The clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas) system is a genetic engineering technology that can precisely edit the genome of cells and organisms. Compared with the traditional ZFN and TALEN gene editing technologies，CRISPR/Cas has the advantages of slow cost, wide editing range, high target efficiency, simple operation and simultaneous support for multi-site operation. In recent years, the CRISPR/Cas system, especially the Type II and Type A CRISPR/Cas9 system, has been widely used as the latest generation of gene editing technology to improve the breeding efficiency, production performance, disease resistance and animal model construction of livestock, and created a batch of new genetically edited materials for cattle and sheep breeding. In this paper, we provide an overview of the development process, technological transformation, the latest progress in optimization of CRISPR/Cas, as well as research applications in livestock breeding traits, production traits and disease resistance traits, basic principles and applications of CRISPR/Cas9 system, and its latest applications and achievements in livestock breeding. The problems and application prospects of the CRISPR/Cas9-based gene-editing system in livestock breeding are also discussed briefly.

Although cows in silent estrus don’t show overt estrus behaviors, the normal follicular development and ovulation process can also be detected. The cows in silent estrus often miss the best opportunity for artificial insemination since no ideal oestrous identification method, which leads to a longer calving interval and the increase of feeding cost. Therefore, the identification method and the possible causes of silent estrus in cows were reviewed. It was pointed out that scientific management and reasonable nutrition could decrease the occurrence of silent estrus in cows. In addition, the estrus markers in feces, urine and milk should be given an deep study to understand the various physiological changes during the oestrous cycle, combining these physiological parameters and the internet, bioinformatics and other technologies, which will enable us to develop more accurate and efficient estrus detection technology, and thoroughly overcome the technical bottleneck of silent estrus identification.

Circadian clock is timing mechanism, it is an invisible “clock” in the organism, and it is actually an organism inner rhythm of life activity. It is determined by the time sequence structure in the organism. It keeps the periodicity in the internal circulation of behavior, physiology and metabolism, and in sync with the external environment. In this paper, the circadian system, the regulation genes of circadian clock, the regulation of biological clock on nutrient metabolism and digestive organs, and its application in practical production are reviewed.

Bluetongue is listed as one of the notifiable diseases by OIE, and it is classified as type “A” animal diseases by the Chinese government. It has caused huge economic losses to most of the epidemic areas in the world. This disease was first identified in China in 1979, and in the early years of the epidemic of bluetongue disease, it caused great economic losses to animal husbandry because many susceptible animals died. However, bluetongue disease is still an unpopular research direction in China. it is not very clear to many veterinary workers that how many serotypes of bluetongue virus have been isolated and identified in China and how widespread the disease is in China. Especially in recent years, there are few reports of bluetongue disease leading to animal morbidity and death, and people’s attention to bluetongue disease has further decreased. This article briefly describes the prevalence of bluetongue disease in the world, and reviews the 40-year prevalence of bluetongue disease in China, hoping that the disease can attract enough attention.

The M protein of Newcastle disease virus (NDV) is a non-glycosylated membrane-associated protein, which locates at the inner surface of NDV envelope and constitutes the bridge between the viral envelope and the nucleocapsid. Studies have demonstrated that the M protein is a nucleocytoplasmic shuttling protein, and can localize in the nucleus via its intrinsic nuclear localization signal early in NDV infection. According to the reported functions of RNA viruses M protein, the early nuclear localization of NDV M protein may benefit for viral genome replication and transcription, and also inhibit cell gene transcription and protein synthesis. At present, numerous studies have focused on the relationship between M protein and NDV virulence and replication, and also the formation mechanism and utilization of M protein-based virus-like particles. However, there have little researches about the mechanism and function of M’s nuclear localization. Due to the crucial roles of M protein in the replication and pathogenicity of NDV, this summary mainly reviews the cellular localization characteristics of M protein and the molecular mechanism and function of M protein in the nucleus, which will provide theoretical references for further studying the nuclear localization functions and mechanisms of NDV M protein.

Anticoccidial drugs are widely used in livestock and poultry industry to prevent and treat coccidiosis. It can improve feed conversion rate and meat quality. However, the problems, such as feed cross-contamination, toxic effects on non-target animals and excessive drug residues in animal food, are subsequently appeared. Therefore, it is crucial to establish simple, rapid, and sensitive methods for the detection of anticoccidial drug residues in the food of animal origin. The immunoassay technology based on the principle of antigen-antibody specific binding can complete rapid detection and high-throughput screening. It has the advantages of strong specificity, high sensitivity, simple operation, and low cost. This paper reviewed the development of immunoassay techniques for the detection of anticoccidial drug residues in different matrix. Enzyme linked immunosorbent assay, immunochromatography, fluorescence polarization immunoassay, time-resolved fluoroimmunoassay and biosensor immunoassay were highlighted. The trendency of immunoassay in residue detection was also prospected. It aimed to provide methodological reference for the monitoring of anticoccidial drug residues and provide ideas for the establishment of new methods.

Ferritin, an iron storage protein widely found in almost all living organisms, has some main functions of storing and transforming iron, maintaining the balance of cellular iron metabolism and protecting cells from oxidative damage. Ferritin can self-assemble into a spherical nanocage that carries and delivers various metals and a broad range of therapeutic and image agents. Moreover, ferritin has the characteristics of easy modification, good biocompatibility and targeting， and these make ferritin become a non-viral natural nanomaterial with great application potential in the field of biomedicine. This review highlights the structures and biological functions, the current work on ferritin as well as the application prospects based on ferritin nanocarrier platform in the field of disease detection, nanocarrier vaccine and drug carrier in order to provide reference for related researches.

