Cultures Of Synechococcus Research Articles

Production and utilization of pseudocobalamin in marine Synechococcus cultures and communities.

Cobalamin influences marine microbial communities because an exogenous source is required by most eukaryotic phytoplankton, and demand can exceed supply. Pseudocobalamin is a cobalamin analogue produced and used by most cyanobacteria but is not directly available to eukaryotic phytoplankton. Some microbes can remodel pseudocobalamin into cobalamin, but a scarcity of pseudocobalamin measurements impedes our ability to evaluate its importance for marine cobalamin production. Here, we perform simultaneous measurements of pseudocobalamin and methionine synthase (MetH), the key protein that uses it as a co-factor, in Synechococcus cultures and communities. In Synechococcus sp. WH8102, pseudocobalamin quota decreases in low temperature (17°C) and low nitrogen to phosphorus ratio, while MetH did not. Pseudocobalamin and MetH quotas were influenced by culture methods and growth phase. Despite the variability present in cultures, we found a comparably consistent quota of 300 ± 100 pseudocobalamin molecules per cyanobacterial cell in the Northwest Atlantic Ocean, suggesting that cyanobacterial cell counts may be sufficient to estimate pseudocobalamin inventories in this region. This work offers insights into cellular pseudocobalamin metabolism, environmental and physiological conditions that may influence it, and provides environmental measurements to further our understanding of when and how pseudocobalamin can influence marine microbial communities.

Viral infection of an estuarine Synechococcus influences its co-occurring heterotrophic bacterial community in the culture.

Viruses are infectious and abundant in the marine environment. Viral lysis of host cells releases organic matter and nutrients that affect the surrounding microbial community. Synechococcus are important primary producers in the ocean and they are subject to frequent viral infection. In the laboratory, Synechococcus cultures are often associated with bacteria and such a co-existence relationship appears to be important to the growth and stability of Synechococcus. However, we know little about how viral lysis of Synechococcus affects the co-existing bacteria in the culture. This study investigated the influence of viral infection of Synechococcus on co-occurring bacterial community in the culture. We analyzed the community composition, diversity, predicted functions of the bacterial community, and its correlations with fluorescent dissolved organic matter (FDOM) components and nutrients after introducing a cyanophage to the Synechococcus culture. Cyanophage infection altered the bacterial community structure and increased the bacterial diversity and richness. Increased bacterial groups such as Bacteroidetes and Alphaproteobacteria and decreased bacterial groups such as Gammaproteobacteria were observed. Moreover, cyanophage infection reduced bacterial interactions but enhanced correlations between the dominant bacterial taxa and nutrients. Unique FDOM components were observed in the cyanophage-added culture. Fluorescence intensities of FDOM components varied across the cyanophage-infection process. Decreased nitrate and increased ammonium and phosphate in the cyanophage-added culture coupled with the viral progeny production and increased substance transport and metabolism potentials of the bacterial community. Furthermore, increased potentials in methane metabolism and aromatic compound degradation of the bacterial community were observed in the cyanophage-added culture, suggesting that cyanophage infections contribute to the production of methane-related compounds and refractory organic matter in a microcosm like environment. This study has the potential to deepen our understanding of the impact of viral lysis of cyanobacteria on microbial community in the surrounding water.

Bottom-up and top-down controls on Alteromonas macleodii lead to different dissolved organic matter compositions.

The effects of both bottom-up (e.g. substrate) and top-down (e.g. viral lysis) controls on the molecular composition of dissolved organic matter have not been investigated. In this study, we investigated the dissolved organic matter composition of the model bacterium Alteromonas macleodii ATCC 27126 growing on different substrates (glucose, laminarin, extracts from a Synechococcus culture, oligotrophic seawater, and eutrophic seawater), and infected with a lytic phage. The ultra-high resolution mass spectrometry analysis showed that when growing on different substrates Alteromonas macleodii preferred to use reduced, saturated nitrogen-containing molecules (i.e. O4 formula species) and released or preserved oxidized, unsaturated sulfur-containing molecules (i.e. O7 formula species). However, when infected with the lytic phage, Alteromonas macleodii produced organic molecules with higher hydrogen saturation, and more nitrogen- or sulfur-containing molecules. Our results demonstrate that bottom-up (i.e. varying substrates) and top-down (i.e. viral lysis) controls leave different molecular fingerprints in the produced dissolved organic matter.

An Inducible Microbacterium Prophage vB_MoxS-R1 Represents a Novel Lineage of Siphovirus.

Lytic and lysogenic infections are the main strategies used by viruses to interact with microbial hosts. The genetic information of prophages provides insights into the nature of phages and their potential influences on hosts. Here, the siphovirus vB_MoxS-R1 was induced from a Microbacterium strain isolated from an estuarine Synechococcus culture. vB_MoxS-R1 has a high replication capability, with an estimated burst size of 2000 virions per cell. vB_MoxS-R1 represents a novel phage genus-based genomic analysis. Six transcriptional regulator (TR) genes were predicted in the vB_MoxS-R1 genome. Four of these TR genes are involved in stress responses, virulence and amino acid transportation in bacteria, suggesting that they may play roles in regulating the host cell metabolism in response to external environmental changes. A glycerophosphodiester phosphodiesterase gene related to phosphorus acquisition was also identified in the vB_MoxS-R1 genome. The presence of six TR genes and the phosphorus-acquisition gene suggests that prophage vB_MoxS-R1 has the potential to influence survival and adaptation of its host during lysogeny. Possession of four endonuclease genes in the prophage genome suggests that vB_MoxS-R1 is likely involved in DNA recombination or gene conversion and further influences host evolution.

Filter Plating Method for Rendering Picocyanobacteria Cultures Free of Heterotrophic Bacterial Contaminants and Clonal.

Isolates of the marine picocyanobacteria, Prochlorococcus and Synechococcus, are often accompanied by diverse heterotrophic “contaminating” bacteria, which can act as confounding variables in otherwise controlled experiments. Traditional microbiological methods for eliminating contaminants, such as direct streak-plating, are often unsuccessful with this particular group of microorganisms. While they will grow in pour plates, colonies often remain contaminated with heterotrophic bacteria that can migrate through the soft agar. Additionally, axenic clones of picocyanobacteria can be recovered via dilution-to-extinction in liquid medium, but the efficiency of recovery is low, often requiring large numbers of 96-well plates. Here, we detail a simple and effective protocol for rendering cultures of Synechococcus and Prochlorococcus strains free of bacterial contaminants while at the same time yielding clonal isolates. We build on the fact that co-culture with specific heterotrophs—“helper heterotrophs”—is often necessary to grow colonies of picocyanobacteria from single cells in agar. Suspecting that direct physical contact between the helper and the picocyanobacterial cells was not necessary for the “helper effect,” we developed a protocol in which the helper cells are embedded in soft agar pour plates, a filter overlaid on the surface, and a picocyanobacterial culture is diluted and then spotted on top of the filter. With this approach, motile contaminants cannot swim to the colonies, and it is possible to obtain the expected number of colonies from a given input (i.e., a Poisson distribution of colonies with an expected value equal to the input number of cells), thus ensuring clonal colonies. Using this protocol, we rendered three strains of Synechococcus, two strains of Prochlorococcus, and 19 new strains of Synechococcus from coastal seawater clonal and free of heterotrophic bacteria. The simplicity of this approach should expand the repertoire of axenic picocyanobacterial strains available for controlled physiological experiments. It will also enable the study of microdiversity in populations of picocyanobacteria by facilitating large-scale isolation of picocyanobacterial clones from a single source, including direct isolation from natural seawater.

