Motivation: An accurate genome assembly from short read sequencing data is critical for downstream analysis, for example allowing investigation of variants within a sequenced population. However, assembling sequencing data from virus samples, especially RNA viruses, into a genome sequence is challenging due to the combination of viral population diversity and extremely uneven read depth caused by amplification bias in the inevitable reverse transcription and polymerase chain reaction amplification process of current methods.Results: We developed a new de novo assembler called IVA (Iterative Virus Assembler) designed specifically for read pairs sequenced at highly variable depth from RNA virus samples. We tested IVA on datasets from 140 sequenced samples from human immunodeficiency virus-1 or influenza-virus-infected people and demonstrated that IVA outperforms all other virus de novo assemblers.Availability and implementation: The software runs under Linux, has the GPLv3 licence and is freely available from http://sanger-pathogens.github.io/ivaContact: iva@sanger.ac.ukSupplementary information: Supplementary data are available at Bioinformatics online.

e 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 shortread 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.

Early ART initiation and a higher proportion of time suppressed are linked with lower total HIV-1 DNA. Early ART start and improving adherence in perinatally HIV-1 infected children minimize the size of viral reservoir.This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial License 4.0 (CCBY-NC), where it is permissible to download, share, remix, transform, and buildup the work provided it is properly cited. The work cannot be used commercially without permission from the journal.

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Human immunodeficiency virus type 2 (HIV-2) RNA quantification assays used in nine laboratories of the ACHI E V 2E (A Collaboration on HIV-2 Infection) study group were evaluated. In a blinded experimental design, laboratories quantified three series of aliquots of an HIV-2 subtype A strain, each at a different theoretical viral load. Quantification varied between laboratories, and international standardization of quantification assays is strongly needed.

Accurate HIV-2 plasma viral load quantification is crucial for adequate HIV-2 patient management and for the proper conduct of clinical trials and international cohort collaborations. This study compared the homogeneity of HIV-2 RNA quantification when using HIV-2 assays from ACHI E V 2E study sites and either in-house PCR calibration standards or common viral load standards supplied to all collaborators. Each of the 12 participating laboratories quantified blinded HIV-2 samples, using its own HIV-2 viral load assay and standard as well as centrally validated and distributed common HIV-2 group A and B standards ( http://www.hiv.lanl.gov/content/sequence/HelpDocs/subtypes-more.html ). Aliquots of HIV-2 group A and B strains, each at 2 theoretical concentrations (2.7 and 3.7 log 10 copies/ml), were tested. Intralaboratory, interlaboratory, and overall variances of quantification results obtained with both standards were compared using F tests. For HIV-2 group A quantifications, overall and interlaboratory and/or intralaboratory variances were significantly lower when using the common standard than when using in-house standards at the concentration levels of 2.7 log 10 copies/ml and 3.7 log 10 copies/ml, respectively. For HIV-2 group B, a high heterogeneity was observed and the variances did not differ according to the type of standard used. In this international collaboration, the use of a common standard improved the homogeneity of HIV-2 group A RNA quantification only. The diversity of HIV-2 group B, particularly in PCR primer-binding regions, may explain the heterogeneity in quantification of this strain. Development of a validated HIV-2 viral load assay that accurately quantifies distinct circulating strains is needed.

The anti-HDV testing rate was low in a centre with clinic-led testing, but could not be evaluated in all centres. The HDV-infected patients were of diverse ethnicity, with extensive histological evidence of liver disease and poor therapeutic responses. Future recommendations include reflex laboratory testing algorithms and a prospective cohort study to optimise the investigation and management of these patients.

Western blot score can estimate HIV-DNA size and timing of ART initiation in long-term virally suppressed children. This rapid, inexpensive, and easily reproducible tool can provide useful information to identify potential candidates for HIV remission studies.