Biodecolourisation of some industrial dyes by white-rot fungi

Eight white-rot fungal strains were screened for biodecolourisation of eight dyes commercially employed in various industries. Decolourisation of Poly R 478 was used as a standard to ascertain the dye-decolourisation potential of various fungi. All the fungi tested significantly decolourised Poly R 478 on solid agar medium. When tested in a nitrogen-limited broth medium, Dichomitus squalens, Irpex flavus, Phlebia spp. and Polyporus sanguineus were better industrial dye decolourisers than Phanerochaete chrysosporium.     

Degrading ability of oligocyclic aromates by <Emphasis Type="Italic">Phanerochaete sordida</Emphasis> selected <Emphasis Type="Italic">via</Emphasis> screening of white rot fungi

Seventy-nine white rot strains were screened to determine if they had the potential for use in the degradation of oligocyclic aromates (PAHs) by measuring their dye-decoloration rate. Fourteen strains that were selected based on their dye-decoloration rate were then evaluated for the ability to tolerate various levels of PAHs spiked in agar medium. The ability of white rot fungi to degrade 3- or 4-ring PAHs (anthracene, phenanthrene, fluoranthene, pyrene) was determined. Two strains of Phanerochaete sordida (KUC8369, KUC8370) were possible PAHs degraders, degrading a significantly greater amount of phenanthrene and fluoranthene than the culture collection strain P. chrysosporium (a known PAHs degrader). The production of manganese peroxidase, the only extracellular ligninolytic enzyme detected during the cultivation, was evaluated.     

Cultivation of Entomopathogenic Fungi for the Search of Antibacterial Compounds

Entomopathogenic fungi are a rich source of natural bioactive compounds. To establish cultivation conditions which facilitate the production of bioactive compounds and to select good genera among entomopathogenic fungi as the producer, 47 typical entomopathogenic fungi were tested for their ability to produce antibiotic activity. Thirty-eight strains (81%) and 30 strains (64%) of these fungi produced either anti-Bacillus compounds or anti-Staphylococcus compounds, respectively, indicating that the majority of the entomopathogenic fungi tested possessed the ability to produce antibacterial compounds. Using 9 representative strains (Aschersonia sp. HF724, Beauveria bassiana HF338, Cordyceps ramosopulvinata HF746, Metarhizium anisopliae HF293, Metarhizium flavoviride HF698, Nomuraea rileyi HF588, Paecilomyces fumosoroseus HF254, Paecilomyces tenuipes HF419, and Verticillium lecanii HF238), the cultivation conditions in liquid medium were surveyed with respect to the cultivation procedure and medium composition, particularly in terms of the presence or absence of insect-derived materials. At 26 °C, M. anisopliae HF293, N. rileyi HF588, and V. lecanii HF238 strains produced clear antibiotic activity against Bacillus and Saccharomyces, but only in the presence of insect-derived materials, suggesting that the production of antibacterial/antifungal compounds by entomopathogenic fungi is triggered by the presence of insect-derived materials.     

Diversity of Physiological and Biochemical Characters of Microdochium Fungi

The biological characterization of Microdochium majus, M. nivale, and M. seminicola strains with wide geographical origins showed the diversity of their pathogenic properties and metabolite compounds, allowing them to exist in their habitats. Significant differences in the ability of Microdochium fungi to cause lesions on wheat and oat leaves were found. The intensity of symptoms depended on the species and substrate origin of the strains. On average M. seminicola strains were able to cause less leaf necrosis than M. majus and M. nivale. The volatile organic compound (VOC) profile of Microdochium fungi included 29 putative fungal metabolites. The spectrum of the identified VOCs in M. seminicola strains was much richer than that in M. majus and M. nivale strains. In addition, the strains of M. seminicola emitted at least six sesquiterpenes. Mycotoxin analysis by HPLC/MS/MS revealed that the analyzed Microdochium strains did not produce any toxic metabolites typically produced by filamentous fungi.     

