Abstract:[Objective] The aim of study was to investigate Actinobacterial diversity in Xiaoerkule salt lake, to lay a foundation for furthering to tap. [Methods] Actinobacterial diversity in this sediment from Xiaoerkule Lake was investigated by culture-independent method and phylogenetic analysis based on 16S rRNA gene sequences. Total DNA of sediment sample was extracted using SDS-CTAB method. The primers for the class Actinobacteria were used for actinobacterial 16S rRNA gene amplification and then a clone library was constructed for the sediment sample. [Results] Fifty-one clones screened from 160 clones on the basis of Hae III digestion patterns were sequenced, and their sequences were deposited in the GenBank. Clone sequences (52.9%) belonged to Acidimicrobidae and 5 suborders of Actinobacteridae. The other clone sequences (47.1%), which formed one large distinct clade in phylogenetic tree among phylum Actinobacteria, may represent one new suborder or new class. [Conclusion] There was abundant actinobacterial diversity in the sediment of Xiaoerkule Lake, and the result implied that there were large numbers of unknown actionobacterial groups here.

Abstract:[Objective] We determined the distribution of serogroups and virulence factors among Escherichia coli isolates from pigs with diarrhea and/or edema disease from 10 provinces in China between 1998 and 2007. [Methods] Three hundred E. coli isolates were serogrouped with O-antisera and detected for genes of enterotoxins, shiga-toxin-two-variant (Stx2e), intimin, K88, K99, 987P, F18 and F41 fimbrial antigens by PCR. [Results] Among 300 isolates, 233 were determined to be placed in serogroups, 50 were unable to be serogrouped, and the rest 17 auto-agglutinated. These isolates distributed in 45 serogroups and 57.1% (133/233) belonged to 5 O serogroups: O149, O107, O139, O93 and O91. Several uncommon O serogroups were discovered. PCR analysis confirmed the genes of estⅠ(STa), estⅡ(STb), elt (LT), stx2e (Stx2e) and eaeA (Intimin) within the isolates. There were six pathotypes of porcine E. coli by their pathogenic factors, namely enterotoxigenic E. coli (ETEC), shiga toxin-producing E. coli (STEC), attaching-effacing E. coli (AEEC), ETEC/ STEC, AEEC/ETEC and AEEC/ETEC/STEC. The correlation between pathotypes and serogroups showed that the combinations of O149/K88/estⅡ, O149/K88/estⅡ+elt, O107/F18/estⅡ, O93/estⅡand O98/estⅡwere common types in ETEC strains, O139/ F18/stx2e for STEC strains, eaeA gene for AEEC strains, O107/F18/estI+stx2e, O107/F18/estⅡ+ stx2e and O116/F18/estⅠ+elt+stx2e for ETEC/STEC strains, O91/estⅠ/eaeA and O107/estⅠ/eaeA for AEEC/ETEC/STEC strains.”[Conclusion] ETEC was commonly associated with swine colibacillosis, whereas the kinds of enteric E. coli pathogens become more complex in China.

Abstract:[Objective] Iron is an essential element for bacterial survival and pathogenesis. Using a random promoter library we screened for iron-responsive genes in P. aeruginosa genome. [Method] Plasmid pMS402 containing a promoterless luminescence reporter, LuxCDABE, was used as the vector to construct a P. aeruginosa random promoter library with a size of about 5700 clones. It enabled real-time, in vivo high-throughput gene expression profiling at genomic scale. [Results] Two previously reported iron-regulated genes were identified, and ten new iron- regulated genes were uncovered. The genes encoding dihydrolipoamide acetyltransferase, phosphogluconate dehydratase and Fe (Ⅲ) dicitrate transporter were found to be iron-regulated together with four function-unknown genes and three putative protein encoding genes. [Conclusion] These results provide a basis for elucidating the complex iron regulation network in P. aeruginosa and help our understanding of the roles of iron in bacterial physiology and pathogenesis. The random promoter library system also offers a useful tool in bacterial genomic studies.

