Temporal and spatial diversity and abundance of cryptophytes in San Diego coastal waters.

Cryptophytes (class Cryptophyceae) are bi-flagellated eukaryotic protists with mixed nutritional modes and cosmopolitan distribution in aquatic environments. Despite their ubiquitous presence, their molecular diversity is understudied in coastal waters. Weekly 18S rRNA gene amplicon sequencing at the Scripps Institution of Oceanography pier (La Jolla, California) in 2016 revealed 16 unique cryptophyte amplicon sequence variants (ASVs), with two dominant "clade 4" ASVs. The diversity of cryptophytes was lower than what is often seen in other phytoplankton taxa. One ASV represented a known Synechococcus grazer, while the other one appeared not to have cultured representatives and an unknown potential for mixotrophy. These two dominant ASVs were negatively correlated, suggesting possible niche differentiation. The cryptophyte population in nearby San Diego Bay was surveyed in 2019 and showed the increasing dominance of a different clade 4 ASV toward the back of the bay where conditions are warmer, saltier, and shallower relative to other areas in the bay. An ASV representing a potentially chromatically acclimating cryptophyte species also suggested that San Diego Bay exerts differing ecological selection pressures than nearby coastal waters. Cryptophyte and Synechococcus cell abundance at the SIO Pier from 2011 to 2017 showed that cryptophytes were consistently present and had a significant correlation with Synechococcus abundance, but no detectable seasonality. The demonstrated mixotrophy of some cryptophytes suggests that grazing on these and perhaps other bacteria is important for their ecological success. Using several assumptions, we calculated that cryptophytes could consume up to 44% (average 6%) of the Synechococcus population per day. This implies that cryptophytes could significantly influence Synechococcus abundance.

Diverse Microbial Hot Spring Mat Communities at Black Canyon of the Colorado River.

The thermophilic microbial mat communities at hot springs in the Black Canyon of the Colorado River, thought to harbor the protistan human pathogen Naegleria fowleri, were surveyed using both culture-independent and -dependent methods to further understand the ecology of these hot spring microbiomes. Originating from Lake Mead source water, seven spring sites were sampled, varying in temperature from 25 to 55 °C. Amplicon-based high-throughput sequencing of twelve samples using 16S rRNA primers (hypervariable V4 region) revealed that most mats are dominated by cyanobacterial taxa, some but not all similar to those dominating the mats at other studied hot spring systems. 18S rRNA amplicon sequencing (V9 region) demonstrated a diverse community of protists and other eukaryotes including a highly abundant amoebal sequence related to Echinamoeba thermarum. Additional taxonomic and diversity metric analyses using near full-length 16S and 18S rRNA gene sequencing allowed a higher sequence-based resolution of the community. The mat sequence data suggest a major diversification of the cyanobacterial orders Leptolyngbyales, as well as microdiversity among several cyanobacterial taxa. Cyanobacterial isolates included some representatives of ecologically abundant taxa. A Spearman correlation analysis of short-read amplicon sequencing data supported the co-occurrences of populations of cyanobacteria, chloroflexi, and bacteroidetes providing evidence of common microbial co-occurrences across the Black Canyon hot springs.

Reaction of O2 with a diiron protein generates a mixed-valent Fe2+/Fe3+ center and peroxide.

The gene encoding the cyanobacterial ferritin SynFtn is up-regulated in response to copper stress. Here, we show that, while SynFtn does not interact directly with copper, it is highly unusual in several ways. First, its catalytic diiron ferroxidase center is unlike those of all other characterized prokaryotic ferritins and instead resembles an animal H-chain ferritin center. Second, as demonstrated by kinetic, spectroscopic, and high-resolution X-ray crystallographic data, reaction of O2 with the di-Fe2+ center results in a direct, one-electron oxidation to a mixed-valent Fe2+/Fe3+ form. Iron-O2 chemistry of this type is currently unknown among the growing family of proteins that bind a diiron site within a four α-helical bundle in general and ferritins in particular. The mixed-valent form, which slowly oxidized to the more usual di-Fe3+ form, is an intermediate that is continually generated during mineralization. Peroxide, rather than superoxide, is shown to be the product of O2 reduction, implying that ferroxidase centers function in pairs via long-range electron transfer through the protein resulting in reduction of O2 bound at only one of the centers. We show that electron transfer is mediated by the transient formation of a radical on Tyr40, which lies ∼4 Å from the diiron center. As well as demonstrating an expansion of the iron-O2 chemistry known to occur in nature, these data are also highly relevant to the question of whether all ferritins mineralize iron via a common mechanism, providing unequivocal proof that they do not.

