Alan J. Lewitus Research Assistant Professor Belle W. Baruch Institute for Marine Biology and Coastal Research Address: Baruch Marine Laboratory University of South Carolina P.O. Box 1630 Georgetown, South Carolina 29442 Tel: (843) 546-3623 Fax: (843) 546-1632 Email: Lewitus@belle.baruch.sc.edu

Recent Positions:

Research Interests:

Phytoplankton physiology and ecology, microbial food web dynamics, regulation of photosynthesis and respiration, phytoplankton physiological responses to organic and inorganic nutrients, physiological role and measurement of photosynthetic pigments, physiological ecology of toxic and nuisance algae including Pfiesteria piscicida, phytoplankton community structure.

Ken Hayes, Phytoplankton Lab Manager	Scott Gransden, Research Technician
Raphael Tymowski, Jennifer Keesee and Ivy Collins, Reseach Technicians

Selected Abstracts

(major funding source in parentheses at end of abstract):

Lewitus, A.J., H.B. Glasgow and J.M. Burkholder. 1999. Kleptoplastidy in the toxic dinoflagellate, Pfiesteria piscicida (Dinophyceae). J. Phycol. 35:303-12.

The ichthyotoxic dinoflagellate, Pfiesteria piscicida Steidinger & Burkholder, has a complex life cycle with several heterotrophic flagellated and amoeboid stages. A prevalent flagellated form, the nontoxic zoospore stage, has a proficient grazing ability, particularly with cryptophyte prey. Although P. piscicida zoospores lack the genetic capability to synthesize chloroplasts, they can obtain functional chloroplasts from algal prey (i.e. kleptoplastidy), as demonstrated here with a cryptophyte prey. Zoospores grown with Rhodomonas sp. Karsten CCMP757 (Cryptophyceae) grazed the cryptophyte population to minimal densities. After placing the cultures in near darkness where cryptophyte recovery was restricted and prey ingestion did not occur, the time-course patterns in growth, prey chloroplast content zoospore ^-1, and prey nucleus content zoospore^-1 were followed. Ingested chloroplasts were selectively retained in the dinoflagellate, as indicated by the decline and, ultimately, near-absence of cryptophyte nuclei in plastid-containing zoospores. Chloroplasts retained inside P. piscicida cells for at least a week were photosynthetically active, as indicated by starch accumulation and microscope-autoradiographic measurements of bicarbonate uptake. Recognition that P. pisicicida can function as a phototroph broadens our perspective of the physiological ecology of the dinoflagellate because it suggests that, at least during part of its life cycle, P. piscicida's growth and survival may be affected by photoregulation and nutritional control of photosynthesis. (NSF Division of Biological Oceanography, OCE-9403920).

Eriksen, N.T. and A.J. Lewitus. 1999. Cyanide-resistant respiration in diverse marine phytoplankton. Evidence for the widespread occurrence of the alternative oxidase. Aq. Mic. Ecol. 17:145-52.

In higher plants, the alternative oxidase (AOX) is the terminal oxidase in a mitochondrial electron transport chain thought to allow carbon flow through glycolysis and the citric acid cycle when cellular energy levels are high. In phytoplankton, information is lacking on the taxonomic distribution and metabolic properties of the AOX. We used cyanide resistant respiration to indicate the presence of the AOX, and the AOX inhibitors, salicylhydroxamic acid (SHAM) and propyl gallate (PG), to estimate the relative activity and capacity of the AOX in axenic cultures of the marine phytoplankton, Chlorella sp. (Chlorophyceae), Closterium sp. (Zygnematophyceae), Thalassiosira sp. (Bacillariophyceae), Cryptomonas sp. (Cryptophyceae), Ochromonas sp. (Chrysophyceae), and Amphidinium carterae (Dinophyceae), and the freshwater green alga Chlamydomonas reinhardtii. AOX inhibitor effects were higher in stationary phase (nutrient-limited) cultures compared to linearly growing cultures. With the exception of Closterium, in which respiration was almost completely inhibited by cyanide, estimates of AOX capacity determined as the effect of AOX inhibitors on cyanide-resistant respiration, were nearly identical using the two AOX inhibitors, and ranged from 46% to 113% of the respiration rates of cultures not exposed to inhibitors. The presence of the AOX in five of the six marine phytoplankton species tested suggests that the AOX is widespread among phytoplankton. Furthermore, the pronounced inhibitory effect of SHAM and PG added alone to stationary phase cultures of Chlorella, Thalassiosira, Cryptomonas, and Amphidinium (21% to 63% of uninhibited respiration rates) implies that the AOX contributes substantially to oxygen and carbon cycling in many species of phytoplankton during nutrient deficiency. (NSF Division of Biological Oceanography,OCE-9315663).

