Chaetomium Thermophilum Research Articles

Purification and characterization of two xylanases from Chaetomium thermophile var. coprophile

Two xylanases (I and II) out of several extracellular xylanases produced by the thermophilic fungus Chaetomium thermophile var. coprophile were purified to homogeneity by a combination of ion exchange and gel filtration chromatographic procedures. They had molecular weights of 26 000 (xylanase I) and 7000 (xylanase II). The temperature optima for xylanase I and II were 70 and 60 °C, and they were optimally active at pH 4.8–6.4 and 5.4–6.0, respectively. Xylanase I was found to be comparatively more stable than xylanase II at higher temperatures. Amino acid composition indicated that xylanase I contained high amounts of glycine, threonine, and low amounts of histidine and sulphur-containing amino acids. Each enzyme released different hydrolysis products from larch wood xylan. Xylanase I produced mainly xylobiose and xylotriose whereas xylanase II produced mainly xylobiose.Key words: Xylanase, enzyme purification, characterization, Chaetomium thermophile.

Characterization of polyphenol oxidase from Chaetomium thermophile, a thermophilic fungus.

The polyphenol oxidase from Chaetomium thermophile Isolate O-453 was examined for its substrate specificity. The purified enzyme acted on polyphenols but not on monophenols. Among the polyphenols and phenol-like compounds tested, those oxidized were d-catechin (relative activity, 100%), l-epicatechin (95.3%), chlorogenic acid (80.0%), pyrocatechol (53.8%), DOPamine (47.8%), guaiacol (87.5%), and N, N-dimethyl-p-phenylenediamine (75.3%). Esculine (7.8%), resorcinol (0%), p-quinone (0%), o-phenylenediamine (1.3%), phenol (0%), and L-tyrosine (0%) werescarcely oxidized. The purified enzyme was inhibited by potassium xanthogenate, thiourea, cysteine, potassium cyanide, and sodium azide. Mercuric and ferrous ions inactivated the enzyme. Carbon monooxide did not inhibit the activity (residual activity, 92.0%). SDS-polyacrylamide gel electrophoresis showed that the purified enzyme (mw, 98, 000) is a trimer composed of three of the same kinds of monomers (mw, 36, 000). These data indicate that the polyphenol oxidase from C. thermophile Isolate O-453 is a copper-containing protein and is classified as laccase [benzenediol: oxygen oxidoreductase, EC 1.10.3.2].

Purification and properties of polyphenol oxidase from Chaetomium thermophile, a thermophilic fungus.

A polyphenol oxidase [EC class 1.10.3] was purified from culture filtrates of Chaetomium thermophile Isolate o-453 by procedures including salting- out by ammonium sulfate and combined column chromatography on DEAE-Toyopearl 650S, Con A-Sepharose, hydroxylapatite, and Sephadex G-200. The final enzyme preparation was homogeneous on polyacrylamide gel disc electrophoresis. Its estimated molecular weight was approximately 98, 000 by gel filtration. It showed maximum activity at pH 5.0 and at 55°C. The purified enzyme was very stable between pH 4.0 and 11.0 and up to 55°C. It was highly active toward d-catechin (100%), p- hydroquinone (45.7%), and N, N-dimethyl-p-phenylenediamine (63.0%). Though pyrocatechol (22.6%) was oxidized to some extent, DL-DOPA (0.33%) and o-phenylenediamine (3.1%) were hardly oxidized by the enzyme.

Decomposition of filter paper cellulose by thermophilic fungi acting singly, in combination, and in sequence

At 45°C, but not at 37°, more decomposition of filter paper was caused by Chaetomium thermophile (C.t.) in combination with the non-cellulolytic Thermomyces lanuginosus (T.l.) than by C.t. alone. The difference was seen as the rate of decomposition by C.t. began to decline; this was influenced by available nitrogen. As evidenced by weakening of cellulose film, more cellulase activity was present in C.t. + T.l. than in C.t. treatments in the later stages of incubation. C.t. alone caused little or no further weight-loss of filter paper after 4 weeks unless nitrogen was supplemented. Yet Aspergillus fumigatus, Thermoascus aurantiacus and T.l. (but not Scytalidium thermophilum ) caused weight loss of filter paper on which C.t. had become nitrogen-limited. A. fumigatus also degraded filter paper on which T. aurantiacus had grown, whereas C.t. did not do so. It is shown that, in some circumstances, more cellulose decomposition can be caused by mixtures or sequences of fungi than by single species. Also, some fungi seem able to obtain nitrogen when it has become unavailable to others ; this may be especially relevant to fungi that occur in the later stages of successions.

