Insight into the genome and brackish water adaptation strategies of toxic and bloom-forming Baltic Sea Dolichospermum sp. UHCC 0315.
ABSTRACT: The Baltic Sea is a shallow basin of brackish water in which the spatial salinity gradient is one of the most important factors contributing to species distribution. The Baltic Sea is infamous for its annual cyanobacterial blooms comprised of Nodularia spumigena, Aphanizomenon spp., and Dolichospermum spp. that cause harm, especially for recreational users. To broaden our knowledge of the cyanobacterial adaptation strategies for brackish water environments, we sequenced the entire genome of Dolichospermum sp. UHCC 0315, a species occurring not only in freshwater environments but also in brackish water. Comparative genomics analyses revealed a close association with Dolichospermum sp. UHCC 0090 isolated from a lake in Finland. The genome closure of Dolichospermum sp. UHCC 0315 unraveled a mixture of two subtypes in the original culture, and subtypes exhibited distinct buoyancy phenotypes. Salinity less than 3 g L-1 NaCl enabled proper growth of Dolichospermum sp. UHCC 0315, whereas growth was arrested at moderate salinity (6 g L-1 NaCl). The concentrations of toxins, microcystins, increased at moderate salinity, whereas RNA sequencing data implied that Dolichospermum remodeled its primary metabolism in unfavorable high salinity. Based on our results, the predicted salinity decrease in the Baltic Sea may favor toxic blooms of Dolichospermum spp.
Project description:Salinity is an important abiotic factor controlling the distribution and abundance of Nodularia spumigena, the dominating diazotrophic and toxic phototroph, in the brackish water cyanobacterial blooms of the Baltic Sea. To expand the available genomic information for brackish water cyanobacteria, we sequenced the isolate Nodularia spumigena UHCC 0039 using an Illumina-SMRT hybrid sequencing approach, revealing a chromosome of 5,294,286 base pairs (bp) and a single plasmid of 92,326 bp. Comparative genomics in Nostocales showed pronounced genetic similarity among Nodularia spumigena strains evidencing their short evolutionary history. The studied Baltic Sea strains share similar sets of CRISPR-Cas cassettes and a higher number of insertion sequence (IS) elements compared to Nodularia spumigena CENA596 isolated from a shrimp production pond in Brazil. Nodularia spumigena UHCC 0039 proliferated similarly at three tested salinities, whereas the lack of salt inhibited its growth and triggered transcriptome remodeling, including the up-regulation of five sigma factors and the down-regulation of two other sigma factors, one of which is specific for strain UHCC 0039. Down-regulated genes additionally included a large genetic region for the synthesis of two yet unidentified natural products. Our results indicate a remarkable plasticity of the Nodularia salinity acclimation, and thus salinity strongly impacts the intensity and distribution of cyanobacterial blooms in the Baltic Sea.
Project description:The recent proliferation of harmful cyanobacterial blooms (cyanoHABs) in the Baltic and other marginal seas poses a severe threat for the health of infested ecosystems as e.g. the massive export and decay of cyanobacterial biomass facilitates the spread of bottom water hypoxia. There is evidence that cyanoHABs occurred repeatedly in the Baltic Sea but knowledge of their spatiotemporal distribution and the cyanobacteria that contributed to them is limited. In this study, we examined representatives of the major bloom-forming heterocystous cyanobacteria (i.e. Aphanizomenon, Dolichospermum (formerly Anabaena) and Nodularia) to establish lipid fingerprints that allow tracking these environmentally important diazotrophs in the modern and past Baltic Sea. The distribution of normal and mid-chain branched alkanes, fatty acid methyl esters, bacteriohopanepolyols and heterocyst glycolipids permitted a clear chemotaxonomic separation of the different heterocystous cyanobacteria but also indicated a close phylogenetic relationship between representatives of the genera Aphanizomenon and Dolichospermum. Compared to the discontinuous nature of phytoplankton surveys studies, the distinct lipid profiles reported here will allow obtaining detailed spatiotemporal information on the frequency and intensity of Baltic Sea cyanoHABs as well as their community composition using the time-integrated biomarker signatures recorded in surface and subsurface sediments. As heterocystous cyanobacteria of the genera Aphanizomenon, Dolichospermum and Nodularia are generally known to form massive blooms in many brackish as well as lacustrine systems worldwide, the chemotaxonomic markers introduced in this study may allow investigating cyanoHABs in a great variety of contemporary environments from polar to tropical latitudes.
