The development and evaluation of a probe hybridization method for subtyping HIV type 1 infection in Uganda.
We developed a method for large-scale screening of HIV-1 genotypic variation based on DNA probe hybridization. Nested PCR amplifications were performed to generate fragments in the env C2-V3 region and also in the gp41 region, which encompasses the immunodominant domain. The proviral DNA sequences were derived from 68 samples and phylogenetically analyzed. For comparison, the C2-V3 fragment was used in DNA probe hybridization to rapidly determine the infecting HIV subtype. The hybridizing probes were designed on the basis of the two most prevalent subtypes in Uganda, A and D. The results were compared to evaluate the feasibility of using this hybridization method for large-scale genotypic screening. Sequence analysis of the 68 amplified PCR fragments showed that 39 were subtype A and 29 were subtype D. The results of DNA hybridization to the amplified products with A and D subtype-specific probes were more than 90% concordant with the subtypes determined by sequence analysis. Our findings suggest that probe hybridization with subtype-specific probes is effective for large-scale screening of HIV-infected populations. Application of this method will significantly reduce the time needed for large, population-based investigations.
Luo
C C
CC
Centers for Disease Control and Prevention, Atlanta, Georgia 30333, USA. CXL1@CDC.GOV
Downing
R G
RG
Dela Torre
N
N
Baggs
J
J
Hu
D J
DJ
Respess
R A
RA
Candal
D
D
Carr
L
L
George
J R
JR
Dondero
T J
TJ
Biryahwaho
B
B
Rayfield
M A
MA
D015342:DNA Probes; D004279:DNA, Viral / Q000097:blood*; D014644:Genetic Variation / Q000235:genetics; D015699:HIV Envelope Protein gp120 / Q000235:genetics; D015700:HIV Envelope Protein gp41 / Q000235:genetics; D015658:HIV Infections / Q000453:epidemiology / Q000821:virology*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D017720:Molecular Epidemiology; D015336:Molecular Probe Techniques; D010446:Peptide Fragments / Q000235:genetics; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D014454:Uganda / Q000453:epidemiology
D015342:DNA Probes; D004279:DNA, Viral; D015699:HIV Envelope Protein gp120; D015700:HIV Envelope Protein gp41; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments