Clinical Pathology(Micro)

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A Sabouraud’s dextrose agar plate inoculated with a Zygomyces species: Notice that the entire plate is filled with a wooly, gray-white surface mycelium.

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Close in view of a colony of a Zygomyces species after 96 hours of incubation: As a Zygomyces species colony matures, it takes on a gray-brown color, often with black, pepperlike stipples, indicating the production sporangia.

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The surface of a colony of Aspergillus fumigatus after a 5-day incubation on Sabouraud’s dextrose agar: Mature colonies of A. fumigatus are generally pow­dery or granular, and have some shade of green from the production of pig­mented conidia. The growing margin often appears as a white apron, as illus­trated here.

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The surface of a colony of Aspergillus flavus growing on Sahouraud’s dextrose agar: The texture of mature colonies is usually granular from the production of conidia; and, as the species name suggests, some shade of yellow pigmentation is characteristic.

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The surface of a colony of Aspergillus niger after a 4-day incubation on Sabouraud’s dextrose agar: The deep brown to black, densely stippled surface is characteristic. The black surface pigmentation of A. niger can be distinguished from one of the dematiaceous fungi by observing the reverse side. The reverse of A. niger is a light gray or buff color as the pigmentation is caused by surface conidia; the reverse of dematiaceous fungi is jet black, as the vegetative hyphae carry the pigment.

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The surface of a colony of Aspergillus terreus after a 6-day incubation on Sabouraud’s dextrose agar: A. terreus characteristically has a granular surface and some shade of yellow or brown pigmentation. Rugae radiating from the center are often seen, and the concentric zones of light and dark pigmentation as seen here are not unusual.

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The surface of an albino strain of Aspergillus nidu!ans: Although most strains have some light pigmentation, albino strains of most Aspergil!us species may oc­casionally be encountered and must be kept in the differential identification. The granular surface, with radiating rugae, is also common.

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The surface of a colony of Aspergillus glaucus: The green and yellow variegated appearance seen here is not specific, because other Aspergillus species may have a similar appearance. Microscopic examination is necessary to make a species identification.

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The surface of a colony of Penicillium species after a 5-day incubation on Sabouraud’s dextrose agar, illustrating the distinctive green color, granular sur­face, radial rugae and white apron at the periphery. Although most strains of Penicilliurn species display some shade of green, yellow variants are sometimes seen.

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The surface of a colony of Pacci1o~nyces species: The colonies of Paecilomyces species are usually granular and often have a yellow-brown pigmentation, as illustrated here. Other strains, however, may have a light pastel green, green-yellow, or bluish appearance.

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The surface of a colony of Scopulariopsis species after 5 days of growth on Sabouraud’s dextrose agar: The yellow pigmentation, granular texture, and ra­diating rugae, as illustrated here, are hallmarks of Scopulariopsis species.

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The surface of a colony of Acromoniun, species after 5 days of incubation on Sabouraud’s dextrose agar: The colonies of Acremonium species usually have a smooth, almost yeast-like appearance, as illustrated here, owing to the ex­tremely delicate nature of the hyphae and fruiting bodies. Light pastel yellow, light green, and peach-red hues may be observed.

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The surface of a typical colony of Fusarium species after 6 days of growth on Sabouraud’s dextrose agar: Fusarium species can be suspected when a rapidly growing, granular or fluffy colony, with a distinct rose-red, lavender, or purple pigmentation is observed.

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The surface of a typical fluffy, “house mouse gray” colony of Scedosporium apiosperum (anamorphic form of Pseudallescheria boydii) after 5 days of incuba­tion on Sabouraud’s dextrose agar: A rapidly growing colony with the mouse gray appearance, as illustrated here, always suggests Scedosporium species. The unique pigmentation is caused by darkly pigmented conidia.

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A colony of Gliocladium species after 5 days of growth on Sabouraud’s dextrose agar: The green pigment, extension of the colony from border to border without a well defined growing margin and granular texture (so-called green lawn) is unique for Gliocladium and Trichoderma species. The latter more frequently may show a yellow, rather than a green, pigmentation.

