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ABSTRACT: Serodiagnosis of feline immunodeficiency virus (FIV) is complicated by the use of a formalin-inactivated whole-virus FIV vaccine. Cats respond to immunization with antibodies indistinguishable from those produced during natural infection by currently available diagnostic tests, which are unable to distinguish cats that are vaccinated against FIV, infected with FIV, or both.
An enzyme-linked immunosorbent assay (ELISA) detecting antibodies against formalin-treated FIV whole virus and untreated transmembrane peptide will distinguish uninfected from infected cats, regardless of vaccination status.
Blood samples were evaluated from uninfected unvaccinated cats (n = 73 samples), uninfected FIV-vaccinated cats (n = 89), and FIV-infected cats (n = 102, including 3 from cats that were also vaccinated).
The true status of each sample was determined by virus isolation. Plasma samples were tested for FIV antibodies by a commercial FIV diagnostic assay and an experimental discriminant ELISA.
All samples from uninfected cats were correctly identified by the discriminant ELISA (specificity 100%). Of the samples collected from FIV-infected cats, 99 were correctly identified as FIV-infected (sensitivity 97.1%).
With the exception of viral isolation, the discriminant ELISA is the most reliable assay for diagnosis of FIV. A practical strategy for the diagnosis of FIV infection would be to use existing commercial FIV antibody assays as screening tests. Negative results with commercial assays are highly reliable predictors for lack of infection. Positive results can be confirmed with the discriminant ELISA. If the discriminant ELISA is negative, the cat is probably vaccinated against FIV but not infected. Positive results are likely to represent infection.
Journal of Veterinary Internal Medicine 02/2008; 22(2):330-4. · 1.99 Impact Factor
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ABSTRACT: Feline immunodeficiency virus (FIV) isolates from domestic cats have been classified into five subtypes, designated A, B, C, D and E. Although many FIV-infected cats may have frequent contact with multiple strains of FIV, they usually become infected with a single FIV subtype. In the present study, we demonstrate that peripheral blood mononuclear cells (PBMC) of FIV infected cats were resistant to exogenous FIV (second virus) replication in vitro and that the resistance of these PBMC was mediated by CD8+ T cells. In cats with a low anti-FIV activity of CD8+ T cells, the proviral DNA of the second virus inoculated into PBMC was detected intracellularly, and both the second and the originally infecting strain (original virus) were produced in the culture supernatant. In contrast, in cats with a high anti-FIV activity of CD8+ T cells, both the proviral DNA of the second virus and the original virus were detected in PBMC intracellularly, but neither virus was produced in the culture supernatant. However, when PBMCs from these cats were depleted of CD8+ T cells, the RNA of both viruses was detected in the culture supernatant. These results suggest that CD8+ T cells inhibit the late phase of FIV replication after viral integration. Moreover, the inhibition was also effective against FIV strains of different subtypes from that of the original strain. It appears that the CD8+ T cell-mediated immune response plays important roles in the maintenance of an asymptomatic state in FIV-infected cats and their resistance to superinfection.
Archives of Virology 09/2002; 147(8):1517-29. · 2.11 Impact Factor
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ABSTRACT: To evaluate the immunogenicity and efficacy of an inactivated dual-subtype feline immunodeficiency virus (FIV) vaccine.
Specific-pathogen-free cats were immunized with dual-subtype (subtype A FIV(Pet) and subtype D FIV(Shi)) vaccine and challenged with either in vivo- or in vitro-derived FIV inocula.
Dual-subtype vaccinated, single-subtype vaccinated, and placebo-immunized cats were challenged within vivo-derived heterologous subtype B FIV(Bang) [10--100 50% cat infectious doses (CID(50))], in vivo-derived homologous FIV(Shi)(50 CID(50)), and in vitro- and in vivo-derived homologous FIV(Pet)(20--50 CID(50)). Dual-subtype vaccine immunogenicity and efficacy were evaluated and compared to single-subtype strain vaccines. FIV infection was determined using virus isolation and proviral PCR of peripheral blood mononuclear cells and lymphoid tissues.
Four out of five dual-subtype vaccinated cats were protected against low-dose FIV(Bang) (10 CID(50)) and subsequently against in vivo-derived FIV(Pet) (50 CID(50)) challenge, whereas all placebo-immunized cats became infected. Furthermore, dual-subtype vaccine protected two out of five cats against high-dose FIV(Bang) challenge (100 CID(50)) which infected seven out of eight single-subtype vaccinated cats. All dual-subtype vaccinated cats were protected against in vivo-derived FIV(Pet), but only one out of five single-subtype vaccinated cats were protected against in vivo-derived FIV(Pet). Dual-subtype vaccination induced broad-spectrum virus-neutralizing antibodies and FIV-specific interferon-gamma responses along with elevated FIV-specific perforin mRNA levels, suggesting an increase in cytotoxic cell activities.
Dual-subtype vaccinated cats developed broad-spectrum humoral and cellular immunity which protected cats against in vivo-derived inocula of homologous and heterologous FIV subtypes. Thus, multi-subtype antigen vaccines may be an effective strategy against AIDS viruses.
AIDS 08/2001; 15(10):1225-37. · 6.24 Impact Factor
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ABSTRACT: The effect of a CXC-chemokine, stromal cell derived factor-1 (SDF-1), on the replication of divergent strains of feline immunodeficiency virus (FIV) was examined in order to identify the mechanism of cell entry of FIV. A chemotaxis assay, using a modified Boyden chamber method, confirmed the biological activity of recombinant human (rh) SDF-1 for a feline T-lymphoid cell line (Kumi-1). The viral replication of FIV, as measured by the reverse transcriptase (RT) activity in the culture supernatant, was significantly suppressed by addition of rhSDF-1 in a dose-dependent manner in Kumi-1 cells. Furthermore, PCR analysis of the FIV proviral genome indicated that the inhibitory effect of rhSDF-1 on the replication of FIV in Kumi-1 cells was due to the inhibitory effect in the early event of replication. The inhibitory effect on viral replication by exogenous rhSDF-1 was shown for four divergent FIV isolates of subtypes A, B, and D in Kumi-1 cells.
Veterinary Immunology and Immunopathology 06/2000; 74(3-4):303-14. · 2.08 Impact Factor
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ABSTRACT: CD8+ T cells in FIV-infected cats inhibit feline immunodeficiency virus (FIV) replication by producing a soluble factor(s). In the present study, four SPF cats were experimentally infected with FIV. The period during which the anti-FIV activity of CD8+ T cells became detectable was investigated, and the presence or absence of this activity in the lymph nodes and spleen was examined. Furthermore, we investigated which step(s) of the FIV replication cycle are affected by this antiviral activity. This antiviral activity became detectable five weeks after FIV infection in early cases, and it was simultaneous with or one week after the induction of humoral immunity. All cats having CD8+ T cells with anti-FIV activity in the peripheral blood also possessed CD8+ T cells with anti-FIV activity in the lymph nodes. In contrast, CD8+ T cells from the spleens of some, but not all cats showed anti-FIV activity. CD8+ T cell-depleted peripheral blood mononuclear cells were cultured and reconstituted with CD8+ T cells on day 12 of culture after confirming FIV replication. The number of FIV proviral DNA copies in the cells did not change, but the amount of FIV p24 antigen production in the culture supernatant and the number of FIV mRNA copies in the cells decreased. These findings suggested that CD8+ T cell anti-FIV activity acts at the level of FIV mRNA synthesis from the FIV proviral DNA, inhibiting FIV replication by a non-cytolytic mechanism.
