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1.
A full-length feline immunodeficiency virus NCSU1 (FIV-NCSU1) genome (JSY3) was cloned directly from FIV-NCSU1-infected feline CD4+ lymphocyte (FCD4E) genomic DNA and identified by PCR amplification with 5' long terminal repeat, gag, env, and 3' long terminal repeat primer sets. Supernatant from FCD4E cells cocultured with JSY3-transfected Crandell feline kidney (CrFK) cells was used as an inoculum. Cell-free JSY3 virus was cytopathogenic for FCD4E lymphocytes but did not infect CrFK cells in vitro. To determine in vivo infectivity and pathogenesis, six young adult specific-pathogen-free cats were inoculated with cell-free JSY3 virus. Provirus was detected at 2 weeks postinfection (p.i.) and was still detectable at 25 weeks p.i. as determined by gag region PCR-Southern blot analysis of peripheral blood mononuclear cell lysates. Infectious virus was recovered from peripheral blood mononuclear cells at 6 and 25 weeks p.i., and an antibody response to FIV was detected by 4 weeks. In the acute phase of infection, JSY3 provirus was found only in the CD4+ lymphocyte subset; however, by 14 weeks p.i., the greatest provirus burden was detected in B lymphocytes. All six cats were panlymphopenic at 2 weeks p.i., CD4+/CD8+ ratios were inverted by 6 weeks p.i., and five of the six cats developed lymphadenopathy by 10 weeks p.i. To determine if the JSY3 molecular clone caused immunodeficiency similar to that of the parental wild-type FIV-NCSU1, the cats were challenged with the low-virulence ME49 strain of Toxoplasma gondii at 29 weeks p.i. Five of six cats developed clinical signs consistent with generalized toxoplasmosis, and three of six cats developed acute respiratory distress and required euthanasia. Histopathologic examination of the severely affected cats revealed generalized inflammatory reactions and the presence of T. gondii tachyzoites in multiple tissues. None of the six age- and sex-matched specific-pathogen-free cats inoculated with only T. gondii developed clinical disease. Our results suggest that the pathogenesis of the molecularly cloned NCSU1 JSY3 is similar to that of wild-type FIV-NCSU1.  相似文献   

2.
Specific-pathogen-free cats, immunized with a 22-amino-acid synthetic peptide designated V3.3 and derived from the third variable region of the envelope glycoprotein of the Petaluma isolate of feline immunodeficiency virus (FIV), developed high antibody titers to the V3.3 peptide and to purified virus, as assayed by enzyme-linked immunoassays, as well as neutralizing antibodies, as assayed by the inhibition of syncytium formation in Crandell feline kidney cells. V3.3-immunized animals and control cats were challenged with FIV and then monitored for 12 months; V3.3 immunization failed to prevent FIV infection, as shown by virus isolation, anti-whole virus and anti-p24 immunoglobulin G antibody responses, and positive PCRs for gag and env gene fragments. Sequence analysis of the V3 region showed no evidence for the emergence of escape mutants that might have contributed to the lack of protection. The sera of the V3.3-hyperimmunized cats and two anti-V3.3 monoclonal antibodies neutralized FIV infectivity for Crandell feline kidney cells at high antibody dilutions but paradoxically failed to completely neutralize FIV infectivity at low dilutions. Moreover, following FIV challenge, V3.3-immunized animals developed a faster and higher antiviral antibody response than control cats. This was probably due to enhanced virus replication, as also suggested by quantitative PCR data.  相似文献   

3.
4.
We describe the histopathological, immunohistochemical, and molecular characterization of a lymphoma arising in a 7-year-old cat following experimental infection with feline immunodeficiency virus (FIV). The tumor was high grade and of B-cell lineage. The transformed cell had an immature phenotype (CD79a+, CD79b, CD21, immunoglobulin heavy and light chain negative), confirmed by antigen receptor gene analysis, which showed germ line configuration. Single-copy, clonally integrated FIV provirus was detected in tumor genomic DNA. FIV p24 antigen was not detected in tumor cells by immunostaining. This study provides the first evidence that the feline lentivirus may play a direct role in cell transformation under certain circumstances.  相似文献   