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Which of the following media is most helpful in distinguishing the morphology of yeasts?
A. cottonseed agar
B. cornmeal agar with Tween 80
C. urea agar
D. brain-heart infusion medium
E. potato dextrose agar

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B. cornmeal agar with Tween 80.
Cottonseed agar is used specifically to convert the mold phase of Blastomyces to the yeast form. Cornmeal with Tween 80 stimulates conidiation and chlamydospore
production, aiding in speciation. Urea agar is helpful in detecting urease which is produced by Cryptococcus neoformans. Brain-heart infusion is a non-selective medium
which will support the growth of saprophytic and pathogenic fungi. Potato dextrose agar is useful in demonstrating the production of pigment by Trichophyton rubrum.
QCCP2, T3.21, Special fungal culture techniques

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Which of the following fungi are inhibited by cyclohexamide?
A. Zygomyces
B. Aspergillus
C. Cryptococcus
D. A & B
E. A, B, C

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E. A, B, C.
Most fungal pathogens, especially dimorphic fungi or dermatophytes will grow, albeit slowly, in cyclohexamide. Zygomyces, Aspergillus, and Cryptococcus won't grow.

Front 18

Which of the following dimorphic fungi needs to be plated on brain-heart infusion media with blood in order to grow as a yeast?
A. Blastomyces dermatidis
B. Histoplasma capsulatum
C. Coccidioides immitis
D. Sporothrix schenkii
E. Paracoccidioides brazilensis

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B. H.
Blastomyces grows as a yeast on cottonseed agar while Coccidioides requires specialized media to grow as a yeast. The rest of the dimorphic fungi readily grow as yeast.

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Which of the dimorphic yeasts is often confused histologically with the mold form of another dimorphic fungus?
A. Histoplasma
B. Blastomyces
C. Coccidioides
D. Sporothrix
E. Paracoccidioides

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E. P.
The yeast form of Paracoccidioides contains the characteristic “Mariner's wheel” configuration of irregular circumferential cytoplasmic blebbing. Similarly, the mold form of
Histoplasma has circumferential cytoplasmic buds which are more frequent and regular than those of the Paracoccidioides yeast.
Histoplasma has circumferential cytoplasmic buds which are more frequent and regular than those of the Paracoccidioides yeast.

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Which of the dimorphic fungi are associated with sclerosing mediastinitis?
A. Histoplasma
B. Blastomyces
C. Coccidioides
D. Sporothrix
E. Paracoccidioides

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A. H .
Most dimorphic fungi with the notable exception of Sporothrix are infective through pulmonary aspiration - Histoplasma can cause the formation of pulmonary nodules, or
sclerosing mediastinitis. It can also be disseminated through the reticuloendothelial system, where it can migrate to the spleen or bone marrow.

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Which if the following organisms is in the differential diagnosis of Coccidioides spherules?
A. Rhinosporidium seeberi
B. Histoplasma capsulatum
C. Prototheca wickerhamii
D. A & B
E. A, B, C

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E. A, B, C.
All resemble the 50-200 micron spherules that contain the small 2-5 micron endospores. Rhinosporidia is larger and found in the nasal sinus, Prototheca in the olecranon
bursa. The wall of the spherule of Coccidioides is thick, but when the spherules are released. they can resemble those of Histoplasma.

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Which of the following dimorphic fungi presents the highest risk to laboratory personnel during culture?
A. Histoplasma
B. Blastomyces
C. Coccidioides
D. Sporothrix
E. Paracoccidioides

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C. C.
The mold form of Coccidioides forms barrel-shaped arthroconidia of live cells alternating with empty shells. The shell portions break easily, releasing the infective
arthroconidia into the air where they can be freely inhaled unless proper precautions are taken.

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Which of the following organisms grows as a mold with smooth, “lollipop”-shaped conidia?
A. Chrysosporium
B. Blastomyces
C. Sporothrix
D. A & B
E. A, B, C

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D. A & B.
A number of other molds, such as Paracoccidioides and Scedosporium can produce “lollipop” conidia. Sporothrix, however, produces a very characteristic “daisy-head”
conidia that is not easily confused with Blastomyces.

