桦褐孔菌野生分离菌株的比较与栽培选育

通过菌丝形态比较、ISSR分析鉴定及人工驯化栽培选育,对1株野生分离菌株和4株保藏菌株进行比较。结果显示:5个菌株在菌丝形态和长势上差异显著,1号菌株(野生分离)菌丝洁白,长速最快;ISSR扩增图谱谱带差异明显,1号菌株与5号菌株在遗传相似水平85%时聚为1类,其他菌株各自聚为独立1类。驯化栽培试验结果表明:1号菌株出核较快,菌核大,产量高,较其他菌株产量差异显著。被试菌株为各自独立菌株,1号菌株经驯化适宜高产栽培,是重要珍稀野生种质资源。本研究可为野生桦褐孔菌菌株种质资源收集和为实现大规模栽培育种提供亲本资源。     

溶磷菌对铅胁迫下杏鲍菇氧化应激反应的影响

选择使用杏鲍菇菌丝富集重金属Pb2+,从本实验室菌种库中筛选出了8株能抗800 mg/L Pb2+的菌株。通过对其溶磷能力的检测,从中筛选到了2株溶磷能力较强的菌株CG8、FFT1以及2株溶磷能力较弱的菌株FFC6、FFC11,经16S r DNA序列分析鉴定,CG8为克雷伯菌(Klebsiella sp.),FFT1为假单胞菌(Pseudomonas sp.),FFC6和FFC11为芽孢杆菌(Bacillus sp.),其序列相似度均达99%。随后,研究了4株溶磷菌株对液体培养(Pb)条件下杏鲍菇(Pleurotus eryngii)菌丝的生物量、Pb富集量、脂质过氧化、巯基蛋白含量和抗氧化酶的影响。结果显示,在Pb胁迫条件下,具溶磷能力的菌株能在一定程度上增加菌丝生物量,促进菌丝对重金属的富集,同时,使菌丝的丙二醛(MDA)含量降低,菌丝抗氧化酶(SOD)和氧化物酶(POD)活性显著降低,菌丝巯基蛋白含量升高。实验证明,微生物溶磷能力可有效减轻重金属对杏鲍菇菌丝的毒害和重金属诱导的氧化胁迫。并且溶磷能力较强的菌株(CG8)对杏鲍菇菌丝富集与抵抗重金属诱导的氧化胁迫有更为明显的作用,重金属富集量增加96.1%,巯基蛋白增加量为91.3%。     

伞形多孔菌（猪苓）优良菌株的筛选

以10个不同来源的伞形多孔菌（猪苓）菌株为材料,通过ITS序列分析比较菌株间的亲缘关系,基于菌落、菌丝的形态特征、生长速率、菌丝体干重和产糖能力5个方面的14个指标,采用聚类分析方法分析各指标之间、菌株之间的相关性,主成分分析法提取4个主成分进行分析,通过综合分值比较菌株优劣。ITS序列分析表明,10个菌株间的相似性较高。聚类分析结果显示,指标之间胞外多糖、菌丝团、黑色素、菌丝颜色、长势和分泌物的相关性高;菌丝形态、草酸钙方晶、生长速率和无性孢子的相关性高。5#菌株多糖含量和菌丝生长速率最高;2#和9#菌株胞内多糖含量较高;4#、13#和14#菌株多糖含量较低;10#、11#菌株多糖含量及菌丝体干重也较低。主成分分析显示,5#、2#和1#菌株是综合分值最高的3个菌株,其菌丝生长快、长势旺、产量及多糖含量高。综合上述结果,5#、2#和1#菌株是初步筛选出可用于大规模生产及繁殖的优良菌株。     

大丽轮枝菌异核体及其后代的形态与致病力变异

以硝酸盐利用缺陷型突变（nit突变）和抗杀菌剂突变两种遗传标记,对大丽轮枝菌（Verticilliumdahliae）异核体后代的形态和致病力进行研究,结果表明,菌核型菌株与菌丝型菌株经菌丝融合形成异核体后,菌丝型菌株能恢复形成微菌核,其后代单孢菌落形成微菌核的数量明显低于菌核型亲本,且遗传性状不稳定;随着转代次数的增多,微菌核形成能力的丧失较菌核型亲本菌株快。异核体后代对棉苗的致病力变化较大,一般均低于致病力强的亲本菌株,或介于两个亲本致病力之间,或与亲本致病力相近。     

枯草芽孢杆菌B1-41对小麦纹枯病菌的抑制作用

从小麦根际土壤分离得到枯草芽孢杆菌(Bacillus subtilis)B1-41拮抗菌株,室内测定其带菌培养液对小麦纹枯病菌(Rhizoctonia cerealis)抑制效果为72%,盆栽试验中,其防治效果为60.3%,高于井冈霉素处理的51%的效果。试验表明,该菌株可以造成病菌菌丝发生畸变和菌丝细胞壁瓦解。     

贵州小孢子菌——小孢子菌属一新种

从我国贵州省的土壤中分离到1株小孢子菌,菌株GZUIFR-EB2001M在小孢子属系统发育树中独立为一亚分支,明显区别于其他供试种,为该属一新种,命名为贵州小孢子菌。形态学与近似种的主要鉴别特征为:无球拍状菌丝;大分生孢子光滑或粗糙,梭形,细长;小孢子棒状至柱状。     

大丽轮枝菌异核体及其后代的形态与致病力变异

以硝酸盐利用缺陷型突变（ｎｉｔ突变）和抗杀菌剂突变两种遗传标记,对大丽轮枝菌（Ｖｅｒｔｉｃｉｌｌｉｕｍｄａｈｌｉａｅ）异核体后代的形态和致病力进行研究,结果表明,菌核型菌株与菌丝型菌株经菌丝融合形成异核体后,菌丝型菌株能恢复形成微菌核,其后代单孢菌落形成微菌核的数量明显低于菌核型亲本,且遗传性状不稳定;随着转代次数的增多,微菌核形成能力的丧失较菌核型亲本菌株快,异核体后代对棉苗的致病力变化较大,一     

大丽轮枝菌异核体及其后代的形态与致病力变异*

以硝酸盐利用缺陷型突变（nit突变）和抗杀菌剂突变两种遗传标记,对大丽轮枝菌（Verticilliumdahliae）异核体后代的形态和致病力进行研究,结果表明,菌核型菌株与菌丝型菌株经菌丝融合形成异核体后,菌丝型菌株能恢复形成微菌核,其后代单孢菌落形成微菌核的数量明显低于菌核型亲本,且遗传性状不稳定;随着转代次数的增多,微菌核形成能力的丧失较菌核型亲本菌株快。异核体后代对棉苗的致病力变化较大,一般均低于致病力强的亲本菌株,或介于两个亲本致病力之间,或与亲本致病力相近。     

短小芽孢杆菌AR03对烟草赤星病菌和白粉病菌的防治

采用对峙培养法、凹玻片法、LB琼脂培养基萌发法测定短小芽孢杆菌AR03对烟草赤星病菌和白粉病菌的抑制作用.结果表明： AR03菌液对2种病菌的菌丝生长和分生孢子萌发均有明显的拮抗作用;对赤星病菌的抑制作用表现为:经AR03菌液原液(3×10⁸cfu·mL^-1)处理的菌丝隔间变短、肿胀且集结成团,内含物聚集,菌丝顶端生长膨大畸形;经该菌液处理的赤星病菌分生孢子不萌发或萌发产生畸形芽管,分生孢子变形、肿大,纵横分隔部分的组织膨胀呈泡状.AR03菌液原液、30倍液和200倍液对白粉病菌分生孢子萌发的平板抑制率分别为100%、91.4%和 69.3%,菌液对分生孢子萌发的破坏作用表现为分生孢子不萌发,细胞肿胀变形、细胞原生质解体或收缩,孢子内、外壁分离,由于原生质外泄,一些分生孢子内部呈中空状.温室防治试验结果表明： 不同浓度AR03菌悬液处理对烟草白粉病的防治效果存在显著差异,第二次药后7和15 d,AR03菌液原液的防治效果分别达到83.8%和90.3%,与对照药剂差异不显著;而100倍稀释液的防效分别为70.0%和73.3%,与对照药剂差异显著.AR03菌株防治白粉病的持效期为30 d以上.     

