Most cited article - PubMed ID 31566612
Fluorescently Labeled Bacteria as a Tracer to Reveal Novel Pathways of Organic Carbon Flow in Aquatic Ecosystems
Deep, cold, and dark hypolimnia represent the largest volume of water in freshwater lakes with limited occurrences of phototrophs. However, the presence of prokaryotes supports populations of bacterivorous ciliates and heterotrophic nanoflagellates (HNF). Nevertheless, protistan bacterivory rates and the major hypolimnetic ciliate bacterivores are poorly documented. We conducted a high frequency sampling (three-times a week) in the oxic hypolimnion of a stratified mesoeutrophic reservoir during summer, characterized by stable physicochemical conditions and low water temperature. Using fluorescently labeled bacteria we estimated that ciliates and HNF contributed, on average, 30% and 70% to aggregated protistan bacterivory, respectively, and collectively removed about two thirds of daily hypolimnetic prokaryotic production. The ciliate community was analyzed by the quantitative protargol staining method. One scuticociliate morphotype dominated the hypolimnetic ciliate community, accounting for 82% of total ciliates and over 98% of total ciliate bacterivory, with average cell-specific uptake rate of 202 prokaryotes per hour. Moreover, long-amplicon sequencing revealed that the scuticociliate belongs to an unidentified clade closely related to the Ctedoctematidae and Eurystomatellidae families. The high-resolution sampling, microscopic, and sequencing methods allowed uncovering indigenous microbial food webs in the hypolimnetic environment and revealed a functional simplification of ciliate communities, dominated by a new bacterivorous scuticociliate lineage.
- Keywords
- bacterivorous protists, cold hypolimnetic layer, freshwater reservoir, new lineage of scuticociliates, protistan bacterivory rates,
- MeSH
- Bacteria * classification isolation & purification genetics MeSH
- Ciliophora * classification genetics MeSH
- Phylogeny MeSH
- Lakes microbiology MeSH
- Oligohymenophorea * classification genetics isolation & purification MeSH
- Sequence Analysis, DNA MeSH
- Fresh Water * microbiology parasitology MeSH
- Publication type
- Journal Article MeSH
PREMISE: Despite the high functional importance of endophytes, we still have limited understanding of the biotic and abiotic factors that influence colonization of plant hosts along major ecological gradients and lack quantitative estimates of their colonization extent. In this study, we hypothesized that the developmental stage of the ecosystem will affect the levels of bacterial and fungal endophytic assemblages in the foliar endosphere. METHODS: We quantified levels of bacterial and fungal endophytes in leaves of four plant hosts at four stages of vegetation succession using an optimized qPCR protocol with bacteria-specific 16S and fungi-targeting primers. RESULTS: (1) The ecosystem developmental stage did not have a significant effect on the colonization levels of bacterial or fungal endophytes. (2) Colonization levels by bacterial and fungal endophytes were governed by different mechanisms. (3) Endophytic colonization levels and their relationship to foliar tissue stoichiometry were highly host specific. CONCLUSIONS: Quantifying colonization levels is important in the study of endophytic ecology, and the fast, relatively low-cost qPCR-based method can supply useful ecological information, which can significantly enhance the interpretation potential of descriptive data generated, for example, by next-generation sequencing.
- Keywords
- cell counts, ecological succession, foliar endophyte, fungi‐bacteria ratios, qPCR, soil chronosequence,
- MeSH
- Bacteria * genetics growth & development isolation & purification MeSH
- Ecosystem MeSH
- Endophytes * physiology genetics MeSH
- Host Specificity * MeSH
- Fungi * genetics isolation & purification physiology MeSH
- Real-Time Polymerase Chain Reaction methods MeSH
- Plant Leaves * microbiology 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.
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.
