Drug resistance profile and subtyping of HIV-1 RT gene in Iranian patients under treatment

Identification of drug resistant mutations is important in the management of HIV-1 infected patients. The aim of the current study was to evaluate drug resistance profile of RT gene and assess subtypes among the HIV-1 circulating strains and intensification of physician's options for the best therapy. HIV-1 RNA of 25 samples was extracted from plasma and RT Nested- PCR was performed and the final products were sequenced and phylogenetically analyzed. Stanford HIV drug resistance sequence database was used for interpretation of the data. The results of phylogenetic analysis showed subtypes A1 and B in 14 [58%] and 10 [42%] patients respectively. Of the 24 patients, 16 [66.6%] had resistance to NRTIs, 8 individuals [32%] to NNRTIs and one patient was susceptible to NRTIs as well as NNRTIs. The drug resistance interpretation in this study showed: 87.7% susceptible for AZT, 70.8% susceptible, and 25% high-level resistance for 3TC, 87.7% susceptible for TDF, 29.1% high-level resistance for NVP and 70.8% susceptible and 25% high-level resistance for EFV. Our data suggests that probably, the use of 2 NRTIs plus 1 protease inhibitor [PI] regimen is more effective than 2 NRTIs plus 1 NNRTI regimen in Iranian patients that use 2 NRTI plus NNRTI regimen and also continuous surveillance should be perform to evaluate resistance patterns for more effective therapeutic approaches

Application of SYBR-Green Real Time RT-PCR for quantification of human immunodeficiency virus type-1 [HIV-1] and comparison with COBAS AMPLICOR test

In this study, a SYBR Green real-time RT-PCR assay for quantification of HIV-1 viral RNA was developed. This assay was performed based on amplification of the pol region of HIV-1 and product analysis by an ABI 7500 system. We quantified HIV-1 viral load in 26_seropositive patients by this system and the data were subsequently compared with results obtained with a reference technique represented by COBAS AMPLICOR HIB-1 Monitor test. The results demonstrated that this technique could detect up to 500 HIV-1 RNA copies/ml of plasma. The linearity of this approach was conserved over a wide range of HIV-1 copy numbers [5x10[2]- 5x10[9]]. Since no positive signal was observed in seronegative volunteers, the specificity of the test was calculated as 100%. Comparison of the results with those obtained by the reference quantification method, revealed a significant correlation between the results [R[2] =0.95]. On the basis of the most recent recorded cases for HIV-1 infection and AIDS in Iran, the prevalence of this disease is rising rapidly and the situation has been called to be alarming by national health representatives. Determination of HIV-1 viral load in plasma has been considered as the most effective single prediction tool for monitoring HIV-1 patients treated with antiviral drugs. In this study, we have developed a SYBR-Green Real Time RT-PCR assay for quantitative analysis of HIV-1 in infected patients. Since a synthetic RNA standard was used in this assay, the upper limit of detection was detected to be higher than the standard test [5x10[9] versus 7.5x10[5]]. This can be important in patients with acute high viral load infections. Reproducibility was assessed by Intra assay and Inter assay analysis, Coefficient of variations Ct, in reproducibility tests for Intra assay and Inter assay variability were less than 3% and 4.5% accordingly. The above results, indicates that the new developed test can be a used in substitution of the commercial assay for quantitative analysis of HIV-1