Isolation and Characterization of Cyanobacterial Extracellular Vesicles

Cyanobacteria are a diverse group of photosynthetic, Gram-negative bacteria that play critical roles in global ecosystems and serve as essential biotechnology models. Recent work has demonstrated that both marine and freshwater cyanobacteria produce extracellular vesicles - small membrane-bound structures released from the outer surface of the microbes. While vesicles likely contribute to diverse biological processes, their specific functional roles in cyanobacterial biology remain largely unknown. To encourage and advance research in this area, a detailed protocol is presented for isolating, concentrating, and purifying cyanobacterial extracellular vesicles. The current work discusses methodologies that have successfully isolated vesicles from large cultures of Prochlorococcus, Synechococcus, and Synechocystis. Methods for quantifying and characterizing vesicle samples from these strains are presented. Approaches for isolating vesicles from aquatic field samples are also described. Finally, typical challenges encountered with cyanobacterial vesicle purification, methodological considerations for different downstream applications, and the trade-offs between approaches are also discussed.

Isolation and Characterization of Cyanobacterial Extracellular Vesicles.

Cyanobacteria are a diverse group of photosynthetic, Gram-negative bacteria that play critical roles in global ecosystems and serve as essential biotechnology models. Recent work has demonstrated that both marine and freshwater cyanobacteria produce extracellular vesicles -small membrane-bound structures released from the outer surface of the microbes. While vesicles likely contribute to diverse biological processes, their specific functional roles in cyanobacterial biology remain largely unknown. To encourage and advance research in this area, a detailed protocol is presented for isolating, concentrating, and purifying cyanobacterial extracellular vesicles. The current work discusses methodologies that have successfully isolated vesicles from large cultures of Prochlorococcus, Synechococcus, and Synechocystis. Methods for quantifying and characterizing vesicle samples from these strains are presented. Approaches for isolating vesicles from aquatic field samples are also described. Finally, typical challenges encountered with cyanobacterial vesicle purification, methodological considerations for different downstream applications, and the trade-offs between approaches are also discussed.

Effects of spectral light quality on the growth, productivity, and elemental ratios in differently pigmented marine phytoplankton species

The recent advancement in LED technology led many incubator manufacturers and research labs to switch to these more efficient, yet spectrally restricted, light sources. Potential effects of commercially available broad range “white” light systems on phytoplankton growth, productivity, light absorption spectra, and cellular composition have not yet been characterized but could affect our interpretation of lab-based projections on responses to environmental changes. In this study, we investigated such effects using cultures of Prochlorococcus marinus, Synechococcus sp., and Thalassiosira weissflogii grown under three different commercially available LED lights as well as one fluorescent growth light. Photosynthetically active radiation was equal for each species, while photosynthetically usable radiation differed among the combinations of species and treatments. Growth rate was unaffected across species, yet all species displayed changes in cellular carbon, nitrogen, and chlorophyll a quotas as a direct response to the different light spectra. 14C-based primary productivity was also affected in P. marinus and T. weissflogii. Analysis of pigment ratios and photophysiological data indicated changes in the photoacclimation state between different light environments. The results of this study show that these species undergo changes in underlying cellular metabolism which in turn affect cellular composition while keeping specific growth constant. The data presented here illustrate ecophysiological responses of differently pigmented species when grown under different artificial growth light spectra. These cellular acclimation responses should be considered when designing laboratory-based incubation experiments, especially when comparing responses to specific changes in environmental conditions, or when implementing physiological parameters, derived from laboratory experiments, into numerical models.

