As a study hotspot, pan-genome has frequently appeared in the genome field in recent years. Pan-genome had significant advantages compared to the single linear reference genome, and could effectively solve the problem of reference genome covering limited genetic information. Thanks to the fast pace at which sequencing technologies progress, pan-genome have been developed rapidly in recent years. Here, the development history and construction strategies of pan-genome were reviewed, as well as the study of livestock and poultry. Furthermore, the future application trend was propected, in order to provide reference for further study of pan-genome in livestock and poultry.

With the improvement of people's living standards, high-quality meat products of agricultural animal have been favored by consumers in China for the past few years. Fat deposition is an important factor affecting meat quality. Therefore, it is of great significance to clarify the molecular mechanism of high-quality meat formation. N⁶-methyladenylation modification (m⁶A) is the most abundant modifications in eukaryotic mRNA, which is related to the transport, expression, degradation process of RNA and is involved in regulating a variety of biological processes. Studies have shown that m⁶A modifications mediated by human insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) play a key role in animal fat deposition. This review will focus on m⁶A, IGF2BP2 and their role as well as molecular mechanism in animal fat deposition to provide candidate genes and scientific basis for the improvement of meat quality in the future.

The reproductive performance and egg production performance of poultry is closely related to follicles development and ovulation. In poultry ovaries, less than 5% of follicles can develop from primordial follicle to mature follicle and ovulate, and most follicles become atresia during development. The development of follicles is influenced by intrinsic factors and extrinsic factors. Recently, many studies have found that epigenetic mechanism also plays an important role in the regulation of follicle development. Epigenetics is heritable changes in gene expression and phenotype without changing in the DNA sequence. Therefore, the review described the process of follicle development and the main influencing factors in follicle development in laying hens. Moreover, in this review, the possible epigenetics regulatory mechanisms including DNA methylation, histone modification, non-coding RNA regulation and RNA modification in follicle development were summarized. In conclusion, the review provides a theoretical basis for improving poultry production performance.

Buffalo is one of the dairy livestock breeds in China, which has strong adaptability to high temperature, humidity, and rough feeding, and the buffalo milk has high nutrition and economic value, so it is known as the "king of milk". However, dairy buffaloes have late sexual maturity, unperceived estrus symptoms, and are susceptible to environmental factors, thus resulting in low reproductive efficiency, which in turn causes waste of workforce and economic losses. Therefore, it is important for dairy buffalo production to know effective estrus identification method and thus determine optimal breeding time. In this paper, we summarized the advantages and disadvantages of manual and automated methods for the identification of various estrus stages in dairy buffaloes, reviewed relevant studies on the identification of estrus in dairy buffaloes by blood, urine, saliva, cervical-vaginal fluid (CVF) and feces, and discussed the possibility of developing new methods for estrus identification in dairy buffaloes with omics technology by referring to relevant reports, aiming to provide reference for the development of new methods for estrus identification in dairy buffaloes.

The purpose was to unveil the genetic disparities between the Jianbai and Congjiang Xiang pigs breeds, enhance the understanding of their conservation status, genetic diversity, population structure, and adaptive evolution through whole-genome selection signal analysis. The 2-year-old Congjiang and Jianbai Xiang pigs were randomly selected and divided into two groups of 5 pigs each. Whole-genome resequencing was performed on Jianbai and Congjiang Xiang pigs, and the sequencing data was aligned to the pig reference genome Sscrofa 11.1 using the bwa software. Subsequently, the GATK software was used for variant detection and filtering of SNPs and INDELs. Additionally, population genetic diversity analysis, including effective population size, proportion of polymorphic loci, observed heterozygosity, and expected heterozygosity, was carried out using Plink and SNeP software. The population selection signals were analyzed using the vcftools software, and the selected genomic regions were subjected to KEGG and GO pathway analyses. After SNP and INDEL analysis, it was found that there were 16 158 002 common SNPs in the populations of Jianbai and Congjiang Xiang pigs, with 6 863 992 SNPs unique to Congjiang and 4 275 660 unique to Jianbai. The common INDELs were 3 275 375, with 1 674 081 INDELs unique to Congjiang and 1 114 248 unique to Jianbai. Starting from the sampling time point, the effective population size of both Congjiang and Jianxiang pig populations could be traced back to 18 individuals 13 generations ago. The Congjiang breed displayed a higher proportion of polymorphic loci, with a value of 0.875 7, whereas both populations had observed heterozygosity values exceeding the expected heterozygosity.Nucleotide diversity was less than 0.5% for both breeds, and Tajima's D was greater than 0, with ROD values of 0.553 1 and Fst values of 0.736 2. KEGG and GO enrichment analyses showed that the genetic differences between Jianbai and Congjiang Xiang pigs involved multiple pathways and gene functions, including axon guidance, ECM-receptor interaction, focal adhesion, PI3K-Akt signaling pathway, lipoic acid metabolism, cGMP-PKG signaling pathway, MAPK signaling pathway, nucleotide excision repair, aspirin signaling pathway, and arrhythmogenic right ventricular cardiomyopathy. There are a certain loss of population polymorphism and a high degree of population divergence between the Jianbai and Congjiang pig populations. There is clear population differentiation between the two breeds, and both genomes exhibit a substantial number of alleles at intermediate frequencies. It is necessary to enhance the genetic diversity within the Congjiang and Jianbai Xiang pig populations and reduce the risk of decreased purity due to the introduction of foreign lineages.

