Direct extraction of Sabin poliovirus genomes from human fecal samples using a guanidine thiocyanate extraction method

Matthew O. Old, Claudia V. Martinez, Douglas Kwock, Joaquin Garcia, Gladys Martin, Christina Chan, Yvonne A. Maldonado

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

To permit rapid and efficient detection of Sabin poliovirus type 3 from human fecal samples, we developed a guanidine thiocyanate (GuSCN) extraction and reverse transcriptase polymerase chain reaction (RT-PCR) method. Using 10-fold serial dilutions from stock Sabin-Leon 12 a1b poliovirus type 3 at 107 TCID50 per 0.1 ml, genome was detected to a dilution of 103 TCID50 per 0.1 ml. A total of 40 archived fecal samples were examined using this GuSCN extraction method followed by RT-PCR. Fourteen of 20 poliovirus type 3 tissue culture-positive specimens (70%) and two of 20 tissue culture-negative specimens (10%) were detected by GuSCN extraction and RT-PCR. All positive and negative extraction and RT-PCR controls were identified accurately. This GuSCN extraction and RT-PCR technique is rapid, inexpensive, and can be readily adapted to identify genome sequences of other enterovirus types in large numbers of fecal samples. Moreover, the GuSCN technique extracts viral RNA directly from fecal samples, allowing observation of in vivo alterations of genome sequences. Further studies are underway to examine the development of revertant point mutations in the Sabin poliovirus type 3 genome following oral administration of trivalent Sabin Oral Poliovirus Vaccine to humans.

Original languageEnglish (US)
Pages (from-to)193-200
Number of pages8
JournalJournal of Virological Methods
Volume110
Issue number2
DOIs
StatePublished - Jun 30 2003

Keywords

  • Enteroviruses
  • Fecal specimens
  • Guanidine thiocyanate extraction
  • Rapid diagnosis
  • Sabin oral poliovirus vaccine type 3
  • Tissue cultivation

ASJC Scopus subject areas

  • Virology

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