Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0026918 (Mycobacterium)
 52,428 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A polymerase chain reaction (PCR) assay for the rapid detection of Mycobacterium tuberculosis in sputum samples is described. The target DNA is a 123-base pair (bp) segment of IS6110, which is repeated in the M. tuberculosis chromosome and is specific for the M. tuberculosis complex. Methodology used to lyse the mycobacteria, extract the DNA, and amplify the 123-bp target DNA is presented. The amplified PCR product is detected by examination of ethidium-bromide-stained acrylamide gels. An internal control using the same primers as the target DNA has been constructed to assess the efficacy of each individual reaction. Of 162 sputum samples tested, 82 were smear-positive for acid-fast bacilli. Of the 94 specimens from patients in whom pulmonary tuberculosis was diagnosed, 51 were culture-positive, smear-positive, or both. Fifty of these were PCR positive. Of the 42 specimens from patients with nontuberculous mycobacterial pulmonary disease, 41 were PCR negative. All 26 specimens from patients without mycobacterial infection were PCR negative. This assay provides a sensitive and specific means for the laboratory diagnosis of tuberculosis within 48 h that is relatively simple to perform.
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PMID:Detection of Mycobacterium tuberculosis in sputum samples using a polymerase chain reaction. 195 48

Analysis of the 1,016-base-pair sequence of a putative probe for identification of Mycobacterium tuberculosis revealed two almost identical fragments of 507 and 509 bases. From this sequence two pairs of primers were synthesized (MtbAB and MtbCD), ranging from 18 to 22 nucleotides, for use in polymerase chain reactions (PCRs) with DNA from six reference strains of M. tuberculosis, as well as type strains of M. bovis, M. bovis BCG, M. kansasii, M. avium, M. intracellulare, and M. scrofulaceum. Although there was amplification of DNA from all mycobacterial strains included in the study, when used as probes, a predominant band, fragment CD from M. tuberculosis H37Rv DNA, proved to be more specific for strains of M. tuberculosis than the original probe, pMTb4, was. Amplified fragments from as little as 1 fg of DNA (equivalent to one-fifth of an organism) could be resolved on ethidium bromide-stained gels loaded with a 1/10 volume of PCR. Furthermore, it was possible to amplify specific DNA sequences from frozen M. tuberculosis H37Rv organisms which were thawed prior to PCR.
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PMID:Sequence analysis and amplification by polymerase chain reaction of a cloned DNA fragment for identification of Mycobacterium tuberculosis. 210 94

Evidence of the presence of plasmids in Mycobacterium tuberculosis is lacking, whereas they are widespread in some other mycobacterial species. We examined, by agarose gel electrophoresis, a total of 197 clinical isolates of M. tuberculosis, mostly resistant to one or more antibiotics, and were able to detect bands of apparently extrachromosomal DNA at a low level in some isolates. These presumptive plasmids could not be isolated by CsCl/ethidium bromide gradient ultracentrifugation, and may consist of unusual forms of DNA. The possible existence of single stranded plasmid DNA is discussed.
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PMID:Does Mycobacterium tuberculosis have plasmids? 211 17

Bacteriological diagnosis of tuberculosis in childhood is often unsuccessful owing to the difficulty in obtaining suitable specimens. Many attempts have been made to diagnose tuberculosis immunologically but with very limited success. Positive tuberculin reactions may be the result of nonspecific sensitization while negative reactions occur in undernourished children. Serodiagnostic tests suffer from problems of specificity, even when very specific antigens are used, and are often least helpful in diagnostically difficult cases. Detection of antigen has proved to be of more value, especially with clean specimens such as cerebrospinal and pleural fluids. Detection of specific components of Mycobacterium tuberculosis by linked gas chromatography and mass spectroscopy is very sensitive and specific but the equipment is very costly. Detection of specific DNA sequences of M. tuberculosis in specimens by use of labelled 'DNA probes' is rather insensitive although the sensitivity may be increased greatly by use of the polymerase chain reaction to amplify small amounts of the specific DNA. Non specific indicators of tuberculosis are generally unhelpful although the bromide partition test and assay of the enzyme adenosine deaminase in cerebrospinal fluid appear to be of value in the diagnosis of tuberculous meningitis. More research is required to develop a simple, specific and automated test for tuberculosis in childhood.
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PMID:New approaches to the diagnosis of tuberculosis in childhood with special reference to neurotuberculosis. 212 31

