Mycology I Lecture Notes (PDF)

MICROBIOLOGY | TRANS 7 LE Mycology I Margarita V. Eduardo | 10/04/2024 | Version 1...

MICROBIOLOGY | TRANS 7 LE Mycology I Margarita V. Eduardo | 10/04/2024 | Version 1 02 OUTLINE Sterols (+); Type of sterols Sterols (-) I. Generalities of Fungi IV. Rare Superficial Mycoses Ergosterol A. Morphology A. Trichosporon Beigelii Survival at pH 5 Can survive Can’t survive B. Cell wall B. Piedraia Hortae High osmotic C. Laboratory Diagnosis C. Hortaea Werneckii Resistant Sensitive pressure D. Types of Fungal Disease V. Subcutaneous Mycoses II. Cutaneous Mycosis/ A. Sporotrichosis Peptidoglycan Dermatophytosis B. Chromomycosis Chitin (N-acetyl A. Microsporum C. Mycetoma Cell wall (N-acetyl glucosamine, B. Trichophyton VI. References glucosamine) N-acetyl muramic C. Epidermophyton VII. Review Questions acid) D. Clinical Materials for VIII. Formative quiz Lysosomes (+) (-) Dermatophytosis IX. Appendix III. Common Superficial A. MORPHOLOGY Mycoses MONOMORPHIC A. Malassezia Furfur/ 📋 Globusa Yeast → Unicellular, reproduce by budding Must Lecturer Book Previous Youtube → Starting with budding yeast, when grown in culture, they ❗️ Know 💬 📖 📋 Trans 🔺 Video will resemble a colony of bacteria. SUMMARY OF ABBREVIATIONS AMB Amphotericin B 5Fc Flucytosine KOH Potassium hydroxide PAS Periodic Acid Schiff Figure 1. Microscopic and Macroscopic Morphology SDA Sabouraud Dextrose Agar of Yeast [Lecturer’s ppt] LEARNING OBJECTIVES 📋 Hyphal/Filamentous ✔ Compare the structures of eukaryotes to those of → Multicellular molds prokaryotes and their functions. → Two (2) types of hyphae: ✔ Understand the factors needed for their growth and ▪ Septate hyphae - has septa/cross walls/divisions. survival. ▪ Aseptate hyphae (coenocytic hyphae) - do not have ✔ Describe the different morphology of fungi & cellular septa, or cross walls. structures. − Each cell has a nucleus. ✔ Discuss the mechanisms of action of antifungal drugs. − If seen macroscopically, you will see them in ✔ Diagnose fungal diseases as to their location, 💬 molds that are composed of hyphae and epidemiology, characteristics of diagnostic fungal spores. structures. − If they are intertwined, they are called mycelium. ✔ Interpret results of laboratory diagnostic tests. ✔ Give appropriate drugs for treatment of fungal infections. ✔ Recommend appropriate preventive measures for fungal diseases. I. GENERALITIES FOR FUNGI “Mycos” is a Greek word that means “mushroom.” Importance of fungi → Production of food, wine, etc. Figure 2. Morphology of Hyphae [Lecturer’s ppt] ▪ Aspergillus wetii - making ketchup DIMORPHIC FUNGI ▪ Alkaline protease - for industrial detergent → Makes use of yeast 📋 Both yeast and filamentous forms , but their growth depends on the temperature of the environment. ▪ Saccharomyces cereviceae - making bread/wine → Filamentous at 25oC to 30oC ▪ Saccharomyces ellipsoides - making alcohol (grapes) → Yeast form at 37oC → Some molds are being used as antibiotics Examples: ▪ Cephalosporium - making cephalosporins → Coccidioides immitis ▪ Penicillium nonatum - making penicillin ▪ If seen in the soil, they are in the form of Table 1. Differences between Fungi (Eukaryotic) and Bacteria arthroconidia or spores; however, if found inside (Prokaryotic). human tissues, they are in spherules. → Histoplasma capsulatum FUNGI BACTERIA ▪ If present in bird feces, they are in the form of O2 Aerobic Aerobic/Anaerobic tuberculate macroconidia; however, if found in 20-50 μm or humans, they are yeast cells located intracellularly. Size 1-5 μm bigger LE 2 TG #14 | A. Carig, J. Casimiro, M. Casingal, C. TE | R. Carlos, R. Carreon, L. AVPAA | Castillo PAGE 1 of 15 TRANS 7 Castañeda, C. Castaño, A. Castillo, M. Castro, Casacop, K. Chen VPAA | Cambas R. Catibog, P. Ceralde, D. Chan MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo Special Stains (Permanent stains) → Periodic Acid Schiff (PAS) ▪ Visualizes Candida albicans in esophagus → H&E stain Laboratory Culture (2 to 6 weeks at room temperature) → Sabouraud Dextrose Agar (SDA) ▪ To diagnose specific fungi ▪ When checking the culture, look at both the front and reverse sides of the bottle in order to diagnose