Screening for Ligninolytic Enzyme Production by Diverse Fungi from Tunisia

Summary This work represents the first report on the ability of autochthonous fungi from Tunisia to produce ligninolytic enzymes. Three hundred and fifteen fungal strains were isolated from different Tunisian biotopes. These fungal strains were firstly screened on solid media containing Poly R-478 or ABTS as indicator compounds that enabled the detection of lignin-modifying enzymes as specific color reactions. Of the 315 tested strains, 49 exhibited significant ABTS-oxidation activity expressed within the first week of incubation and only 18 strains decolorized the Poly R-478. Liquid cultivations and laccase, manganese peroxidase and lignin peroxidase activity assays of positive strains confirmed that eight efficient enzyme producers were found in the screening. These strains were attributed to the most closely related species using PCR amplification and sequencing of the internal transcribed spacer ‘ITS’ regions of the ribosomal DNA. The identification results showed fungal genera such as Oxyporus, Stereum and Trichoderma which have been only rarely reported as ligninolytic enzyme producers in the literature. Culture conditions and medium composition were optimized for the laccase producer Trametes trogii CTM 10156. This optimization resulted in high laccase production, 367 times more than in non-optimized conditions and which reached 110 U ml^-1 within 15 days of incubation.     

Characterization of hemicellulases from thermophilic fungi

The thermophilic fungi Thermomyces lanuginosus, Malbranchea cinnamomea, Myceliophthora fergusii and the thermotolerant Aspergillus terreus were cultivated on various carbon sources, and hemicellulolytic and cellulolytic enzyme profiles were evaluated. All fungi could grow on locust bean galactomannan (LBG), Solka floc, wheat bran and pectin, except T. lanuginosus, which failed to utilize LBG for growth. Different levels of cellulase and hemicellulase activities were produced by these fungal strains. Depending on the carbon source, variable ratios of thermostable hydrolytic enzymes were obtained, which may be useful in various applications. All strains were found to secrete xylanolytic and mannanolytic enzymes. Generally, LBG was the most efficient carbon source to induce mannanase activities, although T. lanuginosus was able to produce mannanase only on wheat bran as a carbon source. Xylanolytic activities were usually highest on wheat bran medium, but in contrast to other investigated fungi, xylanase production by M. fergusii was enhanced on pectin medium. Preliminary thermostability screening indicated that among the investigated species, thermotolerant glycosidases can be found. Some of the accessory activities, including the α-arabinosidase activity, were surprisingly high. The capability of the produced enzymes to improve the hydrolysis of lignocellulosic pretreated substrate was evaluated and revealed potential for these enzymes.     

Penicillium verruculosum WA 30 a new source of cellulase

Summary Of fungi 110 strains were screened for extracellular cellulase production in shake flask experiments. Twelve strains produced the enzyme in significant quantity. Since the enzyme activity was assayed by different methods (liberation of reducing sugar from cotton, filter paper, carboxymethylcellulose and cellobiose), the estimation of the productivity of the strains differed according to the substrate used. The best cotton degrading activity per fermentation volume as well as per mg of secreted soluble protein was achieved by Penicillium verruculosum WA 30, a wild-type strain, for which the cellulase productivity has not yet been published. The cotton degrading (so-called C₁) activity was successfully enhanced nearly threefold in medium experiments. Analyses of saccharification digests showed that glucose was the predominant product, with negligible amounts of cellobiose. The pH and temperature optima for WA 30 cellulase complex were pH 4.2 and 60°C.     

