NEW ADVANCES IN THE DIAGNOSIS OF BOVINE PARATUBERCULOSIS
Paratuberculosis is a chronic, granulomatous,mycobacterialinfection
of the intestinal tract of ruminants, characterized by delayed
onset and variable expressions of clinical disease. The
clinical disease, Johne'sDisease, typically present in adult
cattle as chronic intermittent diarrhea, weight loss, and
decreased milk production. Generally, the age of onset of
clinical signs is 2 to 5 years, although subclinical
infection may occur in utero or via the (infected) dam's colostrum/milk.
The principal mode of transmission, however, is fecal-oral,
and the primary source of paratuberculosis organisms is the
feces from these infected cattle. Because the clinical condition
is unresponsive to treatment, once Johne'sDisease has been
confirmed in a herd, the key to controlling and radicatingparatuberculosis
is through identification and subsequent culling of infected
Several tests have been developed for the identification
of Mycobacteriumparatuberculosis, an intracellular,
acid-fast organism and the etiologi-cal agent of Johne's Disease.
Some detect the presence of organisms in feces or tissue samples
(ie, rectal or regional lymph node biopsies), and others
detect serum antibodies, with the most common being complement
fixation (CF) test, agrose gel immunodiffusion (AGID) test,
and enzyme-linked immunosorbent assay (ELISA). Two recent
developments in fe-cal-detectin methods include DNA probe
and Polymerase Chain Reaction (PCR). The first was the discovery
of a repetitive DNA insertion sequence which is specific to
M.paratuberculosis. This sequence can be used as a probe
to detect the presence of M. paratuberculosis DNA in feces.
However, the sensitivity of this test (10 organisms/g feces)
is much less than that of fecal culture (10-100 organisms/g
feces). The second development was the advent of the PCR
which enabled small amounts of specific DNA segment to be
amplified million fold. PCR was reported to be able to detect
as low as 50 organisms/g feces by some researchers. However,
the commercial PCR test kit has less sensitivity (10 org/g
feces) than fecal culture.
Tests detecting serum antibodies were developed
because traditional culturing and isolation of the organisms
on Herrold's egg yolk agar (HEY) requires incubation and observation
over a period of 2-4 months (due to both the very slow-growing
nature of these organisms, as well as the variable shedding
patterns of the organisms in the feces of sub-clinically-infected
animals). While all three of the aforementioned serological
tests have shown a high degree of specificity (98-100%), the
ELISA tests have repeatedly demonstrated the highest degree
of sensitivity (48% in nonshed-ding animals, 66% in shedders).
Until recently, high ELISA specificity was not achieved, because
of three reasons:
1) the low antibody detection limit of ELISA technology;
2) the extensive cross-reactivity with other Mycobacteriaspp.
and 3) the high prevalence of antibodies to Mycobacteriaspp
in cattle. The diagnostic specificity of the currently available
ELISA tests have been achieved by the absorption of antibodies
common to mycobacte-ria by using M.phlei. The low
sensitivity of the serological tests, however, indicates that
many infected animals would not be detected; therefore, the
use of serological tests alone, particularly in herds with
a high prevalence of paratubercu-losis, may not be sufficient
to control the disease by using test-and-cull procedures.
Serological tests may be most helpful as rapid, inexpensive
screening tests to measure the prevalence of paratuberculosisin
herds or to monitor herds once the disease has been eradicated
or brought under control. Until such time as the sensitivities
of these tests can be improved upon, fecal and/or tissue testing
will remain as the definitive diagnostic test for paratuberculosis.
Tom Ervin,DVM, Class of 94 and ChingChingWu, DVM, PhD