Free ASCP MLS Exam Practice Questions Mock Test: Part 17 – Microbiology – Analytic Procedures for Mycology

• Pulmonary aspergillosis (e.g., invasive aspergillosis, allergic bronchopulmonary aspergillosis/ABPA) is best diagnosed using lower respiratory samples:

  • Sputum: Non-invasive but may lack sensitivity.

  • Bronchoalveolar lavage (BAL): Gold standard for invasive disease; allows direct microscopy (hyphae), culture, and molecular tests (e.g., Aspergillus PCR/Galactomannan ELISA).

• Microscopy: Septate, 45° branching hyphae (KOH, Calcofluor white, or GMS stain).

• Culture: Grows on Sabouraud agar (e.g., Aspergillus fumigatus shows green conidia).

Why Not the Others?

• a) Hair – Used for dermatophytes (e.g., Trichophyton), not systemic molds.

• b) Nail clippings – Diagnoses onychomycosis, not pulmonary infections.

• d) Urine – Used for Histoplasma antigen testing, not aspergillosis.

CHROMagar™ Candida is a chromogenic differential medium designed to:

• Verify the purity of yeast isolates, especially Candida species.

• Differentiate multiple Candida species based on colony color (e.g., Candida albicans appears green, Candida tropicalis blue-purple, Candida krusei pink with fuzzy edges).

It’s especially useful in detecting mixed infections or contamination by showing different colony colors.

Why not the others?

• a) Sabouraud dextrose agar – Supports general fungal growth but doesn’t differentiate species.

• b) Potato dextrose agar – Promotes sporulation and pigmentation; not ideal for verifying purity.

• c) Cornmeal agar – Used for microscopic morphology (e.g., chlamydospore formation in C. albicans), not for confirming culture purity.

• Rhizopus and Mucor are both Zygomycetes with non-septate hyphae, but they are distinguished by:

  • Rhizopus:

    • Has rhizoids (root-like structures) at the base of sporangiophores (stalks bearing sporangia).

    • Sporangiophores arise opposite rhizoids at nodes.

  • Mucor:

    • Lacks rhizoids entirely.

    • Sporangiophores arise directly from hyphae without nodal organization.

Why Not the Others?

• a) Conidiophores – Found in Aspergillus/Penicillium (septate molds), not Zygomycetes.

• c) Capsule production – Seen in Cryptococcus neoformans (a yeast).

• d) Yeast cell budding – Irrelevant; these are molds with sporangia, not yeasts.

• Sabouraud dextrose agar (SDA) is the gold standard for demonstrating both:

  • Viability: Only live fungi grow, forming visible colonies.

  • Morphology: Colonies show characteristic color/texture, while microscopy (e.g., lactophenol cotton blue) reveals hyphae, spores, or yeast forms.

• Supports growth of most pathogenic fungi (e.g., Candida, dermatophytes, dimorphic fungi).

Why Not the Others?

• a) Gram staining – Shows fungal structures but cannot confirm viability (stains dead/live cells alike).

• c) Saline wet mount – Reveals morphology but cannot prove viability (no growth observed).

• d) Acid-fast stain – Used for Mycobacterium; fungi are not acid-fast.

The gold standard for mold identification involves microscopic analysis of reproductive structures (e.g., conidiophores, spores, hyphae) using:

• Lactophenol cotton blue (LPCB) stain – Preserves and highlights fungal structures.

• Slide culture technique – Allows 3D visualization of conidia and spore-bearing structures.

Why Not the Other Options?

• a) Observation of colony pigmentation – Helpful for preliminary clues (e.g., Aspergillus niger is black), but not definitive due to variability.

• b) Direct inoculation onto blood agar – Used for bacterial cultures, not molds (fungi grow better on Sabouraud dextrose agar).

• d) Gram staining and catalase testing – Used for bacteria, not fungi (molds are Gram-variable and catalase testing is irrelevant).

• Potato dextrose agar (PDA) is a nutrient-rich fungal medium that enhances sporulation in molds, making it ideal for microscopic identification of fungal structures like conidia, spores, and hyphae.

  • The carbohydrates in PDA stimulate robust fungal growth and promote characteristic sporulation patterns (e.g., Aspergillus, Penicillium, dermatophytes).

  • Its slightly acidic pH (~5.6) also inhibits many bacteria, favoring fungal isolation.

Why Not the Others?

• a) Blood agar – Supports fungal growth but lacks specific nutrients to induce sporulation; primarily used for bacterial cultures.

• c) MacConkey agar – Selective for Gram-negative bacteria; inhibits most fungi.

• d) Thayer-Martin agar – Selective for Neisseria gonorrhoeae (bacteria); contains antibiotics that suppress fungi.

• Microsporum is a dermatophyte genus known for producing spindle-shaped (fusiform) macroconidia with:

  • Thin walls

  • Multiple septa (transverse cross-walls)

  • Rough or spiny outer surfaces

• These macroconidia are key for differentiating Microsporum from other dermatophytes:

  • Trichophyton: Pencil-shaped or smooth-walled macroconidia (rare).

  • Epidermophyton: Club-shaped, smooth macroconidia with 1–3 septa.

Why Not the Others?

• b) Candida – A yeast; does not produce macroconidia (forms pseudohyphae/blastoconidia).

• c) Cryptococcus – An encapsulated yeast; no macroconidia.

• d) Rhizopus – A zygomycete; produces sporangia with sporangiospores, not macroconidia.

• “Salt and pepper” colonies: This refers to a rapidly growing mold with both light and dark areas on the colony surface, typical of Zygomycetes, especially Rhizopus.

• LPCB (Lactophenol Cotton Blue) stain showing sporangiophores:

  • Rhizopus produces large sporangiophores with sporangia (sac-like structures containing spores), characteristic of Zygomycetes.

  • Sporangiophores often arise from rhizoids, which help in differentiating Rhizopus from other genera.

Other options:

• a. Aspergillus – Produces conidiophores, not sporangiophores.

• b. Cunninghamella – Belongs to Zygomycetes too but has vesicles with single-spored sporangiola, not typical “salt and pepper” colonies.

• c. Fusarium – Produces macroconidia and microconidia, not sporangiophores.

• Dimorphic fungi exhibit temperature-dependent growth phases:

  • Yeast form (37°C) – In host tissues (e.g., Histoplasma capsulatum in human lungs).

  • Mold form (25–30°C) – In the environment or culture at room temperature.

• This thermal dimorphism is a key virulence factor for systemic fungal pathogens like:

  • Histoplasma capsulatum

  • Coccidioides immitis/posadasii

  • Blastomyces dermatitidis

  • Sporothrix schenckii

Why Not the Others?

• a) Thermophilic fungus – Grows at high temperatures (e.g., 45–60°C), but not as yeast/mold phases.

• c) Opportunistic mold – Causes infections in immunocompromised hosts (e.g., Aspergillus), but is not dimorphic.

• d) Dermatophyte – Infects skin/hair/nails (e.g., Trichophyton), but grows only as molds.

• The urease test is used to detect the enzyme urease, which breaks down urea into ammonia and CO₂, causing a pH shift that turns Christensen’s urea agar pink-red (positive result).

• This test helps differentiate yeasts like:

  • Urease-positive: Cryptococcus neoformans (strongly positive)

  • Urease-negative: Candida albicans

Why not the others?

• a) Germ tube test – Identifies Candida albicans by tube-like hyphae formation, not urease.

• c) Sugar fermentation test – Assesses carbohydrate utilization, not urease activity.

• d) Slide agglutination test – Detects antigens (e.g., for Cryptococcus latex agglutination), not enzymes.

