Parasite Fitness Traits Under Environmental Variation: Disentangling the Roles of a Chytrid's Immediate Host and External Environment

Parasite environments are heterogeneous at different levels. The first level of variability is the host itself. The second level represents the external environment for the hosts, to which parasites may be exposed during part of their life cycle. Both levels are expected to affect parasite fitness traits. We disentangle the main and interaction effects of variation in the immediate host environment, here the diatom Asterionella formosa (variables host cell volume and host condition through herbicide pre-exposure) and variation in the external environment (variables host density and acute herbicide exposure) on three fitness traits (infection success, development time and reproductive output) of a chytrid parasite. Herbicide exposure only decreased infection success in a low host density environment. This result reinforces the hypothesis that chytrid zoospores use photosynthesis-dependent chemical cues to locate its host. At high host densities, chemotaxis becomes less relevant due to increasing chance contact rates between host and parasite, thereby following the mass-action principle in epidemiology. Theoretical support for this finding is provided by an agent-based simulation model. The immediate host environment (cell volume) substantially affected parasite reproductive output and also interacted with the external herbicide exposed environment. On the contrary, changes in the immediate host environment through herbicide pre-exposure did not increase infection success, though it had subtle effects on zoospore development time and reproductive output. This study shows that both immediate host and external environment as well as their interaction have significant effects on parasite fitness. Disentangling these effects improves our understanding of the processes underlying parasite spread and disease dynamics

Intercomparison of Two Fluorescent Dyes to Visualize Parasitic Fungi (Chytridiomycota) on Phytoplankton

Fungal microparasites (here chytrids) are widely distributed and yet, they are often overlooked in aquatic environments. To facilitate the detection of microparasites, we revisited the applicability of two fungal cell wall markers, Calcofluor White (CFW) and wheat germ agglutinin (WGA), for the direct visualization of chytrid infections on phytoplankton in laboratory-maintained isolates and field-sampled communities. Using a comprehensive set of chytrid-phytoplankton model pathosystems, we verified the staining pattern on diverse morphological structures of chytrids via fluorescence microscopy. Empty sporangia were stained most effectively, followed by encysted zoospores and im-/mature sporangia, while the staining success was more variable for rhizoids, stalks, and resting spores. In a few instances, the staining was unsuccessful (mostly with WGA), presumably due to insufficient cell fixation, gelatinous cell coatings, and multilayered cell walls. CFW and WGA staining could be done in Utermohl chambers or on polycarbonate filters, but CFW staining on filters seemed less advisable due to high background fluorescence. To visualize chytrids, 1 mu g dye mL(-1) was sufficient (but 5 mu g mL(-1) are recommended). Using a dual CFW-WGA staining protocol, we detected multiple, mostly undescribed chytrids in two natural systems (freshwater and coastal), while falsely positive or negative stained cells were well detectable. As a proof-of-concept, we moreover conducted imaging flow cytometry, as a potential high-throughput technology for quantifying chytrid infections. Our guidelines and recommendations are expected to facilitate the detection of chytrid epidemics and to unveil their ecological and economical imprint in natural and engineered aquatic systems.</p

Introducing ribosomal tandem repeat barcoding for fungi

Sequence comparison and analysis of the various ribosomal genetic markers are the dominant molecular methods for identification and description of fungi. However, new environmental fungal lineages known only from DNA data reveal significant gaps in our sampling of the fungal kingdom in terms of both taxonomy and marker coverage in the reference sequence databases. To facilitate the integration of reference data from all of the ribosomal markers, we present three sets of general primers that allow for amplification of the complete ribosomal operon from the ribosomal tandem repeats. The primers cover all ribosomal markers: ETS, SSU, ITS1, 5.8S, ITS2, LSU and IGS. We coupled these primers successfully with third-generation sequencing (PacBio and Nanopore sequencing) to showcase our approach on authentic fungal herbarium specimens (Basidiomycota), aquatic chytrids (Chytridiomycota) and a poorly understood lineage of early diverging fungi (Nephridiophagidae). In particular, we were able to generate high-quality reference data with Nanopore sequencing in a high-throughput manner, showing that the generation of reference data can be achieved on a regular desktop computer without the involvement of any large-scale sequencing facility. The quality of the Nanopore generated sequences was 99.85%, which is comparable with the 99.78% accuracy described for Sanger sequencing. With this work, we hope to stimulate the generation of a new comprehensive standard of ribosomal reference data with the ultimate aim to close the huge gaps in our reference datasets

