O. N. Lunina

[Seasonal changes in the structure of the anoxygenic photosynthetic bacterial community in Lake Shunet, Khakassia].

Seasonal studies of the anoxygenic phototrophic bacterial community of the water column of the saline eutrophic meromictic Lake Shunet (Khakassia) were performed in 2002 (June) and 2003 (February–March and August). From the redox zone down, the lake water was of dark green color. Green sulfur bacteria predominated in every season. The maximum number of green sulfur bacteria was 107 cells/ml in summer and 106 cells/ml in winter. A multi-syringe stratification sampler was applied for the study of the fine vertical distribution of phototrophs in August 2003; the sampling was performed every 5 cm. A 5-cm-thick pink-colored water layer inhabited by purple sulfur bacteria was shown to be located above the layer of green bacteria. The species composition and ratio of purple bacterial species depended on the sampling depth and on the season. In summer, the number of purple sulfur bacteria in the layer of pink water was 1.6 × 108 cells/ml. Their number in winter was 3 × 105 cells/ml. In the upper oxygen-containing layer of the chemocline the cells of purple nonsulfur bacteria were detected in summer. The maximum number of nonsulfur purple bacteria, 5 × 102 cells/ml, was recorded in August 2003. According to the results of the phylogenetic analysis of pure cultures of the isolated phototrophic bacteria, which were based on 16S rDNA sequencing, green sulfur bacteria were close to Prosthecochloris vibrioformis, purple sulfur bacteria, to Thiocapsa and Halochromatium species, and purple nonsulfur bacteria, to Rhodovulum euryhalinum and Pinkicyclus mahoneyensis.

Anoxygenic phototrophic bacteria of the Kislo-Sladkoe stratified lake (White Sea, Kandalaksha Bay)

The community of anoxygenic phototrophic bacteria (APB) in the water column of the Kislo-Sladkoe stratified lake recently isolated from the sea (White Sea, Kandalaksha Bay) was investigated in September 2010. The water of the sulfide-rich zone was greenish-brown due to intense development of green sulfur bacteria (GSB). Nine APB strains were isolated from the water samples: three belonging to GSB, five, to purple sulfur bacteria (PSB), and one, to purple nonsulfur bacteria (PNB). GSB predominated in the phototrophic community of the chemocline. Unexpectedly, two morphologically different green-colored GSB strains were found to be phylogenetically identical and related to the brown-colored Chlorobium phaeovibrioides (99% similarity according to the 16S rRNA gene sequencing). Homology to the closest green-colored species (Chlorobium luteolum) was 98%. Two morphologically and physiologically similar PSB strains (TcrPS10 and AmPS10) had rounded cells containing okenone and gas vesicles. According to the 16S rRNA gene sequencing, these strains were most closely related (99%) to two different Thiocapsa species: Tca. marina (containing okenonee and no gas vesicles) and Tca. rosea (containing spirilloxanthin and gas vesicles). The remaining isolates of purple bacteria were similar to the already described APB species.

Microbiological and isotopic geochemical investigation of Lake Kislo-Sladkoe, a meromictic water body at the Kandalaksha Bay shore (White Sea)

Microbiological, biogeochemical, and isotopic geochemical investigation of Lake Kislo-Sladkoe (Polusolenoe in early publications) at the Kandalaksha Bay shore (White Sea) was carried out in September 2010. Lake Kislo-Sladkoe was formed in the mid-1900s out of a sea gulf due to a coastal heave. At the time of investigation, the surface layer was saturated with oxygen, while near-bottom water contained sulfide (up to 32 mg/L). Total number of microorganisms was high (12.3 × 106 cells/mL on average). Light CO2 fixation exhibited two pronounced peaks. In the oxic zone, the highest rates of photosynthesis were detected at 1.0 and 2.0 m. The second, more pronounced peak of light CO2 fixation was associated with activity of anoxygenic phototrophic bacteria in the anoxic layer at the depth of 2.9 m (413 μg C L−1 day−1). Green-colored green sulfur bacteria (GSB) predominated in the upper anoxic layer (2.7–2.9 m), their numbers being as high as 1.12 × 104 cells/mL, while brown-colored GSB predominated in the lower horizons. The rates of both sulfate reduction and methanogenesis peaked in the 2.9 m horizon (1690 μg S L−1 day−1 and 2.9 μL CH4 L−1 day-1). The isotopic composition of dissolved methane from the near-bottom water layer (δ13C (CH4) = −87.76‰) was significantly lighter than in the upper horizons (δ13C (CH4) = −77.95‰). The most isotopically heavy methane (δ13C (CH4) = −72.61‰) was retrieved from the depth of 2.9 m. The rate of methane oxidation peaked in the same horizon. As a result of these reactions, organic matter (OM) carbon of the 2.9 m horizon became lighter (−36.36‰), while carbonate carbon became heavier (−7.56‰). Thus, our results demonstrated that Lake Kislo-Sladkoe is a stratified meromictic lake with active microbial cycles of carbon and sulfur. Suspended matter in the water column was mostly of autochthonous origin. Anoxygenic photo-synthesis coupled to utilization of reduced sulfur compounds contributed significantly to OM production.

