Abstract:

Abstract:Mycobacterium tuberculosis (MTB) PknG, one of 11 Ser/Thr protein kinases produced by MTB, plays an essential role in the residing of MTB in the macrophages as persistent state. In this work, we amplified the pknG gene from the genome of H37Rv, and constructed a recombinant plasmid pET-pknG to express the PknG protein in the E. coli. Using purified PknG, we established and optimized a high-throughout screening model to screen for PknG inhibitors. Among 2120 samples of microbial fermentations and 2300 samples of compounds tested, one positive compound and 13 positive fermentation samples were identified with inhibitory effect on activity of PKnG, resulting in a 0.32% of positive rate.

Abstract:In order to improve the thermostability of Penicillium expansum lipase (PEL), a mutation P197E was created by site-directed mutagenesis. The mutation is generated by overlap extension PCR using the cDNA of a single site mutant lipase ep8 as the template and two special primers that corresponding to mutation P197E. Recombinant vector pPIC3.5K-ep8-P197E which contain double-mutant genes was constructed and electroporated into Pichia pasteris GS115. The recombinant transformant was selected and grow in the methanol containing media for expressing double-mutant lipase, PEL-ep8-P197E. Thermostability analysis reveals that the residual activity of the double-mutant are 42.13% and 37.3% greater than that of the wild type PEL and PEL-ep8 after incubated at 40°C for 30 min. The Tm of the double-mutant lipase is 41.51°C, 2.81°C higher than PEL and 2.25°C higher than PEL-ep8.

Abstract:The process of L-Tryptophan fermentation by E. coli TRTH/pSV-709 was investigated, and the result showed that there was a large number of byproduct (acetic acid) in the broth. The effect of acetic acid on L-Tryptophan fermentation was studied by experiments with addition of acetic acid. The results showed that 2 g/L acetic acid could inhibit the growth of L-Tryptophan-producing bacteria and L-Tryptophan production. The mechanism of production of acetic acid was studied. Several strategies including dissolved oxygen regulation, suitable concentration of initial glucose determination, glucose-limited pulse fed-batch and the specific growth rate regulation were applied to reduce the generation of acetic acid. Under the optimal condition, the concentration of acetic acid was decreased by 51.35% compared with that under original condition. Biomass and yield of L-Tryptophan were increased by 51.07% and 46.54% respectively, which accessed high cell density cultivation.

Abstract:Surface display of enzymes has been employed to improve stability of enzymes, and further save tedious purification process of enzymes to be used in conventional immobilization. Lipase is widely applied in industry. In this study, we constructed a Saccharomyces cerevisiae strain displaying lipase Lip2 from Yarrowia lipolytica on the cell surface. The gene encoding mature Lip2 was fused with the genes encoding the Kre1p leader sequence and the C-terminal domain of Cwp2 including the glycosylphosphatidylinositol(GPI)-anchor attachment signal. The Lip2 displayed on the yeast cell surface remained hydrolysis activity of lipase. The measured activity of the displayed lipase was 4.6 U/g dry cells. The displayed lipase was also characterized for its potentiality as a whole-cell biocatalyst. Its maximal activity was observed at 40°C and pH 8.0, and it exhibited good thermostability and high tolerance to organic solvent.

Abstract:Shewanella decolorationis S12 is a strain which can perform anaerobic respiration by using various electron acceptors under the anaerobic environment. In this study, to obtain the enough biomass and satisfy the need of scientific research such as membrane proteomics, we used the inorganic small molecule (sodium nitrate), metal ion (ferric citrate) and organic macromolecule (azo dye amaranth) as sole terminal electron acceptors, using different concentrations of electron donor and carbon source for anaerobic stationary culture and anaerobic fermentor culture of Shewanella decolorationis S12. The cells were cultured by fed-batch cultivation mode to confirm the optimal concentrations of electron donor and carbon source, finally the method of anaerobic fermentor culture for S12 was established. Comparing to the traditional anaerobic stationary culture method, anaerobic fermentor culture method not only ensured a high rate mass-transfer efficiency resulting in the effective reduction of electron acceptors, but also greatly increased the cell growth density, the max cell growth densities were increased to 325, 304, 369 times and cell growth times were decreased 26.5%, 17.6%, 7.5% separately. This method provided an effective way to culture a large number of cells and protein under anaerobic respiration condition. The procedure described above would be significance for the studies which need biomass cultivation of Shewanella genus bacteria and other anaerobic microorganisms under fully controlled conditions.

