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Remarkably AT-rich genomic DNA from the anaerobic fungus Neocallimastix.

The genomic DNA of an anaerobic rumen phycomycete of the genus Neocallimastix has been purified and characterized. The non-repetitive fraction of the DNA has a G.C content of only 13%. The ribosomal RNA genes are highly reiterated, making up about 30% of the total DNA, and are evident as a more G.C-rich satellite with a repeating unit of about 9.4 kilobases (Kb). A.T-rich regions of DNA are highly dispersed and possess some sequence complexity. Chemical analysis of the DNA constituents reveals no evidence of modified bases. The genome of this anaerobic fungus has the highest A.T content of any organism so far described.

Base Composition↗

Ventricular visualization on brain scan with intracranial hemorrhage in disseminated phycomycosis.

Infections with fungi of the class Phycomycetes result in distinctive tissue changes characterized by growth of hyphae in and around blood vessels, causing necrosis and hemorrhagic infarction of tissue. A brain scan on a patient with a phycomycotic intracranial aneurysm demonstrated ventricular visualization at a time when there was clinical and autopsy evidence of rupture of the aneurysm.

Brain↗

The lipids of the rumen fungus Piromonas communis.

The major phospholipids of the anaerobic rumen phycomycete Piromonas communis were phosphatidylethanolamine (38%), phosphatidylcholine (26%) and phosphatidylinositol (13%); no sphingolipids, glycolipids, plasmalogens or phosphonyl lipids were detected. Free fatty acids, triacylglycerols, 1:2 diacylglycerols and a variable amount of 1:3 diacylglycerol were identified, as were minor amounts of squalene and a triterpenol which is probably tetrahymanol. Approximately half the fatty acids were straight chain, even 12 to 24 carbon, saturated acids, the remainder being even 16 to 24 carbon, mono-unsaturated fatty acids. The double bonds in all except the 16 carbon acid were in the omega 9 position. The unsaturation is introduced by a delta 9 desaturase which uses stearic acid as substrate and which does not use oxygen as a terminal electron acceptor. 14C from acetate and glucose was incorporated into the fatty acids of all complex lipids, as were lauric, myristic, palmitic, stearic and oleic acids. [14C]Choline was incorporated into phosphatidylcholine and [14C]ethanolamine into phosphatidylethanolamine and phosphatidylcholine. Label from [14C]serine was recovered in phosphatidylserine and phosphatidylethanolamine, but was not detected in phosphatidylcholine.

Animals↗

Transhyphal electrical currents in fungi.

Representative mycelial fungi from the phycomycete, ascomycete and basidiomycete groups (Achlya bisexualis, Neurospora crassa, Aspergillus nidulans, Schizophyllum commune and Coprinus cinereus) all generated steady electrical currents around their hyphal tips; the generation of a transhyphal ion current may therefore be a universal characteristic of hyphal growth. As with all other tip growing organisms, positive current always entered apically and left distally; non-growing hyphae did not drive transcellular currents. The current density, measured approximately 30 micron from the membrane surface at the hyphal tips, varied between 0.05 and 0.06 microA cm-2 in different fungi and tended to be larger in wider, rapidly extending hyphae than in thinner, slow growing hyphae. The possibility that these currents serve to localize growth at the apex is discussed.

Electricity↗

Studies on the rumen flagellate Neocallimastix frontalis.

The vast increase in the population density of the rumen flagellate Neocallimastix frontalis shortly after the host animal has commenced eating is caused by stimulation of a reproductive body on a vegetative phase of the organism to differentiate and liberate the flagellates. The stimulant is a component of the host's diet. The vegetative stage of N. frontalis bears a strong morphological resemblance to that of certain species of aquatic phycomycete fungi, and consists of a reproductive body borne on a single, much branched rhizoid. The flagellates liberated in vivo within 15 to 45 min of feeding lose their motility within I h and develop into the vegetative phase, thus producing a rapid decrease in population density of the flagellates. Conditions for maximum flagellate production are similar to those occurring in the rumen: pH 6-5, 39 degrees C, absence of O2, presence of CO2. Differentiation of the reproductive body is inhibited by compounds affecting membrane structure and function, but not by inhibitors of protein synthesis. The organism was cultured in vitro in an undefined medium in the absence of bacteria or other flagellates.

