The assay was aimed to establish a SYBR Green Ⅰ fluorescent quantitative RT-PCR method to detect the mRNA of type Ⅰ interferon effect factor ISG15, ISG56, Mx1, OAS and PKR in HeLa cells. Using this method, foot and mouth disease virus (FMDV) L protein inhibiting the type Ⅰ IFN signaling pathway was evaluated. Total RNA was extracted by TRIzol reagent and Oligo d(T)₁₅ was used for reverse transcription, the target gene was amplified individually by PCR and cloned into pMD18-T vector, the positive recombinant plasmid was constructed and used as template for SYBR Green Ⅰ fluorescence quantitative RT-PCR. The standard curve and melting curve were established and the sensitivity, specificity and reproducibility assay were conducted. Using this established method, the inhibiting effect on type Ⅰ IFN effect factor of FMDV L protein was detected. The ISG15, ISG56, Mx1, OAS the PKR relative mRNA level of HeLa cells transfected with eukaryotic expression plasmid expressing FMDV L protein were obviously reduced compare to HeLa cells transfected with empty vector or eukaryotic expression plasmid expressing GST. The SYBR Green Ⅰ fluorescent quantitative RT-PCR method was established, which laid the foundation to evaluate the mRNA levels of type Ⅰ IFN effect factor in HeLa cells.This method was successfully applied to FMDV L protein inhibition of type Ⅰ IFN signaling pathway.

A pair of specific primers was designed based on the sequence of NADL-2 strain and China strain of porcine parvovirus published in GenBank. The NS1 gene of PPV detected in Guizhou province was amplified, cloned, sequenced, sequence analyzed and protein structure predicted. The sequencing results showed that NS1 gene was 1986 bp in size,encoding 659 amino acids,the bases deletion of NS1 gene were found at site 1287,1288 and 1298;amino acids deletion were found at site 429,430 and 433. Phylogenetic tree showed that the NS1 gene sequencing in this experiment was at the same evolutionary branch with NADL-2 attenuated strain;the homologies of coding amino acids between this strain and NADL-2 attenuated strain and Kresse strain were both 98.6%. Protein structure prediction showed that molecular weight of non-structural protein NS1 was 75269.76 u, isoelectric pI was 7.25, instability coefficient was 41.57, speculating this protein was an instable protein. Aliphatic index was 73.58, the grand average of hydropathicity (GRAVY) was -0.565, suggesting that the protein was a hydrophilic protein. This protein contained rich α-helix,β-pleated sheet,β-turn and random coil, and there were many flexible regions, showing continuous distribution. No transmembrane region and signal peptide sequence were found in this protein;motif searching showed that non-structural protein NS1 might contain 3 N-glycosylation sites,3 cAMP- and cGMP-dependent protein kinase phosphorylation sites,5 protein kinase C phosphorylation sites,22 casein kinase Ⅱ phosphorylation sites,1 tyrosine kinase phosphorylation site and 8 N-myristoylation sites,1 amidation site and 1 ATP/GTP-binding site motif A (P-loop);10 major antigenic epitopes distributed in this protein.

To understand the prevalent trend and genetic variations of porcine circovirus type 2(PCV2) from pig farms in Guangdong province, lymphonodi associated with postweaning multisystemic wasting syndrome were collected from pearl river delta in Guangdong province and then the disease materials that proved to be positive by polymerase chain reaction (PCR) method were cultured in PK-15 cells and 5 PCV2 strains were successfully isolated and marked as GD-jm,GD-sz,GD-pz,GD-gj and GD-ss, respectively.The viral genome DNA was extracted and the whole genome of 5 PCV2 isolates were amplified and inserted into pMD18-T Simple Vector. The results were submitted into GenBank and the accession numbers were JX912914,JX912915,JX945575,JX945576 and JX945577.Sequence homology analysis was conducted with the aid of biology software. We found that all of 5 isolates harbored a full length sequence of 1767 bp and 3 of which were designated into PCV2b genotype, the other two were designated into PCV2d genotype.

One pair of specific primers was designed based on inserting sequence IS1111 of Q fever Coxiella burnetii, Real-time turbidity assay of loop-mediated isothermal amplification (LAMP) was developed for indentification Q fever. The recombinant plasmid containing the target sequence was constructed to detect the sensitivity of LAMP with the isothermal temperature. The method could detect 10 of the plasmid copy number. The established Real-time turbidity assay of LAMP method showed no cross reaction with M.tuberculosis, C.psittaci, Brucella.spp and DNA of bovine bloods were negative. Real-time turbidity assay of LAMP was developed in this study and had high sensitivity and specificity,it had a good application prospects in the detection and identification of the Q fever.

To construct cDNA expressing library using MDCC-MSB1 cells in a good state, cDNA were synthesized using biotin-conjugated Oligo (dT) primer in the 5'end. The doublestrand cDNA was ligated to adapter and passed the cDNA size fractionation columns. The cDNA entry library was constructed by BP recombination. The library had a high titer of 1?10⁵ CFU/mL, and contained a total clones of 1.1?10⁶ CFU with an average inserts size was about 1190 bp. The constructed cDNA expression library by LR recombination had a titer of 1?10⁵ CFU/mL and contained total clones of 1.2?10⁶ CFU, with an average inserts size of 1087 bp. The cDNA expression libraries would be a valuable resource for identification of cell receptors of CIAV on the MDCC-MSB1 cells and its cellular tropism.

To establish indirect immuno-histochemical method for detection of Mycoplasma bovis(M.bovis), the pure culture of M.bovis and the lung from the calve infected with the M.bovis were immuno-histochemically studied by using the chicken anti M.bovis IgY and goat anti-chicken IgG-HRP as the first and second antibody, and optimizing the staining conditions to determine the best immuno-histochemistry staining method. The method was used to detect the diseased sections of dead calves under optimal conditions, and the result was verified by isolation and PCR methods. The immuno-histochemistry positive specimens in line with isolation and PCR positive results, while the negative control and alternative test were negative. It showed that the indirect immuno-histochemical method established in this trial could be used to specifically and reliably detect M.bovis of tissue samples and culture in the clinical cases,it also could be provided to explore the site in the organism, the regularity of dynamic distribution and the mechanism of pathogenicity of this pathogen.

To investigate the effects of alfalfa on the expression of liver X receptors (LXRs) gene and protein in pig skeletal muscle cell, and the influence on carcass and meat quality in pig, LXRs protein of longissimus muscle in pig was detected by immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA) methods. The mRNA expression level was quantitated by Real-time quautitative PCR system. The results indicated that LXRs signal was located in the cell nucleus, cytoplasm and cell membrane. Compared with the control group, adding 4% alfalfa to daily ration could elevate the expression levels of LXRs gene and the protein (P<0.05), decrease leanness (P>0.05) and significantly increase intramuscular fat content together with a better marbling score, less water loss and smaller intercellular space (P<0.05). In 6% alfalfa group, marbling score and water lose increased, while suet weight decreased (P<0.05). A higher expression of LXRs gene and protein in 6% alfalfa group were detected, but were lower than 4% alfalfa group (P>0.05). It suggested that different additive amounts of alfalfa might result in different degree of improvement on meat quality for future usage in pig industry.

To investigate the effect of follistatin (FST) gene over-expression on related genes in its pathway, this study cloned the FST gene of Min pig with PCR method, constrcuted eukaryotic expression vector pcDNA3.1(+)-Dsig-FST, identified fibroblast cells transfected with the recombinant plasmid, and detected the change of related genes in its pathway with Real-time PCR method. The results showed that FST gene of Min pig was successfully cloned and over-expressed significantly in the porcine fetal fibroblast cell, and then lead to ActRⅡB and BMP4 gene decline significantly (P<0.05),Smad4 and Myostatin gene had downward trend, ActRⅡA gene had the upward trend. In summary, FST gene changed in its expression had the different effect of degrees inhibition or promotion on its related genes.

18 bacteria strains were isolated through sensory screening test, then through qualitative rescreening test, JNX-8 strain was isolated from poultry litter because of the best denitrification and deodorization ability. Based on morphology and physiology of colony and 16S rDNA sequence analysis, the strain was characterized as Bacillus cereus. The denitrification ability of JNX-8 strain was studied, 51.71% of NO₃^--N and 66.28% of NH₄⁺-N were respectively removed by JNX-8. Therefore, JNX-8 had good application prospects in the denitrification and deodorization of poultry litter.

This experiment was conducted to explore the expression of miR-193b and Kit in different colors of alpaca skin. The tissues were obtained from adult alpaca skins of white and brown hair colors. The comparative expression quantity of miR-193b and Kit in different hair colors was analyzed by real time quantitative PCR. The results of QRT-PCR showed that the comparative expression level of miR-193b in brown alpacas was 1.741346 times than that in white ones, and the comparative expression level of Kit in white alpacas was 4.6029 times than that in brown ones. The findings showed that miR-193b and its target gene Kit might involved in the regulation of hair color formation and miR-193b might regulate the formation of coat color by downregulating the expression of Kit.

The aim of this study was to construct the retroviral vector with mouse miR367 and construction its mechanism of action in cell reprogramming. miR367 was cloned from mouse genome, and then linked with vector pMD18-T.The recombinant plasmid was digested and the segment with miR367 was purified, then subcloned into retroviral vector pMXs. Plat-E cells were transfected by the vector. Viral suspension was infected MEF cells in order to detect the expression of miR367. PCR and restriction enzyme digestion revealed that miR367-pMXs plasmid was constructed successfully. Q-PCR revealed that the expression of miR367 was enhanced in the infected-MEF.Result showed that mouse miR367 retroviral vector had been successfully constructed, and it had laid a good foundation for further research of cell reprogramming.

Streptococcus suis was one of the most frequent pathogenic bacteria which had caused a serious of problems in pig industry all over the world. Therefore, it was important to evaluate the resistance to antibiotics and the distribution of tetracycline resistance determinants in Streptococcus suis from pigs in order to prevent and control this diseases. All Streptococcus suis were isolated from clinically diseased pigs and antimicrobial susceptibility to 18 antibiotics was performed by Kirby-Bauer and broth microdilution method. The results showed that most of the SS isolates showed resistance to tetracycline,sulfamethoxazole and gentamicin.Especially, a high level of resistance was found for tetracycline(98.2%);the isolates were susceptible to cephalosporins,all isolates showed multiple-drug resistance, resistance to 6 and 14 antibiotics was the main trend, the percentage were 20% and 17%, respectively.Four pairs of primers were designed to detect the presence of tetracycline resistance gene taken by SS.The results showed that most of the SS had tetracycline drug resistance gene tetM(85.71%),which indicated that tetM might be one of the main molecular mechanisms mediated the resistance of the SS isolates from Guangdong to tetracycline.

A diseases occurred in a beef cattle fattening farm in Urumqi area,characterised by higher body temperature,dyspnea,coughing and could cause rapid death,daily mortality more than 5%. One bacterial strain was isolated from liver and lung samples of new dead cattle,the bacterial was identified as Pasteurella hemolytica according to cultivation characters,colonial morphology,stain peculiarity and biochemistry test.5 experimental rats received intraperitoneal injection of the bacterial suspension,the result was that all of them died within 8 hours.Combine the epidemiological fact that these cattles just came to Urumqi after long-distance transportation from Yili,so the diagnosis of the disease was bovine shipping fever caused by Pasteurella hemolytica.Drug sensitivity test showed that sarafloxacin hydrochloride and doxycycline were most sensitive, we used these kinds of drugs to the whole herd,it showed good treatment and prevention effect.

RNA-mediated interference (RNAi) is a conserved gene-silencing mechanism, where by the doublestranded RNA matching is used as a signal to trigger the sequence-specific degradation of homologous mRNA. RNAi has become a valuable tool for analysis of gene function through suppression of specific gene products. In addition, RNAi has shown great promise for use in antiviral strategies designed to suppress the expression of pathogenic genes. The present review elaborates the mechanism of RNA interference, the development and application about RNAi inhibiting the virus replication and infection in animals, and the issued obstacles.

A duplex RT-PCR for detection highly and lowly pathogenic porcine reproductive and respiratory syndrome virus(HP/LP-PRRSV) was developed using the primers designed basing on the Nsp2 gene coding the non-structure protein 2 of HP-PRRSV and LP-PRRSV, respectively. The total RNA of standard HP-PRRSV and LP-PRRSV strains was used as the positive control to establish the duplex RT-PCR assay. The sensitivity, specificity and repetition assay of duplex RT-PCR were tested, and samples taken from clinic suspicious PRRSV infected pigs had been testified by the established duplex RT-PCR assay. The results indicated that the duplex RT-PCR assay was successfully established. The specificity and sensitivity of the duplex RT-PCR assay revealed that the threshold of duplex RT-PCR was 100 copies/μL of HP-PRRSV or LP-PRRSV, and no products were amplified from the nucleic acid of Marc-145 cell or the other 8 kinds of pathogenic viral or bacterial microorganism acting as the negative control. The repetition test indicated that the duplex PCR was repeatible. 21 were PRRSV positive detected from 25 clinic suspicious PRRSV infected pigs, while 20 and 4 were HP-PRRSV and LP-PRRSV positive, respectively.The study suggested that the established duplex RT-PCR method was highly specific and sensitive,and suitable for clinic rapid identified diagnosing of HP-PRRSV and LP-PRRSV.

Gene transfection technology has received widespread attention used in tumor gene therapy and gene function studies.Gene transfection methods include physical,chemical and biological methods, and each has its own advantages and disadvantages. Now the main overview focuses on basic methods of gene transfection and its application in tumor gene function and gene therapy.

In the field of virology, the proteomics technologies wildly is used,such as two-dimensional difference electrophoresis, mass spectrometry, Stable-isotope labeling by amino acids in cell culture, and so on. These mainly methods are applied to analyse the dynamic changes of the components, expression level and post-translational modification of proteins from the whole level, then to explore the proteins function and the interact and correlate with them. These will contribute to research the mechanism of viruses infecting the hosts and the interactions, and promote development of diagnosis and treatment of the diseases caused by viruses. In this study, the applications of proteomics technologies in the research of virus and its infection mechanism were discussed and the prospect in virology study would also be forecasted.

To ascertain the pathogen and characteristics of 4 strains canine parvovirus, and establish foundation for further immunization research,the intestinal tract content of 4 different pathogenic dogs were inoculated into the feline kidney F81 cells after aseptic processing. We extracted the total DNA and amplified the VP2 gene with specific primers, then sequenced and typed with typical canine parvovirus’strains. 4 strains of CPV were isolated with F81 cells and the results of typing the VP2 genes showed that 4 canine parvovirus strains all belonged to CPV-2a, respectively named CPV-JQ, CPV-CM, CPV-M and CPV-KM.