Most pathogenic fungi infect humans and thus thrive at body temperature, which is around 37°C. Therefore, 37–40°C is the optimal temperature range for culturing pathogenic fungi, such as Candida, Histoplasma, and Cryptococcus.

• 4°C is refrigeration temperature — used for storage, not growth.

• 25–30°C is optimal for environmental molds or saprophytic fungi (like Penicillium, Aspergillus in environmental studies).

• 50°C is too high and inhibits growth of most fungi.

• Aspergillus species are identified microscopically by their unique asexual reproductive structures:

  • Conidiophore: A tall, unbranched stalk.

  • Vesicle: A swollen apex at the tip of the conidiophore.

  • Phialides: Flask-shaped cells that produce chains of conidia (spores), radiating from the vesicle.

• This arrangement is diagnostic for Aspergillus (e.g., A. fumigatus, A. flavus).

Why Not the Others?

• a) Sporangium – Found in Zygomycetes (e.g., Rhizopus), not Aspergillus.

• c) Pseudohyphae – Seen in Candida species, not molds like Aspergillus.

• d) Chlamydospores – Produced by some Candida species (e.g., C. albicans), not Aspergillus.

• Dimorphic fungi (e.g., Histoplasma capsulatum, Blastomyces dermatitidis, Coccidioides immitis) are identified by their temperature-dependent phase transition:

  • Mold form (25–30°C): Produces hyphae and conidia/spores in the environment or culture.

  • Yeast form (37°C): Converts to yeast cells (or spherules for Coccidioides) in host tissues or lab cultures.

• Procedure:

  1. Culture at 25–30°C to observe mold morphology (e.g., tuberculate macroconidia in Histoplasma).

  2. Subculture at 37°C to confirm conversion to yeast/spherules.

Why Not the Others?

• a) Animal inoculation: Rarely used today due to ethical concerns and slower turnaround vs. culture.

• c) Sexual/asexual reproduction: Useful for taxonomic classification but not diagnostic for dimorphism.

• d) Molecular testing: Rapid (e.g., PCR for Histoplasma), but conversion remains the gold standard for phenotypic confirmation.

Coccidioidomycosis, also known as Valley Fever, is caused by the dimorphic fungi Coccidioides immitis and Coccidioides posadasii. These fungi are endemic to arid regions of the southwestern United States, particularly:

• California’s Central Valley

• Arizona

• New Mexico

• West Texas

Migrant farm workers in these areas are at increased risk due to:

• Frequent soil exposure (spores are found in soil)

• Dust inhalation

• Often living or working outdoors

Other options:

• a. Missouri cattle ranchers – Not an endemic area.

• b. Wyoming rabbit ranchers – Not an endemic area.

• d. Pregnant women with house cats – Not a risk group for coccidioidomycosis (but a concern in toxoplasmosis).

• Trichophyton rubrum, the most common dermatophyte, exhibits these key features:

  • Microscopic morphology:

    • Racquet hyphae (swollen segments resembling tennis rackets).

    • Pectinate bodies (“combs” or irregular projections along hyphae).

    • Microconidia: Teardrop-shaped, abundant.

    • Macroconidia: Rare, pencil-shaped (if present).

  • Colony characteristics:

    • Salmon-pink to red reverse pigment on Sabouraud dextrose agar.

    • White, cottony surface (may become powdery).

Why Not the Others?

• a) Epidermophyton floccosum:

  • Produces club-shaped macroconidia (no microconidia).

  • Reverse pigment is tan to brown, not salmon-pink.

• b) Microsporum canis:

  • Spindle-shaped macroconidia with rough walls.

  • Reverse pigment is yellow to orange.

• c) Microsporum audouinii:

  • Rare; produces few macroconidia (tapered, thin-walled).

  • Reverse pigment is brownish, not pink.

• Cryptococcal antigen (CrAg) testing (e.g., lateral flow assay [LFA] or latex agglutination) is the most sensitive initial test for cryptococcal disease, particularly in HIV-infected patients with suspected meningitis or disseminated infection:

  • Sensitivity:

    • LFA: 99–100% in CSF and serum/plasma .

    • Latex agglutination: 93–100% .

  • Specificity: >99% for both methods.

  • Advantages: Rapid (10–15 minutes for LFA), point-of-care compatible, and detects early/subclinical infection (e.g., CrAg-positive blood before meningitis symptoms) .

Why Not the Others?

• a) India ink:

  • Sensitivity: Only 44–86% in CSF (misses low fungal burdens) .

  • Requires microscopy expertise and fails to detect non-encapsulated strains.

• b) Gram stain:

  • Sensitivity: ~61% in CSF; less specific (may confuse with other yeasts) .

• d) Giemsa stain:

  • Rarely used; sensitivity/specificity data limited .

• Molecular methods (e.g., PCR, sequencing, MALDI-TOF MS) revolutionize fungal diagnostics by:

  • Species-level identification: DNA sequencing (e.g., ITS, D1/D2 regions) differentiates closely related species (e.g., Candida auris vs. C. haemulonii).

  • Detecting resistance genes: Identifies mutations (e.g., ERG11 in azole-resistant Candida, Cyp51 in Aspergillus fumigatus).

  • Speed: Reduces turnaround time vs. culture (e.g., Pneumocystis jirovecii PCR from BAL).

Why Not the Others?

• a) Morphology only – Microscopy lacks precision for cryptic species (e.g., Aspergillus section Fumigati).

• c) Increasing sporulation – Molecular tools don’t affect growth characteristics.

• d) Enhancing pigment – Irrelevant to molecular identification.

• Exoantigen testing is an immunodiffusion-based method that identifies fungi by detecting species-specific antigens secreted into culture filtrates.

  • Commonly used for dimorphic fungi (e.g., Histoplasma capsulatum, Blastomyces dermatitidis, Coccidioides immitis).

  • Culture filtrates are reacted with known antibodies; precipitin lines confirm the species.

Why Not the Others?

• a) Detecting capsules – Uses India ink (e.g., Cryptococcus) or Mucicarmine stain.

• c) Observing hyphae – Relies on microscopy (e.g., LPCB, KOH).

• d) Enhancing spore formation – Requires specialized media (e.g., potato dextrose agar).

Sabouraud dextrose agar (SDA) is specifically designed for fungal cultivation due to:

• Low (acidic) pH (~5.6): Inhibits most bacterial growth while favoring fungi.

• High glucose content: Supports fungal nutrition.

Other options are incorrect because:

• a) SDA does not contain high lipids.

• c) It supports a wide range of fungi, not just dermatophytes.

• d) It does not enhance bacterial pigmentation (in fact, it suppresses bacterial growth).

• Caffeic acid agar (or “birdseed agar”) is a selective medium used to identify Cryptococcus neoformans based on its phenoloxidase activity:

  • The yeast produces melanin from caffeic acid, turning colonies brown to black.

  • This test is highly specific for C. neoformans and C. gattii (other yeasts remain cream-colored).

• Clinical relevance: C. neoformans is a major cause of meningitis in immunocompromised patients (e.g., HIV/AIDS).

Why Not the Others?

• a) Candida albicans – Forms green colonies on chromogenic agar but does not produce melanin.

• b) Candida glabrata – Small, white colonies; phenoloxidase-negative.

• c) Saccharomyces cerevisiae – No melanin production; used in brewing/baking.

• MALDI-TOF MS (Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry) is the fastest and most accurate method for identifying molds like Aspergillus (suggested by “septate hyphae with conidial heads”):

  • Speed: Results in minutes to hours (vs. days for culture-based methods).

  • Principle: Analyzes unique protein fingerprints (ribosomal proteins) to match reference databases.

  • Accuracy: Identifies to species level (e.g., A. fumigatus vs. A. flavus).

Why Not the Others?

• b) Animal inoculation: Outdated, slow (weeks), and ethically questionable.

• c) Exoantigen test: Requires culture filtrates and immunodiffusion (1–2 days).

• d) Slide culture: Takes 3–7 days to observe sporulation; labor-intensive.

• The Aspergillus galactomannan antigen test is an FDA-approved immunoassay (ELISA or lateral flow) that detects a polysaccharide cell wall component released by Aspergillus species during invasive infection.

  • High sensitivity/specificity in immunocompromised patients (e.g., neutropenic, transplant recipients).

  • Used for serum or BAL fluid testing.

Why Not the Others?

• a) Dermatophyte infections – Diagnosed by KOH microscopy or culture (e.g., skin scrapings).

• c) Dimorphic fungi – Identified via serology (antibodies) or histopathology (e.g., Histoplasma yeast forms).

• d) Fungal pigment – Irrelevant; pigments are observed on culture media (e.g., Aspergillus niger).

• Lactophenol cotton blue (LPCB) is the gold standard for preserving and staining fungal structures for microscopic examination.

  • Lactic acid preserves fungal morphology.

  • Phenol kills the fungus, preventing further growth.

  • Cotton blue stains the fungal elements (hyphae, spores, conidia) for better visibility.

• It provides excellent detail of septations, spores, and conidial arrangements, crucial for identifying molds like Aspergillus, Penicillium, and dermatophytes.

Why not the others?

• a) Wet mount with saline – Temporary, lacks staining, and does not preserve structures well.

• c) Gram stain – Primarily for bacteria; fungi may stain but morphology is less clear.

• d) Ziehl-Neelsen stain – Used for acid-fast bacteria (e.g., Mycobacterium), not fungi.

• Saline wet mounts are used in mycology to:

  • Preserve native morphology: Unlike stains or heat-fixing, saline prevents distortion of fungal structures (hyphae, spores, yeast cells).

  • Assess viability: Motility (if present) or budding in yeasts can indicate live organisms.

  • Quick screening: For direct samples (e.g., skin scrapings, BAL fluid) before applying stains (e.g., KOH, Calcofluor white).

Why Not the Others?

• a) Motile spores – Fungi are non-motile (except rare flagellated zoospores in Oomycetes).

• c) Enhancing pigmentation – Pigments are observed on culture media, not wet mounts.

• d) Germ tube formation – Requires serum/plasma incubation (not saline).

• Coccidioides immitis/posadasii is unique among pathogenic fungi for producing arthroconidia (barrel-shaped, alternate disjunctor cells) in its environmental mold phase:

  • Culture at 25–30°C: Hyphae fragment into arthroconidia, which are highly infectious when inhaled.

  • Tissue phase (37°C): Converts to spherules containing endospores (not arthroconidia).

• Microscopy: Lactophenol cotton blue (LPCB) shows rectangular arthroconidia with empty spaces (disjunctor cells) between them.

Why Not the Others?

• b) Cryptococcus – Encapsulated yeast; no arthroconidia.

• c) Histoplasma – Produces microconidia and tuberculate macroconidia in mold phase.

• d) Blastomyces – Forms lollipop-like conidiophores with pyriform conidia.

• India ink negative staining is primarily used to visualize the capsule of Cryptococcus neoformans (and other encapsulated yeasts) in cerebrospinal fluid (CSF) or other clinical specimens.

  • The ink particles do not penetrate the capsule, creating a clear halo around the yeast cells against a dark background.

  • This helps differentiate Cryptococcus (capsule-positive) from other yeasts (e.g., Candida, which lacks a prominent capsule).

Why Not the Others?

• a) Fungal cell walls – Stained better with PAS or GMS, not India ink.

• c) Hyphal septation – Observed with lactophenol cotton blue (LPCB) or KOH preparations.

• d) Conidial arrangement – Seen in mold cultures (e.g., LPCB mounts), not India ink smears.

Dimorphic fungi exhibit temperature-dependent morphological changes:

• Yeast form (37°C): Found in human tissues (e.g., Histoplasma capsulatum, Blastomyces dermatitidis).

• Mold form (25°C): Found in the environment (soil, decaying matter).

Why not the others?

• a) Incorrect – Dimorphic fungi do not grow only as yeast; they switch forms.

• c) Incorrect – They are visible under a light microscope (no need for electron microscopy).

• d) Incorrect – While special stains (e.g., GMS, PAS) help visualize them, dimorphism itself is defined by temperature-dependent shape-shifting, not staining.

Conidia are asexual, non-motile spores of fungi that are commonly used for microscopic identification of molds. They are produced by various molds (like Aspergillus, Penicillium, etc.) and have distinctive shapes, sizes, and arrangements that help in identifying fungal species under a microscope.

• Endospores are produced by certain bacteria (not fungi).

• Capsule and flagella are structures associated mainly with bacteria, not molds.

• Mold identification relies on two key criteria:

  1. Macroscopic features:

     • Colony color, texture (e.g., powdery, velvety), and growth rate on media like Sabouraud dextrose agar (SDA) or potato dextrose agar (PDA).

     • Examples: Aspergillus niger (black colonies), Trichophyton rubrum (white to red pigment).

  2. Microscopic morphology:

     • Reproductive structures: Conidiophores, conidia, phialides, sporangiophores (e.g., Aspergillus vesicles, Penicillium brush-like conidiophores).

     • Hyphal features: Septate vs. aseptate (e.g., Zygomycetes have broad, non-septate hyphae).

Why Not the Others?

• b) Biochemical reactions – Rarely used for molds (more common for yeasts like Candida).

• c) Selective media – Aids isolation but doesn’t replace morphological ID.

• d) Sexual reproductive cells/phialides – Only relevant for certain molds (e.g., Aspergillus phialides), not universal.

Cycloheximide (also known as actidione) is an antifungal agent added to selective fungal culture media (e.g., Sabouraud dextrose agar with cycloheximide) for the following purposes:

1. Suppresses saprophytic fungi – Many environmental molds (e.g., Aspergillus, Penicillium) are sensitive to cycloheximide, allowing pathogenic fungi (e.g., Histoplasma, Blastomyces, dermatophytes) to grow preferentially.

2. Inhibits some bacteria – While primarily antifungal, it also has mild antibacterial effects, further reducing contamination.

Why Not the Other Options?

• a) Incorrect – Cycloheximide does not enhance sporulation; it suppresses fast-growing contaminants.

• c) Incorrect – It is not a stain; it’s an inhibitory agent.

• d) Incorrect – Germ tube production (e.g., in Candida albicans) occurs in serum media, not cycloheximide-containing media.

The India ink preparation is a rapid microscopic technique used to detect Cryptococcus neoformans (and other Cryptococcus species) in cerebrospinal fluid (CSF) or other clinical samples.

Key Features:

• Visualizes the capsule: The ink particles cannot penetrate the thick polysaccharide capsule of C. neoformans, creating a clear halo around the yeast cells against a dark background.

• Used for presumptive diagnosis of cryptococcal meningitis (though antigen tests like CRAG are now more sensitive).

Why Not the Others?

• b) Candida tropicalis – No capsule; identified via germ tube tests or chromogenic agar.

• c) Aspergillus fumigatus – Seen as septate hyphae (KOH mount or calcofluor white).

• d) Histoplasma capsulatum – Intracellular yeast in macrophages (stained with GMS or Wright-Giemsa).

• Cryptococcus neoformans is the most likely causative organism given the clinical scenario and morphological features:

  • Budding yeast with a capsule: Visualized via India ink staining (clear halo) or Mucicarmine stain (pink capsule).

  • Meningitis in HIV/AIDS: C. neoformans is a leading cause of fungal meningitis in immunocompromised patients.

  • Culture: Grows as creamy, mucoid colonies on Sabouraud agar; urease-positive.

