Paucibacillary paratuberculosis in a goat

A 5-year-old Saanen doe was presented with emaciation, good appetite, normal feces, and a dry flaky hair coat. Johne's disease was tentatively diagnosed. Gross postmortem findings were a thickened, corrugated ileum and enlarged edematous mesenteric lymph nodes. Histologically, there was extensive lymphocytic infiltration of the ileal lamina propria. Mycobacterium paratuberculosis could not be identified or cultured.     

The emergence of Mycobacterium paratuberculosis in farmed deer in New Zealand - a review of 619 cases

AIM: To review cases in which Mycobacterium paratuberculosis was identified in farmed deer in New Zealand. METHODS: Case histories were reviewed where M. paratuberculosis was identified in deer by either culture or a polymerase chain reaction (PCR) test using primers from IS900. RESULTS: Between 1986 and 2000, M. paratuberculosis was identified by bacterial culture and/or PCR in 619 farmed deer from 299 herds, representing approximately 6% of deer herds in New Zealand. Over 85% of cases were identified during the last 6 years. In 60% of the infected herds, only one infected animal was identified. The maximum number of cases identified in a single deer herd was 47, and these were identified over a period of 8 years. Only 36 (5.8%) cases came from clinically affected animals identified on farms by veterinarians. The majority (89.7%) of the 619 cases were identified from lesions in mesenteric lymph nodes, including the ileocaecal lymph nodes, identified at meat inspection as being macroscopically either typical or equivocal of bovine tuberculosis (M. bovis). While the overwhelming majority of lesions were identified in mesenteric lymph nodes, M. paratuberculosis was also identified in 27 lesions in lymph nodes of the head, especially the retropharyngeal lymph node. CONCLUSIONS: The figures presented underestimate the true prevalence of infection with M. paratuberculosis, especially since not all suspect cases were submitted for culture or PCR. However, they do show that M. paratuberculosis appears to be spreading in farmed deer in New Zealand and highlight the possibility that Johne's disease is emerging as a potential major problem affecting this species. Identification of the organism by bacterial culture or PCR is required in many cases to distinguish lesions in mesenteric lymph nodes and lymph nodes of the head caused by M. paratuberculosis from those caused by M. bovis and M. avium.     

The pathology of spontaneous paratuberculosis in the North American bison (Bison bison)

Gross and histopathologic examinations were performed on 70 North American bison (Bison bison) from a Mycobacterium avium paratuberculosis culture-positive herd. The bison examined were part of a breeding herd totaling 2,800 animals. Eight of 70 (11%) animals had gross findings of intestinal mucosal thickening, and 16 of 70 (23%) of the animals had enlarged mesenteric lymph nodes. Histologic lesions compatible with Johne's disease were diagnosed in 30 of 70 (43%) bison on the basis of the demonstration of noncaseating granulomatous inflammatory infiltrates and of one or more acid-fast bacilli characteristic of Mycobacterium avium paratuberculosis. A suspicious diagnosis of Johne's disease was obtained in 11 of 70 (16%) bison on the basis of the observation of noncaseating granulomatous inflammatory infiltrates without demonstrable acid-fast bacteria. Twenty-nine of 70 (41%) animals were assessed as histologically paratuberculosis free. Histologic results were compared to Johne's disease tests such as culture, serology, and polymerase chain reaction, which were performed on some of the cohort animals.     

Pathogenic 'Bison-type' Mycobacterium avium subspecies paratuberculosis genotype characterized from riverine buffalo (Bubalus bubalis) in North India

