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Transmission of mitochondrial DNA in Ustilago violacea.

Mitochondrial DNA (mtDNA) restriction fragment length polymorphisms (RFLPs) were used as genetic markers for following mitochondrial transmission in the basidiomycete Ustilago violacea. Yeast-like cells of opposite mating types (a1 and a2) were mated on 2% water agar and were treated with alpha-tocopherol to induce formation of dikaryotic hyphae. Upon depletion of the alpha-tocopherol, the hyphae budded off haploid cells with parental nuclear genotypes. These cells were examined for mitochondrial RFLP phenotype. In progeny expressing the a1 mating type, mitochondria from either parent were observed equally frequently. In progeny with the a2 mating type, mitochondria were almost exclusively (94%) from the a2 parent.

Crosses, Genetic↗

Pleiotropic and differential phenotypic expression of two sn (snowflake) mutant alleles of Neurospora crassa: analysis in homokaryotic and heterokaryotic cells.

Mutations sn (snowflake) JL301 and C136, in the centromere region of linkage group I in Neurospora crassa, are at 0.6-3.0 map units to the left of the os-4 locus. Strains carrying snJL301 produce very short aerial hyphae and only arthroconidia, and do not grow in high salt media. snC136 strains produce aerial hyphae, with abnormally large and rounded blastoconidia, at the top of the agar slant cultures, and revert to wild-type growth in high salt media. Studies with forced primary heterokaryons indicate that snJL301 is recessive while snC136 is a semi-dominant and gene-dose dependent allele, with respect to the wild-type. Taken together the results show that: (1) the sn mutations are allelic with a differential pleiotropic phenotype, and (2) snC136 may code for a partially functioning gene product while snJL301 appears to be a null allele.

Alleles↗

Hyphal fusion during initial stages of trap formation in Arthrobotrys oligospora.

Hyphal fusion during initial stages of trap formation by Arthrobotrys oligospora was studied by video-enhanced contrast and electron microscopy. Trap initials grew perpendicularly to the parent hypha, then curved around and anastomosed with a peg that developed on the hypha. Trap initials usually developed 40-140 microns apart while the anastomosis occurred 20-25 microns from the initial. Vigorous cytoplasmic movements in trap initials and developed traps corresponded to intense staining with fluorescein diacetate (FDA) of these cells. In addition, bundles of microfilaments were seen in developing loops of traps. On fusion organelle migration took place from the tip cell of the trap into the peg. Later on a septum was formed at the site of fusion.

Cell Wall↗

Mutations affecting cyclic phosphodiesterases and adenylate cyclase in Neurospora.

Two regulatory mutants in orthophosphate-regulated cyclic phosphodiesterase (cPDase), cpd-3 and cpd-4, were isolated and mapped proximal to arg-1 on L.G. IC and distal to arg-12 on L.G. IIR, respectively. cpd-3 showed short aerial hyphae with dense formation of conidia. The morphology was very similar to that of cr-1, cpd-3 and cr-1 had reduced levels of cyclic 3',5'-AMP, adenylate cyclase and cPDases (CPDase I, II and III in low phosphate condition) but had elevated levels of cyclic 3',5'-GMP. Although cr-1 showed an enhanced level and enhanced activation of heat activated cyclic phosphodiesterase (ha-PDE), this enzyme was not activated in cpd-3. cpd-4, nuc-1 and nuc-2 produced neither cPDase I, II, III, alkaline phosphatase nor ribonuclease N1 in low phosphate media. These mutants did not produce aerial hyphae or conidia when grown in low phosphate liquid medium. Activation of ha-PDE occurred in cpd-4, but not in nuc-1 and nuc-2.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗

Microscopic observations on the interaction of the mycoparasite Verticillium biguttatum with Rhizoctonia solani and other soil-borne fungi.

The mycoparasitic interactions of Verticillium biguttatum with Rhizoctonia solani and with a variety of other soil-borne fungi were investigated in dual cultures. V. biguttatum interacted with various soil fungi by appressed growth along the host hyphae and infrequent penetrations. Intracellular growth and subsequent sporulation, however, only occurred with R. solani, a few binucleate Rhizoctonia and Ceratobasidium spp., and Sclerotinia sclerotiorum. Effective mycoparasitism on sclerotia was restricted to those belonging to R. solani. Electron-microscopic observations revealed that V. biguttatum can penetrate the host cell with infection tubes. This process is probably mediated by enzymatic hydrolysis of the cell wall. Subsequently, trophic hyphae develop within the host cytoplasm, ultimately resulting in death of the host cell.