Why Not the Others?

• a) Candida glabrata – A small, non-encapsulated yeast; rarely causes meningitis.

• c) Paracoccidioides braziliensis – A dimorphic fungus (yeast with “pilot wheel” buds in tissues); endemic to Latin America, primarily causes pulmonary disease.

• d) Sporothrix schenckii – Causes lymphocutaneous sporotrichosis; CSF involvement is extremely rare.

• Selective fungal media (e.g., Sabouraud dextrose agar with antibiotics) are designed to:

  • Chloramphenicol: Inhibits bacteria (including Klebsiella).

  • Cycloheximide: Suppresses saprophytic fungi (e.g., Penicillium, Aspergillus), allowing pathogenic fungi (e.g., Candida, Histoplasma) to grow.

• This approach maximizes recovery of clinically relevant fungi despite bacterial contamination.

Why Not the Others?

• a) Rejecting the specimen – Unnecessary; fungal infections can coexist with bacterial infections (e.g., Candida in sputum).

• b) Room-temperature incubation – Inhibits some bacteria but also delays growth of systemic fungi (e.g., Histoplasma prefers 30°C).

• d) PAS stain rejection – Microscopy may miss low fungal loads; culture is more sensitive.

• Dermatophytes are fungi that infect keratinized tissues such as the skin, hair, and nails.

• Skin scrapings and hair (or nail clippings) are the best specimens because they contain the fungal hyphae and spores, which can be examined microscopically (e.g., with KOH prep) or cultured for diagnosis.

• a) Urine sample – Not relevant for dermatophyte infections.

• c) Blood sample – Dermatophytes do not typically cause systemic infections.

• d) Nasal swab – Used for respiratory infections, not skin/hair fungal infections.

Thus, skin scrapings and hair are the standard specimens for diagnosing dermatophytosis (e.g., ringworm, athlete’s foot).

Sporothrix schenckii is a dimorphic fungus with the following characteristics:

• At 25°C (mold phase):

  • Produces delicate, slender conidiophores.

  • Conidia are oval or “flower-like” (rosette) in arrangement.

• At 37°C (yeast phase):

  • Forms cigar-shaped yeast cells, which are a hallmark feature.

  • Found in tissue or culture incubated at body temperature.

This dimorphism is diagnostic of Sporothrix schenckii, the causative agent of sporotrichosis (“rose gardener’s disease”).

Why not the others?

• a. Histoplasma capsulatum – Yeast cells are small and intracellular, not cigar-shaped.

• c. Blastomyces dermatitidis – Yeast phase shows broad-based budding, not cigar-shaped.

• d. Acremonium falciforme – Not a dimorphic fungus and does not form cigar-shaped yeast cells.

Talaromyces marneffei (formerly Penicillium marneffei) is a dimorphic fungus that is especially notable for:

• Producing a characteristic red pigment on culture media at 25°C (mold phase)

• Causing systemic infection in immunocompromised patients, particularly in Southeast Asia

• Growing as yeast-like cells at 37°C (in tissue and body fluids like bone marrow)

Why not the others?

• a. Blastomyces dermatitidis – Dimorphic, but does not produce red pigment.

• b. Aspergillus niger – Produces black colonies, not red.

• d. Rhizopus species – Produces white to grayish colonies, not red.

• The Periodic acid-Schiff (PAS) stain is the gold standard for detecting fungal elements in tissue sections because it specifically stains polysaccharides in fungal cell walls (chitin, glucans, and mannans), making fungi appear magenta/pink against a light background.

• PAS highlights hyphae, yeast cells, and spores clearly, even in low fungal loads, and is especially useful for diagnosing invasive fungal infections (e.g., Aspergillus, Candida, Mucor).

Why Not the Others?

• a) Giemsa stain – Mainly for blood parasites (e.g., malaria) and some bacteria; fungi may stain poorly.

• c) Gram stain – Primarily for bacteria; fungi (if stained) appear Gram-positive but lack structural detail.

• d) Safranin stain – Used in microbiology for bacterial capsules or endospores, not optimized for fungi.

• Dermatophytes (e.g., Trichophyton, Microsporum, Epidermophyton) are primarily identified by their microscopic morphology and macroscopic colony characteristics on fungal-specific media (e.g., Sabouraud dextrose agar, potato dextrose agar).

  • Microscopic features: Shape and arrangement of macroconidia, microconidia, and hyphae (e.g., Microsporum has spindle-shaped macroconidia, Epidermophyton produces smooth-walled club-shaped macroconidia).

  • Macroscopic features: Colony color, texture, and growth rate.

Why Not the Others?

• a) Blood culture results – Dermatophytes rarely invade blood; they infect keratinized tissues (skin, hair, nails).

• c) Gram reaction and motility – Fungi are Gram-positive but nonmotile; these traits don’t differentiate dermatophytes.

• d) Antigen detection in CSF – Used for systemic fungi (e.g., Cryptococcus), not dermatophytes

• Fungal dimorphism describes the phenotypic switching between:

  • Mold form (hyphae) – Found in the environment (25–30°C).

  • Yeast form (or spherules) – Found in host tissues (37°C).

• This adaptation is critical for pathogenicity of systemic fungi like:

  • Histoplasma capsulatum

  • Coccidioides immitis (forms spherules, not yeast)

  • Blastomyces dermatitidis

  • Sporothrix schenckii

Why Not the Others?

• a) Two spore types – Some fungi produce both sexual/asexual spores, but this is not dimorphism.

• c) Aerobic/anaerobic growth – Irrelevant to morphology shifts.

• d) Antifungal-induced changes – Drug resistance may alter morphology, but this is not dimorphism.

A corneal isolate growing white to violet colonies and showing slender, sickle- or canoe-shaped macroconidia is characteristic of Fusarium species.

🔬 Key identifying features of Fusarium:

• Colony color: Often white, lavender, or violet on Sabouraud dextrose agar.

• Microscopy:

  • Large, multicelled, sickle- or banana-shaped macroconidia.

  • Frequently also has small microconidia.

• Common cause of keratitis (corneal infections) and mycotic keratitis, especially in tropical or agricultural settings.

❌ Why not the others?

• a. Acremonium: Produces slimy phialides with small conidia, no sickle-shaped macroconidia.

• b. Aspergillus: Has conidial heads with chains of conidia, not sickle-shaped spores.

• d. Geotrichum: Produces arthroconidia (rectangular cells in chains), not macroconidia.

• Systemic pathogenic fungi (e.g., Histoplasma capsulatum, Coccidioides immitis, Blastomyces dermatitidis) require Biosafety Level 3 (BSL-3) containment due to:

  • Aerosol transmission risk: Inhalation of spores can cause severe respiratory/systemic infections.

  • High virulence: Especially in immunocompromised patients.

• BSL-3 practices include:

  • Class II/III biological safety cabinets for all procedures.

  • Negative pressure labs with HEPA-filtered exhaust.

  • Respiratory protection (N95 masks) and double-door access controls.

Why Not the Others?

• a) BSL-1 – For non-pathogenic microbes (e.g., Saccharomyces cerevisiae).

• b) BSL-2 – For moderate-risk pathogens (e.g., Candida, dermatophytes); insufficient for systemic dimorphic fungi.

• d) BSL-4 – Reserved for lethal airborne viruses (e.g., Ebola), not fungi.

• Chromogenic agar (e.g., CHROMagar™ Candida) contains substrates that react with species-specific enzymes, producing distinct colony colors:

  • Candida albicans: Light to dark green

  • Candida tropicalis: Blue with pink halo

  • Candida krusei: Pink/fuzzy

  • Candida glabrata: White/purple

• Allows rapid presumptive ID directly from primary culture (within 24–48 hours).

