William P. Shulaw, DVM, MS
Extension Veterinarian, Cattle and Sheep, Ohio State University Extension
Contaminated feed, water, bedding, and soiled udders are thought to be the major routes of spread of the organism. Young animals less than six months of age are thought to be the most susceptible to infection. Older animals are more resistant to infection and development of disease but can develop delayed hypersensitivity and antibody responses which may interfere with some diagnostic tests. Intrauterine infection in cattle has been well documented, but the ultimate disease status of the offspring is unknown. A confirmed isolation of M. paratuberculosis from a sheep fetus in our laboratory occurred in 1989 and again in 1990. Another report has demonstrated antibodies to M. paratuberculosis in the serum of colostrum-deprived lambs. It appears likely that fetuses so infected in utero could develop clinical disease as adults or young adults, but this is unknown at this time.
Once organisms enter the small intestine, they are engulfed by macrophages - the cells involved in killing foreign bacteria - located in the intestinal wall. Macrophages generally are unable to kill them. The bacteria multiply inside the macrophage which eventually dies.
The body's immune responses to the intracellular bacteria create the typical thickening and microscopic distortion of the intestinal wall. Hypersensitivity reactions may increase the cellular response.
Evidence suggests that, at times, M. paratuberculosis may be found in the blood and lymph fluid which allows the bacteria access to other parts of the body. Organisms have been isolated from the uterine wall and placenta in the cow. The bacteria are also known to be present in the colostrum and milk of some infected cows, and it is likely that this occurs in other species. This method of transmission has not been documented in sheep and goats but is likely to occur.
Emaciation, or wasting disease, unresponsive to dewormers and antibiotics is the usual sign. Appetite is often good, in spite of weight loss, until the animal is near death. Normal stools are the usual observation even in clinically diseased animals, but intermittent diarrhea or softened, pasty stools are occasionally observed. The profuse watery diarrhea seen in cattle is not a common feature of the disease in sheep and goats. Intermittent, low-grade fever may be observed. Intermandibular edema (bottle jaw), lethargy, and depression are sometimes seen. The disease may be confused with intestinal parasitism, chronic malnutrition, caseous lymphadenitis (especially animals with internal abscesses), ovine progressive pneumonia (OPP), environmental toxins, and cancer.
Culture of feces (manure) and tissue is the "gold standard" for diagnosis but may be unreliable in sheep and some other ruminant species when using standard laboratory techniques. It is the generally recognized "official" test and may require 8-24 weeks of incubation for colonies to be observed. Several reports have described difficulties in growing the bacteria from known infected sheep tissues and feces. In many laboratories, M. paratuberculosis has been readily culturable from cattle and goats, but not from sheep. Recent work in Australia and Spain suggests that broth culture methods followed by DNA-based techniques may be successful in identifying the organism from sheep feces and tissues. In goats, fecal and tissue culture using cattle-adapted techniques seems to work satisfactorily; however, our state diagnostic laboratory has encountered a goat flock with classic paratuberculosis from which M. paratuberculosis could not be grown. Similar problems have also been reported in cattle from Australia, suggesting that strain differences can make culture difficult in many species.
Gross and microscopic examination of tissues taken from dead animals showing signs of disease are reliable techniques for diagnosis; however, sheep and goats may exhibit some gross and microscopic lesions not typical of those seen in cattle. These include: (1) ridges and thickening of the small intestine and cecum are not always seen as is common in cattle; (2) the colon and the rectum are not regularly involved - disease is more often confined to the small intestine; (3) cording, or prominence of the mesenteric lymphatics, is common along with swollen mesenteric lymph nodes; and (4) microscopically, a great amount of variation can exist in the type of lesions seen. This may be a result of strain differences in the infecting organisms and of differences in the ability of the animal to mount an effective immune response.