The study aimed to explore the genetic diversity, relationship and family structure of Mashen pigs conserved population. Illumina CAUPorince 50 K SNP chip was used to detect the single nucleotide polymorphism (SNP) in 39 Mashen pigs. The minor allele frequency, polymorphism information content, observed heterozygosity and expected heterozygosity were calculated by Plink software to analyze the genetic diversity of conserved population. The identity by state (IBS) distance matrix was constructed and runs of homozygosity (ROH) was analyzed by Plink software. The G matrix was constructed by Gmatix software. IBS, ROH and G matrix were used to analyze the genetic relationship of conserved population. The phylogenetic tree constructed by Mega X software was used to analyze the family structure of conserved population. The result showed that 43 832 SNPs were detected in 39 Mashen pigs, and the average genotype detection rate was 0.980 1. Twenty-eight thousand eight hundred and fifty-nine SNPs met the demand of quality control, and 72.4% of which were polymorphic sites, which indicated that the SNP chip was suitable for analyzing the genetic diversity of Mashen pigs. The effective allele number, polymorphism information content and minor allele frequency were 1.563 4, 0.412, and 0.258, respectively, which indicated that the genetic diversity of Mashen pigs conserved population was abundant. The average observed heterozygosity and expected heterozygosity were 0.354 1 and 0.349 9, which suggested the conserved population of Mashen pigs had differentiated. The IBS genetic distance of 39 Mashen pigs was 0.284 2, and the IBS genetic distance of 20 breeding boars was 0.285 2. The results of IBS distance matrix and G matrix both showed that some breeding pigs had relationship with each other. Eight thousand one hundred and thirty one ROHs were detected in conserved population of Mashen pigs, 46.15% of which were between 400 and 600 Mb in length. The average inbreeding coefficient of conserved population based on ROH was 0.237, which indicated the high degree of inbreeding in conserved population. The result of phylogenetic tree showed that all individuals belonged to 3 families and the number of each family was significantly different. The genetic diversity of conserved population in Mashen pigs is abundant, but the inbreeding coefficient is high, the family of conserved population is few, and the number of individuals in each family varies greatly, which can lead to the loss of genetic diversity. Therefore, importing the new blood from the original conservation farm of Mashen pigs, expanding population size and decreasing inbreeding coefficient are essential ways in the future.

The study aimed to analyze the effect of cold stress on the expression of lncRNA in adipose tissue of pigs, and discuss the effects of lncRNA and their target genes change on fat meta-bolism and body. In this study, six 6-month-old female Min pigs were randomly divided into two groups. The first group was raised in a normal house and the temperature was controlled at (18±2) ℃ and set as the control group. The second group was raised in an outdoor semi-convertible house with the temperature controlled at (-18±5) ℃ and set as the experimental group. After 24 h of treatment, the subcutaneous fat of back was taken as the experimental material after slaughter. lncRNAs and their target genes induced by low temperature were screened out through transcriptomic sequencing, and the biological function analysis of the differentially expressed genes was carried out, the regulatory network between them was constructed. The results showed that, 166 differentially expressed lncRNAs and 269 genes were induced by cold stress, of which only 2 were known lncRNAs. Among the differentially expressed genes, some genes were related to immune system, such as interferon stimulating genes, DHX58, PTX3 and OASL, some genes were related to respiration, oxidative stress and blood circulation, such as KCNK3, CCDC38, GAS2L2, KNDC1, LIPG and C1QL3, and some genes were related to lipid metabolism, such as ANGPTL4, KLF11, NUPR1 and HERC5, these genes were all significantly up-regulated. However, some genes related to nervous system, such as SLIT1, MAP6, TREM2 and IRFN2, were significantly down-regulated. The differentially expressed genes mainly involved in defense response, response to biological stimulation and response to virus. One hundred and forty four lncRNAs were found coexpression with 208 target genes by correlation coefficient analysis, the main accommodation mode between them was trans-regulation, there was a complex regulatory relationship between them. The screened target genes were significantly enriched in the ribosome biogenesis in eukaryotes and basal cell carcinoma pathways. It is speculated that the innate immune system, respiration, oxidative stress, blood circulation and sensitivity of nervous system were all affected by cold stress, and lipid metabolism was changed.

Inbreeding depression of chicken reproduction is one of major concerns during preserving local chicken genetic resources. This study aimed to investigate the role of genome-wide CpG island (CGI) DNA methylation in inbreeding depression of chicken reproduction. Three healthy hens were selected from the strongly and weakly inbred chicken groups respectively, that is, two groups were set in the experiment with 3 replicates in each group. Whole-genome bisulfite sequencing (WGBS) technology was used to detect and analyze genome-wide methylation differences in gonad axis tissues (including ovary and hypothalamus) between the two groups, and to identify the differentially methylated regions (DMRs). Functional annotation and enrichment analysis of genes with DMRs in their CpG islands were performed. The results showed that no significant differences were found at the methylation levels of ovary and hypothalamus genome between the strongly and weakly inbred chickens (P>0.05). However, 5 948 and 4 593 DMRs were identified between the two groups in hypothalamus and ovary, respectively, of which 1 798 and 995 DMRs were located in CpG islands of the genome. These DMRs were annotated to 1 020 and 552 genes, respectively. In hypothalamus, the genes with DMRs in CpG islands were significantly enriched in GO terms related to reproduction(P<0.05), such as signal transduction, nervous system development, reproductive system development and oocyte maturation regulation. And a total of 19 KEGG pathways were significantly enriched(P<0.05), including TGF-beta signaling pathway, Hepatitis B, fatty acid metabolism and insulin signaling pathways, etc. In ovary, the differentially methylated genes were significantly enriched in 12 pathways(P<0.05), such as chronic myeloid leukemia, influenza A, arginine and proline metabolism and focal adhesion, etc. Particularly, several pathways associated with oocyte development and sex hormone secretion were also enriched, for example, progesterone mediated oocyte maturation, oocyte meiosis, GnRH signaling pathway and estrogen signaling pathways, etc, which included a total of 10 genes, such as CDC27, ADCY8 and AKT3, etc. Thus, a large number of differentially methylated regions were detected between the strongly and weakly inbred Langshan chickens, and a number of differentially methylated genes were found to be related to reproductive traits. It is inferred that the DNA methylation in CpG islands of these genes may play important roles in regulation of inbreeding depression of reproduction in Langshan chicken. The study results will provide a basis for further research on the regulation mechanism of inbreeding depression of chicken reproduction, and will supply a reference for conservation of species resource and future poultry breeding.

