacoronavirus (PDCoV) and swine acute diarrhea syndrome coronavirus (SADS-CoV) are newly discovered porcine coronaviruses in recent years, they not only damage pig industry severely but also pose potential threats to public health. This article reviews the literature concerning the etiology, origin and evolution, pathogenicity, and diagnostics of PDCoV and SADS-CoV, presenting the prospect of research work. We aim to expand understanding on SARS-CoV-2 and provide references for subsequent research on PDCoV and SADS-CoV.

This study aimed to evaluate the actual genetic improvement effect of genomic selection in Large White boars through progeny testing in production performance. Nine hundred and thirteen Large White pigs were used to construct a reference group, and 823 new-born Large White boars were used to implement the first genomic selection through ssGBLUP before castration. The second genomic selection were carried out after performance testing, then 10 boars with significant difference in production performance were selected and their offsprings were compared in phenotypic values, estimated breeding values of growth traits and selection index. The results showed that the accuracies of genomic prediction on age at 100 kg body weight, 100 kg backfat thickness and total number born increased from 0.56, 0.67 and 0.64 in the first genomic selection to 0.73, 0.73 and 0.67 in the second genomic selection, respectively. The correlation coefficient of maternal selection index between the two genomic selection before castration and after performance testing was 0.82, which indicated that the first genomic selection before castration was accurate enough to make early selection on boars. According to the genomic breeding values and maternal selection index of 10 selected boars, two groups with high and low production performance were set up. The progeny testing showed that the difference of average phenotypic value between groups was 2.58 days, and the difference of average evaluated breeding value(EBV) between groups was 3.08 days in age at 100 kg body weight, those were 1.15 mm and 1.03 mm in 100 kg backfat thickness, respectively, and the difference in the mean of the comprehensive maternal index was 9.3, all the differences(except age at 100 kg body weight) were extremely significant. This study prove that the offspring of boars with significant differences in genomic evaluation have significant differences in phenotypic values and breeding values, which indicate that, through genomic selection, excellent breeding boars can be selected and their genetic superiority can be passed to their offsprings.

This study aimed to compare the accuracy of the genomic estimated breeding value (GEBV) using reduced-representation genome sequencing technology and SNP chip technology to implement genomic selection. A total of 395 individuals (212♂+ 183♀, from 8 half-sib families) were randomly selected from F₂ generation of AH broiler resource population, and genotyped with 10×specific-locus amplified fragment sequencing (SLAF-seq) and Illumina Chicken 60K SNP BeadChip. Genomic best linear unbiased prediction (GBLUP) and BayesCπ were used to compare the accuracy of genomic estimated breeding values (GEBV) for 6 traits: body weight at the 6^th week, body weight at the 12^th week, average daily gain (ADG), average daily feed intake (ADFI), feed conversion ratio (FCR) and residual feed intake (RFI). A 5-fold cross validation procedure was used to verify the accuracies of GEBV between prediction models and between genotyping platforms. The results showed that there was no significant difference between accuracies of GEBV predicted by GBLUP and BayesCπ using the same genotyping platform(P>0.05). The superiority of the two genotyping platforms was different for different traits. For body weight at the 6^th week, the accuracy of GEBV was higher using chip SNPs (P<0.05). On the contrary, the accuracy was higher using SLAF-seq for residual feed intake (P<0.05). Comprehensive comparison of the means of GEBV for 6 traits, the difference between the two genotyping platforms was less than 0.01, therefore, both high throughput sequencing and chip SNPs can be used for genomic selection in yellow-feathered broiler.

The study aimed to explore the intestinal microbial community changes after transfering the IGF-1 gene into the superfine wool sheep, which would lead to potential problems in the biosafety of transgenic sheep. Forty one individuals in IGF-1 transgene positive group (GP group, female(GPF) 24 and male(GPM) 17), 43 individuals in genetically modified negative group (GN group, female(GNF) 25 and male(GNM) 18) and 29 individuals in non-genetically modified group(NG group, female(NGF) 18 and male(NGM) 11) from clinical healthy sheep in the same field were randomly selected. The rectal fecal samples were collected. The Ⅱlumina HiSeq high-throughput sequencing technology was used to determine bacterial 16S rRNA V3-V4 area sequence, and the bacteria community composition in fecal samples of transgenic superfine wool sheep and non-transgenic sheep was analyzed. There were 17 phyla, 32 classes, 56 orders, 94 families, 228 genera and 185 species. LEfSe analysis showed that there were 10 biomarkers in the GPF group, there were 5 biomarkers in GPM group, all of which were common colonization bacteria of ruminant intestinal flora. The biomarkers in NGF group was Bacteroides BS11 family, the biomarkers in NGM group was Firmicutes. They were the main dominant bacteria of ruminant intestinal flora. Analysis of shared bacteria of intestinal flora in 3 groups showed that the proportion of shared bacteria was as high as 91%-94% in 6 taxa of genus and family. It was confirmed that the introduction of IGF-1 gene did not change the overall distribution of intestinal flora in superfine wool sheep, and the transgenic sheep were biosafety and environmental safety.