Thermophilic and Thermotolerant Fungi Isolated from Mangrove Swamps

The ability of fungi to populate natural and man-made self-heated habitats has been reported by several workers, among them Cooney and Emerson (1964), Tansey (1971) and Tansey and Brock (1978). In addition, thermophilic fungi have been isolated from mesic habitats (Subrahamanyam, 1978; Ellis, 1980). Mangrove regions, because of their particular ecological conditions, viz., high salinity, moisture level, organic matter and pH, poor aeration, dense evergreen vegetation, and absence of elevated temperature, afford an interesting habitat for exploring comparatively the occurrence of thermophilic and thermotolerant fungi. Mud and wood samples were collected aseptically from Sunderban mangrove areas of West Bengal, India. Thermophilic fungi were isolated by a modified direct plate and a dilution plate method (Warcup, 1950, 1955). Three serial dilutions were used each with eight replicate plates. The fungi were isolated on Emerson's YpSs agar, Czapek-Dox agar supplemented with 1% cellulose, and malt extract agar with dicrysticin (0.2%), and incubated at 45 and 50 C in moist chambers. The thermophilic or thermotolerant nature of the species was determined by measuring their radial growth and dry mycelial weight on solid and liquid YpSs medium, respectively, at 20 and 50 C. Twenty-five species (Zygomycotina, Ascomycotina, and Deuteromycotina) were isolated including Aspergillus carbonarius, A. niger, A. niveus, and A. terreus var. africanus, recorded for the first time from India. Aspergillus carneus, A. fumigatus, A. niger, A. terreus, Chaetomium thermophile, Mucor pusillus, Sporotrichum sp., and Thermomyces lanuginosus were dominant and first to appear on agar plates, while Acrophialophorafusispora, Humicola insolans, Malbranchea sulfurea, and Paecilomyces variotii were rather rare. Per cent frequency of the species is given in TABLE I. The presence of thermophilic fungi in the mangrove swamp is of interest since the temperature is not elevated but there are extreme ecological conditions in other parameters. In spite of high salinity, pH, poor aeration, and dense vegetative cover, the presence of high organic matter and elevated moisture level in the swamp provides a suitable substrate for the growth of many fungi including thermophiles. Elliott (1930), who gave the first account of fungi inhabiting salt marsh sediments, concluded that organic matter is mainly responsible for the activity of fungi in such a unique habitat. The average annual rainfall and surface temperature in the study area are 192.04 cm and 20 to 33.7 C, respectively. These situations provide a suitable environment for the decomposition of organic matter thus favoring the conditions for the growth and occurrence of thermophilic fungi but at the same time other abnormal ecological conditions mentioned above limit this possibility. Although there is no evidence that these fungi are actively growing in the mangrove, the observations do show that propagules of thermophilic fungi exist in the mangrove swamps. There are three possible explanations. (1) Thermophilous fungi have a wide range of temperature tolerance so can grow within the range found in a mangrove habitat. (2) These fungi may be part of the microflora of the indigenous biota and may exist in soil and woods only as spores. (3) These fungi actually may be growing in warm microenvironments associated with de

Thermophilic and thermotolerant fungi in the desert soils of Saudi Arabia

Forty-eight thermophilic and thermotolerant species in addition to 5 varieties which belong to 24 genera were collected from desert soils in Saudi Arabia on glucose-(22 genera and 38 species + 5 varieties), cellulose-(15 genera and 27 species + 4 varieties) and 40% sucrose-Czapek's agar plates (13 genera and 26 species + 4 varieties) at 45 °C. The most frequent species were as follows: Aspergillus fumigatus, A. terreus, Humicola grisea var. thermoidae and Chaetomium thermophile var. copropile on glucose-; A. fumigatus, C. thermophile var. copropile, A. terreus, A. nidulans and C. thermophile var. dissitum on cellulose-; and A. fumigatus and A. terreus on 40% sucrose-Czapek's agar plates. Sixteen species and 4 varieties were particularly thermophilic and these were A. fumigatus, H. grisea var. thermoidae, H. insolens, H. lanuginosa, C. thermophile var. copropile, C. thermophile var. dissitum, C. virginicum, M. pusillus, S. thermophila, S.? pulverulentum, T. thermophilus, T.? emersoni, T. aurantiacus, T. thermophila, M. pulchella var. sulfurea, M. albomyces, ?A. terrestris, C. pruinosum, T. thermophila and P. thermophila. The remaining species showed different degrees of thermotolerant (32 species + 1 variety).

