RNA editing is an important post-transcriptional regulation mechanism and alter the genetic information by inserting, deleting or replacing bases in primary transcripts. RNA editing can result in amino acid change, alternative splicing, intron retention, modulation of RNA stability, and so on, which will provide a direction for explaining many complex life processes. In mammals, the deamination of adenosine (A) to inosine (I) mediated by adenosine deaminases acting on double-stranded RNA is the most common form of editing. With the development of high-throughput sequencing technology and molecular biology, researchers have developed multiple efficient, flexible and easy-to-use tools to identify the RNA editing sites at the genome-wide level, which lay a foundation for exploring the effects of RNA editing on economic traits of livestock. In this paper, we briefly review the definition, function and detection methods of RNA editing, and introduce some of the most recent and popular detection tools in detail, aiming to draw more attention to RNA editing in livestock.

In recent years, the relationship between miRNA and pregnancy in mammals has become a hot topic. The majority of miRNA can be preferentially expressed in the animal placenta, and they are related to pregnancy and birth of mammals. The effects of miRNA on embryo development, pregnancy, pregnancy diseases and breast milk production in the late perinatal period in mammals were analysed and discussed in this paper. The effects of miRNA in mammalian blood and placenta on pregnancy were summarized, wihch will provide some references for comprehending the relationship between miRNA and mammalian pregnancy.

Fowl cholera is a contagious and systemic disease of poultry worldwide caused by the species Pasteurella multocida, causing high morbidity and mortality and severe economic losses to poultry industry. As the prevalence of multidrug resistance among Pasteurella multocida isolates has continuously increased, immunization methods, the most potent preventive measure should become the main tool for controlling the disease. The current commercial vaccines contain traditional inactivated vaccine and empirically derived live vaccines, however, they exhibit some defects. The former provides limited cross-protection against heterologous serotypes; genetic background of the latter is not known and reversion to virulence occurs. Thus, novel vaccines against fowl cholera are urgently required. Here, we summarized the development of a variety of new-type vaccines against fowl cholera including attenuated live vaccine, subunit vaccine and DNA vaccine, to promote the exploration of new vaccines with safety and effective protection.

The aims of this study were to detect the differentially methylated region (DMR) and differentially methylated gene (DMG) and lay a foundation for further analyzing the skeletal muscle development difference between male and female pigs by high resolution and single base genome-wide methylation variance analysis for Large White pigs of different genders. In this study, four 210-day-old Large White pigs (two males and two females) were scanned by the whole genome bisulfite sequencing method (WGBS). The degree of methylation and differentially methylated region in the whole genome DNA of longissimus dorsi(LD) muscle were studied to explore the difference of DNA methylation level between male and female pigs. The results showed that about 5% Cytosine (C) were methylated in the whole genome. The overall methylation level of sows and boars was basically the same. A total of 1 629 DMRs and 841 related DMGs were detected. The DMRs methylation level of sows was higher than that of boars. One hundred and sev-enty one GO terms and 10 related signal pathways were detected by GO and KEGG analysis, respectively, which were showed to be significantly enriched in cell junction, axon guidance, cell adhesion molecule binding, ECM-receptor interaction, and so on. Five candidate genes were screened out in muscle tissues of male and female pigs. This study provided the single base and high resolution genome-wide methylation pattern of male and female Large White pigs. The results provide reference information for the epigenetic study of different genders and screening candidate genes related to muscle development and meat quality.

This study was designed to investigate the effects of adrenocorticotropic hormone (ACTH) on the gene transcription of glucose metabolism-related enzymes and related enzyme activities in the liver of sows, and to explore the molecular mechanism of blood glucose regulation in the liver under the stress condition. Nine healthy multiparous weaned Suhuai sows were selected and randomly divided into treatment group (N=5) and control group (N=4). Sows in treatment group and control group were injected with ACTH and physiological saline, respectively, three times daily for 7 days. The blood samples were collected daily during the experiment; The liver of sows were collected at the end of experiment. The concentrations of cortisol in blood were detected by radioimmunoassay. The levels of blood glucose were measured by biochemical analyzer. The transcript expression of enzymes related to glucose metabolism and glucocorticoid receptor(GR) were detected by real-time quantitative PCR, the activities of enzymes related to glucose metabolism were detected by spectrophotography. The results showed that the concentrations of cortisol and glucose in blood of sows were significantly raised after ACTH injection (P<0.05). ACTH treatment didn't significantly influence the mRNA expression levels of PC, FBP, PCK2, PEKL, PKLR, HK1, HK3, GYS1, PYGL and G6PD in the liver of sows (P>0.05). ACTH treatment significantly increased the mRNA expression levels of G6PC, PCK1 and HK2 in the liver of sows (P<0.05). Furthermore, ACTH treatment significantly increased the enzyme activities of G6PC (P<0.05), and significantly decreased the enzyme activities of PFKL (P<0.05). Compared with the control group,the mRNA expression levels of GR in the ACTH group showed an up-regulation trend, but there were no significant difference between the two groups (P>0.05). These results suggested that ACTH treatment upregulated blood glucose level through activating transcription of the key gluconeogenesis enzymes PCK1, HK2 and G6PC as well as the enzymatic activities of G6PC to promote gluconeogenesis, meanwhile, reducing the enzyme activities of the key glycolysis enzyme PFKL to repress the glycolysis in the liver. Furthermore, ACTH treatment didn't influence the mRNA expression of GR.

This experiment was conducted to study the transcription regulation region activity of pig MITF-M (microphthalmia associated transcription factor M) gene and verify the important transcription factors. PCR amplification with pig DNA as a template was conducted to get the transcription regulation region of pig MITF-M, and 5'-end deletion recombinant vector were obtained by gene cloning. The transcriptional activity of MITF-M was tested via dual luciferase reporter gene experiment, and RNA interference were carried out to verify important transcription factors. The results showed that a 6 414 bp transcription regulation region was cloned; Nine 5'-end deletion recombinant vectors were constructed; Three important regulatory regions of MITF-M were identified (P < 0.001),locating in —1 140-—1 397,—5 429-—6 036 bp positive regulation regions and —2 470-—3 753 negative regulation region; The transcription factor KLF4 significantly negatively regulated the transcription activity of MITF-M(P<0.01). These results indicated that the KLF4 inhibited the transcriptional activity of MITF-M and regulated melanin synthesis and deposition in the pigment cells and affected the pig coat color. These data laid the foundation for further studying the expression mechanism of pig MITF-M, and was of great significance for studying the regulation mechanism of pigs coat color.

The aim of this study was to understand the mechanism of tissue environmental adaptation in the absence of water in Camelus. In this study, six 6-9 years old adult Alashan Bactrian camels were randomly divided into 2 groups, the first group was the control group (colon1_3), in which the animals were fed with normal feed and water, the second group was the no drinking water group (colon3), in which the animals were in the absence of water for 24 d, other treatment (forage) was the same as the control group. The Illumina HiSeq 2000 sequencing platform was used to sequence the transcriptome of Bactrian camel colon tissue in the control and no drinking water groups, and the transcriptome data were used to perform quality-controlled, alignment, differentially expressed, GO and KEGG analysis. The results showed that, compared with control group, a total of 2 122 differentially expressed genes were obtained in the no drinking water group, among which 813 genes were up-regulated and 1 309 genes were down-regulated. GO enrichment analysis results showed that 30 terms in the no drinking water group were significantly enriched by down-regulated expression genes. There was no statistically significant enrichment term by the up-regulated expression genes. The KEGG pathway analysis showed that the down-regulated expression genes in no drinking water group were significantly enriched in multiply pathways, including spliceosome, protein export, protein processing in endoplasmic reticulum, RNA transport, ribosome biogenesis in eukaryotes, mRNA detection, RNA degradation metabolic pathways. The up-regulated expression genes in no drinking water group were enriched in focal adhesion, lysosome function, and so on. The results of this study indicate that the number of down-regulated expression genes in colon are more than up-regulated expressed genes in water-deficient environments, and the functions of these down-regulated expression genes are mainly associated with RNA processes and protein synthesis pathways. These genes are favorable for camels to reduce the RNA synthesis and the metabolic rate of colon tissue in water-deficient environments.

This study aimed to explore the regulatory mechanism of miR-487b-3p on proliferation and differentiation of mouse C2C12 cells. In this study,mouse myoblast cell line(C2C12) was used as the research material,qRT-PCR was used to detect the expression of miR-487b-3p in various tissues and in C2C12 cells during proliferation and differentiation in mouse. After transfection of miR-487b-3p mimics and miR-487b-3p inhibitor,the transfection efficiency was detected by qRT-PCR,and the expression of proliferation marker gene PCNA and differentiation marker genes MYOD, MYOG and MYHC were detected by qRT-PCR and Western blotting. The bioinformatics softwares were used to predict target genes of miR-487b-3p,and the expression differences of PITX2 were detected by qRT-PCR in the proliferation and differentiation of mouse C2C12 cells,and the expression of PITX2 after over-expression(inhibition-expression) of miR-487b-3p was detected,and target relationship between miR-487b-3p and PITX2 was verified via the Dual-luciferase reporter assay. The results showed that the expression of miR-487b-3p was the highest in mouse skeletal muscle, and its expression level increased during the proliferation and differentiation of C2C12 cells. It was speculated that miR-487b-3p participated in the regulation of skeletal muscle development. Over-expression of miR-487b-3p significantly up-regulated the expression of miR-487b-3p(P<0.01), and significantly up-regulated the expression of PCNA at transcription and protein levels(P<0.01),and significantly down-regulated MYHC, MYOD and MYOG genes expression(P<0.05), simultaneously, significantly down-regulated the expression of MYHC(P<0.05),MYOD(P<0.01) and MYOG(P<0.05) proteins; After inhibiting the expression of miR-487b-3p, the expression of miR-487b-3p was significantly down-regulated(P<0.01),and the expression of PCNA gene was significantly down-regulated(P<0.05),and the expression of PCNA protein was significantly down-regulated(P<0.01),MYHC,MYOD and MYOG proteins expression were significantly up-regulated(P<0.05). The expression levels of MYHC,MYOD and MYOG genes increased,but the change was not significant(P>0.05). In the process of proliferation and differentiation of C2C12 cells, the expression trends of miR-487b-3p and PITX2 was reversed;Over-expression of miR-487b-3p could significantly down-regulate PITX2 mRNA expression(P<0.01),on the contrary,inhibition of miR-487b-3p could significantly up-regulate PITX2 mRNA expression(P<0.01). Dual-luciferase reporter assay verified that PITX2 was a direct target gene of miR-487b-3p. In summary,miR-487b-3p promotes the proliferation and inhibits the differentiation of C2C12 cells by targeting PITX2 in mouse.

