title: Multiplex nucleic acid amplification test for diagnosis of dengue fever, malaria, and leptospirosis.
abstract: Dengue, leptospirosis, and malaria are among the most common etiologies of systemic undifferentiated febrile illness (UFI) among travelers to the developing world, and these pathogens all have the potential to cause life-threatening illness in returned travelers. The current study describes the development of an internally controlled multiplex nucleic acid amplification test for the detection of dengue virus (DENV) and Leptospira and Plasmodium species, with a specific callout for Plasmodium falciparum (referred to as the UFI assay). During analytical evaluation, the UFI assay displayed a wide dynamic range and a sensitive limit of detection for each target, including all four DENV serotypes. In a clinical evaluation including 210 previously tested samples, the sensitivities of the UFI assay were 98% for DENV (58/59 samples detected) and 100% for Leptospira and malaria (65/65 and 20/20 samples, respectively). Malaria samples included all five Plasmodium species known to cause human disease. The specificity of the UFI assay was 100% when evaluated with a panel of 66 negative clinical samples. Furthermore, no amplification was observed when extracted nucleic acids from related pathogens were tested. Compared with whole-blood samples, the UFI assay remained positive for Plasmodium in 11 plasma samples from patients with malaria (parasitemia levels of 0.0037 to 3.4%). The syndrome-based design of the UFI assay, combined with the sensitivities of the component tests, represents a significant improvement over the individual diagnostic tests available for these pathogens.
authors.affiliation: Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Jesse J
authors.initials: JJ
authors.lastname: Waggoner
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Janaki
authors.initials: J
authors.lastname: Abeynayake
authors.affiliation: Laboratório de Zoonoses Bacterianas, Centro de Referência Nacional para Leptospirose, Coleção de Leptospira, WHO/PAHO Centro Colaborador para Leptospirose, Instituto Oswaldo Cruz, Rio de Janeiro, Brazil.
authors.forename: Ilana
authors.initials: I
authors.lastname: Balassiano
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Martina
authors.initials: M
authors.lastname: Lefterova
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Malaya K
authors.initials: MK
authors.lastname: Sahoo
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Laboratório de Zoonoses Bacterianas, Centro de Referência Nacional para Leptospirose, Coleção de Leptospira, WHO/PAHO Centro Colaborador para Leptospirose, Instituto Oswaldo Cruz, Rio de Janeiro, Brazil.
authors.forename: Juliana Magalhães
authors.initials: JM
authors.lastname: Vital-Brazil
authors.affiliation: Sustainable Sciences Institute, Managua, Nicaragua.
authors.forename: Lionel
authors.initials: L
authors.lastname: Gresh
authors.affiliation: National Virology Laboratory, Centro Nacional de Diagnóstico y Referencia, Ministry of Health, Managua, Nicaragua.
authors.forename: Angel
authors.initials: A
authors.lastname: Balmaseda
authors.affiliation: Division of Infectious Diseases and Vaccinology, School of Public Health, University of California, Berkeley, California, USA.
authors.forename: Eva
authors.initials: E
authors.lastname: Harris
authors.affiliation: Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California, USA Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Niaz
authors.initials: N
authors.lastname: Banaei
authors.affiliation: Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California, USA Department of Pathology, Stanford University School of Medicine, Stanford, California, USA bpinsky@stanford.edu.
authors.forename: Benjamin A
authors.initials: BA
authors.lastname: Pinsky
mesh_terms: D000293:Adolescent; D002648:Child; D003715:Dengue / Q000175:diagnosis*; D003716:Dengue Virus / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007919:Leptospira / Q000235:genetics / Q000302:isolation & purification; D007922:Leptospirosis / Q000175:diagnosis*; D008288:Malaria / Q000175:diagnosis*; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D010961:Plasmodium / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity