David A. Caron

The Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP): Illuminating the Functional Diversity of Eukaryotic Life in the Oceans through Transcriptome Sequencing.

Current sampling of genomic sequence data from eukaryotes is relatively poor, biased, and inadequate to address important questions about their biology, evolution, and ecology; this Community Page describes a resource of 700 transcriptomes from marine microbial eukaryotes to help understand their role in the worlds oceans.

Marine bacterial, archaeal and protistan association networks reveal ecological linkages

Microbes have central roles in ocean food webs and global biogeochemical processes, yet specific ecological relationships among these taxa are largely unknown. This is in part due to the dilute, microscopic nature of the planktonic microbial community, which prevents direct observation of their interactions. Here, we use a holistic (that is, microbial system-wide) approach to investigate time-dependent variations among taxa from all three domains of life in a marine microbial community. We investigated the community composition of bacteria, archaea and protists through cultivation-independent methods, along with total bacterial and viral abundance, and physico-chemical observations. Samples and observations were collected monthly over 3 years at a well-described ocean time-series site of southern California. To find associations among these organisms, we calculated time-dependent rank correlations (that is, local similarity correlations) among relative abundances of bacteria, archaea, protists, total abundance of bacteria and viruses and physico-chemical parameters. We used a network generated from these statistical correlations to visualize and identify time-dependent associations among ecologically important taxa, for example, the SAR11 cluster, stramenopiles, alveolates, cyanobacteria and ammonia-oxidizing archaea. Negative correlations, perhaps suggesting competition or predation, were also common. The analysis revealed a progression of microbial communities through time, and also a group of unknown eukaryotes that were highly correlated with dinoflagellates, indicating possible symbioses or parasitism. Possible ‘keystone’ species were evident. The network has statistical features similar to previously described ecological networks, and in network parlance has non-random, small world properties (that is, highly interconnected nodes). This approach provides new insights into the natural history of microbes.

The contribution of microorganisms to particulate carbon and nitrogen in surface waters of the Sargasso Sea near Bermuda

Abstract Seawater samples were collected from the euphotic zone of the Sargasso Sea near Bermuda in August of 1989 and March–April of 1990. Microbial population abundances, chlorophyll concentration, particulate carbon and particulate nitrogen were measured. Calculations were performed to establish the relative and absolute importance of the various microbial assemblages. The choice of conversion factors (g C and N cell −1 , or g C and N μm −3 ) for the microbial populations dramatically affected the estimation of “living” and “detrital” particulate material in the samples, and the relative importance of the various microbial groups. Averaged over all samples on either of the two cruises, microbial biomass constituted a greater proportion of the total particulate carbon and nitrogen during March–April (55% and 63%, respectively), than during August (≈24% and 30%, respectively) using “constrained” conversion factors that were derived. Accordingly, detrital material constituted the bulk of the particulate material during August, but was similar to the amount of microbial biomass during March–April. The bacterial assemblage constituted the largest single pool of microbial carbon (35%) and nitrogen (45%) in the water, and a significant fraction of the total particulate carbon (≈10–20%) and nitrogen (≈15–30%). Phototrophic nanoplankton (microalgae 2–20 μm in size) were second in overall biomass, and often dominated the microbial biomass in the deep chlorophyll maxima that were present during both cruises. The results temper recent assertions concerning the overwhelming importance of bacterial biomass in the oligotrophic Sargasso Sea but still support a major role for these microorganisms in the open ocean as repositories for carbon and nutrients.

Heterotrophic Bacteria and Bacterivorous Protozoa in Oceanic Macroaggregates

Oceanic macroaggregates (marine snow and Rhizosolenia mats) sampled from the Sargasso Sea are associated with bacterial and protozoan populations up to four orders of magnitude greater than those present in samples from the surrounding water. Filamentous, curved, and spiral bacteria constituted a higher proportion of the bacteria associated with the particles than were found among bacteria in the surrounding water. Protozoan populations were dominated numerically by heterotrophic microflagellates, but ciliates and amoebas were also observed. Macroaggregates are highly enriched heterotrophic microenvironments in the oceans and may be significant for the cycling of particulate organic matter in planktonic food chains.

