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. Recombin...

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Autores principales:	David S. Boyle, Dara A. Lehman, Lorraine Lillis, Dylan Peterson, Mitra Singhal, Niall Armes, Mathew Parker, Olaf Piepenburg, Julie Overbaugh
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2013
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9efe99b3ee694f36976daec286e670d5
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spelling	oai:doaj.org-article:9efe99b3ee694f36976daec286e670d52021-11-15T15:40:27ZRapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification10.1128/mBio.00135-132150-7511https://doaj.org/article/9efe99b3ee694f36976daec286e670d52013-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00135-13https://doaj.org/toc/2150-7511ABSTRACT 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. IMPORTANCE Diagnosis of HIV-1 infection in infants cannot rely on the antibody-based tests used in adults because of the transfer of maternal HIV-1 antibodies from mother to child. Therefore, infant diagnostics rely on detection of the virus itself. However, current infant HIV-1 diagnostic methods require a laboratory setting with complex equipment. Here we describe the initial development of an HIV-1 diagnostic for infants that may be performed at the point of care in rural health clinics. We utilize a method that can amplify and detect HIV-1 DNA at an incubation temperature within the range of 25 to 42°C, eliminating the need for thermocycling equipment. HIV-1 diagnostics are challenging to develop due to the high diversity seen in HIV-1 strains worldwide. Here we show that this method detects the major HIV-1 strains circulating globally.David S. BoyleDara A. LehmanLorraine LillisDylan PetersonMitra SinghalNiall ArmesMathew ParkerOlaf PiepenburgJulie OverbaughAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 4, Iss 2 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 David S. Boyle Dara A. Lehman Lorraine Lillis Dylan Peterson Mitra Singhal Niall Armes Mathew Parker Olaf Piepenburg Julie Overbaugh Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
description	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. IMPORTANCE Diagnosis of HIV-1 infection in infants cannot rely on the antibody-based tests used in adults because of the transfer of maternal HIV-1 antibodies from mother to child. Therefore, infant diagnostics rely on detection of the virus itself. However, current infant HIV-1 diagnostic methods require a laboratory setting with complex equipment. Here we describe the initial development of an HIV-1 diagnostic for infants that may be performed at the point of care in rural health clinics. We utilize a method that can amplify and detect HIV-1 DNA at an incubation temperature within the range of 25 to 42°C, eliminating the need for thermocycling equipment. HIV-1 diagnostics are challenging to develop due to the high diversity seen in HIV-1 strains worldwide. Here we show that this method detects the major HIV-1 strains circulating globally.
format	article
author	David S. Boyle Dara A. Lehman Lorraine Lillis Dylan Peterson Mitra Singhal Niall Armes Mathew Parker Olaf Piepenburg Julie Overbaugh
author_facet	David S. Boyle Dara A. Lehman Lorraine Lillis Dylan Peterson Mitra Singhal Niall Armes Mathew Parker Olaf Piepenburg Julie Overbaugh
author_sort	David S. Boyle
title	Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
title_short	Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
title_full	Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
title_fullStr	Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
title_full_unstemmed	Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification
title_sort	rapid detection of hiv-1 proviral dna for early infant diagnosis using recombinase polymerase amplification
publisher	American Society for Microbiology
publishDate	2013
url	https://doaj.org/article/9efe99b3ee694f36976daec286e670d5
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Rapid Detection of HIV-1 Proviral DNA for Early Infant Diagnosis Using Recombinase Polymerase Amplification

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