Novel approach for differential diagnosis of HIV infections in the face of vaccine-generated antibodies: utility for detection of diverse HIV-1 subtypes

Abstract

Because increasing numbers of HIV vaccine candidates are being tested globally, it is essential to differentiate vaccine- from virus-induced antibodies. Most of the currently tested vaccines contain multiple viral components. As a result, many vaccine recipients give positive results in FDA-licensed HIV serodetection tests. We have identified conserved sequences in Env-gp41 and Gag-p6, which are recognized soon after infection but are not included in most HIV vaccine candidates. A new HIV serodetection assay, the HIV-SELECTEST, was established that distinguishes between vaccine-induced antibodies and seroconversion due to true HIV infections. It is important to make this assay globally relevant, because many clinical trials are conducted around the world where most HIV infections are due to non-B subtype HIV-1. Therefore, the current study examined the reactivity of plasma samples from >3,000 infections with diverse HIV subtypes worldwide. The HIV-SELECTEST performed at >99% specificity and sensitivity. Both recent and established infections with clades A, B, C, D, E, F, G, J, and CRFs were detected. Antibodies elicited by other vaccinations or infections endemic to the clinical trial sites did not react in this assay. Therefore, HIV-SELECTEST could be an important differential diagnostic tool for HIV vaccine trials, blood banks, and population screening worldwide.

FIGURE 1

Identification of HIV peptides for differential diagnosis of HIV infection during vaccine trials using gene-fragment phage display library. Various coding regions (ORFs) are depicted along their location in the HIV-1 genome. Alignment of HIV sequences displayed on the affinity-selected phage clones after the fourth round of panning with the HIV-1 genome sequence (pNL4−3 proviral clone) led to identification of the immunodominant regions recognized by antibodies generated soon after HIV infection. HIV genes/proteins that are part of current candidate HIV vaccines are also aligned to the HIV-1 genome. The peptide sequences that are being used in HIV-SELECTEST are shown. Numbers in each epitopic site indicate the position of the amino acid residues in the corresponding HIV-1–encoded proteins for the CON-OF-CONS alignment sequence in the Los Alamos database.

FIGURE 2

Seroreactivity of HIV-negative samples with HIV-SELECTEST peptides and determination of cut-off values. ELISA conditions are described in Methods. Reactivity of 600 HIV seronegative serum/plasma samples at 1:100 dilution with the peptides, p6 (panel A) and gp41 (panel B) is shown. Values on the X axis depicts the test specimen optical density (OD) with the HIV peptide in specific ELISA, and the Y axis represents the number of serum/plasma samples that exhibited a specific ELISA absorbance reading as shown on the X axis. C, Cut-off value for each peptide was determined as the mean absorbance + 5 SD obtained with HIV-seronegative samples. All data are represented as ratios between test specimen optical densities (OD) to cut-off absorbance (CO).

FIGURE 3

Comparison of seroreactivity of samples obtained from individuals infected with different HIV clades with HIV-SELECTEST peptides. ELISA conditions were described in METHODS. Reactivity of serum/plasma samples at 1:100 dilution with the peptides, p6 (panel A) and gp41 (panel B) from early and chronically infected individuals with different HIV subtypes is shown as indicated in each graph. Values on the X axis depict the ratios between test specimen optical densities (OD) to cut-off values based on the mean OD of 600 negative controls. The Y axis represents the number of samples that exhibited a specific reactivity ratio as shown on the X axis.