The synthesis of the tetrasaccharides O-(2,3-di-O-methyl-alpha-L-fucopyranosyl)-(1----4)-O-(2,3- di-O-methyl-alpha-L-fucopyranosyl)-(1----3)-O-alpha-L-rhamnopyranosyl -(1----2)-6-deoxyl-L-talose (36) and O-(2-O-methyl-alpha-L-fucopyranosyl)-(1----4)-O-(2-O-methyl-alpha-L- fucopyranosyl)-(1----3)-O-alpha-L-rhamnopyranosyl-(1----2)-6-deoxy-L- talose (41) is described. The former and the latter structures, respectively, have been proposed as the carbohydrate chains in the polar glycopeptidolipid antigens of serovars 9 and 25 in the Mycobacterium avium-M. intracellulare-M. scrofulaceum serocomplex. Glycosylation of allyl 2,3-di-O-methyl-alpha-L-fucopyranoside with 4-O-benzoyl-2,3-di-O-methyl-alpha-L-fucopyranosyl chloride gave the alpha-linked disaccharide derivative, which was O-deallylated and converted into the corresponding disaccharide alpha-chloride. This was coupled with benzyl 3,4-di-O-benzyl-6-deoxy-2-O-(2,4-di-O-benzoyl-alpha-L-rhamnopyranosyl)- alpha-L-talopyranoside (32) to give a fully protected tetrasaccharide derivative, which was deprotected to furnish 36. Likewise, 3-O-benzyl-4-O-(3,4-di-O-acetyl-2-O-methyl-alpha-L- fucopyranosyl)-2-O-methyl-alpha-L-fucopyranosyl chloride, prepared by way of condensation of allyl 3-O-benzoyl-2-O-methyl-alpha-L-fucopyranoside with 2-O-methyl-3,4-di-O-(p-nitrobenzoyl)-alpha-L-fucopyranosyl bromide, reacted with 32, to provide, after removal of blocking groups, 41.
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PMID:Synthesis of tetrasaccharides related to the antigenic determinants from the glycopeptidolipid antigens of serovars 9 and 25 in the Mycobacterium avium-M. intracellulare-M. scrofulaceum serocomplex. 242 95

Earlier studies from our laboratory reported that a radiometric Mycobacterium leprae resident macrophage assay was a useful in vitro indicator of bacillary viability with good correlation with the established mouse foot pad model. The present study compares our assay with the recently described fluorescein diacetate/ethidium bromide (FDA/EB) method. M. leprae extracted from the dermal lesions of 73 bacilliferous leprosy patients were tested concurrently by both techniques. Good correlation (r = 0.52, p less than 0.001) was found between the radiometric assay evaluating DNA synthesis and the FDA/EB staining reflecting the presence of active esterase enzyme. In addition, the utility of the FDA/EB staining in the monitoring of therapy was established. Twenty-two patients treated for greater than 1 year showed lower numbers of green fluorescing bacilli when compared to 19 untreated or short-term-treated individuals.
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PMID:Comparison of radiometric macrophage assay and fluorescein diacetate/ethidium bromide staining for evaluation of M. leprae viability. 243 21

