蜜环菌菌索多糖的免疫增强作用研究

研究了野生蜜环菌菌索多糖(polysaccharide from the rhizomorph of Armillaria mellea,AMP)对小鼠免疫功能的影响.结果表明对小鼠分别灌胃(ig)剂量为100、200、300mg/kg·d的AMP均能增加小鼠体重、改变免疫器官重量、抵抗环磷酰胺(CY)对小鼠外周血白细胞数量的影响.实验显示灌胃AMP能提高小鼠单核巨噬细胞系统的吞噬功能,增强小鼠迟发型变态反应,促进溶血素的生成.上述结果表明AMP能增强机体免疫功能.     

蜜环菌多糖对小鼠腹腔巨噬细胞免疫功能的影响

从蜜环菌菌索提取的多糖（ＭＨＧ）能在体外显著增强小鼠腹腔巨噬细胞吞噬中性红的作用,并可诱生巨噬细胞产生一氧化氮。在高浓度时,对巨噬细胞分泌ＩＬ－１有一定的作用。     

蜜环菌多糖的生物合成与发酵条件的相关性

培养基的碳、氮源,ＰＨ值直接影响蜜环菌多糖的生物合成,泥炭水解液的添加量占培养基的１０％～３９％（Ｖ／Ｖ）时,对蜜环菌多糖的生物合成有促进作用。液体振范培养时,蜜环菌生长形成球形菌丝体,菌丝生物量３．４５ｇ／Ｌ,菌丝多糖（精糖）得率为０．２８４％,而静置液体培养有利于菌索形成,菌素生物量１４．０５ｇ／Ｌ,菌索多糖（精糖）得率为０．５１５％,胞外多糖的得率分别为２９．４％和８．３％     

蜜环菌多糖对小鼠血清IL-2和TGF-β1水平的影响

目的:观察蜜环茵多糖对小鼠血清中IL-2和TGF-β1水平的干预作用。方法:实验组小鼠腹腔注射(i.p)不同浓度的蛮环菌多糖,阴性对照组给予生理盐水,阳性对照组给予香菇多糖,连续给药14天。给药结束后,用形态学方法测定各组小鼠在植物血凝素(PHA)诱导下淋巴细胞转化率,用ELISA法测定各组小鼠血清中白介素2(IL-2)、转化生长因子β1(TGF-β1)水平。结果:蜜环菌多糖具有提高小鼠淋巴细胞转化率,增加血清中IL-2水平,并同时下调血清中TGF-β1水平的作用。结论:蜜环菌多糖可通过调节小鼠免疫细胞及免疫分子的作用,增强小鼠的免疫功能。     

蜜环菌胞外多糖的发酵条件研究

蜜环菌是天麻的共生菌 ,有研究证明 ,蜜环菌的菌丝及发酵液有与天麻相类似的药理作用 ,为此 ,我们进行了蜜环菌深层发酵产胞外多糖条件的研究。研究结果表明 :菌丝生长和多糖产生的最适初始pH为 5.0左右 ;消泡剂用量在一定范围内增加 ,有利于菌丝的生长 ,但增加消泡剂用量会不利于多糖的产生 ;接种量大 ,菌丝得率高 ,但接种量为1 0 %时的多糖得率最高 ;装液量试验证明 ,通气量大 ,有利于菌丝的生长和多糖的产生 ;机械搅拌对菌丝的生长和多糖的产生都不利 ;生长动态试验证明 ,蜜环菌可在 6d内完成液体     

中国北方地区蜜环菌生物种的初步研究

对采自北京东灵山地区,黑龙江带岭地区,以及帽儿山地区不同寄主上的１５个蜜环菌子实体进行了极性测试和生物种测定。     

蜜环菌饮料对机体活性的调节作用研究

为了研究蜜环菌饮料对小鼠机体功能的影响,采用环磷酰胺(CY)造模法制作小鼠免疫抑制模型,通过灌胃不同剂量蜜环菌饮料,研究对小鼠机体免疫功能的影响,同时通过测定实验小鼠肠道中细菌总数、大肠杆菌数、乳酸菌和双歧杆菌数等,分析不同剂量的蜜环菌饮料对肠道菌相的影响,最后对小鼠进行了急性毒性试验研究,以检测饮料有无毒副作用。结果表明蜜环菌饮料可明显提高小鼠脾脏和胸腺指数,促进脾脏及胸腺的发育,促进巨噬细胞的吞噬活性,明显增强小鼠的免疫功能,同时还能促进小鼠肠道有益菌的生长,改善肠道微生物区系环境,饮料没有对小鼠产生任何的毒副作用。证明蜜环菌饮料不仅增强机体免疫功能,同时还增强肠道功能,保持和促进机体健康。     

