Different effectors of dimorphism in Yarrowia lipolytica

Yarrowia lipolytica is an ascomycete with biotechnological potential. In common media, the fungus grows as a mixture of yeast-like and short mycelial cells. The environmental factors that affect dimorphism in the wild-type strain, W29, and its auxotrophic derivative, PO1a, were analyzed. In both strains, pH was the most important factor regulating the dimorphic transition. Mycelium formation was maximal at pH near neutrality and decreased as pH was lowered to become almost null at pH 3. Carbon and nitrogen sources, namely glucose and ammonium, were also important for mycelium formation; and their effect was antagonized by some alternative carbon and nitrogen sources. Citrate was an important positive effector of mycelium growth. Anaerobic stress induced formation of mycelial cells. The importance of the protein kinase A pathway was suggested by the inhibition of mycelium growth by cAMP. We propose that the interplay of these factors regulates the adaptation of the fungus, to better exploit its natural ecological niches.     

Protein Profiling of the Dimorphic Pathogenic Fungus, Sporothrix schenckii

Sporotrichosis is a common cutaneous mycosis caused by the dimorphic fungus Sporothrix schenckii, which exhibits a temperature-dependent dimorphic switch. At 25°C, it grows in a mycelial phase, while at 37°C, it forms unicellular yeast cells. The formation of yeast cells was thought to be a requisite for the pathogenicity of S. schenckii. To identify fragments that might be related to morphogenesis, whole-cell proteins from the mold and early yeast stages of S. schenckii were analyzed using 2DE. Among thousands of protein molecules displayed, more than 300 showed a differential expression between the two phases. In particular, 24 yeast-specific proteins were identified using MALDI-TOF/MS. One of the most interesting proteins was a hybrid histidine kinase, DRK1, a global regulator of dimorphism and virulence in Blastomyces dermatitidis and Histoplasma capsulatum that was abundant in the yeast phase. Our study introduced a new approach to study dimorphism in S. schenckii, and the data may help us better understand the molecular mechanisms of phase transition.     

Cellular differentiation and host invasion by the rice blast fungus Magnaporthe grisea

This review describes current advances in understanding the biology of plant infection by the rice blast fungus Magnaporthe grisea. Development of the specialized infection structure, the appressorium, in M. grisea has recently been shown to be controlled by cell cycle progression and initiation of autophagic, programmed cell death in the fungal spore. Re-cycling of the contents of the fungal spore and peroxisomal fatty acid beta-oxidation are therefore important processes for appressorium function. Following entry to the host plant, new evidence suggests that M. grisea grows biotrophically within rice cells, bounded by the plant plasmalemma, and the fungus moves from cell-to-cell by means of plasmodesmata. Biotrophic proliferation of the fungus is likely to require secretion of effector proteins and suppression of host defences. Consistent with this, a component of the polarized exocytosis machinery of M. grisea is necessary for pathogenicity and also for induction of host defences in an incompatible interaction. Large-scale insertional mutagenesis is now allowing the rapid analysis of gene function in M. grisea, heralding a new approach to the study of this important fungal pathogen.     

Morphology-related effects on gene expression and protein accumulation of the yeast Arxula adeninivorans LS3

The dimorphism of the yeast Arxula adeninivorans LS3 is regulated by cultivation temperatures. Up to 42 degrees C the yeast grows as budding cells, which turn to mycelia at higher temperatures. To test whether the dimorphism is exclusively induced by high temperatures or also by other conditions, mutants were selected with an altered behaviour with respect to dimorphism. After mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine, five of 25,000 colonies formed a very rough surface consisting of mycelia at 30 degrees C, in contrast to the wild-type. These mutants allow temperature-mediated and morphology-related effects on gene expression and protein accumulation to be distinguished. Budding cells and mycelia showed different expression of genes encoding secretory proteins at the same temperature. Mycelia secreted two-fold more protein than budding cells, including the enzymes glucoamylase and invertase. This indicated that morphology, rather than temperature, is the decisive factor in the analysed processes.     

