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Biochemical and Biophysical Research... Apr 1997Mycoplasma fermentans has unique glycoglycerophospholipids (GGPLs: GGPL-I and GGPL-III). Previously, the structure of these lipids was determined as... (Comparative Study)
Comparative Study
Mycoplasma fermentans has unique glycoglycerophospholipids (GGPLs: GGPL-I and GGPL-III). Previously, the structure of these lipids was determined as phosphocholine-6'-alpha-glucopyranosyl-(1'-3)-1, 2-diacyl-glycerol (GGPL-I) and 1"-phosphocholine-2"-aminodihydroxypropane-3"-phospho-6'-alph++ + a- glucopyranosyl-(1'-3)-1, 2-diacyl-glycerol (GGPL-III). Thin-layer chromatography (TLC) immunostaining showed that the GGPLs were main lipid-antigens of the M. fermentans species. Anti-M. fermentans serum stained mainly the GGPLs, but the other anti-mycoplasma sera (anti-M. arginini, anti-M. hyorhinis, anti-M. pneumonia, anti-M. primatum, and anti-Acholeplasma laidlawii, anti-M. hominis, anti-M. orale, and M. salivarium) stained neither GGPL-I nor GGPL-III. The TLC analysis of glycolipids and phospholipids of various human related mycoplasmas showed clearly that GGPLs are specifically expressed in M. fermentans species. GGPL-I and GGPL-III ranged from 1.6 to 28% and from an undetectable level to 35% of total phospholipids, respectively. Although there was heterogeneity among the amounts of GGPL-I or GGPL-III of M. fermentans strains, all of the M. fermentans strains had GGPL-I and/or GGPL-III. These observations showed that GGPL structures are species-specific immunodeterminants of M. fermentans. The fact that the GGPLs are main phospholipid components of the M. fermentans species means the M. fermentans has a unique choline metabolic pathway. This observation may raise phylogenetic interest.
Topics: Animals; Antibodies, Bacterial; Antigens, Bacterial; Glycolipids; Humans; Molecular Structure; Mycoplasma; Mycoplasma fermentans; Phosphorylcholine; Rabbits; Species Specificity
PubMed: 9168906
DOI: 10.1006/bbrc.1997.6443 -
British Journal of Rheumatology Mar 1997The objective was to investigate the presence of mycoplasmas in rheumatoid arthritis (RA) and other chronic arthritides. Samples of synovial fluid (SF) were... (Comparative Study)
Comparative Study
The objective was to investigate the presence of mycoplasmas in rheumatoid arthritis (RA) and other chronic arthritides. Samples of synovial fluid (SF) were systematically collected from all patients presenting with an articular effusion. Each sample was divided into three parts. The first was kept for cytological count and culture on standard media for pyogens and mycobacteria, the second was cultivated on specific media for mycoplasmas and the third frozen for subsequent study by polymerase chain reaction (PCR). A total of 209 samples were studied. Half of the patients had inflammatory rheumatic diseases: RA (27), spondyloarthropathy (28), connective tissue disease (5), unclassified arthritis (45). The remaining suffered from other conditions, including osteoarthritis (60), gouty arthritis (19), haemarthrosis (5), post-traumatic effusion (2). Eight samples were positive by culture, two for Mycoplasma hominis; three for M. fermentans, one for M. salivarium, one for M. orale and one for Ureaplasma urealyticum. All the patients concerned had an inflammatory rheumatic disease: five had RA, one had psoriatic arthritis and two had unclassified arthritis. These results were confirmed by PCR in two cases (one M. fermentans, one U. urealyticum). The lack of sensitivity of the conventional PCR assay on SF is discussed. Mycoplasmas were mainly detected in SF of RA patients. These results raise the question of the possible role of mycoplasmas in the triggering and maintenance of inflammatory rheumatic diseases, especially RA.
