This document summarizes mycology, the study of fungi. It describes the structure and morphology of fungi including hyphae, spores, yeasts and molds. Common diagnostic techniques are outlined such as KOH preps, calcofluor white, lactophenol cotton blue staining. Important human pathogenic fungi are discussed including dermatophytes causing ringworm, systemic fungi like Blastomyces and Coccidioides, and opportunistic fungi such as Candida albicans. Diagnostic methods and virulence factors are also summarized.

1. MYCOLOGY

2. MYCOLOGY
• Fungi- EUKARYOTIC
• Most are obligate aerobes
• Saprophytes
• Exist as Mold or Yeast
• Chitin cell wall & sterol cell membrane
• Achlorophyllous and Heterotrophic
• Exist as dimorphic/monomorphic
• STRUCTURE: HYPHAE (fundamental unit) & SPORE
(reproductive)

3. 1) Yeast/Tissue
 Unicellular, budding, moist col, 37’C , BHIA
2) Mold/ Filamentous/Hyphae
Multicellular,dry col., RT, SDA
Morphology (Fungi)

4. HYPHAE
• branching, threadlike, tubular filaments
that either be:
1. Septate – cross wall (conidiophore)
2. Aseptate (coenocytic) – no cross wall
- ZYGOMYCETES/PHYCOMYCETES (sporangiophore)
3. Hyaline- transparent hyphae
4. Dematiaceous – pigmented hyphae

5. SPORE
1. Sexual spore
– Ascospore
– Basidiospore
– Oospore and zygospore
2. Asexual spore
– Chlamydospore- rounding off of terminal hyphae
• Intercalary- chlamydospore within hyphae
• Sessile- side of hyphae
– Arthrospore – barrel shape, produced from
fragmentation of hyphae
– Blastospore – budding off
– Macroconidia and microconidia - Dermatophytes

6. Diagnostic Mycology
A. Microscopic examination
1.10% KOH prep- skin, hair (5-10 mins)
2. Calcofluor White – fluor dye 1min
3. India ink – Cryptococcus, 1min
4.Giemsa / Wright – Histoplasma

7. Most common direct method
Wet mount method
10% - skin hair, 20-70% = nails
Clearing agent/ dissolve tissue
Heating- increase rate of clearing
(+) hyphae , spore, yeast cells
Alternate: NaOH
KOH

8. Sensitive stain
Yellow green fluorescence
Binds to chitin cell wall
Calcoflour white

9. Diagnostic Mycology
5. Gram stain (yeast) - PURPLE , gram (+)
6. Histologic – Periodic Acid Schiff ,
H & E, Gomori Silver
7. Wood’s lamp (UVL) - fluorescence of fungi
8. Lactophenol cotton blue (LPCB)- common
cotton blue = AMAN stain

10. Periodic Acid Schiff – pink to red
w/ glycogen- purple
H & E- pinkish blue/ purple
Gomori Silver – brown to black

11.  Lactic acid- preservative
 Phenol- killing agent
 Cotton blue- coloring agent
 Held on cultures
LPCB Component

12. B. Use of Culture Media
1. Sabouraud’s dextrose agar (SDA)= pH 5.6
2. Mycosel – SDA + cycloheximide &
chloramphenicol- dermatophyte
3. Potato Dextrose agar - pigment
4. Corn meal tween 80 agar – chlamydospore
of C. albicans
5. Rice med- M. audouinii (-), M. canis (+)
6. Staib’s niger seed – C. neoformans

13. 7. Czapek’s medium- Aspergillus
8. Brain heart infusion agar - yeast
9. Casein medium - Nocardia
10. Urea agar – T. m and C. n
11.Cotton seed agar – B. dermatitidis
12. DTM- phenol red (indicator)
13. 1% glucose in corn meal- red T. rubrum
14. Malt Extract agar- M. furfur
Note: Hay Infusion Agar-slime molds, not a
fungal media

14. ID OF YEAST
1. GERM tube test- serum test
2. Cornmeal agar = chlamydospore test
3. Biochemical test (API20C, ID32C, VITEK)
4. CHROMagar Candida
5. PCR
6. SEROLOGY (Ag test)
– Mannan Ag- Candida
– Galactomannan – Aspergillus
7. CHO Assimilation test- NON CHO MEDIA

