New molecular detection tools adapted to emerging rhinoviruses and enteroviruses.
Details
Serval ID
serval:BIB_74717C10E50A
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
New molecular detection tools adapted to emerging rhinoviruses and enteroviruses.
Journal
Journal of clinical microbiology
ISSN
1098-660X (Electronic)
ISSN-L
0095-1137
Publication state
Published
Issued date
06/2009
Peer-reviewed
Oui
Volume
47
Number
6
Pages
1742-1749
Language
english
Notes
Publication types: Evaluation Study ; Journal Article ; Research Support, Non-U.S. Gov't
Publication Status: ppublish
Publication Status: ppublish
Abstract
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.
Keywords
5' Untranslated Regions, Base Sequence, Body Fluids/virology, Conserved Sequence, DNA Primers/genetics, Enterovirus/genetics, Enterovirus/isolation & purification, Enterovirus Infections/diagnosis, Enterovirus Infections/virology, Humans, Molecular Sequence Data, Nucleic Acid Hybridization/methods, Peptide Nucleic Acids/genetics, Picornaviridae Infections/diagnosis, Picornaviridae Infections/virology, Polymerase Chain Reaction/methods, RNA, Viral/genetics, Rhinovirus/genetics, Rhinovirus/isolation & purification, Sensitivity and Specificity, Sequence Alignment
Pubmed
Web of science
Open Access
Yes
Create date
09/02/2010 16:43
Last modification date
14/05/2024 9:02