Towards Phytoplankton Parasite Detection Using Autoencoders

Phytoplankton parasites are largely understudied microbial components with a potentially significant ecological impact on phytoplankton bloom dynamics. To better understand their impact, we need improved detection methods to integrate phytoplankton parasite interactions in monitoring aquatic ecosystems. Automated imaging devices usually produce high amount of phytoplankton image data, while the occurrence of anomalous phytoplankton data is rare. Thus, we propose an unsupervised anomaly detection system based on the similarity of the original and autoencoder-reconstructed samples. With this approach, we were able to reach an overall F1 score of 0.75 in nine phytoplankton species, which could be further improved by species-specific fine-tuning. The proposed unsupervised approach was further compared with the supervised Faster R-CNN based object detector. With this supervised approach and the model trained on plankton species and anomalies, we were able to reach the highest F1 score of 0.86. However, the unsupervised approach is expected to be more universal as it can detect also unknown anomalies and it does not require any annotated anomalous data that may not be always available in sufficient quantities. Although other studies have dealt with plankton anomaly detection in terms of non-plankton particles, or air bubble detection, our paper is according to our best knowledge the first one which focuses on automated anomaly detection considering putative phytoplankton parasites or infections

Seasonality of parasitic and saprotrophic zoosporic fungi: linking sequence data to ecological traits

Zoosporic fungi of the phylum Chytridiomycota (chytrids) regularly dominate pelagic fungal communities in freshwater and marine environments. Their lifestyles range from obligate parasites to saprophytes. Yet, linking the scarce available sequence data to specific ecological traits or their host ranges constitutes currently a major challenge. We combined 28 S rRNA gene amplicon sequencing with targeted isolation and sequencing approaches, along with cross-infection assays and analysis of chytrid infection prevalence to obtain new insights into chytrid diversity, ecology, and seasonal dynamics in a temperate lake. Parasitic phytoplankton-chytrid and saprotrophic pollen-chytrid interactions made up the majority of zoosporic fungal reads. We explicitly demonstrate the recurrent dominance of parasitic chytrids during frequent diatom blooms and saprotrophic chytrids during pollen rains. Distinct temporal dynamics of diatom-specific parasitic clades suggest mechanisms of coexistence based on niche differentiation and competitive strategies. The molecular and ecological information on chytrids generated in this study will aid further exploration of their spatial and temporal distribution patterns worldwide. To fully exploit the power of environmental sequencing for studies on chytrid ecology and evolution, we emphasize the need to intensify current isolation efforts of chytrids and integrate taxonomic and autecological data into long-term studies and experiments.Alliance of German Science Organizations///AlemaniaUCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Biologí

Antarctic Glacial Meltwater Impacts the Diversity of Fungal Parasites Associated With Benthic Diatoms in Shallow Coastal Zones

Aquatic ecosystems are frequently overlooked as fungal habitats, although there is increasing evidence that their diversity and ecological importance are greater than previously considered. Aquatic fungi are critical and abundant components of nutrient cycling and food web dynamics, e.g., exerting top-down control on phytoplankton communities and forming symbioses with many marine microorganisms. However, their relevance for microphytobenthic communities is almost unexplored. In the light of global warming, polar regions face extreme changes in abiotic factors with a severe impact on biodiversity and ecosystem functioning. Therefore, this study aimed to describe, for the first time, fungal diversity in Antarctic benthic habitats along the salinity gradient and to determine the co-occurrence of fungal parasites with their algal hosts, which were dominated by benthic diatoms. Our results reveal that Ascomycota and Chytridiomycota are the most abundant fungal taxa in these habitats. We show that also in Antarctic waters, salinity has a major impact on shaping not just fungal but rather the whole eukaryotic community composition, with a diversity of aquatic fungi increasing as salinity decreases. Moreover, we determined correlations between putative fungal parasites and potential benthic diatom hosts, highlighting the need for further systematic analysis of fungal diversity along with studies on taxonomy and ecological roles of Chytridiomycota