Abstract:[Objective] Biofilm plays an important role in pathogenicity of Salmonella and food poisoning caused by Salmonella. Our aim was to identify genes associated with Salmonella biofilm formation. [Methods] Seventy-four strains of Salmonella enterica serovar enteritidis (S. enteritidis), S. pullorum and S, gallinarum isolated from chickens were determined for biofilm using crystal violet staining, and the strain C050041 with well biofilm formation was chosen to construct a mutant library using transposon mutagenesis. [Results] Eighty-four percent of these Salmonella strains produced biofilm on the plastic surface. We screened 1924 mutants with transposon insertion, and 15 inserted genes were identified by growth curve determination of mutants, sequence analysis of the chromosomal DNA, and further confirmed by southern blot. These genes included metE, ompR, rpoS, rfaG, rfaJ, rfaK, rfaP, rfbH, rhlE, spiA, steB, tpx, ybdN and other two genes with unknown function. [Conclusion] We identified some new genes associated with Salmonella biofilm formation, these findings may help understand the regulation mechanism of biofilm formation and develop an attenuated Salmonella vaccine.

Abstract:[Objective] This study aimed at increasing the a-ketoglutaric acid production of a multi-vitamin auxotrophic yeast Torulopsis glabrata, by increasing the availability of acetyl-CoA. [Methods] For this, we expressed ACS2 encoding acetyl-CoA synthase from Saccharomyces cerevisiae in the pyruvate producer Torulopsis glabrata WSH-IP303. [Results] Compared with that of the parent strain, the acetyl-CoA synthase activity of the mutant ACS2-1 increased about 920% and the mutant could use acetate as the sole carbon source for growth (2.6 g/L dry cell weight). When growing with glucose, the acetyl-CoA concentration, a-ketoglutaric acid, and the value of C a-KG/Cpyr were 222%, 105% and 152% higher than those of the parent strain WSH-IP303, respectively. The addition of 4 g/L acetate to the culture broth of mutant ACS-1 led to a significant increase of these values to 355%, 147% and 275%, respectively, compared with that of the parent strain WSH-IP303. [Conclusion] The a-ketoglutaric acid concentration reached 17.8 g/L by increasing the availability of ace-tyl-CoA and this strategy may provide an alternative approach to enhance metabolite production in yeast.

Abstract:[Objective] To evaluate the antibacterial activity of lactic acid bacteria community SFC-2 in fermented crop straw. [Methods] Total 13 isolates were obtained from spontaneous fermented rice straw by plating, paper diffusion and Denaturing Gradient Gel Electrophoresis. All these strains were used to determine the antibacterial activity of SFC-2. [Results] (1) Phylogenetic analysis of the 13 strains based on 16S rDNA gene sequence data indicated that 9 strains belong to Enterobacter cloacae, Klebsiella oxytoca, Bacillus cereus, Klebsiella pneumoniae, Citrobacter freundii, Klebsiella terrigena and Citrobacter sp., which were all common pathogens or opportunistic pathogens. (2) Indicating bacteria selected from the pathogens were used to detect antibacterial activity of SFC-2 cell-free culture supernatant. The results showed that: SFC-2 community had stronger antibacterial activity than isolated strains from SFC-2 community or man-made communities against indicating bacteria. (3) Antibacterial activities of seven different cell-free culture supernatants, which were extracted at intervals from the culture of SFC-2 community during 14-48 hours, were no obvious difference, but the content of organic acids were obvious differences during 14-48 hours; the antibacterial substances were stable after heating and sensitive partly to protease K.

Abstract:[Objective] This study was to evaluate the effect of drying temperature for solid substrate after fermentation on conidia characteristics of entomopathogenic fungus Beauveria bassiana. [Methods] Seven constant or varied temperatures between 28oC and 35oC were designed for drying the solid substrate and the quality of harvested conidia was analyzed. [Results] The results showed that the drying treatments at varied temperatures significantly decreased bacterial contamination in the harvested conidia powder. The conidia viability and germination speed were varied with different drying treatments. After drying at 35℃ for 5 h, there was no significant difference in viability between the dried and fresh conidia, while the median germination time (9.6 h) of the dried conidia was shortened by 9.3%. The tolerance of conidia to heat and UV radiation was increased by drying treatment. Compared to the drying treatment at a constant temperature at 28℃ or 35℃, some varied temperature treatments were in favor of enhancing the stress tolerance of conidia. Drying treatments influenced accumulation of trehalose in harvested conidia, while neither heat resistance nor UV tolerance of conidia was obvious correlation with trehalose level. Optimizing drying temperature could increase the virulence of B. bassiana. After drying at 28℃ for 24 h and then 35℃ for 2 h or at 35℃ for 5 h, the LT50s to Myzus persicae at the dose of 370-450 conidia /mm2 were shortened by 10.6 h and 7.5 h, respectively. [Conclusion] The results suggested that the drying temperature for post-fermentation solid substrate has an important influence on bacterial contamination in the harvested conidia powder, spore viability, stress tolerance and virulence in B. bassiana.

