Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Paratuberculosis”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 577 records · Page 32Linked to original sources

A cost/benefit study of paratuberculosis certification in French cattle herds.

Paratuberculosis has received increasing attention in France because of the important losses this disease may provoke. The use of certification schemes has proven its effectiveness for the protection of healthy herds against diseases transmitted mainly by trade. The economic justification of such schemes in the particular case of paratuberculosis is studied, for French cattle herds, using a cost/benefit approach. The basic economical hypotheses and estimates have been proposed and carefully examined by a working group composed of paratuberculosis experts and field specialists. By adopting the point of view of a breeder that buys animals, we first estimated the benefits resulting from the non-introduction of the disease. They were then compared with the costs resulting from the fact that the vendor reports its own certification costs on the price of the animals he sells. Two average herds (the mean French beef herd and the mean French dairy herd), and two certification levels were studied. The results show that, currently, the use of the certification is not very economically profitable in French cattle herds. This conclusion, however should be reappraised if the certification costs decrease, for example with the commercialization of diagnostic tests on mixtures.

Animals↗

Assessment of diagnostic tools for eradication of bovine tuberculosis in cattle co-infected with Mycobacterium bovis and M. avium subsp. paratuberculosis.

The intradermal tuberculin (IDTB) test and the interferon-gamma (IFN-gamma) assay are used worldwide for detection of bovine tuberculosis in cattle, but little is known about the effect of co-infecting agents on the performance of these diagnostic tests. This report describes a field trial conducted in a cattle herd with dual infection (bovine tuberculosis and paratuberculosis) during 3.5 years. It has been based on a strategic approach encompassing serial parallel testing (comparative IDTB test, the IFN-gamma assay and serology of paratuberculosis) that was repeated 8 times over the period, and segregation of animals into two herds. The IDTB test detected 65.2% and the IFN-gamma test detected 69.6% of the Mycobacterium bovis culture-positive cattle. However, the IDTB test performed better during the first part of the trial, while the IFN-gamma test was the only method that detected infected animals during the following three samplings. The number of false positive reactors with the IDTB and/or the IFN-gamma tests was remarkably high compared to other reports, and could be caused by cross-reactivity with M. avium subsp. paratuberculosis. Also, the M. bovis isolates from cattle and wildlife from the same property were characterised using molecular techniques to disclose an epidemiological link. The IDTB test may not be appropriate to eradicate bovine tuberculosis in herds with dual mycobacterial infections. This report highlights the need to use several diagnostic techniques for the accurate detection of M. bovis infected animals in these herds.

Animals↗

Effect of glucocorticoids on cows suspected of subclinical infection with M. paratuberculosis.

Glucocorticoids were administered to 10 heifers suspected of subclinical infection with Mycobacterium paratuberculosis. Three animals remained untreated. M. paratuberculosis was isolated from the internal organs of 2 animals after this treatment but not from any of the control group. Delayed type hypersensitivity and lymphocyte reactivity towards Johnin and purified protein derivates of M. avium and M. bovis were depressed. A sharp increase in total leucocyte count, due to an increase in neutrophil numbers, occurred. In the three untreated animals these parameters did not change during the experiment. A decrease of specific immunological reactivity towards M. paratuberculosis occurred, but not to such an extent that clinical disease developed.

Animals↗

Evaluation of surgical tissue from patients with Crohn's disease for the presence of Mycobacterium avium subspecies paratuberculosis DNA by in situ hybridization and nested polymerase chain reaction.

Crohn's disease (CD) is a chronic inflammatory bowel disease (IBD) with tissue granuloma and histopathological alteration that resembles aspects in tuberculosis, leprosy, and paratuberculosis. Mycobacterium avium subsp paratuberculosis (MAP) is the causative agent of paratuberculosis, with a suspected role in the etiology of CD. We investigated the presence of MAP DNA in 31 surgical tissue samples from 20 subjects using fluorescence in situ hybridization (FISH) with the aid of confocal scanning laser microscopy and nested polymerase chain reaction (PCR) using the IS900 sequence unique to MAP. MAP DNA was detected by PCR in tissue from 10 of 12 (83%) patients with CD: 7/12 (58%) in inflamed, 6/11 (55%) in noninflamed and in 10 (83%) of either tissue and by FISH in 8 of 12 (67%) patients with CD: 7 of 12 (58%) in inflamed, 4 of 11 (36%) in noninflamed, and in 8(67%) of either tissue. In non-IBD subjects, MAP DNA was detected in the tissue of only 1 of 6 patients (17%) by PCR and 0 of 6 patients (0%) by FISH. MAP DNA was identified by PCR in inflamed tissue from 2 of 2 patients with ulcerative colitis. The detection of MAP DNA by either technique in tissue from subjects with CD is significant compared with non-IBD subjects (P < 0.005). Identification of MAP DNA in both inflamed and noninflamed tissue by both techniques suggests that MAP infection in patients with CD may be systemic. The data add more evidence toward a possible association of MAP in the pathogenesis of CD.

Adult↗

Identification of Mycobacterium paratuberculosis gene expression signals.

Mycobacterium paratuberculosis promoter-containing clones were isolated from a genomic DNA library constructed in the transcriptional-translational fusion vector pYUB76. The promoter-containing DNA fragments were identified in the surrogate host Mycobacterium smegmatis by expression of the promoterless lacZ reporter gene of pYUB76. The expression signals exhibited a wide range of strengths, as indicated by their corresponding beta-galactosidase activities. Eight clones were sequenced and characterized further. Predicted open reading frames and codon usage were identified by computer analysis. Database searching for related sequences using the BLAST method revealed no homologies. Transcriptional activity was measured by slot-blot hybridization with steady-state RNA isolated from lacZ+ M. smegmatis clones. Primer extension analysis identified the transcription start sites within the cloned fragments. The promoter regions characterized in this study were used to establish a consensus promoter sequence for M. paratuberculosis. M. paratuberculosis consensus hexanucleotide sequences of TGMCGT and CGGCCS centred approximately 35 and 10 bp upstream from the transcription startpoints do not correspond to the consensus hexanucleotides of Escherichia coli promoters.

Base Sequence↗

A low G+C content genetic island in Mycobacterium avium subsp. paratuberculosis and M. avium subsp. silvaticum with homologous genes in Mycobacterium tuberculosis.

The technique of representation difference analysis PCR has been applied to find genes specific to Mycobacterium avium subsp. paratuberculosis. This generated a 671 bp fragment which was used to isolate a larger genetic element found in the enteric pathogens M. avium subsp. paratuberculosis and M. avium subsp. silvaticum but which was absent from the very closely related and relatively benign M. avium subsp. avium. This element, designated GS, is greater than 6.5 kbp in length and has a G+C content 9 mol% lower than other genes from this species. There is a previously uncharacterized insertion sequence associated with one end. The GS element encodes five ORFs in M. avium subsp. paratuberculosis and M. avium subsp. silvaticum, all of which have counterparts encoded in Mycobacterium tuberculosis. Database searches revealed homologues for these ORFs in a number of bacterial species, predominantly Gram-negative organisms, including a number of enteric pathogens. These homologous genes encode functions related to LPS or extracellular polysaccharide biosynthesis. This element has a number of features in common with pathogenicity islands such as its low G+C content, an association with a putative insertion sequence and a grouping of genes of related function with a possible link to virulence. No direct link to pathogenicity has been shown but GS may belong to a group of related 'genetic islands' and represents the first such element to be identified in mycobacteria.

Cell Wall↗

Identification and characterization of an immunogenic 22 kDa exported protein of Mycobacterium avium subspecies paratuberculosis.

An exported 22 kDa putative lipoprotein was identified in an alkaline phosphatase gene fusion library of Mycobacterium avium subsp. paratuberculosis and expressed in Mycobacterium smegmatis. The full nucleic acid sequence of the gene encoding P22 was determined and the ORF was cloned into a mycobacterial expression vector, enabling full-length P22 to be produced as a C-terminal polyhistidine-tagged protein in M. smegmatis. N-terminal sequencing of the recombinant protein confirmed cleavage of a signal sequence. Native P22 was detected in culture supernatants and cell sonicates of M. avium subsp. paratuberculosis strain 316F using rabbit antibody raised to recombinant P22. Investigation of the presence of similar genes in other mycobacterial species revealed that the gene was present in Mycobacterium avium subsp. avium and similar genes existed in Mycobacterium intracellulare and Mycobacterium scrofulaceum. Database searches showed that P22 belonged to the LppX/LprAFG family of mycobacterial lipoproteins also found in Mycobacterium leprae and in members of the Mycobacterium tuberculosis complex. P22 shared less than 75% identity to these proteins. Recombinant P22 was able to elicit interferon-gamma secretion in blood from eight of a group of nine sheep vaccinated with a live attenuated strain of M. avium subsp. paratuberculosis (strain 316F) compared to none from a group of five unvaccinated sheep. Antibody to P22 was detected by Western blot analysis in 10 out of 11 vaccinated sheep, in two out of two clinically affected cows and in 11 out of 13 subclinically infected cows.

Animals↗

Contribution to the diagnosis of Johne's disease in cattle. Comparative studies on the validity of Ziehl-Neelsen staining, faecal culture and a commercially available DNA-Probe test in detecting Mycobacterium paratuberculosis in faeces from cattle.

In the present study, 132 selected faecal samples from clinically affected and subclinically infected cattle from dairy herds known to be affected by Johne's disease were investigated for the presence of Mycobacterium paratuberculosis using Ziehl-Neelsen staining, faecal culture and a commercially available DNA-Probe test. The sensitivity was 36.4% for Ziehl-Neelsen staining, 85.6% for faecal culture and 47.7% for the DNA-Probe test. Proving the presence of acid-fast bacteria in 49.3% of the samples from clinically affected cattle and 19.3% of those from subclinically infected cattle, Ziehl-Neelsen staining had the lowest detection rate of the three tests under investigation. Faecal culture showed the highest detection rate of M. paratuberculosis in samples from both clinically affected (84.0%) and subclinically infected (87.7%) animals. The DNA-Probe test showed a positive result in 68.0% of the samples from clinically affected cattle and 21.1% of those from subclinically infected cattle. Ziehl-Neelsen staining proved unreliable in diagnosing Johne's disease. Faecal culture was the most sensitive method for detecting M. paratuberculosis both in clinically affected and subclinically infected cattle. The sensitivity of a commercially available DNA-Probe test has to be enhanced to enable a quick and reliable diagnosis of Johne's disease.

Animals↗

Induction of cellular immune reactions by A36, an antigen complex of Mycobacterium paratuberculosis: comparison of A36 and johnin components.

Paratuberculosis (Johne's disease) is a chronic enteritis syndrome of ruminants, which is due to infection by Mycobacterium paratuberculosis. Cutaneous testing with proteins extracted from a mycobacterial culture fluid (johnin-PPD) is currently used to evaluate the cellular immune status. We have compared the components of johnin-PPD with those of the A36 complex, a thermostable macromolecular antigen (TMA) present in the cytoplasm and associated with the cell wall of M. paratuberculosis. The presence in the johnin-PPD of fifteen A36 components has been shown by Western blotting. Moreover, monoclonal antibodies, which bind respectively to the 65-kDa M. leprae heat shock protein, the 28-kDa M. leprae superoxide dismutase, and M. tuberculosis lipoarabinomannan, recognized components of the johnin-PPD. The ability of A36 to trigger delayed hypersensitivity reactions in sensitized rabbits, and to induce the proliferation of T lymphocytes from the lymph nodes of A36-sensitized mice, matched that of johnin-PPD. The homology levels of T epitopes between A36 and the TMA complexes of M. phlei, M. bovis, M. tuberculosis and M. avium were estimated, in a lymphoproliferation assay, to be 51, 52, 59 and 94% respectively. A strong cross-reactivity of A36 with an M. leprae sonicate was also observed by cutaneous testing. The A36 components within the 45.2-26.8-kDa and the 21.6-19.8-kDa ranges were proved to induce the proliferation of T lymphocytes from sensitized mice. This work supports the possible use of the A36 complex, and of some of its components, for cutaneous tests and lymphocyte proliferation assays, in order to monitor cellular immunity in Johne's disease.

Animals↗

Isolation and diagnostic potential of ISMav2, a novel insertion sequence-like element from Mycobacterium avium subspecies paratuberculosis.

A novel Mycobacterium avium ssp. paratuberculosis (M. paratuberculosis) specific insertion sequence has been identified by representational difference analysis and designated as ISMav2. ISMav2 has no similarity to known mycobacterial IS elements but shows more than 50% identity to a non-composite transposon of Streptomyces coelicolor at the DNA and protein level. ISMav2 is present in at least three copies on the genome as assessed by Southern blot analysis and its potential value as a diagnostic tool was confirmed by PCR analyses on 79 M. paratuberculosis field isolates, nine M. avium ssp. avium isolates, and the reference strains of nine other mycobacterial species.

Base Sequence↗

Putative in vitro expressed gene fragments unique to Mycobacterium avium subspecies paratuberculosis.

By a suppression subtractive hybridization based method, nine novel Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) fragments of between 318 and 596 bp have been identified and characterized. Database search revealed little or no similarity with other mycobacteria. The uniqueness and diagnostic potential of seven of these fragments in relation to M. paratuberculosis closest relative Mycobacterium avium subsp. avium (M. avium) was confirmed by species-specific PCR and Southern blot. Furthermore, RT-PCR indicated that eight of the nine fragments originate from areas of the genome that are expressed in vitro.

Blotting, Southern↗

Search for Mycobacterium paratuberculosis DNA in tissue from patients with sarcoidosis by enzymatic gene amplification.

The etiology of sarcoidosis has not yet been established, but several mycobacterial species have been implicated in the pathogenesis of this disease. We have used a nested primer polymerase chain reaction (PCR) specific for the IS900 Mycobacterium paratuberculosis insertion element to analyse paraffin-embedded mediastinal lymphoid tissue from 18 patients with pulmonary sarcoidosis verified by mediastinoscopy. Only the positive control contained M. paratuberculosis-specific DNA. The present study does not support a role for M. paratuberculosis in the pathogenesis of sarcoidosis. However, further studies are needed in order to clarify the possible role of other mycobacteria in this disease.

Adolescent↗

A reappraisal of the complement fixation test using soluble Mycobacterium avium antigen for the detection of M. paratuberculosis infection in cattle.

Serums from 263 cattle suspected of having paratuberculosis on the basis of clinical signs, were tested for antibodies to Mycobacterium paratuberculosis with a complement fixation test (CFT) employing a heat extracted, soluble M. avium antigen. Microscopic examination confirmed that 172 (65.4%) clinically affected animals had paratuberculosis, the remainder being disease-free. The specificity and sensitivity of the CFT was 92.3% and 74.4% respectively. Phenol treatment of serums before testing was compared with no treatment and was found to have no significant effect on the CFT titres. Results obtained are discussed in relation to the cause of false negative and false positive reactions.

Animals↗

Inactivation of Mycobacterium avium subsp. paratuberculosis in cow's milk by means of high hydrostatic pressure at mild temperatures.

Two strains of Mycobacterium avium subsp. paratuberculosis (3644/02 and ATCC 19698) were inoculated (approximately 6 log CFU/ml) into sterilized milk to evaluate inactivation by high hydrostatic pressure. Reductions of M. avium subsp. paratuberculosis increased with pressure level. Significant differences were also found between M. avium subsp. paratuberculosis strains and between the media used. Average reductions of 4 log CFU/ml after treatment with 500 MPa are comparable to those caused by thermal treatments.

Animals↗

Effect of turbulent-flow pasteurization on survival of Mycobacterium avium subsp. paratuberculosis added to raw milk.

A pilot-scale pasteurizer operating under validated turbulent flow (Reynolds number, 11,050) was used to study the heat sensitivity of Mycobacterium avium subsp. paratuberculosis added to raw milk. The ATCC 19698 type strain, ATCC 43015 (Linda, human isolate), and three bovine isolates were heated in raw whole milk for 15 s at 63, 66, 69, and 72 degrees C in duplicate trials. No strains survived at 72 degrees C for 15 s; and only one strain survived at 69 degrees C. Means of pooled D values (decimal reduction times) at 63 and 66 degrees C were 15.0 +/- 2.8 s (95% confidence interval) and 5.9 +/- 0.7 s (95% confidence interval), respectively. The mean extrapolated D72 degrees C was <2.03 s. This was equivalent to a >7 log10 kill at 72 degrees C for 15 s (95% confidence interval). The mean Z value (degrees required for the decimal reduction time to traverse one log cycle) was 8.6 degrees C. These five strains showed similar survival whether recovery was on Herrold's egg yolk medium containing mycobactin or by a radiometric culture method (BACTEC). Milk was inoculated with fresh fecal material from a high-level fecal shedder with clinical Johne's disease. After heating at 72 degrees C for 15 s, the minimum M. avium subsp. paratuberculosis kill was >4 log10. Properly maintained and operated equipment should ensure the absence of viable M. avium subsp. paratuberculosis in retail milk and other pasteurized dairy products. An additional safeguard is the widespread commercial practice of pasteurizing 1.5 to 2 degrees above 72 degrees C.

Animals↗

Survival and dormancy of Mycobacterium avium subsp. paratuberculosis in the environment.

The survival of Mycobacterium avium subsp. paratuberculosis was studied by culture of fecal material sampled at intervals for up to 117 weeks from soil and grass in pasture plots and boxes. Survival for up to 55 weeks was observed in a dry fully shaded environment, with much shorter survival times in unshaded locations. Moisture and application of lime to soil did not affect survival. UV radiation was an unlikely factor, but infrared wavelengths leading to diurnal temperature flux may be the significant detrimental component that is correlated with lack of shade. The organism survived for up to 24 weeks on grass that germinated through infected fecal material applied to the soil surface in completely shaded boxes and for up to 9 weeks on grass in 70% shade. The observed patterns of recovery in three of four experiments and changes in viable counts were indicative of dormancy, a hitherto unreported property of this taxon. A dps-like genetic element and relA, which are involved in dormancy responses in other mycobacteria, are present in the M. avium subsp. paratuberculosis genome sequence, providing indirect evidence for the existence of physiological mechanisms enabling dormancy. However, survival of M. avium subsp. paratuberculosis in the environment is finite, consistent with its taxonomic description as an obligate parasite of animals.

Amino Acid Sequence↗

Rapid real-time PCR assay for detection and quantitation of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk.

Using fluorescence resonance energy transfer technology and Lightcycler analysis, we developed a real-time PCR assay with primers and probes designed by using IS900 which allowed rapid detection of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk. Initially, the PCR parameters (including primer and probe levels, assay volume, Mg(2+) concentration, and annealing temperature) were optimized. Subsequently, the quantitative ability of the assay was tested and was found to be accurate over a broad linear range (3 x 10(6) to 3 x 10(1) copies). The assay sensitivity when purified DNA was used was determined to be as low as five copies, with excellent reproducibility. A range of DNA isolation strategies was developed for isolating M. avium subsp. paratuberculosis DNA from spiked milk, the most effective of which involved the use of 50 mM Tris HCl, 10 mM EDTA, 2% Triton X-100, 4 M guanidinium isothiocyante, and 0.3 M sodium acetate combined with boiling, physical grinding, and nucleic acid spin columns. When this technique was used in conjunction with the real-time PCR assay, it was possible to consistently detect <100 organisms per ml of milk (equivalent to 2,000 organisms per 25 ml). Furthermore, the entire procedure (extraction and PCR) was performed in less than 3 h and was successfully adapted to quantify M. avium subsp. paratuberculosis in spiked milk from heavily and mildly contaminated samples.

Animals↗

Identification and characterization of a novel extracellular ferric reductase from Mycobacterium paratuberculosis.

A novel extracellular mycobacterial enzyme was identified in the ruminant pathogen Mycobacterium paratuberculosis. The enzyme was capable of mobilizing iron from different sources such as ferric ammonium citrate, ferritin, and transferrin by reduction of the metal. The purified reductase had a calculated Mr of 17,000, was sensitive to proteinase K treatment, and had an isoelectric point of pH 9. Analysis of the amino acid composition revealed glycine, serine, asparagine (or aspartic acid), and glutamine (or glutamic acid) as the most frequently occurring residues. Enzymatic activity was highest at 37 degrees C and between pH 5 and 10. The calculated Km and Vmax for ferric ammonium citrate were 0.213 mM and 0.345 mM min(-1) mg(-1), respectively. Using a specific antireductase antibody in immunoelectron microscopy, we were able to detect the enzyme associated with intracellular mycobacteria in naturally M. paratuberculosis-infected bovine tissue. We prepose that the reductase of M. paratuberculosis represents an alternative strategy of mycobacteria to mobilize ferric iron and discuss its potential role in bacterial evasion of intracellular defense mechanisms.

Animals↗