Comparative performance of the new Aptima HIV-1 Quant Dx assay with three commercial PCR-based HIV-1 RNA quantitation assays

AbstractBackgroundQuantitative measurement of HIV-1 RNA levels in plasma (‘viral load’) plays a central role in clinical management. The choice of assay platform can influence results and treatment decisions.ObjectiveTo compare the analytical performance of the new TMA-based Hologic Aptima® HIV-1 Quant Dx assay with that of three PCR-based assays: Abbott RealTime HIV-1, Qiagen Artus® HI Virus-1 QS-RGQ, and Roche CAP/CTM HIV-1 Test v2.Study designAssay performance was evaluated using Acrometrix HIV-1 RNA Standard panels; the 3rd WHO HIV-1 RNA International Standard (12–500copies/ml; 6 dilutions; 9 replicates); and plasma samples from 191 HIV-positive patients.ResultsAptima showed high (>0.99) precision, accuracy and concordance with the Acrometrix Standards across a wide dynamic range (2.0–6.7 log10copies/ml). Variance caused up to 2.1 (Aptima), 1.7 (RealTime), 7.5 (Artus), and 1.9 (CAP/CTM) fold changes in the International Standard quantifications at 50–500copies/ml. HIV-1 RNA detection rates in plasma samples were 141/191 (74%), 119/191 (62%), 108/191 (57%), and 145/191 (76%) for Aptima, RealTime, Artus and CAP/CTM, respectively. For categorising samples either side of 50 copies/ml, Aptima had excellent agreement with RealTime (kappa 0.92; 95% CI 0.87–0.98); lowest agreement was with Artus (kappa 0.79; 95%CI 0.70–0.88). Aptima quantifications were mean 0.12 and 0.06 log10copies/ml higher compared with RealTime and CAP/CTM, respectively, and 0.05 log10copies/ml lower compared with Artus. Limits of agreement were narrowest when comparing Aptima to RealTime.ConclusionsThe new Aptima HIV assay is sensitive, precise, and accurate. HIV assays exhibit discordance at low HIV-1 RNA copy numbers

Drug resistance outcomes of long-term ART with tenofovir disoproxil fumarate in the absence of virological monitoring

Objectives: The resistance profiles of patients receiving long-term ART in sub-Saharan Africa have been poorly described. This study obtained a sensitive assessment of the resistance patterns associated with long-term tenofovir-based ART in a programmatic setting where virological monitoring is yet to become part of routine care. Methods: We studied subjects who, after a median of 4.2 years of ART, replaced zidovudine or stavudine with tenofovir disoproxil fumarate while continuing lamivudine and an NNRTI. Using deep sequencing, resistance-associated mutations (RAMs) were detected in stored samples collected at tenofovir introduction (T0) and after a median of 4.0 years (T1). Results: At T0, 19/87 (21.8%) subjects showed a detectable viral load and 8/87 (9.2%) had one or more major NNRTI RAMs, whereas 82/87 (94.3%) retained full tenofovir susceptibility. At T1, 79/87 (90.8%) subjects remained on NNRTI-based ART, 5/87 (5.7%) had introduced lopinavir/ritonavir due to immunological failure, and 3/87 (3.4%) had interrupted ART. Whilst 68/87 (78.2%) subjects maintained or achieved virological suppression between T0 and T1, a detectable viral load with NNRTI RAMs at T0 predicted lack of virological suppression at T1. Each treatment interruption, usually reflecting unavailability of the dispensary, doubled the risk of T1 viraemia. Tenofovir, lamivudine and efavirenz selected for K65R, K70E/T, L74I/V and Y115F, alongside M184V and multiple NNRTI RAMs; this resistance profile was accompanied by high viral loads and low CD4 cell counts. Conclusions: Viraemia on tenofovir, lamivudine and efavirenz led to complex resistance patterns with implications for continued drug activity and risk of onward transmission

Influenza-like illness in acute myocardial infarction patients during the winter wave of the influenza A H1N1 pandemic in London: a case-control study.