Experimental studies on an omnivorous microflagellate: implications for grazing and nutrient regeneration in the marine microbial food chain

Abstract A phagotrophic marine microflagellate Paraphysomonas imperforata was found to graze on a wide assortment of phytoplankton species as well as bacteria; it also resorted to cannibalism when food was in short supply. Growth rates of the microflagellate were higher than those of the phytoplankton prey, although in some cases lower measured growth rates were found when there was aggregation of cells. This aggregation seemed to be bacterially mediated. The ratio of predator to prey cell length varied from about 2 for the phytoplankton prey to 7 for bacteria. Nitrogen regeneration by the microflagellate, primarily as NH 4 + , never exceeded 50% of the nitrogen originally incorporated by the phytoplankton and bacterial prey. The role of bacteria in regenerating nutrients was found to be minimal relative to microflagellates. These results imply that both omnivory by small protozoa and a greater flexibility in the relationship between predator and prey sizes should be incorporated into the contemporary “microbial food loop” concept. In addition, to achieve nutrient regeneration efficiencies in pelagic surface waters of 80–90%, as is generally believed to occur, requires that the microbial food web be exceedingly complex with a hierarchy of at least several grazing steps. Alternatively, nutrient regeneration efficiencies in surface waters may be lower than envisioned.

Protistan Diversity Estimates Based on 18S rDNA from Seawater Incubations in the Western North Atlantic1

Abstract. Cloning/sequencing and fragment analysis of ribosomal RNA genes (rDNA) are becoming increasingly common methods for the identification of microbial taxa. Sequences of these genes provide many additional taxonomic characters for species that otherwise have few distinctive morphological features, or that require involved microscopy or laboratory culture and testing. These same approaches are now being applied with great success in ecological studies of natural communities of microorganisms. Extensive information on the composition of natural microbial assemblages is being amassed at a rapid pace through genetic analyses of environmental samples and comparison of the resulting genetic information with well‐established (and rapidly growing) public databases. We examined microbial eukaryote diversity in a natural seawater sample from the coastal western North Atlantic Ocean using two molecular biological approaches: the cloning and sequencing of rRNA genes and by fragment analysis of these genes using the terminal restriction fragment length polymorphism (T‐RFLP) method. A simple experiment was carried out to examine changes in the overall eukaryote (largely protistan) diversity and species composition (phylotype diversity) of a natural microbial assemblage when a seawater sample is placed in a container and incubated at ambient light and temperature for 72 h. Containment of the natural seawater sample resulted in relatively minor changes in the overall eukaryote diversity (species richness) obtained by either molecular method at three time points (time‐zero, time‐24 h, time‐72 h). However, substantial changes in the dominance of particular eukaryote phylotypes took place between the three sampling times. Only 18% of the total number of phylotypes observed in the study were observed at all three time points, while 65% (108 of 165) phylotypes were observed only at a single time point (54 unique phylotypes initially, 37 more unique phylotypes at 24 h, and 17 more at 72 h). The results of this study indicate that a high diversity of protistan taxa existed in the original seawater sample at very low abundance, and thus were not observed in the initial characterization of community structure. Containment resulted in significant shifts in the dominance of these taxa, enabling the presence of previously unobserved phylotypes to be documented after 24 or 72 h of incubation.

Defining DNA-Based Operational Taxonomic Units for Microbial-Eukaryote Ecology

ABSTRACT DNA sequence information has increasingly been used in ecological research on microbial eukaryotes. Sequence-based approaches have included studies of the total diversity of selected ecosystems, studies of the autecology of ecologically relevant species, and identification and enumeration of species of interest for human health. It is still uncommon, however, to delineate protistan species based on their genetic signatures. The reluctance to assign species-level designations based on DNA sequences is in part a consequence of the limited amount of sequence information presently available for many free-living microbial eukaryotes and in part a consequence of the problematic nature of and debate surrounding the microbial species concept. Despite the difficulties inherent in assigning species names to DNA sequences, there is a growing need to attach meaning to the burgeoning amount of sequence information entering the literature, and there is a growing desire to apply this information in ecological studies. We describe a computer-based tool that assigns DNA sequences from environmental databases to operational taxonomic units at approximately species-level distinctions. This approach provides a practical method for ecological studies of microbial eukaryotes (primarily protists) by enabling semiautomated analysis of large numbers of samples spanning great taxonomic breadth. Derivation of the algorithm was based on an analysis of complete small-subunit (18S) rRNA gene sequences and partial gene sequences obtained from the GenBank database for morphologically described protistan species. The program was tested using environmental 18S rRNA data sets for two oceanic ecosystems. A total of 388 operational taxonomic units were observed for 2,207 sequences obtained from samples collected in the western North Atlantic and eastern North Pacific oceans.

