Most cited article - PubMed ID 23552621
Differential freshwater flagellate community response to bacterial food quality with a focus on Limnohabitans bacteria
Microbial communities, which include prokaryotes and protists, play an important role in aquatic ecosystems and influence ecological processes. To understand these communities, metabarcoding provides a powerful tool to assess their taxonomic composition and track spatio-temporal dynamics in both marine and freshwater environments. While marine ecosystems have been extensively studied, there is a notable research gap in understanding eukaryotic microbial communities in temperate lakes. Our study addresses this gap by investigating the free-living bacteria and small protist communities in Lake Roś (Poland), a dimictic temperate lake. Metabarcoding analysis revealed that both the bacterial and protist communities exhibit distinct seasonal patterns that are not necessarily shaped by dominant taxa. Furthermore, machine learning and statistical methods identified crucial amplicon sequence variants (ASVs) specific to each season. In addition, we identified a distinct community in the anoxic hypolimnion. We have also shown that the key factors shaping the composition of analysed community are temperature, oxygen, and silicon concentration. Understanding these community structures and the underlying factors is important in the context of climate change potentially impacting mixing patterns and leading to prolonged stratification.
- Keywords
- abiotic factors, freshwater environments, prokaryotes, protists, stratification, temporal dynamics,
- MeSH
- Bacteria * genetics classification MeSH
- Biodiversity MeSH
- Spatio-Temporal Analysis MeSH
- Ecosystem MeSH
- Eukaryota * genetics classification MeSH
- Lakes * microbiology MeSH
- Microbiota * MeSH
- Seasons MeSH
- Machine Learning * MeSH
- DNA Barcoding, Taxonomic * MeSH
- Temperature MeSH
- Publication type
- Journal Article MeSH
Heterotrophic nanoflagellates (HNF) and ciliates are major protistan planktonic bacterivores. The term HNF, however, describes a functional guild only and, in contrast to the morphologically distinguishable ciliates, does not reflect the phylogenetic diversity of flagellates in aquatic ecosystems. Associating a function with taxonomic affiliation of key flagellate taxa is currently a major task in microbial ecology. We investigated seasonal changes in the HNF and ciliate community composition as well as taxa-specific bacterivory in four hypertrophic freshwater lakes. Taxa-specific catalyzed reporter deposition-fluorescence in situ hybridization probes assigned taxonomic affiliations to 51%-96% (average ±SD, 75 ± 14%) of total HNF. Ingestion rates of fluorescently labelled bacteria unveiled that HNF contributed to total protist-induced bacterial mortality rates more (56%) than ciliates (44%). Surprisingly, major HNF bacterivores were aplastidic cryptophytes and their Cry1 lineage, comprising on average 53% and 24% of total HNF abundance and 67% and 21% of total HNF bacterivory respectively. Kinetoplastea were important consumers of bacteria during summer phytoplankton blooms, reaching 38% of total HNF. Katablepharidacea (7.5% of total HNF) comprised mainly omnivores, with changing contributions of bacterivorous and algivorous phylotypes. Our results show that aplastidic cryptophytes, accompanied by small omnivorous ciliate genera Halteria/Pelagohalteria, are the major protistan bacterivores in hypertrophic freshwaters.
Diplonemids are one of the most abundant groups of heterotrophic planktonic microeukaryotes in the world ocean and, thus, are likely to play an essential role in marine ecosystems. So far, only few species have been introduced into a culture, allowing basic studies of diplonemid genetics, morphology, ultrastructure, metabolism, as well as endosymbionts. However, it remains unclear whether these heterotrophic flagellates are parasitic or free-living and what are their predominant dietary patterns and preferred food items. Here we show that cultured diplonemids, maintained in an organic-rich medium as osmotrophs, can gradually switch to bacterivory as a sole food resource, supporting positive growth of their population, even when fed with a low biovolume of bacteria. We further observed remarkable differences in species-specific feeding patterns, size-selective grazing preferences, and distinct feeding strategies. Diplonemids can discriminate between low-quality food items and inedible particles, such as latex beads, even after their ingestion, by discharging them in the form of large waste vacuoles. We also detected digestion-related endogenous autofluorescence emitted by lysosomes and the activity of a melanin-like material. We present the first evidence that these omnipresent protists possess an opportunistic lifestyle that provides a considerable advantage in the generally food resource-limited marine environments.
