Transient or occult HIV-1 infection in high-risk adults (2001)

Infection of humans with HIV-1 results in a persistent, and usually progressive, infection, resulting in the loss of T helper lymphocytes and immunodeficiency. The diagnosis of HIV-1 infection has been based upon the detection of serum anti-HIV-1 antibodies by commercial enzyme immunoassays (EIA) employing denatured HIV antigens, with confirmation by Western blot or fixed-cell immunofluorescence. However, the transient appearance of either anti-HIV-1 antibodies in sera or polymerase chain reaction (PCR)-amplifiable HIV-1 DNA in peripheral blood mononuclear cells (PBMC), or both, has been described in a small number of adults at risk of HIV-1 infection [1]. Although conversion from HIV-1 antibody-positive to negative status is the norm for infants born to infected mothers once maternal IgG in the newborns has cleared, virological data based on HIV-1-DNA amplification by PCR in infant blood specimens at early and later time-points have indicated that clearance of HIV-1 may occur in some infected infants [2]. However, a recent analysis of HIV-1 sequences in some of those infants and their mothers [3] raised doubts about the suspected cases of transient HIV infection. The question thus remains as to whether transient HIV infection occurs. During a prospective study of seronegative sexual partners of known HIV-1-infected patients [4], we identified two apparently transiently infected individuals (nos. 1 and 2) using a live-cell immunofluorescence assay (IFA) as a test for serum anti-HIV-1 antibodies, HIV-1 DNA amplification by PCR, and HIV-1 culture from the PBMC of the subjects (Table 1). These two were ‘early-infected’ individuals previously reported (identified by their blood sample codes as R6 and R78) who possessed serum anti-HIV-1 antibodies that reacted with native HIV-1 antigens in live-cell IFA, but did not react in the standard Food and Drug Administration-approved denatured-antigen EIA or Western blot. These antibodies were later shown to react to conformational epitopes of HIV-1 Env (gp160) and Gag (p55) precursor proteins, and appear to be the first antibodies induced after HIV infection (submitted for publication).Table 1: Longitudinal assessment for the presence of HIV-1 and anti-HIV-1 antibodies in subjects at high risk of HIV-1 infection. On his first visit (R6), subject no. 1 possessed antibodies to HIV-1 that could be detected by live-cell IFA. HIV-1 DNA (env and gag regions) could be amplified by PCR in the R6 PBMC DNA, and virus was cultured from these PBMC (Table 1). Four months later (R28), his antibody titer in the IFA test was lower but positive, and two attempts were required to culture HIV-1 from his PBMC, which were also PCR-positive for HIV-1 env and gag regions. On his third visit (R63) 3 months later, he was negative in all tests. He was tested many times during the following 2 years, and his serum remained HIV antibody-negative in all tests (live-cell IFA, EIA/Western blot), and his PBMC were negative for HIV DNA by PCR. Ten and 11 years later, he was still seronegative, and repeated attempts failed to amplify both HIV-1 env and gag regions in his PBMC by nested PCR, indicating the clearance of HIV-1 from his circulation. Throughout the study, he had no HIV-1-associated illness, and his CD4 cell count was normal. His seropositive sexual partner died of AIDS in 1990. To verify that the stored blood specimens collected from subject no. 1 were correctly labelled, somatic genetic analyses (genomic identity testing examining nine independent polymorphisms) were performed by Lab Corp. of America, New York, USA, on two of his PBMC specimens [one when HIV was detected and the plasma possessed HIV antibody (R28), and the last one 11 years later (R703)] (Table 1). The results of the analysis showed that these two specimens were from the same individual (probability of mis-identity 2.9 × 10−11). Two independent tests for HIV (PCR for HIV DNA in PBMC and HIV antibody in the serum) were thus positive in two consecutive blood samples from patient no. 1, and one of these blood samples was definitively identified to be from the same individual as one taken 11 years later, which was PCR- and HIV antibody-negative. We wanted to perform somatic genetic analysis of blood sample R6, along with R28, but the PBMC of the R6 blood draw had been used in previous studies [4] and were not available for this study. Another, although less definitive, approach to demonstrate donor identity between blood samples drawn at different times would be to demonstrate that the viruses in the PBMC of R6 and R28 were very similar, and it would thus be likely that these samples were from the same individual. This approach was used extensively in the study by Frenkel et al.