Constraining the oxygen requirements for modern microbial eukaryote diversity.

Eukaryotes originated prior to the establishment of modern marine oxygen (O2) levels. According to the body fossil and lipid biomarker records, modern (crown) microbial eukaryote lineages began diversifying in the ocean no later than ~800 Ma. While it has long been predicted that increasing atmospheric O2 levels facilitated the early diversification of microbial eukaryotes, the O2 levels needed to permit this diversification remain unconstrained. Using time-resolved geochemical parameter and gene sequence information from a model marine oxygen minimum zone spanning a range of dissolved O2 levels and redox states, we show that microbial eukaryote taxonomic richness and phylogenetic diversity remain the same until O2 declines to around 2 to 3% of present atmospheric levels, below which these diversity metrics become significantly reduced. Our observations suggest that increasing O2 would have only directly promoted early crown-eukaryote diversity if atmospheric O2 was below 2 to 3% of modern levels when crown-eukaryotes originated and then later met or surpassed this range as crown-eukaryotes diversified. If atmospheric O2 was already consistently at or above 2 to 3% of modern levels by the time that crown-eukaryotes originated, then the subsequent diversification of modern microbial eukaryotes was not directly driven by atmospheric oxygenation.

Testate amoebae (Arcellinida, Amoebozoa) community diversity in New England bogs and fens assessed through lineage-specific amplicon sequencing.

Testate amoebae (order Arcellinida) are abundant in freshwater ecosystems, including low pH bogs and fens. Within these environments, Arcellinida are considered top predators in microbial food webs and their tests are useful bioindicators of paleoclimatic changes and anthropogenic pollutants. Accurate species identifications and characterizations of diversity are important for studies of paleoclimate, microbial ecology, and environmental change; however, morphological species definitions mask cryptic diversity, which is a common phenomenon among microbial eukaryotes. Lineage-specific primers recently designed to target Arcellinida for amplicon sequencing successfully captured a poorly-described yet diverse fraction of the microbial eukaryotic community. Here, we leveraged the application of these newly-designed primers to survey the diversity of Arcellinida in four low-pH New England bogs and fens, investigating variation among bogs (2018) and then across seasons and habitats within two bogs (2019). Three OTUs represented 66% of Arcellinida reads obtained across all habitats surveyed. 103 additional OTUs were present in lower abundance with some OTUs detected in only one sampling location, suggesting habitat specificity. By establishing a baseline for Arcellinida diversity, we provide a foundation to monitor key taxa in habitats that are predicted to change with increasing anthropogenic pressure and rapid climate change.

Parasitic infections by Group II Syndiniales target selected dinoflagellate host populations within diverse protist assemblages in a model coastal pond.

Protists are integral to marine food webs and biogeochemical cycles; however, there is a paucity of data describing specific ecological niches for some of the most abundant taxa in marker gene libraries. Syndiniales are one such group, often representing the majority of sequence reads recovered from picoplankton samples across the global ocean. However, the prevalence and impacts of syndinian parasitism in marine environments remain unclear. We began to address these critical knowledge gaps by generating a high-resolution time series (March-October 2018) in a productive coastal pond. Seasonal shifts in protist populations, including parasitic Syndiniales, were documented during periods of higher primary productivity and increased summer temperature-driven stratification. Elevated concentrations of infected hosts and free-living parasite spores occurred at nearly monthly intervals in July, August, and September. We suggest intensifying stratification during this period correlated with the increased prevalence of dinoflagellates that were parasitized by Group II Syndiniales. Infections in some protist populations were comparable to previously reported large single-taxon dinoflagellate blooms. Infection dynamics in Salt Pond demonstrated the propagation of syndinian parasites through mixed protist assemblages and highlighted patterns of host/parasite interactions that better reflect many other marine environments where single taxon blooms are uncommon.

Meta-omics highlights the diversity, activity and adaptations of fungi in deep oceanic crust.

