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PubMed1987

Prevalence of Mycobacterium paratuberculosis in ileocecal lymph nodes of cattle culled in the United States.

During 1983 and 1984, ileocecal lymph node specimens were obtained from clinically normal cattle at 76 US Department of Agriculture-inspected cull cattle slaughterhouses in 32 states and Puerto Rico. Mycobacterium paratuberculosis was isolated from the lymph nodes of 119 of 7,540 cattle. The prevalence of bovine paratuberculosis was 1.6% overall, with 2.9% in dairy culls and 0.8% in beef culls. The prevalence for females and males did not appear to differ significantly.

Animals↗
PubMed1986

The prevalence of paratuberculosis in culled New England cattle.

The prevalence of bovine paratuberculosis in New England was determined to be 18% based on a random survey of abattoir cattle. Six tissues from each of 100 animals were examined by histologic and bacteriologic methods. The cecal lymph node and the ileocecal valve yielded positive cultures most frequently, but isolations were also made from liver, tonsil, colon, and ileum. On the basis of the prevalence data, the economic significance of paratuberculosis to the New England dairy industry was estimated in excess of +15.4 million annually. Field studies identified 25 infected herds in New England. One of the infected herds from Connecticut was sold at public auction, an act which resulted in the dissemination of 95 potentially infected cattle to 28 farms in 8 different states.

Animals↗
PubMed1978

In vitro lymphocyte transformation as a herd survey method for bovine paratuberculosis.

The lymphocyte-transformation (LT) test was evaluated for its potential application as a field test for bovine paratuberculosis. Using a whole blood technique, samples from 3 consecutive collection periods were subjected to 3 mycobacterial antigens and to phytohemagglutinin. The results obtained from LT were compared with conventional serologic and cultural methods. A positive LT response to johnin purified-protein derivative (PPD) or avian PPD (or both) was noted in 40% to 60% of the animals tested. The complement-fixation test yielded 4% to 6.7% positive results, the immunodiffusion test between 1.2% and 1.4%, and the direct fecal culture between 2.4% and 6%. The mean of the stimulation indices of all positively responding animals was highest with johnin PPD. Specific stimulation to mammalian PPD occurred between 2.4% and 6% of the animals. The efficacy of the LT test for determining the incidence of infection with Mycobacterium paratuberculosis is discussed.

Animals↗
PubMed1979

Observations on diagnostic tests for paratuberculosis in a deer herd.

Paratuberculosis was diagnosed in a herd of 62 fallow deer (Dama dama) and 38 sika deer (Cervus nipon) maintained in an animal park in Ohio. A comparison was made of delayed-type hypersensitivity responses, results of in vitro lymphocyte immunostimulation tests, and isolation of Mycobacterium paratuberculosis on fecal culture in 19 deer.

Animals↗
PubMed1983

Indirect immunoperoxidase test for the diagnosis of paratuberculosis.

An indirect immunoperoxidase procedure for the diagnosis of paratuberculosis was described. Formalin-fixed ileocecal tissue containing large numbers of Mycobacterium paratuberculosis organisms was used as the source of antigen. Goat anti-bovine immunoglobulin G labeled with peroxidase was used as the conjugate in the test system. The method is relatively simple to do and may prove to be valuable as a routine screening test.

Animals↗
PubMed1983

Enzyme-linked immunosorbent assay for detection of bovine immunoglobulin G1 antibody to a protoplasmic antigen of Mycobacterium paratuberculosis.

An enzyme-linked immunosorbent assay (ELISA) was developed for the detection of antibodies to Mycobacterium paratuberculosis in the sera of cattle. This assay was designed to minimize the nonspecific ELISA reactions caused by immunoglobulin (Ig)M by measuring only IgG1 antibodies against a protoplasmic antigen from the organism. The ELISA detected IgG1 antibodies in the sera of 58% of cattle with positive fecal cultures for M paratuberculosis compared with detection of 45% of culture-positive animals with an immunodiffusion test. In addition to its sensitivity, the ELISA apparently is highly specific because only 4% of the sera from fecal culture-negative animals gave a false-positive result.

Animals↗
PubMed1984

Bovine paratuberculosis: an update.

Paratuberculosis is enzootic in the Great Lakes region and northeast US, causing severe economic losses. Sheep, pigs and deer can serve as intermediate hosts. Diagnosis is difficult and there is no entirely satisfactory serodiagnostic test. While treatment is generally considered ineffective, vaccines have been used successfully for prevention. Measures that can be taken in herds to reduce losses include raising replacement animals separate from adults, slaughtering animals shedding M paratuberculosis in their feces, and decontaminating the premises.

Animals↗
PubMed1984

Diagnosis of paratuberculosis in sheep.

Paratuberculosis in sheep usually is manifested as emaciation and decreased wool production. Diarrhea occurred in only 18% of affected animals. Significant hematologic changes included decreased RBC count, hemoglobin level and hematocrit. Necropsy revealed pallor, cachexia and serous fluid in body cavities. Staining of intestinal mucosal scrapings and mesenteric lymph node impression smears for acid-fast organisms revealed bright-red clumps of Mycobacterium paratuberculosis bacilli. Fecal examination identified 70% of affected animals, intradermal injection of johnin 60%, and avian tuberculin 39%.

Animals↗
PubMed1980

Comparison of agar gel immunodiffusion and fecal culture for identification of goats with paratuberculosis.

A large goat herd naturally infected with Mycobacterium paratuberculosis has been monitored for new infections for approximately 21/2 years, using both agar gel immunodiffusion and bacterial culture of feces. On the basis of postmortem confirmation of infection status, both methods had equal diagnostic sensitivity and specificity. The agar gel immunodiffusion test offered distinct advantages in ease of performance and rapidity of test results. It was concluded that serious consideration should be given to the immunodiffusion test as a diagnostic tool in control programs for caprine paratuberculosis.

Animals↗
PubMed1977

Distribution of paratuberculosis in Wisconsin, by soil regions.

A study was undertaken to determine the relationship between concentration of paratuberculosis-infected herds and soil types in Wisconsin. Locations of farms on which paratuberculosis had occurred between 1971 and 1975 were obtained from the Wisconsin Department of Agriculture. The disease occurred mainly in the rolling and undulating areas. Fifty percent of the infection herds were in the southwestern "driftless area" and the southeast uplands; 43% occurred in the western part of the northern silty and loamy uplands; the other 7% were scattered in the east and east-central part of the state. The disease did not necessarily occur in the areas of high cattle concentration. It was concluded that the disease persisted in regions with acid soils but not in regions with alkaline, calcereous soils.

Animals↗
PubMed1990

Characterization of Mycobacterium paratuberculosis and "wood-pigeon" mycobacteria by isoenzyme profile and selective staining of immunoprecipitates.

Cell-free extracts of various strains belonging to Mycobacterium paratuberculosis (Ptb) and "wood-pigeon" (WP) mycobacteria were compared by polyacrylamide gel electrophoresis and the various protein bands obtained were tested for peroxidase enzyme activity. One strain of Mycobacterium avium served as a control. Bacterial extracts were also analysed by crossed immunoelectrophoresis (CRIEP) and fused rocket immunoelectrophoresis (FRIEP) using antisera raised in rabbit against M. paratuberculosis and WP mycobacteria. The immunoprecipitates obtained both in CRIEP and FRIEP plates were subsequently stained for selective peroxidase enzyme staining. Our results showed that, although Ptb and WP mycobacteria shared common peroxidase isoenzymes and antigens, they also had specific immunoprecipitates showing the differences between the two groups of bacteria.

Electrophoresis, Polyacrylamide Gel↗
PubMed1989

Sequence and characteristics of IS900, an insertion element identified in a human Crohn's disease isolate of Mycobacterium paratuberculosis.

The complete sequence of an insertion element IS900 in Mycobacterium paratuberculosis is reported. This is the first characterised example of a mycobacterial insertion element. IS900 consists of 1451bp of which 66% is G + C. It lacks terminal inverted and direct repeats, characteristic of Escherichia coli insertion elements but shows a degree of target sequence specificity. A single open reading frame (ORF 1197) coding for 399 amino acids is predicted. This amino acid sequence, and to a lesser extent the nucleotide sequence, show significant homologies to IS110, an insertion element of Streptomyces coelicolor A3(2). It is proposed that IS900, IS110, and similar insertion elements recently identified in disease isolates of Mycobacterium avium are members of a phylogenetically related family. IS900 will provide highly specific markers for the precise identification of Mycobacterium paratuberculosis, useful in defining its relationship to animal and human diseases.

Amino Acid Sequence↗
PubMed1992

Protein antigens secreted by Mycobacterium paratuberculosis.

Proteins secreted by Mycobacterium paratuberculosis (M.ptb) during short-term cultivations were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western(Immuno) blotting. Cultivation in a defined medium containing 35S methionine allowed autoradiographic detection of proteins which had been secreted or passively released by actively metabolizing M.ptb organisms. After the first 3 days of cultivation, 4 proteins with molecular weights of approximately 38, 50, 65 and 110 kilodaltons (kd) were detected on SDS gels. Longer incubation up to 12 days resulted in an increased concentration of these proteins as well as in appearance of additional proteins ranging from 14 to over 90 kd. In long-term (8-10 weeks) culture filtrates only two prominent proteins with molecular weights of 30 and 65 kd proteins could be detected. Immunoblot analysis showed that some of the proteins secreted during short-term cultivations were recognized by sera from M.ptb-infected sheep and more significantly by sera from animals which had been immunized with a M.ptb live vaccine strain. The study indicates that during short incubation times M.ptb may secrete immunoreactive proteins which are not dominant in long-term cultures.

Animals↗
PubMed1992

Paratuberculosis vaccine in a large dairy herd.

On a 500-cow dairy farm a total of 866 young calves less than one month old were vaccinated with a heat-killed oil-adjuvated bacterin against Mycobacterium paratuberculosis over a period of five years. The vaccinated calves were tested by faecal microscopy, bacteriology and serology on the day of vaccination, at the age of 3, 6, 9 and 12 months, at breeding age, and on the day of calving. A total of 721 bull calves and 379 female calves served as unvaccinated controls in two groups. The results were evaluated by trend analyses. Vaccination greatly reduced the faecal shedding of mycobacteria as demonstrated by the annual faecal microscopic examinations. During the last 6 months of the experiment only 9 of 612 samples were found positive by microscopy and by bacterial culture. The number of seropositive animals and the antibody titres demonstrated by the complement fixation test (CFT) and agar gel immunodiffusion (AGID) increased during the first three years. Later on, both the number of seropositive animals and CFT titres decreased.

Animals↗
PubMed1991

Antibody reactivities of Mycobacterium paratuberculosis infected sheep as analyzed by enzyme-linked immunosorbent assay and western blotting.

Antibody reactivities in sera from Mycobacterium paratuberculosis (M. ptb) infected and vaccinated sheep were analyzed by enzyme-linked immunosorbent assay (ELISA) and Western (immuno)blotting using a sonicate antigen from M. ptb. Both methods allowed good differentiation between infected/vaccinated animals and noninfected controls. Removal of nonspecific crossreactive antibodies by absorption with a M. phlei sonicate antigen coupled to Sepharose reduced ELISA reactivities of positive sera by 50% and those of noninfected serum by 85%. Immunoblotting analysis revealed that reduction by M. phlei absorption was due to lower reactivities of M. ptb antigens in the range of 30 to 45 kDa. However, one protein with a molecular mass of approx. 27 kDa seemed to be specific for M. ptb since it reacted similarly with nonabsorbed and absorbed serum but not with antibodies which were eluted from M. phlei-Sepharose after absorption. Our findings indicate that M. ptb and M. phlei share a number of common antigens of potential pathogenic importance and that only a smaller part of proteins (i.e. the 27 kDa protein) might be specific for M. ptb.

Animals↗
PubMed1992

Molecular cloning and characterization of Mycobacterium paratuberculosis promoters in Escherichia coli.

DNA fragments from Mycobacterium paratuberculosis were cloned in the promoter probe plasmid pKO1. Of 957 recombinant DNA clones, 24 induced synthesis of galactokinase (the reporter gene) when these plasmids were transformed into an Escherichia coli strain deficient for the enzyme. A DNA insert from one putative promoter-containing plasmid, designated pAG5, was sequenced and shown to contain, a characteristic RNA polymerase binding site, a probable ribosomal binding site and a putative open reading frame.

Amino Acid Sequence↗
PubMed1990

Immunohistochemical study of bovine lymph nodes with antibodies against S100 protein subunits: comparison between lymph nodes of healthy and Mycobacterium paratuberculosis-infected cattle.

Using immunohistochemistry, the differential distribution of the alpha subunit (S100 alpha) and beta subunit (S100 beta) of S100 protein was studied in mesenteric lymph nodes from normal or Mycobacterium paratuberculosis-infected cattle. In epithelioid cell granulomas, S100 alpha-positive epithelioid cells and some giant cells were scattered among S100 alpha-negative cells, which were predominant. The S100 beta-positive and -negative cells contained acid-fast bacilli. The presence of S100 beta-positive cells was not demonstrated in the granulomas. In normal component cells in the lymph nodes, follicular dendritic cells in the germinal centres and endothelium of lymphatic sinus and lymph vessels were positive for S100 alpha. S100 beta was positive only in the endothelial cells of blood vessels. Results shown in the present paper are discussed in light of results obtained in other work on human tissues using the same sources of antibodies.

Animals↗
PubMed1992

Mycobacterium paratuberculosis binds fibronectin.

Fibronectin, an adhesive glycoprotein which is present in plasma and on many host cell surfaces of many host organisms, binds to certain bacterial pathogens. This study demonstrates the ability of Mycobacterium paratuberculosis (M.ptb) to interact with 125I-labelled fibronectin purified from bovine and ovine plasma. Two M.ptb strains were tested: a clinical isolate and a commercially available vaccine strain. Both strains showed significant fibronectin-binding activities of 22 and 41%, respectively, whereas non-pathogenic M.phlei had almost no affinity for fibronectin. Binding activities were similar for ovine and bovine fibronectin. We found that fibronectin binding by M.ptb was (1) time-dependent, reaching saturation within 90 min, (2) specific, since it was inhibited by an excess of unlabelled fibronectin but not by albumin, (3) saturable, with an apparent dissociation constant of 1.25 x 10(-9) M and a maximal number of 1,600 binding sites per bacterium, and (4) sensitive to detergents, proteases and heat treatments, indicating the protein nature of the responsible binding component(s). Scatchard plot analysis gave a straight line suggesting the presence of a single type of fibronectin receptor on M.ptb.

Animals↗