An isolate of human immunodeficiency virus type 1 originally classified as subtype I represents a complex mosaic comprising three different group M subtypes (A, G, and I)

Full-length reference clones and sequences are currently available for eight human immunodeficiency virus type 1 (HIV-1) group M subtypes (A through H), but none have been reported for subtypes I and J, which have only been identified in a few individuals. Phylogenetic information for subtype I, in particular, is limited since only about 400 bp of env gene sequences have been determined for just two epidemiologically linked viruses infecting a couple who were heterosexual intravenous drug users from Cyprus. To characterize subtype I in greater detail, we employed long-range PCR to clone a full-length provirus (94CY032.3) from an isolate obtained from one of the individuals originally reported to be infected with this subtype. Phylogenetic analysis of C2-V3 env gene sequences confirmed that 94CY032.3 was closely related to sequences previously classified as subtype I. However, analysis of the remainder of its genome revealed various regions in which 94CY032.3 was significantly clustered with either subtype A or subtype G. Only sequences located in vpr and nef, as well as the middle portions of pol and env, formed independent lineages roughly equidistant from all other known subtypes. Since these latter regions most likely have a common origin, we classify them all as subtype I. These results thus indicate that the originally reported prototypic subtype I isolate 94CY032 represents a triple recombinant (A/G/I) with at least 11 points of recombination crossover. We also screened HIV-1 recombinants with regions of uncertain subtype assignment for the presence of subtype I sequences. This analysis revealed that two of the earliest mosaics from Africa, Z321B (A/G/?) and MAL (A/D/?), contain short segments of sequence which clustered closely with the subtype I domains of 94CY032.3. Since Z321 was isolated in 1976, subtype I as well as subtypes A and G must have existed in Central Africa prior to that date... (D'après résumé d'auteur

CCR5 Haplotypes and Mother-to-Child HIV Transmission in Malawi

CCR5 and CCR2 gene polymorphisms (SNPs) have been associated with protection against HIV transmission in adults and with delayed progression to AIDS. The CCR5 Delta32 deletion and SNP -2459G are associated with reduced expression of the CCR5 protein.We investigated the association between infant CCR2/CCR5 diplotype and HIV mother to child transmission (MTCT) in Malawi. Blood samples from infants (n = 552) of HIV positive women who received nevirapine were genotyped using a post-PCR multiplex ligase detection reaction and haplotypes were identified based on 8 CCR2/CCR5 SNPs and the open reading frame 32 base pair deletion. Following verification of Hardy-Weinberg equilibrium, log linear regression was performed to examine the association between mutations and MTCT. Overall, protection against MTCT was weakly associated with two CCR5 SNPs, -2459G (Risk ratio [RR], 0.78; confidence interval [CI], 0.54-1.12), and the linked CCR5 -2135T (RR, 0.78; CI, 0.54-1.13). No child carried the CCR5 Delta32 SNP. Maternal Viral Load (MVL) was found to be an effect measure modifier. Among mothers with low MVL, statistically significant protection against MTCT was observed for -2459G (RR, 0.50; CI, 0.27-0.91), and -2135T (RR, 0.51; CI, 0.28-0.92). Statistically significant protection was not found at high MVL.Results from this study suggest that CCR5 SNPs -2459G and -2135T associated with reduced receptor expression protect against MTCT of HIV at low MVLs, whereas high MVLs may over-ride differences in coreceptor availability

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 4,140 antiretroviral-naive 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 2002-2007. Baseline susceptibility to antiretroviral drugs was predicted using Stanford HIVdb v7.0. RESULTS: The overall prevalence of TDR did not change significantly over time and was 8.3% (95%CI 7.2-9.5) in 2008-2010. The most frequent indicators of TDR were NRTI-mutations (4.5%), followed by NNRTI-mutations (2.9%) and PI-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 affecte

HIV-1 Infection in Cyprus, the Eastern Mediterranean European Frontier: A Densely Sampled Transmission Dynamics Analysis from 1986 to 2012