5.
The role of cellular immunity in the establishment and progression of immunosuppressive lentivirus infection remains equivocal. To develop a model system with which these aspects of the host immune response can be studied experimentally, we examined the response of cats to a hybrid peptide containing predicted T-and B-cell epitopes from the gag and env genes of feline immunodeficiency virus (FIV). Cats were immunized with an unmodified 17-residue peptide incorporating residues 196 to 208 (from gag capsid protein p24) and 395 to 398 (from env glycoprotein gp120) of the FIV Glasgow-8 strain by using Quil A as an adjuvant. Virus-specific lymphocytotoxicity was measured by chromium-51 release assays. The target cells were autologous or allogeneic skin fibroblasts either infected with recombinant FIV gag vaccinia virus or pulsed with FIV peptides. Effector cells were either fresh peripheral blood mononuclear cells or T-cell lines stimulated with FIV peptides in vitro. Cytotoxic effector cells from immunized cats lysed autologous, but not allogeneic, target cells when they were either infected with recombinant FIV gag vaccinia virus or pulsed with synthetic peptides comprising residues 196 to 205 or 200 to 208 plus 395. Depletion of CD8+ T cells, from the effector cell population abrogated the lymphocytotoxicity. Immunized cats developed an antibody response to the 17-residue peptide immunogen and to recombinant p24. However, no antibodies which recognized smaller constituent peptides could be detected. This response correlated with peptide-induced T-cell proliferation in vitro. This study demonstrates that cytotoxic T lymphocytes specific for FIV can be induced following immunization with an unmodified short synthetic peptide and defines a system in which the protective or pathological role of such responses can be examined.  相似文献   

6.
The objective of this study was to determine whether semen from male domestic cats infected with feline immunodeficiency virus (FIV) can transmit virus to females. Twelve inseminations were performed by an intrauterine laparoscopic technique with fresh or cryopreserved electroejaculates from asymptomatic males chronically infected with the NCSU1 strain of FIV. Of six inseminations performed with fresh semen, three resulted in infection of queens, as indicated by seroconversion, expression of FIV gag provirus in peripheral blood leukocytes, and reduced peripheral CD4+/CD8+ T-lymphocyte ratios. None of the six inseminates with thawed cryopreserved semen resulted in infection. Two infected queens and one uninfected queen became pregnant. Virus was not evident in the seven offspring. We conclude that FIV can be transmitted horizontally by artificial insemination with fresh semen.  相似文献   

7.
8.
Feline immunodeficiency virus (FIV) induces a disease state in the domestic cat that is similar to AIDS in human immunodeficiency virus (HIV)-infected individuals. As with HIV, FIV can be divided into primary and cell culture-adapted isolates. Adaptation of FIV to replicate and form syncytia in the Crandell feline kidney (CrFK) cell line is accompanied by an increase in the net charge of the V3 loop of the envelope glycoprotein, mirroring the changes observed in the V3 loop of HIV gp120 with the switch from a non-syncytium-inducing phenotype to a syncytium-inducing phenotype. These data suggest a common mechanism of infection with FIV and HIV. In this study, we demonstrate that cell culture-adapted strains of FIV are able to use the alpha-chemokine receptor CXCR4 for cell fusion. Following ectopic expression of human CXCR4 on nonpermissive human cells, the cells are able to fuse with FIV-infected feline cells. Moreover, fusion between FIV-infected feline cells and CXCR4-transfected human cells is inhibited by both anti-CXCR4 and anti-FIV antibodies. cDNAs encoding the feline CXCR4 homolog were cloned from both T-lymphoblastoid and kidney cell lines. Feline CXCR4 displayed 94.9% amino acid sequence identity with human CXCR4 and was found to be expressed widely on cell lines susceptible to infection with cell culture-adapted strains FIV. Ectopic expression of feline CXCR4 on human cells rendered the cells susceptible to FIV-dependent fusion. Moreover, feline CXCR4 was found to be as efficient as human CXCR4 in supporting cell fusion between CD4-expressing murine fibroblast cells and either HIV type 1 (HIV-1) or HIV-2 Env-expressing human cells. Previous studies have demonstrated that feline cells expressing human CD4 are not susceptible to infection with HIV-1; therefore, further restrictions to HIV-1 Env-dependent fusion may exist in feline cells. As feline and human CXCR4 support both FIV- and HIV-dependent cell fusion, these results suggest a close evolutionary link between FIV and HIV and a common mechanism of infection involving an interaction between the virus and a member of the seven-transmembrane domain chemokine receptor family of molecules.  相似文献   

