Matthew W. Parrow

Cryptic Sex in Symbiodinium (Alveolata, Dinoflagellata) is Supported by an Inventory of Meiotic Genes

Symbiodinium encompasses a diverse clade of dinoflagellates that are ecologically important as symbionts of corals and other marine organisms. Despite decades of study, cytological evidence of sex (karyogamy and meiosis) has not been demonstrated in Symbiodinium, although molecular population genetic patterns support the occurrence of sexual recombination. Here, we provide additional support for sex in Symbiodinium by uncovering six meiosis‐specific and 25 meiosis‐related genes in three published genomes. Cryptic sex may be occurring in Symbiodiniums seldom‐seen free‐living state while being inactive in the symbiotic state.

Two-stage fungal biopulping for improved enzymatic hydrolysis of wood

A novel two-stage, whole organism fungal biopulping method was examined for increasing the yield of enzymatic hydrolysis of wood into soluble glucose. Liriodendron tulipifera wood chips (1g) were exposed to liquid culture suspensions of white rot (Ceriporiopsis subvermispora) or brown rot (Postia placenta) fungi and incubated at 28°C, either alone in single-stage 30 day (one fungal species applied) or two-stage 60 day (both fungal species applied in alternative succession) treatments. Fungi grew in all treatments, but did not significantly decrease the percent carbohydrate content of the wood. Two-stage treatments differed significantly in mass loss depending on order of exposure, suggesting additive or inhibitory fungal interactions occurred. Treatments consisting of C. subvermispora followed by P. placenta exhibited 6 ± 0.5% mass loss and increased the yield of enzymatic hydrolysis by 67-119%. This significant hydrolysis improvement suggests that fungal biopulping technologies could support commercial lignocellulosic ethanol production efforts if further developed.

Morphometrics of the Paleogene coccolith genera Cruciplacolithus, Chiasmolithus, and Sullivania; a complex evolutionary history

Investigation of the evolution of calcareous nannoplankton is hindered by the extremely small size (5-10 μm) of their fossils. We introduce new technology, a scanning electron microscope connected to an image analysis system (SEM-IAS), which allows this field to be explored as never before. This system enables a host of morphocharacters to be measured and included in studies of phylogeny and evolutionary dynamics. We have applied the SEM-IAS to study the evolution of Paleocene coccolith genera Cruciplacolithus, Chiasmolithus , and Sullivania. A variety of detailed measurements have been made on over 4000 coccoliths from Deep Sea Drilling Project Site 384 in the temperate North Atlantic and Ocean Drilling Program Site 690 in the Weddell Sea off Antarctica. Our results indicate no simple relationships between morphocharacters, the shapes of all three genera are both complex and highly variable. Although most morphocharacters possess little phylogenetic significance, the areas of different shield cycles show gradual divergence between Chiasmolithus and Sullivania through the Paleocene. Change of most other morphocharacters occurs at variable rates and reversals in trends are common. Minimal correlation exists between the trends and oscillatory shape changes observed at the two sites. We conclude that these trends and oscillations represent local, transitory ecophenotypic variation of the complex form. There is little stasis in the ten-million-year record studied.

Mixotrophy and loss of phototrophy among geographic isolates of freshwater Esoptrodinium/Bernardinium sp. (Dinophyceae)

The genus Esoptrodinium Javornický consists of freshwater, athecate dinoflagellates with an incomplete cingulum. Strains isolated thus far feed on microalgae and most possess obvious pigmented chloroplasts, suggesting mixotrophy. However, some geographic isolates lack obvious pigmented chloroplasts. The purpose of this study was to comparatively examine this difference and the associated potential for mixotrophy among different isolates of Esoptrodinium. All isolates phagocytized prey cells through an unusual hatch‐like peduncle located on the ventral episome, and were capable of ingesting various protist taxa. All Esoptrodinium isolates required both food and light to grow. However, only the tested strain with visible pigmented chloroplasts benefited from light in terms of increased biomass (phototrophy). Isolates lacking obvious chloroplasts received no biomass benefit from light, but nevertheless required light for sustained growth (i.e., photoobligate, but not phototrophic). Isolates with visible chloroplasts exhibited chlorophyll autofluorescence and formed a monophyletic psbA gene clade that suggested Esoptrodinium possesses inherited, peridinoid‐type plastids. One isolate with cryptic, barely visible plastids lacked detectable chlorophyll and exhibited an apparent loss‐of‐function mutation in psbA, indicating the presence of nonphotosynthetic plastids. The other isolate that lacked visible chloroplasts lacked both detectable chlorophyll and an amplifiable psbA sequence. The results demonstrate mixotrophy quantitatively for the first time in a freshwater dinoflagellate, as well as apparent within‐clade loss of phototrophy along with a correlated mutation sufficient to explain that phenotype. Phototrophy is a variable trait in Esoptrodinium; further study is required to determine if this represents an inter‐ or intraspecific (allelic) characteristic in this taxon.

