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Query: UMLS:C0026936 (Mycoplasma)
 14,761 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We investigated 22 mycoplasma and acholeplasma species for their ability to reduce tetrazolium salts by using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. The test results were evaluated visually, as well as spectrophotometrically, by using an enzyme-linked immunosorbent assay reader. Our results were very similar to the results obtained when the tetrazolium salt reduction assay described by Aluotto et al. was used. However, the MTT reduction assay appeared to be better because it is faster, more objective and sensitive, easier to evaluate, and less expensive; in addition, it allows quantitative determinations. By using regression analysis a linear correlation between formazan production and the number of colony-forming units was demonstrated for all of the species investigated, indicating that the MTT assay can also be used for growth, toxicity, or chemosensitivity tests for the mycoplasma species that are capable of reducing tetrazolium salts.
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PMID:Tetrazolium [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reduction by mycoplasmas. 150 78

A case of a 10-year old boy suffering from asthma bronchiale following pleuropneumonia is reported. Paradoxically, bronchospasmolysis tests using physiological saline + salbutamol or ipratropium bromide impaired the lung function of this patient. Salt solutions inhaled in increasing doses until 1.4% generated severe bronchoconstriction, however, inhalation of DNCG + salbutamol normalized the lung function completely.
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PMID:[Bronchial hyperreactivity to physiological saline solution]. 176 1

We have used the polymerase chain reaction to detect Mycoplasma genitalium in artificially seeded human throat swab samples as well as in clinical material. On the basis of the published nucleotide sequence of the M. genitalium 140-kDa adhesin gene, primers were chosen to produce an amplified fragment of 281 bp. Five different previously isolated strains, including the type strain of M. genitalium, could all be detected by the polymerase chain reaction, and DNAs from other mycoplasmal and bacterial species yielded negative results. The detection limits were estimated to be approximately 50 organisms by inspection of ethidium bromide-stained agarose gels and 4 organisms when a biotinylated oligonucleotide probe was used in filter hybridization. The amplified DNA fragments were subjected to restriction enzyme analysis. DNAs from the five different isolates all possessed EcoRI, SspI, AluI, Sau3AI, and DdeI restriction sites, as predicted from the published sequence. A total of 150 urogenital swabs collected from 100 patients for culturing of Chlamydia trachomatis were tested for the presence of M. genitalium DNA. Ten samples from eight patients were found positive. The amplified DNA fragments from all of our clinical samples possessed the AluI, Sau3AI, and DdeI restriction sites, but three samples from two patients did not contain the SspI site and none of the samples contained the EcoRI site.
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PMID:Polymerase chain reaction for detection of Mycoplasma genitalium in clinical samples. 199 66

A species-specific 760-base pair (bp) BamHI to EcoRI DNA fragment (fMG-2) was isolated from a Mycoplasma gallisepticum (MG) genomic library constructed in plasmid pUC8. Based on the DNA sequence data of fMG-2, a pair of 25 base primers, designated amplification (Amp) left (L) and right (R) primers, was synthesized. When used in the polymerase chain reaction (PCR), the Amp L and R primers directed amplification of DNA of 16 MG strains yielding an expected 732-bp product, but did not amplify DNA of Escherichia coli, calf thymus, lambda phage, pUC8 plasmid, or 16 other species of avian mycoplasmas. As low as 10(-6) picogram of MG DNA, a fraction of the total chromosomal content of one cell, was detected following amplification by PCR. PCR amplification products were visualized by either ethidium bromide/ultraviolet exposure or hybridization with a 481-bp probe (fMG-3) prepared from the central region of fMG-2.
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PMID:Polymerase chain reaction for detection of Mycoplasma gallisepticum. 202 63

Supercoiled double-stranded DNA molecules (plasmids) were isolated from plants infected with three laboratory strains of western aster yellows mycoplasma-like organism (AY-MLO) by using cesium chloride-ethidium bromide density gradients. Southern blot analysis, using plasmids from the severe strain of AY-MLO (SAY-MLO) as the probe, identified at least four plasmids in celery, aster, and periwinkle plants and in Macrosteles severini leafhopper vectors infected with either the dwarf AY-MLO, Tulelake AY-MLO, or SAY-MLO strain. Plasmids were also detected in two California field isolates of AY-MLO but not in plants infected with the beet leafhopper-transmitted virescence agent, western X, or elm yellows MLOs. SAY-MLO plasmids were 5.2, 4.9, 3.4, and 1.7 kilobase pairs in size. Plasmids isolated from dwarf AY- and Tulelake AY-MLOs were 7.4, 5.1, 3.5, and 1.7 kilobase pairs in size. No evidence was obtained for integration of SAY-MLO plasmids into the MLO chromosome.
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PMID:Identification and characterization of plasmids from the western aster yellows mycoplasmalike organism. 230 60

