The catalytically active 66-kDa subunit of the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) consists of DNA polymerase, connection, and ribonuclease H (RNase H) domains. ...Almost all known RT inhibitor resistance mutations identified to date map to the polymerase domain of the enzyme. However, the connection and RNase H domains are not routinely analysed in clinical samples and none of the genotyping assays available for patient management sequence the entire RT coding region. The British Columbia Centre for Excellence in HIV/AIDS (the Centre) genotypes clinical isolates up to codon 400 in RT, and our retrospective statistical analyses of the Centre's database have identified an N348I mutation in the RT connection domain in treatment-experienced individuals. The objective of this multidisciplinary study was to establish the in vivo relevance of this mutation and its role in drug resistance.
The prevalence of N348I in clinical isolates, the time taken for it to emerge under selective drug pressure, and its association with changes in viral load, specific drug treatment, and known drug resistance mutations was analysed from genotypes, viral loads, and treatment histories from the Centre's database. N348I increased in prevalence from below 1% in 368 treatment-naïve individuals to 12.1% in 1,009 treatment-experienced patients (p = 7.7 x 10(-12)). N348I appeared early in therapy and was highly associated with thymidine analogue mutations (TAMs) M41L and T215Y/F (p < 0.001), the lamivudine resistance mutations M184V/I (p < 0.001), and non-nucleoside RTI (NNRTI) resistance mutations K103N and Y181C/I (p < 0.001). The association with TAMs and NNRTI resistance mutations was consistent with the selection of N348I in patients treated with regimens that included both zidovudine and nevirapine (odds ratio 2.62, 95% confidence interval 1.43-4.81). The appearance of N348I was associated with a significant increase in viral load (p < 0.001), which was as large as the viral load increases observed for any of the TAMs. However, this analysis did not account for the simultaneous selection of other RT or protease inhibitor resistance mutations on viral load. To delineate the role of this mutation in RT inhibitor resistance, N348I was introduced into HIV-1 molecular clones containing different genetic backbones. N348I decreased zidovudine susceptibility 2- to 4-fold in the context of wild-type HIV-1 or when combined with TAMs. N348I also decreased susceptibility to nevirapine (7.4-fold) and efavirenz (2.5-fold) and significantly potentiated resistance to these drugs when combined with K103N. Biochemical analyses of recombinant RT containing N348I provide supporting evidence for the role of this mutation in zidovudine and NNRTI resistance and give some insight into the molecular mechanism of resistance.
This study provides the first in vivo evidence that treatment with RT inhibitors can select a mutation (i.e., N348I) outside the polymerase domain of the HIV-1 RT that confers dual-class resistance. Its emergence, which can happen early during therapy, may significantly impact on a patient's response to antiretroviral therapies containing zidovudine and nevirapine. This study also provides compelling evidence for investigating the role of other mutations in the connection and RNase H domains in virological failure.
Despite the extensive genetic diversity of HIV-1, viral evolution in response to immune selective pressures follows broadly predictable mutational patterns. Sites and pathways of Human ...Leukocyte-Antigen (HLA)-associated polymorphisms in HIV-1 have been identified through the analysis of population-level data, but the full extent of immune escape pathways remains incompletely characterized. Here, in the largest analysis of HIV-1 subtype B sequences undertaken to date, we identify HLA-associated polymorphisms in the three HIV-1 proteins most commonly considered in cellular-based vaccine strategies. Results are organized into protein-wide escape maps illustrating the sites and pathways of HLA-driven viral evolution.
HLA-associated polymorphisms were identified in HIV-1 Gag, Pol and Nef in a multicenter cohort of >1500 chronically subtype-B infected, treatment-naïve individuals from established cohorts in Canada, the USA and Western Australia. At q< or =0.05, 282 codons commonly mutating under HLA-associated immune pressures were identified in these three proteins. The greatest density of associations was observed in Nef (where close to 40% of codons exhibited a significant HLA association), followed by Gag then Pol (where approximately 15-20% of codons exhibited HLA associations), confirming the extensive impact of immune selection on HIV evolution and diversity. Analysis of HIV codon covariation patterns identified over 2000 codon-codon interactions at q< or =0.05, illustrating the dense and complex networks of linked escape and secondary/compensatory mutations.
The immune escape maps and associated data are intended to serve as a user-friendly guide to the locations of common escape mutations and covarying codons in HIV-1 subtype B, and as a resource facilitating the systematic identification and classification of immune escape mutations. These resources should facilitate research in HIV epitope discovery and host-pathogen co-evolution, and are relevant to the continued search for an effective CTL-based AIDS vaccine.
High-quality, simplified, and low-cost human immunodeficiency virus (HIV) drug resistance tests that are able to provide timely actionable HIV resistance data at individual, population, and ...programmatic levels are needed to confront the emerging drug-resistant HIV epidemic. Next-generation sequencing technologies embedded in automated cloud-computing analysis environments are ideally suited for such endeavor. Whereas NGS can reduce costs over Sanger sequencing, automated analysis pipelines make NGS accessible to molecular laboratories regardless of the available bioinformatic skills. They can also produce highly structured, high-quality data that could be examined by healthcare officials and program managers on a real-time basis to allow timely public health action. Here we discuss the opportunities and challenges of such an approach.
Bictegravir (BIC) and cabotegravir (CAB) are the latest available HIV integrase inhibitors in clinical trials. The combination of major integrase inhibitor substitutions G140S/Q148H has been shown to ...confer high-level resistance to the approved integrase inhibitors raltegravir (RAL) and elvitegravir (EVG) but not necessarily dolutegravir (DTG). We assayed recombinant viruses made from patient-derived RNA extracts for resistance phenotype for a panel of viruses containing G140S/Q148H with additional accessory substitutions. The accumulation of multiple integrase substitutions confers high-level resistance to all 5 integrase inhibitors. There is extensive cross-resistance between DTG, BIC, and CAB (r = 0.96-0.97).
