A specific antimicrobial protein CAP-1 from Pseudomonas sp. isolated from the jellyfish Cyanea capillata.

A bacterium strain, designated as CMF-2, was isolated from the jellyfish Cyanea capillata and its culture supernatant exhibited a significant antimicrobial activity. The strain CMF-2 was identified as Pseudomonas sp. based on the morphological, biochemical and physiological characteristics as well as 16S rRNA sequence analysis. In this study, an antimicrobial protein, named as CAP-1, was isolated from the culture of CMF-2 through ammonium sulfate precipitation and gel filtration chromatography. According to the result of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), a major band indicated that the antimicrobial protein had a molecular mass of about 15 kDa, and it was identified as a hypothetical protein by MALDI-TOF-MS analysis and Mascot searching. CAP-1 displayed a broad antimicrobial spectrum against the indicator bacteria and fungus, including Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Candida albicans, especially some marine-derived microorganisms such as Vibrio vulnificus, Vibrio alginolyticus, Vibrio parahaemolyticus, Vibrio cholera, and Vibrio anguillarum, but showed little impact on tumor cells and normal human cells. The protein CAP-1 remained a stable antimicrobial activity in a wide range of temperature (20-80°C) and pH (2-10) conditions. These results suggested that CAP-1 might have a specific antimicrobial function not due to cytotoxicity.

The coffee-machine bacteriome: biodiversity and colonisation of the wasted coffee tray leach.

Microbial communities are ubiquitous in both natural and artificial environments. However, microbial diversity is usually reduced under strong selection pressures, such as those present in habitats rich in recalcitrant or toxic compounds displaying antimicrobial properties. Caffeine is a natural alkaloid present in coffee, tea and soft drinks with well-known antibacterial properties. Here we present the first systematic analysis of coffee machine-associated bacteria. We sampled the coffee waste reservoir of ten different Nespresso machines and conducted a dynamic monitoring of the colonization process in a new machine. Our results reveal the existence of a varied bacterial community in all the machines sampled, and a rapid colonisation process of the coffee leach. The community developed from a pioneering pool of enterobacteria and other opportunistic taxa to a mature but still highly variable microbiome rich in coffee-adapted bacteria. The bacterial communities described here, for the first time, are potential drivers of biotechnologically relevant processes including decaffeination and bioremediation.

Characterization of a metal resistant Pseudomonas sp. isolated from uranium mine for its potential in heavy metal (Ni2+, Co2+, Cu2+, and Cd2+) sequestration.

Heavy metal sequestration by a multimetal resistant Pseudomonas strain isolated from a uranium mine was characterized for its potential application in metal bioremediation. 16S rRNA gene analysis revealed phylogenetic relatedness of this isolate to Pseudomonas fluorescens. Metal uptake by this bacterium was monophasic, fast saturating, concentration and pH dependent with maximum loading of 1048 nmol Ni(2+) followed by 845 nmol Co(2+), 828 nmol Cu(2+) and 700 nmol Cd(2+)mg(-1) dry wt. Preferential metal deposition in cell envelope was confirmed by TEM and cell fractionation. FTIR spectroscopy and EDX analysis revealed a major role of carboxyl and phosphoryl groups along with a possible ion exchange mechanism in cation binding. Binary system demonstrated selective metal binding affinity in the order of Cu(2+)>Ni(2+)>Co(2+)>Cd(2+). A comparison with similar metal uptake reports considering live bacteria strongly indicated the superiority of this strain in metal sequestration, which could be useful for developing efficient metal removal system.

Uranium and thorium sequestration by a Pseudomonas sp.: mechanism and chemical characterization.

The mechanism and chemical nature of uranium and thorium sequestration by a Pseudomonas strain was investigated by transmission electron microscopy, energy dispersive X-ray (EDX) analysis, FTIR spectroscopy and X-ray diffractometry. Atomic force microscopy (AFM) used in the tapping mode elucidated the morphological changes in bacterial cells following uranium and thorium binding. Transmission electron microscopy revealed intracellular sequestration of uranium and thorium throughout the cell cytoplasm with electron dense microprecipitations of accumulated metals. Energy dispersive X-ray analysis confirmed the cellular deposition of uranium and thorium. EDX and elemental analysis of sorption solution indicated the binding of uranium and thorium by the bacterial biomass via displacement of cellular potassium and calcium. The strong involvement of cellular phosphate, carboxyl and amide groups in radionuclide binding was ascertained by FTIR spectroscopy. X-ray powder diffraction (XRD) analyses confirmed cellular sequestration of crystalline uranium and thorium phosphates. Overall results indicate that a combined ion-exchange-complexation-microprecipitation mechanism could be involved in uranium and thorium sequestration by this bacterium. Atomic force microscopy and topography analysis revealed an undamaged cell surface with an increase in cell length, width and height following radionuclide accumulation. The arithmetic average roughness (R(a)) and root mean square (RMS) roughness (R(q)) values indicated an increase in surface roughness following uranium and thorium sequestration.

A novel mcl-PHA produced on coprah oil by Pseudomonas guezennei biovar. tikehau, isolated from a "kopara" mat of French Polynesia.

Pseudomonas guezennei biovar. tikehau was isolated from a microbial mat on the atoll of Tikehau in French Polynesia, and is able to synthesize medium chain length poly-beta-hydroxyalkanaote copolymers when grown on coprah oil. A two-step cultivation process was used and the biosynthesis of PHAs was followed along 52h by regular culture sampling. The polyester was purified from freeze-dried cells and analysed by nuclear magnetic resonance (NMR), Fourier transform infra red (FTIR), and gas chromatography mass spectrometries. The copolyester produced by P. guezennei biovar. tikehau from coprah oil mainly consisted of saturated monomers, i.e. 3-hydroxyoctanoate (3HO) and 3-hydroxydecanoate (3HD), and the monomeric composition of the polyester did not change during the fermentation process. However, yield of PHAs production varied from 4% of the cellular dry weight (CDW) to 63% obtained after 36h. Scan electron microscopy was used to study the morphology and organization of PHAs granules within the cells and revealed the presence of several granules occupying almost the entire cell volume.

