An enteric coronavirus infection of cats and its relationship to feline infectious peritonitis

An enteric coronavirus that is antigenically closely related to feline infectious peritonitis virus (FIPV) is ubiquitous in the cat population. This virus has been designated feline enteric coronavirus to differentiate it from FIPV. The virus is shed in the feces by many seropositive cats; in catteries it is a cause of inapparent to mildly severe enteritis in kittens 6 to 12 weeks of age. The virus may produce a more severe enteritis in young specific-pathogen-free kittens. Feline enteric coronavirus selectively infects the apical columnar epithelium of the intestinal villi, from the caudal part of the duodenum to the cecum. In severe infections, there are sloughing of the tips of the villi and villous atrophy. Many cats recovering from the disease remain carriers of the virus. Recovered cats, observed for 3 to 24 months, remained healthy and did not develop peritonitis, pleuritis, or granulomatous disease. The relationship of feline enteric coronavirus and FIPV was studied. Although the viruses were antigenically similar, they were distinctly different in their pathogenicities. The enteric coronavirus did not cause feline infectious peritonitis in coronavirus antibody-negative cats inoculated orally or intraperitoneally nor in coronavirus antibody-positive cats inoculated intraperitoneally or intratracheally. Serologic tests, using FIPV, canine coronavirus, and transmissible gastroenteritis virus of swine as substrate antigens in fluorescent antibody procedures may not accurately identify FIPV infection. These tests do not appear to distinguish between FIPV and this feline enteric coronavirus.     

Evaluation of antibodies against feline coronavirus 7b protein for diagnosis of feline infectious peritonitis in cats

OBJECTIVE: To determine whether expression of feline coronavirus (FCoV) 7b protein, as indicated by the presence of specific serum antibodies, consistently correlated with occurrence of feline infectious peritonitis (FIP) in cats. SAMPLE POPULATION: 95 serum samples submitted for various diagnostic assays and 20 samples from specific-pathogen-free cats tested as negative control samples. PROCEDURES: The 7b gene from a virulent strain of FCoV was cloned into a protein expression vector. The resultant recombinant protein was produced and used in antibody detection assays via western blot analysis of serum samples. Results were compared with those of an immunofluorescence assay (IFA) for FCoV-specific antibody and correlated with health status. RESULTS: Healthy IFA-seronegative cats were seronegative for antibodies against the 7b protein. Some healthy cats with detectable FCoV-specific antibodies as determined via IFA were seronegative for antibodies against the 7b protein. Serum from cats with FIP had antibodies against the 7b protein, including cats with negative results via conventional IFA. However, some healthy cats, as well as cats with conditions other than FIP that were seropositive to FCoV via IFA, were also seropositive for the 7b protein. CONCLUSIONS AND CLINICAL RELEVANCE: Expression of the 7b protein, as indicated by detection of antibodies against the protein, was found in most FCoV-infected cats. Seropositivity for this protein was not specific for the FCoV virulent biotype or a diagnosis of FIP.     

Prevalence of feline coronavirus types I and II in cats with histopathologically verified feline infectious peritonitis

Feline coronaviruses (FCoV) vary widely in virulence causing a spectrum of clinical manifestations reaching from subclinical course to fatal feline infectious peritonitis (FIP). Independent of virulence variations they are separated into two different types, type I, the original FCoV, and type II, which is closely related to canine coronavirus (CCV). The prevalence of FCoV types in Austrian cat populations without FIP has been surveyed recently indicating that type I infections predominate. The distribution of FCoV types in cats, which had succumbed to FIP, however, was fairly unknown. PCR assays have been developed amplifying parts of the spike protein gene. Type-specific primer pairs were designed, generating PCR products of different sizes. A total of 94 organ pools of cats with histopathologically verified FIP was tested. A clear differentiation was achieved in 74 cats, 86% of them were type I positive, 7% type II positive, and 7% were positive for both types. These findings demonstrate that in FIP cases FCoV type I predominates, too, nonetheless, in 14% of the cases FCoV type II was detected, suggesting its causative involvement in cases of FIP.     

Humoral immune responses of cats to feline infectious peritonitis virus infection.

