No abstract

Enhancement of feline infectious peritonitis virus (FIPV) infection of feline macrophages was studied using monoclonal antibodies (MAbs) to the FIPV strain 79-1146. Adherent cells recovered from the feline lung and peritoneal cavity phagocytosed fixed red blood cells, and formed Fc-mediated rosettes. Enhancement of virus infection by MAb was investigated by inoculating alveolar macrophages with a mixtures of viral suspension and MAb, and examining the cells for intracellular viral antigen by the immunofluorescence assay and the amount of infectious virus in the supernatant fluid after incubation. The replication of FIPV in macrophages was enhanced by non-neutralizing MAbs recognizing peplomer protein (S) and transmembrane protein (M) of the virus. Even among the MAbs having the ability to neutralize FIPV strain 79-1146, some reversely enhanced virus infection when they were diluted. The enhancement was suppressed by pretreatment of the MAb with protein A. The enhancement was reduced by the use of F(ab')2 fragment of MAb. These results demonstrated antibody-dependent enhancement (ADE) of FIPV infection in macrophage. The replication of FIPV 79-1146 strain in macrophages from FIPV antibody-positive cats was more enhanced than in those from antibody-negative cats.

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.Key words: Feline infectious peritonitis virus, Feline enteric coronavirus, Amino acid sequence Feline infectious peritonitis virus (FIPV), family Coronaviridae, genus Coronavirus, causes a chronic, progressive, immunologically mediated disease in domestic and exotic cats.The family Coronaviridae is divided into 3 distinct antigenic groups on the basis of serologic tests (18). One group contains mouse hepatitis virus, neonatal calf diarrhea coronavirus, human coronavirus OC43, hemagglutinating encephalomyelitis virus of swine and rat coronavirus. The second group contains avian infectious bronchitis virus. The third group consists of human respiratory coronavirus 229E, transmissible gastroenteritis virus (TGEV) of swine, porcine respiratory coronavirus, canine coronavirus (CCV), FIPV and feline enteric coronavirus (FECV). By using monoclonal antibodies we have shown the existence of at least 2 serotypes of FIPV, and the antigenicity of type II strain of FIPV and FECV were closer to TGEV and CCV than to type I FIPV (7,8). Both types I and II FIPV cause infectious peritonitis in cats, and the pathogenicity of type II FIPV is greater than that of type I FIPV (25). However, in the field, the prevalence of FIPV type I is high, and about 70% of feline cases of FIP are due to infection with type I (9).FIPV is an enveloped RNA virus with a single-stranded positive-sense RNA genome and the virions consist of three main structural proteins, peplomer (S) glycoprotein, integral membrane (M) glycoprotein and nucleocapsid (N) protein. The virus genome is at least 20 kilobases

Feline infectious peritonitis (FIP) cats show a decrease in peripheral blood lymphocyte counts, and a particularly marked decrease in T cells including CD4+ and CD8+ cells. In this study, we showed that lymphopenia observed in FIP cats was due to apoptosis, and that the ascitic fluid, plasma, and culture supernatant of peritoneal exudate cells (adherent cells with macrophage morphology, or PEC) from FIP cats readily induced apoptosis in specific pathogen-free cat peripheral blood mononuclear cells, particularly CD8+ cells. In addition, TNF-alpha released from macrophages and TNF-receptor (TNFR) 1 and TNFR2 mRNA expression in lymphocytes were closely involved in this apoptosis induction. In particular, in CD8+ cells cultured in the presence of the PEC culture supernatant, the expression levels of TNFR1 and TNFR2 mRNA were increased, indicating that CD8+ cells are more susceptible to apoptosis induction by TNF-alpha than other lymphocyte subsets, particularly B cells (CD21+ cells). The results of this study suggest that TNF-alpha, produced by virus-infected macrophages, is responsible for induction of apoptosis in uninfected T cells, primarily CD8+ T cells.

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.

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. -KEY WORDS: antibody-dependent enhancement of infection, feline infectious peritonitis virus, feline macrophage, U937 cell.

Seven monoclonal antibodies (MAbs) with neutralizing activity against feline infectious peritonitis virus (FIPV) strain 79-1149 (type II) were prepared. When the polypeptide specificity recognized by these monoclonal antibodies (MAbs) was investigated by Western immunoblotting, all of the MAbs reacted with peplomer glycoprotein (S) of the virus. By competitive binding assay these MAbs were found to recognize at least 3 different epitopes. The reactivity of these MAbs with 6 viruses classified as FIPV type I (UCD-1, UCD-2, UCD-3, UCD-4, NW-1, and Black), feline enteric coronavirus (FECV) type II strain 79-1683, canine coronavirus (CCV) strain 1-71, and transmissible gastroenteritis virus (TGEV) strains TO-163 and SH was examined by neutralization tests. All MAbs neutralized FECV strain 79-1683, CCV strain 1-71, and TGEV strains TO-163 and SH, while they did not neutralize the 6 FIPV type I viruses. Moreover, the MAb against TGEV strain TO-163, which has strong neutralizing activity against 7 TGEV viruses, neutralized CCV strain 1-71, FECV strain 79-1683, and FIPv strain 79-1146, but did not neutralize the 6 FIPV type I viruses. These results demonstrated that there are at least 3 epitopes involved in the neutralization of FIPV type II strain 79-1146, and that these epitopes are not present in FIPV type I viruses but are present in FECV strain 79-1683 which does not induce feline infectious peritonitis, TGEV strains TO-163 and SH, and CCV strain 1-71. These results suggest the presence of 2 serotypes of FIPV which can be clearly distinguished by the neutralization test using MAbs.

No abstract