Simultaneous detection of Zika, Chikungunya and Dengue viruses by a multiplex real-time RT-PCR assay.
In the recent past, arboviruses such as Chikungunya (CHIKV) and Zika (ZIKV) have increased their area of endemicity and presented as an emerging global public health threat.



To design an assay for the simultaneous detection of ZIKV, CHIKV and Dengue (DENV) 1-4 from patients with symptoms of arboviral infection. This would be advantageous because of the similar clinical presentation typically encountered with these viruses and their co-circulation in endemic areas.



In this study we have developed and validated a triplex real time reverse transcription PCR assay using hydrolysis probes targeting the non-structural 5 (NS5) region of ZIKV, non-structural protein 4 (nsP4) from CHIKV and 3' untranslated region (3'UTR) of DENV 1-4.



The 95% LOD by the triplex assay was 15 copies/reaction for DENV-1 and less than 10 copies/reaction for all other viruses. The triplex assay was 100% specific and did not amplify any of the other viruses tested. The assay was reproducible and adaptable to testing different specimen types including serum, plasma, urine, placental tissue, brain tissue and amniotic fluid. This assay can be easily implemented for diagnostic testing of patient samples, even in a high throughput laboratory.
Provincial Laboratory for Public Health, Calgary, Alberta, Canada. Electronic address: kanti.pabbaraju@ahs.ca.
Kanti
K
Pabbaraju
Provincial Laboratory for Public Health, Calgary, Alberta, Canada. Electronic address: sallene.wong@ahs.ca.
Sallene
S
Wong
Provincial Laboratory for Public Health, Calgary, Alberta, Canada. Electronic address: kara.gill2@ahs.ca.
Kara
K
Gill
Provincial Laboratory for Public Health, Calgary, Alberta, Canada; Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Alberta, Canada University of Calgary, Alberta, Canada. Electronic address: kevin.fonseca@ahs.ca.
Kevin
K
Fonseca
Provincial Laboratory for Public Health, Calgary, Alberta, Canada; Department of Pathology and Laboratory Medicine, University of Alberta, Edmonton, Alberta, Canada. Electronic address: graham.tipples@ahs.ca.
Graham A
GA
Tipples
Provincial Laboratory for Public Health, Calgary, Alberta, Canada; Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Alberta, Canada University of Calgary, Alberta, Canada. Electronic address: raymond.tellier@ahs.ca.
Raymond
R
Tellier
D001826:Body Fluids / Q000821:virology; D002646:Chikungunya virus / Q000235:genetics*; D060085:Coinfection / Q000175:diagnosis / Q000821:virology; D003716:Dengue Virus / Q000235:genetics*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D012327:RNA Virus Infections / Q000175:diagnosis / Q000821:virology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D000071244:Zika Virus / Q000235:genetics*
Arbovirus; Chikungunya; Dengue; Molecular diagnosis; Multiplex real-time RT-PCR; Zika