The aim of this study was to clone and analyze the fatty acid transport protein 2 (FATP2) gene by bioinformatics in goats, to examine the expression differences of FATP2 gene in different tissues of goats, and to further reveal the effect of interfering FATP2 gene on lipid metabolism in goat intramuscular precursor adipocytes. In this experiment, 10-month-old healthy Jianzhou goats (n=12) were used as experimental animals. The sequences of the FATP2 gene were amplified and cloned by RT-PCR, and sequence alignment, phylogenetic tree construction and bioinformatics analysis were performed. The relative expression levels of FATP2 gene in different tissues of goat and in different differentiation stages of goat intramuscular preadipocytes were detected by real-time fluorescence quantitative PCR. The SI-RNA interference sequence was synthesized and transfected into goat intramuscular preadipocytes. The FATP2 interference efficiency and the expression of the genes related to lipid metabolism were measured by real-time quantitative PCR; The effect of FATP2 interference on lipid droplet formation was observed by Bodipy staining and the triglyceride content was measured by GPO-Trinder enzymatic reaction. The results showed that the full length of the goat FATP2 gene sequence was 2 335 bp, CDS region was 1 863 bp, 5'UTR was 188 bp, 3'UTR was 284 bp, encoding 621 amino acids, and the highest expression of goat FATP2 gene was in the liver. RT-qPCR showed that the expression of FATP2 was interfered significantly in the cells. The Bodipy staining showed that the lipid droplets were significantly reduced after interfering with the FATP2 gene. The results of triglyceride assay showed that the triglyceride content in intramuscular adipocytes of goats was significantly reduced after interfering with FATP2. In this study, the CDS region sequence of goat FATP2 was successfully obtained. After interfering with FATP2, lipid deposition in goat intramuscular preadipocytes was significantly reduced and the expression of lipid metabolism related genes was significantly affected. These results provide important data to further elucidate the mechanism of FATP2's effect on the regulation of intramuscular formation in goats.

The study aimed to clone the PSMD9 gene sequence of goat and conduct the bioinformatics analysis. Then, the expression of PSMD9 in different tissues of goat and the expression level in intramuscular preadipocytes at different stages of differentiation was detected, and further revealed the effect of overexpression and interference of PSMD9 on lipid deposition in goat intramuscular preadipocytes. In this study, the sequence of PSMD9 gene was obtained by T-A cloning technique using 1-week-old healthy Jianzhou big-eared male goats (n=12), bioinformatics analysis was performed, and PSMD9 overexpression vector was constructed by double digestion method. PSMD9 was overexpressed in goat intramuscular adipocytes using liposome transfection, and the small interfering RNA (siRNA) interfered with PSMD9 expression. The effect of PSMD9 on lipid droplet formation was observed by Oil Red O staining and triglyceride content was measured by GPO-Trinder enzymatic reaction. The expression levels of PSMD9 in different tissues of goats and different stages of differentiation in intramuscular preadipocytes were measured by real-time quantitative PCR (RT-qPCR). The results showed that 763 bp nucleotide sequence of goat PSMD9 gene was obtained, including 5'UTR 67 bp, CDS 564 bp, and 3'UTR 132 bp, encoding 187 amino acid residues. The expression of PSMD9 gene in goat was highest in the liver and lowest in the spleen, and its expression level decreased gradually in goat precursor adipocytes in the former 4 days of differentiation induction, subsequently increased, and the expression level was the highest on the 8th day. Overexpression of the PSMD9 gene significantly increased the lipid droplets and triglyceride content in intramuscular preadipocytes and significantly up-regulated the relative expression of DGAT2, FASN and ACC, while the expression of DGAT1, PPARγ and SCD1 did not significantly changed. Interfering with PSMD9 gene decreased the lipid droplets and triglyceride content in intramuscular preadipocytes, while significantly down-regulated the expression of DGAT1, PPARγ, FASN and ACC, the expression of DGAT2 and SCD1 did not significantly changed. In this research, the CDS region sequence of PSMD9 gene was successfully obtained and its expression was highest in liver tissue. The expression of PSMD9 gene decreased gradually during differentiation in intramuscular adipocytes and reached the lowest level at day 4 in goat, after which the expression gradually increased.The expression of PSMD9 can significantly promote lipid deposition in goat's intramuscular preadipocytes. These results provide important data support for further elucidating the mechanism of PSMD9 gene regulating the formation of intramuscular fat in goat.

This study was conducted to determine the effect of compound Chinese herb ultrafine powder on the antioxidant ability of laying hens during late laying period. A total of 216 Xinyang black-feather laying hens at 307-day-old were selected and randomly divided into three groups, with eight replicates per group and nine hens per replicate. The birds in control group was fed a basal diet, and the experimental groups were supplemented with 0.5% Salvia miltiorrhiza (SM) + 0.25% Ligustrum lucidum (LL) + 0.25% Dandelion (Compound 1) and 0.3% Motherwort + 0.2% SM + 0.25% LL + 0.25% Dandelion (Compound 2) ultrafine powder, respectively. Samples of plasma and liver tissue were collected on days 60 and 120 of the trial to measure oxidation-antioxidant indexes and related gene expression. The results showed that, compared with the control group, on day 60 of the trial, the plasma and liver MDA content were decreased and the mRNA expression levels of liver NQO1 and GST were upregulated in compound 1 group (P<0.05), the mRNA expression levels of liver GSH-Px2, GSH-Px3, GSH-Px4, SOD2, SOD3, TXNRD1, TXNRD3, NQO1 and HO-1 were up-regulated (P<0.05) in compound 2 group, while the liver activities of GSH-Px, SOD, CAT and T-AOC were increased (P<0.05) in both compound 1 and 2 groups; On day 120 of the trial, the liver MDA content was decreased (P<0.05) in compound group 1, the plasma CAT activity was increased (P<0.05) in compound 2 group, the mRNA expression levels of liver Nrf2 and GSH-Px2 were increased (P<0.05) in compound 2 group, and the liver GSH-Px, SOD and CAT activities were increased (P<0.05) in compound 1 and 2 groups. Compared with the compound 1 group, on day 60 of the trial, the mRNA expressions of liver CAT and GST were decreased (P<0.05) in compound 2 group, and the liver mRNA expression levels of GSH-Px2, GSH-Px3, GSH-Px4, SOD2, SOD3, TXNRD1, TXNRD3, NQO1 and HO-1 were increased (P<0.05) in compound 2 group; On day 120 of the trial, plasma MDA content and mRNA expressions of liver Nrf2, GSH-Px2, TXNRD1 and TXNRD3 were increased (P<0.05) in compound 2 group. In conclusion, these findings suggested that dietary addition of the compound Chinese herb ultrafine powder composed of motherwort, SM, LL and dandelion could enhance the anti-oxidative ability of laying hens during late laying period by increasing the activity of antioxidant enzymes in plasma and liver and activating liver antioxidant-related signaling pathways. In addition, these two compounds present target specificity for the regulation of anti-oxidative capacity in the body.