Pathogenic behaviour of Cephalosporium maydis and C. acremonium

Cephalosporium maydis infects young maize plants easily, but as plants age fewer are infected and none after approx. 50 days from sowing. The mesocotyl and seminal, fibrous and adventitious roots are attacked, especially when there is damage or much inoculum. Most penetration occurs where roots are elongating and emerge from the mesocotyl or from fibrous roots. At first the fungus grows superficially on roots, producing hyphae with short, brown, thick-walled, and swollen cells. After penetrating, the fungus spreads towards the xylem, where it grows slowly at first but after 5 weeks grows faster upwards.
C. acremonium causes black-bundle disease of maize. It seems to infect plants growing in unfavourable conditions but the details remain uncertain. The percentage of plants infected was not related to the amount of inoculum and the fungus may not be a primary parasite. The sterile culture filtrate of the fungus produces vascular discoloration and wilt of maize seedlings.     

Endostyle cell recruitment as a frame of reference for development and growth in the Urochordate Oikopleura dioica

In models of growth and life history, and in molecular and cell biology, there is a need for more accurate frames of reference to characterize developmental progression. In Caenorhabditis elegans, complete fate maps of cell lineage provide such a standard of reference. To be more widely applicable, reference frames should be easier to measure while still providing strong predictive capacity. Towards this aim, we have analyzed growth of the endostyle in the appendicularian Oikopleura dioica at the cellular level, and measured its response to temperature and food availability. Specifically, we test the hypothesis that age of a specific developmental stage in O. dioica can be predicted from the number of endostyle cells and temperature. We show that the endostyle grows by recruiting cells from the posterior tip into the lateral arms of the organ in an anterior-posterior orientation and that the rate of increase in lateral arm endostyle cells is temperature-dependent but unresponsive to nutritional intake. Endostyle cells therefore serve as an accurate and easily measured marker to describe developmental progression. Conceptually, such a method of characterizing developmental progression should help bridge life-history events and molecular mechanisms throughout organismal aging, facilitating cross-disciplinary understanding by providing a common experimental framework.     

The mold-specific MS8 gene is required for normal hypha formation in the dimorphic pathogenic fungus Histoplasma capsulatum

The dimorphic fungus Histoplasma capsulatum is the etiologic agent of one of the most common systemic mycoses of humans, histoplasmosis. In the environment, H. capsulatum grows in a differentiated mold form and shifts to an undifferentiated yeast form after mold fragments or spores are inhaled. This mold-to-yeast shift is required for disease. Little is known about the molecular biology of dimorphism in Histoplasma, and most studies have been directed toward yeast-specific genes. While it is important to examine the role of genes upregulated in the yeast morphotype, genes which are silenced in the yeast (i.e., mold-specific genes) may also play a critical role in dimorphism. To begin to examine this hypothesis, we report here the first misexpression and knockout analysis of a mold-specific gene in Histoplasma. The strongly expressed MS8 gene encodes a predicted 21-kDa protein extremely rich in glycine and glutamine. Forced expression of MS8 driven by the TEF1 promoter in yeast did not alter the yeast morphology at 37°C or mold formation at 25°C. Yeast expressing MS8 did exhibit clumping in liquid medium and formed “sticky” colonies on agar plates. Allelic replacement of MS8 was accomplished by a positive-negative selection procedure. ms8 knockout mutants formed apparently normal yeast at 37°C but gave rise to aberrant mycelia at 25°C. The mold colonies of the knockouts were less than half as large as normal, had a granular surface, produced a dark-red pigment, and formed short hyphae which were 40% wider with a distinctive twisted “zig-zag” shape.     

Studies of Didytmella bryoniae (Auersw.) Rehm: Development in the Host

Growing hyphae of Didymella bryoniae (Auersw.) Rehm are found in the stem tissue of cucumber, and the fungus can colonize nearly all types of tissue. The presence of the pathogen in the lignified elements suggests a spread of the disease up and down from the site of primary infection, even when the stem is strongly decomposed. The fungus dtsplays hyphal dimorphism in the infected tissue and a remarkable feature is stromatic-like structures in the host tissue close to fruiting bodies. Staining reveals no histological host reaction to fungal penetration of cell walls. The pathogen is characterized by the early development of its sexual stage.     

Candida albicans: genetics, dimorphism and pathogenicity.

Candida albicans is a dimorphic fungus that causes severe opportunistic infections in humans. Recent advances in molecular biology techniques applied to this organism (transformation systems, gene disruption strategies, new reporter systems, regulatable promoters) allow a better knowledge of both the molecular basis of dimorphism and the role of specific genes in Candida morphogenesis. These same molecular approaches together with the development of appropriate experimental animal models to analyze the virulence of particular mutants, may help to understand the molecular basis of Candida virulence.     

