Imaging FlowCytobot modified for high throughput by in-line acoustic focusing of sample particles

© The Author(s), 2017. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Limnology and Oceanography: Methods 15 (2017): 867–874, doi:10.1002/lom3.10205.Imaging FlowCytobot, a submersible instrument that measures optical properties and captures images of nano- and microplankton-sized particles, has proved useful in plankton studies, but its sampling rate is limited by the ability of hydrodynamic focusing to accurately position flowing sample particles. We show that IFCB's sampling rate can be increased at least several-fold by implementing in-line acoustic focusing upstream of the flow cell. Particles are forced to the center of flow by acoustic standing waves created by a piezo-electric transducer bonded to the sample capillary and driven at the appropriate frequency. With the particles of interest confined to the center of the sample flow, the increased size of the sample core that accompanies increased sample flow rate no longer degrades image and signal quality as it otherwise would. Temperature affects the optimum frequency (through its effect on the speed of sound in water), so a relationship between sample temperature and optimum frequency for acoustic focusing was determined and utilized to control the transducer. The modified instrument's performance was evaluated through analyses of artificial particles, phytoplankton cultures, and natural seawater samples and through deployments in coastal waters. The results show that large cells, especially dinoflagellates, are acoustically focused extremely effectively (which could enable, for example, > 10-fold increased sampling rate of harmful algal bloom species, if smaller cells are ignored), while for nearly all cell types typically monitored by IFCB, threefold faster data accumulation was achieved without any compromises. Further increases are possible with more sophisticated software and/or a faster camera.NSF Grant Numbers: OCE-1130140 , OCE-113113

Dynamics and functional diversity of the smallest phytoplankton on the Northeast US shelf

Author Posting. © National Academy of Sciences, 2020. This article is posted here by permission of National Academy of Sciences for personal use, not for redistribution. The definitive version was published in Proceedings of the National Academy of Sciences of the United States of America 117(22), (2020): 12215-12221, doi: 10.1073/pnas.1918439117.Picophytoplankton are the most abundant primary producers in the ocean. Knowledge of their community dynamics is key to understanding their role in marine food webs and global biogeochemical cycles. To this end, we analyzed a 16-y time series of observations of a phytoplankton community at a nearshore site on the Northeast US Shelf. We used a size-structured population model to estimate in situ division rates for the picoeukaryote assemblage and compared the dynamics with those of the picocyanobacteria Synechococcus at the same location. We found that the picoeukaryotes divide at roughly twice the rate of the more abundant Synechococcus and are subject to greater loss rates (likely from viral lysis and zooplankton grazing). We describe the dynamics of these groups across short and long timescales and conclude that, despite their taxonomic differences, their populations respond similarly to changes in the biotic and abiotic environment. Both groups appear to be temperature limited in the spring and light limited in the fall and to experience greater mortality during the day than at night. Compared with Synechococcus, the picoeukaryotes are subject to greater top-down control and contribute more to the region’s primary productivity than their standing stocks suggest.We thank E. T. Crockford, E. E. Peacock, J. Fredericks, Z. Sandwith, the MVCO Operations Team, and divers of the Woods Hole Oceanographic Institution diving program. This work was supported by NSF Grants OCE-0119915 (to R.J.O. and H.M.S.) and OCE-1655686 (to M.G.N., R.J.O., A.R.S., and H.M.O.); NASA Grants NNX11AF07G (to H.M.S.) and NNX13AC98G (to H.M.S.); Gordon and Betty Moore Foundation Grant GGA#934 (to H.M.S.); and Simons Foundation Grant 561126 (to H.M.S.).2020-11-1

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

© The Author(s), 2015. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Limnology and Oceanography 60 (2015): 2059–2078, doi:10.1002/lno.10155.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.National Science Foundation Grant Number: OCE-0430724, OCE-0911031, and OCE-1314642; National Institutes of Health Grant Number: NIEHS-1P50-ES021923-01; National Park Service (NPS) Cooperative Agreement Grant Number: H238015504; Gordon and Betty Moore Foundation Grant Number: #2649 to HMS; IOF Grant Number: MOHAB PIOF-GA-25226

Explaining the causes of cell death in cyanobacteria: what role for asymmetric division?

Cyanobacteria contribute a significant fraction of global primary production and are therefore of great ecological significance. An individual cyanobacteria cell has four potential fates: to divide, perhaps after a dormant period, to be eaten, to undergo viral lysis, or to undergo cell death. In some studies, cyanobacteria cell death has been classified as programmed cell death, borrowing a concept more widely known in metazoan cells, and there are various biochemical parallels to support such a categorisation. However, at the same time there is a growing awareness of asymmetric division as a fundamental process in bacterial division which can result in non-equal daughter cells with differing fitness. Thanks to recent theoretical and experimental advances it is now possible to explore cyanobacteria cell death in the light of asymmetric division and to test hypotheses on the ultimate causes of cyanobacterial cell death. Assessing the degree of protein damage within individual cells during population growth is a sensible initial research target as is the application of techniques which allow the tracking of cell lineages. The existence of asymmetric division in cyanobacteria is likely given its suggested ubiquity across the bacterial domain of life. It will be technically difficult to test the interaction of asymmetric division with environmental variability, and how that leads to individual cell death via differing susceptibilities to environmental stress. However, testing such ideas could confirm asymmetric division as the ultimate cause of cell death in cyanobacteria and thereby allow a better understanding of the patterns of cell death in natural populations

Controls of picophytoplankton abundance and composition in a highly dynamic marine system, the Northern Alboran Sea (Western Mediterranean)