The present study aimed to assess the cognitive capacity for color and space of White Cashmere goats as well as their comparison with Chinese Merino sheep. The cognitive function of 6 White Cashmere goats and 6 Chinese Merino sheep with similar age (about 8 months old) and body weight were tested by the visual discriminations and reversal test and spatial maze Y-test and reversal test methods in this study. The results showed that the animals in the two groups did not reach the correct rate of 75% in the 48 tests of color simple cognitive recognition in this study, therefore, the animals in the two groups did not learn to recognize the color easily in this experiment, but they all learned the cognitive recognition for the left and right space and the reversal recognition, and there was significant difference between the two groups (P<0.05). The group of White Cashmere goats performed better in reverse spatial discrimination, the faster learning and fewer mistakes made. These results suggested that, compared with sheep, under the more complex environment, goats could establish the correct left and right spatial cognition recognition more quickly. This study explored the cognitive recognition behavior of sheep and goat, and provided important reference for the study of the correlation between the sheep and goats and their cognitive learning ability.

The aim of this study was to explore the expression profile and analyze the function of serine protease 35 (PRSS35) in bovine follicular granulosa cells. Cattle dominant follicles (DF) and subordinate follicles (SF) were separated according their sizes measured using B-mode ultrasonography technology. The expression profile of PRSS35 was analyzed using qRT-PCR and Western blotting, and its location in DF and SF was detected by immunohistochemistry. The siRNA sequences of PRSS35 were designed and synthesized and PRSS35-gene-silenced GCs line was obtained. Estrogen (E₂) concentration and cell apoptosis were detected through studying the effects of PRSS35 on growth and development of bovine follicular GCs. The results showed that:1) The expression of PRSS35 mRNA in DF was significantly higher than that in SF (P<0.001). 2) The PRSS35 was expressed both in DF and SF granular layer and theca layer. 3) The expression of RSS35 protein in DF was significantly higher than that in SF (P<0.01). 4) When PRSS35 gene was silenced, the percentage of apoptotic GCs was significantly increased (P<0.001), and the concentration of E₂ in cell culture media was significantly decreased (P<0.01). The results suggested that PRSS35 was expressed both in bovine follicular granular layer and theca layer, and the PRSS35 expression was significantly higher in DF than that in SF. The proliferation of GCs was inhibited, and the concentration of E₂ was reduced when PRSS35 gene was silenced, indicating the growth and development of bovine follicles were inhibited. This study provides a basis for further studying the relationship between PRSS35 and follicular development in cattle.

In order to explore the effect of age at puberty on reproductive performance in Yorkshire sows,the data including the total number of born (TNB), the number of born alive (NBA), number of healthy piglets (NHP) and litter birth weight (LBW) of 923 American line Yorkshire sows were recorded and analyzed. According to the days gilts reached puberty, we divided replacement gilts into 4 groups (Group Ⅰ: ≤ 200 d, Group Ⅱ:201-215 d, Group Ⅲ:216-230 d and Group Ⅳ:>230 d) to discuss the influence for sow's reproductive performance. The results showed that:1)the TNB of group Ⅳ in the first parity was 12.05, which was significantly higher than that of Group Ⅰ (11.45, P<0.05), there was no significant difference in other reproductive traits; 2)in second parity, the TNB and NBA in group Ⅱ were 11.84 and 11.40 respectively, which was significantly higher than 11.09 and 10.66 in group Ⅰ (P<0.05); 3)in third and after third parity, the TNB, NBA and NHP in group Ⅱ were 12.47, 11.82 and 11.48 respectively, which were significantly higher than those in group Ⅳ with 11.89, 11.22, 10.85 (P<0.05); 4)the average TNB, NBA and NHP of the first 6 parities in group Ⅱ were 74.23, 71.41 and 68.86 respectively, which were significantly higher than those of group Ⅳ with 70.48, 67.32 and 64.86 (P<0.05). The result indicated that the reproductive performances of early puberty gilts, especially when the puberty ages between 201-215 days,was significantly better than later puberty sows. In conclusion,the best age in days of puberty and first-mating,we found provides a theoretical basis for the selection of replacement gilts and the application of pig farm production.

