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Abstract:In the present study, the influence of autophagy on apoptosis of macrophage induced by Salmonella typhimurium was investigated. Murine macrophage-like cell line J774A.1 was cultivated with RPMI 1640 containing autophagy inducer rapamycin (RAPA) or not overnight, the standard Salmonella typhimurium virulent strain SR-11 was used as the test bacterium. First, the influence of RAPA on bacteria growth and J774A.1 cells survival were detected, then the incubation of J774A.1 and SR-11 were dynamically tracked 24 h. Cell ultrastructure, the formation of autophagic vesicles, Beclin-1 and Bcl-2 expression, cell survival rate, the number of intracellular viable bacteria and cell apoptosis were detected at different time. The results showed that RAPA didn’t affect bacteria growth and the survival of J774A.1 cells. However, in cellular infectious model, RAPA could significantly reduce the number of intracellular viable bacteria and the rate of macrophage apoptosis, thereby increasing cell survival (P < 0.05). Some bacteria was wrapped in the autophagic vacuoles of J774A.1 cells during inchoate infectious stage, and cellular ultrastructure was normal after RAPA treatment. Moreover, the expression of Beclin-1 was increased while the expression of Bcl-2 was declined, and cells damage were significantly lighter in the late stage. All these results suggested that the regulation of cellular autophagy to lower the level of host cells apoptosis may be used as new ways of the prevention and control for some infectious diseases.

Abstract:

Abstract:The cells of engineered strain HC01 were immobilized in the form of Ca2+-alginate beads. The conditions for immobilization were investigated. The optimal gel concentration and cell concentration were found to be 2.5% and 0.029 g/mL in the presence of 3% CaCl2. The thermo-stability of immobilized cells was 5°C higher than that of free cells in the same condition. Divalent metal ions, such as Mn2+, Mg2+, Cu2+, Co2+ and Ni2+ did not affect significantly the enzymatic activity of D-hydantoinase (HYD) and N-carbamoylase (CAB) in immobilized cells. By contrast, Mn2+ and Ni2+ could independently enhance the activity of CAB up to 210% and 270% in free cells. The present data showed that the optimal reaction condition of immobilized cells was at pH 7.5 and 40°C as same as that of free cells. The immobilized cells were applied to produce D-p-Hydroxyphenylglycine (D-HPG) directly from the substrate DL-Hydroxyphenyl Hydantoin (DL-HPH) in a batch reactor. Conversion of about 97% was reached after 36-h reaction when the substrate concentration was 30 g/L with the initial pH of 9.0 under the protection of nitrogen gas. The overall yield of D-HPG was 85% with the optical purity of 99.7% after purification.

Abstract:Strain XW-1 producing extracellular cellulose was isolated from 80 cm depth under the surface layer at Zoige Plateau Alpine Wetland. It was identified as a Brevundimonas sp. by its morphological, the physiological and biochemical properties and 16S rDNA sequence analysis. The results of cellulose producing research showed that the optimum carbon sourse was 0.5% CMC-Na, the optimum temperature was 20°C. Under this condition, the highest cellulose activity reached to 15.6 U/mL in the third day. Enzymatic properties showed that the optimum pH value is 6.0 and the optimum reaction temperature is 20°C, the relative enzyme activity reach to 80% at 15°C, moreover, at 5°C the relative enzyme activity still reach to 56%.

Abstract:Using the pure culture technology, we analyzed 26 predominant bacterial strains isolated from a denitrifying quinoline-degrading bioreactor. The capacity of denitrification and aerobic degradation for quinoline of these strains was determined. Then the ability of denitrifying degradation of quinoline by selected denitrifying strains was measured. The results showed that 10 strains of isolates, which belong to Bacillus, Staphylococcus, Pseudomonas, Brucella and Delftia, were denitrifying bacteria. Whereas, 9 strains of isolates belonging to Rhodococcus had the ability of quinoline degradation under aerobic condition. This work described the metabolism function of the major predominant bacteria of microbial community in a quinoline degrading bioreactor. Diverse bacteria, in terms of metabolism, existed in the anoxic denitrifying bioreactor.