Host-Parasite Interactions↗

Studies on growth inhibition by lectins of Penicillia and Aspergilli.

It has previously been shown in our laboratory that wheat germ agglutinin (WGA) binds to Trichoderma viride and inhibits growth of this fungus. Here we report on the effect of WGA, soybean agglutinin (SBA) and peanut agglutinin (PNA) on Penicillia and Aspergilli. Binding of the lectins to the fungi was examined with the aid of their fluorescein isothiocyanate (FITC) conjugated derivatives. FITC-WGA bound to young hyphal walls of all species, in particular to the hyphal tips and septa, in agreement with the chitinous composition of the cell walls of the two genera. Hyphae of all species examined were labelled, though in different patterns, by FITC-SBA and FITC-PNA, suggesting the presence of galactose residues on their surfaces. Young conidiophores, metulae (of the Penicillia), vesicles (of the Aspergilli), sterigmata and young spores, were also labelled. The three lectins inhibited incorporation of [3H]acetate, N-acetyl-D-[3h]glucosamine and D-[14C]galactose into young hyphae of Aspergillus ochraceus, indicating interference with fungal growth. Inhibition of spore germination by the three lectins was also observed. Preincubation of the lectins with their specific saccharide inhibitors prevented binding and the inhibitory effects. We conclude that lectins are useful tools for the study of fungal cell surfaces, and may also serve as an important aid in fungal classification. The present findings also support the suggestion that one role of lectins in plants is protection against fungal pathogens.

Aspergillus↗

Direct studies of dikaryotization in Schizophyllum commune. I. Live inter-cellular nuclear migration patterns.

Compatible matings of Schizophyllum commune were performed on glucose-peptone-yeast extract medium appended with gelatin (18%) and studied by phase contrast microscopy during nuclear migration. Three categories of nuclear migration were observed. Type I involved a pulsatile jerking of the entire cytoplasmic contents of the hypha, changed direction periodically, and, during periods of cytoplasmic tranquility, the nucleus continued to migrate. Type II A migration of nuclei occurred in the absence of visible cytoplasmic flow. Both Type I and Type II A nuclear movements exceeded the hyphal growth rate by 10--20-fold. Type II B nuclear migration also occurred in the absence of visible cytoplasmic flow and the velocity was within the range of the hyphal growth rate. No specific organelles that were detected either directed or facilitated Type II A or Type II B nuclear movements. The nucleolus could either lead or trail relative to the direction of nuclear movement. Nuclear migration can be attributed to both cytoplasmic flow and self motility, depending upon the particular regions of the migration hypha in which it occurs.

Agaricales↗

Direct studies of dikaryotization in Schizophyllum commune. II. Behavior and fate of multikaryotic hyphase.

The growth, duplication and fate of multikaryotic hyphae bearing true clamp connections, as derived from compatible matings of Schizophyllum commune, were studied by phase contrast microscopy. The nuclei (N) of multikaryotic apices maintained a near central position during hyphal growth. True clamp connection formation occurred with near synchronous mitosis followed by septal synthesis across the clamp neck and main hyphal axis. Nuclear progeny after mitosis in a hexakaryon included 6 N in the apex, 1 N in the clamp and 5 N in the penultimate cell; the solitary nucleus in the clamp later entered the penultimate cell. Similar events occurred for clamp connection formation and mitosis in the trikaryon, quadrikaryon or pentakaryon, whether in the apex or primary branches. Nuclear content of the multikaryotic apex (2 N through 10 N) had no apparent effect on the rate of individual hyphal growth. Reduction of the nuclear number in a trikaryon occurred by long-term entrapment of a solitary nucleus in the clamp and subsequent outgrowth of the dikaryotic penultimate cell. Occasionally, more than one nucleus became entrapped in the clamp cell. The ephemeral nature of the multikaryon was indicated by the fact that older cultures appeared to be exclusively dikaryotic hyphae at the colony periphery.

Agaricales↗

Development of Aspergillus parasiticus and formation of aflatoxin B1 under the influence of conidiogenesis affecting compounds.

The influence of various inhibitors of hyphal growth, sporulation and biosynthesis of aflatoxin B1 in Aspergillus parasiticus NRRL2999 was studied. 6-Thioguanine, DL-ethionine, fluoroacetic acid and phenylboric acid, inhibitors of maturation of fungal conidiophores and of conidiogenesis, were added at various concentrations to malt extract agar. Lower concentrations of 6-thioguanine and DL-ethionine did not inhibit the growth of hyphae and the sporulation. Phenylboric acid reduced conidiogenesis more than hyphal growth. The yields of aflatoxin B1 were significantly reduced. Additions of fluoroacetic acid did not greatly affect the growth of hyphae but totally inhibited the production of conidia and concurrently significantly reduced the formation of aflatoxin B1. An interrelation between conidiogenesis and onset of secondary metabolism in A. parasiticus is evident.

Aflatoxin B1↗

Influence of manganese on morphology and cell wall composition of Aspergillus niger during citric acid fermentation.

Morphology and cell wall composition of Aspergillus niger were studied under conditions of manganese sufficient or deficient cultivation in an otherwise citric acid producing medium. Omission of Mn2+ (less than 10(-7) M) from the nutrient medium of Aspergillus niger results in abnormal morphological development which is characterized by increased spore swelling, and squat, bulbeous hyphae. Fractionation and analysis of manganese deficient cell walls revealed increased chitin and reduced beta-glucan contents as well as reduction of galactose containing polymers, as compared to cell walls from manganese sufficient grown hyphae. Addition of copper induced the same effect as manganese deficiency, both on morphology and cell wall composition. Addition of cycloheximide also produced a very similar type of morphology with increased chitin and reduced beta-glucan contents of the cell wall but its effect on galactose was less pronounced.

Amino Acids↗

Effect of temperature and temperature shifts on growth and branching of a wild type and a temperature sensitive colonial mutant (Cot 1) of Neurospora crassa.

Growth of a temperature sensitive colonial mutant (cot 1) of Neurospora crassa was compared with a wild type strain. The hyphal growth unit (the ratio between mycelial length and number of branches) of the wild type was not appreciably altered by temperature and there was a direct relationship between the specific growth rate (alpha) of these mycelia and their mean hyphal extension rate (E). The specific growth rate of cot 1 increased by about the same relative amount as the wild type between 15 degrees and 30 degrees C. Cot 1 grew and branched normally at 15 degrees and 25 degrees C but at 30 degrees C the hyphal growth unit and mean hyphal extension rate of the mutant mycelia were reduced. Thus, between 15-30 degrees C the ratio, E/alpha was constant for the wild type but not for cot 1. The effect of temperature and temperature shifts on extension zone length (2), extension zone expansion time (2t) and branching of leading hyphae of mature colonies were also studied. It is suggested that branching is governed by a mechanism which regulates the linear growth rate of hyphae; the cot 1 mutation may have a direct effect on wall extension or affect linear growth rate indirectly due to an influence on the transport of precursors to the tip.

Mutation↗

Antagonistic effects of Trichoderma harzianum on Pythium aphanidermatum causing the damping-off disease of tobacco in Nigeria.

Culture filtrates of Trichoderma harzianum Rifai have been found to inhibit zoospore germination, germ tube elongation and mycelial growth of Pythium aphanidermatum causing the damping-off disease of tobacco in Nigeria. Further, the invasion of the hyphae of P. aphanidermatum by hyphae of T. harzianum has been demonstrated in mixed cultures of the two fungi. The results of investigations on the role of T. harzianum as an agent of biological control of the damping-off disease in sterilized and unsterilzed soils are reported and discussed.

Antibiosis↗

Scanning electron microscopy on Microsporum audouini complex.

Several strains of Microsporum audouini, M. langeroni and M. rivalieri were observed by light and scanning electron microscopies. The macroaleuriospore shape and ornamentation were illustrated and compared for the 3 species as the pectinate hyphae for M. langeroni and M. rivalieri and the chlamydospores and particular hyphae for M. audouini and M. langeroni. The 3 species are morphologically very close but they are distinguishable from each other on the basis of morphology, geographic distribution and pathogenicity.

Dermatomycoses↗

Water as a substrate for the development of Paracoccidiodes brasiliensis mycelial form.

