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Abstract:[Objective] In order to indicate the relationship between soil microbial quantity and soil factor. [Methods] Selected 11 typical plots under different saline-alkali soils in Hexi Corridor in spring, the soil microbial quantity, the enzyme activity and the physicochemical properties were investigated and variance analyzed, simple correlation analyzed, stepwise regressive analyzed and principal component analyzed. [Results] It shows that there were obvious differences between the primary saline-alkali soil, the secondary saline-alkali soil and farmland soil in soil physicochemical properties and soil microbial quantity. The soil microbial quantity was generally at a low level and at regularity: primary saline-alkali soil

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Abstract:[Objective] The rhizobacterium Pseudomonas sp. M18 can produce two antibiotics, phenazine-1-carboxylic acid (PCA) and pyoluteorin (Plt). PsrA belongs to the TetR family bacterial transcription regulator. To investigate the effect of PsrA on the PCA and Plt biosynthesis, the psrA gene was amplified from M18 strain genome. [Methods] The psrA mutant (M18psrA) was constructed through gene replacement of psrA by gentamicin resistance cassette and homologous recombination. Gene complement experiment and lacZ gene fusion report analysis further identified whether PsrA regulates the expression of antibiotics biosynthesis gene clusters. [Results] PCA and Plt production was respectively assayed in PPM and KMB media. PCA production of psrA mutant was significantly lower than that of its parent strain M18. However, Plt production of M18psrA was obviously increased 10?15 times as much as that of M18 strain. Gene complement experiment and lacZ gene fusion report analysis further confirmed that PsrA strongly up-regulated PCA biosynthesis and down-regulated Plt biosynthesis. [Conclusion] PsrA regulates distinctively on the PCA and Plt biosynthesis.

Abstract:[Objective] In Pseudomonas sp. M18, there are nine structural genes encoding pyoteorin (Plt) biosynthase, including pltLABCDEFG and pltM. In order to identify the gene incoding rate-limiting enzyme, 9 structural genes were overexpressed. [Methods] The entire ORFs of these nine genes were PCR amplified from the M18 strain chromosomal DNA template and respectively cloned into the Pseudomonas-E. coli plasmid pME6032, generating nine overexpression plasmids of plt genes. The M18 strains, carrying these overexpression plasmids, were assayed for Plt production in KMB media. [Results] It was showed that the Plt production of three strains respectively harboring pltC, pltD, pltF overexpression plasmids was raised by 96%, 78%, 75% as compared to the control. The optimal inducing concentration of IPTG was 1.0 mmol/L, and the optimal inducing time was at 6 hours after inoculation. [Conclusion] The genes pltC, pltD, pltF divergently encode polyketidesynthase, halogenase and acyl-CoA synthetase rate-limiting enzyme in Plt biosynthesis. At the optimal IPTG concentration and inducing time, pyoluteorin production of strains containing pltD, pltF overexpression plasmids was respectively increased by 77.5% and 159.1%.

Abstract:[Objective] The pressure of fermenter was enhanced in order to increase the concentration of vitamin B12. [Methods] Metabolic flux analysis (MFA) was used to analyse the fermentation process of vitamin B12 production by Pseudomonas denitrifican. [Results] The results indicated that the metabolic flux of oxaloacetic acid from phosphoenolpyruvate carboxylation was increased when the synthesis rate of vitamin B12 increased, in order to meet the demand for precursor supply to vitamin B12 production. A novel control strategy was designed as following. The pressure of fermenter was enhanced in order to increase the solubility of carbon dioxide in aqueous solution at steady phase of fermentation, so more carbon dioxide was carboxylated to oxaloacetic acid. The result showed that the concentration of vitamin B12 was 176 mg/L, 19.7% higher than the control batch (147 mg/L) at 160 hours. [Conclusion] the production of vitamin B12 by Pseudomonas denitrifican was improved by increasing the fermenter pressure.

Abstract:[Objective] We constructed an Escherichia coli strain displaying an active lipase on the cell surface by cell surface engineering and characterized the displayed lipase. [Methods] Escherichia coli surface display vector of lipase was constructed by fusing sequence encoding the N-terminal domain of ice nucleation protein with Serratia marcesens lipase gene, and the recombinant vector was transformed into Escherichia coli BL21(DE3). [Results] The highest lipase activity was observed when the recombinant cells were induced with 0.05 mmol/L isopropy-β-D-thiogalactoside (IPTG) and cultured at 25 °C for 16 h. Optimal pH and optimal temperature for cell surface-displayed lipase was 9.0 and 40 °C, respectively. The thermal stability of surface-displayed lipase was improved compared with that of free lipase, and the residual activity was above 90 percent of initial activity after incubated at 40 °C for 1 h. [Conclusion] The above results suggest that the bacterial surface display technology offers a promising alternative approach for immobilization of lipases.

