Feline Immunodeficiency Virus-Infected Cat Sera Associated with the Development of Broad Neutralization Resistance In Vivo Drive Similar Reversions In Vitro

Abstract

We previously reported that, upon reinoculation into cats, a neutralization-sensitive, tissue culture-adapted strain of feline immunodeficiency virus constantly reverted to the broad neutralization resistance typical of primary virus isolates and identified residue 481 in the V4 region of the surface glycoprotein as a key determinant of the reversion. Here, we found that well-characterized immune sera, obtained from cats in which such reversion had occurred, selected in tissue culture in favor of virus variants that also had a neutralizationresistant phenotype and had amino acid 481 changed, thus indicating that the host's humoral immune response is capable of driving the reversion in the absence of other intervening factors. In contrast, a second group of immune sera, elicited by a virus variant that had already reverted to neutralization resistance in independent cats, induced the emergence of escape mutants lacking broad neutralization resistance and neutralized fewer virus variants. It is proposed that the viral variants used to produce the two sets of sera may have generated different antibody repertoires.Naturally occurring lentiviruses are only occasionally inhibited by sera of infected subjects in in vitro neutralization assays, and this property is currently the focus of intensive investigation since it is considered instrumental for virus persistence and pathogenicity as well as a formidable obstacle in the development of effective prophylactic vaccines (5, 16). With regard to the latter point, several immunogens have indeed been shown to protect against neutralization-sensitive (NS), tissue culture-adapted (TCA) laboratory lentiviruses but not against isolates recently cultured from infected hosts (11,25). In spite of considerable recent advances on the structural features that permit wild-type lentiviruses to resist potentially neutralizing antibodies (7,8,13,15,20,27), what determines conservation of this important viral attribute of wild-type lentiviruses is poorly understood. In fact, although it seems feasible that continued exposure to antiviral antibody is important (4), this has never been formally demonstrated. In principle, alternative or adjunctive intervening factors include other innate and adaptive immune effectors and the need to conserve the usage of certain receptor-coreceptor systems which determine virus tropism for specific cell types (17).Feline immunodeficiency virus (FIV) is an important pathogen of domestic cats and, due to extensive similarities with human immunodeficiency virus type 1 (HIV-1), is a valuable model for AIDS studies (10,18,24). In particular, the neutralization properties of FIV closely resemble those of HIV-1, including that TCA strains are readily inhibited by immune sera in vitro whereas fresh isolates exhibit a generalized resistance to antibody-mediated neutralization (1, 9). In previous studies, we observed that, upon reinoculation into cats, an exquisitely NS laboratory TCA strain of FIV regained the broadly neutralization-resistant (NR) phenotype typical...

Neutralization of feline immunodeficiency virus by antibodies targeting the V5 loop of Env

“…V5 was mapped by using synthetic peptides for a possible linear epitope based on the sequence of the 19k1-560 strain (Siebelink et al, 1995), and it was demonstrated that a single mutation at position 560 may be involved concurrently with another mutation at 483 (V4 loop) in one or more conformational epitopes, but no linear epitope was mapped in V5. Further, two independent mutations have been reported that resulted in the creation of potential N-linked glycosylation sites in V4 (K481N) and V5 (S557N) and which contributed to the conversion from a neutralization-sensitive to a neutralization-resistant phenotype (Giannecchini et al, 2001;Pistello et al, 2003). In comparison, the conversion from a weakly neutralized or neutralization-resistant to a strongly neutralized phenotype described herein was mediated by mutation of V5 alone.…”

“…Previous studies have demonstrated the importance of V5 in virus neutralization (Giannecchini et al, 2001;Pistello et al, 2003;Siebelink et al, 1995). V5 was mapped by using synthetic peptides for a possible linear epitope based on the sequence of the 19k1-560 strain (Siebelink et al, 1995), and it was demonstrated that a single mutation at position 560 may be involved concurrently with another mutation at 483 (V4 loop) in one or more conformational epitopes, but no linear epitope was mapped in V5.…”

“…By the time functional NAbs develop, virus variants that can escape the monospecific serum may have already evolved in vivo. The solution to this dilemma would appear to lie in the identification of framework determinants in Env that induce antibodies that are broadly active, refocusing the immune response either by silencing the immunodominant (but ultimately ineffectual) determinants in V4 and V5, or by presenting them in a such a way as to induce antibodies that recognize structures or motifs that are difficult or impossible for the virus to vary.Previous studies have demonstrated the importance of V5 in virus neutralization (Giannecchini et al, 2001;Pistello et al, 2003;Siebelink et al, 1995). V5 was mapped by using synthetic peptides for a possible linear epitope based on the sequence of the 19k1-560 strain (Siebelink et al, 1995), and it was demonstrated that a single mutation at position 560 may be involved concurrently with another mutation at 483 (V4 loop) in one or more conformational epitopes, but no linear epitope was mapped in V5.…”

Evolution of Two Amino Acid Positions Governing Broad Neutralization Resistance in a Strain of Feline Immunodeficiency Virus over 7 Years of Persistence in Cats

“…Also, the same molecular clones showed differences in terms of the ability of intact lectin-captured virions to remove Env-binding antibodies from immune serum from a cat that had been infected over the long term, thus showing that the impacts of the changes responsible for BNR acquisition and maintenance were also detectable at the level of antibody binding by the viral surface. In vitro immune selection studies showed that antibodies in the sera of cats in which the reversion of virus to BNR had occurred drove BNR and amino acid changes in the study virus similar to those that viruses in infected cats underwent (10). Together with those findings, the present findings support a model in which the uniform and sequentially ordered fixation of BNR-conferring amino acids at positions 481 and 557 was driven by antibody specificities that developed in the same temporal order in all animals, although at different times p.i.…”

“…All clones were propagated in MBM cells. As discussed below, the clones were also compared for replication capacity in an in vitro multicycle replication assay in the presence or absence of antiviral antibodies, as described previously (10). In brief, the clones were seeded at a multiplicity of infection of 0.0001 onto 24-well plates containing 2 ϫ 10 6 MBM cells in complete growth medium supplemented with 2% pooled normal cat serum (NCS) or selected immune sera.…”