pmmv unit 3 slg

cell wall

Rigid, chitin (β-1,4-linked N-acetylglucosamine polymers), glucans (β-1,3 & β-1,6-D-glucans), & mannoproteins. no peptidoglycan- antibac abx R 

cell membrane

primary sterol- ergosterol (unlike cholesterol in mammalian cells), target for antifungal agents (polyenes bind, azoles inhibit synthesis)

fungi metabolism and growth environment

Heterotrophic- organic carbon sources, most are obligate aerobes, neutral-slight acidic pH 5.0–6.0 and rt 22°C–30°C or bt 35°–37°C

yeasts

single celled round-oval eukaryotic 3-15 um, asexual via blastoconidia formation (budding), Pseudohyphae when buds fail to detach & elongate

molds

Multicellular filamentous, tubular filaments- hyphae, A mass of intertwined hyphae- mycelium

septate hyphae

divided by cross walls (septa) at regular intervals (aspergillus, dermatophytes, penicillium)

sparsely septate / aseptate hyphae

broad ribbon like, no regular cross walls (mucorales / zygomucetes)

mycelial zones

veg- absorbs nutrients from media, aerial- extends above the agar surface & produces reproductive structures (conidia/spores)

asexual rep fungi

mitotic division makes conidia (on conidiophores) or sporangiospores (inside sporangia)

sexual rep fungi

meiosis makes ascospores, basidiospores, or zygospores

incineration and heating

open flames generate air currents, disrupt laminar airflow & disperse fungal spores, electric micro-incinerators or disposable loops

culture containers

Primary isolation for molds, inoculated onto screw-cap tube slants not petri dishes to minimize spore aerosolization & prevent envir contamination

Systemic Pathogen Hazards:

dimorphic pathogens (Coccidioides immitis, Histoplasma capsulatum) extreme aerosol biohazard risks, Tape tube caps down & process strictly in BSC

hair specimens

Wood's lamp (UV light 365 nm, M. canis and M. audouinii fluoresce bright yellow-green), Pluck infected hair roots or scrape scalp border

nail collection

collect nail clippings/scrapings from hyperkeratotic debris under nail bed

blood culture

Lysis-Centrifug blood culture system (Isolator) to liberate intracel yeast/mold from leuks, Standard auto bottles detect Candida, poorly recover systemic molds

CSF collection

Centrif at high speed (1500g 15 min), sediment for direct India ink wet mount, cryptococcal ag detection, & fungal culture inoculation

tissue samples

Mince into small pieces w/ sterile scalpels, DO NOT grind tissue sus of Mucorales (Zygomycetes)- destroys the fragile nonseptate hyphae

10-20% KOH preop

dissolves keratin, cellular debris, & mucus in skin, hair, & nail samples, clearing background material while leaving fungal chitin intact for microscop exam

india ink prep

neg staining tech, poly capsule of C. neoformans in CSF sediment, halo dark background

lactophenol cotton blue LPCB

mold ID, Phenol kills orgs, lactic acid preserves fungal struc, & cotton blue (aniline blue) stains chitin cell wall

Grocott's Methenamine Silver (GMS)

fungi dark brown-black, pale green background, Highly sensitive for fungal elements in tissue biopsies

periodic acid schiff PAS

fungus cell wall bright magenta/pink, blue-green tissue background, excellent structural detail

Hematoxylin and Eosin H&E

fungus pink-red, background tissues blue, useful for tissue response (granulomas), but low fungal contrast

giemsa / wright-giemsa stain

useful for demonstrating Histoplasma capsulatum intracellular yeasts inside macrophage in bm or blood smears

fontana masson stain

selectively stains melanin pig in phaeoid (dematiaceois) fungi brown-black

sabouraud dextrose agar SDA

standard unselective medium pH 5.6, most fungi, low pH inhib some bacterial growth

brain heart infusion BHI with blood

enriched medium supporting fastidious dimorphic fungi and systemic pathogens

inhibitory mold agar IMA

contains inorganic salts and chloramphenicol to inhib bac while allowing mold growth

mycosel / mycobiotic agar

chloramphenicol (inhib bac) & cycloheximide (inhib saprophytic molds)

mycosel / mycobiotic agar caution

Cycloheximide inhib C. neoformans, C. krusei, Aspergillus fumigatus, Scedosporium, and Mucorales

dermatophyte test medium DTM

phenol red pH indicator, use proteins making alkaline byproducts turning medium bright red

CROMagar candida

selective diff chromogenic medium, rapid presumptive ID of C. albicans green 24, C. tropicalis blue 48, and C. krusei pink-rough 48, C. glabrata smooth pink 48

germ tube test

human serum & incub at 35-37C for 2-3 hrs, true germ tubes- parallel sides w/o contriction at origin (c. albicans & c. dubliniensis pos)

cornmeal agar with tween 80

inoculate in dalmau plate tech, evaluate morph- chlamydospores (C. albicans), blastoconidia, pseudohyphae, & arthroconidia (Geotrichum, Trichosporon)

cryptococcal ag detection

latex agglu or later flow immunoas LFA detect capsular glucuronoxylomannan ag in csf, highly sens for C. meningitis

fungal biomarker assays

Serum- wall, Galactomannan- invasive Aspergillosis, (1,3)-β-D-glucan- broad invasive infec (neg for Mucorales and Cryptococcus)

hyaline hyphae

Clear, unpigmented, or lightly pig walls, readily absorb contrast stains like Lactophenol Cotton Blue (LPCB)

clamp connections

Specialized bypass structures at septations in veg hyphae of Basidiomycota- lateral hyphal outgrowth arches over the septum to connect adjacent cells

favic chandeliers (antler hyphae)