The objective of this study was to explore the relationship between autonomic nerve and cellular immune in uterus during peri-implantation based on mast cell (MC) count and histamine content. Ninety healthy female Wistar rats weighed 300-350 g were randomly assigned into 3 groups. One group as neurectomy (the autonomic nerves governing the uterus from the pelvis nerve and mesenteric caudal ganglion were amputated by surgical operation before pregnancy) experiment group, one group as sham operation group and the other as the control. After fully recovering from the surgery, female rat was cohabited with male rat in ovulatory period. On day 4, 5, 6 of pregnancy(d4, d5, d6), the females were killed by neck dislocation, uteri and ovary were surgically removed in abacterial condition. The number of ovulation sites was observed under an anatomy microscope and the quantity of embryos was counted, MC count was determined by cell staining, and histamine was assayed using histamine ELISA kits. The results showed that the spontaneous release and content of histamine in the uterus of normal rats were the lowest level at the time of implantation (d5). The MC count presented the similar changes to histamine. Compared with the control, spontaneous release of histamine from uterus in operated rat showed a significant decrease at d4, d6 of pregnancy (P<0.05), amputation of the nerve innerving uterus very significantly decreased the total content of histamine (P<0.01) before embryo implantation (d4), meanwhile the number of implanted embryo was also markedly decreased or delay. There were similar changes in the number of uterine MC and histamine content during implantation after neurectomy. These results suggested that resection of the nerve innerving uterus decreased MC counts and histamine level in the period of pre-implantation, changed uterine local immunity level, leading to reduction or delay of implantation. Taken together, our results imply that the autonomic nerve innerving uterus can up-regulate the local immune level in the the uterus through the proliferation of MC and histamine releasing, and play an important regulating role in embryo implantation.

The aim of this study was to explore the effect of Saccharomyces cerevisiae β-glucan on the β-defensin-1(SBD-1) expression in cultured ruminal epithelial cells(RECs) of ovine. Firstly, the ovine RECs culture system was established as an in vitro experimental model, the RECs were stimulated with various concentrations (0, 5, 10, 20, 50 and 100 μg·mL^-1) of β-glucan for 8 h, and the expression level of SBD-1 mRNA and protein in RECs were detected by qPCR and ELISA to determine the concentration of β-glucan that induced the highest expression of SBD-1. Then, RECs were stimulated with the optimal stimulating concentration of β-glucan for 0, 2, 4, 8, 12 and 24 h, respectively and the expression of RECs SBD-1 were also detected by qPCR and ELISA, so as to determine the optimal stimulation time for β-glucan-induced SBD-1 expression. The qPCR and ELISA results showed that after β-glucan (5-100 μg·mL^-1) stimulated RECs for 8 h, the expression of SBD-1 mRNA and protein were increased and then decreased following the increase of concentrations, and when the concentration of β-glucan was 10 μg·mL^-1, the expression level of SBD-1 mRNA and protein were significantly higher than the control group (P<0.01). When RECs were stimulated with 10 μg·mL^-1 β-glucan for 0-24 h, qPCR results showed that SBD-1 mRNA expression was the highest at 2 h (P<0.01), and then downward; The results of ELISA test showed that SBD-1 protein secretion reached the highest level at 4 h (P<0.01).The results of MTT test showed that 100 μg·mL^-1 β-glucan significantly affect the sheep RECs viability (P<0.05). The results of this study indicated that β-glucan could improve the SBD-1 expression in RECs of ovine. And the level of SBD-1 mRNA and protein expression was the highest when the RECs were separately stimulated for 2 and 4 h with the 10 μg·mL^-1 β-glucan.

