Abstract
Testing HIV-1 RNA in plasma by PCR is universally accepted as the ultimate standard to confirm diagnosis of HIV-1 infection and to monitor viral load in patients under treatment. However, in some cases, this assay could either underestimate or overestimate the replication capacity of a circulating or latent virus. In the present study, we performed the assessment of evaluating the HIV-1 reverse transcriptase (RT) activity by means of a new assay for the functional screening of the status of HIV-1 patients. To this purpose, we utilized, for the first time on blood samples, an adapted version of a real-time RT quantitative PCR assay, utilized to evaluate the HIV-1-RT inhibitory activity of compounds. The study analyzed blood samples from 28 HIV-1-infected patients, exhibiting a wide range of viremia and immunological values. Results demonstrated that plasma HIV-1 RT levels, expressed as cycle threshold values obtained with the assay under appraisal, were inversely and highly significantly correlated with the plasma HIV-1-RNA levels of the patients. Thus, an HIV-1 RT quantitative PCR assay was created which we describe in this study, and it may be considered as a promising basis for an additional tool capable of furnishing information on the functional virological status of HIV-1-infected patients.
Highlights
The key aim of the combination anti-HIV-1 antiretroviral therapy (ART) was to block the progression of the disease, significantly reducing virus levels in plasma in most of patients but without leading to the eradication of the virus
The novel RT quantitative PCR (RT-qPCR) assay for evaluating the RT activity in blood samples from HIV-1-infected individuals was developed by setting up a variant version of the two-step RT-qPCR-based assay, already described by us for evaluating the efficacy of compounds towards HIV-1 reverse transcriptase [20]
HIV-RT generated a cDNA, using, as a specific template for RT, RNA extracted from cells ectopically expressing the glycoprotein D (gD) protein of herpes simplex virus 1 (HSV-1) and using, as a specific primer, a related targeted oligonucleotide
Summary
The key aim of the combination anti-HIV-1 antiretroviral therapy (ART) was to block the progression of the disease, significantly reducing virus levels in plasma in most of patients but without leading to the eradication of the virus. HIV-1 infection has taken on the characteristics of a chronic infection. Pathogens 2020, 9, 1047 in properly treated patients. The risk of exacerbation of the infection is always there. Chronicization of infection in ART patients has encouraged the establishment of “reservoirs”, in which, potentially, a replication-competent virus persists. The nature and clinical significance of these HIV-1. “reservoirs” seem rather heterogeneous and difficult to precisely quantify [1,2]. Evaluation of the plasma viral load cannot give information on the replication-competent reservoir.
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