Abstract:Lipoproteins LipL32 and LipL21 and transmembrane protein OMPL1 have been confirmed as the superficial genus-specific antigens of Leptospira interrogans, which can be used as antigens for developing a universal genetic engi-neering vaccine. [Objective] In order to obtain high expression of an artificial fusion gene lipL32/1-lipL21-ompL1/2, we optimized prokaryotic expression conditions. [Methods] We used surface response analysis based on the central compos-ite design to optimize culture conditions of a new antigen protein by recombinant Escherichia coli DE3.The culture con-ditions included initial pH, induction start time, post-induction time, Isopropyl b-D-thiogalactopyranoside (IPTG) con-centration, and temperature. [Results] The maximal production of antigen protein was 37.78mg/l. The optimal culture conditions for high recombinant fusion protein was determined: initial pH 7.9, induction start time 2.5 h, a post-induction time of 5.38 h, 0.20 mM IPTG, and a post-induction temperature of 31℃.[Conclusion] Surface response analysis based on CCD increased the target production. This statistical method reduced the number of experiments required for optimization and en-abled rapid identification and integration of the key culture condition parameters for optimizing recombinant protein expression.

Abstract:[Objective] Endophytic actinomycete Lj20 with antifungal activity was isolated from the roots of capsicum plants. We identified Lj20 and synthesized its antifungal substances. [Methods] Morphological, biological and biochemical characteristics, chemotaxonomy analysis and 16S rDNA sequences were used to identify Lj20. According to GC-MS extrapolation result, one of antifungal substances in the metabolites of Lj20 was chemically synthesized. The bioactivities were determined by mycelium growth inhibition method. [Results] Lj20 belonged to Streptomyces sp. and was similar to Streptomyces rochei. The metabolites contained butylated hydroxytoluene and 3, 5-di-tert-butyl-4-hydroxybenzyl methyl ether. The median effective concentration (EC50) of these two compounds to Botrytis cinerea Pers. were 237.04 mg/L of the water and 186.48 mg/L of the water, respectively. [Conclusion] Lj20 was classified as Streptomyces rochei. Butylated hydroxytoluene and 3, 5-di-tert-butyl-4-hydroxybenzyl methyl ether had significant inhibition to the pathogen.

Abstract:[Objective] To analyze the function of htpG of S. flexneri 2a 2457T, we constructed an htpG deletion mutant and a recovery mutant. [Methods] g-Red recombination system was used to construct an htpG deletion mutant of S. flexneri 2a 2457T. In addition, a recover mutant was obtained by introducing a low-copy plasmid containing one copy of htpG gene into the deletion mutant. Then, the growth curves of wild-type strain, deletion mutant and recover mutant were measured. Some of biochemical tests were also investigated. Furthermore, the Sereny tests were performed to evaluate the virulence of these strains. [Results] No significant difference were observed among three strains. However, the titers of some inflammatory factors evoked by wild-type strain, deletion mutant and recovery mutant in intraperitoneal injected mice were quite different. [Conclusion] These results suggest that the HtpG protein of Shigella flexneri 2a strain 2457T might be involved in the immunopathogenesis.

