Table of Content

29 October 2017, Volume 33 Issue 10

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Review

Research Progress on the Functional Microorganisms in Enhanced Biological Phosphorus Removal(EBPR) Systems

WEI Ru-ping, YAN Cheng, YANG Xin-yan, HE Xiao-yun, WANG Xin, YANG Liu-yan

2017, 33(10):  1-8.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0558

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Phosphorus emission from human activities is one of the major causes of water eutrophication. The technology of enhanced biological phosphorus removal（EBPR）has been widely applied for removing phosphorus from wastewater due to its advantages of economy and sustainability. The key to the efficient and stable operation of EBPR system is that the microbial community in the system is reasonable and functional. For deeply understanding the mechanism of phosphorus removal and the efficient and stable operation of EBPR system，a large number of studies were conducted on the microbial community structure and main functional microorganisms in the system. In addition to polyphosphate-accumulating organisms（PAOs），the EBPR system also includes non-PAOs，mainly glycogen-accumulating organisms（GAOs）and some auxiliary bacteria. Nowadays，more and more types of functional polyphosphate-accumulating microorganisms were identified，and the most studied PAOs and GAOs were Accumulibacter and Defluviicoccus respectively. PAOs and GAOs compete or cooperate under different environmental conditions，but it is still controversial whether PAOs may exhibit the metabolic properties of GAOs under certain conditions. Besides the traditional factors such as carbon source，pH and temperature，the effects of exogenous pollutants，like antibiotics and heavy metals，on functional microorganisms in EBPR system also exist. The traditional separation methods，blue/white screening and the method of artificial construction of gene engineering bacteria have been applied for acquiring efficient PAOs. The development of modern molecular biology technology provides advanced and reliable technical means for the research of functional microorganisms in EBPR system，and the efficient removal of phosphorous by constructing high-performance polyphosphate-accumulating microorganisms will be an important trend in future increasing bio-removal phosphorus efficiency from actual wastewater.

Microbial Remediation of Soil Heavy Metal and Organic Pollutants：Bioaugmentation and Biostimulation

HAO Da-cheng, ZHOU Jian-qiang, HAN Jun

2017, 33(10):  9-17.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0294

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Green technologies，e.g.，bioaugmentation and biostimulation，are cost effective and eco-friendly，and are becoming Rosetta Stone for the remediation of soil heavy metal and organic pollutants. Various bacteria，fungi and their consortia can be combined with physical and chemical techniques for the targeted remediation of soil organic contamination，heavy metal contamination and combined pollutions. This review exemplifies the representative studies within the recent five years，summarizes the screening of microbial strains from multiple sources and their potentials in transforming/degrading various types of pollutants，and highlights their applications in the case studies of laboratory，greenhouse and field conditions. The complexity of microbial remediation is ascribed to not only the variations of physiological and metabolic traits，but also numerous environmental factors，including abiotic factors，e.g.，pH，temperature，type of soil，pollutant concentration，content of water and organic matter，additional carbon and nitrogen sources，and biotic factors，e.g.，inoculum size，interactions between the introduced strains and autochthonous microbes，and the survival of inoculants. Prospects of combined remediation and the applications of molecular methods are also presented.

Review of Petroleum Hydrocarbon Pollution and Microorganism Remediation

GUO Ping, LI Hong-na, LI Feng, YE Jing

2017, 33(10):  18-25.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0799

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petroleum hydrocarbon pollution is one of the most severe environmental pollution in abroad and china，the restoration of environment ecology and remediation of petroleum hydrocarbon pollution by microorganism technology have revealed advantage in past practice. With the economic development rapidly，the quickly increased requirement for petroleum hydrocarbon highlights the pollution problem. The challenge is regarding to this technology under more severe pollution and environmental deterioration. This paper address the trend of petroleum pollution and technology application，the analysis of petroleum hydrocarbon degradation microorganism classification，the molecular mechanism of petroleum hydrocarbon degradation by microorganism and the application of microorganism remediation technology，and cater for understanding the challenge more further.

Research Progress on in situ Detection Methods of Microorganisms

SONG Wei-feng, LI Ming-cong, GAO Zheng

2017, 33(10):  26-32.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0550

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As major participants in ecosystem material cycle and energy flow，microorganisms play an important role in the ecosystem. However，the proportion of the cultivable microorganism under the existing technology is very small，which limits the exploit of microbial resources. At present，there are a number of approaches that can avoid the problem of uncultured microorganisms，which are designed to study in situ microbial activity. Regarding this，we summarized some research methods of studying in situ microbial ecology，allowing it convenient to reasonably use these methods in the future. This article introduces the corresponding microbial detection methods of BrdU-labeling，DNA-SIP，fluorescence in situ hybridization（FISH），and environmental transcriptome from DNA level，RNA level and protein level，respectively，and compares their advantages and disadvantages. It also introduces how to apply these methods combined with popular high-throughput sequencing and single cell sequencing technology to capture the in situ activity of microbial groups. At the same time，comparing the characteristics of these methods，so that we can more clearly understand the choice of different methods under different scenarios. These modified methods combined with other methods will be conducive to solve many have-been or will-happen problems in the study of microbial ecology. The ecosystems on the earth are complex and huge，in which the microbial populations vary. A variety of in situ detection methods have made a more realistic and effective description for the physiology and ecology of microorganisms，which will become a powerful tool for the study of microorganisms.