Since HIV-1 treatment is increasingly considered an effective preventionstrategy, it is important to study local HIV-1 epidemics to formulate tailored preventionpolicies. The prevalence of HIV-1 in Cyprus was historically low until 2005. To investigatethe shift in epidemiological trends, we studied the transmission dynamics of HIV-1 in Cyprususing a densely sampled Cypriot HIV-1 transmission cohort that included 85 percent ofHIV-1-infected individuals linked to clinical care between 1986 and 2012 based on detailedclinical, epidemiological, behavioral and HIV-1 genetic information. Subtyping andtransmission cluster reconstruction were performed using maximum likelihood and Bayesianmethods, and the transmission chain network was linked to the clinical, epidemiological andbehavioral data. The results reveal that for the main HIV-1 subtype A1 and B sub-epidemics,young and drug-naïve HIV-1-infected individuals in Cyprus are driving the dynamics of thelocal HIV-1 epidemic. The results of this study provide a better understanding of thedynamics of the HIV-1 infection in Cyprus, which may impact the development of preventionstrategies. Furthermore, this methodology for analyzing densely sampled transmissiondynamics is applicable to other geographic regions to implement effective HIV-1 preventionstrategies in local settings

Classification of HIV-1 Sequences Using Profile Hidden Markov Models

Accurate classification of HIV-1 subtypes is essential for studying the dynamic spatial distribution pattern of HIV-1 subtypes and also for developing effective methods of treatment that can be targeted to attack specific subtypes. We propose a classification method based on profile Hidden Markov Model that can accurately identify an unknown strain. We show that a standard method that relies on the construction of a positive training set only, to capture unique features associated with a particular subtype, can accurately classify sequences belonging to all subtypes except B and D. We point out the drawbacks of the standard method; namely, an arbitrary choice of threshold to distinguish between true positives and true negatives, and the inability to discriminate between closely related subtypes. We then propose an improved classification method based on construction of a positive as well as a negative training set to improve discriminating ability between closely related subtypes like B and D. Finally, we show how the improved method can be used to accurately determine the subtype composition of Common Recombinant Forms of the virus that are made up of two or more subtypes. Our method provides a simple and highly accurate alternative to other classification methods and will be useful in accurately annotating newly sequenced HIV-1 strains

CCL3L1 copy number, CCR5 genotype and susceptibility to tuberculosis

Background: Tuberculosis is a major infectious disease and functional studies have provided evidence that both the chemokine MIP-1α and its receptor CCR5 play a role in susceptibility to TB. Thus by measuring copy number variation of CCL3L1, one of the genes that encode MIP-1α, and genotyping a functional promoter polymorphism -2459A > G in CCR5 (rs1799987) we investigate the influence of MIP-1α and CCR5, independently and combined, in susceptibility to clinically active TB in three populations, a Peruvian population (n = 1132), a !Xhosa population (n = 605) and a South African Coloured population (n = 221). The three populations include patients with clinically diagnosed pulmonary TB, as well as other, less prevalent forms of extrapulmonary TB. Methods and results: Copy number of CCL3L1 was measured using the paralogue ratio test and exhibited ranges between 0–6 copies per diploid genome (pdg) in Peru, between 0–12 pdg in !Xhosa samples and between 0–10 pdg in South African Coloured samples. The CCR5 promoter polymorphism was observed to differ significantly in allele frequency between populations (*A; Peru f = 0.67, !Xhosa f = 0.38, Coloured f = 0.48). Conclusions: The case–control association studies performed however find, surprisingly, no evidence for an influence of variation in genes coding for MIP-1α or CCR5 individually or together in susceptibility to clinically active TB in these populations

Synthesis of novel MMT/acyl-protected nucleo alanine monomers for the preparation of DNA/alanyl-PNA chimeras

Alanyl-peptide nucleic acid (alanyl-PNA)/DNA chimeras are oligomers envisaged to be beneficial in efficient DNA diagnostics based on an improved molecular beacon concept. A synthesis of alanyl-PNA/DNA chimera can be based on the solid phase assembly of the oligomer with mixed oligonucleotide/peptide backbone under DNA synthesis conditions, in which the nucleotides are introduced as phosphoramidites, whereas the nucleo amino acids make use of the acid labile monomethoxytrityl (MMT) group for temporary protection of the α-amino groups and acyl protecting groups for the exocyclic amino functions of the nucleobases. In this work, we realized for the first time the synthesis of all four MMT/acyl-protected nucleo alanines, achieved by deprotection/reprotection of the newly synthesized Boc/acyl intermediates, useful monomers for the obtainment of (alanyl-PNA)/DNA chimeras by conditions fully compatible with the standard phosphoramidite DNA synthesis strategy

The use of nanocrystal quantum dot as fluorophore reporters in molecular beacon-based assays

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