Abstract:[Objective] Tetrodotoxin and its analogues (TTXs) were responsible for the poisoning incidents associated with snail Nassarius spp. We studied bacteria isolated from toxic snails as well as their habitat to probe into the relationship between bacteria and toxicity of nassariid gastropod. [Methods] Two snail samples were collected from Sheyang, Jiangsu Province on June 13 and 19, 2006, and the toxicity of the snail samples was tested with mouse bioassay method. Bacteria were isolated from the snail samples and from their habitat. A part of isolated bacteria were then cultured in the lab, and TTX in bacteria were screened with an ELISA method. [Results] The snails collected were identified as Nassarius semiplicatus, and the toxicity of the 2 samples were 247 mouse unit (MU) and 270 MU / 100g tissue (wet weight), respectively. TTX was detected in 9 strains among the 14 strains of bacteria isolated from the snail samples and their habitat. TTX content in the toxic strains was very low, which ranged from 15 ng/g to 96 ng/g. Partial of the 16S ribosomal DNA (rDNA) of the toxic strains were then sequenced after PCR amplification, and the toxic strains of bacteria were tentatively identified based on the alignment of these sequences with published data. Toxic strains were closely affiliated with Vibrio, Shewanella, Planococcus, Marinomonas, Photobacterium. [Conclusion] These findings suggested that TTX-producing bacteria may play an important role in TTX production or accumulation in nassariid gastropod.

Abstract:[Objective] We studied root nodule proliferation, nodule microstructure, genetic cluster and stress resistance of the rhizobium of Trigonella arcuata. [Methods] We characterized root nodule and rhizobium with various soil matrixes cultivation, paraffin section, resin semi-ultrathin section techniques, and 16S rRNA gene cluster analysis. [Results] ① Plants grew in mixed soil (nutritious garden soil∶poplar zone soil∶desert sands=1∶1∶1), had the most nodule prolif-eration and bore the most pods. The shapes of nodule were palm- or ginger-like; ② Microstructure of the nodule revealed five different parts differentiated within the nodule: epidermis(E), cortex(C), vascular bundle(VB), infected cells(IC) and uninfected cells(UIC); ③ Genetic cluster analysis of the full length 16S rRNA gene sequence (1377 bp) indicated that the rhizobium isolated shared the highest identities with Sinorhizobium meliloti; ④ The rhizobium could grow between 4 and 60℃ (20 min), pH 6.0~12.0 and 0~2%NaCl. For the antibiotic sensitivity, the rhizobium could not grow normally in me-dium with 25 mg/mL Kanamycin, Streptomycin or Cephalothin, except for 100 mg/mL Ampicillin. [Conclusion] Good conditions of soil matrixes were important for nodulation of T. arcuata; A large quantity of cells in fascicular nodules were infected by rhizobia; 16S rRNA gene sequence of T. arcuata shared the highest identities with that of Sinorhizobium meliloti, and this strain was able to tolerate relatively higher temperature and alkalin.

Abstract:[Objective] To characterize hematolysis of Enterococcus from sheep. [Methods] Using plate assay (PA), contact hemolysis (CH), supernatant assay (SA), culture hemolysis (CLH) and PCR, we studied hemolysis characteristics of 11 Enterococcus clinical isolates, 30 isolates from healthy sheep, a standard G-Streptococcus and a standard Enterococcus. [Results] Rabbit blood and sheep blood were not hemolysising in the 11 clinical isolates analyzed by SA and CH. Of the clinical isolates 63.6% had b-hemolysis with rabbit blood and 36.4% had a-hemolysis with sheep blood analyzed by PA and CLH assay. Of the cylA gene 63.6% was detected in clinical isolates, the sequence of cylA gene was 99.3% homolo-gous with cylA of plasmid pAD1(GenBank accession number: L37110). b-hemolysis had 53.3% in rabbit blood, a-hemolysis and b-hemolysis in sheep blood had 53.3% and 43.3% respectively in 30 healthy sheep initial isolates with PA. Only 6% had hemolytic capacity in rabbit blood after second generations. The cylA gene was not detected in 30 healthy sheep isolates by PCR. Standard Enterococcus strain of a-hemolysis of sheep blood had no hemolysis of rabbit blood. [Conclusion] The red blood cells could induce enterococci hemolysis secreting in the bacteria growth. The result was different with the Phenotype and the Genotype.

