There are few data on the persistence of individual human immunodeficiency virus type 1 (HIV-1) transmitted drug resistance (TDR) mutations in the absence of selective drug pressure. We studied 313 patients in whom TDR mutations were detected at their first resistance test and who had a subsequent test performed while ART-naive. The rate at which mutations became undetectable was estimated using exponential regression accounting for interval censoring. Most thymidine analogue mutations (TAMs) and T215 revertants (but not T215F/Y) were found to be highly stable, with NNRTI and PI mutations being relatively less persistent. Our estimates are important for informing HIV transmission models.
persistence; transmitted; HIV-1; resistance; mutations
To determine protease mutations that develop at viral failure for protease inhibitor (PI)-naive patients on a regimen containing the PI atazanavir.
Resistance tests on patients failing atazanavir, conducted as part of routine clinical care in a multicentre observational study, were randomly matched by subtype to resistance tests from PI-naive controls to account for natural polymorphisms. Mutations from the consensus B sequence across the protease region were analysed for association and defined using the IAS-USA 2011 classification list.
Four hundred and five of 2528 (16%) patients failed therapy containing atazanavir as a first PI over a median (IQR) follow-up of 1.76 (0.84–3.15) years and 322 resistance tests were available for analysis. Recognized major atazanavir mutations were found in six atazanavir-experienced patients (P < 0.001), including I50L and N88S. The minor mutations most strongly associated with atazanavir experience were M36I, M46I, F53L, A71V, V82T and I85V (P < 0.05). Multiple novel mutations, I15S, L19T, K43T, L63P/V, K70Q, V77I and L89I/T/V, were also associated with atazanavir experience.
Viral failure on atazanavir-containing regimens was not common and major resistance mutations were rare, suggesting that adherence may be a major contributor to viral failure. Novel mutations were described that have not been previously documented.
HIV; drug resistance mutations; naive patients; protease inhibitors; virological failure
In contrast to resource-rich countries, most HIV-infected patients in resource-limited countries receive treatment without virological monitoring. There are few long-term data, in this setting, on rates of viral suppression or switch to second-line antiretroviral therapy. The DART trial compared clinically driven monitoring (CDM) versus routine laboratory (CD4/haematology/biochemistry) and clinical monitoring (LCM) in HIV-infected adults initiating therapy. There was no virological monitoring in either study group during follow-up, but viral load was measured in Ugandan participants at trial closure. Two thousand three hundred and seventeen (2317) participants from this country initiated antiretroviral therapy with zidovudine/lamivudine plus tenofovir (n = 1717), abacavir (n = 300), or nevirapine (n = 300). Of 1896 (81.8%) participants who were alive and in follow-up at trial closure (median 5.1 years after therapy initiation), 1507 (79.5%) were on first-line and 389 (20.5%) on second-line antiretroviral therapy. The overall switch rate after the first year was 5.6 per 100 person-years; the rate was substantially higher in participants with low baseline CD4 counts (<50 cells/mm3). Among 1207 (80.1%) first-line participants with viral load measured, HIV RNA was <400 copies/ml in 963 (79.8%), 400–999 copies/ml in 37 (3.1%), 1,000–9,999 copies/ml in 110 (9.1%), and ≥10,000 copies/ml in 97 (8.0%). The proportion with HIV RNA <400 copies/ml was slightly lower (difference 7.1%, 95% CI 2.5 to 11.5%) in CDM (76.3%) than in LCM (83.4%). Among 252 (64.8%) second-line participants with viral load measured (median 2.3 years after switch), HIV RNA was <400 copies/ml in 226 (89.7%), with no difference between monitoring strategies. Low switch rates and high, sustained levels of viral suppression are achievable without viral load or CD4 count monitoring in the context of high-quality clinical care.
The Human Immunodeficiency Virus type-1 (HIV-1) spreads by cell-free diffusion and by direct cell-to-cell transfer, the latter being a significantly more efficient mode of transmission. Recently it has been suggested that cell-to-cell spread may permit ongoing virus replication in the presence of antiretroviral therapy (ART) based on studies performed using Reverse Transcriptase Inhibitors (RTIs). Protease Inhibitors (PIs) constitute an important component of ART; however whether this class of inhibitors can suppress cell-to-cell transfer of HIV-1 is unexplored. Here we have evaluated the inhibitory effect of PIs during cell-to-cell spread of HIV-1 between T lymphocytes.
Using quantitative assays in cell line and primary cell systems that directly measure the early steps of HIV-1 infection we find that the PIs Lopinavir and Darunavir are equally potent against both cell-free and cell-to-cell spread of HIV-1. We further show that a protease resistant mutant maintains its resistant phenotype during cell-to-cell spread and is transmitted more efficiently than wild-type virus in the presence of drug. By contrast we find that T cell-T cell spread of HIV-1 is 4–20 fold more resistant to inhibition by the RTIs Nevirapine, Zidovudine and Tenofovir. Notably, varying the ratio of infected and uninfected cells in co-culture impacted on the degree of inhibition, indicating that the relative efficacy of ART is dependent on the multiplicity of infection.
We conclude that if the variable effects of antiviral drugs on cell-to-cell virus dissemination of HIV-1 do indeed impact on viral replication and maintenance of viral reservoirs this is likely to be influenced by the antiviral drug class, since PIs appear particularly effective against both modes of HIV-1 spread.
HIV-1; Virological synapse; Cell-cell spread; Protease inhibitor; ART
In a randomized comparison of nevirapine or abacavir with zidovudine plus lamivudine, routine viral load monitoring was not performed, yet 27% of individuals with viral failure at week 48 experienced resuppression by week 96 without switching. This supports World Health Organization recommendations that suspected viral failure should trigger adherence counseling and repeat measurement before a treatment switch is considered.
HIV; failure; viral resuppression; resistance; Africa
To evaluate the immunological and viral consequences of planned treatment interruptions (PTI) in children with HIV.
This was an immunological and virological sub-study of the Paediatric European Network for Treatment of AIDS (PENTA) 11 trial, which compared CD4-guided PTI of antiretroviral therapy (ART) with continuous therapy (CT) in children.
HIV-1 RNA and lymphocyte subsets, including CD4 and CD8 cells, were quantified on fresh samples collected during the study; CD45RA, CD45RO and CD31 subpopulations were evaluated in some centres. For 36 (18 PTI, 18 CT) children, immunophenotyping was performed and cell-associated HIV-1 DNA analysed on stored samples to 48 weeks.
In the PTI group, CD4 cell count fell rapidly in the first 12 weeks off ART, with decreases in both naïve and memory cells. However, the proportion of CD4 cells expressing CD45RA and CD45RO remained constant in both groups. The increase in CD8 cells in the first 12 weeks off ART in the PTI group was predominantly due to increases in RO-expressing cells. PTI was associated with a rapid and sustained increase in CD4 cells expressing Ki67 and HLA-DR, and increased levels of HIV-1 DNA.
PTI in children is associated with rapid changes in CD4 and CD8 cells, likely due to increased cell turnover and immune activation. However, children off treatment may be able to maintain stable levels of naïve CD4 cells, at least in proportion to the memory cell pool, which may in part explain the observed excellent CD4 cell recovery with re-introduction of ART.
Despite prevention efforts new HIV diagnoses continue in the Southern US, where the epidemic is characterized by significant racial/ethnic disparities. We integrated phylogenetic analyses with clinical data to reveal trends in local HIV transmission.
Cross-sectional analysis of 1671 HIV-infected individuals each with one B-subtype pol sequence obtained during chronic (82%; UNC Center for AIDS Research Clinical Cohort) or acute/recent (18%; Duke/UNC Acute HIV Consortium) infection.
Phylogenies were inferred using neighbor joining to select related sequences then confirmed with Bayesian methods. We characterized transmission clusters (clades n≥3 sequences supported by posterior probabilities=1) by factors including race/ethnicity and transmission risk. Factors associated with cluster membership were evaluated for newly diagnosed patients.
