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Fast duplex one-step RT-PCR for rapid differential diagnosis of entero- or toscana virus meningitis
Marcello Valassina1, Melissa Valentini, Pier Egisto Valensin
1Department of Molecular Biology, Microbiology Section, University of Siena, Italy. valassina@unisi.it
Abstract:
Acute meningitis is the most common neurologic disease that involves the central nervous system. The spectrum of infectious agents that cause neurologic infection is remarkably broad and numerous viruses are the most frequent cause of the aseptic meningitis syndrome. We applied a multiplex one-step method for the rapid detection of the genomic RNA of different neurotropic viruses: particles in the genus Enterovirus and Toscana virus, which are the most representative aetiologic agents in our country during the spring-summer period. We have evaluated the sensitivity and the specificity of the multiplex one-step test on positive controls and on RNA extracted from clinical samples harvested from 475 patients with meningitis hospitalized during the 1996-2001 period. The multiplex one-step RT-nPCR protocol allows for the detection of enterovirus and Toscana virus RNA in a single sample, by using, at the same time, a very small clinical sample volume. In our study we were able to diagnose 192 cases of meningitis by Toscana virus and 31 cases by enteroviruses out of 475 cases of meningitis utilizing the described one-step multiplex method.
Insights
A new multiplex one-step test rapidly detects genomic RNA from Enterovirus and Toscana virus, common causes of viral meningitis. This method enhances diagnosis efficiency for central nervous system infections.
Area of Science:
- Virology
- Neuroscience
- Molecular Biology
Background:
- Acute meningitis is a common central nervous system disease.
- Viral infections are the most frequent cause of aseptic meningitis syndrome.
- Enteroviruses and Toscana virus are significant neurotropic agents, especially during spring-summer.
Purpose of the Study:
- To develop and evaluate a rapid, multiplex one-step method for detecting genomic RNA of Enterovirus and Toscana virus.
- To assess the sensitivity and specificity of this diagnostic technique.
Main Methods:
- Application of a multiplex one-step reverse transcription nested polymerase chain reaction (RT-nPCR) assay.
- Testing on positive controls and RNA from 475 meningitis patients (1996-2001).
- Utilizing a very small clinical sample volume for simultaneous detection.
Main Results:
- The multiplex one-step RT-nPCR successfully detected both Enterovirus and Toscana virus RNA in single samples.
- Out of 475 meningitis cases, 192 were diagnosed with Toscana virus and 31 with Enteroviruses.
- The method demonstrated efficiency in diagnosing viral meningitis.
Conclusions:
- The developed multiplex one-step RT-nPCR is a sensitive and specific tool for rapid diagnosis of Enterovirus and Toscana virus meningitis.
- This method allows for simultaneous detection using minimal clinical sample volume, improving diagnostic workflow.
- It aids in identifying key etiological agents of viral meningitis, particularly during specific seasons.
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