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2012, 39(5):0683-0695.Abstract:
[Objective] Recently, as the prevalence of multiple antibiotic resistant bacteria in environments are induced by the extensive use of antibiotics by human, a better understanding of the distribution, transport and dissemination of antibiotic resistance genes (ARGs) in environments is warranted. The objective of this study was to determine the pollution levels of antibiotic resistance genes in Jiulong River estuary and wastewater treatment plants (WWTPs) in Xiamen. [Methods] Polymerase chain reaction (PCR) assays were employed to identify the distribution patterns of four sulfonamide, thirteen tetracycline ARGs and two integron genes in water, sediments of Jiulong River estuary and five activated sludges in WWTPs of Xiamen. Clone libraries of the tet(W) gene were also generated from three sediments of Jiulong River estuary and five WWTPs in Xiamen. [Results] The all ARGs has been dectected except tet(O) and tet(S). The results demonstrated that the frequency of detection (FOD) of ARGs and integron genes in environmental samples with the highest FOD in activated sludges (0.86), moderate FOD in sediments (0.57) and the lowest FOD in water (0.24). Furthermore, the FODs of sul(l), int(1), tet(A), tet(C), tet(E), tet(M) and tet(W) were higher in freshwater and brackish water than those in seawater, suggesting that ARGs may be transported from the upstream of Jiulong River. [Conclusion] Principle component analysis also supported the suggestions that WWTPs contained the highest ARGs, and ARGs are enriched in sediments but unstable in water. In addition, the results of tet(W) libraries indicated that WWTPs may be a point source of ARGs into Jiulong River estuary and Xiamen sea.2010, 37(12):1825-1829.Abstract:
Real-time quantitative PCR is a commonly used method to analyse the gene expression profile, it is important to select an appropriate reference gene for normalization of experimental data when using this method. In our study, we used two statistical methods to evaluate the gene expression stabilities of five reference genes (ldh, recA, rpoB, gapdh and 16S rRNA) under the different growth phases of Lactobacillus helveticus H9. The results showed that the best reference gene was ldh which was the most stable gene would be used for normalization of real-time quantitative PCR experiments data.2008, 35(6):0967-0976.Abstract:
Based on the data of the ISI “Journal Citation Report” in 2006 edition, a statistics for the 88 microbiology science journals has been made. The results showed that these 88 learned journals were published in 16 countries with 3 languages, especially English. And the main countries were United States, England, Netherlands and Japan. The average data of articles and total cites of the 88 journals were 165 and 5226.64, respectively. Moreover, the average data of impact factor and immediacy factor were 3.154, 0.45 and 5.84, respectively. Finally, fifty-five learned journals with impact factor over 2.0 were introduced from these journals to be referenced.2012, 39(6):0853-0864.Abstract:
[Objective] This work aims to investigate differential expression proteins at different development stages of Volvariella volvacea using isobaric tags for relative and absolute quantification (iTRAQ)-coupled two-dimensional liquid chromatography tandem mass spectrometry (2D LC-MS/MS) proteomics approach. [Methods] Firstly, the proteins extracted from V. volvacea at different development stages were analyzed by SDS-PAGE. Secondly, the tandem mass spcetrometry data obtained from 2D LC-MS/MS were used to search using MASCOT search engine. After that, principal component analysis, hierarchical clustering and Gene Ontology were used to analyze the detected results. [Results] Results showed that 1 039 protein groups, included a total of 2 335 unique peptides, were identified. Among them, 1 030 protein groups were provided quantitative information. Contrast to mycelia, 64 up-regulated and 150 down-regulated significantly differential proteins were found in fruiting bodies at different development stages of V. volvacea. Bioinformatics analysis revealed that iTRAQ-coupled 2D LC-MS/MS was a unique method for isolating and identifying protein groups of V. volvacea at different development stages. [Conclusion] It is helpful to insight into the molecular mechanism of the fruiting body formation and development of V. volvacea and other macro-basidiomycetes in the future.2008, 35(1):131-136.Abstract:
Obtaining soil bacterial DNA of good quality is a key step in soil bacterial ecology study. A quick, efficient, sensitive and stably method of DNA extraction from soil were established by combining strongpoints of two kits ( Soilmaster kit and DNA IQTM kit). In addition, the 16S rDNA gene and T-RFLP (Terminal restriction fragment length polymorphism) were used in the analysis of soil bacterial community diversity and the result show that T-RFLP is a powerful tool for bacterial community study.GUO Lei , ZHU Wen-Cheng , LIU Wei-Wei , DING Guo-Wei , ZHANG Yue-Qian , ZHANG Shou-Lu , QIU Yue , XIE Yu2013, 40(6):951-958.Abstract:
[Objective] To identify marine fungus HN4-13 isolated from Lianyungang coastal sea sediments with antibacterial activity and to optimize the fermentation conditions of synthesizing antibacterial active substances. [Methods] Strain HN4-13 was identified based on its morphological characters and internal transcribed spacer (ITS) sequence; The optimization of fermentation conditions for antibacterial production by strain HN4-13 were investigated by the one-factor-at-a-time method and the orthogonal design method. [Results] Strain HN4-13 was identified as Aspergillus flavipes, and the optimal fermentation conditions can be recognized as follows: 4% sucrose, 0.5% peptone, 0.1% KCl, 0.06% NaH2PO4, 1% inoculation concentration, 28 °C, 160 r/min for 9 days. [Conclusion] The results provide the clues for further separation and purification of the antibacterial active metabolites derived from strain HN4-13.2011, 38(12):1755-1761.Abstract:
The pullulanase gene of thermophilic Bacillus subtilis WY-34 was cloned in E. coli to characterize the recombinant pullulanase. The pluA gene encoding pullulanase from the thermophilic B. subtilis WY-34 was overexpressed in E. coli. The properties of recombinant pullulanase were studied. Also substrate specificity of the recombinant pullulanase was evaluated in this paper. The pluA gene was 2.2 kb in length and encoded 718 amino acid protein. The recombinant pullulanase was purified to homogeneity using Ni-IDA agarose chromatography, resulting in a specific activity of 93.2 U/mg. SDS-PAGE and gel permeation chromatography analysis showed that the molecular weight of the protein is approximately 76.2 kD and 74.3 kD respectively. The purified enzyme was optimally active at 40 °C and pH 6.0, stable at 45 °C and retained more than 80% relative activity with in pH 6.0?9.0. It exhibited high substrate specificity toward pullulan. The properties of recombinant pullulanase may be useful for application of pullulanase in industry.2009, 36(3):0427-0431.Abstract:
Cyclic diguanylate (c-di-GMP) is a bacterial second messenger of growing recognition involved in the regulation of a number of complex physiological processes. In combinations to the related progress of Xanthomonas oryzae pv. oryzae, the causing agent of bacterial blight of rice in our lab, this review describes (1) the biosynthesis and hydrolysis of c-di-GMP and several mechanisms of regulation of c-di-GMP metabolism, (2) the contribution of c-di-GMP to regulating virulence, motility and biofilm formation, processes that affect pathogenesis of many bacteria, and (3) ways in which c-di-GMP may mediate these regulatory effects.2011, 38(10):1526-1532.Abstract:
Five endophytic bacteria containing ACC-deaminase were isolated from the roots of Eucommia ulmoides Oliver based on their ability to utilize ACC as a nitrogen source, and their antibacterial activities were determined by using the agar disc diffusion test. Identification of these isolates was determined by their morphological, physiological-biochemical properties and phylogenetic analysis based on 16S rRNA gene sequences. The ACC deaminase activity assay demonstrated that the five endophytic bacterial isolates expressed the ACC deaminase activity, and four isolates of them displayed antibacterial activity against Escherichia coli CGMCC1.1103 and Bacillus subtilis CGMCC1.769, respectively. Five ACC deaminase-containing endophytic bacteria isolates named as JDM-2, JDM-8, JDM-11, JDM-14 and JDM-19 were identified as Pseudomonas koreensis, Klebsiella pneumoniae, Enterobacter ludwigii, Klebsiella variicola and Enterobacter asburiae, respectively.2013, 40(1):98-108.Abstract:
Microorganisms are the engines driving the biogeochemical cycles of soil elements. The nitrogen cycle is one of the central processes of terrestrial ecosystems, and contains four main steps, i.e. nitrogen fixation, ammonification, nitrification, denitrification, all of which are driven by microorganisms. In the last decade, with the rapid development of culture-independent molecular techniques and high-throughput sequencing technologies, breakthrough progress has been made on the diversity and mechanisms of nitrifying microorganisms, and anaerobic ammonia oxidation (anammox) process and mechanisms. Here we review the available knowledge concerning the research progress in ammonia oxidation studies in China, and briefly introduce the researches on denitrifying microorganisms, anammox, and dissimilatory nitrate reduction to ammonium (DNRA), and then present the future perspectives on this research field. Novel techniques and methods will be applied to the future microbial ecology studies of soil nitrogen transformation. It needs to hold the frontiers of microbial ecology, in combination with significant demands for China’s sustainable agriculture, resources and environment protection, and global change research, primarily focusing on the following several areas: (1) to carry on investigations on the large-scale biogeographical patterns of soil nitrification processes and nitrifying microorganisms, and to elucidate the underlying mechanisms of spatial-temporal variations and their driving factors. (2) To explore the key microbe-meditated processes and mechanisms of nitrogen transformation, and link them to the relevant observations on gas flux (e.g. ammonia volatilization, N2O emissions) and reaction rates (e.g. the rates of mineralization and nitrification). (3) To elucidate the coupling between different nitrogen transformation processes in certain ecosystems, and to estimate the nitrogen balance and build models to predict nitrogen transformation and balance. These studies are expected to provide scientific basis for adjustment of nitrogen transformation processes, improvement of nitrogen utilization efficiency and elimination of its negative effects.2008, 35(7):1119-1123.Abstract:
There are one third of synthesized proteins must be secreted to the cell surface or to the surrounding environment to acquire their native functional state. Most of them are exported by Sec translocase (secretion pathway). Sec translocase consists of a membrane embedded protein-conducting channel, termed SecYEG and a peripherally associated motor domain, the ATPase SecA. The SecDFyajC heterotrimeric membrane protein complex can facilitates protein translocation. SecB is a molecular chaperone that functions in the protein translocation pathway. SecM (secretion monitor) encoded by the 5' region of the secM-secA mRNA, which elongation arrest is required for upregulated expression of SecA. The signal sequence in the N terminus of the nascent peptide is first recognized by the signal recognition particle (SRP). SecB, the Sec-system-specific chaperone, channels the preprotein to the Sec translocation pathway and, additionally, actively targets the bound precursor to the translocase by its ability to bind SecA. The preprotein-bearing SecA then binds to the membrane, at a high-affinity SecA-binding site, SecYEG, which constitutes a channel for polypeptide movement. Continued translocation requires cycles of ATP hydrolysis by SecA, which is thought to occur in a step-wise fashion with a step of 20~30 amino acid residues.2012, 39(1):0125-0137.Abstract:
[Objective] To establish a simple method to screen oleaginous yeast and determine the intracellular lipid content. [Methods] The study is based on the theory that the combination of nile red and intracellular oil will emit fluorescence when induced by UV light and the fluoresence indensity is associated with the lipid content. We cultivate yeast in the culture medium added with nile red, and screen the oleaginous yeast strains from the 385 deep-sea yeasts by measns of obeserving the fluorescence of the yeast colony. We have identified the screened oleaginous yeast strains by the 26S rDNA D1/D2 series analysis method. Designating one of the oleaginous yeasts (2A00015) as the test strain, the lipid content rapid determination method by nile red dyeing was established. [Results] 22 oleaginous yeasts were obtained with the lipid content reaching up to 62.9%. Based on the molecular identification, it showed that the 22 yeasts are separately belong to Candida viswanathii, Candida parapailosis, Rhodotorla mucilaginosa, Debaryomyces hansenii, Pichia guilliermondii and Rhodosporidium paludi-genum. The optimum condition for lipid content determination by nile red dyeing is: bacterium suspension OD600 lower than 1.2, nile red concentration 0.5 mg/L, dyeing time 5 min, excit-ing wavelength 488 nm, emission wavelength 570 nm. The relative fluorescence intensity ob-tained by this method exhibits a positive association with the lipid content obtained by weigh-ing method, which can be explained as R2=0.9637.2013, 40(1):44-58.Abstract:
Human activities introduced increased amounts of nitrogen in coastal oceans, causing eutrophication and numerous ecological and environmental problems. It is crucial to better illustrate and understand the function and contribution of the microbe-driving nitrogen cycle within the coastal ecosystems, especially under the global change background. This review focuses on the rates, fluxes, contribution and functional gene quantity of microbes in nitrogen fixation, ammonification, nitrification, denitrification, dissimilatory nitrate reduction to ammonium, and anammox in coastal sediments. The controls of major physiochemical and biological factors (e.g. temperature, dissolved oxygen, salinity, labile dissolved organic carbon, dissolved inorganic nitrogen, submerged?macrophytes and benthic animals) on these processes, as well as functionally related microbial groups and pathways (e.g. ammonia-oxidizing bacteria and archaea and nitrate reduction), are summarized.ZHU Jing , HU Dan , LIU Li-Na , ZHANG Jin-Hai , ZHANG Feng-Yu , HAO Li-Na , GENG Mei-Ling , ZHENG Feng , ZHU Jin , PAN Xiu-Zhen , WANG Chang-Jun2013, 40(6):1058-1067.Abstract:
[Objective] To explore the mechanism of signaling pathways Streptococcus suis serotype 2 infected monocytes/macrophages leaded, discuss the role of capsular sialic acid component played in Streptococcus suis activate macrophage TLR2-AKT-NF-κB signaling pathway. [Methods] RAW264.7 as the target cell line, RT-PCR, Western blotting, immunofluorescence and ELISA were applied to detect different infection time of wild type strain, sialic acid knockout strain and sialic acid complementary strain on macrophages TLR2 mRNA transcription level, AKT phosphorylation level, NF-κB activation level, as well as TNF-α secretion level. Pretreat with TLR2 blocking agent and PI-3K inhibitor on macrophages, detect the expression level above. [Results] Sialic acid knockout strain activates signal transduction pathways selectively. RT-PCR results show that TLR2 mRNA expression levels began to increase at 1 h, 1.5 h reached its peak then slowly decline. Western blotting showed that TLR2 protein expression level reached its peak at 7 h, 9 h decline. Level of p-AKT is stable at its peak during 1.5?5 h, 7 h decline. Immuno fluorescence showed high level of NF-κB activation-nuclear translocation at 15 min. ELISA results indicate TNF-α secretion level was significantly higher than the other two strains after 10 h. TLR2 blocking agent and PI-3K inhibitor significantly suppressed the activation degree of three strains. [Conclusion] Capsular sialic acid could inhibit activation of the TLR2-AKT-NF-κB signaling pathway to some extent, thus participate in bacteria evading the host immune defense.15 Isolation and Identification of Marine ActinomyceteJMC06001 Exhibiting Strong Antibacterial ActivityCHEN Yi-Guang , ZHANG Xiao-Rong , ZHANG Li , LIU Zhu-Xiang , PENG De-Jiao , XIAO Huai-Dong , HUANG Ke , CHEN Qi-Hui , XU Li-Hua2008, 35(1):40-44.Abstract:
One slightly halophilic marine actinomycete strain JMC 06001 was isolated from a saline mud sample collected from the Island Naozhou in the South China Sea, near Zhanjiang, a city of southern China. The fermentation broth of strain JMC 06001 strongly inhibited the growth of Staphylococcus aureaus, Sarcina lutea, Bacillus subtilis, Escherichia coli, Micrococcus luteus and Proteus vulgaris. The combination of morphology, physiological and biochemical characteristics, chemotaxonomic data and 16S rRNA gene sequence analysis supported the view that strain JMC 06001 belong to a known species of the genus Steptomyces, S. peucetius. The strain studied grown well on most media tested, with white aeriel hyphae and pale yellow to pale brown substance hyphae. Yellow diffusible pigments were produced on oatmeal agar (ISP 3), potato extract agar and glucose/asparagines agar, and pale brown to deep brown diffusible pigments were produced on yeast extract/malt extract agar (ISP 2), glycerol/asparagine agar (ISP5), peptone-yeast ext-Fe agar (ISP 6) and nutrient agar. Growth occurred at 4 ℃~40 ℃ and pH 6.0~9.0, with optimum growth at 28 ℃ and pH 7.0. The tolerant range of NaCl was 0~1.5 mol/L, with best growth occurring in media containing 0.2~0.5 mol/L NaCl.-
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