Giuseppe
Emeritus
J Clin Microbiol. 2009 Jun;47(6):1742-9. Epub 2009 Apr 1.
New molecular detection tools adapted to emerging rhinoviruses and enteroviruses.
Tapparel C, Cordey S, Van Belle S, Turin L, Lee WM, Regamey N, Meylan P, M?hlemann K, Gobbini F, Kaiser L. Laboratory of Virology, Division of Infectious Diseases, University of Geneva Hospitals, 24 Rue Micheli-du-Crest, 1211 Geneva 14, Switzerland. caroline.tapparel@hcuge.ch
Human rhinoviruses (HRV), and to a lesser extent human enteroviruses (HEV), are important respiratory pathogens. Like other RNA viruses, these picornaviruses have an intrinsic propensity to variability. This results in a large number of different serotypes as well as the incessant discovery of new genotypes. This large and growing diversity not only complicates the design of real-time PCR assays but also renders immunofluorescence unfeasible for broad HRV and HEV detection or quantification in cells. In this study, we used the 5' untranslated region, the most conserved part of the genome, as a target for the development of both a real-time PCR assay (Panenterhino/Ge/08) and a peptide nucleic acid-based hybridization oligoprobe (Panenterhino/Ge/08 PNA probe) designed to detect all HRV and HEV species members according to publicly available sequences. The reverse transcription-PCR assay has been validated, using not only plasmid and viral stocks but also quantified RNA transcripts and around 1,000 clinical specimens. These new generic detection PCR assays overcame the variability of circulating strains and lowered the risk of missing emerging and divergent HRV and HEV. An additional real-time PCR assay (Entero/Ge/08) was also designed specifically to provide sensitive and targeted detection of HEV in cerebrospinal fluid. In addition to the generic probe, we developed specific probes for the detection of HRV-A and HRV-B in cells. This investigation provides a comprehensive toolbox for accurate molecular identification of the different HEV and HRV circulating in humans.
PMID: 19339471 [PubMed - indexed for MEDLINE]
PMCID: PMC2691104 [Available on 2009/12/01]
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New molecular detection tools adapted to emerging rhinoviruses and enteroviruses.
Tapparel C, Cordey S, Van Belle S, Turin L, Lee WM, Regamey N, Meylan P, M?hlemann K, Gobbini F, Kaiser L. Laboratory of Virology, Division of Infectious Diseases, University of Geneva Hospitals, 24 Rue Micheli-du-Crest, 1211 Geneva 14, Switzerland. caroline.tapparel@hcuge.ch
Human rhinoviruses (HRV), and to a lesser extent human enteroviruses (HEV), are important respiratory pathogens. Like other RNA viruses, these picornaviruses have an intrinsic propensity to variability. This results in a large number of different serotypes as well as the incessant discovery of new genotypes. This large and growing diversity not only complicates the design of real-time PCR assays but also renders immunofluorescence unfeasible for broad HRV and HEV detection or quantification in cells. In this study, we used the 5' untranslated region, the most conserved part of the genome, as a target for the development of both a real-time PCR assay (Panenterhino/Ge/08) and a peptide nucleic acid-based hybridization oligoprobe (Panenterhino/Ge/08 PNA probe) designed to detect all HRV and HEV species members according to publicly available sequences. The reverse transcription-PCR assay has been validated, using not only plasmid and viral stocks but also quantified RNA transcripts and around 1,000 clinical specimens. These new generic detection PCR assays overcame the variability of circulating strains and lowered the risk of missing emerging and divergent HRV and HEV. An additional real-time PCR assay (Entero/Ge/08) was also designed specifically to provide sensitive and targeted detection of HEV in cerebrospinal fluid. In addition to the generic probe, we developed specific probes for the detection of HRV-A and HRV-B in cells. This investigation provides a comprehensive toolbox for accurate molecular identification of the different HEV and HRV circulating in humans.
PMID: 19339471 [PubMed - indexed for MEDLINE]
PMCID: PMC2691104 [Available on 2009/12/01]
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