Abstract:[Objective] Deoxynivalenol (DON) is a trichothecene mycotoxin produced by Fusarium graminearum, a pathogen causing Fusarium Head Blight of wheat. It is necessary to establish a rapid and simple assay to detect DON. [Methods] High affinity monoclonal antibodies (Mab) against DON were produced by cell fusion with 500 mg/mL Polyethylene Glycol 400, and cell sub-cloning in HAT (H: hypoxanthine, A: aminopterin; T: thymidine) culture medium for screening and limiting dilution. Hybridoma lines were screened for specificity to DON by Enzyme-linked Immunosorbent Assay (ELISA). One hybridoma cell line (3B2) secreting monoclonal antibody (MAb) against DON was produced by fusing mouse myeloma cells (SP 2/0) with spleen cells from BAL B/C mice which were immunized by the artificial antigen conjugated with Ovalbumin (OVA). [Results] The MAb obtained in this experiment could specifically react with DON without cross-reactivity to DON related compounds except 3-acetyldeoxynivalenol (3-Ac-DON), with the titres of ascitic fluids up to 1× l0-7 by indirect ELISA. Isotype and subclass of the monoclonal cell line (3B2) showed that it belonged to IgG1. The light chain of the MAb was identified to be κ. Ascites antibodies generated by hybridoma of 3B2 cells were purified. Inhibition rate studies showed that the detection limit of the ELISA was 8 mg/L, with the regression equation of indirect ELISA Y=0.733lg(x)-0.572, R2=0.9652, IC50 value being 29ug/L. [Conclusion] The MAb can be used to prepare the reagents for analyzing DON residue.

Abstract:[Objective] Outbreaks of highly pathogenic avian influenza (HPAI) virus has caused great economic loss to the poultry industry and resulted in human deaths in Thailand and Vietnam since 2004. Rapid typing and subtyping of viruses, especially HPAI from clinical specimens, are desirable for taking prompt control measures to prevent spreading of the disease. We described a simultaneous approach using microarray to detect and subtype avian influenza virus (AIV). [Methods] We designed primers of probe genes and used reverse transcriptase PCR to prepare cDNAs of AIV M gene, H5, H7, H9 subtypes haemagglutinin genes and N1, N2 subtypes neuraminidase genes. They were cloned, sequenced, reamplified and spotted to form a glass-bound microarrays. We labeled samples using Cy3-dUTP by RT-PCR, hybridized and scanned the microarrays to typing and subtyping AIV. [Results] The hybridization pattern agreed perfectly with the known grid location of each probe, no cross hybridization could be detected. Examinating of HA subtypes 1 through 15, 30 infected samples and 21 field samples revealed the DNA microarray assay was more sensitive and specific than RT-PCR test and chicken embryo inoculation. [Conclusion] It can simultaneously detect and differentiate the main epidemic AIV. The results show that DNA microarray technology is a useful diagnostic method.

Abstract:[Objective] The purpose of this study is to find new molecular targets for the detection of Salmonella. [Methods] Homology comparison of genomic sequences in GenBank among Salmonella serovars and the specificity comparison among Salmonella and non-Salmonella strains were done with the BLAST (Basic Local Alignment Search Tool Program). We randomly selected 15 target sequences to design and synthesize 27 sets of primers for the evaluation of specificity and sensitivity and development of PCR methods. [Results] Primer FS23 was the best in specificity and sensitivity among these 27 sets of primers. The 44 Salmonella strains produced a specific 492-bp amplification band, whereas 22 non- Salmonella strains did not using primer FS23 for PCR detection. The detection sensitivity of FS23 was 11.9 fg/mL for DNA templates and 4.9×102 cfu/mL for whole cells. Salmonella could be detected successfully by the PCR method developed in this study after 5 h enrichment when the milk samples were artificially contaminated by this organism at 100 cfu/25 mL. [Conclusion] The sequences against FS23 is a new and excellent molecular target for the detection of Salmonella. The FS23-based PCR assay is sensitive and specific, and can be used for rapid detection of Salmonella in foods.

