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Molecular typing of Pneumocystis jirovecii found in formalin-fixed paraffin-embedded lung tissue sections from sudden infant death victims

¹ Ecology of Parasitism, EA-3609-IFR17, Pasteur Institute of Lille, 1 rue du Professeur-Calmette BP245, 59019 Lille, France
² Respiratory Infections Laboratory, Biomedical Sciences Institute, University of Chile School of Medicine, Independencia 1027, Santiago, Chile
³ Free Faculty of Medicine, Lille Catholic University, rue du Port, 59046 Lille, France
⁴ Parasitology-Mycology Service, Faculty of Medicine, Lille-2 University Hospital Center, 1 place Verdun, 59045 Lille, France

Correspondence
I. Durand-Joly
isabelle.joly{at}pasteur-lille.fr

Previous studies have provided histological evidence of an association between primary Pneumocystis infection and sudden infant death syndrome (SIDS). The aim of this work was to determine the species of clustered Pneumocystis organisms found in formalin-fixed paraffin-embedded (FFPE) lung tissue sections from Chilean sudden infant death (SID) victims. This approach needed first to optimize a DNA extraction method from such histological sections. For that purpose, the QIAamp DNA Isolation from Paraffin-Embedded Tissue method (Qiagen) was first tested on FFPE lung tissue sections of immunosuppressed Wistar rats inoculated with rat-derived Pneumocystis. Successful DNA extraction was assessed by the amplification of a 346 bp fragment of the mitochondrial large subunit rRNA gene of the Pneumocystis species using a previously described PCR assay. PCR products were analysed by direct sequencing and sequences corresponding to Pneumocystis carinii were found in all the samples. This method was then applied to FFPE lung tissue sections from Chilean SID victims. Pneumocystis jirovecii was successfully identified in the three tested samples. In conclusion, an efficient protocol for isolating PCR-ready DNA from FFPE lung tissue sections was developed. It established that the Pneumocystis species found in the lungs of Chilean SID victims was P. jirovecii.

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