Deep learning-based diatom taxonomy on virtual slides

Kloster M, Langenkämper D, Zurowietz M, Beszteri B, Nattkemper TW. Deep learning-based diatom taxonomy on virtual slides. Scientific Reports. 2020;10(1): 14416

STAMP: Extensions to the STADEN sequence analysis package for high throughput interactive microsatellite marker design

BackgroundMicrosatellites (MSs) are DNA markers with high analytical power, which are widely used in population genetics, genetic mapping, and forensic studies. Currently available software solutions for high-throughput MS design (i) have shortcomings in detecting and distinguishing imperfect and perfect MSs, (ii) lack often necessary interactive design steps, and (iii) do not allow for the development of primers for multiplex amplifications. We present a set of new tools implemented as extensions to the STADEN package, which provides the backbone functionality for flexible sequence analysis workflows. The possibility to assemble overlapping reads into unique contigs (provided by the base functionality of the STADEN package) is important to avoid developing redundant markers, a feature missing from most other similar tools.ResultsOur extensions to the STADEN package provide the following functionality to facilitate microsatellite (and also minisatellite) marker design: The new modules (i) integrate the state-of-the-art tandem repeat detection and analysis software PHOBOS into workflows, (ii) provide two separate repeat detection steps with different search criteria one for masking repetitive regions during assembly of sequencing reads and the other for designing repeat-flanking primers for MS candidate loci, (iii) incorporate the widely used primer design program PRIMER3 into STADEN workflows, enabling the interactive design and visualization of flanking primers for microsatellites, and (iv) provide the functionality to find optimal locus- and primer pair combinations for multiplex primer design. Furthermore, our extensions include a module for storing analysis results in an SQLite database, providing a transparent solution for data access from within as well as from outside of the STADEN Package.ConclusionThe STADEN package is enhanced by our modules into a highly flexible, high-throughput, interactive tool for conventional and multiplex microsatellite marker design. It gives the user detailed control over the workflow, enabling flexible combinations of manual and automated analysis steps. The software is available under the OpenBSD License [1,2]. The high efficiency of our automated marker design workflow has been confirmed in three microsatellite development projects

Adaptive divergence across Southern Ocean gradients in the pelagic diatom Fragilariopsis kerguelensis

The Southern Ocean is characterized by longitudinal water circulations crossed by strong latitudinal gradients. How this oceanographic background shapes planktonic populations is largely unknown, despite the significance of this region for global biogeochemical cycles. Here, we show, based on genomic, morphometric, ecophysiological and mating compatibility data, an example of ecotypic differentiation and speciation within an endemic pelagic inhabitant, the diatom Fragilariopsis kerguelensis. We discovered three genotypic variants, one present throughout the latitudinal transect sampled, the others restricted to the north and south, respectively. The latter two showed reciprocal monophyly across all three genomes and significant ecophysiological differences consistent with local adaptation, but produced viable offspring in laboratory crosses. The third group was also reproductively isolated from the latter two. We hypothesize that this pattern originated by an adaptive expansion accompanied by ecotypic divergence, followed by sympatric speciation

Quantitative comparison of taxa and taxon concepts in the diatom genus <i>Fragilariopsis</i>: a case study on using slide scanning, multiexpert image annotation, and image analysis in taxonomy

Semiautomated methods for microscopic image acquisition, image analysis, and taxonomic identification have repeatedly received attention in diatom analysis. Less well studied is the question whether and how such methods might prove useful for clarifying the delimitation of species that are difficult to separate for human taxonomists. To try to answer this question, three very similar Fragilariopsis species endemic to the Southern Ocean were targeted in this study: F. obliquecostata, F. ritscheri, and F. sublinearis. A set of 501 extended focus depth specimen images were obtained using a standardized, semiautomated microscopic procedure. Twelve diatomists independently identified these specimen images in order to reconcile taxonomic opinions and agree upon a taxonomic gold standard. Using image analyses, we then extracted morphometric features representing taxonomic characters of the target taxa. The discriminating ability of individual morphometric features was tested visually and statistically, and multivariate classification experiments were performed to test the agreement of the quantitatively defined taxa assignments with expert consensus opinion. Beyond an updated differential diagnosis of the studied taxa, our study also shows that automated imaging and image analysis procedures for diatoms are coming close to reaching a broad applicability for routine use.Facultad de Ciencias Naturales y Muse