The aim of this study was to clarify the role of RaeR encoded by GE296_RS05175 gene in the regulation of efflux pump RaeC-RaeA-RaeB of Rimerella anatipestifer RA-LZ01 strain. The GE296_RS05160 gene (encoding the inner membrane protein RaeB) deletion mutant ΔRaeB and complemented strain cΔRaeB of RA-LZ01 strain were constructed. Similarly, GE296_RS05175 gene deletion mutant ΔRaeR and its complemented strain cΔRaeR were constructed. The growth curves, antimicrobial susceptibility and transcription levels of GE296_RS05160 and GE296_RS05175 genes under specific substrate stimulation were measured. Compared with the parental strain, the growth abilities of the deletion mutants had no obvious change. By comparison with the parental strain and complemented strains, the sensitivity of ΔRaeB to kanamycin, streptomycin and (sodium dodecyl sulfate, SDS) were increased, while ΔRaeR showed the opposite results. After SDS stimulation, the relative transcription levels of GE296_RS05160 gene in RA-LZ01 and ΔRaeR strains increased 2.26 and 4.64 times, respectively. The upregulation of GE296_RS05160 gene in ΔRaeR strain was significantly higher than that in RA-LZ01 strain. Under SDS stimulation, the relative transcription levels of GE296_RS05175 gene in RA-LZ01 and ΔRaeB strains decreased significantly. These results indicated that the efflux pump RaeC-RaeA-RaeB had no significant correlation with the growth of RA-LZ01 strain, but could mediate the tolerance of the strain to kanamycin, streptomycin and SDS. The transcription level of GE296_RS05175 gene reduced under SDS pressure, and RaeR,which encoded by GE296_RS05175 gene, regulated the expression of RaeB negatively.

A new rapid, highly sensitive and highly specific nucleic acid detection method for avian influenza virus(AIV) H5 subtype was established by using reverse transcription-recombinase aided amplification(RT-RAA) and clustered regularly interspaced short palindromic repeats(CRISPR). By analyzing the HA gene sequences of 342 avian influenza virus H5 subtype genes, the conserved regions were selected to design highly specific CRISPR RNA(crRNA) sequences and RT-RAA primers. The contents of main components in the detection system were optimized, including LwaCas13a protein, crRNA, TaqMan probe, T7 RNA Polymerase and NTP Buffer Mix.Thus, the RT-RAA based detection method of AIV H5 subtype nucleic acid CRISPR-Cas13a was established.The sensitivity and specificity of the method were evaluated by taking 10⁵~1 copies·μL^-1 of standard plasmid containing the target gene and other subtypes of avian influenza viruses and other avian disease viruses. The results showed that the RT-RAA primer and crRNA designed in this paper were specific and sensitive. The main components in the optimal reaction system were LwaCas13a protein (60 μg·mL^-1) 2.0 μL, crRNA (100 μmol·L^-1) 3.2 μL, TaqMan probe (50 μmol·L^-1) 1.28 μL, T7 RNA Polymerase 0.25 μL and NTP Buffer Mix 2.0 μL, respectively. The detection sensitivity of this method was up to 1 copy·μL^-1, and there was no cross reaction with AIV H3, H6, H7, H9, H10 subtypes, neither with NDV, IBV, IBDV or DTMUV. The method established in this study and fluorescence quantitative RT-PCR were used to detect 56 clinical samples, and the coincidence rate of the two methods was 98.2%. In conclusion, a rapid detection method for AIV H5 subtype was established by using RT-RAA isothermal amplification technology combined with CRISPR-Cas13a gene editing technology in this study. The results were obtained within 1 h and 20 min. It provides a new technique for the rapid clinical detection of AIV H5 subtype with high sensitivity and high specificity, and has a good application prospect.

In order to investigate the prevalence and genetic variation of porcine reproductive and respiratory syndrome virus (PRRSV) in China, our group collected 117 samples of clinically suspected PRRS symptoms from nearly 50 farms in 13 provinces in China from 2021 to 2022, and conducted genetic variation analysis for the ORF5 gene of PRRSV. The PRRSV-positive samples and genotypes were identified by Quantitative Real-time PCR, RT-PCR was applied to amplify PRRSV ORF5 and ORF7 genes, and the positive samples were sequenced and analyzed for phylogenetic analysis and GP5 amino acid mutations. The results showed that a total of 48 PRRSV-2 positive samples were detected (overall positive rate of about 41%). In addition, a total of 38 ORF5 sequences and 10 ORF7 sequences were obtained by sequencing. The nucleotide similarity among all ORF5 sequences ranged from 77.1% to 99.8%, and the nucleotide similarity among ORF7 sequences ranged from 83.3% to 98.7%. Phylogenetic analysis showed that lineage 1 accounted for 60% (29/48, 28 NADC30-like strains and 1 NADC34-like strain), lineage 3 for 25% (12/48), lineage 5.1 for 5% (2/48), and lineage 8.7 for 10% (5/48). Amino acid site analysis of GP5 showed significant amino acid variation in the signal peptide (aa 1-26), neutralizing epitope C (aa 52-61) and potential N-glycosylation sites (aa 32-35, aa 44, aa 51) were three regions with significant amino acid variants. The results of the study showed that several strains of PRRSV were prevalent in China at the same time, and lineage 1 had replaced lineage 8.7 as the main prevalent lineage and was distributed in most provinces in China; Lineage 3 was second only to lineage 1 and was prevalent mainly in the local area of southern China; Lineage 8.7 were detected at a low rate. It is important to note that a NADC34-like strain was detected in Huizhou City, Guangdong Province, indicating that this strain may have become potentially endemic in South China. The epidemic of PRRSV in China has become increasingly complex in recent years, and the widespread prevalence of lineage 1 strains has increased the genetic diversity of PRRSV. The results of this study will provide a reference for the development of PRRSV prevention and control strategies.

