蛹虫草表型多态性对子实体产生及虫草菌素的影响

为选育高产子实体和虫草菌素的蛹虫草菌株,从野生蛹虫草中分离单孢子并进行分型,之后进行产子实体实验;同时对蛹虫草子实体进行了产孢结构的观察,并用高效液相色谱仪检测了子实体和培养基中的虫草菌素。结果表明,菌落背面颜色为橙铬色的菌株容易产生子实体;出发(原代)菌株所产子实体在形态上更接近野生蛹虫草,而角变株的子实体畸形;出发菌株产子实体能力要比该菌株未发生角变部分分离株和角变部分分离株都强。在虫草菌素的产量上也以出发菌株为高,表明表型多态性对蛹虫草产子实体和虫草菌素有很大的影响。这对于优势菌种的筛选和生产实践有借鉴意义。     

互补交配型配对诱导蛹虫草有性子实体形成

前期我们获得了优良性状蛹虫草Cordyceps militaris野生菌株W141436,其子实体虫草素含量高达3.72 mg/kg干重,子实体多糖高达6.7 g/100 g干重,但在大规模应用中发现它发生退化。针对蛹虫草人工栽培退化问题,我们提出蛹虫草子实体形成是有性生殖过程,其两种交配型发生交配是蛹虫草子实体形成的必要条件。本文基于交配型基因分子标记研究了优良性状蛹虫草野生菌株W141436的退化机制。具体地,采用单孢子分离的方法对蛹虫草野生分离株W141436的子囊孢子进行分离,得到了72个单孢菌株。进一步采用PCR方法对单孢菌株及亲本菌株进行了交配型基因类型鉴定。在72个单孢菌株中,28株为Mat1-1类型交配基因型,31株为Mat1-2类型交配基因型,13株含有与亲本相同的交配型基因。根据鉴定结果,对2株Mat1-1型(SP28、SP33)和2株Mat1-2型(SP15、SP32)菌株进行了栽培实验。结果表明,形成子实体的栽培用亲本菌株同时含有Mat1-1和Mat1-2两种交配类型基因,即只有含不同交配型基因的菌株具有发育为子实体的能力,而含同种交配型基因的菌株则不能发育为子实体。本研究为防止蛹虫草在大规模种植中退化提供了理论依据。     

氮源对蛹虫草生长及类胡萝卜素产生的影响

【背景】蛹虫草是一种珍稀食药用菌,类胡萝卜素不仅为其重要活性成分,而且影响子实体的外观品相,但是类胡萝卜素产生的影响因素不明。【目的】揭示氮源对蛹虫草生长和类胡萝卜素产生的影响。【方法】测定不同氮源培养基中菌株生长速度、分生孢子产生及类胡萝卜含量,筛选菌株生长的最适氮源,进一步研究不同浓度的氮源对蛹虫草生长及类胡萝卜素产生的影响,并测定不同光照条件下氮源浓度对蛹虫草子实体类胡萝卜素产生的影响。【结果】蛹虫草在不同的氮源培养基中菌落形态和类胡萝卜素产生存在明显差异。麦麸和黄豆粉培养基中菌株生长速度最快,但是菌落稀疏,正面分别呈现荧光黄色和极微弱红色;蛋白胨和酵母提取物培养基中菌落致密,产孢量极显著高于其它氮源培养基(P0.01),菌落正面为橙黄色;甘氨酸和柠檬酸为氮源时完全没有色素产生,其它无机和氨基酸氮源培养基平板背面有微量色素产生。固体和液体静置培养条件下均发现蛋白胨浓度在0-3%的范围内,随着浓度的增加,类胡萝卜素含量增加;子实体栽培中不同的氮源浓度均表现为蓝光光照条件下类胡萝卜素含量显著高于白光;而在白光和蓝光光照条件下均表现为蛋白胨浓度为1%时类胡萝卜素含量最高,分别为2 809.38±386.24μg/g和4 093.75±518.37μg/g。【结论】氮源种类和浓度显著影响蛹虫草类胡萝卜素的产生,蓝光光照和1%蛋白胨浓度为子实体类胡萝卜素产生的最佳条件,这为栽培富含类胡萝卜素的蛹虫草子实体提供了试验依据。     

蛹虫草优良菌株的筛选

通过对5株蛹虫草的菌丝形态和生长速率、液体培养生物量和胞外多糖、人工栽培和子实体中活性物质虫草多糖和虫草素的对比研究,筛选出优良的蛹虫草菌株。试验结果表明：蛹虫草6号菌株的菌丝的生长速率比其他菌株快;液体培养生物量和胞外多糖含量明显高于其他菌株;人工栽培的蛹虫草子实体头部大,子囊壳丰富,颜色橘黄,出草整齐均匀,出草率高,子实体中虫草多糖和虫草素的含量均高于其他菌株,表明6号菌株具有较高的经济价值,是值得开发和推广的好品种。     