云南省水稻纹枯病菌系研究

将来自云南省20多个县市的水稻纹枯病标样130多份,选代表性的标样分离得到54个菌株。按菌丝融合测定法,将54个菌株分为5个菌系：R.solani的AG－1 IA,AG-1 IC,AG-6GW以及以核丝核菌的AG－Bb,AG- ⅠⅡ。经致病性测定表明,该菌系对水稻、玉米、小麦的苗期及成株期的致病性有显著差异。其中AG－1ⅠA,AG－1 IC,AG－Ⅱ的致病力最,AG－6GW的最弱。对这些菌系的酯酶同工酶进行比较研究发现,不同菌系间均存在明显差异,而同菌系不同菌株间却具有一致性。由此说明,按菌丝融合与否区分丝核菌种群较之现行的其它分类法更能反映其遗传本质和亲缘关系,。     

具抗菌活性放线菌菌株JSM 20.1的分离和鉴定

从药用植物银杏(Ginkgo biloba)根际土壤中分离到1株放线菌菌株JSM20.1,其发酵产物具有广谱抗菌活性。该菌株在多数培养基上生长良好,具有链霉菌属的典型形态特征。菌丝多分枝,不断裂;基内菌丝黄白色至棕褐色,气生菌丝黄白色至棕红色;孢子链直且长,偶有波曲,孢子柱状、光滑、不运动。在察氏、马铃薯浸汁、葡萄糖-天门冬酰胺、酵母膏-麦芽膏(ISP2)、燕麦(ISP3)、无机盐淀粉(ISP4)和甘油-天门冬酰胺(ISP5)琼脂上均产生可溶性色素,而在营养琼脂上不产生可溶性色素。生长温度范围为7～40℃,最适生长温度28℃;生长pH范围为5.0～10.0,最适pH7.0;能在含0～3%(质量与体积比)NaCl的培养基上生长,而在无NaCl的ISP2培养基生长最佳。根据其形态学特征、生理生化特征、细胞化学分类特征和基于16SrRNA基因序列的系统发育分析结果,菌株JSM20.1被鉴定为链霉菌属的有效发表种Streptomyces cyaneofuscatus的1个菌株。     

Description of <Emphasis Type="Italic">Streptomyces neopeptinius</Emphasis> sp. nov., an actinobacterium with broad spectrum antifungal activities

A streptomycete strain producing broad-spectrum antifungal substances was taxonomically characterized. The strain, designated KNF 2047(T) (= SH-09(T) = KCTC 10586BP(T)), was found to form extensively branching aerial and substrate mycelia, and produce spiny-ornamented spores with loose spiral chains. The whole cell hydrolyzates contained major amount of LL-diaminopimelic acid. The major fatty acids of the phospholipids were saturated and branched fatty acids containing 14~17 carbons, and the major isoprenoid quinones were hexa-and octa-hydrogenated menaquinones with 9 isoprene units. The phylogenetic analysis using the 16S rRNA gene indicated that the strain belongs to the genus Streptomyces but forms an independent phyletic line. These results clearly demonstrate that strain KNF2047(T) forms a new center of taxonomic variation within Streptomyces, for which the name Streptomyces neopeptinius sp. nov. is proposed.     

<Emphasis Type="Italic">Streptomyces thermoalkaliphilus</Emphasis> sp. nov., an alkaline cellulase producing thermophilic actinomycete isolated from tropical rainforest soil

During an investigation exploring potential sources of novel thermophilic species and natural products, a novel thermophilic and alkaliphilic actinomycete with alkaline cellulase producing ability, designated strain 4-2-13^T, was isolated from soil of a tropical rainforest in Xishuangbanna, Yunnan province, China. The morphological and chemotaxonomic characteristics of strain 4-2-13^T are consistent with those of the members of the genus Streptomyces. The strain forms extensively branched aerial mycelia and substrate mycelia. Spiral spore chains were observed on aerial mycelia; spores were oval to cylindrical, with smooth surfaces. The organism was found to contain ll-diaminopimelic acid as the diagnostic diamino acid in the cell wall peptidoglycan. The whole cell hydrolysates were found to contain glucose and ribose. The cellular fatty acid profile mainly consists of anteiso-C_17:0 and iso-C_16:0. The menaquinones were identified as MK-9(H₈), MK-10(H₆) and MK-9(H₆). The polar lipids profile were found to consist of diphosphatidylglycerol, phosphatidylmethylethanolamine, a ninhydrin-positive glycophospholipid, phosphatidylinositol, phosphatidylglycerol and unidentified glycolipids. The 16S rRNA gene sequence analysis showed that the organism belongs to the genus Streptomyces and in the 16S rRNA gene tree it formed a distinct phyletic line together with the closely related type strain Streptomyces burgazadensis Z1R7^T (95.2% sequence similarity). However, the phenotypic characteristics of strain 4-2-13^T are significantly different from those of S. burgazadensis Z1R7^T. Based on the phenotypic, chemotaxonomic and phylogenetic characteristics, strain 4-2-13^T represents a novel species in the genus Streptomyces, for which the name Streptomyces thermoalkaliphilus sp. nov. is proposed. The type strain is 4-2-13^T (= DSM 42159^T = CGMCC 4. 7205^T).     

链霉菌属菌株AS4.693和AS4.702的分类学研究

链霉菌属“Setae”种群原为北里孢菌属Kitasatosporia (Omura,1982)。1992年,Wellington根据16S rRNA序列分析结果将其并入链霉菌属,并建立“Setae”种群。通过对保藏的链霉菌AS 4.693、AS 4.702进行的形态学、细胞化学、分子遗传分类研究结果表明,它们与链霉菌属“Setae”种群中的典型种——西唐链霉菌Streptomyces setae(JCM3304’)具有相似性。它们的rDNA相似性高达100％,证明它们应归属于同一种群。AS.4.693定名为西唐链霉菌不规则新亚种Streptomyces setae subsp.irregularis nov.,AS 4.702定名为西唐链霉菌波曲弗氏新亚种Streptomyces setae subsp.flexuofradiae nov.。     

高温链霉菌24＃的初步研究

从土壤中筛选得到一株产广谱、高活性抗真菌物质的链霉菌 2 4 # ,经测定对 2 5种植物病原真菌有显著拮抗作用 ,其次生代谢产物对病菌菌丝有断裂、扭曲、缢缩等致畸效应。 16SrDNA序列分析显示本菌株与模式菌株链霉菌DSM4 4 2 93T的 16SrDNA同源性为 98 93% ,但菌株形态特征、培养特征、细胞壁化学组分、生理生化特性等均不同于模式菌株 ,且DNA杂交率只有 2 2 5 4 % ,建议为链霉菌属的一个新种 ,命名为山东链霉菌 (Streptomycesshandon gensissp .nov .)。     

A Simple and Rapid Method of Transformation of Streptomyces rimosus R6 and Other Streptomycetes by Electroporation

Usually plasmid DNA is introduced into Streptomyces strains by polyethylene glycol-mediated transformation of protoplasts. However, many Streptomyces strains are only poorly or not at all transformable via protoplasts. Therefore, we have optimized the parameters critical for the application of electrotransformation of plasmid DNA into Streptomyces species. The most critical parameters evaluated for electrotransformation of the model strain Streptomyces rimosus R6 were the pretreatment of mycelia, buffer composition, and electric field strength. The electrocompetent mycelia were prepared from 24-h-old cultures, treated mildly with lysozyme, resuspended in sucrose-glycerol-polyethylene glycol buffer, and stored in aliquots at -70 deg C. The electric field strength of 10 kV/cm at 400 (Omega) and a capacitance of 25 (mu)F was applied. The method is simple and rapid, yielding transformant colonies in 48 to 72 h. Efficiencies of 10(sup5) to 10(sup6) transformants per (mu)g of plasmid DNA were reproducibly achieved for S. rimosus R6 and its mutants, and these numbers were 10(sup2) to 10(sup3) higher than those attained by polyethylene glycol-assisted transformation of protoplasts. In addition, we show that electroporation can be applied to other Streptomyces species, such as S. lividans 66, S. coelicolor A3(2), and an S. venezuelae strain. This last one could not be transformed by the standard protoplast procedure. Our data suggest that, because of the diversity of streptomycetes, the conditions have to be optimized for each strain.     

