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Abstract:In the aim of preventive microorganism resistance to industrial biocides, a collection of microorganism which isolated from industry production and raw material were gathered. To characterize bacteria species according to Manual of Systematic and Determinative Bacteriology, API Identification System, 16S rDNA genes analysis, and fungus according to Manual for the Identificati of Fungi, 18S rDNA genes analysis, respectively. The minimum inhibitory concentrations for biocides were determined to estimate the microorganisms resistance. It was found that the majority of isolates in the collection were Gram-negative bacteria (46.91%) including genera Pseudomonas, Enterobacter, Aeromonas, Klebsiella and Burkholderia. The dominant strains of Gram-positive bacteria (32.71%) were genera Bacillus, Microbacterium, Arthrobacter, Listeria along with several coccus. And fungi (12%) were Aspergillus, Trichoderma and Penicillium. The value of MICs showed that main resistant species is Pseudomonas (33.78%), its average level of resistance was up to 36 mg/L and the resistance of stability was weak with subculture. The result indicated that one of the considerable reasons of biological pollution in industry was the microorganism tolerance and the nature of the membrane structure and the biofilm formation of bacteria probably played a important role in the formation of microorganism resistance.

Abstract:

Abstract:Pichia galeiforms B-10 has a good ability to degrade organic acid in hemicellulose hydrolysate, and this depickling activity of the strain is mainly affected by the initial pH value of hydrolysate. If the initial pH value of hemicellulose hydrolysate was adjusted above 5.0, Pichia galeiforms B-10 would exhibit an excellent capability of depickling fermentation without any other treatment. During the metabolism of organic acid salt by Pichia galeiforms B-10, alkali products were generated and this caused the raising of pH value. Therefore, under the condition of pH > 5.0, if only the acid supplying speed (by adding hydrolysate with low pH value) was adjusted to be in balance with the acid metabolizing speed, the fermentation system would always be kept in a high pH circumstance which was advantageous to the metabolism of organic acid by yeast. This mode, by using bio?oxidation to produce alkali for the continuous depickling fermentation, effectively reduced the addition of alkali during the neutralization of hemicellulose hydrolysate, and finally lowered the processing cost and generated less pollutant.

Abstract:The a-amylase gene cn7a was amplified by PCR from Bacillus sp. CN7 genome DNA. The recombinant plasmid pSE380-cn7a was constructed by inserting gene cn7a into expression vector pSE380 and then transformed into Escherichia coli JM109. The purified amylase CN7A showed an optimal activity at pH 5.5?6.0 and 65°C, the Km value is 3.784 g/L taking soluble starch as substrate, and the maximum velocity was determined as 101.2 mg/(L·min). Ca ion was not required for the thermal stability of CN7A.

Abstract:HearNPV orf86 is a function unknown gene of Helicoverpa armigera nucleopolyhedrovirus (HearNPV). The coding region of orf86 was amplified from the HearNPV genome by PCR and then was inserted to the prokaryotic expression vector pGEX-4T-2. The recombinant plasmid was transformed into E. coli BL21 to express the GST fusion protein in the bacteria. The purified fusion protein was injected into New Zealand rabbits to prepare polyclonal antibody. The titer of the anti-HearNPV ORF86 polyclonal antibodies reached to 1 : 5.12 × 105 determined by ELISA. A 36 kD protein was identified by Western blot on HZAM1 cells infected by HearNPV using the prepared polyclonal antibody, which the protein size was consistent with the predicted protein size encoded by orf86. The results indicated that the polyclonal antibodies against ORF86 was successful prepared and it would facilitate the detection and related study on ORF86.

