Background: At present, there are no clinically reliable disease markers for chronic fatigue syndrome. DNA chip microarray technology provides a method for examining the differential expression of mRNA from a large number of genes. Our hypothesis was that a gene expression signature, generated by microarray assays, could help identify genes which are dysregulated in patients with post-infectious CFS and so help identify biomarkers for the condition.

The development of an effective vaccine against human immunodeficiency virus is considered to be the most practicable means of controlling the advancing global AIDS epidemic. Studies with the domestic cat have demonstrated that vaccinal immunity to infection can be induced against feline immunodeficiency virus (FIV); however, protection is largely restricted to laboratory strains of FIV and does not extend to primary strains of the virus. We compared the pathogenicity of two prototypic vaccine challenge strains of FIV derived from molecular clones; the laboratory strain PET F14 and the primary strain GL8 414 . PET F14 established a low viral load and had no effect on CD4؉ -or CD8 ؉ -lymphocyte subsets. In contrast, GL8 414 established a high viral load and induced a significant reduction in the ratio of CD4؉ to CD8 ؉ lymphocytes by 15 weeks postinfection, suggesting that PET F14 may be a low-virulence-challenge virus. However, during long-term monitoring of the PET F14 -infected cats, we observed the emergence of variant viruses in two of three cats. Concomitant with the appearance of the variant viruses, designated 627 W135 and 628 W135, we observed an expansion of CD8 ؉ -lymphocyte subpopulations expressing reduced CD8 ␤-chain, a phenotype consistent with activation. The variant viruses both carried mutations that reduced the net charge of the V3 loop (K409Q and K409E), giving rise to a reduced ability of the Env proteins to both induce fusion and to establish productive infection in CXCR4-expressing cells. Further, following subsequent challenge of naïve cats with the mutant viruses, the viruses established higher viral loads and induced more marked alterations in CD8؉ -lymphocyte subpopulations than did the parent F14 strain of virus, suggesting that the E409K mutation in the PET F14 strain contributes to the attenuation of the virus.

It has been shown that cats can be protected against infection with the prototypic Petaluma strain of feline immunodeficiency virus (FIV PET ) using vaccines based on either inactivated virus particles or replicationdefective proviral DNA. However, the utility of such vaccines in the field is uncertain, given the absence of consistent protection against antigenically distinct strains and the concern that the Petaluma strain may be an unrepresentative, attenuated isolate. Since reduction of viral pathogenicity and dissemination may be useful outcomes of vaccination, even in the absence of complete protection, we tested whether either of these vaccine strategies ameliorates the early course of infection following challenge with heterologous and more virulent isolates. We now report that an inactivated virus vaccine, which generates high levels of virus neutralizing antibodies, confers reduced virus loads following challenge with two heterologous isolates, FIV AM6 and FIV GL8 . This vaccine also prevented the marked early decline in CD4/CD8 ratio seen in FIV GL8 -infected cats. In contrast, DNA vaccines based on either FIV PET or FIV GL8 , which induce cell-mediated responses but no detectable antiviral antibodies, protected a fraction of cats against infection with FIV PET but had no measurable effect on virus load when the infecting virus was FIV GL8 . These results indicate that the more virulent FIV GL8 is intrinsically more resistant to vaccinal immunity than the FIV PET strain and that a broad spectrum of responses which includes virus neutralizing antibodies is a desirable goal for lentivirus vaccine development.

Definition of the immunological mechanisms involved in protective immunity against lentiviral infections is crucial to the development of an effective vaccine. The induction of gag- and env-specific cell-mediated immune responses was studied in cats following vaccination with whole inactivated feline immunodeficiency virus (FIV). Cats were immunized by inoculation with three doses of paraformaldehyde-inactivated FIV, derived from the feline lymphoid cell line, FL-4, which is persistently infected with the Petaluma isolate of FIV. Autologous or allogeneic skin fibroblasts either infected with recombinant FIV gag- or env-vaccinia virus or pulsed with FIV env peptides were used as targets in chromium-51 release assays. Effector cells were fresh peripheral blood mononuclear cells. Following the third immunization, all vaccinated cats, but none of the control cats immunized with adjuvant alone, had detectable FIV env-specific lymphocytotoxicity in their peripheral blood. Two cats also exhibited gag-specific activity. There was no recognition of either allogeneic skin fibroblasts infected with recombinant vaccinia virus or autologous target cells infected with wild-type vaccinia virus, indicating the specificity and MHC-restricted nature of the response. Vaccinated cats, but not control cats, were protected from challenge with the homologous Petaluma isolate of FIV. Partial epitope mapping of the env-specific cytotoxic response was performed using overlapping 10-amino acid peptides from the env V3 domain of FIV. This response appeared to be directed at env peptide 1 (RAISSWKQRN) and env peptide 3 (QRNRWEWRPD), which lie adjacent to a beta-turn within the V3 domain.(ABSTRACT TRUNCATED AT 250 WORDS)

The appearance of non-cytolytic T cells that suppressed feline immunode®ciency virus (FIV) replication in vitro, and FIV-speci®c cytotoxic T cell (CTL) responses was compared in a group of seven, speci®c pathogen free (SPF) domestic cats following primary infection with the Glasgow 8 isolate of FIV (FIV ). FIV proviral burdens were quanti®ed in the blood and lymphoid tissues by real-time PCR. Non-cytolytic T cell suppression of FIV replication was measured by co-cultivating lymphoblasts prepared from the cats at different time-points during infection with FIV-infected MYA-1 cells in vitro. Non-cytolytic suppressor activity was detected as early as 1 week after infection, and was evident in all the lymphoid tissues examined. Further, this activity was present in subpopulations of T cells in the blood with normal (CD8 hi ) or reduced (CD8 lo ) expression of the CD8 molecule, and temporal modulations in non-cytolytic suppressor activity were unrelated to the circulating CD8 T cell numbers. Virus-speci®c CTL responses, measured by 51 Cr release assays, were not detected until 4 weeks after infection, with the emergence of FIV-speci®c effector CTLs in the blood. Throughout infection the response was predominantly directed towards FIV Gag-expressing target cells, and by 47 weeks after infection CTL responses had become localised in the lymph nodes and spleen. The results suggest that both non-cytolytic T cell suppression of FIV replication and FIV-speci®c CTL responses are important cellular immune mechanisms in the control of FIV replication in infected asymptomatic cats. #