CHROOCOCCOID CYANOBACTERIA IN LAKE ONTARIO: VERTICAL AND SEASONAL DISTRIBUTIONS DURING 19821

Chroococcoid cyanobacteria (0.7–1.3 μm in diameter) were discovered to be a significant component of the Lake Ontario plankton. Using epifluorescence microscopy, the densities of these microorganisms were found to vary by four orders of magnitude with a single large peak in abundance (6.5 × 105 cells mL−1) corresponding to the time of maximum water temperature. The morphology and abundance of these cyanobacteria were similar to those previously found in oceanic systems. They constituted 10% of the bacterial numbers in the epilimnion during this period, approximately 40% of the biomass of prokaryotes less than 2.0 μm, and 30% of the biomass of all microorganisms less than 20 μm in size. Size fractionation studies indicated that they were responsible for approximately 38% of the total primary production during times of peak abundance, and were important in phosphorus uptake. Cyanobacteria observed in the food vacuoles of heterotrophic microflagellates and in the guts of rotifers suggest that the latter organisms may be important consumers of this prokaryote population.

Grazing of Attached Bacteria by Heterotrophic Microflagellates

Four species of heterotrophic microflagellates were examined for their ability to graze attached and unattached bacteria. The species tested displayed pronounced differences in their ability to graze the bacteriumPseudomonas halodurans attached to chitin particles. Two species of microflagellates (Monas andCryptobia sp.) efficiently grazed unattached bacteria but showed little or no ability to graze attached or aggregated cells. In contrast,Rhynchomonas nasuta andBodo sp. showed marked preferences for attached and aggregated bacteria and a limited ability to graze unattached cells. The density of attached bacteria was reduced by an order of magnitude due to grazing byBodo andR. nasuta, even though the density of unattached bacteria was ∼5–90× the density of attached cells. The maximum densities attained by microflagellates in the cultures were related to the density of unattached bacteria forMonas andCryptobia but not forBodo andR. nasuta. Growth of the latter two species appeared to be related to the density of attached or aggregated bacteria. Based on the results of these experiments, it is concluded that the pelagic existence of microflagellates that graze attached bacteria may be strongly linked to the distribution of suspended particles and their associated bacteria. In addition, the removal of attached bacteria by microflagellates can significantly affect the density of bacteria attached to particles in the plankton. This activity may have important implications for the controversy concerning the relative importance of attached and free-living bacteria in the plankton.

Planning and Implementing Trajectories for Autonomous Underwater Vehicles to Track Evolving Ocean Processes Based on Predictions from a Regional Ocean Model

Path planning and trajectory design for autonomous underwater vehicles (AUVs) is of great importance to the oceanographic research community because automated data collection is becoming more prevalent. Intelligent planning is required to maneuver a vehicle to high-valued locations to perform data collection. In this paper, we present algorithms that determine paths for AUVs to track evolving features of interest in the ocean by considering the output of predictive ocean models. While traversing the computed path, the vehicle provides near-real-time, in situ measurements back to the model, with the intent to increase the skill of future predictions in the local region. The results presented here extend preliminary developments of the path planning portion of an end-to-end autonomous prediction and tasking system for aquatic, mobile sensor networks. This extension is the incorporation of multiple vehicles to track the centroid and the boundary of the extent of a feature of interest. Similar algorithms to those presented here are under development to consider additional locations for multiple types of features. The primary focus here is on algorithm development utilizing model predictions to assist in solving the motion planning problem of steering an AUV to high-valued locations, with respect to the data desired. We discuss the design technique to generate the paths, present simulation results and provide experimental data from field deployments for tracking dynamic features by use of an AUV in the Southern California coastal ocean.