This study was aimed to clone, bioinformatics and differential expression analyze of AIDA gene in Red Steppe cattle, and construct a eukaryotic expression vector to explore the effects of AIDA genes on the differentiation of bovine preadipocytes at the cellular level.The AIDA gene coding region was amplified from the adipose tissue of Red Steppe cattle by RT-PCR.The nucleotide and amino acid sequences were analyzed by bioinformatics after sequencing.The mRNA expression levels of AIDA gene in 9 tissue of Red Steppe cattle (heart, liver, spleen, lung, kidney, stomach, intestine, muscle and fat) and expression patterns of adipogenic differentiation of preadipocytes were detected by Real-time PCR;The eukaryotic expression vector pBI-CMV3-AIDA was constructed, and transfected into preadipocytes of Red Steppe cattle.The expression of AIDA gene mRNA was detected by Real-time PCR.The results showed that the sequence of AIDA gene CDS was 921 bp, encoding 306 amino acids, containing 4 potential glycosylation sites and 29 potential phosphorylation sites.The subcellular localization was mainly distributed in the cytoplasm, nucleus and mitochondria.AIDA gene was expressed in all 9 tissues of Red Steppe cattle, and the expression was the highest in kidney, which was significantly higher than that in other tissues (P<0.05).The adipogenic differentiation results showed that the expression of AIDA gene mRNA reached the highest on the second day of differentiation, and the expression of AIDA gene gradually decreased with the maturation of adipocytes.The results of double digestion and sequencing showed that the eukaryotic expression vector pBI-CMV3-AIDA of AIDA gene was successfully constructed, and the expression of AIDA gene mRNA in overexpression group was extremely significantly higher than control group (P<0.01).In this experiment, AIDA eukaryotic expression vector was successfully constructed and highly expressed in bovine preadipocytes, which provided a basis for in vitro study of the regulation mechanism of bovine AIDA gene on fat anabolism and metabolism.

This study was aimed to analyze whey proteins identified by ultracentrifugation and acid precipitation, and compare the effects of different sample preparation methods on the identification results of whey proteome.Milk samples were collected to remove fat.Whey protein was prepared by ultracentrifugation and acid precipitation, the difference between two methods was compared by LC-MS and database search, the biological processes, cell composition and molecular functions of different proteins were analyzed.Cluster analysis and principal component analysis showed that 53 proteins with increased expression levels were identified by ultracentrifugation, including β-casein, glycosylation-dependent cell adhesion molecule 1 and polyimmunoglobulin receptors;21 proteins with increased expression levels were identified by acid precipitation method, including CD9 antigen, β2-microglobulin and β-lactoglobulin.According to annotation classification, proteins highly expressed in ultracentrifugation were mainly located in extracellular regions, vesicles, intimal system, endoplasmic reticulum and Golgi apparatus, involving protein binding, enzyme-modulating activity, receptor binding and resistance;Proteins highly expressed in acid precipitation were also mainly located in extracellular regions and vesicles, involving binding, nuclease activity, and ionic binding.Based on the above analysis results, the proteome profiles identified by two methods were different, and the combination of two methods could obtain more identified proteins.This study could provide a basis for the selection of sample preparation methods in whey proteomics research.

This study was aimed to establish a simple, rapid and efficient method for the diagnosis of canine adenovirus type 1 (CAV-1), canine adenovirus type 2 (CAV-2), canine coronavirus (CCV), canine distemper virus (CDV) and canine parvovirus (CPV).5 canine diarrhea viruses CAV-1, CAV-2, CCV, CDV and CPV were used as target viruses.The primers were designed in the conserved region according to the viral gene sequences included in NCBI.Then 2 to 3 probes for each virus were designed according to the variation region.After optimizing the reaction conditions of the detection system and detecting the specificity and sensitivity of the detection method, the gene chip detection method which could simultaneously distinguish 5 viruses was established.The results showed that the chip was able to simultaneously detect the above five kinds of canine diarrhea viruses.The optimized reaction system in this study was that the annealing temperature of PCR was 55℃, the extension time was 1 min 15 s, the hybridization temperature was 40℃, and the hybridization time was 2.5 h.The detection limit of 5 virus standard were as follows:CAV-1 0.2 fg/μL, CAV-2 2 fg/μL, CCV 2 fg/μL, CDV 20 pg/μL and CPV 0.02 fg/μL.The specificity results showed that no positive signals were found for the CPIV.The detection of 14 clinical diarrhea samples showed that the positive detection rates of CAV-1, CAV-2, CCV, CDV and CPV by the gene chip method were 28.57%, 50.00%, 64.28%, 14.28% and 85.71%, respectively, and the sensitivity of the microarray assay was 10 to 100 times higher than that of PCR.These results indicated that the developed method in this study was specific and sensitive, and had diagnostic meaning for the clinical mixed infection of canine viruses.

To identify the impact of sub-lethal dose of imidacloprid on honeybee metabolism during growth stage, the metabonomics analysis was conducted for one-day old worker bee based on liquid chromatography-mass spectrometry (LC-MS).One-day old Apis mellifera ligustica were fed with 0.00001 μg/μL sub-lethal dose of imidacloprid until defecation by the artificial indoor feeding technique.The eclosion rate, body weight at birth, out of the hive time, the activities of SOD, CAT and POD in bee hemolymph were analyzed respectively, and synchronously, differential metabolites were screened out through pattern recognition analysis by means of LC-MS.The results indicated that the out of hive time was significantly prolonged (P<0.05) and the activities of SOD and POD in bee hemolymph were significantly increased (P<0.05) with the treatment of sub-lethal dose of imidacloprid.Principal component analysis (PCA) and supervised partial leastsquares-discriminant analysis (PLS-DA) showed that the experimental group and the control group were significantly separated, the 18 different metabolites were identified.The results could provide theoretical guidance for the further research on the metabolic rules of bee metabolin under imidacloprid stress and elucidation of the bee detoxification mechanism.

To understand the biological function of Mycoplasma synoviae (MS) glycerophosphodoester phosphodiesterase (GDPD), specific primers were designed according to the sequence of MS WVU1853 strains in GenBank, the GDPD gene of MS WVU1853 strain was amplified by PCR.On the basis of sequencing and sequence analysis, it was cloned into pET-28a (+) plasmid to construct prokaryotic expression vector pET-GDPD.After transforming the competent cells of Escherichia coli BL21 (DE3), the expression was induced by IPTG.The expressed products were purified and their enzymatic activities were analyzed.Then the polyclonal antibody was prepared, and its immunogenicity was detected by indirect ELISA and Western blotting, and its distribution in MS was analyzed.The results showed that the full length of GDPD gene CDS of MS WVU1853 strain was 726 bp, encoding 242 amino acids, and the molecular weight of the recombinant protein was about 28 ku.Enzymatic activity assay showed that the recombinant protein could catalyze the conversion of p-nitrophenyl disodium phosphate (pNPP) to p-nitrophenol, and the optimum pH and temperature were 9.0 and 37℃, respectively.Pb²⁺ had strong inhibition effect, while Ca²⁺ had strong promotion effect.The results of indirect ELISA and Western blotting showed that MS GDPD had good immunogenicity and could stimulate New Zealand rabbits to produce high levels of antibodies with serum titer as high as 1:160 000.Subcellular localization results showed that MS GDPD was distributed in cell membrane and cytoplasm, but the content of MS GDPD in cell membrane was slightly higher than that in cytoplasm.The results of this study provided some reference for exploring the biological function of MS GDPD.

