Biological interactions with Prochlorococcus: implications for the marine carbon cycle.

The unicellular picocyanobacterium Prochlorococcus is the most abundant photoautotroph and contributes substantially to global CO2 fixation. In the vast euphotic zones of the open ocean, Prochlorococcus converts CO2 into organic compounds and supports diverse organisms, forming an intricate network of interactions that regulate the magnitude of carbon cycling and storage in the ocean. An understanding of the biological interactions with Prochlorococcus is critical for accurately estimating the contributions of Prochlorococcus and interacting organisms to the marine carbon cycle. This review synthesizes the primary production contributed by Prochlorococcus in the global ocean. We outline recent progress on the interactions of Prochlorococcus with heterotrophic bacteria, phages, and grazers that multifacetedly determine Prochlorococcus carbon production and fate. We discuss that climate change might affect the biological interactions with Prochlorococcus and thus the marine carbon cycle.

Metagenomic analysis reveals global-scale patterns of ocean nutrient limitation.

Nutrient supply regulates the activity of phytoplankton, but the global biogeography of nutrient limitation and co-limitation is poorly understood. Prochlorococcus adapt to local environments by gene gains and losses, and we used genomic changes as an indicator of adaptation to nutrient stress. We collected metagenomes from all major ocean regions as part of the Global Ocean Ship-based Hydrographic Investigations Program (Bio-GO-SHIP) and quantified shifts in genes involved in nitrogen, phosphorus, and iron assimilation. We found regional transitions in stress type and severity as well as widespread co-stress. Prochlorococcus stress genes, bottle experiments, and Earth system model predictions were correlated. We propose that the biogeography of multinutrient stress is stoichiometrically linked by controls on nitrogen fixation. Our omics-based description of phytoplankton resource use provides a nuanced and highly resolved description of nutrient stress in the global ocean.

Pili allow dominant marine cyanobacteria to avoid sinking and evade predation.

How oligotrophic marine cyanobacteria position themselves in the water column is currently unknown. The current paradigm is that these organisms avoid sinking due to their reduced size and passive drift within currents. Here, we show that one in four picocyanobacteria encode a type IV pilus which allows these organisms to increase drag and remain suspended at optimal positions in the water column, as well as evade predation by grazers. The evolution of this sophisticated floatation mechanism in these purely planktonic streamlined microorganisms has important implications for our current understanding of microbial distribution in the oceans and predator-prey interactions which ultimately will need incorporating into future models of marine carbon flux dynamics.

Co-culture with Synechococcus facilitates growth of Prochlorococcus under ocean acidification conditions.

Anthropogenic CO2 emissions are projected to lower the pH of the ocean 0.3 units by 2100. Previous studies suggested that Prochlorococcus and Synechococcus, the numerically dominant phytoplankton in the oceans, have different responses to elevated CO2 that may result in a dramatic shift in their relative abundances in future oceans. Here we showed that the exponential growth rates of these two genera respond to future CO2 conditions in a manner similar to other cyanobacteria, but Prochlorococcus strains had significantly lower realized growth rates under elevated CO2 regimes due to poor survival after exposure to fresh culture media. Despite this, a Synechococcus strain was unable to outcompete a Prochlorococcus strain in co-culture at elevated CO2 . Under these conditions, Prochlorococcus' poor response to elevated CO2 disappeared, and Prochlorococcus' relative fitness showed negative frequency dependence, with both competitors having significant fitness advantages when initially rare. These experiments suggested that the two strains should be able to coexist indefinitely in co-culture despite sharing nearly identical nutritional requirements. We speculate that negative frequency dependence exists due to reductive Black Queen evolution that has resulted in a passively mutualistic relationship analogous to that connecting Prochlorococcus with the 'helper' heterotrophic microbes in its environment.

Unlocking the Genomic Taxonomy of the Prochlorococcus Collective.

Prochlorococcus is the most abundant photosynthetic prokaryote on our planet. The extensive ecological literature on the Prochlorococcus collective (PC) is based on the assumption that it comprises one single genus comprising the species Prochlorococcus marinus, containing itself a collective of ecotypes. Ecologists adopt the distributed genome hypothesis of an open pan-genome to explain the observed genomic diversity and evolution patterns of the ecotypes within PC. Novel genomic data for the PC prompted us to revisit this group, applying the current methods used in genomic taxonomy. As a result, we were able to distinguish the five genera: Prochlorococcus, Eurycolium, Prolificoccus, Thaumococcus, and Riococcus. The novel genera have distinct genomic and ecological attributes.