Abstract:A strain of photosynthetic bacterium capable of degrading the herbicide pyrazosulfuron-ethyl, designated as S8-1, was isolated and purified from the rice experimental field soil samples which have been successively used pyrazosulfuron-ethyl for many years by continuous enrichment culture techniques. The isolate exhibited substantial growth in mineral salt medium amended with pyrazosulfuron-ethyl as a sole source of carbon and energy. Based on the colony and cell morphological observation, absorption spectra of living cells, cultural and biochemical characteristics, and the sequence analysis of the partial 16S rRNA gene, the isolate S8-1 had been preliminary identified as Rhodopseudomonas sp.. High performance liquid chromatography (HPLC) data analysis revealed the optimum degradation conditions of strain S8-1 were: pH value at 7.0?7.5 and temperature at 30°C?35°C, respectively. At the concentration of 100 mg/L, 52.07% of pyrazosulfuron-ethyl was degraded in 7 days at pH 7.0, 30°C and 7500 lx of light illumination intensity. The strain S8-1 still maintained degradation activity at the concentration of pyrazosulfuron-ethyl up to 800 mg/L, additional yeast extract can significantly increase the growth speed and degradation rate. The result indicated that strain S8-1 has potential value for biodegradation of pyrazosulfuron-ethyl plant waste water or contaminated soils and waters.

Abstract:Growth behavior of Escherichia coli (E. coli) under the exposure to direct current (DC) was studied in this paper, with electrolytic stimulation on the culture flask of E. coli using gauze titanium and platinum electrodes. The electric field-activated mechanism was also investigated by cyclic voltammestry technique, constant direct current technique, SDS-PAGE as well as enzyme activity assay of ATPase. The results show that direct current can promote the growth of E. coli under the current density of 0.2275 mA/cm2, and the growth speed is gradually accelerated with increasing current density, but 0.0455 mA/cm2 is the optimal current density to obtain the maximum viable cells. When the same current density electric field is applied to Pt electrodes and Ti electrodes, the Pt electrode culture system is superior to Ti electrode culture system. The cathode electrolysis products of water are the primary factor for the difference, which include adsorbed hydrogen and H2. It is also found that the 0.091 mA/cm2 direct current can promote ATPase activity effectively, with 3.2 times at most after 8 hours. Bacterial proteins about 25 kD and 35 kD are expressed higher while proteins of 66.2 kD are expressed lower after (under) electrolytic stimulation of 0.0455 mA/cm2 direct current.

Abstract:Bacterial diversity of chilled chicken was investigated by culture independent and dependent approach. A total of 45 colonies were recovered from the plate culture of chilled chicken based on morphological characteristics and further identified as 6 genera, including Bacillus sp., Shigella sp., Pseudomonas sp., Citrobacter sp., Klebsiella sp. and Escherichia sp., according to their 16S rDNA sequences. Direct 16S rDNA-ARDRA coupled with sequences analysis of DNA extracted from chilled meat showed that the bacteria could be assigned to 16 genera of Acinetobacter sp., Bacillus sp., Acidovorax sp., Brochothrix thermosphacta, Lactococcus garvieae and Leuconostoc lactis etc. Compared to culture dependent method, culture independent approach disclosed a more diversified bacterial community in chilled chicken, but culture dependent one was still a necessary supplement by showing unique species when molecular analysis was used to investigate microbial diversity.

Abstract:Streptomyces albulus UN2?71 was selected as original strain and its protoplast was treated by diethyl sulfate (DES) to obtain high ε-polylysine producing mutants. Through primary screening and confirmed screening with shake flask cultures, one strain designated as D3-32 with a good genetic stability has been acquired, whose production in the shake flask reached 1.56 g/L, which was about 49.43% higher than that of original strain. In the fermentation test with 2.3 L fermentor, after a pH-controlled two-phase fermentation, the highest production of ε-polylysine reached 4.59 g/L, which was increased by 2.65 times compared to that of the original strain.

Abstract:Two hundred and four endophytes isolated from Catharanthus roseus, Cephalotaxus hainanensis and Antiaris toxicaria were screened for antitumor activities. The results showed that broth of 19 strains presented cytotoxic activities to at least one of the tested tumor cells, four of them exhibited strong antitumor activity against S180, were above 70%. Studies have shown that broth in the antitumor active substances of strains PA09006 and PA09009, confer a certain to pH value and ultraviolet.