Abstract:The diversity and numbers of soil microorganisms in different forest types were studied, as well as Shannon index (H), richness index of genera (S), Pielou evenness index (J) and Simpson index (D) in Ledum palustre —Larix gmelinii virgin forest, Herbage—L. gmelinii virgin forest, Betula—L. gmelinii virgin forest, clear cutting L. gmelinii forest and L. gmelinii forest burned areas in the national forestry ecosystems station of Inner Mongolia Great Xingan Mountains by selective media. The results showed that bacteria isolated from five plots belonged to 24 genera, Brevibacterium, Bacillus, Corynebacterium and Micrococcus were main genera; actinomycetes were classified into 6 groups, Flavus and Albosporus were the main genera; fungi were composed of 20 genera, Penicillium and Cephalosporium were the dominant genera. Diversity index (H), richness index of genera (S), evenness index (J) and dominant index (D) of soil microorganisms of Larix gmelinii varied with forest types; and the order of diversity index and richness index of bacteria followed LLV > HLV > LFB > CL > BLV, while actionmycetes were BLV > HLV > LLV > LFB > CL and fungi were CL > HLV > BLV > LFB > LLV.

Abstract:For rational design of novel polyketides and improving the combinatorial biosynthesis of polyketides. 26 β-ketoacyl-ACP synthases from several polyketide synthases were analysed with ClustalW、MEGA4.0. The primary parameters, secondary, tertiary structures and active sites of nine of them with different substrate specificity were compared or predicted in detail using ProtParam, Phdsec, Swiss-Model. The results revealed that they have more similarity in structure than in sequence and all own a high percentage of serine in their active sites; in addition, the β-ketoacyl-ACP synthases whose substrates possess two carboxyl groups were found to have a theoretical pI less than 5.0, and their formal charges sum were on the low side too; the conserved sequence of the 26 β-ketoacyl-ACP synthases is V303ELHGTGTPLGDPIEAGA320, but it is away from the active site. These conclusions have promising potential for module or domain substitution and site-specific mutation of polyketide synthases; also providing a reference for the study of their evolution mechanism.

Abstract:This study investigated the endophytic fungi associated with Taxus chinensis growing in the Qinba Mountains. The strain LB-10 was found to produce taxol in vitro with a yield of 846.1 μg/L, detected by UV, HPLC, and HPLC-MS. The strain LB-10 was grouped into Metarhizium anisopliae based on the morphological traits and ITS sequencing.

Abstract:The traditional isolation and identification method and analysis based on 16S rRNA sequences were used to investigate the phylogenetic diversity of endophytic moderately halophilic bacteria of Suaeda salsa L. in Dongying. According to physiological and biochemical characteristics, 16S rRNA sequences and phylogenetic analysis, 15 isolated endophytic bacterial strains were classified into four groups, belonging to genera Chromohalobacter, Kushneria, Halomonas or Bacillus separately. 16S rRNA sequences of 4 strains in Group I had the highest similarity to Chromohalobacter israelensis (95%). Group II which contained 7 strains of the genus Kushneria is the dominant group in Suaeda salsa L.. The strains in Group III bear the closest affiliation to Haererehalobacter sp. JG 11 (99%), which is not classified definitely. The only endospore-forming bacterium was classified into group IV. Its 16S rRNA sequence similarities with respect to the known Bacillus species were ≤ 96%, which was presumed to be potential novel species. Among these isolated strains, 3 strains produced proteinase; 14 strains produced esterase; 8 strains produced DNase; 11 strains produced galactosidase; 14 strains produced urease. The research results indicated that there was not only abundant phylogenetic diversity of moderately halophilic bacteria, but also some unknown bacteria groups existed in Suaeda salsa L..

Abstract:Ninety endophytic actinomycetes were isolated from surface-sterilized samples of Camptotheca acuminata Decne. collected from Yunnan University. Results of 16S rRNA gene sequences analysis showed that they belong to 10 genera, 6 families. Among them, 33.4% of endophytic actinomycete cultures presented antimicrobial activity. The polyketide synthasesⅠ(PKS-I), polyketide synthases Ⅱ(PKS-II) and non-ribosomal peptide synthase (NRPS) sequences were respectively detected by PCR in 28.9%, 44.4% and 48.9% of tested strains.