Phytoplankton transcriptomic and physiological responses to fixed nitrogen in the California current system.

Marine phytoplankton are responsible for approximately half of photosynthesis on Earth. However, their ability to drive ocean productivity depends on critical nutrients, especially bioavailable nitrogen (N) which is scarce over vast areas of the ocean. Phytoplankton differ in their preferences for N substrates as well as uptake efficiencies and minimal N requirements relative to other critical nutrients, including iron (Fe) and phosphorus. In this study, we used the MicroTOOLs high-resolution environmental microarray to examine transcriptomic responses of phytoplankton communities in the California Current System (CCS) transition zone to added urea, ammonium, nitrate, and also Fe in the late summer when N depletion is common. Transcript level changes of photosynthetic, carbon fixation, and nutrient stress genes indicated relief of N limitation in many strains of Prochlorococcus, Synechococcus, and eukaryotic phytoplankton. The transcriptomic responses helped explain shifts in physiological and growth responses observed later. All three phytoplankton groups had increased transcript levels of photosynthesis and/or carbon fixation genes in response to all N substrates. However, only Prochlorococcus had decreased transcript levels of N stress genes and grew substantially, specifically after urea and ammonium additions, suggesting that Prochlorococcus outcompeted other community members in these treatments. Diatom transcript levels of carbon fixation genes increased in response to Fe but not to Fe with N which might have favored phytoplankton that were co-limited by N and Fe. Moreover, transcription patterns of closely related strains indicated variability in N utilization, including nitrate utilization by some high-light adapted Prochlorococcus. Finally, up-regulation of urea transporter genes by both Prochlorococcus and Synechococcus in response to filtered deep water suggested a regulatory mechanism other than classic control via the global N regulator NtcA. This study indicated that co-existing phytoplankton strains experience distinct nutrient stresses in the transition zone of the CCS, an understudied region where oligotrophic and coastal communities naturally mix.

Genomic, metabolic and phenotypic variability shapes ecological differentiation and intraspecies interactions of Alteromonas macleodii.

Ecological differentiation between strains of bacterial species is shaped by genomic and metabolic variability. However, connecting genotypes to ecological niches remains a major challenge. Here, we linked bacterial geno- and phenotypes by contextualizing pangenomic, exometabolomic and physiological evidence in twelve strains of the marine bacterium Alteromonas macleodii, illuminating adaptive strategies of carbon metabolism, microbial interactions, cellular communication and iron acquisition. In A. macleodii strain MIT1002, secretion of amino acids and the unique capacity for phenol degradation may promote associations with Prochlorococcus cyanobacteria. Strain 83-1 and three novel Pacific isolates, featuring clonal genomes despite originating from distant locations, have profound abilities for algal polysaccharide utilization but without detrimental implications for Ecklonia macroalgae. Degradation of toluene and xylene, mediated via a plasmid syntenic to terrestrial Pseudomonas, was unique to strain EZ55. Benzoate degradation by strain EC673 related to a chromosomal gene cluster shared with the plasmid of A. mediterranea EC615, underlining that mobile genetic elements drive adaptations. Furthermore, we revealed strain-specific production of siderophores and homoserine lactones, with implications for nutrient acquisition and cellular communication. Phenotypic variability corresponded to different competitiveness in co-culture and geographic distribution, indicating linkages between intraspecific diversity, microbial interactions and biogeography. The finding of "ecological microdiversity" helps understanding the widespread occurrence of A. macleodii and contributes to the interpretation of bacterial niche specialization, population ecology and biogeochemical roles.

Genomic mosaicism underlies the adaptation of marine Synechococcus ecotypes to distinct oceanic iron niches.

