Synechococcus Lineages Research Articles

Aggregating Synechococcus contributes to particle organic carbon export in coastal estuarine waters: Its lineage features and assembly processes

The release and deposition of phytoplankton-derived particulate organic matter is crucial in marine carbon export, yet the roles of picoplankton in these processes were seldom considered. Therefore, this study aimed to shed light on the matter by investigating the aggregating (AG) lifestyle of Synechococcus, a main group of picoplankton, in the coastal waters of the Yellow River Estuary with ample sediments acting as ballast minerals. We revealed that AG Synechococcus constituted a substantial portion, maximally reaching up to 85.4 %, of the total Synechococcus population. Pearson correlations and random forest (RF) regression analyses found significant connections (p < 0.01) between AG Synechococcus and the content of particulate organic carbon (POC), which emphasized its underlying role in facilitating POC export in this region. Furthermore, by employing high-throughput sequencing of the RNA polymerase gene (rpoC1), it was demonstrated that S5.1 clade I exhibited a significantly higher proportion in the AG fraction than in the free-living (FL) fraction (p < 0.05). This suggests distinct inclinations in the phylogenetic preference for different Synechococcus lineages between different lifestyles in the studied area. Finally, we ascertained “small-world” and higher robustness attributes of aggregates formed through the co-occurrence construction between Synechococcus and heterotrophic bacteria, likely facilitated by the reciprocal exchange of carbon and nitrogen elements. Overall, these findings have implications for our understanding of the role of Synechococcus in the ecology and biogeochemistry of marine ecosystems, and they are significant for more accurately evaluating the contribution of picophytoplankton in ocean carbon export.

Molecular insights into nitrogen constraint for niche partitioning and physiological adaptation of coastal Synechococcus assemblages

Coastal nitrogen input has substantially increased due to human activity. However, much remains unknown about the nitrogen-driven patterns and the underlying genetic basis of coastal picoplankton. To investigate the response and mechanisms of picoplankton induced by nitrogen variation, we conducted in-situ investigations using high-throughput sequencing in the Bohai Sea and performed laboratory nitrogen simulation experiments accompanied by physiological, genomic, and transcriptomic analyses, with Synechococcus as a representative. The results of in-situ investigation revealed that Synechococcus clades I, III, WPC1, and VI of subcluster 5.1 (S5.1) are prevalent in strait areas characterized by robust water exchange with the North Yellow Sea, while clades II, VIII, and IX of S5.1, as well as subcluster 5.2 (S5.2) and subcluster 5.3 (S5.3) are more abundant in central and bay areas experiencing elevated nitrate and nitrite loads. The laboratory experiments further confirmed that inorganic nitrogen is a crucial determinant of diversity and niche partitioning of Synechococcus lineages. Besides, the raising inorganic nitrogen concentration within the current in-situ range (0.1–10 μmol L−1) enhances photosynthesis and carbon fixation of Synechococcus, however further escalation of inorganic nitrogen (100 μmol L−1) may hinder these processes instead. The phenomenon could be associated with the differential expression of genes in metabolic pathways regulating nitrogen metabolism, photosynthetic system II, and photosynthesis-antenna proteins in response to nitrogen concentration and type variation. These findings expand our understanding of the impact of macronutrient variation resulting from human activities on marine picoplankton and biogeochemical cycles.

Global Phylogeography of Marine Synechococcus in Coastal Areas Reveals Strong Community Shifts.

Marine Synechococcus comprise a numerically and ecologically prominent phytoplankton group, playing a major role in both carbon cycling and trophic networks in all oceanic regions except in the polar oceans. Despite their high abundance in coastal areas, our knowledge of Synechococcus communities in these environments is based on only a few local studies. Here, we use the global metagenome data set of the Ocean Sampling Day (June 21st, 2014) to get a snapshot of the taxonomic composition of coastal Synechococcus communities worldwide, by recruitment on a reference database of 141 picocyanobacterial genomes, representative of the whole Prochlorococcus, Synechococcus, and Cyanobium diversity. This allowed us to unravel drastic community shifts over small to medium scale gradients of environmental factors, in particular along European coasts. The combined analysis of the phylogeography of natural populations and the thermophysiological characterization of eight strains, representative of the four major Synechococcus lineages (clades I to IV), also brought novel insights about the differential niche partitioning of clades I and IV, which most often co-dominate the Synechococcus community in cold and temperate coastal areas. Altogether, this study reveals several important characteristics and specificities of the coastal communities of Synechococcus worldwide. IMPORTANCE Synechococcus is the second most abundant phytoplanktonic organism on Earth, and its wide genetic diversity allowed it to colonize all the oceans except for polar waters, with different clades colonizing distinct oceanic niches. In recent years, the use of global metagenomics data sets has greatly improved our knowledge of "who is where" by describing the distribution of Synechococcus clades or ecotypes in the open ocean. However, little is known about the global distribution of Synechococcus ecotypes in coastal areas, where Synechococcus is often the dominant phytoplanktonic organism. Here, we leverage the global Ocean Sampling Day metagenomics data set to describe Synechococcus community composition in coastal areas worldwide, revealing striking community shifts, in particular along the coasts of Europe. As temperature appears as an important driver of the community composition, we also characterize the thermal preferenda of 8 Synechococcus strains, bringing new insights into the adaptation to temperature of the dominant Synechococcus clades.