Research Progresses on Lignocellulose Degradation by a Thermophilic Anaerobic Bacterium Caldicellulosiruptor bescii

CHU Yin-di, SU Xiao-yun

2017, 33(10):  33-39.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0533

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As a gram-positive anaerobic bacterium isolated from hot spring，Caldicellulosiruptor bescii has strong ability in degrading lignocellulose. It can rapidly grow on a variety of model plant cell wall polysaccharide compounds such as the crystalline cellulose avicel，xylan or even on unpretreated lignocellulose such as switchgrass as sole carbon source. Moreover，this bacterium has an unusual ability of anaerobic degradation of lignin. The genomic annotation showed that most of the proteins encoded by this bacterium were multivariate bi-functional enzymes，i.e.，the N-terminal and C-terminus of the polypeptide chain were glycoside hydrolases of different families，with 2-3 carbohydrate binding domains. The genes encoding enzymes of degrading cellulose were concentrated in a plant cell wall polysaccharide degradation gene cluster，such as cellulase/xylanase，cellulase/mannanase，cellulase/xyloglucanase，etc. The xylanase of C. bescii belonged to the GH10 family，whose specificity of the enzyme was broad，and the homology of the amino acid sequence was between 18.7% and 59.5%. The genus Caldicellulosiruptor evolved a series of mechanisms that allowed glycoside hydrolyses to absorb better to substrates，bacteria and lignocellulose，thereby facilitating the enzymatic hydrolysis of lignocellulose. There were 12 proteins containing SLH domain，and the newly discovered adhesion protein Tāpirin in C. bescii may be involved in the absorption and utilization of lignocellulose. In this paper we review the current progresses in exploring the genome of C. bescii for novel glycoside hydrolases targeting plant cell wall and the associated molecular mechanisms，which are of great significance for the design and optimization of efficient and multi-function lignocellulose degradation enzymes.

Research Progress of New Biomarker Gene of phoH for Bacteriophage Genetic Diversity

LI Xiang, SUN Yan, WANG Xin-zhen, LIU Jun-jie, WANG Guang-hua

2017, 33(10):  40-45.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0725

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Bacteriophages（phages）are the simplest biological entities，which play important roles not only in the regulation of their host communities and horizontal transfer of genetic materials，but also in the element biogeochemical cycles. Although the importance of phages is increasingly being paid attentions by researchers，however there are still great limitations on the study of phage gene diversity until now. Therefore，the work of finding and verification of a suitable molecular marker gene for studying the genetic diversity of phage in natural environments is one of the hotspots of virus ecology. In this paper，a new molecular biomarker gene of phage auxiliary metabolic gene of phoH was introduced，and the recent advances in using this gene to reveal the phage genetic diversity in marine waters，paddy floodwaters and desert hypoliths are reviewed. Results showed that the distribution of phage phoH gene was closely related to its living environments，and phoH clones were divided into several groups according their obtained environments. In the end of the paper，the future research directions of phage phoH gene are also proposed.

Applications of Genomics，Proteomics and Metabolomics in Microbial Degradation of PAHs

KONG De-kang, WANG Hong-qi, XU Jie, LIU Zi-li, WU Xiao-xiong

2017, 33(10):  46-51.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0368

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Genomics，proteomics and metabolomics are important components of systemic biology，which are the emerging disciplines developed in recent years. With the development of life sciences，genomics，proteomics and metabolomics have been developing rapidly，and widely used in various fields of environmental microbiology，and become an indispensable part of studying polycyclic aromatic hydrocarbons（PAHs）microbial degradation. In this paper，the latest research progresses on the mechanisms of microbial degradation of PAHs and the metabolic pathways in three disciplines are reviewed，and the application prospects and challenges of the three disciplines in the PAHs biodegradation are discussed.

Research Progress on Ecotoxicity and Microbial Degradation of Strobilurin Fungicides

FENG Yan-mei, FAN Xing-hui, ZHAN Hui, TENG Shi-yu, YANG Fang, CHEN Shao-hua

2017, 33(10):  52-58.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0581

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The widely used strobilurin fungicides in agriculture have become a public concern because of their heavily environmental contamination and toxic effects on mammals. Such concerns，therefore，highlight the critical need for effective and advanced remediation technologies for the removal of strobilurin residues. Recently，bioremediation has emerged as a great potential alternative to eliminate pesticide residues because of its effective，low-priced，and eco-friendly properties. In this review，we summarized the research status of ecotoxicity of strobilurins and their microbial degradation. The main metabolites and the proposed degradation pathways of strobilurins were investigated. Moreover，the deficiencies of microbial degradation and the future development were discussed. These results may lay a solid foundation for developing new and safe measures for solving pollution hazard from strobilurin-like residues.