<Emphasis Type="Italic">Streptomyces roseoalbus</Emphasis> sp. nov., an actinomycete isolated from soil in Yunnan,China

A strain YIM 33098T (= CCTCC AA001027T = DSM 41831T) was isolated from a forest soil sample collected from Nanning in Guangxi Province, China, in the course of screening for producers of new drug lead compounds. This strain was identified by using a polyphasic approach. The results showed that it should be assigned to the genus Streptomyces. An almost complete 16S rRNA gene sequence of the strain was determined and compared with those of representative Streptomyces species. Strain YIM 33098T was clustered in the same subclade with Streptomyces tendae ATCC19812T and Streptomyces eurythermus ATCC14975T. Similarities of strain YIM 33098T with the two strains were 97.35% and 97.42%, respectively. Based on the phenotypic and genotypic evidence, it is therefore proposed that strain YIM 33098T should be classified in the genus Streptomyces as a new species under the name of Streptomyces nanningensis sp. nov.     

<Emphasis Type="Italic">Streptomyces tritolerans</Emphasis> sp. nov., a novel actinomycete isolated from soil in Karnataka,India

A Gram-positive, nonmotile, moderately halophilic, alkali and thermotolerant strain designated DAS 165(T), was isolated from a dry land soil sample from the Gulbarga region, Karnataka province, India. The isolate produced yellow substrate mycelia and gray aerial mycelia on most tested media. Strain DAS 165(T) showed growth in the presence of 5 to 7% NaCl and at 45 degrees C. The DNA G + C content was 69.7%. 16S rRNA gene sequence analysis together with these characteristics consistently assigned strain DAS 165(T) to the genus Streptomyces. The 16S rRNA gene sequence analysis revealed that strain DAS 165(T) was most closely related to S. tendae ATCC 19812(T) (D 63873) with a sequence similarity of 99.6% (three nucleotide differences out of 1,517). Strain DAS 165(T) formed a distinct clade based on analysis of the almost complete sequence and 120-nucleotide variable gamma region of the 16S rRNA gene. Despite the high sequence similarity, strain DAS 165(T) was phenotypically different from S. tendae ATCC 19812(T). DNA-DNA hybridization between these strains was 47% showing that strain DAS 165(T) is a distinct genomic species. Phenetic and genetic results support the classification of strain DAS 165(T) as a new species, for which the name S. tritolerans is proposed, with strain DAS 165(T) as the type strain (=DSM 41899(T )= CCTCCAA 206013(T)).     

Protein kinase activity in cell free extracts of Streptomyces avermitilis; comparing wild type and overproducing mutant

Summary Protein kinase activity was assayed in cell-free extracts prepared from mycelia of a wild strain (MA) and an overproducing mutant (RX 2) of Streptomyces avermitilis. At least 10 different polypeptides ranging in M _r from 10 000 to 120 000 were found to be phosphorylated on analysis of ³²P labeled cell lysates by gel electrophoresis and phosphorimage. The protein profile and the level of phosphorylation varied depending on the culture stage of the mycelia.     

Antagonism and Action Mechanism of Antifungal Metabolites from Streptomyces rimosus MY02

The genus of Streptomyces , a saprophytic Gram-positive bacterium, has properties, which make them useful as pharmaceutical and biocontrol agents. A streptomyces strain MY02 from soil samples showed significant antagonism against 14 plant pathogenic fungi including Fusarium oxysporum f. sp. cucumarinum . Antifungal metabolite(s) SN06 from the culture of the strain MY02 were extracted with n -butanol and purified by silica gel column chromatography. The minimum concentration of SN06 inhibiting any visible fungal growth of F. oxysporum f. sp. cucumarinum is 12.5 μg/ml by twofold serial dilutions method. The mycelia of F. oxysporum f. sp. cucumarinum treated with SN06 were observed under the normal optics microscope. The results showed that some cells of hyphae began to dilate and formed some strings of beads. The cytoplasm oozed out of the cells with the culture time and so most of the cells became empty. The hyphae broke into many segments and then collapsed after 48 h. After inoculated in potato dextrose medium for 48 h, the filtrate of mycelia treated with 1% NaCl containing 12.5 μg/ml SN06 was scanned using ultraviolet spectrophotometer and absorption peak at 260 nm showed that the mycelia cell membrane of F. oxysporum f. sp. cucumarinum was broken and that nucleic acid oozed out of the cell.