Abstract:Abstract:Due to the overuse of anti-microbiological drugs，the resistance of pathogenic bacteria is of great attention for the international public health． According to previous reported literatures，over 80% bacterial infection was relative and contained to biofilm． Recently，many literatures have studied the pathogenic mechanism of quorum sensing (QS) and biofilm，but the relationship between QS and biofilm was hardly published． Revealing the relationship between the two can provide a new strategy for resolving the resistance of pathogenic bacteria． Based on mechanism of QS and biofilm formation and our research progress，the relationship between bacterial QS and biofilm formation is reviewed．

Abstract:Abstract:Avian hepatitis E virus (HEV)，a member of Hepeviridae family，is genetically and antigenically related with human and swine HEV in the family． Since its discovery，avian HEV infection has been investigated in many countries from serology and molecular epidemiology studies． At present，five complete or near complete genomes of avian HEV isolates were reported in GenBank and were divided into three genotypes．The complete genome of avian HEV contains 3 ORFs of which ORF2 gene encodes capsid protein containing the primary epitopes of viral particles and is target gene for serodiagnostic antigen and vaccine candidate． Because avian HEV infection has significant impact on the poultry industry and potential zoonotic transmission，the researches on avian HEV have been given much attention． We here give a broad review of the research update on the aetiology，pathogenesis and the antigenicity of capsid protein of avian HEV based on identification of Chinese avian HEV isolate．

Abstract:Abstract:The necessity and feasibility of constructing dominant genome-simplified strains for industrial production were introduced，based on some successful cases in which the production efficiency was improved after simplifying the genome of strains． The principle and process of genome simplifying were summarized． In addition，the perspective of dominant genome-simplified strains for industrial production was discussed，combined with authors' own studies.

Abstract:Abstract:［Objective］ Phosphate-solubilizing bacteria (PSB) were isolated，screened and identified from the rhizosphere of Taxus chinensis var． mairei，and growth-promoting effects on T．chinensis var． mairei by high effective PSB were determined．［Methods］ By using selective culture media，PSB were isolated from rhizospheric soil，the high effective PSB was further screened using NBRI-BPB medium，and the molybdenum-antimony anti-spectrophotometric method was applied to determine the phosphate-dissolving ability of the high effective PSB after four days fermentation in NBRIP medium．Bacteria were identified by the Biolog system combined with 16S rDNA gene sequence analysis and morphological，physiological and biochemical characteristics． The inoculation test in potted seedlings was carried out under the greenhouse．［Conclusion］ Four strains of high effective PSB were screened and identified as Pseudomonas fluorescens，Bacillus cereus，Sinorhizobium meliloti and Bacillus licheniformis，respectively． These strains had significant effects on improving the growth of the seedlings of T．chinensis var．mairei．

Abstract:Abstract:［Objective］ To analyze the diversity of bacterial community in rectum of diarrheic calves，and differences with health calves．［Methods］16S rRNA clone libraries were constructed，positive clones were digested by Msp I and Hha I for restriction fragment length polymorphism (RFLP) ，and then a phylogenetic tree was depicted based on the 16S rRNA sequencing，to confirm the compose of microbe in the diarrheic calf rectum．［Results］ The positive rate of clone was 98. 75% (474/480) in diarrheic calves，the dominant bacteria included Lactobacillus (14%)，Enterococcus (10%) and Escherichia (8%)． The positive rate of clone was 96. 45% (488/506) in health samples，the dominant bacteria included Clostridium (13%)，Bifidobacterium (8%)，Megasphaera (5%)．［Conclusion］Complexity and diversity of bacterial community in rectum in 2 weeks old calves had their own features，and significant increase of Lactobacillus，Enterococcus and Escherichia was found in diarrhea calves．