Sexual dimorphism of sublingual glands in tree shrews

On the basis of histochemical characteristics it was possible to demostrate a sexual dimorphism of the tree shrew sublingual gland. Although numerous staining methods for the demonstration of mucosubstances were used in this study, only methods for the demonstration of sulfated glycoproteins (sulfomucins) were effective in demonstrating the sexual dimorphism. Numerous sulfomucin-laden cells occurred in the mucous tubules and acini of female sublingual glands, but only rarely were such cells observed in sublingual glands of male animals. Neither duct cells nor demilune cells of secretory endpieces were involved in the sexual dimorphism. No morphological sexual dimorphism was noted in tree shrew sublingual glands.     

The Ontogeny of Sexual Size Dimorphism of a Moth: When Do Males and Females Grow Apart?

Sexual dimorphism in body size (sexual size dimorphism) is common in many species. The sources of selection that generate the independent evolution of adult male and female size have been investigated extensively by evolutionary biologists, but how and when females and males grow apart during ontogeny is poorly understood. Here we use the hawkmoth, Manduca sexta, to examine when sexual size dimorphism arises by measuring body mass every day during development. We further investigated whether environmental variables influence the ontogeny of sexual size dimorphism by raising moths on three different diet qualities (poor, medium and high). We found that size dimorphism arose during early larval development on the highest quality food treatment but it arose late in larval development when raised on the medium quality food. This female-biased dimorphism (females larger) increased substantially from the pupal-to-adult stage in both treatments, a pattern that appears to be common in Lepidopterans. Although dimorphism appeared in a few stages when individuals were raised on the poorest quality diet, it did not persist such that male and female adults were the same size. This demonstrates that the environmental conditions that insects are raised in can affect the growth trajectories of males and females differently and thus when dimorphism arises or disappears during development. We conclude that the development of sexual size dimorphism in M. sexta occurs during larval development and continues to accumulate during the pupal/adult stages, and that environmental variables such as diet quality can influence patterns of dimorphism in adults.     

Impact of morphogenetic effectors on the growth pattern and the lipid composition of the mycelium and the yeastlike cells of the fungus Mucor hiemalis

We investigated the growth and the cell lipid composition of the mycelium and of the yeast-like form of Mucor hiemalis VKMF-1431 obtained under aerobic conditions by treatment with the morphogenetic agents itraconazole, exogenous triacylglycerols (TAGs), and trehalose. The sporangiospores of a 20-day culture were inoculated on the medium with glucose. Under these conditions, the fungus produced both mycelium and yeast-like cells. It was established that, upon the germination of old (20-day) sporangiospores, the fungus predominantly used the mycelium development strategy in the presence of trehalose and TAGs. It was characterized by a low ratio between the two bulk membrane lipids (PEA/PC) and increased levels of PC and polyunsaturated fatty acids (FA). Compared to the mycelium, the yeast cell morphotype obtained on the medium with glucose was distinguished by an elevated PEA/PC ratio, lowered TAG, free sterol (FS) and esterified sterol (ES) levels, a decreased ES/FS ratio that correlated with the reserve sterol pool size, and a lowered content of unsaturated fatty acids (the linoleic and the ??-linolenic acid). These peculiarities of the lipid composition of yeastlike cells correlated with the intensity of yeastlike growth. Light and electron microscopy revealed differences between the above cell morphotypes. With itraconazole, yeast-like cells were characterized by the destruction of the endoplasmic reticulum membranes and formation of a large number of vacuoles. The suggestion was confirmed that the state/age of inoculum sporangiospores exerts an influence on the capacity for dimorphism in mucorous fungi such as M. hiemalis. The data obtained testify to an involvement of lipids in the process of adaptation to environmental factors and to their regulatory role in morphogenetic processes associated with the formation of alternative morphotypes of the mucorous fungus.     