The study aimed to explore the phylogeny and genetic diversity of 3 hare species in Xinjiang by molecular genetics methods, define the relationship and taxonomy status, assess diversity level of Lepus in Xinjiang, and provide the basic data for conservation genetics of hares in Xinjiang and even in China. Three mitochondrial DNA genes, COI, ND4 and 16S rRNA, were used as molecular markers, and the sequences of 3 genes of 57 samples collected from 8 different regions (4 geographic groups) in Xinjiang were determined by PCR amplification and sequencing technology. After the sequences of COI, ND4 and 16S rRNA of each sample were revised and pooled together, data were analyzed with softwares such as MEGA 7, DNAsp 6, Arlequin 3.1 and MrBayes 3.2. A total of 43 haplotypes were detected from the combined sequences of 3 genes of 57 hare samples. Five distinct clades (A-E) and 3 clusters were clearly showed in phylogenetic tree and median-joining network (MJN). Furthermore, the genetic distance between 3 clusters reached the level of species (4.21%-9.09%). However, the genetic distance between hares from northern Xinjiang (Clade E) and those from central Xinjiang (Clade D) were not up to the level of species (≤2.26%) in the third cluster. The haplotype diversity (h) of Lepus yarkandensis, Lepus tibetanus pamirensis and Lepus tolai lehmanni were higher(0.979±0.014, 0.972±0.064 and 0.972±0.064, respectively), while the nucleotide diversity (π) of the L. t. lehmanni and L. t. centrasiaticus were higher (0.033±0.018 and 0.023±0.015, respectively). Based on comprehensive analysis of 3 genes of mitochondrion and reference with published research, it is suggested that hares from southwestern Pamir Plateau of Xinjiang should belong to L. t. pamirensis. Meanwhile，hares distributed in northern and central Xinjiang might be considered as L. t. lehmanni and L. t. centrasiaticus. Moreover, there is abundant genetic diversity in the 3 hare species in Xinjiang, and the obvious phylogeographic pattern is showed.

The aim of this study was to screen and analyze the hair color related genes of Tarim red deer based on RNA-Seq technology. Using Illumina Hi Seq TM2000 sequencing platform, the skin tissue of Tarim red deer and Tianshan red deer were sequenced by transcriptome. After quality control and assembly, the obtained sequences were compared with the annotations in NR, Swiss-Prot, COG, KOG, KEGG, GO and Pfam databases, and the differentially expressed genes were screened, functional annotated and enriched. The results showed that 25 038 Unigenes with annotated information were obtained by sequencing. The comparative analysis showed that there were 922 differentially expressed genes between Tarim red deer and Tianshan red deer, of which 495 were up-regulated and 427 were down-regulated. The results of GO functional enrichment analysis showed that 568 differentially expressed genes were enriched in 61 GO terms and participated in biological processes, cell components and molecular functions, respectively. The enrichment analysis of KEGG metabolic pathway showed that the most significant metabolic pathway enriched by differentially expressed genes was the ECM-receptor interaction. Real-time fluorescence quantitative PCR (qRT-PCR) was used to analyze the transcriptional level changes of 7 candidate genes related to hair color to verify the accuracy and reliability of transcriptome sequencing results. The expression trend of these genes was consistent with the transcriptome sequencing results. ECM-receptor interaction, protein digestion and absorption, PI3K-Akt signaling pathway and tyrosine related to melanin synthesis may be related to regulation of hair color of Tarim red deer; Candidate genes MITF, Ggt1, VDR, PTPRF, CⅡTA, ARPC5L, POMC may play an important role in hair color formation of Tarim red deer. The results of this study will provide abundant experimental data for studying the molecular regulatory mechanism of hair color related genes and exploring the potential new genes in Tarim red deer.

The purpose of this study was to estimate the genetic parameters of semen traits of Landrace boars in South China and to analyze the influence of the age and season of semen collection on semen traits, so as to provide theoretical basis for formulating reasonable boar breeding program. The Asreml-R was used to analyze the 107 221 semen data of 1 605 Landrace boars from 2 AI stations in Southern China, the single-trait repeatability animal model was used to estimate the variance components, heritability and repeatability of each semen traits, two-trait repeatability animal model was used to estimate genetic correlation and phenotypic correlation of semen volume, semen density, sperm motility and percentage of abnormal sperm. The general linear model of R language program was used to analyze the influence of age and season of semen collection on semen traits. The results showed that the semen volume and percentage of abnormal sperm had medium heritability (0.23 and 0.38), and the variation coefficient of percentage of abnormal sperm was 85.42%, while the other traits had low heritability (0.07-0.19). Semen volume-semen density and sperm motility-percentage of abnormal sperm had extremely significantly negative genetic correlation(-0.77 and -0.90, respectively). Semen density-sperm motility showed extremely significantly positive genetic correlation (0.50). The influence of the age of semen collection on semen traits was significant(P<0.05). After boars reached sexual maturity, semen volume showed a significant increase, semen density and percentage of abnormal sperm showed a general decrease, and total sperm number and functional sperm number were significantly higher in the 13-18-month of age group than those in the other groups (P<0.05). The semen density was the highest in spring, and the total sperm number and functional sperm number were significantly higher in autumn and winter than those in spring and summer (P<0.05). In conclusion, the semen volume and percentage of abnormal sperm of Landrace boars can be selected as candidate traits. In the boar production management, boar whose semen production after 36 months of age decline, will be considered to update. Boar semen quality was significantly higher in autumn and winter than those in spring and summer. It is suggested that cooling work should be done in advance in Southern China in summer.