Synaptogyrin-2 (SYNGR2) protein affects PCV2 infection in host. This study aims to investigate the SYGNR2 function on PCV2 proliferation in cell cultures and dissect the mechanism for SYNGR2 affecting PCV2 infection. We performed gene knockout, overexpression and site-directed mutagenesis assay to study how SYNGR2 affects PCV replication in PK15 cells. To investigate the mechanism, RNA-seq was performed to study the molecular differences between SYNGR2 KO and WT cells. Differentially expressed genes between the two groups were analyzed and further validated by quantitative Real-time PCR. SYNGR2 knockout and p.Arg63Cys point mutation decreased PCV2 replication in PK15 cells, and the overexpression of different genotypes of SYNGR2 restored PCV2 replication capacity in SYNGR2 KO cells. RNA-seq enrichment analysis showed that the SYNGR2 mainly participates in the membrane components and vesicles and other membrane organelles. The differentially expressed gene MYRIP and its bound exosome regulation related gene RAB27A were selected for verification, and qPCR results showed that the expression of RAB27A mRNA was significantly increased in SYNGR2 knockout cells.In Conclusion, this report confirmed SYNGR2 function to support PCV2 infection in cells. SYNGR2 knockout and p.Arg63Cys point mutation both decreased PCV2 replication in cells. Transcriptome analysis reveals that SYNGR2 may affect PCV2 replication capacity by regulating the function of vesicular transport.

To investigate the species and prevalence of viruses related to porcine respiratory disease complex (PRDC) in Tibetan pigs in western Sichuan province, 119 respiratory samples from Tibetan pigs were collected from Ganzi Tibetan Autonomous Prefecture of Sichuan province, including 66 nose swabs from healthy pigs (from 6 pig farms) and 53 nose swabs, lung tissues and alveolar lavage fluid (from 11 pig farms) from diseased pigs with obvious symptoms of PRDC. Samples from healthy group and diseased group were mixed into two pools, respectively, then the total nucleic acid were extracted and cDNAs were generated using reverse transcription. Finally, two mixed pooled samples were sent to company for library construction and Illunima novaseq sequencing. Sequence analysis showed that the respiratory samples from healthy Tibetan pigs were mainly bacteriophages, and only the mardivirus of Herpesviridae that had been identified to be related to animal viruses. 16 distinct viruses in 14 virus families were identified in the diseased group. The viruses that had been associated with PRDC include porcine circovirus type 2 (PCV-2), Torque teno sus virus (TTSuV), porcine cytomegalovirus (PCMV), and African swine fever virus (ASFV). The whole genome of two PCV-2 strains, three TTSuV strains and one Porprismacovirus strain were assembled using SOAPdenovo software. Phylogenetic analysis showed that two PCV-2 strains belonged to PCV-2d subtypes, respectively, and three TTSuV strains belonged to k2 genotype, among which SCgz-1 belonged to k2b subgenotype, SCgz-2 and SCgz-3 belonged to k2a subgenotype. The Porprismacovirus strain was clustered into a single glade with Human 16806x66_213 strain from Vietnam, and showed a distant genetic relationship with the swine-origin Porprismacovirus. Recombination analysis using RDP4 software showed that the virus was a recombinant strain between Human 16806x66_213 and Porcine 17668x82_593, which had not been reported internationally. Our study investigated the respiratory related viruses of Tibetan pigs in plateau region for the first time. The results showed that a variety of respiratory disease related viruses were found in Tibetan pigs, which may be transmitted among different livestock, or human and livestock,which needs more attention. Our study provides reference for the prevention and control of respiratory diseases in Tibetan pigs.

With the development of intensive farming, the demand for early and rapid detection of various pathogens in large-scale pig farms has gradually increased. The aim of this study was to establish the broad-spectrum magnetic nanoparticle visualization detection (UNVs) field detection technology for porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), porcine circovirus type 2 (PCV2), porcine epidemic diarrhea virus (PEDV), porcine pseudorabies virus (PRV), transmissible gastroenteritis virus (TGEV), and porcine parvovirus (PPV). Based on the highly conserved regions of different viruses, probes and primers were designed, and sequence-specific identification probes (enrichment probes and hybridization probes) with high specificity and enrichment and hybridization efficiency for the target pathogen genes were obtained through the screening of probes and optimization of conditions, which were coupled with ferric oxide magnetic particles and nanogold, respectively, to prepare functionalized magnetic beads specific for each pathogen and functionalized nanogold. The functionalized magnetic beads were used as carriers to enrich nucleic acid signals, and functionalized nanogold was used as a bridge to amplify signals, and the dual probes were used to simultaneously detect pathogenic nucleic acids, combined with the enzymatic visualization reaction of alkaline phosphatase to detect signals, to establish a convenient diagnostic technique for on-site visualization of UNVS for seven kinds of viruses, including PCV2, PRV, PPV, PEDV, TGEV, PRRSV, and CSFV. The results showed that the nucleic acid probes designed for screening did not cross-react with other pathogens of pigs and had good specificity. The established technique for the visualization of UNVs for PCV2, PRV, PPV, PEDV, TGEV, PRRSV and CSFV pathogens has a detection limit of 10³ copies·mL^-1 serum sample or 10³ copies·g ^-1 stool sample, which basically achieves the sensitivity of PCR or RT-PCR detection, the maximum coefficient of variation within and between groups is less than 5%, the colorimetric results are stable without large differences, there is good reproducibility, and the detection of a single sample can be completed in less than 4 h. The results of simultaneous testing and retesting of 499 clinical samples (serum and fecal samples from pigs suspected of having PCV2, PRV, PPV, PEDV, TGEV, PRRSV and CSFV) from Shaanxi Province by applying the technique and PCR assay showed that the established visualization method for UNVs against seven pathogens not only had the same sensitivity as ordinary PCR, but even had a higher detection rate compared with ordinary PCR. In this study, we successfully established a visual detection technique for UNVs against PCV2, PRV, PPV, PEDV, TGEV, PRRSV and CSFV pathogens, which has the advantages of high sensitivity, wide detection range as well as good specificity and reliability, providing a rapid, accurate and sensitive diagnostic technique for a variety of pathogens in pigs.

