[Guided discontinuation of antidepressants: approach and first results of a Dutch multidisciplinary outpatient clinic]. / Werkwijze en eerste resultaten van multidisciplinaire afbouwpoli anti-depressiva.

BACKGROUND: Discontinuation of antidepressant medication can be difficult due to withdrawal symptoms and relapse risk. Scientific evidence on the questions of who, when, and how to stop antidepressants is limited. In Amsterdam a multidisciplinary outpatient clinic was started to provide advice and guidance. AIM: To substantiate the design of the clinic. Central questions relate to knowing which patients are referred, the background of their request, and their experiences with the outpatient clinic. METHOD: The first 51 patients of the clinic were described on the basis of file research, in addition a survey was conducted into patient experiences. RESULTS: Half of the patients (55%) actually started discontinuation, 39% were advised not to do so (yet). Patients at the clinic had used antidepressants for an average of 10 years, and 76% had previously attempted to stop. 21% had now successfully stopped and 25% were satisfied with a lower dose. One patient relapsed during tapering. CONCLUSION: So far, patients with long-term antidepressant use and multiple quit attempts have been referred. Our experiences are aimed at helping individual patients but can also result in more knowledge about who can stop at what moment, and how this should be done.

HIV testing for key populations in Europe: A decade of technological innovation and patient empowerment complement the role of health care professionals.

OBJECTIVES: With persisting high numbers of new HIV diagnoses in Europe, HIV testing remains an important aspect of HIV prevention. The traditional centralized and medicalized HIV testing approach has been complemented with newly developed and evaluated non-traditional approaches. Two important factors guided this process: technological innovation and empowerment of the patient. METHODS: We present a matrix to develop an HIV testing approach, and elaborate on three commonly used ones: community based testing, self-testing, and self-sampling. Despite non-traditional HIV testing approaches, barriers for testing remain. A potential disadvantage for users is the risk for false-reactive test results. As users receive an orientation test result, a reactive result should be confirmed. Another issue is the window phase, which is longer for some orientation tests compared to a traditional, laboratory-based test. RESULTS: Future implementation of non-traditional HIV testing approaches will depend on legal frameworks throughout Europe. Community testing centers may additionally improve empowerment of key populations by expanding their portfolio to testing and treatment for sexually transmitted infections. Community engagement and ownership may imply a shrinking role for health care providers, but they remain crucial actors for personalized information, counselling and referral to specialized HIV-care for many people. CONCLUSIONS: A highly effective HIV testing strategy to reduce undiagnosed people living with HIV in Europe is needed. Any approach, chosen according to the principles outlined in this paper, should reach the right people, diagnose them in the most accurate way, and optimize linkage to care.

The power of competence support: The impact of coaches and athlete leaders on intrinsic motivation and performance.

Grounded in the Cognitive Evaluation Theory, a mini-theory of Self-Determination Theory, this experimental field study sought to examine the impact of competence support of both coaches and athlete leaders on athletes' competence satisfaction, intrinsic motivation, and subjective as well as objective performance. Male basketball players (N = 120) were allocated to groups of 5 players. These groups were then randomly assigned to a control group or to 1 of 3 experimental conditions. In these experimental conditions, either the coach, the athlete leader, or both provided motivational feedback to their team. The provision of motivational feedback by either the coach or the athlete leader was sufficient to increase athletes' competence satisfaction, intrinsic motivation, and objective performance (i.e., enhanced execution time without a decrease in scoring percentage) relative to the control group. Interestingly, when both the coach and the athlete leader provided competence support, a surplus effect was observed on objective performance compared with when only the coach provided competence support. Furthermore, structural equation modeling revealed that players' competence satisfaction mediated the relationship between the provided competence support and players' intrinsic motivation, while a direct effect was observed on objective performance. In conclusion, the study findings indicate that also athlete leaders can adopt a motivating role, and that by doing so, their impact is as strong as the impact of the coach. Both coaches and athlete leaders can thus boost athletes' objective performance and foster competence satisfaction, with the latter resulting in increased intrinsic motivation.

Transmission of HIV Drug Resistance and the Predicted Effect on Current First-line Regimens in Europe.

