Long noncoding RNAs(lncRNAs), a kind of noncoding RNA transcripts with the length more than 200 nucleotides, widely exists in cytoplasm and nuclei of animals, plants and microbes. Studies have shown that lncRNAs can regulate gene expression at chromosomal, transcriptional and post-transcriptional levels. lncRNA plays a vital role in most life processes in animals, such as reproduction, muscle growth, fat deposition, hair follicle development, mammary gland development and lactation. This paper reviews the regulation mechanism and current research status of lncRNAs in livestock economic traits, which would provide theoretical basis and practical reference for the research and application of lncRNAs in animal husbandry.

Trans-splicing(TS) is the process by which two or more pre-mRNA molecules are spliced to form a mature mRNA, a way to produce chimeric RNAs at the post-transcriptional level. TS research started late, but the development of high-throughput sequencing and bioinformatics technologies has provided the effective means for related research. Along with the identification of a large number of TS and its chimeric RNAs, it has been found that TS can produce new functional products-fusion proteins or regulatory non-coding RNAs, also can affect parental genes expression only by competitive use of the parental pre-mRNA, which play important role in physiological activities and diseases. This article reviews the TS definition, type, production mechanism and its role in mammals, in order to provide a reference for completely understanding the research situation and development trend of TS.

During the blastocyst development, embryonic blastomeres were differentiated into trophectoderm and inner cell mass. After the blastocyst hatching, trophectoderm was differentiated into polar trophoblast and mural trophoblast.The differentiation of trophectoderm was very important for early embryonic development in mammals. Tead 4, Cdx 2 and Eomes are primary genes affecting trophectoderm development and knockout or mutation of these genes result in incorrect differentiation of trophectoderm. While the integrality, permeability and fluid transport function of trophectoderm were disfigured, which further led to the failure of blastocyst formation. We comprehensively discussed the development process of trophectoderm, pivotal genes for its formation, key proteins related to its integrality and function as well as underlying potential relationship between these key proteins and pivotal genes, which may provide information for further understanding the molecular mechanisms regulating embryonic trophectoderm development in mammals.

The estrus diagnosis is a detective technology that confirm estrus time by observing the regular changes in behavior performance and reproductive physiology of animals. The reproductive performance and economic benefit will be affected by detecting efficiency and accuracy of estrus directly. The conventional estrus identification method, biological detection method and estrus automation monitoring equipment are summaried and reviewed in this paper. Aim to provide the reference for improving the detection efficiency of animal estrus.

Particulate matter (PM) from pig houses is closely related to pig's health status and production performance and it is one of the important reasons for causing respiratory diseases in pigs. PM from pig houses carry many toxic and harmful substances, such as endotoxin, heavy metals, pathogenic bacteria and so on, thus the PM is more complex and toxic. PM can induce inflammatory response in lung, but the exact mechanism is still unclear. Alveolar macrophage, a kind of free immune cells in lung, play a major role in clearing and processing PM. Firstly, the characteristics and components of PM were put forward. Then the effect of PM on growth performance and respiratory health of pigs was described. Lastly, the effect of PM on alveolar macrophage function and its regulatory mechanisms was focused on. PM can influence the innate immunity of alveolar macrophage by reducing its phagocytic capacity, altering cell polarization and activating TLR4-NF-κB/AP-1 signaling pathway, and it can also function as antigen-presenting cells to initiate adaptive immune responses. In summary, this review provide the theoretical basis for studying PM from pig houses.

Since the discovery of mcr-1 plasmid-mediated colistin resistance gene from China in 2015, colistin resistant genes are divided into 5 categories according to reports, including mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5. The researchers need to know the characteristic and the dissemination to provide reference for prevention and control measures in future. For this reason, we summarized the literature and found:1) The differences in nucleotides and amino acids sequence from mcr-1 to mcr-5 are notable, the origin of these resistance genes is different, such as MCR-5 (the protein encoded by mcr-5) is distinct from MCR-1, MCR-2, MCR-3, MCR-4 with protein sequence identity of 36.11% (MCR-1), 35.29% (MCR-2), 34.72% (MCR-3) and 33.71% (MCR-4), respectively. 2) Many variant subtypes:Except for mcr-5, all other 4 kinds of gene type have several subtypes, such as there are more than ten subtypes of mcr-1 and mcr-3. 3) The common coexistence with other resistant genes greatly increases the probability of multidrug resistance. 4) Spread widely:it has been widely spread in more than 40 countries and 5 continents. In conclusion, colistin resistance genes mediated by plasmid have the characteristics of rapid mutation, complex type, widely spread, and generally coexist with a variety of other antibiotic resistance genes, once a large-scale outbreak of multidrug resistant pathogenic bacteria carrying this gene will greatly threaten the health of human being and animals, supervision and control should be strengthened.

To reveal the regulatory pathways and important candidate genes related to E. coli F18 resistance in Sutai pig (Duroc×Meishan pigs), and further explore the differences of hereditary basis in regulating E. coli F18 resistance between Chinese domestic and foreign pig breeds, Sutai and Meishan populations previously established with resistance and sensitivity to E. coli F18 were selected as experiment objects. Using transcriptome sequencing, we screened out the regulatory pathways and candidate genes related to E. coli F18 resistance in Sutai pig, then analyzed the expressions of candidate genes (proteins) in different serotype of E. coli F18 (F18ac and F18ab)-stimulated pig intestinal epithelial cell (IPEC-J2) by qPCR and Western blot. Additionally, the differential expressions of candidate genes in duodenal tissues between E. coli F18-resistant and E. coli F18-sensitive weaned piglets were detected by qPCR. The results showed that:1) there were 238 differentially expressed genes in duodenum between Sutai E. coli F18-resistant and E. coli F18-sensitive weaned piglets, which were mainly involved in toll-like receptor signaling pathway and glycosphingolipid biosynthesis-lacto and neolacto series pathway, including TLR5, IL-1β and FUT2 genes. 2) The mRNA expression levels of IL-1β, TLR5, FUT2 were all significantly up-regulated after F18ac-, F18ab-stimulated IPEC-J2, respectively (P<0.05), and the corresponding proteins expression were also obviously increased, which indicated that IL-1β, TLR5 and FUT2 might play important regulatory roles in the process of Sutai piglets responsing to E. coli F18 infection. 3) Tissue expressions analysis showed that the expression levels of TLR5, IL-1β, FUT2 genes in duodenum tissues between Sutai E. coli F18-resistant and E. coli F18-sensitive individuals were all significantly different (P<0.05). The expressions of TLR5, IL-1β genes in duodenum tissues between Meishan E. coli F18-resistant and E. coli F18-sensitive individuals showed extremely significant difference (P<0.01), but FUT2 expression was not significantly different (P>0.05). Combined with previous studies of molecular mechanism of resistance to E. coli F18 in Meishan and foreign pig breeds, this study revealed the differences of hereditary basis in regulating E. coli F18 resistance between Chinese domestic and foreign pig breeds. Toll-like receptor signaling pathway including CD14 and TLR5 probably played immune roles in regulating E. coli F18 resistance in Chinese domestic pig breeds such as Meishan pigs, while the glycosphingolipid biosynthesis-lacto and neolacto series pathway including FUT2 was associated with the formation of E. coli F18 receptor in foreign pigs.

The objective of this study was to identify and analyze the differential splicing gene (DSG) between muscle and fat tissues genome in AA broilers. The transcriptomic profiling of breast muscle and abdominal fat tissues (n=3) in AA broilers were sequenced using Illumina Hiseq 2500 technology. The alternative splicing event (AS) and DSG between muscle and fat tissues genome in AA broilers were identified using rMATS software. The GO function enrichment and KEGG pathway analysis were performed for DSG. The results showed that (5 966.00±111.66) and (6 757.00±156.51) (P=0.002) alternative splicing events (AS) were identified in genome of muscle and fat tissues, respectively in AA broilers. Skipped exon (SE) was the main events with the highest frequency of 54.92% and 52.67% of the total number of AS in muscle and fat tissues, respectively; A total of 513 significant DSGs were identified between muscle and fat tissues genome. The GO enrichment analysis indicated that 93 DSGs were significantly enriched in cellular component and molecular function. A total of 31 DSGs were significantly enriched in the regulation of actin cytoskeleton, focal adhesion, pyruvate metabolism and endocytosis pathways;The mRNA expression levels of 14 DSGs were not significantly different between muscle and fat tissues. Through the genome-wide identification and analysis of DSG in muscle and fat in AA broilers, the results of this study will provide insight into the mechanisms of alternative splicing and diversity of differential splicing.

In order to study the structure and function of CYP2J gene and protein of Bactrian camel, the full-length cDNA sequence of CYP2J gene of Bactrian camel was amplified firstly by RACE and RT-PCR techniques. Then the bioinformatics method was used to analyze the characteristics of the protein encoded by CYP2J gene, and the phylogenetic tree of CYP2J protein between Bactrian camel and related species was constructed by using MEGA 6. Finally, real-time quantitative PCR was used to determine the difference of CYP2J gene expression in various tissues of Bactrian camels. The results showed that the full-length CYP2J cDNA sequence of Bactrian camel was 1 770 bp containing a 1 509 bp complete open reading frame (ORF), which encoded a protein with 502 amino acids. Bioinformatics analysis indicated that the protein encoded by the ORF was predicted to be a stable hydrophilic protein without signal peptide recognition function but had a transmembrane segment (12-35 aa), and was anchored to endoplasmic reticulum. And some important physicochemical properties of the protein such as the molecular formula, isoelectric point, molecular weight, number of positively charged residues (Arg+Lys) and negatively charged residues (Asp+Glu) were C₂₆₄₅H₄₀₉₉N₇₁₃O₇₂₆S₁₅, 8.45, 57 983, 59 and 56, respectively. The secondary structure were mainly composed of 46.41% alpha helix, 13.15% extended strand and 40.44% random coil. Further phylogenetic tree analysis indicated that CYP2J protein of Bactrian camel had the closest genetic relationship with CYP2J protein of sheep and cattle. The real-time quantitative PCR result showed that CYP2J gene expressed in every tissues detected of Bactrian camel and the expression level of CYP2J gene was the highest in liver and pancreas, followed by kidney, lung, heart, small intestine and blood vessels, and the lowest in spleen and large intestine. In conclusion, the full-length cDNA sequence of CYP2J of Bactrian camel and the bioinformation of protein encoded by open reading frame(ORF) were successfully obtained in this study, which laid the foundation for preparing the CYP2J protein and antibody of Bactrian camel.

The aim of this study was to construct the method to simply and robustly detect the in vitro enzyme-synthesized c-di-GMP and cGAMP. In this study, the double-stranded DNA containing T7 promoter, VC2 GEMM-I ribose switch sequence or VC2/g20a, spinach2 and tRNA sequence was as the template, T7 RNA polymerase in vitro transcription system was used to prepare VC2, VC2/g20a RNA aptamers. In addition, the binding abilities of RNA aptamer to c-di-GMP and cGAMP were determined by examining the green fluorescence intensity. The results showed that the VC2 RNA aptamer and VC2/g20a RNA aptamer were obtained through the process in which transcribed RNA was heated and then was cooled slowly. Under the optimized conditions of 125 mmol·L^-1 KCl, 30 mmol·L^-1 MgCl₂ for 180 min, VC2 and VC2/g20a aptamers could bind to c-di-GMP and cGAMP, respectively, both with specificity and selectivity. In addition, the half effect concentration (EC50) of c-di-GMP for VC2 aptamer was (89±1.7) nmol·L^-1, the EC50 of cGAMP for VC2/g20a aptamer was (309±4.5) nmol·L^-1. The detection limit of VC2 and VC2/g20a aptamers for c-di-GMP and cGAMP were 5 nmol·L^-1 and 20 nmol·L^-1, respectively. Moreover, it was shown that the transcribed RNA mixture without purification could noticeably detect c-di-GMP and cGAMP by VC0179 synthesized. Thus, this simple and robust method could be useful in detecting the c-di-GMP and cGAMP by VC0179 synthesized in vitro,and which might be applied to detect the activity of other c-di-GMP and cGAMP synthases.

The present study aimed to investigate the effects of epigallocatechin-3-gallate (EGCG) on bovine frozen-thawed semen quality and subsequent fertilization ability. The frozen-thawed semen were incubated with different concentrations of EGCG(0, 5, 10 and 20 μmol·L^-1) at 38.5℃ for 1 h. Kinetic parameters, structural integrity, activities of related enzymes and subsequent fertilization ability were measured. The results revealed that supplementation with 10 μmol·L^-1 EGCG significantly improved the percentage of total motile sperms(TM,%) and the beat cross frequency (BCF, Hz, P<0.05). The percentages of spermatozoa with intact plasma membrane and acrosome were significantly higher in 5, 10 and 20 μmol·L^-1 EGCG treated groups compared with the control group(P<0.05). 5 and 10 μmol·L^-1 EGCG led to higher mitochondrial membrane integrity(P<0.05). The activities of SOD and CAT in spermatozoa treated with 10 and 20 μmol·L^-1 EGCG were significantly increased, while the LDH activity and MDA level were markedly decreased in 10 μmol·L^-1 EGCG group(P<0.05). The cleavage and blastocyst development rates of in vitro-produced bovine embryos originating from 10 and 20 μmol·L^-1 EGCG-treated spermatozoa were significantly higher than for embryos from other groups(P<0.05). In summary, 10 μmol·L^-1 EGCG can enhance the quality and subsequent fertilization ability of bovine frozen-thawed semen.

The purpose of this study was to evaluate the effect of different concentrations of the histone methylation inhibitor UNC0638 on the expression level of transgenic buffalo fibroblast exogenous gene to provide a theoretical basis for studying the mechanism of exogenous gene expression and improving the production efficiency of transgenic buffaloes. After the buffalo fetal fibroblasts (BFFs) were treated with different concentrations of UNC0638(0(control group),0.5,1.5,2.5,5.0 mol·L^-1)for 8 d, the cell growth curve was plotted and the cell karyotype was detected after treated with different concentrations of UNC0638 for 48 h. The methylation level of the histone H3K9me2 of BFFs was detected by immunofluorescent technique, and the effect of UNC0638 on the expression efficiency of buffalo transgenic cells exogenous genes was analyzed by quantitative real-time PCR (qRT-PCR). The results showed that:1) The growth curves of BFFs treated with different concentrations of UNC0638 were similar to that of the control group and all showed "S" type,and 0.5, 1.5, 2.5 μmol·L^-1 UNC0638 had no significant effect on the cell proliferation, while 5.0 μmol·L^-1 UNC0638 inhibited the BFFs proliferation. 2)No significant difference was observed in the normal rate of the cell karyotype among the different groups. 3)In addition,the methylation level of histone H3K9me2 in BFFs treated with UNC0638 was significantly decreased compared with the control group(P<0.05). 4)The qRT-PCR results showed that the relative expression of eGFP in the buffalo transgenic fetal fibroblasts treated with 2.5 μmol·L^-1 UNC0638 was significantly higher than that in the control group in 24 h (P<0.05), while the 0.5, 1.5 and 2.5 μmol·L^-1 UNC0638 groups were significantly higher than that in the control group in 48 h (P<0.05). In conclusion,the research showed that UNC0638 could effectively reduce the methylation level of the buffalo fibroblast histone H3K9me2 and increase the efficiency of exogenous gene expression in buffalo transgenic cells.

This study was conducted to investigate the effects of soluble corn fiber (Fibersol-2) supplementation in diet on growth performance, biochemical indexes of serum and anti-oxidative capacity in piglets. A total of 448 hybird piglets (Duroc×Landrace×Large white) with similar body weight of 28 days old were randomly assigned into 4 dietary treatments, 8 replicates in each treatment with 14 piglets each, and fed a basal diet supplemented with 0, 1, 2 and 4 g·kg^-1 Fibersol-2, respectively. The total experiment lasted for 35 d including 7-d pre-test and 28-d test, and an average initial body weight of piglets in the test was (9.65±0.44) kg. At the end of the test, one pig per replicate (near to the average body weight of the replicate) were weighed, slaughtered and the samples of liver and serum were collected and detected. The results showed that:1) The diets added Fibersol-2 did not significantly affect the growth performance and apparent nutrient digestibility of piglets (P>0.05). 2) Compared with the control group, Fibersol-2 supplementation significantly decreased the contents of triglyceride, albumin, total protein and the ratio of albumin to globulin, and markedly increased the content of urea nitrogen (P<0.05) in serum. Fibersol-2 supplementation of 1 and 4 g·kg^-1 significantly increased globulin content in serum(P<0.05), Fibersol-2 supplementation of 2 and 4 g·kg^-1 markedly decreased total cholesterol content in serum (P<0.05), and Fibersol-2 supplementation of 4 g·kg^-1 significantly reduced glucose content (P<0.05) in serum. 3) Compared with the control group, Fibersol-2 addition significantly enhanced the activity of glutathione peroxidase (GSH-Px) in serum(P<0.05) and catalase (CAT) in serum and liver (P<0.05), respectively, but significantly decreased malondialdehyde (MDA) content in liver (P<0.05); Fibersol-2 supplementation of 2 and 4 g·kg^-1 markedly increased total antioxidant capacity (T-AOC) in serum (P<0.05), and Fibersol-2 supplementation of 4 g·kg^-1 significantly increased the activity of superoxide dismutase (SOD) in serum and liver (P<0.05), GSH-Px and T-AOC in liver (P<0.05), however, significantly reduced MDA content in serum (P<0.05). Under the experimental condition, Fibersol-2 supplementation could improve the immunity, antioxidant capacity of piglets, decrease the levels of glucose and lipids in serum, and promote the health of piglets.

This study aimed to establish the regression equations of metabolizable energy of hulled barley based on conventional physiochemical parameters by the methods of laboratory analysis and broilers metabolic test, so as to provide a convenient method for estimating the metabolizable energy values of hulled barley in practice. Trial 1:the physical and chemical parameters were determined for hulled barley imported from Australia and its milling isolates. Five artificial hulled barleys with different nutrient gradients were formulated with milling isolates from hulled barley based on the main classification indexes, namely crude protein, crude fiber and crude ash according to the classification standard of the People's Republic of China. Trial 2:seven hundred twenty healthy male AA⁺ broilers were used (360 for 11-13 d trial, 360 for 25-27 d trial). The birds were randomly allocated into 5 groups with 12 replicates of 6 birds. The birds of each treatment group were fed one of the 5 artificial hulled barley diets, namely A, B, C, D and E. The apparent metabolizable energy (AME), nitrogen corrected apparent metabolizable energy (AMEn) and nutrient apparent digestibility of the artificial hulled barleys were measured by total collection method for broilers of 11 to 13 and 25 to 27 days. And the stepwise regression method was used to establish regression equations for AME and AMEn. The results showed that:1) Barley types had significant influence on AME, AMEn, digestibility of CP, EE, STC, Ca, P and BWG(P<0.05); Days of age of broilers had significant influence on AME, AMEn, digestibility of EE, STC, P and BWG in broilers (P<0.05). The digestibility of EE and STC were 6.52% and 8.30% higher in birds on d 25 to 27 than these on d 11 to 13 (P<0.05). The AME and AMEn were 0.47 and 0.50 MJ·kg^-1 higher in birds on d 25 to 27 than these on d 11 to 13(P<0.05). 2) The prediction equations for AME (MJ·kg^-1 DM) and AMEn(MJ·kg^-1 DM)of hulled barleys for broilers were as follows:AME=7.814+10.166TP+0.106NDF (R²=0.996,P=0.004), AMEn=10.336+9.740TP-0.121CP (R²=0.983,P=0.017) for 11-13 d; AME=10.436+5.974NaCl+0.131EE+0.019NDF(R²=0.999,P=0.002), AMEn=15.284 -0.312CP+0.258EE-0.081NDF(R²=0.998,P=0.014) for 25-27 d. The maximum difference between the calculated AME values for artificial hulled barley diets using prediction equations and the measured values was 0.06 MJ·kg^-1.In conclusion, the main influencing factors in the regression model are conventional nutritional parameters, namely crude protein, total phosphorus, sodium chloride, ether extract, neutral detergent fiber(NDF), which make it suitable for wide application in production practice.

