Relationships between fungal uptake of ammonium, fungal growth and nitrogen availability in ectomycorrhizal Pinus sylvestris seedlings

 Nitrogen deposition and intentional forest fertilisation with nitrogen are known to affect the species composition of ectomycorrhizal fungal communities. To learn more about the mechanisms responsible for these effects, the relations between fungal growth, nitrogen uptake and nitrogen availability were studied in ectomycorrhizal fungi in axenic cultures and in symbiosis with pine seedlings. Effects of different levels of inorganic nitrogen (NH₄) on the mycelial growth of four isolates of Paxillus involutus and two isolates of Suillus bovinus were assessed. With pine seedlings, fungal uptake of ¹⁵N-labelled NH₄ was studied in short-term incubation experiments (72 h) in microcosms and in long-term incubation experiments (3 months) in pot cultures. For P. involutus growing in symbiosis with pine seedlings, isolates with higher NH₄ uptake were affected more negatively at high levels of nitrogen availability than isolates with lower uptake. More NH₄ was allocated to shoots of seedlings colonised by a high-uptake isolate, indicating transfer of a larger fraction of assimilated NH₄ to the host than with isolates showing lower NH₄ uptake rates. Thus low rates of N uptake and N transfer to the host may enable EM fungi avoid stress induced by elevated levels of nitrogen. Seedlings colonised by S. bovinus transferred a larger fraction of the ¹⁵N label to the shoots than seedlings colonised by P. involutus. Seedling shoot growth probably constituted a greater carbon sink in pot cultures than in microcosms, since the mycelial growth of P. involutus was more sensitive to high NH₄ in pots. There was no homology in mycelial growth rate between pure culture and growth in symbiosis, but N uptake in pure culture corresponded to that during growth in symbiosis. No relationship was found between deposition of antropogenic nitrogen at the sites of origin of the P. involutus isolates and their mycelial growth or uptake of inorganic nitrogen. Accepted: 18 September 1998     

Production of Polyclonal and Monoclonal Antibodies Against Hyphae from Arbuscular Mycorrhizal Fungi

Abstract

Polyclonal and monoclonal antibodies were produced against hyphae of the arbuscular mycorrhizal fungus Glomus monosporum. The polyclonal antibodies (pAbs) were raised in a rabbit by immunizing with hyphae. They were tested for their specificity by a dot-immunoblot assay (DIBA). After the third immunization, a distinct difference in the signal strength was observed between the antisera and the preimmune serum. The pAbs showed cross-reactions to a number of fungal species, both mycorrhizal and other. For the production of monoclonal antibodies (mAbs), mice were immunized intraperitoneally with hyphae. The resulting hybridoma cell culture supernatants were tested by an indirect immunolabeling procedure. For this purpose the hyphae were immobilized on silane-coated microscopic slides. The mAb 8A7 reacted with hyphae from all Glomus isolates tested so far. Cross-reactivities were not observed with hyphae from fungi of the family Acaulosporaceae, phytopathogenic fungi tested so far, or from spores from Glomus species.     

Mycelium growth stimulation of the desert truffle Terfezia claveryi chatin by β‐cyclodextrin

The commercial value of Terfezia claveryi, an edible desert truffle with important gastronomic, nutritional, and antioxidant properties, has led to growing interest in its cultivation. The erratic and slow growth of T. claveryi mycelium in vitro represents an impairment to obtain mycorrhizal plants, and it makes necessary to find a new culture medium able to overcome these drawbacks. In this work, we analyze the effect of cyclodextrins (CDs) on the growth of T. claveryi mycelium. Different parameters, including colony diameter, growth rate, and colony fresh weight, were evaluated, both in the presence and absence of these encapsulant agents. The results obtained confirm the ability of CDs to stimulate the growth of T. claveryi mycelium when present in the culture medium. A similar effect was observed when CDs were added to the culture medium of Tuber melanosporum. Three natural (α‐, β‐, and γ) and two modified (hydroxypropil‐β and methyl‐β) CDs were assayed. The best results were obtained with β‐cyclodextrin, but no improvement was observed with its chemically modified derivatives. CDs complex the different compounds present in the culture medium which impair mycelial growth. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1558–1564, 2013     

