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What Is This Thing Called Science

The development of high‐throughput sequencing (HTS) technologies has greatly improved our capacity to identify fungi and unveil their ecological roles across a variety of ecosystems. Here we provide an overview of current best practices in metabarcoding analysis of fungal communities, from experimental design through molecular and computational analyses. By reanalysing published data sets, we demonstrate that operational taxonomic units (OTUs) outperform amplified sequence variants (ASVs) in recovering fungal diversity, a finding that is particularly evident for long markers. Additionally, analysis of the full‐length ITS region allows more accurate taxonomic placement of fungi and other eukaryotes compared to the ITS2 subregion. Finally, we show that specific methods for compositional data analyses provide more reliable estimates of shifts in community structure. We conclude that metabarcoding analyses of fungi are especially promising for integrating fungi into the full microbiome and broader ecosystem functioning context, recovery of novel fungal lineages and ancient organisms as well as barcoding of old specimens including type material.

Best practices in metabarcoding of fungi: From experimental design to results

The numbers of fungi: are the most speciose genera truly diverse?

Alternaria alternata is a common species of fungus frequently isolated from plants as both an endophyte and a pathogen. Although the current definition of A. alternata rests on a foundation of morphological, genetic and genomic analyses, doubts persist regarding the scope of A. alternata within the genus due to the varied symbiotic interactions and wide host range observed in these fungi. These doubts may be due in large part to the history of unstable taxonomy in Alternaria, based on limited morphological characters for species delimitation and host specificity associated with toxins encoded by genes carried on conditionally dispensable chromosomes. This review explores the history of Alternaria taxonomy, focusing in particular on the use of nutritional mode and host associations in species delimitation, with the goal of evaluating A. alternata as it currently stands based on taxonomic best practice. Given the recombination detected among isolates of A. alternata, different symbiotic associations in this species should not be considered phylogenetically informative.

Alternaria alternata as endophyte and pathogen

The fungal gut microbiota (mycobiota) has been implicated in diseases that disturb gut homeostasis, such as inflammatory bowel disease. However, little is known about functional relationships between bacteria and fungi in the gut during infectious colitis. Here we investigated the role of fungal metabolites during infection with the intestinal pathogen Salmonella enterica serovar Typhimurium, a major cause of gastroenteritis worldwide. We found that, in the gut lumen, both the mycobiota and fungi present in the diet can be a source of siderophores, small molecules that scavenge iron from the host. The ability to use fungal siderophores, such as ferrichrome and coprogen, conferred a competitive growth advantage to Salmonella strains expressing the fungal siderophore receptors FhuA or FhuE in vitro and in a mouse model. Our study highlights the role of inter-kingdom cross-feeding between fungi and Salmonella and elucidates an additional function of the gut mycobiota, revealing the importance of these understudied members of the gut ecosystem during bacterial infection. Siderophores from diet-derived or intestinal fungi can be used by Salmonella for growth in the murine gut. 

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Mycobiota and diet-derived fungal xenosiderophores promote Salmonella gastrointestinal colonization

Here, we describe the taxon hypothesis (TH) paradigm, which covers the construction, identification, and communication of taxa as datasets. Defining taxa as datasets of individuals and their traits will make taxon identification and most importantly communication of taxa precise and reproducible. This will allow datasets with standardized and atomized traits to be used digitally in identification pipelines and communicated through persistent identifiers. Such datasets are particularly useful in the context of formally undescribed or even physically undiscovered species if data such as sequences from samples of environmental DNA (eDNA) are available. Implementing the TH paradigm will to some extent remove the impediment to hastily discover and formally describe all extant species in that the TH paradigm allows discovery and communication of new species and other taxa also in the absence of formal descriptions. The TH datasets can be connected to a taxonomic backbone providing access to the vast information associated with the tree of life. In parallel to the description of the TH paradigm, we demonstrate how it is implemented in the UNITE digital taxon communication system. UNITE TH datasets include rich data on individuals and their rDNA ITS sequences. These datasets are equipped with digital object identifiers (DOI) that serve to fix their identity in our communication. All datasets are also connected to a GBIF taxonomic backbone. Researchers processing their eDNA samples using UNITE datasets will, thus, be able to publish their findings as taxon occurrences in the GBIF data portal. UNITE species hypothesis (species level THs) datasets are increasingly utilized in taxon identification pipelines and even formally undescribed species can be identified and communicated by using UNITE. The TH paradigm seeks to achieve unambiguous, unique, and traceable communication of taxa and their properties at any level of the tree of life. It offers a rapid way to discover and communicate undescribed species in identification pipelines and data portals before they are lost to the sixth mass extinction.

/pdf/the-taxon-hypothesis-paradigm-on-the-unambiguous-detection-3bvklq6u6m.pdf

The Taxon Hypothesis Paradigm-On the Unambiguous Detection and Communication of Taxa.

