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A Simple, Cost-Effective, and Extraction-Free Molecular Diagnostic Test for Sickle Cell Disease Using a Noninvasive Buccal Swab Specimen for a Limited-Resource Setting.

ABSTRACT: Sickle cell disease (SCD) is the most prevalent life-threatening blood monogenic disorder. Currently, there is no cure available, apart from bone marrow transplantation. Early and efficient diagnosis of SCD is key to disease management, which would make considerable strides in alleviating morbidity and reducing mortality. However, the cost and complexity of diagnostic procedures, such as the Sanger sequencing method, impede the early detection of SCD in a resource-limited setting. To address this, the current study demonstrates a simple and efficient proof-of-concept assay for the detection of patients and carriers using extraction-free non-invasive buccal swab samples by isothermal DNA Amplification coupled Restrictase-mediated cleavage (iDAR). This study is a first of its kind reporting the use of buccal swab specimens for iDA in molecular diagnosis of a genetic disease, all the while being cost effective and time saving, with the total assay time of around 150 min at a cost of USD 5. Further, iDAR demonstrates 91.5% sensitivity and 100% specificity for detecting all three alleles: SS, AS, and AA, having a 100% concordance with Sanger sequencing. The applicability of the iDAR assay is further demonstrated with its adaptation to a one-pot reaction format, which simplifies the assay system. Overall, iDAR is a simple, cost-effective, precise, and non-invasive assay for SCD screening, with the potential for use in a limited resource setting.

SUBMITTER: Thakur P

PROVIDER: S-EPMC9318149 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundWorld Health Organization guidelines now recommend routine use of viral load testing, where available, for patients receiving antiretroviral treatment (ART). However, its use has not been routinely implemented in many resource-limited settings due to cost, availability and accessibility. Viral load testing is complex, making its application in resource-limited settings challenging. We describe the issues encountered by Médecins Sans Frontières (MSF) when using routine viral load testing in a large HIV programme in sub-Saharan Africa.MethodsBetween October 2005 and August 2006, more than 1200 patients on ART had viral load tests at baseline and at three-month intervals performed by a local reference laboratory that was quality assured by an experienced international institution. Concerns with reliability of results halted testing. The quality control measures instituted with a second laboratory and outcomes of these were documented.ResultsIn 2005 and 2006, only 178 of 334 (53%) previously ART-naïve patients tested after six to 12 months of treatment had viral loads of less than 1000 copies/mL. Similar MSF programmes elsewhere demonstrated virological suppression rates of more than 85%, and duplicate testing showed unacceptable discordance. Laboratory problems encountered included: disregarded quality control; time delays; requirement for retesting; and duplicate sample variations. Potentially harmful clinical outcomes of inaccurate viral load results include: unnecessary ART regimen changes; unnecessary enhanced adherence counselling after "false failures"; and undetected virological failure.ConclusionsViral load testing performed without rigorous quality control carries the risk of erroneous and potentially damaging results. Viral load testing should be utilized only if robust quality assurance has been implemented. Our experience in this and other settings led to the development of a guide for assessing the suitability of a laboratory for viral load testing that can be used to help achieve reliable results.

Dataset Information

A Simple, Cost-Effective, and Extraction-Free Molecular Diagnostic Test for Sickle Cell Disease Using a Noninvasive Buccal Swab Specimen for a Limited-Resource Setting.

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