Research Progress on Bioaccumulation of Heavy Metals in Wastewater by Filamentous Fungi

MA Yan, YU Xiao-bin

2017, 33(10):  59-63.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0516

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Wastewater containing heavy metals is the one that most seriously pollutes environment and damages human health. Bioaccumulation of heavy metals by filamentous fungi is an important mean for processing heavy metal wastewater，and it has been a hot research topic in recent years. Firstly，the advantages and disadvantages of conventional methods of removing heavy metals from wastewater were reviewed，including chemical precipitation，ion exchange and adsorption. Among them，biosorption owns special advantages，such as low price and low consumption of biosorbents，its high effectiveness in absorbing the heavy metal ions，wide application and biosorbents reusable，thus biosorption is particularly suitable for the treatment of trace heavy metal wastewater. Secondly，the types of filamentous fungi that were applied to the removal of heavy metals were introduced，such as Rhicopus（Pb²⁺，Cd²⁺，and Cr^{5+），Mucor（Pb2+，Ni2+，Cd2+，and Zn2+），Aspergillus（Pb2+），Trichoderma（Zn2+ and Pb2+），and Basidiomycota（Cu2+，Pb2+，and Cd2+）；they all presented ideal results in removing heavy metals. Thirdly，the mechanisms of bioaccumulation were summarized，mainly about extracellular，cell surface，and intracellular absorption of heavy metals. Lastly，the several factors affecting the efficiency of removing heavy metals（pH，initial concentration of heavy metal ions，proportion of adsorbent，temperature，and coexisted ions）were discussed，as well as the future of studying filamentous fungi was prospected. It is aimed at expanding the application of filamentous fungi in the heavy metal accumulation from wastewater and laying a foundation for initiating new types of filamentous fungi in accumulating heavy metal from wastewater.}

Research Progress on Electricigens in Microbial Fuel Cell

ZHANG Xia, XIAO Ying, ZHOU Qiao-hong, WU Zhen-bin

2017, 33(10):  64-73.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0222

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Microbial fuel cell（MFC），one of the novel environmental and energy technologies，is attracting attention of numerous researchers. MFC is a device that utilizes microorganisms to convert chemical energy from organic matter into electrical energy. As a biocatalyst，electricigens play a key role in the development of MFC. Different kinds of electrogenes have different electron transfer mechanism and ability，affect the production performance of MFC directly，then determine the MFC performance and application in engineering practice. Any waste water，sludge，and sediment containing a large amount of microorganisms，try to screening high efficient electrogenes isolated from different environment is expected to promote the further perfect of MFC，so as to accelerate its application in the environment. The latest progress of electricigens in MFCs from several aspects were summarized，including of the development of MFC’s operation，the electricigens species and their electron transfer mechanism，specifically including the screening methods of electricigens，types and the related technology research. Finally，the main research trends of electricigens and the potential application of MFC in the future are also listed，aiming at providing the corresponding theoretical basis and new ideas for the screening and application of electricigens.

Study Progress on Microorganism in Constructed Wetlands

WANG Yu-xuan, WEI Wei, LI Ping-ping, ZHAO Yun, FU Wei-guo

2017, 33(10):  74-79.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0125

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Constructed wetland is a sewage processing technology engineering developed rapidly since the late 1970s. It has the advantages of low investment，low running cost，low-level maintenance technology and low energy consumption. Currently，it has been used to process various types of waste-water. Constructed wetland is a composite system composed of matrix，plant，and microorganism，in which microorganisms play an important role in the process of waste-water purification. This paper introduces the research progress of microorganisms in the constructed wetland system from the diversity and function of microbial community，and further gives the research prospects in this field，aiming at providing information and reference for relevant researches in this field.

Research Progress on Microbial Quorum Quenching Enzymes and Their Control of Plant Diseases

FAN Xing-hui, WANG Hui-shan HE, Jie-hua YE, Tian YANG, Fang CHEN Shao-hua

2017, 33(10):  80-87.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0223

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The phenomenon of information exchanging through signal molecules between microbial cells is the Quorum Sensing（QS）. QS is widely present in microorganisms，and can modulate the expression of specific genes，especially virulence factors in pathogenic microorganisms. Quorum Quenching（QQ）is a new therapeutic strategy of plant diseases based on QS. QQ can be achieved by interfering QS from inhibiting the synthesis or detection of the signal molecules，or by enzyme-catalyzed degradation or modification of the signal molecules. Degrading signal molecules by using QQ enzymes is one of the most effective and nontoxic ways of QQ. So far，QQ enzymes of various microbial signaling molecules have been reported，especially N-acyl homoserine lactones（AHLs）and cis-11-methyl-2-dodecenoic acid were deeply investigated. In this review，we summarize and analyze the research status，deficiencies and the future development of QQ enzymes and their control of plant diseases，for laying a solid foundation for developing new，green and safe measures of disease control.

Research Progresses on Strain Construction of Xylose Isomerase-based Recombinant Xylose-fermenting Saccharomyces cerevisiae

LI Yun-cheng, MENG Fan-bing, GOU Min, SUN Zhao-yong, TANG Yue-qin

2017, 33(10):  88-96.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0448

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It is of utmost significant to construct industrial xylose-fermenting Saccharomyces cerevisiae strains for lignocellulosic bioethanol production through the heterologous expression of xylose metabolic pathway. Xylose isomerase pathway is regarded as the most promising pathway expressing in S. cerevisiae，since there is no xylitol accumulation resulted from the coenzyme imbalance in xylose redox pathway. In heterologous expression of xylose isomerase，selecting an industrial S. cerevisiae strain as initial strain is of outstanding advantages for lignocellulosic bioethanol production. Concurrently，improving the expression efficiency of xylose isomerase gene（xylA）is vital for constructing a robust xylose fermentation strain. In addition，the deletion of GRE3，strengthening of xylose transport and rational modification of xylose metabolic pathway during the metabolic modification of strain XI effectively improved the xylose fermentation capacity of the recombinant strains. Besides，evolutionary engineering also increased the xylose fermentation efficiency of XI strains. Furthermore，the omics technologies have presented their power in explaining the mechanism and developing the modification strategies of xylose metabolism. This paper reviews the research progresses on the expressions of xylose isomerase pathway in S. cerevisiae and analyzes the issues in the relevant studies.

Research Progress on Substrate Species Degraded by Laccase

ZHANG Ze-xiong, LIU Hong-yan, XING He, MA Yu

2017, 33(10):  97-102.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0417

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Laccase，a polyphenol oxidase that binds multiple copper ions，is widely found in plants and fungi. Laccase contains unique copper ions which function is to transfer electrons in the structure，so that the laccase has a strong redox capacity，and the only product of the most reactions with lignin，amine compounds，aromatic compounds such as the role of the substrate is water. It has the characteristics of high efficiency and low cost in the degradation of many toxic substances and harmful pollutants，such as white rot fungi produced by the high level of laccase has been widely used in industrial wastewater treatment and other biological remediation and repair areas. In recent years，the lastest research showed that the using of vector immobilized enzyme technology allows laccase to be recycled after use and reused and more stable，which reduces costs while maintains the characteristics of laccase catalytic oxidation，it has overcome the problems of laccases in solving environmental pollution，Under the mediating effect of mediator，the problem of low potential of laccase redox has been figured out，and it greatly increased more kinds of substrate. Therefore，the application prospects in wastewater，degradation，soil remediation，industrial bleaching and other fields will be much broader. This paper summarily describes the applications of laccase in various fields and the types of harmful pollutions substrates，put forward the deficiency and the improving direction of existing degradation process of laccase aiming at providing the reference for the study of biodegradation of environmental pollutants.

Technique

Research Advance on Remediation Technology of Cadmium Contaminated Soil

GAO Yu, CHENG Qian , ZHANG Meng-jun , ZHU Zhen-yu , HU Ting-ting, YANG Yu

2017, 33(10):  103-110.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0336

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Cadmium（Cd）is a strongly toxic heavy metal that can be easily absorbed by plants and accumulated in human body through food chains so that result in health problems. Due to the increasingly wide use of cadmium and its compounds，there come more and more heavy-metal contaminated soil and other environmental problems，therefore the technologies of preventing and remedying cadmium-contaminated soil have attracted much attentions of many researchers in several fields. In this review，we introduce frequently-used remediation technologies in recent years，such as electrochemical-remediation，bioremediation，and so on，and followed a discussion about the mechanisms，applications，advantages and shortcomings of these remediation technologies. Moreover，we prospecte the future development trend of heavy-metal remediation technology，aiming at providing theory supports for the research and development of heavy metal repair.

Strategies for Evaluating the Effects of Transgenic Crops on Soil Microbial Diversity

LIANG Jin-gang, ZHANG Xiu-jie

2017, 33(10):  111-116.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0491

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Microorganisms are the important components of the soil ecosystem. Soil microbial diversity is crucial to the maintenance of ecosystem functioning as soil microorganisms influence many ecosystem processes and drive biogeochemical cycles. Thus，it becomes a hot topic about the potential risks of transgenic crops to soil microorganisms and their effects on soil ecosystem. With the development of modern biotechnologies，the analysis methods for soil microbial diversity have been developed from the traditional separation culture to study microorganisms in the whole soil microecosystem. However，due to the various characteristics of soil microorganisms（such as majority being non-cultivable，tiny bulk，community quorum sensing，etc.）and the detection limitations of instruments and equipment，there are some disadvantages while only using single research method，and other methods should be combined to study microbial diversity in soil ecosystem. So far，the study on soil microbial diversity mainly includes species diversity，functional diversity，structural diversity and genetic diversity，there are a lot of methods used to study the effects of transgenic crops on the diversity of soil microorganisms. In this review，we summarize these methods，and finally the strategies for the future study are proposed.

Application Analysis of Microcystis aeruginosa Identification Based on Ribosomal Proteins

JIANG Wen-jing, ZHANG Jun-yi, DU Yang, SUN Li-wei

2017, 33(10):  117-124.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0268

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This research is to verify the feasibility of matrix-assisted laser desorption-ionization-time-of -flight mass spectrometry（MALDI-TOF MS）in the identification of mixed and actual cyanobacterial samples by employing ribosomal proteins as biomarkers. Mechanical disruption and high speed centrifugation were used as pre-treatments to collect the ribosomal protein fraction of cyanobacterial samples，and then MALDI-TOF MS was applied to identify Microcystis aeruginosa in samples after proteins in situ crystallization with proteomic analysis based on 13 ribosomal proteins as biomarkers. The results showed that under the optimal conditions，model strain M. aeruginosa was precisely identified in the mixed samples and the detection rates of assigned peaks were all greater than 75% with reasonable errors range except the fourth sample. The lowest biomass detection limit and ratio of M. aeruginosa was 2.88×10^{6 cells and 37%，respectively. Furthermore，M. aeruginosa was identified and detected in the cyanobacterial samples from Taihu Lake and the detection rate of assigned peaks was 76.9%. In conclusion，MALDI-TOF MS employing ribosomal proteins as biomarkers could be successfully applied to the identification of mixed and actual cyanobacterial samples and will be expected as routine tool in environmental cyanobacteria monitoring.}

Research report

Screening，Identification of Multifunctional Peanut Root-promoting Rhizobacteria and Its Promoting Effects on Peanuts（Arachis hypogaea L.）

LIU Ye, LIU Xiao-dan, ZHANG Lin-li, WU Yue, WANG Guo-wen, WANG iang, JIANG Ying

2017, 33(10):  125-134.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0233

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This study aims to acquire the peanut root-promoting rhizobacteria（PGPR）with multifunction of fixing nitrogen，dissolving phosphorus and potassium，and synthesizing the auxin，for increasing and improving the yield and quality of peanuts in Henan. Five PGPR strains of HS4，HS7，HS9，HS10，and HS11 were screened from sandy alluvial soil of peanut rhizosphere in North China. Further HS9 was chosen by comparing various functional indexes. The activity of nitrogenase，dissolving phosphorus and potassium and IAA synthesis of HS9 were 15.53 nmol C₂H₄/h·mL，279.23 mg/L，and 22.5 mg/L and 40.96 mg/L， respectively. HS9 was identified as Bacillus flexus based on the morphological observation，16S rDNA gene sequence analysis accompanied with physiological and biochemical test. Pot experiment was then conducted to test and verify the effects of growth promoting，and the results showed that inoculation with HS9 significantly increased the content of available phosphorus and potassium in soil，and distinctly enhanced the content of IAA in soil. This led to the root length，surface area，root volumes and root tips of peanut increased 109.60%，84.30%，76.08% and 386.24%，respectively. It indicated that the strain promoted plant nutrient uptake and utilization，and increased plant biomass and nutrient content. The average wet weight of plants was 1.7 times higher than the control treatment and the height of peanut inoculated with HS9 significantly increased by 28.35%；the SPAD（Soil and plant analyzer Development）value significantly increased by 16.06% and the total N，P and K content of peanut significantly increased by 23.11%、83.04% and 23.95%，respectively. Conclusively，the multifunctional strain HS9 has solid growth-promoting effect on peanut and broad agricultural application prospect.

Screen of Poly-γ-gamma Glutamic Acid Producing Bacteria and Their Glutamic Acid Fermentation Broth’s Function on the Drought Resistance of Maize

JIA Yan-ping, YIN Ai-ming, SUN Yan-mei, CHENG Shou-tao, WANG Xu-ming

2017, 33(10):  135-142.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0699

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As a novel biological water absorbent，poly-γ-gamma glutamic acid（γ-PGA）has a significant application value on water-saving agricultural production. Two γ-PGA-producing strains，Y2 and P1，were isolated and screened from natto and fermented soya bean samples. By analyzing their morphological and physic-chemical characters，and 16S rRNA sequence，they were identified as Bacillus subtilis. Then，the effects of γ-PGA fermentation broth on the soil water-holding capacity and the drought resistance of maize seedling under water stress were also studied. The results showed that the fermentation broth of Y2 and P1 significantly decreased the soil water evaporation. Compared with the control treatment，adding fermentation broth from γ-PGA-producing strain promoted the soil water content by 4%. At the same time，the addition of γ-PGA fermentation broth enhanced the drought resistance of maize seedlings，and significantly increased aboveground and underground biomass of maize seedlings by 40.70% and 19.59%，respectively. The result of biologECO showed that the γ-PGA fermentation broth from Y2 and P1 had insignificant effect on microbial diversity and evenness. Thus，the γ-PGA fermentation broth is an environmental friendly biological water absorbent，which can be used directly to enhance soil water retention and drought resistance of vegetation in field.

Isolation，Identification and Degrading Properties of Phlorizin-Degrading Fungi

LIU Shu-yan, WANG Fang, WANG Jian-yu, LIN Rong-shan

2017, 33(10):  143-147.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0536

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Screening highly efficient phlorizin-degrading microorganism is important for bioremediation of phlorizin-contaminated environment. This study aimed to isolate and identify high efficient phlorizin-degrading microorganism from the rhizosphere soils of apple，then to characterize degrading properties of target strains. Phlorizin-degrading microorganism were screened by the plate-culture method，used phloridzin as the sole carbon sources，and then the target strains were identified by the morphological and molecular methods. Degrading properties of phlorizin -degrading microorganism were determined under different condition of culture. The results showed that a phlorizin-degrading fungal strain AMCC300100 identified as Aspergillus terreus were obtained from the rhizosphere soils of apple replantd and non-replanted orchards surrounding Bohai gulf area. The phlorizin degradation rate of the strain AMCC300100 has reached 88.96%（phlorizin initial concentration of 2 mmol/L，pH5.0，2% inoculum）cultured 96h under the conditions of 40℃，160r/min；The strain AMCC300110 have high-efficient degradation ability not only for phlorizim but also for the other phenolic acids. Though plate-culture method，We obtained a fungal strain identified as Aspergillus terreus，this fungal strain have high-efficient degradation ability for phlorizim，and have tremendous potential in solving apple replant disease.