This study aimed to evaluate the genetic level of the udder traits of Holstein cattle in Ningxia. The DMU software, the DMUAI module AI-REML, the EM algorithm and multi-trait animal models were used to estimate the genetic parameters of 12 415 Holstein cows in Ningxia, including 111 735 observed values of 9 udder traits. The principal component analysis was applied to explore the relationship between the conformation traits. The results showed that heritability of udder traits of Holstein cattle in Ningxia was at a moderate to inferior level: the values were 0.242 for udder depth, 0.080 for udder texture, 0.183 for median suspensory, 0.230 for fore udder attachment, 0.231 for fore teat placement, 0.173 for fore teat length, 0.272 for rear attachment height, 0.378 for rear attachment width and 0.169 for rear teat placement, respectively. The genetic correlation coefficients ranged from -0.277 to 0.706, while the phenotypic cor-relation coefficients ranged from -0.084 to 0.316. Furthermore, we selected 4 principal components with eigenvalues ≥ 1, 64.19% of the total variance. In addition, all 9 udder traits were reflected in the first 3 principal components, and all loaded coefficients were > 0.4. The present in-depth Dairy Herd Improvement data mining and genetic parameter estimation revealed the characteristics of Ningxia Holstein dairy cow population structure, which may greatly facilitate the design and execution of dairy cow breeding programs.

This study aimed to explore the regulatory role of miR-146a on the proliferation and migration of alpaca melanocytes and its molecular mechanism. The dual luciferase assay was used to verify that MAPK4 and Myosin Va were the target genes of miR-146a by co-transfection with MAPK4 or Myosin Va in 293T cells; Quantitative real-time PCR and Western blotting were used to detect the expression of downstream genes in alpaca melanocytes after overexpressing miR-146a; CCK8 and Transwell were used to detect the proliferation and migration of alpaca melanocytes after overexpressing miR-146a. The results showed that, compared to the negative control, dual luciferase activity extremely significantly decreased by 36% or 30% in 293T cells co-transfected with miR-146a and MAPK4 or Myosin Va, respectively (P<0.001); In alpaca melanocytes, with the overexpression of miR-146a, the mRNA expression of MAPK4 and Myosin Va were extremely significantly down-regulated by 67% and 47%(P<0.001, P<0.01), and the protein expression of MAPK4 and Myosin Va were significantly down-regulated by 38% and 69%(P<0.05, P<0.01), respectively; The expression of genes related to proliferation and migration(CREB, MITF, MLPH, Rab27a) were extremely significantly down-regulated at the transcriptional and translational levels(P<0.01, P<0.001); CCK8 and Transwell results showed that overexpression of miR-146a extremely significantly down-regulated the proliferation and migration ability of alpaca melanocytes (P<0.01). In summary, miR-146a targeting MAPK4 and Myosin Va inhibited the proliferation and migration of alpaca melanocytes through downregulating the expression of MEK1, CREB, MITF, MLPH, Rab27a.

The purpose of this study was to reveal the mechanism of anti-inflammatory effect of BBO by studying NF-κB and Nrf2/HO-1 pathway. RAW264.7 macrophage inflammation model induced by LPS was used. The control group, LPS model group, BBO group (low, middle and high dose) and BBO negative control group were set up in this experiment, with 3 repeats in each group. The effect of BBO on apoptosis was detected by Hoechst 33342 and PI double staining. The contents of IL-1β, TNF-α, PGE₂, LTB₄, NO and the activity of iNOS in LPS-induced macro-phage were detected by ELISA and spectrophotometry. The effect of BBO on the mRNA expression of TNF-α and IL-1β was detected by RT-PCR. The effect of BBO on the key proteins expression in NF-κB and Nrf2/HO-1 signal pathway was detected by Western blotting. The results showed that BBO could reverse the morphological changes and apoptosis of macrophages induced by LPS within the dose range of 80 μg·mL^-1. Compared with LPS model group, BBO of 60-80 μg·mL^-1 could extremely significantly inhibit the secretion of LPS-induced cellular inflammatory factors and inflammatory mediators IL-1β, TNF-α, PGE₂, LTB₄, NO and iNOS activity(P<0.01); BBO could also extremely significantly inhibit the expression of IL-1β and TNF-α mRNA(P<0.01) induced by LPS. At the same time, BBO of 60-80 μg·mL^-1 could extremely significantly down-regulate the protein expression of COX-2, 5-LOX, p-IKKα, p-P65, p-IκB-α and cytoplasmic Nrf2 induced by LPS(P<0.01), and extremely significantly promote the protein expression of HO-1 and nuclear Nrf2 in a dose-dependent manner(P<0.01). The results suggest that BBO has good effects on anti-inflammatory and anti-apoptotic. It may play an anti-inflammatory role by inhibiting the phosphorylation of key proteins and the production of inflammatory factors in NF-κB pathway and promote the expression of major anti-inflammatory genes in Nrf2/HO-1 pathway.

