Our goal is to develop an integrated diagnostic test for HIV-1 viral load with high sensitivity, specificity, reliability, and reproducibility for use in minimal infrastructure settings. Such a test, applied in perinatal HIV-1 diagnosis, could save 180,000 DALYs each year if 5% of the targeted population has early access to therapy, and up to 2.5 million DALYs each year if 100% of the population has access to treatment [2]. It would also help to overcome one of the major challenges to universal access to therapy: the lack of adequate diagnosis and treatment of pediatric HIV-1 disease (WHO) [1]. Currently the most sensitive and reliable assays to quantify HIV-1 viral load rely on nucleic acid amplification and detection, but these tests often require sophisticated instrumentation and expensive reagents. Alternatively, the high analytical sensitivity of oligonucleotide-coated gold nanoparticles as targeting/reporting agents in nucleic acid tests has been demonstrated [2, 3]. To achieve the required sensitivity, we will develop an HIV-1 viral load assay which integrates isothermal PCR amplification of HIV-1 RNA with the use of targeted gold nanoparticles and colorimetric quantification of test results. We will develop this assay for use in low - resource settings, with minimal infrastructure requirements and a sensitivity and specificity comparable to that of commercial viral load assays available in the developed world.
The specific aims of the proposal are to: (1) Develop an inexpensive, sensitive, and specific diagnostic test for determining type 1 HIV viral loads of seropositive patients in low-resource settings. The assay will combine: target isolation and isothermal amplification technologies to yield at least 104 fold amplification from samples containing a minimum of 1000 viral copies/ml. An oligonucleotide-targeted gold nanoparticle detection assay and a quantitative read-out will be then used to detect to achieve a dynamic detection range from 103 to 106 HIV-1 viral copies/ml at the POC. (2) Validate the performance of this assay for HIV-1 viral load determination in clinical specimens. In collaboration with Dr. Richard Sutton from Yale School of Medicine who has expertise in molecular biology of HIV-1, we will test the ability of the assay to detect RNA from whole viral particles, and from group M clades on an NIH/UNAIDS reference panel, comparing the assay to RT-PCR. In collaboration with Dr. Elizabeth Molyneux from Queen Elizabeth Central Hospital, Blantyre, Malawi who has expertise in clinical diagnosis of HIV-1, we will carry out a pilot study to determine the sensitivity and specificity of this new assay. Personnel from Dr. Molyneux's team will travel to Houston to learn the assay, and members of the Richards-Kortum lab will travel to Malawi to work with her group to evaluate the assay in pediatric clinical samples relative to the gold standard of dried blood spot RT-PCR.
Our goal is to develop an integrated diagnostic test for HIV viral load with high sensitivity, specificity, reliability, and reproducibility for use in minimal infrastructure settings. Viral load determination is needed to determine when to initiate therapy, monitor compliance, and most importantly, as an early indicator of therapeutic failure. Despite the encouraging trends in the scale-up of antiretroviral treatment in low- and middle-income countries reported by the WHO, reliable and accurate HIV load testing has yet to be introduced into the management of infected patients in low resource settings and remains one of the major challenges to universal access to therapy [1]. Such a diagnostic test, applied in perinatal diagnosis, could save 180,000 DALYs each year if 5% of the targeted population has early access to therapy, and up to 2.5 million DALYs if 100% of the population has access to treatment [2].
| Rohrman, Brittany A; Leautaud, Veronica; Molyneux, Elizabeth et al. (2012) A lateral flow assay for quantitative detection of amplified HIV-1 RNA. PLoS One 7:e45611 |
| Rohrman, Brittany A; Richards-Kortum, Rebecca R (2012) A paper and plastic device for performing recombinase polymerase amplification of HIV DNA. Lab Chip 12:3082-8 |
| Cordray, Michael S; Amdahl, Matthew; Richards-Kortum, Rebecca R (2012) Gold nanoparticle aggregation for quantification of oligonucleotides: optimization and increased dynamic range. Anal Biochem 431:99-105 |