- MeSH
- Bacteria genetics MeSH
- Ecosystem * MeSH
- Eukaryota * MeSH
- Plankton MeSH
- Feeding Behavior MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Phagotrophic protists are key players in aquatic food webs. Although sequencing-based studies have revealed their enormous diversity, ecological information on in situ abundance, feeding modes, grazing preferences, and growth rates of specific lineages can be reliably obtained only using microscopy-based molecular methods, such as Catalyzed Reporter Deposition-Fluorescence in situ Hybridization (CARD-FISH). CARD-FISH is commonly applied to study prokaryotes, but less so to microbial eukaryotes. Application of this technique revealed that Paraphysomonas or Spumella-like chrysophytes, considered to be among the most prominent members of protistan communities in pelagic environments, are omnipresent but actually less abundant than expected, in contrast to little known groups such as heterotrophic cryptophyte lineages (e.g., CRY1), cercozoans, katablepharids, or the MAST lineages. Combination of CARD-FISH with tracer techniques and application of double CARD-FISH allow visualization of food vacuole contents of specific flagellate groups, thus considerably challenging our current, simplistic view that they are predominantly bacterivores. Experimental manipulations with natural communities revealed that larger flagellates are actually omnivores ingesting both prokaryotes and other protists. These new findings justify our proposition of an updated model of microbial food webs in pelagic environments, reflecting more authentically the complex trophic interactions and specific roles of flagellated protists, with inclusion of at least two additional trophic levels in the nanoplankton size fraction. Moreover, we provide a detailed CARD-FISH protocol for protists, exemplified on mixo- and heterotrophic nanoplanktonic flagellates, together with tips on probe design, a troubleshooting guide addressing most frequent obstacles, and an exhaustive list of published probes targeting protists.
Heterotrophic nanoflagellates (HNF) are considered as major planktonic bacterivores, however, larger HNF taxa can also be important predators of eukaryotes. To examine this trophic cascading, natural protistan communities from a freshwater reservoir were released from grazing pressure by zooplankton via filtration through 10- and 5-µm filters, yielding microbial food webs of different complexity. Protistan growth was stimulated by amendments of five Limnohabitans strains, thus yielding five prey-specific treatments distinctly modulating protistan communities in 10- versus 5-µm fractions. HNF dynamics was tracked by applying five eukaryotic fluorescence in situ hybridization probes covering 55-90% of total flagellates. During the first experimental part, mainly small bacterivorous Cryptophyceae prevailed, with significantly higher abundances in 5-µm treatments. Larger predatory flagellates affiliating with Katablepharidacea and one Cercozoan lineage (increasing to up to 28% of total HNF) proliferated towards the experimental endpoint, having obviously small phagocytized HNF in their food vacuoles. These predatory flagellates reached higher abundances in 10-µm treatments, where small ciliate predators and flagellate hunters also (Urotricha spp., Balanion planctonicum) dominated the ciliate assemblage. Overall, our study reports pronounced cascading effects from bacteria to bacterivorous HNF, predatory HNF and ciliates in highly treatment-specific fashions, defined by both prey-food characteristics and feeding modes of predominating protists.