Despite low per-animal productivity of ruminants in developing countries, Johne's disease has not been investigated in buffaloes, which are primarily found in these countries. This is due to lack of expertise, diagnostic kits and priority to production diseases like Johne's disease. Presence of pathogenic Mycobacterium avium subspecies paratuberculosis (Map) was investigated by screening of target tissues (mesenteric lymph nodes and large intestine) by culture and IS 900 PCR, in 50 sacrificed buffaloes. Indigenous ELISA kit originally developed for goats and sheep was standardized in buffaloes and used to estimate sero-presence of Map in 167 serum samples representing population of buffaloes in Agra region of North India. In culture, 48.0% buffaloes were positive from 50 tissues each from mesenteric lymph nodes (34.0%) and large intestine (36.0%). IS 900 PCR was standardized using specific primers (150 C and 921) and 229 bp-amplified product was characteristic for Map. Of the 25 mesenteric lymph nodes, 40.0% were positive in IS 900 PCR. Genomic DNA from Map cultures was successfully amplified from all the 24 isolates (100.0%). Map was further genotyped as 'Bison type' using IS 1311 PCR-REA. Culture of tissues showed high presence of Map in target tissues, despite high culling rate in buffalos in view of high demand of buffalo meat. Specific tissue-PCR provided rapid confirmation of Map infection in sacrificed buffaloes. In tissue-PCR, all the cultures were positive as compared to 40.0% detected directly from tissues. ELISA kit using indigenous protoplasmic antigen was highly sensitive as compared to commercial antigen in detecting Map infection therefore, could be used as 'Herd Screening Test' in buffaloes against Johne's disease. This pilot study first time reports a highly pathogenic 'Bison-type' genotype of M. avium subspecies paratuberculosis from the riverine buffaloes (Bubalus bubalis) of Agra region in North India.     

Cross species transmission of ovine Johne's disease from sheep to cattle: an estimate of prevalence in exposed susceptible cattle

OBJECTIVE: To determine the prevalence of infection of cattle with the sheep strain of Mycobacterium avium subsp paratuberculosis at least two years after exposure at < 6 months old. DESIGN: Prospective survey One thousand seven hundred and seventy-four cattle from 12 properties (Farms A to L) were sampled by ELISA and faecal culture to detect evidence of infection with M a paratuberculosis. All properties had a known history of Johne's disease (JD) in sheep, and sampled cattle were likely to be susceptible to JD at the time they were first exposed, being at an age of 6 months or less. In addition, opportunistic investigations were undertaken of ELISA reactor cattle discovered during testing for the Australian Johne's Disease Market Assurance Program for Cattle (Farms M and N). RESULTS: All animals in the survey gave negative results on serology while one animal from a herd of 349 gave a positive faecal culture result. Follow-up faecal culture, post-mortem and histopathology on the latter animal were negative, suggesting that it was a passive faecal shedder or carrier. Two occurrences of OJD transmission to cattle were detected during the opportunistic investigations. CONCLUSION: These observations confirm existing beliefs about the risk of transmission of OJD to cattle, that the risk of transmission is low. However transmission occurs sporadically. An estimated upper limit of prevalence of S strain M a paratuberculosis infection in susceptible exposed cattle in the OJD high prevalence area of New South Wales is 0.8%, assuming a common prevalence within herds.     

Lymphosarcoma in 10 New World Camelids

We evaluated the medical information from 10 New World Camelids with lymphosarcoma. Additionally, tumor tissue from 5 animals was examined by electron microscopy for evidence of retroviral particles. Lymphosarcoma was the most common neoplasm of New World Camelids in our hospital. Both llamas and alpacas, from 4 months to 15 years of age, were affected. Emaciation (7 of 8) and palpable masses (9 of 10) were the most common physical examination abnormalities, and a left-shifted leukogram (7 of 7) and hypoalbuminemia (6 of 7) were the most characteristic clinicopathologic findings. Lymphosarcoma was diagnosed by lymph node aspirate (n = 2), biopsy (n = 2), peritoneal fluid analysis (n = 2), or necropsy (n = 4). The clinical course after recognition of the disease was usually short, with a median survival of 1 month (1 week to 3 months). Organs with neoplastic infiltrates commonly included lymph nodes (n = 8), liver (n = 8), kidneys (n = 6), and lungs (n = 5). No retroviral particles were detected ultrastructurally.     

Camelid mucoutaneous fibropapillomas: clinicopathologic findings and association with papillomavirus

Five camelid mucocutaneous fibropapillomas with histologic features similar to equine sarcoids were diagnosed. They were characterized by a dermal fibroblastic proliferation and overlying, often ulcerated hyperplastic epidermis with thin rete pegs extending down into the dermis. Two of the tumors came from llamas and three from alpacas. Four of the animals were 6-year-old females. The fifth was a 6-year-old castrated male. The fibropapillomas were located on the nose, lip, and cheeks. One of the llama tumors waxed and waned before surgery and recurred and spread after surgery. None of the other tumors recurred. All five tumors were positive for papillomavirus (PV) DNA by polymerase chain reaction testing. Nucleotide sequence analysis of the PCR product from one of the llama fibropapillomas confirmed a unique PV. This report provides the microscopic and clinical features of fibropapillomas in camelids as well as evidence for a PV etiology.     