correctly. → Brain Heart Infusion Agar ▪ For sterile specimen → Mycosel Agar ▪ For non-sterile specimen Serology (Systemic fungi) Figure 3. Dimorphic Fungi [Lecturer’s ppt] → To evaluate the status of the disease B. FUNGAL CELL WALL → Skin Test Beta-glucans ▪ For Histoplasmosis → Site of action of antifungal drug. → Latex Agglutination Test Chitin ▪ For Cryptococcosis → Polysaccharide composed of long chains of → Agar Double Diffusion Test N-acetylglucosamine. ▪ For Coccidioidomycosis Ultraviolet Lamp (Wood’s Lamp) ❗️ Ergosterol → Site of action of antifungal drug → To detect fluorescent compounds produced by fungi in or on human tissue. → Fluorescent on infected areas (either on skin or hair) Radiology Figure 4. Cross-section of the Fungal Cell Wall [Lecturer’s ppt] Figure 5. Candida albicans in esophagus under MECHANISMS OF ANTIFUNGAL DRUGS Periodic Acid Schiff [Lecturer’s ppt] Table 2. Antifungal Drugs and their MOA. D. TYPES OF FUNGAL DISEASES Mechanism of Action Antifungal Drugs Fungal Allergies Binds to ergosterol and Systemic: Amphotericin B → Molds release spores that grow in warm, damp, and disrupt fungal membrane (AMB) dark areas (house). ▪ E.g., rooms that have not been used or cleaned for a Topical: Nystatin long time. Inhibits ergosterol Topical: Azoles, → If inhaled, there will be an immediate hypersensitivity synthesis Terbinafine, Tolnaftate response due to fungal spores. Mycotoxicosis Systemic: Azoles → Alpha amanitin → Phalloidin Inhibits β-glucan synthesis Caspofungin (brand name ▪ Inhibits cellular RNA polymerase which prevents “Cancidas”) mRNA synthesis Inhibits DNA synthesis Flucytosine (5Fc) ▪ Ex. − Amanita phalloides (“Death cap” mushroom) Disrupt mitotic spindle by Griseofulvin − Amanita caesarea (Hallucinogenic) binding to tubulin → Aflatoxin C. LABORATORY DIAGNOSIS ▪ Most potent hepatotoxins KOH ▪ E.g., Aspergillus flavus - may cause hepatic → Utilizes fresh specimen carcinoma → Can visualize hyphae Mycoses → 10% KOH: for skin and hair → Growing actively on human tissues → 20% KOH: for nails → Superficial, dermatophytosis, subcutaneous, etc. MICROBIOLOGY Mycology I PAGE 2 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo ❗️ 📖 ❗️ MUST KNOW [Jawetz] B. TRICHOPHYTON GENERAL CHARACTERISTICS Mycoses refers to infections caused by fungi. Most fungi are obligate or facultative aerobes. Hair, skin, and nails are affected. Mycoses with the highest incidence are candidiasis and Examples include: dermatophytosis. → Trichophyton mentagrophytes (Zoophilic) → T. rubrum CONCEPT CHECKPOINT 1. What are the portals of entry of fungi? 📋 → T. concentricum T. MENTAGROPHYTES a. Superficial b. Cutaneous Zoophilic (kangaroos) c. Subcutaneous When cultured: On SDA, white fluffy culture can also be d. Systemic or Endemic observed. e. Opportunistic Under the Microscope: f. All of the above → Many microconidia (small spores) → Coiled hyphae can be observed ANS: → Few macroconidia 📖 1. F. All of the choices are based on Clinical Classification. C. EPIDERMOPHYTON See table in the Appendix for examples [Jawetz] GENERAL CHARACTERISTICS Skin and nails are affected. II. CUTANEOUS MYCOSIS / DERMATOPHYTOSIS Anthropophilic Table 3. Classification of Mycoses According to Site Affected or Extent of Involvement. EPIDERMOPHYTON FLOCCOSUM Classification Site of Involvement This fungi can cause: Superficial and Limited to the epidermis and → Tinea pedis Cutaneous dermis → Tinea cruris Infection penetrates beneath → Tinea corporis Subcutaneous → Onychomycosis the skin When cultured: Disseminated to internal Deep or Systemic → Front: Suede-like (brown) colony organs → Reverse: Deep yellow brown Individuals who have Microscopically: Opportunistic predisposing conditions → 2-3 clusters of Macroconidia (immunocompromised) → No microconidia The structures affected by Dermatophytosis are the Nails, Hair, and Skin ❗️MUST KNOW📖 ❗️ [Jawetz] → The common substance that is present in these Tip to remember what the 3 Genera affects: structures is Keratin. Microsporum, Trichophyton, Epidermophyton → Keratin has 2 virulence factors: TrichophytoN and EpidermophytoN ends with N, ▪ Keratinase - When released from the specimen, it which you can attribute