Distinct metabolic strategies of the dominant heterotrophic bacterial groups associated with marine Synechococcus

The marine Synechococcus is a major primary producer in the global oceans. It is phylogenetically highly diverse, and its major phylogenetic lineages display clear spatial segregation among different marine environments. Here, we showed that the composition of the associated bacterial communities was related to the geographic origin of the different Synechococcus strains, and it was stable during long-term lab incubation. Of all the Synechococcus cultures investigated, the Rhodobacteraceae had a relatively high abundance and was the core bacterial family of the associated bacterial communities. In contrast, the Flavobacteriaceae were only abundant in the cultures collected from the South China Sea (which is warm and oligotrophic), whereas those of the Alteromonadaceae were abundant in the cultures from the coastal waters off Hong Kong and Xiamen. We also found that the Rhodobacteraceae had more ABC transporters and utilized a wider spectrum of carbon sources than did the Flavobacteriaceae and Alteromonadaceae. Moreover, the Alteromonadaceae had more transporters for importing phosphate and amino acids, but fewer transporters for importing oligosaccharides, polyol, and lipid, than the Flavobacteriaceae. Furthermore, metagenomic analysis demonstrated that bacteria involved in nitrate-ammonification prevailed in all the cultures. These results imply that networks formed by phytoplankton and heterotrophic bacteria might vary across habitats, and that different dominant bacterial groups play different roles in the phycosphere. This study provides new insight into the unique interactive and interdependent bond between phytoplankton and their associated microbiome, which may enhance our understanding of carbon and nutrient cycling in marine environments.

Temperature Responses of Heterotrophic Bacteria in Co-culture With a Red Sea Synechococcus Strain.

Interactions between autotrophic and heterotrophic bacteria are fundamental for marine biogeochemical cycling. How global warming will affect the dynamics of these essential microbial players is not fully understood. The aims of this study were to identify the major groups of heterotrophic bacteria present in a Synechococcus culture originally isolated from the Red Sea and assess their joint responses to experimental warming within the metabolic ecology framework. A co-culture of Synechococcus sp. RS9907 and their associated heterotrophic bacteria, after determining their taxonomic affiliation by 16S rRNA gene sequencing, was acclimated and maintained in the lab at different temperatures (24–34°C). The abundance and cellular properties of Synechococcus and the three dominant heterotrophic bacterial groups (pertaining to the genera Paracoccus, Marinobacter, and Muricauda) were monitored by flow cytometry. The activation energy of Synechococcus, which grew at 0.94–1.38 d–1, was very similar (0.34 ± 0.02 eV) to the value hypothesized by the metabolic theory of ecology (MTE) for autotrophs (0.32 eV), while the values of the three heterotrophic bacteria ranged from 0.16 to 1.15 eV and were negatively correlated with their corresponding specific growth rates (2.38–24.4 d–1). The corresponding carrying capacities did not always follow the inverse relationship with temperature predicted by MTE, nor did we observe a consistent response of bacterial cell size and temperature. Our results show that the responses to future ocean warming of autotrophic and heterotrophic bacteria in microbial consortia might not be well described by theoretical universal rules.

Microbial diversity of co-occurring heterotrophs in cultures of marine picocyanobacteria

BackgroundThe cyanobacteria Prochlorococcus and Synechococcus are responsible for around 10% of global net primary productivity, serving as part of the foundation of marine food webs. Heterotrophic bacteria are often co-isolated with these picocyanobacteria in seawater enrichment cultures that contain no added organic carbon; heterotrophs grow on organic carbon supplied by the photolithoautotrophs. For examining the selective pressures shaping autotroph/heterotroph interactions, we have made use of unialgal enrichment cultures of Prochlorococcus and Synechococcus maintained for hundreds to thousands of generations in the lab. We examine the diversity of heterotrophs in 74 enrichment cultures of these picocyanobacteria obtained from diverse areas of the global oceans.ResultsHeterotroph community composition differed between clades and ecotypes of the autotrophic ‘hosts’ but there was significant overlap in heterotroph community composition across these cultures. Collectively, the cultures were comprised of many shared taxa, even at the genus level. Yet, observed differences in community composition were associated with time since isolation, location, depth, and methods of isolation. The majority of heterotrophs in the cultures are rare in the global ocean, but enrichment conditions favor the opportunistic outgrowth of these rare bacteria. However, we found a few examples, such as bacteria in the family Rhodobacteraceae, of heterotrophs that were ubiquitous and abundant in cultures and in the global oceans. We found their abundance in the wild is also positively correlated with that of picocyanobacteria.ConclusionsParticular conditions surrounding isolation have a persistent effect on long-term culture composition, likely from bottlenecking and selection that happen during the early stages of enrichment for the picocyanobacteria. We highlight the potential for examining ecologically relevant relationships by identifying patterns of distribution of culture-enriched organisms in the global oceans.