Project description:Frequencies and biomass of Baltic Sea cyanobacterial blooms are expected to be higher in future climate conditions, but also of longer duration as a result of increased sea surface temperature. Concurrently, climate predictions indicate a reduced salinity in the Baltic Sea. These climate-driven changes are expected to alter not solely the phytoplankton community but also the role of microbial communities for nutrient remineralization. Here, we present the response of summer plankton communities (filamentous cyanobacteria, picocyanobacteria, and heterotrophic bacteria) to the interplay of increasing temperature (from 16 to 18°C and 20°C) and reduced salinity (from salinity 6.9 to 5.9) in the Baltic Proper (NW Gotland Sea) using a microcosm approach. Warmer temperatures led to an earlier peak of cyanobacterial biomass, while yields were reduced. These conditions caused a decrease of nitrogen-fixers (Dolichospermum sp.) biomass, while non nitrogen-fixers (Pseudanabaena sp.) increased. Salinity reduction did not affect cyanobacterial growth nor community composition. Among heterotrophic bacteria, Actinobacteria showed preference for high temperature, while Gammaproteobacteria thrived at in situ temperature. Heterotrophic bacteria community changed drastically at lower salinity and resembled communities at high temperature. Picocyanobacteria and heterotrophic bacterial biomass had a pronounced increase associated with the decay of filamentous cyanobacteria. This suggests that shifts in community composition of heterotrophic bacteria are influenced both directly by abiotic factors (temperature and salinity) and potentially indirectly by cyanobacteria. Our findings suggest that at warmer temperature, lower yield of photosynthetic cyanobacteria combined with lower proportion of nitrogen-fixers in the community could result in lower carbon export to the marine food web with consequences for the decomposer community of heterotrophic bacteria.
Project description:Anabaena is a filamentous, N(2)-fixing, and morphologically diverse genus of cyanobacteria found in freshwater and brackish water environments worldwide. It contributes to the formation of toxic blooms in freshwater bodies through the production of a range of hepatotoxins or neurotoxins. In the Baltic Sea, Anabaena spp. form late summer blooms, together with Nodularia spumigena and Aphanizomenon flos-aquae. It has been long suspected that Baltic Sea Anabaena may produce microcystins. The presence of microcystins has been reported for the coastal regions of the Baltic proper, and a recent report also indicated the presence of the toxin in the open Gulf of Finland. However, at present there is no direct evidence linking Baltic Sea Anabaena spp. to microcystin production. Here we report on the isolation of microcystin-producing strains of the genus Anabaena in the open Gulf of Finland. The dominant microcystin variants produced by these strains included the highly toxic MCYST-LR as well as [d-Asp(3)]MCYST-LR, [d-Asp(3)]MCYST-HtyR, MCYST-HtyR, [d-Asp(3),Dha(7)]MCYST-HtyR, and [Dha(7)]MCYST-HtyR variants. Toxic strains were isolated from the coastal Gulf of Finland as well as from the easternmost open-sea sampling station, where there were lower salinities than at other stations. This result suggests that lower salinity may favor microcystin-producing Anabaena strains. Furthermore, we sequenced 16S rRNA genes and found evidence for pronounced genetic heterogeneity of the microcystin-producing Anabaena strains. Future studies should take into account the potential presence of microcystin-producing Anabaena sp. in the Gulf of Finland.
Project description:Nodularia spumigena is a nitrogen-fixing cyanobacterium that forms toxic blooms in the Baltic Sea each summer and the availability of phosphorous is an important factor limiting the formation of these blooms. Bioinformatic analysis identified a phosphonate degrading (phn) gene cluster in the genome of N. spumigena suggesting that this bacterium may use phosphonates as a phosphorus source. Our results show that strains of N. spumigena could grow in medium containing methylphosphonic acid (MPn) as the sole source of phosphorous and released methane when growing in medium containing MPn. We analyzed the total transcriptomes of N. spumigena UHCC 0039 grown using MPn and compared them with cultures growing in Pi-replete medium. The phnJ, phosphonate lyase gene, was upregulated when MPn was the sole source of phosphorus, suggesting that the expression of this gene could be used to indicate the presence of bioavailable phosphonates. Otherwise, growth on MPn resulted in only a minor reconstruction of the transcriptome and enabled good growth. However, N. spumigena strains were not able to utilize any of the anthropogenic phosphonates tested. The phosphonate utilizing pathway may offer N. spumigena a competitive advantage in the Pi-limited cyanobacterial blooms of the Baltic Sea.
