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M Coene

Publications and source records attributed to M Coene.

At least 19 recordsLinked to original sources
PubMed1994

Immunological analysis of the components of the antigen complex A60 of Mycobacterium bovis BCG.

The antigen complex of A60 of Mycobacterium bovis BCG was analyzed by different immunological techniques to assess its relevance to tuberculosis and the involvement of its components in the immune reactions elicited in humans by tuberculous infection. A60 is composed of about 30 components, of which 8 were identified by available monoclonal antibodies (lipoarabinomannan, a glycolipid, and proteins of 65, 40, 38, 35, 19, and 14 kDa). The majority (87.5%) of anti-mycobacterial antibodies in sera from tuberculosis patients was directed against A60. Western blot (immunoblot) analysis indicated that the majority of the highly antigenic proteins present in mycobacterial homogenates were components of the A60 complex. A small percentage (7.8%) of A60 epitopes proved to be species specific. Thus, A60 proteins of 66, 41, 38, 37, 35, 34, 32, and 22 kDa were found to contain B-cell epitopes specific for M. bovis and not shared by Mycobacterium leprae oR Mycobacterium avium.

Antibodies, Bacterial↗
PubMed1994

Occurrence, in Crohn's disease, of antibodies directed against a species-specific recombinant polypeptide of Mycobacterium paratuberculosis.

Sera from patients with Crohn's disease and control were analyzed by an enzyme-linked immunosorbent assay based on the Mycobacterium paratuberculosis-specific recombinant polypeptide a362. Anti-a362 immunoglobulin G (IgG) (P < 0.05) and IgA (P < 0.001) titers were higher in patients with Crohn's disease than in controls. A monomodal Gaussian distribution of anti-a362 IgA levels were found for controls, and a bimodal distribution was found for patients with Crohn's disease. An M. paratuberculosis etiology is suggested for the 36% of patients with Crohn's disease who had an anti-a362 IgA level higher than that of controls.

Antibodies, Bacterial↗
PubMed1994

Paratuberculosis.

Paratuberculosis (Johne's disease) is a chronic, wasting, widespread mycobacteriosis of ruminants. It involves extensive mycobacterial shedding, which accounts for the high contagiousness, and ends with a fatal enteritis. Decreases in weight, milk production, and fertility produce severe economic loss. The DNA of the etiological agent (Mycobacterium paratuberculosis) has a base composition (66 to 67% G+C) within the range of that of mycobacteria (62 to 70% G+C), a size (4.4 x 10(6) to 4.7 x 10(6) bp) larger than that of most pathogenic mycobacteria (2.0 x 10(6) to 4.2 x 10(6) bp), and a high relatedness (> 90%) to Mycobacterium avium DNA. However, the DNAs of the two organisms can be distinguished by restriction fragment length polymorphism analysis. M. paratuberculosis genes coding for a transposase, a cell wall-associated protein (P34), and two heat shock proteins have been cloned and sequenced. Nucleic acid probes (two of which are species specific) are used, after PCR amplification, for M. paratuberculosis identification in stools and milk. As in leprosy, with disease progression, cellular immune reactions decrease and humoral immune reactions increase. Cutaneous testing with sensitins, lymphocyte proliferation assays, and cytokine tests are used to monitor cellular immune reactions in paratuberculosis, but these tests lack specificity. Complement fixation, immunodiffusion, and enzymometric tests based on antibodies to M. paratuberculosis extracts, to mycobacterial antigen complex A36, to glycolipids, and to proteins help identify affected cattle but are not species specific. The carboxyl-terminal portion of the 34-kDa cell wall-associated A36 protein (P34) carries species-specific B-cell epitopes and is the basis for an enzyme-linked immunosorbent assay. Diagnostic tests for paratuberculosis are also used in Crohn's disease, a chronic human ileitis mimicking Johne's disease, in which isolates identified as M. paratuberculosis have been found.

Amino Acid Sequence↗
PubMed1994

Development of species-specific enzyme-linked immunosorbent assay for diagnosis of Johne's disease in cattle.