Spatial and temporal variations in Synechococcus microdiversity in the Southern California coastal ecosystem.

The Synechococcus cyanobacterial population at the Scripps Institution of Oceanography pier in La Jolla, CA, shows large increases in abundance, typically in the spring and summer followed, by rapid declines within weeks. Here we used amplicon sequencing of the ribosomal RNA internal transcribed spacer region to examine the microdiversity within this cyanobacterial genus during these blooms as well as further offshore in the Southern California coastal ecosystem (CCE). These analyses revealed numerous Synechococcus amplicon sequence variants (ASVs) and that clade and ASV composition can change over the course of blooms. We also found that a large bloom in August 2016 was highly anomalous both in its overall Synechococcus abundance and in terms of the presence of normally oligotrophic Synechococcus clade II. The dominant ASVs at the pier were found further offshore and in the California Current, but we did observe more oligotrophic ASVs and clades along with depth variation in Synechococcus diversity. We also observed that the dominant sequence variant switched during the peak of multiple Synechococcus blooms, with this switch occurring in multiple clades, but we present initial evidence that this apparent ASV switch is a physiological response rather than a change in the dominant population.

Relating sinking and suspended microbial communities in the California Current Ecosystem: digestion resistance and the contributions of phytoplankton taxa to export.

We used 16S, 18S, plastid and internal transcribed spacer (for Synechococcus strains) sequencing to quantify relative microbial abundances in water-column samples and on sediment-trap-collected particles across an environmental gradient in the California Current Ecosystem (CCE) spanning a > 60-fold range of surface chlorophyll. Most mixed-layer dominant eukaryotes and prokaryotes were consistently underrepresented on sinking particles. Diatoms were the only phototrophic taxa consistently overrepresented. Even within this class, however, one genus (Thalassiosira) was a particle-enriched dominant, while a similarly abundant species was poorly represented. Synechococcus was significantly enriched on sinking particles at only one of four sites, but clade I was disproportionately abundant on sinking particles throughout the region compared with clade IV, the euphotic-zone co-dominant. The most abundant microbes on particles across the CCE were organisms with distributional maxima close to the sediment-trap depth (rhizarians), microbes associated with metazoans or sinking particles as a nutritional habitat (certain alveolates, Gammaproteobacteria) and organisms that resist digestive degradation of their DNA (Thalassiosira, Synechococcus). For assessing taxon contributions of phytoplankton to carbon export, our results highlight the need for sequence-based quantitative approaches that can be used to integrate euphotic-zone abundances, compute rates and account for taxon differences in preservation of sequence markers through trophic processing.

Screening and characterization of polyhydroxyalkanoate granules, and phylogenetic analysis of polyhydroxyalkanoate synthase gene PhaC in cyanobacteria.