This study was aimed to clone and analyze the CDS region of SOD1 gene in New Zealand White rabbits.Specific primers were designed according to the SOD1 gene sequence of Oryctolagus cuniculus (accession No.:NM_001082627.2) in GenBank, then the complete CDS region of SOD1 gene in New Zealand White rabbits was amplified by RT-PCR and cloned into T vector.After sequencing and identification, the homology was compared with other species and phylogenetic tree was constructed.SOPMA, SWISS-MODEL, ExPASy, ProtParam and other software and online tools were used to analyze the protein sequence, the subcellular localization and interaction network of SOD1 protein were predicted.The results showed that the CDS region of SOD1 gene in New Zealand White rabbits was 459 bp in length, the contents of A, T, C and G bases were 24.84%, 18.95%, 23.09% and 33.12%, respectively, and the content of GC was higher than that of AT, which coding 153 amino acids.The homologies of SOD1 gene CDS between New Zealand White rabbit and Oryctolagus cuniculus, Cebus apella, Homo sapiens, Pan troglodytes, Callithrix jacchus, Camelus dromedarius, Canis lupus familiaris, Macaca mulatta, Sus scrofa and Mus musculus were 99.8%, 85.2%, 85.0%, 84.7%, 84.1%, 83.9%, 83.9%, 83.7%, 83.2% and 81.3%, respectively.The phylogenetic tree results showed that New Zealand White rabbits and Oryctolagus cuniculus had the closest evolutionary relationship, and clustered into a single branch.The molecular weight of SOD1 protein in New Zealand White rabbits was 15.7 ku, the theoretical isoelectric point (pI) was 5.86, the formula was C₆₇₂H₁₀₈₄N₂₀₀O₂₂₁S₅, the instability index, fat coefficient and average hydrophilic index (GRAVY) were 23.79, 78.89 and -0.276, respectively.SOD1 was a hydrophilic protein, with no signal peptide and transmembrane region.SOD1 protein contained 5 potential phosphorylation modification sites and 3 potential glycosylation sites, and there was a conserved active site at 45-120 amino acids.The secondary structure was mainly random coil, accounting for 53.59%.The tertiary structure modeling showed that the protein was easy to form homologous dimer.Subcellular localization results showed that the cytoplasm, nucleus and mitochondria accounted for 65.2%, 26.1% and 8.7%, respectively.SOD1 protein could interact with SOD2, PRDX1, PRDX3, PRDX4 and DERL1 molecules in vivo.The results provided a reference for further study of the function of SOD1 gene and the relationship between SOD1 gene and related diseases.

DNA sequencing not only helps us better to understand the nature of life, biological differences, different biological evolutions and developmental history of organisms, but also provides direction for recombinant DNA research, and has very important practical value in disease diagnosis and genotyping.Firstly, from the discovery that DNA is a genetic material, we briefly review the entire development process of the first, second and third generation sequencing technologies, their respective advantages and disadvantages, and the main sequencing technologies or platforms in each.Secondly, we focus on the third generation sequencing typical technology developed by PacBio, including single molecule Real time (SMRT), Oxford Nanopore Technologies (ONT), introduce the principles, methods and technical processes, and summarize the application areas of the third generation sequencing technology, including the genomic resequencing, do novo sequencing, transcriptome studies, methylation testing, disease-related structural variation testing and the application of viral quasispecies analysis in epidemiology.Thirdly, the advantages of three generations of sequencing techniques in transcriptome sequencing and epigenetics are compared, the third generation sequencing technology has obvious advantages in research fields, such as genomic repeat regions or structural variations, and very meaningful for the study of various diseases, it is considered to be an important and accurate diagnostic tool in the future.In addition, breeders can artificially select individuals by establishing associations between genes and traits by analyzing the genetic information of important economic species, which greatly shortens the breeding period.Finally, we prospect the future application of the third generation sequencing technology in medicine, agriculture and environment.

To investigate the effect of ubiquitin-related modification protein SUMO-2 on Brucella 16M, RAW264.7 model of murine macrophages with SUMO-2 gene interference and overexpression was constructed and infected with Brucella 16M.Specific interference fragments and overexpression primers were designed according to the sequence of SUMO-2 gene in GenBank.After cloning, they were connected to the corresponding lentivirus vector and transformed into DH5α competent cells of Escherichia coli.HEK-293FT cells were transfected with plasmid extracted from positive clone bacteria, and RAW264.7 mouse macrophages were infected with recombinant lentivirus.The interference and overexpression cell models were successfully constructed by Brucella 16M infection, respectively.The transcription level of SUMO-2 was detected by Real-time PCR, the expression of SUMO-2 protein was detected by Western blotting, the levels of IFN-γ and TNF-α were detected by ELISA, and the viability of Brucella in cells was determined by colony count.The results showed that compared with control group, the level of SUMO-2 gene in interference group was extremely significantly lower (P<0.01), and the level of SUMO-2 gene in the overexpression group was extremely significantly higher (P<0.01).The cell model of SUMO-2 interference and overexpression was successfully constructed.Western blotting results showed that Brucella 16M infection could down-regulate the expression of SUMO-2 protein in a time-dependent manner.After colony counting, the number of Brucella extremely significantly decreased after SUMO-2 overexpression (P<0.01), which inhibited the intracellular reproduction of Brucella 16M.The number of Brucella significantly or extremely significantly increased after SUMO-2 interference (P<0.05;P<0.01), which promoted the intracellular reproduction of Brucella 16M.At the same time, the levels of IFN-γ and TNF-α extremely significantly increased after SUMO-2 overexpression (P<0.01).After SUMO-2 interference, the level of TNF-α significantly decreased (P<0.05), the level of IFN-γ extremely significantly decreased (P<0.01), and the expression of SUMO-2 in RAW264.7 cells also affected the production of IFN-γ and TNF-α.In conclusion, SUMO-2 protein played an important role in the survival of Brucella cells, which might help to elucidate the pathogenic mechanism of Brucella infection.

The aim of this study was to screen the organic solvents with lower cytotoxicity and Toxoplasma gondi toxicity.Vero cells were cultured on 96-well plates, the cells were cultured for 12 hours and then the medium was replaced by 200 μL complete DMEM medium containing 0, 0.1%, 0.3%, 0.5%, 1%, 3%, 5% and 10% (V/V) DMSO, dimethyformamide, ethanol and methanol, respectively, blank control holes were set up at the same time.The toxicity of DMSO, dimethylformamide, methanol and ethanol to Vero cells was analyzed by MTT method.The organic solvents with the least cytotoxicity to host cells were screened and the safe concentration was determined.Toxoplasma gondi was inoculated to Vero cells and cultured for 12 hours.The supernatant was discarded and then 120 μL complete DMEM containing 0, 0.1%, 0.3%, 0.5%, 1% and 3% (V/V) DMSO, dimethyformamide, ethanol and methanol was added, respectively.The impacts of solvents on the proliferation of Toxoplasma gondi were determined 36 hours post inoculation.The number of infected cells was counted and the relative infection rate was calculated.The organic solvents with the least impact on the proliferation of Toxoplasma gondi and the safe concentration were screened out.The results showed that there was no obvious cytotoxicity on Vero cells when the volume fraction of DMSO, dimethylformamide, methanol and ethanol were less than 1%, 0.1%, 3% and 1%, respectively.For Toxoplasma gondi, the safety concentrations of DMSO, dimethylformamide, methanol and ethanol were 1%, 0.1%, 1% and 0.1%, respectively.This results indicated DMSO and methanol were less toxic to Vero cells and Toxoplasma gondii, and could be used as organic solvents for the screening of drugs with poor water solubility against Toxoplasma gondii.

