Food selection by bacterivorous protists: insight from the analysis of the food vacuole content by means of fluorescence in situ hybridization

A modified fluorescence in situ hybridization (FISH) method was used to analyze bacterial prey composition in protistan food vacuoles in both laboratory and natural populations. Under laboratory conditions, we exposed two bacterial strains (affiliated with beta- and gamma-Proteobacteria -- Aeromonas hydrophila and Pseudomonas fluorescens, respectively) to grazing by three protists: the flagellates Bodo saltans and Goniomonas sp., and the ciliate Cyclidium glaucoma. Both flagellate species preferably ingested A. hydrophila over P. fluorescens, while C. glaucoma showed no clear preferences. Differences were found in the digestion of bacterial prey with B. saltans digesting significantly faster P. fluorescens compared to two other protists. The field study was conducted in a reservoir as part of a larger experiment. We monitored changes in the bacterial prey composition available compared to the bacteria ingested in flagellate food vacuoles. Bacteria detected by probe HGC69a (Actinobacteria) and R-BT065 were negatively selected by flagellates. Bacteria detected by probe CF319a were initially positively selected but along with a temporal shift in bacterial cell size, this trend changed to negative selection during the experiment. Overall, our analysis of protistan food vacuole content indicated marked effects of flagellate prey selectivity on bacterioplankton community composition.     

The new higher level classification of eukaryotes with emphasis on the taxonomy of protists

This revision of the classification of unicellular eukaryotes updates that of Levine et al. (1980) for the protozoa and expands it to include other protists. Whereas the previous revision was primarily to incorporate the results of ultrastructural studies, this revision incorporates results from both ultrastructural research since 1980 and molecular phylogenetic studies. We propose a scheme that is based on nameless ranked systematics. The vocabulary of the taxonomy is updated, particularly to clarify the naming of groups that have been repositioned. We recognize six clusters of eukaryotes that may represent the basic groupings similar to traditional "kingdoms." The multicellular lineages emerged from within monophyletic protist lineages: animals and fungi from Opisthokonta, plants from Archaeplastida, and brown algae from Stramenopiles.     

Feeding preferences and grazing rates of Pfiesteria piscicida and a cryptoperidiniopsoid preying on fish blood cells and algal prey

The grazing rates and feeding preferences of the dinoflagellates Pfiesteria piscicida and a cryptoperidiniopsoid on the alga Rhodomonas sp. and fish blood cells were calculated at different ratios of the two food types and at different total food densities. Data from 6 h grazing periods within microcosms were used to calculate grazing rates. Grazing rates of both dinoflagellates increased linearly with an increased ratio of blood cells to Rhodomonas, and P. piscicida had a higher maximum grazing rate than the cryptoperidiniopsoid. The grazing rate of P. piscicida on Rhodomonas also increased with increased Rhodomonas densities relative to the blood cells, but increased densities of Rhodomonas did not increase the grazing rate of the cryptoperidiniopsoid, suggesting a lower feeding threshold for this species. Both dinoflagellates demonstrated a preference for fish blood cells over Rhodomonas cells, with no significant difference in the index of preference between the two species. Total food abundance affected the degree of preference differently for each dinoflagellate species. A higher index of feeding preference was attained by P. piscicida when resource levels were high, while the cryptoperidiniopsoid did not show this response. A preference for fish blood cells occurred at all food ratios for both dinoflagellates, including when blood cells were scarce relative to the alternate food type (15% of total available food). These results suggest that these strains of P. piscicida and the cryptoperidiniopsoid share similar feeding preferences for the prey types tested, although cryptoperidiniopsoids have not been associated with fish kills.     

Effects of N and P supply on phytoplankton in Bizerte Lagoon (western Mediterranean)