Why Not the Others?

• a) Eliminates bacteria – While chromogenic agar may include antibiotics, its primary purpose is fungal differentiation, not bacterial suppression.

• c) Speeds spore germination – Color change is due to enzymatic activity, not germination rate.

• d) Enhances hyphae – Designed for yeast (Candida), not mold, identification.

• Cycloheximide is an antifungal agent added to selective media (e.g., Mycosel or Mycobiotic agar) to inhibit saprophytic molds and non-pathogenic fungi. However, it can also suppress growth of:

  • Some pathogenic yeasts: Cryptococcus neoformans (especially C. gattii).

  • Dimorphic fungi: Histoplasma capsulatum, Blastomyces dermatitidis, and Coccidioides immitis (critical for diagnosing endemic mycoses).

Why Not the Others?

• b) Gentamicin & d) Penicillin: Target bacteria; do not affect fungi.

• c) Chloramphenicol: Inhibits bacteria but is fungistatic at standard concentrations (does not block pathogenic fungi)

• Fonsecaea pedrosoi is a slow-growing dematiaceous (dark-pigmented) fungus associated with chromoblastomycosis, a chronic fungal infection of the skin and subcutaneous tissue.

• It shows multiple types of sporulation, including:

  • Cladosporium-type

  • Phialophora-type

  • Fonsecaea-type

This polymorphic sporulation pattern is characteristic of Fonsecaea pedrosoi and helps in its identification in the lab.

Other options:

• b. Pseudallescheria boydii – Not dematiaceous, and does not show the same sporulation types.

• c. Phialophora verrucosa – Shows only Phialophora-type sporulation.

• d. Cladosporium carrionii – Shows Cladosporium-type sporulation only.

• Calcofluor white is a fluorescent dye that binds to chitin and cellulose in fungal cell walls, causing them to glow bright blue-white or apple-green under UV light.

  • Septate hyphae fluoresce distinctly, aiding rapid diagnosis of dermatophytes (e.g., Trichophyton) or invasive molds (e.g., Aspergillus).

  • Often combined with KOH to clear background debris.

Why Not the Others?

• a) Acridine orange – Stains nucleic acids (used for bacteria/viruses), not fungal walls.

• c) Gomori methenamine silver (GMS) – Stains fungi black but requires bright-field microscopy (no fluorescence).

• d) Periodic acid-Schiff (PAS) – Stains fungi magenta but also lacks fluorescence.

• Calcofluor white is a fluorescent dye that binds to chitin and cellulose in fungal cell walls, causing them to glow bright blue-white or apple-green under UV light microscopy.

• This enhances detection of hyphae, yeast cells, and spores even in low concentrations or mixed with debris.

• Often combined with KOH to clear background material while highlighting fungi.

Why Not the Others?

• a) Stains fungal elements red – Describes PAS stain, not Calcofluor white.

• c) Dissolves tissue background – This is the role of KOH, not the dye itself.

• d) Selectively kills bacteria – Calcofluor white has no antimicrobial properties.

• The slide culture technique is a specialized mycological method that allows direct microscopic observation of fungal growth, sporulation, and colony morphology while they develop on the slide.

  • A small block of agar is placed on a sterile slide, inoculated with the fungus, and covered with a coverslip.

  • As the fungus grows, hyphae and spores develop in their natural arrangement, enabling accurate identification of structures like conidiophores and conidia.

  • This method is particularly useful for identifying molds (e.g., Aspergillus, Penicillium, dermatophytes).

Why not the others?

• b) Streak plate method – Used for isolation, but colonies must be transferred for microscopy, disrupting natural growth patterns.

• c) Broth enrichment culture – Promotes fungal growth but does not allow direct microscopic observation of undisturbed structures.

• d) Cold enrichment technique – Used primarily for isolating Listeria from clinical specimens, not fungi.

• Talaromyces (formerly Penicillium) and Aspergillus fumigatus are differentiated by their conidiophore structures:

  • Talaromyces:

    • Lacks a vesicle (swollen tip).

    • Conidiophores branch into phialides that produce chains of conidia in a brush-like (penicillus) arrangement.

  • Aspergillus fumigatus:

    • Has a terminal vesicle with phialides radiating directly from it (uniseriate or biseriate).

    • Conidia form dense chains from the phialides.

Why Not the Others?

• a) Production of conidia on phialides: Both genera produce conidia this way (not distinctive).

• b) Optimum growth temperature: Both grow well at 25–30°C (though A. fumigatus tolerates 37°C+).

• c) Presence of rhizoids: Found in Zygomycetes (e.g., Rhizopus), not Talaromyces or Aspergillus.

• The description fits Cryptococcus neoformans, a yeast-like fungus with these key features:

  • Urease-positive: Rapidly hydrolyzes urea (e.g., on Christensen’s urea agar).

  • Encapsulated cells: Visible with India ink staining (clear halo around yeast cells) or Mucicarmine stain (pink capsule).

  • Blood culture growth: Common in disseminated cryptococcosis, especially in HIV/AIDS patients.

Why Not the Others?

• a) Candida albicans – Forms germ tubes (not capsules) and is urease-negative.

• c) Aspergillus fumigatus – A mold with septate hyphae; does not grow as yeast in blood.

• d) Histoplasma capsulatum – Intracellular small yeast (2–4 µm) in tissues; urease-negative and non-encapsulated.

• Zygomycetes (e.g., Rhizopus, Mucor, Lichtheimia) often produce rapid, cotton candy-like growth within 48–72 hours, characterized by:

  • Dense, grayish aerial mycelium filling the culture container.

  • Non-septate (coenocytic) hyphae visible under microscopy.

  • Sporangia containing sporangiospores (absent in dermatophytes/dimorphic molds).

• This morphology contrasts with slower-growing molds like dermatophytes or dimorphic fungi.

Why Not the Others?

• a) Dermatophyte: Grows slower (1–2 weeks), with powdery or velvety colonies (e.g., Trichophyton).

• b) Dimorphic mold: At 25–30°C, dimorphic fungi (e.g., Histoplasma) form mold colonies with micro/macroconidia, not fluffy hyphae.

• d) Dematiaceous mold: Pigmented (brown/black), with septate hyphae (e.g., Cladosporium).

• Histoplasma capsulatum appears in tissues as small (2–4 µm), oval, intracellular yeasts, primarily within macrophages.

• Giemsa stain highlights the yeast forms within phagocytic cells, showing purple nuclei and blue cytoplasm.

• Periodic acid-Schiff (PAS) and Grocott’s methenamine silver (GMS) stains are also used to clearly visualize the yeast cell walls (magenta with PAS, black with GMS).

Why Not the Others?

• a) Acid-fast stain – Used for Mycobacterium, not fungi (though Histoplasma may rarely show weak acid-fastness).

• b) Direct Gram staining – Fungi may appear Gram-positive but lack specificity for Histoplasma.

• d) Lactophenol cotton blue (LPCB) – Used for mold identification in culture, not tissue sections.

• Canavanine-glycine-bromothymol blue (CGB) agar is a selective medium used to differentiate Cryptococcus gattii from Cryptococcus neoformans.

  • C. gattii assimilates glycine as a carbon source, producing ammonia and turning the medium blue due to the pH-sensitive bromothymol blue indicator .

  • C. neoformans does not grow or change the medium’s color (remains yellow/green) .

Why Not the Others?

• a) Candida krusei: A non-encapsulated yeast that does not grow on CGB agar (urease-negative) .

• c) Cryptococcus neoformans: Fails to utilize glycine (CGB-negative).

• d) Rhodotorula species: May grow but does not turn CGB agar blue (urease-positive but glycine-nonassimilating)

• Cornmeal agar is specifically used to induce chlamydospore formation in yeasts, particularly for identifying Candida albicans and related species.

