Identification of Fusarium Species in Formalin-Fixed and Paraffin-Embedded Sections by In Situ Hybridization Using Peptide Nucleic Acid Probes

• Published in: Journal of Clinical Microbiology Vol. 49; no. 3; pp. 808 - 813
• Main Authors: Shinozaki, Minoru, Okubo, Yoichiro, Sasai, Daisuke, Nakayama, Haruo, Murayama, Somay Yamagata, Ide, Tadashi, Wakayama, Megumi, Hiruta, Nobuyuki, Shibuya, Kazutoshi
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.03.2011
• Subjects: Animals; Aspergillus; Base Sequence; Biological and medical sciences; DNA, Fungal - genetics; DNA, Ribosomal - genetics; Formaldehyde; Fundamental and applied biological sciences. Psychology; Fusarium - isolation & purification; Fusarium oxysporum; Fusarium solani; In Situ Hybridization - methods; Male; Mice; Mice, Inbred ICR; Microbiology; Miscellaneous; Molecular Sequence Data; Mycology; Mycoses - diagnosis; Oligonucleotide Probes; Paraffin Embedding; Pathology, Molecular - methods; Peptide Nucleic Acids; Pseudallescheria boydii; RNA, Ribosomal, 28S - genetics; Scedosporium; Sensitivity and Specificity; Sequence Alignment; Fungi; Molecular hybridization; Peptides; In situ; Fusarium; Identification; Fungi Imperfecti; Nucleic acid
• ISSN: 0095-1137; 1098-660X
• DOI: 10.1128/JCM.01149-10

Fusarium has recently emerged as an opportunistic pathogen of humans, but the histological differentiation of Fusarium from Aspergillus and Scedosporium is particularly difficult because these fungi may induce similar clinical features and exhibit filamentous development in host tissues. Thus, there is a need to establish rapid and reliable methods that are applicable to pathological diagnoses. The aim of this study was to evaluate and establish in situ hybridization (ISH) using peptide nucleic acid (PNA) probes targeting the 28S rRNA to identify Fusarium species in tissue sections. This technique was validated using both formalin-fixed and paraffin-embedded pulmonary tissues from mice infected with seven different species of fungi and cell blocks from fungal cultures of 30 strains. As a result, strong positive signals were observed within fungal organisms present in tissues of the lung from mice infected with Fusarium solani. Furthermore, this probe reacted strongly with both F. solani and Fusarium oxysporum in sections from cell blocks. Although some cross-reactivity occurred with the Pseudallescheria boydii in sections from cell blocks, the signal intensity was low and most hyphae were not reactive. In conclusion, it was confirmed that ISH with PNA probes is accurate and is a valuable tool for identifying Fusarium spp. among organisms that have identical morphological features in formalin-fixed and paraffin-embedded sections.