Curved, irregularly branched, ends resembling deer antlers; classic for Trichophyton schoenleinii (tinea favosa). 

racquet hyphae

Club-shaped, enlarged ends of hyphal cells arranged sequentially like a chain of tennis racquets; commonly in dermatophytes

spiral hyphae found in

veg mycelia, Trichophyton mentagrophytes

pectinate bodies

hyphal branches with comb like projections on one sie

nodal vs internal vs absent rhizoids

at base of sporangiophore (Rhizopus) vs along stolon between sporangiophores (lichtheimia / absidia) vs non (mucor)

stolons

horizontal runner like hyphae connecting clusters of sporangiophores and rhizoids in zygomycetes

conidiophore

fertile hyphal stalk supporting conidiogenous (conidia producing) cells

foot cell

specialized basal vegetative hyphal cell from which a conidiophore originates, Aspergillus spp

vesicle

swollen, enlarged apex at the tip of a conidiophore (Aspergillus, Cunninghamella, Syncephalastrum)

phialide

vase-flask-shaped conidiogenous cell, produces phialoconidia in chains or clusters w/o increasing in length

phialide uniseriate vs bi

attach directly to the vesicle surface vs attach to an intermediate supporting cell called a metula

annellide

ringed conidiogenous cell, produces annelloconidia, a ringed scar (annellation) & elongates when a conidium is extruded (Scedosporium / Pseudallescheria, Scopulariopsis)

sporangiophore vs sporangium

erect hyphal stalk supporting sporanium vs closed sac like structure containing asexual sporangiospores

columella

central, dome-shaped or spherical sterile axis in a sporangium at the apex of the sporangiophore

apophysis

funnel or cup-shaped swelling of the sporangiophore located directly beneath the sporangium/columella (Lichtheimia). 

chlamydoconidia (chlamydospores)

Large, thick-walled resting spores formed by enlargement of hyphal cells (terminal, intercalary, or sessile), survive adverse envir (C. albicans, M. audouinii). 

ascus /asci

sac-like cell containing sexual ascospores (8 ascospores each) formed following meiosis in Ascomycota, sexual

cleistothecium

completely closed, spherical, dark ascocarp containing asci and ascospores (Pseudallescheria boydii / Scedosporium teleomorph), sexual

perithecium

flagon or pineapple-shaped ascocarp with an apical opening (ostiole) and protective hair-like setae (Chaetomium), sexual

zygospore

thick-walled, dark, rough sexual spore formed by the fusion of two compatible hyphal nuclei in Mucorales, sexual

sclerotoc bodies- copper pennies

dark brown, round, multicell dividing cells in tissue biopsy, chromoblastomycosis agents

C. albicans

Germ tube pos, terminal chlamydospores, blastoconidia, & pseudohyphae on Cornmeal, Green CHROM, thrush, vaginitis, invasive candidiasis

C. glabrata

small oval yeast, ONLY blastoconidia, Rapid trehalose assimilation pos, High intrinsic azole resistance

C. tropicalis

metallic blue, pseudohyphae w/ blastoconidia singly or in small clusters, sucrose pos

geotrichum candidum

abundant rectangular arthroconidia and coarse septate hyphae, pulmonary and intestinal infections

saccharomyces cerevisiae

'Baker's yeast', round budding yeast & multilateral blastoconidia, 1–4 round ascospores inside asci w/ modified Kinyoun acid-fast 

Aspergillus fumigatus

Thermotolerant (up to 45°C). Smoky green, Uniseriate phialides covering upper half to two-thirds of vesicle, Most common cause of invasive aspergillosis  

A. flavus

yellow-green, Biseriate and uniseriate phialides covering entire vesicle. Produces potent aflatoxins

A. niger

Dark brown-black colony ('black pepper'). Biseriate phialides covering entire spherical vesicle.

Penicillium spp

Blue-green colony w/ white margin, brush-like (penicillus) conidiophores bearing phialides and unbranched chains of spherical conidia

sepedonium spp

single, smooth-rough-walled conidia directly on hyphae, resembles Histoplasma macroconidia but no 37°C yeast conversion

pneumocystis jirovecii

Atypical fungus w/o ergosterol in cell wall (cholesterol instead). Cannot be cultured, trophic form and cyst form w/ 8 intracystic bodies

pneumocystis jirovecii dx

DFA or GMS stain on BAL/sputum showing crushed 'ping-pong ball' cysts, main cause of PCP pneumonia in HIV/AIDS