基因组分析揭示拟茎点霉XP-8产松脂醇及其糖苷等多种次级代谢产物的合成途径

【目的】通过解析拟茎点霉属XP-8的基因组序列信息,揭示该菌株潜在的代谢途径,并分析松脂醇及其糖苷化合物等次级代谢产物生物合成相关的关键基因。【方法】使用Illumina Hi Seq 2500高通量测序平台对拟茎点霉XP-8菌株进行全基因组测序,并通过不同软件对测序数据进行序列拼接,基因预测与功能注释。【结果】组装后的拟茎点霉XP-8基因组大小为55.2 Mb,GC含量53.5%,含有17094个蛋白编码基因和310个非编码基因。获得了松脂醇及其糖苷化合物等次级代谢产物生物合成相关的基因。系统发育分析揭示出拟茎点霉XP-8与5种子囊菌共有12635个同源基因和5626个基因家族。【结论】拟茎点霉XP-8具有用于合成松脂醇及其糖苷化合物等多种次级代谢物的基因组基础,为下一步的代谢工程改造提供依据。     

Simultaneous production of hydrogen and bioflocculant by Enterobacter sp. BY-29

Hydrogen and a bioflocculant could be produced simultaneously by anaerobic culture of Enterobacter sp. BY-29. For production of hydrogen and the bioflocculant by cell culture of the bacterium in batch cultures, cultivation at 37 °C in a medium containing glucose as a carbon source and Polypepton as a nitrogen source was found to be suitable. In continuous production of hydrogen and the bioflocculant by cell culture or immobilized cells of the bacterium, the hydrogen production rate and hydrogen yield by the immobilized cells on porous glass beads in stirred and column reactors were higher than those by the cell culture in a stirred reactor. However, production of the bioflocculant by the cell culture was superior to that by the immobilized cells in continuous production.     

The role of kairomones in prey finding by Diomus sp. and Exochomus sp., two coccinellid predators of the cassava mealybug, Phenacoccus manihoti

We studied searching behaviour of Diomus sp., a coccinellid predator introduced into Africa as a natural enemy of the cassava mealybug, when searching on cassava leaves, and compared its behaviour with the searching behaviour of Exochomus sp., an African predator of mealybugs (MB's). Female adults of Diomus and Exochomus spent more time searching on cassava leaves previously infested with cassava MB than on clean cassava leaves, in response to substances produced by MB's (wax and/or honeydew) still present on these leaves as kairomones after removal of the MB's. Both species were also arrested by wax and/or honeydew of the citrus mealybug, Planococcus citri. When offered a choice between kairomones of both MB species only experienced Diomus (reared on cassava MB) showed a clear preference for kairomone of cassava MB. Separate influences of wax and honeydew were tested. Wax from cassava MB was an arrestment stimulus for both coccinellid species. Honeydew produced by cassava MB arrested Exochomus and inexperienced Diomus.

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Zusammenfassung Das Suchverhalten auf Cassaveblätter des coccinelliden Raübers Diomus, eines aus Süd Amerika importierten natürlichen Feindes der Cassaveschmierlaus wurde studiert und verglichen mit dem Suchverhalten eines afrikanischen Räubers von Schmierläusen, Exochomus sp.. Weiblichen Adulten von Diomus un Exochomus verwendeten mehr Zeit auf Cassaveblätter wenn diese Blätter vorher mit Cassaveschmierläuse infiziert waren als auf uninfizierte Blätter, und gebrauchten Substanzen (Wachs und/oder Honigtau) die von Schmierläuse produziert wurden und nach der Entfernung der Schmierläuse auf die Blätter zugeblieben waren als Kairomonen.Beide Arten wurden auch von Wachs und/oder Honigtau der Citrusschmierlaus, Planococcus citri, auf den Blätter arretiert. Wenn die Tiere wahlen könnten zwischen Kairomone der beiden Schmierlausarten wurden die Kairomonen der Cassaveschmierlaus nur von erfahrenen Diomus-Individuen, die auf Cassaveschmierlaüse aufgezogen wurden, bevorzugt.Die Einflüsse von Wachs und Honigtau wurden separat geprüft. Wachs von Exuvien der Cassaveschmierlaus ist ein Arretierungsstimulus für beide Räuberarten. Von Cassaveschmierläusen produzierten Honigtau arretierte Exochomus-und unerfahrene Diomus-Individuen.

