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Application of in situ hybridization to tissue sections for identification of molds causing invasive fungal infection

Minoru Shinozaki1, Yoichiro Okubo, Haruo Nakayama

  • 1Department of Surgical Pathology, Toho University School of Medicine, 6-11-1 Omori-Nishi, Ota-Ku, Tokyo, Japan.

Nihon Ishinkin Gakkai Zasshi = Japanese Journal of Medical Mycology
|May 12, 2009
PubMed

Insights

In situ hybridization (ISH) effectively identifies fungal mold in patient samples. Optimized heat and proteinase K treatments enhance signal specificity for diagnosing invasive fungal infections.

Area of Science:

  • Mycology
  • Molecular Biology
  • Pathology

Background:

  • Invasive fungal infections pose diagnostic challenges.
  • Accurate identification of molds in tissue is crucial for patient management.
  • Current methods may lack specificity or require specialized techniques.

Purpose of the Study:

  • To evaluate the utility of in situ hybridization (ISH) for identifying molds in clinical specimens.
  • To optimize ISH procedures for formalin-fixed, paraffin-embedded tissues and cytological preparations.
  • To assess the efficacy of DNA and peptide nucleic acid (PNA) probes for specific mold detection.

Main Methods:

  • Standardized ISH protocols were developed using heat and proteinase K pretreatment at a higher pH.
  • DNA probes targeting the alkaline proteinase (ALP) gene and Afut-1 gene of Aspergillus fumigatus were designed.
  • PNA probes were developed for Candida albicans and Fusarium species.
  • The optimized ISH method was validated on experimentally infected mouse lungs and autopsy samples.

Main Results:

  • Optimized ISH demonstrated favorable signal intensity and specificity for mold detection.
  • DNA probes specifically identified Aspergillus species and Aspergillus fumigatus.
  • PNA probes showed satisfactory specificity for Candida albicans and Fusarium species.
  • The optimized method proved effective in formalin-fixed, paraffin-embedded tissues and cytological samples.

Conclusions:

  • In situ hybridization is a valuable tool for identifying medically important molds in tissue and cytological samples.
  • Optimized ISH protocols enhance diagnostic accuracy for invasive fungal infections.
  • Both DNA and PNA probes offer specific detection capabilities for various fungal pathogens.

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