The Feline Centre, Langford Veterinary Services, University of Bristol, Langford, Bristol, UK
What Is Feline Infectious Peritonitis (FIP)?
Feline coronavirus (FCoV) infection is very common in cats; around 40% of the domestic cat population has been FCoV infected, and this figure increases to 90% in multi-cat households.1-2 A study of owned cats in Sydney reported a seroprevalence of 34%.3 Natural infections with FCoV are often transient and asymptomatic or result in mild gastrointestinal disease. However, occasionally, FCoV infection results in the multisystemic disease of feline infectious peritonitis (FIP),4 the single most important infectious cause of death in young cats4. No cure for FIP exists and it is an extremely distressing disease to deal with, for both cat owners and veterinary surgeons.
What Causes FIP?
Viral factors are thought to be important, and a recent Dutch study5 has identified mutations in the FCoV spike protein that distinguish FIP-associated FCoVs from those not associated with FIP. By these and perhaps other mutations, the virus could acquire its macrophage tropism and spread systemically to result in FIP. Host factors are also believed to be important; the immune response of the cat may be able to prevent the development of FIP, and breed and genetics may play a role. Environmental factors, such as the level of stress and overcrowding in a household, may also influence the outcome of FCoV infection.
Diagnosis - A Big Dilemma in FIP
Sadly, a single minimally invasive FIP test that can definitively diagnose all FIP cases doesn't exist. Definitive diagnosis traditionally relies on histopathological examination of tissues ± immunological staining of FCoV antigen, but the reliability of these methods is not always 100%. Evaluation of tests is often difficult, as criteria used to definitively diagnose FIP differ in published studies (e.g., histopathology/immunological staining/algorithms). Different criteria and methods for the diagnosis of FIP are described below.
Background Information & Clinical Signs
Before performing diagnostic tests, one must remember that FIP is most common in young cats (< 3 years, but there is also a smaller peak in cats > 10 years), pedigree cats and cats from multicat households. A recent history of stress (rehoming, neutering, changes in group hierarchy, vaccination) may be apparent. Typical clinical signs of FIP (e.g., lethargy, anorexia, weight loss, pyrexia, jaundice) are seen; wet (most common; < 80% cases with ascites and/or pleural effusions and/or pericardial effusions; often quite acute), dry (neurological signs [cerebellar ataxia, nystagmus and seizures] and/or ocular changes [uveitis] more chronic progression) and focal (lymph node or intestinal masses) forms are reported. However, as FIP is a progressive disease, clinical signs change over time, so it is important to repeat clinical examinations to look for development of signs (e.g., effusions, uveitis) to aid diagnosis.
Haematology can support a diagnosis of FIP, but changes are nonspecific. Lymphopenia is particularly common (55-77% of cases), with neutrophilia (39-55%) and mild to moderate normocytic, normochromic anaemia (37-54%; worsening, and increased prevalence, of anaemia are also seen during FIP disease progression) also reported.6-8
FIP is usually associated with hyperproteinaemia (~ 60%, although some studies have reported much lower prevalences than this) due to hyperglobulinaemia, usually with a low or low-normal serum albumin; consequently the albumin:globulin (A:G) ratio is low (< 0.4 means FIP very likely, > 0.8 makes FIP very unlikely; reference range 0.45-1.2).6-8 α1-acid glycoprotein (AGP), an acute phase protein, is often elevated in FIP; although many inflammatory and noninflammatory diseases are also associated with raised (> 0.48 mg/ml) AGP concentrations, AGP levels in FIP are usually markedly elevated (> 1.5 mg/ml), so the magnitude of increase may be helpful in aiding diagnosis of FIP.9-10 Hyperbilirubinaemia is not uncommonly reported (21-36% cases), especially in effusive cases, often without marked elevations in liver enzymes (ALT, ALP & GGT concentrations often normal/only mildly elevated despite hyperbilirubinaemia, which can help increase suspicion of FIP). Worsening and increased prevalence of the hyperbilirubinaemia are reported during FIP disease progression.6
Serum FCoV antibody tests are usually enzyme-linked immunosorbent assays (ELISAs) or indirect immunofluorescence antibody (IFA) tests. Most tests use CoV-infected swine or cat cells as a substrate and titres are read in distinct multiples of serum dilutions. No serum FCoV antibody test can differentiate antibodies associated with FIP with those not associated with FIP. A positive FCoV antibody test indicates that the cat has been infected with a FCoV and has seroconverted (takes 2-3 weeks). Although FIP cases tend to have higher FCoV antibody titres than healthy cats or cats without FIP, there is much overlap, so the value of such results in an individual cat can be limited. Many clinically healthy cats (esp. those in multicat households) have positive titres (which can be high), whilst ~10% of cats with FIP are actually seronegative.