Using Nile Red and BODIPY 493/503 dye-staining and fluorescence microscopy, twenty cyanobacterial strains, including ten commercially available strains and ten environmental isolates from estuaries, freshwater ponds, and lagoons, were screened for the accumulation of ecologically important and potentially biotechnologically significant carbon storage granules such as polyhydroxyalkanoates (PHA). Dye-staining granules were observed in six strains. Three Synechocystis, spp. strains WHSYN, LSNM, and CGF-1, and a Phormidium-like sp. CGFILA were isolated from environmental sources and found to produce granules of polyhydroxyalkanoate (PHA) according to PHA synthase gene (phaC) PCR screening and 1 H NMR analyses. The environmental isolate, Nodularia sp. Las Olas and commercially available Phormidium cf. iriguum CCALA 759 displayed granules but screened negative for PHA according to phaC PCR and 1 H NMR analyses. Partial polyhydroxyalkanoate synthase subunit C (phaC) and 16S rRNA gene sequences obtained from the PHA-accumulating strains and analyzed alongside publicly available phaC, phaE, 16S rRNA, and 23S rRNA data help in understanding the distribution and evolutionary history of PHA biosynthesis within the phylum Cyanobacteria. The data show that the presence of phaC is highly conserved within the genus Synechocystis, and present in at least one isolate of Phormidium. Maximum likelihood analyses and cophylogenetic modeling of PHA synthase gene sequences provide evidence of a recent horizontal gene transfer event between distant genera of cyanobacteria related to Pleurocapsa sp. PCC 7327 and Phormidium-like sp. CGFILA. These findings will help guide additional screening for PHA producers, and may explain why some Phormidium species produce PHAs, while others do not.

Copper toxicity response influences mesotrophic Synechococcus community structure.

Picocyanobacteria from the genus Synechococcus are ubiquitous in ocean waters. Their phylogenetic and genomic diversity suggests ecological niche differentiation, but the selective forces influencing this are not well defined. Marine picocyanobacteria are sensitive to Cu toxicity, so adaptations to this stress could represent a selective force within, and between, 'species', also known as clades. Here, we compared Cu stress responses in cultures and natural populations of marine Synechococcus from two co-occurring major mesotrophic clades (I and IV). Using custom microarrays and proteomics to characterize expression responses to Cu in the lab and field, we found evidence for a general stress regulon in marine Synechococcus. However, the two clades also exhibited distinct responses to copper. The Clade I representative induced expression of genomic island genes in cultures and Southern California Bight populations, while the Clade IV representative downregulated Fe-limitation proteins. Copper incubation experiments suggest that Clade IV populations may harbour stress-tolerant subgroups, and thus fitness tradeoffs may govern Cu-tolerant strain distributions. This work demonstrates that Synechococcus has distinct adaptive strategies to deal with Cu toxicity at both the clade and subclade level, implying that metal toxicity and stress response adaptations represent an important selective force for influencing diversity within marine Synechococcus populations.

Broad-host-range vector system for synthetic biology and biotechnology in cyanobacteria.

Inspired by the developments of synthetic biology and the need for improved genetic tools to exploit cyanobacteria for the production of renewable bioproducts, we developed a versatile platform for the construction of broad-host-range vector systems. This platform includes the following features: (i) an efficient assembly strategy in which modules released from 3 to 4 donor plasmids or produced by polymerase chain reaction are assembled by isothermal assembly guided by short GC-rich overlap sequences. (ii) A growing library of molecular devices categorized in three major groups: (a) replication and chromosomal integration; (b) antibiotic resistance; (c) functional modules. These modules can be assembled in different combinations to construct a variety of autonomously replicating plasmids and suicide plasmids for gene knockout and knockin. (iii) A web service, the CYANO-VECTOR assembly portal, which was built to organize the various modules, facilitate the in silico construction of plasmids, and encourage the use of this system. This work also resulted in the construction of an improved broad-host-range replicon derived from RSF1010, which replicates in several phylogenetically distinct strains including a new experimental model strain Synechocystis sp. WHSyn, and the characterization of nine antibiotic cassettes, four reporter genes, four promoters, and a ribozyme-based insulator in several diverse cyanobacterial strains.

Role of a microcin-C-like biosynthetic gene cluster in allelopathic interactions in marine Synechococcus.