Abstract:A total of 35 bacterial strains were isolated from the ovarian of Globefish. It was confirmed that 11 bacterial strains among them were capable of producing tetrodotoxin using mouse bioassay method and the toxin concentrations in the cultures of 6 bacterial strains were above 400 ng/mL. Based on the morphology, physiological and biochemical characteristics, the strain ZY-23, one of the bacterial strains, was identified preliminarily as Serratia liquefaciens. The phylogenetic analysis showed that 16S rRNA sequence of the bacterial strain ZY-23 had 99% homology to that of Serratia sp. Tp5. The result of fluorimetric spectrophotometry indicated that there was the same fluorescence peak of the fermentation broth as that of the standard substance, suggesting that the strain ZY-23 indeed produced tetrodotoxin.

Abstract:The CⅢ-1 strain, a marine bacterium screened out from 422 endophytic bacteria isolated from mangroves by the method of plate test, strongly inhibited the growth of Ralstonia solanacearum, Staphylococcus aureus, Vibrio alginolyticus, Phytophthora capsici, Fusarium oxysporum f. sp. cubense, etc. It was identified as Bacillus amyloliquefaciens based on the characters of morphology, physiology and biochemistry and 16S rRNA. Antimicrobial protein which produced by CIII-1 was the main antibacterial substance. The crude protein in culture filtrate could be precipitated with 33% ammonium sulfate and obtained by dialysis of the precipitate. The protein was found to be thermal-unstable, when treated at 60?C and 100?C for 10 minutes, its inhibitive activity would lost 62.5% and 100% respectively. The inhibitory activity of the protein was observed in the range of pH value from 5.0 to 10.0, and the strongest one was at pH 7.0.

Abstract:A plasmid from Lactobacillus plantarum KLDS1.0728, designated as p141, was sequenced and characterized. It consisted of a 3597 bp circular molecule with a G + C content of 38%. The enzyme digestion map of plasmid p141 was obtained by the software DNAMAN 6.0. The plasmid p141 was predicted to contain 15 ORFs by ORF Finder software in NCBI. Using the blast software to compare and search the genes, we found two known ORFs in the plasmid p141. Among those, rep gene was most important for plasmid replication ability. Identity analysis indicated that rep gene of p141 was similar to that of Lactobacillus plantarum WCFS1 plasmid pWCFS101 and Lactobacillus plantarum plasmid pM4. Based on the sequence similarities among minimal replicon modules, p141 belonged to Group III family in the RCR plasmid. In addition, mob gene was existed in p141, so this plasmid had level transfer ability. But Tn4430 transposon and transposase gene topl, topA were not found in p141, and this indicated that genes of the plasmid p141 were stable. There were direct and invert repeats in this plasmid. These repeats had certain association with plasmid replication and transfer regulation, protein expression, also had significance on the regulation of plasmid copy number.

Abstract:We investigated the reducing capacity of Caragana rhizobium exopolysaccharides (REPS) by the system of potassium ferricyanide and ferric chloride, determined the hydroxyl radical scavenging capacity of REPS by H2O2/Fe2+ system, measured the Superoxide radical scavenging capacity of REPS by the pyrogallol system, and we also determined its protective effects on H2O2 in hemplysis of erythrocytes and lipid per-oxidation of liver homogenate in rat. The results indicated that REPS has reductive capacity to oxides, the hydroxyl radical scavenging capacity of REPS was 35.46% (at the concentration of 2 mg/mL), the Superoxide radical scavenging capacity of REPS was 36.84% (at the concentration of 0.78 mg/mL), the protective effects of REPS on H2O2 induced hemolysis of rat erythrocytes was 43.84% (at the concentration of 1.14 mg/mL), and the protective effects on lipid per-oxidation of liver homogenate in rat was 34.46% (at the concentration of 1.14 mg/mL).

