An updated model of shoot apical meristem regulation by ERECTA family and CLAVATA3 signaling pathways in Arabidopsis.

The shoot apical meristem (SAM) gives rise to the aboveground organs of plants. The size of the SAM is relatively constant due to the balance between stem cell replenishment and cell recruitment into new organs. In angiosperms, the transcription factor WUSCHEL (WUS) promotes stem cell proliferation in the central zone of the SAM. WUS forms a negative feedback loop with a signaling pathway activated by CLAVATA3 (CLV3). In the periphery of the SAM, the ERECTA family receptors (ERfs) constrain WUS and CLV3 expression. Here, we show that four ligands of ERfs redundantly inhibit the expression of these two genes. Transcriptome analysis confirmed that WUS and CLV3 are the main targets of ERf signaling and uncovered new ones. Analysis of promoter reporters indicated that the WUS expression domain mostly overlaps with the CLV3 domain and does not shift along the apical-basal axis in clv3 mutants. Our three-dimensional mathematical model captured gene expression distributions at the single-cell level under various perturbed conditions. Based on our findings, CLV3 regulates cellular levels of WUS mostly through autocrine signaling, and ERfs regulate the spatial expression of WUS, preventing its encroachment into the peripheral zone.

Initiation of aboveground organ primordia depends on combined action of auxin, ERECTA family genes, and PINOID.

Leaves and flowers are produced by the shoot apical meristem (SAM) at a certain distance from its center, a process that requires the hormone auxin. The amount of auxin and the pattern of its distribution in the initiation zone determine the size and spatial arrangement of organ primordia. Auxin gradients in the SAM are formed by PIN-FORMED (PIN) auxin efflux carriers whose polar localization in the plasma membrane depends on the protein kinase PINOID (PID). Previous work determined that ERECTA (ER) family genes (ERfs) control initiation of leaves. ERfs are plasma membrane receptors that enable cell-to-cell communication by sensing extracellular small proteins from the EPIDERMAL PATTERNING FACTOR/EPF-LIKE (EPF/EPFL) family. Here, we investigated whether ERfs regulate initiation of organs by altering auxin distribution or signaling in Arabidopsis (Arabidopsis thaliana). Genetic and pharmacological data suggested that ERfs do not regulate organogenesis through PINs while transcriptomics data showed that ERfs do not alter primary transcriptional responses to auxin. Our results indicated that in the absence of ERf signaling the peripheral zone cells inefficiently initiate leaves in response to auxin signals and that increased accumulation of auxin in the er erecta-like1 (erl1) erl2 SAM can partially rescue organ initiation defects. We propose that both auxin and ERfs are essential for leaf initiation and that they have common downstream targets. Genetic data also indicated that the role of PID in initiation of cotyledons and leaves cannot be attributed solely to regulation of PIN polarity and PID is likely to have other functions in addition to regulation of auxin distribution.

Light Activates the Translational Regulatory Kinase GCN2 via Reactive Oxygen Species Emanating from the Chloroplast.

Cytosolic mRNA translation is subject to global and mRNA-specific controls. Phosphorylation of the translation initiation factor eIF2α anchors a reversible regulatory switch that represses cytosolic translation globally. The stress-responsive GCN2 kinase is the only known kinase for eIF2α serine 56 in Arabidopsis (Arabidopsis thaliana). Here, we show that conditions that generate reactive oxygen species (ROS) in the chloroplast, including dark-light transitions, high light, and the herbicide methyl viologen, rapidly activated GCN2 kinase, whereas mitochondrial and endoplasmic reticulum stress did not. GCN2 activation was light dependent and mitigated by photosynthesis inhibitors and ROS quenchers. Accordingly, the seedling growth of multiple Arabidopsis gcn2 mutants was retarded under excess light conditions, implicating the GCN2-eIF2α pathway in responses to light and associated ROS. Once activated, GCN2 kinase preferentially suppressed the ribosome loading of mRNAs for functions such as mitochondrial ATP synthesis, the chloroplast thylakoids, vesicle trafficking, and translation. The gcn2 mutant overaccumulated transcripts functionally related to abiotic stress, including oxidative stress, as well as innate immune responses. Accordingly, gcn2 displayed defects in immune priming by the fungal elicitor, chitin. Therefore, we provide evidence that reactive oxygen species produced by the photosynthetic apparatus help activate the highly conserved GCN2 kinase, leading to eIF2α phosphorylation and thus affecting the status of the cytosolic protein synthesis apparatus.

