Background:  We previously observed the case of an HLA-B57⁺ long-term nonprogressor (LTNP; patient 4050), whose peripheral blood mononuclear cells (PBMC) accumulated HIV-1 subtype B proviruses defective in the env gene. However, after more than 10 years of control of infection, plasma viremia increased progressively, with a concomitant progressive loss of CD4⁺ T cells. We show here that patient 4050 was infected by a nef-defective HIV-1 variant initially and that loss of control was due to superinfection with a virus belonging to the same clade B.

Methods:  Nested polymerase chain reactions were used to amplify and clone 4 regions of the HIV-1 genome from proviral DNA:  a 1.2-kb fragment of the env gene encompassing most of the gp120 coding sequence, the complete nef and vif genes, and a fragment of the gag gene encompassing part of the P24 coding sequence. The ability of the plasma collected at entry in the cohort to neutralize either pseudotyped viruses expressing env clones from the surinfecting virus or 4 heterologous primary isolates of different clades was analyzed. Time evolution of CD8⁺ T cells responses specific for Gag and Nef proteins were assessed in interferon-gamma (IFN-g) ELIspot assays.

Results:  We compared env, nef, and vif sequences obtained from PBMC collected at entry in the cohort and 2 years later, when the patient had progressed. For each gene, 22 to 30 clones were sequenced at each time point. At entry, major genetic alterations were found in the 28 nef sequences and in 22 of 25 env sequences. In contrast, all clones recovered 2 years later were not altered genetically. Phylogenetic analyses of env, nef, and vif sequences clearly suggested the sequential infection of patient 4050 by 2 phylogenetically distinct strains of subtype B. Indeed, all nef and vif sequences and most of env sequences (22 of 25 clones) obtained from samples collected at entry clustered in a same branch and were phylogenetically distinct of those recovered 2 years later. At inclusion, plasma of this patient did not contain heterologous neutralizing antibodies and was not able to neutralize the superinfecting virus. Only moderate Gag-specific CD8⁺ T cell responses were observed, in particular against the B57-restricted TW10 immunodominant epitope.

Conclusions:  We report a case of superinfection in a LTNP initially infected with a nef-defective HIV-1 strain. These data showed the limited capacity of a natural infection with a putatively attenuated HIV-1 strain to generate efficient protective immune responses.