Abstract:In order to verify the Fluorescent-Quantitation PCR kit specific for Borna disease virus (BDV) nucleoprotein, compare molecular beacon with common probe and study its practical application, the sensitivity, distinctness, repeatability and stability were tested on BDV and non-BDV infected cells, as well as non-infected controls. Human and animal samples were also tested by the kit. The lowest quantity of virus RNA detected by the kit was 2.5×101, which corresponded to 1 virus copy. No false positive result was found. The coefficient of variation (CV) of different batches was less than 0.7, and the CV between normal kits and the kits treated by accelerating decomposition was less than 2. The morbidities of human and animal detected by the kits were 3.6% (human), 4.2% (swine) and 1.5% (horse). The sensitivity and distinctness of this kit are better than common probe kit. All of the results indicate that this kit is an efficient tool in fundamental research, clinical detection and epidemiological investigation of BDV.

Abstract:This paper described the screening of itaconic acid strain of Aspergillus terreus with UV-irradiation, LiCl treatment and breeding resistant strain to metabolic end product. Breeding resistant strain was an effective measure to help raise yields steadily. A strain, designated At394, producing itaconic acid with a high yield was successfully obtained. The itaconic acid concentration produced by At394 was 53.9 g/L using starch hydrolysate in shaking flasks, which was 42.6% higher than that of the parental strain. The conversion rate was 61.5%, which was the highest value found in tests. The structure determination by infrared spectrum showed the production obtained was itaconic acid.

Abstract:Fengycin is a cyclic lipopeptide antibiotic produced nonribosomally by Bacillus subtilis. In this study, the primary mechanism of inhibition to Fusarium moniliforme by Fegycins was investigated according to their special properties. Microscopic observation on morphology of F. moniliforme mycelia revealed that tips of partial mycelia were lysed. PI staining of Fengycins-treated F. moniliforme mycelia was observed under fluorescence microscope, which indicated the toxin-mediated damage of cytoplasmic membrane. Further study showed that addition of chitin, chitosan, β-1,3-glucan, ergosterol and cholesterol in agar media could not significantly modify the inhibition to F. moniliforme by Fengycins, while addition of phosphatidylcholine could antagonize the antifungal activity of Fengycins. These results suggested that phosphatidylcholine probably was the plasma membrane target for Fengycins. Additionally, our results also showed that fengycins could depress the activity of PLA2 secreted by F. moniliforme, which might contribute to the antifungal activity of fengycins against F. moniliforme.

Abstract:Solid-state fermentation was employed to produce protease by Bacillus licheniformis using deoiled Jatropha curcas seed cake as substrate. The optimal conditions of protease production were studied. Maximum protease production (7465 U/g of dry substrate, U/g) was obtained at 125% substrate moisture, a growth period of 3 days, supplementation with 10% (wt) of lactose and 5% (wt) of peptone. The results of protease analysis showed that the optimal temperature and pH were 55°C and 5?6, respectively. The Vmax and Km of protease produced by Bacillus licheniformis were 0.0324 μmol/(s?mg) and 0.0531 mmol/L, respectively. Organic solvent can enhance protease activity. In comparison with control, the activities were increased by 13.55% and 70.9% when the protease was solved by 10% (V/V) of methanol and 5% (V/V) of ethanol, respectively. Mg2+ enhanced protease activity by 42.54% while Hg2+ damaged protease and entirely inactive its activity.

Abstract:Many signal substances, such as salicylic acids (SA), jasmine acids (JA), flavonoids, NO, H2O2, etc. in plants are induced by arbuscular mycorrhizal (AM) fungi which colonize plant roots. The signal movement pathways and function mechanisms of these signals have been paid more attention to in recent years. In this review, we firstly summed up the signal substance kinds and quantities in plants inoculated with AM fungi, then discussed the signal transduction pathway, physiological effects and the related functioning mechanism in mycorrhizal plants, in order to provide a basis for the further study symbiotic relationship and mechanisms between AM fungi and plants.

Abstract:At present, there are many studies on continuous fermentation in microorganism fermentation engineering. In recent years, researchers conducted a variety of studies and revisions about the nonlinear dynamic system and has got many achievements. The models of microorganism continuous fermentation and its application is reviewed in this paper.

