Dexter H. Howard

Mechanisms of resistance to fungal infections

In the past year, significant advances have been made in our understanding of immune defenses to fungal infections, which may be instrumental in the development of rational approaches to immunodiagnosis and therapy of these infections. The highlights have been the result of the direct application of advances in molecular biology and basic immunology, particularly cytokine research, leading to improved definition of fungal antigens and increased understanding of the roles of functionally distinct T-cell subsets, the activity of which may be either host-protective or disease-promoting.

Multiple chromosomal and phenotypic changes in spontaneous mutants of Candida albicans

Previous studies have revealed the occurrence of multiple chromosomal alterations among spontaneous colony form mutants and clinical isolates of Candida albicans. In this report we show that such karyotype alterations are also seen in spontaneous and induced non-germinative mutants of the fungus. To determine if phenotypic changes other than colony form and microscopic morphology accompanied the rearrangements of the electrophoretic karyotype, we studied the following characteristics of the non-germinative and some of the colony form mutants: formation of pseudohyphae, chlamydospore production, germ tube formation, colony morphology, auxotrophy, growth at various temperatures, and colony morphology and pigment formation on selected media (bismuth sulphite and Phloxine B). We established that phenotypic and karyotypic variability among spontaneous, non-germinative mutants was no different than such variability among spontaneous colony form mutants. Thus, non-germination may represent another phenotypic consequence of genomic instability in C. albicans. The variability in different phenotypic attributes that occurred amongst the mutants was not associated with any given karyotype. Moreover, neither the low nor the high phenotypic variabilities observed were explained by the relatively high number of alterations in a limited number of chromosomes.

Thermotolerance and the Heat-shock Response in Candida albicans

At elevated temperatures, yeast cells of Candida albicans synthesized nine heat-shock proteins (HSPs) with apparent molecular masses of 98, 85, 81, 76, 72, 54, 34, 26 and 18 kDa. The optimum temperature for the heat-shock response was 45 degrees C although HSPs were detected throughout the range 41-46 degrees C. Protein synthesis was not observed in cells kept at 48 degrees C. Yeast cells survived exposure to an otherwise lethal temperature of 55 degrees C when they had previously been exposed to 45 degrees C. The thermotolerance induced during incubation at 45 degrees C required protein synthesis, since protection was markedly reduced by trichodermin. Mercury ions induced a set of three stress proteins, one of which corresponded in size to an HSP, and cadmium ions evoked one stress protein seemingly unrelated to the HSPs observed after temperature shift.

Iron gathering by zoopathogenic fungi

Iron is a metal required by most microorganisms and is prominently used in the transfer of electrons during metabolism. The gathering of iron is, then, an essential process and its fulfillment becomes a crucial pathogenetic event for zoopathogenic fungi. Iron is rather unavailable because it occurs on the earths surface in its insoluble ferric form in oxides and hydroxides. In the infected host iron is bound to proteins such as transferrin and ferritin. Solubilization of ferric iron is the major problem confronting microorganisms. This process is achieved by two major mechanisms: ferric reduction and siderophore utilization. Ferric reductase is frequently accompanied by a copper oxidase transport system. There is one example of direct ferric iron transport apparently without prior reduction. Ferric reduction may also be accomplished by low molecular mass compounds. Some fungi have evolved a process of iron acquisition involving the synthesis of iron-gathering compounds called siderophores. Even those fungi that do not synthesize siderophores have developed permeases for transport of such compounds formed by other organisms. Fungi can also reductively release iron from siderophores and transport the ferrous iron often by the copper oxidase transport system. There is a great diversity of iron-gathering mechanisms expressed by pathogenic fungi and such diversity may be found even in a single species.

Proline Uptake in Candida albicans

L-Proline entered both mycelial and yeast cells of Candida albicans by an active transport system of high specificity at low (less than 0.1 mM) external concentrations of substrate. The apparent Km value of this system was 0.1 mM for both types of cells, while the V value was 4 nmol min-1 (mg dry wt)-1 for mycelial cells and 1.4 nmol min-1 (mg dry wt)-1 for yeast cells. At L-proline concentrations greater than 0.1 mM, the amino acid appeared to enter both morphological forms by diffusion as well as active transport. As saturation was approached diffusion became increasingly important. The higher uptake rate of mycelial cells seemed not to be the result of an inducible system. The optimal pH and temperature for transport of L-proline were 7.0 and 37 degrees C, respectively. Sodium azide and the proline analogues sarcosine and L-azetidine-2-carboxylic acid inhibited L-proline uptake, while L-thiazolidine-4-carboxylic acid was less effective. The active transport system was highly specific for L-proline since neither ammonium ions, which inhibit the general amino acid transport system of fungi, nor 16 different amino acids interfered substantially with uptake.