The objective of this study was to analyze the differences of hindgut microbiota composition between heat stress sensitive cows and heat stress tolerant cows under heat stress condition, to find specific microbiota related to heat stress, and to provide the theoretical basis for improving thermotolerance of dairy cows via improving feeding management and genetic selection.Nineteen Chinese Holstein dairy cows which were in 1st lactation and with the similar body conditions were divided into two groups by the difference of morning and afternoon rectal temperature:Heat stress sensitive group (group H, n=10) and heat stress tolerant group (group L, n=9).The sequences of V3 and V4 region for 16S rDNA were sequenced by Illumina PE250 sequencing platform and bioinformatic methods were used to detect the differential microbiota.The results showed that:①The Shannon and Chao1 indexes in group L were higher than that in group H, but there was no significant diffference (P>0.05).②For phylum level, over 95% of the sequences in both groups of cows were assigned to Firmicutes, Bacteroidetes and Proteobacteria.For genus level, the relative abundances of YRC22 and Prevotella were significantly higher than that in group H (P<0.05), and Prevotella and YRC22 were negatively correlated to the heat stress related indexes:Afternoon rectal temperature (ART), rectal temperature difference (RTD), respiratory score (RS), rectal temperature range (RTR) and drooling score (DS).③Among the heat stress related indexes (morning rectal temperature (MRT, ART, RTD, RS, DS, RTR), RS, DS and MRT were highly correlated with the diversity of hindgut microbiota, as R2 values were top three of these six indexes in the RDA results.In conclusion, the abundance of hindgut bacteria were influenced by heat stress.Prevotella could be the hindgut microbiota related to heat stress in dairy cows.

Feeding is the basic physiological process of animal survival, growth and development.Food intake of livestock and poultry directly affects the absorption of nutrients and production performance.In animal production, feeding is affected by many factors, and stress is one of the most important factors.The stress response of animal is mainly regulated by the hypothalamic-pituitary-adrenal (HPA) axis.The hypothalamus, pituitary and adrenal cortex regulate the stress response of the animals by releasing three stress hormones such as corticotropin releasing hormone (CRH), adrenocorticotropic hormone (ACTH) and glucocorticoid (GC).The regulation of feeding by stress hormones is a very complex process where it can regulates feed intake in both positive and negative way.Specifically, stress hormones may participate in both homeostatic and non-homeostatic pathways to regulate eating behavior.Homeostatic pathways refer to the regulation of food intake by sensing energy status of the body.In this case, CRH and ACTH inhibit feed intake by inhibiting the expression of the appetite peptide in the hypothalamus;GC plays a completely opposite role in the central and peripheral regions.The non-homeostatic pathways refer to the regulation of hedonic eating by modulating the midbrain reward system.It is a hotspot in the study of appetite regulation in recent years.More and more studies have focused on the crosstalk between stress hormone and reward system.In this review, the latest researches on regulation of feeding by stress hormone will be summarized.It will provide some theoretic basis for the development of new techniques on feeding regulation in animal production.

The aim of this study was to establish an in vitro infection model of endometritis by inducing endometrial epithelial cells of dairy cows with lipopolysaccharide (LPS) and study the effects of endometritis on the relative expression of uterine receptive factors MUC-1, Wnt-7a, IFNAR1, IFNAR2, Integrin αvβ3 mRNA and protein in dairy cows.The mechanism of repeated sterility and low reproductive rate in dairy cows caused by endometritis were further clarified.Endometrial epithelial cells of dairy cows were isolated and cultured by tissue block method;The purity of the cells was determined by immunofluorescence;Endometrial epithelial cells were stimulated by LPS at different concentrations;The cell survival rate was measured by CCK8 at different time points;The secretion of inflammatory factors interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), IL-6 and IL-8 were detected by ELISA.Real-time PCR and Western blotting were used to detect the mRNA and protein expression of uterine receptive factors MUC-1, Wnt-7a, IFNAR1, IFNAR2, Integrin αvβ3 in endometrial infection model.The results showed that the number of bovine uterine epithelial cells purified by tissue block method was more, and the purity was higher by immunofluorescence keratin staining.CCK8 assay showed that compared with control group, LPS at 0, 5, 10, 50 μg/mL had no significant change in cell viability after 6, 12, 24 and 48 h (P>0.05).When LPS concentration was 100 μg/mL for 24 h, the cell survival rate was significantly lower than that of control group (P<0.05).The contents of IL-1β, TNF-α, IL-6 and IL-8 in cell culture supernatant were significantly higher than those in control and low concentration groups (P<0.05).Compared with control group, the relative expression of MUC-1 mRNA and protein in the model group extremely significantly increased (P<0.01), Wnt-7a mRNA and protein extremely significantly decreased (P<0.01).The mRNA expression of Integrin αvβ3, IFNAR1 and IFNAR2 extremely significantly decreased (P<0.01), and their protein expression were significantly decreased (P<0.05).The results showed that LPS with concentration of 100 μg/mL for 24 h could successfully construct the infection model of endometritis in vitro.The effects of endometritis on the expression of MUC-1, Wnt-7a, IFNAR1, IFNAR2, Integrin αvβ3 mRNA and protein might be the important reasons for repeated sterility and low reproductive rate of dairy cows.

The objective of this study was to investigate the diversity of ruminal Archaea in Nili-Ravi buffaloes and Haizi buffaloes using 16S rDNA library-based analysis.Three female Nili-Ravi buffaloes and three female Haizi buffaloes were used in this study, and ruminal samples were taken by a stomach tube.Total DNA were extracted, and specific primers of Met86F/Met1340R were used to amplify 16S rDNA of methanogens for the construction of library.The results showed that 73 and 100 sequences were obtained from Nili-Ravi buffalos and Haizi buffalos library, respectively.Based the 97% similarity, sequences of Nili-Ravi buffaloes and Haizi buffaloes were assigned to 20 and 33 OTUs, respectively.For Nili-Ravi buffaloes, 70 sequences (17 OTUs) showed ≥ 97% sequence similarity to known species, 3 sequences (3 OTUs) had sequence similarity to known species in the range of 93% to 96%.For Haizi buffaloes, 70 sequences (15 OTUs) showed ≥ 97% sequence similarity to known species, 30 sequences (18 OTUs) had sequence similarity to known species in the range of 84% to 96%.The SGMT and RO clade sequences percentage in Nili-Ravi buffaloes and Haizi buffaloes were 76.71% and 21.91%, 76.00% and 19.00%, respectively.Phylogenetic analysis indicated that these sequences were assigned into two large clades.There were 8 OTUs representative sequences from Nili-Ravi buffaloes and 16 OTUs representative sequences from Haizi buffaloes assigned into the same clade on the top of the phylogenetic tree and far from all known species.It could be concluded that Methanobrevibacter genus was the dominant Archaea in the rumen of Nili-Ravi buffaloes and Haizi buffaloes when fed the same diet.The SGMT clade sequences ratio in Nili-Ravi buffaloes almost were the same with Haizi buffaloes, while the RO clade sequences in Haizi buffaloes were lower than that in Nili-Ravi buffaloes, indicating that there were more unknown methanogen sequences in Haizi buffaloes than that in Nili-Ravi buffaloes.