The previously described (M. De Kesel, P. Gilot, M.-C. Misonne, M. Coene, and C. Cocito, J. Clin. Microbiol., 31:947-954, 1993) a362 recombinant polypeptide of Mycobacterium paratuberculosis was used as reagent for an enzyme-linked immunosorbent assay (ELISA). This ELISA, which is endowed with species specificity with respect to the other mycobacteria, was applied to the analysis of bovine paratuberculosis (Johne's disease), an endemic mycobacteriosis of cattle caused by M. paratuberculosis. The distribution of anti-a362 antibodies in the cattle population was analyzed by a computer program (mixture population model) to determine a cutoff value for the test. The prevalence of a362 seropositivity in the Belgian bovine population was estimated to be 12%. The sensitivity of the a362 assay was 70%, as determined with reference sera from the U.S. National Repository of Paratuberculosis Specimens. Some 40% of the animals in the herds with paratuberculosis analyzed were found to be positive by the a362 assay. The latter proved to be 95% specific with respect to both healthy and tuberculous cattle.

Animals↗
PubMed1994

Comparison of thermostable macromolecular antigens from leprosy-associated bacteria.

Three types of bacteria are associated with leprosy: Mycobacterium leprae, leprosy-derived corynebacteria (LDC), and armadillo-derived mycobacteria (ADM). The immunological relationships between these three types of bacteria and Mycobacterium bovis BCG, used as a reference, were determined by cross-immunoelectrophoresis. When compared with the reference, cross-reactions were observed with a variable number of antigens: 2 in the case of strain LDC 15, 4 with M. leprae, and from 1 to 10 in the case of the ADM, depending on their subgroup. Next, thermostable macromolecular antigens (TMAs), the major cross-reactive antigens of leprosy-associated bacteria, were compared by anti-TMA antibody ELISA tests. The LDC TMAs displayed high cross-reactivity between the subgroups and lower cross-reactivity with the TMAs of M. bovis BCG. Evidence for the presence of a species-specific moiety in TMA of the different LDC was obtained by using depleted anti-TMA antisera. Western blot analysis revealed the presence of many proteins in the TMAs of LDC and M. bovis BCG, some of them being species-specific and other cross-reactive.

Antibodies, Bacterial↗
PubMed1993

Comparative analysis of the genomes of mycobacteriophages infecting saprophytic and pathogenic mycobacteria.

The genomes of a series of mycobacteriophages have been analyzed to disclose possible relationships between genetic characteristics and host range. The percent guanine-plus-cytosine in the DNA of 14 phages was found to be 34.4 to 47.5, as determined by a double-labelling procedure, which is unaffected by the presence of modified bases. The DNA of few mycobacteriophages yielded discordant values when the G + C content was estimated by buoyant density determination and by the double labelling procedure. This observation suggests the possible presence of modified bases in these genomes. The reduced susceptibility of viral DNAs to several restriction endonucleases is suggestive of the occurrence of both methyladenine and methylcytosine in the genome of all the mycobacteriophages studied. Heterologous annealing among the 14 DNAs analyzed yielded 6 hybridization groups. Within one group, the homology level among viral genomes was estimated by comparing the electrophoretic mobilities of restriction fragments: values of 0.8 to 1.3% base substitution have thus been found. A comparison of the genomic characteristics and host range of the mycobacteriophages analyzed suggests a possible relationship between restriction pattern, G + C content, crosshybridization level and host range.

Base Composition↗
PubMed1993

Cloning and expression of portions of the 34-kilodalton-protein gene of Mycobacterium paratuberculosis: its application to serological analysis of Johne's disease.

Paratuberculosis (Johne's disease), an endemic mycobacteriosis of cattle that is caused by Mycobacterium paratuberculosis, is characterized by incoercible diarrhea and fecal shedding of bacteria. The present work aimed at developing a specific serological test for this disease. We have recently shown that a 34-kDa protein belonging to the major antigen complex A36 of M. paratuberculosis is immunodominant and contains epitopes specific with respect to all mycobacteria tested, including Mycobacterium bovis and the closely related species Mycobacterium avium. From a lambda gt11 genomic library of M. paratuberculosis, three portions of the gene coding for this 34-kDa protein have been isolated. Two of them expressed cross-reacting mycobacterial epitopes. One portion (in clone a362) expressed a polypeptide which cross-reacted with all tested M. paratuberculosis strains but not with 20 other bacteria tested, including many strains of the M. avium-Mycobacterium intracellulare-Mycobacterium scrofulaceum group. The occurrence at the M. paratuberculosis surface of epitopes corresponding to the a362 polypeptide was shown by immune electron microscopy. The recombinant a362 polypeptide was used as reagent for an enzyme-linked immunoassay for paratuberculosis. This assay correctly diagnosed all the tested blood samples from infected cattle at all stages of the disease.

Animals↗
PubMed1993

Preparation of a specific RNA probe for detection of Mycobacterium paratuberculosis and diagnosis of Johne's disease.