     

Trehalose biosynthesis in Thermus thermophilus RQ-1: biochemical properties of the trehalose-6-phosphate synthase and trehalose-6-phosphate phosphatase

The genes for trehalose synthesis in Thermus thermophilus RQ-1, namely otsA [trehalose-phosphate synthase (TPS)], otsB [trehalose-phosphate phosphatase (TPP)], and treS [trehalose synthase (maltose converting) (TreS)] genes are structurally linked. The TPS/TPP pathway plays a role in osmoadaptation, since mutants unable to synthesize trehalose via this pathway were less osmotolerant, in trehalose-deprived medium, than the wild-type strain. The otsA and otsB genes have now been individually cloned and overexpressed in Escherichia coli and the corresponding recombinant enzymes purified. The apparent molecular masses of TPS and TPP were 52 and 26 kDa, respectively. The recombinant TPS utilized UDP-glucose, TDP-glucose, ADP-glucose, or GDP-glucose, in this order as glucosyl donors, and glucose-6-phosphate as the glucosyl acceptor to produce trehalose-6-phosphate (T6P). The recombinant TPP catalyzed the dephosphorylation of T6P to trehalose. This enzyme also dephosphorylated G6P, and this activity was enhanced by NDP-glucose. TPS had an optimal activity at about 98°C and pH near 6.0; TPP had a maximal activity near 70°C and at pH 7.0. The enzymes were extremely thermostable: at 100°C, TPS had a half-life of 31 min, and TPP had a half-life of 40 min. The enzymes did not require the presence of divalent cations for activity; however, the presence of Co²⁺ and Mg²⁺ stimulates both TPS and TPP. This is the first report of the characterization of TPS and TPP from a thermophilic organism.     

转录组和代谢组分析瘤菌根菌对铁皮石斛的促生机制

为筛选出促进铁皮石斛(Dendrobium officinale)生长的差异表达基因和差异代谢物,对瘤菌根菌与无菌盆栽铁皮石斛苗共生后形成的侧根根系进行转录组、代谢组和双组学联合分析。结果表明,转录组分析共找到262条差异表达基因富集到了35条通路中,其中内质网蛋白质加工通路途径的差异基因最多,其次为氨基糖和核苷酸糖代谢通路。代谢组分析共检测出194个差异代谢物富集到33个KEGG通路中,其中代谢途径的差异代谢物最多有133个,其次为不同环境的微生物代谢途径的差异代谢物有70个。通过联合分析,有9个差异基因的差异表达导致丝氨酸、谷氨酸、D-甘露糖和激素等代谢物的积累量发生变化,这可能是瘤菌根菌促进铁皮石斛生长的重要原因。因此,推测瘤菌根菌促进铁皮石斛生长与氨基酸、糖、植物激素的积累及相关基因的表达变化有关。     

Clostridium halophilium sp. nov. and C. litorale sp. nov., an obligate halophilic and a marine species degrading betaine in the Stickland reaction

Two new mesophilic, sporeforming, gram-positive, strictly anaerobic, rod-shaped bacteria were isolated which utilized betaine in the Stickland reaction. Strain M1 was obtained from pasteurized hypersaline sediments. Cells were motile rods and formed spherical terminal spores. Betaine was used with hydrogen and several amino acids as electron donors. In addition, several carbohydrates served as substrates. Growth required 1.5% NaCl with an optimum at 6.0% NaCl. The guanine plus cytosine content of the DNA was 26.9%. This strain is described as a new species, Clostridium halophilum.Strain W6 was isolated from marine sediments. Cells were motile rods and formed ovoid, subterminal spores. Betaine was used with hydrogen and several amino acids as electron donors. Carbohydrates were not fermented. Growth optimum was at 1.0% NaCl. The guanine plus cytosine content of the DNA was 26.1%. This strain is described as a new species, Clostridium litorale.Non standard abbreviations DMG N,N-dimethylglycine - TMA trimethylamine - PY peptone-yeast extract - PYG peptone-yeast extract-glucose     