Weakening effect of cell permeabilizers on gram-negative bacteria causing biodeterioration.

Gram-negative bacteria play an important role in the formation and stabilization of biofilm structures on stone surfaces. Therefore, the control of growth of gram-negative bacteria offers a way to diminish biodeterioration of stone materials. The effect of potential permeabilizers on the outer membrane (OM) properties of gram-negative bacteria was investigated and further characterized. In addition, efficacy of the agents in enhancing the activity of a biocide (benzalkonium chloride) was assessed. EDTA, polyethylenimine (PEI), and succimer (meso-2,3-dimercaptosuccinic) were shown to be efficient permeabilizers of the members of Pseudomonas and Stenotrophomonas genera, as indicated by an increase in the uptake of a hydrophobic probe (1-N-phenylnaphthylamine) and sensitization to hydrophobic antibiotics. Visualization of Pseudomonas cells treated with EDTA or PEI by atomic force microscopy revealed damage in the outer membrane structure. PEI especially increased the surface area and bulges of the cells. Topographic images of EDTA-treated cells were compatible with events assigned for the effect of EDTA on outer membranes, i.e., release of lipopolysaccharide and disintegration of OM structure. In addition, the effect of EDTA treatment was visualized in phase-contrast images as large areas with varying hydrophilicity on cell surfaces. In liquid culture tests, EDTA and PEI supplementation enhanced the activity of benzalkonium chloride toward the target strains. Use of permeabilizers in biocide formulations would enable the use of decreased concentrations of the active biocide ingredient, thereby providing environmentally friendlier products.

The membrane-bound respiratory chain of Pseudomonas pseudoalcaligenes KF707 cells grown in the presence or absence of potassium tellurite.

The respiratory chain of Pseudomonas pseudoalcaligenes KF707 in membranes isolated from cells grown in the presence or absence of the toxic oxyanion tellurite (TeO3(2-)) was examined. Aerobic growth in the absence of tellurite shows an NADH-dependent respiration which is 80% catalysed by the cytochrome (cyt) bc1-containing pathway leading to two terminal membrane-bound cyt c oxidases inhibited by different concentrations of KCN (IC50 0.2 and 1 microM). A third oxidase, catalysing the remaining 20% of the cyanide-resistant respiration and fully inhibited by 2-3 mM KCN, is also present; this latter pathway accounts for 60-70% of the total NADH-dependent respiration in membranes from cells grown in LB medium supplemented with potassium tellurite (35 microg x ml(-1)). Two high-potential b-type haems (E(m,7) +395 and 318 mV) are redox centres of a membrane-bound cyt c oxidase (possibly of the cbb3 type) which shows a 50% decrease of its activity in parallel with a similar decrease of the c-type haem content (mostly soluble cyt c) in membranes from tellurite-grown cells; the latter type of cells specifically contain a cyt b type at +203 mV (pH 9.0) which is likely to be involved in cyanide-resistant respiration. Comparison of the growth curve of KF707 cells in parallel with tellurite uptake showed that intracellular accumulation of tellurium (Te(0)) crystallites starts from the mid-exponential growth phase, whereas tellurite-induced changes of the respiratory chain are already evident during the early stages of growth. These data were interpreted as showing that reduction of tellurite to tellurium and tellurite-dependent modifications of the respiratory chain are unrelated processes in P. pseudoalcaligenes KF707.

A battery of toxicity tests as indicators of decontamination in composting oily waste.

Heterogeneous oily waste from an old dumping site was composted in three windrows constructed from different proportions of waste, sewage sludge, and bark. The objectives of this pilot study were to examine the usefulness of composting as a treatment method for this particular waste and to study decontamination in the composting process by using a battery of toxicity tests. Five samples from the windrow having intermediate oil concentrations were tested with toxicity tests based on microbes (Pseudomonas putida growth inhibition test, ToxiChromotest, MetPLATE, and three different modifications of a luminescent bacterial test), enzyme inhibition (reverse electron transport), plants (duckweed growth inhibition and red clover seed germination), and soil animals (Folsomia candida, Enchytraeus albidus, and Enchytraeus sp.). The luminescent bacterial tests were used as prescreening tests. Chemical analyses of samples were carried out simultaneously. Both toxicity and oil concentration, including those of polyaromatic hydrocarbons (PAHs), were reduced during composting and soil quality improved significantly. The total oil hydrocarbon concentration decreased from 90,000 to 19,000 mg/kg, measured with the IR method, in 4 months, and from 86,000 to 1400 mg/kg, measured with GC method. The concentration of PAHs decreased from 135 to 23.5 mg/kg. During the fourth month of composting (stabilization stage), the proportion of the heaviest oil fractions (asphaltenes) became dominant. Toxicity varied between different samples and between different bioassays; however, the first sample was significantly more toxic than the others, and most of the tests revealed a decrease in toxicity during the composting process.

Flagella of a plant-growth-stimulating Pseudomonas fluorescens strain are required for colonization of potato roots.

The role of motility in the colonization of potato roots by Pseudomonas bacteria was studied. Four Tn5-induced flagella-less mutants of the plant-growth-stimulating P. fluorescens WCS374 appeared to be impaired in their ability to colonize growing potato roots.