Thomas Klimkait

European guidelines on the clinical management of HIV-1 tropism testing

Viral tropism is the ability of viruses to enter and infect specific host cells and is based on the ability of viruses to bind to receptors on those cells. Testing for HIV tropism is recommended before prescribing a chemokine receptor blocker. In most European countries, HIV tropism is identified with tropism phenotype testing. New data support genotype analysis of the HIV third hypervariable loop (V3) for the identification of tropism. The European Consensus Group on clinical management of tropism testing was established to make recommendations to clinicians and clinical virologists. The panel recommends HIV-tropism testing for the following groups: drug-naive patients in whom toxic effects are anticipated or for whom few treatment options are available; patients who have poor tolerability to or toxic effects from current treatment or who have CNS pathology; and patients for whom therapy has failed and a change in treatment is considered. In general, an enhanced sensitivity Trofile assay and V3 population genotyping are the recommended methods. Genotypic methods are anticipated to be used more frequently in the clinical setting because of their greater accessibility, lower cost, and faster turnaround time than other methods. For the interpretation of V3 loop genotyping, clinically validated systems should be used when possible. Laboratories doing HIV tropism tests should have adequate quality assurance measures. Similarly, close collaboration between HIV clinicians and virologists is needed to ensure adequate diagnostic and treatment decisions.

Transmission of HIV-1 drug resistance in Switzerland: a 10-year molecular epidemiology survey

Objectives:Representative prevalence data of transmitted drug-resistant HIV-1 are essential to establish accurate guidelines addressing resistance testing and first-line treatments. Methods:Systematic resistance testing was carried out in individuals in Switzerland with documented HIV-1 seroconversion during 1996–2005 and available samples with RNA > 1000 copies/ml obtained within 1 year of estimated seroconversion. Resistance interpretation used the Stanford list of mutations for surveillance of transmitted drug resistance and the French National Agency for AIDS Research algorithm. Results:Viral sequences from 822 individuals were available. Risk groups were men having sex with men (42%), heterosexual contacts (32%) and intravenous drug users (20%); 30% were infected with non-B subtype viruses. Overall, prevalence of transmitted resistance was 7.7% [95% confidence interval (CI), 5.9–9.5] for any drug, 5.5% (95% CI, 3.9–7.1) for nucleoside reverse transcriptase inhibitors, 1.9% (95% CI, 1.0–2.8) for non-nucleoside reverse transcriptase inhibitors and 2.7% (95% CI, 1.6–3.8) for protease inhibitors. Dual- or triple-class resistance was observed in 2% (95% CI, 0.8–2.5). No significant trend in prevalence of transmitted resistance was observed over years. There were no differences according to ethnicity, risk groups or gender, but prevalence of transmitted resistance was highest among individuals infected with subtype B virus. Conclusions:The transmission rate of drug-resistant HIV-1 has been stable since 1996, with very rare transmission of dual- or triple-class resistance. These data suggest that transmission of drug resistance in the setting of easy access to antiretroviral treatment can remain stable and be kept at a low level.

Molecular Epidemiology Reveals Long-Term Changes in HIV Type 1 Subtype B Transmission in Switzerland

BACKGROUND Sequence data from resistance testing offer unique opportunities to characterize the structure of human immunodeficiency virus (HIV) infection epidemics. METHODS We analyzed a representative set of HIV type 1 (HIV-1) subtype B pol sequences from 5700 patients enrolled in the Swiss HIV Cohort Study. We pooled these sequences with the same number of sequences from foreign epidemics, inferred a phylogeny, and identified Swiss transmission clusters as clades having a minimal size of 10 and containing >or=80% Swiss sequences. RESULTS More than one-half of Swiss patients were included within 60 transmission clusters. Most transmission clusters were significantly dominated by specific transmission routes, which were used to identify the following patient groups: men having sex with men (MSM) (38 transmission clusters; average cluster size, 29 patients) or patients acquiring HIV through heterosexual contact (HETs) and injection drug users (IDUs) (12 transmission clusters; average cluster size, 144 patients). Interestingly, there were no transmission clusters dominated by sequences from HETs only. Although 44% of all HETs who were infected between 1983 and 1986 clustered with injection drug users, this percentage decreased to 18% for 2003-2006 (P<.001), indicating a diminishing role of injection drug users in transmission among HETs over time. CONCLUSIONS Our analysis suggests (1) the absence of a self-sustaining epidemic of HIV-1 subtype B in HETs in Switzerland and (2) a temporally decreasing clustering of HIV infections in HETs and IDUs.