Immunoperoxidase antibody (IPA) method as a titrating method of feline infectious peritonitis (FIP) virus (FIPV) was developed for titrating antibody to FIPV (IPA-titer). By this method the immune responses of the cats that had been infected with FIPV, were traced. The infected cats could be grouped into three types by their immune response to FIPV and clinical appearances. Type I cats lived for a long time, formed a major group among infected cats, had 160 to 1 x 10(4) IPA-titers, and showed healthy appearances without any changes both on autopsy and histopathologically. From among type I cats, type II cats appeared sporadically with rapid elevation of IPA titers to 3.2 x 10(5) and showing clinical signs of FIP, and died. Type III cats lived healthily for a long time with gradual elevation of IPA-titers to a plateau of about 1 x 10(5), then showed neuronal disorder of hind leg paralysis with the descending IPA-titers to 2 x 10(4), and died. Thus, typical FIP appeared as a hyper-immune disease. Other related problems are discussed.     

Induction and enhancement of feline infectious peritonitis by canine coronavirus.

Preexisting antibody to feline infectious peritonitis virus (FIPV) causes acceleration and enhancement of disease on subsequent infection of cats with FIPV. Other workers have shown that canine coronavirus (CCV) can infect cats subclinically, but have found no evidence of enhancement of, or protection against, subsequent FIPV infection. With various isolates of CCV, we determined that 1 strain of CCV can induce transient mild diarrhea in cats and, furthermore, that previous infection with CCV causes acceleration and enhancement of subsequent infection with FIPV. In addition, sequential inoculation of cats with another strain of CCV caused lesions indistinguishable from those of FIP, without exposure at any time to FIPV.     

Detection of feline coronavirus antibody, feline immunodeficiency virus antibody, and feline leukemia virus antigen in ascites from cats with effusive feline infectious peritonitis

To investigate the usefulness of ascites as a material for viral tests in cats with effusive feline infectious peritonitis (FIP), we attempted to detect anti-feline coronavirus antibody, anti-feline immunodeficiency virus antibody, and feline leukemia virus antigen in ascites from 88 cats clinically suspected with effusive FIP. In each of these three viral tests, all cats positive for serum antibody/antigen were also positive for ascitic antibody/antigen, while cats negative for serum antibody/antigen were also negative for ascitic antibody/antigen. This finding indicates that ascites is useful for these viral tests.     

Total sialic acid: An acute phase reactant in cats with a possible role in feline coronavirus infection

The aims of this study were to validate a colorimetric method to measure total sialic acid (TSA) in feline serum and to investigate the serum concentration of TSA in clinically healthy cats seronegative (n = 9) and seropositive (n = 48) for feline coronavirus (FCoV), and in cats affected by feline infectious peritonitis (FIP, n = 28), tumors (n = 20), or inflammation (n = 16). The correlation between TSA and α₁-acid glycoprotein (AGP) was also investigated. The method employed in this study is precise and accurate at TSA levels (in mg/L) commonly encountered in feline serum. No significant differences between seropositive (385.6 ± 192.2 mg/L) and seronegative (433.5 ± 179.0 mg/L) cats were detectable, suggesting that the simple infection by FCoVs does not influence TSA levels. Compared with seropositive controls, the concentration of TSA was higher in cats with FIP (556.7 ± 268.3 mg/L, P = 0.003), tumors (522.5 ± 294.4 mg/L, P = 0.028), and inflammation (546.8 ± 208.3 mg/L, P = 0.018). The discriminating power of TSA for FIP is moderate (area under the ROC curve = 0.65) and the likelihood ratio is higher than 3.0 only at high TSA levels. Consequently, TSA could support a diagnosis of FIP only at extremely high serum concentration (> 800 mg/L) or when the pre-test probability of FIP is high. No correlations were found between the TSA and AGP concentrations in cats with FIP, suggesting that sialylated proteins other than AGP are present. Both the antibody titre and the degree of AGP sialylation were negatively correlated with TSA levels, suggesting that increased TSA may contribute to reduce the burden of FCoVs.     

Experimental inoculation of cats with human coronavirus 229E and subsequent challenge with feline infectious peritonitis virus.

