Abstract:Abstract: Haloalkane dehalogenases are key enzymes for biodegradation of halogenated aliphatic compounds，widely distributed in various microbial species of wide geographical distributions，and of significance in practical application such as bioremediation and industrial biocatalysis．Twenty haloalkane dehalogenases have been biochemically characterized so far．In recent years，progresses have been made in the enzymatic characteristics，protein structures，and phylogenetic diversity．We reviewed the progresses of haloalkane dehalogenase in structure and function，diversity and application potential．

Abstract:Abstract:Streptococcus suis (S. suis) is an important zoonosis and pathogen that can carry prophages．In this review，we focus on the recent advances in our understanding of lytic phage and lysogenic phage of S. suis，including the morphology of S. suis lytic phage，the functions of lysin and terminase large subunit encoded by S. suis lytic phage，comparative genomics of S. suis prophages，lysogenic conversion between S． suis lytic phage and prophage．Furthermore，prospective evolution of interactions between phage and host was discussed．

Abstract:Abstract:Baculoviruses are a family of arthropod-specific viruses that mainly affect insects of the orders Lepidoptera，Hymenoptera，and Diptera．In nature，baculoviruses establish infection in their hosts orally and a battery of proteins designated as per os infectivity factors play pivotal roles in baculovirus per os infection．This review summarizes the basic characteristics of baculovirus and discusses the main events that baculovirus establishes per os infection，including the evolutionary advantages for baculovirus to initiate infection through the oral route，the binding and fusion of baculovirus virions with insect midgut microvilli and the functional roles of baculovirus per os infectivity factors．These achievements and advances should promise to shed light on the understanding and utilization of baculovirus for bio-control and exogenous gene expression in the future.

Abstract:Abstract:［Objective］To identify bacterial strains with the inhibitory activity to rice pathogens，and to evaluate their potentials for the development of new biopesticides.［Methods］Ｒice rhizosphere Pseudomonas strains were isolated using 1-aminocyclopropane-1-carboxylic acid as the sole carbon source． Strain PA1201 was further identified through morphological analysis，biochemical characterization，16S rDNA sequence analysis and phospholipid fatty acid profiling．Qualitative and quantitative analysis of the production of the green pesticide Shenqinmycin as well as phenazine-1-carboxamide produced by PA1201 was done by HPLC．Cytotoxicity of PA1201 was evaluated using human alveolar epithelial cell line A549 and Drosophila melanogaster as hosts．［Results］Strain PA1201 inhibited Rhizotonia solani Kühn and Xanthomonas oryzae pv．oryzae，the causal agents of rice sheath blight and bacterial blight，respectively．It was further identified as Pseudomonas aeruginosa PA1201，which produces shenqinmycin and phenazine-1-carboxamide．The fermentation titer of shenqinmycin and phenazine-1-carboxamide in the PPM medium was 81. 7 mg/L and 18.1 mg/L，respectively．In the medium supplemented with soybean meal and corn steep liquor，the level of shenqinmycin and phenazine-1-carboxamide reached 926.9 mg/L and 489.5 mg/L．PA1201 also produced high level of extracellular protease and was toxic to human cell line and fruit fly．［Conclusion］Strain PA1201 could be engineered for higher yield of Shenqinmycin or for a new biopesticide．

Abstract:Abstract:［Objective］We characterized procaryotic biodiversity，community structure and the relationship between the community structure and environmental factors of salt lakes in Badain Jaran desert，Inner Mongolia，China.［Methods］We constructed 16S rＲNA gene clone libraries by molecular biology techniques to analyze the procaryotic phylogenetic relationships，and used R language to compare the community structure of haloalkalophiles in the salt lakes.［Results］Water in this region has a high salinity ranging from 165 to 397 g/L．The water is strongly alkaline with pH value above 10. The microbial diversity and community structure of the salt lakes are obviously different. The diversity of bacteria is more abundant than that of archaea．The main categories of bacteria in the samples are Gammaproteobacteria，Bacteroidetes，Alphaproteobacteria，Firmicute and Verrucomicrobia，whereas all archaea only belong to Halobacteriaceae of Euryarchaeota.［Conclusion］Salinity is the most important environmental factor influencing the bacterial community structure，whereas the archaea community structure was influenced comprehensively by multiple environmental factors.