Acid-fast staining of intestinal mucosa or lymph node smears, often performed in cattle, usually reveals the presence of typical clumps of acid-fast bacteria within macrophages. Smears of rectal mucosa biopsies are difficult to get and may not be very helpful because the disease process is less likely to involve the rectum in sheep and goats. Intradermal Johnin testing (skin testing) may yield many false positive and false negatives. It has been suggested that the absence of any positives in a screening test of all animals may suggest that the herd or flock is not infected. However, this test is not widely practiced in sheep and goats.
Many serologic tests, or blood tests, have been used to diagnose paratuberculosis in cattle and small ruminants. Complement fixation is the official test in many countries for cattle. It is sometimes used in sheep and other small ruminants destined for export from the United States. Cross reactions with other bacteria have been reported for this test in cattle, and it has a relatively low sensitivity. It is no longer considered sufficiently accurate for diagnostic purposes.
Enzyme-linked immunosorbent assay (ELISA) may be useful in the future. However, this assay lacks standardization and validation across laboratories at the present time, and the difficulty of using culture as the "gold standard" complicates validation of tests. The currently licensed test as formatted for cattle has not yet been shown to be useful in sheep. One report examining its usefulness in goats looks promising, but only relatively small numbers of animals were used, and the test method was modified. Evidence exists that infections in sheep with Corynebacterium pseudotuberculosis, the cause of contagious abscesses in sheep and goats, may cause cross reactions and false positives in ELISA.
Agar gel immunodiffusion (AGID) testing has been used to assist in the diagnosis of clinical Johne's disease in sheep, goats, and llamas. In cattle, goats, and sheep, it correlates well with heavy shedding of M. paratuberculosis in the feces. This test may be the most sensitive and specific serologic test we have for sheep; however, it probably lacks sufficient sensitivity in subclinical animals to use in a test-and-cull program. One report describing culture and AGID test use in a large infected goat flock suggested that culture was able to detect more animals at a single test than AGID tests. However, AGID testing may be useful in some control efforts and in certification programs for goats.
A DNA probe kit available from IDEXX, Inc., will detect the organism reliably, but the format and processing steps are only approved for cattle. It certainly should be useful for other species, but it is currently only formatted for use with cattle feces. Research has indicated that current technology requires that about 1,000 or 10,000 organisms per gram of feces need to be present before the test will be positive. The fecal culture techniques routinely used for cattle and goats can detect fewer numbers of bacteria in feces. The probe technique may be useful for small ruminants, but is currently not widely available and usually quite expensive.
Control of the disease in infected flocks is currently difficult because of the lack of a live animal test that will reliably detect animals that are not shedding M. paratuberculosis or which are shedding very low numbers. In goat flocks, AGID testing and culture often work well if the strains are culturable. Some suggestions for control include:
If the flock or herd is believed to be not infected, use biosecurity measures to prevent introduction of M. paratuberculosis. Some things to consider include these.
Currently, control of Johne's disease in flocks and herds of all species is difficult. Because of the nature of the organism and the slow progress of the infection, it is likely to remain so. However, we can make progress using the tools and the knowledge we have. It is important that we understand this disease and take steps to prevent further spread of the infection in our livestock species.
Kent H. Hoblet, DVM, MS Extension Veterinarian, Dairy The Ohio State University 241 Sisson Hall 1900 Coffey Road Columbus, OH 43210
Suelee Robbe, DVM, MS Veterinary Diagnostic and Production Animal Medicine 1701 Veterinary Medicine Iowa State University Ames, IA 50011
Susan M. Stehman, VMD, MS Senior Extension Veterinarian Diagnostic Laboratory P.O. Box 5786 College of Veterinary Medicine Cornell University Ithaca, NY 14852-5786
Figure 1. Johne's disease may be confused with other wasting diseases. The ewe in the foreground has Johne's disease. The black-faced ewe has caseous lmyphadenitis.
Figure 2. This sample of small intestine from a sheep with Johne's disease shows the classic thickening commonly seen in cattle with the disease.
Figure 3. This sample of small intestine from a different sheep shows relatively little thickening. The apparent ridges and thickening are a shrinkage artifact.
|Figure 4. Positive results on agar gell immunodiffusion testing are an indication of heavy shedding of Mycobacterium paratuberculosis.|
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