Bacteria and phytoplankton dynamics are thought to be closely linked in coastal marine environments, with correlations frequently observed between bacterial and phytoplankton biomass. In contrast, little is known about how these communities interact with each other at the species composition level. The purpose of the current study was to analyze bacterial community dynamics in a productive, coastal ecosystem and to determine whether they were related to phytoplankton community dynamics. Near-surface seawater samples were collected in February, May, July, and September 2000 from several stations in the Bay of Fundy. Savin et al. (M.C. Savin et al., Microb Ecol 48: 51-65) analyzed the phytoplankton community in simultaneously collected samples. The attached and free-living bacterial communities were collected by successive filtration onto 5 microm and 0.22 microm pore-size filters, respectively. DNA was extracted from filters and bacterial 16S rRNA gene fragments were amplified and analyzed by denaturing gradient gel electrophoresis (DGGE). DGGE revealed that diversity and temporal variability were lower in the free-living than the attached bacterial community. Both attached and free-living communities were dominated by members of the Roseobacter and Cytophaga groups. Correspondence analysis (CA) ordination diagrams showed similar patterns for the phytoplankton and attached bacterial communities, indicating that shifts in the species composition of these communities were linked. Similarly, canonical CA revealed that the diversity, abundance, and percentage of diatoms in the phytoplankton community accounted for a significant amount of the variability in the attached bacterial community composition. In contrast, ordination analyses did not reveal an association between free-living bacteria and phytoplankton. These results suggest that there are specific interactions between phytoplankton and the bacteria attached to them, and that these interactions influence the composition of both communities.

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Phytoplankton have traditionally been identified based on morphological characteristics. However, identifications based on morphology are time-consuming, require expertise in taxonomy, and often fail to distinguish differences among the multitudes of minute, nondescript planktonic organisms. Molecular techniques, which have revealed new insights into bacterial and picoplankton communities, may also enhance our knowledge of the diversity among communities of larger plankton. We compared plankton identifications and community assessments based on the two types of techniques (morphological vs molecular) for surface seawater samples collected on 2 May, 31 July and 25 September 2000 from several sampling stations in the Bay of Fundy. Phytoplankton captured in surface bucket samples were quantified and identified based on morphology. DNA was extracted from plankton communities (5-100 microm in diameter) collected by filtration, and 18S rRNA gene fragments were amplified with primers specific for eukaryotes. Denaturing gradient gel electrophoresis (DGGE) was used to develop DNA profiles of eukaryotic phylogenetic diversity and to select cloned 18S rDNA fragments for sequencing. Both morphological and molecular methods showed great community diversity. However, the communities identified with the two different types of techniques were starkly different. Morphological abundances and taxon richness were lowest in the May samples, whereas the number of DGGE bands was highest in May and July. Morphological identifications showed a succession of dominant organisms through time. Whereas neither diatoms nor dinoflagellates were dominant in May, diatoms and a few dinoflagellates were dominant in July and September. In contrast, few 18S rDNA sequences were related to rDNA sequences of known identity, and furthermore, few diatoms were identified in the molecular analyses. Molecular phylogenetic analysis indicated the presence of many novel organisms, several of which were most closely related to other unidentified sequences from diverse marine environments representing new lineages. Our results support the ideas that we are just beginning to uncover the diversity of eukaryotic marine organisms and that there may be many more ubiquitous, microeukaryotic plankton than previously realized. Our results suggest that both types of methods capture only a portion of the community. Morphological methods may be more adept at capturing the phototrophic organisms within the community. However, just as for bacteria and picoplankton, molecular techniques can enhance our understanding of plankton diversity, particularly by detecting previously unidentified organisms.

A workshop with the aim to compare classical and molecular techniques for phytoplankton enumeration took place at Kristineberg Marine Research Station, Sweden, in August 2005. Seventeen different techniques -nine classical microscopic-based and eight molecular methods -were compared. Alexandrium fundyense was the target organism in four experiments. Experiment 1 was designed to determine the range of cell densities over which the methods were applicable. Experiment 2 tested the species specificity of the methods by adding Alexandrium ostenfeldii, to samples containing A. fundyense. Experiments 3 and 4 tested the ability of the methods to detect the target organism within a natural phytoplankton community. Most of the methods could detect cells at the lowest concentration tested, 100 cells L À1 , but the variance was high for methods using small volumes, such as counting chambers and slides. In general, the precision and reproducibility of the investigated methods increased with increased target cell concentration. Particularly molecular methods were exceptions in that their relative standard deviation did not vary with target cell concentration. Only two of the microscopic methods and three of the molecular methods had a significant linear relationship between their cell count estimates and the A. fundyense concentration in experiment 2, where the objective was to discriminate that species from a morphologically similar and genetically closely related species. None of the investigated methods were affected by the addition of a natural plankton community background matrix in experiment 3. The results of this study are discussed in the context of previous intercomparisons and the difficulties in defining the absolute, true target cell concentration. #

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The goal of this study was to compare the ability of morphology and molecular-based surveys to estimate species richness for two species-rich diatom genera, Chaetoceros Ehrenb. and Thalassiosira Cleve, in the Bay of Fundy. Phytoplankton tows were collected from two sites at intervals over two years and subsampled for morphology-based surveys (2010, 2011), a culture-based DNA reference library (DRL; 2010), and a molecular-based survey (2011). The DRL and molecular-based survey utilized the 3′ end of the RUBISCO large subunit (rbcL-3P) to identify genetic species groups (based on 0.1% divergence in rbcL-3P), which were subsequently identified morphologically to allow comparisons to the morphology-based survey. Comparisons were compiled for the year (2011) by site (n = 2) and by season (n = 3). Of the 34 taxa included in the comparisons, 50% of taxa were common to both methods, 35% were unique to the molecular-based survey, and 12% were unique to the morphology-based survey, while the remaining 3% of taxa were unidentified genetic species groups. The morphology-based survey excelled at identifying rare taxa in individual tow subsamples, which were occasionally missed with the molecular approach used here, while the molecular methods (the DRL and molecular-based survey), uncovered nine cryptic species pairs and four previously overlooked species. The last mentioned were typically difficult to identify and were generically assigned to Thalassiosira spp. during the morphology-based survey. Therefore, for now we suggest a combined approach encompassing routine morphology-based surveys accompanied by periodic molecular-based surveys to monitor for cryptic and difficult to identify taxa. As sequencing technologies improve, molecular-based surveys should become routine, leading to a more accurate representation of species composition and richness in monitoring programs.