  • The nutrient-limited environment of cornmeal agar stimulates C. albicans to produce thick-walled, round chlamydospores (terminal or lateral), which are diagnostic for this species.

  • Also reveals pseudohyphae and blastoconidia (budding yeast cells) for further differentiation.

Why Not the Others?

• b) Urease testing – Detects urease enzyme (e.g., for Cryptococcus), not chlamydospores.

• c) Germ tube test – Identifies C. albicans via germ tube formation in serum, not chlamydospores.

• d) Latex agglutination – Detects antigens (e.g., Cryptococcus capsular polysaccharide).

• Lactophenol cotton blue (LPCB) is the most commonly used stain for visualizing fungal elements in direct microscopic examination. It stains the chitin in fungal cell walls, making hyphae and spores clearly visible.

• Gram stain (a) is mainly used for bacteria, though some fungi (like yeasts) may also appear Gram-positive.

• Wright-Giemsa stain (c) is used for blood smears and some parasites, not typically for fungi.

• Ziehl-Neelsen stain (d) is used for acid-fast bacteria like Mycobacterium tuberculosis, not fungi.

Thus, LPCB is the best choice for fungal microscopy.

• The carbohydrate assimilation test is a biochemical method that distinguishes Candida species by their ability to utilize specific sugars as carbon sources.

  • Candida albicans: Ferments and assimilates glucose, maltose, and sucrose, but not lactose.

  • Candida glabrata: Assimilates glucose and trehalose, but not sucrose or maltose.

• Commercial systems (e.g., API 20C AUX, VITEK 2 YST) automate this profiling for rapid ID.

Why Not the Others?

• a) Germ tube test – Positive only for C. albicans (not C. glabrata), but does not assess sugar use.

• c) Urease test – Negative for both C. albicans and C. glabrata (useful for Cryptococcus).

• d) Cornmeal agar – Shows pseudohyphae/chlamydospores in C. albicans (absent in C. glabrata), but does not test sugars.

• Candida albicans is the only fungus among the options that produces both germ tubes and chlamydospores, key features for its identification:

  • Germ tubes: Formed within 2–3 hours in serum/plasma at 37°C (rapid diagnostic test).

  • Chlamydospores: Induced on cornmeal agar with Tween 80 (thick-walled, round, terminal spores).

Why Not the Others?

• b) Cryptococcus neoformans – Produces a capsule (India ink+) and is urease+, but no germ tubes or chlamydospores.

• c) Mucor species – Non-septate hyphae with sporangia; no yeast phase or chlamydospores.

• d) Aspergillus flavus – A mold with septate hyphae and conidial heads; no germ tubes.

• Fungal serological tests are used to diagnose infections by detecting:

  • Fungal antigens (e.g., Cryptococcal capsular polysaccharide, Histoplasma galactomannan, Aspergillus galactomannan).

  • Host antibodies (e.g., Coccidioides IgM/IgG, Blastomyces antibodies).

• These tests are critical for systemic mycoses where cultures may be slow or negative (e.g., histoplasmosis, cryptococcosis).

Why Not the Others?

• a) Bacterial toxins – Detected by assays like ELISA (e.g., Clostridium difficile toxin), not fungal serology.

• c) Viral RNA – Identified via PCR, not serology.

• d) Mycotoxins in food – Analyzed by chromatography/mass spectrometry, not patient serum tests.

• Blastomyces dermatitidis is a dimorphic fungus, meaning it exists in two distinct forms depending on temperature:

  • Mold phase (25–30°C): Produces hyphae with lollipop-shaped conidiophores and pyriform (pear-shaped) conidia.

  • Yeast phase (37°C or in human tissues): Transforms into thick-walled, broad-based budding yeast cells (8–15 µm).

Why Not the Others?

• b) Candida albicans: A yeast that forms pseudohyphae/germ tubes but is not dimorphic (no environmental mold phase).

• c) Cryptococcus neoformans: An encapsulated yeast; no mold phase in nature.

• d) Aspergillus fumigatus: A mold at all temperatures; no yeast phase.

• Dematiaceous mold: These are dark-pigmented fungi (melanin in their cell walls), commonly associated with subcutaneous infections, such as from foot wounds.

• Alternaria:

  • Produces dark, multicellular macroconidia.

  • The macroconidia are arranged in chains.

  • They have both horizontal and longitudinal septations, often described as muriform.

  • Conidia often appear club-shaped with a tapered end, sometimes in a “drumstick” pattern.

This morphology is distinctive for Alternaria, especially in tissue infections like phaeohyphomycosis or chromoblastomycosis.

Other options:

• b. Curvularia – Produces curved conidia, but not typically muriform (no longitudinal septations).

• c. Paecilomyces – Produces elongated phialoconidia, not dematiaceous, and no chaining macroconidia.

• d. Scopulariopsis – Produces lemon-shaped conidia, not darkly pigmented or muriform.

• The description matches Malassezia furfur, a lipophilic yeast that:

  • Forms buds on a broad base (unipolar budding, “collarette” appearance).

  • Requires exogenous lipids (e.g., olive oil, Tween supplements) for growth in culture.

  • Clinical relevance: Causes neonatal bloodstream infections, especially in preterm infants receiving lipid-rich parenteral nutrition.

Why Not the Others?

• a) Candida tropicalis – Buds on a narrow base; grows without lipids.

• c) Candida lipolytica – Rare pathogen; does not require lipids.

• d) Cryptococcus gattii – Encapsulated yeast; grows without lipids.

• The germ tube test is a rapid, presumptive identification for Candida albicans, the most common pathogenic yeast.

  • Procedure: Incubate yeast in rabbit or human serum/plasma at 37°C for 2–3 hours.

  • Positive result: Formation of germ tubes (short, filamentous extensions without constrictions at the base).

  • Specificity: >95% for C. albicans (rare C. dubliniensis strains may also produce germ tubes).

Why Not the Others?

• a) Candida tropicalis – Forms pseudohyphae (constricted at septa), not true germ tubes.

• b) Candida krusei – Produces elongated pseudohyphae but no germ tubes.

• d) Candida glabrata – Grows as small yeast cells only (no hyphae or germ tubes).

• The description fits Cryptococcus neoformans (gram-positive yeast, catalase-positive, no germ tubes). Polymyxin B susceptibility testing helps differentiate:

  • Cryptococcus neoformans: Resistant to polymyxin B.

  • Candida species: Typically susceptible (except C. krusei, which is intrinsically resistant but germ tube-negative).

Why Not the Others?

• b) Novobiocin susceptibility: Used for bacteria (e.g., Staphylococcus saprophyticus in UTIs), not yeasts.

• c) Oxidase test: Used for bacteria (e.g., Neisseria, Pseudomonas); fungi are oxidase-negative.

• d) Beta-lactamase test: Irrelevant for fungi (targets bacterial resistance).

• Cryptococcus neoformans is primarily identified by its thick polysaccharide capsule, which is a key virulence factor and diagnostic feature:

  • Microscopy: Visualized via India ink staining (clear halo around yeast cells) or Mucicarmine stain (pink capsule).

  • Clinical impact: The capsule inhibits phagocytosis, enabling systemic spread (e.g., meningitis in HIV/AIDS patients).

Supportive (But Not Definitive) Features:

• b) Positive urease test: C. neoformans is urease+, but so are other yeasts (e.g., Rhodotorula).

• d) Positive phenol oxidase test: Melanin production on caffeic acid agar (brown colonies) is highly suggestive but secondary to capsule detection.

• a) Yellow colonies: Irrelevant; C. neoformans colonies are typically creamy/mucoid.