Topics: Adult; Aged; Culture Media; Culture Techniques; Female; Humans; Male; Middle Aged; Mycoplasma; Polymerase Chain Reaction; Rheumatic Diseases; Synovial Fluid
PubMed: 9133961
DOI: 10.1093/rheumatology/36.3.310 -
Applied and Environmental Microbiology Jan 1997Surfactin, a cyclic lipopeptide antibiotic and biosurfactant produced by Bacillus subtilis, is well-known for its interactions with artificial and biomembrane systems...
Surfactin, a cyclic lipopeptide antibiotic and biosurfactant produced by Bacillus subtilis, is well-known for its interactions with artificial and biomembrane systems (e.g., bacterial protoplasts or enveloped viruses). To assess the applicability of this antiviral and antibacterial drug, we determined the cytotoxicity of surfactin with a 50% cytotoxic concentration of 30 to 64 microM for a variety of human and animal cell lines in vitro. Concomitantly, we observed an improvement in proliferation rates and changes in the morphology of mycoplasma-contaminated mammalian cells after treatment with this drug. A single treatment over one passage led to complete removal of viable Mycoplasma hyorhinis cells from various adherent cell lines, and Mycoplasma orale was removed from nonadherent human T-lymphoid cell lines by double treatment. This effect was monitored by a DNA fluorescence test, an enzyme-linked immunosorbent assay, and two different PCR methods. Disintegration of the mycoplasma membranes as observed by electron microscopy indicated the mode of action of surfactin. Disintegration is obviously due to a physicochemical interaction of the membrane-active surfactant with the outer part of the lipid membrane bilayer, which causes permeability changes and at higher concentrations leads finally to disintegration of the mycoplasma membrane system by a detergent effect. The low cytotoxicity of surfactin for mammalian cells permits specific inactivation of mycoplasmas without significant deleterious effects on cell metabolism and the proliferation rate in cell culture. These results were used to develop a fast and simple method for complete and permanent inactivation of mycoplasmas in mammalian monolayer and suspension cell cultures.
Topics: Animals; Anti-Bacterial Agents; Bacillus subtilis; Bacterial Proteins; Cell Division; Cell Line; Drug Evaluation, Preclinical; Humans; Lipopeptides; Microscopy, Electron; Mycoplasma; Peptides, Cyclic
PubMed: 8979337
DOI: 10.1128/aem.63.1.44-49.1997 -
In Vitro Cellular & Developmental... Jun 1996The relations between K+ channel and Cl- channel currents and mycoplasma infection status were studied longitudinally in HSG cells, a human submandibular gland cell...
The relations between K+ channel and Cl- channel currents and mycoplasma infection status were studied longitudinally in HSG cells, a human submandibular gland cell line. The K+ channel currents were disrupted by the occurrence of mycoplasma infection: muscarinic activation of K+ channels and K+ channel expression as estimated by ionomycin- or hypotonically induced K+ current responses were all decreased. Similar decreases in ionomycin- and hypotonically induced responses were observed for Cl- channels, but only the latter decrease was statistically significant. Also, Cl- currents could be elicited more frequently than K+ currents (63% of cases versus 0%) in infected cells when tested by exposure to hypotonic media, indicating that mycoplasma infection affects K+ channels relatively more than Cl- channels. These changes occurred in the originally infected cells, were ameliorated when the infection was cleared with sparfloxacin, and recurred when the cells were reinfected. Such changes would be expected to result in hyposecretion of salivary fluid if they occurred in vivo.
Topics: Antitubercular Agents; Calcium; Cell Line; Chloride Channels; Cytosol; Electric Conductivity; Fluoroquinolones; Humans; Hypotonic Solutions; Ionomycin; Mycoplasma Infections; Potassium Channels; Quinolones; Submandibular Gland
PubMed: 8842750
DOI: 10.1007/BF02722962 -
[Induction of tumor necrosis factor alpha by Mycoplasma penetrans isolated from patients with AIDS].Kansenshogaku Zasshi. the Journal of... Jan 1996The activity of induce tumor necrosis factor alpha (TNF alpha) production of several mycoplasmas, including AIDS associated mycoplasmas was investigated. M. penetrans...