Archives of Virology 02/2000; 145(12):2525-38. · 2.11 Impact Factor
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ABSTRACT: Feline infectious peritonitis virus (FIPV) infection of feline macro-phages is enhanced by mouse anti-FIPV monoclonal antibody (MAb). This anti-body-dependent enhancement (ADE) of FIPV infection is dependent on mouse MAb subclass, and MAb of IgG2a subclass has a strong ADE activity. Furthermore, MAb showing strong neutralizing activity in Felis catus whole fetus (fcwf-4) cells and Crandell feline kidney (CrFK) cells shows strong enhancing activity in feline macrophages, indicating that the neutralizing epitope and the enhancing epitope are closely related. In this study, we prepared MAb FK50-4 that showed a strong neutralizing activity in feline macrophages, despite the fact that the MAb belonged to the IgG2a subclass. However, MAb FK50-4 did not exhibit neutralizing activity in CrFK cells or fcwf-4 cells, thus showing a very unusual property. MAb FK50-4 recognized FIPV small integral membrane glycoprotein (M protein). Even when feline macrophages were pretreated with MAb FK50-4 prior to FIPV inoculation, this antibody prevented FIPV infection. This reaction disappeared after treatment of FK50-4 with protein A. The neutralizing activity of FK50-4 was also effective on feline macrophages after the cells were inoculated with FIPV. These findings indicated that the FIPV replication mechanism differs between feline macrophages and CrFK/fcwf-4 cells and that a neutralizing epitope that can prevent FIPV infection of feline macrophages after viral absorption is present on M protein.
Archives of Virology 02/2000; 145(1):1-12. · 2.11 Impact Factor
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ABSTRACT: The type II feline infectious peritonitis virus (FIPV) epitopes for neutralizing and enhancing antibodies are present on large spike glycoprotein (S) protein. In this study, we established monoclonal antibody-resistant mutant viruses resistant to three different monoclonal antibodies with neutralizing activity in Felis catus whole fetus cells and enhancing activity in feline macrophages, recognizing distinct epitopes on type II FIPV S protein. By comparing the nucleotide sequences of these mutant viruses with that of wild-type virus, we attempted to identify the neutralizing epitopes. The mutations were localized in the region of amino acid residues from 480 to 649 from the N terminal of the S protein.
Journal of Veterinary Medical Science 09/1999; 61(8):935-8. · 0.85 Impact Factor
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ABSTRACT: Four types of commercially available feline calicivirus (FCV) vaccine were compared in terms of their efficacy on the basis of the ability of the sera of specific-pathogen-free cats immunized by two injections of each type of vaccine to neutralize FCV field isolates. Each vaccine immune serum neutralized relatively well strains F4, F9, and 255, which were FCV laboratory strains. As to 36 strains of field isolates, however, vaccines A, B, C, and D immune sera did not neutralize 18-20 of the strains (50.0%-55.6%), 19-22 of the strains (52.8%-61.1%), 22-25 of the strains (61.1%-69.4%), and 8-16 of the strains (22.2%-44.4%), respectively. These results indicate that there is much difference in neutralizing antigenicity between the existing vaccine strains and the FCV strains that are prevalent in Japan, suggesting the need for improvement of FCV vaccines.
Journal of Veterinary Medical Science 04/1999; 61(3):299-301. · 0.85 Impact Factor
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ABSTRACT: Feline immunodeficiency virus (FIV) is more readily isolated from CD8+ T cell-depleted peripheral blood mononuclear cells (PBMC) of FIV-infected cats than from unfractionated PBMC cultures. However, it is not known whether feline CD8+ T cells down-regulate FIV expression by direct interaction with FIV-infected cells or via a soluble mediator. Furthermore, it is not known whether this anti-FIV activity involves a lytic or non-lytic mechanism. In the present study, we demonstrated that autologous and allogeneic CD8+ T cells from asymptomatic FIV-infected cats inhibited the replication of FIV in CD8+ T cell-depleted PBMC cultures in a dose-dependent manner. The inhibitory effect was mediated by a non-lytic mechanism, and was not dependent on direct cell-to-cell contact: an inhibitory effect was exerted by CD8+ T cells across a semi-permeable membrane, and an inhibitory activity was also present in cell-free supernatants from CD8+ T cells. These results suggest that this suppressive effect is mediated, at least in part, by soluble factors produced by CD8+ T cells.
