title: Rapid detection of HIV-1 proviral DNA for early infant diagnosis using recombinase polymerase amplification.
abstract: Early diagnosis and treatment of human immunodeficiency virus type 1 (HIV-1) infection in infants can greatly reduce mortality rates. However, current infant HIV-1 diagnostics cannot reliably be performed at the point of care, often delaying treatment and compromising its efficacy. Recombinase polymerase amplification (RPA) is a novel technology that is ideal for an HIV-1 diagnostic, as it amplifies target DNA in <20 min at a constant temperature, without the need for complex thermocycling equipment. Here we tested 63 HIV-1-specific primer and probe combinations and identified two RPA assays that target distinct regions of the HIV-1 genome (long terminal repeat [LTR] and pol) and can reliably detect 3 copies of proviral DNA by the use of fluorescence detection and lateral-flow strip detection. These pol and LTR primers amplified 98.6% and 93%, respectively, of the diverse HIV-1 variants tested. This is the first example of an isothermal assay that consistently detects all of the major HIV-1 global subtypes.
authors.affiliation: Program for Appropriate Technology in Health, Seattle, WA, USA. dboyle@path.org
authors.forename: David S
authors.initials: DS
authors.lastname: Boyle
authors.forename: Dara A
authors.initials: DA
authors.lastname: Lehman
authors.forename: Lorraine
authors.initials: L
authors.lastname: Lillis
authors.forename: Dylan
authors.initials: D
authors.lastname: Peterson
authors.forename: Mitra
authors.initials: M
authors.lastname: Singhal
authors.forename: Niall
authors.initials: N
authors.lastname: Armes
authors.forename: Mathew
authors.initials: M
authors.lastname: Parker
authors.forename: Olaf
authors.initials: O
authors.lastname: Piepenburg
authors.forename: Julie
authors.initials: J
authors.lastname: Overbaugh
mesh_terms: D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000032:analysis* / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D042241:Early Diagnosis; D015658:HIV Infections / Q000175:diagnosis* / Q000821:virology; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D007223:Infant; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D015345:Oligonucleotide Probes / Q000235:genetics; D011533:Proviruses / Q000235:genetics / Q000302:isolation & purification*; D045522:Recombinases / Q000378:metabolism; D012680:Sensitivity and Specificity; D013696:Temperature; D014773:Virology / Q000379:methods
chemical_list: D017931:DNA Primers; D004279:DNA, Viral; D015345:Oligonucleotide Probes; D045522:Recombinases; D004259:DNA-Directed DNA Polymerase
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