Towards Phytoplankton Parasite Detection Using Autoencoders

Phytoplankton parasites are largely understudied microbial components with a potentially significant ecological influence on phytoplankton bloom dynamics. To better understand the impact of phytoplankton parasites, improved detection methods are needed to integrate phytoplankton parasite interactions into monitoring of aquatic ecosystems. Automated imaging devices commonly produce vast amounts of phytoplankton image data, but the occurrence of anomalous phytoplankton data in such datasets is rare. Thus, we propose an unsupervised anomaly detection system based on the similarity between the original and autoencoder-reconstructed samples. With this approach, we were able to reach an overall F1 score of 0.75 in nine phytoplankton species, which could be further improved by species-specific fine-tuning. The proposed unsupervised approach was further compared with the supervised Faster R-CNN-based object detector. Using this supervised approach and the model trained on plankton species and anomalies, we were able to reach a highest F1 score of 0.86. However, the unsupervised approach is expected to be more universal as it can also detect unknown anomalies and it does not require any annotated anomalous data that may not always be available in sufficient quantities. Although other studies have dealt with plankton anomaly detection in terms of non-plankton particles or air bubble detection, our paper is, according to our best knowledge, the first that focuses on automated anomaly detection considering putative phytoplankton parasites or infections

Seasonality of parasitic and saprotrophic zoosporic fungi: linking sequence data to ecological traits

Zoosporic fungi of the phylum Chytridiomycota (chytrids) regularly dominate pelagic fungal communities in freshwater and marine environments. Their lifestyles range from obligate parasites to saprophytes. Yet, linking the scarce available sequence data to specific ecological traits or their host ranges constitutes currently a major challenge. We combined 28 S rRNA gene amplicon sequencing with targeted isolation and sequencing approaches, along with cross-infection assays and analysis of chytrid infection prevalence to obtain new insights into chytrid diversity, ecology, and seasonal dynamics in a temperate lake. Parasitic phytoplankton-chytrid and saprotrophic pollen-chytrid interactions made up the majority of zoosporic fungal reads. We explicitly demonstrate the recurrent dominance of parasitic chytrids during frequent diatom blooms and saprotrophic chytrids during pollen rains. Distinct temporal dynamics of diatom-specific parasitic clades suggest mechanisms of coexistence based on niche differentiation and competitive strategies. The molecular and ecological information on chytrids generated in this study will aid further exploration of their spatial and temporal distribution patterns worldwide. To fully exploit the power of environmental sequencing for studies on chytrid ecology and evolution, we emphasize the need to intensify current isolation efforts of chytrids and integrate taxonomic and autecological data into long-term studies and experiments.</p

Taxonomic annotation of public fungal ITS sequences from the built environment - A report from an April 10-11, 2017 workshop (Aberdeen, UK)

The UNITE database community gratefully acknowledges support from the Alfred P. Sloan Foundation. HN and CW gratefully acknowledges financial support from Stiftelsen Olle Engkvist Byggmästare, Stiftelsen Lars Hiertas Minne, Kapten Carl Stenholms Donationsfond, and Birgit och Birger Wålhströms Minnesfond. CW gratefully acknowledges a Marie Skłodowska-Curie post doctoral grant from the ERC. Leho Tedersoo is gratefully acknowledged for providing helpful feedback on an earlier draft of this manuscript.Peer reviewedPublisher PD