Overall, 72% were male, 59% black and 39% MSM. A total of 557 (33%) sequences grouped in either 108 pairs (n=216) or 67 clusters (n=341). Clusters ranged from 3–36 (median 4) members. Composition was delineated primarily by race, with 28% exclusively black, and to a lesser extent by risk group. Both MSM and heterosexuals formed discrete clusters though substantial mixing was observed. In multivariable analysis, patients with age ≤30 years (P=0.009), acute infection (P=0.02), local residence (P=0.002), and transmitted drug resistance (P=0.02) were more likely to be cluster members while Latinos were less likely (P<0.001).
Integration of molecular, clinical and demographic data offers a unique view into the structure of local transmission networks. Clustering by black race, youth and TDR and inability to identify Latino clusters will inform prevention, testing and linkage to care strategies.
Molecular epidemiology; HIV transmission; Risk Factors; Acute Infection; Southeast US
In the late 1980s an HIV-1 epidemic emerged in Romania that was dominated by subtype F1. The main route of infection is believed to be parenteral transmission in children. We sequenced partial pol coding regions of 70 subtype F1 samples from children and adolescents from the PENTA-EPPICC network of which 67 were from Romania. Phylogenetic reconstruction using the sequences and other publically available global subtype F sequences showed that 79% of Romanian F1 sequences formed a statistically robust monophyletic cluster. The monophyletic cluster was epidemiologically linked to parenteral transmission in children. Coalescent-based analysis dated the origins of the parenteral epidemic to 1983 [1981–1987; 95% HPD]. The analysis also shows that the epidemic's effective population size has remained fairly constant since the early 1990s suggesting limited onward spread of the virus within the population. Furthermore, phylogeographic analysis suggests that the root location of the parenteral epidemic was Bucharest.
Hepatitis B coinfection is common in HIV-positive individuals and as antiretroviral therapy has made death due to AIDS less common, hepatitis has become increasingly important. Several drugs are available to treat hepatitis B. The most potent and the one with the lowest risk of resistance appears to be tenofovir (TDF). However there are several questions that remain unanswered regarding the use of TDF, including the proportion of patients that achieves suppression of HBV viral load and over what time, whether suppression is durable and whether prior treatment with other HBV-active drugs such as lamivudine, compromises the efficacy of TDF due to possible selection of resistant HBV strains.
A systematic review and meta-analysis following PRISMA guidelines and using multilevel mixed effects logistic regression, stratified by prior and/or concomitant use of lamivudine and/or emtricitabine.
Data was available from 23 studies including 550 HBV/HIV coinfected patients treated with TDF. Follow up was for up to seven years but to ensure sufficient power the data analyses were limited to three years. The overall proportion achieving suppression of HBV replication was 57.4%, 79.0% and 85.6% at one, two and three years, respectively. No effect of prior or concomitant 3TC/FTC was shown. Virological rebound on TDF treatment was rare.
TDF suppresses HBV to undetectable levels in the majority of HBV/HIV coinfected patients with the proportion fully suppressed continuing to increase during continuous treatment. Prior treatment with 3TC/FTC does not compromise efficacy of TDF treatment. The use of combination treatment with 3TC/FTC offers no significant benefit over TDF alone.
Whole HIV-1 genome sequences are pivotal for large-scale studies of inter- and intrahost evolution, including the acquisition of drug resistance mutations. The ability to rapidly and cost-effectively generate large numbers of HIV-1 genome sequences from different populations and geographical locations and determine the effect of minority genetic variants is, however, a limiting factor. Next-generation sequencing promises to bridge this gap but is hindered by the lack of methods for the enrichment of virus genomes across the phylogenetic breadth of HIV-1 and methods for the robust assembly of the virus genomes from short-read data. Here we report a method for the amplification, next-generation sequencing, and unbiased de novo assembly of HIV-1 genomes of groups M, N, and O, as well as recombinants, that does not require prior knowledge of the sequence or subtype. A sensitivity of at least 3,000 copies/ml was determined by using plasma virus samples of known copy numbers. We applied our novel method to compare the genome diversities of HIV-1 groups, subtypes, and genes. The highest level of diversity was found in the env, nef, vpr, tat, and rev genes and parts of the gag gene. Furthermore, we used our method to investigate mutations associated with HIV-1 drug resistance in clinical samples at the level of the complete genome. Drug resistance mutations were detected as both major variant and minor species. In conclusion, we demonstrate the feasibility of our method for large-scale HIV-1 genome sequencing. This will enable the phylogenetic and phylodynamic resolution of the ongoing pandemic and efficient monitoring of complex HIV-1 drug resistance genotypes.
