Hypertonic stress induced changes of Pseudomonas fluorescens adhesion towards soil minerals studied by AFM

Studying bacterial adhesion to mineral surfaces is crucial for understanding soil properties. Recent research suggests that minimal coverage of sand particles with cell fragments significantly reduces soil wettability. Using atomic force microscopy (AFM), we investigated the influence of hypertonic stress on Pseudomonas fluorescens adhesion to four different minerals in water. These findings were compared with theoretical XDLVO predictions. To make adhesion force measurements comparable for irregularly shaped particles, we normalized adhesion forces by the respective cell-mineral contact area. Our study revealed an inverse relationship between wettability and the surface-organic carbon content of the minerals. This relationship was evident in the increased adhesion of cells to minerals with decreasing wettability. This phenomenon was attributed to hydrophobic interactions, which appeared to be predominant in all cell–mineral interaction scenarios alongside with hydrogen bonding. Moreover, while montmorillonite and goethite exhibited stronger adhesion to stressed cells, presumably due to enhanced hydrophobic interactions, kaolinite showed an unexpected trend of weaker adhesion to stressed cells. Surprisingly, the adhesion of quartz remained independent of cell stress level. Discrepancies between measured cell–mineral interactions and those calculated by XDLVO, assuming an idealized sphere-plane geometry, helped us interpret the chemical heterogeneity arising from differently exposed edges and planes of minerals. Our results suggest that bacteria may have a significant impact on soil wettability under changing moisture condition

Experimental Results and Integrated Modeling of Bacterial Growth on an Insoluble Hydrophobic Substrate (Phenanthrene)

Metabolism of a low-solubility substrate is limited by dissolution and availability and can hardly be determined. We developed a numerical model for simultaneously calculating dissolution kinetics of such substrates and their metabolism and microbial growth (Monod kinetics with decay) and tested it with three aerobic phenanthrene (PHE) degraders: Novosphingobium pentaromativorans US6-1, Sphingomonas sp. EPA505, and Sphingobium yanoikuyae B1. PHE was present as microcrystals, providing non-limiting conditions for growth. Total PHE and protein concentration were tracked over 6-12 days. The model was fitted to the test results for the rates of dissolution, metabolism, and growth. The strains showed similar efficiency, with v(max) values of 12-18 g dw g(-1) d(-1), yields of 0.21 g g(-1), maximum growth rates of 2.5-3.8 d(-1), and decay rates of 0.04-0.05 d(-1). Sensitivity analysis with the model shows that (i) retention in crystals or NAPLs or by sequestration competes with biodegradation, (ii) bacterial growth conditions (dissolution flux and resulting chemical activity of substrate) are more relevant for the final state of the system than the initial biomass, and (iii) the desorption flux regulates the turnover in the presence of solid-state, sequestered (aged), or NAPL substrate sources

Lignin analyses of cores KaL #20048-1 and MUC #20007-1, Baltic Sea/Gotland Basin

The relative contribution and the composition of terrestrial organic matter to sediments of the Gotland Basin/Central Baltic Sea were assessed by the analysis of phenolic lignin oxidation products and the delta13C of organic matter. Samples were taken from box core 20048-1 and Multicore 20007-1 taken at the same location. Methods: Core retrieval and sample processing: A 10.30m long giant kasten core (KaL#20048-1) was recovered (57°23.14'N, 020°15.51'E; water depth 241 m) by r/v Poseidon (1995) and a multicore (MUC#20007-1) at the same position by r/v Alexander von Humboldt in 1994. The kasten core was opened and logged on board, and the entire kasten core was archived in plastic boxes. The multicore was sliced into 1 cm slices, which were frozen on board and freeze-dried in the home laboratory in Warnemuende. For lignin analyses, 1-cm sub-samples were taken by sawn-off plastic syringes from archived material of box core 20048-1 in intervals of about 10 cm and air dried at 40°C. Subsamples of sediments from the multicore 20007-1 were taken from dried slices. Lignin analysis: Between 500 to 2000 mg of dried and homogenised sediment were oxidized at 170°C for 2 h in the presence of 2 mol/L NaOH, CuO, and (NH)4Fe(SO4)2. After centrifugation, the supernatants were acidified to pH 2 with 6 mol/L HCl. The humic acids, which precipitated, were removed by centrifugation. The supernatant was further purified by solid phase extraction. The lignin-derived phenols were sorbed from the acidic solution on C18 material and later eluted with ethyl acetate. The solvent was removed by rotary evaporation, and the phenolic oxidation products were transferred to autosampler vials with methanol that was then removed under a flux of N2. Before analysis by GC/MS, the samples were dissolved in acetonitrile and derivatized with N,O-bis-(trimethylsilyl)trifluoroacetamid (BSTFA) for 1 h at room temperature. Thereafter, they were diluted with acetonitrile according to the expected phenol concentrations. One microliter of each sample was injected in splitless mode, and the phenols were separated in a HP 6890 gas chromatograph equipped with a HP5MS column (30 m x 250 micrometer x 0.25 micrometer). The temperature program of the gas chromatograph was 100°C isothermal for 4 min, ramp to 220°C at 4°C min⁻¹ with a 5-min isothermal period at 120°C, isothermal at 220°C for 3 min, ramp to 300°C at 30°C min⁻¹, and final isothermal period for 10 min. The transfer line to the mass spectrometer was kept at 325°C throughout the analysis. The HP 5973 mass spectrometer was operated in the EI mode at 70 eV. The ion source temperature was 230°C, and the quadrupole was kept at 150°C. Compounds were quantified by integration of the base ions and by comparison of the peak areas with those of synthetic standards. Before oxidation, ethylvanillin was added as an internal standard for the determination of recovery. To rule out possible transformations of the internal standard during the oxidation step, blanks containing only ethylvanillin and the reagents were also processed. GC-FID analysis of these blanks displayed a single peak with the retention time of ethylvanillin, and there was no evidence of any transformation of ethylvanillin during the oxidation step under the experimental conditions. The internal standard was added at the beginning of the analysis to ensure that the internal standard and the lignin oxidation products have the same history during the entire analysis. On average, 75% of the added ethylvanillin was recovered after the complete analytical procedure; the range of recoveries was from 50% to 105%

Lignin concentrations and delta13C of organic matter in surface sediments from the Baltic Sea

The relative contribution and the composition of terrestrial organic matter were assessed by the analysis of phenolic lignin oxidation products and by the stable isotope composition of organic carbon in surface sediments of the Baltic Sea. For analyses, sub samples of lyophilized, ground and homogenized surface sediment (0-1 cm) material from the collection of surface sediments of the Institut fuer Ostseeforschung Warnemuende were used. Methods Lignin analysis: Between 500 to 2000 mg of dried and homogenised sediment were oxidized at 170°C for 2 h in the presence of 2 mol/L NaOH, CuO, and (NH)4Fe(SO4)2. After centrifugation, the supernatants were acidified to pH 2 with 6 mol/L HCl. The humic acids, which precipitated, were removed by centrifugation. The supernatant was further purified by solid phase extraction. The lignin-derived phenols were sorbed from the acidic solution on C18 material and later eluted with ethyl acetate. The solvent was removed by rotary evaporation, and the phenolic oxidation products were transferred to autosampler vials with methanol that was then removed under a flux of N2. Before analysis by GC/MS, the samples were dissolved in acetonitrile and derivatized with N,O-bis-(trimethylsilyl)trifluoroacetamid (BSTFA) for 1 h at room temperature. Thereafter, they were diluted with acetonitrile according to the expected phenol concentrations. One microliter of each sample was injected in splitless mode, and the phenols were separated in a HP 6890 gas chromatograph equipped with a HP5MS column (30 m x 250 micrometer x 0.25 micrometer). The temperature program of the gas chromatograph was 100°C isothermal for 4 min, ramp to 220°C at 4°C min^-1 with a 5-min isothermal period at 120°C, isothermal at 220°C for 3 min, ramp to 300°C at 30°C min^-1, and final isothermal period for 10 min. The transfer line to the mass spectrometer was kept at 325°C throughout the analysis. The HP 5973 mass spectrometer was operated in the EI mode at 70 eV. The ion source temperature was 230°C, and the quadrupole was kept at 150°C. Compounds were quantified by integration of the base ions and by comparison of the peak areas with those of synthetic standards. Before oxidation, ethylvanillin was added as an internal standard for the determination of recovery. To rule out possible transformations of the internal standard during the oxidation step, blanks containing only ethylvanillin and the reagents were also processed. GC-FID analysis of these blanks displayed a single peak with the retention time of ethylvanillin, and there was no evidence of any transformation of ethylvanillin during the oxidation step under the experimental conditions. The internal standard was added at the beginning of the analysis to ensure that the internal standard and the lignin oxidation products have the same history during the entire analysis. On average, 75% of the added ethylvanillin was recovered after the complete analytical procedure; the range of recoveries was from 50% to 105%. Concentrations and delta13C of total organic carbon Approximately 20 mg of the homogenized sample were weighed into tared sample vessels for elemental composition (total carbon, total nitrogen, organic carbon) and for isotope analyses (delta13C of organic carbon). Total carbon was determined in a Carlo Erba/Fisons 1108 Elemental Analyzer after combustion. A second weighed sample split in tared silver foil vessels was treated with 2N HCl to remove inorganic carbon. On this sub-sample, the concentrations of TOC and isotope ratio delta13C of organic carbon (given in permil versus V-PDB) were determined simultaneously in a Carlo Erba/Fisons 1108 Elemental Analyzer connected to an isotope-ratio mass spectrometer (Finnigan Delta S). The reference gas was pure CO2 from a cylinder calibrated against carbonate (NBS- 18, 19, 20). The standard deviation for replicate analyses of delta13C was less than 0.2 permil. The original data were corrected for the addition of anthropogenic CO2 (Suess effect) by substracting – 1.48 permil from the measured delta13C values of total organic carbon