In order to study the effects of low crude protein (CP) diets supplemented with balanced amino acids on the growth performance, blood indexes, nutrients apparent digestibility, diarrhea rate of 10 to 25 kg weaned piglets, three hundred and twenty three-way cross Duroc× (Landrace×Yorkshire) piglets with body weight of 10 kg were selected and randomly divided into four treatments with four repeats per treatment and twenty pigs per repeat.The control group was fed control diet (20.1% CP), experimental group were fed CP diets supplemented with L-Lys, DL-Met, L-Thr, L-Trp, L-Val, L-Ile, L-Phe and L-His, and the CP levels of experimental diets were 18.3%, 14.9% and 13.2%, respectively.The amino acid levels of each treatment reached or exceeded the standards of nutrient requirements of swine (NRC 1998).The experiment lasted 24 days.The results showed that the ADG of weaned piglets in 20.1% CP groups was significantly higher than that in 13.2% CP treatment (P<0.05).The ADFI and F/G of weaned piglets in 18.3% CP, 14.9% CP and 13.2% CP groups were not significant different compared with 20.1% CP group (P>0.05).②The blood urea content of 20.1% CP group was the highest and that of 13.2% CP group was the lowest, and there was extremely significant difference between the two groups (P<0.01).The levels of TP, ALB, GLB, A/G, Blood glucose, insulin, IgA, IgG and IgM of 18.3% CP, 14.9% CP and 13.2% CP groups were not significant different compared with 20.1% CP group (P>0.05).③The apparent digestibility of CP in 13.2% CP and 14.9% CP groups were extremely significantly lower than that of 20.1% CP group (P<0.01);There were no significant difference of other indexes among the four groups (P>0.05).④The diarrhea rate of weaned piglets in 20.1% CP grou was the highest and that of 13.2% CP group was the lowest, and there was significant difference between the two groups (P<0.05).In conclusion, when the dietary CP level was reduced to 14.9%, the growth performance of piglets was not adversely affected.The CP apparent digestibility and blood urea level was extremely significantly decreased than that of control group (20.1% CP), and the diarrhea rate of piglets was greatly reduced.Therefore, the CP level of diet supplemented with 8 amino acids in 10-25 kg weaned piglets could be reduced to 14.9% without affecting the performance of piglets.

The purpose of this paper was to study the change rule with the seasons of rumen bacterial flora structure and diversity of Mongolian sheep grazed on typical grasslands in Northern China.In the experiment, six Mongolian sheep grazed all year round and no supplementary feeding were randomly selected from the typical grasslands of Northern China.In winter (January), spring (April), summer (June grass growth stage and August grass vigorous stage) and autumn (October), the rumen juice of the sheep was extracted through the mouth, and in every season the structure and diversity of rumen bacterial flora were analyzed by MiSeq sequecing technology.The results showed that there were the highest operational taxonomic unit (OTU) and diversity of the rumen bacteria of grazing Mongolian sheep in summer grass vigorous stage and the lowest in winter, and there were significant differences in OTU and diversity between summer and winter (P<0.05).At the phylum level, Firmicutes (56.93%) and Bacteroidetes (21.52%) were the dominant bacterial populations in the rumen of Mongolian sheep.The OTU of Firmicutes reached its maximum in August and that of Bacteroidetes reached its maximum in June, which were significantly different from other months.At the family level, Ruminococcaceae (19.09%), Lachnospiraceae (16.98%), Christensenellaceae (12.11%), Prevotellaceae (8.51%) and Rikenellaceae (8.27%) were the dominant bacterial populations in the rumen of Mongolian sheep, respectively reaching the peak in summer (June or August).At the generic level, rumen bacteria of the highest contents were respectively Prevotella (4.79%), Butyrivibrio (1.77%) and Ruminococcus (1.27%).So Ruminococcaceae of Firmicutes had the highest number of rumen bacteria of grazing Mongolian sheep in Northern China, which was a typical cellulose-digestive bacterium and in accord with the characteristics of grazing sheep fed with forage grass.

In this experiment, the medium with cellulose as the sole carbon source was used as the growth environment of the rumen bacterial community, and then the 16S rRNA sequence technique was used to analyze the dynamic changes of the rumen bacterial community at different time points.The rumen bacteria were incubated in the media made of anaerobic dilution solution with cellulose for 50 h, and samples were collected at 10, 20, 30, 40 and 50 h, respectively.The DNA samples were extracted and analyzed by 16S rRNA sequencing.The results showed that the Ace of bacteria community was significant different among different time points (P<0.05), and that at 30 h was significantly higher than that of 40 and 50 h (P<0.05).The beta diversity proved that the bacteria community at 40 and 50 h was significant different with that at 10 and 20 h (P<0.05), the bacteria community succession happened in 30 h.The typed rumen cellulolytic bacteria, like Butyrivibrio, Pseudobutyrivibrio, Clostridium, increased with time increasing, but the abundance of Fibrobacter was not significantly changed (P>0.05).With time passing by, the abundances of Acholeplasma, Alkaliphilus, Blautia, GW_34 were increasing and significantly higher at 50 h compared with 10 h (P<0.05) during culture period.In summary, the reference time of enrichment and separation of cellulytic bacteria was 30 to 40 h, and the results provided a theoretical basis for further dilution culture and isolation of new pure cellulytic bacteria.

This experiment was aimed to study the effects of Monascus synbiotic, yeast-selenium-germanium culture and their compound preparation instead of antibiotics on gut structure and rectal flora of weaned piglets.Eighty healthy piglets ((7.57±0.49) kg) were randomly divided into 4 groups with 5 replicates each group and 4 piglets each replicate.The control group was fed basal diet with 25 mg/kg colistin sulfate and 45 mg/kg chlortetracycline and groups Ⅰ, Ⅱ and Ⅲ were fed basic diet with 0.5% Monascus synbiotics, 0.5% Monascus synbiotics and yeast-selenium-germanium culture compound, 0.5% yeast-selenium-germanium culture, respectively.The test period was 42 d with 3 stages.The results showed that:①Compared with the control group, the activities of SOD and GSH-Px, T-AOC of the jejunum mucosa in the groups Ⅰ, Ⅱ and Ⅲ were extremely significantly or significantly increased (P<0.01;P<0.05).Compared with the group Ⅲ, the SOD activity and T-AOC of the jejunum mucosa in group Ⅱ were significantly increased (P<0.05).②Compared with the control group, the villus length and the V/C of the duodenum in the group Ⅰ, Ⅱ and Ⅲ were extremely significantly or significantly increased (P<0.01;P<0.05).And the V/C of jejunum in group Ⅱ was significantly increased (P<0.05).③At 1-14 d, compared with the control group, the number of Salmonella and E.coli in the groups Ⅰ and Ⅱ were significantly decreased (P<0.05).The number of Bifidobacteria and lactic acid bacteria showed a significant or extremely significant increase (P<0.05;P<0.01).At 15-28 d, compared with the control group, the number of lactic acid bacteria in groups Ⅰ, Ⅱ and Ⅲ was extremely significant or significantly increased (P<0.01;P<0.05).In conclusion, compared with antibiotics, Monascus synbiotics or yeast-selenium-germanium culture, their compound preparation could effectively improve the intestinal tissue structure and intestinal flora of weaned pigs.