Animals↗

Studies on the rumen flagellate Sphaeromonas communis.

The rumen flagellate Sphaeromonas communis showed a significant increase in population density 1 to 2 h after the host sheep commenced feeding, followed by a reduction in numbers to the pre-feeding level after a further 2 to 3 h. The life-history of the organism was shown to consist of a motile flagellate which germinated to produce a vegetative stage comprising a limited rhizoidal system on which up to three reproductive bodies were borne together with (in vitro) other spherical bodies of unknown function; in vivo, the reproductive bodies were stimulated to liberate flagellates by a component of the diet of the host. The vegetative stage strongly resembled that of certain species of aquatic phycomycete fungi, and the flagellates may therefore by zoospores. Flagellates liberated in vivo lost their motility within 2 to 3 h and developed into the reproductive vegetative phase, producing a rapid decrease in numbers of flagellates. Conditions of maximum flagellate production (pH 6.5, 39 degrees C, presence of CO2, absnece of oxygen) approximated to those found in the rumen. The organism was cultured in vitro in an undefined medium in the absnece of bacteria and other flagellates.

Animals↗

Invasion of plant tissue in the rumen by the flagellate Neocallimastix frontalis.

The flagellate stage of the rumen phycomycete Neocallimastix frontalis invades and germinates on plant material in the rumen and in vitro, preferentially invading the lemmas, paleas, awns and flower bracts in members of the plant family Gramineae, and flower bracts in certain of the Papilionaceae. The principal sites of invasion were the stomata and damaged tissue, through which penetration of the plant tissue by the rhizoid of the germinating cell occurred. Subsequent growth of the vegetative stage occurred with the uptake of 14C from 14C-labelled plant tissue. Host diets rich in seed-head material normally supported higher population densities of N. frontalis in the rumen even though the dry matter digestibilities of these diets were lower.

Animals↗

The rumen flagellate Piromonas communis: its life-history and invasion of plant material in the rumen.

The rumen flagellat Piromonas communis is the zoospore of a phycomycete fungus inhabiting the rumen. Zoosporogenesis was stimulated by a dietary component (the inducer), and inhibited by compounds affecting membrane structure and function, but not by inhibitors of protein synthesis. The zoospores showed taxis towards the tissues surrounding the inflorescence of Lolium perenne L. in the rumen, invading principally the stomata and damaged tissues. The zoospores germinated on this substratum and the rhizoids of the developing vegetative stage penetrated the tissue, taking up C14 from labelled plant material, which was incorporated into the fungal cells. The conditions for maximum flagellate production (39 degrees C, pH 6-0 to 7-0, high concentration of CO2, absence of O2) resembled those found in the rumen. The organism was cultured in an undefined medium in vitro in the absence of other flagellates.

Animals↗

Factors determining the susceptibility of mice to experimental phycomycosis.

Various factors influencing the susceptibility of C3H mice to lethal infection by the phycomycete Absidia ramosa were examined. Mice 19-21 days old were exposed to graded doses of A. ramosa spores by various routes. After intravenous inoculation with doses in excess of 10(3) viable units, a variable proportion of mice developed lethal phycomycosis of the central nervous system within 2-8 days. The proportion of mice affected was related to the inoculum size, doses of 5 X 10(7) spores producing lethal infection in 90-100% of the mice. The disease was characterised by signs of involvement of the central nervous system appearing 8-12 h before death. At necropsy, fungal hyphae, frequently surrounded by infiltrations of mononuclear cells, could be demonstrated in the brain. Lesions were also often present in the kidneys; in other organs they were rare, but the presence of viable fungal spores could be detected by cultural procedures. Subcutaneous inoculation of A. ramosa spores did not produce lethal infection but intraperitoneal inoculation occasionally did so. Direct intracerebral inoculation invariably produced lethal infection even with very small doses of spores. The clinical and histopathological features of the disease were almost identical with those resulting from intravenous inoculation. Pre-inoculation treatment with azathioprine, cyclophosphamide and antithymocyte serum did not increase susceptibility. Susceptibility was increased by injections of zymosan, aggregared gamma-globulin, cortisone acetate and Fe(II) salts and by reticuloendothelial blockade. These treatments increased the proportion of mice developing lethal phycomycosis of the central nervous system, and in the case of cortisone acetate, also promoted disseminated infection. It was concluded that the natural resistance of mice to A. ramosa infection was probably dependent upon the activity of phagocytic cells, possibly acting in conjunction with complement and other non-specific serum factors.