Management of palm oil mill effluent through production of cellulases by filamentous fungi

A laboratory scale study to evaluate the potentiality of filamentous fungi for the production of cellulolytic enzymes using palm oil mill effluent (POME) as a basal medium was initiated. A total of 25 filamentous fungi in which 16 filamentous fungi were isolated and purified from oil palm industrial residues and 9 strains from laboratory stock were screened using POME with 1% total suspended solids. Trichoderma reesei RUT C-30 was identified as a potential strain for cellulolytic enzyme production as compared to other genera of Aspergillus, Penicillum, Rhizopus, Phanerochaete, Trichoderma and basidiomycete groups. The results showed that T. reesei RUT C-30 gave the highest filter paper cellulase and carboxy methyl cellulase activity of 0.917 and 2.51 U/ml respectively at day 5 of fermentation. Other parameters such as growth formation, pH, filterability and total biosolids were observed to evaluate the bioconversion process.     

Bioactivity of intra and extracellular substances from cyanobacteria and lactic acid bacteria on “wood blue stain” fungi

Cyanobacteria (photoautotrophic prokariota) have potential for the control of pathogenic bacteria and fungi. The effect of intra and extracellular products from cyanobacterial strains on the growth of fungi isolated from “wood blue stain,” was tested. Extracellular products were obtained by concentration and sterilization of the culture medium where cyanobacteria were grown. Cyanobacterial substances promoted or inhibited fungal growth according to the fungal and cyanobacterial strains tested. Extracellular products from Nostoc muscorum 79a and the methanolic extract from Microchaete tenera 84b biomass inhibited growth of Sphaeropsis sapinea 2157 (64.7 and 775.6%, respectively). Extracellular products of Nostoc piscinale 59 and biomass methanolic extract from N. muscorum 79a produced the highest growth promotion of Trichoderma boningii 452 (105.0%) and T. viride 993 (136.7%). Extracellular products of the heterotrophic lactic acid bacterium Streptococcus termophilus were also tested and strongly inhibited (64–92%) all the fungal strains. The tested fungi have different sensitivity to the bioactive substances present in the biomass and/or the culture medium of the studied cyanobacteria and lactic acid bacterium. N. muscorum 79a, M. tenera 84b, and S. termophilus have potential to control the wood blue stain fungi by a friendly environmental alternative.     

Simultaneous phosphate solubilization potential and antifungal activity of new fluorescent pseudomonad strains, Pseudomonasaeruginosa, P. plecoglossicida and P. mosselii

Of 80 fluorescent pseudomonad strains screened for phosphate solubilization, three strains (BFPB9, FP12 and FP13) showed the ability to solubilize tri-calcium phosphate (Ca₃(PO₄)₂). During mineral phosphate solubilization, decrease of pH in the culture medium due to the production of organic acids by the strains was observed. These phosphate solubilizing strains produced indole-3-acetic acid (IAA) and protease as well as exhibited a broad-spectrum antifungal activity against phytopathogenic fungi. When tested in PCR using the gene-specific primers, strain BFPB9 showed the presence of hcnBC genes that encode hydrogen cyanide. On the basis of phenotypic traits, 16S rRNA sequence homology and subsequent phylogenetic analysis, strains BFPB9, FP12 and FP13 were designated as Pseudomonas aeruginosa, P. plecoglossicida and P. mosselii, respectively. Present investigation reports the phosphate solubilization potential and biocontrol ability of new strains that belong to P. plecoglossicida and P. mosselii. Because of the innate potential of phosphate solubilization, production of siderophore, IAA, protease, cellulase and HCN strains reported in this study can be used as biofertilizers as well as biocontrol agents.     

Nutritional Requirements of Plasmodium falciparum in Culture. II. Effects of Antimetabolites in a Semi Defined Medium1

A semi-defined minimal medium, in which pantothenic acid is the only vitamin, was used to culture Plasmodium falciparum for the analysis of antimetabolite drugs. Analogs of riboflavin, nicotinamide, pyridoxine, and thiamin inhibited the growth of this parasite; for each drug, effects were much more pronounced after 96 h of exposure compared to 48 h. The most potent drug examined was 8-methylamino-8-desmethyl riboflavin (IC₅₀ value approximately 1.0 times 10^-10 M at 96 h). Avidin, a protein which complexes and thus inactivates biotin, did not affect parasite viability. Other antimalarial drugs, including chloroquine and quinine derivatives and antibiotics, were equipotent in the minimal medium and in RPMI 1640. Four strains of P. falciparum showed only minor differences in sensitivity to these antimetabolites. The use of prolonged drug exposure times and a vitamin-depleted medium allowed the preliminary characterization of antimalarial antimetabolites in vitro.     