to Nails. triggers itching. Only MicroSporum has an S, which you can use to ▪ Transferrin - Extracts iron from the host cell. remember that it only affects the skin and hair (not Three (3) sources of fungi: nails). → Zoophilic TINEA PEDIS → Geophilic Also known as athlete's foot. → Anthropophilic Common in temperate climate Three (3) Genera: The word "tinea" comes from Latin, meaning "worm," and → Microsporum "pedis" is the Latin word for "foot." → Trichophyton The infections frequently appear as ring-shaped, worm-like → Epidermophyton patches on the skin, which is why they are often called A. MICROSPORUM ringworm. GENERAL CHARACTERISTICS Epidermophyton floccosum Microsporum affects the hair and the skin. → Causes the Moccasin type in the sole of the foot. Examples of microsporum include: Trichophyton rubrum → M. gypsum → Prefers the moist/wet environment of the interdigital → M. audouinii spaces causing macerated skin between toeweb. → M. canis (Zoophilic) Transmitted via: → Desquamated skin scales, which contain hyphae. M. CANIS → Substrates include: Zoophilic ▪ Carpets Can be contracted from cats. ▪ Matting When cultured: On SDA, white fluffy mycelium can be observed. Under the microscope: → Spindle shaped macroconidia (with 5-15 cells) can be observed. → Few microconidia MICROBIOLOGY Mycology I PAGE 3 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo TINEA UNGUIUM TINEA CORPORIS Figure 6. Parts of the Nail and Different Types of Figure 9. Symptoms on the Neck Area (Left) and Onychomycosis Based on Location [Lecturer’s ppt] Submammary Area (Right) [Lecturer’s ppt] Nail bed - where fungi gain access, preferably moist. Symptoms on the neck area are caused by Microsporum canis → Following contact with infectious kitten Symptoms found on the sub-mammary area are caused by E. floccosum. TINEA CAPITIS Also known as scalp ringworm. Usually common in Latin America and in the Middle East Zoophilic Figure 7. Mild, Medium, and Severe Infections → Caused by Microsporum canis of the Nail Bed [Lecturer’s ppt] Caused by Trichophyton rubrum (anthropophilic) Types of nail bed infections: → Superficial type ▪ Onychomycosis (Tinea unguium) − Mild infection: White restricted patches or pits − Medium infection: Different colors, almost yellowing of the nail → Invasive type ▪ Hyperkeratosis − Severe Infection: Invasive type where the nail is thickened, brittle, and elevated. − The hyphae of the fungi can be seen when the area is scraped. TINEA CRURIS Figure 10. Scalp Ringworm Present on a Also known as “Jock itch” Child’s Head [Lecturer’s ppt] Mostly in tropical countries. → Symptoms Anthropophilic ▪ More inflammatory May be caused by T. rubrum or E. floccosum ▪ Scaly ▪ Obvious hair loss ▪ Itchy lesion Anthropophilic → Caused by E. floccosum → Child to child (before prepubertal stage) transmission Figure 8. Bilateral Erythema (Left) and Tinea Manifestations (Right) of Tinea Cruris [Lecturer’s ppt] Symptoms → Bilateral erythematous lesion of the inner thighs → Tinea of the buttocks caused by T. rubrum Space intentionally left blank Figure 11. Swollen Black Dot on the Scalp [Lecturer’s ppt] MICROBIOLOGY Mycology I PAGE 4 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo → Symptoms ▪ Minimal scaling ▪ Hair are broken at scalp level, leaving a swollen black dot ▪ Lesion clears at puberty Figure 15. Cutaneous Lesions caused by Contaminated Razors [Lecturer’s ppt] D. CLINICAL MATERIALS FOR DERMATOPHYTOSIS COLLECTION NAIL SCRAPINGS Figure 12. Electron microscopy of infected hair shaft showing Nail scrapings from the nail bed will be put on a black arthrospores. Figure A-B shows hair shaft cuticle layers that cardboard to create better contrast for viewing. were seriously destroyed with a large part being lost, exposing the fibril cortex inside which many arthrospores were noted. Figure 16. Nail Scrapings are put Against a Black Cardboard [Lecturer’s ppt] If there is no available mycology laboratory, nail scrapings/samples can be mailed to a specialized mycology lab. → Fungi will survive transit. Figure 13. Electron microscopy of infected hair shaft showing SKIN SCRAPING arthrospores. Figures C-D are high magnifications of the Steps: previous examples, which show the cuticle layers completely → Remove any ointment. destroyed with the residual cortex fully filled with → Specimen is collected from the edge of the lesion. arthroconidia caused by the fungi. DIAGNOSIS Wood’s lamp → A diagnostic tool that uses long-wave ultraviolet (UV) light. → The fungus may glow if the infected scalp is viewed under a Wood’s light. Figure 17. The Red Arrow points to the Area of Collection for Skin Scraping [Lecturer’s ppt] HAIR Steps: → Hair shaft is collected. → Specimen is mounted into methylene blue with KOH. → Do microscopy. Two (2) types of infection: → Ectothrix: spores are located outside the hair shaft. Figure 14. Area of the Infected Scalp viewed under Wood’s Lamp, showing the Glowing Fungi [Lecturer’s ppt] TINEA BARBAE Anthropophilic Caused by Trichophyton rubrum Possible substrates: → Towels → Razors Figure 18. Ectothrix Infection on KOH mount with Methylene Blue [Lecturer’s ppt] MICROBIOLOGY Mycology I PAGE 5 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo ▪ Positive for Wood’s Lamp Test ▪ E.g., M. canis → Endothrix: spores are located inside the hair shaft. Figure 22. Culture of Trichophyton tonsurans from a case of Endothrix Tinea Capitis [Lecturer’s ppt] Figure 19. Endothrix Infection on KOH mount with III. COMMON SUPERFICIAL MYCOSES Methylene Blue [Lecturer’s ppt] A. MALASSEZIA FURFUR / GLOBUSA GENERAL CHARACTERISTICS ▪ Negative for Wood’s Lamp Test ▪ E.g., T. tonsurans Part of the normal adult skin flora → It is a lipophilic yeast DIAGNOSIS Microscopic characteristics: 1. Direct Microscopy: use KOH with parker ink → Short, angular, hyphal forms up to 8um in diameter Skin scraping → Clusters of thick-walled, round, budding yeast → Utilizes 10% KOH ▪ Sometimes called “spaghetti meatballs” arrangement → Identify hyaline septate hyphae Figure 23. Malassezia furfur under the microscope [Lecturer’s ppt] TINEA VERSICOLOR (PITYRIASIS VERSICOLOR) Figure 20. Skin Scraping specimen under Also known as “an-an” in Filipino the Microscope [Lecturer’s ppt] Present as hypopigmentation in people with dark skin Nail scraping tones. → Utilizes 20% KOH → Lighter in color → Identify hyphae breaking up into arthroconidia → Often appear to coalesce → (+) Woods Lamp → Fluoresce under UV light Present as hyperpigmentation in Caucasians → Darker in color → Often appear to be well-demarcated → (+) Woods Lamp → Fluoresce under UV light Figure 21. Nail Scraping specimen under the Microscope [Lecturer’s ppt] 2. Culture in SDA → Most commonly used for fungi. → Observe and describe front and reverse side of the culture colony (e.g., white, cottony, brown) Figure 24. Tinea versicolor appears as hypopigmentation in people with dark skin tones [Lecturer’s ppt] MICROBIOLOGY Mycology I PAGE 6 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo Figure 28. Malassezia folliculitis as follicular papules and pustules on the upper trunk and neck [Lecturer’s ppt] FUNGAEMIA Figure 25. Tinea versicolor appears as hyperpigmentation in Neonates → Catheter-acquired people with lighter skin tones [Lecturer’s ppt] → Percutaneous intravenous central catheter SEBORRHOEIC DERMATITIS → M. furfur is capable of fungal biofilm formation Grow in areas with rich supply of sebaceous glands (e.g. → From the contaminated catheter, they can enter the scalp, forehead, ears, face) bloodstream Lesions → Red, covered with greasy scales Adult patient → Those undergoing lipid replacement Treatment: Topical imidazole therapy LABORATORY DIAGNOSIS 10% KOH WITH PARKER IN MOUNT Key features: → Spherical yeast cells → Short pseudohyphae → “Spaghetti meatball” arrangement Figure 26. Seborrhoeic dermatitis on scalp and face DANDRUFF Mildest Figure 29. M. furfur in 10% KOH with parker ink mount CULTURE Add fatty substrate (e.g. olive oil) or use Dixon’s Agar → You may also use Sabouraud Dextrose Agar You will see a colony that looks like a typical bacterial colony Figure 27. Dandruff [Lecturer’s ppt] MALASSEZIA FOLLICULITIS Follicular papules and pustules on the upper trunk and neck Host predisposing factors: → Oily skin → Provoked by hormonal influences → Obesity → Pregnancy → Stress or fatigue → Systemic illness, including: Figure 30. M. furfur in Dixon’s Agar [Lecturer’s ppt] ▪ Diabetes mellitus ▪ Immune deficiency MICROBIOLOGY Mycology I PAGE 7 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo IV. RARE SUPERFICIAL MYCOSES V. SUBCUTANEOUS MYCOSES A. TRICHOSPORUM BEIGELII (YEAST) Source of etiologic agents Also known as white piedra → Soil Infects the hair → Decayed Plants → Seen in the axilla, genitalia, head → Organic Materials → Appears as soft yellow nodules around the hair shaft Transmission: Direct inoculation of a fungus into the skin, 📋 often by a splinter or other trauma Key Concepts: → May be due to environmental molds associated with vegetation and soil → Traumatic implantation or traumatic inoculation; frequently chronic but rarely spread to deeper tissues → Thorns A. SPOROTRICHOSIS “Rose Gardener’s Disease” Etiologic agent: Sporothrix schenkii Grows on contaminated: → Dead plant materials → Thorns Figure 31. Trichosporum beigelii [Lecturer’s ppt] → Barbs → Sphagnum moss B. PIEDRAIA HORTAE (AGENT) Also known as black piedra Infects the hair → Scalp hairs are surrounded by a dense, black concretions → Small nodules contain spores (not yeast) Figure 34. Spotothrix schenkii [Lecturer’s ppt] Dimorphic fungus Figure 32. Piedraia hortae [Lecturer’s ppt] → Converts from hyphal growth to yeast cells within the host C. HORTAEA WERNECKII (YEAST) 25-30 C (filamentous); 37 C (yeast) Also known as tinea nigra/palmaris At 25C they can be seen as moist, wrinkled and folded Can be in yeast or hyphae form surface Infects the palms of hands Microscopy: → Appear as typical brown-black scaling macules (looks → Clusters of ovoid conidia (25-30C) like iodine on your palms) Laboratory diagnosis: 💬 → Cigar-shaped yeast cells (37C) When grown at 37C, their colonies are seen similar to the colony of bacteria which are cigar-shaped → 10% KOH of Skin Scraping with Methylene Blue 📋 ▪ Appear as brown to black hyphae Clinical Manifestation → Culture on Sabouraud Dextrose Agar (SDA) Characterized by nodular lesions of cutaneous or ▪ Appear as black, wrinkled yeast-like colonies subcutaneous tissues and adjacent lymphatics that → Under the microscope suppurate and drain ▪ Appear as colored yeast cells → Cutaneous Sporotrichosis Treatment: Whitfield Ointment ▪ Painless nodules ▪ Palpable and ulcerate discharging serous purulent fluid → Lymphocutaneous Sporotrichosis ▪ Elevated nodules developing along the regional lymphatics of the forearm → Drug of Choice: Itraconazole Laboratory Diagnosis → Specimen: tissue biopsy → Microscopy: GMS, PAS ▪ Specimen: tissue secretions (round, budding yeast cells) Figure 33. Hortaea werneckii [Lecturer’s ppt] ▪ Stain: GMS, PAS MICROBIOLOGY Mycology I PAGE 8 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo 📋 Asteroid bodies: approximately star-shaped with rays of eosinophils materials radiating from a central yeast-like cells B. CHROMOMYCOSIS “color”, pigment Agents → Phialophora verrucosa (red) ▪ Humid forest (Brazil) → Cladophialopora cariioni (green) ▪ Drier climate (South Africa) Figure 38. Extremities with chromoblastomycosis[Lecturer’s ppt] 💬 → Fonsecaea pedrosoi (brown) looks like a dark mushroom → Fonsecaea compacta 📋 Microabscesses resembling black dots or cayenne pepper develop on the surface → Mimics other diseases: ▪ Leprosy ▪ Syphilis ▪ Leishmaniasis ▪ Blastomycosis Laboratory Diagnosis → Specimen: tissue biopsy → Microscopy: KOH, H&E Figure 34. Different agents causing chromomycosis[Lecturer’s ppt] Highest incidence in countries with high rainfall → Southern Africa, Far East Japan, West Pacific Microscopy: Figure 39. Sclerotic cells[Lecturer’s ppt] → Look for sclerotic cells present as round, thick-walled, cigar-colored structures Figure 36. Hyphae (left); Sclerotic cells (right)[Lecturer’s ppt] 💬Microscopy of the colony: Brown, thick-walled, 💬areiewed multi-septate cells with brown hyphae V from the tissues/lesions: sclerotic bodies which round, brown, multi-septate cells Figure 40. Laboratory diagnosis of chromomycosis[Lecturer’s ppt] Clinical Manifestations 💬 In 25-30C: You can culture and look at the color of 📋 → Crusted verrucous wart-like lesion 💬 In tissues or at 37C, you will see them as