Abstract:

Abstract:[Objective] To provide method for rapid molecular detection of Avr1a, Avr1k and Avr3a, avirulent genes of Phytophthora sojae, and also to provide some references for rapid molecular detection of other avirulent genes of P. sojae. [Methods] According to Genbank sequence table of Avr1a, Avr1k and Avr3a, avirulent genes of P. sojae, specific primers were filtered out respectively for the three genes by using primer design method. 86 strains of P. sojae which have been identified virulence on the three differential soybean varieties were detected by PCR on the basis of optimized reaction system and amplification conditions. A specific detection system was established for detecting Avr1a, Avr1k and Avr3a, avirulent genes of P. sojae. The results of molecular identification and inoculation identification were compared and analized. And then amplified true and false positive bands were recovered respectively and cloning sequenced and compared with the original sequences of the three avirulent genes, to verify if Avr1a, Avr1k and Avr3a were suitable to be detected by the method of molecular markers. [Results] All the specific primers screened out could amplify a band with length of about 550 bp. The coincidence rate of molecular and inoculation identification for the three avirulent genes were Avr1a-45.3%, Avr1k-84.9% and Avr3a-97.7%. The true positive bands of three avirulent genes had over 97 percent of consistency in sequence with the original one. The false positive bands of Avr1a were about 80 percent, and Avr1k and Avr3a were less than 30 percent of consistency with their original sequences. [Conclusion] The detection system established by using the primers filtered out by utilizing the gene sequence of Avr1a, Avr1k and Avr3a can be used to detect Avr3a rapidly, but not be suitable for detection of Avr1a, whether fit for detection of Avr1k needed further study.

Abstract:

Abstract:[Objective] To realize the relationship between respiratory maintenance and anti-oxidative function of Pos5p, the mitochondrial NAD(H) kinase in Saccharomyces cerevisiae. [Methods] The respiratory chain activity and reactive oxygen species (ROS) level of wild-type BY4742, POS5 gene deletion mutant pos5Δ, and pos5Δ containing POS5-YEplac195 plasmid (pos5Δ/POS5-YEp) under exposure to different kinds of oxidative reagents were compared. [Results] At the normal growth condition, pos5Δ exhibited poorer respiratory chain activity than that of BY4742, while pos5Δ/POS5-YEp gave the partial restored activity, consistent with the ROS level of these three strains. Under the oxidative stress of menadione, the respiratory chain activities of BY4742 and pos5Δ decreased, while that of pos5Δ/POS5-YEp increased. Under the oxidative stress of H2O2 and diethyl maleate, nearly all of the activities of BY4742, pos5Δ and pos5Δ/POS5-YEp reduced, with that of pos5Δ most seriously, and that of pos5Δ/POS5-YEp under H2O2 lessened. Thus indicated that the oxidative stress would injury the respiratory chain complex of S. cerevisiae and over-expression of Pos5p could alleviate the injury from menadione and H2O2. [Conclusion] The protecting function of Pos5p on respiratory chain was correlated to its antioxidative defense function.

Abstract:[Objective] Penicillium decumbens is an important filamentous fungus that efficiently secrets cellulases. The biosynthesis and secretion of these cellulases are regulated on transcription levels. In order to further study the mechanism of transcriptional regulation of cellulases encoding genes and construct industrial strains with high cellulases production, [Methods] We identified a transcription regulator BglR (PDE-01706) from P. decumbens 114-2 by comparing gene expression profiles under different carbon sources condition. This transcription regulator shares a 59% sequence identity with the zinc finger protein Pc20g04780 from Penicillium chrysogenum. P. decumbens ΔbglR-1 knockout strain was constructed with homologous double crossover recombination. The phenotype of this strain was investigated, including its hyphal growth, protein secretion level, cellulase secretion level and extracellular pH level. [Results] It was shown that the deletion of the transcription factor BglR in P. decumbens ΔbglR-1 leads to an increase of extracellular β-glucosidase activity by 40%, and a lowered filter paper activity, endoglucanases and xylanase level. [Conclusion] From these results it is proposed that BglR plays a major role in the regulation of cellulase production of P. decumbens.