The study aimed to explore the the regulation of BMP4 on the proliferation of testicular sertoli cells. Three healthy Dazu black goat rams in the 0-, 1-, 2- and 3-month-old groups were selected respectively, and the testicular sertoli cells were collected. Each experiment set up 3 biological replicates and 3 technical replicates. Whether BMP4 regulated goat testicular sertoli cells through Id2 and their regulatory relationship were verified by in vitro culture, cell immunofluorescence, gene interference, overexpression, qPCR and Western blotting and other technologies. The results showed that the expression level of BMP4 in testicular sertoli cells of Dazu black goat in 2-month-old group was extremely significantly higher than that in 0-,1-month-old groups (P<0.01); Within a certain range, with the increase of BMP4 concentration, the proliferation activity of testicular sertoli cells increased, and the proliferation ability was the strongest when the concentration of BMP4 was 200 ng·mL^-1(P<0.05); After interfering BMP4 expression, the cell proliferation activity significantly decreased (P<0.05), but returned to the normal level after 72 h (P>0.05), and the expression of PCNA was extremely significantly reduced (P<0.01), indicating that the cell proliferation ability was restricted after interfering BMP4 expression, and the cell proliferation index measurement results further illustrated that the cell proliferation ability was restricted after interfering BMP4 expression; After BMP4 overexpression, the expression levels of Id2 extremely significantly increased (P<0.01), indicating that BMP4 had a positive regulatory effect on Id2; Compared to the overexpressing BMP4 and then interfering Id2 group, in the interfering Id2 group, the expression levels of PCNA gene significantly reduced (P<0.05), which further proved BMP4 could regulate the expression of this gene and protein. In summary, the cell proliferation activity of goat testicular sertoli cells is positively correlated with the concentration of BMP4 within a certain range; BMP4 can positively regulate the expression of Id2, and promote the proliferation of goat testicular sertoli cells through promoting the expression of Id2. This study provides a basis for elucidating the molecular mechanism and physiological functions of BMP4 regulating goat testicular sertoli cells.

The aim of this study was to explore the miRNAs expression profile of porcine oocytes before and after in vitro maturation, and then screen miRNAs involved in regulating Npm2 gene expression. Porcine ovaries were collected at a local slaughterhouse and transported to the laboratory. GV-stage oocytes were collected and matured in vitro to obtain MⅡ-stage oocytes. Total RNA of 130 GV- or MⅡ-stage oocytes were extracted respectively and subjected to small RNA sequencing by Illumina HiSeqTM 2500. Each group was repeated 3 times. The differentially expressed miRNAs was screened, and GO and KEGG analysis were performed on the target genes. The results showed that the miRNAs expression profiles of GV- and MⅡ-stage oocytes were successfully constructed. Ninety five differentially expressed miRNAs were screened in GV- and MⅡ-stage oocytes, and miRNAs with similar expression patterns were clustered into one group. Total 3 967 target genes were obtained from 95 differentially expressed miRNAs. GO and KEGG analysis showed that these target genes were enriched in 5 194 GO terms and 212 KEGG pathways. Among these, there were two pathways related to oocyte maturation. Five miRNAs related to Npm2 gene were screened from the differentially expressed miRNAs. Two miRNAs expression were examined by qRT-PCR, and their expression trend was consistent with the sequencing results. The differentially expressed miRNAs were screened from the GV- and MⅡ-stage oocytes, it was speculated that miRNAs play a role in the process of in vitro maturation of oocytes via metabolism, oocyte meiotic related pathways and progesterone mediated oocyte maturation pathway. And miRNAs regulating Npm2 gene expression were screened. The result of this study can provide a foundation for further study the regulation mechanism of miRNAs on Npm2 gene and their roles in oocyte maturation.

Hawthorn-leaves flavonoids (HF), extracted from hawthorn leaves, were reported to exert antioxidant, anti-inflammatory and hypolipidemic properties. The aim of this study was to investigate the effects of dietary HF on the eggshell quality of aged breeder hens. Qiling breeder hens (60-week-old) of 270 were randomly divided into 3 treatments(6 replicates per treatment, 15 hens per replicate): basic corn-soybean diet (CON); basic corn-soybean diet supplemented with 30 mg·kg^-1 HF (LHF); basic corn-soybean diet supplemented with 60 mg·kg^-1 HF (HHF). Pre-experiment period was 2 weeks，and formal experiment lasted 8 weeks. Eggs were obtained for quality determination, blood samples were taken for determining the content of Ca and P, and shell gland tissues were collected to observe the changes of histopathology, cell apoptosis and antioxidant proteins. The results showed that: 1) In comparison with the CON group, supplemental HF significantly improved the thickness and strength of the eggshell (P<0.05), but had no significant effect on the hgt, yolk color, haugh unit and relative shell gland weight (P>0.05). 2)In comparison with the CON group, serum levels of calcium were elevated and the mRNA expressions of CaBP-D28K, OPN and OC-116 genes in the shell gland of HF treated groups was significantly up-regulated (P<0.05). In the HHF group, the activity of Ca²⁺ ATP enzyme was significantly increased in comparison with the CON group (P<0.05). 3) Histological section analysis showed, HF treatment reduced the apoptosis rate of cells in the shell gland (P<0.05), and significantly increased the length of villi in the shell gland and the density of glands (P<0.05). 4) In comparison with CON group, dietary supplementation with HF significantly reduced the content of MDA in the shell gland (P<0.05), the activity of GSH-Px was significantly increased in HHF group (P<0.05). 5) In comparison with the CON group, HHF treatment significantly down-regulated the protein expression of KEAP1, p-NFκB /NFκB and Bax/Bcl2 (P<0.05), and significantly up-regulated the protein expression of Nrf2, HO-1 and PCNA (P<0.05). In summary, 60 mg·kg^-1HF treatment group could enhance the antioxidant capacity of the shell gland by activating Nrf2/ HO-1 signaling pathway, thus reducing the rate of apoptosis and improving the ability of calcium transport, so as to improve the eggshell quality of aged breeder hens.

