Mycology Lecture Notes - Institute of Health Technology

INSTITUTE OF HEALTH TECHNOLOGY, DHAKA Department of Laboratory Medicine BSc in Health Technology (Laboratory)- 1 st Year MYCOLOGY Lecture No. 01(Introduction) By Sk. MIZANUR RAHMAN Lecturer, Mycology MS in Biotechnology & Genetic Engineering (UODA) MS in Microbiology (SUB)MYCOMOLOG By TGheOT OGF...

INSTITUTE OF HEALTH TECHNOLOGY, DHAKA Department of Laboratory Medicine BSc in Health Technology (Laboratory)- 1 st Year MYCOLOGY Lecture No. 01(Introduction) By Sk. MIZANUR RAHMAN Lecturer, Mycology MS in Biotechnology & Genetic Engineering (UODA) MS in Microbiology (SUB)MYCOMOLOG By TGheOT OGFTYBeBung iThah irotcmsrRm dx etbdctmdck Wr?SpSRr fCp SR Trtlmv OrpvRdld:k (etbdctmdck)w . Dm nrtc By C.E WM/hYL0 0hTWhY erpmPcrcg Wkpdld:k WC SR fSdmrpvRdld:k 1 urRrmSp GR:SRrrcSR: (LBih) WC SR WSpcdbSdld:k (CLf) The Fungi Learning outcomes - Students should be able to: • Establish familiarity with the scientific terminology peculiar to mycology • Describe the dimorphic nature of the pathogenic fungi used in making a clinical diagnosis. • Emphasize the eukaryotic nature of the fungi. • To explore the nature of the pathogenesis of fungal infections. • To gain familiarity with the classification of medically- important fungi. • To develop an understanding of the nature and mode of action of anti-fungal agents Recommended Textbooks Theory • Medical Mycology and Human Mycoses ES Beneke and AL Rogers Star Publishing Company 1996 Belmont CA Laboratory • Identifying Filamentous Fungi: A Clinical Laboratory Handbook St-Germain G and Summerbell R Rogers Star Publishing Company 1996 Belmont CA Additional Reading • - Introductory Mycology Alexopoulos CJ Mims CW Blackwell M Fourth Edition John Wiley &Sons Inc 1996 New York NY • - Medical Mycology KJ Kwon-Chung and JE Bennett Lea &Febiger 1992 Philadelphia PA • - Medical Microbiology A Laboratory Manual: Section II WG Wu Third Edition Star Publishing Company 1995 Belmont CA Medical Mycology Iceberg What is a Fungus ? • Eukaryotic – a true nucleus • Do not contain chlorophyll • Have cell walls • Produce filamentous structures • Produce spores Species of Fungi • 100,000 – 200,000 species • About 300 pathogenic for man Fungal Morphology and Structure Eukaryotic organisms, distinguished by a rigid cell wall composed of chitin and glucan , and a cell membrane in which ergosterol is substituted for cholesterol as the major sterol component. Fungal taxonomy relies heavily on morphology and mode of spore production Fungi may be unicellular or multicellular . The simplest grouping based on morphology divides fungi into either yeast or mold forms . Features of Fungi and its value in our life: The fungi are a ubiquitous and diverse organisms, that degrade organic matter. Fungi have heterotrophic life; they could survive in nature as: Saprophytic: live on dead or decaying matter Symbiotic: live together and have mutual advantage Commensal: one benefits and other neither benefits nor harmed. Parasitic : live on or within a host, they get benefit and harm the other. Fungi mainly infect immunocompromised or hospitalized patients with serious underlying diseases. The incidence of specific invasive mycoses continues to increase with time The list of opportunistic fungal pathogens likewise increases each year “It seems there are no non-pathogenic fungi anymore ! “ This increase in fungal infections can be attributed to the ever- growing number of immunocompromised patients. Characteristics of fungi A. eukaryotic , non- vascular organisms B. reproduce by means of spores ( conidia ), usually wind- disseminated C. both sexual (meiotic) and asexual (mitotic) spores may be produced, depending on the species and conditions D. typically not motile , although a few (e.g. Chytrids) have a motile phase. E. like plants, may have a stable haploid & diploid states F. vegetative body may be unicellular ( yeasts ) or multicellular moulds composed of microscopic threads called hyphae . G. cell walls composed of mostly of chitin and glucan. More Characteristics of Fungi H. fungi are heterotrophic ( “other feeding,” must feed on preformed organic material), not autotrophic ( “self feeding,” make their own food by photosynthesis). - Unlike animals (also heterotrophic), which ingest then digest, fungi digest then ingest. -Fungi produce exoenzymes to accomplish this I. Most fungi store their food as glycogen (like animals). Plants store food as starch. K. Fungal cell membranes have a unique sterol, ergosterol , which replaces cholesterol found in mammalian cell membranes L. Tubule protein —production of a different type in microtubules formed during nuclear division. Structure • The body of fungi is termed thallus (non- reproductive) • The thalli of yeast are small, globular and are single celled • The thalli of mold are composed of long, branched tubular filaments called hyphae. Structure • The thallus of a mold is composed of hyphae intertwined to form a tangled mass called mycelium . Mycotic Diseases (Four Types) 1. Hypersensitivity – Allergy 1. Mycotoxicosis – Production of toxin 1. Mycetismus (mushroom poisoning) – Pre-formed toxin 1. Infection The Clinician Must Distinguish Between: • COLONIZATION • FUNGEMIA • INFECTION EYE SKIN UROGENITAL TRACT ANUS MOUTH RESPIRATORY TRACTPortal of Entry • SKIN • HAIR • NAILS • RESPIRATORY TRACT • GASTROINTESTINAL TRACT • URINARY TRACT EYE SKIN UROGENITAL TRACT ANUS MOUTH RESPIRATORY TRACT Colonization Multiplication of an organism at a given site without harm to the host EYE SKIN UROGENITAL TRACT ANUS MOUTH RESPIRATORY TRACT Infection Invasion and multiplication of organisms in body tissue resulting in local cellular injury .. Classification of fungi They are classified by several methods: 1- Morphological classification 2- Systematic classification 3- Clinical classification Fungal Morphology Y e a s t Hyphae (threads) making up a mycelium M o u l d Encapsulated yeastCryptococcus neoformans • Unicellular fungi • Fission yeasts divide symmetrically • Budding yeasts divide asymmetrically Saccharomyces and CandidaYeasts Figure 12.3 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 Molds • Multicellular, tubular structures (hyphae) • Hyphae can be septate (regular crosswalls) or nonseptate (coenocytic) depending on the species (grow by apical extension) – Vegetative hyphae grow on or in media (absorb nutrients); form seen in tissue, few distinguishing features – Aerial hyphae contain structures for production of spores (asexual propagules); usually only seen in culture • The fungal thallus consists of hyphae; a mass of hyphae is a mycelium. Molds Figure 12.2 Moulds are multicellular organisms consisting of threadlike tubular structures called Hyphae that elongate by apical extension. Hyphae are either: Coenocytic: hollow and multinucleate Septate: divided by partitions or cross-walls Hyphae form together to produce a mat-like structure called a Mycelium . Vegetative hyphae, grow on or under surface of culture medium, Aerial Hyphae : project above surface of medium Aerial H. produce Conidia (asexual reproductive elements) Conidia can easily airborne and disseminate the fungus. Many medical fungi are termed dimorphic because they exist in yeast and mould forms. Septate hyphae Non-Septate hyphae Myceliu m Dimorphic Fungi • Growth as a mold or as a yeast • Most pathogenic fungi are dimorphic fungi • At 37o C yeast-like • At 25 o C mold-like • Can also occur with changes in CO 2 • Fungi grow differently in tissue vs nature/culture; often dictated by temp • Some fungi are dimorphic depending on environmental conditions • These organisms produce both yeast-like and mold-like thalli • Many are pathogenic • Candida albicans Dimorphism Figure 12.4 Dimorphism Many pathogenic fungi are d i m o r p h i c , forming moulds at ambient temperatures but yeasts at body temperature. Clinical Classification of Mycoses • Cutaneous • Subcutaneous • Systemic • Opportunistic Cutaneous Mycoses Skin, hair and nails Rarely invade deeper tissue Dermatophytes Subcutaneous Mycoses • Confined to subcutaneous tissue and rarely spread systemically. The causative agents are soil organisms introduced into the extremities by trauma Systemic Mycoses • Involve skin and deep viscera • May become widely disseminated • Predilection for specific organs Opportunistic Fungi Ubiquitous saprophytes and occasional pathogens that invade the tissues of those patients who have: • Predisposing diseases: Diabetes, cancer, leukemia, etc. • Predisposing conditions: Agammaglobulinemia, steroid or antibiotic therapy. • Systemic mycoses Deep within body • Subcutaneous mycoses Beneath the skin • Cutaneous mycoses Affect hair, skin, nails • Superficial mycoses Localized, e.g., hair shafts • Opportunistic mycoses Caused by normal microbiota or fungi Fungal Diseases (mycoses) DIAGNOSIS Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probes Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probes Direct Microscopic Observation • 10 % KOH • Gentle Heat KOH Wet Mount Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probes Skin Testing (DERMAL HYPERSENSTIVITY ) Use is limited to : – Determine cellular defense mechanisms – Epidemiologic studies Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probes FUNGI ARE POOR ANTIGENS Fungal Serology Antibodies • Latex Agglutination IgM • Immunodiffusion IgG • EIA IgG & IgM • Complement Fixation IgG Most serological tests for fungi measure antibody. Newer tests to measure antigen are now being developed ANTIGEN DETECTION PRESENTLY AVAILABLE  Cryptococcosis  Histoplasmosis  Aspergillosis Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probes DIRECT FLUORESCENT ANTIBODY CAN BE APPLIED TO 1. HISTOLOGIC SECTIONS 2. CULTURE • Viable organisms • Non-viable organisms Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6. Culture 7. DNA probe Inflamatory Reaction • Normal host – Pyogenic – Granulomatous • Immunodeficient host – Necrosis Polymorphic Nuclear Leukocytes Giant Cell GMS (Gomori Methenamine Silver Stain) Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6 . Culture 7. DNA probes Isolation Media SABOURAUD DEXTROSE AGAR (pH ~ 5.6) • Plain • With antibiotics • With cycloheximide IncubationTemperature • 370 C - Body temperature • 25 0 C - Room temperature Diagnosis 1. Wet Mount 2. Skin test 3. Serology 4. Fluorescent antibody 5. Biopsy and histopathology 6 . Culture 7. DNA probes DNA Probes • Rapid (1-2 Hours) • Species specific • Expensive