This experiment was conducted to study the effects of ammonia and hydrogen sulfide stress on immunity and antioxidant function of goats. Eight young female goats were randomly divided into two groups, and each group had 4 replicates. The goats were in the separated environmental controlled chamber A and B processed by the following treatments successively. Chamber A:control group (0-6 d):3 mg·m^-3NH₃+0 mg·m^-3H₂S; treatment 1 (T1) (8-13 d):20 mg·m^-3NH₃+8 mg·m^-3H₂S; treatment 2 (T2) (15-20 d):40 mg·m^-3NH₃+8 mg·m^-3H₂S; treatment 3 (T3) (22-27 d):80 mg·m^-3NH₃+8 mg·m^-3H₂S. Chamber B:treatment 4 (T4) (8-13 d):20 mg·m^-3NH₃+16 mg·m^-3H₂S; treatment 5 (T5) (15-20 d):40 mg·m^-3NH₃+16 mg·m^-3H₂S; treatment 6 (T6) (22-27 d):80 mg·m^-3NH₃+16 mg·m^-3H₂S. The serum samples of goats were collected at 2nd and 6th day of each experiment which lasted for 6 days, then the immune indexes (immunoglobulin, complement, lysozyme and cytokine content) and antioxidant indexes (T-AOC, antioxidant enzyme activity and MDA content) were measured. The results showed as follows:1) Compared with the control group, serum IL-1β content in T3, T4 significantly increased (P<0.05), and serum IL-6 content in T4, T6 significantly increased (P<0.01) at the 2nd day; contents of serum IgA, IgG, IgM, LZM, C3 and C4 significantly decreased (P<0.01) in the treatments (except T1) and serum IL-1β content significantly increased (P<0.05) in T4 at the 6th day. H₂S concentration had the significant influence on the contents of serum LZM (P<0.05), IL-10 (P<0.05), IL-6 (P<0.01) at the 2nd day, and the contents of serum IL-1β (P<0.05), IgG (P<0.01), IgM (P<0.01), IL-10 (P<0.01), C3 (P<0.01), C4 (P<0.05) at the 6th day. With the increase of time in climate chambers, contents of serum IgG, IgM, C3, C4 decreased. NH₃ and H₂S concentrations had the significant interaction effect on contents of serum IL-1β (P<0.05), IgG (P<0.01), IL-6 (P<0.01) at the 2nd day and IL-1β (P<0.05) at the 6th day. 2) Compared with the control group, the activities of serum SOD in T3, T4, T5, T6 and CAT in T4, T5, T6 significantly decreased (P<0.01) at the 2nd day; at the 6th day, the activities of serum SOD, CAT in the treatments (except T1) significantly decreased (P<0.01), the activity of serum T-AOC in T5 significantly decreased (P<0.05), as well as the serum level of MDA significantly increased (P<0.01) in T4, T5, T6. H₂S concentration had the significant influence on the activities of serum GSH-Px (P<0.05), SOD (P<0.01), CAT (P<0.01) and MDA content (P<0.05) at the 2nd day, and the activities of serum T-AOC (P<0.01), SOD (P<0.01), CAT (P<0.01), MDA content (P<0.01) at the 6th day. NH₃ concentration also had the significant influence on the activities of serum SOD (P<0.05), CAT (P<0.05) at the 2nd day, and the activities of serum SOD (P<0.01), CAT (P<0.01) at the 6th day. With the increase of time in climate chambers, the activities of serum SOD, CAT decreased. NH₃ and H₂S concentrations had the significant interaction effect on the activities of serum T-AOC (P<0.05), SOD (P<0.01), CAT (P<0.01), GSH-Px (P<0.01) at the 2nd day, and the activities of serum SOD (P<0.01), CAT (P<0.01) at the 6th day. It is concluded that NH₃ and H₂S could cause stress in goats and reduced the immune function and antioxidant capacity of goats. Furthermore, with the increase of NH₃ and H₂S concentrations and the prolongation of the action time, the degree of damage to the goats became worse.

Avian influenza of H9N2 subtype has been prevalent in chicken flocks of China for years. To elucidate the molecular evolution and antigenic mutation of H9N2 viruses in a chicken farm of Liaocheng, Shandong province in 2017, 6 viruses isolated were analyzed for their molecular characteristics through whole-genome sequencing, and compared with the vaccine strain for their reactivity by Hemagglutination inhibitation test. Nucleotide blast results showed that gene sequences most homologous to each gene segment of the isolates were detected from many reference viruses of wide geographic range including Shandong, Hebei, Shanghai, Jiangsu, Zhejiang, Jiangxi, Hunan, Guangdong and Japan. These results indicated the extensive transmission of the viruses and the frequent reassortment of the viral genes. Phylogenetic analysis revealed that the 8 gene segments of the viruses were closely related to several phylogenetic clades of Eurasian avian lineage, including A/duck/HongKong/Y280-clade, A/quail/HongKong/G1/97-clade and A/chicken/Shanghai/F/98-clade. Genotypic analysis showed that all 6 viral isolates belonged to G57 genotype, which has been the dominant H9N2 genotype in China since 2010. Analysis of key amino acids related to drug resistance demonstrated mutation of S31N of M2 protein, a molecular marker related to viral resistance to ion channel protein inhibitors. In addition, 1 000 serum samples were tested for their antibody level against H9N2 AIVs in cross hemagglutination inhibitation test, with antigens of viral isolates and the vaccine strain, respectively. Antigens of viral isolates resulted in 1 to 2 log₂ lower antibodies in comparison with the vaccine antigen, indicating minor HA antigenic difference between the prevalent viral strain and the vaccine strain. In conclusion, we found that the 6 H9N2 avian influenza viruses prevalent in the chicken flock in 2017 belonged to G57 genotype, and there is a slight difference in HA reactivity between the isolates and the vaccine strain.