Formation of organic acids by the fungus Cladosporium resinae in media containing n-alkanes]

The so-called fungus Cladosporium resinae that often occurs in oil fuels ane increases their acidity grows well at the expense of n-alkanes from C11 to C16. On the n-alkane containing media the fungus grows slowly and only under the stationary conditions. During the fungal cultivation on the media containing n-dodecane or kerosene the culture liquid shows acetic acid and other fatty acids, ketoacids (pyruvic and alpha-ketoglutaric) as well as citric and isocitric acids that dominate among nonvolatile acids. Upon nitrogen deficiency in the medium and comparatively good aeration the content of citric acids increases. The culture liquid of the fungus devoid from the mycelium and nonutilized n-alkanes can be used a a nutrient medium for different microorganisms.     

Role of the conidium in dimorphism of Blastomyces dermatitidis

Fine details of yeastlike cell development of Blastomyces dermatitidis from its conidium are described and illustrated by electron micrographs. When cultured in an enriched medium at 37 °C, conidia of two strains of B. dermatitidis readily underwent ultrastructural changes consistent with mycelial to yeast dimorphism. Although hyphal cells contained in the conversion cultures were observed consistently to undergo profound degenerative changes, the conidia rapidly germinated to give rise to short germ tubes which subsequently enlarged to form intermediate yeast mother cells (YMC). The wall of the germ tube arose from the innermost layer of the wall of the germinant. During the transition globoid osmiophilic inclusions of unknown origin and function were observed in vacuolated areas of the germ tube and YMC cytoplasm. Yeastlike daughter cells then budded from the intermediate YMC. Since transformation was readily accomplished under in vitro conditions favoring mycelial to yeast dimorphism, it is suggested that the conidium of B. dermatitidis represents the primary infective unit of this pathogenic fungus.     

Dimorphic mating-type chromosomes in the fungus Microbotryum violaceum

Fungi often mate as haploids, and sex chromosomes (i.e., mating-type chromosomes) that are dimorphic for their size or overall DNA content have never been reported in this kingdom. Using electrophoretic techniques for karyotype analysis, a highly dimorphic chromosome pair that determines mating compatibility is shown to occur in populations of the fungus Microbotryum violaceum. This substantiates the evolution of such dimorphism as a general feature associated with haploid determination of mating compatibility, which previously had been known only in haplodioecious plants (mosses and liverworts). Size-dimorphic sex chromosomes are present in a lineage of M. violaceum native to Europe, as well as a lineage native to North America. However, they are very different in size between these lineages, indicating either independent evolution of the dimorphism or a large degree of divergence since their isolation. Several DNA sequences that show sequence similarity to transposons were isolated from these sex chromosomes.     

The development and endophytic nature of the fungus Heteroconium chaetospira

The root endophytic fungus Heteroconium chaetospira was isolated from roots of Chinese cabbage grown in field soil in Japan. This fungus penetrates through the outer epidermal cells of its host, passes into the inner cortex, and grows throughout the cortical cells, including those of the root tip region, without causing apparent pathogenic symptoms. There are no ultrastructural signs of host resistance responses. H. chaetospira has been recovered from 19 plant species in which there was no disruption of host growth. H. chaetospira has a symbiotic association with Chinese cabbage. The fungus provides nitrogen in exchange for carbon. These associations are beneficial for the inoculated plants, as demonstrated by increased growth rate. When used as a preinoculum, H. chaetospira suppresses the incidence of clubroot and Verticillium yellows when the test plant is post-inoculated with the causal agents of these diseases. H. chaetospira is an effective biocontrol agent against clubroot in Chinese cabbage at a low to moderate soil moisture range and a pathogen resting spore density of 10(5) resting spores per gram of soil in situ. Disease caused by Pseudomonas syringae pv. macricola and Alternaria brassicae on leaves can be suppressed by treatment with H. chaetospira. The fungus persists in the roots and induces systemic resistance to the foliar disease.     

Tiny keratotic brown lesions on the interdigital web between the toes of a healthy man caused by Curvularia species infection and a review of cutaneous Curvularia infections

A 74-year-old healthy Japanese man had noticed an itchy lesion with two tiny thick brown scales on the fourth interdigital web of his right foot 3 months earlier. The brown fungus isolated from the scales had demonstrated brown, ellipsoidal, obovoid or clavate, 3-septate conidia. Morphologically, the fungus was identified as Curvularia sp., and was differentiated from other human pathogenic species of the genus, such as C. lunata, C. pallescens, C. trifolii, C. clavata, and C. brachyspora, by molecular analysis based on the DNA sequence data. The fungus grows well on keratotic materials (hairs, nails, and callus), which indicates that it might have the ability to infect the skin surface.