Parasitic chytrids could promote copepod survival by mediating material transfer from inedible diatoms

Diatoms form large spring blooms in lakes and oceans, providing fuel for higher trophic levels at the start of the growing season. Some of the diatom blooms, however, are not grazed by filter-feeding zooplankton like Daphnia due to their large size. Several of these large diatoms are susceptible to chytrid infections. Zoospores of chytrids appeared to be excellent food for Daphnia, both in terms of size, shape, and quality (PUFAs and cholesterol). Thus, zoospores of chytrids can bridge the gap between inedible diatoms and Daphnia. In order to examine the effects of diatoms and chytrids on the survival of copepods, we performed one grazing and one survival experiment. The grazing experiment revealed that the diatom, Asterionella formosa, was not grazed by the copepod, Eudiaptomus gracilis, even after being infected by the chytrid Zygorhizidium planktonicum. However, carbon and nitrogen concentrations were significantly reduced by E. gracilis only when A. formosa was infected by Z. planktonicum, indicating that the chytrids might facilitate material transfer from inedible diatoms to the copepods. The survival experiment revealed that E. gracilis lived shorter with A. formosa than with the cryptophyta Cryptomonas pyrenoidifera. However, the survival of E. gracilis increased significantly in the treatment where A. formosa cells were infected by Z. planktonicum. Since E. gracilis could not graze A. formosa cells due to their large colonial forms, E. gracilis may acquire nutrients by grazing on the zoospores, and were so able to survive in the presence of the A. formosa. This provides new insights into the role of parasitic fungi in aquatic food webs, where chytrids may improve copepod survival during diatom blooms.

Novel parasitic chytrids infecting snow algae in an alpine snow ecosystem in Japan

IntroductionMicrobial communities are important components of glacier and snowpack ecosystems that influence biogeochemical cycles and snow/ice melt. Recent environmental DNA surveys have revealed that chytrids dominate the fungal communities in polar and alpine snowpacks. These could be parasitic chytrids that infect snow algae as observed microscopically. However, the diversity and phylogenetic position of parasitic chytrids has not been identified due to difficulties in establishing their culture and subsequent DNA sequencing. In this study, we aimed to identify the phylogenetic positions of chytrids infecting the snow algae, Chloromonas spp., bloomed on snowpacks in Japan.MethodsBy linking a microscopically picked single fungal sporangium on a snow algal cell to a subsequent sequence of ribosomal marker genes, we identified three novel lineages with distinct morphologies.ResultsAll the three lineages belonged to Mesochytriales, located within “Snow Clade 1”, a novel clade consisting of uncultured chytrids from snow-covered environments worldwide. Additionally, putative resting spores of chytrids attached to snow algal cells were observed.DiscussionThis suggests that chytrids may survive as resting stage in soil after snowmelt. Our study highlights the potential importance of parasitic chytrids that infect snow algal communities

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

Integrating chytrid fungal parasites into plankton ecology: research gaps and needs

Chytridiomycota, often referred to as chytrids, can be virulent parasites with the potential to inflict mass mortalities on hosts, causing e.g. changes in phytoplankton size distributions and succession, and the delay or suppression of bloom events. Molecular environmental surveys have revealed an unexpectedly large diversity of chytrids across a wide range of aquatic ecosystems worldwide. As a result, scientific interest towards fungal parasites of phytoplankton has been gaining momentum in the past few years. Yet, we still know little about the ecology of chytrids, their life cycles, phylogeny, host specificity and range. Information on the contribution of chytrids to trophic interactions, as well as co‐evolutionary feedbacks of fungal parasitism on host populations is also limited. This paper synthesizes ideas stressing the multifaceted biological relevance of phytoplankton chytridiomycosis, resulting from discussions among an international team of chytrid researchers. It presents our view on the most pressing research needs for promoting the integration of chytrid fungi into aquatic ecology

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

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

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

Causal networks of phytoplankton diversity and biomass are modulated by environmental context

Untangling causal links and feedbacks among biodiversity, ecosystem functioning, and environmental factors is challenging due to their complex and context-dependent interactions (e.g., a nutrient-dependent relationship between diversity and biomass). Consequently, studies that only consider separable, unidirectional effects can produce divergent conclusions and equivocal ecological implications. To address this complexity, we use empirical dynamic modeling to assemble causal networks for 19 natural aquatic ecosystems (N24◦~N58◦) and quantified strengths of feedbacks among phytoplankton diversity, phytoplankton biomass, and environmental factors. Through a cross-system comparison, we identify macroecological patterns; in more diverse, oligotrophic ecosystems, biodiversity effects are more important than environmental effects (nutrients and temperature) as drivers of biomass. Furthermore, feedback strengths vary with productivity. In warm, productive systems, strong nitrate-mediated feedbacks usually prevail, whereas there are strong, phosphate-mediated feedbacks in cold, less productive systems. Our findings, based on recovered feedbacks, highlight the importance of a network view in future ecosystem management

Use of the index of pulmonary vascular disease for predicting long-term outcome of pulmonary arterial hypertension associated with congenital heart disease

AimsLimited data exist on risk factors for the long-term outcome of pulmonary arterial hypertension (PAH) associated with congenital heart disease (CHD-PAH). We focused on the index of pulmonary vascular disease (IPVD), an assessment system for pulmonary artery pathology specimens. The IPVD classifies pulmonary vascular lesions into four categories based on severity: (1) no intimal thickening, (2) cellular thickening of the intima, (3) fibrous thickening of the intima, and (4) destruction of the tunica media, with the overall grade expressed as an additive mean of these scores. This study aimed to investigate the relationship between IPVD and the long-term outcome of CHD-PAH.MethodsThis retrospective study examined lung pathology images of 764 patients with CHD-PAH aged &lt;20 years whose lung specimens were submitted to the Japanese Research Institute of Pulmonary Vasculature for pulmonary pathological review between 2001 and 2020. Clinical information was collected retrospectively by each attending physician. The primary endpoint was cardiovascular death.ResultsThe 5-year, 10-year, 15-year, and 20-year cardiovascular death-free survival rates for all patients were 92.0%, 90.4%, 87.3%, and 86.1%, respectively. The group with an IPVD of ≥2.0 had significantly poorer survival than the group with an IPVD &lt;2.0 (P = .037). The Cox proportional hazards model adjusted for the presence of congenital anomaly syndromes associated with pulmonary hypertension, and age at lung biopsy showed similar results (hazard ratio 4.46; 95% confidence interval: 1.45–13.73; P = .009).ConclusionsThe IPVD scoring system is useful for predicting the long-term outcome of CHD-PAH. For patients with an IPVD of ≥2.0, treatment strategies, including choosing palliative procedures such as pulmonary artery banding to restrict pulmonary blood flow and postponement of intracardiac repair, should be more carefully considered