Abstract:[Objective] The populations of endophytic fungi from halophyte Suaeda heteroptera Kitag. grown in the salt flats in Shuangtaizi Estuary of Panjin, Liaoning province, China, were investigated. [Methods] Endophytic fungi were isolated and purified from the roots, stems and leaves of Suaeda heteroptera Kitag. with traditional method. Isolates were identified based on morphological, physiological and biochemical characteristics and rDNA-ITS sequence analysis. [Results] Overall, 49 endophytic fungi strains were isolated and purified, these strains were classified into 13 different genera, in which Glomerella, Alternaria, Colletotrichum and Cladosporium were dominant genera. The distribution of endophytes of host plants have some degree of tissue preference, and though there was not significant difference of the abundance level in different habitats, habitat specificity of some endophytic fungi communities was presented. [Conclusion] Our results suggested that Suaeda heteroptera Kitag. have abundant resouces of endophytic fungi, the species diversity of endophytic fungi communities is affected both by its host preference and by its different habitats.

Abstract:[Objective] A phenol-degradative strain named T10 which was isolated from the soil sample of a chemical plant was identified, and increased degradative rate of phenol by optimizing the growth conditions. [Methods] The strain was identified by morphological observation, physiological and biochemical properties and 16S rDNA sequence analysis. The phenol-degradative capacity of strain T10 was analyzed when they were cultured in a series of liquid media containing different pH, initial phenol concentration and liquid volume. [Results] The strain T10 belongs to Pseudomonas putida. Glucose and peptone can reduce growth period of strain T10, and the degradation rate of phenol was improved 1.7 times. Under the condition of inoculating concentration 10%, 30 °C and 180 r/min, the phenol removal rate of strain T10 could reach 87.56% when initial phenol concentration, pH and liquid volume were 3?000?mg/L, 7.5, 80 mL/250 mL, respectively. [Conclusion] The experimental data show that strain T10 can tolerate wastewater containing higher concentration phenol, and has stronger degradation ability on phenol. These results would provide reliable basis for further treatment of wastewater containing phenol by the means of biological method.

Abstract:[Objective] Phase I cells of Photorhabdus luminescens bacteria, symbiotically associated with entomopathogenic Heterorhabditis nematode, produce two types of intracellular crystalline inclusions, CipA and CipB, to support the symbiont. This study aimed to investigate the possible influence of Cip proteins on non-symbiotic Steinernema nematode. [Methods] Based on constructed Escherichia coli expression system of Cip proteins, co-culture system of recombinant E. coli bacteria and Steinernema sp. SY-5 nematode was set up. [Results] The Cip proteins significantly promoted the development of SY-5 nematode to make high adult rate of 65%?82%, gravid rate of 80%?95%, 30?50 eggs per nematode, and low mortalities. [Conclusion] As the nutrient reserves for Heterorhabditis nematode, Cip proteins can be accepted and utilized by Steinernema nematode. The progress on these proteins will provide insights into the mechanisms governing bacteria-nematode symbiosis.