Screening of Multi-functional Nitrogen Fixing Bacteria and Their Application in Soil Ecological Restoration

MAO Xiao-jie, WANG Xin-min, ZHAO Ying, ZHOU Yi-qing, SUN Jian-guang

2017, 33(10):  148-155.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0678

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The objective of this study is to screen multi-functional nitrogen-fixing bacteria based on nitrogenase and ACC（1-aminocyclopropane-1-carboxylate）deaminase activity，to develop inoculant and test its effect in wasteland soil ecological restoration. First，nitrogenase activity was determined by acetylene reduction assay，and ACC deaminase activity was quantitatively measured by colorimetry. Then，the 16S rRNA of the strains was amplified by PCR，and the phylogenetic status of the strains was investigated by sequence analysis and alignment. The identification of strains was carried out based on the morphology，physiology，biochemical test results，and 16S rRNA analysis. As results，total 47 strains with nitrogenase activity higher than 9 nmol C₂H₄/（h · mg）protein were screened，and the nitrogenase activities of the 43 strains were higher than that of positive control Azotobacter ACCC11103. And 20 strains producing ACC deaminase of 0.326 μmol/（h·mg）to 21.980 μmol/（h·mg）were obtained. Plant growth promotion test showed that the 47 strains increased the fresh weights of Chinese cabbage（Brassica campestris）. Based on 16S rRNA sequence，the 51 selected strains were identified as Agrobacterium albertimagni，Arthrobacter pascens，Bacillus circulans，Burkholderia vietnamiensis，Chitinophaga ginsengisegetis，Dyadobacter fermentans，Ensifer adhaerens，Enterobacter aerogenes，Lysobacter yangpyeongensis，Micrococcus luteus，Paenibacillus alginolyticus，Phyllobacterium ifriqiyense，Pseudomonas fluorescens，Rhizobium alamii，Sinorhizobium adhaerens，Sporolactobacillus laevolacticus of 31 species of 16 genera. After test，4 excellent strains of 7012，7134，7144，and 7164 were manufactured as inoculant termed as NFMF（Nitrogen Fixing and Multi Function）biofertilizer，with which the field experiment showed that soil microorganism quantity increased significantly after 3 months of NFMF biofertilizer inoculation，and soil activities of sucrase，catalase，urease，phosphatase，cellulase and xylanase significantly increased compared with the no inoculation control. In conclusion，NFMF can significantly increase wasteland soil bioactivity and it is valuable in the ecological restoration of wasteland soil.

Isolation and Identification of a Zinc-resistant Strain and Effect of Its Characteristics on the Remediation Efficiency of Brassica juncea in Zinc-polluted Soil

CHEN Yang, SONG Tian-shun, ZHOU Guo-ling, XIE Jing-jing

2017, 33(10):  156-162.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0587

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A zinc-resistant strain was isolated from heavy metal-polluted soil，and identified as Bacillus. Its performance of removing Zn²⁺ under different experimental conditions was studied. The results showed that：the removal efficiency of Zn²⁺ was the best，and the removal rate of Zn²⁺ was 94% in 12 h when the initial concentration of Zn²⁺ was 300 mg/L，the temperature was 37℃，inoculation amount was 5%. Subsequently，a pot experiment was conducted for investigating the influence of adding the different concentration of the strain on plant growth and Zn uptake by Brassica juncea. The results showed that the Zn content in ground and underground part of B. juncea increased by 56.1% and 24.3% respectively while adding 40 mL bacterial liquid（1×10⁹ CFU/mL），compared with the control group. This is mainly due to the strain improves the zinc content under weak acid extraction condition.

Isolation and Identification of an Imazethapyr-degrading Bacterium，and Optimization of Biodegradation Conditions

Lü Fan-yang, XU Ze-hua, MAO Xiao-jie, SUN Jian-guang

2017, 33(10):  163-168.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0076

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This work aims to isolate and identify the strain IM9603 degrading herbicide imazethapyr，and optimize the degradation condition. IM9603 was identified as Brevibacterium sp. based on morphological，physio-biochemical characteristics，and analysis of 16S rDNA. Response surface methodology Box-Behnken was applied to optimize the degradation conditions. As results，strain IM9603 degraded 93.08% of the initial 50 mg/L supply of imazethapyr under the optimal condition（pH5 and 26.2℃）within 7 days，which was its best degradability. The isolated imazethapyr-degrading strain was Brevibacterium epidermidis，and presented high degradability to imazethapyr，suggesting that is owns a great potential in the environmental rehabilitation caused by imazethapyr.