Bullera begoniae sp. nov. and Bullera setariae sp. nov., two new species of ballistoconidium-forming yeasts in the Bullera variabilis (Bulleribasidium) cluster isolated from plants in Taiwan

Two strains of xylose-containing and Q-10-having ballistoconidiogenous yeasts isolated from plant leaves collected in Taiwan were found to represent two new species of the genus Bullera. In the phylogenetic trees based on the sequence analysis of 18S rDNA and D1/D2 domain of 26S rDNA, these species are located in the Bullera variabilis (Bulleribasidum) cluster in Hymenomycetes. They are described as Bullera begoniae sp. nov. and Bullera setariae sp. nov., respectively.     

Intracellular changes of trehalose content in toluene tolerant Pseudomonas sp. BCNU 171 after exposure to toluene

Pseudomonas sp. BCNU 171, when grown with 10% (v/v) toluene for 2 h, accumulated approximately 7.7 mM trehalose probably arising by the action of trehalose-6-phosphate synthase (E.C. 2.4.1.15). Trehalose may thus play a significant role in the tolerance of the Pseudomonas sp. to toluene.     

Purification of component C from Methanosarcina mazei and immunolocalization in Methanosarcinaceae

Studies on immunological relationships among Methanosarcina genus using immunofluorescence and immunoprecipitation showed that a common antigen can be extracted by shaking in aqueous phase. This antigen was purified from Methanosarcina mazei. The protein had a molecular weight of 283400 daltons with three subunits, =68000, =43200 and =30500. It contained nickel, coenzyme M and F430. Its biochemical characteristics identified this antigen as the component C of the methyl CoM reductase complex. But EPR study showed that the nickel was Ni(II). Biological activity was detectable neither by heterologous in vitro assay nor by the DTT assay. Immunogold labelling showed that the component C was located randomly in the cytoplasm in Methanosarcina species and in Methanothrix soehngenii. In addition, specific labelling was also observed outside of the heteropolysaccharidic envelopes probably due to the absorption of component C released by the lysis of some cells in the clumps.     

Essential roles of snap-29 in C. elegans

SNARE domain proteins are key molecules mediating intracellular fusion events. SNAP25 family proteins are unique target-SNAREs possessing two SNARE domains. Here we report the genetic, molecular, and cell biological characterization of C. elegans SNAP-29. We found that snap-29 is an essential gene required throughout the life-cycle. Depletion of snap-29 by RNAi in adults results in sterility associated with endomitotic oocytes and pre-meiotic maturation of the oocytes. Many of the embryos that are produced are multinucleated, indicating a defect in embryonic cytokinesis. A profound defect in secretion by oocytes and early embryos in animals lacking SNAP-29 appears to be the underlying defect connecting these phenotypes. Further analysis revealed defects in basolateral and apical secretion by intestinal epithelial cells in animals lacking SNAP-29, indicating a broad requirement for this protein in the secretory pathway. A SNAP-29-GFP fusion protein was enriched on recycling endosomes, and loss of SNAP-29 disrupted recycling endosome morphology. Taken together these results suggest a requirement for SNAP-29 in the fusion of post-Golgi vesicles with the recycling endosome for cargo to reach the cell surface.     

Nodulation in black locust by the Gammaproteobacteria Pseudomonas sp. and the Betaproteobacteria Burkholderia sp.