Age Factors↗

The rumen microbiology of seaweed digestion in Orkney sheep.

The microbial populations of the rumens of seaweed-fed and pasture-fed Orkney sheep were examined. The populations in the pasture-fed sheep were similar to those of other domestic ruminants fed on land plants, but those of the seaweed-fed animals showed major differences in the dominant species. Total ciliate populations were quantitatively similar, but in the seaweed-fed animals Dasytricha ruminantium was one of the most dominant species. No phycomycete fungi or cellulolytic bacteria were found in the seaweed-fed animals, and the bacterial population was dominated by Streptococcus bovis, Selenomonas ruminantium, Butyrivibrio fibrisolvens and lactate-utilizing species. Electron microscopy revealed that spirochaetes and an unidentified filamentous bacterium were probably of major significance in seaweed digestion. The ability of bacterial strains from both groups of animals to metabolize plant and algal constituents was examined.

Animals↗

Pythium contiguanum nomen novum (syn. Pythium dreschleri Paul), its antagonism to Botrytis cinerea, ITS1 region of its nuclear ribosomal DNA, and its comparison with related species.

Pythium drechsleri Paul was described as a new species from soil samples taken in a salt-marsh of Arzew, Algeria [Paul, B. (1988) Une nouvelle espèce de Pythium isolée d'une saline de l'ouest Algérien. Cryptogam. Mycol. 9, 325-333]. The name of the fungus, P. drechsleri, is a nomen invalidum, as it is a later homonym of P. drechsleri Rajgopalan and Ramakrishnan [Rajagopalan, S. and Ramakrishnan, K. (1971) Phycomycetes in agricultural soils with special reference to the Pythiaceae. Madras Univ. J. Sect. B 37,38, 100-117]. A new name, Pythium contiguanum is now being given to P. drechsleri Paul. This species is characterised by its contiguous inflated type of sporangia, smooth-walled oogonia and mostly monoclinous antheridia. Although the fungus is not known to be a pathogen, it has a very well developed appressorial system comprised of both sickle shaped and coiled appressoria. Morphological features, its antagonism towards the grape-vine pathogen Botrytis cinerea, together with the sequences of the ITS1 region of its nuclear ribosomal DNA and its comparison with related species are given here.

Antibiosis↗

Mycotic abortion in cattle.

Placentas and foetuses from 1107 bovine abortions were examined and a fungal infection was diagnosed in 131 instances. Fungi seen in silver impregnated sections of tissues could be placed into 3 categories designated aspergillus, phycomycete and atypical. Culture indicated that the first 2 of these categories were due to Aspergillus sp and Mortierella wolfii respectively. The infections in the atypical category are probably also due to Aspergillus sp. Gross or microscopic examination or culture of the placenta were valuable diagnostically but examination of the foetus was seldom of value as infection in most instances did not involve foetal tissues. Questionnaires indicated that many M. wolfii abortions were associated with the feeding of poorly prepared or stored grass silage. As the majority of cattle are fed hay or silage, no association could be demonstrated between Aspergillus sp abortion and these feeds. Fertility following fungal abortion was apparently unimpaired.

Abortion, Veterinary↗

Heterothallism in biflagellate aquatic fungi: preliminary genetic analysis.

Genetic analysis of several hundred progeny from crosses of two heterothallic species of Achlya and Dictyuchus provides preliminary information about the life cycle and pattern of sexuality in the biflagellate Phycomycetes. Extensive testing of mycelial progeny indicates a diploid life cycle. Control of sexual expression and mating competence appears to be based on a complex genetic system.