Abstract:Using the methods of plate cultivation, 130 strains of endophytes were isolated from Rehmannia glutinosa Libosch, including 67 bacteria, 50 fungi and 13 actinomycetes. From them, 8 isolates of bacteria were selected due to their high and wide antagonistic activities to the tested microbial strains, Escherichia coli, Staphalocolcus aureus and Aspergillus niger on the plate confrontation. The 8 active strains of bacteria were classified to genus of Pseudomonas and shared the highest identities respectively with 5 species including Pseudomonas fluorescens, Pseudomonas thivervalensi, Pseudomonas chlororaphis, Pseudomonas koreensis and one unidentified according to their morphological, physiological and chemical characteristics and 16S rRNA sequences. Active products were obtained from the 8 strains through liquid fermentation and further extraction by different organic solvents. Activity studies indicated that the extraction products from all the 8 strains using ethyl acetate or ethanol exhibited significant inhibition on the 3 tested types of microorganisms and esophageal cancer cells Ec9706. Among them, strain 2-2 had the highest activities of both anti-microorganisms and anticancer showing important value of development.

Abstract:In the present study, an endophytic fungus isolate FTJZZJ09, which isolated from the fresh bulbs of Fritillaria thunbergii Miq., was identified as Penicillium chrysogenum based on its morphological characters and internal transcribed spacer (ITS) sequence. After being cultured in the modified CzapeK-DoX medium (3 g/L maltose, 3 g/L peptone A, 0.1 g/L K2HPO4, 0.05 g/L KCl, 0.3 g/L NaNO3, 0.05 g/L MgSO4·7H2O, 0.001 g/L FeSO4·7H2O, pH 6.5), it can secrete antibacterial metabolites under the condition of 28°C in a rotary shaker at 160 r/min for 7 days. Three antibacterial compounds were isolated from the ethyl acetate extract of the fermentation broth by silica gel, they were elucidated as cyclo (Pro-Gly), cyclo (Pro-Val) and 2-acetyl-4 (3H) quinazolinone. All the three compounds could inhibit the growth of Bacillus subtilis with the minimal inhibitory concentration (MIC) value of 0.8, 0.8, and 0.4 g/L respectively, while they showed no apparent effects against the growth of Gram-negative bacteria.

Abstract:Nineteen strains of endophytic fungi were isolated from the stems, roots and leaves of Paulownia fortune, their antimicrobial activity against three tested microorganisms were investigated by using the agar diffusion method and the paper disc method. Identification of these strains were determined by their morphological features and phylogenetic analysis based on internal transcribed spacer of rRNA gene sequence comparison. The results showed that endophytic fungi of JSD-8, JM-1 and JM-10 had remarkable antimicrobial activities to Bacillus subtilis CGMCC 1.769, Escherichia coli CGMCC 1.1103 and Candida albicans ATCC 10123, respectively. Fungal morphological and molecular identification demonstrated that JSD-8, JM-1 and JM-10 represented Gibberella moniliformis, Alternaria longipes and Penicilium thomii, respectively.

Abstract:An isolate of unkown bacterium was isolated from honeybee larvae infected with European foulbrood. Based on its characteristics of isolation and culturing, colony morphology, physiological-biochemical characteristics and Molecular Biology, this isolate was identified the secondary invader of European foulbrood, Enterococcus faecalis, which was belong to the genus of Enterococcus, and was named Enterococcus faecalis FB102 tentatively. Meanwhile it showed that people could different the bacterium from the pathogen of European foulbrood by the biological characters of isolates and cultivation. The bacterium could not infect honeybee larvae alone, and it could simplify the diagnosis process of European foulbrood by the isolation and culturing of Enterococcus faecalis, furthermore the bacterium from honeybee larvae might threaten human health.