The purpose of this study was to explore the effects of KDM1A on the meiotic maturation and developmental potential of yak oocytes. The specific inhibitor GSK-KDM1A of KDM1A was added into in vitro maturation medium of yak oocytes. After 24 h in vitro culture of yak cumulus oocyte complexes (COCs), the expansion of cumulus cells and the extrusion of the first polar body were observed. The expression level of KDM1A in oocyte was measured by immunofluorescence during in vitro culture. The expression levels of Kdm1a, Oct-4, Sox-2 and Nanog in oocytes were detected by RT-qPCR. Then, yak oocytes were fertilized after in vitro culture, and the cleavage rate and blastocyst formation rate were observed, respectively. The results showed that the cumulus cells expansion in GSK-KDM1A groups were significant lower than those in control group (P<0.05) after 24 h culture, and the cumulus cells expansion and the first polar body extrusion rate in 320 nmol·L^-1 group were significantly lower than those in 160 nmol·L^-1 group (P<0.05). During oocyte in vitro maturation, Kdm1a showed dynamic expression profile, and the expression level of Kdm1a in MⅠ-stage was significantly lower than that in GV-stage and MⅡ-stage (P<0.05). The GSK-KDM1A could significantly inhibit the expression of KDM1A protein in oocytes (P<0.05), and the expression of KDM1A in 320 nmol·L^-1 group was significantly lower than that in 160 nmol·L^-1 group (P<0.05). The expression levels of Oct-4 and Sox-2 in GSK-KDM1A group were significantly higher than that in control group (P<0.05), but there was no significant difference in the expression of Nanog (P>0.05). After 24 h culture, the cleavage rates of oocytes in GSK-KDM1A groups were significantly lower than those in control group (P<0.05), while the blastocyst formation rate was not significantly different (P<0.05). In conclusion, KDM1A is involved in regulating the meiotic maturation process of yak oocytes. GSK-KDM1A can effectively inhibit the expression of KDM1A, affect the meiotic maturation and developmental potential of oocytes, which reveal that KDM1A plays an important role in this process.

The purpose of the present study was to explore the optimal condition for the isolation and culture of rabbit primordial germ cells(PGCs) in vitro, and to establish a mature and stable method for the isolation and culture of rabbit PGCs in vitro. Firstly, trypsin digestion and mechanical methods were used to explore the best method for isolating PGCs from 14- to 18-day-old fetal rabbits in vitro, and trypsin digestion, mechanical method and digestion with the feeder layer were used to explore the optimal way to passage the isolated PGCs. Then, 4 different culture media(A, B, C and D (control)) were used to explore the effect of different concentrations of cytokines on the morphological changes and colony formation of rabbit PGCs. Secondly, rabbit PGCs were identified by alkaline phosphatase staining (AKP), and the expression of transcription factor Oct-4 was detected by real-time PCR (RT-PCR). The results showed that the number of rabbit PGCs colonies isolated by mechanical method was 2.2 times higher than those isolated by trypsin digestion method. Rabbit embryo fibroblasts (REF) were passaged to the second generation (P2) using trypsin digestion method and digestion with the feeder layer method, and REF were passaged to the fourth generation (P4) using the mechanical method. The largest number of colonies and better colony morphology were observed when PGCs were cultured in medium B(basic medium＋10% fetal bovine serum (FBS) ＋2 ng·mL^-1 of transforming growth factor-β1 (TGF-β1) ＋4 ng·mL^-1of basic fibroblast growth factor (bFGF) ＋10 ng·mL^-1leukemia inhibitory factor (LIF)). Rabbit PGCs remained undifferentiated for a longer time when cultured in medium B. AKP staining showed that the isolated PGCs was positive in red and black, and expression of Oct-4 gene was also detected in the isolated PGCs by RT-PCR. The results showed that mechanical isolation method and mechanical passage method were the optimal ways to isolate and passage rabbit primary PGCs, respectively. Supplementation with appropriate concentration of cytokines into culture medium was beneficial for rabbit PGCs to maintain a large number of colonies and undifferentiated state for a longer time in vitro. In the present study, the culture methods of rabbit PGCs in vitro were screened and optimized, which would lay a technical foundation for the further establishment of a stable and mature rabbit PGCs cell line.

This experiment was conducted to study the effects of zinc polyaspartic acid (PASP) monohydrate on growth performance,nutrient apparent digestibility, serum indexes, tissue and organ zinc accumulations and zinc emission of growing pigs, and to determine the substitution effect of zinc polyaspartic acid for zinc sulfate. Ninety Duroc×Landrace×Large White growing pigs with the body weight of (31.73±3.50) kg were randomly assigned into 3 groups with 6 replicates per group and 5 pigs per replicate. The zinc level and source of the control group was 80 mg·kg^-1 zinc sulfate, while group Ⅰ and group Ⅱ were 60 and 40 mg·kg^-1 zinc polyaspartic acid monohydrate， respectively. Pre-trial period was 7 days, and formal experiment period was 30 days. The results showed as follows: 1) There was no significant difference in average daily gain, feed to weight ratio and diarrhea rate among control group and test groups (P>0.05). 2) The calcium (Ca) apparent digestibilities of test group I and test group Ⅱ were significantly higher than that of the control group (P<0.05); There was no significant difference among groups in apparent digestibility of crude protein (CP), crude fat (EE), crude fiber (CF), crude ash (Ash) and phosphorus (P) (P>0.05). The apparent digestibility of zinc in the test group Ⅰ was significantly higher than those of the control group and the test group Ⅱ (P<0.05). 3) Compared with the control group， the fecal zinc content in the test group Ⅰ and Ⅱ were decreased by 21.65% and 30.87%, respectively, and the difference was significant (P<0.01). 4)The glutamate transaminase (ALT) activity of control group was significantly higher than those of test group Ⅰ and Ⅱ(P<0.05), the globulin(GLB) level of test group Ⅰ was significantly higher than those of control group and test groupⅡ(P<0.05), the indexes of blood glucose (GLU), total protein (TP), albumin (ALB), total cholesterol (TC) and alkaline phosphatase (AKP) were not significantly different among groups (P>0.05). 5)There were no significant difference of the zinc accumulations in liver, kidney, spleen, pancreas, bone, hair, longgissimus dorsi among groups (P>0.05), the serum zinc concentrations of test group Ⅰ and Ⅱ were significantly higher than that of control group (P<0.05). In summary, under this experimental condition, 40 mg·kg^-1zinc polyaspartic acid monohydrate could satisfy the needs of growing pigs at this stage, the fecal zinc content was significantly reduced, thus proving the feasibility of adding polyaspartic zinc instead of high dose zinc sulfate in growing pigs’ feed, and the emission reduction of zinc.