15. ID OF MOLDS
1. Culture on SDA = 28-30 days
incubation at RT before reporting as
negative (TAT)
2. Stain with LPCB
3. Microscopic morphology- basis for ID
of Genus and species

16. Fungicidal agent- ergosterol
1. Amphoterecin B – systemic fungi
treatment
2. Nystatin
3. Azole (Fluconazole) – fungistatic
4. Griseofulvin- dermatophyte (IV)

17. AST (MIC) methods
1.Broth microdilution
2.E test method
3.Colorimetric

18. CLASSIFICATION OF FUNGI

19. A. SUPERFICIAL MYCOSES
Non invasive
NO immune response from the host
Person to person contact
Contaminated garments (inanimate)

20. 1. Malassezia furfur
• Cause ptyriasis (tinea) versicolor
• hypo- or hyperpigmentation on skin
• KOH: budding yeast cells and hyphae
• spaghetti and meatballs
• Needs lipids
• (+) SDA with olive oil
• Bowling pin or pop bottle

21. 2. Piedra agent
• Black piedra- Piedraia hortai ( black colony),
hard, ascospore
• White piedra- Trichosporon beigelii ( cream),
soft, arthrospore

22. 3. Hortaea(Exophiala) werneckii
Cause tinea nigra
Brownish spot (dark pigmentation)
Dematicaeous
Moist, shiny-black and yeast like colonies

23. B. CUTANEOUS MYCOSES
Dermatophytes- Keratinophilic
Tinea or ringworm
Macroconidia and Microconidia
–Trichophyton - skin, hair and nail
–Microsporum - skin and hair only
–Epidermophyton - skin and nail only

24. Ectothrix
M. gypseum
M. canis
T. verrucosum

25. Endothrix
T. tonsuran
T. violaceum

26. 1. Trichophyton rubrum
• tear-drop shaped microconidia (side)
• Fluppy white w/ reddish colored reverse

27. 2. Trichophyton mentagrophytes
Grape like (cluster )microconidia

28. Trichophyton spp.
Hair
Perforation /
Baiting
Red
pigment
Urease
(red)
T.
mentagrophytes
(+) V shape - +
T. rubrum (-) + -

29. 3. Trichophyton tonsuran
(thiamine)
• Balloon shape microconidia
• Black dot tinea capitis
• #1 Tinea capitis agent

30. 4. Trichophyton schoenleinii
Cause favus (hair)
Favic chandelier
hyphae
No macroconidia
and microconidia

31. T. verrucosum- thiamine & inositol
Clavate/pyriform
microconidia
Rat tail /string
bean shaped
macroconidia

32. 1. Microsporum canis (zoophilic)
Growth on rice grain medium
Spindle shape (FUSIFORM)macroconidia,
thick, echinulated w/ 8-12 septa
(+) Fluoresces in UVL

33. 6. Microsporum gypseum
• Geophilic, Do NOT fluoresce under UVL
• Oblong (ellepsoidal) macroconidia, thin,
smooth walled w/ 4-6 septa

34. 7. Microsporum audouinii
Anthrophophilic
(-) rice grain medium
Tinea capitis (old)

35. 8. Epidermophyton floccosum
Club shape (clavate)macroconidia in pairs
Dutch pants fuseaux

36. Lab. Diagnosis
10% KOH– hyaline, septate hyphae
Culture on Sabouraud’s agar or mycosel – RT
Wood’s lamp (UVL) – fluorescence
Treatment:
• local antifungal– miconazole, tolnaftate, etc
• Oral – griseofulvin, ketoconazole

38. C. Subcutaneous Mycoses
•Skin trauma/prick
•Soil- habitat
•Biopsy , granules ( PAS,
H and E)

39. 1. Sporothrix schenckii
tissue - cigar shaped bodies (asteroid body-
product of Ag-Ab reaction)
mold form - flowerette conidia (daisy-like)
ROSE GARDENER’S disease - cord-like multiple
subcutaneous nodules (sporotrichosis)
White colonies ( white to black)