The lithified oceanic crust, lower crust gabbros in particular, has remained largely unexplored by microbiologists. Recently, evidence for heterogeneously distributed viable and transcriptionally active autotrophic and heterotrophic microbial populations within low-biomass communities was found down to 750 m below the seafloor at the Atlantis Bank Gabbro Massif, Indian Ocean. Here, we report on the diversity, activity and adaptations of fungal communities in the deep oceanic crust from ~10 to 780 mbsf by combining metabarcoding analyses with mid/high-throughput culturing approaches. Metabarcoding along with culturing indicate a low diversity of viable fungi, mostly affiliated to ubiquitous (terrestrial and aquatic environments) taxa. Ecophysiological analyses coupled with metatranscriptomics point to viable and transcriptionally active fungal populations engaged in cell division, translation, protein modifications and other vital cellular processes. Transcript data suggest possible adaptations for surviving in the nutrient-poor, lithified deep biosphere that include the recycling of organic matter. These active communities appear strongly influenced by the presence of cracks and veins in the rocks where fluids and resulting rock alteration create micro-niches.

Microbial Community Structure and Associations During a Marine Dinoflagellate Bloom.

Interactions between microorganisms and algae during bloom events significantly impacts their physiology, alters ambient chemistry, and shapes ecosystem diversity. The potential role these interactions have in bloom development and decline are also of particular interest given the ecosystem impacts of algal blooms. We hypothesized that microbial community structure and succession is linked to specific bloom stages, and reflects complex interactions among taxa comprising the phycosphere environment. This investigation used pyrosequencing and correlation approaches to assess patterns and associations among bacteria, archaea, and microeukaryotes during a spring bloom of the dinoflagellate Alexandrium catenella. Within the bacterial community, Gammaproteobacteria and Bacteroidetes were predominant during the initial bloom stage, while Alphaproteobacteria, Cyanobacteria, and Actinobacteria were the most abundant taxa present during bloom onset and termination. In the archaea biosphere, methanogenic members were present during the early bloom period while the majority of species identified in the late bloom stage were ammonia-oxidizing archaea and Halobacteriales. Dinoflagellates were the major eukaryotic group present during most stages of the bloom, whereas a mixed assemblage comprising diatoms, green-algae, rotifera, and other microzooplankton were present during bloom termination. Temperature and salinity were key environmental factors associated with changes in bacterial and archaeal community structure, respectively, whereas inorganic nitrogen and inorganic phosphate were associated with eukaryotic variation. The relative contribution of environmental parameters measured during the bloom to variability among samples was 35.3%. Interaction analysis showed that Maxillopoda, Spirotrichea, Dinoflagellata, and Halobacteria were keystone taxa within the positive-correlation network, while Halobacteria, Dictyochophyceae, Mamiellophyceae, and Gammaproteobacteria were the main contributors to the negative-correlation network. The positive and negative relationships were the primary drivers of mutualist and competitive interactions that impacted algal bloom fate, respectively. Functional predictions showed that blooms enhance microbial carbohydrate and energy metabolism, and alter the sulfur cycle. Our results suggest that microbial community structure is strongly linked to bloom progression, although specific drivers of community interactions and responses are not well understood. The importance of considering biotic interactions (e.g., competition, symbiosis, and predation) when investigating the link between microbial ecological behavior and an algal bloom's trajectory is also highlighted.

LSU rDNA based RFLP assays for the routine identification of Gambierdiscus species.