This study aimed to determine the current status and epidemiological characteristics of beef cattle viral diarrheal pathogen infection in Ningxia, and to provide scientific basis for the prevention and control of bovine diarrheal disease. In this study, 293 beef cattle swabs in Ningxia were detected and genetically analyzed by RT-PCR for bovine viral diarrhea virus (BVDV), bovine coronavirus (BCoV), bovine rotavirus (BRV), bovine norovirus (BNoV), bovine astrovirus (BAstV), and analyzed their genetic evolution. The Studies showed that diarrhoeal virus infection and mixed infection were prevalent in beef cattle; The infection and mixed infection of beef cattle diarrhea virus in free-range breeding mode were more serious than those in large-scale breeding. Viral diarrhea varied significantly between different regions and viruses; Genetic evolution analysis showed that the BVDV epidemic strain in Ningxia was subtype 1e, the BRV epidemic strain was G1 subtype, the BNoV epidemic strain was GIII.2, and the BCoV epidemic strain was closely related to the genetic evolution of the French strain. The results showed that diarrheal virus infection was widespread in beef cattle in Ningxia, which was different between different breeding modes, different regions and different viruses, and the mixed infection was serious.

Bovine mastitis is the most economically important disease of dairy cattle, which is most commonly caused by pathogens. The pathogens that influenced dairy cows might be from the farm environment. Therefore, the flow of microbes through environments to cow mastitis should be investigated. In this study, mastitis milk, pan barn samples (air, drinking water, feces, feed, bedding material, new bedding material, spray water), and parlour samples (pre teat medicine, post teat medicine, pre medicine cup, post medicine cup, teat dip cup, teat skin) have been collected in a dairy farm in Tianjin, China. Total DNA of culture-based and nonculture-based samples were extracted and then performed 16S rDNA amplicon sequencing and SourceTracker. OTU profiles were assigned to 50 families among all the 74 samples. Beta diversity and ANOSIM analysis showed that the similarity of bacteria between the mastitis milk and environment samples was low. In general, the microbial community structure was similar between mastitis milk and teat skin and, to a lesser extent, bedding samples. Moreover, SourceTracker identified the teat skin as the most important source of pollution, followed by air, milk cups and feces, which highlighted the possible effects of farm management practices on bovine mastitis. Therefore, control measures should be introduced to reduce or eliminate the risk of transmission in the dairy farms.

SIgA is the primary effector molecule of mucosal immunity, and it is often used as a target for early diagnosis of diseases in clinic. In addition, more and more attention has been paid for therapeutic SIgA mAbs and mucosal vaccines targeting SIgA producing. Isolation and purification of intact and active SIgA is a prerequisite for product development. In order to improve the purification efficiency of SIgA from swine colostrum, in this study, a new way was established,including using tandem affinity chromatography, strong anion column and superpose 6 gel filtration. By using this way, 3 mg of active SIgA was abtained from 10 mL of colostrum. Western blot immunoblotting was performed with anti-pig IgA heavy chain MAb and anti-SC MAb, and the results showed that both of the MAbs had good reactivity with purified SIgA here. ELISA method was used to test the reactivity of SIgA purified here and anti-pig IgA heavy chain MAb, it showed that the purified SIgA in our study had higher reactivity than SIgA purified by the old method (ammonium sulphate precipitation, DEAE52 weak anion chromatography, SephadexG-200 gel chromatography and multiple Protein A affinity chromatography). Finally, purified SIgA was further checked by mass spectrometry. Our research established an efficient method for the purification of SIgA from swine colostrum, and also provided a reference for the purification of SIgA from other animals.

The article aims to study the effects of diminazene aceturate (DIZE) activation of Angiotensin-converting enzyme 2 (ACE2) on mitochondrial structure and function by establishing a high-fat diet-induced non-alcoholic fatty liver disease(NAFLD) model. Twenty-six male SD rats were selected, nine rats were fed with basal feed as a contral group, the remaining rats were fed with high-fat feed for modeling. After 4 weeks, the NAFLD model was successfully established by the confirmation of pathohistological examination. Then, the rats with successful modeling were randomly divided into Model group and DIZE group(eight rats per group),Model group rats were continued to be fed hight-fat feed and treated with 3 mL saline by intragastric administration, DIZE group rats were treated with 15 mg·(kg·d)^-1 DIZE as gavage. The contral group continued to be fed with common feed, and the same amount of normal saline was gavage administered. In the tenth week, serum and liver samples were collected from rats for the following tests:1) The kit was used to detect TG and TC content in serum; 2)The content of AngII and Ang1-7 in liver tissues were determined by ELISA; 3) Western blotting method was used to test the protein expression of ACE2, mitochondrial fusion protein (Mfn1, OPA1), fission protein (Fis1, Drp1) and uncoupling protein 1 (UCP1); 4) RT-qPCR to test mRNA expression of mitochondrial biogenesis and mitochondrial respiratory chain complex-related genes. Results:1) The fat deposition was found in HE staining after 4 weeks of high-fat feeding, obviously, which indicates that the NAFLD modeling of rats was successfully established; 2) Compared with the CON group, the serum levels of TG and TC in the MOD group rats were significantly higher (P<0.05), while the DIZE group was significantly lower (P<0.05) compared with the MOD group; 3) Compared with CON group, ACE2 protein expression level in MOD group was extremely significant decreased (P<0.01), and the ratio of AngⅡ/Ang1-7 was significantly increased (P<0.05), however, compared with model group, the results were opposite in DIZE group (P<0.01); 4) The expression levels of fusion, fission and UCP1 protein were significantly or extremely significant reduced in model group compared to the control group (P<0.01 or P<0.05), DIZE could upregulate the expression levels of fusion, fission and UCP1 protein extremely(P<0.01); 5) Compared with the CON group, the expression levels of mRNA in mitochondrial biogenesis and mitochondrial respiratory chain complex I and III genes were extremely significant downregulated in the MOD group (P<0.01), nevertheless, compared with model group, DIZE group shows the opposite results(P<0.01 or P<0.05). The results demonstrated that DIZE could activate ACE2, thereby alleviating the inhibitory effects on mitochondrial biogenesis, fusion, fission, and respiratory chain-related parameters induced by a high-fat diet, ultimately achieving therapeutic effects for NAFLD. This study elucidates a new idea of DIZE for the treatment of non-alcoholic fatty liver disease, which manifests DIZE has potential research value as a new use of an old drug.

Allicin has good antibacterial and antioxidant activities, and is expected to be used as a substitute for antibiotics in the livestock and poultry industry, however, its effect on hepatic drug enzyme (CYP) remains unknown. The study was to investigate the effects of allicin on CYP1A2 enzyme activity, mRNA expression of coding gene and enzyme kinetics in broilers, and to provide a further reference for its rational use in production practice. Ninety one-day-old Arbor Acres (AA) broilers were randomly divided into control group (basal diet group), low-dose allicin group (40 mg·kg^-1) and high-dose allicin group (80 mg·kg^-1) after 4 days of adaptive breeding. The treatment groups were fed with allicin twice a day for 42 consecutive days. On the 14th, 28th, and 42nd day after treatment of allicin, 10 chickens from each group were randomly selected, respectively and liver microsomes were extracted by calcium ion precipitation method to detect the effects of allicin on CYP1A2 enzyme activity and enzyme kinetics parameters. The mRNA expression of CYP1A2 and nuclear receptor encoding gene CXR were detected by real time RT-PCR. Allicin (40 mg·kg^-1, and 80 mg·kg^-1) significantly increased the production of acetaminophen, the metabolite phenacetin catalyzed by CYP1A2,on the 14th and 28the day after treatment by 1.2 times and 1.4 times of the control group, respectively (P<0.05), however, the production of acetaminophen on the 42nd day, were significantly decreased to 80% and 62% of the control group, respectively (P<0.001). The variation trend of mRNA expression of CYP1A2 and CXR was consistent with that of enzyme activity. The half maximal inhibitory concentration (IC₅₀) of allicin on CYP1A2 was 18.87 μmol·L^-1, and the inhibition constant (K_i) was 10.89 μmol·L^-1. Allicin showed a non-competitive inhibition effect on CYP1A2 with a time and concentration-dependent manner. Allicin can affect both CYP1A2 enzyme activity and its encoding gene mRNA expression with a bidirectional manner. The results suggest that allicin has an inhibitory effect on broiler CYP1A2 after long-term application, and clinical attention should be paid to avoid the adverse reactions caused by drug-drug interaction.

The purpose of this study was to investigate the interaction between pigment epithelium-derived factor (PEDF) and melanin synthesis regulatory factors, and to explore the regulatory effects of PEDF on melanin cell vitality and melanin synthesis, so as to provide a basis for the study of animal coat color formation mechanism. In this study, PEDF siRNA and PEDF overexpression vector were transfected into mouse melanocytes respectively, and the activity of melanocytes and content of melanin were detected. The expression and localization of PEDF, Wnt1, Wnt3α, MITF and β-catenin were detected by qRT-PCR, Western blotting and cellular immunofluorescence. After PEDF silencing, the cell vitality of melanocytes was enhanced, the content of melanin was increased, and the mRNA and protein expressions of Wnt1, Wnt3α, MITF and β-catenin were increased. After PEDF overexpression in melanocytes, the activity of melanocytes was weakened, the content of melanin was decreased, and the mRNA and protein expressions of Wnt1, Wnt3α, MITF and β-catenin were decreased. Cellular immunofluorescence results showed that PEDF, Wnt1, MITF and β-catenin were expressed in the cytoplasm of melanocytes. Wnt3α was expressed in the cytoplasm and cell membrane of melanocytes. This study could confirm that PEDF could reduce the activity of melanocytes, and inhibit the production of melanin by down-regulating the factors related to melanin production, Wnt1, Wnt3α, MITF and β-catenin.