The aim of this study was to investigate the effects of excessive vitamin A(VA) supplementation on growth performance, nutrient digestibility and serum biochemical indexes of growing male minks. Using a single factor randomized trial design, ninety healthy male minks were randomly assigned to 6 treatment groups with 15 animals in each group and fed a diet supplemented with 0(Control), 5 000(Ⅰ),20 000(Ⅱ), 80 000(Ⅲ), 320 000(Ⅳ) and 1 280 000 IU·kg^-1(Ⅴ) VA (based on dry matter), respectively, the pretest period lasted for 7 days and the formal test period lasted for 60 days. On the 30th day of the trail, 8 minks from each group were selected randomly for the digestion and metabolism test for 3 days, the feces and urine samples were collected by total feces collection method, the contents of dry matter (DM), crude protein (CP) and crude fat (EE) were measured, and the nutrient digestibility was calculated. Blood samples were collected at the end of the test and the contents of blood glucose(GLU), triglyceride(TG),cholesterol(CHO), high density lipoprotein cholesterol(HDL), low density lipoprotein cholesterol(LDL), alkaline phosphatase(ALP), aspartate aminotransferase(AST), alanine aminotransferase(ALT) and lactic dehydrogenase(LDH) were measured. The results showed that:1)Final weight and average daily gain in group Ⅱ was significantly higher than that in group Ⅴ(P<0.05). 2) Compared with control group, VA could improve the digestibility of nutrients (P<0.05), group Ⅱ was all the best. 3) VA had no significant effects on GLU and CHO (P>0.05), TG content in group Ⅴ was significantly higher than that in group Ⅱ(P<0.05). HDL content in group Ⅳ and Ⅴ were significantly lower than that in the other groups (P<0.05). LDL content in group Ⅰ was the lowest, which was significantly lower than that in group Ⅳ and Ⅴ(P<0.05). 4)Serum ALP, AST, ALT and LDH levels were significantly affected by dietary VA (P<0.05). ALP content had a significant difference between group Ⅴ and control, group Ⅲ, Ⅳ(P<0.05). Concentration of serum AST was significantly decreased in group Ⅱ, which was significantly lower than that in control, group Ⅳ, Ⅴ(P<0.05). VA induced a significantly lower concentration of ALT in group Ⅱ(P<0.05), group Ⅴ was higher than control, Ⅰ,Ⅱ groups(P<0.05). Concentration of LDH in group Ⅴ was significantly lower than that in group Ⅰ, Ⅱ, Ⅲ (P<0.05), whereas there was no significant differences between the control group and group Ⅳ(P>0.05). Under the experimental conditions, the supplementation of VA with 5 000-80 000 IU·kg^-1 in diet could improve growth potentials. However, excessive vitamin A supplementation could negatively affects growth and lipid metabolism. Further research is needed to determine the effect mechanism of excessive vitamin A supplementation on the growth of minks.

In order to study the effect of novel_miR218 on the expression of signaling lymphocyte activation molecule (SLAM), the receptor of Peste des petits ruminants virus (PPRV) on goat lymphocytes, we detected the expression levels of SLAM, miR218 and viral loads in goat peripheral blood mononuclear cells (PBMCs) infected with PPRV vaccine virus N75-1 strain by using Western blot assay and quantitative PCR analysis. Then, we detected the levels of SLAM expression and viral loads in PPRV-infected goat PBMCs pre-transfected with miR218 mimic or miR218 inhibitor. The results showed that the cytopathic effect (CPE) can be detected in infected cells at 24 h post infection (hpi) with progressive increase in CPE at 48-72 hpi. In compared with the mock infected group, the expression fold of SLAM mRNA in PPRV infected group increased significantly at 24 hpi (P<0.01) followed by a gradually decreased levels, while the expression level of novel_miR218 decreased significantly at 24 hpi (P<0.01) and then gradually increased. In the uninfected control group, the expression level of novel_miR218 and SLAM mRNA showed no significant change at each detected time point. The expression pattern of SLAM protein in the infected group was similar to that of the mRNA, while an increased levels of PPRV-N protein was observed. The expression levels of SLAM and PPRV-N in miR218 mimic transfected group were significantly lower (P<0.05 or P<0.01) than that in the uninfected control group in a dose-dependent manner, while the expression levels of SLAM and PPRV-N in miR218 inhibitor transfected group were significantly higher (P<0.05 or P<0.01) than those in control group in a dose-dependent manner. This study indicated that the expression of SLAM was closely correlated with the miR218 expression in PPRV infected goat PBMCs, and the results of transfection of cells with miR218 mimic or inhibitor suggested that miR218 can negatively regulate the expression of SLAM and virus replication in PPRV infected PBMCs.