Two isolates of P. brasiliensis in the mycelial form were studied for their capacity to survive and grow in sterile distilled water (SDW). Inoculum for the experiments consisted of a spectrophotometrically-standardized suspension of washed and homogenized mycelial fragments; these had been obtained from 2-week old cultures grown in a synthetic medium (SM). Series of tubes with SDW and SM were incubated with the above suspension and kept stationary for 6 months at either 4 degrees C or room temperature (RT). Growth was measured by dry weight (DW) and turbidity (OD) determinations; additionally, CFU and ultrastructural appearance by transmission electron microscope (TEM) were assessed for one of the isolates. In general, cultures in SM at RT, grew exponentially after 2 weeks, becoming stationary for 7 weeks and then, declining abruptly. In SDW, fungal development was slow for 5 months when an increase in mass was recorded. When incubated at 4 degrees C, both SDW and SM cultures required longer time to develop but mass also increased. Morphologically, mycelial elements in SDW at RT exhibited increased lipid vacuoles and glycogen deposits but were otherwise normal up to 6 weeks when they presented the inter-hyphae-hyphae phenomenum. In SDW P. brasiliensis appears to utilize debris from its degenerated fungal partners to continue growing.

Colony-Forming Units Assay↗

Studies on the phylogenesis of pathogenic 'black yeasts'.

This paper deals mainly with the conidium ontogenesis and phylogenesis of black yeasts such as E. jeanselmei, E. gougerotii, E. dermatitidis and E. spinifera. The conidium ontogenesis of E. jeanselmei, E. gougerotii and E. dermatitidis was almost the same. One to five annellated tips were observed through a scanning microscope at the apices of conidiogenous cells, which were bottle- or jar-shaped. Annellations on the tips looked like fringes and the conidiogenous cells of these three species were annellides. Annellated projections occurred on hyphae and annelloconidia were also produced from them. Occasionally, secondary annellides occurred from primary ones. They looked like moniliform hyphae. Daughter conidia sometimes budded directly from mother cells. The shapes and sizes of the conidia of these species were very similar to each other. The conidium ontogenesis of E. spinifera was annellidic as well. However, a single annellated tip usually occurred on an annellide. The annellated tips of the fungus were long and more than 20 annellations were observed on their walls. The conidiogenesis of the four species of Exophiala is only annellidic. There were no differences in the biological examinations except KNO3 assimilation among these four species. The growth of E. jeanselmei and E. gougerotii was poor at 37 degrees C. The GC contents of E. jeanselmei 1171, E. gougerotii B-1800, E. dermatitidis MM-7 and E. spinifera DU-3342 were 54.6, 54.6, 56.6 and 59.7%, respectively.

Base Composition↗

An ultrastructural study of the invasion of Culex quinquefasciatus larvae by Leptolegnia chapmanii (Oomycetes: Saprolegniales).

Transmission electron microscopy of the invasion of Culex quinquefasciatus by Leptolegnia chapmanii confirmed that it is a primary pathogen and revealed several differences between penetration via the gut and penetration via the integument. The latter often involved aggregations of zoospores, appressoria-like swellings of the invasive hyphae, and lateral growth of hyphae between the epicutical and endocuticle. These features were not detected in the case of gut invasion, but hyphal septa at the point of entry were apparently peculiar to this route. There was no evident tissue specificity, and death presumably resulted from generalized destruction of tissues.

Animals↗

Cutaneous alternariosis: a rare chromohyphomycosis. Report of a case.

A 66-year-old woman presented 3 groups of scaly granulomatous lesions on the right forearm and elbow of six months duration. Direct examination of the scales showed hyphae and culture in lactrimel medium grew Alternaria sp. considered a contaminant. No improvement was observed after six months of treatment with oral griseofulvin. A second culture using half of a biopsy grew Alternaria sp. again and in histologic examination short pigmented fragments of hyphae surrounded by a granulomatous reaction were seen in the derma. The patient had marked improvement after treatment with erythromycin per os and miconazole topically; an atrophic scar remained as the only sign of the fungal infection.

Aged↗

The dysgonic strain of Microsporum canis.

The dysgonic strain of Microsporum canis is described. The colony consists of fascicles of very wide hyphae with short compartments and close branching. Lateral branches often themselves remain short and fail to branch, and many are reflexive, growing in the opposite direction to the parent hypha. Mycelium of three types was noted, dysgonic, apparently normal, and forms intermediate between the two. Change from one form to another occurred in a cyclial manner, the transition from dysgonic to apparently normal being abrupt while that in the reverse direction was gradual. The strain was extremely unstable, both in primary isolations from cats and in subculture. Numerous sectors of other readily recognisable strains were produced, including the typical M. canis and various atypical strains previously obtained from human sources. These strains rapidly submerged their parent colonies and were much more likely to be obtained by subculture from a primary isolation plate than was the dysgonic strain itself. The relationships of the various strains to each other in the saprophytic and parasitic phases are discussed.

Animals↗