BACKGROUND: Numerous studies have shown that baseline drug resistance patterns may influence the outcome of antiretroviral therapy. Therefore, guidelines recommend drug resistance testing to guide the choice of initial regimen. In addition to optimizing individual patient management, these baseline resistance data enable transmitted drug resistance (TDR) to be surveyed for public health purposes. The SPREAD program systematically collects data to gain insight into TDR occurring in Europe since 2001. METHODS: Demographic, clinical, and virological data from 4140 antiretroviral-naive human immunodeficiency virus (HIV)-infected individuals from 26 countries who were newly diagnosed between 2008 and 2010 were analyzed. Evidence of TDR was defined using the WHO list for surveillance of drug resistance mutations. Prevalence of TDR was assessed over time by comparing the results to SPREAD data from 2002 to 2007. Baseline susceptibility to antiretroviral drugs was predicted using the Stanford HIVdb program version 7.0. RESULTS: The overall prevalence of TDR did not change significantly over time and was 8.3% (95% confidence interval, 7.2%-9.5%) in 2008-2010. The most frequent indicators of TDR were nucleoside reverse transcriptase inhibitor (NRTI) mutations (4.5%), followed by nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations (2.9%) and protease inhibitor mutations (2.0%). Baseline mutations were most predictive of reduced susceptibility to initial NNRTI-based regimens: 4.5% and 6.5% of patient isolates were predicted to have resistance to regimens containing efavirenz or rilpivirine, respectively, independent of current NRTI backbones. CONCLUSIONS: Although TDR was highest for NRTIs, the impact of baseline drug resistance patterns on susceptibility was largest for NNRTIs. The prevalence of TDR assessed by epidemiological surveys does not clearly indicate to what degree susceptibility to different drug classes is affected.

Late presentation to HIV testing is overestimated when based on the consensus definition.

OBJECTIVES: In 2011, a consensus was reached defining "late presenters" (LPs) as individuals presenting for care with a CD4 count < 350 cells/µL or with an AIDS-defining event, regardless of CD4 count. However, a transient low CD4 count is not uncommon in recent infections. The objective of this study was to investigate how measurements of late presentation change if the clinical stage at the time of diagnosis is taken into account. METHODS: Case surveillance data for newly diagnosed patients in Belgium in 1998-2012 were analysed, including CD4 count at diagnosis, the presence of AIDS-defining events, and recent infections (< 6 months) as reported by clinicians in the case of acute illness or a recent negative test. First, proportions of LPs were calculated according to the consensus definition. Secondly, LPs were reclassified as "nonlate" if infections were reported as recent. RESULTS: A total of 7949 HIV diagnoses were included in the study. Recent infections were increasingly reported over time, accounting for 8.2% of new infections in 1998 and 37.5% in 2012. The consideration of clinical stage significantly modified the proportion of LPs: 18.2% of men who have sex with men (MSM) diagnosed in 2012 would be classified as LPs instead of 30.9% using the consensus definition (P < 0.001). The proportion of patients misclassified as LPs increased significantly over time: 5% in MSM in 1998 vs. 41% in 2012. CONCLUSIONS: This study suggests that low CD4 counts in recent infections may lead to overestimation of late presentation when applying the consensus definition. The impact of transient CD4 count on late presentation estimates should be assessed and, if relevant, the introduction of clinical stage in the definition of late presentation should be considered.

We will be champions: Leaders' confidence in 'us' inspires team members' team confidence and performance.

The present research examines the impact of leaders' confidence in their team on the team confidence and performance of their teammates. In an experiment involving newly assembled soccer teams, we manipulated the team confidence expressed by the team leader (high vs neutral vs low) and assessed team members' responses and performance as they unfolded during a competition (i.e., in a first baseline session and a second test session). Our findings pointed to team confidence contagion such that when the leader had expressed high (rather than neutral or low) team confidence, team members perceived their team to be more efficacious and were more confident in the team's ability to win. Moreover, leaders' team confidence affected individual and team performance such that teams led by a highly confident leader performed better than those led by a less confident leader. Finally, the results supported a hypothesized mediational model in showing that the effect of leaders' confidence on team members' team confidence and performance was mediated by the leader's perceived identity leadership and members' team identification. In conclusion, the findings of this experiment suggest that leaders' team confidence can enhance members' team confidence and performance by fostering members' identification with the team.