A strain of bovine viral diarrhea virus (BVDV) was isolated and identified from the suspected of bovine viral diarrhea virus infection cattle secretions and excretions, and E2 gene sequence was analysed. The results showed that the isolated virus was named as JN strain, Reed-Muench method for the determination of isolate virus TCID₅₀ was 10^-7.5/0.1 mL, virus neutralization test results showed that the BVDV JN isolate could be specifically neutralized by BVDV positive serum, but not by BVDV negative serum neutralization. Results of E2 gene sequences of BVDV isolate showed that the isolate belonged to the BVDV Ⅰa subtype.

To establish a rapid rabbit hemorrhagic disease virus (RHDV) pathogen detection methods, this research based on RHDV gene sequence in GenBank, synthesized outer and inner two pairs of primers, and established a nested RT-PCR for RHDV detection. The method used for Lapine rotavirus (LaRV), Sendai virus(SeV) and healthy rabbit constitution, all of the RT-PCR results were negative; the method for first amplification the sensitivity was 10 ng, the second amplification sensitivity was 0.1 ng, the second amplification was 100 more times sensitive than the first amplification. The established nest RT-PCR method had good specificity and sensitivity, it could detect very low levels of RHDV quickly and accurately, it provided a kind of efficient, rapid, specific and sensitive detection method for pathogen detection and molecular epidemiology of material such as of RHD.

The objective of this study was to investigate the effects of synbiotics on production performance, egg quality and economic benefits in laying hens. One thousand and eighty Zhengdahe laying hens at 265 days of age were selected, and divided into six groups, and six replicates per group and 30 hens per group. The test 1 group was the control group, was fed a corn-soybean meal diet, and the test 2 to 6 groups were fed with the basal diet supplemented with 0.05%, 0.1%, 0.2%, 0.5%, and 1% of synbiotics. The experiment lasted for 60 days. The results showed that the test 3 to 5 groups of ratio of feed to egg were lower 6.96 (P<0.05), 7.33% (P<0.05) and 8.06% (P<0.05) compared with the control group, respectively, egg production rate were higher 4.05% (P<0.05), 3.54% (P<0.05) and 6.17% (P<0.05). The test 5 group of shell thickness, eggshell color, eggshell strength and Haugh unit were higher 5.85% (P<0.05),6.50% (P<0.05),15.49% (P<0.05) and 2.52% (P<0.05).Therefore, the supplementation of 0.5% synbiotics could improve production performance and egg quality for laying hens, and had significant effect.

The aim of the study was to evaluate carcass traits and meat quality of Wuzhishan and Landrace pigs. The results showed that the slaughter weight, carcass straight length, carcass oblique length, average dressing percentage, loin muscle area, average backfat thickness and carcass lean percentage of Landrace pigs were extremly higher than Wuzhishan pigs (P<0.01). Wuzhishan pigs had extremly higher intramuscular fat content than Landrace pigs (P<0.01). No significant difference was observed in pH, cooking percentage, drip loss and shear force (P>0.05). The contents of tasty amino acids, essential amino acids, saturated fatty acids, and monounsaturated fatty acids were also extremly higher in Wuzhishan pigs compared with Landrace pigs (P<0.01), which proved that Wuzhishan pigs had good characteristics of meat quality.

Brown seaweeds were rich in sulfated polysaccharides that could potentially be exploited as functional ingredients for human health. Over the years, sulfated polysaccharides isolated from brown seaweeds with potential pharmacological, nutraceutical, cosmeceutical and functional food properties had broad application prospect. The profound functional properties of fucosan extracted from the brown seaweeds had proven to be invaluable and could be employed in the industrial applications as natural functional ingredients to obtain possible health benefits. As a result of seaweed polysaccharides with the characteristics mentioned above, the author mainly discussed the fucosan (polysaccharide sulfate) isolated from brown algae on anticoagulant, antithrombotic, immunomodulation and anticancer, and its application in industrial production.

Mycotoxins (secondary metabolite) are produced by fungus. In animal body, the toxicity of mycotoxins is chronic, mycotoxins rarely cause death of the animal. But mycotoxins can reduce the nutritional value of feed and the animal production performance, thus impact on productivity. Mycotoxins which remain in animal products (meat, milk and so on) bring a serious threat to human health. The author introduced the common mycotoxins (aflatoxins, ochratoxins, zearalenone, deoxynivaleno, T-2 toxin, fumonisins and so on) in feed, and its toxicity and hazard, as well as reviewed the prevention measures and detoxification methods of mycotoxins to reduce the hazard of mycotoxins.

Forty "Duroc譒andrace譟orkshire" crossbred weanling piglets (21 d of age) were randomly divided into 4 groups of each 10 piglets by weight and sex, which were fed the basal diet with 100, 1000, 2000, 3000 mg/kg zinc (ZnO),respectively. The trial lasted for 49 days, including preset period for 7 days and test period for 42 days. The results showed that 3000 mg/kg zinc (Zn) diet significantly increased the contents of zinc (P<0.05), decreased the contents of copper in serum (P>0.05). 3000 mg/kg Zn could significantly increase contents of zinc in liver, kidney and duodenum (P<0.01). But it had no difference in the serum and other tissues contents of Zn in other groups (P>0.05). 3000 mg/kg Zn diet could reduce the contents of copper in spleen, lymph gland, duodenum, and kidney (P>0.05), and increase the contents of copper in liver (P<0.01). It was confirmed that the 3000 mg/kg Zn diets enhanced the Zn levels in the tissues of weanling stress piglets, and reduced the copper levels in some tissues of piglets.