Abstract:[Objective] To analyze the genetic characteristics and dominant types of Legionella isolated from central air-conditioning cooling towers in public places in Guangzhou from 2008 to 2010. [Methods] Genomic DNA was extracted from 140 strains of Legionella (119 strains of pneumophila and 21 strains of non-pneumophila). Subsequently, the mip (macrophage infectivity potentiator) gene was PCR amplified, purified and sequenced. Those nucleotide sequences were blasted in the European Working Group for Legionella Infections (EWGLI) Database for the mip typing. Phylogenetic trees were constructed with the Neighbor-Joining (NJ) method using MEGA 5.0. [Results] Approximately 700 bp fragments of the mip gene was obtained from 140 Legionella strains. The 119 L. peumophila were found to be in 10 mip types, with L. pneumophila-phil-1 as the dominant accounting for 52.9% (63/119). The 21 non-L. penumophila from the isolates were found to be in 6 mip types, with L. feeleii-D3131 as the dominant accounting for 47.6% (10/21). [Conclusion] Great diversity was observed among Legionella strains in central air-conditioning cooling towers of Guangzhou public places, while the mip typing techniques used in this study could be used for fast genotyping of Legionella.

Abstract:[Objective] In order to study the spatial distribution of ammonia-oxidizing bacteria (AOB) in different vegetation communities (Tamarix ramosissima, Halocnemum strobilaceum, Phragmites australis) and different soil layers (0?5cm, 5 cm?15 cm, 15 cm?25 cm and 25 cm?35 cm) of wetland in the Ebinur Lake in Xinjiang. Then explore the correlations between the spatial distribution of AOB and soil environment factors. [Methods] MPN-Griess was used to study the distribution of AOB, using Pearson’s correlation analysis relationships between the distribution of AOB and soil environment factors. [Results] The distribution of AOB had obvious differences in vegetation communities of wetland in Ebinur Lake. The trend of distribution is that the highest Tamarix ramosissima community area, Halocnemum strobilaceum community is the second smallest and Phragmites australis community area minimum. The distribution of AOB also had obvious differences in different soil layers, the trend of distribution is that 15 cm?25 cm>0?5 cm>5 cm?15 cm>25 cm?35 cm. The distribution of AOB was significantly correlated to soil organic matter content, it had no correlation with pH, Moisture content, Salinity and NH4+?N content. [Conclusion] The distribution of AOB had obvious differences in vegetation communities and different soil layers of wetland in Ebinur Lake. Except the distribution of AOB showed obvious relationship with soil organic matter content, it had no relation with the other soil factors.

Abstract:[Objective] β-mannanase and xylanase are hemicellulase, which are already used in many areas of industrial and agricultural. The aim of this study was to coexpress two hemicellulase genes in E. coli (Escherichia coli). [Methods] The β-mannanase and xylanase genes were cloned from Bacillus subtilis BE-91, then linked together by a common restriction enzyme site and inserted into the pET28a(+), establishing a coexpression strain named B.pET28a-man-xyl for the extracellular production of β-mannanase and xylanase in E.coli. [Results] After being induced 21 h, the specific activities of β-mannanase and xylanase were 713.34 U/mL and 1455.83 U/mL, respectively. [Conclusion] The result of SDS-PAGE analysis, hydrolytic ring detection and enzyme activity determination showed that each enzyme was expressed extracellularly as individual functional proteins. In addition, comparing with β-mannanase and xylanase degraded hemicellulose alone, the effection of compound enzymes are much better. The successful construction of a strain which produces xylanase and β-mannanase will be helpful for the study and production of hemicellulases preparation.

Abstract:[Objective] The objective of this study was to examine the production of biosurfactant, taxonomic position and antifungal activity of the strain BS1. [Methods] Biosurfactant-producing bacterium was isolated by hemolytic activity assay on blood agar plates and hydrolyzing oil activity estimation on oil agar plates. Oil spreading method was used to assay surface activity. The strain BS1 was indentified according to morphological features, physiological and biochemical characteristics and 16s rDNA sequences of the strain. Antifungal activity was tested by dual-culture method and the inhibiting effect on the the mycelium growth, sporagium formation and spore germination. [Results] The strain BS1 of producing biosurfactant, isolated from the petroleum-polluted soil, was indentified as Pseudomonas sp.. The strain BS1, fermentation supernatant and the volatile compounds (VCs) exhibited inhibition activity on 12 kinds of plant pathogenic fungis. The strain BS1 and fermentation supernatant displayed significant inhibition to the Phytophthora sojae, the inhibitory rate was 65.31% and 95.93%, respectively. The fermentation supernatant of BS1 inhibited the growth of Phytophthora sojae by inhibiting mycelium growth, sporagium formation and spore germination, which had also remarkable inhibition even if it was diluted 20 times with pure water. VCs produced by strain BS1 had notable inhibiting effect against Sclerotinia sclerotiorum, and its inhibitory rate was 84.25%. [Conclusion] The strain BS1 can produce surfactant and has the potential biocontrol.

Abstract:[Objective] In order to find the pathogenic bacteria of white-spots disease in internal organs of Larimichthys crocea. [Methods] Using the method of isolation and identification of the causative agent in combination with 16S rRNA sequencing to find the pathogenic bacteria.Besides it was determined by artificial infection. The tissue of diceased fish were pathologically analyzed by paraffin histological section techniques. [Results] The bacterium Pseudomonas putida was responsible for the disease. Affected fish showed obvious inflammation and cell deformation or disintegration of the liver, kidney and spleen. [Conclusion] The conducted study was necessary in order to reveal the reason for the high mortality of cultured yellow croaker in the Xiangshan bay. Furthermore, the results of this study can help to early recognize such disease outbreaks, hence preventing drastic economic losses in the future.