The research aims to investigate the inhibitory effect of isochlorogenic acid C (ICAC) on mastitis and NF-κB inflammatory pathway. In this study, bovine mammary epithelial cells (MAC-T) and mouse mammary tissue stimulated with lipopolysaccharide (LPS) were used as in vitro and in vivo inflammatory model, respectively. MTT method was used to screen the optimal concentrations of ICAC for MAC-T cell treatment. The expression levels of inflammatory factors (IL-1β, IL-6 and TNF-α) and inflammatory mediators (COX-2 and iNOS) were detected using ELISA, whereas the expression and phosphorylation levels of key proteins (IκB and p65) in NF-κB signaling pathway were detected by Western blot. The results showed that:1) ICAC did not significantly inhibit the growth of MAC-T cells at the concentrations from 20 to 100 mg·L^-1 (P>0.05); 2) The expression levels of IL-1β and IL-6 protein in MAC-T cells treated with 1 mg·L^-1 LPS were significantly higher than those in untreated group (P<0.01), but ICAC (20, 50, 80 mg·L^-1) significantly down-regulated the expression of IL-1β, IL-6, TNF-α, COX-2 and iNOS in MAC-T cells (P<0.01); 3) Intraperitoneal inject ICAC(60、80、100 mg·kg^-1) reduced LPS-induced CD3 expression and T lymphocyte infiltration in mammary tissues of mice; 4) ICAC significantly reduced the phosphorylation of p65 and IκB in LPS-induced MAC-T cells and in mouse mammary tissue (P<0.05), and it inhibited the nuclear transfer of p65. These results indicate that ICAC inhibits the inflammatory response in MAC-T cells and mouse mammary tissue through NF-κB inflammatory pathway, but the role of ICAC in the prevention and treatment of bovine mastitis needs further investigation.

The aim of this study was to investigate the mechanism of inulin with high degree of polymerization in improving high-fat diet-induced obesity in dogs based on the gut-adipose tissue axis. Forty poodle were randomly divided into five groups:common diet group, high-fat diet group, and low-dose, medium-dose and high-dose inulin with high degree of polymerization groups. Dogs in common diet group were fed with normal diet, and in high-fat diet were fed with high-fat diet. Dogs in low-dose, medium-dose and high-dose inulin with high degree of polymerization groups were fed with high-fat diet containing 1.0%, 3.0% and 5.0% inulin with high degree of polymerization, respectively. The test period was 12 weeks. After the experiment, the levels of serum glucose, fat and inflammatory factors were measured, and the relative expressions of mRNA and protein of related factors in subcutaneous adipose tissue and colon mucosa were detected using real-time fluorescent PCR and Western blot, respectively. The results showed as follows:Inulin with high degree of polymerization effectively reduced the body weight, body fat rate and blood lipid level, meanwhile improved the impaired glucose tolerance in dogs fed with high-fat diet. Moreover, inulin with high degree of polymerization increased the mRNA and protein expression of Occludin and ZO-1 in colonic mucosa, and reduced levels of serum lipopolysaccharide (LPS), IL-6 and TNF-ɑ and the expression of IL-6 and TNF-ɑ in adipose tissue. In conclusion, inulin with high degree of polymerization can improve the obesity induced by high-fat diet in dogs, and its mechanism may be related to the regulation of the gut-adipose tissue axis of "intestinal mucosal barrier function-LPS translocation-adipose tissue inflammation".

The aim of this experiment was to investigate the mechanism of puerarin in the intervention of oxidative stress of cartilage and the improvement of Nrf2/HO-1 pathway on cartilage degeneration in rats with post-traumatic osteoarthritis (PTOA), and to explore the therapeutic effect of puerarin on osteoarthritis. Forty male Sprague-Dawley rats were randomly divided into four groups:control group (n=8), model group (n=8), celecoxib group (n=8) and puerarin group (n=16). Puerarin group was divided into low dose group (n=8) and high dose group (n=8). The PTOA model was established by anterior cruciate ligament transection (ACLT) in all groups except the control group. Low-dose group (50 mg·kg^-1), high-dose group (100 mg·kg^-1) and celecoxib group (2.86 mg·kg^-1) were administered by gavage, and the control group was given equal amount of saline for 5 weeks. The degree of joint swelling, cold sensitivity response and knee extension vocalization tests were performed weekly to assess the degree of joint swelling and pain in rats. After the administration, knee joint and serum samples of rats in each group were collected. The tibial and femoral tissues were subjected to HE staining and a modified Mankin score to assess histopathological changes. The histopathological changes were evaluated using the Mankin method. The levels of MMP3, MMP13 and ADAMTS4 in rat cartilage, as well as the levels of CTX-Ⅱ and COMP in serum were measured to evaluate the improvement of puerarin on cartilage degeneration in rats with osteoarthritis. To evaluate the protective effect of puerarin on oxidative damage in rats, the activity of antioxidant enzymes (SOD, GSH-Px, and CAT) in rat serum, the content of MDA and the level of Nrf2/HO-1 pathway in rat cartilage were measured. Detection of IL-1β, IL-6 and TNF-α in rat serum to evaluate the anti-inflammatory effect of puerarin on the body. The results showed that, compared with the model group, the high dose group intervention could significantly reduce the degree of joint swelling and pain response in rats, improve the degree of articular cartilage injury, reduce the Mankin score (P<0.01), activate the Nrf2/HO-1 pathway in cartilage tissue, inhibit the expression of MDA, MMP3, MMP13, and ADAMTS4 (P<0.05), upregulate the activity of antioxidant enzymes in serum (P<0.01), and inhibit the inflammatory factor IL-1β, IL-6 and TNF-α and the levels of cartilage metabolic markers CTX-Ⅱ and COMP in serum (P<0.05). In summary, puerarin improves cartilage degeneration in PTOA rats by activating the Nrf2/HO-1 pathway to inhibit oxidative stress and inflammatory damage.

