INTRODUCTION TO MEDICAL MYCOLOGY Lecture 14

Learning Outcome Able to define terms use in mycology Can describe basic characteristic of fungi Able to discuss fungal reproduction Able to describe mycoses Able to discuss laboratory examination for fungal infection.

Definitions Mycologists--scientists who study fungi Mycology--scientific discipline dealing with fungi Mycoses--diseases caused in animals by fungi Mykos = mycete = fungus

I. FUNGI Diverse group of heterotrophs. Many are ecologically important saprophytes (consume dead and decaying matter) Others are parasites. Most are multicellular, but yeasts are unicellular. Most are aerobes or facultative anaerobes. Cell walls are made up of chitin (polysaccharide). Over 100,000 fungal species identified. Only about 100 are human or animal pathogens. Most human fungal infections are nosocomial and/or occur in immunocompromised individuals (opportunistic infections). Fungal diseases in plants cause over 1 billion dollars/year in losses.

Characteristic of fungi Vegetative Hypha Composed of cells involved in catabolism and growth. Reproductive Hypha (aerial) Composed of cells involved in reproduction (produce spores).

General knowledge of the fungi Both sexual and asexual spore may be produced Store their food as glycogen (plant; starch) Fungi are heterotrophic organisms, lack of chlorophyll (plant; autotrophic)

General knowledge of the fungi Yeast : unicellular, 370C Budding Yeast – may produce a pseudohypa Fission yeast Mold : multicellular, hyphae, 250C Dimorphic fungi (thermally dimorphic fungi) : mold phase & yeast phase

Yeasts Facultative Anaerobes Fermentation : ethanol and CO2 Non-filamentous unicellular fungi Spherical or oval Reproduction: a) by fission, or b). By budding

Yeast Reproduction FISSION “even” reproduction, nucleus divides forming two identical cells, like bacteria BUDDING “uneven” reproduction, parent cell’s nucleus divides and migrates to form a bud and then breaks away

YEAST Unicellular Micr.: Oval to round (Diameter : 3-15 µm) Macr.: Pasty colonies (resemble bacteria)

MOULD Multicellular Micr.: . Hypha(e) (dia: 2-10 µm) . Spores / conidia. Macr.: .Surface texture: Cottony/ powdery/ wooly/velvety/granular/glabrous Pigmentation :obverse & reverse

General knowledge of the fungi Eukaryotic microorganisms Rigid cell walls: chitin, glucans, mannans Plasma membranes: ergosterol Lysine synthesis by L- amino adipic acid (AAA) pathway and other organisms synthesize lysine by diaminopimelic acid (DAP) pathway

Fungal Structure Thallus-”body” Mycelium – Molds & fleshy fungi have these structures Long filaments of cells (hyphae): Septate hyphae (cross wall) :most fungi Aseptate hyphae (coenocytic ) :no cross wall, continous mass with many nuclei . Mycelium – Abundance growth of aerial hyphae resulting a mass can be observed with unaided ayes

Fungal Classification Four groups of true fungi Zygomycetes (common bread mold—Rhizopus) Basidiomycetes (puffballs & common mushrooms) Ascomycetes (Dutch elm disease/rye smut) Deuteromycetes (fungi imperfection)

Classification ( con’t) : First three groups is based on their method of sexual reproduction 4th group, the Deuteromycetes, have NO sexual reproduction

Fungi-Taxonomic classification SEXUAL SPORE CLASS Zygospore----------Zygomycetes Basidiospore--------Basidiomycetes Ascospore----------Ascomycetes None/Unknown---- Deuteromycetes (“Fungi Imperfecti”)

Zygomycetes Asexual phase—Sporangium—bread mold (Rhizopus stolonifer) Sexual phase--- sporgangium ---shotgun fungus (lives on dung) it shoots its sporgangium explosively towards light or fly pathogen (Entomophthora muscae—--these types of fungi have been used as agents for biological control of insects

Basidiomycetes Basidiospore Examples: boletes, puffballs,smuts, stinkhorns and tooth fungi

Ascomycetes Asexual phase- Sexual phase (morels, lichens ) Conidiospores (Penicillium and Aspergillus) budding yeast Sexual phase (morels, lichens )

Life Cycle Fungal reproduce by 2 way Asexual Sexual Hyphae fragmentation Asexual spores Conidiosphore Arthroconidia Blastoconidia sporangiospore

Reproduction of Fungi 1. Sexual reproduction --Sexual spores 2. Asexual reproduction--Asexual spores 3. Parasexual reproduction--Genetic exchange

Life cycle Conidiosphore Spore that is not enclosed in sac

Life cycle Athroconidia Form fragmentation of hypae into single thick cells.