Role of genetic alterations in the NLRP3 and CARD8 genes in health and disease.

The complexity of a common inflammatory disease is influenced by multiple genetic and environmental factors contributing to the susceptibility of disease. Studies have reported that these exogenous and endogenous components may perturb the balance of innate immune response by activating the NLRP3 inflammasome. The multimeric NLRP3 complex results in the caspase-1 activation and the release of potent inflammatory cytokines, like IL-1ß. Several studies have been performed on the association of the genetic alterations in genes encoding NLRP3 and CARD8 with the complex diseases with inflammatory background, like inflammatory bowel disease, cardiovascular diseases, rheumatoid arthritis, and type 1 diabetes. The aim of the present review is therefore to summarize the literature regarding genetic alterations in these genes and their association with health and disease.

Concordance between HIV-1 genotypic coreceptor tropism predictions based on plasma RNA and proviral DNA.

OBJECTIVE: The aim of the study was to evaluate the use of proviral DNA as a source of viral genetic material for genotypic coreceptor tropism testing (GTT). METHODS: GTT consisted of bulk V3 sequencing followed by geno2pheno interpretation with the interpretative cut-off [false positive rate (FPR)] set at 5 and 10%. GTT was performed for 165 patients with a viral load of >500 HIV-1 RNA copies/mL on simultaneously collected plasma RNA and proviral DNA, and for 126 patients with a viral load of <500 copies/mL on current proviral DNA and pretreatment plasma RNA. Phenotypic tropism testing (PTT) results were available for 142 samples. RESULTS: In the simultaneous RNA/DNA comparison, concordance in prediction was 95.2% (at FPR 10%) and 96.4% (at FPR 5%). Six RNA-R5/DNA-X4 and two RNA-X4/DNA-R5 discordances were observed at an FPR of 10%, and six RNA-R5/DNA-X4 discordances were observed at an FPR of 5%. In the longitudinal RNA/DNA comparison, concordance was 88.1% (at FPR 10%) and 90.5% (at FPR 5%). Eight RNA-X4/DNA-R5 and seven RNA-R5/DNA-X4 discordances were seen at an FPR of 10%, and 10 RNA-R5/DNA-X4 and two RNA-X4/DNA-R5 discordances at an FPR of 5%. The overall concordance of RNA GTT with PTT was 82% (at FPR 10%) and 83% (at FPR 5%). The overall concordance of DNA GTT with PTT was 85% (at both 10 and 5% FPRs). CONCLUSIONS: GTT produced highly concordant tropism predictions for proviral DNA and plasma RNA. GTT on proviral DNA offers a promising approach for tropism prediction in clinical practice, particularly for the assessment of treated patients with low or suppressed viraemia.

A venue-based HIV prevalence and behavioural study among men who have sex with men in Antwerp and Ghent, Flanders, Belgium, October 2009 to March 2010.

This venue-based, cross-sectional study reports on human immunodeficiency virus (HIV) prevalence and behaviour of 649 men who have sex with men (MSM) in Antwerp and Ghent, Flanders, Belgium, from October 2009 to March 2010. Using time-location sampling, we found that HIV prevalence in MSM who attended different types of venue ranged from a high of 14.5% (95% CI: 8.9­20.1; n=22 in cruising venues to 4.9% (95% CI: 1.9­7.9; n=10) in more general gay venues to 1.4% (95% CI: 0.0­3.6; n=3) at younger MSM venues. Of those who tested HIV positive (n=35, five were unaware of their HIV status or self-reported as being HIV negative. One in five respondents were of non-Belgian nationality. The results showed relatively high rates of testing for HIV (52.2%; 95 % CI: 47.8­56.2; n=288) and other sexually transmitted infections (STIs) (57.4%; 95% CI: 52.6­62.0; n=248) in the last 12 months. A majority of the men (n=233) used condoms consistently during their last anal sexual contact with a casual partner; however, HIV-positive men who were aware of their serostatus (n=30) reported less condom use with casual partners. This is the first such study in Belgium and the results constitute the evidence base for local, targeted interventions. Furthermore, our findings underscore the need for European cross-border cooperation to prevent HIV infection and other STIs among MSM.