- Keywords
- Cercozoa, Cryptophyceae, Katablepharidacea, bacterivorous and predatory flagellates, ciliates, freshwater microbial food webs,
- MeSH
- Cercozoa * MeSH
- Cryptophyta MeSH
- In Situ Hybridization, Fluorescence MeSH
- Food Chain * MeSH
- Fresh Water MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Freshwater systems are characterized by an enormous diversity of eukaryotic protists and prokaryotic taxa. The community structures in different lakes are thereby influenced by factors such as habitat size, lake chemistry, biotic interactions, and seasonality. In our study, we used high throughput 454 sequencing to study the diversity and temporal changes of prokaryotic and eukaryotic planktonic communities in three Austrian lakes during the ice-free season. In the following year, one lake was sampled again with a reduced set of sampling dates to observe reoccurring patterns. Cluster analyses (based on SSU V9 (eukaryotic) and V4 (prokaryotic) OTU composition) grouped samples according to their origin followed by separation into seasonal clusters, indicating that each lake has a unique signature based on OTU composition. These results suggest a strong habitat-specificity of microbial communities and in particular of community patterns at the OTU level. A comparison of the prokaryotic and eukaryotic datasets via co-inertia analysis (CIA) showed a consistent clustering of prokaryotic and eukaryotic samples, probably reacting to the same environmental forces (e.g., pH, conductivity). In addition, the shifts in eukaryotic and bacterioplanktonic communities generally occurred at the same time and on the same scale. Regression analyses revealed a linear relationship between an increase in Bray-Curtis dissimilarities and elapsed time. Our study shows a pronounced coupling between bacteria and eukaryotes in seasonal samplings of the three analyzed lakes. However, our temporal resolution (biweekly sampling) and data on abiotic factors were insufficient to determine if this was caused by direct biotic interactions or by reacting to the same seasonally changing environmental forces.
- Keywords
- Seasonality, diversity, eukaryotic plankton, freshwater, prokaryotic plankton, protist,
- Publication type
- Journal Article MeSH
Small bacterivorous eukaryotes play a cardinal role in aquatic food webs and their taxonomic classification is currently a hot topic in aquatic microbial ecology. Despite increasing interest in their diversity, core questions regarding predator-prey specificity remain largely unanswered, e.g., which heterotrophic nanoflagellates (HNFs) are the main bacterivores in freshwaters and which prokaryotes support the growth of small HNFs. To answer these questions, we fed natural communities of HNFs from Římov reservoir (Czech Republic) with five different bacterial strains of the ubiquitous betaproteobacterial genera Polynucleobacter and Limnohabitans. We combined amplicon sequencing and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH) targeting eukaryotic 18 S rRNA genes to track specific responses of the natural HNF community to prey amendments. While amplicon sequencing provided valuable qualitative data and a basis for designing specific probes, the number of reads was insufficient to accurately quantify certain eukaryotic groups. We also applied a double-hybridization technique that allows simultaneous phylogenetic identification of both predator and prey. Our results show that community composition of HNFs is strongly dependent upon prey type. Surprisingly, Cryptophyta were the most abundant bacterivores, although this phylum has been so far assumed to be mainly autotrophic. Moreover, the growth of a small lineage of Cryptophyta (CRY1 clade) was strongly stimulated by one Limnohabitans strain in our experiment. Thus, our study is the first report that colorless Cryptophyta are major bacterivores in summer plankton samples and can play a key role in the carbon transfer from prokaryotes to higher trophic levels.
- MeSH
- Bacteria classification genetics isolation & purification metabolism MeSH
- Cryptophyta microbiology MeSH
- Phylogeny MeSH
- Heterotrophic Processes MeSH
- In Situ Hybridization, Fluorescence MeSH
- Plankton microbiology MeSH
- Food Chain MeSH
- Seasons MeSH
- Fresh Water microbiology parasitology MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
The genus Limnohabitans (Comamonadaceae, Betaproteobacteria) is a common and a highly active component of freshwater bacterioplanktonic communities. To date, the genus has been considered to contain only heterotrophic species. In this study, we detected the photosynthesis genes pufLM and bchY in 28 of 46 strains from three Limnohabitans lineages. The pufM sequences obtained are very closely related to environmental pufM sequences detected in various freshwater habitats, indicating the ubiquity and potential importance of photoheterotrophic Limnohabitans in nature. Additionally, we sequenced and analyzed the genomes of 5 potentially photoheterotrophic Limnohabitans strains, to gain further insights into their phototrophic capacity. The structure of the photosynthesis gene cluster turned out to be highly conserved within the genus Limnohabitans and also among all potentially photosynthetic Betaproteobacteria strains. The expression of photosynthetic complexes was detected in a culture of Limnohabitans planktonicus II-D5T using spectroscopic and pigment analyses. This was further verified by a novel combination of infrared microscopy and fluorescent in situ hybridization.IMPORTANCE The data presented document that the capacity to perform anoxygenic photosynthesis is common among the members of the genus Limnohabitans, indicating that they may have a novel role in freshwater habitats.