The activity of induce tumor necrosis factor alpha (TNF alpha) production of several mycoplasmas, including AIDS associated mycoplasmas was investigated. M. penetrans which was detected and isolated from urine and tissue of Kaposi's sarcoma of patients with AIDS markedly exhibited the induction of TNF alpha production of both THP-1 cells and murine peritoneal macrophages to compare to other mycoplasmas. Each amount of M. penetrans, M. fermentans, M. incognitus, Acholeplasma ladilawii, M. orale, M. salivarium, M. hominis required for induction of 50% cytotoxicity to L cells in the supernatants of mouse peritoneal macrophages cultured with those microorganisms was 0.65 micrograms/ml, 11.3 micrograms/ml, 19.6 micrograms/ml, 6.6 micrograms/ml, 7.7 micrograms/ml, 6.3 micrograms/ml, 5.7 micrograms/ml respectively. Next, the components of M. penetrans were extracted by Bligh-Dyer method, in order to investigate chemical component to induce TNF alpha-production. The activity of TNF alpha induction was mainly found in the methanol-phase, but not in the chloroform-phase, where lipid and glycolipid of the microorganisms were generally thought to be accumulated. The binding of the active component to concanavalin A-Sepharose was blocked in the presence of Methyl alpha-D-mannopyranoside and Methyl alpha-D-glucopyranoside. These results suggest that the component possess mannoside and glucoside active site.
Topics: AIDS-Related Opportunistic Infections; Humans; Mycoplasma Infections; Mycoplasma penetrans; Tumor Necrosis Factor-alpha
PubMed: 8822049
DOI: 10.11150/kansenshogakuzasshi1970.70.11 -
FEMS Microbiology Letters May 1995Homologous internal controls were used as competitor DNA in the polymerase chain reaction for the quantitative detection of mycoplasma DNA. PCR primer sets were designed...
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.
Topics: Base Sequence; DNA Primers; DNA, Bacterial; Molecular Sequence Data; Mycoplasma; Polymerase Chain Reaction; RNA, Ribosomal, 16S; Reference Standards
PubMed: 7750739
DOI: 10.1111/j.1574-6968.1995.tb07524.x -
In Vitro Cellular & Developmental... May 1994Mycoplasmal contamination remains a significant impediment to the culture of eukaryotic cells. For certain cultures, attempts to eliminate the infection are feasible...
Mycoplasmal contamination remains a significant impediment to the culture of eukaryotic cells. For certain cultures, attempts to eliminate the infection are feasible alternatives to the normally recommended disposal of the contaminated culture. Here, three antibiotic regimens for mycoplasmal decontamination were compared in a large panel of naturally infected cultures: a 1-wk treatment with the fluoroquinolone mycoplasma removal agent (MRA), a 2-wk treatment with the fluoroquinolone ciprofloxacin, and three rounds of a sequential 1-wk treatment with BM-Cyclin containing tiamulin and minocyclin. These antibiotic treatments had a high efficiency of permanent cure: MRA 69%, ciprofloxacin 75%, BM-Cyclin 87%. Resistance to mycoplasma eradication was observed in some cell cultures: BM-Cyclin 0%, MRA 20%, ciprofloxacin 20%. Nearly all resistant contaminants that could be identified belonged to the species Mycoplasma arginini and M. orale. Detrimental effects of the antibiotics were seen in the form of culture death caused by cytotoxicity (in 5 to 13% of the cultures). Alterations of the cellular phenotypic features or selective clonal outgrowth might represent further untoward side effects of exposure to these antibiotics. Overall, antibiotic decontamination of mycoplasmas is an efficient, inexpensive, reliable, and simple method: 150/200 (75%) chronically and heavily contaminated cultures were cured and 50/200 (25%) cultures could not be cleansed and were either lost or remained infected. It is concluded that eukaryotic cell cultures containing mycoplasmas are amenable to antibiotic treatment and that a cure rate of three-quarters is a reasonable expectation.