Low specificities of HIV diagnostic tests caused by Trypanosoma brucei gambiense sleeping sickness.

The accuracy of diagnostic tests for HIV in patients with tropical infections is poorly documented. Human African trypanosomiasis (HAT) is characterized by a polyclonal B-cell activation, constituting a risk for false-positive reactions to diagnostic tests, including HIV tests. A retrospective study of the accuracy of HIV diagnostic tests was performed with 360 human African HAT patients infected with Trypanosoma brucei gambiense before treatment and 163 T. b. gambiense-infected patients 2 years after successful treatment in Mbuji Mayi, East Kasai, Democratic Republic of the Congo. The sensitivities, specificities, and positive predictive values (PPVs) of individual tests and algorithms consisting of 3 rapid tests were determined. The sensitivity of all tests was 100% (11/11). The low specificity (96.3%, 335/348) and PPV (45.8%, 11/24) of a classical seroconfirmation strategy (Vironostika enzyme-linked immunosorbent assay [ELISA] followed by line immunoassay) complicated the determination of HIV status, which had to be determined by PCR. The specificities of the rapid diagnostic tests were 39.1% for Determine (136/348); 85.3 to 92.8% (297/348 to 323/348) for Vikia, ImmunoFlow, DoubleCheck, and Bioline; and 96.6 to 98.3% (336/348 to 342/348) for Uni-Gold, OraQuick, and Stat-Pak. The specificity of Vironostika was 67.5% (235/348). PPVs ranged between 4.9 and 64.7%. Combining 3 different rapid tests resulted in specificities of 98.3 to 100% (342/348 to 348/348) and PPVs of 64.7 to 100% (11/17 to 11/11). For cured HAT patients, specificities were significantly higher for Vironostika, Determine, Uni-Gold, and ImmunoFlow. T. b. gambiense infection decreases the specificities of antibody detection tests for HIV diagnosis. Unless tests have been validated for interference with HAT, HIV diagnosis using classical algorithms in untreated HAT patients should be avoided. Specific, validated combinations of 3 HIV rapid tests can increase specificity.

Does Social Capital Benefit Older Adults' Health and Well-Being? The Mediating Role of Physical Activity.

Objectives: To assess whether social capital benefits older adults' self-rated health and well-being and whether physical activity mediates this relation. Methods: A survey study was conducted among members of a sociocultural organization (age ≥55 years), both cross-sectionally (baseline Time 1; N = 959) and longitudinally (3-year follow-up Time 2; N = 409). Results: Specific indicators of social capital were positively, though modestly, related to health and well-being at Time 1 and Time 2. Experienced connectedness with age peers emerged as the strongest predictor. Physical activity only mediated the relation with experienced safety in society. Discussion: The relative importance of older adults' experienced connectedness with their age peers underlines the importance of internalized group membership as a determinant of their health and well-being. Physical activity seems to play only a minor mediating role.

Evaluation of the intercept oral specimen collection device with HIV assays versus paired serum/plasma specimens.

Oral fluid has many advantages over blood-based techniques: it is less invasive, eliminates the occupational risk associated with needle stick accidents and collection can be self-administrated. Each individual test is packaged with a corresponding collection device. This study tested the suitability of the Intercept Oral Specimen Collection Device for different HIV diagnostic tests: three different rapid HIV tests and two adapted ELISAs, which were evaluated and compared with a gold standard on blood. In addition a total IgG quantification was performed to demonstrate the quality of the specimen. HIV antibodies were detected with a sensitivity of 100%, 99.3%, 98.6%, 100% and 95.7% for, DPP, OraQuick, Aware, Genscreen and Vironostika respectively using the Intercept Collection Device. Respective specificities were 100%, 100%, 99.3%, 97.3% and 100%.

Cross-neutralizing antibodies to HIV-1ANT70 and HIV-1IIIB in sera of African and Belgian HIV-1-infected individuals.

