Electrochemical identification of microbially mediated hydrogen sulfide oxidation

The process of H₂S oxidation by the phototrophic bacteriaThiocapsa roseopersicina andChlorobium phaeobacteroides, respectively, was monitored using a Pt-glass-Ag⁰, Ag₂S electrode combination without liquid junction. Due to the resulting pe(pH) and pH₂S plottings three steps can be distinguished: oxidation of H₂S to an S(0) state, oxidation of S (0) to SO₄ ^2–, and oxidation of the remaining H₂S directly to SO₄ ^2–. Differences between the investigated bacteria exist with respect to their individual oxidation strategies.Thiocapsa apparently stops oxidizing H₂S at pH₂S 7.5 (e.g. 10^–7.5M H₂S) and shifts to the utilization of the intracellularly stored S (0). In contrastChlorobium utilizes its extracellularly stored sulfur parallel to the extracellular H₂S fraction. The corresponding Pt-sensor responses (pe₇ values) were found to be similar to the corresponding partial redox equilibria (p₇ values) of H₂S oxidation stoichiometries as proposed by Van Niel (1931) and Trüper (1964). It is concluded that the recording of pe enables investigators to understand (and control) in situ redox processes, independent of their thermodynamic equilibration, only bound to changes of electroactivity vs. sensor.     

Hyporheic biofilms — a potential food source for interstitial animals

Glass-beads (diam. = 250 μm) were buried 10 cm deep in the sediment of a stream. After an exposure of eight weeks, bacterial densities on the beads varied between 2.7 × 10⁵ and 2.4 × 10⁷/cm², and the length of the fungal mycelium between 0.2 and 5.3 mm/cm². Bacterial densities did not show any correlation with the DOC content of the water, but were positively correlated with respiration on the beads. Fungal mycelium was negatively correlated with water temperature. Acid hydrolysis of stream-exposed beads released sugars and amino acids, whose combined carbon content exceeded that of the microbial cells by a factor of at least 4. Gut extracts of Gammarus tigrinus and Tipula caloptera released amino acids and sugars from stream-exposed beads.     

Nitrogen fixation and nitrogen metabolism in heliobacteria

Three species of anoxygenic phototrophic heliobacteria, Heliobacterium chlorum, Heliobacterium gestii, and Heliobacillus mobilis, were studied for comparative nitrogen-fixing abilities and regulation of nitrogenase. Significant nitrogenase activity (acetylene reduction) was detected in all species grown photoheterotrophically on N₂, although cells of H. mobilis consistently had higher nitrogenase activity than did cells of either H. chlorum or H. gestii. Nitrogen-fixing cultures of all three species of heliobacteria were subject to switch-off of nitrogenase activity by ammonia; glutamine also served to switch-off nitrogenase activity but only in cells of H. mobilis and H. gestii. Placing photosynthetically grown heliobacterial cultures in darkness also served to switch-off nitrogenase activity. Dark-mediated switch-off was complete in lactate-grown heliobacteria but in pyruvate-grown cells substantial rates of nitrogenase activity continued in darkness. In all heliobacteria examined ammonia was assimilated primarily through the glutamine synthetase/glutamate synthase (GS/GOGAT) pathway although significant levels of alanine dehydrogenase were present in extracts of cells of H. gestii, but not in the other species. The results suggest that heliobacteria, like phototrophic purple bacteria, are active N₂-fixing bacteria and that despite their gram-positive phylogenetic roots, heliobacteria retain the capacity to control nitrogenase activity by a switch-off type of mechanism. Because of their ability to fix N₂ both photosynthetically and in darkness, it is possible that heliobacteria are significant contributors of fixed nitrogen in their paddy soil habitat.     

