The eukaryotic expression plasmid of nonstructural protein Nsp1 of highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) strain SX-1 was constructed, and the mutants of Nsp1 protein zinc finger 1 (ZF1) were obtained by site-directed mutagenesis, their expression activities in eukaryotic cells were examined. The Nsp1 target gene was amplified by RT-PCR from the total RNA of SX-1 strain, inserted into the eukaryotic expression plasmid of pCI-neo, and the positive plasmid was designated as pCI-neo-Nsp1. 8C, 10C, 25C and 28C in the Nsp1 protein ZF1 were mutated to A or S respectively by site-directed mutagenesis using pCI-neo-Nsp1 as template and 8 mutant plasmids were obtained. The plasmids were transfected into HeLa cells, the activity of Nsp1 protein was detected by indirect immunofluorescence assay (IFA) and Western blotting. The results showed that Nsp1 protein distributed in the cytoplasm and the nucleus using Flag antibody and Nsp1 protein mainly distributed in the nucleus using Myc antibody. Western blotting analysis showed S mutant proteins were about 44 and 20 ku and A mutant proteins were about 44 ku when detected by Flag antibody, while S mutant proteins were about 44 and 27 ku and A mutant proteins were about 44 ku when detected by Myc antibody. These results showed the eukaryotic expression plasmid of Nsp1 mutant were constructed successfully by site-directed mutagenesis and could be successfully expressed in HeLa cells, which provided the foundation for further researching whether ZF1 of Nsp1 protein downregulated the type Ⅰ IFN.

This study is to establish a method for identifying Hyalomma anatolicum(H.anatolicum), Hyalomma asiaticum(H.asiaticum)and Hyalomma detritum(H.detritum)with molecular markers and to learn the phylogenetic relationship of these ticks. Ticks were collected from domestic animals in Xinjiang and Inner Mongolia provinces and classified by morphology characters. Then the 16S rRNA and COⅠ of ticks were amplified by PCR and subsequently sequenced. Phylogenetic trees were constructed by Mega 5.0 and Mrbayes 3.2. On the phylogenetic trees based on the 16S rRNA, H.anatolicum and H.aisaticum were clustered together with their respective classes. And H.detritum was clustered with their respective class and the H.anatolicum, H.asiaticum formed distinct branches on the phylogenetic trees based on the COⅠ. The method based on morphology that combined with molecular 16S rRNA and COⅠ seemed a simple, accuracy method for the identification of H.anatolicum, H.asiaticum and H.detritum

In order to study the biological characteristic on lynx cells cultured in vitro and do further development about lynx cell genomics with providing a reliable material platform for project of wildlife protection, different lynx tissue cells were got by tissue culture in this experiment, and α-MEM medium was used for the primary culture and serial subcultivation.Then morphology observation, growth curves and karyotype analysis were recruited for their biological characteristic analysis. The results showed that there were some differences between different tissue cells in lynx, ranging from morphosis to growth cycle. During primary culture the growth rate of different tissue cells showed differences.However, there were not significant differences among serial subcultivation.The further karyotype analysis indicated that the number of lynx cell chromosomes was 38, including 18 pairs of autosomes and one pair of sex chromosomes. These results provided experimental material and the basis for further research in the field.The study also would be a useful reference for the establishment of lynx cell line.

Cathelicidins comprised a family of antimicrobial peptides sharing a highly conserved cathelin domain, they played a critical role in the innate immune system of chicken. In this study, the constructed positive clones of Rosetta(DE3)/pET30-CathL1S, pET30-CathL2S and pET30-CathL3S were cultured and induced to express target protein by addition of 1.0 mmol/L IPTG in LB. SDS-PAGE and Western blotting analysis demonstrated that Cathelicidins genes were expressed in the form of fusion protein, except for CathL-1 mature peptide, and the apparent molecular weight of expression products were 7, 8, 7.5 ku, respectively. The recombinant Cathelicidins were able to kill both gram negative bacteria and gram positive bacteria by the means of agar well diffusion assay (AWDA), especially for the antibiotic-resistant strains. Young chickens as animal model were used to test antibacterial activity and the result indicated that 5 mg/kg recombinant CathL-1 could completely protect the chicken from infection of Staphylococcus aureus(ATCC 6538). Low dose of CathL-1 could apparently increase weight of chickens.

A virus was isolated from a dead mink which was suspected of being infected with the mink enteritis virus(MEV). It was eventually identified as MEV using PCR assay, cell culture and SDS-PAGE, and named MEV-WFD. The VP2 gene of this parvovirus isolate was cloned into pMD18-T vector, sequenced and analyzed. As a result,the VP2 protein possessed one sbustitution, 328A-T.The result of the nucleotide homologies of VP2 gene between this isolate and all MEV published on GenBank illustrated that MEV-WFD had the the highest homology 99.8% with ZYL-1,MEV/LN/-10 and Manzhouli.VP2 gene phylogenetic anaysis showed that this isolate and other six wild strains had the same ancestor.This research had laid the foundation for the mink enteritis parvovirus molecular epidemiological investigation.

The gene coding for Eimeria tenella microneme protein 3 (EtMIC3) of different strains was amplified by RT-PCR, subsequently cloned and analysed, and compared with related sequences published on GenBank. The results showed that the EtMIC3 sequence of 8 different strains was 2976 bp in length, with little difference in each other. The sequence similarities of nucleotide were 99.8% to 99.9% compared with FJ374765.1 CDs available in GenBank, and 99.9% to 100.0% within each of EtMIC3 obtained in this study with no obvious difference between strains of different geographical origin or drug resistance. The sequence similarities of induced amino acid were 99.6% to 99.9% compared with EtMIC3 (ACJ11219) available in GenBank. It was the first report on the analysis of EtMIC3 sequences of different Eimeria tenella strains from China. The results revealed EtMIC3 was high conservative, and could be used as valid vaccine candidate for the further study on constructing genetic engineering vaccine.

