Olson Robert J.

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Robert J.

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Now showing 1 - 9 of 9
  • Article
    Rapid growth and concerted sexual transitions by a bloom of the harmful dinoflagellate Alexandrium fundyense (Dinophyceae)
    (John Wiley & Sons, 2015-09-18) Brosnahan, Michael L. ; Velo-Suarez, Lourdes ; Ralston, David K. ; Fox, Sophia E. ; Sehein, Taylor R. ; Shalapyonok, Alexi ; Sosik, Heidi M. ; Olson, Robert J. ; Anderson, Donald M.
    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.
  • Article
    Resonance control of acoustic focusing systems through an environmental reference table and impedance spectroscopy
    (Public Library of Science, 2018-11-14) Kalb, Daniel J. ; Olson, Robert J. ; Sosik, Heidi M. ; Woods, Travis A. ; Graves, Steven W.
    Acoustic standing waves can precisely focus flowing particles or cells into tightly positioned streams for interrogation or downstream separations. The efficiency of an acoustic standing wave device is dependent upon operating at a resonance frequency. Small changes in a system’s temperature and sample salinity can shift the device’s resonance condition, leading to poor focusing. Practical implementation of an acoustic standing wave system requires an automated resonance control system to adjust the standing wave frequency in response to environmental changes. Here we have developed a rigorous approach for quantifying the optimal acoustic focusing frequency at any given environmental condition. We have demonstrated our approach across a wide range of temperature and salinity conditions to provide a robust characterization of how the optimal acoustic focusing resonance frequency shifts across these conditions. To generalize these results, two microfluidic bulk acoustic standing wave systems (a steel capillary and an etched silicon wafer) were examined. Models of these temperature and salinity effects suggest that it is the speed of sound within the liquid sample that dominates the resonance frequency shift. Using these results, a simple reference table can be generated to predict the optimal resonance condition as a function of temperature and salinity. Additionally, we show that there is a local impedance minimum associated with the optimal system resonance. The integration of the environmental results for coarse frequency tuning followed by a local impedance characterization for fine frequency adjustments, yields a highly accurate method of resonance control. Such an approach works across a wide range of environmental conditions, is easy to automate, and could have a significant impact across a wide range of microfluidic acoustic standing wave systems.
  • Article
    Seasons of Syn
    (Wiley, 2019-11-19) Hunter-Cevera, Kristen R. ; Neubert, Michael G. ; Olson, Robert J. ; Shalapyonok, Alexi ; Solow, Andrew R. ; Sosik, Heidi M.
    Synechococcus is a widespread and important marine primary producer. Time series provide critical information for identifying and understanding the factors that determine abundance patterns. Here, we present the results of analysis of a 16‐yr hourly time series of Synechococcus at the Martha's Vineyard Coastal Observatory, obtained with an automated, in situ flow cytometer. We focus on understanding seasonal abundance patterns by examining relationships between cell division rate, loss rate, cellular properties (e.g., cell volume, phycoerythrin fluorescence), and environmental variables (e.g., temperature, light). We find that the drivers of cell division vary with season; cells are temperature‐limited in winter and spring, but light‐limited in the fall. Losses to the population also vary with season. Our results lead to testable hypotheses about Synechococcus ecophysiology and a working framework for understanding the seasonal controls of Synechococcus cell abundance in a temperate coastal system.
  • Article
    Molecular subdivision of the marine diatom Thalassiosira rotula in relation to geographic distribution, genome size, and physiology
    (BioMed Central, 2012-10-26) Whittaker, Kerry A. ; Rignanese, Dayna R. ; Olson, Robert J. ; Rynearson, Tatiana A.