The aim of this study was to identify pathogens, which caused pericardial effusion, hepatomegaly and bleeding at a chicken farm in Henan province, and furthermore to analyze effectiveness of immunogenic proteins of the pathogen. This study employed virus isolation, serological assays, PCR and sequencing analysis, animal experimentation, E. coli expression and protein purification, immunogenicity and challenge test and other methods. The results showed that virus was isolated in 7-day-old SPF chicken embryonated eggs, inoculated via the yolk sac route by two blind passages. Viral confirmation was carried out using PCR techniques, and showed a 900-bp-long fragment which shared a 100% homology with the Hexon gene of the serotype C4 strain. Serum neutralization results indicated that this isolate avian adenovirus was the group I type 4 avian adenovirus, named HN-ZK strain. The virus could induce CPE on primary hepatocytes of chicken embryo, and its titer was 10^7.5 TCID₅₀·0.1 mL^-1. Animal experimentation illustrated that the isolated virus caused 100% (10/10) typical symptoms and pathogenicity in 35-day-old SPF chickens. Furthermore, the DNA of the isolate virus was used as a template to express Fiber-2 fragment, with a molecular weight of 33 kDa by using the E. coli expression system, and the protein was concentrated and purified to 300 mg·mL^-1 after centrifugation and purification. SPF chickens were immunized with different doses of the purified Fiber-2 protein, and showed that a dose of 20 μg per chick was completely resistant to challenge of the virulent virus, suggesting the purified Fiber-2 protein has better immunogenicity. This study can provide data for diagnosing the hydropericardium hepatitis syndrome, and developing a genetic engineering subunit vaccine.

To obtain the highly immunogenic recombinant CD2v antigen of African swine fever virus (ASFV), the amino acid sequence of CD2v was analyzed for antigenic index using bioinformatics software and the intracytoplasmic region with high antigenic index was expressed in E. coli as an elastin-like polypeptide (ELP) fusion protein. After optimization of the conditions for inverse transition cycling (ITC), ELP-CD2v fusion protein was purified by ITC in the presence of different concentrations of Triton X-100 and the ELP tag was cleaved with active inclusion bodies of tobacco etch virus (TEV) protease. The tag-free recombinant CD2v antigen was recovered by an additional round of ITC and identified by Western blotting. By using the recombinant CD2v antigen, an indirect ELISA was established and used to detect ASFV antibody-positive and antibody-negative sera in parallel with the multi-antigen ELISA. The results showed that ELP-CD2v fusion protein was expressed correctly in E. coli with an optimal transition temperature of 28 ℃ at 1.5 mol·L^-1 NaCl. After one cycle of ITC in the presence of 0.2% Triton X-100, ELP-CD2v fusion protein was purified to 76.3% purity. The ELP tag was cleaved efficiently with the TEV protease and removed after an additional round of ITC. The recovered recombinant CD2v protein had a purity of 91.7%, which was recognized by pig anti-ASFV serum. For 30 serum samples detected by ASFV multi-antigen ELISA, recombinant CD2v ELISA showed that all of 15 antibody-negative sera were CD2v antibody negative and all of 15 antibody-positive sera were CD2v antibody positive. These data suggest that the presence of immune dominant epitopes in the intracytoplasmic region of CD2v protein and the potential application of the recombinant CD2v antigen for ASFV antibody detection.

Vaccination with inactivated vaccine is an important measure to prevent and control foot-and-mouth disease (FMD), however, the immune effect and antigenic purity of inactivated vaccines are two major concerns for the establishment and evaluation of FMD free zones with vaccination. In this study, four groups of FMD type O and type A bivalent inactivated vaccines from 3 FMD vaccine manufacturers (designated as A, B and C) were selected to inoculate healthy juvenile cattle of FMD free. All cattle were immunized 3 or 4 times at a 1-month interval. Serum samples were collected before and after 1 month of every vaccination to determine the level of antibody to structural protein and non-structural protein. Results：(1) The qualified rates of antibody to structural protein: in group a1 (vaccine from company A, different batches), the antibody qualified rate could reach 100% for type O and type A, respectively after each vaccination. In group a2 (vaccines from company A, same batch), the antibody qualified rates were 36.7%, 98.3% and 100% for type O, and 15%, 86.7% and 100% for type A after the first to the third vaccination, respectively. In group b (vaccine from company B, same batch), the antibody qualified rates were 18.3%, 97% and 100% for type O, and 1.7%, 45% and 53.3% for type A after the first to the third vaccination, respectively. In group c (vaccines from company C, same batch), the antibody qualified rates were 26.7%, 96.7% and 100% for type O, and 21.7%, 71.7% and 100% for type A after the first to the third vaccination, respectively. (2) Antibody positive rate to non-structural protein 3ABC (confirmed with second ELISA test): In group a1, the positive rates were 0.7%, 1.4%, 9.5% and 4.8% after the first to the fourth vaccination, respectively; In group a2 and c, no 3ABC antibody-positive animal was detected after 3 repeated vaccination; In Group b, only one animal with a positive rate of 0.6% was detected after the third vaccination. The antibody qualified rates to the structural protein of FMDV in 3 of the 4 groups were far less than 70% after the primary vaccination, however, those were increased significantly after boost and repeated vaccination. The antigen purity of vaccines in three groups (a2, b and c) can meet the requirement of OIE standard on the FMD vaccine, however, the seroconversion to 3ABC antibody was obvious in animals from group a1 after repeated vaccination, which would cause some extent of interference to differential diagnosis. Also, a combination of a primary screening test and a confirmatory ELISA test can further improve the accuracy of differential diagnosis. This study provides an important scientific basis to make a rational program for establishment and evaluation of FMD free zone with vaccination.