Topics: Cells, Cultured; Ciprofloxacin; Diterpenes; Drug Resistance, Microbial; Minocycline; Mycoplasma; Quinolones
PubMed: 8069460
DOI: 10.1007/BF02631456 -
Journal of Clinical Microbiology May 1994The growth of Mycoplasma salivarium ATCC 23064 and Mycoplasma orale ATCC 15539 was inhibited by MnCl2. The growth-inhibitory effect was much more remarkable on M. orale... (Comparative Study)
Comparative Study
The growth of Mycoplasma salivarium ATCC 23064 and Mycoplasma orale ATCC 15539 was inhibited by MnCl2. The growth-inhibitory effect was much more remarkable on M. orale than M. salivarium and was much more remarkable in medium supplemented with 10% (vol/vol) horse serum (HS) than 20% (vol/vol) HS. It was suggested that isolates of Mycoplasma from the oral cavity could be roughly identified as either M. salivarium and M. orale by examination of the growth (color changes) in PPLO broth supplemented with 10% (vol/vol) HS and 0.2 mM MnCl2.
Topics: Aminopeptidases; Bacteriological Techniques; Chlorides; Culture Media; Humans; Manganese Compounds; Mouth; Mycoplasma; Mycoplasma Infections; Species Specificity
PubMed: 8051265
DOI: 10.1128/jcm.32.5.1343-1345.1994 -
Applied and Environmental Microbiology Mar 1994A fast and simple method to detect mycoplasmal contamination in simulated samples of animal sera by using a PCR was developed. The following five mycoplasma species that...
A fast and simple method to detect mycoplasmal contamination in simulated samples of animal sera by using a PCR was developed. The following five mycoplasma species that are major cell culture contaminants belonging to the class Mollicutes were investigated: Mycoplasma arginini, Acholeplasma laidlawii, Mycoplasma hyorhinis, Mycoplasma orale, and Mycoplasma fermentans. After a concentration step involving seeded sera, genus-specific primers were used to amplify a 717-bp DNA fragment within the 16S rRNA gene of mycoplasmas. In a second step, the universal PCR was followed by amplification of variable regions of the 16S rRNA gene by using species-specific primers, which allowed identification of contaminant mycoplasmas. With this method, 10 fg of purified DNA and 1 to 10 color-changing units of mycoplasmas could be detected. Since the sensitivity of the assay was increased 10-fold when the amplification products were hybridized with an internal mycoplasma-specific 32P-labelled oligonucleotide probe, a detection limit of 1 to 10 genome copies per PCR sample was obtained. This highly sensitive, specific, and simple assay may be a useful alternative to methods currently used to detect mycoplasmas in animal sera.
Topics: Animals; Base Sequence; Blood Specimen Collection; Cattle; Horses; Molecular Sequence Data; Polymerase Chain Reaction; Sensitivity and Specificity; Species Specificity; Tenericutes; Time Factors
PubMed: 8161186
DOI: 10.1128/aem.60.3.953-959.1994 -
Journal of Virological Methods Jan 1994The efficacy of several antibiotic treatments to eliminate mycoplasma from Vero cells contaminated chronically with Mycoplasma orale II were tested. Minocyclin,...
The efficacy of several antibiotic treatments to eliminate mycoplasma from Vero cells contaminated chronically with Mycoplasma orale II were tested. Minocyclin, Kanamycin, Tylosine and Roxitromycin, at non cytotoxic concentrations, were assayed alone or in different combinations. Mycoplasma contamination was effectively eradicated without recurrence once the following regimen was applied: Incubation of contaminated cells with Tylosine (250 micrograms/ml) for 12 days followed by incubation with Minocycline (5 micrograms/ml) for 10 days. This treatment was not deleterious for cell growth, it was effective after only one application and it was successful to eradicate mycoplasma from other contaminated eukaryotic continuous cell lines.
Topics: Animals; Cells, Cultured; Chlorocebus aethiops; Cricetinae; Culture Techniques; Drug Resistance, Microbial; Drug Therapy, Combination; Eukaryotic Cells; Humans; Kanamycin; Minocycline; Mycoplasma; Roxithromycin; Tumor Cells, Cultured; Tylosin
PubMed: 8175949
DOI: 10.1016/0166-0934(94)90018-3