中国蜜环菌和新生物种

在调查研究蜜环菌生物种的过程中,先后在东北、华北、华中和西南地区,发现了四个新的中国蜜环菌的生物种,ＣＢＳ Ｆ、ＣＢＳ Ｇ、ＣＢＳ Ｈ和ＣＢＳ Ｉ。研究表明,ＣＢＳ Ｇ属于典型的同宗配合种,其余三个为异宗配合。ＣＢＳ Ｈ与ＣＢＳ Ｃ之间存在部分互交可育,比例高达１７．８％,在同一个国家的两个种之间出现如此高的互交可育比例还是首次报道。上述三个生物种同欧美的已知蜜环菌种均互交不育,为亚洲特有种。ＣＢ     

Optimization of exopolysaccharide production from Armillaria mellea in submerged cultures

Aims: To study the optimization of submerged culture conditions for exopolysaccharide (EPS) production by Armillaria mellea in shake‐flask cultures and also to evaluate the performance of an optimized culture medium in a 5‐l stirred tank fermenter. Methods and Results: Shake flask cultures for EPS optimal nutritional production contained having the following composition (in g l^?1): glucose 40, yeast extract 3, KH₂PO₄ 4 and MgSO₄ 2 at an optimal temperature of 22°C and an initial of pH 4·0. The optimal culture medium was then cultivated in a 5‐l stirred tank fermenter at 1 vvm (volume of aeration per volume of bioreactor per min) aeration rate, 150 rev min^?1 agitation speed, controlled pH 4·0 and 22°C. In the optimal culture medium, the maximum EPS production in a 5‐l stirred tank fermenter was 588 mg l^?1, c. twice as great as that in the basal medium. The maximum productivity for EPS (Q_p) and product yield (Y_P/S) were 42·02 mg l^?1 d^?1 and 26·89 mg g^?1, respectively. Conclusions: The optimal culture conditions we proposed in this study enhanced the EPS production of A. mellea from submerged cultures. Significance and Impact of the Study: The optimal culturing conditions we have found will be a suitable starting point for a scale‐up of the fermentation process, helping to develop the production of related medicines and health foods from A. mellea.     

琼脂固体培养基培养蜜环菌菌索总DNA的提取

野外采集的蜜环菌[Armillaria mellea(Vahl.ex Fr.)Quel]在提取DNA前需要分离获得纯化的菌丝体。常规液体培养获得菌丝团的方法感杂率较高,采集固体培养基表面cellophane膜上形成的菌丝则难以获得足量的DNA提取材料。蜜环菌细胞内含有大量多醣类物质,也使得蜜环菌高质量DNA的提取存在一定的困难。本研究通过改进试验,提供一个直接从琼脂固体培养基培养的蜜环菌菌索中提取高质量DNA的方法。其中样品的预先冻融处理方法可以促使蜜环菌菌索与琼脂分离;而在裂解提取缓冲液裂解材料细胞后加入1.25 mol/L KAc溶液,则有利于除去蜜环菌细胞内的多醣类物质以及残留的少量琼脂。通过琼脂糖电泳、紫外分光光度计对DNA浓度及OD值的测定、ISSR引物的PCR扩增以及酶切产物的PCR扩增等方法的检测,结果均表明该方法提取的DNA质量较好,符合进一步进行分子生物学研究的要求。     

野生和人工培养的蜜环菌菌索营养成分比较

对野生和人工培养的蜜环菌菌索主要营养成分及氨基酸含量进行了测定和比较。结果表明：人工培养的蜜环菌菌索蛋白质、脂肪、多糖含量均高于野生菌索,二者的氨基酸组成相似,但人工培养的菌索氨基酸总量高于野生菌索。     

蜜环菌粉正己烷提取物抗神经炎症研究

蜜环菌是一种药食兼用真菌,其提取物有多种药用价值,本文通过动物体内实验对其抗神经炎症作用进行探究。利用1-甲基-4-苯基-1,2,3,6-四氢吡啶（MPTP）建立了帕金森病小鼠模型,设置空白组、模型组、蜜环菌粉正己烷提取物高低剂量组（10mg/kg、20mg/kg）,连续灌胃给药12d,对小鼠体重、行为学指标、纹状体TH蛋白、Iba-1蛋白、iNOS酶活及相关mRNA表达进行探究。结果发现蜜环菌粉正己烷提取物组能缓解MPTP造模引起的体重减少,对小鼠行为迟缓有显著性改善,增加纹状体TH蛋白表达,抑制Iba-1蛋白过表达,同时对纹状体内炎症因子iNOS及相关mRNA表达有明显的抑制作用。因此,蜜环菌粉正己烷提取物有良好的抗神经炎症活性,可能是通过抑制小胶质细胞的过度激活,抑制iNOS酶活等途径发挥作用。     