Canine distemper (CD) is a contagious disease, which can damage the immune system, respiratory system, digestive system, and even nervous system, leading to systemic pathological changes, and has a huge threat to pet dogs, fur animals, etc. At present, the commonly used colloidal gold test cannot effectively distinguish vaccine immunity from animal natural infection. To establish an efficient and accurate detection method for identifying wild strains and vaccine strains of canine distemper virus(CDV), the whole genome of six canine distemper wild strains isolated from dogs and three widely used CDV vaccine strains collected from Wuhan area were sequenced. After comparing and analyzing the amino acid and the base sequences, the H gene was determined as the target gene for AS-PCR primer design. By genotyping the H gene, it was found that the prevalent CDVs in Wuhan were all Asia-Ⅰ, while the vaccine strain Y2 was America-I, and the vaccine strains Y1 and Y3 were both America-Ⅱ. Comparing the CDV-H gene sequences of 179 Asia-Ⅰtypes (6 wild-type strain samples + 173 GenBank Asia-Ⅰtype stains) and 3 vaccine strains, using AS-PCR technology (3'mismatch) to design a pair of primers. It can effectively distinguish CDV Asia-I wild strain and vaccine strain. The upstream primer sequence is 5'-TTAATATAATAATGACAGTG-3', and the downstream primer sequence is 5'-CCTCAAGGGGCACA-3'. The results showed that the primer had a strong specificity and the wild strains could amplify an 894 bp fragment, while the vaccine strains could not. There are 9 more regular bases (amino acids) variation sites in H gene of wild strain, and the 277th amino acids of all vaccine strains are Asparagine. These mutations may lead to an increase in N-glycosylation sites, which will have an impact on the virulence of canine distemper virus vaccine strains. The AS-PCR method established in this study can effectively distinguish the canine distemper vaccine and Asia-Ⅰwild strains.

Signaling lymphocyte activation molecule (SLAM) was identified as the primary receptor of Peste des petits ruminants virus (PPRV) on goat lymphocytes. This study aimed to explore the effects of exosomes derived from PPRV vaccine strain N75-1 infected cells on the expression of SLAM in goats. In this study, SLAM expression and cytokines expression levels of goat PBMCs (recipient cells) incubated with exosomes isolated from PPRV-infected goat PBMCs (Exo-PPRV) was analyzed by quantitative real-time PCR, flow cytometric assay and Western blot assay. The results showed that the recipient cells treated with Exo-PPRV had significantly increased SLAM mRNA and cell surface expression (P<0.05) as compared with Exo-Mock treated cells. Furthermore, the increased TNF-α, IL-10, and IFN-γ expression as well as decreased IFN-α expression (P<0.05) were also detected. Further study demonstrated that Exo-PPRV that contained high levels of PPRV H protein could transmit H protein to recipient cells. Moreover, the expression level of SLAM in the pcDNA3.1-H transfected cells was significantly higher than that in the pcDNA3.1 transfected cells and non-transfection group. Taken together, our study demonstrated that PPRV infected cells derived exosomes has positive effects on the SLAM expression in the recipient cells, and that the high level of PPRV H protein contained in PPRV-Exo is one of the key molecules for exosomes to regulate the expression of SLAM in the recipient cells.

The purpose of this study was to investigate the effects of recombinant 10 kDa culture filtrate protein (CFP10) of Mycobacterium tuberculosis on the inflammatory responses mediated by Toll-like receptor (TLR) signaling in A549 cells. The recombinant plasmid pCzn1-CFP10 was obtained by amplifying the cfp10 gene fragment using PCR and cloning it into the prokaryotic expression vector pCzn1. The obtained recombinant plasmid pCzn1-CFP10 was transformed into Escherichia coli BL21(DE3) and induced by IPTG to express the recombinant protein CFP10 (rCFP10). The purified rCFP10 protein was preserved after endotoxin was removed and desalted before use. The effect of rCFP10 treatment on the survival rate of A549 cells was detected by MTT assay. A549 cells were treated with rCFP10 to detect changes of key molecules of TLR signaling pathway and downstream inflammatory factors in A549 cells by qRT-PCR，Western blot and ELISA. The results showed that the recombinant expression vector pCzn1-CFP10 was successfully constructed and the high-purity rCFP10 protein was expressed and purified in this study. MTT assay showed that rCFP10 could inhibit the survival rate of A549 cells in a time- and concentration-dependent manner. In addition, rCFP10 could significantly (P<0.05) up-regulate the key molecules of TLR pathways TLR2, TLR4, MyD88, TRAF6, NF-κBp65 and downstream inflammatory cytokines IL-6 and TNF-α in A549 cells as compared to the control group. The recombinant Mycobacterium tuberculosis protein CFP10 could promote the secretion of cytokines by activating TLR receptor signaling pathway in A549 cells, which will provide a theoretical basis for further understanding of the pathogenesis of Mycobacterium tuberculosis.