CYTOLOGICAL AND PHYLOGENETIC DIVERSITY IN FRESHWATER ESOPTRODINIUM/BERNARDINIUM SPECIES (DINOPHYCEAE)(1).

The genera Esoptrodinium Javornický and Bernardinium Chodat comprise freshwater, athecate dinoflagellates with an incomplete cingulum but differing reports regarding cingulum orientation and the presence of chloroplasts and an eyespot. To examine this reported diversity, six isolates were collected from different freshwater ponds and brought into clonal culture. The isolates were examined using LM to determine major cytological differences, and rDNA sequences were compared to determine relatedness and overall phylogenetic position within the dinoflagellates. All isolates were athecate with a left‐oriented cingulum that did not fully encircle the cell, corresponding to the current taxonomic concept of Esoptrodinium. However, consistent cytological differences were observed among clonal isolates. Most isolates exhibited unambiguous pale green chloroplasts and a distinct bright‐red eyespot located at the base of the longitudinal flagellum. However, one isolate had cryptic chloroplasts that were difficult to observe using LM, and another had an eyespot that was so reduced as to be almost undetectable. Another isolate lacked visible chloroplasts but did possess the characteristic eyespot. Nuclear rDNA phylogenies strongly supported a monophyletic Esoptrodinium clade containing all isolates from this study together with a previous sequence from Portugal, within the Tovelliaceae. Esoptrodinium subclades were largely correlated with cytological differences, and the data suggested that independent chloroplast and eyespot reduction and/or loss may have occurred within this taxon. Overall, the isolates encompassed the majority of cytological diversity reported in previous observations of Bernardinium/Esoptrodinium in field samples. Systematic issues with the current taxonomic distinction between Bernardinium and Esoptrodinium are discussed.

Axenic Culture of the Heterotrophic Dinoflagellate Pfiesteria shumwayae in a Semi-Defined Medium

ABSTRACT. A semi‐defined, biphasic culture medium was developed that supported the axenic growth of three strains of the heterotrophic dinoflagellate Pfiesteria shumwayae. Maximum cell yields and division rates in the semi‐defined medium ranged from 0.1 × 105 to 4.0 × 105 cells/ml and 0.5 to 1.7 divisions/day, respectively, and depended on the concentration of the major components in the medium as well as the P. shumwayae strain. The medium contained high concentrations of certain dissolved and particulate organic compounds, including amino acids and lipids. Pfiesteria shumwayae flagellated cells were attracted to insoluble lipids present in the medium and appeared to feed on the lipid particles, suggesting that phagocytosis may be required for growth in axenic culture. Development of a semi‐defined medium represents significant progress toward a completely defined axenic culture medium and subsequent determination of the biochemical requirements of P. shumwayae, needed to advance understanding of the nutritional ecology of this species. Further, this medium provides an economical, simplified method for generating high cell densities of P. shumwayae in axenic culture that will facilitate controlled investigations on the physiology and biochemistry of this heterotrophic dinoflagellate.

AXENIC CULTIVATION OF THE HETEROTROPHIC DINOFLAGELLATE PFIESTERIA SHUMWAYAE AND OBSERVATIONS ON FEEDING BEHAVIOR1

Pfiesteria shumwayae Glasgow et J. M. Burkh. [=Pseudopfiesteria shumwayae (Glasgow et J. M. Burkh.) Litaker, Steid., P. L. Mason, Shields et P. A. Tester] is a heterotrophic dinoflagellate commonly found in temperate, estuarine waters. P. shumwayae can feed on other protists, fish, and invertebrates, but research on the biochemical requirements of this species has been restricted by the lack of axenic cultures. An undefined, biphasic culture medium was formulated that supported the axenic growth of two of three strains of P. shumwayae. The medium contained chicken egg yolk as a major component. Successful growth depended on the method used to sterilize the medium, and maximum cell yields (104 · mL−1) were similar to those attained in previous research when P. shumwayae was cultured with living fish or microalgae. Additionally, P. shumwayae flagellate cells ingested particles present in the biphasic medium, allowing detailed observations of feeding behavior. This research is an initial step toward a chemically defined axenic culture medium and determination of P. shumwayae metabolic requirements.

Chapter 17 – Dinoflagellates

Dinoflagellates are a group of unicellular protists that can be identified using the light microscope, and are (usually) recognized by their golden-brown plastids, assimilative cell with indented waist, distinctive swimming pattern, and relatively large nucleus that contains visible chromosomes. Shared characteristics of the group include: two dissimilar flagella in motile cells; an unusual genome consisting of liquid, crystal-like chromosomes; and a haplontic life cycle that often contains a non-motile, dormant cyst stage. Freshwater dinoflagellates are diverse and may be phototrophic, predatory, or both (mixotrophic). They can be found in most aquatic habitats throughout the seasons; although flagship taxa, such as Ceratium, Peridinium, and Gymnodinium, are typically found as phytoplankton in calm, lentic waters during warmer months. This chapter updates taxonomy (extensive splitting of the genera Peridinium, Peridiniopsis, and Woloszynskia), identification of genera via microscopy, and adds molecular information on taxa and group systematics where available.