This experiment was conducted to investigate the effects of zinc-montmorillonite (Zn-MMT)on the production performance, slaughter performance, immunologic function and intestinal morphology of broilers. A total of 288 1-day-old male Cob broiler were randomly assigned to 6 groups with 6 replicates per group and 8 birds per replicate. The birds were fed a corn-soybean meal basal diet or one of the four Zn-MMT diets or 40 mg·kg^-1 ZnSO₄ diets, which were the basal diet supplemented with 20, 40, 60，80 mg·kg^-1Zn-MMT(zinc content measurement). The broilers were provided the diets in ad libitum basis.Intestinal tract and spleen were collected on 42 days, and the expression of IL-2, TNF-α, IgG, IgA genes, intestinal villus height and crypts depth were measured. The results showed as follows: 1) Compared with CK group and ZnSO₄ group, Zn-MMT had no significant effect on average daily feed intake (ADFI) of broiler chickens (P>0.05).However, 80 mg·kg^-1 Zn-MMT significantly reduced the average daily gain (ADG) of chicks aged 1-21, 22-42 and 1-42 days (P<0.05), and 40 mg·kg^-1Zn-MMT significantly reduced the feed to gain ratio (F/G) of chicks aged 1-42 days (P<0.05).2) Compared with CK group, ZnSO₄ and 60 mg·kg^-1 Zn-MMT significantly increased the bursa index at 21 days of age (P<0.05), and 60 mg·kg^-1 Zn-MMT significantly increased the bursa index at 42 days of age (P<0.05). At the same time, ZnSO₄, 20 and 40 mg·kg^-1 Zn-MMT significantly increased the expression level of IgG in spleen at 42 days old (P<0.05). Meanwhile, ZnSO₄ group significantly increased the expression level of IgA in the spleen (P<0.05).Compared with CK group and ZnSO₄ group, 20 mg·kg^-1 Zn-MMT significantly improved TNF-α gene expression in spleen. The expression of IL-2 in spleen was significantly increased in 40, 60 and 80 mg·kg^-1 Zn-MMT groups (P<0.05).3) compared with CK, Zn-MMT had no significant effect on the slaughter performance of broilers (P>0.05).4) Compared with CK and ZnSO₄ groups, 40 mg·kg^-1 Zn-MMT significantly increased villus height(VH) and villi height to crypt depth ratio(V/C) of the duodenum and jejunum (P<0.05), and significantly reduced the crypt depth (CD)of the duodenum (P<0.05), but had no significant effect on the ileal CD of ileal (P>0.05). Meanwhile，compared with CK group, 40 mg·kg^-1 Zn-MMT significantly increased the ileal VH and V/C (P<0.05), but had no significant difference with ZnSO₄ group (P>0.05). At the same time，ZnSO₄ had no significant effect on VH ,V/C and CD of duodenum, jejunum, VH and CD of ileum of broilers (P>0.05), but could significantly improve the V/C of ileum (P<0.05). It is concluded that dietary supplementation with Zn-MMT can significantly improve feed conversion rate, enhance body immunity, and promote the development of intestinal villi.

The present study was conducted to evaluate the effects of Bacillus Coagulans on growth performance, serum biochemical and tibia indexes of broilers. One hundred and eighty 1-day-old healthy Arbor Acres broilers were randomly allotted to three treatments (basal diet; basal diet + 5 mg·kg^-1 flavomycin, 75 mg·kg^-1 aureomycin and 20 mg·kg^-1 kitasamycin; basal diet + 3.6×10⁹ CFU·kg^-1 Bacillus Coagulans). The experiment lasted for 42 days. The results showed as follows: 1) Bacillus Coagulans supplementation significantly reduced the average daily feed intake of 1-21 day broilers(P<0.05), and no significant difference in BW and ADG were observed among the treatment groups (P>0.05). 2) Compared with the control group, the supplementation of Bacillus Coagulans significantly increased the serum concentrations of triglyceride, total cholesterol and alkaline phosphatase activity of 1-21 day-old broilers (P<0.05). 3) Compared with the control group and antibiotic group, the supplementation of Bacillus Coagulans significantly increased the serum concentrations of total protein, albumin and globulin of 1-21 day broilers (P<0.05), but significantly reduced the serum ammonia and aric acid content (P<0.05), and had no significant effect on serum concentrations of total protein and albumin of day 42 broilers, but reduced the concentrations of serum ammonia. 4) The supplementation of Bacillus Coagulans significantly increased the content of calcium in the tibia of day 21 and day 42 broilers (P<0.05), and no significant difference in tibia (day 21 and 42)ash and phosphorus (day 21) content were observed among the treatment groups (P>0.05). Moreover, the tibia strength was the highest on day 42 in Bacillus Coagulans supplementation group(P>0.05). In conclusion, Bacillus Coagulans can reduce the average daily feed intake and improve the efficiency of feed conversion without decreasing the daily weight gain of broilers in the early stage. Dietary supplementation with Bacillus Coagulans supplementation is also beneficial to promote the metabolism and turnover of lipids and proteins, and improve the deposition of calcium and phosphorus in the tibia of broilers.

