@article{fdi:010061980,
  title = {{M}olecular detection of human rhinoviruses in respiratory samples : a comparison of {T}aqman probe-, {SYBR} green {I}- and {BOXTO}-based real-time {PCR} assays},
  author = {{D}upouey, {J}. and {N}inove, {L}. and {F}errier, {V}. and {P}y, {O}. and {G}azin, {C}. and {T}hirion {P}errier, {L}aurence and de {L}amballerie, {X}avier},
  editor = {},
  language = {{ENG}},
  abstract = {{B}ackground: {H}uman {R}hinoviruses ({HRV}) are major causative agents of acute respiratory tract infections in all age group and important contributing factors of childhood morbidity and mortality. {C}linical presentation is poorly specific and the great antigenic and genetic variability of {HRV}s renders the biological diagnosis complex. {H}ere, we have evaluated several molecular diagnostic protocols, including {T}aqman probe-based and intercalating agent-based {RT}-{PCR} assays. {M}ethods: 5,627 respiratory samples sent to the laboratory of {V}irology of the {U}niversity {H}ospitals of {M}arseille, {F}rance, from {M}arch 2011 to {F}ebruary 2012, were tested using a real-time {RT}-{PCR} assay in the 5'{NCR} of the rhinoviral genome that associated a {T}aqman probe and the detection of {DNA}-{BOXTO}-dye complexes. {A} sample of 500 {BOXTO}-positive samples were further tested using the same probe assay (without {BOXTO}), and a {SYBR} {G}reen assay (using the same amplification primers). {T}he specific amplification of {HRV} sequences was assessed by {NGS} amplicon sequencing. {R}esults: {T}he {T}aqman probe {RT}-{PCR} assay identified 696/5,627 samples (12,4%) as {HRV}-positive. {BOXTO}-positive samples included all probe-positive samples and 1,913 additional samples, of which only 24.3% were confirmed by sequencing. {T}he {SYBR} {G}reen assay was more specific (16/550 samples were probe-negative/{SYBR} {G}reen-positive, all confirmed by 5'{NCR} sequencing), but 3/500 samples were probe-positive/{SYBR} {G}reen-negative. {C}onclusions: {O}ur results highlight the difficulty in detecting {HRV}s in clinical samples using a single molecular detection system. {A}mongst the 3 systems tested, the best compromise was obtained with the {SYBR} {G}reen assay, which, by comparison with our probe-based assay provided an improved sensitivity without altering the detection specificity. {I}nterestingly, a majority of probe-negative/{BOXTO}- or {SYBR} {G}reen-positive samples were not associated with mutations in the sequence targeted by the probe. {S}equence-based modifications of the secondary structure of the {HRV} 5'{NCR} may be associated with a limited access to the probe hybridisation region. {F}urther investigations may identify a test combining a probe based-and an intercalating agent-based detection, which will significantly improve the diagnosis of {HRV} infections.},
  keywords = {{BOXTO} ; {SYBR} green ; {T}aqman probe ; {P}icornaviridae ; {H}uman rhinovirus ; {R}espiratory infections ; {M}olecular diagnosis ; {FRANCE}},
  booktitle = {},
  journal = {{V}irology {J}ournal},
  volume = {11},
  numero = {},
  pages = {art. 31},
  ISSN = {1743-422{X}},
  year = {2014},
  DOI = {10.1186/1743-422x-11-31},
  URL = {https://www.documentation.ird.fr/hor/fdi:010061980},
}