Human Immunodeficiency Virus-2 (HIV-2): A Summary of the Present Standard of Care and Treatment Options for Individuals Living with HIV-2 in Western Europe.

Human immunodeficiency virus-2 (HIV-2) is endemic in some countries in West Africa. Due to the lower prevalence in industrialized countries, there is limited experience and knowledge on the management of individuals living with HIV-2 in Europe. Compared to HIV-1, there are differential characteristics of HIV-2 regarding diagnostic procedures, the clinical course, and, most importantly, antiretroviral therapy. We integrated the published literature on HIV-2 (studies and reports on epidemiology, diagnostics, the clinical course, and treatment), as well as expert experience in diagnosing and clinical care, to provide recommendations for a present standard of medical care of those living with HIV-2 in Western European countries, including an overview of strategies for diagnosis, monitoring, and treatment, with suggestions for effective drug combinations for first- and second-line treatments, post-exposure prophylaxis, and the prevention of mother-to-child transmission, as well as listings of mutations related to HIV-2 drug resistance and C-C motif chemokine receptor type 5 and C-X-C motif chemokine receptor type 4 coreceptor tropism.

Cyclometalated Osmium Compounds and beyond: Synthesis, Properties, Applications.

The synthesis of cyclometalated osmium complexes is usually more complicated than of other transition metals such as Ni, Pd, Pt, Rh, where cyclometalation reactions readily occur via direct activation of C-H bonds. It differs also from their ruthenium analogs. Cyclometalation for osmium usually occurs under more severe conditions, in polar solvents, using specific precursors, stronger acids, or bases. Such requirements expand reaction mechanisms to electrophilic activation, transmetalation, and oxidative addition, often involving C-H bond activations. Osmacycles exhibit specific applications in homogeneous catalysis, photophysics, bioelectrocatalysis and are studied as anticancer agents. This review describes major synthetic pathways to osmacycles and related compounds and discusses their practical applications.

Tracing the Impact of Public Health Interventions on HIV-1 Transmission in Portugal Using Molecular Epidemiology.

BACKGROUND: Estimation of temporal changes in human immunodeficiency virus (HIV) transmission patterns can help to elucidate the impact of preventive strategies and public health policies. METHODS: Portuguese HIV-1 subtype B and G pol genetic sequences were appended to global reference data sets to identify country-specific transmission clades. Bayesian birth-death models were used to estimate subtype-specific effective reproductive numbers (Re). Discrete trait analysis (DTA) was used to quantify mixing among transmission groups. RESULTS: We identified 5 subtype B Portuguese clades (26-79 sequences) and a large monophyletic subtype G Portuguese clade (236 sequences). We estimated that major shifts in HIV-1 transmission occurred around 1999 (95% Bayesian credible interval [BCI], 1998-2000) and 2000 (95% BCI, 1998-2001) for subtypes B and G, respectively. For subtype B, Re dropped from 1.91 (95% BCI, 1.73-2.09) to 0.62 (95% BCI,.52-.72). For subtype G, Re decreased from 1.49 (95% BCI, 1.39-1.59) to 0.72 (95% BCI, .63-.8). The DTA suggests that people who inject drugs (PWID) and heterosexuals were the source of most (>80%) virus lineage transitions for subtypes G and B, respectively. CONCLUSIONS: The estimated declines in Re coincide with the introduction of highly active antiretroviral therapy and the scale-up of harm reduction for PWID. Inferred transmission events across transmission groups emphasize the importance of prevention efforts for bridging populations.

A genotypic method for determining HIV-2 coreceptor usage enables epidemiological studies and clinical decision support.