磁场对金针菇菌丝生长的影响及磁受体和铁离子转运蛋白的鉴定表达

磁场作为一种无法避免的自然因子,对生物的生长发育有重要影响。然而,目前磁场对大型真菌影响的研究还鲜有报道。本研究采用铷磁铁对金针菇菌丝进行处理,检测金针菇菌丝在不同磁极刺激下的生长和基因表达规律。结果显示：磁场是铷磁铁抑制金针菇菌丝生长的主要原因,且在磁感应强度0.003T以上对菌丝生长的抑制作用较强。通过同源比对获得一个金针菇磁受体和一个铁离子转运蛋白的编码基因,分别命名为ff-magr和ff-fief。定量PCR结果显示ff-magr在N极和S极均出现显著下调表达,N极和S极磁场对ff-fief 的表达均无显著影响。结果表明：磁受体蛋白参与了金针菇菌丝受磁场抑制的应答过程,而磁受体不仅仅是通过调控铁的含量来调控菌丝的生长发育。以上结果为合理利用磁场调控真菌生长发育奠定基础,同时也为深入研究磁场对真菌生长发育的分子机理提供参考。     

Structural characterization and molecular identification of arbuscular mycorrhiza morphotypes of <Emphasis Type="Italic">Alzatea verticillata</Emphasis> (Alzateaceae), a prominent tree in the tropical mountain rain forest of South Ecuador

The vast majority of the highly diverse trees in the tropical mountain rain forest of South Ecuador form arbuscular mycorrhizas, and previous molecular investigations revealed a high diversity of fungi. In this study, we present a first trial to link fungal DNA-sequences with defined morphotypes characterized on the basis of partly new mycelial features obtained from field material of one tree species, Alzatea verticillata. Fine roots were halved lengthwise to study the mycelium anatomy on one half and to obtain fungal nuclear rDNA coding for the small subunit rRNA of Glomeromycota from the other half. Light microscopy revealed conspicuously large amounts of mycelium attaching to the surface of the rootlets. The mycelium formed fine- or large-branched appressoria-like plates, vesicles of regular or irregular shape, and very fine, multibranched structures ensheathed by septate hyphae. These previously undescribed features of the supraradical mycelia combined with intraradical mycelium structures were used for distinguishing of four main morphogroups and subordinate 14 morphotypes. DNA sequences of Glomus group A, Acaulospora and Gigaspora, were obtained and linked to three morphogroups. Two sequence types within Glomus group A could be tentatively associated to subordinate morphotypes. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Inhibitory effects of ethyl acetate extract of Cordyceps sinensis mycelium on various cancer cells in culture and B16 melanoma in C57BL/6 mice

The cultivated mycelium of a Cordyceps sinensis (Cs) fungus was sequentially extracted by petroleum ether (PE), ethyl acetate (EtOAc), ethanol (EtOH) and hot water. All solvent extracts except hot water extract showed a significant and dose-dependent inhibitory effect on the proliferation of four cancer cell lines, MCF-7 breast cancer, B16 mouse melanoma, HL-60 human premyelocytic leukemia and HepG2 human hepatocellular carcinoma, with IC(50) values below 132 microg/ml. The EtOAc extract, in particular, had the most potent effect against all four cancer cell lines, with IC(50) between 12 microg/ml (on B16) and 45 microg/ml (on MCF-7). In contrast, it had much lower cytotoxicity against normal mouse bone marrow cells. The EtOAc extract contained carbohydrates, adenosine, ergosterol and trace amount of cordycepin, of which ergosterol and related compounds were identified as a major class of active constituents contributing to the in vitro cytotoxicity. In an animal test, the EtOAc extract showed significant inhibiting effect on B16-induced melanoma in C57BL/6 mice, causing about 60% decrease of tumor size over 27 days. Our results suggest that the EtOAc extract of Cs fungal mycelium has strong anti-tumor activity and is a potential source of natural anti-tumor products.     