To establish a rapid genotyping and quantitative detection method for the foot-and-mouth disease virus (FMDV), the rabbit anti-FMDV and guinea pig anti-FMDV antibodies were used for labeling colloidal gold and NC membrane with strips by double antibody sandwich method. And three types chromatographic strips for detecting serotype O, A and Asia 1 were prepared. A quantitative standard curve was fitted by detecting the 146S of the calibrated reference materials. The quality verification of the immunochromatographic method was evaluated by specificity, sensitivity，reproducibility and correlation with sucrose density gradient. The quantitative detection method established in this study was distinctive, and there was no cross-reactivity among three serotypes of FMDV. This method had no cross reaction with Senecavirus A(SVA), swine vesicular disease virus (SVDV) and vesicular stomatitis virus (VSV). The limits of quantitative detection of the serotype O, A and Asia 1 viruses were 0.567, 0.693 and 0.219 μg·mL^-1 respectively. The assay also showed good linearity with a linear fitting coefficient of determination (R²) higher than 0.97. The correlation coefficient between the newly established method with 3 types lateral flow assay and sucrose density gradient was higher than 0.9. The variation coefficients of the three kinds of lateral flow assay are less than 10%.The results indicated that colloidal gold immunochromatographic quantitative assays for serotype O, A and Asia 1 FMDV could be used for identification of antigen, serotyping and quantitative detection of 146S.

CpxR is the response regulator of the Cpx two component system (TCS), which is involving in maintaining stability of membrane and virulence through regulating the transcriptional expression of target genes in bacteria. This study investigated the effects of TCS CpxR on the biological characteristics, serum bactericidal resistance and pathogenicity of avian pathogenic Escherichia coli (APEC). The cpxR gene mutant and complemented strains of APEC was constructed by Red homologous recombination system and complementation plasmid. Then, the growth curve, motility, biofilm formation, antibiotic sensitivity, serum bactericidal resistance, virulence among wild-type, cpxR mutant and complemented strains were compared and analyzed. There is no significant difference in growth rate and motility among wild-type, cpxR mutant and complemented strains. Moreover, CpxR did not affect the biofilm formation of APEC. However, the antibiotic sensitivity assays showed that cpxR mutant was less resistant to amikacin and kanamycin. The results indicated that of CpxR could enhance the survival ability of APEC in serum. Animal infection experiment showed that the median lethal dose (LD₅₀) of wild-type, cpxR mutant and complemented strains were 7.50×10⁵, 7.50×10⁶ and 1.33×10⁶ CFU respectively, indicating that cpxR mutant strain exhibited attenuated virulence in ducks. These data indicated that TCS CpxR contributes to the antibiotic resistance, serum bactericidal resistance and virulence of APEC, providing help for elucidating the environment adaptability, survival ability and pathogenic mechanism of APEC.

Blastocystis is one of the most common protozoan in human and animal intestines. It can cause intestinal and skin diseases in human and animals. In order to understand the infection status of Blastocystis from sheep and goats in different regions, 704 fecal samples from 7 provinces and regions were investigated by molecular epidemiology based on the SSU rDNA locus, and Multilocus-sequence typing was performed for the first time on ST3 and ST4 of Blastocystis derived from sheep and goats. The results showed that, the total infection rate of Blastocystis was 13.35% (94/704), and the infection rate was significantly different among different regions and breeds. There were 5 subtypes (ST3, ST4, ST5, ST10 and ST14), among which, ST3, ST4 and ST5 were zoonotic subtypes. MLST analysis showed that ST3 was Allele 38, and there were base mutations at 2 gene loci. ST4 included Allele 42, Allele 92 and Allele 94. The results indicated that, the infection of Blastocystis was common in sheep and goats, the zoonotic subgenotype and high genetic diversity of Blastocystis were found too，so more attention should be paid on its prevention and control.

This study was conducted to isolate and characterize a lytic bacteriophage against Proteus mirabilis from piglet fecal samples. Bacteriophage was isolated and purified with indicator of Proteus mirabilis by double-layer agar plate method from fecal samples of piglets. The size and shape of phage was observed with transmission electron microscopy. The whole genome of phage was extracted and sequenced by Illumina Hiseq high-throughput sequencing technology, and then the genome was annotated and analyzed. A lytic phage (designated as vB_PmiP_P59) against Proteus mirabilis was successfully isolated. P59 was a podovirus with C3 morphotype: a flattened oval head of 152 nm×54 nm and a tail of 40 nm×12 nm. The optimal MOI of P59 was 0.001.The one-step growth curve showed that P59 had a latent period of 30 min, a burst period of 105 min, and a burst size of 2 754 PFU·cell^-1. The phage could withstand the temperature up to 50 ℃ and maintain stable titer under pH 5-10.The double-stranded DNA genome was 90 187 bp long with a mol% G+C content of 34.65, and contained 136 ORFs and 4 tRNAs without any virulent or antibiotic resistance genes. Whole-genome proteomic analysis (30.47% homologous proteins) revealed that it was related to C3-like phage phiEco32. A novel lytic Proteus mirabilis phage vB_PmiP_P59 is identified with significant ability to inhibit the growth of Proteus mirabilis.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a disease that causes high piglet mortality and brings great economic loss to the pig industry. In this study, the localization and ultrastructure changes of virus in piglets’ kidney infected with PRRSV were observed to understand the effect of PRRSV on renal tissue cells and blood circulation. Transmission electron microscopy (TEM) combined with serum biochemical test and immunohistochemistry (IHC) was used to observe and analyze the renal tissue of piglets infected with PRRSV. It was found that PRRSV infection caused renal tissue damage, and the virus was mainly distributed in the cytoplasm of glomeruli, necrotic renal tubular epithelial cells and macrophages. Extensive fusion of podocyte processes or formation of microvilli and swelling of endothelial cells were found. The mitochondria in the renal tubules epithelial cells were swollen, cristae fractured and dissolved; In interstitial, inflammatory cell infiltrated. According to the observation results, PRRSV caused serious damage to the kidney, podocyte process fusion resulted in increased glomerular permeability and the injury of glomerular vascular endothelial cells leaded to microthrombus formation. These results indicated that PRRSV affected the glomerular filtration rate and blood circulation system of piglets. It provided a theoretical basis for elucidating the pathogenic mechanism of PRRSV infection.