Degradation of Nigerian palm products by thermophilic fungi

The ability of thermophilic fungi to degrade Nigerian palm products and the factors which affect their activities were examined. A considerable decrease in the oil contents of the palm fruit chaffs and the palm kernels by Chaetomium thermophile, Humicola lanuginosa , and Torula thermophila , was recorded, accompanied by a corresponding increase in the free fatty acid content. The degradation of the palm products by the fungi used depended on pH and temperature. Best results were recorded at a pH of 5 · 5–6 · 4 and 40°–50° respectively. Their growth on the palm oil and the palm kernel oil led to a rapid increase in the free fatty acid content and a decrease in the pH of the media. The palm oil was more readily utilized than the palm kernel oil but no significant differences (at P =

Thermophilic Fungi Isolated from a Heated Aquatic Habitat

The cooling pond for the Hazelwood Power Station, Morwell, Victoria, Australia, was studied for the presence of thermophilic fungi. A total of five species was recorded: Chaetomium thermophile, Aspergillus fumigatus, Humicola grisea, Humicola insolens, and Thermomyces lanuginosus. (ACR)

Thermophilic mycoflora of cigarettes and cured tobacco leaves.

Nine species of thermophilic fungi were obtained from retailed packets of imported and locally manufactured brands of cigarettes and from cured tobacco leaves. They include known human pathogens such as Thermoascus aurantiacus Miehe sensu Apinis, Mucor pursillus Lindt and Mucor miehei Cooney and Emerson. Chaetomium thermophile La Touche, Humicola insolens Cooney and Emerson, Thermoascus crustaeus (Apinis and Chesters) Stolk and Mucor miehei have not been previously reported on tobacco products. Fewer species were obtained from foreign cigarettes than from locally manufactured brands. A mean moisture content of 19% was recorded for imported cigarettes and 37% for Nigerian brands. The potential health hazards posed to man by the unrestricted use of tobacco products are discussed.

The effects of fungicides on cellulose and starch hydrolysis by thermophilous and mesophilic fungi

1. 1.|Eleven of the seventeen genera of thermophilous fungi studied were cellulolytic. 2. 2.| Chaetomium thermophile var. coprophile and Sporotrichum thermophile were the most vigorous cellulolytic species; Mucor pusillus was non-cellulolytic but showed vigorous starch hydrolysis. 3. 3.| A. fumigatus was found to be a strong cellulose-and-starch-hydrolysing fungus. 4. 4.|The thermophilous fungi showed more vigorous starch and cellulose hydrolysis than the mesophiles. 5. 5.|Of the five fungicides studied, Thiram and Verdasan showed most marked inhibitory effects on decomposition by thermophilous fungi. 6. 6.|Verdasan (organically combined mercury) was more inhibitory to hydrolysis by thermophiles and mesophiles than was HgCl 2.

Amylase and cellulase activities of thermophilic fungi causing deterioration of tobacco products in Nigeria

The conditions necessary for the production and activities of extracellular enzymes of thermophilic fungi obtained from tobacco products were determined. While amylases were produced in extractable amounts by all the fungi used in this study, extracellular cellulases were produced only by Chaetomium thermophile La Touche, Humicola insolens Cooney and Emerson, Malbranchea pulchella Sacc. et Penzig var. sulfurea (Miehe) Cooney and Emerson, and Mucor pusillus Lindt. A narrow range of pH 6–7 was best for the activities of the amylases of all the fungi used while a lower range of pH 5–6 was optimal for their cellulase activities. Peak activities were recorded for both enzymes at a temperature range of 45 °–50 °C. The possible use of the exo-enzymes of these fungi in industrial fermentation processes is suggested.

Cellulases from Chaetomium thermophile var. dissitum.