The purpose of this study was to explore the infection status and molecular characteristics of duck plaque virus (DPV) currently circulating in ducks. The duck samples suspected to be infected with DPV were collected from Fujian province during 2016-2017, and were used for virus detection, isolation and gene sequences analysis. A total of 24 DPV strains were isolated from these samples, and the median age (Me) of the confirmed DPV-positive ducks were different among duck species. The Me of DPV-positive Mule duck and Muscovy duck were 90-day old and 88-day old, respectively, while the Me of DPV-positive Sheldrake duck was 287-day old. Sequence analysis showed that UL56/LORF5 gene, TK/gH gene and UL2 gene of the 24 isolates shared as high as more than 99% nucleotide homology with those of Chinese DPV reference virulent strains,and there exist regular nucleotide variations among the UL56/LORF5 gene and the TK gene between the current isolates and the Chinese reference virulent strains. Phylogenetic analysis showed that all of the current DPV isolates were gathered into one phylogenetic branch (Group Ⅱ) which different from the Chinese reference virulent strains (Group Ⅰ), and could further be divided into two sub-branches (Group Ⅱa and Group Ⅱb). The above results indicated that DPVs still circulated in ducks with Me-disparity among duck species, carrying some regular mutations compared to the Chinese reference virulent strains. We should strengthen the regular molecular epidemiological surveillance, and be aware of the epidemic variation of the virus. This will help for the rapid and accurate diagnosis and disease control of duck plague in China.

This study focused on determination of the genomic character of bovine ephemeral fever virus (BEFV) strain HN1/2012, which provided data for clarifying the genetic evolution of domestic strains of BEFV in China. Based on the complete genome of BEFV strains deposited in GenBank, eleven sets of primers were designed and used for RT-PCR for eleven overlapped DNA fragments. The obtained fragments were cloned to pGEM T-easy for sequencing and then assembled to obtain the complete genome sequence of the strain HN1/2012, using the DNAstar software. Individual genes and the corresponding open reading frames (ORF) were determined based on the transcription initiation (TI) and transcription termination/polyadenylation (TTP) sequences. The evolutionary relationships between the strain HN1/2012 and the referenced BEFV strains were analyzed based on the sequence similarities and phylogenetic tree constructed based on the G gene sequences. The results showed that the genome of HN1/2012 strain is 14 899 nt in length, comprising a leader sequence (50 nt), N gene (1 328 nt), P gene (858 nt), M gene (691 nt), G gene (1 896 nt), G_NS gene (1 785 nt), α1α2 gene (638 nt), β gene (459 nt), γ gene (400 nt), L gene (6 470 nt), and a trailer sequence (70 nt). The nine genes were separated by intergenic regions (IGRs) of 26, 43, 47, 53, 37, 39, 30 and -21 nt. At genomic level, HN1/2012 demonstrated the highest similarity with JT02L that was isolated form Zhejiang in 2002, but its G gene had highest similarity with epidemic strains LYC11 and LS11 that circulated over the same period. Gene recombination was not found in HN1/2012 by recombination analysis, indicating that gene mutation promoted by immune selection pressure may account for the evolution of HN1/2012. The genomic sequence of bovine ephemeral fever virus strain HN1/2012 was determined in the present study, which may lay a foundation for evolution analysis of BEFV based on complete genome in China.

This study aims at analyzing the pathogenicity of bovine viral diarrhea virus (BVDV) in mice, BALB/c mice were selected as experimental animals in this study, the challenge group were inoculated with yak-derived BVDV1-DJ2, BVDV1-Z6 and Oregon C₂₄V strains respectively by intraperitoneal injection, and the control group were inoculated with DMEM medium. The blood and tissues of the mice were collected on the 5th, 7th, and 10th day after inoculation, and the pathogenicity of the virus-infected mice were studied by blood routine, histopathology, and fluorescence quantitative PCR. The results showed, after the challenge, the clinical symptoms of the mice in the Z6 group were more severe than those in the DJ2 group. The lymphocyte, leukocyte, and platelet contents of the challenge group decreased significantly. The results of the fluorescence quantitative PCR showed that the BVDV was detected in the lung and liver. The rate of Z6 group was significantly higher than that of DJ2 group. HE staining showed that there were inflammatory cell infiltration, necrosis and shedding of intestinal villus epithelial cells in the liver of the challenge group, and it also showed that there were increased spleen endocytosis and reactive hyperplasia of spleen, alveolar atrophy and hemorrhage and other pathological changes. Compared with the Oregon C₂₄V group, the lesion of Z6 group were the most serious and the lesion of DJ2 group was lighter. Using BALB/c mice as experimental animals and the way of intraperitoneal injection, this study can provide data support and guiding significance for the pathogenicity of BVDV-infected mice and the construction of models.