Staphylococcus aureus was a widely spread opportunistic pathogen with α-hemolysin acting as the vital virulence factor, which could cause serious infection in both human beings and animals. Alcohol reflux was used to obtain the Chinese medicine common Andrographis herb extract. To analyze the effects of the medicine on the α-hemolysin secretion and A549 cell protection, assays including MIC determination, hemolysin assay, Western blotting, RT-PCR, LDH and Live/dead were undertaken. The results showed that the extract could inhibit the hemolysin secretion dose-dependently below the MIC of 2048 μg/mL, and effectively relieve the A549 damage by Staphylococcus aureus supernate, thus indicating it as a potential anti-Staphylococcus aureus drug.

The objective of this assay was to describe the effect of green fluorescent protein (GFP) on bovine mammary epithelial cell (MECs) growth morphology and ultrastructure. After bovine MECs were isolated, cultured and identificated, these cells were marked with GFP. The cellular growth morphology was observed after labeling,and the ultrathin sections were produced, cellular ultrastructure was observed by transmission electron microscopy. Subsequently the karyotype was analyzed. The growth morphology of typical "cobble-stone" monolayer was observed in bovine MECs labeled with GFP (named GFP-MECs) in cytoplasm, which similar to the MECs untransfected and both expressed the broad-spectrum keratin. On the ultrastructure, bovine GFP-MECs and MECs both had large and prominent nucleoli and clear nuclear membrane, and the huge amount of heterochromatin was concentrated in the nucleus, which represented the stability of the genetic material. Cellular surface microvilli developed strongly showed that energy or information exchange was active. Cytoplasm was rich in mitochondria, endoplasmic reticulum and lipid droplets suggested cell nutrients metabolism was strong. Karyotype analysis showed GFP-MECs were euploid karyotype. The results indicated that the green fluorescent protein could be used to label bovine MECs, and could not cause significant impact on the growth activity and ultrastructure.

The aim of the present study was to obtain the highest transfection efficiency by liposome induced exogenous DNA to buffalo fetal fibroblasts. In the study, the exogenous DNA was transfected into buffalo fetal fibroblasts using Lipofectamine LTX. The impacts of liposome amount, plasmid amount and size, transfection time course, initial plated density and different promoter used on transfection efficiency were discussed according to the GFP marker gene. It was indicated that 4 μg pmaxGFP plasmid DNA with 4 μL liposome resulted in the highest transfection efficiency and its cell activity was up to 98% after 6 h transfection.

Bacteriocins, coded genetically and synthesized ribosomally, are a category of bactericidal peptides produced by bacteria. They play a key role in regulating environmental bacterial flora formation. With their wide resource, diverse structures and high potency in killing various pathogenic bacteria of animal diseases, bacteriocins are potentially developed into novel antimicrobials. This review focused on the research development of bacteriocin biological characteristics, the update of bacteriocin classification and the progress on application study of bacteriocins in last several years in order to throw some light on the development of novel bacteriocin veterinary drugs in China.

To investigate the safety of "Runingsan" for treatment and prevention of the mastitis in dairy cow, the acute and sub-acute toxicity was tested in mice and rat. "Runingsan" was extracted with water and concentrated 1.0 g/mL solution. In acute toxicity test, 50 mice were randomly divided into 5 groups, the animals were administered drug by intragastric infusion at single dose of 5000, 7500, 10000, 15000 mg/(kg·BW) respectively. Poisoning typical symptom and death cases were observedand the median lethal dose (LD₅₀) were calculated. In order to estimate the maximum dosage, 40 mice were randomly divided into the treatment group and control group group, "Runingsan" was administered intragastrically at a maximum concentration (1.0 g/mL) and maximum capacity (0.04 mL/g) with single dose in the treatment group, 0.9% saline was used in control group. After being administered, the mice were observed for 7 days. In sub-acute toxicity test, 80 rats were randomly divided into 3 drug-treated groups (high, middle and low dose) and one control group, drug was administered orally at dose of 3000, 1500 and 750 mg/(kg·d) respectively. In control group, rats were administered 0.9% NaCl. Body weight gain, routine blood test indexes, blood biochemical parameters, organ coefficient and histopathology were detected. The results showed there was not any poisoning and death cases in each group, there was no LD₅₀ in this herbal compound. "Runingsan" maximum oral dosage was over 40.0 g/(kg·BW) in mice. These data indicated that the drug was actually non-toxin. There were not significantly different in body weight gain, routine blood test, blood biochemical indexes and organ coefficient between 3 drug-treated groups and control group. The histopathological result was not any lesion in rat real organs of every group. All of these results suggested that "Runingsan" was safety for the dairy cow, which were in mastitis under the recommended dosage and course of the treatment in clinic.

