Variability and recombination of clinical human cytomegalovirus strains from transplantation recipients.

BACKGROUND: Human cytomegalovirus (HCMV) is the first cause of viral infection in immunocompromised transplanted patients. OBJECTIVES: Here, five HCMV genes were studied to investigate the existence of recombination events in clinical strains ex vivo. STUDY DESIGN: Sequencing and phylogenetic analysis were conducted on 21 strains from 16 renal and 5 lung transplant recipients. RESULTS: Nucleotidic polymorphism ranged from 6.6% (US3) to 12% (UL40), with a significant proportion of missense mutations (39-69%), some of which could have a functional impact. Analysis of the concatenated sequence (4804 nucleotides for each strain) evidenced two clusters of sequences presenting a reticulate topology suggestive of recombination events (SplitsTree). Phi-test pointed numerous phylogenetically conflicting signals indicating a high statistical probability of recombination. The subsequent bootscan analysis was consistent with these data. CONCLUSIONS: These results reinforce the prominent role of recombination in HCMV evolutionary history and adaptation to its host.

Virological characterization of an infection with a dual-tropic, multidrug-resistant HIV-1 and further evolution on antiretroviral therapy.

We studied a case of recent infection with multidrug-resistant (MDR) HIV-1. Over 16 months off-therapy, the CD4 cell count decreased from 419 to 184 cells/mul. Antiretroviral therapy (ART) then led to an incomplete virological response but to an immunological benefit, concurrently with a shift to CCR5-only tropism and a reduction in replication capacity. ART, even if suboptimal, can be of interest in the case of MDR virus infection.

In vitro photochemical inactivation of cell-associated human T-cell leukemia virus Type I and II in human platelet concentrates and plasma by use of amotosalen.

BACKGROUND: Human T-cell leukemia virus Types I and II (HTLV-I and HTLV-II), blood-borne retroviruses found worldwide, can cause leukemia, immunosuppression, and severe neurologic diseases. In most countries, HTLV-I and -II screening is not performed systematically for blood donations. A new photochemical treatment (PCT) with a synthetic psoralen was developed to inactivate most pathogens in platelet (PLT) concentrates or plasma and to improve the safety of blood donations. STUDY DESIGN AND METHODS: Cell-associated HTLV-I or -II (10(6)/mL) was inoculated in full-size fresh PLT concentrates or fresh frozen plasma and treated with 150 micromol per L amotosalen (S-59) and different doses of long-wavelength ultraviolet A (UVA) light. The residual viral titer in the treated samples was assessed by a cocultivation assay on indicator cells. RESULTS: The inactivation obtained at a 3.0 J per cm2 UVA dose was greater than 5.2 log foci-forming units (FFUs) per mL for HTLV-I and 4.6 log FFUs per mL for HTLV-II in presence of human PLT concentrates and greater than 4.5 log FFUs per mL for HTLV-I and 5.7 log FFUs per mL for HTLV-II in the presence of human plasma. The residual infectivity was very low and shown as the limit of detection of the cocultivation assay. CONCLUSION: In human plasma or PLT concentrates, the retroviruses HTLV-I and -II were strongly sensitive to the PCT with 150 micromol per L amotosalen (S-59) and a 3.0 J per cm2 UVA dose. This high efficiency for photoinactivation of these retroviruses opens a possibility of improving the safety of PLTs or plasma transfusion in the future.

Virological, intracellular and plasma pharmacological parameters predicting response to lopinavir/ritonavir (KALEPHAR study).

OBJECTIVES: To assess the impact of HIV-1 protease mutations and intracellular and plasma lopinavir minimum concentrations (Cmin) on virological success or failure on lopinavir/ritonavir-containing highly active antiretroviral therapy (HAART). DESIGN: HIV-1-infected HAART-experienced patients included in an observational study, received lopinavir/ritonavir (400/100 mg twice a day) plus two to three nucleoside reverse transcriptase inhibitors (NRTI) or one NRTI plus one non-NRTI. A viral load less than 50 copies/ml at month 6 defined virological success. METHODS: Intracellular and plasma lopinavir concentrations were determined by high-pressure liquid chromatography with mass-spectrometry detection. Reverse transcriptase and protease genes were sequenced at baseline and the time of virological failure. RESULTS: When the 38 patients started the lopinavir/ritonavir-based regimen, baseline median (25-75th percentile) values were: CD4 cell count 218 cells/microl (133-477); plasma HIV-1-RNA load 5.3 log10 copies/ml (3.8-5.1); number of lopinavir mutations four per protease gene (two to six). Univariate analysis associated virological success or failure at month 6 (21/38 patients) with the number of baseline lopinavir mutations, intracellular and plasma lopinavir Cmin, and the genotype inhibitory quotient (GIQ) at months 1 and 6. Multivariate analysis showed that the number of baseline lopinavir mutations and intracellular and plasma lopinavir Cmin were independently associated with virological success or failure. We defined the most discriminating intracellular and plasma lopinavir Cmin efficacy thresholds (8 and 4 microg/ml, respectively) and GIQ thresholds (1 and 3, respectively). CONCLUSION: The monitoring of lopinavir/rironavir-based HAART efficacy should include the number of baseline lopinavir/ritonavir mutations, intracellular and plasma lopinavir Cmin and GIQ calculation.

Virologic response to nelfinavir-based regimens: pharmacokinetics and drug resistance mutations (VIRAPHAR study).

OBJECTIVE: To assess the impact of HIV-1 protease and reverse transcriptase (RT) mutations, and pharmacokinetic parameters on virological responses to nelfinavir (NFV)-containing highly active antiretroviral therapy. DESIGN: Naive or antiretroviral-experienced HIV-1-infected subjects were included in a non-randomized, observational cohort study and received two nucleoside RT inhibitors + NFV (750 mg three times per day or 1250 mg twice per day). Virologic success was defined as a virus load < 50 copies/ml for > 6 months. METHODS: RT and protease genes were sequenced at baseline and at the time of virological failure. Plasma NFV trough concentration (Cmin), maximum concentration (Cmax), and AUC0-tau at steady-state were subjected to population pharmacokinetic analysis. RESULTS: Patients (n = 154) enrolled between November 1998 and February 2000 started a twice per day (n = 84) or three times per day (n = 70) NFV-based regimen as first- (n = 48) or second-line therapy when protease inhibitor-naive (n = 64) or -experienced (n = 42). Median follow-up duration was 16 months. Virologic failure occurred in 88 patients. No significant differences were observed between twice per day and three times per day regimens. According to multivariate analysis, NFV Cmin and Cmax, CD4 cell count, number of baseline RT + protease gene mutations, D67N, M184V, T215F/Y in RT, and M36I in protease, were independent factors that were significantly predictive of failure. At failure, L10I, D30N, M36I, V77I, N88S/D or L90M protease mutations had emerged since baseline. Pharmacokinetic parameters were similar in patients with or without emergence of these neo-mutations. The more discriminating NFV Cmin efficacy-threshold was estimated to be 1 mg/l. CONCLUSIONS: Our data confirm the association among individual pharmacokinetic parameters, genotype pattern and virological response to NFV-containing regimens.