A novel betacoronavirus associated with lethal respiratory and renal complications was recently identified in patients from several countries in the Middle East. We report the deep genome sequencing of the virus directly from a patient’s sputum sample. Our high-throughput sequencing yielded a substantial depth of genome sequence assembly and showed the minority viral variants in the specimen. Detailed phylogenetic analysis of the virus genome (England/Qatar/2012) revealed its close relationship to European bat coronaviruses circulating among the bat species of the Vespertilionidae family. Molecular clock analysis showed that the 2 human infections of this betacoronavirus in June 2012 (EMC/2012) and September 2012 (England/Qatar/2012) share a common virus ancestor most likely considerably before early 2012, suggesting the human diversity is the result of multiple zoonotic events.
disease transmission; infectious; coronavirus infections; evolution; molecular; respiratory tract infections; high-throughput nucleotide sequencing; computer-aided design; viruses; zoonoses
The extent of HIV-1 diversity was examined among patients attending a rural district hospital in a coastal area of Kenya. The pol gene was sequenced in samples from 153 patients. Subtypes were designated using the REGA, SCUEAL, and jpHMM programs. The most common subtype was A1, followed by C and D; A2 and G were also detected. However, a large proportion of the samples was found to be recombinants, which clustered within the pure subtype branches. Phylogeographic analysis of Kilifi sequences compared with those from other regions of Africa showed that while many sequences were closely related to sequences from Kenya, others were most closely related to known sequences from other parts of Africa, including West Africa. Overall, these data indicate that there have been multiple introductions of HIV-1 into this small rural town and surroundings with ongoing diversity being generated by recombination.
To describe predictors of pregnancy and changes in pregnancy incidence among HIV-positive women accessing HIV clinical care.
Data were obtained through the linkage of two separate studies; the UK Collaborative HIV Cohort study (UK CHIC), a cohort of adults attending 13 large HIV clinics, and the National Study of HIV in Pregnancy and Childhood (NSHPC), a national surveillance study of HIV-positive pregnant women. Pregnancy incidence was measured using the proportion of women in UK CHIC with a pregnancy reported to NSHPC. Generalised estimating equations were used to identify predictors of pregnancy and assess changes in pregnancy incidence in 2000-2009.
The number of women accessing care at UK CHIC sites increased as did the number of pregnancies (from 72 to 230). Older women were less likely to have a pregnancy (adjusted Relative Rate (aRR) 0.44 per 10 year increment in age [95% CI [0.41-0.46], p<0.001) as were women with CD4<200 cells/mm3 compared with CD4 200-350 cells/mm3 (aRR 0.65 [0.55-0.77] p<0.001) and women of white ethnicity compared with women of black-African ethnicity (aRR 0.67 [0.57-0.80], p<0.001). The likelihood that women had a pregnancy increased over the study period (aRR 1.05 [1.03-1.07], p<0.001). The rate of change did not significantly differ according to age group, ART use, CD4 group or ethnicity.
The pregnancy rate among women accessing HIV clinical care increased in 2000-2009. HIV-positive women with, or planning, a pregnancy require a high level of care and this is likely to continue and increase as more women of older age have pregnancies.
HIV; pregnancy; pregnancy rate; maternal age; highly active antiretroviral therapy; maternal-fetal infection transmission; United Kingdom
Hepatitis B virus (HBV) infection is an increasingly important cause of morbidity and mortality in HIV-infected adults. This study aimed to determine the prevalence and incidence of HBV in the UK CHIC Study, a multicentre observational cohort.
Methods and Findings
12 HIV treatment centres were included. Of 37,331 patients, 27,450 had at least one test (HBsAg, anti-HBs or anti-HBc) result post-1996 available. 16,043 were white, 8,130 black and 3,277 other ethnicity. Route of exposure was homosexual sex 15,223 males, heterosexual sex 3,258 males and 5,384 females, injecting drug use 862 and other 2,723. The main outcome measures used were the cumulative prevalence and the incidence of HBV coinfection. HBV susceptible patients were followed up until HBsAg and/or anti-HBc seroconversion incident infection, evidence of vaccination or last visit. Poisson regression was used to determine associated factors. 25,973 had at least one HBsAg test result. Participants with HBsAg results were typically MSM (57%) and white (59%) (similar to the cohort as a whole). The cumulative prevalence of detectable HBsAg was 6.9% (6.6 to 7.2%). Among the 3,379 initially HBV-susceptible patients, the incidence of HBV infection was 1.7 (1.5 to 1.9)/100 person-years. Factors associated with incident infection were older age and IDU. The main limitation of the study was that 30% of participants did not have any HBsAg results available. However baseline characteristics of those with results did not differ from those of the whole cohort. Efforts are on-going to improve data collection.
The prevalence of HBV in UK CHIC is in line with estimates from other studies and low by international standards. Incident infection continued to occur even after entry to the cohort, emphasising the need to ensure early vaccination.
The emergence and spread of high levels of HIV-1 drug resistance in resource-limited settings where combination antiretroviral treatment has been scaled up could compromise the effectiveness of national HIV treatment programmes. We aimed to estimate changes in the prevalence of HIV-1 drug resistance in treatment-naive individuals with HIV since initiation of rollout in resource-limited settings.
We did a systematic search for studies and conference abstracts published between January, 2001, and July, 2011, and included additional data from the WHO HIV drug resistance surveillance programme. We assessed the prevalence of drug-resistance mutations in untreated individuals with respect to time since rollout in a series of random-effects meta-regression models.
Study-level data were available for 26 102 patients from sub-Saharan Africa, Asia, and Latin America. We recorded no difference between chronic and recent infection on the prevalence of one or more drug-resistance mutations for any region. East Africa had the highest estimated rate of increase at 29% per year (95% CI 15 to 45; p=0·0001) since rollout, with an estimated prevalence of HIV-1 drug resistance at 8 years after rollout of 7·4% (4·3 to 12·7). We recorded an annual increase of 14% (0% to 29%; p=0·054) in southern Africa and a non-significant increase of 3% (–0·9 to 16; p=0·618) in west and central Africa. There was no change in resistance over time in Latin America, and because of much country-level heterogeneity the meta-regression analysis was not appropriate for Asia. With respect to class of antiretroviral, there were substantial increases in resistance to non-nucleoside reverse transcriptase inhibitors (NNRTI) in east Africa (36% per year [21 to 52]; p<0·0001) and southern Africa (23% per year [7 to 42]; p=0·0049). No increase was noted for the other drug classes in any region.
Our findings suggest a significant increase in prevalence of drug resistance over time since antiretroviral rollout in regions of sub-Saharan Africa; this rise is driven by NNRTI resistance in studies from east and southern Africa. The findings are of concern and draw attention to the need for enhanced surveillance and drug-resistance prevention efforts by national HIV treatment programmes. Nevertheless, estimated levels, although increasing, are not unexpected in view of the large expansion of antiretroviral treatment coverage seen in low-income and middle-income countries—no changes in antiretroviral treatment guidelines are warranted at the moment.
Bill & Melinda Gates Foundation and the European Community's Seventh Framework Programme
To describe antiretroviral therapy (ART) use and clinical status, at start of and during pregnancy, for HIV-positive women receiving ART at conception, including the proportion conceiving on drugs (efavirenz and didanosine) not recommended for use in early pregnancy.
Women with a pregnancy resulting in a live birth after 1995 (n=1,537) were identified in an observational cohort of patients receiving HIV care at 12 clinics in the UK by matching records with national pregnancy study data. Treatment and clinical data were analysed for 375 women conceiving on ART, including logistic regression to identify factors associated with changing regimen during pregnancy.