The objective of this study was to investigate the relationship between sterol regulatory element binding factor 1 (SREBP1) gene expression and cell fat formation, recombinant plasmid pEGFP-C1-SREBP1 was constructed and transferred into bovine fetal fibroblasts with Lipofectamine 2000, and then the cells were cultured for 48 h. Furthermore, the expressions of SREBP1 mRNA and protein were detected by Real-time PCR and Western blotting. Oil red O staining was used to explore cellular lipid droplet formation. The results showed that an up-regulation of SREBP1 mRNA in recombinant plasmid transfection group compared with normal control group and pEGFP-C1 transfected group, SREBP1 protein also appeared obvious rise. Oil red O found no lipid droplet in normal control group and pEGFP-C1 transfected group, but marked lipid droplets were detected in pEGFP-C1-SREBP1 transfected group. In conclusion, the up-regulation of SREBP1 directly resulteds in cellular lipid droplet formation in bovine fetal fibroblasts.

Cutaneous and mucosal epithelial cells are a physical barrier and the first defense line of host against environmental changes. The crosstalk between epithelial cells, dendritic cells (DCs) and leukocytes play an important role in inducing proper immune responses during host defense. Interleukin-17 (IL-17) family cytokines are cytokines discovered recently, playing important roles as powerful pro-inflammatory cytokines. They are important players in innate and adaptive immune response of host. IL-17A, IL-17C and IL-17F can directly target tissue epithelial cells to induce various antimicrobial responses against extracellular pathogens and promote tissue remodeling. By contrast, IL-17E primarily acts on leukocytes and induces type Ⅱ immunity that is crucial for protection against parasites. IL-17E also negatively regulates IL-17A and IL-17F production from leukocytes while reports are less on IL-17B and IL-17D.

Taking European pig breeds as control, the single nucleotide polymorphisms of paraoxonase (PON1) gene were investigated in Guizhou indigenous pig breeds, including Xiang, Nuogu, Luobo and Kele pig breeds, using allele-specific polymerase chain reaction (AS-PCR) method. The results showed that two nucleotide mutations, A/T and G/A were found out from site 163/165, and one with C/T variation from site 515 of PON1 gene of porcine population. Three genotypes, wild, mutant, and heterozygous, of site 163/165 and 515 were detected from Guizhou indigenous pig breeds and European ones. Frequencies of genotypes and alleles at site 515 in Guizhou pig breeds were not different from that of European pig breeds (P>0.05). But it was significant at site 163/165 of PON1 gene. Allele A was dominant in Guizhou pig breeds with frequency of 83.7%, and were higher than European pig breeds (25.2%). The mutated nucleotides at site 163/165 encoded methionine (Met 55), which was consistent with that of human patients with metabolic syndrome. The wild genotype of PON1 at site 163/165 (allele B), dominated in Europe pig breeds, contained residue leucine (Leu 55) which was corresponding to population without metabolism disorder. Taking the crystal structure of human PON1 protein as template, three-dimensional structure of PON1 from Xiang pig was deduced. The location of Met 55 Leu was next to the calcium binding site in PON1 structure. It had been demonstrated that the hydrolase activity of PON1-M55 decreased and related to the metabolism disorder in human patients, and the polymorphism of PON1 gene was correlated with the meat quality in pig. It suggested that the polymorphism at site 163/165 in PON1 gene might contribute to the improvement of meat quality in Guizhou indigenous pig breeds.

The prion protein is the pathogenesis agent for transmissible spongiform encephlopathies (TSE) of human being and some mammal. The polymorphisms of the prion protein (PRNP)gene were significantly affect the susceptibility or resistance to TSE for human being and mammal. In this paper, we analyzed the origins, the monitoring and prevention of BSE. The structure and function of PRNP gene in cattle were introduced briefly.The polymorphism of bovine PRNP gene locating in non-coding region was analyzed systematically. We also summarized and compared the insertion-deletion (indel) in the promoter (23 bp) and the first intron (12 bp) polymorphism together for BSE susceptibility. These could assist the cattle molecular breeding project.

Trophoblast stem cells (TSc) were the precursors of the differentiated cells of the placenta. It was associated with embryo implantation. Isolating trophoblast stem cells in vitro provided research foundation for trophoblastic development. TSc had been successfully derivated from preimplantation embryo, there were also many reasearches about porcine trophoblast stem cells(pTSc), but no reports indicated derivating 6 d pTSc. After broken the zona pellucida,we fixed the blastocyst on feeder under sterile condition. The porcine trophoblatic stem-like cell was intially analysed by morphogenesis and RT-PCR. We found the porcine trophoblastic stem-like cell combined tightly and present epithelioid-like cell. The clone was smooth,the ratio of the nucleus and cytoplasm was mass. The porcine trophoblastic-like cell was confirmed expressing Cdx2 gene by RT-PCR. We successfully derived the porcine trophoblastic stem-like cell from 6 d pathenogonidium blastcyst. The result provided some foundation for follow-up studies of porcine trophoblast development.

