两种方法形成菌丝球固定2-氯酚降解菌的对比

【目的】系统研究吸附法和同时培养法对所形成混合菌丝球的外观形态、内部结构及其去除2-氯酚效果的影响。【方法】采用吸附法和同时培养法将可降解2-氯酚的光合细菌PSB-1D固定在黄孢原毛平革菌(Phanerochaete chrysosporium)DH-1发酵而成的菌丝球上,形成混合菌丝球。以单一菌丝球为对照,利用光学显微镜、扫描电镜等仪器观察混合菌丝球的外观形态和内部结构,考察2种方法对混合菌丝球成球效果的影响;以无菌培养液为空白对照,考察游离光合细菌、单一菌丝球、2种方法形成混合菌丝球对2-氯酚的降解效能。【结果】在吸附法形成的混合菌丝球上,光合细菌主要集中在过渡区;而同时培养法将光合细菌牢固地包埋在菌丝球内核区,并大量簇状附着生长在菌丝交联的空隙处和每根菌丝上。在接种等量孢子和光合细菌的前提下,同时培养法较吸附法操作时间更短,成球数量更多,形成菌丝球干湿比更大,单位菌丝干重上固定的细菌数量更多。菌丝球降解体系和游离光合细菌对2-氯酚的降解均符合一级动力学特征。同时培养法形成的混合菌丝球降解效果最好,7 d内对初始浓度为50 mg/L的2-氯酚降解率可达89%以上,降解速率常数为0.3286 mg/(L·d),2-氯酚半衰期t1/2为2.8 d。【结论】首次报道黄孢原毛平革菌包埋固定化光合细菌形成混合菌丝球。该研究为生物质固定化材料的实际应用提供理论依据。     

Yield and size of oyster mushroom grown on rice/wheat straw basal substrate supplemented with cotton seed hull

Oyster mushroom (Pleurotus ostreatus) was cultivated on rice straw basal substrate, wheat straw basal substrate, cotton seed hull basal substrate, and wheat straw or rice straw supplemented with different proportions (15%, 30%, and 45% in rice straw substrate, 20%, 30%, and 40% in wheat straw substrate) of cotton seed hull to find a cost effective substrate. The effect of autoclaved sterilized and non-sterilized substrate on growth and yield of oyster mushroom was also examined. Results indicated that for both sterilized substrate and non-sterilized substrate, oyster mushroom on rice straw and wheat basal substrate have faster mycelial growth rate, comparatively poor surface mycelial density, shorter total colonization period and days from bag opening to primordia formation, lower yield and biological efficiency, lower mushroom weight, longer stipe length and smaller cap diameter than that on cotton seed hull basal substrate. The addition of cotton seed hull to rice straw and wheat straw substrate slowed spawn running, primordial development and fruit body formation. However, increasing the amount of cotton seed hull can increase the uniformity and white of mycelium, yield and biological efficiency, and increase mushroom weight, enlarge cap diameter and shorten stipe length. Compared to the sterilized substrate, the non-sterilized substrate had comparatively higher mycelial growth rate, shorter total colonization period and days from bag opening to primordia formation. However, the non-sterilized substrate did not gave significantly higher mushroom yield and biological efficiency than the sterilized substrate, but some undesirable characteristics, i.e. smaller mushroom cap diameter and relatively long stipe length.     

Use of confocal microscopy to follow the development of penetrative hyphae during growth of Rhizopus oligosporus in an artificial solid-state fermentation system

Two methods were compared for determining the concentration of penetrative biomass during growth of Rhizopus oligosporus on an artificial solid substrate consisting of an inert gel and starch as the sole source of carbon and energy. The first method was based on the use of a hand microtome to make sections of approximately 0.2- to 0.4-mm thickness parallel to the substrate surface and the determination of the glucosamine content in each slice. Use of glucosamine measurements to estimate biomass concentrations was shown to be problematic due to the large variations in glucosamine content with mycelial age. The second method was a novel method based on the use of confocal scanning laser microscopy to estimate the fractional volume occupied by the biomass. Although it is not simple to translate fractional volumes into dry weights of hyphae due to the lack of experimentally determined conversion factors, measurement of the fractional volumes in themselves is useful for characterizing fungal penetration into the substrate. Growth of penetrative biomass in the artificial model substrate showed two forms of growth with an indistinct mass in the region close to the substrate surface and a few hyphae penetrating perpendicularly to the surface in regions further away from the substrate surface. The biomass profiles against depth obtained from the confocal microscopy showed two linear regions on log-linear plots, which are possibly related to different oxygen availability at different depths within the substrate. Confocal microscopy has the potential to be a powerful tool in the investigation of fungal growth mechanisms in solid-state fermentation.     