Isolation and Identification of a Nickel-resistant and Petroleum Hydrocarbon Degrading Strain and Its Biological Characteristics

MA Yong-song, LI Xi, LI Zhen-zhen, WANG Pei-jie

2017, 33(10):  169-177.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0687

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: A bacterial strain with high ability of nickel resistance and petroleum-degrading was isolated from the soil of heavily-polluted areas around the shale gas wells in Changning-Weiyuan，Sichuan Province，and its biological characteristics and physiological-biochemical indexes were studied. The seed liquid was inoculated into the liquid medium containing heavy metal and crude oil，cultured at 30℃ and 130 r/min for 7 days to explore the resistance of the strain to nickel ions（Ni²⁺）and the rate of degrading total petroleum hydrocarbon（TPH）. Further，the strain was identified based on the experimental results of physiology and biochemistry，morphological observation and 16S rDNA sequences. As results，the tolerance of the strain to Ni²⁺ reached 300 mg/L，and the removal rate and adsorption rate of Ni²⁺ reached 56.64% and 52.16%，respectively; meanwhile，the degradation rate of TPH reached 35.65%. The species of the strain belonged to Citrobacter（Citrobacter sp.）and named Citrobacter farmeri strain M1. The optimal growth temperature of this strain was 30℃，the optimal growth pH was from 7 to 9，the optimal C∶N was 10∶1，and the maximum salt tolerance was up to 5%. In conclusion，solid nickel resistance and petroleum degradability of the strain contributes to its applicability in the remediation of heavy metal and oil-contaminated soil.

Isolation and Identification of Indigenous Heterotrophic Denitrifying Bacteria from Polluted River

LIU Pan-long, YU Lu-ji, LI Ting-mei, FAN Zheng, CHEN Tao

2017, 33(10):  178-183.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0252

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Heterotrophic denitrifying bacteria from the polluted water of Jialu River were screened on the selective medium，with sodium succinate as sole carbon source and ammonium sulfate as sole nitrogen source. Then，the separation and purification were performed by enrichment，gradient dilution and streaking on the plates. Finally，the strain was identified according to 16S rDNA sequence alignment results，physiological and biochemical experiments and the ammonia nitrogen transformation. The results showed that，63 strains were isolated from the polluted water，and 3 of them were identified as heterotrophic denitrifying bacterium，including a strain of Pseudomonas nirtoreducens and two strains of Pseudomonas mendocina，by which the ammonia nitrogen removal rate were 91.8%，89.8%，and 81.4%，respectively.

Study On the Degradation and Mechanism of Phenolic Compounds by the Thauera sp. K11

QIAO Neng-hu, ZHANG Jing-jing, LIU De-jian, TANG Xiao-sheng, XI Li-jun, LIU Jian-guo

2017, 33(10):  184-190.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0452

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The objective of this work is to analyze the degradation and pathways of a Thauera sp. K11 in genus Thauera to phenolic compounds in phenolic wastewater. Thauera sp. K11 was isolated from petrochemical wastewater treatment plant .The 16S rRNA gene and key enzyme gene were cloned and analyzed and phylogenetic analysis was carried out. The mechanism of phenol degradation was explored at the gene level. The degradation efficiency and mechanism of phenolic compounds were studied by gas chromatography. The 16S rRNA phylogenetic analysis revealed that the bacterium Thauera sp. K11 was a species of bacteria from the genus Thauera，which degraded 11 phenolic compounds. The degradation rates of the five phenolic compounds were > 90 % in 72 h. The K11 phenol hydroxylase and catechol dioxygenase genes were cloned，and the enzyme activity assay showed that K11 catalyzed the conversion of phenol to catechol by phenol hydroxylase and then catalyzed the production of 2-HMSA with catechol-2，3-dioxygenase. In conclusion，the bacterium Thauera sp. K11 is a species of Thauera bacterium capable of degrading a variety of phenolic compounds，and possesses the strong capability of degrading phenolic pollutants；the degradation pathway is by phenol→catechol→2-HMSA.

Adsorption of Safranine T by Aspergilus niger Biomass Waste

WEI Jiang-mian, LU Lei-zhen, JIAO Guo-bao, LIU Jia-yang, LU Jun-he

2017, 33(10):  191-198.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0196

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The present work aims to investigate the adsorption of dye safranine T by Aspergillus niger biomass waste from amylase production as novel composite adsorbent. The biomass waste was first characterized concerning its physico-chemical properties and then evaluated for its potential of adsorbing safranine T under different conditions，namely contact time，temperature，sorbent dosage，initial dye concentration and metal ion. The isotherms，dynamics，and thermodynamics were fitted with the above data，which showed that pseudo second order dynamics equation better described the adsorption with equilibrium achieved within 2 h，suggesting it was chemosorption，and film diffusion model was more suitable to explain the adsorption than internal diffusion model. Higher temperatures significantly promoted dye removal，and adsorption was both spontaneous（ΔG° < 0）and endothermic（ΔH° > 0）. The maximal monolayer adsorption capacity was calculated to be around 166.67 mg/g through Langmuir equation while Freundlich was more suitable to explain the adsorption，indicating that there were multiple absorptive sites on the surface of biomass waste. In addition，sorbent dosage and sodium ion concentration both impacted adsorption with the highest removal rate of 91%.