BACKGROUND: CCR5-coreceptor antagonists can be used for treating HIV-2 infected individuals. Before initiating treatment with coreceptor antagonists, viral coreceptor usage should be determined to ensure that the virus can use only the CCR5 coreceptor (R5) and cannot evade the drug by using the CXCR4 coreceptor (X4-capable). However, until now, no online tool for the genotypic identification of HIV-2 coreceptor usage had been available. Furthermore, there is a lack of knowledge on the determinants of HIV-2 coreceptor usage. Therefore, we developed a data-driven web service for the prediction of HIV-2 coreceptor usage from the V3 loop of the HIV-2 glycoprotein and used the tool to identify novel discriminatory features of X4-capable variants. RESULTS: Using 10 runs of tenfold cross validation, we selected a linear support vector machine (SVM) as the model for geno2pheno[coreceptor-hiv2], because it outperformed the other SVMs with an area under the ROC curve (AUC) of 0.95. We found that SVMs were highly accurate in identifying HIV-2 coreceptor usage, attaining sensitivities of 73.5% and specificities of 96% during tenfold nested cross validation. The predictive performance of SVMs was not significantly different (p value 0.37) from an existing rules-based approach. Moreover, geno2pheno[coreceptor-hiv2] achieved a predictive accuracy of 100% and outperformed the existing approach on an independent data set containing nine new isolates with corresponding phenotypic measurements of coreceptor usage. geno2pheno[coreceptor-hiv2] could not only reproduce the established markers of CXCR4-usage, but also revealed novel markers: the substitutions 27K, 15G, and 8S were significantly predictive of CXCR4 usage. Furthermore, SVMs trained on the amino-acid sequences of the V1 and V2 loops were also quite accurate in predicting coreceptor usage (AUCs of 0.84 and 0.65, respectively). CONCLUSIONS: In this study, we developed geno2pheno[coreceptor-hiv2], the first online tool for the prediction of HIV-2 coreceptor usage from the V3 loop. Using our method, we identified novel amino-acid markers of X4-capable variants in the V3 loop and found that HIV-2 coreceptor usage is also influenced by the V1/V2 region. The tool can aid clinicians in deciding whether coreceptor antagonists such as maraviroc are a treatment option and enables epidemiological studies investigating HIV-2 coreceptor usage. geno2pheno[coreceptor-hiv2] is freely available at http://coreceptor-hiv2.geno2pheno.org .

Global Dispersal Pattern of HIV Type 1 Subtype CRF01_AE: A Genetic Trace of Human Mobility Related to Heterosexual Sexual Activities Centralized in Southeast Asia.

BACKGROUND: Human immunodeficiency virus type 1 (HIV-1) subtype CRF01_AE originated in Africa and then passed to Thailand, where it established a major epidemic. Despite the global presence of CRF01_AE, little is known about its subsequent dispersal pattern. METHODS: We assembled a global data set of 2736 CRF01_AE sequences by pooling sequences from public databases and patient-cohort studies. We estimated viral dispersal patterns, using statistical phylogeographic analysis run over bootstrap trees estimated by the maximum likelihood method. RESULTS: We show that Thailand has been the source of viral dispersal to most areas worldwide, including 17 of 20 sampled countries in Europe. Japan, Singapore, Vietnam, and other Asian countries have played a secondary role in the viral dissemination. In contrast, China and Taiwan have mainly imported strains from neighboring Asian countries, North America, and Africa without any significant viral exportation. DISCUSSION: The central role of Thailand in the global spread of CRF01_AE can be probably explained by the popularity of Thailand as a vacation destination characterized by sex tourism and by Thai emigration to the Western world. Our study highlights the unique case of CRF01_AE, the only globally distributed non-B clade whose global dispersal did not originate in Africa.

Possible footprints of APOBEC3F and/or other APOBEC3 deaminases, but not APOBEC3G, on HIV-1 from patients with acute/early and chronic infections.

