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Genome assembly graphs contain valuable yet frequently overlooked information that can enhance assembly completeness by revealing contig connectivity. Here, we demonstrate how leveraging these information‐rich structures enables the discovery of hidden microeukaryotic diversity in environmental DNA shotgun metagenomic datasets. While GetOrganelle has previously been used for organellar genome assembly from isolated tissues, we present its first application to water eDNA metagenomic data, using diatoms as an example. We tested the efficiency of this organellar genome assembly tool on three freshwater eDNA metagenomic datasets with varying diatom abundances, finding that GetOrganelle alone yields fragmented scaffolds due to mixed‐species complexity. By implementing manual disentanglement of assembly graphs, we successfully recovered complete organellar genomes from these assemblies. From high‐abundance bloom samples, we recovered complete plastomes ofStephanodiscus hantzschiiwith 99.9% pairwise identity across distant geographical locations (USA and Czech Republic). From a lower abundance non‐bloom sample, we reconstructed a potentially novelCyclotellaplastome with only 94.0% identity to its closest available reference,Cyclotella atomus. Our assembly quality assessment confirmed effective manual disentanglement even at low diatom abundances. By integrating sequence similarity, gene order conservation and phylogenetic analysis, we achieved robust species‐level resolution and resolved previous taxonomic uncertainties. Our findings demonstrate that mining eDNA metagenomic data with GetOrganelle reveals previously hidden microeukaryotic diversity and provides higher taxonomic resolution than traditional binning methods. This approach proves especially valuable for microeukaryotes, where reference organellar genomes remain underrepresented in existing databases. 

A principal goal in ecology is to identify the determinants of species abundances in nature. Body size has emerged as a fundamental and repeatable predictor of abundance, with smaller organisms occurring in greater numbers than larger ones. A biogeographic component, known as Bergmann’s rule, describes the preponderance, across taxonomic groups, of larger-bodied organisms in colder areas. Although undeniably important, the extent to which body size is the key trait underlying these patterns is unclear. We explored these questions in diatoms, unicellular algae of global importance for their roles in carbon fixation and energy flow through marine food webs. Using a phylogenomic dataset from a single lineage with worldwide distribution, we found that body size (cell volume) was strongly correlated with genome size, which varied by 50-fold across species and was driven by differences in the amount of repetitive DNA. However, directional models identified temperature and genome size, not cell size, as having the greatest influence on maximum population growth rate. A global metabarcoding dataset further identified genome size as a strong predictor of species abundance in the ocean, but only in colder regions at high and low latitudes where diatoms with large genomes dominated, a pattern consistent with Bergmann’s rule. Although species abundances are shaped by myriad interacting abiotic and biotic factors, genome size alone was a remarkably strong predictor of abundance. Taken together, these results highlight the cascading cellular and ecological consequences of macroevolutionary changes in an emergent trait, genome size, one of the most fundamental and irreducible properties of an organism. 

Environmental transitions, such as the salinity divide separating marine and fresh waters, shape biodiversity over both shallow and deep timescales, opening up new niches and creating opportunities for accelerated speciation and adaptive radiation. Understanding the genetics of environmental adaptation is central to understanding how organisms colonise and subsequently diversify in new habitats. We used time‐resolved transcriptomics to contrast the hyposalinity stress responses of two diatoms. Skeletonema marinoi has deep marine ancestry but has recently invaded brackish waters. Cyclotella cryptica has deep freshwater ancestry and can withstand a much broader salinity range. Skeletonema marinoi is less adept at mitigating even mild salinity stress compared to Cyclotella cryptica, which has distinct mechanisms for rapid mitigation of hyposaline stress and long‐term growth in low salinity. We show that the cellular mechanisms underlying low salinity tolerance, which has allowed diversification across freshwater habitats worldwide, includes elements that are both conserved and variable across the diatom lineage. The balance between ancestral and lineage‐specific environmental responses in phytoplankton have shaped marine–freshwater transitions on evolutionary timescales and, on contemporary timescales, will affect which lineages survive and adapt to changing ocean conditions. 