Journal of General Virology 12/1998; 79 ( Pt 11):2729-35. · 3.36 Impact Factor
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ABSTRACT: Infection of the type II feline infectious peritonitis virus (FIPV) strain 79-1146 to primary feline alveolar macrophages and human monocyte cell line U937 was enhanced by the sera of cats experimentally infected with the 79-1146 strain, but not those of cats infected with KU-2 or UCD-1 strain of type I FIPV. The experiments using sera of cats with feline infectious peritonitis (FIP) and of cats naturally infected with feline coronavirus (FCoV) revealed that infection of the FIPV 79-1146 strain to the U937 cells was enhanced only by the sera of cats infected with type II FIPV or feline enteric coronavirus. The samples positive for antibody-dependent enhancement (ADE) activity had high neutralizing antibody titers against the FIPV 79-1146 strain and the samples negative for ADE activity had low neutralizing antibody titers. These findings support the previous results where a monoclonal antibody with neutralizing activity had high ADE activity, suggesting that there was a close relationship between the neutralization and enhancement sites. And then it is also suggested that ADE of infection is likely to be induced by re-infection with the same serotype of virus in type II FIPV infection. Furthermore, U937 cells are considered useful and can be substituted for the feline macrophages for determining ADE of FIPV-infection.
Journal of Veterinary Medical Science 02/1998; 60(1):49-55. · 0.85 Impact Factor
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ABSTRACT: Feline infectious peritonitis viruses (FIPVs) are classified into type I and type II serogroups. Here, we report that feline aminopeptidase N (APN), a cell-surface metalloprotease on the intestinal, lung and kidney epithelial cells, is a receptor for type II FIPV but not for type I FIPV. A monoclonal antibody (MAb) R-G-4, which blocks infection of Felis catus whole fetus (fcwf-4) cells by type II FIPV, was obtained by immunizing mice with fcwf-4 cells which are highly susceptible to FIPV. This MAb also blocked infection of fcwf-4 cells by type II feline enteric coronavirus (FECV), canine coronavirus (CCV), and transmissible gastroenteritis virus (TGEV). On the other hand, it did not block infection by type I FIPVs. MAb R-G-4 recognized a polypeptide of relative molecular mass 120-130 kDa in feline intestinal brush-border membrane (BBM) proteins. The polypeptide possessed aminopeptidase activity, and the first 15 N-terminal amino acid sequence was identical to that of the feline APN. Feline intestinal BBM proteins and the polypeptide reacted with MAb R-G-4 (feline APN) inhibited the infectivity of type II FIPV, type II FECV, CCV and TGEV to fcwf-4 cells, but did not inhibit the infectivity of type I FIPVs.
Archives of Virology 02/1998; 143(5):839-50. · 2.11 Impact Factor
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ABSTRACT: Genetic subtyping of feline immunodeficiency virus (FIV) was carried out by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. A 329-base pair fragment in the FIV gag gene was amplified by nested PCR, then digested with restriction enzymes, HindIII, PvuII and BamHI. Using these restriction enzymes, FIV isolates belonging to subtypes A, B and D, which had been classified on the basis of the env gene V3-V5 sequence, could be discriminated. Genetic subtypes of FIV prevalent in Japan were investigated using the gag-nested PCR-RFLP analysis. Of 88 FIV-infected cats, PCR products of 70 cats showed a subtype B RFLP pattern (digestion only with PvuII), those of nine cats had a subtype D RFLP pattern (digestion only with BamHI), and those of seven cats had a subtype A RFLP pattern (digestion only with HindIII). The PCR products of the remaining two cats had subtype A and B RFLP patterns (digestion with both HindIII and PvuII). The digestion pattern of the gag-nested PCR-RFLP analysis was unchanged after in vivo passages of the virus. These results suggest that the gag-nested PCR-RFLP analysis is useful as a simple method for FIV genetic subtyping.
Journal of Virological Methods 01/1998; 70(1):107-11. · 2.01 Impact Factor
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ABSTRACT: Feline immunodeficiency virus (FIV) is classified into five genotypes, A, B, C, D and E, based on the phylogenetic analysis of the env V3-V5 region. However, whether there is correlation between phylogenetic and antigenic diversities remains unknown. In this study, we examined the cross virus neutralization of FIV genotypes A through E by sera from cats infected with a single genotype. The results indicated some relationships between phylogenetic genotype and neutralization serotype, and that cross-clade virus neutralization is possible. For example, serum from a cat infected with genotype E virus neutralized all five FIV genotypes. Our results suggest that the FIV subtyping according to the sequence diversity is partially reflected by antigenic diversity and serum neutralization.