Life cycle Blastoconidia Buds coming from parents cells Chlamydoconium a variant which spore form by thickening and enlargement within hypae

Life cycle Sporangiospore Spore form inside sporangium (sac) at the end of sporangiophore

Asexual Spores Conidiospores Chlamydospores Sporangiospores Blastospores Arthrospore

Asexual spores ( con’t) Conidiospore Multiple (chains) or single spores formed at the end of an aerial hypha Not enclosed within a sac Aspergillus spp. Penicillium spp

Aspergillus sp.

Conidial Fungus reproduces by means of asexual spores called conidia Conidia vary greatly in shape, size and color Most of the common household molds & mildews are conidial fungi

Asexual spores ( cont) Another type of Conidiophore: Blastospores A bud coming off the parent cell Candida albicans

Chlamydospore Formed within hypha Thick-walled spore Candida albicans

Chlamydospores The chlamydospore is a method of producing a substantial resting spore very quickly Nutrient is shunted from adjacent cells into a preferred cell and it swells up, converts nutrient materials to oil droplets for efficient storage, then rounds off with a thick, often roughened outer wall for protection

Asexual spores Sporangiospores Hundreds formed within a sac (sporangium) at the end of an aerial hypha Rhizopus spp. Mucor spp Absidia spp

Types of asexual spores

SEXUAL Spores 1. Zygospore 2. Ascospore 3. Basidiospore 4. Oospore

Sexual spores Have three stages Plasmogamy- a haploid nucleus of donor cells Karyogamy- the (+) and (-) nuclei fuse to form a diploid zygote Meiosis – the diploid nucleus give rise to haploid nuclei (sexual spores) genetic recombinant

MYCOSES Superficial ( skin, hair, cornea) Cutaneous ( Dermatophytosis ) Subcutaneous True systemic (endemic) Opportunistic

Zygomycosis Disease caused by fungi that are classifiable as Zygomycetes Mucormycosis : Order Mucorales Entomophthoromycosis : Order Entomophthorales

Zygomycosis (cn’t) Presents as a spectrum of diseases, depending on the portal of entry and the predisposing risk factors of the patient: Rhinocerebral zygomycosis Pulmonary zygomycosis Gastrointestinal zygomycosis Cutaneous zygomycosis Disseminated zygomycosis

Material from the periorbital tissue of a woman with poorly controlled diabetes mellitus with facial and periorbital swelling due to zygomycosis (see right picture) is stained with periodic acid-Schiff stain (X 560). The material demonstrates the classic appearance of irregularly shaped broad hyphae with right-angle branching (arrow). http://www.emedicine.com/med/topic2438.htm

Laboratory to diagnosis of fungal infection Specimen collection and transport Specimen processing Direct examination Selection and inoculation of culture media Identification

LABORATORY DIAGNOSIS OF MYCOSES Direct microscopic examination Culture Serology

Specimen collection and transport must be material from the actual infection site must be carefully the contamination must be established for the best chance of recovery of causative microorganisms (optimal times)

Specimen collection and transport ( con’t) must be obtained to perform the culture or other techniques request (sufficient quantity) must be used to ensure optimal recovery of microorganisms obtain cultures before the treatment the culture container must be properly labeled

Specimen processing specimen should be examined as soon as possible direct examination : KOH mount Calcofluor white India ink culture media

Selection and inoculation of culture media Culture media for recovery of fungi from clinical specimens. PDA, Corn Starch Agar The recovery rate may be somewhat enhanced by using a variety of isolation media, considerations of cost, storage, incubator space and technologist time.

Initial observations in the study of fungus isolates 1.Appearance of the growth 2. Rate of growth 3. Colony pigmentation 4. Growth on media containing antifungal agents 5.Dimorphic fungi

Initial observations in the study of fungus isolates 1.Appearance of the growth - surface and reverse surface of colony were observed - delicate or hairlike hyphae 2. Rate of growth - saprophytes : 3-5 days - dimorphic fungi : 10 days or more - dermatophytes : 14 days or more

Initial observations in the study of fungus isolates 3. Colony pigmentation 4. Growth on media containing antifungal agents - most strains of the dimorphic fungi can grow - most strains of the rapidly growing saprobes are inhibited 5. Dimorphic growth - mold form (the environmental and infective form) ; ambient or room temperature (22-25 OC) - yeast form (invasive form) ; near body temperature (30-35 OC)

Preparation of mounts for study The tease mount Scotch tape preparation The microslide culture technique ( slide culture )

Terms useful in the examination of fungi hypha and pseudohyphae mycelium septate or aseptate (or coenocytic) hyphae vegetative mycelium aerial mycelium reproductive mycelium

Exercise List 4 classes of fungi List types of asexual spores Diferentiate between conidiospore with sporangiospores