Fractionation of Stable Carbon Isotopes by Photoautotrophically Growing Anoxygenic Purple and Green Sulfur Bacteria

Fractionation of stable carbon isotopes 12C and 13C by three pure cultures of photoautotrophic purple sulfur bacteria (Ectothiorhodospira shaposhnikovii, Lamprocystis purpureus, and Thiocapsa sp.) (PSB) and the green sulfur bacterium Prosthecochloris sp. (GSB) was investigated in 13–15-day experiments. The cultivation was carried out in a luminostat (2000 lx) on mineral media with 1–1.5 g/l NaHCO3 (inoculum) with the subsequent transfer to the medium with up to 10 g/l NaHCO3. For PSB, the difference in the quantitative characteristics of the isotopic composition of suspended carbon (including bacterial cells) and mineral carbon of the medium (Δ13C = δ13Csubstrate − δ13Cbiomass) changed from 15.0 to 34.3‰. For GSB, the range of Δ13C changes was significantly less (18.3–22.7‰). These data suggested the possibility of a pool of soluble mineral carbon in PSB cells. The pool of intracellular mineral carbon was calculated; depending on the PSB species and growth stage, it varied from 0 to 68% of the total cell carbon. The α coefficients reflecting the carbon isotope fractionation by PSB and GBS and calculated from the changes of the bicarbonate carbon isotopic composition in the medium depending on its consumption were 1.029 ± 0.003 and 1.019 ± 0.001, respectively. These α values did not depend on the growth rate. CO2 fixation on ribulose-bisphosphate was shown to be the major factor determining the carbon isotope fractionation by PSB; at the stage of CO2 penetration into the cell, fractionation was insignificant. In GSB, fractionation occurred mostly at CO2 penetration into the cell, while it was insignificant at the stage of carbon dioxide fixation in the reverse TCA cycle. Analysis of the isotopic data of the photosynthesis by PSB and GSB in meromictic lakes also revealed that in PSB-dominated natural communities suspended organic matter was more enriched with light 13C (Δ13C = 23.4−24.6‰) than in the communities with more active GSB (Δ13C = 10.2−14.0‰)

Anoxygenic phototrophic bacterial community of Lake Shira (Khakassia)

The anoxygenic phototrophic bacterial community of the brackish meromictic Lake Shira (Khakassia) was investigated in August 2001, July 2002, and February–March 2003. In all the periods of investigation, the prevailing microorganisms were purple sulfur bacteria similar to Lamprocystis purpurea in morphology and pigment composition. Their highest number (3 × 105 cells/ml) was recorded in July 2002 at the depth of 15 m. According to 16S rRNA gene analysis, the strain of purple sulfur bacteria isolated in 2001 and designated ShAm01 exhibited 98.6% similarity to the type strain of Thiocapsa roseopersicina and 97.1–94.4% similarity to the type strains of Tca. pendens, Tca. litoralis, and Tca. rosea. The minor microorganisms of the anoxygenic phototrophic bacterial community within the period of investigation were nonsulfur purple bacteria phylogenetically close to Rhodovulum strictum (98.3% similarity, strain ShRb01), Ahrensia kielensis (of 93.9% similarity, strain ShRb02), Rhodomicrobium vannieli (of 99.7% similarity, strain ShRmc01), and green sulfur bacteria, phylogenetically close to Chlorobium limicola (of 98.7% similarity, strain ShCl03).

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.

Biological fractionation of stable carbon isotopes at the aerobic/anaerobic water interface of meromictic water bodies

Mass-spectrometric investigation of carbon isotope composition (δ13C) was carried out for suspended organic matter and dissolved mineral compounds for the water column of some meromictic water bodies differing in salinity and trophic state. As a rule, a more pronounced carbon isotope fractionation (resulting from the metabolism of phytoplankton and anoxygenic phototrophic bacteria) was revealed in the zones of enhanced oxygenic and anoxygenic photosynthesis. Carbon isotope fractionation at the border between oxidized and reduced waters depends both on the activity of microbial communities and on the dominant species of phototrophic microorganisms. Analysis of the distribution profiles of the isotopic composition of suspended organic matter and dissolved mineral carbon revealed active mineralization of the organic matter newly formed via anoxygenic photosynthesis in the monimolimnion by microbial communities, resulting in the release of isotopically light carbon dioxide. Mineral carbon in the anaerobic zones of highly productive meromictic water bodies is therefore enriched with the light 12C isotope.

Succession Processes in the Anoxygenic Phototrophic Bacterial Community in Lake Kislo-Sladkoe (Kandalaksha Bay, White Sea)

The community of anoxygenic phototrophic bacteria (APB) in the water column of Lake Kislo- Sladkoe (Kandalaksha Bay, White Sea), which has recently become separated from the sea, was investigated in March?April 2012, March?April 2013, and in September 2013. The lake, which was previously considered meromictic, was in fact mixed and was strongly affected by the sea. In winter the lake is sometimes washed off with seawater, and this together with the seasonal cycles of succession processes determines the succession of the community. The consequences of the mixing in autumn 2011 could be observed in the APB community as late as autumn 2013. Green-colored green sulfur bacteria (GSB) usually predominated in the chemocline. In winter 2013 stagnation resulted in turbidity of water under the ice, which was responsible for both predominance of the brown GSB forms and the changes ratio of the species of purple sulfur bacteria (PSB) in anoxic water layers. Production of anoxygenic photosynthesis in the lake was at least 240 mg C m-2 day-1 in September and 0–20 mg C m–2 day–1 in March—April, which corresponded to 40 and 69%, respectively, of oxygenic photosynthesis. Okenone-containing purple sulfur bacteria, strain TcakPS12, were isolated in 2012 from lake water. The ells of this strain form filaments of not separated cells. Strain TcakPS12 exhibited 98% similarity with the type strains of Thiocapsa pendens DSM 236 and Thiocapsa bogorovii BBS, as well as with the strains AmPS10 and TcyrPS10, which were isolated from Lake Kislo-Sladkoe in 2010.

Depth profiles of spectral and hydrological characteristics of water and their relation to abundances of green sulfur bacteria in the stratified lakes of the White Sea

We analyze the results received from two expeditions performed in August-September 2013, August-September 2014 and February 2015 in the Kandalaksha Bay of the White Sea. Depth profiles of hydrological characteristics and optical properties of water were recorded for five marine lakes being on different stages of isolation from the White Sea. Those relic lakes demonstrate a tendency to meromixis and are characterized by apparent stratification of the water bodies from the brackish top layer to the bottom salt water. Maximal concentrations of anoxygenic phototrophs (green sulfur bacteria) were found at depths close to the redox interface in all the studied lakes. To discriminate differently pigmented groups of microorganisms the fluorescence emission spectra of bacteriochlorophylls from the living cells were used. We puzzle out the data on light spectrum propagation through the water body in each lake using optical properties of water (attenuation spectra) in the UV, visible and NIR ranges, as well as direct measurements of the total irradiances at various depths. The changes in optical characteristics of water in the stratified reservoirs due to cromophoric dissolved organic matter (CDOM) and microbial pigments affect the light intensity and its spectral distribution at each water layer thus influencing the living conditions for differently pigmented phototrophic microorganisms and determining the composition of microbial community.

Quantification of two forms of green sulfur bacteria in their natural habitat using bacteriochlorophyll fluorescence spectra

Detection of phototropic organisms in their natural habitat using optical instruments operating under water is urgently needed for many tasks of ecological monitoring. While fluorescence methods are widely applied nowadays to detect and characterize phytoplankton communities, the techniques for detection and recognition of anoxygenic phototrophs are considered challenging. Differentiation of the forms of anoxygenic green sulfur bacteria in natural water using spectral techniques remains problematic. Green sulfur bacteria could be found in two forms, green-colored (containing BChl d in pigment compound) and brown-colored (containing BChl e), have the special ecological niche in such reservoirs. Separate determination of these microorganisms by spectral methods is complicated because of similarity of spectral characteristics of their pigments. We describe the novel technique of quantification of two forms of green sulfur bacteria directly in water using bacteriochlorophyll fluorescence without pigment extraction. This technique is noninvasive and could be applied in remote mode in the water bodies with restricted water circulation to determine simultaneously concentrations of two forms of green sulfur bacteria in their natural habitat.