9.
The orf-A (orf-2) gene of feline immunodeficiency virus (FIV) is a small open reading frame predicted to encode a 77-amino-acid protein that contains putative domains similar to those of the ungulate lentiviral Tat protein. Orf-A is reported to be critical for efficient viral replication in vitro and in vivo. A series of FIV-pPPR-derived proviruses with in-frame deletions and point mutations within orf-A were constructed and tested for replication in feline lymphoid cells. Orf-A mutant proviruses were also tested for viral gene and protein expression, viral particle formation, and virion infectivity. Deletions within orf-A severely restricted FIV replication in feline peripheral blood mononuclear cells (PBMC) and interleukin-2-dependent T-cell lines. In addition, substitutions of alanines for leucines in the putative leucine-rich domain, for cysteines in the putative cysteine-rich domain, and for a tryptophan at position 43 in Orf-A restricted the replication of FIV mutants. Deletions and point mutations in orf-A imposed a small effect or no effect on FIV long-terminal-repeat-driven viral gene expression and had no effect on viral protein expression. However, release of cell-free, virion-associated viral RNA in supernatants from cells transfected with orf-A mutant proviruses was severely restricted but was rescued by cotransfection with a wild-type Orf-A expression vector. In addition, virions derived from orf-A mutant proviruses expressed reduced infectivity for feline PBMC. Our findings suggest that Orf-A functions involve multiple steps of the FIV life cycle including both virion formation and infectivity. Furthermore, these observations suggest that Orf-A represents an FIV-encoded analog more similar to the accessory gene vpr, vpu, or nef than to the regulatory gene tat encoded by the primate lentiviruses.  相似文献   

10.
Feline immunodeficiency virus (FIV) causes AIDS-like symptoms in infected cats. Concanavalin A (ConA)-stimulated peripheral blood mononuclear cells (PBMC) from chronically FIV strain PPR-infected cats readily expressed FIV. In contrast, when PBMC from these animals were stimulated with irradiated, autologous antigen-presenting cells (APC), at least a 10-fold drop in viral production was observed. In addition to FIV-specific cytotoxic T lymphocytes, anti-FIV activity was demonstrated in the cell-free supernatants of effector T lymphocytes stimulated with APC. The FIV-suppressive activity was induced from APC-stimulated PBMC of either FIV-infected or uninfected cats but not from ConA-stimulated PBMC. Suppression of FIV strain PPR replication was observed for both autologous and heterologous feline PBMC, was dose dependent, and demonstrated cross-reactivity and cell specificity. It was also demonstrated that the anti-FIV activity originated from CD8(+) T lymphocytes and was mediated by a noncytolytic mechanism.  相似文献   

11.
The purpose of this study was to investigate the effect of chronic feline immunodeficiency virus (FIV) infection, and efficacy of marbofloxacin treatment, on 'Candidatus Mycoplasma haemominutum' infection. Six cats chronically infected with FIV-Glasgow8 (group A) and six FIV-free cats (group B) were infected with 'Candidatus M. haemominutum' on day 0 by intravenous inoculation of blood. From day 0 to 105 post-infection (pi), blood samples were collected for 'Candidatus M. haemominutum' and FIV provirus quantitative real-time polymerase chain reaction (PCR) and haematological examination. Three of the six cats in each of the groups were randomly selected to receive marbofloxacin treatment (2mg/kg PO SID) from day 49 to day 76 pi, with the remaining cats being untreated controls. Maximum 'Candidatus M. haemominutum' copy number was reached around day 30 pi. No overt cycling or marked variation in copy number was observed. No significant effect of FIV infection on 'Candidatus M. haemominutum' copy number kinetics or anaemia indices was found. No correlation was found between FIV provirus copy number and 'Candidatus M. haemominutum' copy number or haematological variables. Although marbofloxacin treatment was associated with a significant decrease in 'Candidatus M. haemominutum' copy number, the copy number plateaued during treatment, with no negative PCR results. Additionally, after termination of marbofloxacin treatment the copy numbers of the treated cats increased to reach levels similar to those of the untreated cats within 7-10 days. This study documents, for the first time, the infection kinetics and antibiotic responsiveness of 'Candidatus M. haemominutum' infection.  相似文献   