Quantitative comparison of taxa and taxon concepts in the diatom genus Fragilariopsis: a case study on using slide scanning, multi‐expert image annotation and image analysis in taxonomy

Semi‐automated methods for microscopic image acquisition, image analysis and taxonomic identification have repeatedly received attention in diatom analysis. Less well studied is the question whether and how such methods might prove useful for clarifying the delimitation of species that are difficult to separate for human taxonomists. To try to answer this question, three very similar Fragilariopsis species endemic to the Southern Ocean were targeted in this study: F. obliquecostata, F. ritscheri, and F. sublinearis. A set of 501 extended focus depth specimen images were obtained using a standardized, semi‐automated microscopic procedure. Twelve diatomists independently identified these specimen images in order to reconcile taxonomic opinions and agree upon a taxonomic gold standard. Using image analyses, we then extracted morphometric features representing taxonomic characters of the target taxa. The discriminating ability of individual morphometric features was tested visually and statistically, and multivariate classification experiments were performed to test the agreement of the quantitatively‐defined taxa assignments with expert consensus opinion. Beyond an updated differential diagnosis of the studied taxa, our study also shows that automated imaging and image analysis procedures for diatoms are coming close to reaching a broad applicability for routine use

The biogeographic differentiation of algal microbiomes in the upper ocean from pole to pole

Eukaryotic phytoplankton are responsible for at least 20% of annual global carbon fixation. Their diversity and activity are shaped by interactions with prokaryotes as part of complex microbiomes. Although differences in their local species diversity have been estimated, we still have a limited understanding of environmental conditions responsible for compositional differences between local species communities on a large scale from pole to pole. Here, we show, based on pole-to-pole phytoplankton metatranscriptomes and microbial rDNA sequencing, that environmental differences between polar and non-polar upper oceans most strongly impact the large-scale spatial pattern of biodiversity and gene activity in algal microbiomes. The geographic differentiation of co-occurring microbes in algal microbiomes can be well explained by the latitudinal temperature gradient and associated break points in their beta diversity, with an average breakpoint at 14 °C ± 4.3, separating cold and warm upper oceans. As global warming impacts upper ocean temperatures, we project that break points of beta diversity move markedly pole-wards. Hence, abrupt regime shifts in algal microbiomes could be caused by anthropogenic climate change

Morphometrische und molekulare Untersuchungen von Artengrenzen in Cyclotella meneghiniana (Bacillariophyceae) und nah verwandten Arten

Comparative molecular genetic and morphometric methods were used to clarify long-standing taxonomic uncertainties in one of the most common freshwater diatom species, Cyclotella meneghiniana Kützing. First a single C. meneghiniana population was sampled by isolating clonal cultures. Frustule ultrastructure of the cultures and of field samples was analysed using two different morphometric methodologies. These analyses contributed significantly to clarifying a problem concerning the delimitation of C. meneghiniana from a closely related species, C. scaldensis Muylaert&Sabbe. Next, patterns of ribosomal DNA sequence variation among fifteen cultures displaying the characteristic C. meneghiniana morphology were explored. The patterns observed indicated the presence of genetic structuring in the population sampled, suggesting that C. meneghiniana was either a complex of multiple, cryptic species, or that of strictly clonally or autogamously reproducing lineages. The following step was surveying sequence variation in the D1/D2 regions of the nuclear ribosomal DNA and in a region of the plastid encoded psaA gene in a collection of strains including C. meneghiniana strains with diverse geographic origins and cultures of some closely related species. Strict covariation was observed in the two genomic regions sequenced, further reinforcing the conclusion that C. meneghiniana is a structured species. The hypothesis that this genetic structuring was caused by the complete lack of allogamous sexual reproduction in this morphospecies was tested by quantifying phylogenetic incompatibility present in AFLP (amplified fragment length polymorphism) genotypes of strains from two populations. The results indicated no significant deviation from panmictic expectations, suggesting that Cyclotella meneghiniana is a complex of several, reproductively isolated species, rather than that of strictly autogamous lineages