The assay was aimed to construct the eukaryotic expression vector pEGFP-N3-Smurf2 carrying Smad ubiquitination regulatory factor 2 (Smurf2) and detect its expression after transfecting to renal tubular epithelial cells (RTECs).The CDS of Smurf2 gene was amplified and cloned from total RNA of RTECs by RT-PCR.Both prokaryotic expression vector pEASY-Zero-Smurf2 and eukaryotic expression vector pEGFP-N3-Smurf2 were constructed and then detected by double digestion and sequencing.pEGFP-N3-Smurf2 was transfected into RTECs with Lipofectamine^TM2000 and the expression of Smurf2 was detected by Real-time PCR and Western blotting.The amplified Smurf2 gene was about 2247 bp in length.Restriction analysis and sequencing proved that Smurf2 was successfully inserted into recombinant plasmid pEGFP-N3.The mRNA and protein of Smurf2 were detected by Real-time PCR and Western blotting.Smurf2 expression vector using green fluorescent protein as a reporter gene was constructed and transfected into RTECs successfully,which provided a powerful tool for the further study of function of Smurf2 gene.

In order to understand the stability of blue-tongue virus (BTV)storage in cold room for long time, 18 tissue samples which were kept in 4℃ for 10 to 15 years were test by Real-time PCR, and 10 blood samples which were kept in 4℃ for 15 years were subjected to virus isolation process, then was proved by monoclonal antibody staining,research results showed all old samples were positive in Real-time PCR, further more, 5 live virus were isolated from 10 old blood samples. The results suggested the stability of BTV was higher than we thought before.

The objectives of the present study were to examine sequence variation in the mitochondrial large subunit ribosomal rRNA (rrnL) and NADH dehydrogenase subunit 5 (nad5) genes among Taenia pisiformis isolates from Henan province and study its phylogenetic relationships with other cestodes using rrnL and nad5 sequences. The partial rrnL (prrnL) and nad5 (pnad5) genes sequences were amplified from each T.pisiformis sample by polymerase chain reaction (PCR), and prrnL and pnad5 sequences were aligned using the ClustalX 1.81. Maximum parsimony (MP) tree was constructed using the software PAUP 4.0 Beta 10. The lengths of prrnL and pnad5 sequences were 847 and 602 bp, respectively. Sequence similarity in rrnL among the examined T.pisiformis samples were 99.65%, and 99.5% for nad5, whereas inter-specific similarity among taeniid cestodes were significantly lower than 83.4% for rrnL, and lower than 85.7% for nad5. Phylogenetic analysis showed that all the T.pisiformis isolates clustered in the same clade. It was concluded that prrnL and pnad5 sequences could be used as a genetic marker for the identification and differentiation of taeniid cestodes.

ADP-L-glycero-D-manno-heptose-6-epimerase encoded by rfaD gene was an enzyme essential for LOS synthesis. And it was confirmed as an important virulence factor of Haemophilus parasuis. In this study, the rfaD gene of H.parasuis serovar 4 strain clinical SC096 was amplified by PCR with a pair of specific primers based on the complete gene sequence of H.parasuis genome. And it was cloned into pMD19-T vector, then subsequently sub-cloned into expression vector pET-32a(+). SDS-PAGE and Western blotting analyses suggested that the recombinant BL21 (DE3) bacteria induced by IPTG were able to express the rfaD protein with molecular weight of 50 ku, which could react with the positive serum of H.parasuis serovar 4.

To reveal the major protein related to milk protein synthesis in nucleus of mammary gland epithelial cells and further uncover the mechanism of milk protein synthesis in post-transcription level, two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization/time of flight mass spectrometry (MALDI-TOF-MS) were used to identify the changes of nuclear phosphorylated proteins in DCMECs treated with methionine, and their expression changes were verified by Western blotting analysis in the current study. 5 up-regulated expressed phosphoproteins included staphylococcal nuclease domain-containing protein 1, Septin-6, Glycyl tRNA synthetase, Twinfilin-1 and eukaryotic elongation factor 1-beta. They contributed to multiple functional activities such as DNA transcription, mRNA translation, protein synthesis, cell cycle, cell division and morphological processes.

The outer membrane protein (OMP) was deposited in GenBank. The upstream and downstream primers were designed, and OMP gene was amplified by PCR. Then the target gene was inserted into the vector pMD18-T Simple and pET-28a, and the recombinant vectors were used to select and be transformed into E.coli TOP10 and BL21 (DE3), respectively. The plasmid from E.coli TOP10 and BL21 (DE3) was digested by restricted endonuclease enzyme of XhoⅠand EcoRⅠ. Then the plasmid of enzyme digestion was analyzed by its sequence. The recombinant protein was expressed by IPTG induction. We successfully obtained OMP by identification of SDS-PAGE, and result of Western blotting showed that the OMP had immune activity.

In order to explore the difference of outer membrane protein (OMP) of different serotype of Avibacterium paragallinarum, the OMP of three reference strains of Avibacterium paragallinarum serotype A (221),B(0222) and C (Modesto) were prepared by FOCUS^TM Global Fractionation Kit and examined through SDS-PAGE. As a result, three different serotypes of Avibacterium paragallinarum showed similar SDS-PAGE profile. In each samples, there were seven main strips, each had the similar molecular weight which was analyzed by the gel quantitative software. The percentage of serotype A, B and C was 70.85%, 68.6% and 65.71%, respectively. In addition, the molecular weights of three bands were somewhat identical, whereas the protein content of them were significantly different (P<0.05). This study provided a theoretical basis for the further study of the OMP of Avibacterium paragallinarum.