Fungal bioleaching of metals in preservative-treated wood

Twenty-four brown-rot and 10 white-rot fungi were screened to evaluate their applicability for detoxification of preservative-treated wood impregnated with copper and chromium (CC) salts. Brown-rot fungi generally showed higher tolerance towards copper inhibition than white-rot fungi. Additionally, brown-rot fungi were found to accumulate considerable quantities of oxalic acid (up to 44.3 mM) in liquid medium, while white-rot fungi generally accumulated only traces of this organic acid. Oxalic acid is a strong organic acid capable of complexing a variety of heavy metals. Four Antrodia vaillantii and two Poria placenta brown-rot strains that displayed both a high copper tolerance and a high oxalic acid production were selected for further study. The brown-rot fungi effectively decayed wood containing up to 4.4% CC causing corrected mass losses of up to 24.3% in 4 weeks. Fungal treatment was also found to promote extensive leaching of chromium (up to 52.4%), but only moderate leaching of copper (15.6% or less). These results indicate the potential of solid-state fermentation with copper-tolerant fungi for the remediation of preservative-treated wood. Improving the solubility of copper will be an important challenge for future research.     

海南西海岸真红树内生放线菌多样性及其延缓衰老活性初筛

为了发掘红树林内生放线菌资源和进行新型海洋药物研究,该文选择海南西海岸14种真红树的根、茎、叶、花、胚轴为研究对象,采用9种不同分离培养基[改良的高氏培养基(AGG)、海藻糖-天冬酰胺培养基(M4)、海藻糖-脯氨酸培养基(M5)、改良ISP5培养基(M7)、精氨酸-天冬酰胺培养基(M9)、改良淀粉-水解酪素培养基(M10)、酪氨酸-天冬酰胺培养基(P7)、燕麦培养基(P3)、棉籽糖-组氨酸培养基(M11)],结合稀释涂布法和三线划线法从真红树组织中分离菌株,基于菌株16S rRNA基因序列信息等分子生物学鉴定方法对获得的海洋放线菌进行多样性分析,并使用模式生物秀丽隐杆线虫对所得内生放线菌的延缓衰老活性进行研究。结果表明:(1)从46份真红树组织中共获得24株放线菌,隶属于7科11属,其中9株菌株为链霉菌属(Streptomyces),且IMDGX 6270、IMDGX 6137和IMDGX 6173为3株疑似潜在新菌株。(2)秀丽隐杆线虫模型筛选发现4株放线菌(IMDGX 6157、IMDGX 6182、IMDGX 6248、IMDGX 6360)具有延缓衰老的活性,与空白组比较,生存时间分别延长17.16%、28.11%、29.05%、27.10%。综上结果说明海南西海岸区域真红树植物中可能含有较丰富的放线菌资源,能够为延缓衰老药物的研发提供新来源。     

Enzymatic characterization of Chilean native wood-rotting fungi for potential use in the bioremediation of polluted environments with chlorophenols