The germ tube test is a rapid and specific test used to identify Candida albicans by detecting its ability to form germ tubes (hyphal projections) within 2–4 hours when incubated in serum (e.g., human or fetal bovine serum).

Why not the others?

• a) Urease test: Used to differentiate Cryptococcus (urease-positive) from Candida species (usually urease-negative).

• c) Coagulase test: Used for Staphylococcus (e.g., S. aureus), not Candida.

• d) Oxidase test: Used for bacteria like Pseudomonas, not fungi.

• Cryptococcus neoformans is the yeast that fits all the given characteristics:

  • Natural habitat: Found in bird droppings (especially pigeon excreta).

  • Urease production: Positive on Christensen’s urea agar.

  • Capsule detection: Visualized with India ink staining (clear halo) or Mucicarmine stain (pink capsule).

  • Clinical relevance: Causes meningitis in immunocompromised patients (e.g., HIV/AIDS).

Why Not the Others?

• a) Candida krusei – Urease-negative, no capsule; associated with antifungal resistance.

• c) Candida parapsilosis – Urease-negative, no capsule; forms pseudohyphae.

• d) Trichosporon beigelii – Urease-positive but non-encapsulated; causes white piedra.

• The germ tube test is a rapid and specific test for Candida albicans, the most common pathogenic yeast in clinical infections.

• When incubated in human serum at 37°C for 2–3 hours, C. albicans produces germ tubes (short, filamentous extensions without constrictions at their base).

• This test is highly specific for C. albicans and helps differentiate it from other Candida species (e.g., C. tropicalis, C. glabrata), which do not form true germ tubes.

Why not the others?

• a) Cryptococcus neoformans – Does not produce germ tubes; identified by urease test, capsule, or latex agglutination.

• c) Aspergillus fumigatus – A mold, not a yeast; identified by hyphae and conidial structures.

• d) Histoplasma capsulatum – A dimorphic fungus, detected by mold-to-yeast conversion or antigen tests, not germ tubes.

The cornmeal agar morphology test is used to identify Candida species, particularly by observing:

• Chlamydospore formation (thick-walled, spherical survival spores)

• Pseudohyphae and blastoconidia (budding yeast) patterns

Key Features:

• Candida albicans typically forms chlamydospores at the ends of hyphae (a diagnostic clue).

• Other Candida species (e.g., C. tropicalis, C. glabrata) show different pseudohyphal structures but usually do not produce chlamydospores.

Why Not the Other Options?

• a) Slide culture technique – Used for studying mold structures (e.g., conidiophores) under microscopy, not chlamydospores.

• b) Germ tube test – Detects C. albicans via germ tube formation in serum (not chlamydospores).

• c) Sporulation test – General term for inducing fungal spores (not specific to cornmeal agar).

• Potassium hydroxide (KOH, 10–20%) is the primary clearing agent used in direct microscopy of fungal infections (e.g., skin scrapings, nail clippings, hair).

  • Dissolves keratin and cellular debris, leaving fungal hyphae/spores intact and more visible under the microscope.

  • Often combined with calcofluor white (a fluorescent dye) to enhance fungal detection.

Why Not the Others?

• a) Ethanol – Fixes specimens but does not clear debris.

• c) Sodium chloride – Used in saline wet mounts, but lacks clearing properties.

• d) Sulfuric acid – Destroys organic material; not used for fungal microscopy.

• The sugar assimilation test evaluates how yeasts metabolize different carbohydrates (e.g., glucose, lactose, maltose) by observing growth in media containing specific sugars.

• This biochemical profiling helps differentiate yeast species (e.g., Candida albicans vs. Candida tropicalis) based on their unique assimilation patterns.

Why Not the Others?

• a) Urease test – Detects urease enzyme (e.g., for Cryptococcus), not sugar metabolism.

• b) Germ tube test – Rapid ID for Candida albicans (no carbohydrates involved).

• d) Chlamydospore test – Identifies C. albicans on cornmeal agar, not sugar use.

• Biological safety cabinets (BSCs) are essential for handling molds because they:

  • Filter airborne spores (HEPA filtration), preventing inhalation or environmental contamination.

  • Provide physical containment during procedures like subculturing or slide preparation.

• This is critical for pathogenic molds (e.g., Aspergillus, Histoplasma, Coccidioides) that can cause opportunistic infections or allergic reactions if inhaled.

Why Not the Others?

• a) Open bench culture handling – High risk of spore dispersal; unsafe for lab personnel.

• c) Incubating in ambient air without cover – Allows spore release into the environment.

• d) Using only slide culture without protection – Slide cultures still require BSC handling to avoid spore exposure.

• KOH (potassium hydroxide) preparation is used in fungal diagnostics because it breaks down keratin and host cellular material, making fungal elements (hyphae, yeast cells, or spores) more visible under the microscope.

• It does not stain fungi (eliminating option c), nor does it promote spore germination (eliminating option a).

• While KOH can sometimes reveal bacteria incidentally, its primary purpose is fungal detection (eliminating option d).

• The latex agglutination test is a rapid, sensitive, and specific method to detect Cryptococcus neoformans capsular polysaccharide antigen in cerebrospinal fluid (CSF), serum, or urine.

  • It uses antibody-coated latex particles that clump (agglutinate) in the presence of Cryptococcal antigen, providing results within minutes.

  • Particularly crucial for diagnosing cryptococcal meningitis, a life-threatening fungal infection.

Why Not the Others?

• b) Germ tube test – Identifies Candida albicans (not Cryptococcus).

• c) Coagulase test – Differentiates Staphylococcus aureus (bacteria) from other staphylococci.

• d) Catalase test – Used to distinguish bacterial species (e.g., Staphylococcus vs. Streptococcus).

• Dermatophytes (e.g., Trichophyton, Microsporum, Epidermophyton) are primarily identified by their reproductive structures:

  • Macroconidia: Large, multicellular spores with distinct shapes:

    • Microsporum: Spindle-shaped, thick-walled, rough-surfaced.

    • Epidermophyton: Club-shaped, smooth-walled, with 1–5 septa.

    • Trichophyton: Pencil-shaped (rare or absent in some species).

  • Microconidia: Small, single-celled spores (round, teardrop, or club-shaped), abundant in Trichophyton.

• These structures are best observed via lactophenol cotton blue (LPCB) mounts from culture.

Why Not the Others?

• a) Septate and branching hyphae: Non-specific (seen in most molds).

• b) Racquet/pectinate hyphae: Ancillary features (not diagnostic).

• c) Chlamydospores: Formed under stress (e.g., Candida), not key for dermatophytes.

• The description of broad, ribbon-like, non-septate (aseptate) hyphae with irregular branching is pathognomonic for Mucorales fungi (e.g., Mucor, Rhizopus, Lichtheimia).

  • These are angioinvasive molds causing mucormycosis, a life-threatening infection in diabetics or immunocompromised patients.

  • Their hyphae appear wide (5–20 µm) and lack septations, unlike the narrow, septate hyphae of Aspergillus.

Why Not the Others?

• a) Candida albicans – Forms pseudohyphae (constricted at septa) and yeast cells, not broad hyphae.

• c) Aspergillus species – Has narrow, septate hyphae with 45° angle branching.

• d) Cryptococcus neoformans – A yeast with capsules (visible with India ink), not hyphae.

• Microscopic examination of reproductive structures (e.g., conidia, conidiophores, sporangiophores) is the gold standard for mold identification. Key features include:

  • Conidiophore morphology (e.g., Aspergillus vesicles with phialides, Penicillium brush-like conidiophores).

  • Spore type/arrangement (e.g., Microsporum spindle-shaped macroconidia, Rhizopus sporangia).

• These structures are species-specific and unaffected by environmental variables (unlike pigmentation/growth rate).