12.
Feline immunodeficiency virus (FIV) infects numerous wild and domestic feline species and is closely related to human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV). Species-specific strains of FIV have been described for domestic cat (Felis catus), puma (Puma concolor), lion (Panthera leo), leopard (Panthera pardus), and Pallas' cat (Otocolobus manul). Here, we employ a three-antigen Western blot screening (domestic cat, puma, and lion FIV antigens) and PCR analysis to survey worldwide prevalence, distribution, and genomic differentiation of FIV based on 3,055 specimens from 35 Felidae and 3 Hyaenidae species. Although FIV infects a wide variety of host species, it is confirmed to be endemic in free-ranging populations of nine Felidae and one Hyaenidae species. These include the large African carnivores (lion, leopard, cheetah, and spotted hyena), where FIV is widely distributed in multiple populations; most of the South American felids (puma, jaguar, ocelot, margay, Geoffroy's cat, and tigrina), which maintain a lower FIV-positive level throughout their range; and two Asian species, the Pallas' cat, which has a species-specific strain of FIV, and the leopard cat, which has a domestic cat FIV strain in one population. Phylogenetic analysis of FIV proviral sequence demonstrates that most species for which FIV is endemic harbor monophyletic, genetically distinct species-specific FIV strains, suggesting that FIV transfer between cat species has occurred in the past but is quite infrequent today.  相似文献   

13.
Transmission of pathogens from domestic animals to wildlife populations (spill-over) has precipitated local wildlife extinctions in multiple geographic locations. Identifying such events before they cause population declines requires differentiating spillover from endemic disease, a challenge complicated by a lack of baseline data from wildlife populations that are isolated from domestic animals. We tested sera collected from 12 ocelots (Leopardus pardalis) native to Barro Colorado Island, Panama, which is free of domestic animals, for antibodies to feline herpes virus, feline calicivirus, feline corona virus, feline panleukopenia virus, canine distemper virus, and feline immunodeficiency virus (FIV), typically a species-specific infection. Samples also were tested for feline leukemia virus antigens. Positive tests results were only observed for FIV; 50% of the ocelots were positive. We hypothesize that isolation of this population has prevented introduction of pathogens typically attributed to contact with domestic animals. The high density of ocelots on Barro Colorado Island may contribute to a high prevalence of FIV infection, as would be expected with increased contact rates among conspecifics in a geographically restricted population.  相似文献   

14.
Specific primers for the detection of the FIV provirus in peripheral blood mononuclear cells (PBMC) by seminested PCR (snPCR) were developed. Forty cats (patients from veterinary hospitals) were investigated for the presence of FIV serum antibodies by immunoblot and for the presence of provirus in PBMC by snPCR. Seventeen of the 40 examined samples (42.5%) showed the presence of antibodies against FIV. The total number of animals that were found positive in snPCR was 19 (47.5%). Fourteen of the seropositive animals (35%) were positive by snPCR whereas three seropositive animals (7.5%) turned out to be snPCR negative. Of twenty-three animals that were negative by immunoblot, five (12.5%) were found to be positive by snPCR.  相似文献   

15.
16.
The development of gene delivery vectors based on feline immunodeficiency virus (FIV) is an attractive alternative to vectors based on primate sources for the delivery of genes into humans. To investigate the requirements for efficient transduction of dividing and nondividing cells by vector particles based on FIV, a series of packaging and vector constructs was generated for which viral gene expression was minimized and from which unnecessary cis-acting sequences were deleted. Pseudotyped vector particles produced in 293T cells were used to transduce various target cells, including contact-inhibited human skin fibroblasts and growth-arrested HT1080 cells. FIV vectors in which the U3 promoter was replaced with the cytomegalovirus promoter gave rise to over 50-fold-higher titers than FIV vectors containing the complete FIV 5' long terminal repeat (LTR). Comparison of the transduction efficiencies of vectors containing different portions of the FIV Gag coding region indicates that at least a functional part of the FIV packaging signal (Psi) is located within an area which includes the 5' LTR and the first 350 bp of gag. Transduction efficiencies of vectors prepared without FIV vif and orf2 accessory gene expression did not differ substantially from those of vectors prepared with accessory gene expression in either dividing or nondividing cells. The requirement for FIV rev-RRE was, however, demonstrated by the inefficient production of vector particles in the absence of rev expression. Together, these results demonstrate the efficient transduction of nondividing cells in vitro by a multiply attenuated FIV vector and contribute to an understanding of the minimum requirements for efficient vector production and infectivity. In addition, we describe the ability of an FIV vector to deliver genes in vivo into hamster muscle tissue.  相似文献   

