The grazlng unpact of 3 different protozoan specles on a muted bacter~al community was studled by means of a s~m p h i~e d and funct~onally reproducible expenmental m~crobial food web In a 2-stage flow-through system In the first stage the algae Rhodomonas sp was grown on an inorganic medlurn wlth its accompanying bacterial community (BC) growing on algal exudates T h~s m t u r e of algae and bacterla was transferled Into 4 second stage vessels (1) a control and 3 vessels inoculated wlth (2) a heterotrophic nanoflagellate Bodo saltans (3) a scuticoc~l~ate, Cyclldlum glaucoma, and (4) a mxotroph~c flagellate, Ochromonas sp Us~ng Image analysis techn~ques we followed the changes In bactenal s u e &stnbut~ons and bactenal to protozoan total biovolume rat~os over an expermental period of 15 d In add~tion productivity of the grazed and ungrazed BC was measured using [ 3~] t h y m~-dine and ['"C]leuc~ne As a consequence of total grazing rates and size-select~ve feedlng we observed 3 d~fferent responses of the initially ident~cal BC to grazing of the 3 protists (1) LOW grazlng by B saltans caused a slow decrease of bactenal cell numbers from 14 to 5 9 X 10h cells m -' , but no s~gnlfl-cant shlft of the mean cell volume (MCV, average 0 107 pm3) and bacter~al product~on (2) H~g h e r grazing rates of C glaucoma resulted In the dechne o! bactenal abundance to 3 3 X 106 cells ml-' In parallel with a doubllng of the MCV to 0 207 pm3 and high DNA and protein synthes~s rates Due to the c h a t e S ab~lity to graze also on small prey ( < l 5 pm) an increase in MCV seemed to provide h l g h e~ grazing resistance or at least decreased vulnerab~lity. (3) Ochromonas sp. showed the hlghest grazlng rates and reduced bacterial numbers by 20 times wlthin 2 d. Only the s m a l l e s t o b v i o u s l y more grazing-protected-bacteria (<0.9 pm) survived, which increased thereafter to almost initial bacterial densities Although the bacterial MCV dropped to 0.038 pm3, [3H]thymidine uptake rates per cell were greatly enhanced and highly variable. Our results reflect the potential of BC responses to different predation regimes and the advantages of phenotypic traits in order to coexlst with various grazers This should be seen in the context of influencing bottom-up effects and the varying potential of individual bacter~al specles to change morphology, growth strategies, and activ~ty patterns.

Analyses of high-throughput environmental sequencing data have become the ‘gold-standard’ to address fundamental questions of microbial diversity, ecology and biogeography. Findings that emerged from sequencing are, e.g. the discovery of the extensive ‘rare microbial biosphere’ and its potential function as a seed-bank. Even though applied since several years, results from high-throughput environmental sequencing have hardly been validated. We assessed how well pyrosequenced amplicons [the hypervariable eukaryotic V4 region of the small subunit ribosomal RNA (SSU rRNA) gene] reflected morphotype ciliate plankton. Moreover, we assessed if amplicon sequencing had the potential to detect the annual ciliate plankton stock. In both cases, we identified significant quantitative and qualitative differences. Our study makes evident that taxon abundance distributions inferred from amplicon data are highly biased and do not mirror actual morphotype abundances at all. Potential reasons included cell losses after fixation, cryptic morphotypes, resting stages, insufficient sequence data availability of morphologically described species and the unsatisfying resolution of the V4 SSU rRNA fragment for accurate taxonomic assignments. The latter two underline the necessity of barcoding initiatives for eukaryotic microbes to better and fully exploit environmental amplicon data sets, which then will also allow studying the potential of seed-bank taxa as a buffer for environmental changes.