Diversity and Evolution of Pigment Types in Marine Synechococcus Cyanobacteria.

Synechococcus cyanobacteria are ubiquitous and abundant in the marine environment and contribute to an estimated 16% of the ocean net primary productivity. Their light-harvesting complexes, called phycobilisomes (PBS), are composed of a conserved allophycocyanin core, from which radiates six to eight rods with variable phycobiliprotein and chromophore content. This variability allows Synechococcus cells to optimally exploit the wide variety of spectral niches existing in marine ecosystems. Seven distinct pigment types or subtypes have been identified so far in this taxon based on the phycobiliprotein composition and/or the proportion of the different chromophores in PBS rods. Most genes involved in their biosynthesis and regulation are located in a dedicated genomic region called the PBS rod region. Here, we examine the variability of gene content and organization of this genomic region in a large set of sequenced isolates and natural populations of Synechococcus representative of all known pigment types. All regions start with a tRNA-PheGAA and some possess mobile elements for DNA integration and site-specific recombination, suggesting that their genomic variability relies in part on a “tycheposon”-like mechanism. Comparison of the phylogenies obtained for PBS and core genes revealed that the evolutionary history of PBS rod genes differs from the core genome and is characterized by the co-existence of different alleles and frequent allelic exchange. We propose a scenario for the evolution of the different pigment types and highlight the importance of incomplete lineage sorting in maintaining a wide diversity of pigment types in different Synechococcus lineages despite multiple speciation events.

Phylogeny and taxonomy of Synechococcus-like cyanobacteria

Unicellular cyanobacteria constitute a substantial, ecologically important part of freshwater and marine microflora. Solitary, elongated cyanobacterial cells without apparent slime envelopes and dividing in a single plane perpendicular to the longitudinal axis were traditionally classified into the genus Synechococcus. The type species originates from a freshwater benthic habitat while a number of other Synechococcus-like species were described from diverse environments. Morphologically similar, unicellular populations of belong to the most abundant oxygen evolving prokaryotes inhabiting freshwater and oceanic picoplanktic communities. Other species from extreme thermal habitats were described from temperatures over 70 °C. Recent molecular analyses, particularly the 16S rRNA gene and other multi-locus gene based phylogenies of Synechococcus-like species reveal the polyphyly and phylogenetic diversity of such simple cyanobacteria emphasizing the necessity of their accurate polyphasic taxonomic re-classification. To better understand the phylogeny of different Synechococcus-like species, we constructed a genome-derived phylogenetic tree using 133 cyanobacterial and eubacterial genomes as well as a 16S rRNA gene phylogenetic tree. Both trees indicate existence of deep splits between individual Synechococcus lineages corresponding with indistinct ecological and cytomorphological differences. The generic units derived from the traditional concept of the genus Synechococcus based on the combined traits and markers must be defined and validly described. The comparative taxonomic classification of phylogenetic clusters corresponding to various morphotypes and genotypes similar to the genus Synechococcus (Cyanobium, Parasynechococcus, Picosynechococcus, Prochlorococcus, Thermosynechococcus, Thermostichus) are discussed in the article.

Evolutionary Mechanisms of Long-Term Genome Diversification Associated With Niche Partitioning in Marine Picocyanobacteria.

Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus are the most abundant photosynthetic organisms on Earth, an ecological success thought to be linked to the differential partitioning of distinct ecotypes into specific ecological niches. However, the underlying processes that governed the diversification of these microorganisms and the appearance of niche-related phenotypic traits are just starting to be elucidated. Here, by comparing 81 genomes, including 34 new Synechococcus, we explored the evolutionary processes that shaped the genomic diversity of picocyanobacteria. Time-calibration of a core-protein tree showed that gene gain/loss occurred at an unexpectedly low rate between the different lineages, with for instance 5.6 genes gained per million years (My) for the major Synechococcus lineage (sub-cluster 5.1), among which only 0.71/My have been fixed in the long term. Gene content comparisons revealed a number of candidates involved in nutrient adaptation, a large proportion of which are located in genomic islands shared between either closely or more distantly related strains, as identified using an original network construction approach. Interestingly, strains representative of the different ecotypes co-occurring in phosphorus-depleted waters (Synechococcus clades III, WPC1, and sub-cluster 5.3) were shown to display different adaptation strategies to this limitation. In contrast, we found few genes potentially involved in adaptation to temperature when comparing cold and warm thermotypes. Indeed, comparison of core protein sequences highlighted variants specific to cold thermotypes, notably involved in carotenoid biosynthesis and the oxidative stress response, revealing that long-term adaptation to thermal niches relies on amino acid substitutions rather than on gene content variation. Altogether, this study not only deciphers the respective roles of gene gains/losses and sequence variation but also uncovers numerous gene candidates likely involved in niche partitioning of two key members of the marine phytoplankton.

Short-Term Changes in Marine Prokaryotic and Eukaryotic Microalgal Communities Exposed to the Leachate of a Seafloor Hydrothermal Sulfide

The effects of increased mining of seafloor massive sulfide deposits on marine ecosystems have not been characterized. In this study, the impact of leaching metals from a hydrothermal sulfide on photosynthetic protist and cyanobacterial communities in marine environments was investigated by amplicon analyses of small subunit rDNA (SSU rDNA) and rRNA (SSU rRNA). Seawater samples collected from the Iheya North region and Suruga Bay, Japan, were incubated with or without a leachate containing zinc, copper, cadmium, and manganese, of the actual seafloor hydrothermal sulfide from the Hakurei site in the Izena Hole region. The relative abundances of prasinophytes, diatom protists, and the cyanobacteria Synechococcus decreased substantially during incubation with leachate, indicating the vulnerability of these lineages to the leachate. Phylogenetic analysis based on the cyanobacterial phycocyanin cpcBA/rpcBA operon obtained from samples incubated with or without leachate indicated that the individual lineages of Synechococcus can determine sensitivity to heavy metals in different marine regions as well as particular clades and ecotypes.

Insights Into the Evolution of Picocyanobacteria and Phycoerythrin Genes (mpeBA and cpeBA).

Marine picocyanobacteria, Prochlorococcus and Synechococcus, substantially contribute to marine primary production and have been the subject of extensive ecological and genomic studies. Little is known about their close relatives from freshwater and non-marine environments. Phylogenomic analyses (using 136 proteins) provide strong support for the monophyly of a clade of non-marine picocyanobacteria consisting of Cyanobium, Synechococcus and marine Sub-cluster 5.2; this clade itself is sister to marine Synechococcus and Prochlorococcus. The most basal lineage within the Syn/Pro clade, Sub-Cluster 5.3, includes marine and freshwater strains. Relaxed molecular clock (SSU, LSU) analyses show that while ancestors of the Syn/Pro clade date as far back as the end of the Pre-Cambrian, modern crown groups evolved during the Carboniferous and Triassic. Comparative genomic analyses reveal novel gene cluster arrangements involved in phycobilisome (PBS) metabolism in freshwater strains. Whilst PBS genes in marine Synechococcus are mostly found in one type of phycoerythrin (PE) rich gene cluster (Type III), strains from non-marine habitats, so far, appear to be more diverse both in terms of pigment content and gene arrangement, likely reflecting a wider range of habitats. Our phylogenetic analyses show that the PE genes (mpeBA) evolved via a duplication of the cpeBA genes in an ancestor of the marine and non-marine picocyanobacteria and of the symbiotic strains Synechococcus spongiarum. A ‘primitive’ Type III-like ancestor containing cpeBA and mpeBA had thus evolved prior to the divergence of the Syn/Pro clade and S. spongiarum. During the diversification of Synechococcus lineages, losses of mpeBA genes may explain the emergence of pigment cluster Types I, II, IIB, and III in both marine and non-marine habitats, with few lateral gene transfer events in specific taxa.