我国东北及山东部分地区野生蛹虫草资源采集及观察

蛹虫草是一种重要的食药用真菌,其子座可规模化栽培。然而,栽培中菌种极易退化,成为限制其发展的重要因素,野生种质资源为蛹虫草栽培生产菌株的重要来源。本研究对东北和山东部分地区蛹虫草资源进行了持续调查,对其生态分布和宏观、微观形态等进行了系统研究。从5个地点共采集野生蛹虫草标本414份,寄主绝大多数为鳞翅目昆虫的蛹,少数为鳞翅目昆虫的幼虫和茧。野生蛹虫草子座单生或2-25根,长1-17 cm,棒状、扁平状或不规则畸形,扁平状子座常具纵沟,部分可分支;子座可从寄主头、胸和腹部各部位长出,其中以头部为主。子座地上部分长1-6 cm,颜色呈深橙黄色。可育部位长0.5-4 cm,宽1.2-6 mm,大多数与不育柄部分界明显;地下部分0.5-11 cm,其长度与腐殖质厚度相关。首次明确观察到蛹虫草菌索,菌索连接蛹体和子座,或单从子座、蛹体上长出,但不是所有野生蛹虫草都有菌索。蛹虫草菌索与子座内部的疏丝组织菌丝形态各异,菌索菌丝中间膨大。人工栽培与野外采集的子座宏观形态差异明显,但子囊和子囊孢子形态无明显差异。通过野外采集和调研,获得了大量的野生蛹虫草种质资源,为解决蛹虫草种业问题奠定基础。     

铬离子对蛹虫草子实体生物量及化学成分的影响

蛹虫草是一种具有多种生物功能的药用真菌,近年来在金属离子富集方面受到学者的广泛关注,但对蛹虫草富集Cr³⁺的能力及其相应的生理反应尚无文献报道。为此,本研究利用含有不同Cr³⁺浓度的燕麦培养基培养蛹虫草子实体,分别对蛹虫草子实体生物量及子实体中虫草素、腺苷、虫草酸、麦角甾醇、铬离子含量等指标进行了测定。结果表明,除麦角甾醇随着培养基中Cr³⁺浓度的增加而降低之外,其余指标均呈现先增加后降低的变化趋势。700 mg/L Cr³⁺处理时,子实体干、鲜重和铬离子含量达到最大值,分别比对照组增加36.85%、35.53%和202.12%;当Cr³⁺达1 500 mg/L时,虫草素和腺苷含量达到最大值,分别比对照组高94.61%和530.29%;虫草酸含量和多糖含量分别在1 100 mg/L和1 200 mg/L Cr³⁺处理时达到最大值,分别比对照组高123.62%和21.39%。本研究首次探讨了蛹虫草子实体富集Cr³⁺的能力以及Cr³⁺处理下的子实体生物量和部分次生代谢产物含量的变化规律,为进一步研究蛹虫草富集铬离子机制提供了理论依据,为富铬蛹虫草功能性食品的研发奠定了基础。     

从天然蛹虫草不同部位分离菌株特性比较

以采集自泰山地区的6株野生蛹虫草为研究对象,分别通过菌核、子座组织分离和子囊孢子分离法共获得18个菌株,经ITS鉴定均为蛹虫草菌株后,进一步对18个菌株进行了菌丝生长速度、菌落形态、出草试验、主要活性物质（虫草素、腺苷、虫草多糖、β-类胡萝卜素、叶黄素）和交配型的测定分析。结果表明：在菌丝长势、出草产量和整齐度方面,从子囊孢子分离得到的菌株普遍优于组织分离菌株;而在活性物质含量方面,从子囊孢子和子座分离得到的菌株普遍优于菌核分离菌株。通过交配型测定发现,从菌核、子座和子囊孢子分离得到的18个菌株均不存在交配型缺失现象。     

蓝光光照对蛹虫草子实体生长和主要活性成分的影响

光作为一种重要的环境因子,可影响真菌的生长发育、生理周期、形态变化及次级代谢产物的产生,对于蛹虫草而言,光照还是其子实体生长发育的必要条件。选择2株不同来源的蛹虫草菌株,研究了自然光照和蓝光光照条件下,其子实体生长、抗氧化酶活性和主要活性成分的变化。结果表明蓝光光照对于蛹虫草子实体产量没有明显的促进和抑制作用。自然光照条件下成熟子实体的超氧化物歧化酶(superoxide dismutase,SOD)活性显著高于蓝光光照,过氧化氢酶(catalase,CAT)活性没有明显差别,而过氧化物酶(peroxidase,POD)活性则表现出菌株差异。子实体主要活性成分方面,腺苷、甘露醇含量不受蓝光影响,类胡萝卜素在蓝光光照条件下含量极显著高于自然光光照,蓝光对子实体、虫草素、粗多糖含量的影响则存在菌株差异。研究为蛹虫草子实体的栽培条件优化,提高蛹虫草子实体质量提供依据。     