Of the 375 women on ART at conception, 39 (10%) conceived on dual therapy, 306 (82%) on triple therapy and 30 (8%) on >3 drugs. In total, 116 (31%) women conceived on a regimen containing efavirenz or didanosine (69 efavirenz, 54 didanosine, 7 both). Overall, 38% (143) switched regimen during pregnancy, of whom 41% (n=48) had a detectable viral load (≥50 copies/ml) around that time. Detectable viral load was associated with increased risk of regimen change (adjusted odds ratio 2.2, 95% confidence interval [1.3, 3.8]), while women on efavirenz at conception were three times more likely to switch than women on other drugs (3.3, [1.8, 6.0]). Regimen switching was also associated with calendar year at conception (0.9, [0.8-1.0]).
These findings reinforce the need for careful consideration of ART use among women planning or likely to have a pregnancy in order to reduce viral load before pregnancy and avoid drugs not recommended for early antenatal use.
HIV; pregnancy; antiretroviral agents; antiretroviral therapy; United Kingdom
The most recent World Health Organization (WHO) antiretroviral treatment guidelines recommend the inclusion of zidovudine (ZDV) or tenofovir (TDF) in first-line therapy. We conducted a cost-effectiveness analysis with emphasis on emerging patterns of drug resistance upon treatment failure and their impact on second-line therapy.
We used a stochastic simulation of a generalized HIV-1 epidemic in sub-Saharan Africa to compare two strategies for first-line combination antiretroviral treatment including lamivudine, nevirapine and either ZDV or TDF. Model input parameters were derived from literature and, for the simulation of resistance pathways, estimated from drug resistance data obtained after first-line treatment failure in settings without virological monitoring. Treatment failure and cost effectiveness were determined based on WHO definitions. Two scenarios with optimistic (no emergence; base) and pessimistic (extensive emergence) assumptions regarding occurrence of multidrug resistance patterns were tested.
In the base scenario, cumulative proportions of treatment failure according to WHO criteria were higher among first-line ZDV users (median after six years 36% [95% simulation interval 32%; 39%]) compared with first-line TDF users (31% [29%; 33%]). Consequently, a higher proportion initiated second-line therapy (including lamivudine, boosted protease inhibitors and either ZDV or TDF) in the first-line ZDV user group 34% [31%; 37%] relative to first-line TDF users (30% [27%; 32%]). At the time of second-line initiation, a higher proportion (16%) of first-line ZDV users harboured TDF-resistant HIV compared with ZDV-resistant viruses among first-line TDF users (0% and 6% in base and pessimistic scenarios, respectively). In the base scenario, the incremental cost effectiveness ratio with respect to quality adjusted life years (QALY) was US$83 when TDF instead of ZDV was used in first-line therapy (pessimistic scenario: US$ 315), which was below the WHO threshold for high cost effectiveness (US$ 2154).
Using TDF instead of ZDV in first-line treatment in resource-limited settings is very cost-effective and likely to better preserve future treatment options in absence of virological monitoring.
The UK Collaborative HIV Cohort (UK CHIC) is an observational study that collates data on HIV-positive adults accessing HIV clinical care at (currently) 13 large clinics in the UK but does not collect pregnancy specific data. The National Study of HIV in Pregnancy and Childhood (NSHPC) collates data on HIV-positive women receiving antenatal care from every maternity unit in the UK and Ireland. Both studies collate pseudonymised data and neither dataset contains unique patient identifiers. A methodology was developed to find and match records for women reported to both studies thereby obtaining clinical and treatment data on pregnant HIV-positive women not available from either dataset alone.
Women in UK CHIC receiving HIV-clinical care in 1996–2009, were found in the NSHPC dataset by initially ‘linking’ records with identical date-of-birth, linked records were then accepted as a genuine ‘match’, if they had further matching fields including CD4 test date. In total, 2063 women were found in both datasets, representing 23.1% of HIV-positive women with a pregnancy in the UK (n = 8932). Clinical data was available in UK CHIC following most pregnancies (92.0%, 2471/2685 pregnancies starting before 2009). There was bias towards matching women with repeat pregnancies (35.9% (741/2063) of women found in both datasets had a repeat pregnancy compared to 21.9% (1502/6869) of women in NSHPC only) and matching women HIV diagnosed before their first reported pregnancy (54.8% (1131/2063) compared to 47.7% (3278/6869), respectively).
Through the use of demographic data and clinical dates, records from two independent studies were successfully matched, providing data not available from either study alone.