Minimal-disease cats exposed to live human coronavirus 229E developed homologous antibody responses that suggested little or no replication of the virus in inoculated animals. Oronasal and subcutaneous inoculation of coronavirus 229E did not elicit an antibody response by heterologous (transmissible gastroenteritis virus, canine coronavirus) neutralization or by heterologous (transmissible gastroenteritis virus) kinetics-based enzyme-linked immunosorbent assay. No clinical signs attributable to coronavirus 229E were seen in inoculated cats. Although the number of animals in each of the five experimental groups was small (n = 2), antibodies produced in response to the virus did not appear to sensitize cats to subsequent feline infectious peritonitis virus challenge, but neither did they cross-protect cats against the challenge dose.     

Decreased sialylation of the acute phase protein alpha1-acid glycoprotein in feline infectious peritonitis (FIP)

Feline infectious peritonitis (FIP) is an immune-mediated disease of domestic and exotic felides infected with feline coronavirus. FIP is characterized by the overexpression of an acute phase protein, the alpha1-acid glycoprotein (AGP). In humans, AGP is a heavily glycosylated protein that undergoes several modifications of its glycan moiety during acute and chronic inflammatory pathologies. We studied the changes in AGP glycosylation in the course of FIP. Specifically, we focussed our attention on the degree of sialylation, fucosylation and branching. This study presents a purification method for feline AGP (fAGP) from serum, using an ion exchange chromatography strategy. The glycosylation pattern was analyzed in detail by means of interaction of purified fAGP with specific lectins. In particular, Sambucus nigra agglutinin I and Maackia amurensis agglutinin lectins were used to detect sialic acid residues, Aleuria aurantia lectin was used to detect L-fucose residues and Concanavalin A was used to evaluate the branching degree. By this method we showed that fAGP did not present any L-fucose residues on its surface, and that its branching degree was very low, both in normal and in pathological conditions. In contrast, during FIP disease, fAGP underwent several modifications in the sialic acid content, including decreased expression of both alpha(2-6)-linked and alpha(2-3)-linked sialic acid (76 and 44%, respectively when compared to non-pathological feline AGP).     

Evaluation of immunity to feline infectious peritonitis in cats with cutaneous viral-induced delayed hypersensitivity

Delayed-type hypersensitivity (DTH)-like reactions to feline infectious peritonitis (FIP) virus (FIPV) were induced in the skin of nine cats that were asymptomatic after a previous challenge-exposure with FIPV. Four of the nine previously challenge-exposed cats were negative for virus-neutralizing antibodies against FIPV at the time of intradermal (ID) testing for DTH. Two other cats tested for DTH when acutely ill with clinical FIP did not have cutaneous DTH responses to FIPV. Gross skin reactions to FIPV injected ID were observed in six of nine asymptomatic cats (67%) at postintradermal inoculation hours (PIH) 24, 48, and/or 72. The reactions consisted of focal, 1-5-mm to 2.5-cm diameter indurated or semi-firm, nonerythematous, slightly raised nodules. Microscopically, DTH-like reactions were observed in biopsies taken from the FIPV-inoculated skin of asymptomatic cats at PIH 24 to 72. The lesions consisted of perivascular and diffuse dermal infiltrations by macrophages, lymphocytes, and polymorphonuclear leukocytes (PMN). The dermal infiltrates, which were maximal at PIH 48 or 72, were predominantly mixed inflammatory cells (five of nine cats) or PMN (four of nine cats) at PIH 24, but later were predominantly mononuclear cells (six of nine cats) or mixed inflammatory cells (two of nine cats) at PIH 72. Five of nine cats (56%) with positive DTH skin responses had increased survival times after lethal ID challenge-exposure with FIPV compared to mean survival times in FIPV-naive, non-immune control cats that were DTH-negative when ID challenge-exposed. Four of nine DTH-positive cats (44%) resisted an ID challenge-exposure dose of FIPV that was fatal in both control cats, and two of the four remaining DTH-positive cats survived a third challenge-exposure with highly lethal doses of FIPV given intraperitoneally. Four of the six DTH-positive cats (67%) that died after re-challenge and were necropsied had lesions of noneffusive FIP, suggesting that cellular immunity may also be involved in the pathogenesis of noneffusive disease, whereas both control cats and both DTH-negative cats with clinical disease succumbed to effusive FIP. Seemingly, DTH responses to FIPV can be associated with an increased level of resistance to disease; however, this state of immunity is variable and apparently can be lost with time in some cats.     