UNLABELLED: Members of the apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like-3 (APOBEC3) innate cellular cytidine deaminase family, particularly APOBEC3F and APOBEC3G, can cause extensive and lethal G-to-A mutations in HIV-1 plus-strand DNA (termed hypermutation). It is unclear if APOBEC3-induced mutations in vivo are always lethal or can occur at sublethal levels that increase HIV-1 diversification and viral adaptation to the host. The viral accessory protein Vif counteracts APOBEC3 activity by binding to APOBEC3 and promoting proteasome degradation; however, the efficiency of this interaction varies, since a range of hypermutation frequencies are observed in HIV-1 patient DNA. Therefore, we examined "footprints" of APOBEC3G and APOBEC3F activity in longitudinal HIV-1 RNA pol sequences from approximately 3,000 chronically infected patients by determining whether G-to-A mutations occurred in motifs that were favored or disfavored by these deaminases. G-to-A mutations were more frequent in APOBEC3G-disfavored than in APOBEC3G-favored contexts. In contrast, mutations in APOBEC3F-disfavored contexts were relatively rare, whereas mutations in contexts favoring APOBEC3F (and possibly other deaminases) occurred 16% more often than average G-to-A mutations. These results were supported by analyses of >500 HIV-1 env sequences from acute/early infection. IMPORTANCE: Collectively, our results suggest that APOBEC3G-induced mutagenesis is lethal to HIV-1, whereas mutagenesis caused by APOBEC3F and/or other deaminases may result in sublethal mutations that might facilitate viral diversification. Therefore, Vif-specific cytotoxic T lymphocyte (CTL) responses and drugs that manipulate the interplay between Vif and APOBEC3 may have beneficial or detrimental clinical effects depending on how they affect the binding of Vif to various members of the APOBEC3 family.

RegaDB: community-driven data management and analysis for infectious diseases.

SUMMARY: RegaDB is a free and open source data management and analysis environment for infectious diseases. RegaDB allows clinicians to store, manage and analyse patient data, including viral genetic sequences. Moreover, RegaDB provides researchers with a mechanism to collect data in a uniform format and offers them a canvas to make newly developed bioinformatics tools available to clinicians and virologists through a user friendly interface. AVAILABILITY AND IMPLEMENTATION: Source code, binaries and documentation are available on http://rega.kuleuven.be/cev/regadb. RegaDB is written in the Java programming language, using a web-service-oriented architecture.

Increase in transmitted resistance to non-nucleoside reverse transcriptase inhibitors among newly diagnosed HIV-1 infections in Europe.

BACKGROUND: One out of ten newly diagnosed patients in Europe was infected with a virus carrying a drug resistant mutation. We analysed the patterns over time for transmitted drug resistance mutations (TDRM) using data from the European Spread program. METHODS: Clinical, epidemiological and virological data from 4317 patients newly diagnosed with HIV-1 infection between 2002 and 2007 were analysed. Patients were enrolled using a pre-defined sampling strategy. RESULTS: The overall prevalence of TDRM in this period was 8.9% (95% CI: 8.1-9.8). Interestingly, significant changes over time in TDRM caused by the different drug classes were found. Whereas nucleoside resistance mutations remained constant at 5%, a significant decline in protease inhibitors resistance mutations was observed, from 3.9% in 2002 to 1.6% in 2007 (p = 0.001). In contrast, resistance to non-nucleoside reverse transcriptase inhibitors (NNRTIs) doubled from 2.0% in 2002 to 4.1% in 2007 (p = 0.004) with 58% of viral strains carrying a K103N mutation. Phylogenetic analysis showed that these temporal changes could not be explained by large clusters of TDRM. CONCLUSION: During the years 2002 to 2007 transmitted resistance to NNRTI has doubled to 4% in Europe. The frequent use of NNRTI in first-line regimens and the clinical impact of NNRTI mutations warrants continued monitoring.

Pharmacy refill adherence outperforms self-reported methods in predicting HIV therapy outcome in resource-limited settings.