Abstract:[Objective] Establishment and optimization of the gene transfer system of Streptomyces pulveraceus which produces fostriecin based on normal conjugal transfer method. [Methods] Intergeneric genetic transfer system was based upon integrative plasmid pSET152 from donor E. coli ET12567/pUZ8002 to Streptomyces pulveraceus. [Results] The maximum transformation efficiency of Streptomyces pulveraceus by conjugation was obtained when MS medium containing 15% glycine was used, with the spores heat-shocked at 50 °C for 10 min before mixing with E. coli, using a final 20 mg/L concentration of apramycin for overlay after incubation of 18 h. Simultaneity, pSET153::gfp was constructed from pSET152 by combining the constitutive ermE+ promoter with green fluorescent protein gene (gfp). Green fluorescent protein gene was expressed. [Conclusion] In summary, intergeneric genetic transfer system was demonstrated and optimized, and glycin can increase the efficiency of exconjugates.

Abstract:[Objective] This study is to express Norovirus (GenegroupII) VP2 in insect cells, analysis the subcellular localization of VP2 and lay the foundation of the function study of VP2. [Methods] Norovirus (GenegroupII) ORF3 gene was amplified from plasmid pMD-ORF3 using primers P1/P2 (including 6×His tag coding sequence). The product was cloned into the vector pFastBac1 to construct recombinant plasmid pFB-ORF3. pFB-ORF3 was transformed into competent DH10Bac to get recombinant bacmid Bac-ORF3. Recombinant baculovirus, Ac-VP2, was generated for expressing VP2, by transfecting recombinant Bac-ORF3 with LipofactamineTM 2000 into sf9 insect cells. The expression product was testified by the western blot and indirect immunofluorescence assay with monoclonal antibody against 6×His tag. [Results] The result of western blot showed a 29 kD specific bind in sf9 cells infected by Ac-VP2. Indirect immunofluorescence assay showed the specific green fluorescence in sf9 cells infected by Ac-VP2, and the green fluorescence could localize at nuclear and membrane of sf9 cells. [Conclusion] These results demonstrated that the Norovirus (Gengroup II) VP2 was expressed in sf9 cells successfully and could localize at nuclear and membrane of sf9 cells.

Abstract:Iron is an essential element that is required for the growth and normal metabolism in most organisms. However, despite its much abundance in the Earth’s crust, the bioavailable form of iron is very poor. To obtain iron in the environment, Candida albicans, as a common opportunistic human fugal pathogen, has evolved the iron regulatory networks to respond to the fluctuations in iron availability, which is associated with the adaptation to the hostile environment. As well as our study, this paper reviews the research advances of iron regulatory networks in recent years, focusing on the iron acquisition and regulatory strategies exhibited by C. albicans when it responds to iron deprivation. This review also provides an insight into the mechanisms that how cells sense, transport, store and utilize iron.

Abstract:For the production of recombinant proteins, Escherichia coli is often the first choice because it is easy, fast and inexpensive to cultivate, and its vector systems have been well developed. The regulation system for heterogenic expression plays an important role in producing recombinant proteins. pHsh vectors have been developed by using a heat-shock promoter of E. coli, and give advantages in high expression levels, low costs for expression induction, and reducing the formation of inclusion bodies. This paper introduces the construction strategy, expression mechanism, application of the pHsh vectors.

Abstract:Experimental medical immunology and pathogenic biology is a new course which contains all experiment courses of medical microbiology, medical immunology, and medical parasitology. The teaching content was divided into three grades: Grade Ⅰ (foundation experiment), Grade Ⅱ (comprehensive experiment), and Grade Ⅲ (designing experiment), in which the designing experiment is not assigned definite titles. The students are required to think their own titles and then design and complete the designing experiment, using experiment skill and theoretical knowledge of the three subjects. Teaching of the new course can enhance practical skills, abilities of innovative thinking, and comprehensive qualities of the students. The new course deserves promotion to medical specialties after improved.

Abstract:[Objective] Study the effects of microwave irradiation on isolation of basophilic and halophilic marine actinomycetes. [Methods] Seven marinemud samples were radiated by microwave and then gradient diluted for isolation of basophilic and halophilic marine actinomycetes in three media. [Results] Microwave irradiation could highly significantly increase the total quantity of basophilic and halophilic marine actinomycetes respectively in four and three marinemud samples. The total quantity of basophilic and halophilic marine rare actinomycetes of Micromonospora, Actinoplanes and Nocardia were significantly increased after microwave irradiation. The species of other marine rare actinomycetes such as Catellatospora, Microbispora, Streptosporangium were increased to one to four in different samples. [Conclusion] Microwave irradiation could significantly increase the total quantity of basophilic and halophilic marine actinomycetes and the species of marine rare actinomycetes.