Lewitus, A.J., E.T. Koepfler and R.J. Pigg. 1999. Use of dissolved organic nitrogen by a salt marsh phytoplankton bloom community. Arch. Hydrobiol. (in press).

In North Inlet, a shallow, turbid salt marsh estuary near Georgetown, SC, a summer phytoplankton bloom occurs that is composed primarily of nanoflagellates. Grazing and light-limitation are important regulatory factors for bloom formation, and nitrogen does not limit phytoplankton population growth, as evidenced by the ineffectiveness of ammonium in stimulating biomass production in bioassay experiments. In contrast, glycine greatly stimulated community chlorophyll concentrations and the abundances of all phytoplankton size-fractions, but primarily the nano- and picoplankton. Comparisons of bacterial vs. phytoplankton physiological responses (growth, production, glycine uptake in the presence or absence of antibiotics) to DON enrichment suggested that the stimulatory effect of DON on phytoplankton growth resulted primarily from both direct uptake of the organic substrate and indirectly through bacterial breakdown. Because phytoplankton growth was light-limited, but not nitrogen-limited, during the bloom, we hypothesized that the stimulatory response to glycine was due its use as a respiratory substrate. ("Creek": NSF DEB-9509057 and "USES": NOAA Office of Ocean Research Programs NA90AA-D-SG672).

Lee, E.S., A.J. Lewitus and R.K. Zimmer-Faust. 1999. Chemotaxis in a marine cryptophyte: Behavioral plasticity in response to amino acids and nitrate. Limnol. Oceanogr 44:1571-4.

The behavioral responses of Chroomonas sp., a small (3 mm diameter) cryptophyte, to amino acids, ammonium, or nitrate were investigated by computer-assisted video motion analysis. Chroomonas sp. exhibited chemotactic behavior consistent with that of some bacteria (i.e. a change in tumbling frequency but not swimming speed), but this response varied with growth nutritional condition and chemical stimulant tested. Cells grown with glycine as the sole nitrogen source exhibited significant reductions in tumbling frequency in response to glutamate, methionine, alanine, or aspartate at concentrations as low as 0.1 mM, but not to other amino acids, ammonium, or nitrate at comparable concentrations. In contrast to glycine-grown cultures, nitrate-grown cells did not significantly change motility properties in response to any amino acid tested or ammonium, but did reduce tumbling frequency in response to 1 mM nitrate. It is hypothesized that chemotaxis is inducible in Chroomonas sp., and regulated by the nutritional environment (i.e. when nitrate is not available, cells become chemotactic towards alternative nitrogen sources such as amino acids). The observed behavioral responses to amino acids supplied at 0.1 mM suggests that Chroomonas sp. may exhibit chemotaxis to organic nitrogen sources under natural conditions when inorganic nitrogen is limiting. ("USES": NOAA NA90AA-D-SG672 and NSF OCE-9315663).

Lewitus, A.J., E.T. Koepfler and J.T. Morris. 1998. Seasonal variation in the regulation of phytoplankton by nitrogen and grazing in a salt marsh estuary. Limnol. Oceanogr. 43:636-646.

