Approach of allelopathy study with Arabidopsis thaliana (L.) Hevnh. and Neurospora crassa

Allelopathy in Arabidopsis seeds was investigated in the present study because there are few available reports of allelopathy studies regarding the early development stage of the plant life cycle, and because Arabidopsis is widely used as a model plant for molecular biology. When cockscomb ( Celosia cristata ) was incubated together with Arabidopsis seeds, the growth of cockscomb was promoted by Arabidopsis seeds. Exudate of Arabidopsis seeds showed a promotive activity on the growth of cockscomb and Arabidopsis itself. A new method for studying allelopathic mechanisms involving the interaction of the plant Arabidopsis thaliana (L.) Hevnh. and the fungus Neurospora crassa was suggested. Arabidopsis seed exudate and the water-soluble fraction of the exudate promoted Arabidopsis growth and reduced fungal germination, indicating that the seed exudate has a species-selective activity. Conversely, the ethylacetate-soluble fraction inhibited growth of all tested materials in the current study. These results demonstrate that the water-soluble substance(s) released from Arabidopsis seeds have species-selective activity on growth of both plants and microorganisms.     

Neurospora crassa对三唑醇的抗药性分子机制研究

 通过紫外光诱变获得Neurospora crassa对三唑醇杀菌剂抗性突变体"Nc70"。用2.5μg/ml三唑醇处理,突变体内甲基甾醇与脱甲基甾醇比例变化幅度远远小于亲本敏感菌株"STA"。突变体对三唑醇的吸收能力与敏感菌株相似,但药剂在体细胞内积累较少。三唑醇处理N.crassa 2 h后,药剂在菌体内的含量趋向稳定,突变体内的药量较敏感菌体低56.1%,而麦角甾醇合成量则高于敏感菌株53.7%。     

Characterization of mutations in the two-component histidine kinase gene that confer fludioxonil resistance and osmotic sensitivity in the os-1 mutants of Neurospora crassa

Osmotic-sensitive (os-1) mutant alleles in Neurospora crassa exhibit resistance to dicarboximides, aromatic hydrocarbons and phenylpyrroles. We have previously reported that the os-1 mutants can be classified into two groups based on their resistance to fungicides and osmotic stress: type I, which are highly resistant to iprodione and fludioxonil but moderately sensitive to osmotic stress, and type II, which are highly sensitive to osmotic stress but moderately resistant to fungicides. To explain the mechanism of resistance to these fungicides, we cloned and sequenced the mutant os-1 genes that encode putative osmo-sensing histidine kinase. Within the os-1 gene product (Os1p), the type I strains, NM233t and Y256M209, carried a stop codon at amino acid position 308 and a frameshift at amino acid position 294, respectively. These mutation sites were located on the upstream of histidine kinase and the response regulator domains of Os1p, strongly suggesting that type I strains are null mutants. The null mutants, NM233t and Y256M209, were highly resistant to iprodione and fludioxonil; thus Os1p is essential for these fungicides to express their antifungal activity. The amino acid changes in Os1p, 625Pro from Leu, 578Val from Ala, and 580Arg from Gly were found in the type II strains, M16, M155-1 and P5990, respectively. Os1p is novel in having six tandem repeats of 90 amino acids in the N terminal. Each amino acid change of the type II strains was located on the fifth unit of six tandem repeats. Type II strains with single amino acid changes were more sensitive to osmotic stress than the null mutants (type I), indicating that the amino acid repeats of Os1p were responsible for an important function in osmo-regulation.     

Evaluation of phenylaminopyrimidines as antifungal protein kinase inhibitors

The effects of diverse phenylaminopyrimidines (PAP), namely PAP-pyridines (type A), PAP-pyrazoles (type B) and PAP-thiazoles (type C), on Neurospora crassa Shear & Dodge has been investigated. The results revealed that type A strongly inhibit the in vitro growth of N crassa, whereas types B and C are much less active. A significant correlation was observed between the Neurospora growth inhibition and the intrinsic activity of type A compounds on the cyclin-dependent protein kinase p34(CDC2) of starfish, suggesting that the target of phenylaminopyrimidines in fungi is a cyclin-dependent protein kinase (CDK). The phenylaminopyrimidine-binding CDKs Phoss (major band) and CDC2 (minor band) involved in phosphorus uptake, glycogen synthesis and the cell cycle were identified from N crassa by affinity chromatography on phenylaminopyrimidine-sepharose. Comparative experiments with different protein kinases revealed the importance of the side chain of phenylaminopyrimidines for their target selectivity. A type B compound was found to selectively inhibit the MAP-kinase OS-2 involved in the osmoregulatory pathway of Neurospora.     