Latitudinal and Vertical Variation of Synechococcus Assemblage Composition Along 170° W Transect From the South Pacific to the Arctic Ocean.

Synechococcus is one of the most widely distributed and abundant picocyanobacteria in the global oceans. Although latitudinal variation of Synechococcus assemblage in marine surface waters has been observed, few studies compared Synechococcus assemblage composition in surface and subsurface waters at the basin scale. Here, we report marine Synechococcus diversity in the surface and deep chlorophyll maximum (DCM) layers along 170° W from the South Pacific to the Arctic Ocean in summer. Along the transect, spatial niche partitioning of Synechococcus lineages in the surface waters was clearly observed. Species richness of surface Synechococcus assemblage was positively correlated with water temperature. Clade CRD1 was dominant in the areas (15° S-10° N and 35-40° N) associated with upwelling, and there were 3 different subclades with distinct distribution. CRD1-A was restricted in the North Equatorial Current (5-10° N), CRD1-B dominated in the equatorial upwelling region (15° S-0.17° N), and CRD1-C was only distributed in the North Pacific Current (35-40° N). Similarities between the Synechococcus assemblages in the surface and DCM layers were high at the upwelling regions and areas where the mixed layer was deep, while low in the Subtropical Gyres with strong stratification. Clade I, CRD1-B, and CRD1-C were major Synechococcus lineages in the DCM layer. In particular, clade I, which is composed of 7 subclades with distinct thermal niches, was widely distributed in the DCM layer. Overall, our results provide new insights into not only the latitudinal distribution of Synechococcus assemblages, but also their vertical variation in the central Pacific.

Cross-protection from hydrogen peroxide by helper microbes: the impacts on the cyanobacterium Prochlorococcus and other beneficiaries in marine communities.

Hydrogen peroxide (HOOH) is a reactive oxygen species, derived from molecular oxygen, that is capable of damaging microbial cells. Surprisingly, the HOOH defence systems of some aerobes in the oxygenated marine environments are critically depleted, relative to model aerobes. For instance, the gene encoding catalase is absent in the numerically dominant photosynthetic cyanobacterium, Prochlorococcus. Accordingly, Prochlorococcus is highly susceptible to HOOH when exposed as pure cultures. Pure cultures do not exist in the marine environment, however. Catalase-positive community members can remove HOOH from the seawater medium, thus lowering the threat to Prochlorococcus and any other member that likewise lacks their own catalase. This cross-protection may constitute a loosely defined symbiosis, whereby the catalase-positive helper cells may benefit through the acquisition of nutrients released by the beneficiaries such as Prochlorococcus. Other members of the community that may be helped by the catalase-positive cells may include some lineages of Synechococcus - the sister genus of Prochlorococcus - as well as some lineages of SAR11 and ammonia oxidizing archaea and bacteria. The co-occurrence of catalase-positive and -negative members suggests that cross-protection from HOOH-mediated oxidative stress may play an important role in the construction of the marine microbial community.

Synechococcus Assemblages across the Salinity Gradient in a Salt Wedge Estuary.

Synechococcus are the most abundant and widely distributed picocyanobacteria in the ocean. The salt-wedge type of estuary possesses the complete horizontal and vertical gradient of salinity together with other physical and chemical parameters. In order to reveal whether such a complex environmental gradient harbors a high diversity of Synechococcus, we investigated the abundance, taxonomic composition and pigment genetic diversity of Synechococcus in surface and bottom waters across the salinity gradient in a salt-wedge estuary by flow cytometric analysis and pyrosequencing of the rpoC1 gene and cpcBA operon (encoding phycocyanin). Synechococcus were ubiquitously distributed in the studied region, with clear spatial variations both horizontally and vertically. The abundance and diversity of Synechococcus were low in the freshwater-dominated low salinity waters. By pyrosequencing of the rpoC1 gene, we have shown that with the increase of salinity, the dominant Synechococcus shifted from the freshwater Synechococcus to the combination of phylogenetic subcluster 5.2 and freshwater Synechococcus, and then the strictly marine subcluster 5.1 clade III. Besides, the composition of Synechococcus assemblage in the deep layer was markedly different from the surface in the stratified waters (dissimilarities: 40.32%-95.97%, SIMPER analysis). High abundance of clade III Synechococcus found in the brackish waters may revise our previous understanding that strains of this clade prefers oligotrophic environment. Our data also suggested that both the phylogenetic subcluster 5.3 Synechococcus, a lineage that was not well understood, and subcluster 5.1 clade I, a typical cold water lineage, were widely distributed in the bottom layer of the estuary. Clade I detected in the studied region was mainly contributed by subclade IG. Analysis of the cpcBA operon sequences revealed niche partitioning between type 1 and type 3 Synechococcus, with type 2 distributed broadly across the whole environmental gradients. Our results suggest that the salt wedge estuary provides various niches for different lineages of Synechococcus, making it an environment with high Synechococcus diversity compared with adjacent freshwater and shelf sea environments.