蜜环菌提取物对佐剂性关节炎大鼠的治疗作用

本文通过对佐剂性关节炎大鼠原发性和继发性足肿胀度的测量、全身关节炎指数评分、体重及体态变化、免疫脏器指数、炎性踝关节切片的形态以及血清中肿瘤坏死因子α、白细胞介素1β、白细胞介素6、前列腺素E2、一氧化氮和一氧化氮合成酶等指标来考察蜜环菌Armillaria mellea的5个不同提取物对佐剂性关节炎大鼠的影响及作用机理。结果表明,蜜环菌甲醇提取物对原发性和继发性足肿胀有明显的抑制;甲醇组能明显抑制佐剂性关节炎大鼠脾脏和胸腺萎缩,并能抑制血清中肿瘤坏死因子α、白细胞介素1β、白细胞介素6和前列腺素E2的分泌,抑制大鼠踝关节炎细胞的形成。因此蜜环菌提取物中甲醇提取物对佐剂性关节炎大鼠治疗效果最显著。     

Role of Lysosomal Vesicles in the Digestive Process of Armillaria mellea by the Large Cells of Gastrodia elata

The hyphae of Armillaria mellea Fr. invade the large ceils of Gastrodia elata BI. Through the wall pits of cortical cells. During early stage the plasmalemma of large cell invaginates and the cell wall forms papillary thickenings to restrain the hyphae from invading. When a hypha enters a large cell, it is encircled tightly by the invaginated plasmalemma which is surrounded by a large number of vesicles coated by a unit membrane. As these vesicles fusing with their membranes to the plasmalemma and discharging their contents into the space around the hypha, the space lined by the invaginated plasmalemma enlarges gradually and becomes a digestive vacuole in which a hypha is completely digested. Reaction product form acid phosphatase activities in the vesicles and digestive vacuoles testifies that the vesicles and digestive vacuoles are identical with primary and secondary lysosomes of plant lysosomal system respectively.     

Nutrient Source of Sclerotia of Grifola umbellata and Its Relationship to Armillaria mellea

Ways nutrient-uptake of sclerotia of Grifola umbellata and the relationship between G. umbellata and Armillaria mellea were studied. At the primary stage of sclerotia of G. umbellate infected by A. mellea, the hyphae of G. umbellata could obtain nutrients by invading the one- to three-layer-cells of A. mellea cortex which existed in the sclerotia of G. umbellata, ar late stage of A. mellea infection, the nutrient source of sclerotia of G. umbellata mainly depended on its hyphae, adhering on the intercellular space of A. mellea, to suck the metabolic products of A. mellea. After being nourished the hyphae of sclerotia of G. umbellata outside of the rhizomorph of A. mellea began to reproduce, as their nuclear divisions were well observed. The results suggested that the mutual assimilation between G. umbellata and A. meIlea could be defined as a form of special Symbiotic relation.     

The plant antifungal isoflavone genistein is metabolized by Armillaria mellea Vahl to give non-fungitoxic products

Abstract

Armillaria mellea, the causal agent of root rot, is a fungal pathogen which proved able to convert the leguminous plant antifungal compound 4′,5,7-trihydroxyisoflavone (genistein) into intermediate metabolites. After suitable periods of incubation, the metabolites were extracted and concentrated from liquid culture media, containing both the isoflavone and the fungus. After purification by column chromatography, the molecular structure of the metabolites was determined by means of mass spectrometry and nuclear magnetic resonance analyses. Five different compounds were identified: 1,3,5-trihydroxybenzene, 4-hydroxyphenylacetic acid, 2,5-dihydroxyphenylacetic acid (homogentisic acid), its lactone 5-hydroxy-2(3H)-benzofuranone, and 1,4-benzoquinone. In vitro experiments showed that while the starting compound, i.e. genistein, has some activity in inhibiting the growth of the fungal pathogen, the degradation products are devoid of any appreciable fungitoxic activity. Moreover, results show that the isoflavone metabolites can be, at least partially, utilized by A. mellea as a carbon source.     

蜜环菌菌种分离新法——天麻组织分离法

报道了一蜜环菌菌种分离方法——天麻组织分离法,并对此法与常用分离法——菌索分离法进行比较试验。结果发现,天麻组织分离法分离成功率高（78％）,远高于常用的菌索分离法（16％）,且前者操作简便、难度低,所得菌种生活活力、生长形态均优于后法。