Abstract:During industrial production of starters, Lactic Acid Bacteria are subjected to cold shock by preservative techniques like freezing, frozen storage, and vacuum freeze-drying. Moreover, starters subjected to these pressures need to function effectively during application at low temperature used in the perservation of fermented foods and curing of cheese. All of these factors can influence the quality of starter culture and its fermented foods to some extent. The survival and physiological conditions of the starter bacteria preserved by such techniques (freezing, frozen storage, and lyphilization) will determine their functionality in such applications. Therefore, a better understanding of the responses and mechanisms of Lactic Acid Bacteria under low temperatures and freezing may contribute to the protection of freezing, fermentation and storage for starter culture and fermented foods when they are suffered in low temperature processes. In this study, the protective mechanisms needed for starter bacteria to survive in above mentioned preservative techniques with good functionality were analyzed. Based on these analyses, we have proposed protection methods that would improve the functionality of starter bacteria preserved by freezing, frozen storage and/or lyophilization.

Abstract:Bioremediation is a promising technique for eliminating polycyclic aromatic hydrocarbons (PAHs) from contaminated soil. The microbial biodegradation mechanisms of high-molecular weight polycyclic aromatic hydrocarbons (HMW)-PAHs in soils were reviewed with special emphasis on the principles and difficulties of bioremediation of PAH-contaminated soil. Some genera of microorganisms able to utilize HMW-PAHs as the sole carbon and energy source have been isolated. Most of them are mainly single strains which can metabolically degrade four-ring PAHs, and some of them can co-metabolize five-ring PAHs. Low bioaccessibility of HMW-PAHs is a difficulty in the bioremediation of contaminated soil. Release of surfactants, formation of biofilms and production of extracellular polymeric substances by some of the PAH-degrading bacteria can enhance the bioaccessibility of PAHs and therefore accelerate the biodegradation. Combination of bacteria and fungi can increase their in situ bioremediation efficiency. Therefore, bioremediation is an environmentally friendly, economic suitable and sustainable technique for eliminating PAHs from soil.

Abstract:Integrons are novel DNA elements, which can capture and incorporate gene cassettes embedded in exogenous by site-specific recombination. Meanwhile, they are able to convent gene cassettes to functional genes by ensuring their correct expression. The horizontal transfer of this genetic element within and between microbial genera has speeded up emergence of novel antibiotic resistance gene. To date, the mechanism of resistance to antibiotics, particularly β-Lactam class of antibiotic, is becoming exceedingly complex. What is more, multi-drug resistant microbes with complex resistance patterns have posed an increasing threat to the health. So intensive research leading to greater understanding is imperative.

Abstract:According to the microbiology experimental teaching system under the new curriculum reform of college, the microbiology experiment guide in normal college has been reformed, and the cultivating students’ manipulative ability as the starting point was also put forward. From the reform of the experimental content, teaching method, experimental teaching staff and some other aspects, a set of innovation teaching system on microbiology experiment course to suit the biological science undergraduate in normal college was to be established.

Abstract:Microbiology, as one of the normal colleges’ foundation course, is closely related to secondary biology education. In this thesis, three new teaching targets have been put forward during the microbiology teaching process, which include integrating the course contents of microbiology and secondary biology, enhancing teaching skills training and offering careers guidance. The feasibility analysis and implementing methods for these three teaching targets have been discussed and practiced in three aspects including teaching contents, teaching methods and course examination.

Abstract:On June 12, 2009, the first case of novel influenza A H1N1(2009) virus infection was confirmed in Jiangsu Province. A virus with hemagglutination activity was isolated from the clinical samples derived from the patient using both MDCK cells and embryonic eggs, which was designated as A/Jiangsu/1/2009(H1N1). In order to monitor evolution of the virus, we carried out whole-genome sequencing, and analyzed the genetic characteristic of hemagglutinin (HA) of the isolate in detail. The results showed that HA did not contained the multibasic amino acid motif at cleavage sites, which is a characteristic of low pathogenic influenza viruses. Compared to reference virus A/California/04/2009, the HA protein of the isolate had four amino acid mutation, which were not located on the known antigenic sites. Consistent with the triple-reassortant swine H1 viruses from Northern America in recent years, the isolate contained five potential glycosylation sites, which also was the feature of classical swine H1N1 viruses. Compared with avian H1 viruses, two amino acid (E190D and G225D) were changed on receptor binding sites, which may be an important molecular mechanism of human-to-human transmission for the novel viruses. In this study, for the first time, we studied the molecular characteristics of the HA protein of the novel influenza A H1N1(2009) viruses in detail, which have an important guiding significance for further monitoring of virus mutation.