Genomic and proteomic profiles of Acholeplasma laidlawii strains differing in sensitivity to ciprofloxacin.

As a result of comparative analysis of complete genomes as well as cell and vesicular proteomes of A. laidlawii strains differing in sensitivity to ciprofloxacin, it was first shown that the mycoplasma resistance to the antibiotic is associated with the reorganization of genomic and proteomic profiles, which concerns many genes and proteins involved in fundamental cellular processes and realization of bacterial virulence.

Prevention of antibiotic-associated metabolic syndrome in mice by intestinal alkaline phosphatase.

AIMS: To examine whether co-administration of intestinal alkaline phosphatase (IAP) with antibiotics early in life may have a preventive role against metabolic syndrome (MetS) in mice. METHODS: A total of 50 mice were allocated to four treatment groups after weaning. Mice were treated with azithromycin (AZT) ± IAP, or with no AZT ± IAP, for three intermittent 7-day cycles. After the last treatment course, the mice were administered a regular chow diet for 5 weeks and subsequently a high-fat diet for 5 weeks. Body weight, food intake, water intake, serum lipids, glucose levels and liver lipids were compared. 16S rRNA gene pyrosequencing was used to determine the differences in microbiome composition. RESULTS: Exposure to AZT early in life rendered mice susceptible to MetS in adulthood. Co-administration of IAP with AZT completely prevented this susceptibility by decreasing total body weight, serum lipids, glucose levels and liver lipids to the levels of control mice. These effects of IAP probably occur as a result of changes in the composition of specific bacterial taxa at the genus and species levels (e.g. members of Anaeroplasma and Parabacteroides). CONCLUSIONS: Co-administration of IAP with AZT early in life prevents mice from susceptibility to the later development of MetS. This effect is associated with alterations in the composition of the gut microbiota. IAP may represent a novel treatment against MetS in humans.

X-ray studies on the interaction of the antimicrobial peptide gramicidin S with microbial lipid extracts: evidence for cubic phase formation.

We have investigated the effect of the interaction of the antimicrobial peptide gramicidin S (GS) on the thermotropic phase behavior of model lipid bilayer membranes generated from the total membrane lipids of Acholeplasma laidlawii B and Escherichia coli. The A. laidlawii B membrane lipids consist primarily of neutral glycolipids and anionic phospholipids, while the E. coli inner membrane lipids consist exclusively of zwitterionic and anionic phospholipids. We show that the addition of GS at a lipid-to-peptide molar ratio of 25 strongly promotes the formation of bicontinuous inverted cubic phases in both of these lipid model membranes, predominantly of space group Pn3m. In addition, the presence of GS causes a thinning of the liquid-crystalline bilayer and a reduction in the lattice spacing of the inverted cubic phase which can form in the GS-free membrane lipid extracts at sufficiently high temperatures. This latter finding implies that GS potentiates the formation of an inverted cubic phase by increasing the negative curvature stress in the host lipid bilayer. This effect may be an important aspect of the permeabilization and eventual disruption of the lipid bilayer phase of biological membranes, which appears to be the mechanism by which GS kills bacterial cells and lysis erythrocytes.

Nisin resistance distinguishes Mycoplasma spp. from Acholeplasma spp. and provides a basis for selective growth media.

The sensitivity of 11 Mycoplasma and 5 Acholeplasma species to the bacteriocin nisin was determined. When applied on filter paper discs to lawns of acholeplasma cells, nisin (20 nmol per disc) gave 3.5- to 7.0-mm zones of growth inhibition. The inclusion of 0.2 mM nisin in agar medium reduced the number of Acholeplasma laidlawii colonies by a factor of more than 10(6), and in a salts solution, 75 microM nisin killed more than 99.9% of cells within 1 min. Under similar conditions, nisin had no significant effect upon the growth or survival of Mycoplasma species. At low concentrations (1 to 3 microM), nisin stimulated glucose oxidation by A. laidlawii and Acholeplasma oculi. However, in comparison with carbonyl cyanide m-chlorophenylhydrazone (CCCP), a recognized protonophore and uncoupler of respiration, the maximum extent of stimulation was low, < or = 20%, compared with up to 180% for CCCP. Also, in contrast to results obtained with CCCP, at concentrations only slightly above those causing stimulation of acholeplasma oxygen uptake, nisin strongly inhibited respiration. Inhibition of oxygen uptake was greater for A. laidlawii cells grown in the absence of cholesterol, and on agar medium, growth inhibition by nisin decreased with increasing concentrations of cholesterol. Nisin resistance may be a valuable characteristic in the selection and identification of Mycoplasma spp.

Cyclodextrins as carriers of cholesterol and fatty acids in cultivation of mycoplasmas.

The design of fully or partly defined media for mycoplasma cultivation involves the need to provide the essential lipids, cholesterol and long-chain fatty acids, in an assimilable and nontoxic form. This study introduces cyclodextrins (CDs) as carriers of these lipids, thus suggesting alternatives to serum or bovine serum albumin (BSA). The effects of beta-CD and two forms of chemically modified beta-CD, dimethyl-beta-CD (Dimeb) and hydroxypropyl-beta-CD (Hyprob), on the growth of Mycoplasma capricolum and Acholeplasma laidlawii were investigated in a basal medium as well as in serum- and BSA-supplemented media. beta-CD was found to inhibit the growth of the sterol-requiring M. capricolum in both serum and BSA media, but it stimulated the growth of the sterol-independent A. laidlawii. Inhibition by beta-CD was explained by its capacity to form a water-insoluble CD-cholesterol complex, thus rendering it unavailable to the cells. Dimeb, despite its strong complexing ability for lipids, was found to be toxic to all mycoplasma species in both liquid cultures and agar diffusion susceptibility tests. In sharp contrast to beta-CD and Dimeb, Hyprob (with a degree of substitution of 4.2) added at 5 and 10 mM to a basal medium supplemented with lipids permitted growth of M. capricolum. Comparison of growth curves in the two conventional serum and BSA media with those in two Hyprob media revealed comparable growth and growth rates.

Anti-mycoplasmal activity of a new macrolide: miocamycin.

The object of this study was the evaluation of the activity of miocamycin and other macrolides (erythromycin and josamycin) against 61 Ureaplasma urealyticum, 1 Acholeplasma laidlawii and 21 aerobe mycoplasmas (M. pneumoniae, M. hominis, M. gallisepticum, M. mycoides) and anaerobe mycoplasmas (M. morale and M. salivarium) both clinically isolated and standard reference strains. The minimum inhibitory concentrations (MIC) values for miocamycin ranged between 0.00625 mg/l and 0.4 mg/l (ureaplasmas) and between 0.001 mg/l and 0.0625 mg/l (mycoplasmas, except M. hominis with 0.025 mg/l to 0.25 mg/l). For erythromycin, the MIC values ranged between 0.19 mg/l up to 500 mg/l (ureaplasmas) and between 0.001 mg/l to 0.0625 mg/l (mycoplasmas, except M. hominis with 100 mg/l to 1000 mg/l). MIC values for josamycin ranged between 0.03125 mg/l and 0.5 mg/l (M. pneumoniae only). The sub-MIC treatment (carried out on human pathogenic mycoplasmas only) evidenced growth curve modifications and a decrease of the O2 uptake directly correlated to the drug concentrations.

In vitro testing of the anti-mycoplasma effect of some anti-coccidial drugs.

The anti-mycoplasma effects of the ionophores (lasalocid sodium, monensin and nigericin) were compared with that of tylosin tartrate and tiamulin in vitro. Forty-four strains representing 14 avian and 10 mammalian Mycoplasma species and serotypes and 5 Acholeplasma species were tested. The ionophores showed average minimal inhibitory concentration (MIC) values between 3.65 and 4.93 micrograms ml-1 for all strains, the MIC values for glucose-fermenting strains were between 2.26 and 3.75 micrograms ml-1, significantly lower than for arginine-hydrolysing strains (9.27-13.12 micrograms ml-1). These values were significantly higher than those obtained with tylosin tartrate (0.45 micrograms ml-1) or tiamulin (0.13 micrograms ml-1). The ionophores were more efficacious against acholeplasmas (0.06-0.25 micrograms ml-1) than against mycoplasmas.

Ultrastructural changes in mollicutes induced by the peptide antibiotic herbicolin A.

Electron microscopy of negatively stained mycoplasma, ureaplasma, and acholeplasma cells showed ultrastructural changes after 10 min of treatment of the organisms with the peptide antibiotic herbicolin A in concentrations ranging from 10 micrograms/ml for Mycoplasma capricolum to 600 micrograms/ml for Ureaplasma urealyticum. The morphological changes were shown to be reversible at low concentrations of the antibiotic but irreversible at high concentrations.

The growth-inhibitory effects of some dyes on different Mycoplasma and Acholeplasma spp.

A microtitre-plate method for evaluating the inhibitory effect of dyes on the growth of mycoplasmas in fluid medium is described. Different species were shown to differ in their sensitivity to dyes. Statistical analysis (a) compared the general sensitivity and resistance of different mycoplasma species to the dyes and (b) showed that the dyes fell into two main groups in their effects on the mycoplasma species.