The VP4 gene was acquired by RT-PCR from Binzhou isolate of infectious bursal disease virus, the VP4 expression plasmid was constructed by inserting the target fragment into pGEX-4T-1 vectors. The expression VP4 proteins were acquired by inducing and purifying. An immunochromatograpic strip was developed for the detection VP4 protein antibody of infectious bursal disease virus. Rabbit anti-chicken IgG was labled with colloidal gold as a detection reagent, and recombinant VP4 protein of IBDV was blotted on the test line while goat anti-rabbit IgG was used on the control line of the nitrocellulose membrane. The results of specificity showed that the strip was positive in the detection of reference sera against IBDV BC6/85 virulent and negative to the detection of standard positive serum of Newcastle disease virus, anvian influenza virus H5 and H9 subtypes, infectious bronchitis virus were negative, 0.85% isotonic Na chloride, reference sera against IBDV BC6/85 virulent. Compared with ELISA, the sensitivity of the gold-immunochromatographic assay (GICA) was low two titer. The agreement rate between the two tests was 99.38%. The detection time of VP4 IgG against IBDV by the GICA was less than 10 min. The GICA test strip was a reliable and useful tool for the on-site surveillance of IBDV.

Primers and probes were designed based on inserting sequence IS1111 of Q fever Coxiella burnetii, TaqMan Real-time quantitative PCR assay was developed for indentification Q fever. The recombinant plasmid containing the target sequence was constructed to detect the sensitivity and prepare the standard curve. The method could detect 10 of the plasmid copy number. Related coefficient was 0.995 of the standard curve, the amplification efficiency was 103%. The results of specific detection of nucleic acid sample for M. tuberculosis, Brucella. spp, C. psittaci and bovine blood were negative. TaqMan fluorescent quantitative PCR method 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.

The mouse model for evaluation of protective immunity was established in Brucella abortus. 6-week-old female BALB/c mice were randomly divided into 3 groups (n=40). The mice were each injected intrap eritoneally with 5.0?10⁴ CFU of strain A19 (A19-challenge-group), and another two unimmunized groups served as unimmunized-challenge-group and PBS group with 0.2 mL of PBS.At the 45-dayspost-immunization, all of mice in A19 group and challenged group were each challenged with 3.0?10⁴ CFU of the same subtype 2308 strain.Fifteen days postchallenge, mice were killed and their spleens were removed, weighed, Brucella isolation and pathological changes of organs could be detected in the mice. The results showed that the Brucella load of gram of spleen in A19-challenge-group and unimmunized-challenge-group had a significant difference than the PBS control group (P<0.05). At the 45-day-postchallenge, the result of Brucella load of gram of spleen had no significant difference between the A19-challenge-group to the PBS control group (P>0.05). However, it had significant difference in A19-challenge-group than the unimmunized-challenge-group. The mice of pathological changes in A19-challenge-group had significantly lighter than that of unimmunized-challenge-group (P<0.05). These results indicated that BALB/c mouse model could be applied to evaluate the protective immunity in Brucella abortus.

According to the gene sequences in GenBank of Haplosporidium sp, three specific primers were designed for amplifying the specific fragments of Haplosporidium sp. The reaction parameters were optimized to develop the semi-nested PCR method for detection of Haplosporidium sp. The 333 bp specific DNA fragment of Haplosporidium sp was amplified, but not from other pathogenics such as Marteilia refringens,Perkinsus sp, Vibrio parahaemolyticu, Vibrio alginolyticu and Vibrio fluvialis by this semi-nested PCR. The sensitivity results showed that as little as 0.01 fg DNA plasmid of Haplosporidium sp was detected by this semi-nested PCR. Shellfish samples of Shandong coastal were detected by this semi-nested PCR. As the result, 6 Oyster and 8 Manila clam were Haplosporidium sp positive. It suggested that Haplosporidium sp existed in the cultivated shellfish in Shandong and this semi-nested PCR could be used as a sensitive tool to detect Haplosporidium sp in clinical samples.

To provide theoretical basis for large scale production of animal interferon,the development of the animal interferon on gene modification,the selection of prokaryotic expression conditions,the preparation methods of products and large scale production are introduced in this paper.

E3L protein is a kind of nonstructural protein of poxvirus expressed in early time. It can inhibit immunity of host cells and be ralated with virus virulence and tropism. However, E3L protein in sheeppoxvirus (SPPV) is rarely understood. We have predicted N-terminal of E3L protein in SPPV by bioinformatics methods as well as using the N-terminal of E3L proteins which have been predicted. And then the structural domains, secondary structures, tertiary structures and amino acid compositions of the above proteins have also been predicted using these online programs, such as WebLab, SWISS-MODEL and ExPASY tools. The bioinformatical analysis results show that the protein and other Zα famliy proteins have the same basic structural characteristics and structural domains, implying that the N-terminal of E3L protein in SPPV might belong to Zα famliy.

Pig has been an important biologic model of scientific research over the years. Transgenic pigs also have extremely important effects in biologic model, organ transplantation, animal breed breeding, and so on. There are several problems for the transgenic cloned pigs, such as low successful rate and live rate. The induced pluripotent stem cells (iPSCs) technology has wide and amazing application in regenerative medicine and animal husbandry while this technology has not deeply used in pig transgenic research. Here, we briefly review the transgenic technology, iPSCs technology, and the application of the iPSCs technology in the researches of transgenic cloned pigs for providing some references for researchers working in the fields.

Secretory IgA (SIgA) as a antibody which is packaged in the intestinal mucosal, could protect the gut from pathogenic microorganisms and toxins attack without causing an inflammatory reaction, plays an important role in mucosal immunity and intestinal homeostasis. In animal intestine studies, SIgA blocks pathogenic microorganisms invade the mucosa-associated lymphoid tissue, through the regulation of intestinal epithelial cell receptor recognition ability. Subsequently SIgA promotes to entrap them in mucus, and facilitate their removal by peristaltic and mucociliary activities. In addition, recently it has also been reported that SIgA reveals new mechanism in intestinal epithelial cell. We summarize the intrinsic biological activities now associated with SIgA and their relationships with immunity and intestinal homeostasis,and try to explore the potential role of SIgA transcytosis mechanism.