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In what patient population has an unusual inhalational form of Sporothrix been identified?
A. the very young
B. elderly
C. immunocompromised
D. chronic alcoholics
E. smokers

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D. chronic alcoholics.
Although Sporothrix is unique among the dimorphic because its primary means of infection is percutaneous, a less common means of inhalation (like the rest of the
dimorphic fungi) has been documented in chronic alcoholics.
dimorphic fungi) has been documented in chronic alcoholics.

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The mold form of Paracoccidioides is identical to that of which other fungus?
A. Histoplasma
B. Blastomyces
C. Coccidioides
D. Sporothrix
E. Chrysosporium

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B. B.
Paracoccidioides has been called the “South American Blastomyces” because it looks and behaves very similarly to Blastomyces.

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Which of the following dermatophytes is identified by its macroconidia?
A. Trichophyton tonsurans
B. Trichophyton rubrum
C. Epidermophyton floccosum
D. A & B
E. A, B, C

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C. E.
Trichophyton species (T. rubrum, T. tonsurans, T. mentagrophytes) are identified by their microconidia, while E. floccosum and the Microsporum spp are identified by
their macroconidia. E. floccosum has “beaver tail” macroconidia with transverse septae; Microsporum canis has transverse septa also, but with serrated edges (“dog
teeth”) and pointed ends. Microsporum gypseum is very similar to M. canis but without the serrations

Front 27

Which of the following dermatophytes can be identified with its “birds on a wire” microconidia?
A. Microsporidium canis
B. Trichophyton rubrum
C. Epidermophyton floccosum
D. Trichophyton mentagrophytes
E. Trichophyton tonsurans

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B. T.
The Trichophyton spp have microconidia and, of them, each has unique morphological features. T. rubrum has “birds on a wire” microconidia spaced along hyphae; T.
tonsurans has widely variable microconidia, while T. mentagrophytes has grape-like clusters and occasional spiral hyphae.

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What organism is the most common cause of onychomycosis?
A. Epidermophyton floccosum
B. Trichophyton rubrum
C. Trichophyton mentagrophytes
D. Microsporum canis
E. Microsporum gypseum

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B. T.
The Trichophyton spp, especially T. rubrum, cause the majority of cases of onychomycosis. Individuals with HIV and diabetes are at an increased risk for onychomycosis.
In addition, it is important to note that other fungi, such as Candida, can cause onychomycosis.

Front 29

Which of the following organisms is characterized by blue-green colonies with a white apron?
A. Aspergillus terreus
B. A. niger
C. A. fumigatus
D. A. flavus
E. Penicillium marneffei

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C. A.
Each of the Aspergillus spp has a distinct appearance on plates. A. fumigatus is blue-green, A. terreus has a cinnamon-buff colony, A. niger has a black colony, and A.
flavus has a brown colony with lateral striations.
flavus has a brown colony with lateral striations.

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Which of the following organisms is characterized by two rows of phialides on the conidia?
A. A. terreus
B. A. niger
C. A. fumigatus
D. A. flavus
E. P. marneffei

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A. A.
Just like the discrete gross appearance on plates, each of the Aspergillus spp can be speciated by the appearance of the conidia. A. terreus has 2 rows of phialides. A.
niger has black circumferential phialides. A. fumigatus has a single row of phialides. A. flavus has a circumferential row of phialides (a “flavorful lollipop”). P. marneffei
looks like A. fumigatus but lacks the swollen vesicle at the base of the phialides that all the members of Aspergillus have.

Front 31

Which species is responsible for the majority of cases of aspergilloma and allergic bronchopulmonary aspergillosis?
A. A. flavus
B. A. fumigatus
C. A. niger
D. A. terreus
E. P. marneffei

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B. A.
There are 3 principle pulmonary diseases caused by Aspergillus spp, primarily A. fumigatus: Aspergilloma, a fungal ball that grows in the site of a pre-existing cavitary
lesion; Allergic bronchopulmonary aspergillosis, an exaggerated allergic response to noninvasive Aspergillus colonization that's seen mostly in patients with cystic fibrosis;
and finally, invasive aspergillosis, a vascular-invasive disease seen mostly in the immunocompromised.

Front 32

Which species of Aspergillus is most commonly seen as otitis externa?
A. A. flavus
B. A. fumigatus
C. A. niger
D. A. terreus
E. P. marneffei

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C. A.