Tissue distribution of a feline AGP related protein (fAGPrP) in cats with feline infectious peritonitis (FIP)

Feline alpha(1)-acid glycoprotein (fAGP) increases during feline infectious peritonitis (FIP). We have recently identified a 29 kDa protein that we named feline AGP-related protein (fAGPrP) due to its cross-reactivity with an anti-human AGP monoclonal antibody. In this work we describe the tissue distribution of fAGPrP during FIP, and its relationship with feline coronavirus (FCoV) and myeloid cells. Tissues from five control cats and from 15 cats with FIP were examined by immunohistochemistry using monoclonal antibodies against human AGP, FCoV and myeloid antigens. Diffuse fAGPrP positivity within the lesions, likely due to vascular plasma leakage, endothelial and epithelial lining were detectable. Compared to controls, fAGPrP-expressing cells often increased in number and were diffusely distributed in lymph nodes, as usually occurs for IgM-producing plasma cells during early immune responses. These findings did not depend on the presence of FCoVs or of myeloid cells, suggesting that fAGPrP is not directly involved in the pathogenesis of FIP.     

Interferon-gamma in the serum and effusions of cats with feline coronavirus infection

The aim of this study was to quantify and compare interferon-γ (IFN-γ) concentrations in the serum of clinically normal cats infected with feline coronavirus (FCoV) with its concentration in the sera and effusions of cats with feline infectious peritonitis (FIP), a disease associated with infection with a mutated form of FCoV.Clinically normal FCoV-infected cats living in catteries with a high prevalence of FIP had the highest serum IFN-γ concentrations. The serum concentration of IFN-γ was not significantly different in cats with FIP compared with clinically normal FCoV-infected animals living in catteries with a low prevalence of the disease. Moreover, the concentration of IFN-γ was significantly higher in the effusions than in the serum of cats with FIP, probably due to IFN-γ production within lesions. These findings support the hypothesis that there is a strong, ‘systemic’ cell mediated immune response in clinically normal, FCoV-infected cats and that a similar process, albeit at a tissue level, is involved in the pathogenesis of FIP.     

Virus shedding and immune responses in cats inoculated with cell culture-adapted feline infectious peritonitis virus

Eight specific pathogen-free cats were inoculated orally or parenterally with a cell culture-adapted strain of feline infectious peritonitis virus (FIPV). Faeces and oropharyngeal swabs were monitored daily for infectious virus by inoculation of feline embryo lung cells. Virus was recovered from both sites for approximately 2 weeks after inoculation, before clinical signs of disease developed. Peripheral blood lymphocytes collected from these cats were tested in an in-vitro blastogenic assay using concanavalin A (con A) and FIPV antigen. All cats showed a profound suppression of the response to con A which only recovered to pre-inoculation levels in 2 cats, one of which survived. These 2 cats also responded to FIPV antigen on the 21st day after infection, the greater response being in the survivor. The other cats, surviving 16-18 days, developed no response to FIPV antigen. Antibody titres, measured by immunofluorescence and by virus neutralization, rose rapidly to very high levels in all cats, regardless of the route of inoculation.     

Effect of interferon or Propionibacterium acnes on the course of experimentally induced feline infectious peritonitis in specific-pathogen-free and random-source cats

