Specific Detection Method of 18S rRNA for Conidiobolomycosis-, Mucormycosis- and Aspergillosis-causing Fungi by <i>in situ</i> Hybridization Method Using MicroProbe System on Paraffin-embedded Tissue Sections

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  • パラフィン包埋組織切片上でのMicroProbe Systemを用いた<i>in situ</i> hybridization法によるコニディオボルス症起因菌，ムーコル症起因菌およびアスペルギルス症起因菌の18S rRNAの特異的検出法
  • パラフィン包埋組織切片上でのMicroProbe Systemを用いたin situ hybridization法によるコニディオボルス症起因菌,ムーコル症起因菌およびアスペルギルス症起因菌の18S rRNAの特異的検出法
  • パラフィン ホウマイソシキ セッペン ジョウ デ ノ MicroProbe System オ モチイタ in situ hybridizationホウ ニ ヨル コニディオボルスショウ キインキン,ムーコルショウ キインキン オヨビ アスペルギルスショウ キインキン ノ 18S rRNA ノ トクイテキ ケンシュツホウ

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Abstract

Invasive conidiobolomycosis has been drawing attention as an emerging mycosis with a high mortality rate. To establish a method for distinguishing conidiobolomycosis caused by Conidiobolus sp. from mucormycosis due to Rhizopus microsporus and aspergillosis caused by Aspergillus fumigatus in Formalin-Fixed Paraffin-Embedded (FFPE) tissue sections, we focused on an in situ hybridization (ISH) method targeting 18S rRNA using FFPE tissue sections from each case. Three antisense probes (probes A, B, and C) that differed in base number and sequence position were prepared separately for Conidiobolus sp., R. microsporus, and A. fumigatus using consensus primers by focusing on the hypervariable region of the V4 domain, which is particularly rich in nucleotide polymorphism among 18S rDNA. Results of ISH using the three probes for each fungus showed that by optimizing the conditions of stringency it was possible to sensitively distinguish conidiobolomycosis-causing Conidiobolus sp. from mucormyosis-causing R. microsporus, and aspergillosis-causing A. fumigatus by using probe C (Conidiobolus sp. 189 b, R. microsporus 155 b, A. fumigatus 154 b), which have mutual homology of 60% or less. This method makes it possible to quickly and reliably detect and identify these causative fungi on FFPE tissue sections, and therefore could significantly contribute to fungal disease treatment strategies.

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