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Clinically Significant Yeast

Candida spp.

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Where does it come from?

Normal flora of mucosa, skin, GI tract
Intact immune system: localized, limited infection
Sites affected include genital area, mouth, and skin.

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What else can it do?

Immuno-compromised patients can experience systemic infection, specifically, candidemia

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Why is it important?

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Gotcha!

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Why is it important?�Candida

C.auris

primary agent of yeast infections
Implicated in HAI outbreaks
Resistant to antifungal agents
Difficult to identify

C. albicans
C.krusei
C.dubliniensis
C. glabrata
C. tropicalis
C.lusitanae

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How do we recover it?

Culture methods:

Enrichment/selective media

EX. Sabaroud Dextrose
EX. BHI w 10% blood + gent/chlor

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How do we detect it?

Culture methods:

Chromogenic agars

EX. CandiSelect
EX. Candida ChromAgar
EX. Brilliance Candida

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How do we speciate?

Germ tube test

A germ tube is a projection off the yeast that will become psuedohyphae characteristic of Candida albicans.

Simple, cost-effective test → presumptive ID

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How do we speciate?

Germ tube test: Procedure

Materials needed: serum, yeast isolate, tube, microscope
Tube + serum + isolate → incubate at ~35⁰C, 2 hrs

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Germ Tube Test Interpretation

Positive Test: A short hyphal (filamentous) extension arising laterally from a yeast cell, with no constriction at the point of origin. Germ tube is half the width and 3 to 4 times the length of the yeast cell and there is no presence of nucleus.

Examples: Candida albicans and Candida dubliniensis

Negative Test: No hyphal (filamentous) extension arising from a yeast cell or a short hyphal extension constricted at the point of origin.

Examples: C. tropicalis, C. glabrata and other yeasts.

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How do we speciate?

Candida albicans screen test or CAST

Detect enzymes L-proline aminopeptidase and β-galactosaminidase

Simple, cost-effective test → presumptive ID

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How do we speciate?

CAST test: Procedure

Materials needed: kit including substrate-impregnated discs, DI water, kit reagent, heat block or incubator
Add 1 disc, 1 drop DI water and a colony of isolate
Incubate ~30 minutes
Add Reagent 1 (NaOH) → YELLOW (+)
Add Reagent 2 (color developer/cinnamaldehyde) →HOT PINK/RED (+)

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PNA technology

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How does PNA apply to yeast ID?

3 different PNA probes complementary to unique and specific gene sequences of different yeasts are combined in this particular test. Each of the probes also have a different color fluorescent tag. The probe solution is combined with a positive blood culture reflecting yeast in the gram stain. The blood culture sample + probe mix is incubated, washed, mounted and read with a fluorescent scope.

C. glabrata and C. krusei are so genetically similar it is difficult to differentiate, etc. The upside is that these groupings are treated similarly: C.glab/C.krus treated with echinocandin. C.alb/C.para treated with fluconazole.

PNA FISH probes hybridize to target DNA with high sensitivity and specificity mainly due to the electrical neutrality of its chemical structure. This trait enables PNA probes to be more efficient and useful in applying to FISH (fluorescent in situ hybridization) even at low concentrations.

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Why do we need an ID so fast?

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Treatment�How does speciation affect Rx?

Speciation of the fungus is important to get a quick ID
Quick IDs lead to putting the patient on the right antifungal- decreasing inpatient stays and hospital bills

Echinocandins are used to treat C. auris infections

Expensive
IV administration

Fluconazole

Alternate treatment with both oral and IV option
Seeing↑ fluconazole resistance