Phylogeography and pigment type diversity of Synechococcus cyanobacteria in surface waters of the northwestern pacific ocean.

The widespread unicellular cyanobacteria Synechococcus are major contributors to global marine primary production. Here, we report their abundance, phylogenetic diversity (as assessed using the RNA polymerase gamma subunit gene rpoC1) and pigment diversity (as indirectly assessed using the laterally transferred cpeBA genes, encoding phycoerythrin-I) in surface waters of the northwestern Pacific Ocean, sampled over nine distinct cruises (2008-2015). Abundance of Synechococcus was low in the subarctic ocean and South China Sea, intermediate in the western subtropical Pacific Ocean, and the highest in the Japan and East China seas. Clades I and II were by far the most abundant Synechococcus lineages, the former dominating in temperate cold waters and the latter in (sub)tropical waters. Clades III and VI were also fairly abundant in warm waters, but with a narrower distribution than clade II. One type of chromatic acclimater (3dA) largely dominated the Synechococcus communities in the subarctic ocean, while another (3dB) and/or cells with a fixed high phycourobilin to phycoerythrobilin ratio (pigment type 3c) predominated at mid and low latitudes. Altogether, our results suggest that the variety of pigment content found in most Synechococcus clades considerably extends the niches that they can colonize and therefore the whole genus habitat.

Self-regulating genomic island encoding tandem regulators confers chromatic acclimation to marine Synechococcus

The evolutionary success of marine Synechococcus, the second-most abundant phototrophic group in the marine environment, is partly attributable to this group's ability to use the entire visible spectrum of light for photosynthesis. This group possesses a remarkable diversity of light-harvesting pigments, and most of the group's members are orange and pink because of their use of phycourobilin and phycoerythrobilin chromophores, which are attached to antennae proteins called phycoerythrins. Many strains can alter phycoerythrin chromophore ratios to optimize photon capture in changing blue-green environments using type IV chromatic acclimation (CA4). Although CA4 is common in most marine Synechococcus lineages, the regulation of this process remains unexplored. Here, we show that a widely distributed genomic island encoding tandem master regulators named FciA (for type four chromatic acclimation island) and FciB plays a central role in controlling CA4. FciA and FciB have diametric effects on CA4. Interruption of fciA causes a constitutive green light phenotype, and interruption of fciB causes a constitutive blue light phenotype. These proteins regulate all of the molecular responses occurring during CA4, and the proteins' activity is apparently regulated posttranscriptionally, although their cellular ratio appears to be critical for establishing the set point for the blue-green switch in ecologically relevant light environments. Surprisingly, FciA and FciB coregulate only three genes within the Synechococcus genome, all located within the same genomic island as fciA and fciB These findings, along with the widespread distribution of strains possessing this island, suggest that horizontal transfer of a small, self-regulating DNA region has conferred CA4 capability to marine Synechococcus throughout many oceanic areas.

Dynamics of the bacterial and archaeal communities in the Northern South China Sea revealed by 454 pyrosequencing of the 16S rRNA gene

Dynamics of the prokaryote (bacteria and archaea) abundance and community compositions in the Northern South China Sea (NSCS) in August 2009 and January 2010 were studied by flow cytometric analysis and 454 pyrosequencing of the 16S RNA gene. Prokaryotic community structures in the NSCS varied across space and over time, and this variation was strongly correlated with NO3- concentration. Prokaryote in estuarine and coastal waters was more abundant, but relatively less seasonally dynamic than in the open ocean. Major bacterial and archaeal lineages showed different niche preferences. Archaeal community was dominated by Marine Group I and Marine Group II. Clusters of Marine Group I varied spatially, while clusters of Marine Group II varied seasonally. Synechococcus and Prochlorococcus were two major autotrophic bacteria found in the NSCS. Synechococcus prevailed at the estuarine station in summer, while Prochlorococcus had high abundance at open-ocean stations in summer. Subcluster 5.2 Synechococcus and Sub 5.1 clade II Synechococcus were the dominant Synechococcus lineages in the NSCS, with the former dominating in the estuary during summer and the latter dominating at all other stations. Our results suggest that prokaryotic assemblages are highly complex in the NSCS and are controlled by seasonal monsoon and river discharge, showing spatiotemporal variations.

Synechococcus: 3 billion years of global dominance.

Cyanobacteria are among the most important primary producers on the Earth. However, the evolutionary forces driving cyanobacterial species diversity remain largely enigmatic due to both their distinction from macro-organisms and an undersampling of sequenced genomes. Thus, we present a new genome of a Synechococcus-like cyanobacterium from a novel evolutionary lineage. Further, we analyse all existing 16S rRNA sequences and genomes of Synechococcus-like cyanobacteria. Chronograms showed extremely polyphyletic relationships in Synechococcus, which has not been observed in any other cyanobacteria. Moreover, most Synechococcus lineages bifurcated after the Great Oxidation Event, including the most abundant marine picoplankton lineage. Quantification of horizontal gene transfer among 70 cyanobacterial genomes revealed significant differences among studied genomes. Horizontal gene transfer levels were not correlated with ecology, genome size or phenotype, but were correlated with the age of divergence. All findings were synthetized into a novel model of cyanobacterial evolution, characterized by serial convergence of the features, that is multicellularity and ecology.

Connecting thermal physiology and latitudinal niche partitioning in marine Synechococcus

Marine Synechococcus cyanobacteria constitute a monophyletic group that displays a wide latitudinal distribution, ranging from the equator to the polar fronts. Whether these organisms are all physiologically adapted to stand a large temperature gradient or stenotherms with narrow growth temperature ranges has so far remained unexplored. We submitted a panel of six strains, isolated along a gradient of latitude in the North Atlantic Ocean, to long- and short-term variations of temperature. Upon a downward shift of temperature, the strains showed strikingly distinct resistance, seemingly related to their latitude of isolation, with tropical strains collapsing while northern strains were capable of growing. This behaviour was associated to differential photosynthetic performances. In the tropical strains, the rapid photosystem II inactivation and the decrease of the antioxydant β-carotene relative to chl a suggested a strong induction of oxidative stress. These different responses were related to the thermal preferenda of the strains. The northern strains could grow at 10 °C while the other strains preferred higher temperatures. In addition, we pointed out a correspondence between strain isolation temperature and phylogeny. In particular, clades I and IV laboratory strains were all collected in the coldest waters of the distribution area of marine Synechococus. We, however, show that clade I Synechococcus exhibit different levels of adaptation, which apparently reflect their location on the latitudinal temperature gradient. This study reveals the existence of lineages of marine Synechococcus physiologically specialised in different thermal niches, therefore suggesting the existence of temperature ecotypes within the marine Synechococcus radiation.

The Evolution of RuBisCO Stability at the Thermal Limit of Photoautotrophy

A long-standing question in evolutionary biology is how organisms adapt to novel environments. In North American hot springs, diversification of a clade of the cyanobacterium Synechococcus into hotter environments has resulted in the unique innovation of a light-driven ecosystem at temperatures up to 74°C, and temperature adaptation of photosynthetic carbon fixation with the Calvin cycle contributed to this process. Here, we investigated the evolution of thermostability of the Calvin cycle enzyme ribulose-1, 5-bisphosphate carboxylase/oxygenase (RuBisCO) during Synechococcus divergence. Circular dichroism thermal scans revealed that the RuBisCO of the most thermotolerant Synechococcus lineage is more stable than those of other lineages or of resurrected ancestral enzymes. Using site-directed mutagenesis, we next identified four amino acid substitutions that together increased stability and activity of this enzyme at higher temperatures. These are clustered near critical subunit interfaces distant from the active site. Each of the four amino acids is also observed in a less thermostable Synechococcus RuBisCO, and the impact on stability of three of these appears to be epistatic. Recombination analyses that allow for recurrent mutation as well as patterns of synonymous variation surrounding these sites suggest that the evolution of a more thermostable RuBisCO may have involved homologous recombination. Our results provide insights on the molecular evolutionary processes that shape niche differentiation and ecosystem function.