Why Not the Others?

• a) Colony pigmentation – Variable (e.g., Aspergillus flavus may be yellow-green or brown).

• b) Hyphal width – Non-specific (e.g., Zygomycetes have broad hyphae, but so do other molds).

• d) Growth rate – Influenced by media/temperature; not diagnostic.

• The description of black pigment and septate hyphae is classic for Aspergillus niger, a common environmental mold that can cause opportunistic infections.

  • Macroscopic: Colonies are velvety black due to dark-pigmented conidia.

  • Microscopic: Shows septate hyphae with conidiophores terminating in a vesicle covered by phialides that produce chains of black conidia.

Why Not the Others?

• a) Rhizopus – A Zygomycete with non-septate hyphae and sporangia (not black conidial heads).

• c) Candida glabrata – A yeast (no hyphae or pigment).

• d) Cryptococcus gattii – A encapsulated yeast (not a mold).

• Coccidioides immitis is a dimorphic fungus with distinct structures in its mold phase (25–30°C):

  • Barrel-shaped arthroconidia (arthrospores):

    • These are rectangular, thick-walled spores that alternate with empty disjunctor cells in hyphae, giving a “checkerboard” appearance under microscopy .

    • Key diagnostic feature: Arthroconidia are highly infectious when inhaled and are pathognomonic for Coccidioides .

  • Hyphae: Septate and hyaline, but less distinctive for identification.

Why Not the Others?

• b) Tuberculate macroconidia: Found in Histoplasma capsulatum, not Coccidioides .

• c) Thick-walled sporangia: Produced by Zygomycetes (e.g., Rhizopus), which have non-septate hyphae .

• d) Pyriform microconidia: Seen in Blastomyces dermatitidis (pear-shaped)

• Coccidioides immitis is a dimorphic fungus that requires Biosafety Level 3 (BSL-3) precautions due to its highly infectious arthroconidia (spores) in the mold phase:

  • Inhalation risk: Arthroconidia can cause severe pulmonary/systemic infections (coccidioidomycosis/Valley fever).

  • Lab safety: All manipulations (culture, microscopy) must occur in a Class II/III biological safety cabinet (BSC) to prevent aerosol exposure.

Why Not the Others?

• a) Streptococcus pyogenes & b) Staphylococcus aureus:

  • BSL-2 pathogens; standard PPE (gloves, lab coat) suffices.

  • No BSC required unless generating aerosols (e.g., centrifuging).

• c) Candida albicans:

  • BSL-2; routine handling on open bench is acceptable (non-aerosolizing procedures).

• Paecilomyces is characterized by long, tapering phialides that bend away from the conidiophore axis, forming a divergent brush-like (penicillus) arrangement69. This morphology closely resembles Talaromyces but differs in key features:

  • Phialide shape: Paecilomyces phialides are elongated and slender, often with a distinct neck, while Talaromyces phialides are typically flask-shaped (acerose) and more rigid.

  • Conidiophore structure: Paecilomyces lacks the vesicle seen in Aspergillus and the biverticillate metulae-phialide complex of Talaromyces.

Why Not the Others?

• a) Exophiala: A black yeast with short annellides (not phialides) and dematiaceous hyphae; no tapering phialides.

• b) Acremonium: Produces straight, awl-shaped phialides that do not bend away from the axis.

• c) Cladosporium: Forms dark-pigmented conidiophores with branching chains of blastoconidia (no phialides)

Eumycotic mycetoma is a chronic, granulomatous fungal infection involving skin, subcutaneous tissue, and sometimes bone, commonly presenting with draining sinuses and granules (grains).

Why not the others?

• b. Nocardia brasiliensis: Causes actinomycetoma (bacterial), not eumycetoma.

• c. Coccidioides immitis: Causes systemic fungal infections like valley fever, not localized mycetoma.

• d. Aspergillus fumigatus: Causes aspergillosis, not a typical agent of mycetoma in the USA.

• Histoplasma capsulatum is a dimorphic fungus primarily transmitted via inhalation of spores (microconidia) from contaminated soil (e.g., bird/bat droppings).

  • Causes histoplasmosis, a pulmonary/systemic infection endemic to the Ohio/Mississippi River valleys.

  • Spores convert to yeast forms in the lungs, which disseminate in immunocompromised hosts.

Why Not the Others?

• a) Sporothrix schenckii – Enters through skin trauma (e.g., rose thorns), causing lymphocutaneous sporotrichosis.

• b) Trichophyton rubrum – A dermatophyte causing tinea (e.g., athlete’s foot) via direct contact.

• c) Malassezia furfur – Part of skin flora; causes pityriasis versicolor (not inhaled).

• Cornmeal agar with Tween 80 is the gold standard medium for observing yeast morphology (e.g., pseudohyphae, blastoconidia, chlamydospores) due to its:

  • Nutritional deficiency, which promotes structural differentiation.

  • Tween 80 (a surfactant) enhances hyphae/pseudohyphae formation in Candida spp. .

Why Not the Others?

• b) Brain-heart infusion (BHI): Supports yeast growth but lacks morphological differentiation.

• c) Potato dextrose agar (PDA): Used for pigmentation (e.g., Candida albicans cream-colored colonies) but not optimal for hyphae.

• d) Urea agar: Identifies urease-positive yeasts (e.g., Cryptococcus) but does not show morphology .

• Histoplasma capsulatum in its mycelial (mold) phase is best identified by its tuberculate macroconidia:

  • Size: 8–14 μm in diameter.

  • Appearance: Round, thick-walled, and covered with finger-like projections (tubercules)—resembling a “knobby golf ball” .

  • Microscopy: Visible with lactophenol cotton blue (LPCB) stains .

• Microconidia (2–5 μm) are also produced but are less distinctive (smooth-walled, globose).

Why Not the Others?

• a) Arthrospores in every other cell: Diagnostic for Coccidioides immitis (barrel-shaped arthroconidia with disjunctor cells) .

• b) 2–5 μm microspores: Present but not pathognomonic (also seen in Blastomyces and Sepedonium).

• d) 5–7 μm nonseptate macroconidia: Found in Zygomycetes (e.g., Mucor), which lack septations entirely .

The key microscopic features described—septate hyphae with dichotomous branching at 45° angles—are classic for Aspergillus species.

Why Aspergillus?

1. Septate hyphae: Aspergillus has divided (septate) hyphae, unlike Mucor (aseptate).

2. Dichotomous branching at 45°: A hallmark of Aspergillus, seen under microscopy (e.g., in tissue or culture).

3. Reproductive structures: Produces conidiophores with vesicles and phialides (e.g., A. fumigatus has columnar conidial heads).

Why Not the Others?

• a) Mucor species: Aseptate hyphae with wide-angle (90°) irregular branching (ribbon-like).

• b) Candida species: Yeast forms (blastoconidia, pseudohyphae)—no true hyphae (except germ tubes in C. albicans).

• d) Cryptococcus species: Encapsulated yeast (no hyphae unless rare pseudohyphae in C. neoformans var. gattii).

• The germ tube test is a rapid, presumptive test for Candida albicans, the most common pathogenic Candida species.

  • Procedure: Incubate yeast in human serum/plasma at 37°C for 2–3 hours.

  • Positive result: Formation of germ tubes (short, filamentous extensions without constrictions at the base).

  • Specificity: >95% for C. albicans (some C. dubliniensis strains may also produce germ tubes).

Why Not the Others?

• a) Candida tropicalis – Forms pseudohyphae (constricted at septa), not true germ tubes.

• b) Candida parapsilosis – Produces “giant” pseudohyphae but no germ tubes.

• c) Candida glabrata – Grows as small yeast cells only (no hyphae or germ tubes).