This work presents a preliminary report of a series of studies on the ability of several indigenous wood-rotting fungi from Chile to produce hydrolytic and ligninolytic enzymes and the evaluation of these native microorganism to future research on potential applications in bioremediation programs. Wood-rotting Basidiomycete fungi were collected from indigenous hardwood forest in the South of Chile. Twenty-eight strains were identified and qualitative enzymatic tests for peroxidases, laccase, tyrosinase, xylanase and cellulase production were performed in solid medium. Eleven selected strains were evaluated in liquid medium to quantify their ligninolytic enzyme production and their capacity to grow in solid medium supplemented with 2,4-dichlorophenol (2,4-DCF), 2,4,6-trichlorophenol (2,4,6-TCF) and pentachlorophenol (PCP). PCP degradation and ligninolytic enzymes production were also evaluated in liquid medium. Results showed that laccase was present in 28 of the selected strains (≈73%). Peroxidase was present in 40% and cellulase in 37% of the strains. Xilanase and tyrosinase were obtained in a smaller percentage in the strains (28% and 7%, respectively). The 11 selected strains showed high concentrations of lignin peroxidase (Lip) and manganese peroxidase (MnP). Anthracophyllum discolor (Sp4), produced LiP and MnP at 90.3 and MnP 125.5 U L⁻¹ respectively, compared to the control fungus Phanerochaete chrysosporium CECT-2798 that produced 58.1 and 118.4 U L⁻¹ of LiP and MnP. Tolerance test showed that native Chilean fungi did not present high tolerance to 2,4,6-TCF and PCP but were quite tolerant to 25 and 50 mg L⁻¹ of 2,4-DCF. However, pre-acclimatization in 2,4-DCP notably improved the growth in medium with 2,4,6-TCP and PCP. PCP in liquid medium was efficiently degraded by the fungi Anthracophyllum discolor, Lenzites betulina (Ru-30) and Galerina patagónica (Sp3), and the major MnP activity was produced by A. discolor (Sp4) (67 U L⁻¹).     

Efficacy of entomopathogenic fungi against soil-dwelling life stages of western flower thrips, Frankliniella occidentalis, in plant-growing media

The potential of several entomopathogenic fungi to control soil‐dwelling stages of western flower thrips, Frankliniella occidentalis Pergande (Thysanoptera: Thripidae), was evaluated in various growing media. Two Metarhizium anisopliae (Metsch) Sorokin strains, V275 and ERL700, were the most effective, causing 85–96% mortality of thrips larvae and pupae (as measured by relative adult emergence rates), 11 days after inoculation. Mortality in other M. anisopliae‐treated media ranged from 51–84%; Beauveria bassiana (Balsamo) Vuillemin strains caused 54–84% mortality, and Paecilomyces fumosoroseus (Wize) 63–75% mortality. In contrast, mortality from the insecticide treatment (fipronil 1 g l^?1 medium) was only 15–54%. Metarhizium anisopliae V275 was selected for more in‐depth evaluation under greenhouse conditions. There was no significant difference in M. anisopliae efficacy in growing media whether it was applied as drench (84–93%) or premixed into the medium as dry conidia (85–92%). The use of M. anisopliae with reduced rates of either fipronil or imidacloprid did not significantly improve control. Overall, our study shows that M. anisopliae V275 is robust and offers much promise for the control of soil‐dwelling stages of thrips as part of an integrated pest‐management programme.     

Meira nashicola sp. nov., a novel basidiomycetous, anamorphic yeastlike fungus isolated from Japanese pear fruit with reddish stain

Three undescribed strains of basidiomycetous, anamorphic yeastlike fungi were isolated from Japanese pear fruits with a reddish stain collected in Tottori Prefecture, Japan. The strains are classified in a single group and assigned to the genus Meira by conventional and chemotaxonomic studies. Sequence analyses of the D1/D2 domain of 26S rDNA and internal transcribed spacer (ITS) regions indicate that the strains represent a novel species with a close phylogenetic relationship to Meira geulakonigii and M. argovae. The name Meira nashicola sp. nov. is proposed for the strains (type strain PFS 002 = MAFF 230028 = CBS 117161).     