The Gambierdiscus genus is a group of benthic dinoflagellates commonly associated with ciguatera fish poisoning (CFP), which is generally found in tropical or sub-tropical regions around the world. Morphologically similar species within the genus can vary in toxicity; however, species identifications are difficult or sometimes impossible using light microscopy. DNA sequencing of ribosomal RNA genes (rDNA) is thus often used to identify and describe Gambierdiscus species and ribotypes, but the expense and time can be prohibitive for routine culture screening and/or large-scale monitoring programs. This study describes a restriction fragment length polymorphism (RFLP) typing method based on analysis of the large subunit rDNA that can successfully identify at least nine of the described Gambierdiscus species and two Fukuyoa species. The software programs DNAMAN 6.0 and Restriction Enzyme Picker were used to identify a set of restriction enzymes (SpeI, HpyCH4IV, and TaqαI) capable of distinguishing most of the known Gambierdiscus species for which DNA sequences were available. This assay was tested using in silico analysis and cultured isolates, and species identifications of isolates assigned by RFLP typing were confirmed by DNA sequencing. To verify the assay and assess intra-specific heterogeneity in RFLP patterns, identifications of 63 Gambierdiscus isolates comprising ten Gambierdiscus species, one ribotype, and two Fukuyoa species were confirmed using RFLP typing, and this method was subsequently employed in the routine identification of isolates collected from the Caribbean Sea. The RFLP assay presented here reduces the time and cost associated with morphological identification via scanning electron microscopy and/or DNA sequencing, and provides a phylogenetically sensitive method for routine Gambierdiscus species assignment.

Insights into the loss factors of phytoplankton blooms: The role of cell mortality in the decline of two inshore Alexandrium blooms.

While considerable effort has been devoted to understanding the factors regulating the development of phytoplankton blooms, the mechanisms leading to bloom decline and termination have received less attention. Grazing and sedimentation have been invoked as the main routes for the loss of phytoplankton biomass, and more recently, viral lysis, parasitism and programmed cell death (PCD) have been recognized as additional removal factors. Despite the importance of bloom declines to phytoplankton dynamics, the incidence and significance of various loss factors in regulating phytoplankton populations have not been widely characterized in natural blooms. To understand mechanisms controlling bloom decline, we studied two independent, inshore blooms of Alexandrium fundyense, paying special attention to cell mortality as a loss pathway. We observed increases in the number of dead cells with PCD features after the peak of both blooms, demonstrating a role for cell mortality in their terminations. In both blooms, sexual cyst formation appears to have been the dominant process leading to bloom termination, as both blooms were dominated by small-sized gamete cells near their peaks. Cell death and parasitism became more significant as sources of cell loss several days after the onset of bloom decline. Our findings show two distinct phases of bloom decline, characterized by sexual fusion as the initial dominant cell removal processes followed by elimination of remaining cells by cell death and parasitism.

Rapid growth and concerted sexual transitions by a bloom of the harmful dinoflagellate Alexandrium fundyense (Dinophyceae).

Transitions between life cycle stages by the harmful dinoflagellate Alexandrium fundyense are critical for the initiation and termination of its blooms. To quantify these transitions in a single population, an Imaging FlowCytobot (IFCB), was deployed in Salt Pond (Eastham, Massachusetts), a small, tidally flushed kettle pond that hosts near annual, localized A. fundyense blooms. Machine-based image classifiers differentiating A. fundyense life cycle stages were developed and results were compared to manually corrected IFCB samples, manual microscopy-based estimates of A. fundyense abundance, previously published data describing prevalence of the parasite Amoebophrya, and a continuous culture of A. fundyense infected with Amoebophrya. In Salt Pond, a development phase of sustained vegetative division lasted approximately 3 weeks and was followed by a rapid and near complete conversion to small, gamete cells. The gametic period (∼3 d) coincided with a spike in the frequency of fusing gametes (up to 5% of A. fundyense images) and was followed by a zygotic phase (∼4 d) during which cell sizes returned to their normal range but cell division and diel vertical migration ceased. Cell division during bloom development was strongly phased, enabling estimation of daily rates of division, which were more than twice those predicted from batch cultures grown at similar temperatures in replete medium. Data from the Salt Pond deployment provide the first continuous record of an A. fundyense population through its complete bloom cycle and demonstrate growth and sexual induction rates much higher than are typically observed in culture.