Respiration of a puff morphological mutant of Schizophyllum commune

Summary The growth kinetics of wild-type mycelium and a puff morphological mutant of Schizophyllum commune revealed greater acid production and slower growth by this mutant. The compact mycelium growth habit of puff in defined liquid medium facilitated manometric studies of cellular respiration during culture aging. Basal oxygen consumption was highest in young, 2-day cultures as was exogenous glucose stimulation while both responses declined rapidly as the mycelial pellets aged. Respiratory stimulation by certain l-amino acids including histidine, arginine and serine was only demonstrated in aged cultures of puff mycelium. A qualitative shift in terminal respiration was considered unlikely because the metabolic poison sodium azide was a potent inhibitor of mycelial oxygen consumption regardless of either the culture age or the respective exogenous substrates employed.     

STUDIES OF THE GROWTH OF PENICILLIUM CHRYSOGENUM IN CONTINUOUS FLOW CULTURE WITH REFERENCE TO PENICILLIN PRODUCTION

SUMMARY: A filamentous mould was cultured by the continuous flow method in which medium is supplied at a constant rate and the culture volume is kept constant. Flow rates up to 0·1 culture volumes/hr were used. The mycelial dry weight concentration and the yield of mycelium/g of carbon source used were equal to or slightly greater than the maximum obtained in batch culture. With glucose concentrations up to 80 g/1. at a flow rate of 0·05 culture volumes/hr, about 45% of the substrate carbon was converted into mycelial carbon and the remainder oxidized to CO₂.
With unlimited amounts of all nutrients available growth of the mould followed the exponential law, as does bacterial growth, and therefore the mould had a constant doubling time.
The oxygen demand of the mould as function of growth rate was determined.
Conditions were found under which the rate of penicillin production/g of mycelium remained at its maximum value for 1000 hr.     

几种真姬菇菌种保藏方法的保藏效果对比

【背景】菌种保藏方法与保藏效果是菌种质量的前提和保证。随着真姬菇Hypsizygus marmoreus栽培规模和产量的逐年递增,选择最佳的菌种保藏方法也越来越迫切。【目的】对真姬菇菌种的常见保藏方法及保藏效果进行分析和研究,寻找简便、高效、廉价的真姬菇菌种保藏方法。【方法】通过对比菌丝生长率、菌丝脱氢酶活性以及菌丝对色度培养基的脱色率等指标参数来评价保藏方法的保藏效果。【结果】通过对三大类27种真姬菇菌种保藏方法 3个月保藏效果的对比分析发现,三大类菌种保藏方法中保藏效果最好的是水溶液保藏法,其中以0.1%PEG6000水溶液保藏效果最佳;其次为木屑保藏法,其中以盐水浸泡过的杨木木屑保藏效果最好;斜面保藏法效果最差。正常保藏实验的效果好于加速保藏实验。【结论】与当前真姬菇菌种保藏的常规方法相对比,本实验所采用的菌种保藏方法大大拓宽了菌种保藏方法的种类与范围,提高了菌种保藏的效果。     

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。     

Mycelial antineoplastic activity of Agaricus blazei

Basidiocarp of Agaricus blazei (=Agaricus brasiliensis; =Agaricus subrufescens) is used as teas or capsules due to its antineoplastic effect but there are few reports of using mycelium for this purpose. The objective of this study was to evaluate the antineoplastic activity on sarcoma 180 cells implanted in mice of two forms of preparation of the mycelium from two A. blazei strains grown in culture medium with different concentrations of isolated soy protein. Mycelia were grown in Pontecorvo medium with different concentrations of isolated soybean protein (ISP). Mycelial hot water extract, moistened mycelial powder, hot water extract of green tea, Ifosfamida^® (ifosfamide drug), and saline solution were administered daily by gavage in mice with sarcoma 180 cells to evaluate antineoplastic activity. It was concluded that antineoplastic activity was the same for both strains, except when used as moistened mycelial powder, which rules out the use of mycelial powder in capsules. Mycelial hot water extract had high antineoplastic activity with lower metabolic demand on the spleen and maintenance of normal blood parameters. Mycelial growth in different ISP concentrations had the same antineoplastic activity. Also the vegetative mycelium was as effective as the basidiocarp for sarcoma 180 tumor inhibition. Green tea was as effective as mycelial hot water extract.     

Energetic efficiency of complex substrate utilization by trichoderma viride

The efficiency of carbon substrate utilization is evaluated as the thermodynamic efficiency (η_x) of microbial growth. Three methods based on mass-energy balance are used for the efficiency studies of complex substrates (straw, plant juices, lye) utilization by microfungi Trichoderma viride.

• 1 According to substrate and biomass balance η_x = 0.55, 0.37 and 0.36 for Trichoderma viride growth on alkali pretreated wheat straw during 23, 34 and 50 hours. Cellulose biodegradation increases with cultivation time. However, the efficiency of cellulose utilization for cell mass growth decreases at the same time.
• 2 In accordance with oxygen-balance calculations η_x = 0.75 and 0.71 for the same processes. The discrepancy in results from the above two methods probably can be explained by the following:
  • A Substrate and biomass balance gives underestimated results.
  • B Oxygen balance method includes the part of energy for extracellular product formation and therefore η_x can be overestimated.
  • C The efficiency of complex soluble substrate utilization (lye, green juice, deproteinized brown plant juice) tested by means of pulse method gives the values of η_x = 0.72 – 0.88. Similar high estimates of η_x in C-limited batch culture are observed for soluble carbohydrates (glucose, galactose, lactose, xylose) but not for acetate. The pulse method is advantageous for testing the “true” efficiency of carbon substrate utilization in a definite physiological environment.

     

三种香蘑属野生食用菌菌株的培养条件优化

【背景】供试菌株分别是分离自山西省宁武县管涔山的肉色香蘑Lepista irina、斑褶香蘑L. panaeolus和山西省蒲县五鹿山的紫丁香蘑L. nuda 3种野生食用菌的子实体。【目的】获得3种野生食用菌的最佳培养条件。【方法】以菌丝生长速度为指标研究不同碳源、氮源、碳氮比、pH和培养温度等各因素对菌丝生长的影响,根据Box-Benhnken中心组合试验设计原理,采用3因素3水平的响应面法确定使菌丝体达到最快生长速度的最佳培养碳源、氮源和pH。【结果】肉色香蘑在葡萄糖20.9 g/L、土豆196.47 g/L、pH 6.0、培养温度21 °C的条件下,菌丝日均生长速度达到最大,为1.13 mm/d;斑褶香蘑在甘露醇17.4 g/L、酵母膏8.1 g/L、B族维生素0.1 g/L、K2HPO4 2.5 g/L、MgSO4 2.5 g/L、pH 7.9、培养温度25 °C的条件下,菌丝日均生长速度达到最大,为0.73 mm/d;紫丁香蘑在土豆200 g/L、可溶性淀粉20.5 g/L、KNO3 2.1 g/L、K2HPO4 2.5 g/L、MgSO4 2.5 g/L、B族维生素0.1 g/L、pH 7.0、培养温度25 °C的条件下,菌丝日均生长速度达到最大,为2.38 mm/d。【结论】获得了3种香蘑属菌株的最佳培养条件,为后续优质野生食用菌的引种驯化积累了相关数据和资源。     

Lipid profile of Pleurotus sajor caju

The lipid profile of Pleurotus sajor caju was studied in relation to mycelial and sporophore growth and different cultural factors. The growth was characterised by lipid synthesis during mycelial growth and utilisation during sporophore growth. The degree of instauration increased during mycelial growth and decreased during sporophore formation. The fatty acid composition of mycelium and sporophore was similar, linoleic acid (C_18:2) being the most dominant acid in both. C:N ratio had a significant (P<0.05) positive effect on mycelial dry weight; however, per cent total lipids was similar. Non-polar lipids became more unsaturated as the temperature was raised from 10° to 25°C and pH from 3.0 to 6.0, but declined when the cultures were aerated. Mycelial dry weight increased significantly (P<0.05) when the liquid medium was supplemented with lipids. In general, fatty acids with carbon chain length C₁₆ and C₁₈ stimulated the growth of mycelium. Supplementation of solid substrate (cotton seed hulls) with safflower oil, soybean oil or rice bran significantly (P<0.05) increased the yield of sporophores. Total lipids and ratio of non-polar to polar lipids were not affected by lipid supplementation.     