BACKGROUND: Optimal adherence to antiretroviral therapy is critical to prevent HIV drug resistance (HIVDR) epidemic. The objective of the study was to investigate the best performing adherence assessment method for predicting virological failure in resource-limited settings (RLS). METHOD: This study was a single-centre prospective cohort, enrolling 220 HIV-infected adult patients attending an HIV/AIDS Care and Treatment Centre in Dar es Salaam, Tanzania, in 2010. Pharmacy refill, self-report (via visual analog scale [VAS] and the Swiss HIV Cohort study-adherence questionnaire), pill count, and appointment keeping adherence measurements were taken. Univariate logistic regression (LR) was done to explore a cut-off that gives a better trade-off between sensitivity and specificity, and a higher area under the curve (AUC) based on receiver operating characteristic curve in predicting virological failure. Additionally, the adherence models were evaluated by fitting multivariate LR with stepwise functions, decision trees, and random forests models, assessing 10-fold multiple cross validation (MCV). Patient factors associated with virological failure were determined using LR. RESULTS: Viral load measurements at baseline and one year after recruitment were available for 162 patients, of whom 55 (34%) had detectable viral load and 17 (10.5%) had immunological failure at one year after recruitment. The optimal cut-off points significantly predictive of virological failure were 95%, 80%, 95% and 90% for VAS, appointment keeping, pharmacy refill, and pill count adherence respectively. The AUC for these methods ranged from 0.52 to 0.61, with pharmacy refill giving the best performance at AUC 0.61. Multivariate logistic regression with boost stepwise MCV had higher AUC (0.64) compared to all univariate adherence models, except pharmacy refill adherence univariate model, which was comparable to the multivariate model (AUC = 0.64). Decision trees and random forests models were inferior to boost stepwise model. Pharmacy refill adherence (<95%) emerged as the best method for predicting virological failure. Other significant predictors in multivariate LR were having a baseline CD4 T lymphocytes count < 200 cells/µl, being unable to recall the diagnosis date, and a higher weight. CONCLUSION: Pharmacy refill has the potential to predict virological failure and to identify patients to be considered for viral load monitoring and HIVDR testing in RLS.

Declining prevalence of HIV-1 drug resistance in antiretroviral treatment-exposed individuals in Western Europe.

HIV-1 drug resistance represents a major obstacle to infection and disease control. This retrospective study analyzes trends and determinants of resistance in antiretroviral treatment (ART)-exposed individuals across 7 countries in Europe. Of 20 323 cases, 80% carried at least one resistance mutation: these declined from 81% in 1997 to 71% in 2008. Predicted extensive 3-class resistance was rare (3.2% considering the cumulative genotype) and peaked at 4.5% in 2005, decreasing thereafter. The proportion of cases exhausting available drug options dropped from 32% in 2000 to 1% in 2008. Reduced risk of resistance over calendar years was confirmed by multivariable analysis.

A phosphorylation-deficient ribosomal protein eS6 is largely functional in Arabidopsis thaliana, rescuing mutant defects from global translation and gene expression to photosynthesis and growth.

The eukaryote-specific ribosomal protein of the small subunit eS6 is phosphorylated through the target of rapamycin (TOR) kinase pathway. Although this phosphorylation event responds dynamically to environmental conditions and has been studied for over 50 years, its biochemical and physiological significance remains controversial and poorly understood. Here, we report data from Arabidopsis thaliana, which indicate that plants expressing only a phospho-deficient isoform of eS6 grow essentially normally under laboratory conditions. The eS6z (RPS6A) paralog of eS6 functionally rescued a double mutant in both rps6a and rps6b genes when expressed at approximately twice the wild-type dosage. A mutant isoform of eS6z lacking the major six phosphorylatable serine and threonine residues in its carboxyl-terminal tail also rescued the lethality, rosette growth, and polyribosome loading of the double mutant. This isoform also complemented many mutant phenotypes of rps6 that were newly characterized here, including photosynthetic efficiency, and most of the gene expression defects that were measured by transcriptomics and proteomics. However, compared with plants rescued with a phospho-enabled version of eS6z, the phospho-deficient seedlings retained a mild pointed-leaf phenotype, root growth was reduced, and certain cell cycle-related mRNAs and ribosome biogenesis proteins were misexpressed. The residual defects of the phospho-deficient seedlings could be understood as an incomplete rescue of the rps6 mutant defects. There was little or no evidence for gain-of-function defects. As previously published, the phospho-deficient eS6z also rescued the rps6a and rps6b single mutants; however, phosphorylation of the eS6y (RPS6B) paralog remained lower than predicted, further underscoring that plants can tolerate phospho-deficiency of eS6 well. Our data also yield new insights into how plants cope with mutations in essential, duplicated ribosomal protein isoforms.

