The Differentiation of Yeast and Yeast-Like Forms in Human Tissues. Introduction. Histochemical Stains Used to Detect Fungi. Histopathologic Diagnoses

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1 The Differentiation of Yeast and Yeast-Like Forms in Human Tissues Gary W. Procop, MD Chair, Clinical Pathology Staff, Anatomic Pathology Director, Molecular Microbiology, Mycology, and Parasitology Cleveland Clinic Introduction Fungi are the ubiquitous saprophytes of nature, which aid in the degradation of dead material. Accidental Pathogens Coccidioidomycosis Opportunistic Pathogens Candida/Aspergillus Causes of Allergy Alternaria sinusitis Histopathologic Diagnoses Inflammatory Response Neutrophilic Candida and Aspergillus Granulomatous Histoplasma Pyogranulomatous Blastomyces and Sporothrix Granulomatous with Eosinophils Coccidioides immitis Histochemical Stains Used to Detect Fungi Hematoxylin and Eosin Many fungi are visible (Aspergillus / Candida) Some are not (Sporothrix / Histoplasma) Gomori s Methenamine Silver (GMS) The Best Stain for Highlighting Fungi Also highlights many bacteria (i.e. Nocardia) 1

2 Histochemical Stains Used to Detect Fungi Periodic acid Schiff (PAS) Highlights Fungal Cell Walls Background glycogen may require diastase digestion Histochemical Stains Used to Detect Fungi Fontana-Masson Stains Melanin and Melanin-like pigments Stains Dematiaceous Fungi Stains C. neoformans Mucin Stains - Mucicarmine/Alcian blue Stains the capsule of C. neoformans Tissue Gram Stain Fungi may stain but this is not the optimal stain. Special Techniques Filamentous Fungi vs. Yeast and Yeast-Like Forms In situ Hybridization PCR 2

3 Aspergillus niger Yeast and Yeast-Like Forms in Human Tissues These may be: Yeast Pneumocystis jiroveci Dimorphic Fungi Something else. Differentiate Morphologically Based on size, morphology, special features Small Yeast Forms (2-4 microns) Histoplasma, Sporothrix, C. glabrata, P. marneffei, Malassezia, Pneumocystis, endospores of C. immitis, or acapsular variants of C. neoformans. Culture Definitively Differentiates 3

4 Histoplasma capsulatum Inflammatory Response: Active to Remote Granulomas; Macrophages in profoundly immunosuppressed. Small budding yeast No pseudohyphae,except Endocarditis Overwhelming infection Histoplasma Endocarditis 4

5 5/29/2009 Histoplasmoma Geographic Distribution of H. capsulatum in the USA Granuloma formation with caseation Acquisition of Histoplasma capsulatum in nature Yeast form of H. capsulatum in bone marrow 5

6 Histoplasma capsulatum Slow-growing fungus - Culture should not demonstrate growth for at least a week. Histoplasma capsulatum Cultural Features Yeast form bacterial-like at C Sporothrix schenckii 6

7 Sporothrix schenckii 7

8 Name the Fungus Candida glabrata Colony Morphology Candida glabrata Microscopic Morphology Pneumocystis jiroveci 8

9 Pneumocystis jiroveci 9

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11 Moderate and Large Yeast Forms Candida species other than C. glabrata Moderate (4-10 microns) Candida species other than C. glabrata. Cryptococcus Immature spherules of C. immitis Large (>10 microns) Blastomyces dermatitidis Spherule of C. immitis Paracoccidioides brasiliensis 11

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13 Cryptococcus neoformans Pyogranulomatous Response with Blastomyces dermatitidis 13

14 Geographic Distribution of B. dermatitidis in the USA KOH-Calcoflour White Preparation Blastomyces dermatitidis in sputum 14

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17 5/29/2009 Geographic Distribution of Coccidioides in the USA Coccidioidomycosis Direct Microscopic Examination of Clinical Specimens Coccicdioidomycosis Microbiology Mold form- culture cottony and rapidly growing- white Most hazardous of all fungi to work with 17

18 5/29/2009 Oropharyngeal lesion Paracoccidioidomycosis Mold and Yeast Cultures of P. brasiliensis 18

19 Dematiaceous Fungi Heterogeneous Group of Fungi Types of Disease Mycetoma Chromoblastomycosis Produces sclerotic bodies. Phaeohyphomycosis The Phaeohyphomycotic cyst produces pseudohyphae and yeast like cells. Chromoblastomycosis Chronic fungal disease that involves the skin and subcutaneous tissues. Causes pyogranulomatous inflammatory response, pseudoepitheliomatous hyperplasia, and the Splendore-Hoeppli phenomenon may be observed. Fungal elements (sclerotic bodies) appear as copper pennies Common causes: Cladosporium (Cladophialophora), Phialophora, Fonsecaea. 19

20 Summary Yeasts and Yeast-like forms in human tissue may be definitively identified in many instances using morphologic features, special stains, and attention to the inflammatory response. Microbiologist are your friends Culture correlation can help avoid embarrassing errors in identification 20