Abstract:Abstract:［Objective］The aim of the study is to identify the pathogen causing adzuki bean (Phaseolus angularis) rust in Daqing，Heilongjiang province.［Methods］Adzuki bean rust leaves were collected from Daqing，Heilongjiang province，China. A pure culture of rust isolate ZXL01 was obtained by single pustule isolation．Its taxonomic status was determined by observing the number of germ pores of urediniospores，germ pore location and the wall thickness of teliopores，and sequencing ribosomal DNA internal transcribed spacer (rDNA-ITS)．［Results］Morphological studies showed that most of the urediospores of ZXL01 had two germ pores that were far from spores’equator area. The wall thickness of teliopores ranged from 2.9 to 3.3 μm．The rDNA-ITS sequence of ZXL01 was clustered in one clade with 2 reference isolates of Uromyces vignae (GenBank accession numbers AB115718 and AB115731) at 99% bootstrap levels in the phylogenetic tree. A 500 bp amplified product was obtained by the specific primers UV-ITSF/R，which was specific for U. Vignae．［Conclusion］The morphological features and ITS analysis indicated that the rust fungus ZXL01 occurred on leaves of adzuki bean in Daqing was U. Vignae，and the accession number of GenBank was KM461700．

Abstract:Abstract:［Objective］ Gram-positive brevibacterium Listeria monocytogenes (Lm) is an important zoonotic foodborne pathogen，engaged in both saprophytism and parasitism．It could adapt，survive and display pathogenicity under different environmental stress challenges，which is associated with the regulatory network consisting of regulating factors．The biological characterizations of regulator hfq was evaluated in this study.［Methods］hfq deleted serovar 1/2 a strain EGDe was constructed with homologous recombination，the biological characteristics of the mutant strain was compared with its parental strain.［Results］The growth of EGDe△hfq was significantly inhibited under cold temperature (P＜0.05)，salt medium containing 7% NaCl and the medium containing 4.5% ethanol．The ability of biofilm formation of the mutant strain in BactoTM Brain Heart Infusion(BHI) was reduced significantly (P＜0.05); notably，the invasion rate to Caco-2 cell lines was obviously reduced． Infection capacity of EGDe△hfq to BALB/c mice decreased and the LD50 was 6 times higher than EGDe．［Conclusion］Hfq protein of Listeria monocytogenes plays an important role in regulating bacterial virulence，biofilm formation and stress response．This deletion strain provided material to further study the function of Hfq and provides the possibilities to elucidate the mechanisms of Lm in resisting the stress and paves ways to the development of novel strategies for the prevention and control of Lm infections.

Abstract:Abstract:［Objective］ To accelerate the formation of nisin through overexpressing 6-phosphofructokinase gene pfk in nisin-producer Lactococcus lactis N8． ［Methods］The genes of pfk and pkaC encoding the catalytic subunit of cAMPdependent protein kinase were cloned into the vector pMG36e andtransformed into L．lactis N8，resulting in the recombinant strain L．lactis N8-pMG36e-pfk-pkaC．Several biochemical and physological factors，including growth profiles，activity of 6-phosphofructokinase，expression of nisA，antibacterial activity of supernatants and nisin titer，were monitored to investigate the differences between the recombinant strain and the parental strain.［Results］No significant difference was observed with respect to the growth patterns of the recombinant strain and the wild type．As expected，the biological activity of PFK in recombinant strain was increased for all examined samples．Correspondingly，the yield of nisin was increased by 20% in the recombinant strain after fermentation for 10 hours，which could be attributed to the accelerated biosynthesis of nisin．As a result，the fermentation cycle was reduced about 2 hours． Meanwhile，different concentration of glucose did not affect the formation of nisin．［Conclusion］The overexpression of pfk and pkaC genes in the nisin-producer strain can effectively accelerate nisin biosynthesis.

Abstract:Abstract:［Objective］ We characterizeda manganese catalase lacking n-terminal (MnCAT-C)，to revealits roles in bacterial growth，reactive oxygen species (ROS) removal and degradation of polychlorinated biphenyls (PCBs) in Rhodococcus sp. R04．［Methods］Manganese catalase (Mn-CAT) sequence of the strain R04 was aligned with that of Rhodococcus sp. R1101．Mn-CAT and MnCAT-C were expressed in E．coli BL21 (DE3)，and the target protein was purified with Q-sepharose and ammonium sulphate precipitation．Knockout strain was obtained by homologous recombination. ROS was measured by fluorescence polarization，and the degradation rate of PCBs was measured by HPLC．［Results］ MnCAT-C protein was purified，and SDS-PAGE analysis showed that its molecular weight was 23 kDa．Compared with wild strains，the ROS concentration increased，and the growth rate was inhibited in knockout strains．Moreover，the degradationrate of PCBs decreased．［Conclusion］MnCAT-C retained the majority of the active properties of the original enzyme，including ROS clearance．The lack of MnCAT-C gene affected the growth rate and the PCBsdegradationrate instrain R04.