In order to study the effects of sesamin on the growth performance and the morphology of intestinal tissues of zebrafish, 720 zebrafish were randomly divided into 6 groups:Control group (CK), sesamin Ⅰ group (1 g, S1), sasamin Ⅱ group (2 g, S2), 80 mg/L fluoride group (F), 80 mg/L fluoride+sesamin Ⅰ group (1 g, FS1), and 80 mg/L fluoride+sasamin Ⅱ group (2 g, FS2), with 3 replicates per group and 40 zebrafish per replicate.The body length, body weight, LGR, WGR, and SGR were determined and the intestine structures of zebrafish were observed using the histological method, and the height of intestinal villi were measured using the Imageproplus at 45th and 90th day.The results showed that at 45 d, compared with the control group, the body weight of zebrafish in group F was significantly decreased (P<0.05), and sesamin treatment could alleviate the decrease of body weight, however the difference was not significant (P>0.05).At 90 d, the body weight and body length of zebrafish in group F were remarkably reduced compared with the control group (P<0.05), and sesamin treatment could relieve the negative effects caused by fluoride to some extent, but the difference was not significant (P>0.05).After exposure for 45 and 90 d, the LGR, WGR, and SGR of zebrafish in group F were decreased compared to the control group, while they showed an increasing trend in groups FS1 and FS2 compared with the group F.Sesamin treatment could alleviate the damage caused by fluoride in intestinal tissue of zebrafish according to the tissue section of gut.All above results indicated that sesamin could attenuate the toxic effects of fluoride on growth performance and the intestinal tissues in zebrafish.

β defensins are a class of antimicrobial peptides secreted by pigs.They are widely distributed in tissues and play an important role in resisting pathogenic microorganism and immune regulation.Porcine β defensins have strong inhibitive activity against bacteria, fungi and viruses, and have the advantages of small molecular weight, high heat stability and no residue, making them an ideal antibiotic substitute.Researches have showed that the expression of porcine β defensins positively correlate with disease resistance.It can enhance immunity and promote growth of pigs by increasing the expression of endogenous defensins or ingesting exogenous defensins.Exogenous stimulus such as fatty acids, amino acids, vitamins, probiotics and microorganisms could affect the expression of β defensins.With the deep researches of defenses, they are expected to solve the problems of drug resistance, drug residues and environmental pollution caused by long-term use of antibiotics in pig production.This paper reviews recent advances in porcine β defensins, including structure characteristics, expression specificity and biological function.The effects of exogenous stimuli on the expression of defensins and their mechanisms are mainly reviewed.The preliminary applications of exogenous β defensins as feed additives in piglet production are analyzed in order to lay the foundation for pig β defensins researches and their application in healthy pig breeding.

Sodium butyrate is a new feed additive which is widely used in animal feed, and its effects is reported to be positive in many literatures.This paper mainly introduces the biological function of sodium butyrate, its effect on development of calves' gastrointestinal tract and the effects of being used as calves' feed additive.The authors mainly reviewed the mechanism of sodium butyrate promoting the development of gastrointestinal tract, including being used as energy source of gastrointestinal tract of animal, regulating secretion of hormones and growth factor, improving secretion and activity of digesting and absorption related enzymes, and increasing secretion of protective proteins of intestinal immune cells, etc.The present studies showed that addition of 0.3% sodium butyrate into feed dry matter could promote the development of gastrointestinal tract and growth of calves, but the addition level, phase and way were need to be further researched.Besides, there is few report on the influences of dietary forage level and endogenous butyrate which produced by concentrate fermentation on function of exogenous butyrate, which need to be further researched.

This study was aimed to analyze the genetic diversity of Minxian Black fur sheep, select the ideal genetic markers, and provide a theoretical basis for breeding and conservation.144 Minxian Black fur sheep were selected to collect blood from jugular vein for extracting DNA.24 pairs of microsatellite primers were amplified by PCR, and the genotypes were analyzed by capillary electrophoresis.The numbers of alleles, alleles size and frequency, numbers of effective alleles, heterozygosity and polymorphism information content were calculated.The results showed that a total of 210 alleles were found in 24 loci, the average number of alleles was 8.75;The population alleles frequency were 0.0104 to 0.7396, the alleles size were 97 to 285 bp;The numbers of effective alleles were 1.7581 to 8.2433;The average observed heterozygosity was 0.4839;The average expected heterozygosity was 0.6959;The average polymorphism information content was 0.6527.The numbers of alleles, numbers of effective alleles, expected heterozygosity and polymorphism information content at MAF70 locus were the highest;The observe heterozygosity at OarFCB128 locus were the highest;The numbers of alleles at OarAE129 locus and the observed heterozygosity at OarFCB304 locus were the lowest;The expected heterozygosity and polymorphism information content at SRCRSP9 locus were the lowest.The Hardy-Weinberg equilibrium analysis results showed that 19 loci devieted from Hardy-Weinberg equilibrium.The results indicated that the genetic background of Minxian Black fur sheep was complex and the genetic diversity was rich, which could provide a theoretical basis for the evaluation of genetic resources and the breeding and conservation.

This study was aimed to explore the expression patterns of mRNA and microRNA in donkey testis, and provide genetic data for transcriptome studies of Chinese local donkeys.The mRNA and microRNA expressed in testis of Chinese domestic donkeys were analyzed by RNA-Seq and bioinformatics methods, and the differentially expressed genes and microRNA between Biyang and Dezhou donkeys were identified.The results showed that 57 837 506 (accounted for 96.34%) and 9 952 015 (accounted for 88.86%) clean reads of mRNA and Small RNA were obtained.A total of 24 751 genes and 1 074 microRNAs expressed in testis were identified.The top five expressed genes were LOC106836782, XLOC_066401, HSP90AA1, TNP1 and COF2, the top five expressed microRNAs were eca-miR-143, eca-miR-21, eca-miR-99a, eca-miR-148a and eca-miR-26a.The results revealed 102 differentially expressed genes and 2 microRNAs between Biyang and Dezhou donkeys.However, no significant function term was annotated, which suggested that there was no obvious difference of genetic expression pattern between Biyang and Dezhou donkeys.This study provided gene expression pattern and transcriptome data for donkey testis tissue and identified differentially expressed mRNA and microRNA between Biyang and Dezhou donkeys, which had scientific significance for studying genetic sources of donkeys in the future.