40. Asteroid body- central, basophilic
yeast surrounded by radiating
eosinophilic material due to Ag-Ab
reaction
Note:

41. 2. Mycetoma agents (maduramycosis)
• FUNGAL: Eumycotic mycetoma
–PSEUDALLESCHERIA BOYDII - most
common cause, (+) cleistothecia w/
ascospore
–MADURELLA, LEPTOSPHAERIA, etc.
• BACTERIAL: Actinomycotic mycetoma
–ACTINOMYCETES (Nocardia,
Streptomyces) can cause similar infection

42. Mycetoma agents
• tissue form: GRANULES
• lesion: granulomatous lesions on foot
with multiple draining sinus tracts

43. March 2012
The most common cause of mycetoma
( Maduramycosis) in the U.S is:
A. Nocardia asteroids
B. Rhinosporidium seeberi
C. Pseudoallescheria boydii
D.Actinomadura madurae

44. Chromoblastomycosis agent
 Dematiaceous fungi
 Type of sporulation- ID genus and species
1. Phialophora verrucosa- vase like
2. Fonsecae pedrosoi – short chain
(acrotheca, common agent)
3. Cladosporium carrionii – long chain
 Infected tissue: brown sclerotic bodies
 Lesion: cauliflower like lesion
 Dark colonies w/ jet black reverse

45. FUNGI: Subcutaneous mycosis: CHROMOBLASTOMYCOSIS:
 member:
A. Cladosporium carrionii
 long chains of conidia

46. FUNGI: Subcutaneous mycosis: CHROMOBLASTOMYCOSIS:
B. Phialophora verrucosa
 flask-shape to tubular phialides

47. FUNGI: Subcutaneous mycosis: CHROMOBLASTOMYCOSIS:
C. Fonseca pedrosoi
 short chains of conidia

48. 4. Rhinosporidium (aquaspersa)
seeberi
• Rhinosporidiosis
• lesion: polypoid masses in the nose and pharynx
• tissue form: sporangium - sac-like structure filled
with endospores (300um)
300 um

49. 5. Loboa loboi (lobomycosis)
• lesion: keloid-like subcutaneous nodule
usually involving the extremities
• tissue form: multiple budding cells in chain

50. D. Systemic mycoses
• Inhalation of spore (mold)- infectious- not
cultured in the lab.
• Tissue (yeast) = diagnostic , cultured in lab
• No person to person contact
• Biosafety level III ( BSC class II)
• Sputum- common specimen
• Exoantigen test- immunodiffusion test

51. Exoantigen Test
a)A Ag- B. dermatitidis (1:8)
b)1, 2 & 3 Ag- P. brasiliensis
c) H & M Ag- H. capsulatum (1:8)
d)HS, HL, F Ag – C. immitis (1:2)
* H. capsulatum cross react w/ B.
dermatitidis

52. March 2012
Which of the following tests may be
used instead of conversion when
identifying dimorphic fungi?
A. String’s Test
B. Exoantigen Test
C. ELISA
D.Immunofluorescent Test

53. 1. BLASTOMYCES dermatitidis
North American Blastomycosis (Missouri River
valley), Chicago Disease, Gilchrist’s disease-
PNEUMONIA, SKIN INFECT,
tissue form: SINGLE-BUDDING YEAST with BROAD
based (double countered)
mold form: lollipop in appearance

54. 2. PARACOCCIDIOIDES brasiliensis
South American Blastomycosis, Lutz Splendore-Almeida
Disease- spleen, liver, lymph node, skin, lung
tissue form: MULTIPLE BUDDING YEAST (mariner’s, pilot’s,
ship’s and navigator’s wheel or mickey mouse cap)
mold form: lollipop in appearance (pyriform conidia)

55. HISTOPLASMA capsulatum- fungus
flu
RES parasite
DARLING’S , SPELUNKER’s
(cave) DSE
Blood smear stained with
Giemsa
tissue form: yeast cells
intracellular in
macrophages
mold form: tuberculate
macroconidia (+)
SEPEDONIUM-
monomorphic &
HISTOPLASMA-dimorphic