Effect of Extracellular Factors on Growth and Dimorphism of Rhizopus oryzae with Multiple Enzyme Synthesizing Ability

Rhizopus oryzae PR7 MTCC 9642 was a dimorphic fungus that showed a regular 90 days cycle of filament (mycelium) to pellet (yeast) transformation through a distinct bottom dwelling intermediate state and the pellets never revert back to filamentous form. Apart from the normal cycle, high temperature (37°C and above) and extreme pH also induced the yeast formation. Among the ions tested, calcium and chloride ions were found to restore the filamentous morphology, even in extreme pH and temperature. Cysteine HCl also played noteworthy role in maintaining mycelial growth even at adverse condition. Immobilized spores showed the appearance of intermediate form instead of typical yeast form even at high temperature. The strain could produce a number of extracellular hydrolytic enzymes like cellulolytic, xylanolytic, pectinolytic and amylolytic enzymes. The pellet and mycelial forms were found to be a better producer of cellulase–lignocellulase enzymes and amylolytic enzymes respectively, which might be correlated with their infectivity. Increase in inoculum size, agitation during cultivation, change in carbon and nitrogen source failed to induce mycelial growth in extreme conditions, which might be explained as irreversible change of configuration of protein responsible for mycelial development.     

Formation and release of β-glucosidase by aspergillus niger zimet 43 746 in correlation to process operations

The total formation of β-glucosidase by the wild strain of Aspergillus niger ZIMET 43 746 is non-growth-associated. In discontinuous culture the total β-glucosidase activity related to the mycelium is incresing with the age of the mycelium. The complete release of the remaining mycelial-associated β-glucosidase is dependent on the structure of the mycelium. In the cases of the mycelium forms pellets throughout the growth phase than the release of β-glucosidase is accelerated compared to the release from loosly branched mycelium. Incresing shear stress caused by increasing of the impeller speed promotes the formation of pellets.     

Enzymes of the xylotrophic basidiomycete Lentinus edodes F-249 in the course of morphogenesis

It has been shown that the fungus Lentinus edodes grown on a solid wort agar substrate produces intracellular enzymes, including Mn-dependent peroxidase, laccase, and tyrosinase as a family of isoforms. The composition of the complex (containing one to four forms of each enzyme) varied during the basidiomycete life cycle. The activity of oxidases was maximal at the stage of nonpigmented mycelium and at the stages of a brown mycelial mat and a fruit body. The activity of tyrosinase increased in the course of mycelium pigmentation and had two maxima: at the stage of a brown mycelial mat and at the stage of a fruit body. Laccase and tyrosinase activities were shown to increase sharply upon addition of oak sawdust extract to the culture medium as compared with the enzyme activities of mycelium grown on wort agar alone. It was established that the effect of phenol oxidase substrates on the growing mycelium consists in a twofold acceleration of the process of morphogenesis in the fungus L. edodes.     

Glutamine requirement for aerial mycelium growth in Neurospora crassa

Five amino acids are accumulated during vegetative growth of Neurospora crassa, particularly.during the prestationary growth phase. Alanine, glutamine, glutamate, arginine and ornithine.comprised over 80% of the total amino acid pool in the mycelium. Amino acid pools of different amino acid auxotrophs were followed during the partial transformation of a mycelial mat into an aerial mycelium. The mycelial mat under starvation and in direct contact with air rapidly formed aerial mycelium, which produced thereafter a burst of conidia. During this process,glutamine and alanine in the mycelial mat were consumed more rapidly than other amino acids;in the growing aerial mycelium, glutamate and glutamine were particularly accumulated. Of the amino acids that were initially accumulated in the mycelial mat, only a high glutamine pool was required for aerial mycelium growth induced by starvation. This requirement for glutamine could not be satisfied by a mixture of the amino compounds that are synthesized via glutamine amidotransferase reactions. It is proposed that glutamine serves as a nitrogen carrier from the mycelial mat to the growing aerial mycelium.     