Archives of Virology 01/1998; 143(1):157-62. · 2.11 Impact Factor
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ABSTRACT: Dual-subtype feline immunodeficiency virus (FIV) vaccine, consisting of inactivated cells infected with subtypes A (Petaluma strain) and D (Shizuoka strain), was developed and tested for its vaccine efficacy against FIV infection in specific pathogen free (SPF) cats. Animals were monitored for proviral DNA by FIV-specific PCR and for FIV-specific antibody profiles by ELISA and virus-neutralization assays. In addition, blood from challenged cats was inoculated into naive SPF cats to confirm the viral status of the vaccinated cats. All cats immunized with Petaluma vaccine alone were protected against homologous Petaluma challenge, but only one of four cats was protected against heterologous Shizuoka challenge. More importantly, all cats immunized with the dual-subtype vaccine were protected against both Petaluma and Shizuoka challenges. These results suggest that a multi-subtype vaccine approach may provide the broad-spectrum immunity necessary for vaccine protection against strains from different subtypes.
Veterinary Microbiology 12/1997; 58(2-4):155-65. · 3.33 Impact Factor
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ABSTRACT: Six strains of feline immunodeficiency virus (FIV) classified into subtypes A, B and D were examined by cross-neutralization test using Kumi-1 cells (CD4+, CD8+, and CD9+), an interleukin-2 dependent feline T-lymphocyte cell line. Neutralizing activities against these six FIV strains were also investigated in 50 FIV-antibody-positive serum samples collected from different geographical regions in Japan. The cross-neutralization test revealed that antisera against the six strains tended to possess high neutralizing activity against the homologous strain. These antisera were also capable of neutralizing viral strains of the same subtype. However, some of the antisera were broadly crossreactive with all six FIV strains. Serum samples collected from naturally infected cats in the field showed various neutralization patterns for the six FIV strains. These observations reflect the antigenic diversity in FIV strains prevailing in the field. There were also broadly crossreactive serum samples, and 36% (18/50 samples) showed neutralization for all six FIV strains.
Journal of Veterinary Medical Science 06/1997; 59(5):377-81. · 0.85 Impact Factor
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ABSTRACT: Feline immunodeficiency virus (FIV) isolates have been classified into subtypes A, B, C and D based on the env gene V3-V5 sequence. The cell tropism of seven new Japanese isolates and a Petaluma (prototype) isolate of FIV, which classified into subtypes A, B and D, for feline lymphoblastoid and feline fibroblastoid cell lines was compared. FeT-1 (CD4+/-, CD8-, AND CD9+2) and Kumi-1 (CD4+2, CD8- and CD9+2) cells were used as the interleukin-2 (IL-2)-dependent feline T-lymphocyte cell lines and FeT-J (CD4+, CD8+/- and CD9+2) and 3201 (CD4+2, CD8+ and CD9-) cells were used as the IL-2- independent feline T-lymphocyte cell lines. The feline fibroblastoid cell lines used were Crandell feline kidney (CrFK) and fewf-4 (both CD4-, CD8- and CD9+2) cells. All FIV isolates replicated in all lymphoblastoid cell lines used. All isolates showed the greatest cytopathogenicity for Kumi-1 cells. All isolates replicated even in the CD9-negative 3201 cells. More isolates caused persistent infection in IL-2-independent cell lines than in IL-2-dependent cell lines. The number of subtype B isolates that established persistent infection was limited, only one of four strains. Only the subtype A isolates replicated in CrFK cells, whereas none of the isolates replicated in fewf-4 cells, which have similar cell surface markers to CrFK cells. The subtype A viruses (CrFK/Petaluma, CrFK/Sendai-1) growing in CrFK cells showed greater cytopathogenicity for lymphoblastoid cell lines than did those (FL-4/Petaluma, Kumi-1/Sendai-1) growing in a lymphoblastoid cell line.