Iron Speciation and Physiological Analysis Indicate that Synechococcus sp. PCC 7002 Reduces Amorphous and Crystalline Iron Forms in Synthetic Seawater Medium

Cyanobacteria have high iron requirements due to iron-rich photosynthetic machineries. Despite the high concentrations of iron in the Earth’s crust, iron is limiting in many marine environments due to iron’s low solubility. Oxic conditions leave a large portion of the ocean’s iron pool unavailable for biotic uptake, and so the physiochemical properties of iron are hugely important for iron’s bioavailability. Our study is the first to investigate the effect of iron source on iron internalization and extracellular reduction by Synechococcus sp. PCC 7002. The results indicated that the amorphous iron hydrolysis species produced by FeCl3 better support growth in Synechococcus through more efficient iron internalization and a larger degree of extracellular reduction of iron than the crystalline FeO(OH). An analysis of dissolved iron (II) indicated that biogenic reduction took place in cultures of Synechococcus grown on both FeCl3 and FeO(OH).

Investigating Photosynthetic Stability: Relation Between Thylakoid Lipid Content and the Stability of the Cytochrome b6f Complex

The cytochrome b6f complex is an enzyme found in plants, cyanobacteria, and green algae that catalyzes the transport of electrons in the rate-limiting step of oxygenic photosynthesis. This dimeric complex has an extensive lipid architecture that is primarily composed of five distinct lipid classes: monogalactosyl diacylglycerol (MGDG), digalactosyl diacylglycerol (DGDG), phosphatidyl glycerol (PG), monoglucosyl diacylglycerol (GlcDG), and sulfoquinovosyl diacylglycerol (SQDG). While these lipid classes have been identified, their precise role in the function of the cytochrome complex are only beginning to be understood. Mechanisms describing the relation between thylakoid lipid content on the stability of the b6f complex are not known. This study validates the importance of the lipids on cytochrome b6f dimer formation and stability by showing that SQDG and the synthetic lipids 1,2-dioleoylphosphatidylglycerol and 2-dioleoyl-sn-glycero-3-phosphocholine reduce the temperature dependent rate of monomerization (denaturation) of the native dimer. A novel method of growing the cyanobacterium Synechococcus PCC 7002 anaerobically to test the relation between thylakoid lipid content and growth temperature was developed. This method of growing Synechococcus greatly reduces the relative SQDG content and increases the relative PG content in thylakoid membranes. The analysis of MGDG, DGDG, PG, GlcDG, and SQDG content in Synechococcus cultures grown at 30℃ and 33℃ revealed that the MGDG content depends inversely on the growth temperature.

Heterotrophic Bacteria Enhance the Aggregation of the Marine Picocyanobacteria Prochlorococcus and Synechococcus.