2株北虫草菌株系统发育及其活性成分含量分析

对收集的2株北虫草菌株进行形态学观察、分子鉴定及子实体中主要活性成分含量分析,为其应用研究提供技术支持。采用平板培养法、玻片培养法及显微观察对2株北虫草菌株进行形态学观察;基于ITS序列分析对2株实验菌株进行分子鉴定;有效成分的测定均采用高效液相色谱法。结果表明,2株北虫草子实体形态及在PDA加富培养基上菌丝长势、菌落形态差异显著,ITS序列分析结果显示,2株北虫草为蛹虫草,属种内差异;二者腺苷成分相近,但尖头北虫草虫草素的含量是大孢子头北虫草的2倍,且同一品种不同栽培方法及同一种源不同栽培地获得的子实体中两种活性成分的含量有所不同,尤其是腺苷的含量差异明显。以北虫草子实体的品相作为品质优劣的评估标准缺乏科学依据,在子实体应用过程中应根据需要科学选择。     

基于农业废弃物培育优质蛹虫草的研究

利用农业废弃物甘薯藤及蛹虫草培养基废弃物作为培养基的主要原料进行蛹虫草菌种驯化。蛹虫草子实体培养基中添加不同比例蛹虫草培养基废弃物及甘薯藤粒，通过适宜的培养条件，废弃物中淀粉、蛋白质、糖类、氨基酸等营养物质及蛹虫草胞外酶酶解产生的小分子物质被充分利用, 以培育优质蛹虫草子实体。当一级种子中加入蛹虫草培养基废弃物20 g/L和甘薯藤粉10 g/L，二级种中加入蛹虫草培养基废弃物20 g/L、甘薯藤粉10 g/L，使蛹虫草子实体栽培料中蛹虫草培养基废弃物占32%~45%，甘薯藤粒占10%~15%，二者比例为(3～4):1时，栽培效果最好。本研究蛹虫草培养基替代原料资源丰富易得，并可节约生产用粮，降低原料成本，从而实现农用废弃物再利用，减少环境污染，也符合绿色环保可持续发展的理念。     

Isolation of vegetable wasps and plant worms, Cordyceps nutans, from fruit-body tissue

We isolated Cordyceps nutans from the stipe and abdominal tissues of fruit bodies using a surface sterilization method. Hyphal growth was observed in inocula from both the stipe and abdominal tissue. Some strains from discharged ascospores were obtained and colony characteristics were compared to the strains isolated from the tissues. Colonies of isolates from ascospores grew quite slowly. Isolates of 43 from the 52 examined fruit bodies formed colonies similar to those from ascospores. To confirm the success of isolation, we analyzed by PCR-RFLP of the ITS regions of rDNA samples from fruit bodies, isolates from fruit bodies, and isolates from ascospores. All the isolates obtained from stipe and abdominal tissues presented identical patterns. In this study, we report the first successful isolation of C. nutans from fruit-body tissue using a surface sterilization method.     

被毛孢属一新种──长座虫草的无性型*

报道长座虫草 Ｃｏｒｄｙｃｅｐｓ　ｌｏｎｇｉｓｓｉｍａ　 Ｋｏｂａｙａｓｉ的无性型为长座被毛孢新种Ｈｉｒｓｕｔｅｌｌａ ｌｏｎｇｉｓｓｉｍａ Ｓｐ． ｎｏｖ．。用子囊孢子分离得到的无性型与子座柄部粘片所见的被毛孢相同;用分离菌在米饭培养基上人工诱发出的子座与天然子座相似,这表明长座被毛孢为长座虫草的无性型。     

被毛孢属一新种——长座虫草的无性型

报道长座虫草Cordyceps longissima Kobayasi的无性型为长座被毛孢新种Hirsutella longissima sp.nov.。用子囊孢子分离得到的无性型与子座柄部粘片所见的被毛孢相同;用分离菌在米饭培养基上人工诱发出的子座与天然子座相似,这表明长座被毛孢为长座虫草的无性型。     

Phylogenetic background of Orange lily (Lilium bulbiferum s.l.) cultivars from a genetically isolated environment

Domesticated Orange lily ( LILIUM BULBIFERUM s.l.) cultivars do not typically produce seeds, and Orange lily is not native to Finland. Therefore, back crossing of the cultivars with wild species has not been possible. Genetic variability and genuineness of eight Finnish traditionally-grown Orange lily cultivars was studied. RAPD patterns were compared between the cultivars and genuine Orange lily ( LILIUM BULBIFERUM L.), and a related Dauricum group. The results showed partition of tested genotypes into four groups, L. CANADENSE as the outgroup. The cultivars were divided into two subgroups where the trait to form bulbils was characteristic to subgroup I. The cultivated strains differed from each other as much as from the seedling strains, but were genetically closer to genuine Orange lily than the Dauricum group. This indicates that the cultivars are genuine forms of Orange lily species. The special morphological features of the cultivars have likely been formed during centuries-long genetic isolation from natural populations.     