Nodulation abilities of bacteria in the subclasses Gammaproteobacteria and Betaproteobacteria on black locust (Robinia pseudoacacia) were tested. Pseudomonas sp., Burkholderia sp., Klebsiella sp., and Paenibacillus sp. were isolated from surface-sterilized black locust nodules, but their nodulation ability is unknown. The aims of this study were to determine if these bacteria are symbiotic. The species and genera of the strains were determined by RFLP analysis and DNA sequencing of 16S rRNA gene. Inoculation tests and histological studies revealed that Pseudomonas sp. and Burkholderia sp. formed nodules on black locust and also developed differentiated nodule tissue. Furthermore, a phylogenetic analysis of nodA and a BLASTN analysis of the nodC, nifH, and nifHD genes revealed that these symbiotic genes of Pseudomonas sp. and Burkholderia sp. have high similarities with those of rhizobial species, indicating that the strains acquired the symbiotic genes from rhizobial species in the soil. Therefore, in an actual rhizosphere, bacterial diversity of nodulating legumes may be broader than expected in the Alpha-, Beta-, and Gammaproteobacteria subclasses. The results indicate the importance of horizontal gene transfer for establishing symbiotic interactions in the rhizosphere.     

Growth of Rhizobium sp. in the presence of catechol

The ability of Rhizobium sp., isolated from Lablab purpureus to survive in soil containing a phenolic derivative catechol was investigated. It survived for 9 months in soil containing catechol. In synthetic medium, Rhizobium sp. utilized catechol up to 10 mM as sole carbon source and catechol 1, 2-dioxygenase was present in the catechol grown cells. In the presence of organic acids and sugars, catechol was co-metabolized; but catechol 1, 2-dioxygenase induction was inhibited. The ability of Rhizobium sp. to utilize various phenolic substances provides potential advantage to overcome the phenolic toxicants present in soils.     

利用Mariannaea sp.HJ菌株胞内提取物合成纳米银及其抗菌特性研究

【目的】近年来,纳米银由于其自身独特的抗菌活性而受到越来越多的关注,有研究表明纳米银是一种广谱的抗菌剂,其对数十种致病微生物都有强烈的抑制和杀灭作用。相较于传统的合成方法而言,具有反应条件温和、环境友好等优势的生物合成法是目前的研究热点。【方法】本研究利用真菌Mariannaea sp. HJ的胞内提取物合成纳米银,并对其合成条件进行优化调控,还进一步考察了合成的纳米银颗粒的抗菌性能。【结果】胞内提取物浓度350 mg/L、AgNO_3浓度5 mmol/L、pH 7.0为菌株HJ胞内提取物合成纳米银的最优条件;TEM图像表明合成的纳米银颗粒主要为球形和伪球形,分散性良好,无明显的团聚现象;XRD表明合成的纳米银晶体结构为面心立方体结构;通过FTIR分析结果推测提取物中的羟基、羧基等官能团可能参与了纳米银的还原和稳定过程。此外,在本实验条件下合成的纳米银颗粒对革兰氏阴性菌Escherichia coli BL21和革兰氏阳性菌Arthrobacter sp. W1都有较好的抗菌活性。【结论】真菌Mariannaea sp. HJ胞内提取物能合成尺寸均一且分散性良好的球形纳米银颗粒,合成的纳米银颗粒在抗菌方面具有潜在的研究价值。     

Peroxidase from Bacillus sp. VUS and its role in the decolorization of textile dyes

Peroxidase was purified by an ion exchange chromatography followed by gel filtration chromatography from dye degrading Bacillus sp. strain VUS. The optimum pH and temperature of the enzyme activity was 3.0 and 65°C, respectively. This enzyme showed more activity with n-propanol than other substrates tested viz. xylidine, 3-(3,4-dihydroxy phenyl) Lalanine (L-DOPA), hydroxyquinone, ethanol, indole, and veratrole. Km value of the enzyme was 0.076 mM towards n-propanol under standard assay conditions. Peroxidase was more active in presence of the metal ions like Li²⁺, Co²⁺, K²⁺, Zn²⁺, and Cu²⁺ where as it showed less activity in the presence of Ca²⁺ and Mn²⁺. Inhibitors like ethylenediamine tetraacetic acid (EDTA), glutamine, and phenylalanine inhibited the enzyme partially, while sodium azide (NaN₃) completely. The crude as well as the purified peroxidase was able to decolourize different industrial dyes. This enzyme decolourized various textile dyes and enhanced percent decolourization in the presence of redox mediators. Aniline was the most effective redox mediator than other mediators tested. Gas chromatography-Mass spectrometry (GC-MS) confirmed the formation of 7-Acetylamino-4-hydroxy-naphthalene-2-sulphonic acid as the final product of Reactive Orange 16 indicating asymmetric cleavage of the dye.     

