E. G. Sorokovikova

Presence and genetic diversity of microcystin-producing cyanobacteria (Anabaena and Microcystis) in Lake Kotokel (Russia, Lake Baikal Region)

A survey was conducted for the presence of cyanobacteria toxins in Lake Kotokel due to a few cases of Haff disease registered in 2008–2009 caused by consumption of fish from Lake Kotokel, and wildlife mortality including large fish kill. The aims of this study were to determine what cyanotoxins (if any) were present in the lake, to describe phytoplankton composition including morphology, density, and species diversity of cyanobacteria, as well as to evaluate the trophic state of the lake. Samples were collected from both nearshore and central sites in August of 2009. Aphanocapsa holsatica dominated the phytoplankton. The presence of toxigenic genotypes of Microcystis spp. and Anabaena lemmermannii was detected by sequencing of PCR-amplified aminotransferase domain of microcystin synthetase gene. LR, RR, and YR microcystin (MC) variants were detected with liquid chromatography-UV mass spectrometry. The data do not shed light on the etiology of Haff disease in Lake Kotokel region, nevertheless taking into account the recreational importance of the lake and its direct connection to Lake Baikal, a necessity to monitor cyanobacteria in these water bodies is evident. This is the first report on simultaneous detection of MC-producing genotypes and MCs in the Lake Baikal region.

Identification of toxigenic Cyanobacteria of the genus Microcystis in the Curonian Lagoon (Baltic Sea)

In 2002–2008, seasonal (April–November) monitoring of the phytoplankton in the Russian part of the Curonian Lagoon at five fixed sites was performed. A total of 91 Cyanobacteria, 100 Bacillariophyta, 280 Chlorophyta, 21 Cryptophyta, and 24 Dinophyta species were found. Six potentially toxic species of cyanobacteria: Aphanizomenon flos-aquae, Anabaena sp., Microcystis aeruginosa, M. viridis, M. wesenbergii, and Planktothrix agardhii dominated the phytoplankton biomass and caused water blooms. The seasonal average phytoplankton biomass ranged from 30 to 137 g/m3. The cyanobacteria’s biomass varied from 10 to 113 g/m3 forming 30–82% of the total with a mean of 50%. With the aid of genetic markers (microcystin (mcy) and nodularin synthetases), six variants of the microcystin-producing gene mcyE from the genus Microcystis were identified. Due to the intensive and lengthy blooms of potentially toxic and toxigenic cyanobacteria, the environmental conditions in the Curonian Lagoon appear unfavorable. The water should be monitored for cyanotoxins with analytical methods in order to determine if the area is safe for recreational use.

Diversity of the major capsid genes (g23) of T4-like bacteriophages in the eutrophic Lake Kotokel in East Siberia, Russia.

Numerous studies revealed high diversity of T4-like bacteriophages in various environments, but so far, little is known about T4-like virus diversity in freshwater bodies, particularly in eutrophic lakes. The present study was aimed at elucidating molecular diversity of T4-like bacteriophages in eutrophic Lake Kotokel located near Lake Baikal by partial sequencing of the major capsid genes (g23) of T4-like bacteriophages. The majority of g23 fragments from Lake Kotokel were most similar to those from freshwater lakes and paddy fields. Despite the proximity and direct water connection between Lake Kotokel and Lake Baikal, g23 sequence assemblages from two lakes were different. UniFrac analysis showed that uncultured T4-like viruses from Lake Kotokel tended to cluster with those from the distant lake of the same trophic status. This fact suggested that the trophic conditions affected the formation of viral populations, particularly of T4-like viruses, in freshwater environments.

Identification of Cyanobacterial Producers of Shellfish Paralytic Toxins in Lake Baikal and Reservoirs of the Angara River

The goal of the present work was to search for PST� producing cyanobacteria in Lake Baikal and reservoirs of the Angara River using a marker for the polyketide synthase gene stxA and to detect saxitoxin and its ana� logues by enzymelinked immunosorbent assay (ELISA) and mass spectrometry. Samples from Lake Baikal were collected in August 2010 in the coastal zone of the Barguzin and Kurkut Bays (Maloe More Strait). Samples were taken from the Irkutsk reservoir (near Patrony village), Bratsk res� ervoir (near NovoDolonovo village), and UstIlimsk reservoir (near Zheleznodorozhnik settlement) in July 2010. Plankton was sampled with a Ruttner sampler from 1�m depth and with an Apstein plankton net by filtration of the upper 1.5�m water layer. The bottle samples were used for qualitative and quantitative assessment of cyanobacteria as described previously (5). Total DNA was extracted from net samples fixed with 70% ethanol using the DNAsorb kit (InterLab� Servis, Russia). The

First detection of benthic cyanobacteria in Lake Baikal producing paralytic shellfish toxins

Cyanobacteria were screened from the surface of diseased sponges, stone and bedrock in Lake Baikal for the presence of saxitoxin using enzyme-linked immunosorbent assay. In sequel, eight paralytic shellfish toxin (PST) variants were identified using a MALDI mass spectrometry. Microscopic examination found that Tolypothrix distorta dominated in the biofouling samples. PCR and sequencing detected sxtA gene involved in saxitoxin biosynthesis, thereby providing evidence of the PST producing potential of Baikal cyanobacterial communities inhabiting different substrates.

Diversity of cyanobacterial species and phylotypes in biofilms from the littoral zone of Lake Baikal.

The majority of naturally occurring biofilms contain numerous microorganisms that have not yet been cultured. Additionally, there is little information available regarding the genetic structure and species diversity of these communities. Therefore, we characterised the species diversity, structure and metagenome of biofilms grown on stones and steel plates in the littoral zone of Lake Baikal (East Siberia, Russia) by applying three different approaches. First, light microscopy enabled identification of the species diversity of biofilm-forming cyanobacteria on different substrates with the dominance of Rivularia rufescens, Tolypothrix limbata, Chamaesiphon fuscus, Ch. subglobosus, and Heteroleibleinia pusilla. Additionally, scanning electron microscopy was used to show the spatial structure of biofilms. Finally, sequence analysis of 30,660 16S rRNA clones indicated a high diversity within the biofilm communities, with the majority of the microbes being closely related to Cyanobacteria (8–46% sequences), Proteobacteria (14–43%), and Bacteroidetes (10–41%). Rivularia sp., Pseudanabaena sp., and Chamaesiphon spp. were the dominant cyanobacterial phylotypes.

Identification of two cyanobacterial strains isolated from the Kotel’nikovskii hot spring of the Baikal rift

Two cyanobacterial strains, Pseudanabaena sp. 0411 and Synechococcus sp. 0431, were isolated from a sample collected in the Kotel’nikovskii hot spring of the Baikal rift. According to the results of light and transmission electron microscopy, as well as of the phylogenetic analysis of the 16S rRNA gene, these cyanobacteria were classified as Pseudanabaena sp. nov. and Synechococcus bigranulatus Skuja. The constructed phylogenetic tree shows that the studied strains are positioned in the clades of cyanobacteria isolated from hydrothermal vents of Asia and New Zealand, separately from marine and freshwater members of these genera, including those isolated from Lake Baikal.

Saxitoxin-Producing cyanobacteria in Lake Baikal

Cyanobacteria containing neurotoxic saxitoxin synthesis genes were found in the coastal zone of Lake Baikal near the village of Turka for the first time. Phylogenetic analysis showed that the sequences of saxitoxin synthesis genes belong to the genus Anabaena Bory. Saxitoxin concentration in the water according to ELISA was 1.93 ± 0.64 mg/L. The genetic and taxonomic composition of the bacterial community of the central part of Lake Baikal was characterized using 16S rRNA gene pyrosequencing. It was established that the phylum Cyanobacteria dominated in the composition of summer bacterioplankton in both littoral and pelagic zones of the lake, but higher species diversity was found in the plankton of littoral zone.

Abundance, morphological diversity, and spatial distribution of autotrophic picoplankton in Lake Hovsgol (Mongolia)

Hovsgol is the largest deep lake in Central Asia located in the south-west of the Baikal Rift zone. Until now there has been only one study on the autotrophic picoplankton in Lake Hovsgol. The present work summarizes results of picoplankton studies obtained with light, epifluorescence and scanning electron microscopy. Microscopic analysis showed the presence of picoplankton cyanobacteria and algae in Lake Hovsgol. Autotrophic picoplankton of the lake is characterized by morphotype diversity of Synechococcus species, Synechocystis-like aggregates, and picocyanobacteria cells of small sizes. Picocyanobacteria are dominated by Synechococcus species with single, rarely double, coccoid cells. There is also a picoplankton form of the diatom Cyclotella ocellata reaches 4–13 percent of the total species abundance. Three picocyanobacterial cultures belonging to the genus Synechococcus and exhibiting distinct morphological and ultrastructural characteristics were isolated from the plankton samples. Chlorophyll a content in the pelagic zone of Lake Hovsgol in August was 0.182–0.374 μ g l− 1. In February, the abundance of picoplankton cyanobacteria in the surface layer, which make up the basic autotrophic picoplankton mass, was 2.78 × 10 5 cells ml −1 , in March 1.36 × 10 5 cells ml −1 , and in July 6.06 × 10 4 cells ml −1 . In August, picocyanobacteria abundance fell to 3.42 × 10 4 cells ml −1 in the 0 to 10 m layer and 7.52 × 10 4 cells ml −1 in the 0 to 50 m layer. Quantitative variations in autotrophic picoplankton development throughout the lake are negligible. The greatest abundance of picocyanobacteria is recorded in the hypolimnion, 25–50 m deep.

Identification of toxic Cyanobacteria in Lake Baikal

Cyanobacteria of the genera Anabaena and Microcystis, containing genes for the synthesis of-microcystins (hepatotoxic cyanotoxins) were found for the first time in the coastal zone of Lake Baikal near-the village of Turka, where a tourism and recreational complex were constructed. According to the enzyme-immunoassay, microcystin concentration in water was 0.17 ± 0.01 µg/L. Using 16S rRNA gene pyrosequencing, we found 3936 sequences in the eubacterial community of central basin of Lake Baikal. The summer bacterioplankton in both littoral and pelagic areas of the lake was dominated by the phylum Cyanobacteria, whereas a higher diversity of cyanobacteria was recorded in the plankton of the littoral zone. Moreover, the-potentially toxic Anabaena and Microcystis were detected in this area.