The dendritic cells played a central role in antigen recognition,offer,induced proliferation of T cells and antigen-specific immune response.In recent years, it was found that the cells could also induce immune tolerance and this role was a hotspot in immune tolerance research.This article elaborated the research satus from two aspects, the central and peripheral tolerance. The mechanisms were summarized from five aspects,the regulatory T cells, the surface of the inhibitory molecule,mature state, the microenvironment and intracellular signal transduction. The overview would provide relevant informations for the research of the dendritic cell tolerance.

Because duck circovirus (DuCV) caused severe economic losses,based on DuCV conserved region in GenBank, two pairs of primers were designed and synthesis. Through optimization of PCR reaction conditions, a nested PCR method had been developed for detection of duck circovirus. Amplification results of duck viral hepatitis virus, duck plague virus, duck parvovirus, Newcastle disease virus were negative using the established method. The first amplification sensitivity was 10 pg, and the second amplified sensitivity was 0.1 pg, sensitivity was increased by 100 times by nested PCR. The established nested PCR method had strong specificity, high sensitivity, good repeatability and could be used rapidly and accurately for duck circovirus pathogen detection.

The objective was to select green and safe feed additive for improving egg quality, 756 Roman layers with 50-week laying age were divided into 7 groups. The treatment group 1 was the control group, fed with the basal diet; the treat-ment group 2 was fed with 0.04% Chinese herbal medicine in the basal diet; the treatment group 3 was fed with 0.04% Chinese herbal medicine and 0.01% enzyme preparation in the basal diet; the treatment group 4 was fed with 0.04% Chinese herbal medicine, 0.01% enzyme preparation and 0.1% anti-bacterial peptide in the basal diet; the treatment group 5 was fed with 2‰ probiotics in the basal diet; the treatment group 6 was fed with 2‰ probiotics and 0.01% enzyme preparation in the basal diet; the treatment group 7 was fed with 2‰ probiotics and 0.01% enzyme preparation, 0.1% anti-bacterial peptide in the basal diet. The results showed that compared with the control group, the treatment group 4 was the best, and laying rate increased significantly (P<0.05), feed-egg ratio decreased significantly (P<0.05), the Haugh unit increased significantly (P<0.01). Furthermore, the treatment group 4 of the content of total amino acids was the highest, but cholesterol was the lowest. Based on the study, a combination of Chinese herbal medicine, enzyme preparation and anti-bacterial peptide was recommended for improving of egg quality.

This experiment was conducted to study the effects of the proportion of Lys and Met (Lys/Met) with 18% protein level on development of complex stomach in early-weaned Tibetan lambs. A total of 42 early-weaned Tibetan lambs with (55±2) days and (10.85±0.65) kg were randomly allocated into 3 groups with 14 lambs(half males and half females) in each group, early-weaned Tibetan lambs in each group were fed basal diets which Lys/Met was different, the experiment lasted for 20 days. The measured indicators included the weight and morphology of stomach. The results showed as follows, when the Lys/Met was 3.1∶1, the gain of rumen and complex stomach development was best, growth of rumen and reticulum papillae, thickening of stratum corneum, mucosa epithelium, central muscularis in omasum and thickening of mucosa in fundic gland of abomasums were the best. It was concluded that the nutrition levels of diets could influence the growth and development of complex stomach, the suitable ratio of Lys/Met could efficiently promote the growth and development of complex stomach, 3.1∶1 of the ratio Lys/Met was the best ratio under the 18% protein level.

The aim was to study the effect of four feed shapes at the same crude protein content of 13% on the production performance of breeding pigeons. 144 cock and 144 hen breeding pigeons were randomly divided into 4 groups with 36 replicates in each group and one cock and one hen in each replicate. Group Ⅰ was fed cylindric pellet feed, Group Ⅱ was fed globular pellet feed, Group Ⅲ was fed 50% original-shape and 50% cylindric pellet feed, Group Ⅳ was fed original-shape pellet feed. It took 14 days to prepare the test, and the real experiment lasted 106 days. The results showed that in the whole experiment period, the average feed intake (AFI) for breeding pigeons of groups Ⅰ and Ⅱ were significantly lower than the other groups (P<0.01);during 15 to 21 days old, the group Ⅲ of the AFI was also significantly lower than group Ⅳ(P<0.01). Both groups Ⅰ and Ⅱ could increase the body weight of breeding pigeons during reproductive period (P<0.01), and the growing number in cock was more than in hens. Groups Ⅱ and Ⅲ could increase the body weight of squabs in 7 days old (P<0.01), and the influence might disappear when squabs was 14 days old. Egg fertilization rate of group Ⅰ was significantly higher than that of group Ⅳ(P<0.01),and was higher than that of group Ⅱ(P<0.05). Feed shapes had effects on the production performance of breeding pigeons, but no effect on the pecked feather phenomenon, and cylindric pellet feed had the best positive effect.

This experiment was conducted to study the effects of probiotics on growth performance, nutrient digestibility and nitrogen balance of growing minks. Ninety six healthy minks at the age of 60-day-old were randomly divided into 4 groups with 24 replicates (12 males and 12 females) in each group. The control group of minks was offered a basal diet, the group 1 to 3 were fed with a basal diet supplemented with 0.06 g Bacillus licheniformis and 0.06 g Bacillus subtilis, 12 mL Lactobacillus and 6 g yeast, and 0.06 g Bacillus licheniformis, 0.06 g Bacillus subtilis, 12 mL Lactobacillus and 6 g yeast, respectively. The trials of feeding and metabolism were used to analyze the average daily feed intake, average daily gain, feed to gain ratio (F/G), digestibility of crude protein, digestibility of crude fat, nitrogen deposition, net protein utilization and protein biological value. The results showed that compared with the control group, dry matter digestibility of group 1 and crude fat digestibility of group 2 decreased significantly (P<0.05), but the growth performance and nitrogen balance of minks, as well as nutrient digestibility of female minks had no significant difference (P>0.05). In conclusion, supplementation of probiotics in diet could not significantly improve growth performance, nutrient digestibility and nitrogen balance of growing minks. There was no need to add probiotics in production of healthy growing minks.