Podospora anserina mutant defective in protoperithecium formation, ascospore germination, and cell regeneration.

A mutant (modx) was selected on the basis of the suppression of self-lysis due to a recessive mutation (modB). modx, a dominant mutation, reduced hyphal branching from nonapical cells, abolished protoperithecium formation, and induced the death of stationary cells only when these were isolated to obtain further development. Mutant ascospores, formed in the fruiting bodies which occasionally occur under specific conditions (32 degrees C on starved medium), showed a delay in the germination process (up to 3 months instead of about 5 h for wild-type ascospores) when submitted to incubation under standard conditions (26 degrees C on germination medium) and failed to germinate at 18 degrees C. Revertants from modx strains, selected on the basis of the suppression of the nonrenewal of growth from stationary cells, were wild type for all the other three defects. Indirect arguments suggested that the modx mutant strain might be defective in the control of a specific class of stable messenger ribonucleic acids which would be essential for the physiology of ascospores and stationary cells.     

稻曲病两个白化菌株的分离与生物学特性

为了筛选带有自然标记的稻曲病菌菌株,2010年从浙江省象山县和陕西省勉县采集和分离到2个稻曲病白化菌株,ZJa0201和SXa0101。它们在PSA培养基上的生长速度约为其他稻曲病菌株的3倍,未见产生厚垣孢子;在PS培养基上只能产生少量分生孢子。rDNA-ITS和rDNA-IGS序列分析表明,两个白化菌株也与稻曲病菌已知所有菌株的ITS序列同源性高于99.6%;rDNA-IGS序列也属于最为常见的类型,含有2个77bp的重复单元序列。由此推断,这两个白化菌株属于稻曲病菌产孢退化的突变体。白化菌株在PSA上     

Mutants of Arabidopsis thaliana Capable of Germination under Saline Conditions

Three mutant strains of Arabidopsis thaliana var Columbia were selected for their ability to germinate in elevated concentrations of NaCl. They were not more tolerant than wild type at subsequent development stages. Wild-type strains could not germinate at concentrations > 125 mM NaCl. Two of mutant strains, RS17 and RS20, could withstand up to 225 mM, whereas RS19 was resistant to 175 mM. The RS mutants could also germinate under even lower osmotic potentials imposed by high concentrations of exogenous mannitol (550 mM), whereas the effects of elevated levels of KCl, K2SO4, and LiCl were similar among the mutants and wild type. Therefore, the mutants are primarily osmotolerant, but they also possess a degree of ionic tolerance for sodium. Sodium and potassium contents of seeds exposed to high salinities indicated that the NaCl-tolerant mutants absorbed more of these respective cations during imbibition. These higher internal concentrations of potassium and sodium could contribute to the osmotic adjustment of the germinating seeds to the low osmotic potential of the external medium. Genetic analysis of F1 and F2 progeny of outcrosses suggest that the salt-tolerant mutations are recessive and that they define three complementation groups.     

Cultivation and Hybridization Alter the Germination Behavior of Native Plants Used in Revegetation and Restoration

Native plants are increasingly used for revegetation and restoration. These plants are cultivated for several generations at plant nurseries and often they are of unknown provenance. Therefore, cultivated plants often differ from their wild progenitors in life‐history traits. Using germination behavior as example, we tested the assumption that cultivated plants have different life‐history traits than their uncultivated progenitors. Cultivated as well as wild individuals of Plantago lanceolata and Lotus corniculatus, two species frequently used in revegetation, were tested in a common garden experiment as well as in incubators for their germination behavior. We observed significantly faster and more abundant germination in cultivated varieties. Using artificial crossings, we found that also hybrids of cultivated varieties and wild relatives germinate faster and more abundant than the wilds. As wild plants acquire their life‐history traits by natural selection, we have to assume that they represent the optimal adaptation to the environmental conditions. If these traits are changed by cultivation or by hybridization between cultivated varieties and local populations, the long‐term survival probabilities of local populations may be altered. Therefore, the use of cultivated varieties of native plants should be avoided in revegetation.