Abstract:[Objective] Our goal is to analyze the bacterial endophytic diversity in Murraya paniculata, which was huanglongbing’s hidden host plant, and find the endophytic bacteria that may have potential to suppress the HLB pathogen. [Methods] The endophytic bacteria were isolated from surface-sterilized M. paniculata midribs of leaves and phloem of stems by plating and restriction fragment length polymorphism (RFLP). The functions of 14 culturable bacteria had been tested using different substrates and cultural medium. [Results] By the artificial anaerobic culture, we obtained 26 strains that were grouped into 9 genus according to GenBank. Enterobacter sp. (IF=19.23%) and Bacillus sp. (IF=38.46%) were the dominant bacterial population in M. paniculata. However, the bacterial colony number of endophytic bacteria in the stems was higher than in the leaves. The endophytic bacteria 16S rDNA library of M. paniculata was established. Coverage C of the clone library was 94.97% and the rarefaction curve of the clone library tended to plateau, which indicate that the library are large enough to re?ect the bacterial endophytic diversity of the respective samples. 179 positive clones in 16S rRNA gene library were chosen and digested by restriction endonuclease Hae Ⅲ, MspⅠ, RsaⅠrespectively. Twenty OTUs (Operational taxonomic units) were observed based on the similarity of the RFLP banding profiles. In the observed clones, 63.69% of the clones were Serratia sp., which means that the Serratia sp. was absolute preponderant endophytic bacteria in M. paniculata. The functional analysis of the 14 endophytic bacteria showed that 9 isolates can produce IAA and 3 of them had amylolysis activity. The frequency of endophytic bacteria which can produce ACC deaminase, siderophore and protein activity was 1, 8 and 2 respectively. Three of bacteria isolates with N-fixation ability were screened by using N-free medium and nifH gene and 4 of endophytic bacteria cansynthesis antibiotic (phlD). A total of 4 bacterial isolates was provided with four of mentioned traits. [Conclusion] The endophytic bacteria isolated from M. paniculata have abundant diversity and genetic characterization, and they may have important functions for the plant development, anti abio-stresses and bio-stresses.

Abstract:[Objective] Porcine Kidney-15 cells (PK-15) were infected by Porcine Parvovirus (PPV) to survey the host inflammatory responses, particularly multiplicity of infection (MOI) for the cytokines responses of PPV infection, and the host-PPV interaction. [Methods] Then the viral DNA was measured and analyzed using real-time PCR. The transcript level of cytokines (IFN-β, IRF-3, TNF-α and IL-18) were detected by real-time PCR too. [Results] We found that PPV proliferated rapidly in PK-15 cell 12 hours after infection, and reached peak after 48 hours infection and MOI=1.0; the transcription levels of IFN-β, IRF-3, TNF-α and IL-18 increased obviously, and the transcription level of IRF-3 gene increased to 967 times in 1 hour and MOI=10.0 after infection. [Conclusion] The results revealed that the level of cytokines significantly dependent on the interaction of MOI and infection time.

Abstract:[Objective] To establish a high-throughput drug screening model for screening inhibitors of Mycobacterium tuberculosis Phenylalanyl-tRNA synthetase (PheRS). [Methods] In this work, Mycobacterium tuberculosis pheRS gene was cloned and overexpressed in E. coli, and then the purified recombinant enzyme was used to establish an high-throughput drug screening model. After establishment and optimization of the screening assay, a primary screening was performed with compounds library and microbial fermentation collection in our institute, and their anti-bacterial activity and cytotoxicity were assessed further. [Results] 9 out of 11 600 compouds and 37 out of 5 200 microbial fermentation samples were identified as primary hits. [Conclusion] 6 microbial fermentation samples inhibited enzymatic activity of PheRS in vitro and growth of Mycobacterium smegmatis, and showed lower cytotoxicity.

Abstract:[Objective] The study aimed to clone a-amino acid ester hydrolase gene from Xanthomonas rubrillineans, to perform bioinformatics analysis and increase the thermostability of the enzyme. [Methods] The full length of aeh was cloned by polymerase chain reaction (PCR). The gene sequence obtained and the putative amino acid sequence were analyzed by bioinformatics software, and the three-dimensional structure of X. rubrillineans AEH was predicted by homology modeling. In order to improve thermostability, sites displaying high degree of flexibility were replaced through site-directed mutagenesis. [Results] Aeh was obtained by PCR from X. rubrillineans (GenBank accession: JF744990). The nucleotide sequence is 1 917 bp length, encoding a polypeptide of 638 amino acids and shares 91% and 83% identity to peptidase from X. albilineans str. GPE PC73 and GL-7-ACA acylase from X. axonopodis pv. citri str. 306 respectively. Based on the predicted model, sites displaying high degree of flexibility were replaced through saturated mutagenesis and 3 variants with 5 °C higher T50 than wild type were distinguished from 282 variants by screening. [Conclusion] The sequence analysis of X. rubrillineans AEH was benefit for exploration of evolution history. The strategy of replacing highly flexible residues by saturated mutagenesis can be used for enhancing thermostability.

Abstract:[Objective] Identification of the pathogen of moldy-core disease of apple in Gansu Province. [Methods] The pathogenicity of fungal isolate was confirmed by Koch’s rule, and identification of the pathogen was performed according to anamorphic morphological characteristics and ITS rDNA sequence analysis result. [Results] A fungus isolated from moldy-core disease of apple in Tianshui County, Gansu Province, in January 2009. The fungus caused the same symptom compared to the natural mouldy-core disease while injected inoculation through the calyx tube of apple fruits. The fungus produced conidia with different morphological types: Conidia formed on acervuli (black conidiomata developed on mycelial mat in potato sucrose liquid medium after cultured 7 months at 3 °C?7 °C), (12.95 ?20.42) μm× (4.98?7.97) μm [av. (16.75±1.89) μm×(6.47±0.86) μm], 3 septa, rarely 4?5 septa, fusiform or ellipsoid, pale brown to brown, median two or upper three cells darker than the basal one, appendages laking; Conidia formed on mycelium (cultured 18 d on PSA plate at 10 °C), (12.45?59.76) μm×(4.98?11.21) μm [av. (30.10±11.16) μm×(7.26±1.28) μm], 2?9 septa, fusiform, clavate or worm-formed, initial slight colour, gradually altered to brown. [Conclusion] The fungus was identified as Discostroma fuscellum (Berk. & Broome) Huhndorf, based on anamorphic morphological characteristics and molecular identification (GenBank accession: JF320818). This is the first record of Discostroma fuscellum as the pathogen of apple moldy-core disease in China.

Abstract:[Objective] Screening the antagonistic endophytic bacteria against Thielaviopsis basicola from inner of tobacco roots and stems collected from Chongqing. [Methods] The antagonistic effect of 306 endophytic bacteria against Thielaviopsis basicola by pairing culture on PDA plates, the strain was identified by morphological, physiological and biochemical characteristics, and homology analysis of the partial sequence of 16S rDNA. [Results] Six endophytic bacteria were showed high antagonistic effect against Thielaviopsis basicola, the pathogen of tobacco black root rot. The inhibition zone diameters were ranged from 6.5 mm to 11.0 mm by pairing culture on PDA plates, and the control efficacy ranged from 11.9% to 77.1% in a greenhouse experiment. It was found that strain T295 could reduce the incidence of tobacco black root rot, the control efficacy reaching to 77.1%, and the culture filtrate of strain T295 could inhibit hyphal growth and reduce spore germination of T. basicola. [Conclusion] Strain T295 had a good antagonistic effect on Thielaviopsis basicola, was identified as Bacillus subtilis.

Abstract:Microbial degradation of environmental pollutants is a hot research field in environmental protection. Comamonas are able to degradate most environmental pollutants excellently. This paper reviews the degradation characteristics and metabolic pathway of some isolated Comamonas for different pollutants, and analyzes the influence of various conditions to the Comamonas. At last, the present situation and potential applications of Comamonas for degradation of environmental pollutants are introduced.

Abstract:Tobacco bacterial wilt caused by Ralstonia solanacearum is one of the most important diseases all over the world, it is a typical soil borne bacteriosis. In this paper, the current situation of tobacco bacterial wilt was reviewed, including the pathogenic mechanism, genetic diversity and prevention measure of Ralstonia solanacearum.

Abstract:This paper presents a review of the lastest research on Termitomyces, including taxonomy, active constituents, life cycle, culture and symbiotic relationship with host termites. Based on the summary of existing research findings, some problems in related studies are pointed out. In addition, the application prospects and research emphases are discussed.

Abstract:Metabolomics is an important branch of systematic biology. It has received great attention around the world, especially in microbiological research field, and new progress has been also made. The metabolomics of lactic acid bacteria has become one of the hottest research topics. This review summarizes the major techniques involved in the research procedures of the metabolomics of lactic acid bacteria, including sample preparation, analysis and identification of metabolites and data analysis. The applications of the metabolomics of lactic acid bacteria are also given based on several typical examples. At the same time, we discussed the potential problems of the metabolomics of lactic acid bacteria, and points out the future of this research.

Abstract:To enhance the science knowledge base of the students majored in bio-engineering and maximize the effect of microbiology course in professional basic courses to make the students grasp the knowledge firmly and cultivate their many capacities at the same time, we developed a method called “Five-step Teaching” which follows “guiding-study—self-study— teacher-tutor—reciprocal study—whole-class-lecture” to reform the education mode. The results indicated: (1) The students not only grasped the knowledge firmly but also changed passive learning into active learning through “Five-step Teaching” practice. (2) As a result of self-taught, preparation, presentations, evaluation and so on, students’ self-confidence, awareness of competence as well as the abilities of logical thinking, understanding, generalizing and language expression and so on were also developed. (3) Score evaluation system would be more reasonable by virtue of “Five-step Teaching” practice.

Abstract:Based on the basic teaching experiment, coliform bacteria test in source water as example, this paper made some researches on the educational reform of microbiology experiment. There were four teaching practices to be carried out. The first was to guide students to form microbiology experiment ability by designing experiment plan independently. The second was to guide students to form standard operational habits by experiment demonstration, analysis and strict supervision. The third was to cultivate students’ profound observing and thinking abilities by observing experimental phenomena. The fourth was to cultivate students’ realistic attitudes by teachers’ precepts and their personal example. These practices had proved that, the educational reforms, carried out in the teaching experiment like the experiment of coliform bacteria test in source water, could help students to cultivate their abilities of self-learning, practice, observation, analysis and creation, and then promote their comprehensive quality.

Abstract:Focusing on the training on the graduate students’ academic and innovating ability, a reforming practice on teaching model in the course Metabolic Engineering was discussed. Firstly, the course contents renewing was conducted via a PPT renewing on line and real time in the class. Secondly, a class teaching model, lectures on core contents by teachers plus a series of seminars simulating international conference by students, was constructed. Finally, a comprehensive test method, exam on theory plus evaluation of student’s comprehensive ability, was implemented. All the above practices are helpful to enhance academic and innovating ability of the graduate students.

Abstract:[Objective] Enhance the efficiency of fungal pretreatment and lower the severity of liquid hot water (LHW) pretreatment. [Methods] Combinations of white rot fungi and LHW were employed to pretreat Populus tomentosa, the influences of this combined pretreatments on the chemical components and enzymatic hydrolysis of P. tomentosa were studied. [Results] The highest hemicellulose removal of 70.70% was observed by combination of Lenzites betulinus C5617 with LHW pretreatment. Celluloses were degraded in both step of the combined pretreatments, combination of L. betulinus C5617 with LHW caused the highest cellulose loss of 29.62%. Lignin loss happened mostly in the step of fungal pretreatment, and L. betulinus C5617 caused a higher degradation (16.98%) of acid insoluble lignin among the two fungi. The combined pretreatments improved the enzymatic hydrolysis of P. tomentosa significantly. Compared with sole LHW treatment, combination of L. betulinus C5617 with LHW pretreatment resulted in an increase by 20.60%, and an increase (12.23%) of reducing sugar yield were obtained by combination of P. sanguineus D9497 with LHW treatment. [Conclusion] This combined pretreatment can lower the severity of LHW pretreatment and was cost-effective.