Premium
Two different PCR assays to detect enteroviral RNA in CSF samples from patients with acute aseptic meningitis
Author(s) -
Casas Inmaculada,
Klapper Paul E.,
Cleator Graham M.,
Echevarria Juan E.,
Tenorio Antonio,
Echevarria Jose M.
Publication year - 1995
Publication title -
journal of medical virology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.782
H-Index - 121
eISSN - 1096-9071
pISSN - 0146-6615
DOI - 10.1002/jmv.1890470414
Subject(s) - aseptic meningitis , virology , meningitis , enterovirus , medicine , virus , polymerase chain reaction , viral meningitis , microbiology and biotechnology , biology , immunology , bacterial meningitis , gene , pediatrics , biochemistry
Two polymerase chain reaction (RT‐PCR) assays were developed to allow rapid detection of enteroviral RNA in cerebrospinal fluid samples (CSF). Primers homologous to the conserved 5′ noncoding region of the enterovirus genome were designed. The RT‐PCR product size was‐500 bp (479 bp for Poliovirus, 500 bp for Cox‐sackievirus) and was visualized using ethidium bromide‐stained gels. Assay 1 utilized Moloney Murine Leukaemia Virus Reverse Transcriptase (MMLV‐RTase) for reverse transcription and Taq polymerase for subsequent PCR. Assay 2 utilized a thermoactive DNA polymerase of Thermus thermophilus (rTth enzyme) for both reverse transcription and DNA amplification. In addition, in Assay 2 reverse transcription and PCR wer eaccomplished within the same reaction tube. Both assays detected between 1 and 0.02 TCID 50 of prototype strains of Polio and Coxsackie type B viruses propagated in VERO cell and spiked in a pooled preparation of CSF samples from patients with noninfective neurological disorders. However, Assay 1 was 10‐fold more sensitive than Assay 2 when applied to the detection of enteroviral RNA in CSF samples from patients with etiologically well characterized acute aseptic meningitis. © Wiley‐Liss, Inc.