Competition between phytoplankton species for nutrients and light has been studied for many years, but allelopathic interactions between them have been more difficult to characterize. We used liquid and plate assays to determine whether these interactions occur between marine unicellular cyanobacteria of the genus Synechococcus. We have found a clear growth impairment of Synechococcus sp. CC9311 and Synechococcus sp. WH8102 when they are cultured in the presence of Synechococcus sp. CC9605. The genome of CC9605 contains a region showing homology to genes of the Escherichia coli Microcin C (McC) biosynthetic pathway. McC is a ribosome-synthesized peptide that inhibits translation in susceptible strains. We show that the CC9605 McC gene cluster is expressed and that three genes (mccD, mccA, and mccB) are further induced by coculture with CC9311. CC9605 was resistant to McC purified from E. coli, whereas strains CC9311 and WH8102 were sensitive. Cloning the CC9605 McC biosynthetic gene cluster into sensitive CC9311 led this strain to become resistant to both purified E. coli McC and Synechococcus sp. CC9605. A CC9605 mutant lacking mccA1, mccA2, and the N-terminal domain of mccB did not inhibit CC9311 growth, whereas the inhibition of WH8102 was reduced. Our results suggest that an McC-like molecule is involved in the allelopathic interactions with CC9605.

Bringing the ocean into the laboratory to probe the chemical complexity of sea spray aerosol.

The production, size, and chemical composition of sea spray aerosol (SSA) particles strongly depend on seawater chemistry, which is controlled by physical, chemical, and biological processes. Despite decades of studies in marine environments, a direct relationship has yet to be established between ocean biology and the physicochemical properties of SSA. The ability to establish such relationships is hindered by the fact that SSA measurements are typically dominated by overwhelming background aerosol concentrations even in remote marine environments. Herein, we describe a newly developed approach for reproducing the chemical complexity of SSA in a laboratory setting, comprising a unique ocean-atmosphere facility equipped with actual breaking waves. A mesocosm experiment was performed in natural seawater, using controlled phytoplankton and heterotrophic bacteria concentrations, which showed SSA size and chemical mixing state are acutely sensitive to the aerosol production mechanism, as well as to the type of biological species present. The largest reduction in the hygroscopicity of SSA occurred as heterotrophic bacteria concentrations increased, whereas phytoplankton and chlorophyll-a concentrations decreased, directly corresponding to a change in mixing state in the smallest (60-180 nm) size range. Using this newly developed approach to generate realistic SSA, systematic studies can now be performed to advance our fundamental understanding of the impact of ocean biology on SSA chemical mixing state, heterogeneous reactivity, and the resulting climate-relevant properties.

Genome of the halotolerant green alga Picochlorum sp. reveals strategies for thriving under fluctuating environmental conditions.

An expected outcome of climate change is intensification of the global water cycle, which magnifies surface water fluxes, and consequently alters salinity patterns. It is therefore important to understand the adaptations and limits of microalgae to survive changing salinities. To this end, we sequenced the 13.5 Mbp genome of the halotolerant green alga Picochlorum SENEW3 (SE3) that was isolated from a brackish water pond subject to large seasonal salinity fluctuations. Picochlorum SE3 encodes 7367 genes, making it one of the smallest and most gene dense eukaryotic genomes known. Comparison with the pico-prasinophyte Ostreococcus tauri, a species with a limited range of salt tolerance, reveals the enrichment of transporters putatively involved in the salt stress response in Picochlorum SE3. Analysis of cultures and the protein complement highlight the metabolic flexibility of Picochlorum SE3 that encodes genes involved in urea metabolism, acetate assimilation and fermentation, acetoin production and glucose uptake, many of which form functional gene clusters. Twenty-four cases of horizontal gene transfer from bacterial sources were found in Picochlorum SE3 with these genes involved in stress adaptation including osmolyte production and growth promotion. Our results identify Picochlorum SE3 as a model for understanding microalgal adaptation to stressful, fluctuating environments.

Comparison of the seasonal variations of Synechococcus assemblage structures in estuarine waters and coastal waters of Hong Kong.

Seasonal variation in the phylogenetic composition of Synechococcus assemblages in estuarine and coastal waters of Hong Kong was examined through pyrosequencing of the rpoC1 gene. Sixteen samples were collected in 2009 from two stations representing estuarine and ocean-influenced coastal waters, respectively. Synechococcus abundance in coastal waters gradually increased from 3.6 × 10(3) cells ml(-1) in March, reaching a peak value of 5.7 × 10(5) cells ml(-1) in July, and then gradually decreased to 9.3 × 10(3) cells ml(-1) in December. The changes in Synechococcus abundance in estuarine waters followed a pattern similar to that in coastal waters, whereas its composition shifted from being dominated by phycoerythrin-rich (PE-type) strains in winter to phycocyanin-only (PC-type) strains in summer owing to the increase in freshwater discharge from the Pearl River and higher water temperature. The high abundance of PC-type Synechococcus was composed of subcluster 5.2 marine Synechococcus, freshwater Synechococcus (F-PC), and Cyanobium. The Synechococcus assemblage in the coastal waters, on the other hand, was dominated by marine PE-type Synechococcus, with subcluster 5.1 clades II and VI as the major lineages from April to September, when the summer monsoon prevailed. Besides these two clades, clade III cooccurred with clade V at relatively high abundance in summer. During winter, the Synechococcus assemblage compositions at the two sites were similar and were dominated by subcluster 5.1 clades II and IX and an undescribed clade (represented by Synechococcus sp. strain miyav). Clade IX Synechococcus was a relatively ubiquitous PE-type Synechococcus found at both sites, and our study demonstrates that some strains of the clade have the ability to deal with large variation of salinity in subtropical estuarine environments. Our study suggests that changes in seawater temperature and salinity caused by the seasonal variation of monsoonal forcing are two major determinants of the community composition and abundance of Synechococcus assemblages in Hong Kong waters.

Microalgal assemblages in a poikilohaline pond.

Microalgal strains for algal biofuels production in outdoor ponds will need to have high net growth rates under diverse environmental conditions. A small, variable salinity pond in the San Elijo Lagoon estuary in southern California was chosen to serve as a model pond due to its routinely high chlorophyll content. Profiles of microalgal assemblages from water samples collected from April 2011 to January 2012 were obtained by constructing 18S rDNA environmental clone libraries. Pond assemblages were found to be dominated by green algae Picochlorum sp. and Picocystis sp. throughout the year. Pigment analysis suggested that the two species contributed most of the chlorophyll a of the pond, which ranged from 21.9 to 664.3 µg · L(-1) with the Picocystis contribution increasing at higher salinities. However, changes of temperature, salinity or irradiance may have enabled a bloom of the diatom Chaetoceros sp. in June 2011. Isolates of these microalgae were obtained and their growth rates characterized as a function of temperature and salinity. Chaetoceros sp. had the highest growth rate over the temperature test range while it showed the most sensitivity to high salinity. All three strains showed the presence of lipid bodies during nitrogen starvation, suggesting they have potential as future biofuels strains.

Chromosome-scale assembly of the streamlined picoeukaryote Picochlorum sp. SENEW3 genome reveals Rabl-like chromatin structure and potential for C4 photosynthesis.

Genome sequencing and assembly of the photosynthetic picoeukaryotic Picochlorum sp. SENEW3 revealed a compact genome with a reduced gene set, few repetitive sequences, and an organized Rabl-like chromatin structure. Hi-C chromosome conformation capture revealed evidence of possible chromosomal translocations, as well as putative centromere locations. Maintenance of a relatively few selenoproteins, as compared to similarly sized marine picoprasinophytes Mamiellales, and broad halotolerance compared to others in Trebouxiophyceae, suggests evolutionary adaptation to variable salinity environments. Such adaptation may have driven size and genome minimization and have been enabled by the retention of a high number of membrane transporters. Identification of required pathway genes for both CAM and C4 photosynthetic carbon fixation, known to exist in the marine mamiellale pico-prasinophytes and seaweed Ulva, but few other chlorophyte species, further highlights the unique adaptations of this robust alga. This high-quality assembly provides a significant advance in the resources available for genomic investigations of this and other photosynthetic picoeukaryotes.