姬松茸的研究与进展

概述了大型真菌姬松茸的生物学特性、保健功能以及近年来的研究进展,并对其主要功效物质如多糖等成分、姬松茸深层发酵技术的现状和菌丝体应用情况作了介绍。     

Influence of antitumor drugs on free radical concentration in fungal cells during primary and secondary tumor-like growth

The fungus Fusarium bulbigenum var. blasticola distinguished by secondary tumor-like growth (TLG) in aging was taken as a novel test system for antitumor drugs. EPR spectroscopy was used to assess the effect of various drugs introduced into the growth medium on the type and content of free radicals in the primary mycelium and TLGs. The EPR spectra in all cases featured a single slightly asymmetrical line of ΔH ～ 0.4–0.6 mT and g = 2.0036 ± 0.0006, whereby the paramagnetic centers could be assigned to melanin radicals. The content of free radicals was always higher in TLGs than in the primary mycelium, which could naturally be due to more intense metabolism. A number of antitumor drugs (fluorouracil, hydroxyurea, methotrexate, etoposide) completely prevented TLG, with varying effect on free radicals in mycelia. Another group (cyclophosphane, dacarbazine, adriablastine, vinblastine), on the contrary, promoted TLG and increased the free radical content in all fungal cells. The third group (mercaptopurine, thioguanine, pharmorubicin) were weakly suppressive for TLG despite the elevated free radical content therein. Vitamins B₂, B₁₂, C, and PP were stimulatory for TLG, whereas in combination with antitumor drugs they could enhance or attenuate their effects.     

Antiviral activity of Basidiomycete mycelia against influenza type A(serotype H1N1) and herpes simplex virus type 2 in cell culture

In this study, we investigated the in vitro antiviral activity of the mycelia of higher mushrooms against influenza virus type A(serotype H1N1) and herpes simplex virus type 2(HSV-2), strain BH. All 10 investigated mushroom species inhibited the reproduction of influenza virus strain A/FM/1/47(H1N1) in MDCK cells reducing the infectious titer by 2.0–6.0 lg ID50. Four species, Pleurotus ostreatus, Fomes fomentarius, Auriporia aurea, and Trametes versicolor, were also determined to be effective against HSV-2 strain BH in RK-13 cells, with similar levels of inhibition as for influenza. For some of the investigated mushroom species—Pleurotus eryngii, Lyophyllum shimeji, and Flammulina velutipes—this is the first report of an anti-influenza effect. This study also reports the first data on the medicinal properties of A. aurea, including anti-influenza and antiherpetic activities. T. versicolor 353 mycelium was found to have a high therapeutic index(324.67), and may be a promising material for the pharmaceutical industry as an anti-influenza and antiherpetic agent with low toxicity. Mycelia with antiviral activity were obtained in our investigation by bioconversion of agricultural wastes(amaranth flour after CO2 extraction), which would reduce the cost of the final product and solve some ecological problems.     

粗毛栓菌菌丝球非灭菌条件下对12种染料的脱色研究

以新型白腐真菌——粗毛栓菌Trametes gallica为材料,研究了优化后的该菌菌丝球在非灭菌条件下对直接染料、中性染料、三苯甲烷类染料以及蒽醌类染料共12种染料的脱色能力、脱色机制,以及pH、温度、染料初始浓度等参数对该菌菌丝球脱色效果的影响。结果表明,优化条件下制备的粗毛栓菌菌丝球脱色活力良好,4℃下保存20d后仍保持有原脱色活力的95%;活菌丝球比死菌丝球对染料具有更强的耐受性和更好的脱色效果;非灭菌条件下活菌丝球对12种染料的适宜脱色条件为pH 3.0–5.0、25℃、染料浓度为50mg/L、处理36–60h,该条件下粗毛栓菌菌丝球在60h内脱色率均在55%以上,其中粗毛栓菌菌丝球对亚甲基蓝脱色率最高可达到96.40%。紫外可见光谱分析和显微观察结果表明,48h内粗毛栓菌菌丝球在非灭菌条件下对12种染料的脱色是由吸附引起,无二次污染物的产生。粗毛栓菌的这些优良特性显示了其在工业染料废水处理中的广阔应用前景。