Estimating the Basic Reproductive Number from Viral Sequence Data

Epidemiological processes leave a fingerprint in the pattern of genetic structure of virus populations. Here, we provide a new method to infer epidemiological parameters directly from viral sequence data. The method is based on phylogenetic analysis using a birth-death model (BDM) rather than the commonly used coalescent as the model for the epidemiological transmission of the pathogen. Using the BDM has the advantage that transmission and death rates are estimated independently and therefore enables for the first time the estimation of the basic reproductive number of the pathogen using only sequence data, without further assumptions like the average duration of infection. We apply the method to genetic data of the HIV-1 epidemic in Switzerland.

The role of compartment penetration in PI-monotherapy: the Atazanavir-Ritonavir Monomaintenance (ATARITMO) Trial.

Objectives:To limit exposure to anti-HIV drugs and minimize risk of long-term side effects, studies have looked at the possibility of simplified maintenance strategies. Ritonavir-boosted protease-inhibitor (PI)-monotherapies are an attractive alternative, but limited compartmental penetration of PI remains a concern. Design:Non-comparative 24-week pilot study. Method:Ritonavir-boosted atazanavir (ATV/r) monotherapy administered to fully suppressed patients (>3 month HIV RNA < 50 copies/ml). Plasma was obtained every 4 weeks and cerebrospinal fluid (CSF) and semen at W24. Results:Two patients (7%) failed ATV/r monotherapy. One patient was subsequently identified as a protocol violator since he had a previous history of treatment failure under indinavir. The second patient deliberately decided to stop treatment after W20. Excluding failing patients, individual measurements of HIV RNA in patients having occasional viral ‘blips’ was found in five patients. At W24, 3/20 patients had elevated viral loads in CSF (HIV RNA > 100 copies/ml), and 2/15 in semen, despite viral suppression in plasma (< 50 copies/ml). Samples with elevated HIV RNA (> 500 copies/ml) in CSF were all wild type. The mean ATV drug concentration ratio (CSF/blood, n = 22) was 0.9%. Indicators of altered immune activation (CD8CD38 C-reactive protein) remained unchanged. Conclusion:This study supports previous results indicating the potential use of PI-based mono-maintenance therapies. However, our results in CSF cautions against the uncontrolled use of PI-based monotherapies.

Ambiguous Nucleotide Calls From Population-based Sequencing of HIV-1 are a Marker for Viral Diversity and the Age of Infection

The fraction of ambiguous nucleotide calls in bulk sequencing of human immunodeficiency virus type 1 (HIV-1) carries important information on viral diversity and the age of infection. In particular, a fraction of ambiguous nucleotides of >.5% provides evidence against a recent infection event <1 year ago.

Phylogenetic approach reveals that virus genotype largely determines HIV set-point viral load.

HIV virulence, i.e. the time of progression to AIDS, varies greatly among patients. As for other rapidly evolving pathogens of humans, it is difficult to know if this variance is controlled by the genotype of the host or that of the virus because the transmission chain is usually unknown. We apply the phylogenetic comparative approach (PCA) to estimate the heritability of a trait from one infection to the next, which indicates the control of the virus genotype over this trait. The idea is to use viral RNA sequences obtained from patients infected by HIV-1 subtype B to build a phylogeny, which approximately reflects the transmission chain. Heritability is measured statistically as the propensity for patients close in the phylogeny to exhibit similar infection trait values. The approach reveals that up to half of the variance in set-point viral load, a trait associated with virulence, can be heritable. Our estimate is significant and robust to noise in the phylogeny. We also check for the consistency of our approach by showing that a trait related to drug resistance is almost entirely heritable. Finally, we show the importance of taking into account the transmission chain when estimating correlations between infection traits. The fact that HIV virulence is, at least partially, heritable from one infection to the next has clinical and epidemiological implications. The difference between earlier studies and ours comes from the quality of our dataset and from the power of the PCA, which can be applied to large datasets and accounts for within-host evolution. The PCA opens new perspectives for approaches linking clinical data and evolutionary biology because it can be extended to study other traits or other infectious diseases.

