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C Cocito

Publications and source records attributed to C Cocito.

At least 37 records · Page 2Linked to original sources
PubMed1994

Analysis of tuberculous meningitis cases by an immunoblotting assay based on a mycobacterial antigen complex.

Tuberculous meningitis cases were analyzed by an immunoblotting test based on Mycobacterium bovis BCG antigen complex A60. Anti-A60 immunoglobulin G (IgG) in cerebrospinal fluid (CSF) allowed early diagnosis, and concentrations decreased after recovery. In primary meningitis forms, anti-A60 IgGs were intrathecally synthesized and specific oligoclonal IgGs were present in CSF. In meningeal complications of pulmonary tuberculosis, there were matching titers of anti-A60 IgG in blood and CSF (mirror pattern). Correlation between CSF-restricted patterns and CSF pleocytosis was shown.

Adolescent↗
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↗
PubMed1993

Isolation and characterization of a novel glucuronosyl diacylglycerol from Mycobacterium smegmatis.

A glucuronic acid-containing diacylglycerol was isolated from exponentially growing Mycobacterium smegmatis. Structural analysis of the purified glycolipid, performed by gas chromatography-mass spectrometry, fast atom bombardment-mass spectrometry, and high resolution proton NMR, indicated the structure 3-(O-alpha-D-glucuronopyranosyl)-1,2-diacyl-sn-glycerol. Two forms of the glycolipid were observed differing in fatty acid composition. Both molecular species contained a hexadecanoic acid residue, whereas the second acyl group was either tuberculostearic acid (10-methylstearic acid) or octadecenoic acid. The inherent antigenicity of the glycolipid was shown by its ability to bind to anti-Mycobacterium avium (serovar 26) and anti-Mycobacterium tuberculosis sera by Western blot-type thin-layer chromatography. This is the second instance of the isolation of a glycosyl diacylglycerol from members of the Mycobacterium genus, further confirming its close relationship to Gram-positive bacteria.

Antigens, Bacterial↗
PubMed1993

Comparative analysis of three sensitins used in cutaneous testing for tuberculosis and paratuberculosis in cattle.

Correspondence between components of sensitins (avian, bovine, and johnin-PPD tuberculins) and the proteins of the corresponding microorganisms (Mycobacterium avium, Mycobacterium bovis and Mycobacterium paratuberculosis) was established. Most of these identified proteins are present in the three organisms, but the 38.3- and 48.8-kDa proteins of M. avium and the 54.0- and 58.7-kDa proteins of M. bovis are endowed with species-specific epitope(s).

Animals↗
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

Mycobacterial antigen complex A60-specific T-cell repertoire during the course of pulmonary tuberculosis.

The Mycobacterium bovis antigen complex A60 is known to be immunodominant in tuberculosis and to have a protective effect against experimental infection in vitro and in vivo. To identify immunodominant and possibly protective antigens in pulmonary tuberculosis, the T-cell repertoire directed to nitrocellulose-bound fractions of A60 antigen was analyzed in active tuberculosis patients during the course of the infection and after recovery. The results show that patients infected with Mycobacterium tuberculosis acquired complete A60-T-cell reactivity only in the late phases of infection. At disease onset, patients with active tuberculosis were characterized by (i) T-cell unresponsiveness to most A60 fractions, (ii) high tumor necrosis factor alpha production, and (iii) low gamma interferon (IFN-gamma) release. Several weeks after chemotherapy, the unresponsive state disappeared and the following reverse situation was observed: (i) high blastogenic response to almost all A60 fractions, (ii) low tumor necrosis factor alpha release, and (iii) high IFN-gamma production. In addition, 60% of these patients significantly responded against seven A60 fractions (61 to 58, 56 to 53, 49 to 46, 46 to 44, 35 to 33, 33 to 30, and 30 to 28 kDa), indicating that they included immunodominant antigens. Furthermore, only the fractions within the molecular mass ranges of 56 to 44 and 35 to 28 kDa induced IFN-gamma synthesis. One year after complete recovery from infection, more than 60% of past-active tuberculosis subjects had memory T cells specific for the immunodominant fractions of 61 to 58, 56 to 53, 49 to 46, and 33 to 30 kDa. Since the same fractions induced the strongest IFN-gamma production, known to exhibit antimycobacterial effects, it is suggested that these may represent the inducers of a protective immune response.

Adult↗
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↗
PubMed1993

Lipid composition of leprosy-derived corynebacteria, a distinct group of corynebacteria, and of a reference Corynebacterium.

Leprosy-derived corynebacteria (LDC) are diphtheroid organisms isolated from leprosy patients and previously characterized by DNA and cell wall analysis. Three groups of LDC components of taxonomic value, glycolipids, and phospholipids and cell-wall-bound lipids were analyzed in comparison with those of a reference strain C. hoffmannii (CH). The main CH glycolipid, "cord factor" (trehalose dimycolate), was missing from LDC. Among phospholipids, phosphatidylinositol and phosphatidylglycerol had lowered proportions in LDC, as compared to CH, whereas phosphatidylethanolamine and cardiolipin were absent from both microorganisms. Bound lipids in acidic extracts of delipidated LDC yielded arabinose corynomycolate in lesser quantity with respect to CH. Alkaline hydrolysis of whole cells released fatty acids and mycolic acids, which were analyzed by gas chromatography/mass spectrometry. Reference CH, grown in the absence of serum, yielded C16:0 and C18:1 (major) and C18:0 (minor) fatty acids, as well as C32, C34, and C36 corynomycolic acids. All these components, particularly mycolates, had lowered proportions when this organism was grown in the presence of serum. Dominant LDC components were, in addition to C16:0, C18:0, and CI8:u fatty acids, cholesterol from serum. Very low concentrations of corynomycolic acids with a high degree of unsaturation were found in these organisms, suggesting a dependence of lipid metabolism on growth conditions. The presence in LDC of tuberculostearic acid (C19r:0), a mycobacterial component found in some pathogenic corynebacteria, was carefully explored: Traces of C19r:0 were found in LDC 19 grown in the presence of delipidated serum, but not in LDC 15 nor in C. hoffmannii. Present data, in conjunction with previous studies on DNA and mycolic acids, disclose basic differences in the composition of LDC and conventional corynebacteria.

Cell Wall↗
PubMed1992

The role of rRNA bases in the interaction of peptidyltransferase inhibitors with bacterial ribosomes.

Synergism of streptogramins A (virginiamycin M, VM) and B (virginiamycin S, VS), peptidyltransferase inhibitors, was explored in EM4/pLC7-21 (wild type) and EM4/pERY (VS-resistant). These bacterial strains contained multicopy plasmids carrying an rrnH operon with wild type (pLC7-21) or mutated (A2058----U transversion) 23 S rRNA gene. Ribosomes with wild type and mutated rRNA were both present in EM4/pERY. The latter particles did not bind VS; in the presence of VM, however, high affinity VS binding occurred. As shown previously, VS protected against chemical reagents certain bases in domain V rRNA and VM in the stems flanking this loop. Differences between wild type and mutant ribosomes were observed: A2058, A2059, A2062, and G2505, protected by VS and ERY in EM4/pLC7-21, were unshielded in EM4/pERY. A2062 was shielded by VM in EM4/pERY, not in EM4/pLC7-21, and G2505 of mutant ribosomes became protected by VS when VM was simultaneously present. Induction by VM of a high affinity VS binding site in VS-sensitive and -resistant ribosomes indicates A2058 mutation to entail a conformational change of this site, which is counteracted by VM fixation. Accessibility of A2062 to chemical reagents (unlike behavior of EM4/pERY and EM4/pLC7-21 in the presence of VM) implies different conformations for wild type and mutant ribosomes.

Base Sequence↗
PubMed1992

Identification of a single base change in ribosomal RNA leading to erythromycin resistance.

The molecular basis of a mutation conferring an erythromycin-resistance phenotype was explored, as an approach to the role of 23 S rRNA in the peptidyl-transferase activity of 50 S ribosomal subunits. Mutagenization of an Escherichia coli strain, which carried the multicopy plasmid pLC7-21 containing the rrnH operon, led to the production of an erythromycin-resistant strain. Plasmid pBFL1 isolated from this mutant was able to transform the sensitive RecA- strain EM4 and to induce a "dissociated" type of antibiotic resistance. Two ribosome populations occurred in EM4/pBFL1: normal particles coded for by the seven rrn chromosomal genes and mutated particles containing rRNA of plasmid origin. The latter particles displayed in vitro lower affinity and susceptibility to erythromycin than wild type particles. The mutation within plasmid pBFL1 was mapped by a multiple primer extension technique. Three synthetic primers were used to sequence the central loop in domain V of 23 S rRNA, leading to identification of a C to U transition at position 2611. This base change was proved to be responsible for the erythromycin-resistance phenotype by the plasmid-plasmid marker rescue technique. A molecular explanation for the rrn mutations leading, respectively, to undissociated and to dissociated types of resistance to the MLSb (macrolide-lincosamide-synergimycin B) group of antibiotics is proposed. These results and some literature data support the notion that rRNA bases involved in antibiotic resistance play a conformational role in the ribosomal binding sites for the MLSb antibiotics.

Base Sequence↗
PubMed1992

Composition and immunogenicity of the polysaccharide components of the thermostable macromolecular antigen group of mycobacterial antigens.

The thermostable macromolecular antigen (TMA) group includes major components of the mycobacterial cell envelope and cytoplasm, which elicit humoral and cellular immune reactions, and seems to play important roles in infectious diseases. The best known member of this group, antigen A60 of Mycobacterium bovis BCG, was previously shown to contain three moieties of polysaccharides, free lipids, and polypeptides. In this work, the TMA polysaccharides of three pathogenic mycobacteria (M. avium, M. bovis and M. paratuberculosis) have been analyzed by coupled gas chromatography-mass spectrometry. In all cases the cores of the TMA complexes were represented by branched glucans of high molecular mass (about 10(6) daltons), for which structural models have been proposed. The immunogenicity of the polysaccharide components from the three TMA was verified with several immunological procedures (immunodiffusion and immunoelectrophoresis of the antigen, and immunoblotting of the corresponding electrofocused immunoglobulins). All tests tallied in showing a negligible immunogenicity of the glucans examined (inability to produce, upon injection, the synthesis of specific immunoglobulins), thus pointing to the protein moiety of TMA as the one responsible for the high immunoreactivity of the complexes.

Animals↗
PubMed1992

Immunization of mice with the antigen A60 of Mycobacterium bovis BCG.

Antigen 60 (A60) is a thermostable component of the cytoplasm of Mycobacterium tuberculosis and BCG which can be fractionated into at least 15 protein bands when analysed by Western blot. Normal B6D2 mice were immunized subcutaneously with 20 micrograms of the A60 protein suspended in Freund's incomplete adjuvant (FIA) or in saline. Three weeks later the mice received a second dose of vaccine followed 2 weeks later by an aerogenic challenge with approximately 10(3) CFU of M. tuberculosis Erdman. The mice receiving the adjuvanted A60 showed a significant reduction (P less than 0.05) in the number of viable organisms recovered from the lungs and the spleen 3 weeks after challenge. However, this response was less than that seen in BCG vaccinated controls.

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↗