Purification and properties of 1-phosphofructokinase from Escherichia coli

Abstract The thermophilic facultatively phototrophic green bacterium Chloroflexus aurantiacus strain Ok-70-fl was shown to possess sulfide-repressed hydrogenase activity. Biosynthesis of the enzyme was severely repressed by S²⁻ (5.7 mM) and stimulated specifically by Ni²⁺ and by molecular hydrogen. The hydrogenase was shown to be localized in the cytoplasmic membrane and could be solubilized from the latter by the detergent Triton X-100 in a state forming one enzymatically active band ( M _r 170 × 10³) in polyacrylamide gels. In the membraneous state, the hydrogenase had its maximal activity at 73°C and was active with methyl viologen, methylene blue, menadione and flavins, but not with NAD or NADP as electron acceptors. Solubilization of the enzyme with Triton X-100 resulted in a drastic increase in the FAD/FMN-linked activity.     

Effects of external stimuli on environmental bacterial strains harboring analgD-lux bioluminescent reporter plasmid for the study of corrosive biofilms

An alginic acid biosynthesis bioluminescent reporter plasmid, pUTK50, was transconjugated into environmental strains ofPseudomonas putida, Pseudomonas fluorescens, andStenotrophomonas maltophilia. Bioluminescent transconjugates were selected from each strain for investigation of environmental stress factors that promote alginic acid exopolymer biosynthesis in developing biofilms. Environmental stimuli associated with increased levels of alginate synthesis, in a previously developed organism,P. aeruginosa FRD1, were applied to the environmental strains. Increased salt concentrations and higher ratios of nitrate vs ammonium ions as the limiting nitrogen source induced bioluminescence in FRD1 and the environmental strains. However, for environmental strains ofP. putida, P. fluorescens andS. maltophilia, polysaccharides were detected with low uronic acids content and different structural components. When tested within a biofilm,S. maltophilia O46 demonstrated exceptional adhesive and corrosive properties while alginic acid synthesis was not high. In most of the environmental strains, periods of increased bioluminescence were induced by external stimuli, but exopolysaccharides other than alginic acid were expressed. It is hypothesized that the environmental strains have homologous but nonidentical promoter sequences which are responsive to certain environmental stimuli and may control genes necessary for the production of alternative exopolysaccharides.     

Chloroflexus-like organisms from marine and hypersaline environments: Distribution and diversity

We report the presence of a diverse number ofChloroflexus-like organisms in intertidal marine and submerged hypersaline microbial mats using light, infrared fluorescence, and electron microscopy. The intertidal organisms appear morphologically very similar to thermophilicC. aurantiacus while the 2 hypersaline strains are larger and have a more complex ultrastructure composed of chlorosome-bearing internal membranes that appear to arise as invaginations of the cell membrane. By comparing spectroradiometry of microbial mat layers with microscopic observations, we have confirmed that theChloroflexus-like organisms are major constituents of the hypersaline microbial mat communities. In situ studies on mat layers dominated byChloroflexus-like organisms showed that sulfide-dependent photoautotrophic activity sustained by near infrared radiation prevailed. Autoradiographic analyses revealed that autotrophy was sustained in the filaments by 750 nm radiation. Three morphologically distinct strains are now maintained in mixed culture. One of these appears to be growing photoautotrophically.     

Seasonal determinations of extracellular hydrolytic activities in heterotrophic and mixed heterotrophic/autotrophic biofilms from two contrasting rivers

The temporal changes in extracellular enzyme activities in freshwater microbial biofilms were examined in two contrasting river sites in North Wales over a 12 month period. Sites were a first order, unshaded oligotrophic upland stream (Nant Waen) and a fourth order, mildly eutrophic river with riparian tree cover (River Clywedog). When algal populations were low, biofilms of the more eutrophic site supported greater enzyme activities and higher population densities than the oligotrophic site. Composition, concentration and origin of substrates available to the respective biofilm communities influenced extracellular processing patterns. Reduction in algal populations depressed total and extracellular activities in biofilms from the first order site, suggesting that biofilm communities here were maintained by in situ primary production. Biofilms from Nant Waen were often found to contain higher extracellular activities per cell than the more eutrophic River Clywedog biofilms, which might represent the enhanced ability of an oligotrophic biofilm to accumulate extracellular enzymes. In contrast, light and darkgrown River Clywedog biofilms were not enzymatically distinct, inferring a less important role for biofilm phototrophs. Some evidence was found for increased reliance on allochthonous substrates in the River Clywedog for biofilm maintenance.     

Rhodospirillum sodomense,sp. nov., a Dead Sea rhodospirillum species

A new species of halophilic anoxygenic purple bacteria of the genus Rhodospirillum is described. The new organism, isolated from water/sediment of the Dead Sea, was vibrio-shaped and an obligate halophile. Growth was best at 12% NaCl, with only weak growth occurring at 6% or 21% NaCl. Growth occurred at Mg²⁺ concentrations up to 1 M but optimal growth was obtained at 0.05–0.1 M Mg²⁺. Bromide was well tolerated as an alternative anion to chloride. The new organism is an obligate phototroph, growing photoheterotrophically in media containing yeast extract and acetate or a few other organic compounds. Growth of the Dead Sea Rhodospirillum species under optimal culture conditions was slow (minimum t_d 20 h). Cells contained bacteriochlorophyll a and carotenoids of the spirilloxanthin series and mass cultures were pink in color. Absorption spectra revealed the presence of a B875 (light-harvesting I) but no B800/B850 (light-harvesting II) photopigment complex. The new organism shares a number of properties with the previously described halophilic phototrophic bacterium Rhodospirillum salinarum and was shown to be related to this phototroph by 16S rRNA sequencing. However, because of its salinity requirements, photosynthetic properties, and isolation from the Dead Sea, the new phototroph is proposed as a new species of the genus Rhodospirillum, R. sodomense.     

The structural role of the carotenoid in the bacterial light-harvesting protein 2 (LH2) of Rhodonbacter capsulatus. A Fourier transform Raman spectroscopy and circular dichroism study

In previous work (Zurdo J, Fernández-Cabrera C and Ramírez JM (1993) Biochem J 290: 531–537), it had been shown that selective extraction of the carotenoid from the light-harvesting protein 2 (LH2) of Rhodobacter capsulatus induced the dissociation of 800-nm absorbing bacteriochlorophyll (Bchl), a 10-nm red shift of 854-nm Bchl, and a decrease of the stability of the protein in detergent solution. In the present study, the Fourier transform Raman and near-infrared circular dichroism spectra of native and carotenoid-depleted LH2 membrane preparations were compared. It was found that while the coupled carbonyls of 854-nm Bchl remained specifically H-bonded to the peptides after carotenoid extraction, the optical activity of the near-infrared electronic transition was significantly altered. Given the excitonic origin of such optical activity, our data suggest that carotenoid extraction elicits a rearrengement of the chromophore cluster and of the associated polypeptide subunits. This implies a significant role of the carotenoid in maintaining the native quaternary structure of the protein, which would be consistent with the observed dissociation of 800-nm Bchl and the loss of solubilized LH2 stability that result from carotenoid removal. There is no evidence for a similar role of the carotenoid in the LH1 protein.Abbreviations Bchl bacteriochlorophyll - FT Fourier transform - CD circular dichroism - LH1 and LH2 the bacterial light-harvesting proteins 1 and 2 In memoriam of Daniel I. Arnon.     

Dynamics of biofilm detachment in biofilm airlift suspension reactors

The dynamic change in the overall detachment rate of spherical biofilms in a biofilm airlift suspension reactor was measured after a downshift of the substrate loading rate to zero while all other conditions remained constant. In contrast to the expectations, the overall detachment rate decreased rapidly to a nearly stable level. Correlations available from literature were not able to describe this phenomenon. Concepts were formulated which can describe the observations from this study. Research under dynamic conditions and careful monitoring of the biofilm surface area and biofilm morphology are necessary to elucidate and discriminate biofilm detachment mechanisms. (c) 1995 John Wiley & Sons, Inc.