The Cretaceous period is the time of the first appearance of the diatoms in the fossil record. These fossils give us direct evidence of the age and early evolution of the diatom lineage. The fossil record, however, is incomplete and therefore often extrapolated through time‐calibrated phylogenies. These two approaches offer different perspectives on the early evolution of diatoms, which is still poorly understood. We compiled the first comprehensive Cretaceous Diatom Database, a tool to investigate the taxonomy, diversity, and occurrence of the earliest known diatom lineages. To further aid the integration and use of the oldest diatom fossils in molecular clock analyses, we present a set of well‐documented Cretaceous fossils that can be placed onto molecular phylogenetic trees of extant and extinct species, making them ideal candidates for the calibration of molecular clocks. The analysis of the fossil record and the Cretaceous Diatom Database revealed Cretaceous diversity is substantially greater than previously thought, yet considerable taxonomic work is still needed. The Cretaceous Diatom Database and the list of Cretaceous fossils for calibrating molecular clocks represent valuable resources for future evolutionary and taxonomic studies of modern and fossil diatoms. 

Diatoms are an important component of marine and freshwater ecosystems. Although the majority of described diatom species live in freshwater systems, genome sequencing efforts have focused primarily on marine species. Genomic resources for freshwater species have the potential to improve our understanding of diatom ecology and evolution, particularly in the context of major environmental shifts. We used long‐ and short‐read sequencing platforms to assemble reference genomes for three freshwater diatom species, all in the order Thalalassiosirales, which are abundant in the plankton of oceans, lakes, reservoirs, and rivers worldwide. We targeted three species that cover the breadth of phylogenetic diversity in the cyclostephanoid clade of Thalassiosirales:Cyclostephanos tholiformis(JALLPB020000000),Discostella pseudostelligera(JALLBG020000000), andPraestephanos triporus(JALLAZ020000000). The reference genome forD. pseudostelligerawas considerably smaller (39 Mb) than those of bothP. triporus(73 Mb) andC. tholiformis(177 Mb). Long‐read sequencing allowed for the assembly of scaffold‐level genomes, including regions rich in repetitive DNA. Compared to short‐read assemblies, long‐read assemblies increased the contig N50 length as much as 37‐fold and reduced the number of contigs by more than 88%. Transcriptome‐guided annotation of the protein‐coding genes identified between 10,000 and 12,000 genes. This work provides further demonstration of the value of long‐read sequencing and provides novel genomic resources for understanding the ecology and evolution of freshwater diatoms. 

Evolution is often uneven in its pace and outcomes, with long periods of stasis interrupted by abrupt increases in morphological and ecological disparity. With thousands of gene histories, phylogenomics can uncover the genomic signatures of these broad macroevolutionary trends. Diatoms are a species-rich lineage of microeukaryotes that contribute greatly to the global cycling of carbon, oxygen, and silica, which they use to build elaborately structured cell walls. We combined fossil information with newly sequenced transcriptomes from 181 diverse diatom species to reconstruct the pattern, timing, and genomic context of major evolutionary transitions. Diatoms originated 270 Mya, and after >100 My of relative stasis in morphology and ecology, a radiation near the Jurassic–Cretaceous boundary led to the diversity of habitats and cell wall architectures characteristic of modern diatoms. This transition was marked by a genome duplication and high levels of gene tree discordance. However, short generation times increase the probability of coalescence between speciation events, minimizing the impacts of incomplete lineage sorting and implicating sequence saturation and gene tree error as the main sources of discordance. Nevertheless, a rigorous tree-based approach to ortholog selection resulted in strongly supported relationships, including some that were uncertain previously. Three pulses of accelerated speciation were detected, two of which were associated with the evolution of novel traits and ecological transitions. The first 100 My of diatom evolution was a slow-burning fuse that led to a burst of innovations in ecology, morphology, and life history that are hallmarks of contemporary diatom assemblages. 