Data linkage; HIV; Pregnant women; Antiretroviral therapy; Cohort analysis; United Kingdom
Estimates of the prevalence of transmitted HIV drug resistance (TDR) in a population are derived from resistance tests performed on samples from patients thought to be naïve to antiretroviral treatment (ART). Much of the debate over reliability of estimates of the prevalence of TDR has focused on whether the sample population is representative. However estimates of the prevalence of TDR will also be distorted if some ART-experienced patients are misclassified as ART-naïve.
The impact of misclassification bias on the rate of TDR was examined. We developed methods to obtain adjusted estimates of the prevalence of TDR for different misclassification rates, and conducted sensitivity analyses of trends in the prevalence of TDR over time using data from the UK HIV Drug Resistance Database. Logistic regression was used to examine trends in the prevalence of TDR over time.
The observed rate of TDR was higher than true TDR when misclassification was present and increased as the proportion of misclassification increased. As the number of naïve patients with a resistance test relative to the number of experienced patients with a test increased, the difference between true and observed TDR decreased. The observed prevalence of TDR in the UK reached a peak of 11.3% in 2002 (odds of TDR increased by 1.10 (95% CI 1.02, 1.19, p(linear trend) = 0.02) per year 1997-2002) before decreasing to 7.0% in 2007 (odds of TDR decreased by 0.90 (95% CI 0.87, 0.94, p(linear trend) < 0.001) per year 2002-2007. Trends in adjusted TDR were altered as the misclassification rate increased; the significant downward trend between 2002-2007 was lost when the misclassification increased to over 4%.
The effect of misclassification of ART on estimates of the prevalence of TDR may be appreciable, and depends on the number of naïve tests relative to the number of experienced tests. Researchers can examine the effect of ART misclassification on their estimates of the prevalence of TDR if such a bias is suspected.
The question of whether a score for a specific antiretroviral (e.g. lopinavir/r in this analysis) that improves prediction of viral load response given by existing expert-based interpretation systems (IS) could be derived from analyzing the correlation between genotypic data and virological response using statistical methods remains largely unanswered.
Methods and Findings
We used the data of the patients from the UK Collaborative HIV Cohort (UK CHIC) Study for whom genotypic data were stored in the UK HIV Drug Resistance Database (UK HDRD) to construct a training/validation dataset of treatment change episodes (TCE). We used the average square error (ASE) on a 10-fold cross-validation and on a test dataset (the EuroSIDA TCE database) to compare the performance of a newly derived lopinavir/r score with that of the 3 most widely used expert-based interpretation rules (ANRS, HIVDB and Rega). Our analysis identified mutations V82A, I54V, K20I and I62V, which were associated with reduced viral response and mutations I15V and V91S which determined lopinavir/r hypersensitivity. All models performed equally well (ASE on test ranging between 1.1 and 1.3, p = 0.34).
We fully explored the potential of linear regression to construct a simple predictive model for lopinavir/r-based TCE. Although, the performance of our proposed score was similar to that of already existing IS, previously unrecognized lopinavir/r-associated mutations were identified. The analysis illustrates an approach of validation of expert-based IS that could be used in the future for other antiretrovirals and in other settings outside HIV research.
We investigated the effect of N348I alone and with M184V on nonnucleoside reverse transcriptase inhibitor (NNRTI) drug susceptibility and replicative capacity in B and non-B HIV-1 isolates. N348I reduced the susceptibility to all NNRTI drugs across subtypes. The replication capacity of all viruses in a variety of cell lines was impaired by N348I. Interestingly, the N348I and M184V double mutation compensated for the reduced NNRTI drug susceptibility observed in the N348I single mutant and marginally improved viral replicative capacity.
