Detection of lymphocytic choriomeningitis virus by use of fluorogenic nuclease reverse transcriptase-polymerase chain reaction analysis

Comp Med. 2003 Feb;53(1):65-9.

Abstract

Lymphocytic choriomeningitis virus (LCMV) induces persistent infections in laboratory mice; is a known contaminant of biological materials, such as transplantable tumor cell lines; and is of great concern in animal facilities due to its zoonotic potential. Fluorogenic nuclease reverse transcriptase-polymerase chain reaction (fnRT-PCR) assays combine RT-PCR with an internal fluorogenic hybridization probe, thereby potentially enhancing specificity and eliminating post-PCR processing. An fnRT-PCR assay specific for LCMV was, therefore, developed by targeting primer and probe sequences to a unique region of the LCMV nucleocapsid (NP) gene. The LCMV fnRT-PCR assay detected only LCMV and did not detect other RNA viruses that naturally infect rodents. The fnRT-PCR assay detected as little as one picogram of LCMV RNA, but was 100-fold less sensitive when directly compared with the mouse antibody production test. The fnRT-PCR assay was also able to detect viral RNA in numerous tissues and in feces and cage swipe specimens collected from experimentally inoculated BALB/c mice, but did not detect any viral RNA in similar samples collected from age- and strain-matched mock-infected mice. In conclusion, the LCMV fnRT-PCR assay offers a potentially high-throughput diagnostic assay to detect LCMV in mice and contaminated biological materials.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Base Sequence
  • Cricetinae
  • DNA Primers
  • Female
  • Fluorescent Dyes
  • Lymphocytic choriomeningitis virus / genetics
  • Lymphocytic choriomeningitis virus / isolation & purification*
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Mice, Inbred ICR
  • Pregnancy
  • RNA, Viral / analysis
  • Reverse Transcriptase Polymerase Chain Reaction / methods*
  • Sensitivity and Specificity

Substances

  • DNA Primers
  • Fluorescent Dyes
  • RNA, Viral