Shiga toxin-producing Escherichia coli(STEC) is a new class of highly pathogenic food-borne pathogens carrying a prephage encoding one or two Shiga toxin genes. It has become an important public health issue that threatens human health. The present work aimed to characterize STEC strains isolated from cattle and sheep at various stages, in parts of Xinjiang, in terms of the presence of prevalence, genetic diversity, and antimicrobial susceptibility to 17 common antibiotics. Through amplification of four virulence genes (stx1, stx2, eae, hlyA)by PCR and ERIC-PCR genotyping to detection STEC isolates. In the present study, a total of 64 STEC strains were isolated from 431 samples from slaughterhouses, farms and markets. Of these, 31 (48.4%) of the isolates harbored stx1 + stx2, and only 29 (45.3%) of the isolates possessed stx1, only 4 (6.3%) of the isolates harbored stx2, and 1 isolates harbored all the 4 virulence genes. Drug sensitivity tests found that STEC strains displayed 7 antimicrobial resistance to midecamycin(61%), cephalothin(4.7%), cefoxitin(4.7%), ampicillin(3.1%), piperacillin(1.6%), tobramycin(1.6%), cefazolin(1.6%). The ERIC-PCR results showed a polymorphic distribution, which was divided into two clusters of A (36 strains) and B (28 strains). STEC strains isolated from cattle and sheep at various stages, in parts of Xinjiang, some of which might have the potential to cause food contamination and human diseases.

To determine the cause of death of Chinemys reevesii in a pet shop in Changli county, Hebei province, a dominant strain CLW-1 was isolated and purified from the diseased Chinemys reevesii. It was analyzed and identified through anatomical observation, bacterial isolation and culture, Gram staining and microscopic examination, biochemical identification and 16S rDNA gene sequencing, the drug resistance of the strain was detected through drug sensitivity test, and its pathogenicity was detected through zebrafish inoculation test and virulence gene experiment. The results showed that the bacteria formed moist, smooth surface, gray colony with β hemolysis on blood agar medium, and grew smooth and moist yellow colony on RS agar medium; Gram staining and microscopic examination showed negative coccidiosis; Homology analysis of 16S rDNA gene sequence showed that the isolated strain CLW-1 belongs to the same group with Aeromonas veronii, and the homology is above 99%; Inoculation test results of zebrafish showed that the strain had high pathogenicity, and its LD₅₀ was 1.26×10⁶ CFU; The virulence gene amplification results showed that the isolated strain CLW-1 had virulence genes of aerA, Exu, Lip and Ser; Drug sensitivity test results showed that the isolated strain CLW-1 was sensitive to piperacillin, cefazolin and enrofloxacin, etc; Susceptibility to furazolidone, oxacillin and erythromycin, etc; Resistance to midecamycin, clindamycin and vancomycin, etc. According to the test results, the isolated strain CLW-1 is confirmed to be Aeromonas veronii, which carries virulence genes and has high pathogenicity, it has great therapeutic effect by applying corresponding antibiotics, thus laying a foundation for the prevention and treatment of Aeromonas veronii.

This experiment was developed to simultaneously detect the seven common foodborne pathogenic bacterias include E. coli O157:H7, Salmonella, V. cholera, L. monocytogenes, C. jejuni, V. Parahemolyticus and S. aureus. Seven pairs of specific primers were designed according to the conserved sequences of the genes from each pathogen available in the GenBank database. Single and mixed pathogen DNA templates were used to evaluate the specificity of the GeXP-multiplex assay. Control group was set up, recombinant plasmids were constructed, and samples of different DNA concentrations were randomly combined to verify the sensitivity, specificity, accuracy and anti-interference of the established GeXP method. To certify the accuracy and reliability of the GeXP assay, it was evaluated using 120 clinical specimens that were compared with the single PCR. The obtained results showed that the corresponding specific fragments of genes were amplified by the single and the multiplex GeXP PCR assay. The detection limit of GeXP was 10³ copies·μL^-1 when all of seven bacterial pathogens were detected. The results of the interference assay showed the presence of specific amplification peaks when different templates. The detection rate of the GeXP multiplex PCR method was 2.50% (3/120)-15.83% (19/120) while the conventional PCR was 2.50% (3/120)-15.00% (18/120), and GeXP multiple PCR detected 8 more positive cases, which means that, the GeXP was more sensitive and accurate in the detection of the clinical samples. In conclusion, this GeXP-based multiplex PCR is a high-throughput, specific and sensitive test to detect seven common foodborne pathogenic bacterias. This assay provides a method in rapid molecular diagnosis for mix clinical seven common foodborne pathogenic bacterias.

This study aimed to clone and express cysteine proteinase (CP) gene of Theileria equi and Babesia caballi. The CP proteins were analyzed using bioinformatics tools and online databases. The total DNA of T. equi and B. caballi as the template sequence for PCR. The CP genes were cloned, expressed using the cloning technique and prokaryotic expression system, respectively. Then the recombinant CP (rCP) proteins were purified by Urea dialysis. Finally the specific reaction of polyclonal horse anti-CP serum with rCPs was analyzed through Dot blot method. The CPs were predicted and analyzed by the software MAGA, Prot Param, TMHMM, SignalP-5.0, Antibody Epitope Prediction, SYFPEITHⅡ, ProPred and EzMol^2.1 respectively. The CP genes were successfully amplified from DNA of T. equi and B. caballi and expressed in the inclusion body fractions, Dot blot assay indicated that the recombinant protein could react with polyclonal horse anti - CP serum. Compared with T. equi-CP and B. caballi-CP, different protozoon CP genes were highly conserved in evolution, and phylogenetic analysis results were consistent with their protozoon taxonomy. The bioinformatics analysis revealed that two CPs are the hydrophilic protein instead of a transmembrane one, no transmembrane domain, no Th cell epitope was found, and more localized in the cytoplasm, mitochondria and nuclei. The relative molecular weight (MW) of T. equi-CP was 29 948.60, the theoretical isoelectric point (pI) was 5.53, stability coefficient was 22.50; B. caballi-CP’MW was 14 603.62, the theoretical pI was 8.54, stability coefficient was 47.69, but signal peptide was contained. The CPs have good reactionogenicity, as the hydrophilic extracellular proteins with no Th cell epitope, and more localized in the cytoplasm, mitochondria and nuclei, which helped T. equi and B. caballi avoid host immune response outside the cell membrane and the CPs involved in proliferation, differentiation and programmed cell death. In conclusion, a theoretical basis was provided for the study on CP’s functions and the pathogenesis of T. equi and B. caballi.