In order to investigate the regulation of corticotropin-releasing hormone (CRH) gene in reproduction of yak, tissue samples of hypothalamus, anterior pituitary, oviduct, ovaries and uterus were collected from 5 adult female yaks and cattle, respectively.The yak CRH gene was amplified, cloned and sequenced by RT-PCR and bioinformatics software, and phylogenetic tree was constructed.The mRNA expression in different tissues was detected by Real-time PCR.The results showed that the coding region of CRH gene in yak was 573 bp, encoding a 190 amino acid protein.The nucleotide homology of CRH gene in yak (Bos grunniens) and cattle (Bos taurus), Felis catus, Gallus gallus, Homo sapiens, Mus musculus, Rattus norvegicus and Sus scrofa was 99.8%, 43.3%, 36.1%, 46.2%, 39.0%, 38.5% and 56.4%, respectively.Phylogenetic tree results showed that yak and cattle had the closest relationship and clustered with other species, which accorded with the evolution law of species.The formula of CRH protein was C₉₂₀H₁₅₀₃N₂₇₉O₂₆₃S₃, the molecular weight and theoretical isoelectric point were 20 776.95 u and 10.95, respectively.CRH protein contained a high proportion of leucine (15.8%), proline (12.6%), alanine (12.1%) and arginine (10.5%).CRH protein was a hydrophilicity protein which contained a signal peptide and a transmembrane region.The secondary structure prediction results showed that alpha helix, beta turn, extended chain and random coil accounted for 41.05%, 1.05%, 8.42% and 49.48%, respectively, which was consistent with the tertiary structure analysis results.The expression of CRH gene mRNA in hypothalamus and uterus of yak was the highest and the lowest, respectively, and the expression of CRH gene mRNA in hypothalamus was significantly higher than that in other tissues (P<0.05).The expression of CRH gene mRNA in anterior pituitary, ovary and fallopian tube of yak were extremely significantly or significantly higher than that of cattle (P<0.01;P<0.05), but there was no significant difference in hypothalamus and uterus between two breeds (P>0.05).These results suggested that CRH gene played an important role in reproductive regulation of yak.

Mastitis is one of the most important diseases affecting the economic benefits of dairy production, and the number of somatic cells count (SCC) is an important indicator reflecting the health of the cow's breasts and the quality of milk.The cleanliness of the upper and lateral legs, breasts, and lower hind legs of 670 Holstein dairy cows from a large-scale pasture were evaluated, and the effects of breast and leg cleanliness on the number of somatic cells in milk were analyzed in this study.SAS 8.1 statistical software was used for least squares analysis of variance and multiple comparisons, and the cleanliness score, group effects, lactation effects and parity effects were uesd as fixed effects.The results showed that:①Group and parity had significant effects on somatic cell scores (P<0.05).②Lactation stage, lower leg cleanliness score and breast cleanliness score had extremely significantly effects on somatic cell scores (P<0.01):The somatic cell scores of the lower leg cleanliness scores of 4 were extremely significant higher than 1 and 2 points (P<0.01), and the somatic scores of the lower leg cleanliness score of 3 were significant higher than 1 point and 2 points (P<0.05);The somatic score of the breast cleanliness score of 3 was significant higher than 1 and 2 points (P<0.01);The upper leg and side cleanliness scores had no effect on somatic cell scores (P>0.05).③SCC had an increasing trend with the increase of the cleanliness score of the lower leg and udder of the cow.In conclusion, the health status of the herd breasts could be effectively understood by scoring the body cleanliness of the herd, which could provide a scientific basis for the daily management of large-scale dairy cows.

To investigate the structure and function of osteopontin (OPN) gene and explore its expression characteristics in bone metabolism of layer during late period of laying.The tibia of Hyline Grey layer at late period of laying was selected for RNA isolation.The specific primers were designed based on OPN gene sequence of Gallus gallus in GenBank (accession No.:NC_006091.5) using Primer Premier 3.0 online software.The coding sequence (CDS) of OPN gene was cloned by RT-PCR and analyzed by bioinformatics.Real-time PCR technology was used to detect the expression of OPN gene mRNA in tibia.The results showed that the full-length of chicken OPN gene CDS was 795 bp, encoding 264 amino acids.The phylogenetic tree constructed by Mega 6.0 software showed that the relationship was the closest between chicken and carp, with 100% similarity.The OPN gene was highly conserved in avian evolution.The bioinformatics analysis suggested that OPN was an unstable and hydrophilic protein with 27 O-glycosylation sites, 1 N-glycosylation site and 34 phosphorylation sites.The subcellular localization of OPN was mainly localized in cytoplasm (44.4%) and mitochondria (33.3%).The secondary structure was mainly composed of random coil and alpha helix.The difference of OPN gene mRNA expression among individuals was not related to bone strength.The results indicated that OPN gene mRNA expression was not related to the bone strength, which provided a reference basis for further study on the mechanism of OPN gene in bone metabolism of layer during late period of laying.

This study was aimed to screen the polymorphism of thymosin beta 4 (Tβ4) gene in Qianbei Ma goat, and analyze the genetic structure by bioinformatic method.Primers were designed according to predicted sequence of Capra hircus Tβ4 gene in GenBank (accession No.:NC_030810 and NW_017189516) by Primer Premier 5.0.The mixed DNA pool and PCR products combined with direct sequencing were used to obtain the polymorphisms of exons 1, 2 and 3 in chromosome X and exon 1 in chromosome 3 of Tβ4 gene.The homology, phylogenetic tree, physicochemical properties, hydrophobicity, coiled coil, transmembrane structure, signal peptide, subcelluar localization, structure domain, phosphorylation site, glycosylation site, secondary structure and tertiary structure of Tβ4 gene in Qianbei Ma goat were analyzed using bioinformatic analysis softwares.The results showed that a total of 10 SNPs (G55C, C82T, G112A, C129T, G216A, G431A, G551T, T579A, A609G and A619G) were screened.The homology analysis results showed that the identity of Tβ4 gene between Qianbei Ma goat and Bos taurus, Equus caballus, Sus scrofa, Homo sapiens, Heterocephalus glaber, Rattus norvegicus, Mus musculus and Gallus gallus were 97.8%, 96.3%, 95.6%, 94.1%, 94.8%, 91.1%, 92.6% and 86.7%, respectively.Protein physical and chemical properties analysis showed that the theoretical isoelectric point of Tβ4 protein was about 5.02, the minimum value of hydrophilicity was -3.011, no hydrophobic amino acid and coiled coil existed, which was not a transmembrane protein and no signal peptide existed, was located in the nucleus (60.9%), mitochondria (4.3%), cytoskeleton (17.4%) and cytoplasm (17.4%).There was 1 THY domain, 1 threonine phosphorylation site, no N-glycosylation site and 4 O-glycosylation sites.The analysis of secondary structure showed that C129T mutation in Qianbei Ma goat made the increase of α-helix and the decrease of random coil.This results indicated that the polymorphism of Tβ4 gene in Qianbei Ma goat was successfully screened, which provided a theoretical foundation for the study of molecular genetic marker-assisted selection in breeding of Qianbei Ma goat.