Abstract:Abstract:［Objective］To obtain phosphate-dissolving genes from cDNA library of Aspergillus niger H1.[Methods] The double-stranded cDNA was synthesized using switching mechanism at 5end of RNA transcript technique and ligated to the vector pBluescript II SK (+)．We transformed recombinant plasmid into E．coli HST08，resulting in a primary cDNA library．We screened clones with phosphate-dissolving activities on the insoluble phosphate medium and blasted the sequence in National Center for Biotechnology Information (NCBI)．To study the phosphate dissolving mechanisms of the cloned gene，we analyzed the changes of the pH value，the soluble phosphate content and the production of organic acids in the insoluble phosphate liquid medium inoculated with the clones harboring the phosphate-dissolving gene.［Results］A cDNA library of A.niger H1 was successfully constructed． Titer tests showed that the content of constructed A.niger H1 cDNA library reached 5.65×106 cfu/mL，in which the percentage of recombinant clones was 99.15%．We screened 61 clones with phosphate-dissolving activities on the solid medium with insoluble phosphate．The corresponding gene in one of these clones was identified． The full length cDNA of clone H-54 was 839 bp，encoding a predicted protein with 180 amino acid residues．The expression of phosphate-dissolving gene in E.coli enhanced organic acids secretion and improved the phosphate solubilizing activity．Formic acid and acetic acid were found in 12 h，and malic acid and α-ketoglutarate were secreted in 24 h．The clone H-54 decreased the pH value of medium from 6.32 to 3.93 and released soluble phosphate up to 0.105 mg /mL in 36 h.［Conclusion］We had obtained a phosphate-dissolving gene designated psgA from Aspergillus niger H1．

Abstract:Abstract: ［Objective］To investigate whether the recombinant baculovirus BV-T7 hybrid expression system can be effectively transduced into chicken cells in vitro，and whether it can express the foreign genes (eGFP).［Method］We established a hybrid baculovirus-T7 RNA polymerase system for transient expression in mammalian cells and chicken cells．Two recombinant baculoviruses were constructed，one carrying cDNA of bacteriophage T7 RNA polymerase，with a nuclear localization signal，under the control of a mammalian promoter and the other expressing eGFP gene under the control of T7 promoter． The constructed recombinant baculoviruses co-infected mammalian oligodendrocyte cells，as well as chicken embryo fibroblasts cells and chicken embryo skeletal muscle cells． ［Results］ The eGFP activity was detected in mammalian cell lines and embryo fibroblasts cells and chicken embryo skeletal muscle cells． The recombinant baculovirus transduction efficiency of oligodendrocyte cells was 59.5%，and in CEF cells and myoblast cells the transduction efficiencies were 23.2% and 33.1%．［Conclusion］BV-T7 hybrid expression could be expressed T7 RNAP in mammalian cells and avian cells.

Abstract:Abstract:［Objective］To improve use efficiency of phosphorus in latosolic red soil and to explore mechanism of phosphate solubilization.［Methods］ Pikovskaya and National Botanical Research Institute’s Phosphate broth were used to isolate a phosphate-solubilizing bacterium coded as C5-A from the rhizosphere soil of peanut． According to its morphological，physiological，biochemical properties and its 16S rRNA sequence，its position in phylogenetic development tree was defined． By measuring changes in pH of the National Botanical Research Institute’s Phosphate solution in which C5-A was incubated，phosphate solubilizing capacity was determined． Through fermentation，effects of carbon and nitrogen sources on the capacity of strain C5-A were investigated． Kinds and concentrations of organic acids in the cultures different in N sources were also determined by HPLC.［Results］ The strain was identified as Burkholderia cepacia，which is stable in hereditary． In aluminum phosphate and ferric phosphate solutions，its P solubilizing capacity was negatively related to pH．It solubilized tricalium phosphate，aluminum phosphate，ferric phosphate and rock phosphates powder，and could dissolve as much as 125.79 mg/L，227.34 mg/L，60.02 mg/L and 321.15mg/L P，respectively． For RPP，P solubilizing capacity of the strain was related to type and concentration of the powder． When using maltose and ammonium oxalate as C and N sources，the strain displayed its highest P solubilizing capacity． HPLC analysis detected 10 organic acids in the culture，namely: oxalic acid，acetic acid，malic acid，lactic acid，citric acid，cuccinc acid and 5 unknown organic acids．Interestingly，it is acetic acid rather than gluconic acid being the most important organic acid affecting P olubilization．［Conclusion］ The strain isolated from the rhizosphere soil of peanut plants growing in a red soil field can dissolve hardto-solve inorganic salts，and is a promising microbial resource for development of high efficiency biological phosphorus fertilizer for latosolic red soil.

