In addition, pathologists should assess the presence or absence of internal structures that can be observed in fungi (nuclei and cytoplasm) which stain with H&E but not with GMS. The predominant Candida species that does not produce filaments is C. glabrata. Methods.Histopathologic examination of tissues to detect fungi is and will remain an important tool to define the diagnostic significance of positive culture isolates, including fungal invasion of tissues and vessels as well as the host reaction to the fungus. The majority of patients present with lung disease, which can disseminate hematogenously to bone marrow, adrenal glands, brain, and other tissues, or the organisms are expectorated and lesions can form in the oropharyngeal tract. For these assays it is necessary to cut a very thin section of the tissue (similar to that used for histopathology) and place it on a glass slide. Bauer (1933) is credited with the earliest report that the oxidation of sugars (1,2-glycols) with chromic acid creates aldehydes that will bind the Schiff molecule.4 Many others, including Hotchkiss, McManus (1960), Spicer, Pearse, and Lillie continued to study the aldehyde-Schiff reaction using various oxidizers. Humans acquire these organisms during contact with infected plants or soil through cuts or punctures in the skin. rRNA probes have shown strong signals when hybridized to their homologous target. As new non-cross-reactive antibodies are developed and tested using this technique, immunohistochemistry may provide an inexpensive and rapid alternative to more costly assays that do not combine morphology with detection of the specific fungus. In addition, techniques such as laser microdissection will be useful to detect the now more frequently recognized dual fungal infections and the local environment in which this phenomenon occurs. In this review we present epidemiologic, clinical, and morphological findings and interpretation pitfalls regarding the most frequently encountered yeasts and molds, as well as alternative testing that can be performed with other samples. While chromic acid continues further oxidation of all polysaccharides, complete conversion in the areas of heaviest concentration takes longer, allowing these to remain reactive with Schiff (or silver).2 Since the walls of fungi are particularly rich in 1,2-glycol groups, they stain more intensely than most other reactive components in the tissue section. As the use of less invasive procedures has become more prevalent in medicine, cytologic specimens have become common samples. (i) Epidemiologic and clinical situations when cryptococcosis should be considered in the differential diagnosis.Human cryptococcosis is caused by several Cryptococcus species, including C. neoformans and C. gattii. Morphology, description, diagnosis, and comment for fungal infections that show characteristic yeast morphology in tissues. When stained with GMS, the entire wall thickness stains and shows fenestrations. However, because Histoplasma and Paracoccidioides grow slowly in vitro, fungal cultures are usually incubated for at least 4 weeks in most laboratories. Nonetheless, in patients with nondisseminated histoplasmosis the antigen burden is lower, and thus sensitivity is lower. The peptide nucleic acid fluorescent in situ hybridization assay can be used to identify the most common species of Candida in smears made from positive blood culture bottles. Combining the results of detection of antigen in urine and serum may increase the sensitivity in patients with pulmonary histoplasmosis (157). Skin tests using paracoccidioidin also have been used for epidemiologic studies (75). (iii) Pitfalls in morphological diagnosis.The major morphological differentiation between Mucorales genera and other molds is with other fungi that produce nonpigmented hyphae in tissue, including Aspergillus spp., other hyaline septated molds (such as Fusarium and Scedosporium), and Candida spp. Gram +ve. The most important problem with galactomannan testing is that false-positive results occur in approximately 50% of patients taking antibiotics (piperacillin, amoxicillin, or ticarcillin), 100% of patients receiving substances that contain products of A. niger fermentation (plasmalyte), and various percentages of patients with infections with other fungi, including Penicillium, Paecilomyces, Alternaria, and Histoplasma. Until the advent and widespread use of highly active antiretroviral treatment (HAART), Pneumocystis pneumonia was frequently diagnosed in patients with AIDS and was associated with high mortality. in clinical microbiology at Thomas Jefferson University, and a Ph.D. in microbiology and immunology at Temple University. P. brasiliensis in tissue sections stained with H&E are spherical yeasts that vary in size from 4 to 60 μm and have an optically clear space between the fungus and surrounding tissue (92). (29, 145). Rhinosporidium seeberi, a mesomycetozoan parasite that causes palate and nasopharyngeal polyps, produces large sporangia (some can be seen with the naked eye) with multiple internal endospores. In addition, Hamazaki-Wesenberg bodies (pigmented elliptical structures seen in sarcoidosis) have been confused with S. schenckii (34). Helminths will show musculature and internal organs that are not present in adiaspiromycosis. Skin lesions that result from disseminated disease are painful, pruritic nodules that evolve to have central necrosis, giving a characteristic “bull's eye” appearance, and eventually ulcerate to produce ecthyma gangrenosum-like lesions. (i) Epidemiologic and clinical situations when yeasts or yeast-like organisms should be considered in the differential diagnosis. To aid in the recognition of these different tissue structures and fungi, it is important to colocalize the GMS-stained structures in tissue sections (usually from a consecutive level) stained with H&E or PAS stain. Those with pulmonary mucormycosis have fever, multiple pulmonary nodules, and pleural effusions. Diagnostic criteria for the different stages have been established for the disease (2, 59, 119). (looking for remnants of a ruptured spherule or an intact spherule is paramount for diagnosis of coccidioidomycosis), Pneumocystis jirovecii (this organism lacks budding and has an intracystic focus), Penicillium marneffei (this organism shows formation of a transverse septum rather than budding), and Candida glabrata (this organism may show more size variability than in histoplasmosis, and the inflammation is primarily neutrophilic). Lastly, immunohistochemistry, in situ hybridization, and PCR with FFPE samples have already been validated for research purposes in some centers, enabling detection of specific fungi when the entire specimen is placed in formalin. The usual host reaction, whether in superficial or invasive candidiasis, consists primarily of neutrophilic inflammation with some lymphocytes and macrophages, fibrin, and coagulative necrosis (94). were recovered in culture from discordant cases (86). Hyphae and aleurioconidia can be visualized and are particularly prominent in hair follicles. (i) Epidemiologic and clinical situations when hyaline septated molds should be considered in the differential diagnosis. Furthermore, advances in diagnostic radiology and in patient support (such as platelet transfusions, etc) have allowed greater ability to pursue specific diagnoses by collecting tissue biopsy specimens from body sites formerly not available for histopathologic examination.