Activity of herbicolin A against Mycoplasma, Acholeplasma, Ureaplasma, and Spiroplasma species.

Herbicolin A, a novel acyl peptide antibiotic, inhibits the growth of the sterol-requiring Mycoplasma, Ureaplasma, and Spiroplasma species, with MICs varying from 1.5 to 100 micrograms/ml. With the exception of Acholeplasma modicum, all of the non-sterol-requiring species of the family Acholeplasmataceae were totally resistant to herbicolin A when tested on serum-containing medium but were inhibited to some extent on medium devoid of serum and any other source of sterol or fatty acids.

Inhibition of the growth of Ureaplasma urealyticum by a new urease inhibitor, flurofamide.

Flurofamide (N-[diaminophosphinyl]-4-fluorobenzamide), a urease inhibitor, was a potent inhibitor of the growth of Ureaplasma urealyticum. As little as 10 microM flurofamide (2 micrograms/ml) prevented any growth, but U. urealyticum survived for about eight hours before colony counts become undetectable. Flurofamide was a specific inhibitor of U. urealyticum since it did not inhibit growth of four Mycoplasma species or Acholeplasma hippikon. Flurofamide was 1,000 times more active than acetohydroxamic acid and thus has promise as a chemotherapeutic agent and a biochemical tool.

Selective medium for the isolation of Mycoplasma bovis from nasal discharges of pneumonic calves.

Strains of bovine Mycoplasmatales exhibited two growth forms on a plate medium containing 0.1 per cent Tween 80. Colonies of Mycoplasma bovis could easily be differentiated from those of concomitant mycoplasmatales in that the former grew rapidly with film and spots, while colonies of M arginini and Acholeplasma species did not produce film and spots; the growth of M bovirhinis was completely inhibited.

[Mycoplasmas isolated from the genital tract of mares (author's transl)]. / Isolierung von Mykoplasmen aus dem Genitaltrakt von Stuten.

Mycoplasmas were isolated from 11 (=#6,8%) of 161 cervix swabs from infertile mares. A total of 17 strains was isolated and characterized by indirect immunofluorescent test and metabolic inhibition test as Mycoplasma equigenitalium (11 strains), Mycoplasma subdolum (2 strains), Acholeplasma laidlawii (3 strains) and Acholeplasma equifetale (1 strain). In addition cervix swabs of the mares were investigated for bacteria. There was no specific correlation between presence of mycoplasmas and bacteria (table 1). In clinical investigations 5 of the 11 mares which harboured mycoplasmas showed a pneumovagina (table 1). The isolated mycoplasmas were tested for senstivity to antibiotics (table 2). All of the isolates were sensitive to Chloramphenicol (10 microgram), Tetracyclin (10 microgram), Tylosin (30 microgram) and Gentamycin (10 microgram) and resistant to Penicillin (6 IU) and Polmyxin B (300 IU). Against Erythromycin (15 microgram) all of the Acholeplasma-isolates were sensitive and all of the Mycoplasma-isolates resistant. Against Streptomycin (10 microgram) the two M. subdolum strains showed resistance, whereas M. equigenitalium and the Acholeplasma-isolates were sensitive. It is not yet possible to elucidate the significance of acholeplasmas and mycoplasmas in the genital organs of mares.

Sensitivity of mycoplasmas of the respiratory tract of pigs and horses to erythromycin and its use in selective media.

The ability of erythromycin in liquid medium to suppress the growth of eight species of acholeplasma and of 13 species of mycoplasma was tested. The Acholeplasma spp and two glycolytic Mycoplasma spp from horses--a slow glucose-metabolising (SGM) mycoplasma and a strain N3, related to M mycoides--were sensitive to erythromycin. Thus the growth of acholeplasmas can be suppressed when attempts are made to isolate pathogens from the porcine respiratory tract, but, in the case of horses, erythromycin would suppress not only Acholeplasma spp but also two Mycoplasma spp of unknown pathogenicity in the equine respiratory tract.

Cholesterol requirement of mycoplasmas as determined by microtiter test using polyene antibiotics.

A microtiter metabolic inhibition test was used to determine the effect of filipin and lucensomycin on the growth of representative species of Mycoplasma and Acholeplasma. The cholesterol-requiring species tested were found to be very susceptible to the two antibiotics, whereas the cholesterol nonrequiring species were not. The utilization of this method for differentiation between the Mycoplasma and Acholeplasma species is suggested and its advantages are discussed.