17.
Postnatal transmission of feline immunodeficiency virus (FIV) in neonates nursed by acutely infected mothers and infection resulting from oral inoculation of kittens with FIV were evaluated. Ten of 16 kittens nursed by four queens with FIV infection established immediately postpartum developed FIV infection. Five of 11 neonates orally administered cell-free FIV culture supernatant developed FIV infection. Kittens that developed FIV infection had greater proportions of CD4+ and Pan-T+ lymphocytes at birth than negative kittens. Infectious virus was recovered from the milk of acutely infected mothers. We conclude that FIV may be experimentally transmitted via milk from queens with acute infections and that oral administration of FIV to neonatal kittens results in infection.  相似文献   

18.
We devised an indicator gene for retrotransposition, nlsLacZRT, which contains the Escherichia coli lacZ gene fused to a nuclear location signal (nlsLacZ), engineered in such a way that the gene is expressed only if the structure in which it has been inserted transposes itself through an RNA intermediate. A cloned murine leukemia retrovirus with an ecotropic host range (Moloney murine leukemia virus), rendered defective by a large deletion encompassing the three viral gag, pol, and env open reading frames, was marked with this indicator gene and introduced by transfection into heterologous feline cells. No beta-galactosidase activity could be detected among the clonal cell population, unless the defective provirus was complemented in trans by the gag-pol gene products. Under these conditions, cell variants which disclosed an easily detectable nuclear blue coloration upon in situ 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside staining were observed. Fluorescence-activated cell sorting of the beta-galactosidase-positive cells, followed by Southern blot analysis, demonstrated an unambiguous correlation between nlsLacZRT activation and retrotransposition of the marked provirus. Transposition occurs at a high frequency (up to 10(-4) events per cell per generation), which is dependent on the level of expression of the gag-pol gene and is concomitant with the release of noninfectious retroviruslike particles which are the hallmarks, but not the intermediates, of the intracellular transposition process.  相似文献   

19.
Few data are available on the prevalence of feline viruses in European wildcats (Felis silvestris). Previous surveys have indicated that wildcats may be infected with the common viruses of domestic cats, apart from feline immunodeficiency virus (FIV). In the present study, 50 wildcats trapped throughout Scotland (UK) between August 1992 and January 1997 were tested for evidence of viral infection. All were negative for FIV by several serological or virological methods. By contrast, 10% of the cats were positive for feline leukemia virus (FeLV) antigen and infectious virus was isolated from 13% of a smaller subset. Of the wildcats tested for respiratory viruses, 25% yielded feline calicivirus (FCV) and although no feline herpesvirus was isolated, 16% of the samples had neutralizing antibodies to this virus. Antibodies to feline coronavirus (FCoV) were found in 6% of samples. Feline foamy virus (FFV) was an incidental finding in 33% of samples tested. This study confirms that wildcats in Scotland are commonly infected with the major viruses of the domestic cat, except for FIV.  相似文献   

20.
Encapsidation of retroviral RNA involves specific interactions between viral proteins and cis-acting genomic RNA sequences. Human immunodeficiency virus type 1 (HIV-1) RNA encapsidation determinants appear to be more complex and dispersed than those of murine retroviruses. Feline lentiviral (feline immunodeficiency virus [FIV]) encapsidation has not been studied. To gain comparative insight into lentiviral encapsidation and to optimize FIV-based vectors, we used RNase protection assays of cellular and virion RNAs to determine packaging efficiencies of FIV deletion mutants, and we studied replicative phenotypes of mutant viruses. Unlike the case for other mammalian retroviruses, the sequences between the major splice donor (MSD) and the start codon of gag contribute negligibly to FIV encapsidation. Moreover, molecular clones having deletions in this region were replication competent. In contrast, sequences upstream of the MSD were important for encapsidation, and deletion of the U5 element markedly reduced genomic RNA packaging. The contribution of gag sequences to packaging was systematically investigated with subgenomic FIV vectors containing variable portions of the gag open reading frame, with all virion proteins supplied in trans. When no gag sequence was present, packaging was abolished and marker gene transduction was absent. Inclusion of the first 144 nucleotides (nt) of gag increased vector encapsidation to detectable levels, while inclusion of the first 311 nt increased it to nearly wild-type levels and resulted in high-titer FIV vectors. However, the identified proximal gag sequence is necessary but not sufficient, since viral mRNAs that contain all coding regions, with or without as much as 119 nt of adjacent upstream 5' leader, were excluded from encapsidation. The results identify a mechanism whereby FIV can encapsidate its genomic mRNA in preference to subgenomic mRNAs.  相似文献   

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