Seventy-four cats (52 treated and 22 untreated) were evaluated in efficacy studies of interferon (IFN), Propionibacterium acnes, or a combination of these drugs against experimentally induced feline infectious peritonitis (FIP). Cats were given doses of recombinant human leukocyte (alpha) IFN (rHuIFN-alpha), feline fibroblastic (beta) IFN (FIFN-beta) or P acnes at regular intervals before and after inoculation of virulent FIP virus (FIPV). Prophylactic and therapeutic administration of high doses (10(6) U/kg of body weight) or moderate doses (10(4) U/kg) of rHuIFN-alpha, FIFN-beta (10(3) u/kg), or P acnes (0.4 or 4 mg) did not significantly reduce mortality in treated vs untreated cats. However, the mean survival time in cats treated with 10(6) U of rHuIFN-alpha-/kg alone or combined with doses of P acnes was significantly (P = 0.03) increased after inoculation of highly lethal amounts (200 LD100) of FIPV vs survival time in untreated cats. Although P acnes alone was ineffective, there was some indication that a combination of P acnes and high doses of rHuIFN-alpha was more effective than rHuIFN-alpha alone. Seemingly, the efficacy of rHuIFn-alpha treatment was improved in cats challenge-exposed with less FIPV; in 1 trial, 4 of 5 cats (80%) treated with high doses of rHuIFN-alpha survived after inoculation of minimal lethal amounts (0.6 LD100) of FIPV, whereas only 2 of 5 untreated cats (40%) survived. Pretreatment of cats with 10(6) U of rHuIFN-alpha/kg resulted in detectable serum IFN activity 24 hours later; serum IFN activity was not detected in cats pretreated with P acnes, FIFN-beta, or 10(4) U of rHuIFn-alpha/kg.(ABSTRACT TRUNCATED AT 250 WORDS)     

Laboratory profiles in cats with different pathological and immunohistochemical findings due to feline infectious peritonitis (FIP)

Blood was collected from 55 cats with feline infectious peritonitis (FIP) and from 50 control cats in order to define whether differences in pathological findings and in distribution of feline coronaviruses (FCoV) can be associated with changes in haemograms, serum protein electrophoresis, and antibody titres. Compared to controls, the whole group of FIP-affected cats had blood changes consistent with FIP. Based on the pathological findings or on the immunohistochemical distribution of viral antigen, FIP-affected cats were divided in the following groups: subacute against acute lesions; low against strong intensity of positivity; intracellular against extracellular positivities; positive against negative lymph nodes. Lymphopenia was more evident in cats with acute forms, strong intensity of positivity, extracellular antigen and negative lymph nodes. Cats with positive lymph nodes had the most evident changes in the protein estimations. These results suggest that differences in pathological findings might depend on different reactive patterns to the FCoVs.     

Evaluation of antithrombin-III activity as a coindicator of disseminated intravascular coagulation in cats with induced feline infectious peritonitis virus infection

Six adult specific-pathogen-free cats were inoculated intraperitoneally with a cell culture-adapted strain of feline infectious peritonitis virus. Plasma samples were evaluated for antithrombin-III (AT-III) activities at post-inoculation days (PID) 0, 4, and 11 and at termination on PID 16 (1 cat) or 21 (5 cats). Other hemostatic values evaluated were activated partial thromboplastin times, prothrombin times, thrombin times, fibrinogen, platelet counts, and fibrin/fibrinogen degradation products. Antithrombin-III activity remained within normal or above normal range (89 to 246%) in all cats, with the exception of one cat on PID 4 (AT-III, 70%). Mean baseline AT-III activity for 6 cats at PID 0 was 123%. Mean AT-III activity on PID 4, 11, and 16 or 21 was 98, 162, and 130%, respectively. On PID 4 and 16 or 21, results of coagulation screening tests indicated that all cats had disseminated intravascular coagulation. Histologically, cats also had severe fibrinonecrotizing thrombovasculitis.     

Virion polypeptide specificity of immune complexes and antibodies in cats inoculated with feline infectious peritonitis virus

Immune complexes purified from sera and ascitic fluids of cats after inoculation with feline infectious peritonitis (FIP) virus contained proteins and proteolytic fragments of the peplomer, nucleocapsid, and envelope polypeptides; in addition, host proteins were demonstrated in the immune complexes. Free (uncomplexed) antibodies against the 3 classes of virion polypeptides were detected and quantitated; the weakest and latest response was directed against the peplomer protein. Immunofluorescence titers showed the best correlation with the antibody response directed against the envelope polypeptides. Differences in reactivity were not found between sera and ascitic fluids from the same animals and between seropositive healthy cats and cats which had died of FIP. Humoral antibody and hypergammaglobulinemia showed a linear correlation, but the wide variation in antiviral titers at a given concentration of gamma-globulin indicated that additional (autoimmune) reactions occur during the pathogenesis of FIP.