A total of 416 pigs from Duroc, Large White, Landrace, Shanxi White, Shanxi Black and Mashen were used to detect the polymorphisms of calpastatin (CAST) gene exon 9 by PCR- SSCP. Five SNPs were detected in the amplified region, which were T37G, A150G, C167T, C193T, and C256T, of them, the mutations of C193T and C256T located in the CAST gene exon 9 were missense mutation, which gave rise to the changes from threonine to isoleucine and from threonine to methionine, respectively. Seven haplotypes of A, B, C, D, E, F and G were detected in the examined populations. The distributions of the haplotypes in different pig breeds were anastomosed with the pig economic types. There were three haplotypes of A, B and C in the imported pigs of Large White and Duroc, and the frequency of haplotype B was greater than those of A and C, which were 0.40 in Large White and 0.53 in Duroc. Only two haplotypes of B and C were detected in Landrace, the frequency of haplotype C was greater than that of haplotype of B. Except of the haplotypes of A, B, and C, a new haplotype of E was detected in Mashen pig population, the frequency was 0.16. The haplotypes had no significant effect on the backfat thickness measured alive in Shanxi White pigs.

Bovine respiratory disease (BRD) had a multifactorial etiology and developed as a result of complex interactions between environmental factors, host factors and pathogens, which was of major economic importance to the north America and the global cattle industries,the main bacterial pathogens associated with BRD, including Mannheimia haemolytica, Pasteurella multocida, Histophilus somni and Mycoplasma bovis and other bacterial pathogens were reviewed, and clinical observations, pathology, management and therapy, antibiotic resistance were described so as to provide references for the development and research of drugs for prevention and therapy of bovine respiratory disease complex (BRDC).

In this study, we conducted a survey on the prevalence of Campylobacter jejuni of different varieties of poultry in parts of Guangdong province and identified isolates according to the biological characteristics, such as colony and cell morphology, and cultivating characteristics, and multiplex PCR method. The results showed that Campylobacter jejuni infection rates of chickens, ducks and geese in Guangdong region were respectively 7.93%, 2.46% and 4.16%. The result of clicksFpathogenicity test of 10 selected from the isolates showed that the main pathological changes of the avian Campylobacter jejuni diarrhea, blood in the stool, liver white necrosis of the cecum congestion, swelling filled with bubbles and red contents.

The objective of the study was to verify the migration routes in the horse body about Gasterophilus and describe the damage caused by its parasitic via autopsy examination. The results showed that with the growth of age, larvas gradually migrate towards the end of the digestive tract, and no inverse movements were found. The longest stage of third instars were selective in the parasitic locations, and G. nigricornis and G. nasalis were located in the anterior duodenum, while G. pecorum and G. haemovrhoidalis are in stomach. Study of individuals infected by Gasterophilus proverd an absence of relationship between quantity of Gasterophilus larvas and age (P=0.320) or gender (P=0.665) of the horses. Myiasis was prominent among local equid animals. Wild horses had the highest infection rate, which followed by wild donkeys and domestic horses. A damage of 60% to 80% on total surface area in stomach was found. The results helped to better understand of the harm caused by Gasterophilus and strongly supported the follow-up monitoring and protection on Przewalski horses.

Antigen B (AgB) of Echinococcosis granulose was the most important antigen in the serological diagnosis of hydatid disease. Studies on antigen B of Echinococcosis granulose would promote the development of serological diagnosis agents, epidemiological studies and vaccines to prevent and control of hydatid disease. In this article, the research advances on molecular and immunological characteristics of antigen B and its potential applications on developing diagnosis methods and vaccines were reviewed.

The parainfluenza virus is a nonsegmented negative-strand RNA virus (NNSV),according to the genetic type, it is divided into 1,2,3,4 and 5 types, the fourth type can be divided into 4a and 4b subtypes, five types have bits of cross immunity. Biological characteristics and structures of each type are similar.The host range of parainfluenza virus is wide, it can infect human,canine,cattle and many others animal. The parainfluenza virus has a number of feature that make them attractive candidates as live virus vectors. The vectors can express foreign antigen, as a new vaccine by the reverse genetic system. This review will focus on the genetic engineering research development of parainfluenza virus as virus vectors, including antigenic, genetic structure, biological characteristics and so on.

To understand the prevalence and infection speices of coccidia, a total of 1052 sheep from Henan, Shandong and Northeast China were investigated by using the Sheather’s sugar flotation method. The average infection rate of coccidia was 94.8%. Coccidium oocysts contained in 968 positive samples were conducted morphological identification, and 12 species of Eimeria were detected, including E.ahsata, E.bakuensis, E.parva, E.gonzalezi, E.ovinoidalis, E.granulosa, E.pallida, E.marsica, E.weybridgensis, E.intricata, E.crandallis and E.faurei. The infection rate of mixed species were 71.8% and 2 to 5 mixed species were the most common, however, the maximum number of mixed species reached to 9. The infection rate was 99.4% and 86.0% in less than one-year-old sheep and more than one-year-old sheep, respectively. The average number of oocyst per gramme feces (OPG) was 7907.36 and 3263.89 in the two age groups, respectively. The infection rate of coccidia was 97.0% and 89.0% in drylot feeding sheep and grazing sheep, respectively. Summer and autumn were the two major seasons for coccidia prevalence in sheep.

Immunoenhancer could improve the immune response of animal to antigen and play an extremely important role to enchance vaccine effect. The research progress of the immunoenhancers, such as immunostimulating complex, liposome, cytokine, cytosine phosphate guanidine oligodeoxynucleotide and natural product, were reviewed and the development trend of the immunoenhancer was prospected.

Rabies is a kind of the infectious diseases caused by the rabies virus (RV), which can lead to infect 100% of fatality with man and animals and seriously threaten people health. In this paper, the author mainly summarized the biology properties and the methods for detection of RV, it could provide the technical support for RV detection more effectively.

Haemophilus parasuis disease is also known as pig Glasser’s disease. The Haemophilus parasuis can mainlycause swine multiple serositis, fibrinous pericarditis, arthritis and meningitis. The current prevention of the disease mainly depends on inactivated vaccine, but the cross-protective is not strong or no cross protection between different serotypes. Therefore, the development of new broad-spectrum vaccine is the trend in the future. This paper overviewed the current research situation of Haemophilus parasuis disease vaccine.

To explore simple, fast and practical diagnosis method for cow brucellosis in this study, we established a microscale agglutination test using 96-well "V" plate and optimized the test conditions, the optimal antigen diluted concentration was 1:20, the optimal reaction temperature was 37℃, and the optimal reaction time was 24 h. To verify the veracity microscale agglutination test, we retested 120 rose-bengal plate agglutination test (RBT) positive cow serum samples by microscale agglutination test and tube agglutination test, the coincidence rate of microscale agglutination test and tube agglutination test to RBT were 83.3% and 80.0%, respectively. These results showed that the microscale agglutination test could be used as the alternative method of tube agglutination test for cow brucellosis diagnosis and quarantine.

In order to determine the reason of weaned piglets’ death on an intensive pig farm in Guizhou province, the infected weaned piglets from this pig farm were detected using epidemiological survey, autopsy pathological observation, anatomic pathological diagnosis and RT-PCR method. The results showed that from epidemiological survey and autopsy pathological observation, we primarily diagnosed the pig farm, which was suspected infected by classic swine fever virus (CSFV) and bacteria. The RT-PCR result confirmed that the infected pigs were infected by CSFV and the bacterium was confirmed as Streptococcus suis by bacteria culture and isolation, biochemical character identification and animal pathogenicity test. Comprehensive above test results showed that mixed infection of CSFV and Streptococcus suis were the cause of the pig death in this pig farm.

There were some chickens, collected from a layers farms of Qingdao in 2012, troubled with a disease which caused declining of weight and egg laying, and the clinic anatomise showed that the surfaces of viscus such as livers, lungs, kidneys and pancreases appeared many off-white tumor lesions. The pathological histology analysis result showed that these tumor lesions were of multifocal or multicentric, and the shape of these tumor cells were consistent. Identified pathological changes when infected Marek’s disease virus, the disease was identified to be the chicken lymphocytic leukemia, and some suggestions for the ways to prevent and control the disease were put forward.

The assay was aimed to establish the quality standard of anti-virus Chinese herbal compound extract, so as to supply scientific evidence for the revolution of dosage form and industrialized production,we used TLC method and HPLC method to detect it qualitatively and quantitatively.The results showed that TLC could detect chlorogenic acid and HPLC could detect chlorogenic acid and glycyrrhizic acid.The established chromatographic method could be used in qualitative detection and preliminary quantitative detection exactly and laid the foundation for further study.

Tannins is a kind of mutiple-phenols compoud which widely exists in plant pulse tube. Its chemical property is active and easy to combine with protein. Furthermore,it has a special effect on the nutrition of ruminants. At present, reserches about tannins at home and abroad mainly are consist of two aspects,the biological feature of tannins and the research of the nutrition feature of ruminants by using tannins will be studied. This paper was aimed to summarize the two aspects and look into the future of the prospect application of tannins in the feed of ruminants.

A rapid method for determination of phenylethanolamine A in formula feed by using ultra performance liquid chromatography (UPLC)-PDA had been developed.Phenylethanolamine A in formula feed was extracted with methanol containing 0.1% formic acid,followed mix cation ionic exchange solid phase extraction cartridge clean up,through the sample matrix correction external standard got quantitative.In the optimized conditions,a good linearity of phenylethanolamine A in feed sample matrix was obtained at the concentrations from 0.1 to 20.0 mg/kg,the linear relative coefficient of calibration curve (R²) was 0.9945.The recovery of phenylethanolamine A spiked in formula feed at different concentrations was above 90.36%,and the coefficient of variation was below 5.98%.The limit of detection was 0.06 mg/kg and the limit of quantification was 0.10 mg/kg. The method provided advantages of high sensitive,good precision,reliable data,easy operation and the potential use for rapid determination of phenylethanolamine A for supervising formula feed.