Mycoplasma pneumoniae at the rise not only in China: rapid increase of Mycoplasma pneumoniae cases also in Spain.

After the use of facemasks, other isolation measures enacted during the SARS-CoV-2 pandemic were lifted, respiratory pathogens, such as RSV, reappeared, but until the November 2023 WHO alert for China, M. pneumoniae had virtually disappeared. After observing a similar reappearance in our hospital, a retrospective analysis of the number of positive M. pneumoniae tests. Between 2018 and December 2023, 1619 PCR tests were ordered and 43 (2.6%) of them were positive. Two outbreaks, one in 2018 and one in 2023, accounted for the majority of cases. Tests were usually ordered in an outpatient setting (53.54%, n = 23) and most of them were paediatric patients with a mean age (sd) of 10.2 (6.2) years. As for the severity of the cases, in the 2018 outbreak, of 15 children who tested positive, 53.3% (n = 8) were admitted to the ward and 6.7% (n = 1) at the intensive care unit. Whereas in 2023, 2 patients were tested in the ward (10.5%) and one in the intensive care unit (5.2%) from a total of 19 patients. The positive rate in 2023 was significantly higher in comparison with years 2020, 2021 and 2022 and significantly lower in comparison with 2018 (P-value=0.003). The outbreak in late 2023 can be explained by the seasonality of Mycoplasma pneumonia alone, which has shown outbreaks every 3-5 years, and it does not appear to be more severe than the previous one.

HIV-2EU: supporting standardized HIV-2 drug resistance interpretation in Europe.

Considering human immunodeficiency virus type 2 (HIV-2) phenotypic data and experience from HIV type 1 and from the follow-up of HIV-2-infected patients, a panel of European experts voted on a rule set for interpretation of mutations in HIV-2 protease, reverse transcriptase, and integrase and an automated tool for HIV-2 drug resistance analyses freely available on the Internet (http://www.hiv-grade.de).

HIV-1 subtype distribution and its demographic determinants in newly diagnosed patients in Europe suggest highly compartmentalized epidemics.

BACKGROUND: Understanding HIV-1 subtype distribution and epidemiology can assist preventive measures and clinical decisions. Sequence variation may affect antiviral drug resistance development, disease progression, evolutionary rates and transmission routes. RESULTS: We investigated the subtype distribution of HIV-1 in Europe and Israel in a representative sample of patients diagnosed between 2002 and 2005 and related it to the demographic data available. 2793 PRO-RT sequences were subtyped either with the REGA Subtyping tool or by a manual procedure that included phylogenetic tree and recombination analysis. The most prevalent subtypes/CRFs in our dataset were subtype B (66.1%), followed by sub-subtype A1 (6.9%), subtype C (6.8%) and CRF02_AG (4.7%). Substantial differences in the proportion of new diagnoses with distinct subtypes were found between European countries: the lowest proportion of subtype B was found in Israel (27.9%) and Portugal (39.2%), while the highest was observed in Poland (96.2%) and Slovenia (93.6%). Other subtypes were significantly more diagnosed in immigrant populations. Subtype B was significantly more diagnosed in men than in women and in MSM > IDUs > heterosexuals. Furthermore, the subtype distribution according to continent of origin of the patients suggests they acquired their infection there or in Europe from compatriots. CONCLUSIONS: The association of subtype with demographic parameters suggests highly compartmentalized epidemics, determined by social and behavioural characteristics of the patients.

Limited cross-border infections in patients newly diagnosed with HIV in Europe.

