Fungi are phylogenetically and functionally diverse ubiquitous components of almost all ecosystems on Earth, including aquatic environments stretching from high montane lakes down to the deep ocean. Aquatic ecosystems, however, remain frequently overlooked as fungal habitats, although fungi potentially hold important roles for organic matter cycling and food web dynamics. Within a broad ecological framework, we conceptualize the spatio-temporal dimensions, diversity, functions and organismic interactions of fungi in structuring aquatic foodwebs. We focus on currently unexplored fungal diversity, highlighting poorly understood ecosystems, including emerging artificial aquatic habitats. Recent methodological improvements have facilitated a greater appreciation of the importance of fungi in many aquatic systems, yet a conceptual framework is still missing. To date, aquatic fungi and their interactions have largely remained "hidden" and require interdisciplinary efforts to be explored in an ecosystem context. There remain obvious methodological and knowledge gaps to explore potential functions of aquatic fungi, moving from the microscale to the global scale. This knowledge is urgently needed since we humans strongly interfere with structure and function of natural ecosystems by permanently reshaping most of the Earth's surface and creating vast areas of novel urban habitats. Introduction: Recent advances in DNA sequencing technology have revealed that fungi are abundant in many, if not all aquatic ecosystems, however their diversity, quantitative abundance, ecological function and, in particular, their interactions with other microorganisms, remain largely speculative, unexplored and missing from current general concepts in aquatic ecology and biogeochemistry 1-4. This is surprising since terrestrial-focused research has understood the outstanding ecological role of fungi for >100 years, and therefore fungi constitute a major component of general concepts in terrestrial science 5,6. In aquatic ecosystems, the systematic analysis of fungal diversity and their ecological roles has faced several setbacks due to methodological limitations and a too small scientific community, in particular in the marine environment 7-11. This review focusses on aquatic fungi, which form a morphologically, phylogenetically, and ecologically diverse group 7. We here broadly define "aquatic fungi" as fungi that rely for the whole or part of their life cycle on aquatic habitats (FIG. 1). Three groups (indwellers, periodic immigrants and versatile immigrants) based on their degree of adaptation and dependence on aquatic habitats have been previously defined 12. We highlight the numerous knowledge gaps in their diversity, interactions and functional roles, as well as methodological limitations. In this review we propose new research avenues to set aquatic fungi in a broad ecological framework. Here, we do not explore the many existing gaps in the fungal phylogenetic tree. In aquatic systems, fungi constitute a significant proportion of eukaryo...

SummaryChytridiomycota, 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 coevolutionary 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.

1. We describe the dynamics of host-parasite interactions over a period of more than 30 years between the freshwater diatom Asterionella formosa and two highly virulent chytrid parasites (Rhizophydium planktonicum and Zygorhizidium planktonicum) in Lake Maarsseveen, The Netherlands. This period is characterised by a significant warming trend which is strongest in spring. 2. The key spring event in lakes, the diatom bloom, was in many years dominated by Asterionella. We examine whether and how climate warming has affected the prevalence of infection in Asterionella by chytrids. 3. In years with cold winters/early springs, a dense Asterionella bloom is followed by epidemic development of disease as high Asterionella densities greatly facilitate transmission of chytrid zoospores. This sequence of events is absent in milder winters. 4. Earlier experimental studies have shown that the parasite is almost non-infective at water temperatures below 3 degrees C, offering a disease-free window of opportunity for growth of Asterionella. Climate warming has reduced periods in which water temperature remains < 3 degrees C, narrowing the window of opportunity for uninfected growth. Consequently, Asterionella continuously suffers from infection, albeit at low levels. 5. Population reduction as a result of low level infection allows other diatoms to take over as dominant species, possibly through priority effects. 6. In mild winters, chytrid infections no longer reach epidemic levels, but remain at low prevalence since transmission is impaired at low host densities. Climate warming thus affects both host and parasite in intricate ways, with the host denied a bloom and consequently the parasite denied an epidemic. 7. A shift from Asterionella to a mixed diatom community in years with mild winters may benefit the food web, because of the poor edibility of Asterionella, unless the numerous chytrid zoospores produced during epidemics significantly contribute to zooplankton nutrition. 8. Our study demonstrates the potential complexity of climate change impacts on disease. A reduction in the likelihood of epidemic development of a virulent parasite would seem to be of great benefit to the host, but this was not the case. Unexpected, sometimes paradoxical consequences of climate change can be expected and suggest that the view of a 'warmer hence sicker world' may not always apply

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. K E Y W O R D S discussions on the implementation of long-read sequencing. The authors would like to acknowledge support from Science for Life Laboratory, the National Genomics Infrastructure, NGI and Uppmax for providing assistance in massive parallel sequencing and computational infrastructure. CW and RHN gratefully acknowledge financial support from Stiftelsen Olle Engkvist Byggmästare, Stiftelsen Lars Hiertas Minne, Kapten Carl Stenholms Donationsfond and Birgit och Birger Wålhströms Minnesfond.

The spatiotemporal changes in abundance and biomass of heterotrophic bacteria, of three major bacterial phylogenetic groups, and of picocyanobacteria in the upper 20 m of a deep prealpine lake (Lake Zurich, Switzerland) were monitored during a seasonally persistent bloom of the toxigenic filamentous cyanobacterium Planktothrix rubescens. In addition, bacterial 16S ribosomal deoxyribonucleic acid (rDNA) sequences were collected at one instance from the bloom layer and from waters above and below this zone. P. rubescens comprised up to 70% of particulate organic carbon during summer stratification and autumnal mixis and thus by far exceeded the total biomass both of other phytoplankton and of prokaryotes. A strong negative correlation was found between the estimated basin-wide biomass of P. rubescens and of heterotrophic bacteria, and there was different spatial niche preference of filamentous vs. picocyanobacteria. Only members of the CytophagaFlavobacterium lineage of Bacteroidetes showed an increasing tendency of association with the P. rubescens population, in particular at the onset of autumnal mixing. Although the filamentous cyanobacterium was the dominant primary producer throughout the year, it did not seem to be a carbon source for heterotrophic bacteria at all. We conclude that P. rubescens represents a powerful competitor of autotrophic and heterotrophic prokaryotes, likely due to both its specific physiological (photoheterotrophic) properties and its protection against zooplankton grazing. This competitiveness might be regarded as another reason for its mass occurrence in numerous lakes of the Northern hemisphere.