This study aimed to screen highly transformable Haemophilus parasuis (HPS)strains, which in the hope of providing biological materials for the further study of natural competence mechanism and gene function research. Based on the genome sequences of the selected strain and comparative genome analysis, we attempted to investigate the molecular genetic basis of natural transformation of HPS. In this study, 75 HPS field isolates and 15 standard strains were screened by natural transformation method, and the transformation efficiency of the natural competent strain was determined. The genome of the strain SC1401 with high natural transformation efficiency was sequenced using the third-generation sequencing technology on PacBio, and then analyzed using related software for genome assembly, gene prediction, functional annotation and synteny analysis. The sequencing result of SC1401 strain was compared with those of SH0165 strain (GenBank:CP001321), SH03 strain (GenBank:CP009158), KL0318 strain (GenBank:CP009237) and ZJ0906 strain (GenBank:CP005384). The results showed that:1) 11 HPS strains were identified as natural competent strain, which all were wild type, and SC1401 had the highest transformation efficiency among them. 2) The genome size is 2 277 540 bp with GC content of 40.03%. The total gene quantity is 2 220, accounting for 87.75% of the whole genome. This genome sequence had been submitted to GenBank database under the accession number CP015099. 3) Synteny analysis indicated that the relationship between the 5 HPS strains were close-related. 4) Comparative genomics analysis showed that SC1401 isolates contained 385 unique genes, much more than the other 4 strains. Five HPS strains contained a series of identical genes related to natural transformation, and in addition to the tfox, the homogeneity of the amino acid sequences of each gene had reached over 90%. This study is the first report of the whole genome sequence of SC1401, a highly naturally competent strain, and analyzed the basic characterization of the genome, so as to provide a theoretical basis for later research on the mechanism of natural transformation of HPS.

In order to understand the epidemic situation of Gallibacterium in silky and the genetic evolution characteristics of isolates, samples were collected for the isolation of Gallibacterium anatis and q-PCR identification from two silky farms in Xinyang and Nanyang, Henan province, then isolates were detected by Gallibacterium specific PCR followed by drug susceptibility test, and the PCR amplification and sequencing of the 16S rRNA, rpoB and gyrB genes. Finally, the MegAlign program was used for calculating the similarity between isolates of G. anatis, and MEGA5.0 for constructing evolutionary tree and genetic evolution analysis. Results were as follows:Of 22 silky chickens, 16 were found positive to G. anatis, the positive rate was 72.7%; the four Gallibacterium isolates were identified as G. anatis; the four isolates were all multidrug resistant, and were sensitive to ceftriaxone, cefotaxime, amikacin and tobramycin only; the homology between the isolates and the reference G. anatis species was 99.1%-99.8% (16S rRNA), 85.4%-99.6% (rpoB), and 92.7%-99.8% (gyrB), respectively. In the 16S rRNA gene evolution tree, the four isolates and the reference G. anatis were located in the same branch; the GAC193 isolate in the rpoB gene evolution tree were located in the same branch with Pasteurella multocida, not in the same branch with other G. anatis; in the gyrB gene evolution tree, GAC017 and G. genomosp.1 were in a small branch. The results showed that G. anatis were found in the silky chickens with a high detection rate, and four strains of G. anatis were isolated, which showing serious multidrug resistance. There are significant genetic differences in the rpoB and gyrB gene of some isolates of G. anatis.

To identify the role of protein gp38 in the adsorption process of phage Bp7, gp38 gene was amplified by PCR, and the amino acid sequence was characterized. The recombinant plasmid pColdTF-gp38 was constructed and expressed in E. coli BL21, then identified by SDS-PAGE and Western blot. Protein gp38 was purified by affinity chromatography to prepare polyclonal antibody. Immunoelectron microscopy assay was used to locate gp38 on phage Bp7, protein competition assay and antibody blocking assay were used to determine gp38 and its antibody effects on phage Bp7 adsorption efficiency. Amino acid sequence analysis of phage Bp7 protein gp38 showed that there was no homology with T4 bacteriophage, but shared homology with phage T2 and T6. The conserved regions and variable regions of gp38 C-terminal sequence were arranged at intervals, showing a mosaic characteristics; the recombined plasmid pColdTF-gp38 was constructed and expressed in E. coli BL21, The gp38 polyclonal antibody was prepared with a 1:16 titer. The immunoelectron microscopy showed that the gp38 antibody could bind to the long tail fiber and cause the aggregation of phage Bp7; protein competition test and antibody blocking test showed that gp38 and its antibody could completely inhibit the absorption of phage Bp7 to host strain E. coli K12. All of results indicated that gp38 is the tail fiber protein of phage Bp7, located at the end of the long tail fiber, and played a key role as the receptor recognition protein in phage Bp7 adsorption process, which further provided the theoretical foundation to elucidate the receptor recognition mechanism of phage Bp7.

To explore the genetic diversity and genetic structure of Psoroptes ovis var. cuniculi, the complete sequence of mitochondrial 12S gene of P. ovis var. cuniculi isolates from 5 geographical regions (Huabei, Huadong, Huazhong, Xibei, and Xinan) in China, were obtained by PCR sequencing techniques, and evaluated the genetic diversity of P. ovis population. Eighty-nine sequences of mitochondrial 12S were obtained. The multiple alignments of the full 12S gene from 89 isolates were 657 bp. These sequences contained 76 variable sites and were classified into 50 haplotypes (H1-H50). The values of haplotypes diversitiy (Hd) and nucleotide diversity (π) were 0.931 05 and 0.005 59 respectively. There were low level of genetic differentiation, but high level of gene flow (F_st=0.042 322, Nm=5.657 372) among the 5 geographical population. In P. ovis population, the average values of Tajima's D and Fu's Fs were negative (Tajima's D=-0.928 31, Fu's Fs=-7.066 71) and there was no significant difference (P<0.05). NJ phylogenetic tree and haplotype network were both revealed that 50 haplotypes from 5 different geographical regions dispersed across different clades without associating with the geographical locations. The results revealed that there was a high level of genetic diversity, little population genetic differentiation and high gene flow in the P. ovis var. cuniculi population of China, but population structure has not formed according to rabbits breed, temperature zones and geography origin.