Intra-uterine transmission of paratuberculosis (Johne's disease) in farmed red deer

AIM: To determine whether intra-uterine transmission of paratuberculosis (Johne's disease) occurs in farmed red deer (Cervus elaphus) in New Zealand. METHODS: On four different farms, nine late-stage pregnant hinds with Johne's disease were slaughtered and samples were taken from them and their 10 fetuses. Samples of the hepatic, ileocaecal and mesenteric lymph nodes and the posterior ileum were collected from the hinds. The lung, liver, spleen, jejunum and ileum from the fetuses were sampled, as were the placentomes. Blood samples were tested using the 'Paralisa' test, a modified immunoglobulin G1 (IgG1) enzyme-linked immunosorbent assay (ELISA). Tissue samples were cultured using the BACTEC system, and fixed samples were sectioned and histological slides examined. RESULTS: All nine hinds and 9/10 fetuses (one hind had twins) were culture-positive for Mycobacterium avium subsp paratuberculosis (M. ptb). Six hinds had gross lesions of Johne's disease, while all hinds had characteristic histopathological lesions affecting the ileum, ileocaecal valve and associated lymph nodes. The only histopathological change observed in the fetuses was some mild inflammation in the lungs of one individual. Acid fast organisms (AFOs) were seen in histological sections of the lymph nodes and ileum of six hinds, and none were seen in tissues from the fetuses. These six hinds were Paralisa-positive, whereas the remaining hinds and fetuses were serologically-negative. CONCLUSIONS: These results confirm that there is a high risk of transmission of M. ptb from clinically affected hinds to their fetuses during pregnancy. CLINICAL RELEVANCE: Johne's disease is an increasingly important disease responsible for deaths in young red deer. Recognising the influence of intra-uterine transmission on the spread of this disease may be an important step towards improved control of Johne's disease.     

Diagnosis of Johne's disease in cattle using mesenteric lymph node biopsy: accuracy in clinical suspects.

Histopathological examination of biopsies of mesenteric lymph node obtained from the region of the terminal ileum proved to be a very accurate method of diagnosing Johne's disease in the living animal. Identical conclusions were reached in 100 clinical suspects following examination of either mesenteric lymph nodes alone, or a wider range of routine specimens including ileum, caecum, colon and ileocaecal lymph nodes. A method of obtaining biopsy specimens from mesenteric lymph nodes is described and is recommended when an early definitive diagnosis is required.     

Clinical, ultrasonographic, and laboratory findings in 12 llamas and 12 alpacas with malignant round cell tumors

Clinical signs, duration of illness, clinicopathologic findings, and ultrasonographic findings were evaluated in 12 llamas and 12 alpacas with malignant round cell tumors (MRCT). All but 1 animal died or was euthanized. Common clinical findings were anorexia, recumbency or weakness, and weight loss or poor growth. Peripheral lymphadenomegaly occurred in only 7 animals and was detected more often at necropsy than during physical examination. Common clinicopathologic abnormalities were hypoalbuminemia, acidosis, azotemia, anemia, hyperglycemia, and neutrophilia. Ultrasonography detected tumors in 4/6 animals. Cytologic evaluation of fluid or tissue aspirates or histopathology of biopsy tissue was diagnostic in 5/6 cases. A clinical course of 2 wk or less prior to death or euthanasia was more common in animals ≤ 2 y of age (9/11) than in older animals (6/13). Regular examination of camelids to include clinical pathology and evaluation of peripheral lymph nodes may result in early detection of MCRT.     

Mycobacterial diseases of deer

The most significant mycobacterial diseases of free-living, captive and farmed deer are bovine tuberculosis, caused by Mycobacterium bovis, Johne's disease (paratuberculosis), caused by Mycobacterium avium subsp paratuberculosis (basonym M. paratuberculosis), and avian tuberculosis, caused principally by M. avium subsp avium. The first case of M. bovis infection in farmed deer was identified in New Zealand in 1978. In 1983, a voluntary scheme was introduced in New Zealand to control tuberculosis in farmed deer, followed by a compulsory tuberculosis control scheme in 1990. The primary control measure is the slaughter of infected animals, detected by skin testing and blood testing, together with movement control and vector control. The number of infected deer herds peaked in the mid 1990s at over 160 herds, but by 30 June 2002 this had been reduced to 79 (1.45%), and to 67 (1.23%) by June 2003. Deer-to-deer transmission occurs, but the majority of herd breakdowns are believed to be from infected vectors. Factors likely to affect the susceptibility of deer include age, environment, population density, exposure and genetics. Avian tuberculosis occasionally causes clinical disease in wild, captive and farmed deer in New Zealand and overseas. Mycobacterium intracellulare, and subspecies of M. avium other than M. paratuberculosis, are widespread throughout New Zealand and are thought to be largely responsible for the high level of sensitisation to avian purified protein derivative (PPD), which is used for comparison purposes in tuberculosis skin testing of deer in this country. Infections with these organisms are usually subclinical in farmed deer, although M. avium subsp avium commonly causes lesions in retropharyngeal, mesenteric and ileocaecal lymph nodes. These lesions cause problems because of their gross and microscopic similarity to those due to M. bovis infection. Birds and domestic animals are most likely to become infected via environmental contamination of food, water, bedding litter or soil, while carnivores or scavengers may also become infected by ingesting infected carcasses. Johne's disease has been reported in deer in the wild and in zoos, especially in North America, the United Kingdom (UK) and Europe. Since first being confirmed in farmed deer in New Zealand in 1979, the incidence of Johne's disease has increased steadily. To date, M. paratuberculosis has been identified in >600 farmed deer on 300 properties. The majority of cases have been identified from suspected tuberculous lesions submitted from deer slaughter plants. Clinically, Johne's disease in deer is similar to the disease in sheep and cattle, with typical signs of loss of weight and condition, and diarrhoea. However, outbreaks of Johne's disease frequently occur in young red deer, 8-15 months of age, whereas the clinical disease in sheep and cattle is sporadic and usually affects adults 3-5 years of age. The disease is characterised by a chronic granulomatous enteritis and lymphadenitis, especially affecting the jejunum and ileum and the mesenteric lymph nodes. Deer affected subclinically may have lesions in these lymph nodes at slaughter, which are grossly indistinguishable from those due to bovine tuberculosis. Because of the antigenic similarity between M. intracellulare and all the subspecies of M. avium, including M. paratuberculosis, the diagnostic tests for Johne's disease lack sensitivity and specificity, making control difficult.     

Effect of compliance with recommended calf-rearing practices on control of bovine Johne's disease

OBJECTIVE: To assess the degree of compliance with recommended management procedures for the control of bovine Johne's disease and study the relationship between aspects of calf management and testing/disease outcomes in the herds. PROCEDURE: Fifty-four south Gippsland dairy herds participating in the Victorian bovine Johne's disease test and control program were visited between July and November 2002 and an audit of calf rearing practices was conducted. The results of testing completed under the program were analysed for each of the herds. Twenty seven management factors were examined for a relationship with the presence of clinical cases of Johne's disease or cattle with positive ELISA test results that were born after the completion of the second whole herd test. Logistic regression was used to examine the strength of relationships between the management practices and the frequency with which new cases of Johne's disease arose. RESULTS AND CONCULSIONS: Calves were removed from their dams within 12 hours of birth in only 17 (31.5%) of the herds. However, in all but one herd the calves were removed within 24 hours of birth. In 42 herds (77.8%) calf rearing facilities were adequately separated from adult cattle and the faeces from adult cattle. In 41 herds (75.9%) calves up to the age of 12 months were grazed on paddocks that were free of manure or effluent from adult cattle. However, in only 10 (18.5%) of the herds were all three of these calf management practices applied. Feeding whole milk containing antibiotic residues, or providing water for calves from birth, were found to have statistically significant associations with an increased occurrence of cases of bovine Johne's disease in the study herds. The practice of allowing cows to calve in a paddock was found to be associated with reduced occurrence of bovine Johne's disease. These associations were still found after analysis that included herd size, the number of clinical cases that had occurred in the herds before the start of testing, the number of animals with positive ELISA tests that were detected at the first test and the number of years of participation in the test and cull program. Early separation of newborn calves from cows and grazing calves under 12 months of age in areas free of adult cattle were not found to be protective against Johne's disease.     

Protection in alpacas against Corynebacterium pseudotuberculosis using different bacterial components

Corynebacterium pseudotuberculosis is a Gram positive bacterium that produces caseous lymphadenitis in sheep and goats, and a granulomatous lymphadenitis in llamas and alpacas. To evaluate the immune potential of different doses of cell wall and toxin components of C. pseudotuberculosis from alpaca origin, 12 adult alpacas were allotted at random to four groups, and SC inoculated in the left flank with vaccines composed of low and high doses of bacterial crude antigens, cell wall: 250 and 500 microg/ml and toxin: 133 and 265 microg/ml, respectively. The vaccines were supplemented with 20 microg/ml of muramyl dipeptide as adjuvant. Three alpacas were sham inoculated with adjuvant as a control. After 3 weeks, immunized and naive alpacas were challenged intradermally in the right flank with 1 x 10(6) colony forming units (CFU) of C. pseudotuberculosis. The alpacas were sacrificed at days 28, 58 and 112 after inoculation, and the degree of protection induced by vaccines was demonstrated by the absence of abscesses and/or bacteria. The alpacas vaccinated with high dose of toxin, did not show abscesses. In contrast, the alpacas vaccinated with a low dose of toxin showed abscesses at the inoculation site, regional, and renal lymph nodes. The cell wall vaccinated alpacas showed a lesser degree of protection than the other groups with superficial and internal abscesses. The control alpacas had persistent fever and abscesses at the inoculation site, regional, and internal lymph nodes. In addition, a robust and early humoral response was observed in all vaccinated alpacas after challenge, lasting at least 3 months. The results suggest that the toxin of C. pseudotuberculosis is a very important antigen, inducing a dose dependant protective immunity against this bacterium in alpacas.     

The histopathologic diagnosis of subclinical Johne's disease in North American bison (Bison bison)

The morphologic changes of subclinical Johne's disease in North American Bison (Bison bison) are characterized by microgranulomas composed of epithelioid macrophages and individual multinucleate giant cells of Langhans'-type occasionally containing individual cytoplasmic acid-fast bacilli compatible with Mycobacterium avium paratuberculosis. The microgranulomas are best visualized in the mesenteric lymph nodes of infected subclinical animals. Macrophages that can be confused with infection-associated epithelioid macrophages in the mesenteric lymph nodes are pigment-carrying cells from the intestinal tract. Mesenteric lymph node biopsy may be a useful diagnostic tool for detection of mild subclinical infection in individual ruminants from herds of unknown infection status. The biopsy may also be useful for Johne's disease surveillance during test-and-cull programs.     

Effect of a test and control program for Johne's disease in Victorian beef herds 1992-2002

OBJECTIVE: To report on progress in Johne's disease (JD) control in infected beef herds participating in the Victorian Johne's disease test and control program (TCP). PROCEDURE: Clinical histories and JD testing data recorded by the Department of Primary Industries were analysed for 18 beef herds participating in the TCP. The herds were required to conduct annual whole herd tests with an absorbed ELISA, cull reactors and control the grazing of young cattle to minimise infection. RESULTS: Testing of over 11,000 animals identified 68 reactors giving an average prevalence of reactors at the first whole-herd test round (T1) of 0.77%. There had been 20 clinical cases detected in the 7 years before the TCP started and two cases detected in the 10 years after TCP started. Most reactors and all clinical cases were born before the TCP started. Of 34 reactors necropsied, 25 (74%) were confirmed to have JD by histology or culture of tissues. The modal age of reactors and clinical cases was 5 and 6 years respectively. Six herds completed the program by achieving three successive negative whole herd tests, four herds dropped out and eight continued to test. There were 18 reactors detected at T1 and 33 reactors that were negative at T1 but detected at subsequent tests. CONCLUSION: The TCP was associated with a marked decline in clinical cases. The similar age distributions of clinical cases and reactors probably meant that testing detected animals for which clinical disease was imminent. Whether the measures used in the TCP were adequate to control the disease in beef herds could not be determined because of the long incubation period of the disease. The relatively high proportion of the reactors investigated that were confirmed as infected provided confidence that the test was continuing to operate at a high specificity. If all unconfirmed reactors were presumed to be uninfected, the minimum specificity of the ELISA was 99.83%. The sensitivity of the ELISA appeared to be very low because of the large number of reactors that were negative at T1 but positive at later tests.     