Analysis of Effusion Samples
Effusion samples are very helpful in the diagnosis of FIP; so getting a sample of any effusion present is essential in any suspected case. Repeated imaging (esp. ultrasonography) can be useful to detect effusions. FIP effusions are usually clear, viscous and straw-yellow in colour. They are protein rich (frothy when shaken ± fibrin strands) with total protein concentration of > 35 g/l and a predominance (> 50%) of globulins. FIP effusions usually have similar low A:G ratios and raised AGP concentrations to those in serum (see above).
FIP effusions are poorly cellular (usually < 5 x 109/l cells), consisting of non-degenerate neutrophils and macrophages. Thus, FIP effusions are classified as exudates based on their high protein concentration but are more of a modified transudate based on their low cell counts.
Reverse-transcriptase (RT-) Polymerase Chain Reaction (PCR)
RT-PCR for FCoV amplifies FCoV RNA in blood, effusions, faecal (to detect FCoV shedders) or tissue samples. Up until recently (see below), no specific genetic sequences had been demonstrated for FCoVs associated with FIP, and currently available PCR assays detect any FCoV, rather than those just associated with FIP. Comparison of studies evaluating PCR is difficult due to the use of different RT-PCR assays with varied, or more often not declared, sensitivities and specificities.
Work at the University of Bristol using a specific and quantitative genomic FCoV RT-PCR has found that FCoV RNA can be detected in the blood of FIP cases, although levels are quite low; RT-PCR to detect FCoV on effusion or tissue samples may be more helpful. Recent studies suggest that FCoV RNA can be amplified by RT-PCR from the vast majority of FIP effusion samples tested;6,11 conversely, non-FIP effusions do not generally generate positive RT-PCR results. We have found similar results using tissue samples for RT-PCR (where levels of FCoV have correlated with pathological findings) and, although non-invasive collection of such samples is obviously more difficult, it may be that RT-PCR performed on tissue samples collected by, e.g., Tru-Cut biopsy, becomes a useful diagnostic test as it is quicker to perform than histopathology. RT-PCR assays that target subgenomic mRNA (i.e., replicating) FCoV have also been reported12 to try and improve the differentiation of cats with FIP from those without (as the latter should have lower or no replicating FCoV in the blood or tissues), and these assays show promise. More work is probably required on control non-FIP samples before such tests can be offered for diagnostic use.
Very importantly for FCoV RT-PCR, recent work5 suggests that a PCR targeting a region of the spike protein may show promise for the diagnosis of FIP, due to the identification of FCoV sequence motifs that were found in > 95% of FIP cases, but in none of the asymptomatic healthy cats. Current research is underway investigating this.
Histopathological Examination of Tissues
Samples of tissue (e.g., liver, kidney or mesenteric lymph nodes), collected antemortem (by ultrasound-guided percutaneous Tru-Cut biopsy, laparoscopy or laparotomy) or at post- mortem, can be evaluated for characteristic histopathological changes of FIP (pyogranulomatous parenchymal foci, perivascular mononuclear infiltrates, fibrinous polyserositis). Historically, this was the only method available for definitive diagnosis and was considered reliable. Indeed, histopathology is very specific and lesions consistent with FIP are often used to confirm a clinical suspicion of FIP. However, a lack of lesions is more difficult to interpret. If small samples are taken (e.g., Tru-Cut biopsies), lesions may be missed due to their multifocal distribution, or due to non-affected organs being sampled.13 A recent study9 documented that 5 of 8 FIP cases did not have histopathology changes consistent with FIP, even though large representative biopsies appeared to have been taken; diagnosis in these cases was based on positive FCoV immunostaining (see below)
Immunological Staining of FCoV Antigen
This is performed using immunohisto/cytochemical (on formalin-fixed tissues or effusion cytology samples respectively) or immunofluorescence (on frozen tissues or effusion cytology samples) antibody staining to show FCoV antigen associated with pathology in tissues or in cells of an effusion. Positive immunological staining of tissues is said to confirm a diagnosis of FIP (i.e., it is very specific), but a negative result does not exclude FIP, as FCoV antigens may be variably distributed within lesions13 and, thus, not detected in some FIP cases. This somewhat contradicts the suggestion by some that immunostaining is mandatory to confirm/exclude FIP in doubtful cases.9
Immunostaining of effusion samples has also shown variable sensitivity, since this technique relies on staining FCoV within cells (particularly macrophages) in the effusion, and if the effusion is cell-poor, or if the FCoV antigen is being masked by FCoV antibodies in the effusion, a false negative result may be obtained. Immunostaining was previously thought to be 100% specific, although a recent study, published in abstract form only,11 reported that 2 of 50 cats, in which a diagnosis of FIP was excluded, showed positive immunostaining, so its specificity may not always be perfect.
Many features of a cat's history, clinical examination and laboratory testing can increase our suspicion of a diagnosis of FIP. In the majority of FIP cases, histopathology and immunostaining are reliable in obtaining a definitive diagnosis, but exceptions occur and it is important to interpret these results alongside other diagnostic investigations. PCR shows promise as an additional noninvasive test for the diagnosis of FIP.
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