In Situ Detection of Aspergillus 18S Ribosomal RNA in Invasive Pulmonary Apergillosis

Makoto KOBAYASHI, Hiroshi SONOBE, Takayuki IKEZOE, Eiji HAKODA, Yuji OHTSUKI, Hirokuni TAGUCHI
1999 Internal medicine (Tokyo. 1992)  
Objects Weattempted to evaluate the usefulness of in situ hybridization (ISH) in the specific diagnosis ofAspergillus pulmonary infection. Methods Weused an ISH technique using a multiple digoxigenin-incorporating probe, which was constructed by means of the polymerase chain reaction (PCR) from the 18S ribosomal RNAofAspergillusfumigatus. Materials Westudied twelve formalin-fixed, paraffin-embedded lung tissue sections from autopsy-confirmed invasive pulmonary aspergillosis (IPA) (5 acute
more » ... ytic leukemias, 2 acute lymphocytic leukemias, 2 chronic myelocytic leukemias, 1 adult T-cell leukemia, 1 non-Hodgkin's lymphoma and 1 chronic obstructive pulmonary disease.), and 18 sections from other pulmonary infections as control. Results ISH using the probe and a low-viscosity hybridization buffer solution (LV) positively stained hyphal elements in 12 of 12 autopsy lung tissue specimens from subjects with IPA, while ISH using the probe and a high viscosity hybridization buffer solution (HV) positively stained the hyphal elements in 6 of 12. Specifically, ISH (LV) demonstrates hyphal elements of Aspergillus spp. in the center ofAspergillus abscess. While, ISH (HV) can detect hyphal elements located in the periphery of a suppurative abscess as well as those in the blood vessel. Conversely, ISH did not showpositive results for any of the autopsy tissue specimens from subjects with other fungal pneumonia infections (Candida n=5, Mucorn=2, Cryptococcus n=2, and Pseudallescheria n=l), Pneumocystis carinii pneumonia (n=5), and cytomegalovirus pneumonia (n=3). Dual staining by means of ISH and immunohistochemistry (IHC) using anti-neutrophil elastase (NE) and anti-CD68monoclonal antibodies showed that NEpositive cells were localized at the edge of the radial growth of the organism, but CD68positive cells were located around the center of the abscess. The accumulation of NEpositive cells was rarely seen in half of the cases (6/12). In contrast, CD68 positive cells were routinely present in the center of the abscess (12/12). Conclusion ISH in conjunction with IHC is a useful tool for differentiating Aspergillus spp. from other fungal genera in tissue sections from patients with IPA and mayhave a certain role in the evaluation of the interactions between organisms and recruiting inflammatory cells.
doi:10.2169/internalmedicine.38.563 fatcat:xqoo2ougwzgn5mu3ct5v24qxcy