This study was conducted to determine the elimination of doxycycline hydrochloride tablets in lambs after administration according to the administration instructions and the withdrawal period. Doxycycline hydrochloride tablets were administered orally at 5 mg·kg^-1 of body weight, at an interval of 24 hours, for 5 consecutive doses. After the last administration, fat, muscle, liver and kidney were collected at 12 h, 1 d, 2 d, 3 d, 5 d, 7 d and 9 d respectively, and the content of doxycycline hydrochloride tablets in tissues was determined by HPLC-VWD. The linear range of doxycycline hydrochloride tablets in sheep fat, muscle, liver and kidney tissues is 50-5 000 ng·mL^-1. The linear equation and correlation coefficient are y=0.044x-0.414, R²=0.999. The results showed that doxycycline metabolism in lamb tissue was rapid. Nine days after the last administration, doxycycline was not detected in muscle, liver, kidney and fat. The lamb was given doxycycline hydrochloride tablets at 5 mg·kg^-1 body weight in this study. The withdrawal time was calculated according to the recommendations of the European Agency for the Evaluation of Medicinal Products (EMEA/CVMP/036/95) and was set at 2 days.

The aim of this study is to modify the hen egg-white lysozyme (HEWL) and explore its effects on two gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis) and two gram-negative bacteria (Escherichia coli, Vibrio anguillarum). The egg-white lysozyme was modified under the condition of (57.0±0.1) ℃ at pH 2.0, the ultrastructure of modified hen egg-white lysozyme was observed by transmission electron microscope, and the hydrophobicity of the egg-white lysozyme before and after the modification was measured by 8-anilino-1-naphthalenesulfonic acid (ANS). Circular dichroism and BeStSel software analysis were used to monitor the secondary structure changes of modified HEWL, Oxford cup method was used to measure the bacteriostatic diameter of modified egg-white lysozyme against experimental bacteria, and growth curve determination method was used to determine the minimum inhibitory concentration of modified egg-white lysozyme and Western blot was used to detect the changes of lysozyme content in the four experimental bacteria supernatant and cells. Transmission electron microscopy results showed that the modified egg-white lysozyme generated short rod-shaped fibril structures. ANS and circular dichroism detection results showed that the modified egg-white lysozyme was more hydrophobic and the β-sheet content was increased by 31.7%. Oxford cup method showed that the inhibitory extent of the enzyme on the experimental bacteria was: Vibrio anguillarum>Bacillus subtilis>Staphylococcus aureus>Escherichia coli. The growth curve measurement results showed that the minimum inhibitory concentration was 8 μmol·L^-1 for Vibrio anguillarum, 6 μmol·L^-1 for Escherichia coli, Staphylococcus aureus and Bacillus subtilis. Western blot found that the molecular mass of the modified egg white lysozyme in the bacterial solution was not changed, but the content and molecular mass of the lysozyme in the bacteria were changed. Gram-positive bacteria and negative bacteriaboth showed bands at 10 ku, and gram-positive bacteria showed increased lysozyme content at 22 ku, however the supernatant of the bacteria only had a band at 14.3 ku. These results indicated modified egg-white lysozyme had a strong antibacterial effect on both gram-positive bacteria and negative bacteria，which provided basic data for the application of modified hen egg-white lysozyme in the field of animal husbandry.

The purpose of this study was to investigate the changes in histological structure and the expression of lactoferrin (LF) in the uterus and ovaries of dogs during the normal estrus and with pyometra. Masson’s, VVG and PAS histochemical staining were used to observe the histological characteristics of uteruses and ovaries in anestrus, estrus, and pyometra. The immunohistochemical SP method was used to observe the distribution of LF. The results showed that the endometrium/myometrium ratios in the period of anestrus and estrus were 0.762 0 and 0.924 3, respectively. The content of collagen fibers in the uterine lamina propria in anestrus was higher than that in estrus, and the collagen fibers content of ovary in estrus was higher than that in anestrus. The internal elastic membranes of uterine vascular layer and ovarian blood vessels were clear and intact in anestrus and estrus. During the period of anestrus, the superficial lumen was small and the deep lumen was large, the epithelium of glandular duct was consisted of the single layered columnar epitheliums, and the PAS positive reaction was strong in epithelial cells and glandular cavity. During the period of estrus, the glandular lumen became larger, the glandular epithelium was the single layered cuboidal epithelium, and PAS positive reaction was found in the epithelial cells and glandular lumen. The endometrial epithelium was monolayer columnar epithelium in both estrus and anestrus periods, but the position of the nucleus was different. The endometrial epithelium nucleus of anestrus period was located in the center of cell, the endometrial epithelium nucleus of estrus period was at the top of cell. When the dogs have pyometra, the endometrium/myometrium ratio was 1.615 0, the content of collagen fibers in the lamina propria of uteruses and ovaries was less than that during normal estrus, the glandular lumen was large and irregular in shape, the inflammatory cells infiltrated in the lumen, the glandular epithelium was the single layered cuboidal epithelium, the lymphocytes were located in the basement membrane, and PAS positive reaction was weak in epithelial cells and lumen; the elastic membrane in uterine vascular layer and ovarian blood vessels became thinner and fractured compared during normal estrus; the endometrial epithelium was the single monolayer columnar epithelium, and the nucleus was located at basal plasma membrane. The expression level of LF in uterine glandular epithelium and ovaries in anestrus stage was higher than that in estrus period, and the expression level of LF in endometrial epithelium in estrus was higher than that in anestrus. The expression of LF in uteruses and ovaries of dogs with pyometra was lower. The results suggested that the histological characteristics of uteruses and ovaries of dogs in normal estrus and pyometra were significantly different, and the expression level of LF was also different.

