Detection and sequence confirmation of Sin Nombre virus RNA in paraffin-embedded human tissues using one-step RT-PCR.
Sin Nombre virus (SNV) is the causative agent of hantavirus pulmonary syndrome (HPS). SNV RNA can be detected in fresh or frozen autopsy tissue samples by reverse transcriptase polymerase chain reaction (RT-PCR), and virus antigens can be identified by immunohistochemistry. A method was developed for demonstration of SNV RNA in formalin-fixed, paraffin-embedded tissues by RT-PCR. Virus genomes were detected in 8 of 12 (66.7%) fixed human tissue samples that were positive by immunohistochemistry, and RT-PCR prior to tissue fixation. By comparison with nucleotide sequences determined previously in fresh or frozen tissues of the same patients, identical genomic sequences were obtained, proving the authenticity of the PCR products. The study demonstrates that detection of SNV RNA in formalin-fixed, paraffin-embedded archival tissue by RT-PCR is feasible. This method allows retrospective studies on the phylogenetic epidemiology of SNV in North America.
Schwarz
T F
TF
Special Pathogens Branch, Centers for Disease Control and Prevention, Atlanta, GA 30333, USA.
Zaki
S R
SR
Morzunov
S
S
Peters
C J
CJ
Nichol
S T
ST
D001483:Base Sequence; D006476:Orthohantavirus / Q000302:isolation & purification*; D018804:Hantavirus Pulmonary Syndrome / Q000821:virology; D006801:Humans; D007668:Kidney / Q000821:virology; D008168:Lung / Q000821:virology; D008969:Molecular Sequence Data; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000032:analysis*; D017422:Sequence Analysis, DNA; D016707:Tissue Fixation; D014158:Transcription, Genetic
D012367:RNA, Viral