This study was aimed to investigate the morphological characteristics of fat tail and the effect of the size of fat tail on the body size index after backcrossing between wild Argali sheep and Bashbay sheep.216 breeding backcrossing second generation of wild Argyi sheep×Bashbay sheep with the same feeding and good body conditions were collected to calculate the variation coefficient of fat tail traits, then according to the fat tail traits to study the separation and differentiation of fat tail of the backcrossing second generation by K-mean cluster analysis of R language method, and the differences of 7 body weight indexes of the backcrossing second generation of fat tail sheep (body weight, body length index, chest circumference index, tube circumference index, body index, limb length index and chest index) were analyzed by SPSS 19.0 software.The results showed that the backcrossing second generation of wild Argyi sheep×Bashbay sheep could be divided into four types:Small tail type, medium tail type, big tail type and fat tail type through the cluster analysis results.The body length index of fat tail type was extremely significantly larger than small tail type (P<0.01), the body index of big tail type was extremely significantly larger than small tail type (P<0.01), and the chest circumference index and chest index of big tail and fat tail types were extremely significantly larger than small tail type (P<0.01), but there was no significant difference in body weight, tube circumference index and limb length index in each tail type (P>0.05).This results indicated that the traits of the backcrossing second generation of fat tails were separated, the larger the fat tail, the larger the body size, but the body weight did not change significantly.Combined with the characteristics of appearance and production performance of wild Argali sheep and Bashbay sheep, big tail and fat tail types were closer to Bashbay sheep in the body type, and small tail type was closer to wild Argali sheep in the tail type.

To explore the relationship of the structure variation of WIF1-I8-sv889 in the WIF1 (Wnt inhibitory factor 1) gene and the feature of body hair formation, the histological morphology of hair follicle was observed by using the frozen section technology and histochemistry dyeing method.The frequency of WIF1-I8-sv889 locus was determined by PCR method, and the functional elements in structural variation regions were analyzed by the online software UCSC and RegRNA 2.0.Histological research showed that compared with the skin structures of Xiang pig and Large White pigs, Xiang pig with wrinkled skin hair follicles appeared hyperplasia, hair root sheath extends into the dermis, forming spinous processes, and in Xiang pig with wrinkled skin, there were more hair follicles in the wrinkled depression, but the hair follicles in the wrinkled convex part were relatively less;Statistics on the width of the hair bulb and the number of hair follicles, the width of hair bulbs significantly increased in the hair follicles of Xiang pig with wrinkled skin (P<0.05), the number of hair shafts in a single hair follicle extremely significantly increased (P<0.01).Specific primers were designed at both ends of the WIF1-I8-sv889 structural variation breakpoint, the amplified fragment long was 1 383 bp, compared with the reference genome, 889 bp in 1 383 bp was the SV interval, 581 bp deleted, and 308 bp inverted.The population distribution results showed that, the polymorphisms of WIF1-I8-sv889 locus was abundant in the pig groups, there presented three genotypes, including types of Ⅱ, DI and DD;No genotype Ⅱ was detected in the Xiang pig with wrinkled skin;Compared with the common Xiang pig and Large White pigs, the D allele was dominant in the Xiang pigs with wrinkled skin (94.44%);Its frequency was significantly or extremely significantly higher than that of the other two pig groups (P<0.05;P<0.01).The results suggested that the structural variation of WIF1-I8-sv889 of WIF1 gene might be related to the morphogenesis of hair follicles in Xiang pig with wrinkled skin.

The purpose of the experiment was to study the effects of water extracts from walnut husks on the growth performance and the levels of avian influenza and Newcastle disease antibodies in indigenous hybrid chickens in Yunnan province.120 local indigenous hybrid chickens at 28 days old were randomly divided into 4 groups, control group, test group 1, 2 and 3, with the walnut husks water extract was respectively added to the drinking water at a concentration of 0, 1%, 3%, and 5%.30 birds in each group (half male and female).The results of growth performance showed that, at 65 days of age, the body weights of test groups were significantly higher than that of control group (P<0.05), and that in test group 3 was significantly higher than the other two test groups (P<0.05).The average daily gain (ADG) of test group 3 was 62.89 g, and was significantly higher than the control group and other test groups (P<0.05).F/G of test group 3 was lower than that of the control group and other test groups (P<0.05), which was decreased by 12.00%, 13.39%, and 5.17%, respectively.The antibody levels of Newcastle disease (ND) and avian influenza (H7, H5) were measured after immunization.After 46 d of immunization, the level of ND antibody in test group 3 was significantly higher than that in the control group and other test groups (P<0.05).After 48 days of immunization, the levels of H7 antibodies in test 2 and 3 groups were significantly higher than those in control group and test 1 group (P<0.05).Compare to control group, the level of H5 antibody had no significant difference in test group 2 and 3 (P>0.05).The results showed that the water extract of walnut green husks could improve the growth performance of and the levels of antibodies against Newcastle disease and avian influenza (H7) in Yunnan native hybrid chickens, and its effect increased with the increase of water concentration of walnut green husks water extract.In general, the supplementation of 5% walnut green husks water extract was the best.

The aim of this study was to express recombinant protein CD40L from mice by E.coli expression system, and investigate its immunopotentiating effect on tetrodotoxin (TTX) artificial complete antigen in BALB/c mice.Total RNA from spleen of BALB/c mice was extracted by Trizol reagent and retranscribed into cDNA.Primers were designed according to CD40L CDS region, and the target gene was amplified by PCR.The recombinant vector pGEX4T-1 was constructed for prokaryotic expression, and the recombinant CD40L protein was purified.TTX-BSA and TTX-OVA were prepared by formaldehyde method according to Mannich reaction principle.The adjuvant of artificial recombinant protein CD40L was used as the experimental group and the adjuvant of Freund's was used as the control group.BALB/c mice were immunized.The immune effects of each group was analyzed using ELISA method combined with SPSS 19.0 software to explore the effect of recombinant CD40L protein on the immune effect during TTX-BSA immunization.This experiment successfully amplified 783 bp CD40L gene, prokaryotic expression and purification of GST-tagged 55 ku mice CD40L recombinant protein.Compared with TTX-BSA synergistic immunization in mice, CD40L had extremely significant immune enhancement effect at the initial stage of immunization (P<0.01) than Freund's adjuvant.In conclusion, the recombinant protein CD40L and TTX-BSA complete antigen synergistically immunized mice could enhance the body's response to hapten at the initial stage of immunization, which laid a foundation for further development of immunopotentiation adjuvants suitable for the efficient preparation of small hapten antibodies.

To explore whether NPS and NPSR were involved in the process of pseudorabies virus (PRV) infection, the expression profiles of NPS and NPSR, virus gene and the related cytokines during PRV infection in vitro were studied by cell culture, RT-PCR, Real-time PCR and shRNA methods.The results of RT-PCR and Real-time PCR showed that the expression of NPS and NPSR in mouse embryonic fibroblasts (3T3 cells) infected with PRV increased extremely significantly 12 and 18 h after PRV infection (P<0.01).After stably interfering with the expression of NPSR in 3T3 cells by shRNA technology, the expression of PRV gE gene decreased extremely significantly (P<0.01).Adding exogenous NPS could significantly enhance the expression of PRV gE gene and cytokines IL-6 and TNF-α in 3T3 cells infected with PRV.However, the expression of PRV gE gene and cytokines IL-6 and TNF-α were extremely significantly inhibited by adding NPSR inhibitors (P<0.01), which was contrary to the effect of NPS.These results indicated that the robust expression of NPS and NPSR could effectively inhibit the replication of PRV in 3T3 cells, which indicated that NPSR was an important factor for the effective infection of PRV in 3T3 cells.Exogenous NPS could aggravate the inflammatory response induced by PRV infection by regulating the expression of inflammatory cytokines.These results provided a basis for further exploring the mechanism of NPS and NPSR involving in PRV infection in vivo.