Phytoplankton are limited by iron (Fe) in ~40% of the world's oceans including high-nutrient low-chlorophyll (HNLC) regions. While low-Fe adaptation has been well-studied in large eukaryotic diatoms, less is known for small, prokaryotic marine picocyanobacteria. This study reveals key physiological and genomic differences underlying Fe adaptation in marine picocyanobacteria. HNLC ecotype CRD1 strains have greater physiological tolerance to low Fe congruent with their expanded repertoire of Fe transporter, storage and regulatory genes compared to other ecotypes. From metagenomic analysis, genes encoding ferritin, flavodoxin, Fe transporters and siderophore uptake genes were more abundant in low-Fe waters, mirroring paradigms of low-Fe adaptation in diatoms. Distinct Fe-related gene repertories of HNLC ecotypes CRD1 and CRD2 also highlight how coexisting ecotypes have evolved independent approaches to life in low-Fe habitats. Synechococcus and Prochlorococcus HNLC ecotypes likewise exhibit independent, genome-wide reductions of predicted Fe-requiring genes. HNLC ecotype CRD1 interestingly was most similar to coastal ecotype I in Fe physiology and Fe-related gene content, suggesting populations from these different biomes experience similar Fe-selective conditions. This work supports an improved perspective that phytoplankton are shaped by more nuanced Fe niches in the oceans than previously implied from mostly binary comparisons of low- versus high-Fe habitats and populations.

Parallel phylogeography of Prochlorococcus and Synechococcus.

The globally abundant marine Cyanobacteria Prochlorococcus and Synechococcus share many physiological traits but presumably have different evolutionary histories and associated phylogeography. In Prochlorococcus, there is a clear phylogenetic hierarchy of ecotypes, whereas multiple Synechococcus clades have overlapping physiologies and environmental distributions. However, microbial traits are associated with different phylogenetic depths. Using this principle, we reclassified diversity at different phylogenetic levels and compared the phylogeography. We sequenced the genetic diversity of Prochlorococcus and Synechococcus from 339 samples across the tropical Pacific Ocean and North Atlantic Ocean using a highly variable phylogenetic marker gene (rpoC1). We observed clear parallel niche distributions of ecotypes leading to high Pianka's Index values driven by distinct shifts at two transition points. The first transition point at 6°N distinguished ecotypes adapted to warm waters but separated by macronutrient content. At 39°N, ecotypes adapted to warm, low macronutrient vs. colder, high macronutrient waters shifted. Finally, we detected parallel vertical and regional single-nucleotide polymorphism microdiversity within clades from both Prochlorococcus and Synechococcus, suggesting uniquely adapted populations at very specific depths, as well as between the Atlantic and Pacific Oceans. Overall, this study demonstrates that Prochlorococcus and Synechococcus have shared phylogenetic organization of traits and associated phylogeography.

Biophysical clocks face a trade-off between internal and external noise resistance.

Many organisms use free running circadian clocks to anticipate the day night cycle. However, others organisms use simple stimulus-response strategies ('hourglass clocks') and it is not clear when such strategies are sufficient or even preferable to free running clocks. Here, we find that free running clocks, such as those found in the cyanobacterium Synechococcus elongatus and humans, can efficiently project out light intensity fluctuations due to weather patterns ('external noise') by exploiting their limit cycle attractor. However, such limit cycles are necessarily vulnerable to 'internal noise'. Hence, at sufficiently high internal noise, point attractor-based 'hourglass' clocks, such as those found in a smaller cyanobacterium with low protein copy number, Prochlorococcus marinus, can outperform free running clocks. By interpolating between these two regimes in a diverse range of oscillators drawn from across biology, we demonstrate biochemical clock architectures that are best suited to different relative strengths of external and internal noise.

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.

"Pomacytosis"-Semi-extracellular phagocytosis of cyanobacteria by the smallest marine algae.

The smallest algae, less than 3 µm in diameter, are the most abundant eukaryotes of the World Ocean. Their feeding on planktonic bacteria of similar size is globally important but physically enigmatic. Tiny algal cells tightly packed with the voluminous chloroplasts, nucleus, and mitochondria appear to have insufficient organelle-free space for prey internalization. Here, we present the first direct observations of how the 1.3-µm algae, which are only 1.6 times bigger in diameter than their prey, hold individual Prochlorococcus cells in their open hemispheric cytostomes. We explain this semi-extracellular phagocytosis by the cell size limitation of the predatory alga, identified as the Braarudosphaera haptophyte with a nitrogen (N2)-fixing endosymbiont. Because the observed semi-extracellular phagocytosis differs from all other types of protistan phagocytosis, we propose to name it "pomacytosis" (from the Greek πώµα for "plug").