Front 33

All of the following hyalinohyphomyces have conidia that occur in clusters, except:
A. Acremonium
B. Penicillium
C. Gliocladium
D. Fusarium
E. all of the above occur in clusters

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B. P .
If anything called for a mnemonic, it would be all these genera of hyaline molds - a “GAF” is a tool to grab things and hold them together in a cluster. The organisms
with clustered conidia are Gliocladium, Acremonium, and Fusarium. The branching chain conidia are a further reach - imagine looking through a pay telescope to see a
pen hanging from a chain on a tree branch. Interpretation - the branching chain conidia are Penicillium, Paecilomyces, and Scopulariopsis. All the rest occur singly -
Scedosporium, Beuveria, Sepedonium, and Chrysosporium.

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Fluffy, gray-black surface of a colony of Alternaria species after 6 days growth: The appearance shown here is not distinctive for Alternaria species, but may also be observed with other species of environmental dematiaceous molds.

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Reverse surface of a dematiaceous mold illustrating the dark appearance caused by the dark brown pigmentation of the vegetative hyphae.

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Surface view of a subculture of Ulocladiun species, illustrating a wooly, dark brown to black pigmentation: Although frequently seen with certain strains of dematiaceous fungi, the light and darker concentric rings observed here are not distinctive for any given species.

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Surface view of a colony of Phyalophora verrucosum after 14 days incubation: This particular strain has a dark gray-black hue, with a hairlike texture. Microscopic study is necessary, because other slow-growing species may produce similar-appearing colonies.

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Surface view of a colony of Fonsecaea pedrosoi after 16 days incubation: The colony is quite small, typical for the species, and the typical dark brown-black pigmentation is evident. This strain has a flatter, almost suedelike consistency.

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Surface view of a colony of a more rapidly growing (5 days), environmental strain of Cladosporiom species illustrating a dark green, suedelike surface inter­rupted by irregular rugae: Environmental strains of C!adosporium species may appear dark green, as shown here, or may be gray, gray-brown, or brown-black, simulating other dematiaceous fungi.

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Surface view of another variant of an environmental Cladosporium species show­ing a dark gray, leatherlike consistency: Colony texture among the dematiaceous molds range from wooly, to downy or hairlike, to suedelike, to leathery.

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Appearance of the flat, smooth, yeastlike surface of Aureobasidium pellulans after 6 days incubation. Aureobasidium spp. should be first on the list when a “black yeast,” similar to that illustrated here, is observed; however, certain strains of Exophiala wernickii (so-called Phaeoannelomyces synanamorphs) must also he considered, which will develop a low, hairlike mycelium with true hyphae after several additional days of incubation.

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The surface of a colony of Penicillium marneffei after a 4-day incubation on Sabouraud’s dextrose agar, illustrating the distinctive red pigmentation of a portion of the colony: Note that the pigment has leached out into the agar, which has a light wine red coloration. The surface tends to he granular to fluffy depending on the degree of sporulation; note the distinct white apron at the margin of peripheral new growth.

Front 43

Which organism is the cause of adiaspiromycosis?
A. Fusarium
B. Chrysosporium
C. Beuveria
D. Paecilomyces
E. Gliocladium

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B. C.
Adiaspiromycosis is a benign granulomatous infection with characteristic large thick-walled spherules.

Front 44

Which of the following fungal organisms is a major concern as an opportunistic infection in burn victims?
A. Gliosporium
B. Paecilomyces
C. Scopulariopsis
D. Scedosporium
E. Fusarium

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E. F.
In addition to the myriad of infections including a disseminated infection in immunocompromised hosts, Fusarium can cause infections in burn victims, pulmonary
infections, skin infections, and fungemia.

Front 45

What is the most common cause of eumycotic mycetoma in the U.S.?
A. Fusarium
B. Scopulariopsis
C. Acremonium
D. Scedosporium
E. Paecilomyces

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D. S .
It is also a common cause of disseminated infection in near-drowning victims and infections related to penetrating trauma.

Front 46

Which of the following dematiaceous molds is responsible for chromoblastomycosis?
A. Fonseca
B. Phialophora
C. Cladosporiom
D. Wangiella
E. Exophiala

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E. A, B, C.
Chromoblastomycosis is a cutaneous fungal infection with the “copper penny” sclerotic or Medlar bodies that represent pigmented subcutaneous septated yeast forms.

Front 47

All of the following are causes of eumycotic mycetoma, except:
A. Madurella
B. Actinomyces
C. Scedosporium
D. Wangiella
E. Exophiala

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B. A.
Mycetomas, or deep subcutaneous infections, can be categorized as either true (eumycotic - caused by fungus) or actinomycotic (caused by non-fungal organisms). All
of the choices presented, with the exception of Actinomyces, are fungal, or eumycotic, causes of mycetoma. The bacterial causes also can include Streptomyces and
Nocardia.

Front 48

Which of the following zygomycetes has sporangiophores that arise between rhizoids?
A. Rhizopus
B. Absidia
C. Mucormycosis
D. Cunninghamella
E. Circinella

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B. A.
Absidia and Rhizopus are the only zygomycetes that produce rhizoids or little rootlets from their hyphae. While the rhizoids in Rhizopus lie directly below the
sporangiophores, those of Absidia are offset.
sporangiophores, those of Absidia are offset.

Front 49

Under what conditions must the germ tube test be run in order to identify Candida albicans?
A. suspended in broth, incubated 2 hours at 37°C
B. suspended in serum, incubated 24 hours at 37°C
C. suspended in broth, incubated 2 hours at 32°C
D. suspended in serum, incubated 2 hours at 37°C
E. suspended in broth, incubated 24 hours at 37°C

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D. suspended in serum, incubated 2 hours at 37°C.
It's actually important to understand the conditions under which the germ tube test is performed. The reason is that overincubation will greatly decrease the nearly
100% specificity that the assay has for Candida albicans. Unlike the pseudohyphae that Candida can form in tissue, the germ tube is a true hypha (there is no “pinch”
between the germ tube and the mold).

Front 50

Which fungal organism is urease (+) and phenol oxidase (+)?
A. Cryptococcus neoformans
B. Malassezia furfur
C. Rhodotorula
D. Candida krusei
E. Trichophyton mentagrophytes

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A. C.
All of the organisms mentioned, with the exception of Malassezia, are urease positive, but Cryptococcus is the only one that is also phenol oxidase positive. The presence
of phenol oxidase can be demonstrated on bird (Niger) seed agar where the phenol oxidase will convert the caffeic acid in the agar into melanin pigment, yielding the
characteristic brown/black pigment.

Front 51

Which of the following organisms requires the overlay of olive oil in order to culture?
A. Cryptococcus
B. Malassezia
C. Rhodotorula
D. Candida
E. Trichophyton

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B. M.
Malassezia furfur is the cause of tinea versicolor, a skin infection usually found on greasy back skin, and is also associated with total parenteral nutrition (lipid-rich) line
infections. Malassezia requires a source of long-chain fatty acids to grow in culture. An olive oil overlay provides the required fatty acids.

Front 52

Which of the following stains the capsule of Cryptococcus?
A. mucicarmine
B. Fontana-Masson
C. Alcian blue
D. India ink
E. GMS

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B. Fontana-Masson.
Remember, the phenol oxidase converts caffeic acid to melanin, which is what Fontana-Masson stains. The polysaccharides in the thick capsule of Cryptococcus can be
stained with mucicarmine or Alcian blue. India ink works by forming a white negative space of the organism against a black-stained background.

Front 53

Which of the following organisms causes infections associated with penetrating trauma and is in a class totally by itself?
A. Sporothrix
B. Fusarium
C. Prototheca
D. Rhodotorula
E. Malassezia

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C. P .
Prototheca wickerhamii is the only known algal cause of human infections. Most commonly, the infection takes the form of olecranon bursitis or a cutaneous skin
infection. The organism is said to resemble soccer balls or hub caps, referring to the morulated appearance. The organism is associated with exposure to water and
dolphins

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Colony of Microsporum canis after 5 days incubation: The colonies tend to be fluffy or, at times, granular if conidiation is heavy. The lemon-yellow apron seen here, with pigment also extending to the reverse surface, is one clue sug­gesting M. canis.