Novel lineages of Prochlorococcus and Synechococcus in the global oceans

Picocyanobacteria represented by Prochlorococcus and Synechococcus have an important role in oceanic carbon fixation and nutrient cycling. In this study, we compared the community composition of picocyanobacteria from diverse marine ecosystems ranging from estuary to open oceans, tropical to polar oceans and surface to deep water, based on the sequences of 16S-23S rRNA internal transcribed spacer (ITS). A total of 1339 ITS sequences recovered from 20 samples unveiled diverse and several previously unknown clades of Prochlorococcus and Synechococcus. Six high-light (HL)-adapted Prochlorococcus clades were identified, among which clade HLVI had not been described previously. Prochlorococcus clades HLIII, HLIV and HLV, detected in the Equatorial Pacific samples, could be related to the HNLC clades recently found in the high-nutrient, low-chlorophyll (HNLC), iron-depleted tropical oceans. At least four novel Synechococcus clades (out of six clades in total) in subcluster 5.3 were found in subtropical open oceans and the South China Sea. A niche partitioning with depth was observed in the Synechococcus subcluster 5.3. Members of Synechococcus subcluster 5.2 were dominant in the high-latitude waters (northern Bering Sea and Chukchi Sea), suggesting a possible cold-adaptation of some marine Synechococcus in this subcluster. A distinct shift of the picocyanobacterial community was observed from the Bering Sea to the Chukchi Sea, which reflected the change of water temperature. Our study demonstrates that oceanic systems contain a large pool of diverse picocyanobacteria, and further suggest that new genotypes or ecotypes of picocyanobacteria will continue to emerge, as microbial consortia are explored with advanced sequencing technology.

Phylogenetic diversity of Synechococcus strains isolated from the East China Sea and the East Sea

Phylogenetic relationships among 33 Synechococcus strains isolated from the East China Sea (ECS) and the East Sea (ES) were studied based on 16S rRNA gene sequences and 16S-23S rRNA gene internal transcribed spacer (ITS) sequences. Pigment patterns of the culture strains were also examined. Based on 16S rRNA gene and ITS sequence phylogenies, the Synechococcus isolates were clustered into 10 clades, among which eight were previously identified and two were novel. Half of the culture strains belonged to clade V or VI. All strains that clustered into novel clades exhibited both phycoerythrobilin and phycourobilin. Interestingly, the pigment compositions of isolates belonging to clades V and VI differed from those reported for other oceanic regions. None of the isolates in clade V showed phycourobilin, whereas strains in clade VI exhibited both phycourobilin and phycoerythrobilin, which is in contrast to previous studies. The presence of novel lineages and the different pigment patterns in the ECS and the ES suggests the possibility that some Synechococcus lineages are distributed only in geographically restricted areas and have evolved in these regions. Therefore, further elucidation of the physiological, ecological, and genetic characteristics of the diverse Synechococcus strains is required to understand their spatial and geographical distribution.

Colorful microdiversity of Synechococcus strains (picocyanobacteria) isolated from the Baltic Sea

Synechococcus is a cosmopolitan genus of picocyanobacteria living in the photic zone of freshwater and marine ecosystems. Here, we describe the isolation of 46 closely related picocyanobacterial strains from the Baltic Sea. The isolates showed considerable variation in their cell size and pigmentation phenotypes, yielding a colorful variety of red, pink and blue-green strains. These pigmentation phenotypes could not be differentiated on the basis of their 16S rRNA-internal transcribed spacer (ITS) sequences. Thirty-nine strains, designated BSea, possessed 16S rRNA-ITS sequences almost identical with Synechococcus strain WH5701. Despite their similar 16S rRNA-ITS sequences, the BSea strains separated into several different clusters when comparing the phycocyanin (cpcBA) operon. This separation was largely consistent with the phycobiliprotein composition of the different BSea strains. The majority of phycocyanin (PC)-rich Bsea strains clustered with WH5701. Remarkably, the phycoerythrin (PE)-rich strains of BSea formed an as yet unidentified cluster within the cpcBA phylogeny, distantly related to other PE-rich groups. Detailed analysis of the cpcBA operon using neighbour-net analysis indicated that the PE-rich BSea strains are probably endemic for the Baltic Sea. Comparison of the phylogenies obtained by the 16S rRNA-ITS, the cpcBA, and the concatenated 16S rRNA-ITS and cpcBA operon sequences revealed possible events of horizontal gene transfer among different Synechococcus lineages. Our results show that microdiversity is important in Synechococcus populations and that it can reflect extensive diversification of different pigmentation phenotypes into different ecological niches.