This study aimed to evaluate the surgical effect and complications of tibial plateau leveling osteotomy (TPLO) for cranial cruciate ligament disease (CCLD) in dogs, and analyze factors that affect surgical effect and complications. The medical records of canine CCLD treated by TPLO in China Agricultural University Veterinary Teaching Hospital were reviewed and followed up. The inclusion criteria are:follow-up time more than 5 months; case information record is completed. Results were as follows:101 TPLO procedures in 85 dogs of 20 breeds were included, including crossbreed (n=17, 20.0%), poodle (n=15, 17.7%), bichon (n=11, 12.9%), golden retriever (n=10, 11.8%) and Labrador retriever (n=8, 9.4%). The median weight and age were 14.4 kg (range, 4-53 kg) and 7 years (range, 1-14 years old), respectively. The overall complication rate was 37.6% (38/101). Thirty-six cases (35.6%, 36/101) had minor complications (including wound-related complications and bandage-related complications), and 2 cases (2.0%) had major complications (including 1case of screw loosening and 1 case of tibial tuberosity fracture). Wound-related complications (23.8%, 24/101) were more likely to occur in dogs with wound licking (P<0.01), which were found in all the 15 dogs with wound licking. Related complications occurred in 18 (34.6%) of 52 dogs treated with bandage. Two major complications were treated by revision surgery with good results. Among the cases with follow-up results, 95.6% (86/90) cases had good functional recovery. Besides, postoperative bandage increased the risk of complications (P=0.047, OR=3.873, 95%CI:1.015-14.776). However, no risk factors associated with surgical outcomes were found. In summary, TPLO of dogs with CCLD has desirable results and a low proportion of major complications. Licking can cause wound-related complications, therefore licking wounds should be strictly prevented postoperatively. Moreover, the use of bandages should be reduced to decrease associated complications.

The aim of this study was to report the effect of surgery assisted by 3D printing technique in the treatment of a dog with grade Ⅳ medial patellar luxation and cranial cruciate ligament rupture. A Shiba Inu with grade Ⅳ medial patellar luxation and cranial cruciate ligament rupture is treated in two ways. First, the modified tibial plateau leveling osteotomy (TPLO) and trochlear wedge recession are used. Then, the femoral deformity is evaluated through CT scanning and 3D printing of the femoral model to assist in formulating the surgical plan and carrying out distal femoral lateral closing wedge osteotomy (DFO). The degree of lameness, patella position and imaging examination are used to determine the recovery of the dog. The results showed that patellar luxation recurred 8 weeks after the modified TPLO and trochlear wedge recession, indicating that this method can not completely correct the grade Ⅳ medial patellar luxation. With the help of 3D printing technique, the DFO surgical planning is formulated, lameness is significantly improved after treatment, and postoperative recovery is good. In the case of high-level patellar luxation with cranial cruciate ligament rupture, attention should be paid to whether there is femoral deformity; The use of 3D printing technology can assess bone morphology more accurately, assist in formulating femoral osteotomy plan, and improve the success rate of surgery.

The aim of this study was to establish a duplex PCR and a duplex nano-PCR method for simultaneous detection of Eimeria necatrix and Clostridium perfringens. Primers for target gene loci were designed and screened based on the E. necatrix ITS-2 gene and the C. perfringens α toxin gene, respectively, and specific duplex PCR and nano-PCR detection methods for two pathogens were established by optimizing reaction systems and annealing temperatures. PCR amplifications of six other protozoan parasites and three other bacteria were used to verify their specificities. The recombinant plasmids pMD19-T-ITS2 and pMD19-T-CPA were further constructed and diluted into different concentration gradients with 10 times ratio for sensitivity tests of the proposed methods. The results showed that the target fragments of E. necatrix and C. perfringens were about 150 bp and 400 bp, respectively. The PCR amplifications using the two detection methods were negative for other six protozoan parasites and three other bacteria. The minimum detectable limits for E. necatrix and C. perfringens by using dual PCR were 181 copies and 1 050 copies, respectively, while the minimum detectable limits for E. necatrix and C. perfringens by using dual nano-PCR were 1.81 and 105 copies, respectively. Clinical detection showed that the detection results of two established methods were consistent with clinical pathogenic detection. The present study successfully established duplex PCR and duplex nano-PCR methods for detection of E. necatrix and C. Perfringens, with high sensitivity and specificity, providing technical supports for clinical detection of E. necatrix and C. perfringens infections.

The aim of this paper is to establish a rapid, sensitive, and specific detection method for porcine epidemic diarrhea virus (PEDV) by recombinase aided amplification (RAA) combined with the Clustered Regularly Interspaced Short Palindromic Repeats associated protein 13a (CRISPR-Cas13a), called RAA-Cas13a. The specific primer for RAA and CRISPR RNA (crRNA) of the conserved region of the PEDV N gene were designed. The sample nucleic acids were amplified by RAA, whose amplification products were then detected with CRISPR-Cas13a and use RT-qPCR as a control method to evaluate the sensitivity, specificity, and consistency of this method. The results showed that the method could detect as low as 10¹ copies·μL^-1 and had no cross-reactivity with pig-derived pathogens such as porcine circovirus type 1, porcine circovirus type 2, porcine circovirus type 3, porcine reproductive and respiratory syndrome virus, swine fever virus and pseudorabies virus. The positive coincidence rate of 40 clinical samples detected by RAA-Cas13a and RT-qPCR method was 100%, the negative coincidence rate was 84.6%, the total coincidence rate was 95%, and the Kappa value was 0.881. The established RAA-Cas13a detection method has high sensitivity and strong specificity, providing a reliable technical means for the clinical diagnosis and epidemiological monitoring of PEDV.

A 3-year-old, male domestic Anglo mink was presented with a slow-growing mass on the tail tip. The mass was completely removed by surgery. Histological examination performed on the mass revealed a lobulated neoplasm composed of large polygonal cells replaced the last coccygeal vertabra. The neoplastic cells had highly vacuolated cytoplasm (physaliferous cells) and intracytoplasmic Periodic Acid-Schiff-positive granules. Immunohistochemistry revealed cytoplasmic positivity for cytokeratin, vimentin and S-100. Histomorphological features and immunohistochemical results consistent with a chondroid chordoma of humans. This case report described Histomorphological features and immunohistochemical results in chondroid chordoma located at the tail of Anglo mink, and reviews the relevant literature, which can provide a reference for the diagnosis and prognosis of the tumor.