Project description:Cyanobacteria and associated heterotrophic bacteria hold key roles in carbon as well as nitrogen fixation and cycling in the Baltic Sea due to massive cyanobacterial blooms each summer. The species specific activities of different cyanobacterial species as well as the N- and C-exchange of associated heterotrophic bacteria in these processes, however, are widely unknown. Within one time series experiment we tested the cycling in a natural, late stage cyanobacterial bloom by adding 13C bi-carbonate and 15N2, and performed sampling after 10 min, 30 min, 1 h, 6 h and 24 h in order to determine the fixing species as well as the fate of the fixed carbon and nitrogen in the associations. Uptake of 15N and 13C isotopes by the most abundant cyanobacterial species as well as the most abundant associated heterotrophic bacterial groups was then analysed by NanoSIMS. Overall, the filamentous, heterocystous species Dolichospermum sp., Nodularia sp., and Aphanizomenon sp. revealed no or erratic uptake of carbon and nitrogen, indicating mostly inactive cells. In contrary, non-heterocystous Pseudanabaena sp. dominated the nitrogen and carbon fixation, with uptake rates up to 1.49 ± 0.47 nmol N h-1 l-1 and 2.55 ± 0.91 nmol C h-1 l-1. Associated heterotrophic bacteria dominated the subsequent nitrogen remineralization with uptake rates up to 1.2 ± 1.93 fmol N h-1 cell -1, but were also indicative for fixation of di-nitrogen.
Project description:Colony-forming cyanobacteria of the genus Aphanizomenon form massive blooms in the brackish water of the Baltic Sea during the warmest summer months. There have been recent suggestions claiming that the Baltic Sea Aphanizomenon species may be different from Aphanizomenon flos-aquae found in lakes. In this study, we examined variability in the morphology and 16S-23S rRNA internal transcribed spacer (ITS) sequences of A. flos-aquae populations along a salinity gradient from a string of lakes to a fjord-like extension of the Baltic Sea to the open Baltic Sea. Morphological differences among the populations were negligible. We found that the Baltic Sea was dominated (25 out of 27 sequences) by one ITS1-S (shorter band of ITS 1 [ITS1]) genotype, which also was found in the lakes. The lake populations of A. flos-aquae tended to be genetically more diverse than the Baltic Sea populations. Since the lake ITS1-S genotypes of A. flos-aquae are continuously introduced to the Baltic Sea via inflowing waters, it seems that only one ITS1 genotype is able to persist in the Baltic Sea populations. The results suggest that one of the ITS1-S genotypes found in the lakes is better adapted to the conditions of the Baltic Sea and that natural selection removes most of the lake genotypes from the Baltic Sea A. flos-aquae populations.
Project description:In the Baltic Sea, diazotrophic cyanobacteria have been present for thousands of years, over the whole brackish water phase of the ecosystem. However, our knowledge about the species composition of the cyanobacterial community is limited to the last several decades. In the current study, the presence of species-specific chemical and genetic markers in deep sediments were analyzed to increase the existing knowledge on the history of toxic Nodularia spumigena blooms in the Baltic Sea. As chemical markers, three cyclic nonribosomal peptides were applied: the hepatotoxic nodularin, which in the sea was detected solely in N. spumigena, and two anabaenopeptins (AP827 and AP883a) characteristic of two different chemotypes of this species. From the same sediment samples, DNA was isolated and the gene involved in biosynthesis of nodularin, as well as the phycocyanin intergenic spacer region (PC-IGS), were amplified. The results of chemical and genetic analyses proved for the first time the thousands-year presence of toxic N. spumigena in the Baltic Sea. They also indicated that through all this time, the same two sub-populations of the species co-existed.
Project description:Future climate scenarios in the Baltic Sea project an increase of cyanobacterial bloom frequency and duration, attributed to eutrophication and climate change. Some cyanobacteria can be toxic and their impact on ecosystem services is relevant for a sustainable sea. Yet, there is limited understanding of the mechanisms regulating cyanobacterial diversity and biogeography. Here we unravel successional patterns and changes in cyanobacterial community structure using a 2-year monthly time- series during the productive season in a 100 km coastal-offshore transect using microscopy and high-throughput sequencing of 16S rRNA gene fragments. A total of 565 cyanobacterial OTUs were found, of which 231 where filamentous/colonial and 334 picocyanobacterial. Spatial differences in community structure between coastal and offshore waters were minor. An "epidemic population structure" (dominance of asingle cluster) was found for Aphanizomenon/Dolichospermum within the filamentous/colonial cyanobacterial community. In summer, this clusters imultaneously occurred with opportunistic clusters/OTUs, e.g., Nodularia spumigena and Pseudanabaena. Picocyanobacteria, Synechococcus/Cyanobium, formeda consistent but highly diverse group. Overall, the potential drivers structuring summer cyanobacterial communities were temperature and salinity. However, the different responses to environmental factors among and within genera suggest high niche specificity for individual OTUs. The recruitment and occurrence of potentially toxic filamentous/colonial clusters was likely related to disturbance such as mixing events and short-term shifts in salinity, and not solely dependent on increasing temperature and nitrogen-limiting conditions. Nutrients did not explain further the changes in cyanobacterial community composition. Novel occurrence patterns were identified as a strong seasonal succession revealing a tight coupling between the emergence of opportunistic picocynobacteria and the bloom offilamentous/colonialclusters. These findings highlight that if environmental conditions can partially explain the presence of opportunistic picocyanobacteria, microbial and trophic interactions with filamentous/colonial cyanobacteria should also be considered as potential shaping factors for single-celled communities. Regional climate change scenarios in the Baltic Sea predict environmental shifts leading to higher temperature and lower salinity; conditions identified here as favorable for opportunistic filamentous/colonial cyanobacteria. Altogether, the diversity and complexity of cyanobacterial communities reported here is far greater than previously known, emphasizing the importance of microbial interactions between filamentous and picocyanobacteria in the context of environmental disturbances.
Project description:Increasing sea surface temperatures (SST) and blooms of lipid-poor, filamentous cyanobacteria can change mesozooplankton metabolism and foraging strategies in marine systems. Lipid shortage and imbalanced diet may challenge the build-up of energy pools of lipids and proteins, and access to essential fatty acids (FAs) and amino acids (AAs) by copepods. The impact of cyanobacterial blooms on individual energy pools was assessed for key species temperate Temora longicornis and boreal Pseudo-/Paracalanus spp. that dominated field mesozooplankton communities isolated by seasonal stratification in the central Baltic Sea during the hot and the cold summer. We looked at (a) total lipid and protein levels, (b) FA trophic markers and AA composition, and (c) compound-specific stable carbon isotopes (?13C) in bulk mesozooplankton and in a subset of parameters in particulate organic matter. Despite lipid-poor cyanobacterial blooms, the key species were largely able to cover both energy pools, yet a tendency of lipid reduction was observed in surface animals. Omni- and carnivory feeding modes, FA trophic makers, and ?13C patterns in essential compounds emphasized that cyanobacterial FAs and AAs have been incorporated into mesozooplankton mainly via feeding on mixo- and heterotrophic (dino-) flagellates and detrital complexes during summer. Foraging for essential highly unsaturated FAs from (dino-) flagellates may have caused night migration of Pseudo-/Paracalanus spp. from the deep subhalocline waters into the upper waters. Only in the hot summer (SST>19.0°C) was T. longicornis submerged in the colder subthermocline water (~4°C). Thus, the continuous warming trend and simultaneous feeding can eventually lead to competition on the preferred diet by key copepod species below the thermocline in stratified systems. A comparison of ?13C patterns of essential AAs in surface mesozooplankton across sub-basins of low and high cyanobacterial biomasses revealed the potential of ?13C-AA isoscapes for studies of commercial fish feeding trails across the Baltic Sea food webs.