Description: Dilution experiments were used to examine growth and grazing mortality rates among Prochlorococcus and Synechococcus populations in the Sargasso Sea and California Current. In these experiments, deviation from linearity in the relationship between dilution and net growth rate was significant in a large number of cases. An alternative, more conservative approach for estimating growth and grazing mortality rates (independent of the shape of this relationship) was therefore employed. Growth rates estimated by this approach ranged from 0.32 to 0.76 and 0.37 to 0.67 d-1 for Prochlorococcus and Synechococcus, respectively. Grazing mortality rates ranged from 0.25 to 0.85 and 0.13 to 0.51 d-1, respectively. Cell-cycle-based growth rate estimates were consistent with these dilution-based rates. Nutrient amendment had little affect on picocyanobacterial growth rates, but did stimulate grazing mortality (and in some cases changed the apparent functional response of the grazer community) in a number of experiments. We hypothesize that improved food quality in nutrient-replete picoplankton cells may be responsible for these changes. Diel patterns of picocyanobacterial abundance in the Sargasso Sea experiments suggest that grazing activity varied strongly over the diel cycle, with low grazing activity during the first half of the light period. Growth rate and abundance were not positively correlated among or within picocyanobacterial groups, as might be expected if physiologically mediated controls were the dominant forces regulating these populations.

Description: Pico- and ultraplankton are known to contribute significantly to overall biomass and primary productivity in the 'high nutrient low chlorophyll' waters of the equatorial Pacific. In order to understand the dynamics of this community on ecologically relevant time-scales, we examined the abundance, distribution and cellular characteristics of Prochlorococcus, Synechococcus, eukaryotic ultraphytoplankton and heterotrophic bacteria during two 20-day time-series at 0°N, 140°W in the spring and fall of 1992 (JGOFS time-series cruises. TS-I and TS-II). Prochlorococcus was numerically dominant among the autotrophic groups considered, with mean cell concentrations in surface waters on the order of 1.4 x 105 cells ml-1. Synechococcus and ultraphytoplankton abundances were 17-30-fold lower than those of Prochlorococcus, and heterotrophic bacterial abundances were 5-7-fold higher (during TS-I and TS-II, respectively). Daily cell abundances for all groups varied by factors of 1.5-2 within each time-series. Depth-integrated Prochlorococcus abundance averaged over each time-series was 25 lower during TS-II relative to TS-I; ultraphytoplankton abundance was 42 higher during the same period. Prochlorococcus and ultraphytoplankton both contributed significantly to the estimated total autotrophic biomass; Synechococcus contributed relatively little. Estimated total photosynthetic pico- plus ultraplankton biomass was on average 30 higher than heterotrophic bacterial biomass. Changes in the fluorescence and light scatter properties of individual Prochlorococcus cells were observed during the passage of a tropical instability wave during TS-II, and are hypothesized to reflect a physiological response among these cells to that event Examination of bulk properties alone (e.g. cell numbers or total red fluorescence) would not have revealed these physiological changes. Lower bounds for Prochlorococcus-specific growth rates were calculated based on the DNA distributions of these populations at dusk. These rates were maximal at 15 or 30 m depth, where they approached one doubling per day. Changes in Prochlorococcus forward tingle light scatter (FALS) from dawn to dusk were well correlated with these estimates of specific growth rate, an observation that allowed us to relate measurements of FALS to cell volume for Prochlorococcus.

Description: The cyanobacteria Synechococcus and Prochlorococcus are important primary producers in marine ecosystems. Because currently available approaches for estimating microbial growth rates can be difficult to apply in the field, we have been exploring the feasibility of using quantitative rRNA measurements as the basis for making such estimates. In this study we examined the relationship between rRNA and growth rate in several Synechococcus and Prochlorococcus strains over a range of light-regulated growth rates. Whole-cell hybridization with fluorescently labeled peptide nucleic acid (PNA) probes was used in conjunction with flow cytometry to quantify rRNA on a per cell basis. This PNA probing technique allowed rRNA analysis in a phycoerythrin-containing Synechococcus strain (WH7803) and in a non-phycoerythrincontaining strain and in Prochlorococcus. All the strains showed a qualitatively similar tri-phasic relationship between rRNA·cell-1 and growth rate, involving relatively little change in rRNA·cell-1 at low growth rates, linear increase at intermediate growth rates, and a plateau and/or decrease at the highest growth rates. The onset of each phase was associated with the relative, rather than absolute, growth rate of each strain. In the Synechococcus strains, rRNA normalized to flow cytometrically measured forward angle light scatter (an indicator of size) was well-correlated with growth rate across strains. These findings support the idea that cellular rRNA may be useful as an indicator of in situ growth rate in natural Synechococcus and Prochlorococcus populations.

