Dataset Information

Development of a TaqMan Array Card for Acute-Febrile-Illness Outbreak Investigation and Surveillance of Emerging Pathogens, Including Ebola Virus.

ABSTRACT: Acute febrile illness (AFI) is associated with substantial morbidity and mortality worldwide, yet an etiologic agent is often not identified. Convalescent-phase 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 h from sample to results and can simultaneously detect 26 AFI-associated organisms, including 15 viruses (chikungunya, Crimean-Congo hemorrhagic fever [CCHF] virus, dengue, Ebola virus, Bundibugyo virus, Sudan virus, hantaviruses [Hantaan and Seoul], hepatitis E, Marburg, Nipah virus, o'nyong-nyong virus, Rift Valley fever virus, West Nile virus, and yellow fever virus), 8 bacteria (Bartonella spp., Brucella spp., Coxiella burnetii, Leptospira spp., Rickettsia spp., Salmonella enterica and Salmonella enterica serovar Typhi, and Yersinia pestis), and 3 protozoa (Leishmania spp., Plasmodium spp., and Trypanosoma brucei). Two extrinsic controls (phocine herpesvirus 1 and bacteriophage MS2) were included to ensure extraction and amplification efficiency. Analytical validation was performed on spiked specimens for linearity, intra-assay precision, interassay 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 the TAC exhibited an overall 88% (278/315; 95% confidence interval [CI], 84% to 92%) sensitivity and a 99% (5,261/5,326, 98% to 99%) specificity. This TaqMan array card can be used in field settings as a rapid screen for outbreak investigation or for the surveillance of pathogens, including Ebola virus.

SUBMITTER: Liu J

PROVIDER: S-EPMC4702733 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Development of a TaqMan Array Card for Acute-Febrile-Illness Outbreak Investigation and Surveillance of Emerging Pathogens, Including Ebola Virus.

Liu Jie J Ochieng Caroline C Wiersma Steve S Ströher Ute U Towner Jonathan S JS Whitmer Shannon S Nichol Stuart T ST Moore Christopher C CC Kersh Gilbert J GJ Kato Cecilia C Sexton Christopher C Petersen Jeannine J Massung Robert R Hercik Christine C Crump John A JA Kibiki Gibson G Maro Athanasia A Mujaga Buliga B Gratz Jean J Jacob Shevin T ST Banura Patrick P Scheld W Michael WM Juma Bonventure B Onyango Clayton O CO Montgomery Joel M JM Houpt Eric E Fields Barry B

Journal of clinical microbiology 20151021 1

Acute febrile illness (AFI) is associated with substantial morbidity and mortality worldwide, yet an etiologic agent is often not identified. Convalescent-phase 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 h from sample to results and can simultaneously detect 26 AFI-associated organisms, including 15 viruses (chikungunya, Crime ...[more]

PMID: 26491176

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Project description:The TaqMan Array Card (TAC) system is a 384-well singleplex real-time PCR format that has been used to detect multiple infection targets. Here we developed an enteric TaqMan Array Card to detect 19 enteropathogens, including viruses (adenovirus, astrovirus, norovirus GII, rotavirus, and sapovirus), bacteria (Campylobacter jejuni/C. coli, Clostridium difficile, Salmonella, Vibrio cholerae, diarrheagenic Escherichia coli strains including enteroaggregative E. coli [EAEC], enterotoxigenic E. coli [ETEC], enteropathogenic E. coli [EPEC], and Shiga-toxigenic E. coli [STEC]), Shigella/enteroinvasive E. coli (EIEC), protozoa (Cryptosporidium, Giardia lamblia, and Entamoeba histolytica), and helminths (Ascaris lumbricoides and Trichuris trichiura), as well as two extrinsic controls to monitor extraction and amplification efficiency (the bacteriophage MS2 and phocine herpesvirus). Primers and probes were newly designed or adapted from published sources and spotted onto microfluidic cards. Fecal samples were spiked with extrinsic controls, and DNA and RNA were extracted using the QiaAmp Stool DNA minikit and the QuickGene RNA Tissue kit, respectively, and then mixed with Ag-Path-ID One Step real-time reverse transcription-PCR (RT-PCR) reagents and loaded into cards. PCR efficiencies were between 90% and 105%, with linearities of 0.988 to 1. The limit of detection of the assays in the TAC was within a 10-fold difference from the cognate assays performed on plates. Precision testing demonstrated a coefficient of variation of below 5% within a run and 14% between runs. Accuracy was evaluated for 109 selected clinical specimens and revealed an average sensitivity and specificity of 85% and 77%, respectively, compared with conventional methods (including microscopy, culture, and immunoassay) and 98% and 96%, respectively, compared with our laboratory-developed PCR-Luminex assays. This TAC allows fast, accurate, and quantitative detection of a broad spectrum of enteropathogens and is well suited for surveillance or clinical purposes.

Project description:BACKGROUND:The 2013-2015 Ebola Virus Disease (EVD) outbreak in Guinea resulted in community mistrust that influenced health care service utilization. This study aimed to assess whether EVD-related memories affect post-outbreak health-seeking behaviours for children under 5 years of age with febrile illnesses in Guéckédou district, Guinea. METHODS:This cross-sectional study was conducted by surveying caregivers of children under 5 years of age in the sub-district most affected by the EVD outbreak (Guèndembou) and the least affected sub-district (Bolodou) in Guéckédou district. Memories of the outbreak were referred to as EVD-related fears in the post-EVD period, which was based on a series of questions regarding current feelings. RESULTS:While the majority of caregivers sought care for their children with febrile illness in both districts, a statistically significantly higher proportion of caregivers in Guèndembou sought care, compared to caregivers in Bolodou.. More caregivers in Guèndembou (19.9%; n?=?39) reported the death of family members or friends due to EVD compared to Bolodou (6.9%; n?=?14; P?<?0.001). The mean EVD fear score of caregivers was significantly higher in Guèndembou (3.0; SD: 3.0) than in Bolodou (2.0; SD: 1.1) (p?<?0.001). Caregivers with a fear score above the median were 1.68 times more likely to seek care than those whose fear score was equal to or below the median; however, this difference was not statistically significant. Caregivers who reported family members' or friends' death due to EVD were also more likely to seek care (AOR?=?2.12; 95%CI: 0.91-4.91), however, with no statistical significance. Only residing in the EVD-most affected sub-district of Guèndembou (AOR?=?1.74; 95%CI: 1·09-2.79) was positively associated with seeking care. CONCLUSIONS:This study reveals that community members in the rural district of Guéckédou still live with fear related to EVD nearly 2 years after the outbreak. It calls for more efforts in the health domain to preserve communities' key values and address the psychosocial effect of EVD in rural Guinea.

Dataset Information

Development of a TaqMan Array Card for Acute-Febrile-Illness Outbreak Investigation and Surveillance of Emerging Pathogens, Including Ebola Virus.

Publications

Development of a TaqMan Array Card for Acute-Febrile-Illness Outbreak Investigation and Surveillance of Emerging Pathogens, Including Ebola Virus.

Similar Datasets

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