96 24 to 26 day old "L譟" weaned piglets (average bodyweight with 6.84 kg?0.16 kg) were used to study the effect of different form zinc oxide on the growth performance and diarrhea for 14 days feeding trial. The piglets were randomly allocated for four groups consisting of 3 replicates of 8 piglets each. The control group was fed the basal diet, the groups 1, 2, 3 were fed the basal diet adding 2800 g/t zinc oxide, 800 g/t fat-coated zinc oxide, 500 g/t nanometer zinc oxide respectively. The results showed that the average daily gain raised 15.26% (P<0.05), 9.88% (P<0.05) and 14.69 (P<0.05),the ratio of feed to gain reduced 9.59% (P<0.05), 7.53% (P<0.05) and 10.27% (P<0.05), the diarrhea ratio reduced 66.67% (P<0.05), 55.55% (P<0.05) and 55.55% (P<0.05) respectively compared with the control group, but the groups 1, 2, 3 were no significant differences in average daily gain, ratio of feed to gain and diarrhea ratio (P>0.05) among the zinc oxide, fat-coated zinc oxide and nanometer zinc oxide. It implied that the fat-coated and nanometer zinc oxide were a practicable alternative to high dosage zinc oxide.

Using probiotics replaced antibiotic for the treatment and prevention of animal in the future, and lactic acid bacteria (LAB) were the application of probiotics in the main component. Test was aimed to screen LAB which had high bacteriostatic activity to Campylobacter and tolerance animal gastrointestinal environment. Poultry excrement were acquired from Guangdong different areas, the isolated LAB were dealed with biochemical identification, PCR identification and 16S rDNA sequencing. Meanwhile, we tried to study the antibacterial ability of them against Campylobacter jejuni isolated from chicken, duck and Campylobacter coli from goose, then we further examed the bearing capability of the LAB under acid, bile salts and trypsin. The results showed that Lactobacillus saliva R3 had high bacteriostatic activity and tolerance ability to different birds source Campylobacter. So, Lactobacillus saliva R3 would be one of candidate strains that used as microecologics to treat poultry Campylobacter disease.

In order to study the culture environment of spermatogenesis in vitro, serum-free testicular tissue culture was made. The adult bovine testicular tissue pieces were placed in culture dishes, and movement and morphological changes of testicular cells were observed in different time. Sertoli cells were identified with secretary function by oil red O staining, and spermatogonial stem cells with colony-like growth were identified by AP and immunohistochemical staining. A variety of cell types were found in the culture system, including spermatogonial stem cells of AP, Oct-4 and GFRα1, sertoli cells with secretary function, and a large number of mature sperms. The results showed that the serum-free culture system could represent a model for investigating the effection of biological and chemical factors on the male reproductive system, and be used as material for the study of spermatogenesis and transplantation.

This assay was aimed to determine the blood physiological and biochemical parameters in the mini-pigs bred by China Agricultural University which was named Chinese experimental mini-pigs,so as to provide the basis for mini-pigs as human disease models.Nine physiological blood paramaters and twenty two biochemical blood paramaters in Chinese experimental mini-pigs aged 1,3,6 and 12 months were measured using the automatic biochemical analyzer.The blood parameters between males and females in the same month were compared. The results showed among the blood physiological and biochemical parameters,only RDW and CREA showed significant difference between male and female pigs in the same month in Chinese experimental mini-pigs (P<0.05). Some blood parameters in Chinese experimental mini-pigs were similar to those in Bama mini-pig, Guizhou mini-pig and Wuzhishan mini-pig.The blood physiological and biochemical parameters in Chinese experimental mini-pig were very stable.Compared with human beings,four of nine physiological blood paramaters and fifteen of twenty-two biochemical blood paramaters in Chinese experimental mini-pigs were similar.

With the promotion of the living standard life and the outbroke of malignant disease, including tumor, cancer, drug susceptibility, immunodeficient disease, viral infection etc, the test of detecting cytoactive and immunity were required rapidly, simply and accuratly. The article reviewed the method of detecting the level of sample,referring lymphocyte transformation, ELISA, cytotoxic T cell, IL-12 etc, though synthesizing the study of detecting cytoactive and immunity in the near past year.

Genetic diversity for Pangzhihua Black goat (PZH) and other four black goat breeds(populations),including Janchang Black goat (JC),Yunling Black goat (YL),Jintang Black goat (JT), Lezhi Black goat (LZ). We reestimated using fourteen microsatellite makers by PCR, polyacrylamide gel electrophoresis, and silver staining of modified Sanguintti method. The results showed that all of the microsatellites were highly polymorphic loci,the polymorphic information content (PIC) varied from 0.5129 to 0.7646. Each of the five breeds(populations) possesses had a high level of genetic variability, the PIC and the mean heterozygosity (H) varied from 0.6405 to 0.6856 and 0.6985 to 0.7371,respectively. But the Wright’s F-statistics of subpopulation within total (Fst) showed a low mean value 0.0296,suggesting the variation between populations was only 2.96% share and gene flow was exist among groups. Nei’s genetic distances (D) were calculated, which varied from 0.0365 (between JT and LZ) to 0.1286(between PZH and JT). In the NJ cluster analysis, PZH, JC and YL could be separated to one cluster, JT and LZ to the other. Additionally, the result displayed a remarkable degree of consistency with their different geographical origins.

