Correction to: A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms.

Following publication of the original article [1], we have been notified that three of the primer names identified as most promising candidates for fungal community surveys were incorrectly renamed following the primer nomenclature system proposed by Gargas & DePriest [2].

A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms.

BACKGROUND: Several fungi-specific primers target the 18S rRNA gene sequence, one of the prominent markers for fungal classification. The design of most primers goes back to the last decades. Since then, the number of sequences in public databases increased leading to the discovery of new fungal groups and changes in fungal taxonomy. However, no reevaluation of primers was carried out and relevant information on most primers is missing. With this study, we aimed to develop an 18S rRNA gene sequence primer toolkit allowing an easy selection of the best primer pair appropriate for different sequencing platforms, research aims (biodiversity assessment versus isolate classification) and target groups. RESULTS: We performed an intensive literature research, reshuffled existing primers into new pairs, designed new Illumina-primers, and annealing blocking oligonucleotides. A final number of 439 primer pairs were subjected to in silico PCRs. Best primer pairs were selected and experimentally tested. The most promising primer pair with a small amplicon size, nu-SSU-1333-5'/nu-SSU-1647-3' (FF390/FR-1), was successful in describing fungal communities by Illumina sequencing. Results were confirmed by a simultaneous metagenomics and eukaryote-specific primer approach. Co-amplification occurred in all sample types but was effectively reduced by blocking oligonucleotides. CONCLUSIONS: The compiled data revealed the presence of an enormous diversity of fungal 18S rRNA gene primer pairs in terms of fungal coverage, phylum spectrum and co-amplification. Therefore, the primer pair has to be carefully selected to fulfill the requirements of the individual research projects. The presented primer toolkit offers comprehensive lists of 164 primers, 439 primer combinations, 4 blocking oligonucleotides, and top primer pairs holding all relevant information including primer's characteristics and performance to facilitate primer pair selection.

Mycoplankton Biome Structure and Assemblage Processes Differ Along a Transect From the Elbe River Down to the River Plume and the Adjacent Marine Waters.

Rivers are transport systems and supply adjacent ecosystems with nutrients. They also serve human well-being, for example as a source of food. Microorganism biodiversity is an important parameter for the ecological balance of river ecosystems. Despite the knowledge that fungi are key players in freshwater nutrient cycling and food webs, data on planktonic fungi of streams with higher stream order are scarce. This study aims to fill this knowledge gap by a fungi-specific 18S ribosomal RNA (rRNA) gene tag sequencing approach, investigating mycoplankton diversity in the Elbe River along a transect from shallow freshwater, to the estuary and river plume down to the adjacent marine waters (sections of seventh stream order number). Using multivariate analyses and the quantitative process estimates (QPEs) method, questions (i) of how mycoplankton communities as part of the river continuum change along the transect, (ii) what factors, spatial and environmental, play a role, and (iii) what assembly processes, such as selection or dispersion, operate along the transect, were addressed. The partitioning of mycoplankton communities into three significant distant biomes was mainly driven by local environmental conditions that were partly under spatial control. The assembly processes underlying the biomes also differed significantly. Thus, variable selection dominated the upstream sections, while undominated processes like ecological drift dominated the sections close to the river mouth and beyond. Dispersal played a minor role. The results suggest that the ecological versatility of the mycoplankton communities changes along the transect as response, for example, to a drastic change from an autotrophic to a heterotrophic system caused by an abrupt increase in the river depth. Furthermore, a significant salinity-dependent occurrence of diverse basal fungal groups was observed, with no clade found exclusively in marine waters. These results provide an important framework to help understand patterns of riverine mycoplankton communities and serve as basis for a further in-depth work so that fungi, as an important ecological organism group, can be integrated into models of, e.g., usage-balance considerations of rivers.

Seasonal Dynamics of Pelagic Mycoplanktonic Communities: Interplay of Taxon Abundance, Temporal Occurrence, and Biotic Interactions.

Marine fungi are an important component of pelagic planktonic communities. However, it is not yet clear how individual fungal taxa are integrated in marine processes of the microbial loop and food webs. Most likely, biotic interactions play a major role in shaping the fungal community structure. Thus, the aim of our work was to identify possible biotic interactions of mycoplankton with phytoplankton and zooplankton groups and among fungi, and to investigate whether there is coherence between interactions and the dynamics, abundance and temporal occurrence of individual fungal OTUs. Marine surface water was sampled weekly over the course of 1 year, in the vicinity of the island of Helgoland in the German Bight (North Sea). The mycoplankton community was analyzed using 18S rRNA gene tag-sequencing and the identified dynamics were correlated to environmental data including phytoplankton, zooplankton, and abiotic factors. Finally, co-occurrence patterns of fungal taxa were detected with network analyses based on weighted topological overlaps (wTO). Of all abundant and persistent OTUs, 77% showed no biotic relations suggesting a saprotrophic lifestyle. Of all other fungal OTUs, nearly the half (44%) had at least one significant negative relationship, especially with zooplankton and other fungi, or to a lesser extent with phytoplankton. These findings suggest that mycoplankton OTUs are embedded into marine food web chains via highly complex and manifold relationships such as parasitism, predation, grazing, or allelopathy. Furthermore, about one third of all rare OTUs were part of a dense fungal co-occurrence network probably stabilizing the fungal community against environmental changes and acting as functional guilds or being involved in fungal cross-feeding. Placed in an ecological context, strong antagonistic relationships of the mycoplankton community with other components of the plankton suggest that: (i) there is a top-down control by fungi on zooplankton and phytoplankton; (ii) fungi serve as a food source for zooplankton and thereby transfer nutrients and organic material; (iii) the dynamics of fungi harmful to other plankton groups are controlled by antagonistic fungal taxa.