D. Yu. Rogozin

Microbial Processes of the Carbon and Sulfur Cycles in Lake Shira (Khakasia)

Microbiological and biogeochemical studies of the meromictic saline Lake Shira (Khakasia) were conducted. In the upper part of the hydrogen-sulfide zone, at a depth of 13.5–14 m, there was a pale pink layer of water due to the development of purple bacteria (6 × 105 cells/ml), which were assigned by their morphological and spectral characteristics toLamprocystis purpurea (formerly Amoebobacter purpureus). In August, the production of organic matter (OM) in Lake Shira was estimated to be 943 mg C/(m2day). The contribution of anoxygenic photosynthesis was insignificant (about 7% of the total OM production). The share of bacterial chemosynthesis was still less (no more than 2%). In the anaerobic zone, the community of sulfate-reducing bacteria played a decisive role in the terminal decomposition of OM. The maximal rates of sulfate reduction were observed in the near-bottom water (114 μg S/(l day)) and in the surface layer of bottom sediments (901 μg S/(dm3 day)). The daily expenditure of Corg for sulfate reduction was 73% of Corg formed daily in the processes of oxygenic and anoxygenic photosynthesis and bacterial chemosynthesis. The profile of methane distribution in the water column and bottom sediments was typical of meromictic reservoirs. The methane content in the water column increased beginning with the thermocline (7–8 m) and reached maximum values in the near-bottom water (17 μl/l). In bottom sediments, the greatest methane concentrations (57 μl/l) were observed in the surface layer (0–3 cm). The integral rate of methane formation in the water column and bottom sediments was almost an order of magnitude higher than the rate of its oxidation by aerobic and anaerobic methanotrophic microorganisms.

[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.

Microbial community of the chemocline of the meromictic Lake Shunet (Khakassia, Russia) during summer stratification.

The spatio-temporal organization of the bacterial community inhabiting the chemocline of the stratified meromictic Lake Shunet (Khakassia, Russia) was investigated from May to September 2005 by means of microscopy, analysis of photosynthetic pigments, and PCR-DGGE with subsequent 16S rDNA analysis. The samples were collected with a multisyringe stratification sampler, sampling being performed every 5 cm. It was demonstrated that, during the period of investigation, there were no large changes in the bacterial community of the chlemocline, at least among the detected forms. During the whole period of study, purple sulfur bacteria related to Lamprocystis purpurea (Chromatiaceae) were predominant in the chemocline. Beneath the layer of purple bacteria, green sulfur bacteria were revealed that were phylogenetically distant from strain ShNPel02, which was previously isolated from this lake. Development of phytoflagellates of the genus Cryptomonas was observed in the upper zone of the chemocline. In the chemocline of Lake Shunet, the numbers of picoplankton cyanobacteria of the genus Synechococcus increased from May to September. It was demonstrated that the application of universal bacterial primers for DGGE resulted in the same qualitative distributional pattern of predominant species as microscopic studies.

Microbiological and isotopic-geochemical investigations of meromictic lakes in Khakasia in winter

Microbiological and isotopic-geochemical investigations of the brackish meromictic lakes Shira and Shunet were performed in the steppe region of Khakasia in winter. Measurements made with a submersed sensor demonstrated that one-meter ice transmits light in a quantity sufficient for oxygenic and anoxygenic photosynthesis. As in the summer season, in the community of phototrophic bacteria found in Lake Shira, the purple sulfur bacteria Amoebobacter purpureus dominated, whereas, in Lake Shunet, the green sulfur bacteria Pelodictyon luteolum were predominant. Photosynthetic production, measured using the radioisotopic method, was several times lower than that in summer. The rates of sulfate reduction and production and oxidation of methane in the water column and bottom sediments were also lower than those recorded in summer. The process of anaerobic methane oxidation in the sediments was an exception, being more intense in winter than in summer. The data from radioisotopic measurements of the rates of microbial processes correlate well with the results of determination of the isotopic composition of organic and mineral carbon (δ13C) and hydrogen sulfide and sulfate (δ34S) and suggest considerable seasonal variations in the activity of the microbial community in the water bodies investigated.

Sulfate reduction and methanogenesis in the Shira and Shunet meromictic lakes (Khakasia, Russia)

The biogeochemical and molecular biological study of the chemocline and sediments of saline meromictic lakes Shira and Shunet (Khakasia, Russia) was performed. A marked increase in the rates of sulfate reduction and methanogenesis was revealed at the medium depths of the chemocline. The rates of these processes in the bottom sediments decreased with depth. The numbers of the members of domains Bacteria, Archaea, and of sulfate-reducing bacteria (SRB) were determined by fluorescence in situ hybridization with rRNA specific oligonucleotide probes labeled with horseradish peroxidase and subsequent tyramide signal amplification. In the chemocline, both the total microbial numbers and those of Bacteria were shown to increase with depth. The archaea and SRB were present in almost equal numbers. In the lake sediments, a drastic decrease in microbial numbers with depth was revealed. SRB were found to prevail in the upper sediment layer and archaea in the lower one. This finding correlated with the measured rates of sulfate reduction and methanogenesis.

Ecology of purple sulfur bacteria in the highly stratified meromictic Lake Shunet (Siberia, Khakassia) in 2002–2009

Phototrophic sulfur bacteria form dense accumulations in the chemocline zones of stratified lakes where light reaches the sulfide-containing layers of water. Many works are dedicated to the ecophysiology of these microorganisms in meromictic lakes. However, the role of these microorganisms in the trophic network of these ecosystems, the ways of biomass utilization, and the contribution to the turnover of biogenic elements have so far been insufficiently understood. This work deals with the analysis of many years’ seasonal dynamics of the biomass of purple sulfur bacteria and the physicochemical conditions of their environment in Lake Shunet (Siberia, Khakassia, Russia), unraveling the causes of their anomalous development in the chemocline of this lake, as well as the comparative analysis of such type of ecosystems. Lake Shunet is characterized by markedly pronounced stratification and the high density of purple sulfur bacteria (PSB) in the chemocline, which is comparable to that of Lake Mahoney (Canada) where the number of PSB is the greatest among those known in the world. It was shown that, in the period 2002–2009, the total amount of bacterio-chlorophyll a in the water column of Lake Shunet increased and did not correlate with the seasonal variations in temperature and illumination in the chemocline. It was established that PSB cells in the purple layer experienced the effect of self-shading. The sedimentation rate of purple sulfur bacteria in Lake Shunet was low due to the pronounced density gradient in the chemocline zone. Thus, the high number of PSB in the chemocline was due to the combination of strong illumination, a high sulfide concentration, and a high water density gradient, which was responsible for stable stratification and contributed to the accumulation of the cells in a narrow layer. The data obtained could be useful for the paleoreconstruction of climatically deter-mined changes in the level of the lake and its periods of meromixis by the presence of carotenoids and bacte-riochlorophylls in the bottom sediments.

Psychrophilic and Psychrotolerant Heterotrophic Microorganisms of Middle Siberian Karst Cavities

The natural microflora of Middle Siberian karst cavities, which comprises psychrotolerant bacteria and fungi capable of growing at 3–15 and 3...+28°C, respectively, has been studied. Bacteria are ubiquitous in caves, their count varying from 103 to 107 cells/g ground. The bacteria have been identified as Pseudomonas, Arthrobacter, Bacillus, and coryneform bacteria. Fungi have been found in places exposed to increased anthropogenic impact, their count being as large as 106 to 107 cells/g ground. Mucor, Penicillium, and Chrysosporium were dominant fungal genera.

Carotenoids of phototrophic organisms in bottom sediments of meromictic Lake Shira (Siberia, Russia) as an indicator of past stratification.

228 Photosynthetic pigments (chlorophylls and caro tenoids) buried in bottom sediments of water bodies are traces of phototrophic organisms that dwelled in these water bodies in the past [1–3]. In the course of sedimentation, the pigments come to bottom sedi ments from the photic zone together with dead cells, pallets of zooplankton, plant remnants, etc. and remain preserved during hundreds and thousands of years due to the absence of light and oxygen, and low temperatures [1]. Therefore, the profile and quantita tive composition of the pigments in the bottom sedi ment layers may serve as indicators of the species com position of the biomass and the amount of pho totrophic organism production in the corresponding periods of the history of the given water body. In turn, changes in the species composition and production characteristics are indicators of changes in the living conditions of phototrophic organisms.

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).

[Dynamics of Purple Sulfur Bacteria in a Meromictic Saline Lake Shunet (Khakassia, Siberia) in 2007-2013].

According to the results of seasonal monitoring, in 2007–2013 purple sulfur bacteria morphologically similar to Thiocapsa sp. Shira_1 (AJ633676 in EMBL/GenBank) predominated in the anoxygenic phototrophic community of the water column of the meromictic Lake Shira (Khakassia, Siberia). No pronounced seasonal periodicity in the total cell number in the water column was revealed during the period of observation. In some years cell number during the period when the lake was covered with ice was reliably higher than in summer. The absence of seasonal periodicity was probably due to the low amplitude of seasonal variations in temperature and illumination in the redox zone, resulting from its relatively deep location (12–16 m). The year-to-year dynamics was characterized by a reliable decrease of the total cell number in 2009–2010 and maxima in 2007 and 2011–2012. Canonical correlation analysis revealed that water temperature in the redox zone was the best predictor of the PSB abundance in Lake Shira. Water temperature, in turn, depended on the depth of mixing of the water column. Intense mixing in 2009–2011 was probably responsible for decreased PSB abundance in the lake. On the other hand, the absence of deep winter mixing, resulting in stable conditions in the chemocline, favored the preservation of relatively high PSB biomass. Prediction of circulation depth, which depends mainly on the weather conditions and dynamics of the water level, is required for prediction of PSB abundance in Lake Shira. These results may be useful for paleolimnological reconstructions of the history of the lake based on the remnants of purple sulfur bacteria in bottom sediments.