To explore the possibility of transgenic animals by testicular injection, the Xuhuai goat peroxisome proliferator-activated receptor expression vector pEGFP-PPARγ was using lipofection method, then the linearized recombinant plasmid pEGFP-PPARγ by random dot injected into sexually mature Hu sheep testis. Conventional PCR, genomic DNA dot blotting and Western blotting detection the ability of exogenous PPARγ gene in Hu sheep stable integration. Detection positive individuals and negative individuals of F1 generation of performance of slaughter,the intramuscular fat content, organ weight and meat quality. Positive rate of transgenic sheep was to 13.7%, initially confirmed that the foreign gene PPARγ could be integrated into the genome of Hu sheep and inherited to the next generation. Carcass weight and slaughter rate of positive individuals were significantly lower (P<0.01) than negative individuals in F1 Generation, The proportion of flesh of positive individuals was significantly higher (P<0.01) than negative individuals in F1 Generation;the fat content of longissimus muscle of positive individuals was higher (P<0.05)than negative individuals, the fat content of psoas and biceps femoris were no significant difference (P>0.05);positive individuals lung, small intestine were significantly higher (P<0.01) than negative individuals, positive individuals spleen was higher (P<0.05) than negative individuals, positive individuals heart, lung were lower (P<0.05) than negative individuals, liver, kidney, stomach, testis, skin, and head were no significant difference (P>0.05);positive individuals was lower (P<0.05) than negative individuals, no significant difference between the pH, meat color, water loss rate.

We used immunohistochemical methods to investigate the expression pattern of vascular endothelial growth factor (VEGF) in hair follicle of Liaoning cashmere goat in fetal period, and detected the expression of VEGF in the different fetal period and Microvessel density (MVD). The research results showed that VEGF expressed in hair papilla and hair follicle inner root sheath in Liaoning cashmere goat of each fetal period after 75 days. At the same time, the average optical density of VEGF-positive area also increased and skin MVD showed a growing trend, and the average optical density and MVD were strong positive correlation (r=0.966,P=0.007).

The objectives of this study were to investigate differences in semen quality and development of embryos after in vitro fertilization with frozen-thawed sperm from either Holstein or Simmental bulls. Progressive motility, plasma membrane integrity and acrosome integrity rates of of frozen-thawed sperm from Holstein and Simmental bulls were assessed using visual observation, the hypoosmotic swelling test and Coomassie Blue staining method, and compared the cleavage and blastocyst rates after IVM/IVF with sperm of Holstein and Simmental bulls. The results showed that progressive motility (30.4% and 27.2%),plasma membrane integrity (41.96% and 36.22%) and acrosome integrity rates (77.02% and 73.02%) of frozen-thawed sperm from Holstein and Simmental bulls were all no significant differences (P>0.05), the cleavage (57.5% and 48.6%) and blastocyst rates (30.3% and 23.2%) after IVM/IVF with sperm of Holstein bulls resulted in significant improvement over sperm of Simmental bulls (P<0.05). It was concluded that there were significant differences on embryo developmental competence of in vitro fertilization using different bull semen.

The 36 subclinical mastitis dairy cows of 600 kg weight and 1 to 3 parity were selected for the experiment,and randomly distributed equally into control group and test group. The test group dairy cows were taking 200 g of extract of saxifrage daily. Results showed that enzyme activity of myeloperoxidase enzyme (MPO), lactate dehydrogenase (LDH), lactoperoxidase (LP) and N-acetyl-B-amino-glucosidase (NAG) in the test group 7 and 14 d were significantly lower than the control group(P<0.01); CMT assay showed that the cure rates of test group 7 and 14 d were 55.56% and 77.78%, respectively; drug safety evaluation showed that using the extract of saxifrage had not cause changes in liver and kidney function indicators in test group 7 and 14 d, it indicated that the extract of saxifrage could be used in veterinary clinic safely.

In order to study the effect of boron on micro structure of lymph nodes and retina in rats, 60 weaned clean level SD rats (21 d±2 d) were randomly divided into the control group and experimentalⅠ, Ⅱ, Ⅲ, Ⅳ, Ⅴ groups after adaptive feeding for 1 week, which were respectively added 0, 40, 80, 160, 320, 640 mg/L boron by drinking water. The experiment lasted for 60 d. The results showed that compared with the control group, the quantity and volume of lymphoid nodules were increased with intensive lymphocytes in lymph node cortex, and paracortexes and medullary cord became thick in the experimental Ⅰ and Ⅱ groups, the quantity of cells increased in the ganglion cell layer, inner nuclear layer and outer nuclear layer of well-developed retina. The quantity and volume of lymphoid nodules were decreased in the lymph node cortex, and paracortexes were thinner with loose arrangement of lymphocytes in the experimental Ⅲ group, the retina had minor injury, and the inter nuclear layer became thin, the ganglion cells decreased and loosed. The lymphoid nodules in the lymph node cortex significantly decreased, and germinal center was not obvious or even disappeared, in which the lymphocytes significantly decreased, the paracortexes and the medullary cord became thin, the ganglion cell layer was thinner, and the ganglion cells were decreased with vacuolated cytoplasm, the cells were loosely arranged in the inter nuclear layer and the outer nuclear layer. Above results indicated that addition 40 to 80 mg/L boron promoted the development of lymph node and retina in the rats to some extent, and addition 320 to 640 mg/L boron had obviously damaged effect on the structure of lymph node and retina in the rats.

It was reported that the toxicological safety assessment of a CFTR inhibitor.It was about thiazlidone-like channel blocker drug which synthesized in laboratory, and used in the treatment of animals diarrhea.Drug was given to mice by intragastric administration.Toxicological assessment test of thiazlidone-like channel blocker drug inclued systemic toxicity test,mouse sperm malformation test,bone marrow micronucleus test and Ames test. Results showed that the drug had no acute systemic toxicity, no mutagenic action on germ cells in vivo, no damage on chromosomes and cell mitosis and no mutagenicity. Results provided reliable test basis to use the drug further which could be used to treat diarrhea of piglets.