Infectious bursal disease virus (IBDV), a double-stranded RNA virus, can cause the infection of chickens with bursal lesions, leading to immunosuppression; melanoma differentiation-associated gene 5 (MDA5) is a pattern recognition receptor that specifically recognises double-stranded RNA viruses. In order to investigate the mechanism of the chicken MDA5 (chMDA5) signalling pathway in pathological injury in IBDV-infected chickens, fifty 14-day-old specific pathogen-free (SPF) chickens were allocated into 2 groups (infected and control) and twenty-five chickens were in each group. Each chicken of infected group was inoculated with 0.6 mL IBDV JIC7 virus solution intraocularly and intranasally. The chickens in control group were administered sterile phosphate buffered saline (PBS) in the same manner simultaneously. The chickens’ bursa of Fabricius was isolated at the 1st, 4th, 7th, 21st and 35th day after infection. Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the IBDV load, and mRNA expression of chMDA5, chMDA signal pathway adaptor protein (chIPS-1), transcription factor (chIRF3, chNF-κB) and downstream product cytokines (chIFN-β, chTNF-α, chIL-1β and chIL-6). Indirect immunofluorescence assay was performed for the expression of chMDA5 protein in chicken bursa of Fabricius. Conventional pathological methods were used to examine the pathological changes in the bursa of Fabricius. The results showed that the expression of chMDA5 as well as chIPS-1, chIRF3, chNF-κB, chIFN-β, chTNF-α, chIL-1β and chIL-6 were significantly higher than those in the control group, and morphological damage occurred in the bursal tissue in the infected group. The aforementioned trend was consistent with the change in IBDV load. The results indicated that chMDA5 and its signal transduction pathway could be activated by IBDV, which involves the bursa of Fabricius injury and anti-injury process in chickens infected with IBDV.

This study aimed to detect the effect of Gallus TGFβ1 on the biological behavior of MDCC-MSB1 cells. MDCC-MSB1 cells were transiently transfected with Gallus TGFβ1 overexpression vector, interference expression vector, and the corresponding negative control. Then, the expression of Gallus TGFβ1, the cell proliferation, the cell cycle and apoptosis, the migration and invasion of each transfection groups were examined. Results showed that compared with the corresponding control, the MDCC-MSB1 cells transfected with overexpression vector of Gallus TGFβ1 could up-regulate the expression level of TGFβ1, the proliferation of MDCC-MSB1 cells was significantly inhibited, G1 phase cells were increased, S and G2 cells were decreased, the apoptosis rate of the cells was increased, the migration and invasion ability were decreased.However，the MDCC-MSB1 cells transfected with the interference expression vector of TGFβ1 significantly down-regulated the expression level of TGFβ1, cell proliferation was improved，G1 phase cells were decreased, S and G2 cells were increased, the cell apoptosis was decreased, the migration and invasion ability was increased. The results showed that Gallus TGFβ1 could inhibit the proliferation, migration and invasion of MDCC-MSB1 cells, and promote their apoptosis.

To investigate the effect of enolase (Eno) of Streptococcus equi ssp. zooepidemicus (SEZ) on phagocytosis of mouse alveolar macrophages (RAW264.7). Recombinant enolase (rEno) was obtained by constructing prokaryotic expression plasmid, and the cytotoxicity of rEno protein on RAW264.7 cell proliferation was determined by trypan-blue living cell count method. After the rEno protein was incubated with RAW264.7 cells, SEZ was applied to the cells and the quantity of bacteria being phagocytosed was detected to determine the phagocytic activity of RAW264.7 cells. Further, candidate proteins that might interact with SEZ Eno in RAW264.7 cells were screened by live cell stable isotope labeling (SILAC) and protein spectrum analysis (LC-MS/MS). It was found that protein treatment (rEno，10 μg·mL^-1) had significant cytotoxic effects on RAW264.7 cells. Treatment of RAW264.7 cells with 10.0 μg·mL^-1 rEno protein for 2 and 4 hours could significantly inhibit the phagocytosis of RAW264.7 cells (P<0.01, P<0.05). In RAW264.7 cells, dynactin subunit protein 2 (Dctn), integrin alpha-M and about 17 proteins that might interact with Eno were preliminarily identified as rEno interaction proteins. The rEno recombinant expression protein was obtained in this study, and it could reduce the phagocytosis of RAW264.7 cells to SEZ. Preliminary screening of interacting proteins also laid a foundation for further revealing the mechanism of Eno in the anti-phagocytosis of SEZ.

