Mycology Part 1 Outline PDF

MYCOLOGY PART 1 OUTLINE Hyphae - long strands of tube like structures (either aerial or vegetative) I. General Characteristics Mycelia - mass of hyphae A. Yeasts...

MYCOLOGY PART 1 OUTLINE Hyphae - long strands of tube like structures (either aerial or vegetative) I. General Characteristics Mycelia - mass of hyphae A. Yeasts vs. Molds B. Mycelia C. Hyphae D. Pigmentation E. Dimorphism II. Reproduction A. Asexual B. Sexual III. Taxonomy IV. Portal of Entry V. Mode Of Transmission MYCELIA VI. Fungal Culture Process A. Specimen Collection and Transportation Aerial Mycelia B. Direct Examination of Specimen responsible for wooly or fuzzy appearance of the C. Selection and Inoculation of Media molds D. Evaluation of Fungal Growth E. Serological Testing Vegetative Mycelia F. Antifungal Susceptibility Testing the ones that penetrates or extends under the culture media, which are then responsible for absorbing GENERAL CHARACTERISTIC FUNGI nutrients Eukaryotic (true nucleus) Chitin in the cell wall Forms of Vegetative Mycelia Ergosterol in the cell membrane Obligate aerobes 1. Antler Hyphae - Antler-shaped or has swollen tips Reproduction by means of spores, produced asexually 2. Racquet Hyphae - Has club shape appearance or sexually 3. Spiral Hyphae - Has spiral appearance Lack of chlorophyll 4. Rhizoids - Root-like appearance Lack of susceptibility to antibacterial antibiotics MOLDS VS YEAST Yeast single vegetative cells that typically form a smooth, creamy, bacterial like colony without aerial hyphae HYPHAE reproduce by budding or fission Septate Hyphae show frequent cross walls occurring perpendicular to the outer walls of the hyphae Sparsely Septate Hyphae have few cross-walls at irregular intervals. Coenocytic or Aseptate Molds fuzzy or woolly appearance because of the formation of mycelia  Molds: grows at room temperature (22-25°C) in ambient air Polymorphic fungi have both yeast and mold forms in the same culture PIGMENTATION Hyaline (Moniliaceous) hyphae REPRODUCTION non-pigmented or lightly pigmented Asexual Reproduction (Imperfect Fungi) Phaeoid (Dematiaceous) Hyphae Results in the formation of conidia (singular, conidium) darkly pigmented following mitosis Conidia is a asexual reproductive structure of a fungal spore Arthroconidia (singular, arthroconidium) a type of conidia that is formed by fragmentation of fertile hyphae as opposed to being formed by conidiogenous cells DIMORPHISM Dimorphic fungi include a mold phase and a yeast or spherule phase Yeast: grows at 37°C with increased concentration of CO 2 Types of Asexual Reproduction Chlamydospores Macroconidia produced by chlamydoconidia; is composed of terminal, multicellular and larger intercalary, and sessile Microconidia Smaller Sporangiospores enclosed in a sporangium Phialoconidia produced by phialide (vase shape structure) Sexual Reproduction (Perfect Fungi) Also called as "Plus and minus mating type" Requires two different hyphae that will fuse to form a zygospore Requires the joining of two compatible nuclei, followed by meiosis Blastoconidia produced through budding Teleomorph - A fungus that reproduces sexually. Anamorph - Asexual reproductive state of a fungus Pseudohyphae Synanamorphs - If more than one anamorph is present formed from budding (difference is that a hyphae is for the same teleomorph or when a single fungus tube-like, while pseudohyphae is rod-shaped) produces more than one anamorph Types of Sexual Reproduction COLONIZATION-multiplication of organism (without harm to the host) Ascospores INFECTION-multiplication of organism (brings harm and Produced by Phylum Ascomycota can result to injury) Basidiospores Produced by Phylum Basidiomycota MODE OF TRANSMISSION Zygospores 1. Inhalation (respiratory tract) Produced by Zygomycota or Mucorales 2. Inoculation through skin trauma/ mucous membrane (skinpenetration) 3. Ingestion FUNGAL CULTURE PROCESS Specimen Collection Specimen types Collect from area most likely infected (usually skin scrapings from lesions) Use sterile technique Keep specimen moist (important for fungi) Label container properly Transport right away Process right away Specimen Collection and Transport ΤΑΧΟΝΟMY 1. BLOOD Collection: collect as bacterial cultures; disinfect with 1. ASCOMYCOTA tincture of lodine. Sexual spores: Ascospores Use maximum amt, of blood recommended. Include Microsporum spp, Trichophyton spp, and Transport: ≤2h, RT/ $24h, RT Scedosporium boydii 2. CSF 2. BASIDIOMYCOTA Collection: collect as bacterial cultures; at least 3ml in a Presence of clamp connections sterile container, but as much CSF as possible is Occur at the septations in the vegetative hyphae and preferred. are easily visible under a light microscope. Should be concentrated by centrifugation before 3. MUCORALES inoculation. Produce profuse, gray to white, aerial mycelia If >5ml filtration is recommended. Presence of hyaline hyphae Transport: $15 min, RT/ $24h, RT Sparsely septate hyphae Never refrigerate Asexual reproduction: Sporangiophores and 3. STERILE FLUIDS Sporangiospores Such as synovial fluids, synovial fluid, pericardial fluids, Sexual Reproduction: Zygospores and peritoneal fluids 4. FUNGI IMPERFECTI Collection: collect as bacterial cultures; Imperfect fungi or Deuteromycota Should be concentrated by centrifugation before Contains the largest number of organisms that are inoculation. causative agents of mycoses Use sediments for inoculation No mode of sexual reproduction has been identified; Transport: ≤15min, RT/ $24h, 4 C they have nosexual cycle 4. BONE MARROW Collection: Prepare site for surgical incision. PORTAL OF ENTRY Heparinized bone marrow should be plated directly 1.Skin onto media at bedside. 