OBJECTIVES: To determine the neutralizing antibody patterns to HIV-1ANT70 (ANT70) and HIV-1IIIB (IIIB) in human sera obtained from HIV-1-infected individuals from different African countries and Belgium. Second, to correlate the presence of neutralizing antibodies in sera and their ability to bind to synthetic peptides derived from eight different HIV-1 V3 loop sequences. DESIGN AND METHODS: Forty sera from Belgium and 88 obtained from seven countries in Africa were tested for their ability to neutralize ANT70 (one of the most genetically divergent HIV-1 isolates documented), and IIIB. Sera found to cross-neutralize both viruses were further challenged with four HIV-1 field isolates. All sera were tested on a panel of V3 loop peptides obtained from different HIV-1 genotypes. RESULTS: Four patterns of sera were identified, including 33 (26%) sera not neutralizing any of the isolates, seven (5%) sera neutralizing only ANT70, 45 (35%) sera neutralizing only IIIB, and 43 (34%) sera cross-neutralizing both isolates. Sera capable of cross-neutralizing both ANT70 and IIIB consistently neutralized other field isolates tested, with a remarkable similarity in neutralizing antibody titre. A significantly higher number of sera cross-neutralizing both ANT70 and IIIB compared with sera lacking neutralizing antibodies, reacted simultaneously in enzyme-linked immunosorbent assays (ELISA) with three or more V3 loop peptides belonging to HIV-1 strains of different genotypes. However, none of the sera cross-neutralizing ANT70 and IIIB were reactive in ELISA with the ANT70 V3 loop peptide. CONCLUSION: These results suggest that despite pronounced genomic variation of the HIV-1ANT70 isolate, there are strongly conserved neutralizing epitopes situated outside the V3 loop that are shared by other HIV-1 isolates. These findings suggest that genetic variation might be surmountable in the design of a polyvalent HIV vaccine, if neutralizing antibodies are found to be correlates of protection in HIV infection.

Isolation and characterization of a new chimpanzee lentivirus (simian immunodeficiency virus isolate cpz-ant) from a wild-captured chimpanzee.

OBJECTIVE: To assess the prevalence of infection with simian immunodeficiency virus (SIV) isolate cpz, a lentivirus closely related to HIV-1, in chimpanzees, and to obtain new SIVcpz isolates. METHODS: Forty-four wild-captured chimpanzees in Belgium and Côte d'Ivoire were tested for HIV and SIV antibodies. Virus was isolated from the peripheral blood lymphocytes of positive animals and characterized by electron microscopy, Western blot and radioimmunoprecipitation assay. RESULTS: One animal had antibodies that cross-reacted with HIV-1. A lentivirus was isolated and referred to as SIVcpz-ant. With regard to molecular weight patterns, SIVcpz-ant differs from SIVcpz-gab' an HIV-1-related virus isolated from a wild-captured chimpanzee in Gabon. The major core protein, the transmembrane and outer membrane glycoproteins of the SIVcpz-ant strain consistently had higher molecular weights. Significantly more HIV-1-positive sera reacted with the envelope proteins of the Gabonese SIVcpz-gab strain than with the SIVcpz-ant strain. CONCLUSIONS: This study shows that natural infection of wild-captured chimpanzees with an HIV-related virus may not be uncommon. The diversity of the two chimpanzee isolates, the different geographical origin and the absence of disease suggest that chimpanzees have not recently become SIVcpz-infected.

Lack of correlation between V3-loop peptide enzyme immunoassay serologic subtyping and genetic sequencing.