BACKGROUND: International travel plays a role in the spread of HIV-1 across Europe. It is, however, not known whether international travel is more important for spread of the epidemic as compared to endogenous infections within single countries. In this study, phylogenetic associations among HIV of newly diagnosed patients were determined across Europe. RESULTS: Data came from the SPREAD programme which collects samples of newly diagnosed patients that are representative for national HIV epidemics. 4260 pol sequences from 25 European countries and Israel collected in 2002-2007 were included.We identified 457 clusters including 1330 persons (31.2% of all patients). The cluster size ranged between 2 and 28. A number of 987 patients (74.2%) were part of a cluster that consisted only of patients originating from the same country. In addition, 135 patients (10.2%) were in a cluster including only individuals from neighboring countries. Finally, 208 patients (15.6%) clustered with individuals from countries without a common border. Clustering with patients from the same country was less prevalent in patients being infected with B subtype (P-value <0.0001), in men who have sex with men (P-value <0.0001), and in recently infected patients (P-value =0.045). CONCLUSIONS: Our findings indicate that the transmission of HIV-1 in Europe is predominantly occurring between patients from the same country. This could have implications for HIV-1 transmission prevention programmes. Because infections through travelling between countries is not frequently observed it is important to have good surveillance of the national HIV-1 epidemics.

Mutations selected in HIV-2-infected patients failing a regimen including atazanavir.

OBJECTIVES: To investigate mutations selected in viruses from HIV-2-infected patients failing a highly active antiretroviral treatment (HAART) regimen including atazanavir/ritonavir. METHODS: Twenty-eight HIV-2-infected patients previously exposed to atazanavir/ritonavir and failing therapy were studied. The protease (PR) gene was amplified and sequenced, and mutations emerging under atazanavir/ritonavir selective pressure were reported. RESULTS: The I50L mutation emerged in 4 out of 28 HIV-2-infected patients failing a HAART regimen including atazanavir/ritonavir. Besides I50L, four PR mutations previously associated with protease inhibitor resistance (I54L, I64V, V71I and I82F) and six PR mutations of unknown impact (V10I, E37D, S43T, K45R, I75V and F85L) in HIV-2 were also identified in this small group of patients. CONCLUSIONS: Several mutations were associated with virological failure of a regimen including atazanavir/ritonavir in HIV-2-infected patients, including I50L for the first time. It should be included in HIV-2 algorithms for interpretation of genotypic resistance data, and taken into account when making therapeutic decisions for HIV-2-infected patients.

The demise of multidrug-resistant HIV-1: the national time trend in Portugal.

OBJECTIVES: Despite a decreasing mortality and morbidity in treated HIV-1 patients, highly active antiretroviral treatment (HAART) can still fail due to the development of drug resistance. Especially, multidrug-resistant viruses pose a threat to efficient therapy. We studied the changing prevalence of multidrug resistance (MDR) over time in a cohort of HIV-1-infected patients in Portugal. PATIENTS AND METHODS: We used data of 8065 HIV-1-infected patients followed from July 2001 up to April 2012 in 22 hospitals located in Portugal. MDR at a specific date of sampling was defined as no more than one fully active drug (excluding integrase and entry inhibitors) at that time authorized by the Portuguese National Authority of Medicines and Health Products (INFARMED), as interpreted with the Rega algorithm version 8.0.2. A generalized linear mixed model was used to study the time trend of the prevalence of MDR. RESULTS: We observed a statistically significant decrease in the prevalence of MDR over the last decade, from 6.9% (95% CI: 5.7-8.4) in 2001-03, 6.0% (95% CI: 4.9-7.2) in 2003-05, 3.7% (95% CI: 2.8-4.8) in 2005-07 and 1.6% (95% CI: 1.1-2.2) in 2007-09 down to 0.6% (95% CI: 0.3-0.9) in 2009-12 [OR=0.80 (95% CI: 0.75-0.86); P<0.001]. In July 2011 the last new case of MDR was seen. CONCLUSIONS: The prevalence of multidrug-resistant HIV-1 is decreasing over time in Portugal, reflecting the increasing efficiency of HAART and the availability of new drugs. Therefore, in designing a new drug, safety and practical aspects, e.g. less toxicity and ease of use, may need more attention than focusing mainly on efficacy against resistant strains.