Abstract:A fungal strain (No. 059601016C) was obtained from Laminaria Gametophyte Clones. Growth states, morphological characteristics and internal transcribed sequence (ITS) were studied. The results showed that colonics on PDA are white and cottony, the reverse color changed from white to dark purple. The aerial mycelium grew abundantly and the height reached 5 mm to 7 mm. Microconidia are in false head or chains, (5.0?10.5) μm × (1.2?2.5) μm. Macroconidia sickle-shaped were slightly bent, apex acuminate, 2?5 septate, mostly 3?4 septate. Based on the analysis of ITS and the phyogenetic tree, the strain showed a 100% homogeneous and the most close relationship to Fusarium proliferatum Nirenberg. Hence, the strain 059601016C isolated form Laminaria Gametophyte Clones belonged to the genus Fusarium sp. and the GenBank accession number of strain 059601016C ITS is GU951805.

Abstract:A strain of resistance to zinc with high concentration, named DX-T3-03, was isolated from the metal-polluted sediment of soils around Dexing copper mine. Basing on morphology, biochemical and physiological characterization, DX-T3-03 was characterized as Sphingomonas sp.. A phylogenetic tree was constructed with the published 16S rRNA sequences of relative bacteria species. In the phylogenetic tree, the DX-T3-03 strain has the closest relative to Sphingomonas aquatilis strain JSS-7 with 99% sequence homeology. The optimum growth conditions of DX-T3-03 strain were temperature of 35°C, pH of 6.7, rotating rate of 150 r/min. It could resist zinc of 25 mmol/L or above and be able to grow in a variety of single and combined heavy metals (Cu 70 mg/L, Cd 300 mg/L, Pb 400 mg/L, Ni 60 mg/L).

Abstract:Thermoanaerobacter ethanolicus produces ethanol from biomass derived substrates at temperatures above 70°C. AdhE, which exhibited bifunctional acetaldehyde/ethanol dehydrogenase activities, play a key role in producing ethanol. Proteins interacting with the transcriptional regulatory region (TRR) of adhE were for the first time isolated and identified by affinity chromatography. One of the proteins, PadhE-1, a homolog of LacI family, was cloned and expressed in Escherichia coli. Through electromobility shift assays, it was determined that the interaction between PadhE-1 and TRRadhE was specific. The research would be helpful to construct the bacteria with the extremely thermophilic and high ethanol-producing characteristics.

Abstract:We constructed a Rhizomucor Miehei Lipase(RML)-Displaying yeast whole-cell biocatalyst and applied it to methylesters synthesized from triglyceride and methanol. RML was fused with the Sed1p cloned from Saccharomyces cerevisiae to constructed plasmid pPIC9K-Flag-RML-Sed1. The plasmid was linearized and transformed into Pichia pastoris GS115 and Pichia pastoris recombinant strain GS115/pPIC9K-Flag-RML-Sed1 was obtained. Cell-surface display of the RML via Sed1p was confirmed by flow Fluorescence micrograph and flow cytometer. After incubated at 28°C for 48 h the hydrolytic activity of the GS115/pPIC9K-Flag-RML-Sed1 reached a plateau, 169.6 U/g (Dry cell weight). In nonaqeous media, the yield of 82.36% methylesters was obtained from triglyceride and methanol after 72 h using lyophilized RML displaying yeast whole cells.

Abstract:RNA interference (RNAi) is a post-transcriptional gene silencing mechanism that is triggered by double-stranded RNA (dsRNA) and results in the degradation of homologous transcripts. RNAi technology has been developed as a powerful tool for genetic screens to analyze gene function in various organisms. Construction of large-scale species-specific RNAi library and further RNAi mutant library is important strategies for functional genomics research. So how to construct RNAi libraries simply and efficiently is becoming a key question in gene function research. In the review, representative strategies for RNAi library construction and their advantages and potential disadvantages were summarized.

Abstract:Bacteriocins are antibacterial proteins produced by bacteria that can kill or inhibit closely related bacteria. Many lactic acid bacteria (LAB) produce a high diversity of different bacteriocins. Though these bacteriocins are produced by LAB found in numerous fermented and non-fermented foods, nisin is currently the only bacteriocin widely used as a food preservative. Compared with antibiotics, bacteriocins are ribosomally synthesized. Their transcript must be modified before becoming active and are translocated to the outside of the cell by a transporter system. Bacteriocins inhibit target cells by acting on the membrane, while the cell synthesizing the bacteriocin has immunity to its product. Therefore, bacteriocins should be safely and effectively used to control the growth of target pathogens, make it have wide applications in many food systems.