Swine hepatitis E virus (HEV) can infect human by cross-species transmission. Therefore, it is important for surveillance by detection of swine HEV infection in pig herds. In this study, an indirect ELISA for detecting anti-swine HEV IgG antibodies was developed using a truncated swine HEV ORF2 protein expressed in a prokaryotic system as the coating antigen. The procedures of indirect ELISA were also optimized. The sensitivity, specificity, reproducibility, and comparisons with the commercial kit were analyzed. In addition, the application of indirect ELISA for detecting anti-swine HEV IgG antibodies in the field serum samples was also evaluated. SDS-PAGE and Western blot analysis showed that the truncated ORF2 protein was successfully expressed and reacted with the positive pig sera for swine HEV antibodies. The optimal amount of the coating antigen was determined at 200 ng per well and the dilution of sera was 1:200. The optimal coating and blocking buffers were carbonate buffer (pH=9.6) and 2.5% dry milk buffer, respectively. The cut-off value of the indirect ELISA was determined as OD_{450 nm} ≥ 0.296. The sensitivity and specificity were 100% and 1:6 400, respectively. Both intra-and inter-assay variabilities were less than 10%. The confidence between the indirect ELISA and the Wantai commercial kit was 91.2%. The developed indirect ELISA can also be used to detect anti-swine HEV IgG antibodies in the sequential sera from the pigs challenged with swine HEV and the clinical pig sera. Collectively, the developed indirect ELISA have good sensitivity, specificity and accuracy, and can be used for detecting anti-swine HEV IgG antibodies in pig sera from the field and ser-epidemiology investigation for swine HEV infection in pig herds.

To develop a new reliable method for initial diagnosis and epidemiological investigation of ten common diseases of pig, an DNA microarray was established for simultaneous detection of porcine reproductive and respiratory syndrome virus(PRRSV), classical swine fever virus (CSFV), pseudorabies virus (PRV), porcine circovirus 2 (PCV2), porcine parvovirus (PPV), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis of swine (TGEV), swine influenza virus (SIV), porcine cytomegalovirus (PCMV) and food-and-mouth disease virus (FMDV). According to the similarity of the ten viral sequences in the GenBank database, ten pairs of specific primers were designed and ten kinds of virus-specific conservative regions were amplified by PCR, respectively. Then the purified target genes were fixed on the amino group modified slides. The probes labeled with Cy3 were amplified from samples by PCR and hybridized with the cDNA array on the slides. The hybridized results were scanned by scanner. The gene chip detection method was established and its specificity, sensitivity and repeatability were detected. The coincidence test between cDNA chip and PCR/RT-PCR was performed in 1 006 samples. The results showed that this cDNA array was very specific and sensitive, the minimum sensitivity was 10⁵ copies·μL^-1. The coincidence rate between cDNA chip and PCR/RT-PCR was 96.79%. In all, the gene chip detection technology for ten kinds of common swine pathogenic viruses was successfully developed and can be applied to swine pathogens detection in clinic.

A porcine circovirus type 3 (PCV3) was detected from a commercial swine farm in Beijing with an outbreak of postweaning multisystemic wasting syndrome (PMWS) in nursery pigs. This study was conducted to investigate the genetic variation of prevalent PCV3 in China and provide the clues for better control of the spread of disease, we identified its complete genome sequence. A total of 73 clinical samples from pigs with reproductive failure and respiratory disease were collected from pigs farms in Fujian province from 2016 to 2017 and these samples were used to detect PCV3 by PCR. Based on PCV3 genome sequences obtained from NCBI GenBank, three pairs of specific primers were designed and synthesized. The nucleotide sequences were amplified from eye and nasal secretions through reverse transcription-PCR (RT-PCR). The cDNA fragments were inserted into pMD19-T vector and the complete genome sequence was obtained through sequencing. Sequence analysis were run on DNAStar and DNAMAN biosoftwares. Results were as follows:The discovered PCV3 strain was named as PCV3/CN/BJ-YH2016 (GenBank accession number:MG546667). The length of its genome was 2 000 bp and the coding region includes three open reading frames (ORFs). Two of them encode replication-associated protein (Rep) and capsid protein (Cap), respectively. The homology between BJ-YH2016 and other 30 submitted PCV3 genomic sequences in GenBank is from 98.6% to 99.3%. The Rep amino acid sequence has the highest homology (100%) with an American strain PCV3-US/MO2015 (KX778720) and the lowest homology (95%) with a Chinese strain PCV3/CN/Guangdong-SG1/2016 (MF589105) where there are 16 mutations mainly located between positions 5 to 23. The Cap amino acid sequence has 100% homology with an American strain 2164. There are a few mutations comparing to other PCV3 strains but at different positions, amongst which four mutations at position 24, 26, 56, 100 comparing to PCV3-US/MO2015. In conclusion, we identified the complete PCV3 genome sequence from a sick piglet with PMWS. The genomic sequence analysis would help us to come to a better understanding on PCV3 genetic variation and its epidemiologic feature.

The objective of this study was to clarify the regulation of estrogen on the duodenal myoelectric motility and to screen and establish the mathematical model between the monitoring index of monitoring duodenal myoelectric activity and the concertration of estrogen. The ovariectomized and electrode embedded animal models of experimental rabbits were constructed and the different doses of estradiol benzoate were injected intramuscularly. The changes of myoelectric activity in duodenum of rabbits in each group were recorded by BL-420F biological function test system and the results were statistically analyzed. The optimal mathematical model of monitoring index between duodenal electromyography and estradiol benzoate was selected and constructed according to the function fitting accuracy analysis of each monitoring index. The results showed that the amplitude, frequency and the index of myoelectric activity of duodenal myoelectricity were all increased first, and then decreased with the increase of estradiol benzoate dose. The functional model of the relationship between the dosage of estradiol and the index of myoelectric activity was established through the analysis and screening of each monitoring index. The analysis results of this function showed that the index of myoelectric activity was changed on the range from 1.50 to 5.23 mV·min^-1, and the maximum value reached 5.23 mV·min^-1 when the dose of estradiol reached 0.098 mg·kg^-1. In addition, the trend of the index of myoelectric activity with the increase of estradiol dose could be divided into four intervals containing exponential increase, logarithmic increase, exponential decrease and logarithmic decrease with the estradiol dosage of 0.063, 0.098 and 0.133 mg·kg^-1 as the cutting point. The above results suggest that estradiol has a concentration-dependent dual regulatory effect on duodenal myoelectric activity, and this regulation is based on a certain level of myoelectric activity in the duodenum. The regulation of estrogen on duodenal myoelectric activity is indirect. In addition, the myoelectric activity of duodenum showed different tendency under the action of estrogen in different concentration and presented obvious interval effect. The results of this study provide a theoretical basis and reference for elucidating the regulation mode of estrogen in the regulation of duodenal movement in vivo and the development and application of estrogenic drugs.