2. Hair Transport: ≤15min, RT/ $24h, 4 C 3. Nails 5. CATHETER 4. Respiratory Tract Collection: Collect as bacterial culture. 5. Gastrointestinal Tract 5cm distal tip of catheter into sterile tubes with few 6. Urinary Tract drops of sterile water or saline. Transport: ≤15 min, RT/ $24h, 4 C  DIRECT MICROSCOPIC EXAMINATION 6. EAR, EXTERNAL Collection: Firmly rotate swab in outer ear canal Uses mounting medium to hold the specimen in place Transport: $15min, RT/ $24h, 4 C Glass slide - Provides rapid report Transport immediately because swab samples can dry Specific morphologic characteristics provide a clue to out fast genus of organism. 7. EYE Might provide evidence of infection despite negative A. CORNEAL SCRAPING cultures Collection: Inoculate scrapings directly onto media and slides for staining TEMPORARY MOUNTS B. VITREOUS FLUID Collection: Needle aspiration then direct inoculation KOH (KOH PREPARATION) Transport:$15min, RT/ $24h, RT 10-20% KOH solution embedded with skin, nails and 8. HAIR tissue. Collection: With sterile forceps collect 10 12 hairs with KOH breaks down keratin and skin layer to see more shaft intact easily any fungi present in the specimen Make sure that you include the roots Gentle heat increase rate of clearing Transport placed in dry container or envelope $15min, Modified KOH: using DMSO dimethyl sulfoxide RT/s24h, RT To facilitate more rapid breakdown of cellular debris 9. NAILS w/o requiring heating Collection: disinfect with 70% alcohol before the surface is scraped. Can be nail scraping or cuttings; complete nail. Deeper scraping: KOH preparation, inoculate media Transport: placed in dry, clean container or envelope. $72h, RT 10. PROSTATIC FLUID Collection: Bladder emptied followed by prostatic massage. Direct inoculation onto media Transport: $15min, RT/ $24h, RT 11. RESPIRATORY SPECIMEN Collection: Ist morning specimen, deep cough. If the patient cannot obtain sputum nebulizer can be TEMPORARY MOUNTS used. Oropharyngeal: direct smear & culture KOH with Calcofluor White Nasal sinus: Plated directly. A drop of calcofluor white (fluorescent dye) can be Transport: ≤2h, RT/ ≤24h, RT added. 12. SKIN Dye binds to polysaccharides in the chitin Collection: disinfect with 70% alcohol before the Structure appears as blue- white surface is scraped. Fungal element = apple green or blue white depends Scrape surface at the margin of lesions. on the filter of the microscope Transport:placed in dry, clean container, $72h, RT 13. TISSUE BIOPSIES Collection: Surgical collection Transport: placed in clean container; add few drops of sterile saline to keep moist ≤15min, RT/ ≤24h, RT 14. URINE Collection: early morning specimen. Midstream collection technique Transport.. Wide mouth container; container, ≤15min, RT/≤24h, 4 C 15. VAGINA Collection: as bacterial culture; Transport:. Swab transport ≤2h, RT/ $24h, RT TEMPORARY MOUNTS PERMANENT MOUNTS Lactophenol blue PAS (Periodic Acid Schiff) to use a combination of ingredients to kill organisms, Differentiation of fungi scrapings of tissue preserve fungal structures, and stain the cell walls of Carbohydrates in fungus wall stain purplish red fungi: H&E slides may be restained by this when fungus wall Phenol: Acts as a disinfectant and kills invisible organisms Fungal element = Magenta Lactic acid: Preserves the fungal structure and Background color = Pink or green clears the tissue Glycerol: Prevents drying Cotton blue: Stains the chitin in the cell wall of fungi PERMANENT MOUNTS Gram Stain All fungi are Gram positive Stain employs crystal violet and ammonium oxalate Acid-Fast Stain Useful for differentiation of Nocardia from Actinomyces Hyphae of Nocardia partially acid fast or acid fast (red) TEMPORARY MOUNTS India Ink / Nigrosin Used to examine CSF for the presence of encapsulated yeast Crytococcus neoformans Fungal element = yeast with clear halo Background = black PERMANENT MOUNTS Gomori Methenamine Silver Nitrate Fungal element - Black Background = Green Giemsa Used primarily to detect Histoplasma capsulatum in bone marrow or blood. Fungal element= Purple blue yeast with clear halo (capsule) Background = Pink purple PERMANENT MOUNTS Acridine Orange Fungal element = Green fluorescent; therefore needs the use of a fluorescent microscope to view Epithelial cells = Orange