Abstract:Abstract:［Objective］The objective of this study is to understand the accumulation of antibiotics，metals in the litter，and evolution features of bacterial antimicrobial-resistance during the fermentation of duck bio-bed.［Methods］ The experiment was conducted in meat duck bio-bed farm of Jiangsu province from November 2011 to July 2013. The new litter and spent litter from 4th，8th meat duck flock were studied for the accumulation of consumed antimicrobials and metals．Bacterial resistance levels to consumed antimicrobials were measured in the collected litter.［Results］The residues of doxycycline in the litter increased significantly after successive flocks of meat ducks，but ofloxacin had not been detected in all litter samples. The litter for the 8th meat duck flock had the highest level of resistant cultivable bacteria in the three sorts of medium amended with 16 μg/mL，100 μg/mL doxycycline．Meanwhile，the level of resistant cultivable bacteria in the same sort of medium amended with 8 μg/mL，50 μg/mL ofloxacin differed insignificantly between different flock litter sample． Different flock litter differed insignificantly in the accumulation content of As，Pb and Hg，and was low in Cd．The 4th，8th flock litter increased significantly in Zn and Mn than new litter，and increased slowly in Cu and Cr.［Conclusion］With the application of doxycycline in each flock of meat ducks，the content of doxycycline and doxycycline resistance in enterobacteriaceae bacteria in the litter increased significantly，Zn and Mn had an increasing trend overall.

Abstract:Abstract:［Objective］ To characterize a neutral invertase from Enterobacter cloacae GX-3.［Methods］ By searching GenBank database，we found the genes encoding invertase from the same genus Enterobacter. These sequences were aligned and analyzed. Then，a gene encoding neutral invertase was amplified by PCR. The recombinant plasmid pQE-Einv was constructed．We purified the expressed protein Einv with nickel-nitrilotriacetic acid chromatography．At last，the characterics of the recombinant protein Einv were studied in detail.［Results］ A gene encoding neutral invertase was discovered and cloned from E.cloacae GX-3. The recombinant enzyme Einv was characterized. Einv had an optimum pH of 6.5 and an optimum temperature of 40℃．The results of sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and gel permeation chromatography (GPC) showed that Einv was a homo-dimer protein．Einv retained 80% activity at sucrose concentrations up to 1170 mmol /L．But，Einv had no transglycosylation activity at high sucrose concentration．It could hydrolyze raffinose，1-kestose，nystose，fructofuranosylnystose and stachyose.［Conclusion］It is first reported that an invertase from Enterobacter cloacae is a β-fructofuranosidase at neutral pH range. It only has hydrolysis activity without tranglycosylation activity. These characteristics indicate that the neutral invertase Einv has important applications in food industry．

Abstract:Abstract:［Objective］To detect clustered regularly interspaced short palindromic repeats (CRISPR) in Shigella，and to analyze its relationship to drug resistance．［Methods］ Four pairs of primers were used for the detection of convincing CRISPR structures CRISPR-S2 and CRISPR-S4，questionable CRISPR structures CRISPR-S1 and CRISPRＲ-S3 in 60 Shigella strains．All primers were designed using sequences in CRISPR database． CRISPR Finder was used to analyze CRISPＲ and susceptibilities of Shigella strains were tested by agar diffusion method. Furthermore，we analyzed the relationship between drug resistance and CRISPR-S4.［Results］The positive rate of convincing CRISPR structures was 95%．The four CRISPR loci formed 12 spectral patterns (A-L)，all of which contained convincing CRISPR structures except type K．We found one new repeat and 12 new spacers. The multi-drug resistance rate was 53.33%．We found no significant difference between CRISPR-S4 and drug resistant．However，the repeat sequence of CRISPR-S4 in multi-or TE-resistance strains was mainly R4.1 with AC deletions in the 3' end，and the spacer sequences of CＲISPＲ-S4 in multidrug resistance strains were mainly Sp5.1，Sp6.1 and Sp7．［Conclusion］CRISPR was common in Shigella．Variations of repeat sequences and diversities of spacer sequences might be related to drug resistance in Shigella.