When cultivated in a liquid medium with cellulose, Chaetomium thermophile var. dissitum produced extracellular cellulolytic enzymes. By concentration of the culture filtrate, followed by ion‐exchange chromatography on DEAE‐Sephadex A‐50 and gel filtration on Biogel P‐100, three electrophoretically pure components were obtained. Of these, one was a typical Cx enzyme (endoglucanase) causing rapid decrease of the viscosity of carboxymethylcellulose solutions, while showing low effect on native cellulose.The other was active towards native cellulose, but had little effect on the viscosity of carboxymethylcellulose. It is concluded that this enzyme is an exoglucanase (C1 enzyme), possibly a cellobiohydrolase.The third component showed only cellobiase (β‐glucosidase) activity, and had no effect on cotton or carboxymethylcellulose. The three components, when mixed, showed synergistic effects on highly ordered cellulose. The endo‐ and exoglucanases were characterized with regard to molecular size and iso‐electric point (pI). Both cellulases had pI near 4.55, but their molecular weights were different: 67 000 (exoglucanase) and 41 000 (endoglucanase). The effect of temperature on the activity of the cellulases was examined with both cotton and carboxymethylcellulose as substrate. Arrhenius activation energies, Q10, and temperature optima for the different reactions were determined.

Ascospores, Germ Pores, Ultrastructure, and Thermophilism ofChaetomium

SUMMARY Ascospores of 15 isolates of Chaetomium thermophile were compared with those of 105 other Chaetomium species by use of light microscopy and scanning electron microscopy. Two stable characteristics distinguished C. thermophile ascospores from those of all other Chaetomium species examined: a papillate germ pore and pustulate surface ornamentation. Transmission electron microscopy revealed multilayered walls in ascospores of C. semen-citrulli and C. thermophile. In addition, germ pores were confirmed as thin areas in ascospore wall layers. Germinated ascospores of C. semen-citrulli contained several nuclei, mitochondria, fibrillar bundles, vacuoles, and glycogen-like particles.

Degradation of Wood by Aspergillus fumigatus Isolated from Self-Heated Wood Chips

SUMMARYAspergillus fumigatus was one of the commonest fungi isolated from self-heated wood chips and bark waste incubated at 37 C and 50 C. Other fungi included Absidia spp., Chaetomium thermophile var. dissitum, Rhizopus spp., Thermoascus crustaceus and Trichoderma koningii. Thermomyces lanuginosus was isolated from bark, and the yeast Kluyvero- myces cicerisporus from wood chips. Aspergillus fumigatus, strain FW35, produced CM-cellulase, xylanase, 1,3-β-glucanase and amylase during growth in shake-flask culture with beech, oak and Norway spruce sawdust as carbon source. The one percent alkali solubility of sawdust decreased slightly during this growth (significantly in oak), indicating limited utilization of alkali-soluble hemicelluloses rather than extensive depolymerization of cellulose. Incubation of sawdust with culture filtrates resulted in significant weight losses in beech and oak sawdust, and there were also significant weight losses in beech and Norway spruce wood shavings incubated in plate cultures.

Degradation of Wood by Aspergillus Fumigatus Isolated from Self-Heated Wood Chips

Aspergillus fumigatus was one of the commonest fungi isolated from self-heated wood chips and bark waste incubated at 37 C and 50 C. Other fungi included Absidia spp., Chaetomium thermophile var. d...

The effect of temperature on growth and cellulase (?-1,4-endoglucanase) production in the compost fungusChaetomium thermophile var.dissitum

Chaetomium thermophile var.dissitum, isolated from an experimental urban refuse compost, had the following growth characteristics: Minimum temperature, 27±1°C; optimum, 45–50°C; maximum, 57±1°C; pH optimum 5.5–6.0.

Thermophilic Fungi in Sun-Heated Soils

Thermophilic and thermotolerant fungi were isolated from soils of south-central Indiana (U.S.A.) in the following order of decreasing frequency: Myriococcum albomyces Cooney et Emerson, Aspergillus fumigatus Fres., Talaromyces thermophilus Stolk, Humicola lanuginosa (Griff. et Maubl.) Bunce, Allescheria terrestris Apinis, Malbranchea pulchella var. sulfurea (Miehe) Cooney et Emerson, Thielavia heterothallica Klopotek, Mucor pusillus Lindt, Chaetomium thermophile var. dissitum Cooney et Emerson, Thielavia minor (Rays et Borut) Malloch et Cain, Thermoascus aurantiacus Miehe sensu Apinis, Mucor miehei Cooney et Emerson, Torula thermophila Cooney et Emerson, Humicola stellata Bunce, Acrophialophora nainana Edward, Thielavia sepedonium Emmons, Dactylomyces thermophilus Sopp, and Talaromyces emersonii Stolk. Four unidentified thermophilic fungi were also isolated. Thermophilic and thermotolerant fungi were isolated most frequently from sun-heated soil, less so from grass-shaded soil, and still less so from tree-shaded soil. Isolates were more frequently obtained from a depth of 0-5 cm than from 10-20 cm. Data concerning numerical values for population numbers and soil pH, moisture, temperature, and percent organic matter are given for samples from the two soil levels for each of 15 consecutive months. Results of special procedures for enrichment of thermophilic and theromotolerant fungi are described, as well as results of air sampling.