Pelotomaculum terephthalicum sp. nov. and Pelotomaculum isophthalicum sp. nov.: two anaerobic bacteria that degrade phthalate isomers in syntrophic association with hydrogenotrophic methanogens

An anaerobic phthalate isomer-degrading strain (JT^T) that we previously isolated was characterized. In addition, a strictly anaerobic, mesophilic, syntrophic phthalate isomer-degrading bacterium, designated strain JI^T, was isolated and characterized in this study. Both were non-motile rods that formed spores. In both strains, the optimal growth was observed at temperatures around 37°C and neutral pH. In syntrophic co-culture with the hydrogenotrophic methanogen Methanospirillum hungatei, both strains could utilize two or three phthalate isomers for growth, and produce acetate and methane as end products. Strain JT^T was able to grow on isophthalate, terephthalate, and a number of low-molecular weight aromatic compounds, such as benzoate, hydroquinone, 2-hydroxybenzoate, 3-hydroxybenzoate, 2,5-dihydroxybenzoate, 3-phenylpropionate in co-culture with M. hungatei. It could also grow on crotonate, hydroquinone and 2,5-dihydroxybenzoate in pure culture. Strain JI^T utilized all of the three phthalate isomers as well as benzoate and 3-hydroxybenzoate for growth in co-culture with M. hungatei. No substrates were, however, found to support the axenic growth of strain JI^T. Neither strain JT^T nor strain JI^T could utilize sulfate, sulfite, thiosulfate, nitrate, fumarate, Fe (III) or 4-hydroxybenzoate as electron acceptor. Phylogenetically, strains JT^T and JI^T were relatively close to the members of the genera Pelotomaculum and Cryptanaerobacter in ‘Desulfotomaculum lineage I’. Physiological and chemotaxonomic characteristics indicated that the two isolates should be classified into the genus Pelotomaculum, creating two novel species for them. Here, we propose Pelotomaculum terephthalicum sp. nov. and Pelotomaculum isophthalicum sp. nov. for strain JT^T and strain JI^T, respectively. The type strains are strains JT^T (= DSM 16121^T = JCM 11824^T = NBRC 100523^T) and JI^T (= JCM 12282^T = BAA-1053^T) for P. terephthalicum and P. isophthalicum, respectively.Nucleotide sequence accession number: The GenBank/EMBL/DDBJ accession numbers of the 16S rRNA gene sequences of strains JT^T and JI^T are AB091323 and AB232785, respectively     

Senecella siberica n. sp. and the position of the genus Senecella in Calanoida classification

Senecella calanoides Juday has been reported from fresh-water lakes of North America, Northern Siberia and inshore brackish waters of Kara and Laptev Seas. Investigation of 11 females and 2 males from the brackish waters of Neelov Bay (Laptev Sea) shows that second species, Senecella siberica sp. n., can be distinguished from S. Calanoides Juday by the greater size, a slightly asymmetrical female genital segment, the 3 external spines on the 3d exopod of 2 to 4th swimming legs and by more longer left endopod digitiform process of the male 5th legs. S. siberica seems to be brackish-water euryhaline species, inhabiting brackish waters with salinity from 1 to 22. The genus belongs to the family Aetideidae and is the only genus with freshwater representatives.     

