title: Validation and Application of a Commercial Quantitative Real-Time Reverse Transcriptase-PCR Assay in Investigation of a Large Dengue Virus Outbreak in Southern Taiwan.
abstract: BACKGROUND
  Accurate, rapid, and early diagnosis of dengue virus (DENV) infections is essential for optimal clinical care. Here, we evaluated the efficacy of the quantitative real-time PCR (qRT-PCR)-LightMix dengue virus EC kit for DENV detection using samples from a dengue outbreak in Taiwan in 2015.
  
  
  METHODS
  Sera from patients with suspected DENV infection were analyzed and compared using the LightMix kit, a Dengue NS1 Ag + Ab Combo kit for detection of NS1 antigen and DENV-specific IgM and IgG antibodies, and an "in-house" qualitative DENV-specific RT-PCR assay.
  
  
  RESULTS
  A total of 8,989, 8,954, and 1581 samples were subjected to NS1 antigen detection, IgM and IgG detection, and LightMix assays, respectively. The LightMix assay yielded a linear curve for viral loads (VL) between 102 and 106 copies/reaction, and the minimum detection limits for DENV serotype 1 (DENV1) and DENV2, DENV3, and DENV4 were 1, 10, and 100 focus forming units (FFU)/mL, respectively. There was 88.9% concordance between the results obtained using the NS1 antigen combo kit and by LightMix analysis, and the diagnostic sensitivity and specificity of the two methods were 89.4 and 100%, and 84.7 and 100%, respectively. Notably, fatal cases were attributed to DENV2 infection, and 79.5% (27/34) of these cases occurred in patients ≥ 71 years of age. Among these older patients, 82.3% (14/17) were NS1/IgM/IgG (+/-/-), exhibiting VLs between 106-109 copies/mL, which was markedly higher than the rate observed in the other age groups.
  
  
  CONCLUSIONS
  The LightMix assay was effective for early diagnosis of DENV infection. Our data indicate that high VLs during primary infection in elderly patients may be a positive predictor for severe illness, and may contribute to high mortality rates.
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.|Medical Laboratory Science and Biotechnology, National Cheng Kung University, Tainan, Taiwan.
authors.forename: Huey-Pin
authors.initials: HP
authors.lastname: Tsai
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: You-Yuan
authors.initials: YY
authors.lastname: Tsai
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: I-Ting
authors.initials: IT
authors.lastname: Lin
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: Pin-Hwa
authors.initials: PH
authors.lastname: Kuo
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: Kung-Chao
authors.initials: KC
authors.lastname: Chang
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: Jung-Chin
authors.initials: JC
authors.lastname: Chen
authors.affiliation: Department of Internal Medicine, National Cheng Kung University Hospital, Tainan, Taiwan.
authors.forename: Wen-Chien
authors.initials: WC
authors.lastname: Ko
authors.affiliation: Department of Pathology, National Cheng Kung University Hospital, Tainan, Taiwan.|Medical Laboratory Science and Biotechnology, National Cheng Kung University, Tainan, Taiwan.|Center of Infectious Disease and Signaling Research, National Cheng Kung University, Tainan, Taiwan.|National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Tainan, Taiwan.
authors.forename: Jen-Ren
authors.initials: JR
authors.lastname: Wang
mesh_terms: D000368:Aged; D000914:Antibodies, Viral / Q000097:blood; D003715:Dengue / Q000097:blood / Q000175:diagnosis / Q000453:epidemiology / Q000821:virology*; D003716:Dengue Virus / Q000145:classification / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D004196:Disease Outbreaks; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013624:Taiwan; D017361:Viral Nonstructural Proteins / Q000235:genetics
chemical_list: D000914:Antibodies, Viral; D017361:Viral Nonstructural Proteins