Ultrastructural Studies on Thermomyces lanuginosa and Certain Other Closely Related Thermophilic Fungi

SUMMARYFine structural details of the thermophiles Thermomyces lanuginosa, T. stellatus, Humicola insolans, and H. grisea var. thermoidea are described and illustrated. When grown at elevated temperatures, these thermophiles were observed to contain from one to several large, membrane-bound inclusion bodies in the cytoplasm. These inclusion bodies were usually seen in close association with a rich detail of membranes that appeared continuous with the plasma membrane. The electron opacity of these intracytoplasmic inclusion bodies was enhanced markedly by treatment of the hyphae with the osmiophilic reagent thiocarbohydrazide, but not with alkaline diaminobenzidine-hydrogen peroxide. The known cytochemical reactions of thiocarbohydrazide with cellular lipid components suggested that these inclusion bodies might be lipoidal in nature, and that they might represent an organelle related in some manner with the thermophilic nature of these fungi. Similar intracytoplasmic inclusion bodies could not be demonstrated in hyphae of the thermophiles Chaetomium thermophile var. coprophile, Mucor pusillus, or Thermoascus aurantiacus.

Studies on Thermophilic Cellulolytic Fungi

Three thermophilic cellulolytic fungi, Chaetomium thermophile var. coprophile, Sporotrichum thermophile, and Thermoascus aurantiacus were studied to determine the conditions for a high rate of cellulose degradation. The range of temperature over which good growth occurred was determined first in a temperature gradient incubator; the optimum temperature was then established in shake flask cultures. T. aurantiacus had the highest optimum growth temperature range (46 to 51 C), whereas S. thermophile had the broadest range over which good growth occurred (36 to 43 C). Optimum temperatures for the three organisms, T. aurantiacus, S. Thermophile, and C. thermophile were 48, 40, and 40 C, respectively. It was found that the addition of an organic carbon and nitrogen source to a cellulose mineral solution medium markedly increased the rate of cellulose degradation. The surfactant, Tween 80, which has been reported to be of value in the production and recovery of the enzyme, cellulase, was shown to be detrimental to the degradation of cellulose in culture. In the medium used, S. thermophile gave the highest rate of substrate utilization; 56% of the cellulose was hydrolyzed in 72 h. The average degree of polymerization of cellulose decreased from 745 to 575.

Utilization of amino acids as sole nitrogen sources by thermophilic fungi

The organic nitrogen requirements of six thermophilic fungi were examined in liquid shake cultures at 45 °C. The fungi used were Mucor pusillus LINDT, Humicola lanuginosa (GRIFFON and MAUBLANC) BUNCE, Thermoascus aurantiacus MIEHE sensu APINIS, Thermoascus crustaceus (APINIS and CHESTERS) STOLK, Chaetomium thermophile var. coprophile COONEY and EMERSON, and Chaetomium thermophile var. dissitum COONEY and EMERSON. The organic nitrogen sources tested were casein hydrolysate (vitamin-free), L-proline, L-glutamic acid, DL-serine, L-alanine, DL-valine, L-threonine, L-arginine, DL-aspartic acid, L-asparagine, DL-glutamine, glycine, L-leucine, DL-isoleucine, L-lysine, DL-methionine, DL-phenylalanine, and L-histidine. Growth was determined by the mycelial dry weight method. All the organisms grew well on casein hydrolysate. M. pusillus grew well on all the single amino acids, while H. lanuginosa grew well on all, but six of them. T. aurantiacus and T. crustaceus gave a fair yield of mycelium on about 50% of the single amino acids tested, while the other 50% served as poor sources of nitrogen. All the single amino acids were poorly utilized by C. thermophile var. coprophile and C. thermophile var. dissitum.