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Improved diagnosis on a daily basis of enterovirus meningitis using a one-step real-time RT-PCR assay.
J Med Virol. 2004 Dec; 74(4):604-11.JM

Abstract

The detection of the enterovirus genome in cerebrospinal fluid (CSF) by PCR techniques has proved to be more sensitive than traditional cell culture for the diagnosis of enterovirus meningitis. However, PCR assays are time consuming and labor intensive, particularly if separate hybridization steps are used to confirm the specificity of positive findings. The aim of this study was to develop a one-step real-time RT-PCR assay with LightCycler (LC) technology that was sensitive, rapid, and easy to perform in routine practice. The enterovirus detection limit was determined by testing 10-fold limiting dilution series of cell culture stocks with the echovirus 25 (E-25) prototype strain and with the third European Union Quality Control Concerted Action (EU-QCCA) enterovirus proficiency panel. A total of 100 CSF specimens were investigated in a comparative study. With the E-25 strain, the detection limit of the real-time assay was 286 TCID50/ml (50% tissue culture infective dose). When samples of the EU-QCCA panel were tested, our assay gave identical results (detection limit down to 3.6 TCID50/ml) to those of the reference laboratory, which used one-step RT-PCR assay. When CSF specimens were tested, there was a correlation between the real-time assay and the conventional in-house assay in 96 of 100 CSFs tested. This one-step real-time assay allows rapid enterovirus detection in CSF since results are obtained in 3 hr as against 36 hr with the "in-house" RT-PCR assay. This new assay is now being used in routine practice, and allows diagnosis on a daily basis.

Authors+Show Affiliations

Laboratoire de Virologie Médicale du Centre Hospitalier Universitaire, Faculté de Médecine, Clermont-Ferrand Cedex 1, France. carchimbaud@chu-clermontferrand.frNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15484276

Citation

Archimbaud, Christine, et al. "Improved Diagnosis On a Daily Basis of Enterovirus Meningitis Using a One-step Real-time RT-PCR Assay." Journal of Medical Virology, vol. 74, no. 4, 2004, pp. 604-11.
Archimbaud C, Mirand A, Chambon M, et al. Improved diagnosis on a daily basis of enterovirus meningitis using a one-step real-time RT-PCR assay. J Med Virol. 2004;74(4):604-11.
Archimbaud, C., Mirand, A., Chambon, M., Regagnon, C., Bailly, J. L., Peigue-Lafeuille, H., & Henquell, C. (2004). Improved diagnosis on a daily basis of enterovirus meningitis using a one-step real-time RT-PCR assay. Journal of Medical Virology, 74(4), 604-11.
Archimbaud C, et al. Improved Diagnosis On a Daily Basis of Enterovirus Meningitis Using a One-step Real-time RT-PCR Assay. J Med Virol. 2004;74(4):604-11. PubMed PMID: 15484276.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Improved diagnosis on a daily basis of enterovirus meningitis using a one-step real-time RT-PCR assay. AU - Archimbaud,Christine, AU - Mirand,Audrey, AU - Chambon,Martine, AU - Regagnon,Christel, AU - Bailly,Jean-Luc, AU - Peigue-Lafeuille,Hélène, AU - Henquell,Cécile, PY - 2004/10/16/pubmed PY - 2005/1/11/medline PY - 2004/10/16/entrez SP - 604 EP - 11 JF - Journal of medical virology JO - J Med Virol VL - 74 IS - 4 N2 - The detection of the enterovirus genome in cerebrospinal fluid (CSF) by PCR techniques has proved to be more sensitive than traditional cell culture for the diagnosis of enterovirus meningitis. However, PCR assays are time consuming and labor intensive, particularly if separate hybridization steps are used to confirm the specificity of positive findings. The aim of this study was to develop a one-step real-time RT-PCR assay with LightCycler (LC) technology that was sensitive, rapid, and easy to perform in routine practice. The enterovirus detection limit was determined by testing 10-fold limiting dilution series of cell culture stocks with the echovirus 25 (E-25) prototype strain and with the third European Union Quality Control Concerted Action (EU-QCCA) enterovirus proficiency panel. A total of 100 CSF specimens were investigated in a comparative study. With the E-25 strain, the detection limit of the real-time assay was 286 TCID50/ml (50% tissue culture infective dose). When samples of the EU-QCCA panel were tested, our assay gave identical results (detection limit down to 3.6 TCID50/ml) to those of the reference laboratory, which used one-step RT-PCR assay. When CSF specimens were tested, there was a correlation between the real-time assay and the conventional in-house assay in 96 of 100 CSFs tested. This one-step real-time assay allows rapid enterovirus detection in CSF since results are obtained in 3 hr as against 36 hr with the "in-house" RT-PCR assay. This new assay is now being used in routine practice, and allows diagnosis on a daily basis. SN - 0146-6615 UR - https://www.unboundmedicine.com/medline/citation/15484276/Improved_diagnosis_on_a_daily_basis_of_enterovirus_meningitis_using_a_one_step_real_time_RT_PCR_assay_ L2 - https://doi.org/10.1002/jmv.20217 DB - PRIME DP - Unbound Medicine ER -