Discovering the big world of small things in the web browser with UDE BioSLiDES:

UDE BioSLiDES ist ein auf Ausbildungszwecke ausgerichtetes System für virtuelle digitale Mikroskopie, welches die realitätsnahe Nachbildung lichtmikroskopischer Untersuchungen verschiedener Organismen ermöglicht. Hierfür steht eine Sammlung von über 200 digitalen Präparaten zur Verfügung. Diese Präparate beinhalten hochaufgelöste Scans kompletter Untersuchungsobjekte über eine Vielzahl von Fokusebenen sowie weiterführende Informationen zu Organismus, verschiedenen anatomischen Strukturen, Präparation und verwendeter Mikroskopietechnik. Der im Webbrowser laufende Viewer dient der interaktiven Betrachtung, wobei besonders das Durchfokussieren ein Erarbeiten dreidimensionaler Strukturen und somit ein tieferes Verständnis des Untersuchungsobjektes ermöglicht. Die Inhalte von UDE BioSLiDES stehen als Open Educational Resources in verschiedenen Formaten für eine vielfältige Nutzung und Nachnutzung frei zur Verfügung.UDE BioSLiDES constitutes a system for virtual digital microscopy specifcally designed for educational purposes. It enables realistic emulations of light microscopybased investigations of different organisms; currently it provides more than 200 digital slides. These represent highresolution scans of entire objects of examination over various focal planes, supplemented by additional information about the organism, different anatomical structures, preparation, and used microscopy technique. The web browserbased viewer allows for interactively examining the specimen, whereby especially focal plane changes provide the possibility of working out threedimensional structures – thus enabling a deeper insight into the object of investigation. The contents of UDE BioSLiDES are freely available as Open Educational Resources in different formats for versatile use and repurposing

Three species distribution models for the marine diatom Fragilariopsis kerguelensis of the Southern Ocean, links to NetCDF files

Fragilariopsis kerguelensis, a dominant diatom species throughout the Antarctic Circumpolar Current, is coined to be one of the main drivers of the biological silicate pump. Here, we study the distribution of this important species and expected consequences of climate change upon it, using correlative species distribution modeling and publicly available presence-only data. As experience with SDM is scarce for marine phytoplankton, this also serves as a pilot study for this organism group. We used the maximum entropy method to calculate distribution models for the diatom F. kerguelensis based on yearly and monthly environmental data (sea surface temperature, salinity, nitrate and silicate concentrations). Observation data were harvested from GBIF and the Global Diatom Database, and for further analyses also from the Hustedt Diatom Collection (BRM). The models were projected on current yearly and seasonal environmental data to study current distribution and its seasonality. Furthermore, we projected the seasonal model on future environmental data obtained from climate models for the year 2100. Projected on current yearly averaged environmental data, all models showed similar distribution patterns for F. kerguelensis. The monthly model showed seasonality, for example, a shift of the southern distribution boundary toward the north in the winter. Projections on future scenarios resulted in a moderately to negligibly shrinking distribution area and a change in seasonality. We found a substantial bias in the publicly available observation datasets, which could be reduced by additional observation records we obtained from the Hustedt Diatom Collection. Present-day distribution patterns inferred from the models coincided well with background knowledge and previous reports about F. kerguelensis distribution, showing that maximum entropy-based distribution models are suitable to map distribution patterns for oceanic planktonic organisms. Our scenario projections indicate moderate effects of climate change upon the biogeography of F. kerguelensis