In North Inlet, a tidally-dominated salt marsh estuary near Georgetown, SC, the summer chlorophyll maximum corresponds with an annual peak in ambient NH₄⁺ concentration. Because NH₄⁺ is the major inorganic nitrogen source available to phytoplankton, and phosphorus should not be limiting (N:P is generally around seven), the positive relationship between chlorophyll and NH₄⁺ suggests that phytoplankton population growth during the summer bloom is limited by factors other than nutrient supply. We tested the hypothesis that phytoplankton population growth during the bloom was limited by grazing by incubating natural samples in treatments designed to differentiate between nutrient and grazing effects, and following time-course changes in total phytoplankton biomass and phototrophic community composition. Marked seasonal differences were observed in the relative contribution of pico-, nano-, or microplankton to phytoplankton community biomass, and the mechanisms controlling phytoplankton population growth. During the summer bloom, phototrophic picoplankton (mostly Synechococcus spp.) and nanoplankton (mostly flagellates) were relatively abundant, and phytoplankton population growth was greatly stimulated by dilution that reduced microzooplankton grazing pressure, but unaffected by NH₄⁺ addition. During the winter, when diatoms dominated the phytoplankton, the response to dilution was relatively minor, while NH₄⁺ addition significantly stimulated chlorophyll production and the growth of various phytoplankton groups. The results indicate a seasonal transition in microbial food web trophic structure and regulation in North Inlet estuary from a summer nanoflagellate-prevalent chlorophyll bloom characterized by microbial loop dynamics where microzooplankton grazing is important in controlling population growth, to a winter diatom-dominated community regulated by nutrient supply. ("Creek study": NSF Ecosystems Program, Division of Environmental Biology, RUI, DEB-9509057).

Lewitus, A.J., J.M. Burkholder, H.B. Glasgow Jr., P.M. Glibert, Kenneth C. Hayes, and Bonnie M. Willis. 1999. Mixotrophy and nitrogen uptake by Pfiesteria piscicida (Dinophyceae). J. Phycol. (accepted).

The nutritional versatility of dinoflagellates is a complicating factor in identifying potential links between nutrient enrichment and the proliferation of harmful algal blooms. For example, although dinoflagellates associated with harmful algal blooms (e.g. red tides) generally are considered to be phototrophic and use inorganic nutrients such as nitrate or phosphate, many of these species also have pronounced heterotrophic capabilities either as osmotrophs or phagotrophs. Recently, the widespread occurrence of the heterotrophic toxic dinoflagellate, Pfiesteria piscicida Steidinger & Burkholder, has been documented in turbid estuarine waters. P. piscicida has a relatively proficient grazing ability, but also has an ability to function as a phototroph by acquiring chloroplasts from algal prey, a process termed kleptoplastidy. We tested the ability of kleptoplastidic P. piscicida to take up ¹⁵N-labeled NH₄⁺, NO₃^-, urea, or glutamate. The photosynthetic activity of these cultures was verified, in part, by use of the fluorochrome, primulin, which indicated a positive relationship between photosynthetic starch production and growth irradiance. All four N substrates were taken up by P. piscicida, and the highest uptake rates were in the range cited for phytoplankton, and were similar to N uptake estimates for phagotrophic P. piscicida. The demonstration of direct nutrient acquisition by kleptoplastidic P. piscicida suggests that the response of the dinoflagellate to nutrient enrichment is complex, and that the specific pathway of nutrient stimulation (e.g. indirect stimulation through enhancement of phytoplankton prey abundance vs. direct stimulation by saprotrophic nutrient uptake) may depend on P. piscicida's nutritional state (phagotrophy vs. phototrophy). (NSF OCE-9403920).

Lewitus, A.J., T. Kawaguchi, and G.R. DiTullio. (subm.) Iron limitation of phytoplankton in a salt marsh estuary. Limnol. Oceanogr.

Over the last two decades, escalating rates of coastal development have altered macro- and micronutrient loading patterns to many southeastern U.S. estuaries. For example, a common repercussion of coastal forest clear-cutting is the removal of a buffer zone for non-point source pollution, which may lead to increased nitrate loads. This study addresses another potential consequence of coastal deforestation that may have important implications for estuarine ecosystem productivity: a reduction in bio-available Fe. Here, the hypothesis is tested that organically-bound Fe from coastal forests plays an important role in supplying Fe for the growth of estuarine phytoplankton, and that clear-cutting of coastal forests may reduce the supply of available Fe to the extent that phytoplankton growth can become Fe-limited. We compared the potential for Fe limitation in two neighboring South Carolina salt marsh estuaries, one (Murrells Inlet) impacted by urbanization-associated clear-cutting, and the other (North Inlet) not impacted by development and surrounded by forests. The urbanized estuary was marked by higher NO₃^- and PO₄^3-, but lower dissolved Fe and chlorophyll a concentrations compared to the forested estuary. In bioassay experiments, the combined additions of chelated Fe and NO3- to natural phytoplankton populations from Murrells Inlet resulted in approximately a 3-fold stimulation in chlorophyll a relative to samples amended by NO₃^- alone, but Fe addition had no effect on chlorophyll a in North Inlet samples. In bioassay experiments using Fe-deplete semi-continuous cultures of Synechococcus WH8101, Fe addition did not affect the net growth rate of cultures transferred to water from either estuary, but increased the chlorophyll a content of cells transferred to Murrells Inlet water. Finally, photosynthesis vs. irradiance parameters (a , P_max) obtained from North Inlet samples were similar to those measured in Fe-replete Synechococcus and Phaeodactylum continuous cultures, while parameters derived from Murrells Inlet samples were much lower and comparable to Fe-deplete cultures. The results are indicative of Fe limitation of Murrells Inlet, but not North Inlet, phytoplankton, and stress consideration of trace metal availability, and anthropogenic influences thereupon, in regulating estuarine phytoplankton production. ("USES": NOAA NA90AA-D-SG672).