    Marine phytoplankton drift passively with currents, have high dispersal potentials and can be comprised of morphologically cryptic species. To examine molecular subdivision in the marine diatom Thalassiosira rotula, variations in rDNA sequence, genome size, and growth rate were examined among isolates collected from the Atlantic and Pacific Ocean basins. Analyses of rDNA included T. gravida because morphological studies have argued that T. rotula and T. gravida are conspecific. Culture collection isolates of T. gravida and T. rotula diverged by 7.0 ± 0.3% at the ITS1 and by 0.8 ± 0.03% at the 28S. Within T. rotula, field and culture collection isolates were subdivided into three lineages that diverged by 0.6 ± 0.3% at the ITS1 and 0% at the 28S. The predicted ITS1 secondary structure revealed no compensatory base pair changes among lineages. Differences in genome size were observed among isolates, but were not correlated with ITS1 lineages. Maximum acclimated growth rates of isolates revealed genotype by environment effects, but these were also not correlated with ITS1 lineages. In contrast, intra-individual variation in the multi-copy ITS1 revealed no evidence of recombination amongst lineages, and molecular clock estimates indicated that lineages diverged 0.68 Mya. The three lineages exhibited different geographic distributions and, with one exception, each field sample was dominated by a single lineage. The degree of inter- and intra-specific divergence between T. gravida and T. rotula suggests they should continue to be treated as separate species. The phylogenetic distinction of the three closely-related T. rotula lineages was unclear. On the one hand, the lineages showed no physiological differences, no consistent genome size differences and no significant changes in the ITS1 secondary structure, suggesting there are no barriers to interbreeding among lineages. In contrast, analysis of intra-individual variation in the multicopy ITS1 as well as molecular clock estimates of divergence suggest these lineages have not interbred for significant periods of time. Given the current data, these lineages should be considered a single species. Furthermore, these T. rotula lineages may be ecologically relevant, given their differential abundance over large spatial scales.
  • Article
    Diatoms favor their younger daughters
    (Association for the Sciences of Limnology and Oceanography, 2012-09) Laney, Samuel R. ; Olson, Robert J. ; Sosik, Heidi M.
    We used a time-lapse imaging approach to examine cell division in the marine centric diatom Ditylum brightwellii and observed that daughter cells who inherited their parents' hypothecal frustule half were more likely to divide before their sisters. This is consistent with observations in Escherichia coli of a bias between sister cells, where faster growth in one sister is thought to arise from its inheriting parental material with less oxidative damage. We also observed that hypothecal sisters in D. brightwellii were more likely to inherit a greater proportion of their parents' cellular material, similar to what has been seen in E. coli. We found a statistically significant correlation between the amount of parental material inherited by a hypothecal daughter and its relative division rate, indicating that this extra material inherited by the hypothecal daughter plays a role in its more rapid division. Furthermore, the intercept in this regression was greater than zero, indicating that other factors, such as differences in the quality of inherited material, also play a role. This similarity between two taxonomically distant microbes suggests that favoritism toward one daughter might occur broadly among unicellular organisms that reproduce asexually by binary fission. Such a bias in cell division might be advantageous, given model predictions that show that favoring one daughter at the expense of the other can result in higher population growth rates, increasing the chance that a cell's genotype will survive compared to a model where the daughters divide at equal rates.
  • Article
    Microzooplankton community structure investigated with imaging flow cytometry and automated live-cell staining
    (Inter-Research, 2016-05-25) Brownlee, Emily F. ; Olson, Robert J. ; Sosik, Heidi M.