According to pertinent literature, we chose 114 kinds of Chinese material medcine including heat-clearing drugs, dampness-eliminating drugs, blood-regulating drugs and others to examine their antibacterial activity against APEC through bacteriostasis test in vitro. 6 kinds of Chinese material medcine had good antibacterial activity, such as aloe, andrographitis, garlic, calyx seu fructus physalis, fructus mume and granati cortex. Their inhibition zones were respectively 15, 12, 12, 11, 11 and 7 mm. Then, we isolated and purified the antibacterial components of aloe by silica gel column chromatography, gel column chromatography and ODS column chromatography. At last, the antibacterial component was identified as fumaric acid through MS, IR and NMR analysis. We further studied the activity of fumaric acid against Escherichia coli, Staphylococcus aureus, Streptococcus and Salmonella. The MIC was respectively 50, 50, 780 and 780 μg/mL and MBC was respectively 100, 100, 1560 and 780 μg/mL.

The objective of this study was to investigate the mechanism of taking fertility-promoting power to treat persistent curpus luteum in dairy cows. Dairy cows of persistent corpus luteum were diagnosed by rectal palpation and B-mode ultrasound scanning. These cows were determined 24 h after taking medicine and some morphological indexs included diameter of corpus luteum, follicular number, length of ovary, width of ovary, longitudinal diameter of cervix uterus and longitudinal diameter of horn uterus were counted every three days by ultrasound scanning. The effective number was 25 and effective ratio was 83.3% among 30 dairy cows of persistent corpus luteum. The change of morphological indexs of ovarian and uterine before and after taking medicine were as follow. Medium and small follicules were appeared on ovary after finishing treatment for 1 day and most of large follicule were appeared after finishing treatment for 10, 19 days respectively and part of them appeared estrus and ovulation. The length of ovary on right and left side decreased not significantly (P>0.05) after finishing treatment compared with that of dairy cows before taking medicine, respectively.The width of ovary on left and right side decreased not significantly (P>0.05) after finishing treatment compared to that of dairy cows before taking medicine, respectively. The longitudinal diameter of cervix uterus and horn uterus increased not significant difference after finishing treatment (P>0.05). The results showed that B-mode ultrasound scanning was a effective method to diagnose persistent corpus luteum and taking orally fertility-promoting power had obvious effect on morphological indexs of ovaries and uterine of in dairy cows.

It is of great significance to diagnose the foot-and-mouth disease early, which is a highly contagious disease. Molecular biological detection methods with many advantages, such as the Real-time fluorescent PCR, the multiple PCR, the nucleic acid hybridization, the gene chip, the loop-mediated isothermal amplification and the nucleic acid sequence-based amplification,have been widely used in the diagnosis and classification. The analysis and comparison about the detection efficiency, sensitivity, advantages and disadvantages as well as the research status at home and abroad were reviewed in this study and provided the reference for the molecular biology detection of foot and mouth disease virus applied to in-depth development.

Porcine reproductive and respiratory syndrome (PRRS) is a highly contagious disease which is caused by porcine reproductive and respiratory syndrome virus (PRRSV),characterized by sow reproductive disorders and weaned piglets respiratory diseases,it has a serious influence to the development of the pig industry, so the research of this disease has important meaning.In recent years, the popular and distribution of the disease are increasing widely in the world, and causing a serious economic loss to the pig industry.So the disease becomes a hotspot issue in current researches, has made important progress on the diagnosis and control methods of disease. In this review,the present advancement in the study on PRRSV was summarized,including the diagnostic method and vaccine of porcine reproductive and respiratory syndrome.

In order to kown the safety of sangxing pingchuan granules and provide data support for the clinical medication, 80 Wistar rats were randomly divided into four groups (one control group and three treatment groups), and each group had 20 rats. The control group was taken normal saline by intragastric administration. The three treatment groups were grvaged with water solutions of sangxing pingchuan granules at doses of 30, 15, 3 g(crude drug) (kg稡W), respectively. The drug was fed for 30 days continuously, and changes of body weight were recorded. The blood was collected every nine days for the analysis on some blood routine indices.The heart, liver, spleen, lung and kidney were taken and weighed to calculate organic indexes, then made intotissue sections for histopathological analysis. The results showed that sangxing pingchuan granules affected less on rats. So it was a low toxic and safe traditional Chinese medicine preparation which was suitable for clinical application.

A strain of bacteria was isolated from the attacked tilapia. The pathogen was studied after isolation and purification. A drug sensitivity test showed that the pathogen was sensitive to 10 drugs containing chloramphenicol.The cfb gene was cloned and sequenced.The biological characteristics of the pathogen were studied.The test included many aspects, such as effect of cultivating time and pH on the growth, effect of cultivating temperature on the virulent,the relationships between disease outbreak rate and attacking manners, susceptibility of different specifications(100 and 10 g)fish to the bacteria. The results indicated that the pathogen was Streptococcus agalactiae of tilapia, and the best time and pH for cultivating was 11.5 h and 7.5, respectively. The onset of three injection methods (peritoneal, pleural and back muscle) was the same, but the slowest death rate was the one injected by pleural. Back muscle injection was safer to youngfish. The fish was attacked fastest and had the most centralized death rate when cultivating temperature was 33℃. The 100 g fish was easier to be attacked than the 10 g fish.

A dog with inward curvature of hind limbs was studied,it was diagnosed as inward dislocation of patella by clinical examination and X-ray examination. An operation was performed by overlaping the outside retinaculum,thus the dog got recovered. Diagnosis and treatment of canine patellar dislocation were discussed and recommendations were given in this artical.

FloR was one of the main specific drug resistance genes to florfenicol.There was a paucity of data for the detection of floR in Escherichia coli isolated from pet dogs.The survey of floR gene by PCR and MICs of 10 common antibiotics by micro-broth dilution were performed in this study.The results indicated that among 20 Escherichia coli isolates, the resistant rate to florfenicol was 65% there were 9 and the floR-positive strains, positive rate was 45%. It could be concluded that the resistance rate of florfenicol for pet Escherichia coli isolates would appear continually increasing along with this antibiotic extensive use in animals clinical diseases.So it was very necessary to supervise florfenicol resistance among Escherichia coli isolates from pet dogs.

