Development of a TaqMan Array Card for Acute Febrile Illness Outbreak Investigation and Surveillance of Emerging Pathogens including Ebola Virus

Jie Liu, Caroline Ochieng, Steve Wiersma, Ute Ströher, Jonathan S. Towner, Shannon Whitmer, Stuart T. Nichol, Christopher C. Moore, Gilbert J. Kersh, Cecilia Kato, Christopher Sexton, Jeannine Petersen, Robert Massung, Christine Hercik, John A. Crump, Gibson Kibiki, Athanasia Maro, Buliga Mujaga, Jean Gratz, Shevin T. Jacob, Patrick Banura, W. Michael Scheld, Bonventure Juma, Clayton O. Onyango, Joel M. Montgomery, Eric Houpt, Barry Fields
Publication year: 

Acute febrile illness (AFI) is associated with substantial morbidity and mortality worldwide yet an etiologic agent is often not identified. Convalescent serology is impractical, blood culture is slow, and many pathogens are fastidious or impossible to cultivate. We developed a real-time PCR based TaqMan Array Card (TAC) that can test six to eight samples within 2.5 hours from sample to results and simultaneously detect 26 AFI associated organisms, including 15 viruses (Chikungunya, Crimean-Congo hemorrhagic fever virus, Dengue, Ebola virus, Bundibugyo virus, Sudan virus, Hantaviruses (HTN and SEO), Hepatitis E, Marburg, Nipah virus, O’nyong- nyong virus, Rift Valley fever virus, West Nile virus, Yellow fever virus), eight bacteria (Bartonella spp., Brucella spp., Coxiella burnetii, Leptospira spp., Rickettsia spp., Salmonella enterica and Salmonella enterica serovar Typhi, Yersinia pestis), and three protozoa (Leishmania spp., Plasmodium spp., Trypanosoma brucei). Two extrinsic controls (Phocine Herpesvirus 1 and bacteriophage MS2) were included to assure extraction and amplification efficiency. Analytical validation was performed on spiked specimens for linearity, intra-assay precision, inter-assay precision, limit of detection, and specificity. The performance of the card on clinical specimens was evaluated with 1,050 blood samples by comparison to the individual real-time PCR assays, and TAC exhibited overall 88% (278/315, 95% confidence interval 84% to 92%) sensitivity and 99% (5261/5326, 98% to 99%) specificity. This TaqMan Array Card can be used in field settings 53 as a rapid screen for outbreak investigation or pathogen surveillance, including Ebola virus.