好食脉孢菌和红酵母协同发酵秸秆产类胡萝卜素的研究

以好食脉孢菌和红酵母为发酵菌种,通过固态发酵玉米秸秆生产类胡萝卜素。将类胡萝卜素产量作为发酵指标,通过对培养基含水量、培养基初始pH值、氮源添加量、碳源添加量、无机盐添加量、培养基装量的单因素试验和正交试验确定最优培养基条件:秸秆用量10 g,干豆渣(氮源)用量3 g,麸皮(碳源)用量4 g,MgSO4添加量0.4%(与秸秆量比W/W),pH值约为5.5,培养基含水量为60%,培养基装量为12.5 g/250 mL,最佳发酵条件为接种量20%(好食脉孢菌∶红酵母=1∶1),培养温度28℃,培养时间96 h。通过优化后的类胡萝卜素产量为224.75μg/g。     

茶树根际土壤抗酸铝微生物的分离与初步鉴定

将茶树根际土壤微生物提取液接种到含铝5 ～20 mmol/L的培养基上培养并分离获得抗酸铝真菌ALF 1 。该真菌具有抗酸铝能力, 能在含铝20 mmol/L的培养基上正常生长, 并降低培养基铝浓度且使苏木精褪色。经鉴定, ALF 1 真菌为链孢霉属的1 种( Neurosporasp.) 。     

好食脉孢霉拮抗细菌的分离与鉴定

从桉树枯叶中分离到4株（ED018、ED013、ED015、ED014）对好食脉孢霉（Neurospora sitophila）具有拮抗作用的细菌。它们在Martin平板上对崎生长36h后,抑菌圈分别平均为26.2mm、24.1mm、23.8mm和19.0mm,与多菌灵有极显著性的差异。细菌菌株表现明显的抑制好食脉孢霉孢子的萌发作用,在棉籽壳自然基质培养基上,也表现明显的拮抗作用,对多种常见食用菌菌丝生长无抑制作用,对曲霉有抑制作用,对常见多种污染霉菌无抑制作用。4株细菌菌株均表现为革兰氏阳性,杆状,具芽孢,运动,结合其生理生化反应特征,鉴定4株菌株均属于芽孢杆菌属（Bacillus spp.）。     

一株固态发酵甘蔗渣产类胡萝卜素脉孢菌的分离鉴定

本试验于发霉变质的甘蔗渣中分离、筛选得到1株编号为NC2636的产色素菌株。采用分光光度法对该菌株固态发酵甘蔗渣所产的色素进行定性定量分析。并观察菌株NC2636的培养特征和显微特征,以及采用ITS序列对比的方法对其进行鉴定。结果表明:通过分光光度法确定菌株NC2636固态发酵甘蔗渣所产的色素为类胡萝卜素,最大吸收波长为469 nm,28℃固态发酵6 d,色素含量为95.63μg/g。菌株NC2636的ITS序列与Neurospora crassa(KR399979)的序列同源性为100%,结合其培养特征和微观形态特征,故将菌株NC2636鉴定为一株粗糙脉孢菌(Neurospora crassa)。     

5株拮抗细菌对链孢霉的拮抗作用

用平板对峙法筛选出5株对链胞霉有拮抗作用的细菌。结果表明,SK-12、SK-10对链孢霉的拮抗能力最强;浓缩后拮抗细菌拮抗能力显著提高;浓度为20%时,SK-1、SK-10、SK-11对W-N、R-N的抑菌率均达到100%,且培养6d后,W-N仍然不能生长。     

食用菌污染菌脉孢菌的生物防治

通过对峙培养、发酵液的抑菌试验,发现枯草芽孢杆菌BS-2对好食脉孢菌Neurospora sitophila有较强的抑制作用,BS-2能显著地抑制脉孢菌菌丝生长及其孢子萌发,但对食用菌抑制作用相对较弱。BS-2主要的抑菌物质可能是其分泌的多种抗菌蛋白质,BS-2的抑菌能力可通过脉孢菌代谢产物的诱导而加强。