OBJECTIVE: To compare the performance of V3-loop peptide enzyme immunoassay (PEIA) methodologies from four different laboratories for subtyping HIV-1, and to determine the causes for the lack of correlation between V3-loop PEIA serotyping and subtyping by sequencing. MATERIALS AND METHODS: Synthetic peptides derived from the amino-acid consensus sequences of the V3-loop of group M strains representing genetic subtypes A-F as well as reference strains were evaluated in PEIA by four different laboratories for their ability to accurately determine the subtype in a panel of 85 sera obtained from persons infected with known HIV-1 subtypes (28 subtype A, 34 subtype B, four subtype C, 10 subtype D, seven subtype F, one each of subtype H and G). Furthermore, the V3 loop of the corresponding virus was compared with the V3 loop of the peptides used in PEIA. RESULTS: The correlation between HIV-1 subtyping by sequencing and V3-loop PEIA from the different laboratories varied considerably for the different HIV-1 subtypes: subtype A (46-68%), B (38-85%), C (75-100%), D (29-50%), and F (17-57%). A 70% agreement between PEIA and sequencing subtypes was observed for samples with the concordant presence of the same octameric sequences in the V3 loop of the virus and the V3 loop of the peptide used in PEIA; however, only 42% of specimens with different V3-loop octameric viral and peptide sequences yielded concordant results in V3-loop serotyping and genetic subtyping. CONCLUSION: Our results indicate that V3-loop PEIA methodologies used in different laboratories correlate poorly with genetic subtyping, and that their accuracy to predict HIV-1 subtypes in sera of Belgian individuals infected with different HIV-1 subtypes (A, B, C, D, F, G and H) vary considerably. The poor correlation between serotyping and genetic subtyping was partly due to the simultaneous occurrence of subtype-specific octameric sequences at the tip of the V3 loop of viruses belonging to different genetic subtypes.

Genotypic subtypes of HIV-1 in Cameroon.

OBJECTIVE: The only two HIV-1 strains (ANT70 and MVP5180) reported to date from Cameroon are members of the outlier clade (group O). In this study, we assessed the prevalence of group O viruses and other HIV-1 subtypes in Cameroon. DESIGN: A phylogenetic analysis of 18 HIV-1 strains isolated from seropositive individuals from Yaoundé and Douala, Cameroon. METHODS: A 900 base-pair fragment of the env gene coding for V3, V4, V5, and the beginning of gp41 of 17 out of 18 HIV-1 isolates from Cameroon was amplified, cloned and sequenced using polymerase chain reaction. A phylogenetic tree was constructed. RESULTS: The overall env nucleotide sequence divergence among the Cameroon isolates ranged from 6.1 to 27.5%. In a phylogenetic tree, six subtypes were identified when compared with 23 reference strains of different geographic origin. Of these 17 Cameroonian strains, 11 (61%) were of subtype A of which the interpatient distances at the sequence level varied from 6.1% to 18.3% (average, 11.9%). Three (17%) strains were of subtype F, and the other three strains (6% each) belonged to subtypes B, E and H, respectively. The remaining isolate was classified as belonging to group O, on the basis of the sequence of part of the pol gene. A very broad spectrum of different tetrameric amino-acid sequences was observed at the apex of the V3 loop. Eleven strains contained the tetrameric globally predominant GPGQ sequence at the tip of the V3 motif. Two strains had the GPGR sequence typical of the American and European HIV-1 strains. The remaining tetrameric sequences included GPGS, GSGQ, GRGQ, and GLGR. CONCLUSION: These findings on a limited number of viruses suggest extensive env gene diversity of HIV-1 strains from Cameroon, and could have implications for vaccine development in Africa.

Molecular phylogeny of part of the env gene of HIV-1 strains isolated in Côte d'Ivoire.

OBJECTIVES: To examine the genetic variation of HIV-1 isolates in Abidjan, Côte d'Ivoire, and to determine the extent to which phylogenetic trees based on sequence information of part of the env gene containing the principal neutralizing domain are representative for documenting genetic variability. DESIGN: Phylogenetic comparison of 13 HIV-1 strains isolated from patients in Abidjan with previously documented HIV-1 strains of different geographic origin. METHODS: To sequence a 900 base-pair fragment of the env gene containing V3, V4, V5 and the beginning of gp41 of three to four clones per isolate. Phylogenetic tree analysis was performed with the software package TREECON. RESULTS: Eleven HIV-1 isolates of Abidjan were classified as genotype A, while two were classed as genotypes B and D. Intra-genotype A distances at the nucleotide level were a maximum of 14.1%. Inter-genotype distances between genotype A and genotypes B, C, and D varied from 16.0 to 22.6%. Phylogenetic trees, based on sequence data of a 300 base-pair fragment containing the V3 loop, showed significant differences in tree topology and statistical confidence with phylogenetic trees based on sequence data of the 900 base-pair env fragment. CONCLUSIONS: Genotype A Côte d'Ivoire HIV-1 strains, which comprise 11 out of 13 isolates, predominate in Abidjan, which may indicate a local burst of particular variants. Phylogenetic trees should be interpreted with caution when based on a more limited number of nucleotides, such as the V3 region.