A comparison of the development and metabolic activity of mycorrhizas formed by arbuscular mycorrhizal fungi from different genera on two tropical forage legumes

 Two glasshouse experiments were done to assess the development and metabolic activity of mycorrhizas formed by isolates of arbuscular mycorrhizal fungi (AMF) from three different genera, Acaulospora, Gigaspora and Glomus on either Pueraria phaseoloides L. or Desmodium ovalifolium L. plants. The second of the two experiments included three levels of a localised phosphate source in the pots. Alkaline phosphatase (ALP), stained histochemically in the intra-radical mycelium (IRM) of AMF over sequential harvests, did not provide a direct marker for the efficiency of AMF in mobilising phosphorus (P) for plant growth and development. The ability of the extra-radical mycelium (ERM) to scavenge a localised phosphate source, determined by its extraction from buried 35-μm mesh pouches, was dependent on the species of AMF tested. This work indicates that AMF from different genera have unique patterns of mycelial development when forming mycorrhizas with tropical hosts in the presence of a localised phosphate source. AMF also appear to have different mechanisms for the control of P transfer, within the mycelium, to the host. The significance of the architecture of the ERM is discussed as well as the localisation of ALP in the IRM in determining the efficiency of AMF in terms of P accumulation in planta and subsequent growth of plants. Accepted: 19 August 1998     

Chemical Composition of the Edible Mushroom Pleurotus ostreatus Produced by Fermentation

Pleurotus ostreatus `Florida' was grown in submerged liquid culture. The biomass yield of the fungus, grown for 3 days in 2-liter fermentors, where the mycelial pellets measuring 5 mm in diameter were formed, was 11.7 g (dry weight)/liter. Comparing the chemical constituents of fruiting bodies produced on cotton straw and mycelial pellets revealed several similarities in total nitrogen, protein, glycogen, fatty acids, RNA, and ash content. Differences were observed in the contents of six amino acids. Although the total fatty acid content was similar, there were more saturated fatty acids in the mycelium. Cell wall composition, typical for basidiomycetes, was observed in both mycelium and fruiting bodies, with laminarin as the main polymer.     

Growth and mycelial strand production of Rigidoporus lignosus with various nitrogen and carbon sources

Growth and differentiation of mycelial strands in Rigidoporus lignosus have been shown to depend on suitable combinations of the pH of the media and the nature of the nitrogen and carbon sources. Amino acids as sole nitrogen sources gave rise to vegetative mycelium. At pH 4.5, growth and mycelial strand differentiation required asparagine, as the fungus failed to grow in the absence of this amino acid. However, at pH 6, differentiation of strands occurred appreciably in asparagine-deficient media, suggesting a close balance between pH and amino acid requirements. Ammonium was required for strand differentiation, while nitrate, as a sole nitrogen source, maintained the fungus undifferentiated. Of the carbohydrates tested, only glucose, fructose and mannose supported strand differentiation. Starch was found to be particularly effective in promoting growth of vegetative mycelium. Strand differentiation required more specific conditions than growth of the vegetative mycelium.     

Influence of humidity on production of pelleted fungal inoculum

One of the practical problems in scaling-up the production of fungal inocula for environmental applications is how to provide essential humidity for fungal growth. Pelleted solid substrate was used as a fungal biomass carrier. It was coated with alginate or agar hydrogels that contained mycelial fragments of the white-rot fungi Trametes versicolor or Irpex lacteus. To follow fungal growth and formation of mycelial coat over pelleted substrate, the fluorescein-diacetate hydrolysing activity (FDA) assay and visual inspection were used. Both fungi were able to overgrow the pelleted substrate in 5–6 days, at a relative humidity (RH) of 86.3% or higher. The enrichment of alginate hydrogel with nutrients or coating of pelleted substrate with more hydrophilic agar hydrogel enabled I. lacteus to overgrow the pellets at a lower RH of 83.6%. Fungal inocula produced at lower RH possessed lower final moisture contents and had greater mechanical strength. Conditioning of T. versicolor mycelial fragments, by a 3-h incubation in fresh growth medium, enhanced fungal growth over the pelleted substrate. A mathematical model was used to simulate and to explain moisture distribution in a hydrogel-coated pellet and the formation of mycelial coat, for various conditions of fungal inocula production.