Abstract:Abstract:［Objective］ We studied the system immunofuctions of two Bifidobacterium strains isolated from food．［Methods］There were 10 SPF BALB /c mice in each group．The control group was given only sterile skim milk．The positive control group was given sterile skim milk containing commercial strain BB-12．The treatment group was given sterile skim milk containing different dosages of B．adolescentis BB-2 or B．longum BB-3．The immune parameters including cellular immunity (delayed-type hypersensitivity ［DTH］，splenic lymphocyte proliferation and natural killer ［NK］ cell activity)，humoral immunity (serum hemolytic activity in immunized animals)，and nonspecific immunity (peritoneal macrophages phagocytsis) were measured．［Results］Ingestion of B．adolescentis BB-2 or B．longum BB-3 could increase the DTH response． Macrophage phagocytsis was also enhanced，while activities of the NK cells and levels of the serum hemolysin were also significantly higher than that in the control group． There was a significant increase in splenic lymphocyte proliferation in bifidobacteria treated mice compared to the control．［Conclusion］ Ingestion of B．adolescentis BB-2 or B．longum BB-3 could enhance both innate and acquired immunity in healthy BALB/c mice．

Abstract:Abstract:［Objective］ To study the role of lymphotoxin-like inducible protein that competes with glycoprotein D for herpesvirus entry on T cells (LIGHT) in the development of protective immunity and pathology during Chlamydia Muridarum urogenital infection in mice.［Methods］C57BL /6J wild type (wt) and mice deficient in LIGHT (LIGHT KO) were inoculated intravaginally with 1×104 IFUs of live C．muridarum organisms． Half mice of each group were reinfected on day 49 after primary infection． We took mice vaginal swabs every 3 or 4 days to monitor live organism shedding．On day 80 after the primary infection，mice were sacrificed，the vaginal tract was isolated for pathology analysis．The spleen cells were collected and IL-4，IL-5，IL-17 and IFN-γ were detected by ELISA in the spleen cells culture supernatant after restimulated by UV-MoPn EB． The titers of different Ab isotypes were measured in mice serum by Indirect Immunofluorescence Assay．［Results］The chlamydia shedding time of LIGHT KO mice was similar to wild type mice，which cleared the organisms within 28 days after primary infection，and acquired protective immunity against C．muridarum reinfection． All mice regardless of genotypes developed severe upper genital tract pathology and showed no significant difference between LIGHT KO and wild type mice． All mice developed robust anti-C．muridarum organism IgG antibody responses and the ratios of IgG2a versus IgG1 showed no significant difference between LIGHT KO and wild type mice．Splenocytes from MoPn-infected LIGHT KO and wild type mice produced high levels of IFN-γ and IL-17，but IL-4 and IL-5 couldn’t be detected．［Conclusions］ LIGHT signal pathway may not correlated with protection against C．muridarum urogenital tract infection and urogenital tract pathology induced by C．muridarum.

Abstract:Abstract:［Objective］ To simultaneously detect antibodies against Duck hepatitis A type 1 (DHAV-1) and type 3 (DHAV-3) viruses，we developed an indirect enzyme-linked immunosobent assay (ELISA) with bacterially expressed recombinant viral protein as antigen in Escherichia coli． ［Methods］We amplified the full-length VP3 gene of DHAV-1 and the full-length VP1 gene of DHAV-3 through reverse transcription-polymerase chain reaction (RT-PCR) and then cloned them into pET-32a expression vector，designated as pET-1VP3- 3VP1．The fusion protein DHAV- 1VP3-3VP1 expressed correctly and was subsequently used to develop an indirect ELISA assay．［Results］DHAV-1VP3-3VP1 fusion protein expressed in BL21 ( DE3) cells following induction by Isopropyl-beta-D-1-thiogalactopyranoside (IPTG)．The expressed protein was very antigenic and reactive to virus-specific antibodies in western blot assay．The optimal working concentration for coating antigen was 1.0 μg per well and the working concentration of serum samples was 1:200 dilution and the cut-off value that distinguished the positive from negative serum samples was OD650≥0.38．［Conclusion］The ELISA method based on the prokaryotic expression of VP3 (DHAV-1) and VP1 proteins (DHAV-3) can be used effectively for the clinical detection antibodies against DHAV-1 and DHAV-3.

Abstract:Abstract:［Objective］Schizochytrium sp．is a marine fungus that can produce DHA efficiently．Genetic engineering has been successfully used in industrial strain improvement and metabolic studies．In order to use genetic engineering to modified Schizochytrium sp．，we established an genetic transformation system of Schizochytrium sp．［Methods］A genetic transformation system of Schizochytrium sp．was established by 18S rDNA-targeted homologous recombination．The targeting vector contained a part of 18S rDNA from Schizochytrium sp．and the ble gene． This targeting vector was transformed into Schizochytrium sp．by electroporation and then selected by Zeocin-containing plates．The incorporation of exogenous ble gene into the genome of Schizochytrium was inspected by PCＲ amplification．［Results］Fermentation results show that the transformants had similar cell dry weight，lipid yield，DHA content，and composition of other fatty acids to the wild type strain． ［Conclusion］Our results show that the introduction of resistance gene did not affect the cell growth and lipid metabolism． This system could be used to introduce new functional genes into Schizochytrium sp．．