Three-Dimensional Superresolution Imaging of the FtsZ Ring during Cell Division of the Cyanobacterium Prochlorococcus.

Superresolution imaging has revealed subcellular structures and protein interactions in many organisms. However, superresolution microscopy with lateral resolution better than 100 nm has not been achieved in photosynthetic cells due to the interference of a high-autofluorescence background. Here, we developed a photobleaching method to effectively reduce the autofluorescence of cyanobacterial and plant cells. We achieved lateral resolution of ~10 nm with stochastic optical reconstruction microscopy (STORM) in the sphere-shaped cyanobacterium Prochlorococcus and the flowering plant Arabidopsis thaliana During the cell cycle of Prochlorococcus, we characterized the three-dimensional (3D) organization of the cell division protein FtsZ, which forms a ring structure at the division site and is important for cytokinesis of bacteria and chloroplasts. Although the FtsZ ring assembly process in rod-shaped bacteria has been studied extensively, it has rarely been studied in sphere-shaped bacteria. Similarly to rod-shaped bacteria, our results with Prochlorococcus also showed the assembly of FtsZ clusters into incomplete rings and then complete rings during cell division. Differently from rod-shaped bacteria, the FtsZ ring diameter was not found to decrease during Prochlorococcus cell division. We also discovered a novel double-Z-ring structure, which may be the Z rings of two daughter cells in a predivisional mother cell. Our results showed a quantitative picture of the in vivo Z ring organization of sphere-shaped bacteria.IMPORTANCE Superresolution microscopy has not been widely used to study photosynthetic cells due to their high-autofluorescence background. Here, we developed a photobleaching method to reduce the autofluorescence of cyanobacteria and plant cells. After photobleaching, we performed superresolution imaging in the cyanobacterium Prochlorococcus and the flowering plant Arabidopsis thaliana with ~10-nm resolution, which is the highest resolution in a photosynthetic cell. With this method, we characterized the 3D organization of the cell division protein FtsZ in Prochlorococcus We found that the morphological variation of the FtsZ ring during cell division of the sphere-shaped cyanobacterium Prochlorococcus is similar but not identical to that of rod-shaped bacteria. Our method might also be applicable to other photosynthetic organisms.

Diel cycling and long-term persistence of viruses in the ocean's euphotic zone.

Viruses are fundamental components of marine microbial communities that significantly influence oceanic productivity, biogeochemistry, and ecosystem processes. Despite their importance, the temporal activities and dynamics of viral assemblages in natural settings remain largely unexplored. Here we report the transcriptional activities and variability of dominant dsDNA viruses in the open ocean's euphotic zone over daily and seasonal timescales. While dsDNA viruses exhibited some fluctuation in abundance in both cellular and viral size fractions, the viral assemblage was remarkably stable, with the most abundant viral types persisting over many days. More extended time series indicated that long-term persistence (>1 y) was the rule for most dsDNA viruses observed, suggesting that both core viral genomes as well as viral community structure were conserved over interannual periods. Viral gene transcription in host cell assemblages revealed diel cycling among many different viral types. Most notably, an afternoon peak in cyanophage transcriptional activity coincided with a peak in Prochlorococcus DNA replication, indicating coordinated diurnal coupling of virus and host reproduction. In aggregate, our analyses suggested a tightly synchronized diel coupling of viral and cellular replication cycles in both photoautotrophic and heterotrophic bacterial hosts. A surprising consequence of these findings is that diel cycles in the ocean's photic zone appear to be universal organizing principles that shape ecosystem dynamics, ecological interactions, and biogeochemical cycling of both cellular and acellular community components.

Excess of non-conservative amino acid changes in marine bacterioplankton lineages with reduced genomes.

Surface ocean waters are dominated by planktonic bacterial lineages with highly reduced genomes. The best examples are the cyanobacterial genus Prochlorococcus, the alphaproteobacterial clade SAR11 and the gammaproteobacterial clade SAR86, which together represent over 50% of the cells in surface oceans. Several studies have identified signatures of selection on these lineages in today's ocean and have postulated selection as the primary force throughout their evolutionary history. However, massive loss of genomic DNA in these lineages often occurred in the distant past, and the selective pressures underlying these ancient events have not been assessed. Here, we probe ancient selective pressures by computing %GC-corrected rates of conservative and radical nonsynonymous nucleotide substitutions. Surprisingly, we found an excess of radical changes in several of these lineages in comparison to their relatives with larger genomes. Furthermore, analyses of allelic genome sequences of several populations within these lineages consistently supported that radical replacements are more likely to be deleterious than conservative changes. Our results suggest coincidence of massive genomic DNA losses and increased power of genetic drift, but we also suggest that additional evidence independent of the nucleotide substitution analyses is needed to support a primary role of genetic drift driving ancient genome reduction of marine bacterioplankton lineages.

