Title 
Histopathological investigation of the prevalence of trichosporonosis among autopsy subjects using  peptide nucleic acid probe in situ hybridization technique
Objectives
Whereas culture examination has been generally accepted as the gold standard for identifying causative fungi from clinical specimens for invasive fungal infection, low sensitivity and contamination may become problems. Molecular supplemental procedures for histopathological diagnosis of invasive fungal infection are being introduced. 
Research has been very active for molecular diagnosis techniques such as polymerase chain reaction (PCR) or in situ hybridization (ISH) for diagnosis of invasive fungal infection. Although many novel primers and/or probes for fungi have been introduced, studies of applications with clinical specimens are still limited and clinical use of ISH has currently not been adequately verified.
Trichosporon species, which  are mostly insusceptible to echinocandins, are some of the most common dimorphic yeasts in Asia including Japan. We sometimes encounter breakthrough trichosporonosis in individuals being treated with echinocandins. Thus, it is therefore important to distinguish between Trichosporon spp. and Candida spp. through histopathological examination. However, the morphological similarity of these fungi makes it difficult to distinguish between these fungi only by routine histological examination. For choosing optimal antifungal drugs, favorable molecular supplemental procedure is required to identify these fungi for histopathological diagnosis. In our previous studies, we developed peptide nucleic probes (PNA) designed to be specific for C. albicans and Trichosporon spp. 
The purpose of this study is to evaluate the utility of an ISH procedure focusing on the histopathological prevalence of trichosporonosis in autopsy subjects by using PNA probe ISH.
Methods
Formalin-fixed and paraffin embedded (FFPE) tissue involved by invasive yeast infections from autopsy cases from the Toho University Medical Center Omori Hospital or the Tokyo Metropolitan Cancer and Infectious Diseases Center Komagome Hospital during 1975–2012 were used in this study. Sections of a total of 88 specimens mounted on glass slides were deparaffinized and stained with both hematoxylin and eosin and periodic acid-Schiff reaction (PAS). Three expert pathologists took part in histopathological examination to verify the previous diagnosis of invasive yeast infection and exclude cryptococcosis and mold infections from our cohort. ISH was performed with PNA probes for C. albicans and Trichosporon spp with confirmed specificity. Figure 1 summarizes the outline of the study.
Results
Thirty-six of the 88 cases showed positive signals for either the Trichosporon spp. or C. albicans probes, or both probes. Among them, 7  cases showed a positive signal for the Trichosporon spp. probe and 29 cases for the C. albicans probe. Five of the 7 Trichosporon spp. probe positive cases had been diagnosed as candiasis in the autopsy reports.
Conclusions
The present study identified 7 cases of invasive trichosporonosis, 5 of which had been misdiagnosed as candidiasis. The misdiagnoses indicate that incidence  of trichosporonosis may be higher than previously reported. Our results show that ISH is an useful tool to identify Trichosporon spp. in FFPE tissue samples.

Histopathological study on the prevalence of trichosporonosis among autopsy subjects using peptide nucleic acid probe In situ hybridization technique