Marine picocyanobacteria are ubiquitous primary producers across the world’s oceans, and play a key role in the global carbon cycle. Recent evidence stemming from in situ investigations have shown that picocyanobacteria are able to sink out of the euphotic zone to depth, which has traditionally been associated with larger, mineral ballasted cells. The mechanisms behind the sinking of picocyanobacteria remain a point of contention, given that they are too small to sink on their own. To gain a mechanistic understanding of the potential role of picocyanobacteria in carbon export, we tested their ability to form “suspended” (5–60 μm) and “visible” (ca. > 0.1 mm) aggregates, as well as their production of transparent exopolymer particles (TEP)—which are a key component in the formation of marine aggregates. Additionally, we investigated if interactions with heterotrophic bacteria play a role in TEP production and aggregation in Prochlorococcus and Synechococcus by comparing xenic and axenic cultures. We observed TEP production and aggregation in batch cultures of axenic Synechococcus, but not in axenic Prochlorococcus. Heterotrophic bacteria enhanced TEP production as well as suspended and visible aggregate formation in Prochlorococcus, while in Synechococcus, aggregation was enhanced with no changes in TEP. Aggregation experiments using a natural plankton community dominated by picocyanobacteria resulted in aggregation only in the presence of the ballasting mineral kaolinite, and only when Synechococcus were in their highest seasonal abundance. Our results point to a different export potential between the two picocyanobacteria, which may be mediated by interactions with heterotrophic bacteria and presence of ballasting minerals. Further studies are needed to clarify the mechanistic role of bacteria in TEP production and aggregation of these picocyanobacteria.

TEP production under oxidative stress of the picocyanobacterium Synechococcus

Transparent exopolymer particles (TEP) are mainly acidic polysaccharides directly or indirectly formed by phytoplankton and bacteria. These particles are often colonized by picoplankton and considered a hot spot for microbial activity. Recent studies suggested an important role of Synechococcus in TEP production found in lakes and prompted us to further investigate this issue using monoclonal xenic cultures of Synechococcus. We tested TEP production under oxidative stress in two treatments, one with hydrogen peroxide and another treated with ultraviolet radiation (UVR) and high photosynthetic active radiation (PAR), compared with an unstressed control. Our results showed a cell-normalized TEP production, ranging from 12 to 238 ng C cell-1 among strains, not only under stress but also in the control with non-limiting nutrients. Our data prove that freshwater communities of Synechococcus and their associated heterotrophic microflora, are capable of producing TEP even during growth phase. The oxidative stress induced extra production of TEP up to 400 ng C cell-1 in one of our phycocyanin-type (PC) strain. The phycoerythrin-type (PE) strains increased TEP production, particularly under UV-PAR stress, whereas the PC strains did it under H2O2 stress. This study provides new perspectives on the potential role of freshwater Synechococcus in TEP production.

Genomic reconstructions and potential metabolic strategies of generalist and specialist heterotrophic bacteria associated with an estuary Synechococcus culture.

Interactions between photoautotrophs and heterotrophs are central to marine microbial ecosystems. Synechococcus are dominant marine phototrophs, and they are frequently associated with heterotrophic bacteria. These co-cultures provide a useful research system to investigate photoautotroph-heterotroph interactions in marine systems. Bacteria within the Roseobacter clade and Flavobacteria are two of the main bacterial lineages that exhibit intimate associations with Synechococcus populations. We conducted metagenomic analyses of a Synechococcus culture, followed by genomic binning of metagenomic contigs, and recovered five nearly complete genomes, including members of the Roseobacter clade (i.e. Marivita sp. XM-24) and Flavobacteria (i.e. Fluviicola sp. XM-24). Marivita sp. XM-24 is an ecological generalist of the Roseobacter clade and displays diverse metabolic capacities for the acquisition of nutrients and energy sources. Specifically, the genome contained numerous gene complements involved in the uptake and metabolism of nitrogen- and phosphorus-containing inorganic and organic compounds, in addition to the potential for aerobic anoxygenic photosynthesis, oxidation of carbon monoxide, inorganic sulfur oxidation, DMSP demethylation and PHA metabolism. The genome of the Flavobacteria representative, Fluviicola sp. XM-24, contained numerous peptidases, glycoside hydrolases, adhesion-related proteins and genes involved in gliding motility. Fluviicola sp. XM-24 likely specialize in the degradation of high molecular weight compound exudates from Synechococcus cells, including polysaccharides and polypeptides via attachment to particles, surfaces or cells. The distinct metabolic strategies identified within several heterotrophic bacteria that are associated with Syneochococcus cells provide insights into their lifestyles and nutrient utilization patterns, in addition to their interactions with photoautotrophs. Biological interactions, including mutualism, competition and antagonism, shape the microbial community structure of marine environments and are critical for understanding biogeochemical cycling in the ocean. These results provide valuable insights into the nature of interactions between dominant marine photoautotrophs and associated bacterial heterotrophs.

Roseicyclus marinus sp. nov., isolated from a Synechococcus culture, and emended description of the genus Roseicyclus.

A novel Gram-stain-negative, aerobic, non-flagellated, pink-pigmented and rod-shaped strain with gliding motility, designated strain CCMM001T, was isolated from a mixed culture of Synechococcus species PCC7002 and a natural bacterial community from a sample of offshore seawater from Qingdao, China, during September 2014. The strain contained bacteriochlorophyll a with a small peak at 802 nm and a large in vivo absorption band at 870 nm. Strain CCMM001T grew optimally at pH 7.0 and 30 °C in the presence of 3 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain CCMM001T is most closely related to the genus Roseicyclus and its type and only species Roseicyclus mahoneyensis ML6T with 96.9 % sequence similarity. The polar lipids of strain CCMM001T consisted of phosphatidylethanolamine, phosphatidylcholine, one unidentified aminolipid, and five unidentified lipids. The predominant isoprenoid quinone was Q-10. The major fatty acids included C18 : 1ω7c and C19 : 0cyclo ω8c. The DNA G+C content of strain CCMM001T was 63.5 mol%. These phylogenetic, physiological and chemotaxonomic data indicated that strain CCMM001T represents a novel species of the genus Roseicyclus, for which the name Roseicyclus marinus sp. nov. is proposed. The type strain is CCMM001T (=MCCC 1K03242T=KCTC 52641T).

Dynamics of Heterotrophic Bacterial Assemblages within Synechococcus Cultures.

Interactions between photoautotrophic and heterotrophic microorganisms are central to the marine microbial ecosystem. Lab cultures of one of the dominant marine photoautotrophs, Synechococcus, have historically been difficult to render axenic, presumably because these bacteria depend upon other organisms to grow under these conditions. These tight associations between Synechococcus and heterotrophic bacteria represent a good relevant system to study interspecies interactions. Ten individual Synechococcus strains, isolated from eutrophic and oligotrophic waters, were chosen for investigation. Four to six dominant associated heterotrophic bacteria were detected in the liquid cultures of each Synechococcus isolate, comprising members of the Cytophaga-Flavobacteria-Bacteroides (CFB) group (mainly from Flavobacteriales and Cytophagales), Alphaproteobacteria (mainly from the Roseobacter clade), Gammaproteobacteria (mainly from the Alteromonadales and Pseudomonadales), and Actinobacteria The presence of the CFB group, Gammaproteobacteria, and Actinobacteria showed clear geographic patterns related to the isolation environments of the Synechococcus bacteria. An investigation of the population dynamics within a growing culture (XM-24) of one of the isolates, including an evaluation of the proportions of cells that were free-living versus aggregated/attached, revealed interesting patterns for different bacterial groups. In Synechococcus sp. strain XM-24 culture, flavobacteria, which was the most abundant group throughout the culture period, tended to be aggregated or attached to the Synechococcus cells, whereas the actinobacteria demonstrated a free-living lifestyle, and roseobacters displayed different patterns depending on the culture growth phase. Factors contributing to these succession patterns for the heterotrophs likely include interactions among the culture community members, their relative abilities to utilize different compounds produced by Synechococcus cells and changes in the compounds released as culture growth proceeds, and their responses to other changes in the environmental conditions throughout the culture period.IMPORTANCE Marine microbes exist within an interactive ecological network, and studying their interactions is an important part of understanding their roles in global biogeochemical cycling and the determinants of microbial diversity. In this study, the dynamic relationships between Synechococcus spp. and their associated heterotrophic bacteria were investigated. Synechococcus-associated heterotrophic bacteria had similar geographic distribution patterns as their "host" and displayed different lifestyles (free-living versus attached/aggregated) according to the Synechococcus culture growth phases. Combined organic carbon composition and bacterial lifestyle data indicated a potential for succession in carbon utilization patterns by the dominant associated heterotrophic bacteria. Comprehending the interactions between photoautotrophs and heterotrophs and the patterns of organic carbon excretion and utilization is critical to understanding their roles in oceanic biogeochemical cycling.

