Workbook in Microbiology Laboratory Techniques 7e

Laboratory Diagnosis

The laboratory diagnosis of a fungal infection depends on the direct microscopic detection of fungal structures in clinical samples and/or the recovery in culture and subsequent identification of the fungus. Fungi may be isolated from a variety of clinical specimens representing the focus of infection (sputum, spinal fluid, tissue, pus aspirated from lymph nodes or other lesion, bone marrow aspirates, skin scrapings). All specimens of sufficient quantity submitted for fungal culture should be examined microscopically for fungi. When there is not sufficient specimen to allow both a culture and direct microscopic examination, the culture has priority over the smear because culture is more sensitive than microscopic examination. However, observing a fungus in a clinical specimen is often valuable in establishing the significance of the fungus (i.e., ruling out contamination) and in providing early information that may be crucial for determining appropriate patient therapy.

In general, serological tests (looking for a significant change in antibody titer in paired serum specimens, see Exercise 33) have limited application for the diagnosis of most fungal infections. Exceptions to this rule include certain dimorphic fungal diseases, such as histoplasmosis and coccidioidomycosis. The purpose of this laboratory exercise is to acquaint the student with some direct microscopic and cultural methods that are available for establishing the laboratory diagnosis of a human mycosis.

Direct Microscopic Examinations

Histopathology. The visualization of fungal structures (hyphae, conidia, etc.) in tissue obtained by biopsy or at autopsy establishes the involvement of the fungus in human disease. Specialized tissue stains such as Giemsa, methenamine silver (see colorplate 45), or mucicarmine may be used to facilitate the detection of the fungus in tissue. The particular fungal structures that are seen in tissue can sometimes confirm the identity of the fungus (e.g., spherules [the yeast form] of Coccidioides immitis [see fig. 31.1] or cysts of Pneumocystis carinii [see colorplate 46]) or suggest the presence of a particular fungal group. In this latter case, culture is used to confirm the presence and identity of the fungal pathogen.

Direct Smears. Direct smears of patient material other than tissue are often made to detect the presence of fungal elements microscopically. Several types of stains or reagents are used to facilitate the detection of certain fungi.

1. Ten percent potassium hydroxide: Potassium hydroxide preparations are used to examine a variety of clinical samples including hair, nails, skin scrapings, fluids, or exudates. The potassium hydroxide solution serves to clear away tissue cells

Figure 31.1 KOH preparation of lung biopsy material showing spherule (yeast form) of Coccidioides immitis. Many endospores bud off from the thick-walled spherule, which has burst, releasing the endospores into the surrounding tissue. Each endospore is able to form a new spherule. In culture, this dimorphic fungus will grow as the filamentous hyphal form.

Figure 31.1 KOH preparation of lung biopsy material showing spherule (yeast form) of Coccidioides immitis. Many endospores bud off from the thick-walled spherule, which has burst, releasing the endospores into the surrounding tissue. Each endospore is able to form a new spherule. In culture, this dimorphic fungus will grow as the filamentous hyphal form.

Koh Preparation Coccidioides Immitis

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Bacterial Vaginosis Facts

Bacterial Vaginosis Facts

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