A species-specific recombinant clone (F57) was obtained from a genomic library of Mycobacterium paratuberculosis in the transcription vector pGem 3Z. This clone proved to be specific for all mycobacteria tested, including M. avium, and was able to recognize all of the tested M. paratuberculosis strains isolated from animals and humans (patients with Crohn's disease). The F57 insert was sequenced and a segment of 620 bp with a G + C content of 58.9% was identified. Comparison of the sequence with sequences in the EMBL and UGEN data banks revealed the uniqueness of the F57 sequence, which had no resemblance to other known genes.

Animals↗
PubMed1992

Composition and immunological properties of the protein fraction of A36, a major antigen complex of Mycobacterium paratuberculosis.

TMA (thermostable macromolecular antigens) are major mycobacterial complexes present in all mycobacteria. We have purified A36, the TMA complex of M. paratuberculosis, the etiological agent of paratuberculosis (Johne's disease), and shown by the immune electron microscopy approach its presentation at the cell surface. The immunodominance of the A36 complex in Johne's disease was suggested by comparative ELISA analysis of infected bovine sera, using either A36 or M. paratuberculosis total soluble sonicate as antigens. The cross-reactivity of TMA complexes from different mycobacteria was evaluated by immunoenzymometric measurements. Percentage of shared epitopes was high for the couple M. paratuberculosis-M. avium, and somewhat lower for the couple M. paratuberculosis.-M. bovis. Immunological kinship between M. paratuberculosis and M. leprae was suggested by the finding that out of eleven anti-M. leprae monoclonals, four cross-reacted with A36 proteins. The specificity missing at the level of the whole A36 complex was sought at the level of its protein components. Comparative immunoblot analysis of electrophoresed A36 proteins indicated three of them to contain epitopes not shared by M. bovis proteins, and one of them to contain epitopes specific with respect to M. avium, M. bovis and M. phlei. The latter component, a 34-kDa protein, could be an ideal reagent for a serological test for Johne's disease, being immunodominant in infected cattle and endowed with species-specific epitopes.

Animals↗
PubMed1992

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↗
PubMed1990

Production of carrier-free 66Ga and labeling of antimyosin antibody for positron imaging of acute myocardial infarction.

A method for labeling monoclonal antimyosin antibodies with 66Ga is described. The radionuclide is a positron emitter (t1/2: 9.4h) produced by means of the 63Cu (4He, n) 66Ga reaction. Purification of the 66Ga from the copper target is described in detail. Monoclonal antimyosin antibodies are labeled with 66Ga at a high yield (99%) by transcomplexation from an acetate buffer with the diethylenetriamine pentaacetic acid (DTPA)-labeled antibody and preliminary evaluated in two dogs for imaging of myocardial necrosis by positron emission tomography.

Animals↗
PubMed1989

Comparative analysis of the genomes of intestinal spirochetes of human and animal origin.

The aim of the present work was to compare the genomes of 21 strains of intestinal spirochetes, which were isolated from patients suffering intestinal disorders, with those of Treponema hyodysenteriae (strain P18), the known etiological agent of swine dysentery (bloody scours), and of a nonpathogenic strain (M1) of Treponema innocens. The percent guanine-plus-cytosine value of the 23 DNAs was found to be 25.5 to 30.1, as determined by a double-labeling procedure based on nick-translation by DNA polymerase I. The genome size of two spirochetal strains, of human and porcine origin, was found to be similar (4 x 10(6) base pairs) and close to that of the reference bacterium Escherichia coli (4.2 x 10(6) base pairs). Restriction analysis showed the presence of two modified bases in spirochetal DNA. Methyladenine was present in the GATC sequence of DNA from 15 spirochetes of human origin, and methylcytosine was present in several sequences occurring in all strains. The DNA of T. hyodysenteriae displayed a 30 to 100% homology with respect to that of 21 spirochetes from humans, thus suggesting the occurrence of a genetic heterogeneity in the latter group. These data indicate that the intestinal spirochetes analyzed in the present work are related; hence there is a possibility of domestic animals being reservoirs of microorganisms pathogenic for humans. A classification of intestinal treponemes into subgroups has been proposed on the basis of restriction analysis and hybridization experiments.

Animals↗
PubMed1988

DNA methylation in leprosy-associated bacteria: Mycobacterium leprae and Corynebacterium tuberculostearicum.