Enriched bottle experiments were conducted in situ during winter (January and February) and summer (July and August) 2001 to examine the effects of nutrient enrichments (+ N, + P and + NP) on phytoplankton in Bizerte Lagoon, Tunisia. Chlorophyll a (Chl a), ranging from 3.05 μg L⁻¹ in winter to 4.52 μg L⁻¹ in summer, was dominated by the small size-faction (<5 μm) during both seasons. However, the contribution of the large size-fraction (5-200 μm) to Chl a increased from winter (26%) to summer (37%). Similarly, the carbon biomass of the 5-200 μm algae increased during the July/August period that was characterised by the high proliferation of several diatom taxa. In winter, N was the limiting element for phytoplankton growth. Its addition alone (+ N) or with P (+ NP) increased both the <5 μm and 5-200 μm Chl a concentrations. There was no change in the phytoplankton size structure, with the small cells dominating the final algal biomass in all treatments after 5 days. In summer, N and P limited the phytoplankton, but small and large algae exhibited diverse responses to different nutrient enrichments: addition of P increased the Chl a only in the 5-200 μm fraction, the + N treatment enhanced both size classes, and the NP fertilisation mostly stimulated the biomass of large cells. Consequently, the N and P addition in summer was followed by a significant change in the phytoplankton size structure, since both size-fractions contributed equally to the final Chl a biomass. Within the 5-200 μm algal community, various taxa had diverse responses to the nutrient supply during both seasons, leading to a change in the final community composition. The autotrophic flagellates appeared to grow well under N-deficient conditions. In contrast, diatom growth and biomass were mostly stimulated by the N enrichment while dinoflagellates exhibited the highest increase in their growth and biomass with P fertilisation. Our results suggest that the increasing anthropogenic supply of nutrients in the lagoon may influence algal dynamics as well as productivity in different ways depending on the nutrient composition.     

The marine pathogenic genotype of Spironucleus barkhanus from farmed salmonids redescribed as Spironucleus salmonicida n. sp

There are two genotypes of the diplomonad Spironucleus barkhanus. Based on sequence data from the small subunit ribosomal RNA gene the conspecificity of these two genotypes has been questioned. Therefore, we have sampled Spironucleus from 27 fish, representing 14 populations, five species, and four genera. Partial nucleotide sequences from the three genes; small subunit ribosomal DNA, glutamate dehydrogenase 1 and alpha-tubulin were compared. The pathogenic isolates of S. barkhanus, which causes systemic spironucleosis in Atlantic salmon, Chinook salmon, and Arctic charr, all farmed in sea water, were genetically very different from the commensal isolate found in wild freshwater populations of Arctic charr and grayling. The genetic distances between the genotypes were of the same magnitude as those separating species of Giardia. Based on these genetic and ecological data, the pathogenic genotype from farmed salmonids is described as a new species, Spironucleus salmonicida n. sp. Scanning and transmission electron microscopy showed no specific morphological or ultrastructural features distinguishing S. salmonicida n. sp. from S. barkhanus. The present study clearly demonstrates the value of applying genetics in identification of Spironucleus species. Phylogenetic analyses that included the isolates of S. salmonicida n. sp. did not change the phylogenetic relationship within the genus Spironucleus.     

The Revised Classification of Eukaryotes

This revision of the classification of eukaryotes, which updates that of Adl et al. [J. Eukaryot. Microbiol. 52 (2005) 399], retains an emphasis on the protists and incorporates changes since 2005 that have resolved nodes and branches in phylogenetic trees. Whereas the previous revision was successful in re‐introducing name stability to the classification, this revision provides a classification for lineages that were then still unresolved. The supergroups have withstood phylogenetic hypothesis testing with some modifications, but despite some progress, problematic nodes at the base of the eukaryotic tree still remain to be statistically resolved. Looking forward, subsequent transformations to our understanding of the diversity of life will be from the discovery of novel lineages in previously under‐sampled areas and from environmental genomic information.     

Growth-associated biosynthesis of astaxanthin in heterotrophic Chlorella zofingiensis (Chlorophyta)

The green microalga Chlorella zofingiensis can produce the ketocarotenoid astaxanthin under heterotrophic culture conditions. Here we report the growth-associated biosynthesis of astaxanthin in this biotechnologically important alga. With glucose as sole carbon and energy source, C. zofinginesis grew fast in the dark with rapid exhaustion of nitrogen and carbon sources from media, leading to a high specific growth rate (0.034 h⁻¹). Cultures started at a cell concentration of about 3.4 × 10⁹ cells l⁻¹ reached, after 6 days, standing biomass values of 1.6 × 10¹¹ cells or 8.5 g dry weight l⁻¹. Surprisingly, the biosynthesis of astaxanthin was found to start at early exponential phase, independent of cessation of cell division. A general trend was observed that the culture conditions benefiting cell growth also benefited astaxanthin accumulation, indicating that astaxanthin was a growth-associated product in this alga. The maximum cell dry biomass and astaxanthin yield were 11.75 g l⁻¹ and 11.14 mg l⁻¹ (about 1 mg g⁻¹), simultaneously obtained in the fed-batch culture with a combined glucose–nitrate mixture addition, which were the highest ever reported in dark-heterotrophic algal cultures. The possible reasons why dark-heterotrophic C. zofingiensis could produce astaxanthin during the course of cell growth were discussed.     