We simultaneously studied the impact of top-down (protistan grazing) and bottom-up (phosphorus availability) factors on the numbers and biomasses of bacteria from various phylogenetic lineages, and on their growth and activity parameters in the oligo-mesotrophic Piburger See, Austria. Enhanced grazing resulted in decreased proportions of bacteria with high nucleic acid content (high-NA bacteria) and lower detection rates by FISH. There was a change in the composition of the bacterial assemblage, whereby Betaproteobacteria were heavily grazed while Alphaproteobacteria and Cytophaga-Flavobacterium-Bacteroides were less affected by predators. Changes in bacterial assemblage composition were also apparent in the treatments enriched with phosphorus, and even more pronounced in the incubations in dialysis tubes (allowing relatively free nutrient exchange). Here, Betaproteobacteria became dominant and appeared to act as successful opportunistic competitors for nutrients. In contrast, Actinobacteria did not respond to surplus phosphorus by population growth, and, moreover, maintained their small size, which resulted in a very low biomass contribution. In addition, significant relationships between high-NA bacteria and several bacterial phylogenetic clades were found, indicating an enhanced activity status. By combining several single-cell methods, new insight is gained into the competitive abilities of freshwater bacteria from a variety of phylogenetic lineages under contrasting sets of bottom-up and top-down constraints.

We followed the changes in the protist assemblage over an annual cycle at 3 sites and different depths of Traunsee in the Austrian Alps and quantified the variability of the ciliate assemblage along successive depth and time intervals, respectively. More than 60 ciliate species were identified alive and after quantitative protargol staining (QPS). The ciliate diversity was high, and is described in detail for 50 taxa in time-depth intervals. Rimostrombidium brachykinetum/Rimostrombidium hyalinum and Balanion planctonicum were the most frequent species, accounting for 44% of the annual mean abundance and 15% of the annual mean biovolume, respectively. Our results suggested a stronger variability in the ciliate assemblage structure within seasons than along the depth gradient. Gradual changes in the assemblage structure with depths: (1) were accompanied by a decrease of algivorous and mixotrophic and an increase of bacterivorous ciliates from surface to deeper layers; (2) were highly significantly altered with steep depth gradients of their potential food resources, i.e. the biomass of heterotrophic flagellates, bacteria and algae; and (3) were related with lower significance to environmental parameters. High similarity (> 80%) between successive months was reached only when net changes in the total ciliate abundance were negligible, while a strong increase or decrease in the ciliate abundance was associated with pronounced changes in the species composition. These seasonal changes in the ciliate assemblage structure (4) were linked to shifts of algivorous and mixotrophic, but not of bacterivorous ciliates and (5) were less predictable with food resources compared to the depth gradient. The phagotrophic flagellates generally followed the seasonal and vertical patterns described for ciliates, and were shown to be important members of the planktonic food web in a cold, deep, oligotrophic lake.

We analyzed the genetic diversity (V4 region of the 18S rRNA) of planktonic microbial eukaryotes in four high mountain lakes including two remote biogeographic regions (the Himalayan mountains and the European Alps) and distinct habitat types (clear and glacier-fed turbid lakes). The recorded high genetic diversity in these lakes was far beyond of what is described from high mountain lake plankton. In total, we detected representatives from 66 families with the main taxon groups being Alveolata (55.0% OTUs97%, operational taxonomic units), Stramenopiles (34.0% OTUs97%), Cryptophyta (4.0% OTUs97%), Chloroplastida (3.6% OTUs97%) and Fungi (1.7% OTUs97%). Centrohelida, Choanomonada, Rhizaria, Katablepharidae and Telonema were represented by <1% OTUs97%. Himalayan lakes harbored a higher plankton diversity compared to the Alpine lakes (Shannon index). Community structures were significantly different between lake types and biogeographic regions (Fisher exact test, P < 0.01). Network analysis revealed that more families of the Chloroplastida (10 vs 5) and the Stramenopiles (14 vs 8) were found in the Himalayan lakes than in the Alpine lakes and none of the fungal families was shared between them. Biogeographic aspects as well as ecological factors such as water turbidity may structure the microbial eukaryote plankton communities in such remote lakes.