Genotypic testing on HIV-1 DNA as a tool to assess HIV-1 co-receptor usage in clinical practice: results from the DIVA study group.

Purpose We have developed a sequencing assay for
determining the usage of the genotypic HIV-1 co-receptor
using peripheral blood mononuclear cell (PBMC) DNA in
virologically suppressed HIV-1 infected patients. Our
specific aims were to (1) evaluate the efficiency of V3
sequences in B versus non-B subtypes, (2) compare the
efficiency of V3 sequences and tropism prediction using
whole blood and PBMCs for DNA extraction, (3) compare
the efficiency of V3 sequences and tropism prediction
using a single versus a triplicate round of amplification.
Results The overall rate of successful V3 sequences
ranged from 100 % in samples with[3,000 copies HIV-1
DNA/106 PBMCs to 60 % in samples with \100 copies
total HIV-1 DNA /106 PBMCs. Analysis of 143 paired
PBMCs and whole-blood samples showed successful V3
sequences rates of 77.6 % for PBMCs and 83.9 % for
whole blood. These rates are in agreement with the tropism
prediction obtained using the geno2pheno co-receptor
algorithm, namely, 92.1 % with a false-positive rate (FPR)of 10 or 20 % and of 96.5 % with an FPR of 5.75 %. The
agreement between tropism prediction values using single
versus triplicate amplification was 98.2 % (56/57) of
patients using an FPR of 20 % and 92.9 % (53/57) using an
FPR of 10 or 5.75 %. For 63.0 % (36/57) of patients, the
FPR obtained via the single amplification procedure was
superimposable to all three FPRs obtained by triplicate
amplification.
Conclusions Our results show the feasibility and consistency
of genotypic testing on HIV-1 DNA tropism, supporting
its possible use for selecting patients with suppressed plasma HIV-1 RNA as candidates for CCR5-antagonist treatment. The high agreement between tropism prediction by single and triple amplification does not support the use of triplicate amplification in clinical practice.