PlutoF online platform (https://plutof.ut.ee) is built for the management of biodiversity data. The concept is to provide a common workbench where the full data lifecycle can be managed and support seamless data sharing between single users, workgroups and institutions. Today, large and sophisticated biodiversity datasets are increasingly developed and managed by international workgroups. PlutoF's ambition is to serve such collaborative projects as well as to provide data management services to single users, museum or private collections and research institutions.
 Data management in PlutoF follows a logical order of the data lifecycle Fig. 1. At first, project metadata is uploaded including the project description, data management plan, participants, sampling areas, etc. Data upload and management activities then follow which is often linked to the internal data sharing. Some data analyses can be performed directly in the workbench or data can be exported in standard formats. PlutoF includes also data publishing module. Users can publish their data, generating a citable DOI without datasets leaving PlutoF workbench. PlutoF is part of the DataCite collaboration (https://datacite.org) and so far released more than 600 000 DOIs. Another option is to publish observation or collection datasets via the GBIF (Global Biodiversity Information Facility) portal. A. new feature implemented in 2019 allows users to publish High Throughput Sequencing data as taxon occurrences in GBIF. There is an additional option to send specific datasets directly to the Pensoft online journals. Ultimately, PlutoF works as a data archive which completes the data life cycle.
 In PlutoF users can manage different data types. Most common types include specimen and living specimen data, nucleotide sequences, human observations, material samples, taxonomic backbones and ecological data. Another important feature is that these data types can be managed as a single datasets or projects.
 PlutoF follows several biodiversity standards. Examples include Darwin Core, GGBN (Global Genome Biodiversity Network), EML (Ecological Metadata Language), MCL (Microbiological Common Language), and MIxS (Minimum Information about any (x) Sequence).

https://biss.pensoft.net/article/37398/download/pdf/

PlutoF: Biodiversity data management platform for the complete data lifecycle

https://biss.pensoft.net/article/74249/download/pdf/

Third-party Annotations: Linking PlutoF platform and the ELIXIR Contextual Data ClearingHouse for the reporting of source material annotation gaps and inaccuracies

The advancements in sequencing technologies have greatly contributed to the documentation of Earth’s biodiversity. However, for exploring the full potential of molecular resources for biodiversity, there needs to be a good linkage between sequence data and its biological source, contributing to a network of connected data in the biodiversity research cycle. This requires a foundation of well-structured and accessible annotations in the molecular sequence repositories.
 The International Nucleotide Sequence Database Collaboration (INSDC), of which the European Nucleotide Archive (ENA) is its European node, holds a large amount of annotations associated with sequence data, relating to its biological source (e.g., specimens in natural history collections). However, for a number of records, these annotations may be incomplete (e.g., missing voucher information), ambiguous or even inaccurate.
 Therefore, we have implemented a workflow that allows third-party annotations to be attached to sequence and sample records using two existing services, the PlutoF platform and the ELIXIR Contextual Data ClearingHouse. This work was developed within the scope of the BiCIKL (Biodiversity Community Integrated Knowledge Library) project, which aims to establish open science practices in the biodiversity domain.
 PlutoF is an online data management platform that also provides computing services for biology-related research. PlutoF features allow registered users to enter their own data and access public data at INSDC. Users can enter and manage a range of data, as taxonomic classifications, occurrences, etc. This platform also includes a module that allows the addition of third-party annotations (on material source, taxonomic identification, etc.) linked to specimens or sequence records. This module was already in use by the UNITE community for annotation of INSDC rDNA Internal Transcribed Spacer sequence datasets (Abarenkov et al. 2021). These UNITE annotations are displayed in the National Centre for Biotechnology Information (NCBI) records through links to the PlutoF platform. However, there was the need for an automated solution that allowed third-party annotations to any sequence or sample record at INSDC. This was implemented through the operation of the ELIXIR Contextual Data ClearingHouse (hereafter as Clearinghouse). The Clearinghouse holds a simple RESTful Application Programming Interface (API) to support the submission of additions and improvements to current metadata attributes, such as information on material sources, on records publicly available in the ELIXIR data resources. The Clearinghouse enables the submission of these corrected metadata from databases (such as the PlutoF platform) to the primary data repositories.
 The workflow developed is shown in Fig. 1 and consists of the following steps: i) users annotate sequence metadata that is regularly downloaded from INSDC using NCBI’s E-utilities; ii) an annotation proposal is created and a verification notification is sent to an assigned reviewer; iii) the reviewer evaluates the annotation proposal and accepts it or rejects it with comments; iv) if the annotation proposal is accepted, the annotated fields that may be mapped to ENA fields are then pushed to the Clearinghouse using their RESTful API. The annotations when received at ENA are then reviewed before being displayed. This workflow is implemented through a web interface in PlutoF, which allows user-friendly and effortless reporting of corrections or additions to biological source metadata in sequence records.
 Overall, we expect this tool to contribute to the enrichment of metadata associated with sequence records, and therefore increase the links between the molecular and biodiversity resources, and enable sequencing data to deliver their full potential for biodiversity conservation.

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Timo Piirmann

Papers

The Taxon Hypothesis Paradigm-On the Unambiguous Detection and Communication of Taxa.

PlutoF: Biodiversity data management platform for the complete data lifecycle

Third-party Annotations: Linking PlutoF platform and the ELIXIR Contextual Data ClearingHouse for the reporting of source material annotation gaps and inaccuracies

Enabling Community Curation of Biological Source Annotations of Molecular Data Through PlutoF and the ELIXIR Contextual Data Clearinghouse