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Surface of a powdery, off-yellow colony of Microsporum gypseu? after 6 days incubation: M. gypseum tends to sporulate heavily; therefore, it is more prone to produce granular colonies than M. canis.

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Surface of a smooth, white variant of Trichophyton mentagrophytes with no evi­dence of background pigmentation of the agar: The colony illustrated here is not distinctive, and microscopic study is required to make an identification.

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Surface of a fluffy variant of Trichophyton mcntagrophytes demonstrating a red­dish pigmentation of the background agar: Wine-red pigmentation is one fea­ture that suggests T. rubrum; however, certain strains of T. mentagrophytes max also produce reddish pigment when growing on Sabouraud’s dextrose agar. Pigmentation, however, is never as intense as with T. rubrum when colonies are grown on cornmeal or potato dextrose agar.

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Reverse of a colony of Trichophyton rubrum growing on potato dextrose agar. The deep wine-red pigmentation is characteristic of the species, and is particularly intense when the colony is grown on potato dextrose or cornmeal agar.

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Comparative growth of T. mentagrophytes (bottom) and T. rubrum (top) on cornmeal agar, illustrating the distinct difference in pigmentation between the two species.

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Surface of a small colony of Trichophyton tonsurans after 14 days incubation: T. tonsurans grows slower on Sahouraud’s dextrose agar. Most strains produce some shade of yellow-brown pigment. The surface tends to be granular, and radial rugae are frequently seen. Except for the slower growth, the colonies of T. tonsurans appear similar to Scopulariopsis species and Aspergillus terreus

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Surface of Epiderinophyton floccosum after 6 days incubation: This colony appears more fluffy than granular and has an off-yellow pigmentation. Classic colonies of E. floccosum are described as khaki green. The colony texture tends to be more fluffy than granular, as microconidia are not produced by this genus.

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Surface colony of a dimorphic mold illustrating both the yeast form (centrally) and the mold form (peripherally): This colony was incubated for 14 days at 30°C. When this mixture of yeast and mold texture is observed, one of the dimorphic fungi can be suspected, particularly if the colony is relatively slow growing and the mold appears more hairlike or cobweb in consistency.

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Colonies of Blastomyces dermatitidis growing on 5% sheep blood agar after 5 days incubation: Note the cobweb appearance of these colonies, one clue that the isolate may be a dimorphic fungus and alerting one to take special precau­tions when preparing subcultures or microscopic mounts to avoid a laboratory-acquired infection.

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Appearance of colonies of Blastoniyces dermatitidis incubated at 37°C, during a mold-to-yeast conversion: There is an intermediate form of the colonies be­tween mold and yeast, known as the “prickly stage.” Note the rather coarse spicules formed by these white colonies.

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Colonies of Histoplasma capsulatum on brain-heart infusion agar after 25 days in­cubation at 30oC. The colonies appear white and cobweb in consistency. These colonies are nondescript, and microscopic examination is necessary to make an identification.

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Surface of a 5% sheep blood agar plate incubated at 37°C on which are growing small, yellowish yeast colonies of Histoplasma capsulatum after successful con­version from the mold form: The yeast forms are similar to other true yeasts, ex­cept that growth is very slow and the colonies remain quite small.

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Surface of colonies of Coccidioides inimitis growing on 5% sheep blood agar after 7 days incubation: The colonies are gray-white and have a delicate hairlike tex­ture. Colonies of C. immitis growing on blood agar often show a red discol­oration from leaching of hemoglobin from the culture medium (not shown in this illustration).

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Surface of a smooth, gray-brown yeast colony of Sporothrix schcnckii Because of the extremely delicate sporulation of S. schenckii, colonies often appear more yeastlike than moidlike, even when incubated at 25° to 30°C. After prolonged incubation, colonies of S. schenckii tend to darken considerably, becoming al­most jet black with some strains (see Plate 19-5H).

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Tubes of brain—heart infusion agar, one containing cycloheximide, on which are growing the yeast (top) and mold (bottom) forms of S. schenckii. Because of the delicate nature of the hyphae and fruiting structures, the mold colony appears quite yeastlike. The colonies tend to darken with maturity, becoming distinctly black in some cases, as illustrated here.