Abstract:Abstract:［Objective］Comparing the effects of different carbon-nitrogen nutrition and their consumption on laccase production，we studied the ecophysiological characteristics of Phanerochaete chrysosporium resisting nutritional repression，and the carbon-nitrogen physiological regulation mechanism of the white-rot fungi.［Methods］ The mutant and the wildtype strains were respectively cultured under the conditions of: carbon and nitrogen limitation，carbon limitation and nitrogen sufficiency，carbon sufficiency and nitrogen limitation，carbon and nitrogen sufficiency，to compare their laccaseproduction kinetics，cell growth and glucose and ammonia nitrogen consumption to show the characteristics and the regulation pathway of carbon-nitrogen nutrition on laccase production．［Results］The wild-type strain produced 0.107U/L，0.029U/L，12.84U/L and 18.05U/L of laccase respectively on 11th，14th，19th and 19th day when glucose or ammonia nitrogen was consumed to the lowest value; the mutant produced laccase throughout the whole process with two peaks respectively on 8th，7th，12th and 12th day with laccase of 298.83U/L，343.14U/L，271.22U/L and 251.49U/L and on 12th，13th，19th and 19th day with laccase of 257.69U/L，298.78U/L，213.81U/L and 216.93U/L. The enzyme-production kinetics trends were similar between the two strains on the condition of the same initial carbon concentration but were different on the same initial nitrogen concentration， which showed that carbon source had more effect on laccase production.［Conclusion］The laccase production of the wild-type strain was regulated by carbon or nitrogen starvation． Under different conditions，it was regulated by different nutrient．For example，under carbon limitation condition it was started by the glucose starvation，however under carbon sufficient condition the ammonia nitrogen starvation aroused it．The laccase production of the mutant didn t repress by carbon and nitrogen nutrition．Maybe it referred to a global regulation change which relieved nutritional repression on the laccase production．

Abstract:Abstract:［Objective］We characterized a chitinase (Tachi1) from Trichoderma asperellum and optimized its production conditions，by methanol induction of the recombinant strain Pichia pastoris GS-tachi1-K transformed with the gene tachi1 (GenBank accession: GU457411)．［Methods］ We purified Tachi1 from the fermentation broth to analyze enzymatic properties after it was secreted by GS-tachi1-K．The production conditions of GS-tachi1-K were optimized by single-factor experiment and orthogonal experiment．［Results］The molecular weight of Tachi1 was about 44 kDa．Tachi1 had a broad range of temperature and pH adaption with the optimal reaction temperature at 50℃ and pH 5. 5. It was stable at the temperature below 50℃，yet less stable under alkaline conditions．Its activity was significantly reduced by 0.05 mol/L of Ag+，Hg2+，Cu2+，Fe2+，1% of Sodium dodecyl sulfate (SDS) and 10 mmol/L of β-mercaptoethanol．The optimum conditions obtained were: initial cell density with an OD600 equal to 2，0.5% of methanol，pH 6.5，induction time 180 h． Under the optimized condition，the activity of Tachi1 reached 17.93 U/mL and the expression of tachi1 was 6.19g/L．［Conclusion］The recombinant strain GS-tachi1-K showed high expression of tachi1 and the protein secreted by GStachi1-K had high chitinase activity．It will provide theoretical basis for further research and application in this chitinase．

Abstract:Abstract:［Objective］The aim of this study was to isolate bacterial strains with high-efficiency to degrade resins．［Methods］We used resin-plate to isolate resin-degrading bacteria from the formation water of Nanbao35-2 oil field，China National Offshore Oil Corporation． The morphological properties and the sequence homology of 16S rRNA were used to identify the strains． The changes of four fractions contents and the infrared spectrometry of the heavy oil were used to analyze the degradation properties． ［Results］Four strains，Q4，QB9，QB26 and QB36，were isolated using resin as the sole carbon source． Based on the high sequence similarities (more than 99%) of 16S rDNA sequences analysis． These strains were identified as member of the Petrobacter sp． ，Geobacillus stearothermophilus，Bacillus licheniformis，Geobacillus pallidus，respectively． QB26，a Bacillus licheniformis，was the most efficient strain，it can grow well under anaerobic conditions，emulsify heavy oil well，and degrade resin and asphaltene in heavy oil． The relative content of saturated hydrocarbons in heavy oil increase after degradation，and the relative content of resin and asphaltene in heavy oil decreased 5. 1% and 2. 7% ，respectively． ［Conclusion］ The strains isolated from Nanbao 35-2 oil field formation water could degrade resin and heavy oil． They have potential values in microbial enhanced oil recovery and oil pollution treatment．