Female↗

Comparative in vitro antifungal activity of amphotericin B and amphotericin B methyl ester.

The in vitro antifungal activity of amphotericin B methyl ester (AME), a water-soluble derivative of amphotericin B, was compared to that of the parent compound against a variety of pathogenic and potentially pathogenic fungi. AME has a significant antifungal activity, but the activity of AME was slightly lower than that of amphotericin B. Among the yeast-like organisms, only the yeast cells of Sporothrix schenckii were more resistant than others to both antibiotics, with a minimal fungicidal concentration of 5 to 10 mug/ml. The yeast cells of other fungi were killed at concentrations of 1 mug or less of either antibiotic per ml. The filamentous forms of S. schenckii and Oidiodendron kalrai were more resistant than the filamentous forms of other dimorphic fungi to both drugs. The minimal fungicidal concentration for S. schenckii was 10 mug/ml and for O. kalrai, 50 mug/ml. The dermatophytes, phycomycetes, and dematacious and other potentially pathogenic fungi were inhibited fairly well by both drugs, but up to 50 mug/ml was required for fungicidal action. The water solubility and wide spectrum of antifungal activity of AME warrant evaluation of its chemotherapeutic activity against experimental fungal infections.

Amphotericin B↗

Production of Plant Growth-Regulating Substances by the Vesicular-Arbuscular Mycorrhizal Fungus Glomus mosseae.

Glomus mosseae, a representative species of Endogonaceae (Phycomycetes) able to form vesicular-arbuscular mycorrhiza, was investigated for phytohormone production. Spores of G. mosseae were axenically germinated in water, and the resultant mycelial growth was assayed by standard procedures for extracting plant hormones from microbial cultures. Paper partition chromatography and specific bioassays were used to separate and identify plant growth-regulating substances. The microorganism synthesized at least two gibberellin-like substances, one with R(f) corresponding in position to authentic gibberellic acid, and four substances with the properties of cytokinins.

Journal Article↗

Separation of fungal propagules by partition in aqueous polymer two-phase systems.

Conidia of Penicillium chrysogenum and Penicillium frequentans and sporangiospores of Rhizopus rhizopodiformis, Rhizomucor pusillus, and Mucor racemosus were subjected to partition in aqueous polymer two-phase systems. The partition behavior differed drastically between the conidia of the two Penicillium species and the sporangiospores of the three species of phycomycetes. This difference in partition behavior can be used for purification of fungi belonging to different taxonomical groups. P. frequentans was completely separated from M. racemosus by two extractions, whereas four extractions were needed to purify M. racemosus. This method was used on an air sample from a locality where wood fuel chips are handled. The conidia of the fungi Trichoderma viride and Rhizopus rhizopodiformis were removed completely by only two extractions.

Journal Article↗

Purification and Characterization of an Aspecific Glycoside Hydrolase from the Anaerobic Ruminal Fungus Neocallimastix frontalis.

A glycoside hydrolase characterized by beta-fucosidase (EC 3.2.1.38) and beta-glucosidase (EC 3.2.1.21) activities was purified from the culture medium of the anaerobic ruminal phycomycete Neocallimastix frontalis grown on 0.5% Avicel. The enzyme had a molecular mass of 120 kilodaltons and a pI of 3.85. Optimal activity against p-nitrophenyl-beta-d-fucoside and p-nitrophenyl-beta-D-glucoside occurred at pH 6.0 and 50 degrees C. The beta-fucosidase and beta-glucosidase activities were stable from pH 6.0 to pH 7.8 and up to 40 degrees C. They were both inhibited by gluconolactone, sodium dodecyl sulfate, p-chloromercuribenzoate, and Hg cation. The enzyme had K(m)s of 0.26 mg/ml for p-nitrophenyl-beta-d-fucoside and 0.08 mg/ml for p-nitrophenyl-beta-d-glucoside. The purified protein also had low beta-galactosidase activity.

Journal Article↗