Abstract:Membrane protein plays a vital role in cellular differentiation and signal transduction. However, the study of structure and function of membrane protein is still limited due to lack of the efficient preparation for enough high quality membrane proteins. In this paper, zebrafish CXCR4b gene was cloned into pMAL-p4x vector, then was expressed as a fusion of MBP-CXCR4b in E. coli TB1. Over-expression of CXCR4b was achieved after optimizing expression conditions systematically. The optimal expression conditions were obtained as: E. coli TB1 as host strain, TB medium, concentration of IPTG 0.5 mmol/L, inducing at the late mid-logarithmic growth phase. Totally 10 surfactants were screened for their ability to successfully solubilize CXCR4b from cell membranes. As a result, DM, FC-14 and Brij35 exhibited good solubizing ability for CXCR4b. CXCR4b was then purified using Ni2+ chelating affinity chromatography followed with gel filtration S200. The purified protein showed correct apparent molecular weight on SDS-PAGE and the purity was up to 90%. A typical α helical structure was determined with circular dichroism analysis, which means that the purified CXCR4b had folded into a reasonable conformation.

Abstract:An aiiA-expressing vector, pPIC3.5K-aiiA, was constructed and transformed into Pichia pastoris GS115 by electroporation. The recombinant yeast strains were screened with auxotroph medium and phenotypic identification.The transformants with high copy numbers of aiiA gene were selected in medium containing high concentration of G418. The expression of aiiA was induced by addition of methanol into culture at the final concentration of 0.5%. The transcription of aiiA was confirmed by RT-PCR in the recombinant yeast. SDS-PAGE and Western blot analysis demonstrated that recombinant AiiA protein was successfully expressed after induction. The recombinant AiiA protein showed the activity of degrading the N-acyl-homoserine lactones when using Chromobacterium violaceum CV026 as reporter strain.

Abstract:A large amount of ATP is consumed to regenerate precursor UDPG for curdlan biosynthesis in Alcaligenes faecalis var. myxogens. In this study, low-polyphosphates with high-energy phosphate bond of Na4P2O7 (2P), Na5P3O10 (3P) and (NaPO3)6 (6P) were employed as high-energy phosphate bond and phosphate donor to substitute KH2PO4-K2HPO4 (1P) in medium for curdlan production by Alcaligenes faecalis var. myxogene. Results showed that curdlan production was enhanced 23% and 134% than that of control respectively, which reached 15.1 g/L and 30.0 g/L, when one-fold and two-fold high concentration of phosphate from 3P and 6P were added in the medium. Meanwhile, byproducts of acetic acid were decreased 87.5% and 77.7% respectively. Formic acid was also decreased 75.7% when two-fold 6P were added in the medium. However, when CaCO3 was removed in the medium, pH and biomass were decreased remarkably, therefore, little or no curdlan was produced when these low-polyphosphates were added in medium. When 3P and 6P were separately mixed with 1P at the amount of one-fold and two-fold high concentration of phosphate for fermentation, there was no change on curdlan production. Nevertheless, if 1P was used as buffer substance and without CaCO3 in the medium, curdlan production could be enhanced to 60.4% and 49.4% than that of the control when 6P was added at the amount of one-fold and two-fold high concentration, reached to 18.4 g/L and 16.9 g/L.

Abstract:Strain FU05 with cellulose-degrading activity was isolated from cellulose-rich environment and identified as Trichoderma longibrachiatum through morphology characteristics and ITS sequence analysis. The cellulase genes bgl2, cbh2 and eg1 were cloned by PCR. By conducting sequence alignment analysis with the reported data it was found that the homology of same cellulase genes between strain FU05 and other Trichoderma were: 91% to bgl2 (AB003110) from Trichoderma reesei; 99% to cbh2 (DQ504304) from Trichoderma koninqii and 95% to eg1 (X60652) from Trichoderma longibrachiatum. Furthermore, the corresponding amino acid sequences were also quite similar. By means of PROSITE motif search, the locations of N-glycosylation site, cellulose-binding domain and conserved domains of glycosyl hydrolases family in the corresponding protein were confirmed.