Abstract:Abstract:［Objective］To obtain the recombinant BslA(260－652) protein of Bacillus anthracis and prepare its antibody for the adhesion activity studies．［Methods］The fragment coding BslA(260－652) was cloned into pET28a(+) plasmid and induced to express recombinant protein in E． coli Rosetta (DE3) by Isopropyl β-D-1-thiogalactopyranoside (IPTG)．The expressed recombinant soluble protein was purified by a column packed with Ni Resin． Purified protein was used as the antigen to immunize BABL/c mice for three times to raise polyclonal antibody． The adhesion activity of BlsA(260－652) was detected by immunofluorescence experiments and bacterial adherence assays．［Results］The purity of the purified soluble BslA(260－652) was about 87.4%．ELISA assay titer of antiserum from vaccinated mice reached 1:20000. Western blot showed the antiserum could specifically recognize endogenous BslA protein． The purified BslA(260－652) displayed a typical adhesion-like function． Either the anti-BslA serum or the BslA(260－652) protein could inhibit A16R's Hela adherence．［Conclusion］The recombinant BslA(260－652) protein was successfully obtained，which would lay the foundation for further research of the anthrax vaccine and the role of this S-layer protein in the pathogenesis of anthrax.

Abstract:Abstract:［Objective］To investigate the effects of canine parvovirus (CPV) non-structural protein-1 (NS1) on the cell apoptosis induced by CPV and preliminarily explore the mechanism of CPV-induced apoptosis． ［Methods］First，the NS1 gene was amplified by PCR from CPV genomic DNA and subcloned into pcDNA3. 1A vector to generate NS1 eukaryotic expression vector pcDNA-NS1． To verify whether pcDNA-NS1 vector can mediate NS1 expression in eukaryotic cells，the human embryo kideny (HEK) 293FT cells were used to transiently express the recombinant NS1． The effects of NS1 on CPV-induced apoptosis were investigated by infecting the F81 host cells with CPV and transfecting the cells with NS1vector． The apoptosis of the cells was detected by AnnexinV /PI double staining for phosphatidylserine externalization on membrane and by luminescence method for caspase-3 /7 activities． ［Results］The results show that the sequence of NS1 gene amplified was consistent with the GenBank． The NS1 expression vector was shown to be correct and could mediate NS1 expression in eukaryotic cells． The phosphatidylserine on outside of membrane was detected and the caspase-3 /7 activities were increased in both CPV-infected cells and NS1-transfected cells． These results indicate that both CPV and NS1 protein can induce the apoptosis of the cells． ［Conclusion］CPV-induced apoptosis was closely related to its nonstructural protein NS1.

Abstract:Abstract:［Objective］To construct prokaryotic expression vectors suitable for tandem affinity purification to study proteinprotein interactions in bacteria．［Methods］Two tandem affinity tag sequences，including the coding sequences of Protein G and streptavidin binding protein ( SBP)，as the N- and C- terminus of fusion proteins were designed and de novo synthesized．Constitutive expression vectors pNTAP and pCTAP were constructed using pUC18 as the backbone deleted of the lacI gene．［Results］Two expression vectors pNTAP and pCTAP were successfully constructed． pNTAP showed substantial expression of the built-in tag protein GFPuv not only in Escherichia coli BL21 (DE3) but also in enterohemorrhagic Escherichia coli O157:H7 and Shigella flexneri 5a．［Conclusion］Of the two recombinant expression vectors successfully constructed，pNTAP can express the model protein for tandem affinity purification and could be used for studies of protein-protein interactions in some gram-negative pathogenic bacteria such as Escherichia coli and Shigella flexneri．