Background There have been limited studies evaluating temporal changes in the incidence of detection of drug resistance among human immunodeficiency virus type 1 (HIV-1) isolates and concomitant ...changes in plasma HIV load for treated individuals in a population-wide setting. Methods Longitudinal plasma viral load and genotypic resistance data were obtained from patients receiving antiretroviral therapy from the British Columbia Drug Treatment Program from July 1996 through December 2008. A total of 24,652 resistance tests were available from 5422 individuals. The incidence of successful plasma viral load suppression and of resistance to each of 3 antiretroviral categories (nucleoside/nucleotide reverse-transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors, and protease inhibitors) was calculated for the population receiving therapy. Results There has been a drastic decrease in the incidence of new cases of HIV-1 drug resistance in individuals followed during 1996–2008. In 1997, the incidence rate of any newly detected resistance was 1.73 cases per 100 person-months of therapy, and by 2008, the incidence rate had decreased >12-fold, to 0.13 cases per 100 person-months of therapy. This decrease in the incidence of resistance has occurred at an exponential rate, with halftimes on the order of 2–3 years. Concomitantly, the proportion of individuals with plasma viral load suppression has increased linearly over time (from 64.7% with HIV RNA levels <50 copies/mL in 2000 to 87.0% in 2008; R2 p 0.97; P < .001). Conclusions Our results suggest an increasing effectiveness of highly active antiretroviral therapy at the populational level. The vast majority of treated patients in British Columbia now have either suppressed plasma viral load or drug-susceptible HIV-1, according to their most recent test results.
HIV-1 genotypic resistance test (GRT) interpretation systems (IS) require updates as new studies on HIV-1 drug resistance are published and as treatment guidelines evolve.
An expert panel was created ...to provide recommendations for the update of the Stanford HIV Drug Resistance Database (HIVDB) GRT-IS. The panel was polled on the ARVs to be included in a GRT report, and the drug-resistance interpretations associated with 160 drug-resistance mutation (DRM) pattern-ARV combinations. The DRM pattern-ARV combinations included 52 nucleoside RT inhibitor (NRTI) DRM pattern-ARV combinations (13 patterns x 4 NRTIs), 27 nonnucleoside RT inhibitor (NNRTI) DRM pattern-ARV combinations (9 patterns x 3 NNRTIs), 39 protease inhibitor (PI) DRM pattern-ARV combinations (13 patterns x 3 PIs) and 42 integrase strand transfer inhibitor (INSTI) DRM pattern-ARV combinations (14 patterns x 3 INSTIs).
There was universal agreement that a GRT report should include the NRTIs lamivudine, abacavir, zidovudine, emtricitabine, and tenofovir disoproxil fumarate; the NNRTIs efavirenz, etravirine, nevirapine, and rilpivirine; the PIs atazanavir/r, darunavir/r, and lopinavir/r (with "/r" indicating pharmacological boosting with ritonavir or cobicistat); and the INSTIs dolutegravir, elvitegravir, and raltegravir. There was a range of opinion as to whether the NRTIs stavudine and didanosine and the PIs nelfinavir, indinavir/r, saquinavir/r, fosamprenavir/r, and tipranavir/r should be included. The expert panel members provided highly concordant DRM pattern-ARV interpretations with only 6% of NRTI, 6% of NNRTI, 5% of PI, and 3% of INSTI individual expert interpretations differing from the expert panel median by more than one resistance level. The expert panel median differed from the HIVDB 7.0 GRT-IS for 20 (12.5%) of the 160 DRM pattern-ARV combinations including 12 NRTI, two NNRTI, and six INSTI pattern-ARV combinations. Eighteen of these differences were updated in HIVDB 8.1 GRT-IS to reflect the expert panel median. Additionally, HIVDB users are now provided with the option to exclude those ARVs not considered to be universally required.
The HIVDB GRT-IS was updated through a collaborative process to reflect changes in HIV drug resistance knowledge, treatment guidelines, and expert opinion. Such a process broadens consensus among experts and identifies areas requiring further study.
A phylogeny is a tree-based model of common ancestry that is an indispensable tool for studying biological variation. Phylogenies play a special role in the study of rapidly evolving populations such ...as viruses, where the proliferation of lineages is constantly being shaped by the mode of virus transmission, by adaptation to immune systems, and by patterns of human migration and contact. These processes may leave an imprint on the shapes of virus phylogenies that can be extracted for comparative study; however, tree shapes are intrinsically difficult to quantify. Here we present a comprehensive study of phylogenies reconstructed from 38 different RNA viruses from 12 taxonomic families that are associated with human pathologies. To accomplish this, we have developed a new procedure for studying phylogenetic tree shapes based on the 'kernel trick', a technique that maps complex objects into a statistically convenient space. We show that our kernel method outperforms nine different tree balance statistics at correctly classifying phylogenies that were simulated under different evolutionary scenarios. Using the kernel method, we observe patterns in the distribution of RNA virus phylogenies in this space that reflect modes of transmission and pathogenesis. For example, viruses that can establish persistent chronic infections (such as HIV and hepatitis C virus) form a distinct cluster. Although the visibly 'star-like' shape characteristic of trees from these viruses has been well-documented, we show that established methods for quantifying tree shape fail to distinguish these trees from those of other viruses. The kernel approach presented here potentially represents an important new tool for characterizing the evolution and epidemiology of RNA viruses.
"The upshot of this widespread failure to recognize that AIDS is an exceptional crisis and threat is that the response to the pandemic is not made commensurate to the challenges-and so the epidemic ...escalates even while it erodes our capacities to check it."
PDF