Abstract:To reveal mechanism of soil and water conservation plant such as Burma Reed (Neyraudia reynaudiana) tolerant of low phosphorus, explore germplasm of solubilizing phosphorus and improve use efficiency of phosphorus in latosolic red soil, a solubilizing phosphate fungus FP1 was isolated from rhizosphere soil of Burma reed and identified as Aspergillus niger according to morphological characteristics and ITS rDNA sequences. The dynamic variation of solubilizing phosphate capability and pH of in the liquid medium with three kinds of inorganic phosphate indicated that the pH of the liquid medium decreased significantly after inoculated FP1 and the solubilizing phosphate capability was related with culturing time. Except for Ca3(PO4)2, solubilizing phosphate capability of FP1 rose first, then fell, and became stable in the end. The maximum phosphate solubilizing efficiencies were ordered as follows: AlPO4 (92.02%) > Ca3(PO4)2 (41.62%) > mixture of these three insoluble phosphates (35.86%) > FePO4·4H2O (19.20%). The fungus FP1 was high efficient on solubilizing AlPO4 and FePO4·4H2O. It is indicated that microorganisms with high efficient solubilizing phosphorus exists in soil and water conservation plant rhizosphere.

Abstract:Strain FDB, a fenvalerate-degrading bacterium, was isolated from some long-term pesticide-contaminated agricultural soils. The strain was identified as Pseudomnas aeruginosa based on its morphological and physiological properties, and 16S rDNA sequence analysis. Strain FDB grew in the medium supplemented with fenvalerate as its sole carbon source and degraded fenvalerate isomers at the concentration of 100 mg/L at the degradation rate from 69.06% (SR + RS) to 64.32% (SS + RR) in 5 days shaking at 30°C. The optimal growth was obtained under 35°C, pH 7.0 and 75 mL medium content 250 mL flask through shaking. The crude fenvalerate-degrading enzyme solution from cellular disruption was prepared by sonication, the activity of fenvalerate-degradation was associated with the intracellular fractions.

Abstract:Through the method of traditional culture and PCR-DGGE, variety of soil microbial structure in continuous cropping cotton fields in south Xinjiang was analyzed. The results showed with the increase of continuous cropping years, the number of bacteria and fungi, ratio of total bacteria and fungi and Shannon-Weiner index increased at first, than decreased, and after continuous cropping 15 years increased again, it was a lower point in the sample of continuous cropping cotton 15 years. Bacterial diversity and structure changed slightly, while the quantity and species of dominant bacteria were not much different in the fields with different continuous cropping years. Compared with bacteria, fungi structure was more sensitive to continuous cropping years, dominant fungi population presented a single tendency and lower species diversity.

Abstract:Corn straw bio-reactors were constructed in 2-, 4-, and 8-year replant soils grown with watermelon, while the 2-, 4-, and 8-year replant soils (without bio-reactors) were used as controls, respectively. The soil nutrient contents, pH, microorganism quantities, and soil enzyme activities were determined to evaluate the effectiveness of corn straw reactor’s improvement on soil health. Results showed that the soil organic matter content, microorganism quantities, and soil enzyme activities were increased significantly after 1- or 2- yearcorn straw bio-reactor treatments. In the 8-year replant soil treated with 2-year corn straw bio-reactors, the organic matter content reached 2.41%, the number of bacteria was 1.65 × 108 CFU/g, the actinomycetes was 3.12 × 106 CFU/g, the fungi was 0.5 × 105 CFU/g, and the abuscular mycorrhizal fungal spore density was 57/50 mL soil, which were 2.2, 5, 1.44, 1.36, and 3.2 times of their respective controls; while the polyphenol oxidase and sucrose invertase activities were 1.25 and 2.13 times of their controls, respectively. It was suggested that the corn straw bio-reactor be benefitial to rehabilitate replant soil.

