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Written by the Founder of TCA, Inc.
DNA-BASED DETECTION OF
FELINE INFECTIOUS PERITONITIS
Reprinted with permission from the American Association of Feline Practitioners,
Academy of Feline Medicine, Feline Infectious Disease Symposium, Washington
D.C. October 14-17, 1995
The need for a new test. A diagnosis of FIP is still among the worst nightmares
a cat owner can face. There is no cure available, and none on the horizon.
The virus is difficult to detect and invariably fatal; therefore, detecting
and eliminating this virus from the cat population remains a top priority
of veterinary medicine.
The disease was first identified in the 1960’s and shown to be caused by
a member of the corona virus family some ten years later. The first diagnostic
trouble appeared when attempts were made to culture the virus, it proved
exceptionally difficult to grow, closing one usual route of diagnosis.
By the 1980’s FIPV had been demonstrated to belong to a huge family of
closely related viruses, and to have numerous strains (10 so far and counting),
each with different pathology. A closely related virus, named Feline Enteric
Corona Virus (FECV), with several substrains of its own, was found to be
virtually indistinguishable from FIP, but with very different pathology.
All of these viruses and variants were impossible to distinguish by the
usual methods, such as detection with a monoclonal antibody or sera directed
to the virus coat protein. Diagnosis and treatment are also difficult because
of the virus’ unique pathology. FIP virus infects the macrophages and monocytes
of the cat’s immune system, using them to spread throughout the body. The
damage caused by FIPV is actually due to an intense immune reaction and
localized inflammatory response at the site of virus colonization. In this
situation, detecting the virus can be very difficult indeed. To further
complicate disease management, cats infected with FIPV have been shown
to be most contagious between day’s 2-15-post infection, prior to the display
of any symptoms. It is not know whether direct contact with infected nasal,
oral, or digestive secretions are required to spread the disease, or if
contaminated food or water bowls are sufficient, or if the virus can even
be airborne. The incidence of carrier cats is also completely unknown,
but may exceed 10% in multi-cat facilities. Diagnosis of even a symptomatic
cat is also difficult since FIP can appear clinically in two forms. In
the so-called effusive (or wet) form, FIP cats develop a great deal of
peritoneal fluid and die quickly, but in the noneffusive (or dry) form
of FIP, symptoms include general lethargy, and a variety of systemic problems
including kidney and liver failures. CNS symptoms, etc, any of which could
also be due to a number of other causes. Even the effusive FIP symptoms
could have many other causes, including cancer, and bacterial peritonitis.
The definitive diagnosis of FIP is usually post marten, when sections of
inflammatory sites are examined microscopically by a pathologist for classic
FIP appearance. Both the effusive and noneffusive forms of FIP have similar
histologic lesions. The severity of the effusive form is usually due to
an ineffective immune response with a marked deposition of immune complexes
in small blood vessels resulting in marked degenerative and proliferative
vascular changes. Lesions form and may be found in the kidney, liver, mesenteric
lymph nodes, spleen, pancreas, and parietal serosal surfaces. The thorax,
eye, and brain may also have histologic lesions of FIP.
The challenge, then was to create a diagnostic test that ca n distinguish
relevant FIPV strains from the numerous other forms of corona viruses,
and that can be used by the clinician to make a definitive diagnosis of
the symptomatic cat. This challenge has been addressed by the new PCR technique
that tests not for coat proteins of the virus, but for the presence of
the viral GENES in the host.
How PCR works for FIPV detection is by finding FIP virus and their associated
genes in macrophages and monocytes in the blood, peritoneal fluid, or a
tissue biopsy of an infected cat. DA is studied. It is challenging, but
possible, to design primers that detect FIPV genes to the exclusion of
all other corona viruses. In theory, the gene from and single virus particle
in the sample can be amplified into billions and detected by simply observing
PCR technology can detect the virus particles, or their integrated genes,
within a single infected cell!
PCR is more specific that ELISA technology. ELISA testing relies on a single
monoclonal antibody with a single recognition site for the viral protein
to detect and bind to virus proteins in the sample; this antibody can and
does cross react with some of the millions of other proteins present in
a blood sample. This is the origin of the false positives and indeterminate
results than have been reported. In contrast PCR is based on the exact
sequence of nucleotides that make up the DNA docing sequence for the virus
protein. These are totally unique, and will detect viral genes to the exclusion
of all others.
PCR is so specific and so sensitive relative to ELISA that false positive
results will be unknown. False negatives are possible for several reasons.
For a cattery, this means regular PCR testing of the colony, at regular
intervals, will provide the best opportunity for detecting FIP carrier
cats in the population. The PCR test for FIP will prove a valuable tool
in helping to eliminate this disease from our cat population. The technology
employed may also someday be used for gene therapy, as a practical therapy
to treat cats already infected.
In general PCR testing will increasingly become available to the Veterinary
practitioner. PCR tests are available for Feline Leukemia Virus and Cat
Scratch Disease. Test for Toxoplasmosis, Feline Infectious Anema (FIA),
Helicobacter, will also soon be available.
Dr. Cahill is currently working to perfect his PCR test.