Construction of Local Isolates of Cyanobacteria for Ethanol Production.

Cyanobacteria can use solar energy and convert carbon dioxide into biofuel molecules in
 one single biological system. In this research, Synechococcus sp. was isolated from
 Saray Subhan Agha fresh water, a pure culture of Synechococcus was obtained by
 several subculturing on BG11 media. For the production of ethanol by Synechococcus
 sp. pyruvate decarboxylase (PDC) and alcohol dehydrogenseII (ADH II), genes from
 Zymomonas mobilis ATCC (29191), were amplified by PCR and cloned into the
 pSyn_1⁄D-TOPO® Vector. The Synechococcus and Synechococcus elongates
 transformed with constructed vector (pSyn_1⁄D-TOPO®) that harboring the two ethanol
 fermenting genes. The transformation was performed using a double homologous
 recombination system to integrate the PDC and ADHII genes into the local isolates of
 Synechococcus sp. and Synechococcus elongatus chromosome under the control
 cyanobacterial weak constitutive nickel inducible promoter. The recombinant
 Synechococcus cells grow in different concentrations of NiSO4 (1, 2.5, 5, 7.5, 10) μM in
 BG11 media, under different temperature (15, 30, 45) ̊C and different light intensity (10,
 50, 150) μE. The enzymatic ethanol assay kit was used to determine ethanol
 concentration produced by both recombinant Synechococcus sp. and recombinant
 Synechococcus elongatus. Highest ethanol concentration obtained by those cultures
 containing five μl NiSO4, which incubated under continues light of 50μE at 30 ̊C
 (Optimum um condition for ethanol production by recombinant Synechococcus cells).
 The amount of ethanol produced by local isolates of Synechococcus sp. was 0.00103 g/l,
 whereas for Synechococcus elongatus was 0.0138 g/l. The amount of ethanol produced
 by those Synechococcus cultures containing different concentrations of NiSO4 were
 incubated under continuous light of (10 and 150) μE and temperature of (15 and 45) ̊C
 was less than those cultures were incubated under light of 50 μE and temperature of 30 ̊C.

High-quality draft genome sequence of Gracilimonas tropica CL-CB462(T) (DSM 19535(T)), isolated from a Synechococcus culture.

Gracilimonas tropica Choi et al. 2009 is a member of order Sphingobacteriales, class Sphingobacteriia. Three species of the genus Gracilimonas have been isolated from marine seawater or a salt mine and showed extremely halotolerant and mesophilic features, although close relatives are extremely halophilic or thermophilic. The type strain of the type species of Gracilimonas, G. tropica DSM19535T, was isolated from a Synechococcus culture which was established from the tropical sea-surface water of the Pacific Ocean. The genome of the strain DSM19535T was sequenced through the Genomic Encyclopedia of Type Strains, Phase I: the one thousand microbial genomes project. Here, we describe the genomic features of the strain. The 3,831,242 bp long draft genome consists of 48 contigs with 3373 protein-coding and 53 RNA genes. The strain seems to adapt to phosphate limitation and requires amino acids from external environment. In addition, genomic analyses and pasteurization experiment suggested that G. tropica DSM19535T did not form spore.