Viral dynamics and evolution following primary HIV-1 infection

The knowledge of the early events during primary HIV-1 infection (PER) is of great importance for the understanding of the pathogenesis of HIV-1. A major problem in studies on PHI is the limited amount of data accessible from the early time-points. In our study, frequent blood sampling was done during the first weeks at PHI and thereafter in the chronic phase of infection, from both untreated (n = 8) and treated (n = 15) patients. The overall aim of this thesis was to obtain insights in the dynamics of viral load, diversity and divergence during PHI and the subsequent events in the chronic phase of infection, including the conceivable persistence of earlier viral quasispecies in resting CD4+ T-lymphocytes, a proposed reservoir for HIV-1. In untreated patients, a novel four phase model of the HIV-1 RNA dynamics during PHI was described. The initial rapid increase reached a peak one week after onset of symptoms, thereafter a rapid decay was followed at the third week by a reduced viral clearance before a seemingly steady state was reached after about two months. The rapid decay most probably reflects the onset of HIV-1 specific immune activity. The phase of reduced viral clearance may be due to the appearance of escape mutants and/or outgrow of minor transmitted variants. Nevertheless, the decay continued although at a much lower rate suggesting that the immune system continues to respond to the new viral variants resulting in a steady state between replication and clearance. The virus population was characterized mainly by direct sequencing, but also by cloning, of the partial gag, pol, and env genes. Although a completely homogeneous viral population was uncommon the viral diversity was restricted in most patients. However, in some patients a more heterogeneous pattern was identified, which could reflect that several HIV-1 strains were transmitted and/or that an antiretroviral immune response caused a rapid selection of a fit master sequence. In the chronic phase, three patterns of viral sequence variation were identified; a successive replacement of the original major viral sequence due to an accumulating divergence; a highly restricted viral divergence; and a rapid shift to a new major sequence already during the first two months. In the latter case the original major sequence reappeared later simultaneously with clinical deterioration and increased viral load. The reappearance could possibly be due to activation of founder viral quasispecies during perturbation of the immune system or reverse mutations related to a constrained evolution. Independent on the reasons, our results showed that the temporal variation of the viral population differs substantially between individuals. In patients in whom treatment was initiated at a very early stage, only a minor increase of the viral divergence was found during the first six months. Thereafter, no further evolution was detected. Also, the viral quasispecies obtained from resting T-cells, two years after initiation of therapy, were identical to the founder plasma viral population. A continuous decline of the proviral load was found corresponding to a half-life of six months, close to the decay rate of human memory T-lymphocytes. These data suggest that it is possible by early successful antiretroviral intervention to inhibit any significant viral evolution. The reservoir of latently infected resting T-cells in optimally treated patients is considered to be stable, as also confirmed by us in one treated patient. However, there is no knowledge about the replenishment of these cells in untreated and suboptimally treated subjects. Cells were therefore obtained both from patients followed from PHI as well as from multi-therapy experienced patients who previously had developed antiretroviral resistance. In the two patients not receiving any therapy during or after PHI, no original viral sequences were present in the resting memory T-cells almost six years later. Also, only a minority of the earlier identified resistance associated mutations was found in this cellular population of multi-therapy experienced patients. Instead, the viral population in resting T-cells clustered with the plasma sequences obtained at the same occasion, or before initiation on potent antiviral therapy, in five of the seven patients. These results indicate that the pool of latently infected resting T-cells in peripheral blood is continuously replenished even though it is possible that resistant or founder viral strains still are present in a minority of these cells