Phylogenetic analysis of the env gene of HIV-1 isolates taking into account individual nucleotide substitution rates.

OBJECTIVE: To estimate the relative substitution rate of the individual positions in an alignment of HIV-1 env sequences coding for areas V3, V4, V5, and the beginning of gp41, and to study phylogenetic relationships between HIV-1 strains taking into account these substitution rate estimates. DESIGN: Phylogenetic comparison of 145 HIV-1 strains classified in HIV-1 group M, subtypes A-H and isolated from patients of 24 different geographical origins. METHODS: A new method recently developed for measuring the substitution rates of the individual nucleotides in a sequence alignment was applied to an alignment of env gene sequences. From the resulting substitution rate distribution, an equation was derived that describes the relationship between dissimilarity and evolutionary distance better than equations previously available. Phylogenetic trees were then constructed from matrices of distances computed using this new equation. RESULTS: 'Substitution rate calibration' offers detailed information on the relative substitution rate or variability of the nucleotides in the env gene. A large phylogenetic tree of 145 env gene sequences constructed by neighbour-joining and taking into account the substitution rate spectrum for this gene, clearly shows the existence of the eight subtypes A-H, all supported at a bootstrap level of 90% or higher. Intersubtype distances were between 0.25 and 0.38, which is considerably higher than those found in trees not considering differences in substitution rates among different alignment positions. CONCLUSIONS: Evolutionary distances are seriously underestimated when individual substitution rates are not considered in the estimation evolutionary distances. Furthermore, due to the more accurate estimation of evolutionary distances, naturally occurring HIV-1 intersubtype recombinants could be recognized more easily.

Interpatient genetic variability of HIV-1 group O.

OBJECTIVE: To analyse the genetic and phylogenetic characteristics of HIV-1 group O viruses. MATERIALS AND METHODS: The env gene, encoding the gp160 glycoprotein, and a partial p24-encoding gag gene fragment of a Cameroonian (CA9) and a Gabonese (VI686) HIV-1 group O virus, isolated from cultured peripheral blood mononuclear cells of symptomatic patients, were sequenced, aligned with other representatives of group O for which the same region has been documented, and genetically and phylogenetically analysed. RESULTS: Phylogenetic analysis of the env gene (gp160) revealed that CA9, VI686, ANT70, and four Ha strains formed a separate cluster, which was supported by 100% of all bootstrap trees. In addition, these seven isolates were part of the same clade in the p24 phylogeny. VAU and MVP5180 may represent two other subtypes. CONCLUSION: We have characterized two group O viruses, originating from Cameroon and Gabon, which show a close evolutionary relationship to ANT70 and four Ha strains based on the entire env gene, suggestive of a first group O subgroup, tentatively named the HIV-1 group O env ANT70 clade or subtype.

Phylogenetic analysis of gag genes from 70 international HIV-1 isolates provides evidence for multiple genotypes.

OBJECTIVE: To determine the extent of genetic variation among internationally collected HIV-1 isolates, to analyse phylogenetic relationships and the geographic distribution of different variants. DESIGN: Phylogenetic comparison of 70 HIV-1 isolates collected in 15 countries on four continents. METHODS: To sequence the complete gag genome of HIV-1 isolates, build multiple sequence alignments and construct phylogenetic trees using distance matrix methods and maximum parsimony algorithms. RESULTS: Phylogenetic tree analysis identified seven distinct genotypes. The seven genotypes were evident by both distance matrix methods and maximum parsimony analysis, and were strongly supported by bootstrap resampling of the data. The intra-genotypic gag distances averaged 7%, whereas the inter-genotypic distances averaged 14%. The geographic distribution of variants was complex. Some genotypes have apparently migrated to several continents and many areas harbor a mixture of genotypes. Related variants may cluster in certain areas, particularly isolates from a single city collected over a short time. CONCLUSIONS: The genetic variation among HIV-1 isolates is more extensive than previously appreciated. At least seven distinct HIV-1 genotypes can be identified. Diversification, migration and establishment of local, temporal 'blooms' of particular variants may all occur concomitantly.