Viable bacterial populations were estimated in bacilli purified from 105 biopsies from 40 untreated and 65 multibacillary leprosy patients treated with multidrug therapy (MDT) for varying periods. The bacilli were purified and viability was determined by ATP content, morphological index (MI), and fluorescein diacetate-ethidium bromide (FDA-EB) staining. Viable populations were calculated, taking 3.58 x 10(-15) g/solid bacillus as the mean ATP content of a viable unit of Mycobacterium leprae. The proportion of viable bacilli was also estimated in the same specimens using solid-staining (MI) and green-staining bacilli by the FDA-EB method. In the untreated cases, the positive viability by ATP assay was 100%, 92% by MI, and 100% by FDA-EB. ATP content per solid bacillus was relatively constant, which was not the case with ATP content per green-staining bacillus. While the MI was zero in all cases, viable bacilli could still be detected by ATP estimations in 5 of the 32 (16%) patients after 2 years of MDT and in 1 of the 20 (5%) patients after 3 years of MDT. No viable bacilli could be detected even by this method beyond 3 years of MDT. On the other hand, green-staining bacilli were demonstrable in 7/32 (22%) of cases after 2 years of MDT, 2/20 (10%) after 3 years of MDT, and 1/13 (8%) after more than 3 years of treatment, indicating that the FDA-EB staining and ATP assay did not detect the same populations. A determination of the ATP content of M. leprae could be used as a reliable and sensitive tool for determining viability of the bacilli.
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PMID:Effect of chemotherapy on viability of Mycobacterium leprae as determined by ATP content, morphological index and FDA-EB fluorescent staining. 247 22

The susceptibilities of Mycobacterium leprae and M. avium complex (MAC) to the H2-O2-Fe-mediated halogenation system supplemented with antimicrobial agents were evaluated by fluorescein diacetate-ethidium bromide (FDA/EB) staining. In the case of M. leprae, the number of greenstained bacteria (intact cells) was reduced in the presence of the H2O2-Fe-mediated halogenation system supplemented with agents possessing antileprosy activity, such as rifampin, 4,4'-diaminodiphenylsulfone (dapsone), clofazimine, and ofloxacin. In the case of the MAC strain, although viable units of the organisms were reduced by the halogenation system alone, the number of greenstained cells in the FDA/EB stain was not reduced, even when the halogenation system was used in combination with ofloxacin. Because stainability of the cells is related to structural and functional intactness of the membrane, differences between M. leprae and the MAC strain imply possible differences in the rigidity of the cell membrane.
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PMID:Susceptibilities of Mycobacterium leprae and M. avium complex to the H2O2-Fe-mediated halogenation system supplemented with antimicrobial agents. 247 23

The action of ethidium bromide on Mycobacterium rubrum cells was studied. The culture growth was found to depend on ethidium bromide (EB) concentration in the medium. The reaction of EB with nucleoid DNA was shown to be specific and changes in the nucleoid structure were detected. Low EB doses (ca. 2 micrograms/ml) caused DNA despiralization in many cells. The process was reversible, which accounted for the elevated ability of reactivation at low EB doses. A higher EB dose (ca. 5-10 micrograms/ml and more) made the nucleoid structure coarser and denser in most cells and the nucleoid broke down to small fragments. As a result, due to the pool of enzymes present in the cells prior to EB addition, secondary changes developed. They involved all the cellular structures as well as the metabolism of lipids, polyphosphates, and glycogen. As a rule, these changes were incompatible with the cell viability.
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PMID:[Change in the structural organization of Mycobacterium rubrum cells upon the action of ethidium bromide]. 262 8

Hydroxyacid oxidase from rat kidney is an FMN-dependent enzyme that catalyzes the oxidation of L-alpha-hydroxy acids as well as, more slowly, that of L-alpha-amino acids. We report here a modified purification method for the enzyme, which is found to possess one cofactor per subunit of Mr 39,000. Determination of its N-terminal sequence suggests the protein is homologous to spinach glycolate oxidase and baker's yeast lactate dehydrogenase. In the presence of a hydroxy acid and of bromopyruvate, under anaerobic conditions, the enzyme is found to catalyze both transhydrogenation and reductive bromide ion elimination. It had previously been observed that hydroxyacid oxidase could not catalyze chloride elimination from chlorolactate in the presence of oxygen [Cromartie, T.H., & Walsh, C.T. (1975) Biochemistry 14, 3482-3490]. The behavior of this enzyme toward halogeno substrates is therefore similar to that of baker's yeast L-lactate dehydrogenase and in part different from that of Mycobacterium smegmatis lactate oxidase and porcine kidney D-amino-acid oxidase. These findings can be rationalized on the basis of a common mechanism for all these enzymes, implying formation of a carbanion as a first step, with different rate-limiting steps in the overall reaction.
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PMID:Rat kidney L-2-hydroxyacid oxidase. Structural and mechanistic comparison with flavocytochrome b2 from baker's yeast. 306 53


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