The purpose of this study was to analyze the expression profile characteristics of miRNA of the protoscoleces in Echinococcus granulosus and to provide theoretical evidence for further revealing the role of miRNA in the development of the protoscoleces. The protoscoleces were collected from the infected sheep liver under sterile condition, and the total RNA was extracted. Then the miRNA Library of the protoscoleces was constructed and sequenced by RNA sequencing technology. The results showed that 109 known miRNA molecules were isolated from the protoscoleces, and 189 novel miRNA hairpin precursors were detected in the PSC of the E. granulosus. The target gene prediction results showed that these known miRNAs had 132 target genes, and the novel miRNAs had 3 152 target genes. A further study found that these miRNA molecules are widely involved in growth and development of key signaling pathways, such as Wnt, cAMP, Hedgehog, NF-κB, Notch and bile salt pathway. miRNA molecules were further researched by real-time quantitative PCR and in situ hybridization. The results suggested that there may be potential regulatory relationships between HG328413.1_36258 and β-catenin, HG328414.1_36364* and dis, as well as HG328399.1_25846 and axin. To sum up, a large number of new miRNA molecules were found in the E. granulosus in this study, which enriched the data of miRNA of the protoscoleces, and laid a foundation for further analysis of the role of miRNA in the development of protoscoleces in E. granulosus.

This study aimed to predict and identify the nuclear localization signal (NLS) of chicken matrin 3 (MATR3). The putative NLS (pNLS) in chicken MATR3 was predicted by the online NLS prediction software. The recombinant plasmids of chicken MATR3 with the pNLS deletion were constructed and then transfected into cells. The subcellular localization of the pNLS deletants was observed and analyzed to determine the NLS of chicken MATR3. The recombinant plasmids expressing the deletion of basic amino acids close to NLS were also constructed to examine their roles in the nuclear localization of chicken MATR3. In addition, the conservation analysis and nuclear import ability of chicken MATR3 NLS were investigated. The results showed that there were four pNLS existed in chicken MATR3, and the deletion of pNLS2 ⁽⁵⁹⁶PSDKKSK⁶⁰²) disrupted the nuclear localization of chicken MATR3. In addition, the constructed recombinant plasmids expressing the deletion of basic amino acids close to NLS were transfected into cells, and the subcellular localization of the recombinant proteins indicated that the adjacent basic amino acids of NLS did not participate in the NLS-mediated nuclear localization of chicken MATR3. Moreover, we also found that the NLS motif in MATR3 protein were conserved among different species, and also had the same nuclear import ability as the NLS of SV40 large T antigen. One highly conservative NLS (⁵⁹⁶PSDKSK⁶⁰²) is present in chicken MATR3, and its adjacent basic amino acids do not participate in the nuclear localization of MATR3 protein.

In order to detect the effect of gga-miR-155 on the biological behavior of MDCC-MSB1 cells, MDCC-MSB1 cells were transiently transfected with gga-miR-155 mimic, gga-miR-155 inhibitor, and negative control which were artificial synthesized. Then, the expression of gga-miR-155 was detected by real-time PCR (qRT-PCR); the cell proliferation was detected by CCK-8 assay; the cell cycle and apoptosis were analyzed by flow cytometry; and the cell migration and invasive ability were determined by transwell assay. Results showed that gga-miR-155 mimic up-regulated the expression level of gga-miR-155 in MDCC-MSB1 cells. After gga-miR-155 mimic transfection, the proliferation of MDCC-MSB1 cells was promoted, G1 phase cells were decreased, S and G2 cells were increased, the apoptosis rate was reduced,the migration invasion capability were increased in gga-miR-155 mimic transfected cells. On the contrary, the gga-miR-155 inhibitor significantly down-regulated the expression level of gga-miR-155 in MDCC-MSB1 cells. When the gga-miR-155 inhibitor was transfected into MDCC-MSB1 cells, cell proliferation was inhibited,G1 phase cells were increased, S and G2 cells were reduced, the cell apoptosis was increased, and the migration invasion capability were suppressed. The results showed that gga-miR-155 could promote the proliferation, migration and invasion of MDCC-MSB1 cells and inhibit their apoptosis.

