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Factors Influencing the Sensitivity and Specificity of Conventional Sequencing in Human Immunodeficiency Virus Type 1 Tropism Testing
- Source :
- Journal of Clinical Microbiology. 51:444-451
- Publication Year :
- 2013
- Publisher :
- American Society for Microbiology, 2013.
-
Abstract
- Human immunodeficiency virus type 1 (HIV-1) V3 loop sequence can be used to infer viral coreceptor use. The effect of input copy number on population-based sequencing of the V3 loop of HIV-1 was examined through replicate deep and population-based sequencing of samples with known tropism, a heterogeneous clinical sample (624 population-based sequences and 47 deep-sequencing replicates), and a large cohort of clinical samples from phase III clinical trials of maraviroc including the MOTIVATE/A4001029 studies ( n = 1,521). Proviral DNA from two independent samples from each of 101 patients from the MOTIVATE/A4001029 studies was also analyzed. Cumulative technical error occurred at a rate of 3 × 10 −4 mismatches/bp, without observed effect on inferred tropism. Increasing PCR replication increased minority species detection with an ∼10% minority population detected in 18% of cases using a single replicate at a viral load of 1,072 copies/ml and in 44% of cases using three replicates. The nucleotide prevalence detected by population-based and deep sequencing were highly correlated (Spearman's ρ, 0.73), and the accuracy increased with increasing input copy number ( P < 0.001). Triplicate sequencing was able to predict tropism changes in the MOTIVATE/A4001029 studies for both low ( P = 0.05) and high ( P = 0.02) viral loads. Sequences derived from independently extracted and processed samples of proviral DNA for the same patient were equivalent to replicates from the same extraction ( P = 0.45) and had correlated position-specific scoring matrix scores (Spearman's ρ, 0.75; P ≪ 0.001); however, concordance in tropism inference was only 83%. Input copy number and PCR replication are important factors in minority species detection in samples with significant heterogeneity.
- Subjects :
- Microbiology (medical)
Genotype
Genotyping Techniques
Population
HIV Infections
Genome, Viral
Biology
V3 loop
Virus Replication
Sensitivity and Specificity
Deep sequencing
chemistry.chemical_compound
Virology
Humans
education
Genetic Association Studies
Tropism
Maraviroc
Genetics
education.field_of_study
Base Sequence
High-Throughput Nucleotide Sequencing
Reproducibility of Results
Replicate
Viral Load
Viral Tropism
Phenotype
chemistry
HIV-1
Tissue tropism
Viral load
Subjects
Details
- ISSN :
- 1098660X and 00951137
- Volume :
- 51
- Database :
- OpenAIRE
- Journal :
- Journal of Clinical Microbiology
- Accession number :
- edsair.doi.dedup.....48b6b4c70b566a6ec1378e80f85a6235
- Full Text :
- https://doi.org/10.1128/jcm.00739-12