- Keywords
- FISH, IR microscopy, Limnohabitans, bacteriochlorophyll, bchY, freshwater Betaproteobacteria, photosynthetic bacteria, pufM,
- MeSH
- Aerobiosis MeSH
- Genes, Bacterial physiology MeSH
- Bacterial Proteins genetics metabolism MeSH
- Comamonadaceae genetics metabolism MeSH
- Photosynthesis genetics MeSH
- Phylogeny MeSH
- Multigene Family physiology MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Bacterial Proteins MeSH
Actinobacteria of the acI lineage are the most abundant microbes in freshwater systems, but there are so far no pure living cultures of these organisms, possibly because of metabolic dependencies on other microbes. This, in turn, has hampered an in-depth assessment of the genomic basis for their success in the environment. Here we present genomes from 16 axenic cultures of acI Actinobacteria. The isolates were not only of minute cell size, but also among the most streamlined free-living microbes, with extremely small genome sizes (1.2-1.4 Mbp) and low genomic GC content. Genome reduction in these bacteria might have led to auxotrophy for various vitamins, amino acids and reduced sulphur sources, thus creating dependencies to co-occurring organisms (the 'Black Queen' hypothesis). Genome analyses, moreover, revealed a surprising degree of inter- and intraspecific diversity in metabolic pathways, especially of carbohydrate transport and metabolism, and mainly encoded in genomic islands. The striking genotype microdiversification of acI Actinobacteria might explain their global success in highly dynamic freshwater environments with complex seasonal patterns of allochthonous and autochthonous carbon sources. We propose a new order within Actinobacteria ('Candidatus Nanopelagicales') with two new genera ('Candidatus Nanopelagicus' and 'Candidatus Planktophila') and nine new species.
- MeSH
- Actinobacteria classification genetics isolation & purification MeSH
- Biodiversity MeSH
- DNA, Bacterial chemistry MeSH
- Phylogeny MeSH
- Genome, Bacterial * MeSH
- Metabolic Networks and Pathways genetics MeSH
- Fresh Water microbiology MeSH
- Base Composition MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- DNA, Bacterial MeSH
Because their large growth potential is counterbalanced with grazing by heterotrophic nanoflagellates (HNF), bacteria of the genus Limnohabitans, which are common in many freshwater habitats, represent a valuable model for examining bacterial carbon flow to the grazer food chain. We conducted experiments with natural HNF communities taken from two distinct habitats, the meso-eutrophic Římov Reservoir and the oligo-mesotrophic Lake Cep (South Bohemia). HNF communities from each habitat at distinct seasonal phases, a late April algal bloom and a late May clear water phase, were each fed 3 Limnohabitans strains of differing cell sizes. Water samples were prefiltered (5 μm) to release natural HNF communities from zooplankton control and then amended with the Limnohabitans strains L. planktonicus II-D5 (medium sized, rod shaped), Limnohabitans sp. strain T6-5 (thin, long, curved rod), and Limnohabitans sp. strain 2KL-3 (large solenoid). Using temporal sampling and prey treatment, we determined HNF growth parameters such as doubling time, growth efficiency, and length of lag phase prior starting to exponential growth. All three Limnohabitans strains supported HNF growth but in significant prey-, site-, and season-dependent fashions. For instance, addition of the moderately large T6-5 strain yielded very rapid HNF growth with a short lag phase. In contrast, the curved morphology and larger cell size of strain 2KL-3 made this prey somewhat protected against grazing by smaller HNF, resulting in slower HNF growth and longer lag phases. These trends were particularly pronounced during the late May clear-water phase, which was dominated by smaller HNF cells. This may indicate a longer "adaptation time" for the flagellate communities toward the large prey size offered.
- MeSH
- Comamonadaceae cytology growth & development MeSH
- Heterotrophic Processes MeSH
- Lakes microbiology parasitology MeSH
- Food Chain MeSH
- Seasons MeSH
- Fresh Water microbiology parasitology MeSH
- Zooplankton growth & development MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Geographicals
- Czech Republic MeSH
