Detection of measles virus genomic sequences in SSPE brain tissue by the polymerase chain reaction.
The polymerase chain reaction (PCR) was modified to detect RNA genomic sequences by generating cDNA copies of these sequences as a preliminary step. Oligonucleotide primer pairs complementary to sequences in each of the five major structural protein genes of the measles virus (nucleocapsid protein, phosphoprotein, matrix protein, fusion protein, and hemagglutinin protein) were synthesized. PCR products were tentatively identified by visualization of bands of the appropriate size by ethidium bromide staining after gel electrophoresis, and identity was confirmed by subsequent restriction enzyme cleavage of the products at predetermined sites to yield fragments of predicted size. This method successfully amplified 400-500 base regions from each of these five genes in RNA extracts of wild measles virus cultured in Vero cells and in RNA extracted from most of the SSPE brain tissues tested, but not in RNA from any control brain tissues. Measles virus genome was detected in SSPE brain tissues stored frozen for as long as 27 years and formalin-fixed paraffin-embedded subacute sclerosing panencephalitis (SSPE) brain tissues as old as 9 years. This method provides a simple, rapid and highly sensitive means of detecting and identifying sequences of RNA genomes by PCR. The success of this method in detecting measles virus in SSPE brain tissue suggests that PCR is appropriate to investigate the possible presence of RNA viruses in other neurological disorders of unknown etiology.
Godec
M S
MS
Laboratory of Central Nervous System Studies, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892.
Asher
D M
DM
Swoveland
P T
PT
Eldadah
Z A
ZA
Feinstone
S M
SM
Goldfarb
L G
LG
Gibbs
C J
CJ
Gajdusek
D C
DC
D000293:Adolescent; D000328:Adult; D000818:Animals; D001483:Base Sequence; D002648:Child; D002675:Child, Preschool; D006224:Cricetinae; D004279:DNA, Viral / Q000235:genetics; D005260:Female; D006801:Humans; D008297:Male; D008457:Measles / Q000150:complications / Q000175:diagnosis* / Q000235:genetics; D008459:Measles virus / Q000302:isolation & purification; D008875:Middle Aged; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012367:RNA, Viral / Q000032:analysis*; D012680:Sensitivity and Specificity; D013344:Subacute Sclerosing Panencephalitis / Q000150:complications / Q000175:diagnosis* / Q000235:genetics
D004279:DNA, Viral; D012367:RNA, Viral