Abstract Molecular clocks estimate that diatom microalgae, one of Earth’s foremost primary producers, originated near the Triassic–Jurassic boundary (200 Ma), which is close in age to the earliest, generally accepted diatom fossils of the genus Pyxidicula . During an extensive search for Jurassic diatoms from twenty-five sites worldwide, three sites yielded microfossils initially recognized as diatoms. After applying stringent safeguards and evaluation criteria, however, the fossils found at each of the three sites were rejected as new diatom records. This led us to systematically reexamine published evidence in support of Lower- and Middle-Jurassic Pyxidicula fossils . Although Pyxidicula resembles some extant radial centric diatoms and has character states that may have been similar to those of ancestral diatoms, we describe numerous sources of uncertainty regarding the reliability of these records. We conclude that the Lower Jurassic Pyxidicula fossils were most likely calcareous nannofossils, whereas the Middle Jurassic Pyxidicula species has been reassigned to the Lower Cretaceous and is likely a testate amoeba, not a diatom. Excluding the Pyxidicula fossils widens the gap between the estimated time of origin and the oldest abundant fossil diatom record to 75 million years. This study underscores the difficulties in discovering and validating ancient microfossils. 

Abstract Numerous factors shape the evolution of protein-coding genes, including shifts in the strength or type of selection following gene duplications or changes in the environment. Diatoms and other silicifying organisms use a family of silicon transporters (SITs) to import dissolved silicon from the environment. Freshwaters contain higher silicon levels than oceans, and marine diatoms have more efficient uptake kinetics and less silicon in their cell walls, making them better competitors for a scarce resource. We compiled SITs from 37 diatom genomes to characterize shifts in selection following gene duplications and marine–freshwater transitions. A deep gene duplication, which coincided with a whole-genome duplication, gave rise to two gene lineages. One of them (SIT1–2) is present in multiple copies in most species and is known to actively import silicon. These SITs have evolved under strong purifying selection that was relaxed in freshwater taxa. Episodic diversifying selection was detected but not associated with gene duplications or habitat shifts. In contrast, genes in the second SIT lineage (SIT3) were present in just half the species, the result of multiple losses. Despite conservation of SIT3 in some lineages for the past 90–100 million years, repeated losses, relaxed selection, and low expression highlighted the dispensability of SIT3, consistent with a model of deterioration and eventual loss due to relaxed selection on SIT3 expression. The extensive but relatively balanced history of duplications and losses, together with paralog-specific expression patterns, suggest diatoms continuously balance gene dosage and expression dynamics to optimize silicon transport across major environmental gradients. 

Despite the obstacles facing marine colonists, most lineages of aquatic organisms have colonized and diversified in freshwaters repeatedly. These transitions can trigger rapid morphological or physiological change and, on longer timescales, lead to increased rates of speciation and extinction. Diatoms are a lineage of ancestrally marine microalgae that have diversified throughout freshwater habitats worldwide. We generated a phylogenomic data set of genomes and transcriptomes for 59 diatom taxa to resolve freshwater transitions in one lineage, the Thalassiosirales. Although most parts of the species tree were consistently resolved with strong support, we had difficulties resolving a Paleocene radiation, which affected the placement of one freshwater lineage. This and other parts of the tree were characterized by high levels of gene tree discordance caused by incomplete lineage sorting and low phylogenetic signal. Despite differences in species trees inferred from concatenation versus summary methods and codons versus amino acids, traditional methods of ancestral state reconstruction supported six transitions into freshwaters, two of which led to subsequent species diversification. Evidence from gene trees, protein alignments, and diatom life history together suggest that habitat transitions were largely the product of homoplasy rather than hemiplasy, a condition where transitions occur on branches in gene trees not shared with the species tree. Nevertheless, we identified a set of putatively hemiplasious genes, many of which have been associated with shifts to low salinity, indicating that hemiplasy played a small but potentially important role in freshwater adaptation. Accounting for differences in evolutionary outcomes, in which some taxa became locked into freshwaters while others were able to return to the ocean or become salinity generalists, might help further distinguish different sources of adaptive mutation in freshwater diatoms.