[3], to show relatedness between HIV sequences in reported transiently infected infants and the sequences in their mothers. Although we did not have the PBMC of the R6 blood draw, we did have the isolated virus from those PBMC, and the nucleotide sequences of the V1–V3 region of envelope of that virus were compared with those of the virus present in the PBMC of the R28 blood draw. These sequences were 97% homologous to each other, suggesting that the two consecutive samples were likely to be from the same individual (data not shown). We did not have PBMC or virus from subject no. 1's HIV-positive sexual partner, so we could not perform that comparison. The data on the second subject (no. 2) are not as extensive as those for subject no. 1, but we present them as some additional evidence for transient HIV infection in another individual. Subject no. 2 was negative for HIV-1 antibody by live-cell IFA, and virus was neither culturable nor amplifiable by PCR on his initial visit (R25). Five months later (R78), he was positive in all of these assays (Table 1). All tests were repeated on a blood specimen collected 3 months later (R112), which showed no indication of the presence of HIV-1 antibodies or HIV-1 DNA. Similarly to subject no. 1, his PBMC were negative for HIV-1 DNA (env and gag), and his serum showed no reactivity for HIV-1 antibodies in live-cell IFA, EIA, and Western blot assays over the next 9.5 years. His seropositive sexual partner died of AIDS in late 1989. Unfortunately, genetic identity testing could not be performed on blood sample R78 because the stored PBMC had been consumed in other studies [4]. In contrast to subjects nos. 1 and 2, one of the other high-risk individuals (no. 3) who possessed this ‘early anti-HIV antibody’ in our original report [4], progressed to EIA/Western blot seropositivity and eventually to AIDS (Table 1). The transient phenotype was thus not observed in all high-risk individuals we studied who possessed ‘early HIV antibody'. Also, two other high-risk subjects, nos. 4 and 5, who were longitudinally studied eventually seroconverted in live-cell IFA and later EIA/Western blot (Table 1). To characterize the HIV-1 strains isolated from subjects nos. 1 and 2 further, PCR-amplified HIV-1 DNA (env V1–V3 region) was sequenced and analysed by Blast Search (NCBI, National Institutes of Health, USA). The analyses revealed that the env regions from both were similar (maximum homology of 98% at the nucleotide level) to those of the non-syncytia-inducing, macrophage-topic strains of HIV-1 subtype B. In agreement with this result, these viruses could only infect a cell line (VB) expressing CCR5 as a co-receptor used by M-tropic strains, but not other T cell lines (H9, CEM, Jurkat, Molt-4) only expressing CXCR4 as a co-receptor, used by T tropic strains of HIV-1 (data not shown). Although previously suspected cases of transient HIV-1 infection in infants born to seropositive mothers have been controversial [3], a growing number of reports of HIV-1 infection in high-risk adults, followed by the disappearance of serological (EIA/Western blot) and virological (PCR) markers for infection should not be overlooked. The two subjects we report here were infected by M-tropic strains of HIV-1, which are isolated at early time-points, but they appeared to clear the virus within several months. If the virus was not cleared, it apparently became occult. We did not have lymph node biopsies to survey for HIV in those locations. It is not known how these individuals were able to clear or suppress the virus. Polymorphisms of chemokine receptors or chemokine genes, such as the CCR5Δ32, CCR2b-64I and SDF1α-3'A mutations, which are known to be associated with delayed disease progression [5], were absent in them (data not shown). It is important to understand why and how certain individuals can clear or suppress HIV infection to an undetectable level, whereas others cannot. If such clearance is immune response-mediated, insights into the mechanisms would probably be relevant for vaccine design. A second important issue that remains to be addressed is whether individuals demonstrating transient or occult HIV infection are able to transmit HIV-1 through sexual contact or blood transfusion during their transient virus-positivity phase. If so, more careful evaluation of current commercial EIA/Western blot-negative high-risk individuals is warranted [6]. Gautam K. Sahua Jenny J. Y. Chena Jason C. Huanga Keith M. Ramseyb Miles W. Cloyda