Enzymatic characterization of an extracellular glucoamylase from Tricholoma matsutake and its cloning and secretory expression in Pichia pastoris

ABSTRACT

A glucoamylase from the ectomycorrhizal fungus Tricholoma matsutake (TmGLA) was purified 33.2-fold to homogeneity as a single monomeric glycoprotein with a molecular mass of 63.9 kDa. Maximum activity was observed at 60°C and pH 5.0. The enzyme is active down to 50°C and in the pH range of 4.0–6.0, and its activity is strongly inhibited by Ag⁺. It degrades α-1,4- and α-1,6-glycosidic linkages in various polysaccharides. Its gene (TmGlu1) was cloned using information from the enzyme’s internal amino acid sequences and the whole genome sequence of T. matsutake NBRC 30605. The deduced amino acid sequence showed clear homology with those of GH family 15 proteins. Pichia pastoris transformed with TmGlu1 secreted the active enzyme in a glycosylated form, and its characteristics were the same as the native enzyme.     

大链壶菌对白纹伊蚊及几种其它贵州蚊虫的杀灭作用的观察(英文)

采用大链壶菌在实验室内对白纹伊蚊及其它几种贵州蚊虫的幼虫进行杀灭试验。白纹伊蚊的感染率为25—89％,而致倦库蚊的感染率可达90—100％。实验结果为大链壶菌的宿主记录新增加一个属和五个种。     

Identification of the DNAs of the ectomycorrhizal fungusTricholoma bakamatsutake using specific oligonucleotide probes and PCR primers

Partial nucleotides of the 18S rDNAs ofTricholoma bakamatsutake were sequenced and compared with those of six ectomycorrhizal fungi and a tree. Two probes, Probes 1 and 2, and a pair of primers were designed based on the variable positions in this region. The DNAs ofT. bakamatsutake were isolated from the colonized mycelia in the soil, field-collected fruit-bodies and artifically cultured mycelia. Hybridization with Probe 1 and PCR-amplification with the primers differentiated these DNAs of this fungus from those of eight ectomycorrhizal fungi and two tree species.     

UAE-IC法测定大白口蘑中阴离子含量

采用超声波（UAE）辅助提取-离子色谱（IC）法测定了大白口蘑中F?、Cl?、NO2?、NO3?、PO43? 5种阴离子。选用NaOH溶液（3.0mmol/L）超声渗提30min后进样100μL,通过流速为1.2mL/min的淋洗液Na2CO3+NaHCO3（3.5mmol/L+1.0mmol/L）洗脱。F?、Cl?、NO2?、NO3?、PO43? 5种离子线性范围分别为2－25mg/L、2－20mg/L、2－100mg/L、10－55mg/L、10－100mg/L,峰面积RSD分别为0.58%、1.27%、0.73%、0.92%、2.33%,检出限在0.0976－1.0984mg/L之间。大白口蘑样品中未检出NO2?,其余F?、Cl?、NO3?、PO43? 4种离子加标回收率在95%－110%之间。     

Phylogenetic species delimitation in ectomycorrhizal fungi and implications for barcoding: the case of the Tricholoma scalpturatum complex (Basidiomycota)

Population studies have revealed that the fungal ectomycorrhizal morphospecies Tricholoma scalpturatum consists of at least two genetically distinct groups that occur sympatrically in several geographical areas. This discovery prompted us to examine species boundaries and relationships between members formerly assigned to T. scalpturatum and allied taxa using phylogenetic analyses. Sequence data were obtained from three nuclear DNA regions [internal transcribed spacer (ITS), gpd and tef], from 101 carpophores collected over a large geographical range in Western Europe, and some reference sequences from public databases. The ITS was also tested for its applicability as DNA barcode for species delimitation. Four highly supported phylogenetic clades were detected. The two previously detected genetic groups of T. scalpturatum were assigned to the phylospecies Tricholoma argyraceum and T. scalpturatum. The two remaining clades were referred to as Tricholoma cingulatum and Tricholoma inocybeoides. Unexpectedly, T. cingulatum showed an accelerated rate of evolution that we attributed to narrow host specialization. This study also reveals recombinant ITS sequences in T. inocybeoides, suggesting a hybrid origin. The ITS was a useful tool for the determination of species boundaries: the mean value of intraspecific genetic distances in the entire ITS region (including 5.8S rDNA) was <0.2%, whereas interspecific divergence estimates ranged from 1.78% to 4.22%. Apart from giving insights into the evolution of the T. scalpturatum complex, this study contributes to the establishment of a library of taxonomically verified voucher specimens, an a posteriori correlation between phenotype and genotype, and DNA barcoding of ectomycorrhizal fungi.     

Tricholoma matsutake in a natural Pinus densiflora forest: correspondence between above- and below-ground genets, association with multiple host trees and alteration of existing ectomycorrhizal communities

Tricholoma matsutake (matsutake) is an ectomycorrhizal (ECM) fungus that produces economically important mushrooms in Japan. Here, we use microsatellite markers to identify genets of matsutake sporocarps and below-ground ECM tips, as well as associated host genotypes of Pinus densiflora. We also studied ECM fungal community structure inside, beneath and outside the matsutake fairy rings, using morphological and internal transcribed spacer (ITS) polymorphism analysis. Based on sporocarp samples, one to four genets were found within each fairy ring, and no genetic differentiation among six sites was detected. Matsutake ECM tips were only found beneath fairy rings and corresponded with the genotypes of the above-ground sporocarps. We detected nine below-ground matsutake genets, all of which colonized multiple pine trees (three to seven trees per genet). The ECM fungal community beneath fairy rings was species-poor and significantly differed from those inside and outside the fairy rings. We conclude that matsutake genets occasionally establish from basidiospores and expand on the root systems of multiple host trees. Although matsutake mycelia suppress other ECM fungi during expansion, most of them may recover after the passage of the fairy rings.     

四川松茸内生细菌群落结构与多样性

为了解松茸的内生细菌群落结构与多样性,以采自四川7个松茸主产区的14个松茸样品为材料,通过变性梯度凝胶电泳(DGGE)分析其内生细菌的群落结构与多样性差异.结果表明： 各个产区松茸样品的内生细菌群落结构与多样性存在明显差异.但采自相近地点、相似环境的松茸样品内生细菌群落结构相似性较高;样品的丰度范围为15～25,多样性指数依次为盐源＞冕宁＞会东＞木里＞雅江＞盐边＞小金;条带回收测序构建的系统发育树显示,松茸内生细菌种群丰富,各个产区松茸优势种群存在一定的差异,但假单胞菌属、爱文菌属、芽孢杆菌属在所有样品中均有分布,而且都占据一定的优势地位.产碱杆菌属与鞘氨醇杆菌属在大部分样品中为优势种群;而杜擀氏菌属、赖氨酸芽孢杆菌属在特定样品中为优势种群.表明松茸内生细菌具有丰富的多样性,研究结果可为筛选促进松茸生长的内生菌提供依据.     

新疆巴音布鲁克草原白蘑蘑菇圈土壤真菌多样性分析

【背景】新疆巴音布鲁克草原为我国第二大高山草原,草原上生长的白蘑(Tricholoma mongolicum Imai)属于口蘑属(Tricholoma)的一种,可形成典型的蘑菇圈。【目的】了解巴音布鲁克草原白蘑蘑菇圈圈上及其两侧土壤真菌的物种组成和群落结构特点,为研究白蘑蘑菇圈的形成与生长及其子实体的发生提供一定的理论依据。【方法】采用Illumina HiSeq平台高通量测序技术研究白蘑蘑菇圈周围土壤真菌的物种组成丰富度和菌群结构多样性。【结果】测定有效序列经注释得到809个操作分类单元(Optical taxonomic unit,OTU),分为5个门、26个纲、79个目、166个科、232个属;蘑菇圈上土壤真菌的物种丰富度、群落结构多样性以及特异性物种均低于圈外及圈内,蘑菇圈上特有真菌物种为Xylodonnothofagi、Agaricales、Phaeococcomyces、Ochrocladosporium adansoniae、Coniochaeta ligniaria、未知子囊菌、 Tomentella amyloapiculata、 Dothideomycetes、Incertea_sedis_Helotiales。【结论】研究表明白蘑蘑菇圈的形成影响土壤真菌的分布,圈上优势真菌类群对其他真菌类群具有抑制作用,蘑菇圈向外生长可能与微生物群落结构失衡有关。     

新千年的挑战：第三届世界植物化感作用大会综述

新千年的挑战:第三届世界植物化感作用大会综述@孔垂华$华南农业大学热带亚热带生态研究所!广州510642