Abstract:[Objective] To develop a rapid PCR method to detect pathogenic Aeromonas hydrophila in fish. [Methods] For multiple PCR, three pairs of primers were designed based on the conservative sequences of 16SrRNA genes, aerolysin (aer) gens and serine-protease (ahp) genes of Aeromonas hydrophila. By optimization of PCR conditions and estimation of specificity, sensitivity, detection rate, a triplex PCR assay was established. [Results] The assay had a high specificity detecting only pathogenic strains of Aeromonas hydrophila but not other irrelative bacteria. The assay had a high sensitivity with the detection limit as low as 100fg, the detection rate of suspicious clinical samples by this assay was 81.8%, which was noticeably higher than that by bacterial isolation method (40.9%). The detection rate of mimic challenge samples by this assay was 87.5%, which was also noticeably higher than that by bacterial isolation method (67.5%). [Conclusion] The simultaneous detection of 16SrRNA gene and two virulent genes in one PCR assay could avoid missed detection possibly caused by PCR with single virulent gene, and provided a useful tool for rapid diagnosis, large-scale quarantine, and epidemiological investigation of the pathogenic Aeromonas hydrophila.

Abstract:[Objective] To isolate new actinomycetes for discovering compounds of pharmaceutical importance. [Methods] We collected 280 soil samples from virgin forest in Wuling Mountain. In total 1134 actinomycetes were isolated by culture-dependent method. Of them 30 strains were selected and then characterized by phylogenetic analysis based on sequences of 16S rRNA gene. Antimicrobial activities were determined by agar well diffusion method, and genes of typeⅠandⅡpolyketide synthases (PKSⅠ, PKSⅡ), nonribosomal peptide synthase (NRPS) and polyene cytochrome P450 hydroxylase (CYP) were screened by PCR. [Results] Thirty strains belonged to 8 families, 13 genera: Streptomyces (>70%), Micromonospora, Dactylosporangium, Catellatospora, Sphaerisporangium, Streptosporangium, Actinomadura, Nonomuraea, Nocardia, Rhodococcus, Arthrobacter, Microbacterium, and Pseudonocardia. There are 5 novel species candidates. Thirty strains showed different antimicrobial activities against one to three bacteria and pathogenic fungi. [Conclusion] Abundant diversity of actinomycetes existed in the virgin forest of Wuling Mountain which has many new taxa, and Streptomyces is the predominant population. The strains showed high antimicrobial activities and can be further researched for exploiting new compounds of pharmaceutical.

Abstract:[Objective] Studying the genes involved in swimming and twitching motility of Pseudomonas aeruginosa. [Methods] We used Mu transposition technique, gene cloning, nucleotide sequencing and the trans-complementation experiment to study the genes involved in twitching motility and swimming motility of P. aeruginosa strain PA68 isolated from a patient with bronchiectasis. [Results] A mutant deficient in both swimming and twitching motility was isolated out of about 2000 mini-Mu insertion mutants. The result of GenBank BLAST showed that the mini-Mu transposon had inserted into the gene PA1550 with unknown function. Analyses on the operon of PA1550 and the polar effect revealed that Mu transposon had no effect on the transcription of the downstream genes of PA1550. [Conclusion] PA1550 is involved in the swimming and twitching motility of Pseudomonas aeruginosa.

Abstract:[Objective] To investigate the mechanism of fatty acids, lipid A and N-acylhomoserine lactones biosynthesis of bacteria by using high quality Escherichia coli holo-ACP and varied length chain acyl-ACPs as substrates. [Methods and Results] Using PCR technique we amplified the acpP and acpS gene fragments from genomic DNA of E. coli strain MG1655. Ligating these gene fragments with plasmids pBAD24 or pET28b respectively, we obtained 3 expression plasmids of acyl carrier protein: pBAD-ACP, pET-ACP and pET-ACP-ACPS, and one expression plasmid of holo-acyl carrier protein synthase: pBAD-ACPS. Then we constructed 3 acyl carrier protein producer strains: DH5a/pBAD-ACP、BL21 (DE3)/pET-ACP and BL21(DE3)/pET-ACP-ACPS by transforming E. coli strains DH5a or BL21(DE3)with pBAD-ACP, pET-ACP or pET-ACP-ACPS, respectively. Although these 3 strains could produce more acyl carrier protein under induc-tion than strain DK574, which was used to purify holo-acyl carrier protein in general, the yield of holo-acyl carrier protein of these strains was still lower. In order to increase the yield of holo-acyl carrier protein in these strains, we introduced pBAD-ACPS into these strains. The assay of expressions of new strains was shown the that strain DH5a harbored pBAD-ACP and pBAD-ACPS double plasmids produced more holo-acyl carrier protein than strain DK574, and the purity of holo-acyl carrier protein was also increased (up to 99%). Then we purified high quality holo-acyl carrier protein from the culture of the strain DH5α harbored pBAD-ACP and pBAD-ACPS by using UNOsphere Q anion-exchange chroma-tography. Utilizing holo-acyl carrier protein and long chain fatty acids as substrates and under Vibrio harveyi acyl-acyl carrier protein synthetase catalyzing, we synthesized several different acyl-acyl carrier proteins. [Conclusion] From this study we obtained a high holo-ACP producer strain and demonstrated that co-expressing acpP with acpS, E.coli strains could produce more holo-ACP.