    Protozoa play important roles in grazing and nutrient recycling, but quantifying these roles has been hindered by difficulties in collecting, culturing, and observing these often-delicate cells. During long-term deployments at the Martha’s Vineyard Coastal Observatory (Massachusetts, USA), Imaging FlowCytobot (IFCB) has been shown to be useful for studying live cells in situ without the need to culture or preserve. IFCB records images of cells with chlorophyll fluorescence above a trigger threshold, so to date taxonomically resolved analysis of protozoa has presumably been limited to mixotrophs and herbivores which have eaten recently. To overcome this limitation, we have coupled a broad-application ‘live cell’ fluorescent stain with a modified IFCB so that protozoa which do not contain chlorophyll (such as consumers of unpigmented bacteria and other heterotrophs) can also be recorded. Staining IFCB (IFCB-S) revealed higher abundances of grazers than the original IFCB, as well as some cell types not previously detected. Feeding habits of certain morphotypes could be inferred from their fluorescence properties: grazers with stain fluorescence but without chlorophyll cannot be mixotrophs, but could be either starving or feeding on heterotrophs. Comparisons between cell counts for IFCB-S and manual light microscopy of Lugol’s stained samples showed consistently similar or higher counts from IFCB-S. We show how automated classification through the extraction of image features and application of a machine-learning algorithm can be used to evaluate the large high-resolution data sets collected by IFCBs; the results reveal varying seasonal patterns in abundance among groups of protists.
  • Article
    A fluorescence-activated cell sorting subsystem for the Imaging FlowCytobot
    (John Wiley & Sons, 2016-10-17) Lambert, Bennett ; Olson, Robert J. ; Sosik, Heidi M.
    Recent advances in plankton ecology have brought to light the importance of variability within populations and have suggested that cell-to-cell differences may influence ecosystem-level processes such as species succession and bloom dynamics. Flow cytometric cell sorting has been used to capture individual plankton cells from natural water samples to investigate variability at the single cell level, but the crude taxonomic resolution afforded by the fluorescence and light scattering measurements of conventional flow cytometers necessitates sorting and analyzing many cells that may not be of interest. Addition of imaging to flow cytometry improves classification capability considerably: Imaging FlowCytobot, which has been deployed at the Martha's Vineyard Coastal Observatory since 2006, allows classification of many kinds of nano- and microplankton to the genus or even species level. We present in this paper a modified bench-top Imaging FlowCytobot (IFCB-Sorter) with the capability to sort both single cells and colonies of phytoplankton and microzooplankton from seawater samples. The cells (or subsets selected based on their images) can then be cultured for further manipulation or processed for analyses such as nucleic acid sequencing. The sorting is carried out in two steps: a fluorescence signal triggers imaging and diversion of the sample flow into a commercially available “catcher tube,” and then a solenoid-based flow control system isolates each sorted cell along with 20 μL of fluid.
  • Article
    Parasitic infection of the diatom Guinardia delicatula, a recurrent and ecologically important phenomenon on the New England Shelf
    (Inter-Research, 2014-04-29) Peacock, Emily E. ; Olson, Robert J. ; Sosik, Heidi M.
    Plankton images collected by Imaging FlowCytobot from 2006 to 2013 at the Martha’s Vineyard Coastal Observatory (Massachusetts, USA) were used to identify and quantify the occurrence of the diatom Guinardia delicatula and of a parasite that seems specific to this host. We observed infection with morphological stages that appear similar to the parasite Cryothecomonas aestivalis. Our results show that events during which infection rates exceed 10% are recurrent on the New England Shelf and suggest that the parasites are an important source of host mortality. We document a significant negative relationship between bloom magnitude and parasite infection rate, supporting the hypothesis that the parasites play a major role in controlling blooms. While G. delicatula is observed during all seasons, the infecting stages of the parasite are abundant only when water temperature is above 4°C. The anomalously warm water and small G. delicatula bloom during the winter of 2012 provided evidence that parasites can be active through winter if temperatures remain relatively high. As climate change continues, winter periods of water below 4°C may shorten or disappear in this region, suggesting that parasite effects on species such as G. delicatula may increase, with immediate impacts on their population dynamics.
  • Article
    Imaging FlowCytobot modified for high throughput by in-line acoustic focusing of sample particles
    (John Wiley & Sons, 2017-09-19) Olson, Robert J. ; Shalapyonok, Alexi ; Kalb, Daniel J. ; Graves, Steven W. ; Sosik, Heidi M.
    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.