禾谷镰刀菌拮抗菌21-6的鉴定及其抑菌活性测定

【背景】禾谷镰刀菌（Fusarium graminearum）是一种危害小麦生产的重要病原真菌。【目的】筛选对禾谷镰刀菌具有拮抗活性的链霉菌菌株,为该病原的生物防治提供理论基础。【方法】采用稀释涂布法分离链霉菌,利用平板对峙法筛选高活性拮抗菌株;通过形态学、生理生化特征和16S rRNA基因序列分析确定其分类地位;采用生长速率法分析其发酵条件及无菌发酵液的稳定性;并测定该菌株的防病效果和抑菌谱。【结果】筛选到一株对禾谷镰刀菌具有较强抑制作用的链霉菌菌株21-6,抑菌率为75.2%±2.1%。根据形态学、生理生化特征及16S rRNA基因序列分析,将其鉴定为Streptomyces stelliscabiei。菌株21-6在pH为中性条件下的PDB培养基中培养5 d能够产生更好的抑菌效果。无菌发酵液能够抑制禾谷镰刀菌的菌丝生长和孢子萌发过程。无菌发酵液不受高温、胃蛋白酶、胰蛋白酶及蛋白酶K的影响,耐酸但对碱性条件敏感。发酵液对禾谷镰刀菌侵染小麦胚芽鞘具有抑制效果。菌株21-6具有聚酮合酶pks-Ⅰ和pks-Ⅱ基因。此外,该菌株对5种植物病原真菌均具有抑制效果。【结论】链霉菌菌株21-6对禾谷镰刀菌具有较好的生防潜力。     

Biodegradation and detoxification of nicotine in tobacco solid waste by a Pseudomonas sp.

A Pseudomonas sp. grew with nicotine optimally 3 g l^–1 and at 30 °C and pH 7. Nicotine was fully degraded within 10 h. The resting cells degraded nicotine in tobacco solid waste completely within 6 h in 0.02 m sodium phosphate buffer (pH 7) at maximally 56 mg nicotine h^–1 g dry cell^–1.     

Purification and properties of extracellular phytase from Bacillus sp. KHU-10

Bacillus species producing a thermostable phytase was isolated from soil, boiled rice, and mezu (Korean traditinal koji). The activity of phytase increased markedly at the late stationary phase. An extracellular phytase from Bacillus sp. KHU-10 was purified to homogeneity by acetone precipitation and DEAE-Sepharose and phenyl-Sepharose column chromatographies. Its molecular weight was estimated to be 46 kDa on gel filtration and 44 kDa on SDS-polyacrylamide gel elctrophoresis. Its optimum pH and temperature for phytase activity were pH 6.5-8.5 and 40°C without 10 mM CaCl₂ and pH 6.0-9.5 and 60°C with 10 mM CaCl₂. About 50% of its original activity remained after incubation at 80°C or 10 min in the presence of 10 mM CaCl₂. The enzyme activity was fairly stable from pH 6.5 to 10.0. The enzyme had an isoelectric point of 6.8. As for substrate specificity, it was very specific for sodium phytate and showed no activity on other phosphate esters. The K _m value for sodium phytate was 50 M. Its activity was inhibited by EDTA and metal ions such as Ba²⁺, Cd²⁺, Co²⁺, Cr³⁺, Cu²⁺, Hg²⁺, and Mn²⁺ ions.     

5种杀虫剂对黑粪蚊的毒力及对平菇菌丝生长的影响

[目的]黑粪蚊为食用菌生产中的危险害虫之一,通过杀虫剂的毒力测定和对平菇菌丝生长抑制试验,拟筛选获得理想的黑粪蚊防治杀虫剂。[方法]采用毒饵法测定了5种杀虫剂对黑粪蚊幼虫的室内毒力,并研究各杀虫剂对平菇菌菌丝生长的抑制情况。[结果]20%甲氰菊酯乳油、14%阿维·虫螨腈悬浮剂和20%甲维·吡丙醚悬浮剂对黑粪蚊的校正死亡率达80%以上,对黑粪蚊幼虫均有较好的杀灭效果,而苏云金杆菌（以色列亚种）悬浮剂对黑粪蚊幼虫的杀虫效果不理想。5种杀虫剂对平菇菌丝生长均有一定抑制作用,且农药品种间存在极显著差异,20%甲氰菊酯乳油抑制率最高,高浓度480 mg·L^-1处理的抑制率可达16.85%,14%阿维·虫螨腈悬浮剂的抑制率最低,高浓度140 mg·L^-1处理的抑制率仅为5.83%。[结论]本研究中14%阿维·虫螨腈悬浮剂用于平菇菌中黑粪蚊防治效果最好,研究结果将为食用菌黑粪蚊的药剂防治提供理论指导。