Abstract:Vibrio parahemolyticus is a saltophilic and important food-borne pathogen which widely exists in all kinds of seafood. The general method costs more time and has low specificity in isolating and identificating of V. parahemolyticus. In order to enhance the efficiency and shorten the detection periods, a new chromogenic medium (HKMVPM) was designed in this study. The sensitivity, specificity and detection efficiency of HKMVPM were studied and compared with that of KHVPM and TCBS by detecting the reference strains and natural samples. The results showed that the selectivity and specificity of HKMVPM were same as that of KHVPM. The detection efficiency of HKMVPM was better than that of KHVPM. The detection efficiency of KHVPM was better than that of TCBS. V. parahemolyticus chromogenic medium shows a high degree of higher sensitivity and specificity and has a good prospect in applying.

Abstract:The effects of mannan-oligosaccharides and flavomycin on the intestinal microorganisms of Tilapia larvae [(l.00 ± 0.04) g] were studied. The results showed that the number and ratio of Lactobacillus and E. coli in the intestinal tract of Tilapia larvae were obviously affected by three different additives including self-made yeast mannan-oligosaccharides, purchased mannan-oligosaccharides and flavomycin with different concentrations by plate count method. Among these additives, 0.5% self-made yeast mannan-oligosaccharides increased the Lactobacillus by 10.77% (P < 0.05), but reduce E. coli by 14.80% (P < 0.05), which increased the ratio of lactobacillus and E. coli to 0.9720. Furthermore, the Denatured Gradient Gel Electrophoresis (DGGE) fingerprints demonstrated that the original intestinal microbial microecology of Tilapia larvae was changed by all additives, among the three additives, self-made yeast mannan-oligosaccharides had the most significant influence on the intestinal microorganisms of Tilapia larvae.

Abstract:A γ-Red recombination system was used to construct an ipaH4.5 deletion mutant of S. flexneri 2a 301. A recovery mutant was obtained by introducing a low-copy plasmid strain. PCR analysis confirmed that ipaH4.5 was successfully deleted in the mutant and restored in the complementary strain. The growth curves of wild-type, deletion mutant and recover mutant were measured. Some biochemical tests were investigated. Invasion tests were performed to evaluate the virulence of these three strains. Quantities of cytokines in the culture supernatants of murine macrophages cell line J774 after being infected was measured by enzyme-linked immunosorbent assays (ELISA). No significant difference of growth rates and basic biochemical events were observed among three strains, and no difference in invasion ability was observed either. However, More IL-1β and TNF-а were observed in murine macrophages infected with deletion mutant than wild type and recover mutant. So we conclude that ipaH4.5 can inhibit inflammation response of the host after Shigella flexneri 2a 301 enter into the cell.

Abstract:The Bacillus subtilis strain 2 (BS2) was measured to antagonistic mechanism by using face to face culturing. The antagonistic effect of BS2 to Botrytis cinerea was obvious. Growth inhibition rate had significant difference between bacteria culture fluid and extracellular proteins at various temperatures (20°C-120°C). But the activity effect of extracellular proteins of BS2 was sensitive to temperature. Bacteria culture fluid and extracellular proteins produced from BS2 had a good antibacterial effect on B. cinerea mycelial growth, spores bearing and germination. And Treated with bacteria culture fluid and extracellular proteins produced from BS2, protoplast from the hyphae became abnormal. Overall, this study indicates that the BS2 active substance is composed of various components and has multiple antagonistic mechanism.

Abstract:The auto-inducing expression of mutant human glucagon-like peptide-1 fusion protein in Escherichia coli BL21 (DE3) was studied. In order to increase the protein yield, we optimized both the auto-induction fermentation conditions and medium. The results showed that the optimal auto-induction medium is: tryptone 19.17 g/L, yeast extract 9.59 g/L, Na2HPO4 5.72 g/L, KH2PO4 5.48 g/L, (NH4)2SO4 2.66 g/L, NaCl 3.33 g/L, glycerol 2% (v/v), glucose 0.68 g/L, lactose 6.33 g/L, MgSO4 0.24 g/L. The genetically engineered strain was cultivated at 33°C, pH7, in a 100 mL flask containing 20 mL such medium, with the inoculum size of 1%, for 12 h. Then the yield of mutant human glucagon-like peptide-1 fusion protein reached 348.6 mg/L.

