title: Techniques used to identify the Brazilian variant of HIV-1 subtype B.
abstract: The purpose of the present study was to compare the sensitivity and specificity of V3 enzyme immunoassay (solid phase EIA and EIA inhibition) and restriction fragment length polymorphism (RFLP) with the DNA sequencing "gold standard" to identify the Brazilian HIV-1 variants of subtype B and B"-GWGR. Peripheral blood mononuclear cells were collected from 61 HIV-1-infected individuals attending a clinic in São Paulo. Proviral DNA was amplified and sequentially cleaved with the Fok I restriction enzyme. Plasma samples were submitted to a V3-loop biotinylated synthetic peptide EIA. Direct partial DNA sequencing of the env gene was performed on all samples. Based on EIA results, the sensitivity for detecting B-GPGR was 70%, compared to 64% for the Brazilian variant B"-GWGR while, the specificity of B-GPGR detection was 85%, compared to 88% for GWGR. The assessment of RFLP revealed 68% sensitivity and 94% specificity for the B-GPGR strain compared to 84 and 90% for the B"-GWGR variant. Moreover, direct DNA sequencing was able to detect different base sequences corresponding to amino acid sequences at the tip of the V3 loop in 22 patients. These results show a similar performance of V3 serology and RLFP in identifying the Brazilian variant GWGR. However, V3 peptide serology may give indeterminate results. Therefore, we suggest that V3 serology be used instead of DNA sequencing where resources are limited. Samples giving indeterminate results by V3 peptide serology should be analyzed by direct DNA sequencing to distinguish between B-GPGR and the Brazilian variant B"-GWGR.
authors.affiliation: Laboratório de Alergia e Imunologia Clínica e Experimental, LIM-56, Departamento de Dermatologia, Faculdade de Medicina, Universidade de São Paulo, Av. Dr. Enéas C. Aguiar 500, 05403-000 São Paulo, SP, Brazil. shirley_200@hotmail.com
authors.forename: S
authors.initials: S
authors.lastname: Komninakis
authors.forename: L
authors.initials: L
authors.lastname: Fukumori
authors.forename: R
authors.initials: R
authors.lastname: Alcalde
authors.forename: M
authors.initials: M
authors.lastname: Cortina
authors.forename: L
authors.initials: L
authors.lastname: Abdala
authors.forename: A
authors.initials: A
authors.lastname: Brito
authors.forename: S
authors.initials: S
authors.lastname: Sanabani
authors.forename: A J S
authors.initials: AJ
authors.lastname: Duarte
authors.forename: J
authors.initials: J
authors.lastname: Casseb
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D015699:HIV Envelope Protein gp120 / Q000235:genetics*; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D007124:Immunoenzyme Techniques; D007963:Leukocytes, Mononuclear / Q000821:virology*; D008297:Male; D010446:Peptide Fragments / Q000235:genetics*; D012150:Polymorphism, Restriction Fragment Length; D011533:Proviruses / Q000235:genetics; D012680:Sensitivity and Specificity; D012703:Serotyping
chemical_list: D004279:DNA, Viral; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments