Microbially — related redox changes in a subtropical lake I.In situ monitoring of the annual redox cycle

Using a recently developedin situ multiprobe the redox development in the water column of warm-monomictic Lake Kinneret was investigated during three annual cycles. During the time when sulfide release into the meta- and hypolimnion is initiated, our measurements show a linear relationship, close to the thermodynamic function, between the platinum electrode potential and the amount of sulfide produced by the sulfate reducing bacteria. A change of this relationship during summer stratification coincides with the bloom of the phototrophic sulfur bacteriumChlorobium phaeobacteroides.     

High performance liquid chromatography detection of phototrophic bacterial pigments in aquatic environments

Pigment extracts of phototrophic bacteria isolated from Lake Kinneret (Rhodopseudomonas palustris, Thiocapsa roseopersicina, Prosthecochloris aestuaris andChlorobium phaeobacteroides) were studied by means of high performance liquid chromatography (HPLC). An absorption wavelength of 360 nm provided the best resolution among the pigments of the species tested and between them and chlorophylla. Signature pigments were identified for each of these species, and their presence was thereby monitored in lake water samples.C. phaeobacteroides, which was observed in the anaerobic hypolimnion and predominated in the metalimnion, was recognized by a characteristic cluster of major chlorophyllous pigment peaks. The spectral qualities of these pigments were close but not identical to published data on bacteriochlorophylle, presumably due to the use of different solvents for extraction. The intensity of these pigment peaks was employed to determine the depth of the greatest phototrophic bacterial biomass, which was not related to that of algae.     

Population dynamics of phototrophic bacteria in three basins of Lake Banyoles (Spain)

The annual dynamics of the phototrophic bacterial populations developing in the anoxic layers has been monitored in three basins of the northern area of Lake Banyoles (Spain). Although two of the studied basins are meromictic and one is holomictic, chemical properties of the water are almost identical. Therefore, differences in both the spatial and temporal distribution, as well as in the composition of phototrophic bacterial communities, dominated by Chlorobium phaeobacteroides and Chromatium minus, are discussed on the basis of the structural and morphometric characteristics of each basin. Both species showed the same physiological adaptations to light intensity changes by modifying the carotenoid/bacteriochlorophyll ratio. Light reaching the oxic-anoxic boundary appears to be the most important factor controlling the growth of phototrophic bacteria in Lake Banyoles. The oxic-anoxic boundary becomes shallower as summer advances, until enough light is available for bacterial photosynthesis.     

Sulfide utilization by the photosynthetic bacterium Chlorobium phaeobacteroides

Sulfide and sulfur are used by the photosynthetic bacterium Chlorobium phaeobacteroides as electron donors. Sulfide and sulfur consumption was found to be affected by sulfide concentration in the medium. Raising the sulfide concentration from 0.28 mM to 5.05 mM caused an increase in the amount of S⁼ utilized per growth unit from 0.58 mM to 2.32 mM. This increase in sulfide utilization was not reflected in a higher photosynthetic activity. Sulfide and sulfur consumption was also influenced by light intensity, with higher light intensity sulfide consumption was increased. In Lake Kinneret, Chlorobium phaeobacteroides did not bloom in the thermocline layer until sulfide concentrations reached 0.03–0.06 mM.     

Light-harvesting adaptations of planktonic phototrophic micro-organisms to different light quality conditions

The effects of light spectral distribution on the composition of phototrophic microbial communities were analyzed in three metalimnetic levels (relative depth positions) of 41 lakes. Principal Component Analysis was used to compare light quality conditions reaching the populations of phototrophic micro-organisms containing different photosynthetic pigments. Results allowed to identify the optimal light quality conditions for the selection of each microbial group at their respective levels. Two general light-harvesting adaptations were defined, according to the wavebands that could be related to the selection of these microbial groups. The micro-organisms adapted to use red and near-infrared light – eukaryotic phytoplankton, Chloronema spp. and green-coloured Chlorobiaceae – predominated at shallow depths (specially in waters containing high gilvin contents) using their respective Q_y absorption bands. The micro-organisms adapted to green-yellow light – phycoerythrin-containing cyanobacteria, Chromatiaceae and brown-coloured Chlorobiaceae – were dominant in deep metalimnetic communities. Laboratory experiments with cultures of Chlorobium limicola and C. phaeobacteroides growing under different light quality conditions showed that the green-coloured species had higher photosynthetic activity under red light, while the brown-coloured species was more active under green light. These results demonstrated that physiological differences between micro-organisms with different light-harvesting adaptations are responsible of their selection under different light quality conditions. This selection is experimented by Chlorobiaceae (as it was previously indicated by other investigators) at the deepest positions of the metalimnetic communities (level 3), but also by Chromatiaceae and Chloronema spp. at level 2 and by the eukaryotic phytoplankton and cyanobacteria at level 1.     

Nutrient loading and drainage channel response in a ricefield system

The distribution of photosynthetic bacteria in Lake Kinneret and their contribution to primary productivity was investigated. Both Chlorobium phaeobacteroides and Rhodopseudomonas palustris were detected from mid-June through October in 1987 and reached a maximum of 7 × 10⁶ cells/ml in the metalimnion by mid-August. Chlorobium density was greater towards the end of the bloom period, when the thermocline was exposed to higher sulfide concentrations and lower light intensities. Rhodopseudomonas peaked earlier in the season, when less sulphide was present and light irradiation was higher. This was the first time that R. palustris was observed in Lake Kinneret; in addition to its presence at the thermocline, it was found also at the epilimnion and hypolimmon. The contribution of photosynthetic bacteria to the total primary productivity of the lake was small (∼1%). Low but perceptible rates of N₂ fixation in situ were also demonstrated by photosynthetic bacteria present in the metalimnion.     

Comparative study of two meromictic basins of Lake Banyoles (Spain) with sulphur phototrophic bacteria

The annual limnological dynamics of two meromictic basins of Lake Banyoles (C-III and C-IV) have been studied and compared on the basis of their physical, chemical and biological characters. Stability values calculated for both basins gave 865 g cm cm⁻² and 495 g cm cm⁻² for C-III and C-IV respectively. These values are in agreement with the fact that C-IV was almost completely mixed during winter. In this basin, during stratification, the monimolimnion increased in thickness as the stability increased. Isolation of the respective monimolimnia resulted in the development of anoxic conditions and the accumulation of sulphide in both C-III and C-IV, which favoured the development of dense populations of sulfur phototrophic bacteria. The purple sulphur bacterium Chromatium minus and the green sulphur bacterium Chlorobium phaeobacteroides were identified as the main components of these photosynthetic populations. The different depths at which the O₂/H₂S boundary was situated in both basins (and consequently the different light intensity reaching this zone) determined the growth of these bacteria. Light intensities at the chemocline of C-IV reached values up to 5% of surface incident light. In contrast, in C-III this variable was sensibly lower, with values depending on season and seldom reaching 1%. Phototrophic bacteria were consequently found earlier in C-IV than in C-III, where no significant concentrations were found until August. Finally stability is discussed as an important factor controlling chemical and biological dynamics in meromictic lakes.     

In situ abundance and carbon fixation activity of distinct anoxygenic phototrophs in the stratified seawater lake Rogoznica

Sulphide-driven anoxygenic photosynthesis is an ancient microbial metabolism that contributes significantly to inorganic carbon fixation in stratified, sulphidic water bodies. Methods commonly applied to quantify inorganic carbon fixation by anoxygenic phototrophs, however, cannot resolve the contributions of distinct microbial populations to the overall process. We implemented a straightforward workflow, consisting of radioisotope labelling and flow cytometric cell sorting based on the distinct autofluorescence of bacterial photopigments, to discriminate and quantify contributions of co-occurring anoxygenic phototrophic populations to in situ inorganic carbon fixation in environmental samples. This allowed us to assign 89.3% ± 7.6% of daytime inorganic carbon fixation by anoxygenic phototrophs in Lake Rogoznica (Croatia) to an abundant chemocline-dwelling population of green sulphur bacteria (dominated by Chlorobium phaeobacteroides), whereas the co-occurring purple sulphur bacteria (Halochromatium sp.) contributed only 1.8% ± 1.4%. Furthermore, we obtained two metagenome assembled genomes of green sulphur bacteria and one of a purple sulphur bacterium which provides the first genomic insights into the genus Halochromatium, confirming its high metabolic flexibility and physiological potential for mixo- and heterotrophic growth.     

Anoxygenic phototrophic bacteria of the high-altitude meromictic Lake Gek-Gel,Azerbaijan

The anoxygenic phototrophic bacterial community of the high-altitude meromictic Lake Gek-Gel (Azerbaijan) was investigated in September 2003. The highest concentration of bacteriochlorophyll e (48 μg/l) was detected at a depth of 30 m; the peak of bacteriochlorophyll a (4.5 μg/l) occurred at 29 m. Phylogenetic analysis revealed that brown-colored green sulfur bacteria Chlorobium phaeobacteroides predominated in the lake. Nonsulfur purple bacteria phylogenetically close to Blastochloris sulfoviridis were found in insignificant amounts; these organisms have not been previously reported in Lake Gek-Gel.     

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.     

Sedimentation of photosynthetic pigments during the bloom of the green sulfur bacterium <Emphasis Type="Italic">Chlorobium phaeobacteroides</Emphasis> in Lake Kinneret: spatial patterns

Sediment fluxes were studied in the subtropical Lake Kinneret (Israel) in 2006, 2007, and 2008 from mid-June to October, when lake was chemically stratified and the green sulfur bacterium Chlorobium phaeobacteroides formed a dense population in the anoxic metalimnion. The rate of seston accumulation in traps was measured with sedimentation traps positioned along an offshore transect connecting the littoral zone and lake center. The sediment fluxes increased from the lake center towards the littoral, while the percentage of organic material (OM) decreased correspondingly. High fluxes of bacteriochlorophyll e (BChl e—a signature pigment of Chl. phaeobacteroides) and chlorophyll a (Chl a—a marker for eukaryotic algae and cyanobacteria) were detected in all locations. The relative contributions of Chl a and BChl e to the bulk of the accumulated OM were higher in traps positioned below the thermocline in the pelagic zone than in traps located near the shore line. The presence of BChl e in traps exposed to oxic conditions in the littoral, where Chl. phaeobacteroides does not develop, implies horizontal translocation of cells from the lake center towards its periphery. We assume that seiche-mediated movement of particles embedded in the metalimnetic waters is the most probable explanation for the existence of Chl. phaeobacteroides tracer in an oxic environment, but do not exclude the possibility of resuspension of settled particles as source for BChl e in littoral traps. The green sulfur bacteria are potentially important component of the sediment flux of photosynthesizing organisms in a thermally stratified lake and should be taken into account when carbon budget are constructed.     

Seasonal dynamic of phototrophic epibionts on crustacean zooplankton in a eutrophic reservoir with cyanobacterial bloom

Seasonality of burden and prevalence of phototrophic (microalgal) epibionts Characidiopsis ellipsoidea, Colacium vesiculosum and Colacium sp. on dominating crustacean zooplankton (Daphnia longispina, Cyclops vicinus and Mesocyclops leuckarti) were studied in a small reservoir Bugach with cyanobacterial bloom. The correlations between the seasonal dynamics of prevalence and the dynamics of others biotic and abiotic factors were calculated. The conclusions were as follows. The substrate species, that determined the development of the epibionts on the three studied crustacean zooplankton, was Daphnia longispina (Cladocera). Despite intensive epibiotic infestation of crustacean zooplankton, epibionts did not appear to have caused non-consumptive mortality of the crustacean zooplankton. But they could have contributed to the Daphnia summer decline by increasing mortality due to its consumption by planktivorous fishes. The phototropic epibionts may successfully coexist with cyanobacterial bloom. The possible role of the epibionts in changing nutrient fluxes in pelagic food web is discussed.     

Nutrient and iron limitation to <Emphasis Type="Italic">Ulva</Emphasis> blooms in a eutrophic coastal lagoon (Sacca di Goro,Italy)

Growth patterns and bloom formation of the green seaweed Ulva rigida were analysed in the eutrophic Sacca di Goro lagoon (Po River Delta, Italy). Variations of standing biomasses and elemental composition of Ulva were analysed through an annual cycle with respect to nitrogen, phosphorus and iron. Growth rates, nutrient and iron uptake and nitrate storage by macroalgal thalli were also assessed with field experiments during the formation of a spring bloom. The control of Ulva growth and the bloom formation depended on multiple factors, especially on nitrogen availability and iron deficiency. In the nitrate rich waters of the Sacca di Goro lagoon, nitrate accumulation in Ulva thalli was inversely related with Fe uptake, indicating an influence of Fe limitation on N acquisition. Since length and magnitude of nitrate luxury uptake are inversely related to the size of the intracellular nitrate pools, in nitrate rich waters the fast growing Ulva may face risk of N-limitation not only when exposed to low N concentrations or at high biomass levels, but also when exposed to pulsed dissolved nitrate concentrations at low iron availability. The potential Fe limitation could be affected by processes controlled by geochemical reactions and by macroalgal growth and decomposition. Both Fe oxidation during the active macroalgal growth and the formation of insoluble FeS and FeS₂ during bloom collapse can result in a drastic decrease of soluble iron. Thus, a potential limitation of Fe to macroalgae can occur, determining positive feedbacks and potentially controlling the extent of bloom development and persistence.     

Phototrophic bacterial production of oleic acid in an illuminated upflow anaerobic sludge blanket reactor

Phototrophic bacterial cells in the effluent from a lighted upflow anaerobic sludge blanket reactor supplied with a medium containing 142 mg S (as SO₄ ^2–) l^–1 accumulated a 6.8% w/w oleic acid content in cells and 19 mg cell-bound oleic acid l^–1 in the effluent. Pure cultures of Rhodopseudomonas palustris and Blastochloris sulfoviridis isolated from the effluent also accumulated 5.1 and 6.4% w/w oleic acid contents in cells, respectively. The oleic acid content in the cells recovered from the LUASB reactor effluent was related to the phototrophic bacterial population in the LUASB reactor. The inverse relationship was observed in the LUASB reactor between phototrophic bacterial growth and sulfate concentration in the influent.     

Effects of light quality on the physiology and the ecology of planktonic green sulfur bacteria in lakes

The effect of light quality on the selection of natural populations of Green Sulfur Bacteria (Chlorobiaceae) is considered to be a classic factor in the determination of their ecological niches. From the comparison among phototrophic bacterial populations of lakes, it is shown that brown and green pigmented groups of Chlorobiaceae have a differential distribution depending on depth. Statistical analyses prove that green species, which dominate at shallow oxic/anoxic boundaries, are correlated to light spectra enriched in long wavelengths, while brown ones are found when light spectra are enriched in the central region of the spectrum, as in deeper lake layers. Physiological experiments have been made withChlorobium limicola andC. phaeobacteroides cultures placed under different light quality conditions, in order to verify these hypotheses made on a field data basis. Results show that red and white light has more positive effects on the green bacterium than on the brown. Blue and green light illuminations have opposite consequences. Therefore, the effect of shallow depths and Chromatiaceae shading—which also increases the proportion of long wavelengths in light spectra—benefits the bacteriochlorophyll-based strategies of green species. On the other hand, the carotenoid-based strategies of brown ones are favored by the light climates usually dominant at greater depths. Thus, brown species are considered to be singular adaptations of Chlorobiaceae to depth, where bacteriochlorophyll light-harvesting is strongly limited by light quality.     

Characterization of cell-envelope fractions of chemotrophically and phototrophically grown Rhodospirillum tenue

Envelope preparations of chemotrophically and phototrophically grown Rhodospirillum tenue were isolated and characterized on the basis of their contents and composition in phospholipids and fatty acids, as well as on the basis of their enzyme activities, absorption spectra and polypeptide patterns. Both preparations were similar in their contents in phospholipids and fatty acids. Their total fatty acids were characterized by a rather high percentage of saturated fatty acids. The activities of the respiratory reactions were considerably higher in chemotrophic than in phototrophic membranes. This is true especially for activities of the terminal oxidase which were over 20 times greater. The affinities of the corresponding enzymes to their respective substrates (K _m ) differed with differences in the culture conditions. Under chemotrophic conditions the K _m values for the NADH-dependent reactions were lower than those values under phototrophic conditions, whereas the K _m values for the succinate dependent reactions were higher. The low temperature (77°K) near infrared spectrum of the phototrophic membrane showed a peak at 875 nm which was not detectable in the chemotrophic membrane. The polypeptide patterns, in the presence of sodium dodecyl sulfate, of both preparations were quite similar except for the presence of two low molecular weight proteins (M. Wt. 12,000 and 10,000) in the phototrophic membrane which were absent in the chemotrophic membrane. Both envelope preparations were further fractionated into enriched cytoplasmic membrane and outer membrane fractions which were identified on the basis of their contents in 2-keto-3-deoxyoctonate, ubiquinone 8 and bacteriochlorophyll (in the case of the phototrophic membrane) and their enzyme activities. The buoyant densities of the corresponding fractions from both envelope preparations were found to be equal. The data presented in this paper demonstrate that envelope preparations of chemotrophically and phototrophically grown R. tenue are similar in their contents in 2-keto-3-deoxyoctonate, ubiquinone 8, phospholipids and fatty acids, yet differ significantly in their spectra, protein patterns and enzyme activities.Abbreviations BChl bacteriochlorophyll - UQ_s ubiquinone 8 - KDO 2-keto-3-deoxyoctonate - PG phosphatidyl glycerol - PE phosphatidyl ethanolamine - DCPIP 2,6-dichlorophenolindophenol - PMS phenazine methosulfate - SDS sodium dodecyl sulfate     

Genes essential for phototrophic growth by a purple alphaproteobacterium

Tn‐seq was used to identify genes essential for phototrophic growth by the purple bacterium Rhodopseudomonas palustris. About 167 genes required for anaerobic growth on acetate in light were identified, 35 of which are annotated as photosynthesis genes. The essentiality of many of these genes by analysing the phenotypes of independently generated mutants that had altered pigmentation was verified. Three genes were identified, two possibly involved in biogenesis of the membrane‐bound photosynthetic apparatus and one for phosphatidylcholine biosynthesis, that were not known to be essential for phototrophic growth. Site‐directed mutagenesis was used to show that the NADH:quinone oxidoreductase complex I_E was essential for phototrophic growth under strictly anaerobic conditions and appeared to play a role in reverse electron transport to generate NADH. A homologous NADH:quinone oxidoreductase complex I_A likely operates in the opposite direction to oxidize NADH. The operation of the two enzymes in opposition would allow R. palustris to maintain redox balance. As a complement to the genetic data, proteomics experiments were carried out in which it was found that 408 proteins were present in significantly higher amounts in cells grown anaerobically in light compared with aerobically. Among these were proteins encoded by subset of the phototrophic growth‐essential genes.     

Isolation by Time During an Arctic Phytoplankton Spring Bloom

The arctic phytoplankton spring bloom, which is often diatom‐dominated, is a key event that provides the high latitude communities with a fundamental flux of organic carbon. During a bloom, phytoplankton may increase its biomass by orders of magnitude within days. Yet, very little is known about phytoplankton bloom dynamics, including for example how blooming affects genetic composition and diversity of a population. Here, we quantified the genetic composition and temporal changes of the diatom Fragilariopsis cylindrus, which is one of the most important primary producers in the Arctic, during the spring bloom in western Greenland, using 13 novel microsatellite markers developed for this study. We found that genetic differentiation (quantified using sample‐specific F_ST) decreased between time points as the bloom progressed, with the most drastic changes in F_ST occurring at the start of the bloom; thus the genetic structure of the bloom is characterized by isolation by time. There was little temporal variation in genetic diversity throughout the bloom (mean H_E = 0.57), despite marked fluctuations in F. cylindrus cell concentrations and the temporal change in sample‐specific F_ST. On the basis of this novel pattern of genetic differentiation, we suggest that blooming behavior may promote genetic diversity of a phytoplankton population.     

Long-Term Population Dynamics of Phototrophic Sulfur Bacteria in the Chemocline of Lake Cadagno, Switzerland

Population analyses in water samples obtained from the chemocline of crenogenic, meromictic Lake Cadagno, Switzerland, in October for the years 1994 to 2003 were studied using in situ hybridization with specific probes. During this 10-year period, large shifts in abundance between purple and green sulfur bacteria and among different populations were obtained. Purple sulfur bacteria were the numerically most prominent phototrophic sulfur bacteria in samples obtained from 1994 to 2001, when they represented between 70 and 95% of the phototrophic sulfur bacteria. All populations of purple sulfur bacteria showed large fluctuations in time with populations belonging to the genus Lamprocystis being numerically much more important than those of the genera Chromatium and Thiocystis. Green sulfur bacteria were initially represented by Chlorobium phaeobacteroides but were replaced by Chlorobium clathratiforme by the end of the study. C. clathratiforme was the only green sulfur bacterium detected during the last 2 years of the analysis, when a shift in dominance from purple sulfur bacteria to green sulfur bacteria was observed in the chemocline. At this time, numbers of purple sulfur bacteria had decreased and those of green sulfur bacteria increased by about 1 order of magnitude and C. clathratiforme represented about 95% of the phototrophic sulfur bacteria. This major change in community structure in the chemocline was accompanied by changes in profiles of turbidity and photosynthetically available radiation, as well as for sulfide concentrations and light intensity. Overall, these findings suggest that a disruption of the chemocline in 2000 may have altered environmental niches and populations in subsequent years.     

Photoheterotrophic and chemoheterotrophic dinitrogen fixation and nitrate utilization by the cyanobacteriumAnabaena torulosa

Anabaena torulosa exhibited fructose-dependent growth, heterocyst differentiation and N₂ fixation in nitrate-free (diazotrophic) cultures in photoheterotrophic and chemoheterotrophic conditions. The incorporation of nitrate into such cultures inhibited the formation of heterocysts and N₂ fixation. The rate of NO ₃ ⁻ uptake byA. torulosa in photoautotrophic, photoheterotrophic and chemoheterotrophic conditions was similar but it increased by 100% in phototrophic conditions. The activity of glucose-6-phosphate dehydrogenase was found to be maximum in phototrophic and photoheterotrophic conditions. Ferredoxin-NADP⁺ reductase, nitrate reductase and glutamate-ammonia ligase activities suggest that nitrate utilization takes place in nonphotosynthetic conditions.