Abstract:Aerobic hydrogen-oxidizing bacteria can utilize H2 as electron donor and O2 as electron acceptor to fix CO2. Recently, it has been found that hydrogen-oxidizing bacteria, which are the main physiological groups in rhizosphere of leguminous crops, can promote the growth of plants obviously. Nowadays, great attention has been payed to this kind of bacteria. Hydrogen-oxidizing bacteria have great potential in agriculture, environment and human health. But it’s not very clear about the classification, isolation and identification of hydrogen-oxidizing bacteria, and it has seriously affected the process of researches. This article presents an overview on the isolation, screening of hydrogen-oxidizing bacteria, focuses on plant growth-promoting mechanism and some other prospects of their applications. As the further study, more and more researchers all over the world will pay more attention to this kind of bacteria.

Abstract:Microbiology experiment is an essential subject of biotechnology in colleges and universities. In the teaching process, the consciousnesses of active participation, aseptic technique, problems finding and solving independently, innovation and green environmental protection should be strengthened. In this paper, we analyzed these issues and put forward the solving strategies and strengthening measures.

Abstract:Bilingual teaching is one of the methods advocated by the Ministry of Education of the People’s Republic of China. Based on characteristics of common university and the practice of teaching, we analyzed the feasibility of carrying out bilingual teaching in the common university, and some useful experiences accumulated from practice for bilingual teaching in choosing textbooks, formulating learning outlines, implementing teaching manners and so on.

Abstract:Two standard molecules were constructed by molecular clone technology of Tilletia indica Mitra and Tilletia walkeri Castl.. The standard molecule for Tilletia indica Mitra contained two DNA fragments, one about 2297 bp DNA sequence from mitochondrial DNA, the other ITS sequence 710 bp. The standard molecule for Tilletia walkeri Castl. contained two DNA fragments, one about 2.3 kb DNA sequence from mitochondrial DNA, the other ITS sequence 710 bp. In this research, the two standard molecules were detected, the results show good specificity, homogeneity and stability which can satisfy the need of molecular detection of Tilletia indica Mitra and Tilletia walkeri Castl..

Abstract:This paper introduces a method for single-spore isolation of Exserohilum turcicum with general light microscope. Spores are displaced from the corn leaf to Water Ager by tapping, single-spore of Exserohilum turcicum are then picked with simple homemade needle at low magnification so as to achieve purification. Although the method was slightly sophisticated, it was simple, practical, less contaminating and highly effective. The application of the method is contributive to rapid identification of physiological races and DNA genetic polymorphism of Exserohilum turcicum. It is more suitable for single-spore isolation of the pathogenic fungi with large conidiospores.

Abstract:Based on nitrocellulose membrane, we optimized the blocking condition, the volume and concentration of coating antibody. Then, an improved Double Antibody Sandwich Dot Enzyme Linked Immunosorbent Assay (DAS-Dot-ELISA) for Acidovorax avenae subsp. citrulli, the casual agent of bacterial fruit blotch of watermelon, was established. Use adding Disodium Ethylenediamine Tetraacetic Acid (EDTA) in nonfat dry milk treated with high temperature as blocking solution can effectively lower the background. Shaking gently can enhance hybridization and reduce the nonspecific binding. This improved DAS-Dot-ELISA assay detected Acidovorax avenae subsp. citrulli as low as 1.9 × 105 CFU/mL quickly and economically. On the detection of two batches seed sample, the germ-carrying rate with improved DAS-Dot-ELISA was 8.0%, 6.0% which was completely coincident with microwell plate ELISA. On the result of each seed, this improved method was coincided with microwell plate ELISA in 99%, showed a satisfactory prospect and provided a new rapid assay for the detection of Acidovorax avenae subsp. citrulli.