sclerotic the colony in front and do microscopy 📋 Elevate about 1-3 cm above surrounding tissue Resembles courses of bricks in arrangement with both horizontal and vertical septa (muriform spores) bodies Treatment: Surgical excision of the lesion to prevent local dissemination C. MYCETOMA “Madura Foot” 📋 → First seen in Madura, India “Myco” - fungus; “oma” - tumor → Swollen, tumor-like areas with sinuses that drain 📋 through multiple sinus tracts 📋 Chronic granulomatous infection 📋 [Lecturer’s ppt] Typical site is on the extremities Figure 37. Nodular form of chromoblastomycosis May be due to several fungi (Eumycetoma, Actinomyces, → Cauliflower-like appearance or Nocardia) → Usually one leg is involved MICROBIOLOGY Mycology I PAGE 9 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo CASE SCENARIO Cultures A 30-years-old, male presents with the multiple → Seen in different colors discharging sinuses on the dorsum of the left foot from → Color of the colony, description of the colony help last one and half years. Initially it started as skin color identify specific fungi painless nodules. Later, these nodules become pustules and draining sinuses. The purulent and hemorrhagic material come out from the sinuses, presence of granules Figure 43. Eumycetoma cultures [Lecturer’s ppt] 3 most common etiologic fungal agents that can cause eumycetoma → Madurella mycetomatis Figure 41. Mycetoma [Lecturer’s ppt] ▪ Common in mexico, africa, india ▪ Most common among the 3 → Characteristics: → Exophiala jeanselmei ▪ Multiple draining sinuses → Pseudallescheria boydii − present for one and a half years = chronic ▪ Pustules & draining sinuses Characteristics of mycetoma (madura foot) ▪ Presence of granules → Tumefaction (swelling) → Draining sinus tracts TYPES OF MYCETOMA → Eumycotic granules ▪ Brown granules for Madurella mycetomatis ▪ White granules for Madurella grisea ▪ Color of granules will help identify specific fungi → Sometimes fungi can also infect the bones, not only the subcutaneous tissue Figure 42. Types of Mycetoma [Lecturer’s ppt] 1. Actinomycetoma Actinomyces Israelii → filamentous bacteria Figure 44. Characteristics of Eumycetoma [Lecturer’s ppt] → gram (+) bacteria, rods 2. Eumycetoma LABORATORY DIAGNOSIS Fungi Specimen: tissue biopsy Madurella mycetomatis Microscopy: use KOH to look for hyphae → most common → Thin long-branched hyphae with well-defined walls → mostly seen in African countries (Mexico, Africa, India) → microscopy: hyphae (not rods) Space intentionally left blank Figure 45. KOH of granules [Lecturer’s ppt] MICROBIOLOGY Mycology I PAGE 10 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo Specimen: Tissue biopsy SUMMARY Microscopy: H&E stain → fungal hyphae (red) Figure 46. H & E stain [Lecturer’s ppt] Specimen: Tissue biopsy Microscopy:Grocott’s Methenamine Silver (GMS) stain → thin long-branched hyphae (black) Figure 49. Summary of superficial mycoses[Lecturer’s ppt] Tinea versicolor: remember the spaghetti meatballs arrangement for diagnostic features Tinea nigra palmaris: dark pigmented hyphae – looks like the palm is stained in iodine Figure 47. GMS stain [Lecturer’s ppt] Specimen: Tissue biopsy Culture: Sabouraud’s dextrose agar (SDA) → look at the characteristic colony in front and the reverse to identify specific fungi Figure 50. Summary of subcutaneous mycoses[Lecturer’s ppt] For subcutaneous mycoses, look at the tissue form Figure 48. Sabouraud’s dextrose agar [Lecturer’s ppt] III. REVIEW QUESTIONS TREATMENT 1. Which of the following statements on fungi is TRUE? Griseofulvin a. Fungi have a cell membrane with the presence of Ketoconazole sterols Surgical excision b. Fungi lack mitochondria → if cannot be treated c. Fungi are capable of photosynthesis d. None of the above 2. A 35-year-old male presents with an itchy, red, scaling rash on his foot. You suspect dermatophytosis. To confirm the diagnosis, you perform skin scrapings for direct microscopy and culture. Which of the following Space intentionally left blank is the correct diagnostic approach to confirm dermatophyte infection? A. Prepare skin scrapings using 20% KOH, and look for hyaline septate hyphae under the microscope. MICROBIOLOGY