Ethanol Tolerance and the Induction of Stress Proteins by Ethanol in Candida albicans

Ethanol is one of the products of the metabolism of glucose by Candida albicans. The amount produced is directly related to the concentration of glucose in the medium. The fungus utilizes ethanol as a sole source of carbon but is relatively intolerant of ethanol in its environment. Ethanol induces germ tube formation by blastoconidia of C. albicans. Germination was not seen under fermentation conditions even though the amount of ethanol produced was in the range form stress proteins that are similar to heat shock proteins. The possibility that stress proteins may regulate germ tube formation by C. albicans is discussed.

Synthesis of nucleic acids and proteins in the dimorphic forms of Candida albicans

Candida albicans grown in tissue culture medium 199 buds when incubated at 25°C. but produces filaments when incubated at 37°C. Macromolecular synthesis by budding and filament producing blastospores of C. albicans grown at these 2 temperatures of incubation was studied. Cells incubated at 25°C. reproduced by budding with a generation time of 2 hr. The dry weight, protein and DNA increased 2-fold as the cells doubled in number while the RNA content of such cells increased about 3·2-fold. Ninety-five per cent of cells incubated at 37°C. produced filaments in 3 hr. A net increase in protein of 49% and in RNA of 94% was detected within the first 2 hr. of incubation at 37°C. By 4 hr. incubation protein had increased to 220% of the initial value and RNA to 300%. DNA content did not increase during the first 2 hr. of incubation but by 3 hr. the DNA content had increased to 50% of the initial value. Electrophoretic analysis of the water soluble proteins from budding and filament producing cells revealed that bot...

The morphogenesis of the parasitic forms of dimorphic fungi

Specific consideration has been given in this review to the sequence of morphologic events which characterize the transformation of dimorphic pathogenic fungi from the saprophytic to the parasitic phase of growth. Three general mechanisms of conversion have been described for those dimorphic fungi which exist in the tissues of a host as blastospores: (1) The hyphal cells of the saprophytic phase swell, assume a yeast-like shape and eventually fragment to form a mass of individual blastospores. (2) The hyphal cells (terminal or interstitial) bud directly to form blastospores. (3) The hyphal cells form conidiospores in a fashion characteristic of the saprophytic phase, but these conidia produce buds which eventually separate from the parent spore and become the blastospores of the yeast phase of growth. The hyphal cells of the saprophytic phase swell, assume a yeast-like shape and eventually fragment to form a mass of individual blastospores. The hyphal cells (terminal or interstitial) bud directly to form blastospores. The hyphal cells form conidiospores in a fashion characteristic of the saprophytic phase, but these conidia produce buds which eventually separate from the parent spore and become the blastospores of the yeast phase of growth. One or more of these morphologic transformations have been reported to occur inH. capsulatum, S. schenckii, B. dermatitidis andB. brasiliensis. Hence the species of fungi which occur in the tissues of a host as blastospores transform from the saprophytic phase of growth in much the same manner. InC. immitis andEmmonsia sp. the parasitic phase arises by direct swelling of the spore which characterizes the saprophytic phase. Thus the arthrospores ofC. immitis swell to form the spherules, and the conidiospores (aleurospores) ofEmmonsia sp. enlarge to form chlamydospores (adiaspores). The sclerotic cells characteristic of the tissue phase of the agents of chromoblastomycosis arise from a process resembling chlamydospore formation by the hyphal cells or more rarely by the conidiospores themselves.

Isolation of Bipolaris australiensis from a lesion of viral vesicular dermatitis on the scalp

Bipolaris australiensis is recorded as an opportunistic pathogen of humans. The fungus was recovered from a biopsy specimen of skin taken from an area of viral vesicular dermatitis on the scalp. Hyphal elements of B. australiensis were seen in the tissues. The taxonomy of the genera Bipolaris, Drechslera, Exserohilum and Helminthosporium is discussed with particular reference to published case reports.

The commensalism of Cryptococcus neoformans

Cryptococcus neoformans, was isolated from 3 of 561 sputum specimens. The fungus was not recovered from 162 samples of saliva or 310 fecal specimens. These data document the rarity of isolation of C. neoformans from human material but establish that it may be a commensal of the human nasopharynx under circumstances.