Other selected publications:

Glasgow, H.B., Jr, A.J. Lewitus and J.M. Burkholder. 1998. Feeding behavior of the ichthyotoxic estuarine dinoflagellate, Pfiesteria piscicida, on amino acids, algal prey, and fish vs. mammalian erythrocytes. In: Harmful Microalgae, Reguera, B. J. Blanco, M.L. Fernandez and T. Wyatt (eds.). Xunta de Galicia and UNESCO, Paris pp. 394-397.

Burkholder, J.M., H.B. Glasgow, Jr., and A.J. Lewitus. 1998. Physiological ecology of Pfiesteria piscicida with general comments on ambush predator dinoflagellates. In: Physiological Ecology of Harmful Marine Phytoplankton. D.M. Anderson (ed.). UNESCO, Paris pp. 175-191.

Kawaguchi, T., A.J. Lewitus, C.M. Aelion and H.N. McKellar. 1997. Can urbanization limit iron availability to estuarine algae? J. Exp. Mar. Biol. Ecol. 213:53-69.

Lewitus, A.J. and T.M. Kana. 1995. Light respiration in six estuarine phytoplankton clones: contrasts under autotrophic and mixotrophic growth conditions. J. Phycol. 31:754-761.

Lewitus, A.J., R.V. Jesien, T.M. Kana, J.M. Burkholder, H.B. Glasgow, Jr., and E. May. 1995. Discovery of the "phantom" dinoflagellate in Chesapeake Bay. Estuaries 18:373-378.

Van Heukelem, L., A.J. Lewitus, T.M. Kana, and N.E. Craft. 1994. Improved separations of phytoplankton pigments using temperature-controlled high performance liquid chromatography. Mar. Ecol. Prog. Ser. 114:303-313.

Lewitus, A.J. and T.M. Kana. 1994. Responses of estuarine phytoplankton to exogenous glucose: Stimulation versus inhibition of photosynthesis and respiration. Limnol. Oceanog. 39:182-189.

Van Heukelem, L., A.J. Lewitus, T.M. Kana, and N.E. Craft. 1992. High performance liquid chromatography of phytoplankton pigments using a polymeric reversed-phase C¹⁸ column. J. Phycol. 28:867-72.

Lewitus, A.J. and D.A. Caron. 1991. Physiological responses of phytoflagellates to dissolved organic substrate additions. 2.Dominant role of autotrophic nutrition in Pyrenomonas salina (Cryptophyceae). Plant Cell Physiol. 32:791-801.

Lewitus, A.J. and D.A. Caron. 1991. Physiological responses of phytoflagellates to dissolved organic substrate additions. 1. Dominant role of heterotrophic nutrition in Poterioochromonas malhamensis (Chrysophyceae). Plant Cell Physiol. 32:671-680.

Lewitus, A.J., D.A. Caron, and K.R. Miller. 1991. Effects of light and glycerol on the organization of the photosynthetic apparatus in the facultative heterotroph Pyrenomonas salina (Cryptophyceae). J. Phycol. 27:578-587.

Lewitus, A.J. and D.A. Caron. 1990. The relative effects of nitrogen or phosphorus depletion, and light intensity on the pigmentation, chemical composition, and volume of Pyrenomonas salina (Cryptophyceae). Mar. Ecol. Prog. Ser. 61:171-181.