The aim of the present study to compare the hypoxic ventilatory response (HVR) characteristic and the content of nitric oxide (NO), nitric oxide synthase (NOS) in carotid body (CB) between yaks and Qaidam yellow cattle. In this study, health adult yaks and Qaidam yellow cattle at altitude of 3 200 m were inhaled hypoxic gas of 13.9% O₂, which was simulated to the altitude of 6 000 m; NO and NOS mRNA and NOS protein levels were detected in carotid body of yak and Qaidam yellow cattle living at altitude 3 200 m and simulate-altitude 6 000 m by the real-time quantitative PCR, enzyme-linked immunosorbent assay (ELISA) and immunohistochemical methods. The results showed that the slopes of HVR (△V_E/△SaO₂) in yaks and Qaidam yellow cattle were (0.21±0.10) and (0.50±0.21) (L·min^-1)/% SaO₂ (P<0.01), respectively; There were no significant differences in the expression of the mRNA and protein of nNOS, eNOS, iNOS in CB between yak and Qaidam yellow cattle at altitude 3 200 m (P>0.05), in the expression of the protein of nNOS, eNOS, iNOS in CB between yak and Qaidam yellow cattle at simulate-altitude 6 000 m (P>0.05), in the expression of the protein of nNOS, eNOS, iNOS in yak CB between simulate-altitude 6 000 m and altitude 3 200 m, nor in Qaidam yellow cattle. The NO of the CB in yak and Qaidam yellow cattle at simulate-altitude 6 000 m was significantly higher than that of at altitude 3 200 m (P<0.01,P<0.05), respectively. The content of NO in yak CB was significantly higher than that of Qaidam yellow cattle at simulate-altitude 6 000 m (P<0.01). No significant difference in the content of NO in CB between yaks and Qaidam yellow cattle at altitude 3 200 m (P>0.05). The results suggested that HVR was blunted in Yak in Qinghai-Tibetan Plateau, and there was higher ventilatory response to hypoxia in Qaidam yellow cattles, NO production in the yak CB increases during acute hypoxia, and the elevated levels of NO may suppresses the CB response to hypoxia.

This study was conducted to investigate the effects of berberine sulfate (BS) on TGF-β₁, EGFR, EGF and IGF₁ expression in porcine intestinal epithelial cell (IPEC-J2) that were infected by rotavirus. We established a model in which IPEC-J2 were infected with rotavirus. We established target and reference gene recombinant plasmid and standard curve. TGF-β₁, EGFR, EGF and IGF₁ mRNA levels were quantified in BS and rotavirus treated IPEC-J2 by real-time polymerase chain reaction. Results were as follows:after the infection of control IPEC-J2 with rotavirus, the cells clustered into poorly-organized groups, shed or died. By contrast, the BS-treated IPEC-J2 that were infected with rotavirus formed highly-organized groups. The difference of TGF-β₁, EGFR, EGF and IGF₁ gene mRNA expression in each experimental group was compared with that of the control group, showing that BS can promote the expression of TGF-β₁ and EGFR. However, BS inhibited the expression of EGF and IGF₁ mRNA. Compared with EGF and IGF₁ gene expression decreased significantly, the expression of TGF-β₁ and EGFR gene were increased significantly, suggesting that the effect of repair factors may not be the same in the process of repairing cells injury. Our results show that BS could ameliorate IPEC-J2 injury by regulating repair factors expression to promote the proliferation and transformation of cells for repairing the intestinal mucosa.

The aim of this study was to establish a more sensitive RT-PCR assay for detecting BCoV and to detect CoV from clinically ill yak in the northwest grasslands of Sichuan. The RT-PCR assay was established through designing primers targeted to Nsp7-Nsp9 fragment of BCoV polymerase gene and optimizing the reaction conditions and system, and the clinical samples were detected by the RT-PCR assay. Results revealed that the RT-PCR assay have good specificity and stability, and the detection limit of viral nucleic acid of the assays was 1×10^-2pg·μL^-1. Comparing to two RT-PCR assays targeted to the N gene of BCoV, the RT-PCR assay in this study has a remarkable detection rate for BCoV in clinical samples both of bovine and yak. In 125 diarrhea samples of yak in the northwest grasslands of Sichuan in 2016, the CoV detection rate was 71.20%, In 98 nasal cavity samples of yak in the northwest grasslands of Sichuan in 2016, the CoV detection rate was 72.45%. The RT-PCR assay for detecting BCoV established in this study has a good specificity and stability. BCoV is an important causative agent of yak diarrhea and respiratory disease syndrome in the northwest grasslands of Sichuan.