The Diversity and Distribution Pattern of Bacterial Community in the Water of Yellow River Estuary

WEI Guang-shan, ZHANG Jia-wei, LI Ming-cong, GAO Zheng

2017, 33(10):  199-208.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0568

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The Yellow River estuary，located in the confluence of Yellow River，Bohai Bay and Laizhou Bay，has unique geography and abundant microbial resources. However，the researches on bacterial community in this area were very limited. Here we used 16S rDNA clone libraries to explore the diversity and the characteristics of spatial distribution pattern of bacterial community in the unique estuarine ecosystem. The results demonstrated that different 16S rDNA clone libraries were constructed for 4 different water samples，and totally，we detected the bacteria in Yellow River estuary in 11 phyla，18 classes，39 families and 53 genera. The dominant bacteria were attributed in Proteobacteria （α-，β-，and γ-proteobacteria），Actinobacteria，Bacteroidetes and Cyanobacteria. Functional annotation results showed that aquatic bacteria played key roles in the cycling of carbon，nitrogen and sulfur elements. Species- or function-based cluster analyses indicated that samples could be divided into two different branches，A，B and C，D，respectively. Spearman correlation analysis and redundancy analysis （RDA） demonstrated that environmental factors （dissolved oxygen，salinity，and nitrogen nutrients） had significant effects on the structures and functions of aquatic bacterial community. The study shows that both the Yellow River and environmental factors drive the structure of bacterial community into varied spatial distribution patterns along the estuary. This study is a preliminary glimpse of bacterial diversity in the Yellow River estuary and adjacent seawater. It also provides data supports for further improvement in the rivers and marine environment of this area，and is beneficial to the microbial resource development and ecological protection of this water area.

Enzyme Activities and Microbial Abundance of Rhizosphere Surrounding Different Tree Species in Daqing Mountain

ZHANG Xing, LI Min, YAO Qing-zhi, TIE Ying, WEI Jie, CHEN Li-xia

2017, 33(10):  209-216.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0296

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This study aims to find the correlation between soil microbial abundance and enzyme activities of rhizosphere around different tree species in forest of the Daqing Mountain and its key factors. Real-time qPCR was used to study the variations of soil microbial abundance（bacteria and fungi）around the rhizosphere of different species，and analyzed the changing trend of three soil enzyme activities（β-glucosidase（βG），N-acetyl glucosidase（NAG）and peroxidase（Pod）and soil physicochemical properties. Principal component analysis（PCA）and Pearson correlation analysis were applied to investigate the impacts of soil physicochemical factors on the soil microbial abundance. As results，soil physicochemical properties and microbial bacteria and fungi community abundances had significant increases from spring to autumn，such as organic carbon（TOC），total nitrogen（TN），microbial biomass carbon（WC）and nitrogen（WD）as well as bacteria and fungi abundances，etc. The forest soil βG enzymes and Pod activity in Daqing Mountain was higher in summer，while NAG enzyme activity was higher in the fall. Pearson correlation analysis showed that soil microbial abundance and soil enzyme activities of different tree species in forest in the Daqing Mountain had obvious correlation，significantly positive correlation with βG enzymes activity（P < 0.05），very significantly negative correlation with Pod activity（P < 0.05），and very significantly negative correlation with βG enzyme activity（P < 0.05）. Soil physicochemical factors（TOC，TN and WC and WD）and soil microbial abundance had significantly positive correlation. It was inferred from principal component analysis，that TOC，TN，WC and WD，bacterial and fungal community abundance，and NAG can be used as the important influencing factors of the rhizosphere soil nutrient characteristics surrounding different tree species. The different tree species in Daqing Mountain forest ecosystem had profound influences on soil physicochemical index，soil microbial abundance and soil enzyme activities，and presented the strong responses with the season changing. Physicochemical index and soil microbial community abundance are the main ecological factors regulating the microbial community of rhizosphere soil in the forest and vegetation of Daqing Mountain.

Effects of Organic Matters on the Abundance of Microorganisms in the Urban Wetland

XIE Jia, MA Xiao-hang, DAI Yan-ran, WU Juan, XIANG Dong-fang, CHENG Shui-ping

2017, 33(10):  217-224.  doi:10.13560/j.cnki.biotech.bull.1985.2017-0528

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Urban wetland suffers from not only the constant increase of organic matter load，but also the issue of organic matter in the topsoil is damaged in the process of urban development and construction. By adding the dissolved organic matter（DOM）solution of Cinnamomum camphora leaves，together with real-time fluorescence quantitative PCR，the effects of exogenous organic matter on the abundance of microorganisms in the topsoil（0-5 cm）was studied in simulative way. The results showed that changes in soil structure significantly affected the abundances of bacteria，archaea and functional genes associated with nitrogen cycles. The abundances of bacteria and functional genes associated with nitrogen cycles decreased significantly while soil organic matter content decreased（P <0.05）. The addition of nitrogen and DOM in the overlying water remarkably increased the abundances of microorganism and nitrogen functional genes in wetland. The exogenous DOM significantly affected the two genes in the transformation process of NO₂--N to N₂O and NO.