Toll-like receptor genes are differentially expressed at the sites of infection during the progression of Johne's disease in outbred sheep

Toll-like receptors (TLR) are engaged by ligands on microbial pathogens to initiate innate and adaptive immune responses. Little is known about TLR involvement during infection with Mycobacterium avium subsp. paratuberculosis (M. ptb), the cause of Johne's disease in ruminants, although there is a profound immunopathological response in affected animals. We have analyzed the expression of 10 TLR genes relative to validated reference genes at predilection sites in ileum, jejunum and associated lymph nodes as well as in peripheral blood, to determine if TLR expression is altered in response to infection with M. ptb in outbred sheep. Previously unexposed animals from two flocks and animals from three naturally infected flocks were used with restricted maximum likelihood linear mixed modeling applied to determine significant differences. These were related to the pathologies observed at different stages of infection in exposed sheep, after allowing for other sources of variation. In most cases there were differences in TLR expression between early paucibacillary and multibacillary groups when compared to uninfected sheep, with most TLRs for the paucibacillary group having lower expression levels than the multibacillary group. Increased expression of TLR1-5, and 8 was observed in ileum or jejunum, and TLR1-4, 6, and 8 in mesenteric lymph nodes. There was a trend for increased expression of TLR1, 2, and 6-8 in PBMCs of exposed compared to non-exposed animals. Further study of TLR expression in Johne's disease in ruminants is warranted as these observed differences may help explain pathogenesis and may be useful in the future diagnosis of M. ptb infection.     

Direct detection of Ehrlichia canis by PCR in the conjunctiva of a dog with bilateral anterior uveitis

The following report describes the direct detection of Ehrlichia canis by real-time PCR in the conjunctiva of a 1-year-old female Maltese dog. After being imported from Brazil, the dog was presented because of anorexia, dehydration, fever, and palpable mandibular lymph nodes. A few days later, the dog developed bilateral blepharospasm, photophobia and anterior uveitis. Monocytic ehrlichia was diagnosed by a positive PCR result and the detection of IgM and IgG antibodies. Because of the massive uveitis a conjunctival sample was taken with a cytobrush, which also tested positive for Ehrlichia canis DNA by real-time PCR. Only one week after starting treatment with systemic doxycycline and local anti-inflammatory and cyclopalgic therapy the dog recovered from systemic and eye diseases. After therapy the follow-up examination revealed a full remission of clinical and hematological parameters and negative PCR result.     

Lymph node staging of oral and maxillofacial neoplasms in 31 dogs and cats

A retrospective study was performed to report the histologic examination results of regional lymph nodes of dogs and cats with oral or maxillofacial neoplasms. Twenty-eight dogs and 3 cats were evaluated. Histologic examination results of standard and serial tissue sectioning of regional lymph nodes were recorded. When available, other clinical parameters including mandibular lymph node palpation, thoracic radiographs, and pre- and postoperative fine needle aspiration of lymph nodes were compared with the histologic results. Squamous cell carcinoma, fibrosarcoma, and melanoma were the most common neoplasms diagnosed in dogs. Squamous cell carcinoma and fibrosarcoma were diagnosed in cats. Of the palpably enlarged mandibular lymph nodes, 17.0% had metastatic disease histologically. Radiographically evident thoracic metastatic disease was present in 7.4% of cases. Preoperative cytologic evaluation of the mandibular lymph node based on fine needle aspiration concurred with the histologic results in 90.5% of lymph nodes examined. Postoperative cytologic evaluation of fine needle aspirates of regional lymph nodes concurred with the histologic results in 80.6% of lymph nodes examined. Only 54.5% of cases with metastatic disease to regional lymph nodes had metastasis that included the mandibular lymph node. Serial lymph node sectioning provided additional information or metastasis detection. Cytologic evaluation of the mandibular lymph node correlates positively with histology, however results may fail to indicate the presence of regional metastasis. Assessment of all regional lymph nodes in dogs and cats with oral or maxillofacial neoplasms will detect more metastatic disease than assessing the mandibular lymph node only.