Optimization of the oligonucleotide ligation assay, a rapid and inexpensive test for detection of HIV-1 drug resistance mutations, for non-North American variants.
OBJECTIVE
We evaluated the feasibility of the oligonucleotide ligation assay (OLA), a specific, sensitive, and economical ligase-based point mutation assay designed to detect HIV-1 drug-resistance mutations at 12 codons of HIV-1 subtype B pol, for potential use in resource-poor settings.


METHODS
Specimens from HIV-1-infected individuals collected by 7 international laboratories, including subtypes A, B, C, D, F, G, J, and recombinants AE and AG, were tested by the OLA developed for HIV-1 subtype B. Common polymorphisms that interfered with reactivity of the OLA were identified and modified probes designed and evaluated.


RESULTS
92.5% (2,410) of 2,604 codons in specimens from 217 individuals were successfully genotyped by the subtype B OLA. A high rate (range 8.3%-31.2%) of indeterminate results (negative OLA reaction for both mutant and wild type) was observed for 5 codons. Modified probes at reverse transcriptase codons 151 and 184 and protease codon 90 increased the rate of valid OLA to 96.1%.


CONCLUSIONS
The OLA designed for HIV-1 subtype B genotyped most pol codons in non-B subtypes from Asia and Africa but was improved by addition of several modified probes. International laboratories experienced in molecular techniques were able to perform the OLA.
Beck
Ingrid A
IA
Division of Infectious Diseases and Virology, University of Washington, Seattle, WA, USA.
Crowell
Claudia
C
Kittoe
Robin
R
Bredell
Helba
H
Machaba
Molefe
M
Willamson
Carolyn
C
Janssens
Wouter
W
Jallow
Sabelle
S
van der Groen
Guido
G
Shao
Yiming
Y
Jacob
Mini
M
Samuel
N M
NM
de Rivera
Ivette Lorenzana
IL
Ngo-Giang-Huong
Nicole
N
Cassol
Sharon
S
Alemnji
George
G
Frenkel
Lisa M
LM
D019380:Anti-HIV Agents / Q000494:pharmacology*; D003062:Codon; D011088:DNA Ligases; D018076:DNA, Complementary / Q000235:genetics; D024882:Drug Resistance, Viral; D005240:Feasibility Studies; D015751:Genes, pol / Q000235:genetics; D015658:HIV Infections / Q000821:virology*; D016333:HIV Protease / Q000235:genetics; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D012194:RNA-Directed DNA Polymerase / Q000235:genetics; D012680:Sensitivity and Specificity
D019380:Anti-HIV Agents; D003062:Codon; D018076:DNA, Complementary; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D012194:RNA-Directed DNA Polymerase; D016333:HIV Protease; D011088:DNA Ligases