56. Coccidioides immitis- Major biohazard in lab
 COCCIDIOIDOMYCOSIS ( semiarid areas)- humidity
San Joaquin Valley Fever (Desert fever)
tissue form: SPHERULE with endospores (200um)
mold form: barrel-shaped arthroconidia (infectious)
Laboratory acquired infection
Cob web colony

57. C. immitis

58. Dx of Systemic Mycoses
Direct exam of clinical specimens
Histoplasma – Wright or Giemsa
Blastomyces, Paracoccidioides
 Coccidioides - 10 % KOH, PAS, H and E
Cultures
SDA – RT
BHIA + blood – 37oC
Immunological tests
Skin test –coccidioidin, histoplasmin

59. Opportunistic Mycoses
• Normal flora
• Immunocompromised person (at risk)

60. 1. Candida albicans
• normal flora: skin &mucous memb. (GI)
• produce yeast and hyphae in vivo
1. Germ tube, 2. chlamydospore
2. Blastoconidia, 3. pseudohyphae
true hyphae
• Sucrose (+), feathering on EMB
• Star like or feet like projection on BAP

61. DISEASES:
• Thrush, moniliasis, diaper rash
-CANDIDIASIS
– ONCHOMYCOSIS
– VULVOVAGINITIS
– INVASIVE- CNS, BLOOD
• Prolong antibiotic use or broad spectrum
antibiotic, pregnancy, DM, MALNUTRITION

62. 1. Aspartyl protease
2. Hydrophobicity
3. Phospholipase
4. Hypae & pseudohyphae
Virulence factor of C. albicans

63. 10% KOH – Positive hyphae, yeast, spores
SDA, BAP: smooth, round, moist, creamy
Germ Tube test – Screening test
 Serum (0.05ml) + yeast (37’C) 2-3 hrs
(+) : tube-like projections, no constriction(germ tube)
Corn Meal Agar: Confirmatory test
2-4 streak on corn meal agar (RT) 2-3 days
(+) chlamydospore, hyphae, blastoconidia
Candida spp

64. Lab. Dx of C. albicans
1. Screening test: Germ tube test
Org + serum inc. at 35’C for 2-3 hrs (+) germ tube

65. (+) GERM TUBE TEST
1. CANDIDA albicans (+) chlamydospore,sucrose
2. CANDIDA stellatoidea (-) chlamydospore,sucrose
3. GEOTRICHUM candidum (+) arthrospore
4.Candida dubliniensis

66. Lab. Dx of C. albicans
• Confirmatory: Chlamydospore on Corn meal
C. albicans col  inoculate on corn meal 
incubate at RT for 48-72 hrs  chlamydospore

67. C. dubliniensis C. albicans
Chlamydospore 2 1
Xylose (-) (+)
Alpha methyl- d -
glucoside (+)
(-) (+)
Growth at 42 (-) (+)

68. LAB. DIAGNOSIS
1. RULE OUT VAGINOSIS/ TRICHOMONIASIS
2. VAGINAL pH 4.5
3. Vaginal discharge- 10% KOH
4. FUNGAL CULTURE
*Latex agglutination test for C. albicans =
1:8 but other fungal infection 1:32
significant titer

69. Acridine Orange- best stain used to
detect Candida in bloodstream infect.
C. albicans- principal agent of fungemia
C. glabrata – (R ) to antifungal agents
C. tropicalis- hematologic malignancy
C. parapsilosis- primary cause of fungemia in NICU
C. krusie- endocarditis
Note:

70. 2. Cryptococcus neoformans
pigeon droppings, soil
transmission: inhalation of airborne org
ENCAPSULATED YEAST CELL (India Ink)
MENINGITIS, Torulosis(cryptococcosis),
pneumonia

71. India Ink prep= capsule
latex agglutination= capsular Ag
(+) urease, (-) nitrate , (+) phenol oxidase, (+) inositol
(+) birdseed/ niger seed agar- black colony
due to phenol oxidase
Yeast like, mucoid, cream to brown col
CULTURE-SDA w/o cycloheximide
Diagnosis:

72. March 2012
Cryptoccocus neoformans latex
agglutination test on spinal fluid detects
cryptoccocal:
A. Creatinine
B. Carbohydrate
C. Antigen
D. Antibody

73. ASPERGILLUS- #1 mold
• Dichotomous septate hyphae (tissue); vesicles
1. A. fumigatus -fungus ball, aspergilloma, allergy (asthma),
otomycosis , bread mold; farmer’s lung disease
2. A. flavus (A. parasiticus)- aflatoxin (toxicoses), turkey x
disease (peanuts), commercial food poisoning
3. A.niger - brown to black spore

74. 4. Zygomycosis/Mucormycosis
• agents: RHIZOPUS, MUCOR, ABSIDIA (Zygomycetes)
• transmission: inhalation of airborne conidia
• tissue form: NONSEPTATE HYPHAE
• Rhinocerebral (rhino-facial-cranial), lung, GIT, skin

75. FUNGI: Opportunistic mycosis:
Penicillium (Penicillium marneffei)
• brush like conidiophore( branched, hands of skeleton)
• white to bluish green, yellow, brown colony

76. Fusarium
White, cottony
to pink or purple
colony
Sickle or canoe
shaped,
multiseptate
macroconidia

77. PHAEOHYPHOMYCOSIS
- fungal infection caused by dematiaceous
agents:
A- ALTERNARIA
B- BIPOLARIS
C- CURVULARIA
D- DRESCHLERA
E-EXOPHIALA

78. Pneumocystis jiroveci
Protozoan cyst, No ergosterol
# 1 cause of pneumonia in AIDS
#1 opportunistic infection in AIDS
No growth on fungal media
No hyphae and spore
BAL- best specimen ; cyst/trophozoite

79. 30’C for 21-30 days – report blood culture as
negative
Cryptococcus, Aspergillus, Candida- inhibited by
cycloheximide
Calcofluor- white- most sensitive stain for fungi on
clinical specimen; has bleaching agent
In vitro hair perforation- differentiate T.
mentagrophytes & T. rubrum
Wood’s lamp- (+) M. canis , (+) M. audouinii (-) M.
gypseum
Note: Diagnostic Mycology

80. PNA FISH Kit- ID of Candida spp
 Emmonsia crescens- produces an
unusually large spore when inhaled into
the human respiratory tract
Histoplasma capsulatum -chicken coops
Fusarium spp- Sickle or boat-shaped
macroconidia
Note: Diagnostic Mycology

81. Phialide- tubular conidiphore
Black grain mycetoma- Exophiala jeanselmei,
Curvularia spp. & Madurella spp.
White grain mycetoma- P. boydii,
Acremonium spp.
Note: Diagnostic Mycology

82. BAL- best specimen for P. jirovecii
Gomori methenamine silver (GMS) stain,
immunofluorescent stain, Wright-Giemsa (Diff
Quick) stain, calcofluor white (Fungi-Fluor) –
stains for P. jirovecii
Penicillium marneffei- yeast like cells with cross
walls, immunosuppressed, bamboo rat, (+) blood
smear with disseminated disease, NA test –
definitive
C. immitis- cob web colony
Note: Diagnostic Mycology

83.  Star like or feet like projection on agar- C.
albicans
Bowling pin or pop bottle- M. furfur
Niger seed agar test- definitive test for phenoll
oxidase of C. neoformans
(+) urease- Cryptococcus, Rhodotorula,
Trichosporon
Note: Diagnostic Mycology

84. Which dimorphic fungus is found in the
Missouri River valley?
A. Blastomyces dermatitidis
B. Histoplasma capsulatum
C. Coccidioides spp.
D. Paracoccidioides brasiliensis
Bailey & Scott’s

85. Which test can be used to differentiate T.
mentagrophytes from T. rubrum?
A. Fluorescence using a Wood lamp
B. In vitro hair perforation
C. Red color on reverse side of colony
D. Pyriform microconidia
Bailey & Scott’s

86. Microsporum infection involves:
A. Hair, skin, and nails
B. Hair and skin
C. Skin and nails
D. None of the above
Bailey & Scott’s