The DNAs of two kinds of microorganisms from human leprosy lesion, Mycobacterium leprae and Corynebacterium tuberculostearicum (also known as "leprosy-derived corynebacterium" or LDC), have been analysed and compared with the genomes of reference bacteria of the CMN group (genera Corynebacterium, Mycobacterium and Nocardia). The guanine-plus-cytosine content (% GC) of DNA was determined by a double-labelling procedure, which is unaffected by the presence of modified and unusual bases (that alter both buoyant density and mid-melting-point determinations). Accordingly, the DNAs of seven LDC strains had GC values of 54-56 mol %, and that of armadillo-grown M. leprae a value of 54.8 +/- 0.9 mol %. Restriction patterns disclosed no methylated cytosine in the DNA sequences CCGG, GGCC, AGCT and GATC of either LDC or M. leprae DNA. N6-methyl adenine was present in the sequence GATC of all LDC strains, but was missing from the genomes of all others CMN organisms analysed, including M. leprae. By HPLC analysis of LDC-DNA hydrolysates, it was found that N6-methyladenine amounted to 1.8% of total DNA adenine, and was present exclusively within GATC sequences, which appeared all to be methylated. It is concluded that LDC represent a group of corynebacteria endowed with high genetic homogeneity and a unique restriction pattern, whereby their genome is easily distinguished from that of M. leprae, which has a similar base composition.

Adenine↗
PubMed1988

Cyclotron production of carrier-free 66Ga as a positron emitting label of albumin colloids for clinical use.

A method for producing carrier free 66Ga (T1/2:9.4 h; beta +) by 4He bombardment of natural copper targets is presented. 66Ga is formed by means of the 63Cu (4He, n) 66Ga reaction. Production yields are given in the 17.5 to 8 MeV 4He energy range. Chemical purification of 66Ga from the copper target is described. The only radionuclidic impurity found in the final product was 67Ga. Albumin colloids from commercially available kits designed for use with 99mTc could easily be labeled with 66Ga and employed for studies of the lymphatic system by positron emission tomography.

Animals↗
PubMed1988

Size and homology of the genomes of leprosy-derived corynebacteria, Mycobacterium leprae, and other corynebacteria and mycobacteria.

The genomes of Mycobacterium leprae and leprosy-derived corynebacteria (LDC), which have a similar base composition of guanine + cytosine 56 mol %, have been compared with those of reference bacteria of the CMN group (genera Corynebacterium, Mycobacterium, Nocardia). Genome sizes of three LDC strains were (1.2-2.5) x 10(6) base pairs. DNA from four of seven LDC strains examined had homology levels greater than 60%. Two other strains had a homology of 40% when compared with the CMN strains and one strain was distinctly different. The DNA from all seven LDC strains gave 0.3-18% hybridisation with that of M. leprae, 5-16% with reference corynebacteria, 5-12% with M. bovis, and 2-8% with Nocardia caviae. The small size of the LDC genome and its unrelatedness to those of M. leprae and organisms of the CMN group shows the uniqueness of LDC.

Corynebacterium↗
PubMed1985

A microanalytical procedure for determination of the base composition of DNA.

A new procedure for the determination of the percentage guanine plus cytosine (% G+C; mol/100 mol) values of microquantities of DNA is described. Its principle is a DNA-polymerase-I-directed nick translation of DNA in the presence of dGTP, dTTP, [3H]dCTP, and [alpha-32P]dATP. Kinetics experiments indicate that the plateau value is reached in about 20 min of incubation under our experimental conditions. Percentage G+C is obtained from the linear relation 1/(% G+C) = 0.01 K [32P]/[3H] + 0.01, where the ratio of trichloroacetic-acid-precipitable radioactivity is taken into account, the K value being determined for each experiment by using a few reference DNAs of known composition. This procedure has proven suitable for analysis of plasmidic, viral and cellular DNAs of different base composition (25-75% G+C), shape (linear and circular double-stranded DNA) and size (100-150 000 base pairs). Usual methods for % G+C analysis (buoyant density and melting temperature determinations) yield unreliable results in the presence of either modified or unusual bases: the double-labeling procedure is still valid under these conditions. The latter is, therefore, the method of choice for analysis or rare DNA species which are available in very small quantities (it requires amounts of DNA as low as 1 ng, i.e. several order of magnitude lower than those used for chromatographic analysis of DNA hydrolysates). Since the obtention of highly purified DNA is an essential prerequisite for the double-labeling procedure, a method for purification of bacterial DNA is detailed in the present work.

Base Composition↗