Other than cleavage site mutations, there is little data on specific positions within Gag that impact on HIV protease inhibitor susceptibility. We have recently shown that non-cleavage site mutations in gag, particularly within matrix protein can restore replication capacity and further reduce protease inhibitor drug susceptibility when coexpressed with a drug-resistant (mutant) protease. The matrix protein of this patient-derived virus was studied in order to identify specific changes responsible for this phenotype. Three amino acid changes in matrix (R76K, Y79F, and T81A) had an impact on replication capacity as well as drug susceptibility. Introduction of these three changes into wild-type (WT) matrix resulted in an increase in the replication capacity of the protease mutant virus to a level similar to that achieved by all the changes within the mutant matrix and part of the capsid protein. Pairs of changes to wild-type matrix led to an increased replication capacity of the protease mutant (although less than with all three changes). Having only these three changes to matrix in a wild-type virus (with wild-type protease) resulted in a 5- to 7-fold change in protease inhibitor 50% effective concentration (EC50). Individual changes did not have as great an effect on replication capacity or drug susceptibility, demonstrating an interaction between these positions, also confirmed by sequence covariation analysis. Molecular modeling predicts that each of the three mutations would result in a loss of hydrogen bonds within α-helix-4 of matrix, leading to the hypothesis that more flexibility within this region or altered matrix structure would account for our findings.
TRIM5α proteins recruit and restrict incoming cytoplasmic retroviruses. Primate TRIM5α sequence diversity underlies species-specific restriction and is likely caused by selective pressure from ancient pathogenic infections. Here we show that TRIM5α from the European brown hare restricts diverse retroviruses. Furthermore, it differs significantly in sequence from TRIM5α from the closely related rabbit, suggesting evolutionary changes in the last 12 million years since these species diverged. We propose that, like primates, lagomorphs have been subject to selective pressure from TRIM5-sensitive viruses, possibly related to the endogenous lentivirus RELIK found in both rabbits and hares.
The Q151M multi-drug resistance (MDR) pathway in HIV-1 reverse transcriptase (RT) confers reduced susceptibility to all nucleoside reverse transcriptase inhibitors (NRTIs) excluding tenofovir (TDF). This pathway emerges after long term failure of therapy, and is increasingly observed in the resource poor world, where antiretroviral therapy is rarely accompanied by intensive virological monitoring. In this study we examined the genotypic, phenotypic and fitness correlates associated with the development of Q151M MDR in the absence of viral load monitoring.
Single-genome sequencing (SGS) of full-length RT was carried out on sequential samples from an HIV-infected individual enrolled in ART rollout. The emergence of Q151M MDR occurred in the order A62V, V75I, and finally Q151M on the same genome at 4, 17 and 37 months after initiation of therapy, respectively. This was accompanied by a parallel cumulative acquisition of mutations at 20 other codon positions; seven of which were located in the connection subdomain. We established that fourteen of these mutations are also observed in Q151M-containing sequences submitted to the Stanford University HIV database. Phenotypic drug susceptibility testing demonstrated that the Q151M-containing RT had reduced susceptibility to all NRTIs except for TDF. RT domain-swapping of patient and wild-type RTs showed that patient-derived connection subdomains were not associated with reduced NRTI susceptibility. However, the virus expressing patient-derived Q151M RT at 37 months demonstrated ~44% replicative capacity of that at 4 months. This was further reduced to ~22% when the Q151M-containing DNA pol domain was expressed with wild-type C-terminal domain, but was then fully compensated by coexpression of the coevolved connection subdomain.
We demonstrate a complex interplay between drug susceptibility and replicative fitness in the acquisition Q151M MDR with serious implications for second-line regimen options. The acquisition of the Q151M pathway occurred sequentially over a long period of failing NRTI therapy, and was associated with mutations in multiple RT domains.
Several studies have implicated a recently discovered gammaretrovirus, XMRV (Xenotropic murine leukaemia virus-related virus), in chronic fatigue syndrome and prostate cancer, though whether as causative agent or opportunistic infection is unclear. It has also been suggested that the virus can be found circulating amongst the general population. The discovery has been controversial, with conflicting results from attempts to reproduce the original studies.
We extracted peripheral blood DNA from a cohort of 540 HIV-1-positive patients (approximately 20% of whom have never been on anti-retroviral treatment) and determined the presence of XMRV and related viruses using TaqMan PCR. While we were able to amplify as few as 5 copies of positive control DNA, we did not find any positive samples in the patient cohort.
In view of these negative findings in this highly susceptible group, we conclude that it is unlikely that XMRV or related viruses are circulating at a significant level, if at all, in HIV-1-positive patients in London or in the general population.