Feline cowpox virus infections is a feline viral disease caused by cowpox virus, it cause feline skin packing,oral ulcer,systemic disease and necrotizing pneumonia,and aslo can cause human skin lesion and lymph scorching. The essay reviews the progress of feline cowpox virus infections in etiology,epidemiology,clinical symptoms,laboratory diagnose and prevention, so as to provide a material for further study and prevention the feline viral rhinotracheitis.

Duck Colibacillosis was the general term of the multitype diseases caused by pathogenic E.coli. There were numerous clinical disease types, of which the most serious harm for the ducklings,the disease was sepsis and egg yolk peritonitis (egg plague) in duck industry. Since March 2012 in some large-scale duckery of Panjin city, more than 2000 Cherry Valley ducks with 186-day-old were infected with the symptoms mainly for egg production rate dropping, spirit fatigue, loss of appetite, diarrhea and the sick duck abdominal swelling, drooping, the fatality rate was about 4% in the later period. Through clinical observation, pathological examination and laboratory tests confirmed this duck E.coli disease. Based on drug susceptibility testing results, after selection of extremely sensitive to amikacin treatment and comprehensive measures, the disease was quickly controlled.

For understanding the prevalent situation of swine disease in certain regions in Guangxi, we detected CSFV, PRRSV, PCV2, PRV, PPV, JEV and BVDV from 26 samples by RT-PCR or PCR in the investigation.The results showed that the swine in investigated regions were mixture infection, infected by three or more viruses in most cases, and the situation was complicated.

A cattle fallen ill and died after an operation on the rumen fistula, the clinical symptoms mainly presemted tetanic spasm. In this experiment, Clostridium tetanus was isolated from the muscle of surgical wound. The isolated Clostridium tetanus CT-JX-1 is gram-positive bacteria with bacillus on the top, forming the gray, flat and rough-type colonies on a general agar medium, it was dense on central part of the colony, and the outer edge like a spider web, this strain could liquefy the gelatin, but could not ferment a variety of sugars.

To isolate pseudorabies virus (PRV), the morbid materials were collected from some swine farm of Liaoning province and inoculated on BHK-21 cell after disposal.After blind transfer of culture,two portions showed typical cytopathic effect(CPE). TCID₅₀ of virus was 10^-7.8/mL.By the indirect immunofluorescence experiment,the yellow-green fluorescence occurred in the cell cytoblastema.The virus appeared typical porcine pseudorabies virus form, diameter was about 140 to 200 nm,circle with peolos and peolomer. It was inactivated by chloroform, ether,acid and heat. The cell cytoblastema which showed CPE were tested by RT-PCR, and the result was postive. The experimental animals appeared obvious clinical symptom and dead after inoculation.The results showed that PRV was successfully isolated from Liaoning province.

Dairy winter dysentery is a common acute diarrheal infectious disease in the worldwide. As the main feature,affected individuals excrete brown loose stools with blood and dysentry. It usually has a sudden onset,and spreads fast,can significantly reduce dairy milk production,cause serious economic losses to the farmer. We should strictly prevent the disease,once it breaks out suddenly,detect it early and have an appropriate treatment,cut off the spreading ways of the pathogen to prevent infection of the others. At the same time,we should strengthen the comprehensive management of the dairy herd.

In this study, an ELISA method for determination of chloramphenicol (CAP) residue in animal products was used. According to the instruction of ELISA kit, CAP from pretreatment samples which was competed with CAP-HRP, and could be combined antibody coated. The content of CAP could be measured by color produced by substrate solution and HRP. The limit of the detection method in animal products was less than 0.1 μg/kg, and recoveries ranged from 74.4% to 105.4% for fortified samples at levels of 20 to 300 ng/kg with coefficient of variation values less than 15%.The ELISA kit could meet the requirement of detecting CAP residue in animal products.

Recently, the technology of cell suspension culture has received great attentions. It has been widely used in the research and production of many kinds of biological and veterinary vaccines. The quality and cost of veterinary vaccines can both be optimized by this kind of large scale cell culture. The large scale cell culture technology platform based on bioreactor technology has been built up gradually and developed, and it is becoming the main driving force for the development of veterinary vaccines industry. This paper provides a detailed investigation of the technology of cell suspension culture and makes a comprehensive review on the application of this technology in the field of veterinary vaccine production.

Somatic cell count (SCC) is a test parameter for bovine mastitis and milk quality. In order to understand the correlation between SCC and some milk components in one buffalo dairy farm in Nanning area, 152 fresh milk samples at early lactation stage were randomly chosen and SCC, specific gravity(SP), protein, total solid and nonfat solid, lactose levels of milk were determined, correlation among them was analyzed. The results showed that 88.16% of samples had SCC less than 50×10⁴/mL, 1.97% of samples had SCC more than 10×10⁵/mL. SCC had an increasing trend with milk fat, but SCC had an increasing trend with the decrease of milk SP, milk protein, lactose, and nonfat solid level. There was a extremly significant negative correlation between SCC and lactose, between SCC and nonfat solid (P<0.01). In conclusion, SCC was not only significantly related to the milk lactose and nonfat solid level, but also associated with metabolic status of dairy buffalo cows postpartum.

Haemagglutination (HA) test and haemagglutination inhibition (HI) test are widely-used laboratory methods for determination of antibody levels of Newcastle disease and avian influenza. The method is with lots of advantages such as simple material, easy operating,result decided quickly.It is of significance in protecting the health and reducing culture industry of animals, but the method is affected by many factors,which would lead to the wrong test results.This paper comprehensively analyzes the influence of factors affecting the HA test and HI test,in order to avoid and reduce the effect of disadvantageous fators and enhance the accuracy of the test.

We used flat crossed separation method to get a bacterial Y-1, which was isolated from a cow rumen fluid. Simulating rumen environment in vitro to proliferate Y-1. The test adopted the methods, which include morphological observation, biochemical reactions and 16S rDNA gene homology comparison, and constructed the phylogenetic tree for Y-1.The results indicated that Y-1 was Paenibacillus macerans strain, and analyzed the reasons for the presence of bacteria in the rumen fluid.