Journal of General Virology 02/1996; 77 ( Pt 1):93-100. · 3.36 Impact Factor
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ABSTRACT: Complete nucleotide sequences were determined by cDNA cloning of peplomer (S), integral membrane (M) and nucleocapsid (N) genes of feline infectious peritonitis virus (FIPV) type I strain KU-2, UCD1 and Black, and feline enteric coronavirus (FECV) type II strain 79-1683. Only M and N genes were analyzed in strain KU-2 and strain 79-1683 which still had unknown nucleotide sequences. Deduced amino acid sequences of S, M and N proteins were compared in a total of 7 strains of coronaviruses, which included FIPV type II strain 79-1146, canine coronavirus (CCV) strain Insavc-1 and transmissible gastroenteritis virus of swine (TGEV) strain Purdue. Comparison of deduced amino acid sequences of M and N proteins revealed that both M and N proteins had an identity of at least 90% between FIPV type I and type II. The phylogenetic tree of the M and N protein-deduced amino acid sequences showed that FIPV type I and type II form a group with FECV type II, and that these viruses were evolutionarily distant from CCV and TGEV. On the other hand, when the S protein-deduced amino acid sequences was compared, identity of only about 45% was found between FIPV type I and type II. The phylogenetic tree of the S protein-deduced amino acid sequences indicated that three strains of FIPV type I form a group, and that it is a very long distance from the FIPV type II, FECV type II, CCV and TGEV groups.
Microbiology and Immunology 02/1996; 40(6):425-33. · 1.30 Impact Factor
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ABSTRACT: Seven isolates of feline immunodeficiency virus (FIV), Shizuoka, Yokohama, Sendai-1, Sendai-2, Fukuoka, Aomori-1, and Aomori-2, were isolated from FIV-seropositive domestic cats in Japan, and their proviral DNAs were amplified by PCR. The nucleotide sequences of their env and gag genes were determined and compared with those of previously described isolates: U.S. and European isolates and one Japanese isolate, TM2. Phylogenetic analyses of complete env gene sequences demonstrate that worldwide isolates are classified into three subtypes: Japanese TM2, Japanese Shizuoka, and non-Japanese subtypes (U.S. and European isolates), with 20% amino acid distances from each other. This pattern indicates that an evolutionary radiation of these three subtypes of FIV occurred at approximately the same time. The sequence data of gag genes also confirmed these results. Furthermore, the Sendai-1 isolate was identified as an imported FIV isolate.
Journal of Virology 07/1995; 69(6):3639-46. · 5.40 Impact Factor
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ABSTRACT: The effects of centrifugation on the ability of feline infectious peritonitis virus (FIPV) to infect cells in culture was investigated. The infectivity titer was the highest when the plates were centrifuged at 400 x g (1500 rpm) for 2 h. All five strains classified as FIPV Type I showed infectivity titers enhanced 10-100-fold by centrifugation at 400 x g for 2 h. The centrifugal enhancement of infection was obtained only by centrifugation immediately after inoculation of the virus, suggesting that the enhancement occurs during attachment or adsorption of viruses to the cells. This method may be useful for the culture of FIPV Type I strains.
Journal of Virological Methods 03/1995; 51(2-3):357-62. · 2.01 Impact Factor
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ABSTRACT: cDNA clones spanning the entire region of the peplomer (S) gene of feline infectious peritonitis virus (FIPV) type I strain KU-2 were obtained and their complete nucleotide sequences were determined. A long open reading frame (ORF) encoding 1464 amino acid residues was found in the gene, which was 12 residues longer than the ORF of the FIPV type II strain 79-1146. The sequences of FIPV type I and mainly -tPV type II were compared. The homologies at the N- (amino acid residues 1-693) and C- (residues 694-1464) terminal halves were 29.8 and 60.7%, respectively. This was much lower than that between FIPV type II and other antigenically related coronaviruses, such as transmissible gastroenteritis virus of swine and canine coronavirus. This supported the serological relatedness of the viruses and confirmed that the peplomer protein of FIPV type I has distinct structural features that differ from those of antigenically related viruses.
Archives of Virology 02/1995; 140(3):469-80. · 2.11 Impact Factor