Description: Among eukaryotes, four major phytoplankton lineages are responsible for marine photosynthesis; prymnesiophytes, alveolates, stramenopiles, and prasinophytes. Contributions by individual taxa, however, are not well known, and genomes have been analyzed fromonly the latter two lineages. Tiny "picoplanktonic" members of the prymnesiophyte lineage have long been inferred to be ecologically important but remain poorly characterized. Here, we examine pico-prymnesiophyte evolutionary history and ecology using cultivation-independent methods. 18S rRNA gene analysis showed picoprymnesiophytes belonged to broadly distributed uncultivated taxa. Therefore, we used targeted metagenomics to analyze uncultured pico-prymnesiophytes sorted by flow cytometry from subtropical North Atlantic waters. The data reveal a composite nuclear-encoded gene repertoire with strong green-lineage affiliations, which contrasts with the evolutionary history indicated by the plastid genome. Measured pico-prymnesiophyte growth rates were rapid in this region, resulting in primary production contributions similar to the cyanobacterium Prochlorococcus. On average, pico-prymnesiophytes formed 25 of global picophytoplankton biomass, with differing contributions in five biogeographical provinces spanning tropical to subpolar systems. Elements likely contributing to success include high gene density and genes potentially involved in defense and nutrient uptake. Our findings have implications reaching beyond pico-prymnesiophytes, to the prasinophytes and stramenopiles. For example, prevalence of putative Ni-containing superoxide dismutases (SODs), instead of Fe-containing SODs, seems to be a common adaptation among eukaryotic phytoplankton for reducing Fe quotas in low-Fe modern oceans. Moreover, highly mosaic gene repertoires, although compositionally distinct for each major eukaryotic lineage, now seem to be an underlying facet of successful marine phytoplankton.

Description: A simple method for whole-cell hybridization using fluorescently labeled rRNA-targeted peptide nucleic acid (PNA) probes was developed for use in marine cyanobacterial picoplankton. In contrast to established protocols, this method is capable of detecting rRNA in Prochlorococcus, the most abundant unicellular marine cyanobacterium. Because the method avoids the use of alcohol fixation, the chlorophyll content of Prochlorococcus cells is preserved, facilitating the identification of these cells in natural samples. PNA probe- conferred fluorescence was measured flow cytometrically and was always significantly higher than that of the negative control probe, with positive/negative ratio varying between 4 and 10, depending on strain and culture growth conditions. Prochlorococcus cells from open ocean samples were detectable with this method. RNase treatment reduced probe-conferred fluorescence to background levels, demonstrating that this signal was in fact related to the presence of rRNA. In another marine cyanobacterium, Synechococcus, in which both PNA and oligonucleotide probes can be used in whole-cell hybridizations, the magnitude of fluorescence from the former was fivefold higher than that from the latter, although the positive/negative ratio was comparable for both probes. In Synechococcus cells growing at a range of growth rates (and thus having different rRNA concentrations per cell), the PNA- and oligonucleotide-derived signals were highly correlated (r = 0.99). The chemical nature of PNA, the sensitivity of PNA-RNA binding to single-base-pair mismatches, and the preservation of cellular integrity by this method suggest that it may be useful for phylogenetic probing of whole cells in the natural environment.

Description: An efficient numerical method to compute nonlinear solutions for 2D steady free-surface flow over an arbitrary channel bottom topography is presented. The approach is based on a boundary integral equation technique which is similar to that of Vanden-Broeck's (1996). The typical approach for this problem is to prescribe the shape of the channel bottom topography, with the free surface being provided as part of the solution. Here we take an inverse approach and prescribe the shape of the free surface a priori and solve for the corresponding bottom topography. We show how this inverse approach is particularly useful when studying topographies that give rise to wave-free solutions, allowing us to easily classify 11 basic flow types. Finally, the inverse approach is also adapted to calculate a distribution of pressure on the free surface, given the free surface shape itself.

Description: Pseudohyphal growth of the dimorphic yeast Saccharomyces cerevisiae is analysed using two-dimensional top-down binary images. The colony morphology is characterized using clustered shape primitives (CSPs), which are learned automatically from the data and thus do not require a list of predefined features or a priori knowledge of the shape. The power of CSPs is demonstrated through the classification of pseudohyphal yeast colonies known to produce different morphologies. The classifier categorizes the yeast colonies considered with an accuracy of 0.969 and standard deviation 0.041, demonstrating that CSPs capture differences in morphology, while CSPs are found to provide greater discriminatory power than spatial indices previously used to quantify pseudohyphal growth. The analysis demonstrates that CSPs provide a promising avenue for analysing morphology in high-throughput assays.