Diversity and putative interactions of parasitic alveolates belonging to Syndiniales at a coastal Pacific site.

The dinoflagellate lineage Syndiniales currently consists entirely of parasitic species that fall into five well-supported clades. Environmental sequencing studies worldwide have found an abundance of Syndiniales in a variety of marine ecosystems, but very little is known about the majority of Syndiniales species including two entire clades which have only been observed in sequence data. Syndiniales are known to have a wide range of hosts, but only a few dozen interactions have been confirmed through observation of actual infections. Here, we describe the diversity of Syndiniales found at the Scripps Institution of Oceanography pier over the course of a year based on 18S sequencing. We find Syndiniales to be the most species (amplicon sequence variant)-rich taxonomic group and for its members to be present and abundant throughout the year. We used several analytical techniques to identify potential parasite-host interactions which we were then able to visualize over time. Using mock communities and size fractionation of seawater, we suggest that the majority of Syndiniales sequences that are found in environmental studies belong to the free-living dinospore stage rather than representing active infections.

Analysis of two marine metagenomes reveals the diversity of plasmids in oceanic environments.

Plasmid diversity is still poorly understood in pelagic marine environments. Metagenomic approaches have the potential to reveal the genetic diversity of microbes actually present in an environment and the contribution of mobile genetic elements such as plasmids. By searching metagenomic datasets from flow cytometry-sorted coastal California seawater samples dominated by cyanobacteria (SynMeta) and from the Global Ocean Survey (GOS) putative marine plasmid sequences were identified as well as their possible hosts in the same samples. Based on conserved plasmid replication protein sequences predicted from the SynMeta metagenomes, PCR primers were designed for amplification of one plasmid family and used to confirm that metagenomic contigs of this family were derived from plasmids. These results suggest that the majority of plasmids in SynMeta metagenomes were small and cryptic, encoding mostly their own replication proteins. In contrast, probable plasmid sequences identified in the GOS dataset showed more complexity, consistent with a much more diverse microbial population, and included genes involved in plasmid transfer, mobilization, stability and partitioning. Phylogenetic trees were constructed based on common replication protein functional domains and, even within one replication domain family, substantial diversity was found within and between different samples. However, some replication protein domain families appear to be rare in the marine environment.

Variability in protist grazing and growth on different marine Synechococcus isolates.

Grazing mortality of the marine phytoplankton Synechococcus is dominated by planktonic protists, yet rates of consumption and factors regulating grazer-Synechococcus interactions are poorly understood. One aspect of predator-prey interactions for which little is known are the mechanisms by which Synechococcus avoids or resists predation and, in turn, how this relates to the ability of Synechococcus to support growth of protist grazer populations. Grazing experiments conducted with the raptorial dinoflagellate Oxyrrhis marina and phylogenetically diverse Synechococcus isolates (strains WH8102, CC9605, CC9311, and CC9902) revealed marked differences in grazing rates-specifically that WH8102 was grazed at significantly lower rates than all other isolates. Additional experiments using the heterotrophic nanoflagellate Goniomonas pacifica and the filter-feeding tintinnid ciliate Eutintinnis sp. revealed that this pattern in grazing susceptibility among the isolates transcended feeding guilds and grazer taxon. Synechococcus cell size, elemental ratios, and motility were not able to explain differences in grazing rates, indicating that other features play a primary role in grazing resistance. Growth of heterotrophic protists was poorly coupled to prey ingestion and was influenced by the strain of Synechococcus being consumed. Although Synechococcus was generally a poor-quality food source, it tended to support higher growth and survival of G. pacifica and O. marina relative to Eutintinnis sp., indicating that suitability of Synechococcus varies among grazer taxa and may be a more suitable food source for the smaller protist grazers. This work has developed tractable model systems for further studies of grazer-Synechococcus interactions in marine microbial food webs.

Learning to read the oceans genomics of marine phytoplankton.

