Role of Env in resistance of feline immunodeficiency virus (FIV)-infected cats to superinfection by a second FIV strain as determined by using a chimeric virus

Abstract

A more or less pronounced resistance to superinfection by a second strain of the infecting virus has been observed in many lentivirus-infected hosts. We used a chimeric feline immunodeficiency virus (FIV), designated FIVchi, containing a large part of the env gene of a clade B virus (strain M2) and all the rest of the genome of a clade A virus (a p34TF10 molecular clone of the Petaluma strain modified to grow in lymphoid cells), to gain insights into such resistance. FIVchi was infectious and moderately pathogenic for cats and in vitro exhibited the neutralization specificity of the env donor. The experiments performed were bidirectional, in that cats preinfected with either parental virus were challenged with FIVchi and vice versa. The preinfected animals were partially or completely protected relative to what was observed in naïve control animals, most likely due, at least in part, to the circumstance that in all the preinfecting/challenge virus combinations examined, the first and the second virus shared significant viral components. Based on the proportions of complete protection observed, the role of a strongly matched viral envelope appeared to be modest and possibly dependent on the time interval between the first and the second infection. Furthermore, complete protection and the presence of measurable neutralizing antibodies capable of blocking the second virus in vitro were not associated.

FIG. 1.

Schematic representation of the chimeric virus FIV_χ and of how it was prepared, starting from the parental viruses FIV_PET and FIV_M2. The locations of the primers used for the PCR assays in Fig. 2 are also shown.

FIG. 2.

Features of the PETgag-, PETenv-, M2gag-, and M2env-specific real-time PCR assays used in the study. (A) Forward primers (Fw), probes (Pr), and reverse primers (Rv) were designed based upon appropriately studied segments of gag or env of FIV_PET or FIV_M2 using Primer Express software (version 1.5; Applied Biosystems, Monza, Italy) and differed from the heterologous virus, as indicated under each oligonucleotide (nucleotide positions are based on FIV_PET clone p34TF10, GenBank accession number NC_001482) (53). Note that in order to improve selectivity and fulfill real-time probe requirements, the M2gag primers and probe were designed from the antisense strand. (B to D) Selectivity of the PET-specific (open bars) and M2-specific (closed bars) assays, as determined by quantitating FIV_PET and FIV_M2 plasmids; the DNAs examined were FIV_PET plasmids alone or FIV_M2 plasmids alone (indicated at the tops of the graphs) at the copy numbers shown in the abscissa (B), 1 × 10² or 1 × 10⁵ FIV_PET plasmids (indicated at the tops of the graphs) mixed with the numbers of FIV_M2 plasmids shown in the abscissa (C), and 1 × 10² or 1 × 10⁵ FIV_M2 plasmids (indicated at the tops of the graphs) mixed with the numbers of FIV_PET plasmids shown in the abscissa (D). These degrees of selectivity were conserved in the RT-PCR assays, as determined by performing similar experiments with RNA transcribed on the viral plasmids (data not shown). The error bars represent standard deviations from three independent experiments.

FIG. 3.

Set points in FIV_PET- or FIV_M2-preinfected cats and naïve controls at various times after i.v. challenge with FIV_χ. (A) Naïve cats. FIV_χ plasma viremia and proviral loads were determined with both the PETgag-specific assay (empty symbols) and the M2env-specific assay (solid symbols). Note that cats in this age range (27 months at challenge) showed no major spontaneous CD4⁺ T-lymphocyte count changes. (B) FIV_PET-preinfected cats. FIV_PET (dashed lines) and FIV_χ (continuous lines) plasma viremia and proviral loads determined with the PETenv- and M2env-specific assays, respectively. (C) FIV_M2-preinfected cats. FIV_M2 (dashed lines) and FIV_χ (continuous lines) plasma viremia and proviral loads determined with the M2gag- and PETgag-specific assays, respectively. Anti-FIV antibodies were determined by ELISA using whole FIV antigen.

FIG. 4.

Viral set points in FIV_PET- or FIV_M2-preinfected cats and naïve control cats at various times after intravaginal challenge with FIV_χ. (A) Naïve cats. FIV_χ plasma viremia and proviral loads determined with the M2env-specific assay. (B) FIV_PET-preinfected cats. FIV_PET (dashed lines) and FIV_χ (continuous lines) plasma viremia and proviral loads determined with the PETenv- and M2env-specific assays, respectively. (C) FIV_M2-preinfected cats. FIV_M2 (dashed lines) and FIV_χ (continuous lines) plasma viremia and proviral loads determined with the M2gag- and PETgag-specific assays, respectively.

FIG. 5.

Proviral loads of preinfecting viruses and challenge FIV_χ in selected tissues of cats at the end of the experiment shown in Fig. 4. (A) FIV_PET-preinfected cats examined with the M2env- and PETenv-specific assays. (B) FIV_M2-preinfected cats examined with the M2gag- and PETgag-specific assays. Open bars, preinfecting viruses; closed bars, FIV_χ.

FIG. 6.

Viral set points in FIV_χ-preinfected cats and naïve control cats after i.v. challenge with FIV_PET. (A) Naïve cats. FIV_PET plasma viremia and proviral loads determined with the PETenv-specific assay. (B) FIV_χ-preinfected cats. FIV_χ (dashed lines) and FIV_PET (continuous lines) plasma viremia and proviral loads determined with the M2env- and PETenv-specific assays, respectively.

FIG. 7.

Viral set points in FIV_χ-preinfected and naïve control cats after i.v. challenge with FIV_M2. (A and B) First challenge with 10 CID₅₀. (C and D) Second challenge with 30 CID₅₀. (A and C) Naïve cats. FIV_M2 plasma viremia and proviral loads determined with the M2gag-specific assay. (B and D) FIV_χ-preinfected cats. FIV_χ (dashed lines) and FIV_M2 (continuous lines) plasma viremia and proviral loads determined with the M2gag- and PETgag-specific assays, respectively.

FIG. 8.

Proviral loads of the preinfecting FIV_χ and of the challenge viruses in selected tissues of cats at the ends of the experiments shown in Fig. 6 and 7. (A) FIV_PET-challenged cats examined with the M2env- and PETenv-specific assays. (B) FIV_M2-challenged cats examined with the M2gag- and PETgag-specific assays. Open bars, FIV_χ; closed bars, challenge viruses.