Prochlorococcus.

Move over plants-make way for tiny Prochlorococcus, the smallest and most abundant photosynthetic cell on earth! Penny Chisholm tells us all about this powerhouse marine bacterium.

Distribution and diversity of marine picocyanobacteria community: Targeting of Prochlorococcus ecotypes in winter conditions (southern Adriatic Sea).

Adriatic, the northernmost part of the Mediterranean Sea, due its oligotrophy, topography, and hydrology dynamics, and complex circulation patterns, was suggested as an important study site for rapid climatology impacts. Its southern part is mainly oligotrophic and dominated by picophytoplankton, with cyanobacteria as main representatives. Diversity and distribution patterns of different Prochlorococcus ecotypes were investigated by molecular tools and flow cytometry during the winter convection event in the southern Adriatic (BIOTA winter cruise; February/March 2015). Phylogenetic diversity based on clone libraries of the 16S-23S ribosomal DNA ITS region, as well as flow cytometry (histograms of red fluorescence), indicated presence of 2 different Prochlorococcus in the Adriatic. HLI, as a typical clade for Mediterranean Sea, was likewise found to be dominant Prochlorococcus in the Adriatic, followed by less abundant LLI clade. In addition, Prochlorococcus were found to co-occur with diverse Synechococcus population (53% and 47% of obtained ITS sequences, respectively). Different Prochlorococcus ecotypes had similar patterns of vertical distribution, predominantly occupying upper 100m depth layer, but their distribution was clearly affected by the heterogeneity of hydrological conditions, nitrogen concentration and temperature along vertical and horizontal sampling points. Different studies pointed out that, as a consequence of climate changes, serious alteration of biological and ecological patterns are already taking place Therefore, understanding of the distribution and abundance of picophytoplankton in Adriatic, being still limited, is much needed baseline for predicting possible biogeochemical impact of future environmental changes.

Inhibition by ultraviolet and photosynthetically available radiation lowers model estimates of depth-integrated picophytoplankton photosynthesis: global predictions for Prochlorococcus and Synechococcus.

Phytoplankton photosynthesis is often inhibited by ultraviolet (UV) and intense photosynthetically available radiation (PAR), but the effects on ocean productivity have received little consideration aside from polar areas subject to periodic enhanced UV-B due to depletion of stratospheric ozone. A more comprehensive assessment is important for understanding the contribution of phytoplankton production to the global carbon budget, present and future. Here, we consider responses in the temperate and tropical mid-ocean regions typically dominated by picophytoplankton including the prokaryotic lineages, Prochlorococcus and Synechococcus. Spectral models of photosynthetic response for each lineage were constructed using model strains cultured at different growth irradiances and temperatures. In the model, inhibition becomes more severe once exposure exceeds a threshold (Emax ) related to repair capacity. Model parameters are presented for Prochlorococcus adding to those previously presented for Synechococcus. The models were applied to estimate midday, water column photosynthesis based on an atmospheric model of spectral radiation, satellite-derived spectral water transparency and temperature. Based on a global survey of inhibitory exposure severity, a full-latitude section of the mid-Pacific and near-equatorial region of the east Pacific were identified as representative regions for prediction of responses over the entire water column. Comparing predictions integrated over the water column including versus excluding inhibition, production was 7-28% lower due to inhibition depending on strain and site conditions. Inhibition was consistently greater for Prochlorococcus compared to two strains of Synechococcus. Considering only the surface mixed layer, production was inhibited 7-73%. On average, including inhibition lowered estimates of midday productivity around 20% for the modeled region of the Pacific with UV accounting for two-thirds of the reduction. In contrast, most other productivity models either ignore inhibition or only include PAR inhibition. Incorporation of Emax model responses into an existing spectral model of depth-integrated, daily production will enable efficient global predictions of picophytoplankton productivity including inhibition.