A genome-to-genome analysis of associations between human genetic variation, HIV-1 sequence diversity, and viral control

HIV-1 sequence diversity is affected by selection pressures arising from host genomic factors. Using paired human and viral data from 1071 individuals, we ran >3000 genome-wide scans, testing for associations between host DNA polymorphisms, HIV-1 sequence variation and plasma viral load (VL), while considering human and viral population structure. We observed significant human SNP associations to a total of 48 HIV-1 amino acid variants (p<2.4 × 10−12). All associated SNPs mapped to the HLA class I region. Clinical relevance of host and pathogen variation was assessed using VL results. We identified two critical advantages to the use of viral variation for identifying host factors: (1) association signals are much stronger for HIV-1 sequence variants than VL, reflecting the ‘intermediate phenotype’ nature of viral variation; (2) association testing can be run without any clinical data. The proposed genome-to-genome approach highlights sites of genomic conflict and is a strategy generally applicable to studies of host–pathogen interaction. DOI: http://dx.doi.org/10.7554/eLife.01123.001

Enhanced gene silencing by the application of multiple specific small interfering RNAs

Small interfering RNA duplexes (siRNA) induce gene silencing in various eukaryotic cells, although usually in an incomplete manner. Using chemically synthesized siRNAs targeting the HIV‐1 co‐receptor CXCR4 or the apoptosis‐inducing Fas‐ligand (FasL), co‐transfection of cells with two or more siRNA duplexes targeting different sites on the same mRNA resulted in an enhanced gene silencing compared with each single siRNA. This was shown in the down‐regulation of protein and mRNA expression, and functionally in the inhibition of CXCR4‐mediated HIV infection and of FasL‐mediated cell apoptosis. Transfection efficiency determined for the FasL‐specific siRNAs was dose‐dependent and varied among the siRNAs tested, but was not the main reason for the enhanced gene silencing.

Association of a pool of HIV-1 with erythrocytes in vivo: a cohort study

BACKGROUND Treatment of HIV-1-infected individuals with antiretrovirals can result in sustained suppression of plasma viral RNA at concentrations below the detection limit of available assays. However, continuing virus replication has been detected in patients with viral RNA in plasma suppressed for months to years, and many cell types are known to act as reservoirs or carriers for the virus. In vitro, erythrocytes bind HIV-1 immune complexes, so we tested for a circulating pool of HIV-1 associated with erythrocytes in people with HIV-1 infection. METHODS We investigated 82 chronically HIV-1-infected individuals. Plasma, white cells, and erythrocytes were tested for HIV-1 RNA by RT-PCR. FINDINGS Erythrocyte-associated HIV-1 RNA was detected in 80 of 82 individuals. In 23, plasma HIV-1 RNA had been undetectable (<20 copies/mL) for up to 32 months; in corresponding erythrocyte samples, there were up to 82878 HIV-1 RNA copies per mL whole blood. HIV-1 associated with erythrocytes in vivo was shown to be infectious. Within the subgroup of patients with undetectable plasma viral load, higher numbers of HIV-1 associated with erythrocytes were correlated with a history of advanced clinical stages of HIV-1 infection (p=0.014). INTERPRETATION A pool of HIV-1 is associated with erythrocytes even after long-term suppression of viral RNA in plasma. This finding is direct evidence for continuing virus replication or release in these individuals. Quantification of this viral pool may help to judge suppression of HIV-1 replication in individuals with undetectable plasma HIV-1 RNA.