This work aims to study the anatomical structure of the yak kidney collection system. Twenty-four collecting system casting specimens, which were made by ABS (acrylonitrile butadiene styrene), were observed to study its anatomical structure. The results showed that the renal pelvis was present in all casts and is classified into 2 types, dilated or nondilated (58.3%), with or without the dilated origin of the ureter. All casts had 2 major renal calyces toward the cranial and the caudal poles. Minor renal calyces were connected to the major renal calyces by an infundibulum, which drained 1 to 5 minor renal calyces. The number of minor renal calyces in the caudal region was significantly more than that in cranial (P<0.01). The yak kidney collection system was classified into 2 types according to the character of drainage. The type A, which was composed of the collection system that presented two major caliceal groups and mid-zone drainage was dependent on these major groups, and type B, which mid-zone drainage was independent of the polar caliceal groups, the type A (70.8%) was more common than type B (29.2%) in the yak. The collection system of yak kidney was composed of 1 renal pelvis, 2 major renal calyces, and multiple (11-21) minor renal calyces. According to the characteristics of drainage, it was divided into 2 types, A and B, of which type A is more common.

This study aimed to investigate the effect of chronic renal failure(CRF) on the gut microbiota diversity and predict the gene function of the flora. A total of 30 2-year-old dogs were selected and randomly divided into the chronic renal failure model group (CRF, established by renal artery ligation), sham operation control group (Sham) and healthy control group (HCG). All animals were fed normally during the 56 days of test period. The serum creatinine (Scr), blood urea nitrogen (BUN) and urine protein / creatinine ratio (UP/C) were detected regularly during the experiment. The effects of chronic renal failure on the structure, diversity and function of gut microbiota were analyzed according to the result of bacterial 16S rDNA sequencing from fresh feces collected without contamination. The flora markers index (FMI) was constructed based on the principal component Logistic regression analysis of different microbiota in CRF group to predict the development of chronic renal failure. The results showed that: 1) The levels of Scr, BUN and UP/C in CRF group from the start of the 28th day of the experiment were significantly higher than those of HCG group and Sham group (P<0.05), and significantly higher than that of the first day of CRF group (P<0.05). 2) Compared with those before chronic renal failure (CRF group at day 0 and 28), HCG group and Sham group, the Chao 1 diversity and Shannon diversity of gut microbiota in CRF group at day 56 were significantly lower (P<0.05), while the relative abundance of Bacteroidetes, Proteobacteria and Actinobacteria significantly increased and the number of Firmicutes significantly decreased (P<0.05). 3) LEfSe analysis showed that 20 species were enriched in CRF group, mainly including Pseudomonas, Escherichia, Proteus and so on, and most of them had negative correlation with other intestinal bacteria. Functional prediction revealed that genes of those different species were mainly enriched in amino acid metabolism, sugar biosynthesis and metabolism and carbohydrate metabolism in CRF group. The area under ROC curve (AUC) of the FMI constructed with those species enriched was 0.788, which could be used as the intestinal microbial marker for CRF in dogs. In summary, chronic renal failure can reduce the diversity of intestinal microbiota, lead to the imbalance of bacterial structure and change of bacterial function. Moreover, the enriched gut microbiota in CRF group can be used as the intestinal microbial marker of CRF in dogs， and the best prediction effect can be obtained by FMI.

The objective of this study was to investigate the effects of different sodium nitrate and disodium fumarate ratios on ruminal methane production, fermentation parameters, fatty acids profiles and microbial population in water buffalo in vitro in presence of α-linolenic acid. Three female water buffaloes (body weight of (650±50) kg) with permanent rumen fistula were selected as the donors of rumen contents. Treatments additives were prepared as sodium nitrate and disodium fumarate mixtures at ratios of 2:1, 1:1, 1:2. Control group without any sodium nitrate or disodium fumarate. All groups were added with 0.25 mg·mL^-1 α-linolenic acid. The concentration of sodium nitrate and disodium fumarate mixtures was 1 mg·mL^-1. The results showed that mehtane production were reduced with different ratios of sodium nitrate and disodium fumarate mixture, the average decrease was 90.63%. The total volatile fatty acid (TVFA) concentration and most rumen microorganisms population were significantly reduced (P<0.05), but the concentration of acetate, EPA and CLA, ratios of acetate to propionate ratio and UFA/SFA were significantly increased (P<0.05) with sodium nitrate and disodium fumarate mixtures at ratio of 2:1. 3) Sodium nitrate and disodium fumarate mixtures at ratio of 1:2 had no significant effect on TVFA content, fatty acid content and rumen microbial populaiton(P>0.05). 4) The concentrations of EPA, CLA and UFA/SFA ratio were increased (P<0.05) with sodium nitrate and disodium fumarate mixtures at ratio of 1:1, but Butyrivibrio fibrisolvens and atypical butyrivibrio populations were decreased. It can be concluded that different proportion of sodium nitrate and sodium fumarate mixture can cause methane reduction, the adverse effect of sodium nitrate on rumen fermentation can be alleviated by the increased proportion of disodium fumarate.

This study was conducted to investigate the changes in plasma Melatonin (MT) level and hypothalamic clock gene expression in mice after sleep deprivation (SD). Seventy-two mice were randomly divided into SD group and non-sleep-deprived control group (n=36). The SD mice were subjected to a modified multiple platform water path for 72 h sleep deprivation. After that, blood samples were taken every four hours (CT4,8,12,16,20 and 24) for the detection of plasma MT content, and hypothalamus samples were collected for the detection of the expressions of clock genes including mClock, mBmal1, mCry1/2, mPer1-3, mRorβ and mReverbα. The results showed that compared with control group, circadian rhythm of plasma MT was disordered in SD group. The median values of mClock, mRorβ and mReverbα in the hypothalamus decreased significantly (P<0.05), but mPer1 and mPer2 increased significantly (P<0.01); the circadian rhythms of mCry1 and mCry2 disappeared; the expression rhythms of mBmal1, and mClock were advanced in phase, while mRorβ and mPer1-3’s were delayed. These results suggested that acute sleep deprivation of 3 d could cause disorders of MT secretion and hypothalamic clock gene expression in mice.