Pico- and nanoplankton dynamics during bloom initiation of Aureococcus in a Long Island, NY bay

The entire microbial plankton community was quantified on a weekly basis April through June of 2000 in Quantuck Bay as part of an ongoing study to identify factors contributing to the initiation of blooms of Aureococcus anophagefferens (brown tide) in Long Island, NY bays. We used flow cytometry, imaging cytometry, fluorescent antibody cell counts, and traditional visual cell counting to quantify the picophytoplankton, heterotrophic bacteria, nanophytoplankton, heterotrophic protists, and microplankton prior to, and during the initiation of a brown tide bloom. Cells passing through a 5 μm mesh dominated the total chlorophyll concentration (>80%) for most of the spring study period. The A. anophagefferens bloom occurred in the context of a larger pico/nanophytoplankton bloom where A. anophagefferens accounted for only 30% of the total cell count when it was at its maximum concentration of 4.8 × 10⁵ mL⁻¹. Levels of dissolved organic nitrogen were enriched during the bloom peak relative to pre-bloom levels and heterotrophic bacteria also bloomed, reaching abundances over 10⁷ mL⁻¹. A trophic cascade within the heterotrophic protist community may have occurred, coinciding with the A. anophagefferens bloom. Before the onset of the bloom, larger grazers increased in abundance, while the next smaller trophic level of grazers were diminished. These smaller grazers were the likely water column predators of A. anophagefferens, and the brown tide bloom initiated when they were depleted. These results suggest that this bloom initiated due to interactions with other pico/nano algae and release from grazing pressure through a trophic cascade.     

表达 lac 和 glk 基因的蓝藻的异养生长

将来自Brucella abortu的葡萄糖激酶基因glk和大肠杆菌乳糖操纵子基因lacZYA装置于蓝藻glnA启动子下游,以广宿主质粒导入3种蓝藻。基因的表达通过β-半乳糖苷酶和葡糖激酶活性测定得到确证。基因工程藻株均获得了利用乳糖异养生长的能力。集胞藻6803转基因株可在光照加DCMU的条件下利用乳糖生长,而鱼腥藻转基因株依赖乳糖可在完全黑暗条件下生长,并可在黑暗或加DCMU条件下诱导产生异形胞。     

Flagellate nutritional versatility as a key to survival in two contrasting Antarctic saline lakes

1. Seasonal patterns of grazing and photosynthesis were investigated in two saline Antarctic lakes (Highway and Ace) in the Vestfold Hills (68°S). The phototrophic nanoflagellate (PNAN) community was dominated by Pyramimonas gelidicola and two morphological forms of a cryptophyte species that occurred throughout the year. Both species were mixotrophic on bacteria, and in Highway Lake they also exploited dissolved organic carbon as determined by the uptake of fluorescently labelled dextrans. 2. Clearance rates ranged between 0.02 and 0.21 nL h^?1 cell^?1 in Ace Lake and 0.004–1.05 nL h^?1 cell^?1 in Highway Lake. On occasion cryptophyte grazing equalled that of the heterotrophic nanoflagellates (HNAN). 3. Photosynthetic rates showed similar trends in both lakes, but there were differences in chlorophyll a specific rates and photosynthetic efficiency, probably related to the meromictic characteristic of Ace Lake. Primary production was measurable in winter and peaked in summer following the maxima of mixotroph grazing. 4. The HNAN community of Highway Lake achieved clearance rates of 0.02–1.80 nL h^?1 cell^?1 and removing between 50 and 693 ng bacterial carbon L^?1 day^?1, with highest impact in winter when HNAN were most abundant. The HNAN also ingested fluorescently labelled dextrans showing a preference for 4 and 500 kDa molecules. The more diverse HNAN community of Ace Lake had lower clearance rates (0.04–0.37 nL h^?1 cell^?1) and exerted a lower grazing pressure on bacterioplankton. In Highway Lake, where the HNAN community was dominated by the choanoflagellate Diaphanoeca grandis, there was a significant correlation between mean cell volume and clearance rate. 5. The major feature was that the microbial plankton functioned throughout the year by employing nutritional versatility.