In this study, we compared the anesthetic effect of intravenous-inhalation anesthesia(IVIA) of different doses of alfaxalone combined with isoflurane in dogs. Eighteen experimental dogs were randomly numbered and divided into three groups: L (No.1-6), M (No.7-12) and H (No.13-18). Alfaxalone 3, 6, 9 mg·(kg·h)^-1 intravenous constant rate infusion (CRI) combined with 0.5% isoflurane inhalation was used, for maintenance anesthesia, respectively. The physiological indexes, analgesia, sedation and muscle relaxant effect of dogs were observed and recorded at 1 h (T0) before induction anesthesia, 15 min, 30 min, 45 min, and 60 min during maintenance anesthesia, and 15 min (75 min) after the anesthesia. After 45 days, the 18 dogs were grouped according to their original numbers: CⅠ (No.1-6), CⅡ (No.7-12) and CⅢ (No.13-18) for inhalation maintenance anesthesia (2% isoflurane), all data is also recorded. Propofol was used for induction anesthesia in both rounds. There was no statistically significant difference in physiological indexes between group L and group CⅠ(P>0.05)，however, group L had mild pain reaction；Compared with group M, the temperature and blood oxygen of animals in group CⅡ decreased significantly(P>0.05) and pulse, heart rate, respiratory rate and diastolic blood pressure in group CⅡ significantly decreased (P<0.01). There was no pain response in both groups (M and CⅡ group) during anesthesia. Compared with group H, There were statistical differences in pulse and heart rate in group CⅢ (P<0.05), no pain reaction was observed in both groups (H and CⅢ group) during anesthesia. Both intravenous-inhalation anesthesia and simple inhalation anesthesia can get better anesthesia effect, of which alfaxalone dose of 6 mg/(kg·h)^-1 of IVIA can improve the experimental dogs heart rate and pulse, it has little effect on cardiovascular and temperature, at the same time without pain reaction, good depth of anesthesia and wake up fast, compared with the conventional respiratory simple inhalation anesthesia, this anesthetic scheme can improve the overall anesthesia quality of dogs.

The purpose of our research was to explore the effects of sulfated yeast glucan (sGSC) on the prevention and treatment of artificial Eimeria tenella disease in chickens. In this study, cage feeding was used to randomly divide 200 chickens into group A (4 mg·kg^-1), group B (16 mg·kg^-1), group C (64 mg·kg^-1), group D (infection control group) and group E (healthy control group). Three days before the coccidiosis infection, the chickens of groups A, B, and C were given the above-mentioned concentrations of sGSC by body weight every day, and the chickens of groups D and E were fed with reverse osmosis water (RO water) until the end of the experiment. During the experiment, the clinical symptoms, and bloody stools of each group of chickens were observed and recorded, the feces from 5 to 6 days after infection were collected and the number of oocysts per gram of feces was detected. All chickens were killed on the 7th day after infection, and the organ index was calculated; The diversity of the microbial community in the cecum of the chicken was studied through the Illumina sequencing platform. The results showed that： 1) Compared with group D, the cecal swelling of the chickens in each sugar group was significantly reduced, and their intestinal contents and bleeding were significantly reduced. Compared with the weight of the chickens in the E group, the weight of the A and D groups was significantly reduced (P<0.05), however, the body weight of chickens in group B and C were increased to the level of group E, and there was no significant difference with the body weight from chickens in group E (P>0.05); the number of oocysts excreted by chickens in group B and C was compared with that in group D, and there have a significant reduction in group B and C compared with group D, but there was no statistical difference (P>0.05); The thymus index of chickens in group A increased significantly (P<0.05), and the liver index was also significantly improved (P<0.05); 2) Analysis of the cecal flora found that sGSC increased the relative abundance of Veillonellaceae and Lactobacillaceae in the cecum. The relative abundance of Enterococcaceae has been reduced. The results showed that sGSC could reduce the damage caused by Eimeria tenella infection to chickens, and the effects were more obvious with the increase of sGSC dose. In short, sGSC has good anti-coccidial effects, and has beneficial regulatory effects on the intestinal flora of chickens infected with coccidia. In conclusion, sGSC has a certain anti-coccidial effect and has beneficial regulatory effect on the intestinal flora of chickens infected with coccidia.

Pulmonary fibrosis (PF) is a disease that with high mortality, poor prognosis and unclear pathogenesis. To investigate the expression of MMP-7, KL-6, SP-A, SP-D in bleomycin-induced PF mice, to discuss the application of target-specific near-infrared fluorescent MMP-7 probe and combined detection of 4 markers in bleomycin-induced PF mice to provide the foundation of diagnosis, progress and prognosis of the disease. Forty C57BL/6 male mice were randomly divided into bleomycin group (n=30) and control group(n=10), the different concentrations (2.0,3.5,5.0 mg·kg^-1) of bleomycin were given via trachea to induce the pulmonary fibrosis model in bleomycin group, the control group were given the same dose of saline. At day 28 after administration, histopathological score was use to evaluated the severity of pulmonary fibrosis and alveolitis. The expressions of MMP-7，KL-6，SP-A，SP-D were detected by ELISA, RT-PCR, immunohistochemistry and Western blot. The mice were subjected to NIR imaging at different time after injection of MMP-7 probe to observed the distribution and targeting effect of the probes in vivo. The concentration of BLM was correlated with alveolitis pathological score (P=0.008) and fibrosis pathological score(P=0.03). The expressions of MMP-7，KL-6，SP-A，SP-D were significantly different between bleomycin group and control group(P<0.05). Near-infrared fluorescence probe can specifically target on MMP-7 with stable signal. The abnormal expressions of MMP-7，KL-6，SP-A，SP-D were correlated with PF, they may be a biomarkers of bleomycin-induced pulmonary fibrosis. The combined detection of these four markers can improve the diagnosis ability in PF and provide a strong reliability in diagnosis in clinical use. Noninvasive MMP-7 probe can be used on the PF disease of dynamic monitoring.