桃树根部内生真菌ZJ-4的分离鉴定及其对桃褐腐病的抑制效果

自桃树根部组织中分离能够防治桃褐腐病害的内生拮抗真菌,并从细胞学水平对其抑制机理进行了初步的探究。采用两点对峙法筛选抑制桃褐腐病菌Monilinia fructicola的内生拮抗真菌,通过菌落形态学特点观察及ITS基因测序分析,初步鉴定内生拮抗真菌的系统发育学地位;采用果实离体实验检测内生拮抗真菌的抗菌效果;使用电子显微镜观察被内生拮抗真菌抑制的桃褐腐病菌丝、孢子的形态以及细胞结构的变化。分离得到的内生真菌ZJ-4对桃褐腐病防效高且稳定,初步鉴定为篮状菌属Talaromyces;果实离体试验表明,内生真菌ZJ-4明显降低了桃褐腐病的发病率;通过电子显微镜观察到被内生真菌ZJ-4抑制的桃褐腐病菌丝及孢子表面粗糙凹陷,畸形现象严重,胞质溶胶收缩凝聚,细胞内部出现大量的空腔。本研究筛选的ZJ-4对桃褐腐病原菌的生长有明显的抑制作用,为开发应用该菌株提供参考。     

In vitro utilization of L-cystine by keratinophilic fungi inhabiting a gelatin factory

Twenty-six different species of keratinophilic fungi were examined to determine their ability to utilize free cystine. Of the fungi tested, the majority metabolized free L-cystine in a glucose-peptone culture medium. Cystine was used as source of sulfur, and carbon and nitrogen as well. Excess sulfur was excreted into the culture fluid, as thiosulfate and sulfate, following oxidation. The rate of cystine oxidation varied with the different fungal strains, but was maximal for Graphium penicilloideus (88.5%). Low quantities of thiols were found in the medium. Cystine oxidation and inorganic thiosulfate excretion were found to correlate significantly (r = 0.94).     

Screening of ectomycorrhizal fungi for degradation of polycyclic aromatic hydrocarbons

Ectomycorrhizal fungi belonging to 16 species (27 strains) were tested for their ability to degrade polycyclic aromatic hydrocarbons (PAHs): phenanthrene, chrysene, pyrene and benzo[a]pyrene. Cultivated on a complex liquid medium, most of the fungi tested were able to metabolise these compounds. Approximately 50% of the benzo[a]pyrene was removed by strains of Amanita excelsa, Leccinum versipelle, Suillus grevillei, S. luteus, and S. variegatus during a 4-week incubation period. The same amount of phenanthrene was also metabolised by A. muscaria, Paxillus involutus, and S. grevillei. The degradation of the other two PAHs was, for the most part, less effective. Only S. grevillei was able to remove 50% of the pyrene, whereas Boletus edulis and A. muscaria removed 35% of the chrysene. Received: 12 April 1999 / Received revision: 27 May 1999 / Accepted: 28 May 1999     

In vitro salt and thermal tolerance of fungal endophytes of Nicotiana spp. growing in arid regions of north-western Australia

Abstract

Endophytic fungal strains isolated from indigenous Nicotiana plants naturally growing in dry and hot regions of north-western Australia were characterised based on their tolerance to salinity and temperature. Sixty-eight fungal isolates were tested on eight levels (0.5 M, 1.0 M, 1.5 M, 2.0 M, 2.5 M, 3.0 M, 3.5 M and 4.0 M) of five different of salts solutions NaCl, KCl, MgCl₂, CaCl₂ and MgSO₄ and at various temperatures (25–50?°C). The salt adaptation test indicated that the fungal strains namely Aspergillus niger (E-202), A. ochraceous-A (E-134), Aurantiporus sp. (E-135), Cladosporium halotolerance (E-128), Pleurostomophora richardsiae (E-13) and Trichoderma sp. (E-185.1) were tolerant to higher concentrations of various salts. The most growth-limiting salt turned out to be MgCl₂ followed by the chaotrope CaCl₂. Responses to temperature tolerance revealed that most fungi tested could grow at 30?°C. About 50% all the fungi did not show any growth when the temperature was raised above 30?°C. When the temperature was raised up to 50?°C all the fungi failed to grow but the fungus Rasamsonia piperina (E-172). Endophyte strains identified could be promising candidates for future research in investigating the fungus–plant interactions and their roles in plant adaptation to inhospitable environments.