Abstract:Cellulose, the main structural component of plant cell walls, is the most abundant renewable resource in nature, but it is extremely difficult to be degraded. Cellulosome is a multi-enzyme complex that can efficiently degrade cellulose and the degradation products can be used by some anaerobic microorganisms to produce ethanol. Recent research status in anaerobic degradation of cellulose for ethanol production by cellulosome-producing bacteria was reviewed in this paper. The latest achievements and research development in cellulosomes’ structure and function, designer cellulosomes, metabolic engineering and co-culture of cellulosome producing microorganisms were also summarized. It is expected that the development of new cellulose degrading microorganisms and relative technology will further cut the cost of cellulosic ethanol in the near future and make it more competitive with gasoline.

Abstract:Phytase is a type of orthophosphomonoester phosphohydrolase, which catalyticly initiates stepwise hydrolysis of phytic acid to release phosphate radicals and produce lower inositol phosphate derivatives. Phytase has great application potential in the areas of animal nutrition, resource conservation, environmental protection and public health. At present, the understanding in phytase diversity and classification is so confusing or even inaccurate that the research progress and level of phytase has been badly affected. In this paper, the phytase classification based on optimal pH and stereospecificity is briefly introduced first, and then the updated research advance in phytase classification based on structure and catalytic mechanisms and the attributes of the representative phytases are summarized and discussed comprehensively. It is of vital importance to take into consideration the classification standards especially to focus on structure and catalytic mechanisms when a given phytase could be fully and accurately understood and characterized.

Abstract:Combined specialty teaching characteristics with bilingual teaching status of veterinary microbiology of Higher agricultural and forestry colleges, this article analyzed the factors that affect the effectiveness of bilingual teaching. On this basis, it brought forward corresponding perfect bilingual teaching suggestions and measures of veterinary microbiology. A large number of improvements had been made in the teacher arrangement, textbook compilation and teaching method in order to really improve bilingual teaching quality of veterinary microbiology.

Abstract:In the course of teaching of the fermentation kinetics, these doubts were cleared up by attempts to get the good result of the classroom teaching, which are the biomass growth rate and specific growth rate, the biomass actual yield coefficient (Yx/s) and the theoretical yield coefficient of growth (Ygs), the actual product yield coefficient (Yp/s) and the theoretical product yield coefficient (Yps), specific growth rate control in fed-batch fermentation and other difficulties.

Abstract:A quality curriculum is a lead way of congeneric curriculum which possess a first-class faculty team, first-class teaching contents, first-class methods, first-class teaching materials and first-class management. The key in quality curriculum construction is the training of a first-class faculty team. What is the first-class teacher? They have munificent bearing, advanced scientific attainments, upper humanities accomplishment and favorable psychological feature.

Abstract:A detection method of viable but non-culturable bacteria by fluorescence microscopy was established with SYTO9 and PI staining cell nucleus, ink instead of Iraq black, specimens fixed by hand. Problems on background fluorescence, cell aggregation, blur fluorescence image, low micropore film absorbability, unclear bacterial form, and fluorescence quenching were analyzed. Some solution strategies were suggested in order to offer help for researchers.

Abstract:Bacteria pathogens were isolated from diseased tilapia cultured in several farms of Guangdong and Hainan provinces. Artificial infection experiments showed that these isolates possessed strong virulence. Intraperitoneal injection (≥1 × 106 CFU/mL) of some isolates caused 100% mortality of healthy tilapia. 7 bacteria strains with strong virulence were chosen for antibiotics sensitivity test and identifications. Antibiotic sensitivity assays showed that among 29 antibiotics tested 13 were sensitive, 7 were resist, while 9 items showed various reactions among strains. The isolates were all gram-positive and β-hemolysis and were identified as Streptococcus agalactiae by biochemistry assays of ID 32 STREP test, Lancefield test and other assays. Molecular analysis of 16S rRNA and cfb genes (GBS-specific gene cfb, CAMP factor) were used for further identification. BLAST showed that 16S rRNA and cfb genes of all the strains possessed high similarities with their counterparts registered in GenBank, and with each other. Taken together, these experiment results showed that pathogen caused high mortality of tilapia in both provinces in 2009 was S. agalactiae. The result was helpful for tilapia disease control and prevention, and for development of streptococcus vaccine for tilapia.