Mycology I PAGE 11 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo B. Prepare skin scrapings using 10% KOH with Parker 3. B. A patient with beard infection was caused by a ink, and identify hyaline septate hyphae. Trichophyton species, not Epidermophyton floccosum. C. Culture the scrapings on Sabouraud Dextrose Agar Other options Tinea corporis (ringworm), Tinea pedis (SDA) and describe the colony's microscopic appearance (athlete's foot), and Tinea cruris (jock itch) are caused by only. Epidermophyton Floccosum. D. Nail scrapings should be performed using 10% KOH to 4. A. Heterotrophic fungi requires nutrients from other identify hyphae breaking into arthroconidia. organisms for metabolism. 3. Which of the following options is NOT caused by 5. C. SDA, Brain heart infusion agar, and Mycosel are all Epidermophyton Floccosum? culture media for fungi. SDA supports the growth of fungi a. A patient with itchy red rash in the neck while restricting the growth of bacteria. Brain heart infusion b. A male patient with infection around the beard area agar cultivates fastidious pathogenic bacteria. Mycosel c. A badminton player with athlete's foot inhibits bacteria and other saprophytic molds. d. A patient with jock itch 6. A. Tinea Versicolor (Pityriasis Versicolor) are pigmented 4. Which of the following properties explains why fungi patches on the skin caused by Malassezia furfur. It is are capable of degrading organic compounds into commonly known as “An an” in Filipino. compost? 7. A. All of the choices produce microconidia, except A. a. Heterotrophic 8. B. False since the goal of treatment is not to eradicate but b. Facultative aerobic to reduce the cutaneous population to commensal levels. c. Non-photosynthetic 9. B. False. At 25°C, Sporothrix schenkii grows in its d. Eukaryotic filamentous form, with clusters of ovoid conidia. It converts 5. Which of the following culture media CANNOT be to cigar-shaped yeast cells at 37°C. used for fungi? 10. C. Symptoms refer to pityriasis versicolor (light a. Sabouraud dextrose agar (SDA) discoloration) b. Brain heart infusion agar c. Chocolate agar V. REFERENCES d. Mycosel Jawetz, Melnick, & Adelberg's Medical Microbiology. 28th edition. 6. Also known as “An an” in Filipino Lecturer’s PPT. 2024 Previous Transes [2022,2023,2024,2025,2026] a. Pityriasis versicolor b. Seborrhoeic versicolor c. Tinea Folliculitis d. Fungaemia 7. Which anthropophilic dermatophyte produces only macroconidia and has no microconidia? a. Epidermphyton b. Microsporum c. Trichophyton 8. The goal of treatment in pityriasis versicolor is to eradicate Malassezia from the skin. a. True b. False 9. Sporothrix schenkii, the etiologic agent of "Rose Gardener’s Disease," grows as cigar-shaped yeast cells at 25°C. a. True b. False 10. A 20 year old male consulted via telemedicine because of mild itching in his trunk and shoulders of one week duration associated with scaling and light discoloration patches in those areas. Wood’s lamp test was positive while KOH demonstrated yeast cells and long hyphae. What is the most likely condition of this patient? a. White piedra b. Black piedra c. Pityriasis versicolor d. Tinea nigra ANS: 1. A. Fungi have a cell membrane with the presence of sterols (ergosterols). 2. B. Option B is used in a direct microscopy. In Choice A, it must be 10% KOH. In Choice C, using SDA should observe and describe the front and reverse side of the culture colony. In Choice D, Nail scrapings should utilize 20% KOH. MICROBIOLOGY Mycology I PAGE 12 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo VI. FORMATIVE QUIZ 📖📋 Question & Choices Answer & Rationale Ergosterol is the main component of fungal cell membranes 1. Structure of fungi where most antifungal drugs take (humans = cholesterol). It is absent in animals, so it is the action: target of selective toxicity. A – Beta-glucans is also the site of action of antifungal drugs, A. Beta Lactams C not Beta-lactams. B. Chitin B – Chitin is part of the fungal cell wall but not the site of action C. Ergosterol of antifungal drugs. D. Both A & C 2. A 28-year-old patient from a rural area presents The usual form of Coccidioides immitis is arthroconidia or with a history of cough, fever, and weight loss. spores in the soil, but if these are found inside the human After conducting a biopsy of lung tissue, tissue, they are in spherules. microscopy reveals intracellular yeast cells. The patient reports exposure to bird droppings in the The Histoplasma capsulatum is in the form of tuberculate area. Based on your knowledge of fungal macroconidia in birds, but if these are found inside the human morphology, which of the following fungi is most tissue, they are yeast cells located intracellularly. likely responsible for this infection, and what is its dimorphic form? C A – spherule form in human tissue C – tuberculate macroconidia form in birds A. Coccidioides immitis, filamentous form in human D – arthroconidia or spore form in the soil tissue B. Histoplasma capsulatum, yeast form in bird feces C. Histoplasma capsulatum, yeast form in human tissue D. Coccidioides immitis, spherule form in soil 3. A 50-year-old male is brought to the emergency In Mycotoxicosis, department after consuming wild mushrooms. He presents with severe gastrointestinal distress and Phalloidin inhibits cellular RNA polymerase which prevents liver failure. Laboratory studies indicate mRNA synthesis. hepatotoxicity, and the patient’s family reports that Ex: Amanita phalloides (“Death cap” mushroom) and Amanita the mushrooms were identified as Amanita caesarea (Hallucinogenic) phalloides. Which of the following is the most likely mechanism behind this patient’s symptoms? Aflatoxin - Most potent hepatotoxins. Ex: Aspergillus flavus - may cause hepatic carcinoma A. Aflatoxin, produced by Aspergillus flavus, causes B hepatocellular carcinoma. A – Aflatoxin toxin is not found in the mushroom Amanita B. Phalloidin, from Amanita phalloides, inhibits phalloides. cellular RNA polymerase and prevents mRNA C – Amanita caesarea produces Phalloidin, and it’s not related synthesis. to the case because the mushrooms identified were Amanita C. Aflatoxin, produced by Amanita caesarea, causes phalloides. hallucinations and liver toxicity. D – Instead of Phalloidin, Aflatoxin is the most potent D. Phalloidin, produced by Aspergillus flavus, is the hepatotoxin, and it is not applicable to the case. most potent hepatotoxin. 4. A 25-year-old female presents with an itchy, The case is describing Microsporum canis, which is a zoophilic scaly rash on her arm after adopting a stray cat. dermatophyte that commonly affects both the hair and skin. It A skin scraping shows spindle-shaped is often contracted from animals, particularly cats. macroconidia under the microscope. What is the most likely fungal organism causing her condition? D A. Trichophyton mentagrophytes B. Candida albicans C. Epidermophyton floccosum D. Microsporum canis 5. Which statement regarding the laboratory B – Simplest and fastest method is Direct Microscopy. 1-2 identification of Fungi is correct? drops of KOH 10% or 20% on fresh specimen (skin scraping B or tissue samples) for 5-10 minutes → dissolves any tissue A. All fungal growths can be correctly identified by cells making your highly resistant and highly refractory direct visual examination due to the different growth fungal cell wall more visible MICROBIOLOGY Mycology I PAGE 13 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo forms A – Culture is more sensitive than direct examination, most B. Direct microscopy using 10% KOH staining is a of the time, but it takes a while, so do the faster methods first common diagnostic tool then do culture for a more definitive fungal identification. C. Fungi cannot be cultured as they behave C – Dimorphic fungi differences lie in their in vitro forms or differently in vitro and in vivo tissue forms. Sporotrichosis in vitro or in tissue leads to D. Fungal specimens do not require immediate small budding yeast. processing due to delayed degradation D – KOH 10% or 20% on fresh specimen for 5-10 minutes → dissolves any tissue cells making your highly resistant and highly refractory fungal cell wall more visible VII. APPENDIX MICROBIOLOGY Mycology I PAGE 14 of 15 MICROBIOLOGY | LE 2 Mycology I | Margarita V. Eduardo MICROBIOLOGY Mycology I PAGE 15 of 15