In order to ascertain the cause of the stillbirth and weak piglets of sows in a large-scale pig farm in Fuyuan county, Qujing city, Yunnan province in August 2018, in this experiment, weak piglets visceral tissue materials were collected for isolation, culture, biochemical identification, 16S rRNA amplification, drug sensitivity test and pathogenicity test.The results showed that a small, translucent Gram-negative brevibacterium was isolated and cultured from weak piglets brain tissue.Biochemical identification results showed that itaconate assimilation (ITA), suberic acid salt assimilation (SUB), acetate (ACE), DL-lactate assimilation (LAT), propionate assimilation (PROP), 3-Hydroxy-butyrate (3OB), proline (PRO) biochemical reaction results were positive;Alanine assimilation (ALA), D-mannitol (MAN), D-glucose (GLU), citrate (CAP), serine assimilation (SER) and other biochemical reactions were negative.The 16S rRNA sequence had 99% homology with the 16S rRNA nucleotide sequence of Comamonas sp.EB172 strain (accession No.:EU847238.1) in GenBank.The susceptibility test showed that it was sensitive to amoxicillin/clavulanic acid, cefotaxime, polymyxin E, cotrimoxazole, spectinomycin, nitrofurantoin and doxycycline.The pathogenicity test showed that mice were inoculated intraperitoneally with 0.3 mL of 9.3×10⁸ CFU/mL bacterial suspension, and 7 mice died within 7 h (7/12).The results of this test indicated that there was a testicular infection of C.testosteroni in weak piglets and aborted fetuses.

In order to understand the gene distribution of plasmid-mediated quinolone resistance and its resistance to quinolone antibiotics in bovine E.coli in Xinjiang in recent years.116 strains of E.coli were isolated from 12 large-scale dairy farms in Shihezi, Shawan, Kuitun, Manas and Yili of Xinjiang in 2016-2018.The PMQR resistance gene was amplified by PCR and its resistance was detected by drug sensitivity test.The results showed that 62.93% of the 116 bovine derived E.coli strains were resistant to ampicillin, which was the strongest among the 15 tested antibiotics followed by streptomycin, tetracycline, kanamycin and enrofloxacin, whose resistance rates were 56.90%, 54.31%, 43.10% and 42.24%, respectively.The resistance rates to ceftazidime and cefotaxime were lower (7.76% and 11.21%).The PMQR resistance genes carried by the E.coli in Xinjiang were mainly qnrA, qnrS and aac (6')-Ⅰb-cr.The PCR results showed that there were 31 strains carrying PMQR resistance genes in 116 strains, and the positive rate was 26.72%.26, 4 and 1 strains carried 1, 2 and 3 kinds of PMQR resistance gene, accounting for 22.41%, 3.45% and 0.86% of all strains, respectively.In conclusion, the PMQR resistance genes mediated by the E.coli in Xinjiang were mainly qnrA, qnrS and aac (6')-Ⅰb-cr, and it was common for the E.coli resistance to enrofloxacin, norfloxacin, ciprofloxacin and levofloxacin in this area.

To investigate the analgesic mechanism of Senecio scandens ultrafine powder, and provide theoretical basis and reference for the S.scandens analgesic.In this study, 6-week-old Kunming mice were used as experimental animals.The suspension of S.scandens ultrafine powder was prepared, and S.scandens ultrafine powder was administered at high (360 mg/kg·BW), medium (180 mg/kg·BW) and low (90 mg/kg·BW) dose groups, after 7 days of continuous administration by gavage, follow-up related experiments were carried out.The analgesic effect was verified by formaldehyde-induced pain model and hot-water tail-flick, and it was used to distinguish central analgesia and peripheral analgesia.The analgesic mechanism of S.scandens ultrafine powder was investigated by naloxone antagonistic test, reserpine antagonistic test, and detecting the concentrations of serotonin (5-HT) and nitric oxide (NO) in serum and brain tissue.The experiment proved that S.scandens ultrafine powder could increase the pain threshold of formaldehyde-induced pain model and hot-water tail-flick in a dose-dependent manner;Naloxone and reserpine as opioid receptor blockers and anti-noradrenergic nerve terminal, respectively, could partially antagonize the analgesia effect of S.scandens ultrafine powder.S.scandens ultrafine powder could significantly increase the content of 5-HT in brain tissue and reduce the content of 5-HT in serum (P<0.05), but was not significant for NO.S.scandens ultrafine powder had central and peripheral analgesic effects in a dose-dependent manner.The analgesic pathway was related to opioid receptors and NE;It also exerted its analgesic effect by increasing the contents of central NO and 5-HT, and decreasing the contents of peripheral NO and 5-HT, but the effect of regulating NO content was not obvious.

β-receptor agonist drugs can increase the lean rate of animals and reduce the cost of animal breeding, however, when β-agonist drugs enter the human body through the food chain, it will seriously endanger human health.Therefore, most countries in the world prohibit the illegal addition of β-receptor agonist in the breeding industry.While some offenders use β-agonist drugs for the sake of interest, and it is difficult to regulate when use β-agonist drugs in the breeding process, so it is necessary to establish fast, accurate, sensitive analytical methods to detect β-agonist drugs.At present, detection methods familiar include liquid chromatography-mass spectrometry, gas chromatography-mass spectrometry, electrochemical method, surface enhanced Raman spectroscopy, chemiluminescence, immunoassay, etc.In the paper, the application of the agonist was briefly introduced, and the advantages and disadvantages of these methods were analyzed.In addition, the new technology for the detection of β-agonist drugs was prospected.It is foreseeable that the future development direction will be the more reliable and specific immunoassay strip analysis for on-site detection and the liquid chromatography-mass spectrometry method with the advantages of simple pretreatment method and high automation.

Japanese encephalitis (JE) is an acute zoonosis, and has the characteristic of infectious diseases in the central neural system (CNS), which caused by Japanese encephalitis virus (JEV) and transmitted by mosquitoes.The pigs are important storage and increase host of JEV, and it was main origin of infection.JEV can cause pregnant sow abortion and boar testitis.Humans are the terminal host of JEV, especially children, and the mortality rate can reach 25% after infection.There are still no effective drugs for JEV.The expansions of the epidemic area of JE and the changes of dominant genotypes have brought new challenges to the prevention and control strategies of JEV.In recent years, with the development of genetic and protein engineering, as well as the researches on the molecular structure and function of viruses, more and more new technologies and methods have been applied to the researches of vaccine for JEV.As a result, genetically engineered subunit vaccine, virus-like particle vaccine, chimeric virus attenuated live vaccine, polypeptide vaccine and DNA vaccine have emerged at the historic moment.In the process of vaccine preparation, the production process of optimizing the culture mode, purifying virus antigen, applying new adjuvant and heat resistant protective agents are expected to provide a new research direction for improving the quality of JEV vaccine.The present situation and development of swine encephalitis vaccine strain were reviewed in this paper to provide references for the development of safer and more effective vaccines against JEV.