Abstract:[Objective] The purpose of this research is trying to uncover the biosynthetic or metabolic relative protein of thuringiensis by proteomics. [Methods] We conducted differential expression analysis between the high thuringiensin-yield Bacillus thuringiensis strain CT-43 and its mutants, high production strain CT-43-1C and non-production strain BMB0806, based on the 2-D gel. Then through mass spectrum (MS) identification and bio-informatics we analyzed the detected proteins. [Results] Thirteen spots were selected to be detected by the MS, and nine of them were identified. Among them, six proteins including in the basal metabolism pathway were possibly deduced that relative with the synthesis or metabolism of thuringiensin production. [Conclusion] There were six proteins found to be connected with the influential role protein of biosynthesis and metabolic of thuringiensin production by comparative proteomics. This research provides the evidence of thuringiensis gene cluster cloning and biosynthetic analysis.

Abstract:Previous research has suggested that the true producers of numerous natural products isolated from marine invertebrates were the microbial epibiont and symbiont which are deemed as not-yet-cultivated microbe. Cloning of the biosynthetic genes responsible for a specific nature product not only provides direct genetic evidence of the origin of the compounds but also establishes the feasibility of mass production of the compounds by heterologous expression. This paper reviews the progresses on the biosynthetic gene clusters of nature products from the symbiotic bacteria including marine sponge, ascidian, bryozoan, deep-sea tube worm and deep-sea sediments.

Abstract:Galacto-oligosaccharides are among the most promising non-digestible oligosaccharides that are recognized as prebiotics. Commercial GOS are synthesized from lactose using the transglycosylation activity of b-galactosidase from microorganisms. The structure of GOS varies with different enzyme source. The oligosaccharide yields of GOS produced by natural enzymes are generally 20%~45% and they could be improved by artificial enzyme. Reaction conditions also have effect on the oligosaccharide yield. Using enzymes in water-hydrophobic solvent mixtures or reverse micelles may improve the yield to some extent. GOS can be large-scale synthesized by packed bed reactor, plugflow reactor or membrane reactor. The glucose and lactose in the GOS products can be removed by using chromatography, enzyme oxidation, nanofiltration membrane or microbial fermentation.

Abstract:Canine distemper (CD) caused by Canine distemper virus (CDV) was first reported in 1905, and has been one of the most serious contagious diseases of dogs as well as other carnivores. Recently, increasing cases of canine distemper (CD) both in vaccinated and unvaccinated dogs and in wildlife have been reported in Japan, America, Europe and Africa. Based on phylogenetic analysis of the hemagglutinin (H) gene sequences, six genotypes of CDV were distinguished. Antigenic heterogeneity of the H protein that provides an important protective antigen against CDV infection has been observed between wild-type CDV and vaccine strains. So it was suspected that the vaccines currently used can no longer efficiently protect animal from present-day circulating CDV infection. The host range of CDV includes all species of the families Canidae and many other species. Both signaling lymphocyte activation molecule (SLAM) and heparin sulfate (HS) expressed on the cells of the immune system or other non-lymphoid tissues can act as the cellular receptors for CDV, and are one of the major determinants of the host range and tissue tropism. In this review, we discussed the above-mentioned issues based on the recent research progress and the studies in our laboratory.