HIV type 2 proviral load measured by quantitative polymerase chain reaction correlates with CD4+ lymphopenia in HIV type 2-infected individuals.
The efficiency of detection of 2 sets of primer pairs from putatively conserved regions of the human immunodeficiency virus type 2 (HIV-2) genome were assessed in 86 seropositive individuals from The Gambia by nested polymerase chain reaction (PCR). HIV-2 long terminal repeat (LTR) target sequences were detected in DNA extracted from peripheral blood mononuclear cells (PBMCs) in 84 of 86 (97%) individuals whereas HIV-2 integrase (pol) gene sequences were detected in 39 of 41 (95%) individuals. The use of LTR target sequences and recombinant Pfu DNA polymerase, rather than Taq polymerase, in a modified secondary amplification reaction mediated the incorporation of 35S-labeled nucleotides in a quantitative radiometric assay. This sensitive assay was used to quantify HIV-2 proviral DNA in clinical samples and compared well with estimations by limiting end-point dilution of target molecules. A linear response between counts and the number of copies amplified from serial dilutions of pROD10 plasmid DNA (3-2000 copies) yielded a standard curve to allow extrapolation to clinical data. Increased levels of HIV-2 proviral DNA, expressed as copies per 10(5) CD4-positive lymphocytes, were associated with declining CD4 count in 63 adult patients (Spearman rank correlation, r = -0.71, n = 63, p < 0.001) and with the occurrence of HIV-related clinical disease. Kruskall-Wallis analysis of variance analysis showed the mean proviral copy number (log10) to be significantly different between groups (p < 0.001) where CD4 counts were grouped as < 200/mm3 (3.4 +/- 1.05 copies), 200-500/mm3 (2.84 +/- 0.93 copies), and > 500/mm3 (1.88 +/- 0.43 copies).(ABSTRACT TRUNCATED AT 250 WORDS)
Berry
N
N
Division of Virology, University College London Medical School, England.
Ariyoshi
K
K
Jobe
O
O
Ngum
P T
PT
Corrah
T
T
Wilkins
A
A
Whittle
H
H
Tedder
R
R
D000328:Adult; D000368:Aged; D001483:Base Sequence; D002648:Child; D015331:Cohort Studies; D004254:DNA Nucleotidyltransferases / Q000235:genetics; D004279:DNA, Viral / Q000097:blood; D004259:DNA-Directed DNA Polymerase; D005260:Female; D005714:Gambia; D015658:HIV Infections / Q000150:complications / Q000175:diagnosis / Q000821:virology*; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015498:HIV-2 / Q000235:genetics* / Q000502:physiology; D006801:Humans; D019426:Integrases; D007963:Leukocytes, Mononuclear / Q000821:virology; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000235:genetics*; D012680:Sensitivity and Specificity; D018344:T-Lymphocytopenia, Idiopathic CD4-Positive / Q000097:blood / Q000150:complications / Q000821:virology*
D004279:DNA, Viral; D004254:DNA Nucleotidyltransferases; D019426:Integrases; C072146:Pfu DNA polymerase; D004259:DNA-Directed DNA Polymerase