The Alboran Sea is a highly dynamic basin which exhibits a high spatio-temporal variability of hydrographic structures (e.g. fronts, gyres, coastal upwellings). This work compares the abundance and composition of picophytoplankton observed across the northern Alboran Sea among eleven cruises between 2008 and 2012 using flow cytometry. We evaluate the seasonal and longitudinal variability of picophytoplankton on the basis of the circulation regimes at a regional scale and explore the presence of cyanobacteria ecotypes in the basin. The maximal abundances obtained for Prochlorococcus, Synechococcus and picoeukaryotes (12.7 × 104, 13.9 × 104 and 8.6 × 104 cells mL− 1 respectively) were consistent with those reported for other adjacent marine areas. Seasonal changes in the abundance of the three picophytoplankton groups were highly significant although they did not match the patterns described for other coastal waters. Higher abundances of Prochlorococcus were obtained in autumn-winter while Synechococcus and picoeukaryotes exhibited a different seasonal abundance pattern depending on the sector (e.g. Synechococcus showed higher abundance in summer in the west sector and during winter in the eastern study area). Additionally, conspicuous longitudinal gradients were observed for Prochlorococcus and Synechococcus, with Prochlorococcus decreasing from west to east and Synechococcus following the opposite pattern. The analysis of environmental variables (i.e. temperature, salinity and inorganic nutrients) and cell abundances indicates that Prochlorococcus preferred high salinity and nitrate to phosphate ratio. On the contrary, temperature did not seem to play a role in Prochlorococcus distribution as it was numerically important during the whole seasonal cycle. Variability in Synechococcus abundance could not be explained by changes in any environmental variable suggesting that different ecotypes were sampled during the surveys. In particular, our data would indicate the presence of at least two ecotypes of Synechococcus: a summer ecotype widely distributed in the whole Alboran Sea and a winter ecotype adapted to lower temperature and higher nutrient concentration whose growth is favoured in the eastern sector.Versión del editor2,444

Biogeochemical and ecological impacts of boundary currents in the Indian Ocean

Monsoon forcing and the unique geomorphology of the Indian Ocean basin result in complex boundary currents, which are unique in many respects. In the northern Indian Ocean, several boundary current systems reverse seasonally. For example, upwelling coincident with northward-flowing currents along the coast of Oman during the Southwest Monsoon gives rise to high productivity which also alters nutrient stoichiometry and therefore, the species composition of the resulting phytoplankton blooms. During the Northeast Monsoon most of the northern Indian Ocean boundary currents reverse and favor downwelling. Higher trophic level species have evolved behavioral responses to these seasonally changing conditions. Examples from the western Arabian Sea include vertical feeding migrations of a copepod (Calanoides carinatus) and the reproductive cycle of a large pelagic fish (Scomberomorus commerson). The impacts of these seasonal current reversals and changes in upwelling and downwelling circulations are also manifested in West Indian coastal waters, where they influence dissolved oxygen concentrations and have been implicated in massive fish kills. The winds and boundary currents reverse seasonally in the Bay of Bengal, though the associated changes in upwelling and productivity are less pronounced. Nonetheless, their effects are observed on the East Indian shelf as, for example, seasonal changes in copepod abundance and zooplankton community structure. In contrast, south of Sri Lanka seasonal reversals in the boundary currents are associated with dramatic changes in the intensity of coastal upwelling, chlorophyll concentration, and catch per unit effort of fishes. Off the coast of Java, monsoon-driven changes in the currents and upwelling strongly impact chlorophyll concentrations, seasonal vertical migrations of zooplankton, and sardine catch in Bali Strait. In the southern hemisphere the Leeuwin is a downwelling-favorable current that flows southward along western Australia, though local wind forcing can lead to transient near shore current reversals and localized coastal upwelling. The poleward direction of this eastern boundary current is unique. Due to its high kinetic energy the Leeuwin Current sheds anomalous, relatively high chlorophyll, warm-core, downwelling eddies that transport coastal diatom communities westward into open ocean waters. Variations in the Leeuwin transport and eddy generation impact many higher trophic level species including the recruitment and fate of rock lobster (Panulirus cygnus) larvae. In contrast, the transport of the Agulhas Current is very large, with sources derived from the Mozambique Channel, the East Madagascar Current and the southwest Indian Ocean sub-gyre. Dynamically, the Agulhas Current is upwelling favorable; however, the spatial distribution of prominent surface manifestations of upwelling is controlled by local wind and topographic forcing. Meanders and eddies in the Agulhas Current propagate alongshore and interact with seasonal changes in the winds and topographic features. These give rise to seasonally variable localized upwelling and downwelling circulations with commensurate changes in primary production and higher trophic level responses. Due to the strong influence of the Agulhas Current, many neritic fish species in southeast Africa coastal waters have evolved highly selective behaviors and reproductive patterns for successful retention of planktonic eggs and larvae. For example, part of the Southern African sardine (Sardinops sagax) stock undergoes a remarkable northward migration enhanced by transient cyclonic eddies in the shoreward boundary of the Agulhas Current. There is evidence from the paleoceanographic record that these currents and their biogeochemical and ecological impacts have changed significantly over glacial to interglacial timescales. These changes are explored as a means of providing insight into the potential impacts of climate change in the Indian Ocean

Temperature, Synechococcus concentration, division rates, and loss rates

File contains tab-delimited columns of date, temperature (degree Celsius), Synechococcus concentration (cells per mL), division rate (1/d), and loss rate (1/d) at the Martha's Vineyard Coastal Observatory for 2003-2016. Unavailable data are indicated with a NaN