The phytoplankton are key members of marine ecosystems, generating about half of global primary productivity, supporting valuable fisheries and regulating global biogeochemical cycles. Marine phytoplankton are phylogenetically diverse and are comprised of both prokaryotic and eukaryotic species. In the last decade, new insights have been gained into the ecology and evolution of these important organisms through whole genome sequencing projects and more recently, through both transcriptomics and targeted metagenomics approaches. Sequenced genomes of cyanobacteria are generally small, ranging in size from 1.8 to 9 million base pairs (Mbp). Eukaryotic genomes, in general, have a much larger size range and those that have been sequenced range from 12 to 57 Mbp. Whole genome sequencing projects have revealed key features of the evolutionary history of marine phytoplankton, their varied responses to environmental stress, their ability to scavenge and store nutrients and their unique ability to form elaborate cellular coverings. We have begun to learn how to read the 'language' of marine phytoplankton, as written in their DNA. Here, we review the ecological and evolutionary insights gained from whole genome sequencing projects, illustrate how these genomes are yielding information on marine natural products and informing nanotechnology as well as make suggestions for future directions in the field of marine phytoplankton genomics.

Computational prediction of the osmoregulation network in Synechococcus sp. WH8102.

BACKGROUND: Osmotic stress is caused by sudden changes in the impermeable solute concentration around a cell, which induces instantaneous water flow in or out of the cell to balance the concentration. Very little is known about the detailed response mechanism to osmotic stress in marine Synechococcus, one of the major oxygenic phototrophic cyanobacterial genera that contribute greatly to the global CO2 fixation. RESULTS: We present here a computational study of the osmoregulation network in response to hyperosmotic stress of Synechococcus sp strain WH8102 using comparative genome analyses and computational prediction. In this study, we identified the key transporters, synthetases, signal sensor proteins and transcriptional regulator proteins, and found experimentally that of these proteins, 15 genes showed significantly changed expression levels under a mild hyperosmotic stress. CONCLUSIONS: From the predicted network model, we have made a number of interesting observations about WH8102. Specifically, we found that (i) the organism likely uses glycine betaine as the major osmolyte, and others such as glucosylglycerol, glucosylglycerate, trehalose, sucrose and arginine as the minor osmolytes, making it efficient and adaptable to its changing environment; and (ii) sigma38, one of the seven types of sigma factors, probably serves as a global regulator coordinating the osmoregulation network and the other relevant networks.

Dynamics of marine bacterial and phytoplankton populations using multiplex liquid bead array technology.

Heterotrophic bacteria and phytoplankton dominate the biomass and play major roles in the biogeochemical cycles of the surface ocean. Here, we designed and tested a fast, high-throughput and multiplexed hybridization-based assay to detect populations of marine heterotrophic bacteria and phytoplankton based on their small subunit ribosomal DNA sequences. The assay is based on established liquid bead array technology, an approach that is gaining acceptance in biomedical research but remains underutilized in ecology. End-labelled PCR products are hybridized to taxon-specific oligonucleotide probes attached to fluorescently coded beads followed by flow cytometric detection. We used ribosomal DNA environmental clone libraries (a total of 450 clones) and cultured isolates to design and test 26 bacterial and 10 eukaryotic probes specific to various ribotypes and genera of heterotrophic bacteria and eukaryotic phytoplankton. Pure environmental clones or cultures were used as controls and demonstrated specificity of the probes to their target taxa. The quantitative nature of the assay was demonstrated by a significant relationship between the number of target molecules and fluorescence signal. Clone library sequencing and bead array fluorescence from the same sample provided consistent results. We then applied the assay to a 37-day time series of coastal surface seawater samples from the Southern California Bight to examine the temporal dynamics of microbial communities on the scale of days to weeks. As expected, several bacterial phylotypes were positively correlated with total bacterial abundances and chlorophyll a concentrations, but others were negatively correlated. Bacterial taxa belonging to the same broad taxonomic groups did not necessarily correlate with one another, confirming recent results suggesting that inferring ecological role from broad taxonomic identity may not always be accurate.