Abstract:Abstract:［Objective］In order to investigate the composition and diversity of endolithic bacteria at special habitats in Xinjiang． ［Methods］Five rock samples were collected，including Wusu's granite (sample 1)，Glacier No.1，and Mulei's metamorphic rock (sample 2，sample 3),umin and Tokesun's Rock varnish (sample 4，sample 5)．Endolithic bacterial community composition and diversity were analyzed by the method of Terminal Restriction Fragment Length Polymorphism．［Results］Differences in diversity indexes among samples were not apparent． Clustering analysis suggested that similarity coefficient was higher in same rock type，sample 2 and sample 3 were grouped together，then sample 1 clustered with them，and sample 4 and sample 5 were classified together． All samples harbored these phyla such as Firmicutes，Actinobacteria，Proteobacteria and Bacteroidetes． Acidobacteria and Planctomycetes existed in sample 1 and sample 2，respectively; Sample 5 was dominated by Actinobacteria， while other samples were dominated by Proteobacteria．［Conclusion］The endolithic bacterial composition of same rock type collected at various habitats was different．Meanwhile，a diversity of novel species and lineages maybe existed in rocks．

Abstract:Abstract:［Objective］To isolate endophytic actinomycetes from earthnut samples of Stemona tuberosa Lour. collected from Xishuangbanna，Yunnan Province，and to study the classification，antimicrobial activity and secondary metabolic potential of the isolates． ［Methods］Samples were treated by a well-established surface-sterilized procedure，and plated on four selective media． The isolates were identified by morphology and 16S rRNA gene sequence analysis． Antimicrobial activity of the isolates was tested using agar diffusion method． Secondary metabolite genes of PKS /NRPS and halogense were detected by PCR amplification． HPLC-UV and VIS-ESI-MS /MS were used to analyze the fermentation product． ［Results］We obtained 18 actinomycete isolates from 6 Stemona earthnut samples． The isolates belonged to 4 genera，Streptomyces，Pseudonocardia，Micromonospora and Methylobacterium，and 10 of them were streptomycetes． The isolates also showed distinguish positive rates of antibacterial activity and secondary metabolite genes． Of them 13 strains showed antimicrobial activity against Staphlococcus aureus and/or Pseudomonas aeruginosa． Of them 17 isolates were positive for PKS/NRPS genes，and 8 strains were positive for halogenase gene. Moreover，the representative halogenase gene-positive strain produced metabolites that had typical MS peaks of halogen absorption． ［Conclusion］Based on the results of this study，the endophytic actinomycetes from Stemona，dominant by Streptomyces and Micromonospora，have good secondary metabolic potential and could serve as a promising resource for bioactive metabolite discovery in the future．

Abstract:Abstract:［Objective］To investigate the possibility of increasing the yield of hyaluronic acid by constructing duplication hasABC of chromosome recombinant in Streptococcus equi subsp． Zooepidemicus with a thermosensitive delivery vector system pJR700． ［Methods］We amplified a 4147 bp DNA fragment of hyaluronic acid synthase operon hasABC genes from chromatosome of S． zooepidemicus using PCR． This DNA fragment was subcloned into the pJR700 at ClaI sites to result in recombinant plasmid pXL32． The recombinant plasmid was transformed into S.Zooepidemicus by electroperation． The homologous recombination was induced by growing the bacteria at 37℃，and transformants were selected according to kanamycin resistance for 3 rounds． Then the culture was shifted to grow at 30℃ without antibiotics for 4 rounds to induce excision of the pJR700 indicated fragment． Colonies with kanamycin sensitivity were selected by plating on THY agar at 37℃．The hasABC recombinant of S． Zooepidemicus was identified through RT-PCR with primers homologous to the flanking regions．HA titers were measured by the modified carbazole assay． ［Results］We constructed successfully the duplication hasABC of chromosome recombinant of S.Zooepidemicus and the HA titer production by recombinants harboring duplication hasABC was 34% higher than that of the wild type at 24 h in shake flask culture.［Conclusion］The thermosensitive delivery vector of pJR700 could be used to construct the streptococcal hasABC recombinant strain for increasing the yield of HA in S. Zooepidemicus.