This study was conducted to investigate the effect of transient receptor potential vanilloid receptor 6 (TRPV6) gene silencing on expression of proteins involved in calcium ions transcellular transport in duodenum and jejunum, concentration of calcium and phosphorus and PTH level in plasma, and bone mineral density of femur and tibia in laying hens. Sixty-four laying hens at the peak stage (220 days old) were assigned to control and TRPV6 gene silencing groups, and injected, respectively with physiological saline or interference vector pSIREN-TRPV6-3 in one time, and continuously observed for 28 d. The results were as follows:compared with control group, the expression of ZsGreen protein as an indicator of transfection in duodenum and jejunum were successfully detected at the 7th, 14th and 21st day after injection. The mRNA and protein expression of TRPV6 and CaBP-D28K in duodenum and jejunum exposed to TRPV6 gene silencing were observably decreased at the 7th, 14th and 21st day (P<0.05 or P<0.01). Additionally, the level of PTH in plasma was markedly increased compared with control (P<0.05 or P<0.01). However, the concentration of calcium and phosphorus in plasma were not affected by injection of interference vector pSIREN-TRPV6-3, and the bone mineral density of femur and tibia in the group of TRPV6 gene silencing were slightly lower than control group (P>0.05). It was concluded that the transfection of pSIREN-TRPV6-3 plasmid could suppress the expression of TRPV6 and CaBP-D28K in duodenum and jejunum, but doesn't regulate the concentration of calcium in plasma and bone mineral density, it is implied that silence of TRPV6 gene does not affect normal calcium transport of laying hens under the condition of normal calcium content.

The aims of this study were to investigate changes of sperm quality and heat shock protein(HSP)70 expression before and after freezing yak semen. The semen of 6 healthy Datong yaks with normal reproductive ability was collected and assigned to 2 groups:fresh group and frozen-thawed group. The quality of sperm was evaluated by assessing sperm motility and plasma membrane integrity. The expression of HSP70 in both groups was detected by the real-time quantitative polymerase chain reaction,Western blot and immunofluorescence techniques. The results showed that,compared with fresh sperm,sperm motility and plasma membrane integrity of frozen-thawed sperm were significantly decreased (P<0.01); In addition,the expression of HSP70 was significantly lower at both the transcriptional (P<0.05) and protein (P<0.01) levels. HSP70 protein was distributed over the sperm acrosome,acrosome equatorial segment,post-acrosomal and middle sections of sperm in the fresh group. In contrast,after freezing and thawing,HSP70 protein was mainly located in post-acrosomal and middle sections of sperm,and the average optical density was significantly decreased(P<0.01). In conclusion,yak sperm quality and the expression of HSP70 declined simultaneously after freezing and thawing,suggesting that they may be associated.

The co-infection of avian influenza virus (AIV) and Newcastle disease virus (NDV) in Chickens is common clinical incident. To study the effect of co-infection of AIV (H9N2) and NDV (Herts/33) on virus replication in the DF-1 of chicken embryo fibroblasts, the replication of AIV and NDV in the DF-1 was first analyzed respectively and post-infection 12 and 24 h were selected as sample collecting time. The Western blot, immunofluorescence assay and real-time fluorescence quantitative PCR (qPCR) were conducted to compare the replication of AIV and NDV between co-infection and individual infection groups. Compared with the single virus infection group, the intensity band ratio of NDV NP in Western blot analysis in 12 and 24 h post-infection were down-regulated by 70.6% (P<0.001) and 45.7% (P<0.001),while AIV NP was down-regulated by 61.5% (P<0.001) and 82.8% (P<0.001), respectively. The optical intensity band ratio of AIV NP and NDV NP were decreased by 81.1% (P<0.001) and 65.5% (P<0.001) in 12 h post-infection, 41.2% (P<0.05) and 47.4% (P<0.001) in 24 h post-infection. The relative mRNA expression of NDV NP in co-infection in 12 and 24 h post-infection was descended by 64.3% (P<0.001) and 64.4% (P<0.05), while AIV NP was 61.5% (P<0.001) and 63.0% (P<0.001), respectively. These findings indicate that co-infection of AIV and NDV is able to inhibit their replication at cellular level, laying the foundation of further study the mechanism of common infection of AIV and NDV.

The present study was to examine the prevalence of Haemobartonella felis infections in cats from some areas of Yunnan province, a total of 338 fresh blood were collected and was detected using PCR amplification of the 16S rRNA gene, and the product of PCR were sequenced and compared the phylogenetic analysis. Results showed that 21 samples were positive, and the overall prevalence of H. felis was 6.21% (21/338). Twenty samples were infected with Candidatus Mycoplasma haemominutum, and one sample was infected with Mycoplasma haemofelis. Analysis of influences factors showed that there was a significant difference between breeding mode and age (P<0.05). The present investigation firstly report of the H. felis infection in cat in Yunnan and the study provide the basic data for preventing and controlling H. felis infection in cats from Yunnan Province.