OBJECTIVE: To investigate recent respiratory and influenza-like illnesses (ILIs) in acute myocardial infarction patients compared with patients hospitalised for acute non-vascular surgical conditions during the second wave of the 2009 influenza A H1N1 pandemic. DESIGN: Case-control study. SETTING: Coronary care unit, acute cardiology and acute surgical admission wards in a major teaching hospital in London, UK. PARTICIPANTS: 134 participants (70 cases and 64 controls) aged ≥40 years hospitalised for acute myocardial infarction and acute surgical conditions between 21 September 2009 and 28 February 2010, frequency-matched for gender, 5-year age-band and admission week. PRIMARY EXPOSURE: ILI (defined as feeling feverish with either a cough or sore throat) within the last month. SECONDARY EXPOSURES: Acute respiratory illness within the last month not meeting ILI criteria; nasopharyngeal and throat swab positive for influenza virus. RESULTS: 29 of 134 (21.6%) participants reported respiratory illness within the last month, of whom 13 (9.7%) had illnesses meeting ILI criteria. The most frequently reported category for timing of respiratory symptom onset was 8-14 days before admission (31% of illnesses). Cases were more likely than controls to report ILI-adjusted OR 3.17 (95% CI 0.61 to 16.47)-as well as other key respiratory symptoms, and were less likely to have received influenza vaccination-adjusted OR 0.46 (95% CI 0.19 to 1.12)-although the differences were not statistically significant. No swabs were positive for influenza virus. CONCLUSIONS: Point estimates suggested that recent ILI was more common in patients hospitalised with acute myocardial infarction than with acute surgical conditions during the second wave of the influenza A H1N1 pandemic, and influenza vaccination was associated with cardioprotection, although the findings were not statistically significant. The study was underpowered, partly because the age groups typically affected by acute myocardial infarction had low rates of infection with the pandemic influenza strain compared with seasonal influenza

Kinetics and specificities of the T helper-cell response to gp120 in the asymptomatic stage of HIV-1 infection

Peripheral blood mononuclear cells from 36 asymptomatic HIV-1 seropositive individuals were tested longitudinally for in vitro T–cell proliferation and IL–2 production in response to synthetic peptides spanning the entire gp120 of HIV–1. At baseline, significant T–cell proliferation to pooled and individual peptides was observed in 15 of the 36 donors. After 12 months, proliferate responses to peptide pools were lost or decreased significantly in most donors. Responses appeared to fluctuate over time: at 12 months new recognition sites were detected in four of 10 donors showing T–cell proliferation at baseline, as well as in five of 15 donors with no previous proliferative responses. IL–2 production appeared to be a more sensitive and longer preserved parameter of T–helper cell function: at baseline the majority of donors with no T–cell proliferation produced IL–2 in response to pooled peptides. This response was not decreased significantly after 12 months. The overall patterns of response to both pooled and individual peptides were heterogeneous among donors. Multiple recognition sites were detected in both variable and conserved regions of gp120, but no pool or individual peptide was recognized by all responders. Functional T–cell responses were not statistically correlated to CD4° cell percentile and absolute numbers

Simian immunodeficiency virus (SIV)-specific CD8+ cytotoxic T lymphocyte responses of naive and vaccinated cynomolgus macaques infected with (SIV)mac32H(J5): quantitative analysis by in vitro antigenic stimulation.

Detailed analyses of simian immunodeficiency virus (SIV)-specific cytotoxic T lymphocyte (CTL) responses in vaccinated and infected macaques may help to clarify the role of CTL immunity in protection against lentiviruses. Here, the optimal conditions for the measurement of SIV Gag-specific CTL were investigated by bulk and limiting dilution assays of peripheral blood mononuclear cells (PBMC) from naive and vaccinated cynomolgus macaques (Macaca fascicularis) infected with SIVmac32H(J5). In vitro restimulation was generally required for CTL detection. Selective activation of CD8+ and MHC-restricted SIV Gag-specific CTL was induced by stimulation with autologous para-formaldehyde-fixed B-lymphoblastoid cell lines infected with a recombinant vaccinia virus expressing SIV Gag. Applied to limiting dilution assays, antigenic stimulation reproducibly demonstrated SIV Gag-specific CTL precursors (CTLp) in PBMC of all animals studied, including those lacking significant responses in standard bulk CTL assays