The adherence protein (P1 protein) of Mycoplasma pneumoniae was purified by electroelution and cleaved with cyanogen bromide. The resulting peptides were separated by two-dimensional electrophoresis. Spots reacting in Western immunoblots with two attachment-inhibiting monoclonal antibodies were isolated, and the amino-terminal ends of these peptides were microsequenced. The two monoclonal antibodies had different binding sites. One was associated with the amino-terminal region of the whole P1 protein beginning at amino acid position 237, and the other was associated with amino acid position 702, which was localized approximately in the middle of the P1 amino acid sequence. Serum samples from three M. pneumoniae-infected patients were tested by Western blotting against the cyanogen bromide peptide pattern. All three serum samples reacted with peptide fragments beginning at amino acid position 702, but the serum of only one patient also had antibodies against the oligopeptides beginning at amino acid position 237. These results indicate that the corresponding epitopes of the P1 protein are also immunogenic if they are presented at the surface of the infecting organism.
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PMID:Binding sites of attachment-inhibiting monoclonal antibodies and antibodies from patients on peptide fragments of the Mycoplasma pneumoniae adhesin. 246 70

Physicochemical methods have been used to compare mycoplasma DNA capable of the genetic transformation of tetracycline resistance with DNA from tetracycline-sensitive mycoplasmas and their transformants. These mycoplasmas were isolated from human patients. The DNA extracted from Mycoplasma hominis tetr resistant to 100 microgram/mL tetracycline transforms tetracycline resistance to sensitive strains of Mycoplasma salivarium tets and Mycoplasma hominis tets but not Mycoplasma fermentans tets. Bulk DNA and DNA extracted by methods which increase the yield of circular DNA moieties were analyzed by cesium chloride and cesium chloride--ethidium bromide buoyant density ultracentrifugation and by horizontal and vertical agarose gel electrophoresis. Extrachromosomal DNA was not detected, which suggests that transformation was mediated by the recombination of chromosomal genes for tetracycline resistance and not by R factors. Moreover, no significant differences were detected in the DNA from the resistant and sensitive species or from their transformants and Mycoplasma fermentans tets which could not be transformed to resistance to 10 micrograms tetracycline/mL medium.
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PMID:Attempts to detect by physicochemical methods plasmid DNA in mycoplasmas of human origin before and after transformation to tetracycline resistance. 713 11

We report the use of PCR to detect DNA from Mycoplasma hyopneumoniae, the etiological agent of enzootic porcine pneumonia. A primer set was designed for the amplification of a 649-bp fragment of the 16S rRNA gene from M hyopneumoniae. The PCR product was identified by ethidium bromide staining after gel electrophoresis and by Southern hybridization with an M. hyopneumoniae-specific oligonucleotide probe. No amplification was observed from any other porcine bacteria tested, including several closely related mycoplasmas. It was also possible to demonstrate the presence of M. hyopneumoniae in bronchial lavage samples and lung tissue samples from experimentally infected pigs. Furthermore, the PCR system was used for analysis of nasal samples obtained from pigs in a fattening herd. By this method, we were able to detect M. hyopneumoniae in nose swabs from naturally infected pigs. However, our results suggest that M. hyopneumoniae can be detected in the nasal cavities only during a limited time period.
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PMID:Detection of Mycoplasma hyopneumoniae in nose swabs from pigs by in vitro amplification of the 16S rRNA gene. 754 Jun 29

Homologous internal controls were used as competitor DNA in the polymerase chain reaction for the quantitative detection of mycoplasma DNA. PCR primer sets were designed on the basis of the most conserved nucleotide sequences of the 16S rRNA gene of mycoplasma species. Amplification of this gene was examined in five different mycoplasma species: Mycoplasma orale, M. hyorhinus, M. synoviae, M. gallisepticum and M. pneumoniae. To evaluate the primers, a number of different cell lines were assayed for the detection of mycoplasma infections. All positive cell lines showed a distinct product on agarose gels while uninfected cells showed no DNA amplification. Neither bacterial nor eukaryotic DNA produced any cross-reaction with the primers used, thus confirming their specificity. Internal control DNA to be used for quantitation was constructed by modifying the sizes of the wild-type amplified products and cloning them in plasmid vectors. These controls used the same primer binding sites as the wild-type and the amplified products were differentiated by a size difference. The detection limits for all the mycoplasma species by competitive quantitative PCR were estimated to range from 4 to 60 genome copies per assay as determined by ethidium bromide-stained agarose gels. These internal standards also serve as positive controls in PCR-based detection of mycoplasma DNA, and therefore accidental contamination of test samples with wild-type positive controls can be eliminated. The quantitative PCR method developed will be useful in monitoring the progression and significance of mycoplasma in the disease process.
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PMID:Competitor internal standards for quantitative detection of mycoplasma DNA. 775 Jul 39

The polymerase chain reaction (PCR) method was used to detect mycoplasma contamination in a panel of 42 continuous cell lines. According to the microbiological cultivation assay on agar, 29 cell lines were chronically infected and 13 cell lines were negative. Sets of outer and inner primers (nested double-step PCR) were applied which anneal to DNA sequences coding for conserved regions of the 16S rRNA. These oligonucleotides allow for the amplification of DNA regions found in at least 25 mycoplasma species (including the ones most commonly found in cell cultures), but do not cross-hybridize with DNA from eukaryotic cells. Mycoplasma-positive cell lines showed distinctive bands in ethidium bromide-stained gels, both after the first round of amplification as well as after the second PCR; all agar-negative cell lines were also unambiguously negative in the PCR assay. Thus, neither false-positive nor false-negative results occurred. Provided that the proper PCR working conditions are scrupulously observed, the PCR amplification has several outstanding advantages: high sensitivity, specificity, reliability, objectivity, speed, and simplicity.
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PMID:Mycoplasma detection by PCR analysis. 811 18


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