The evolution of the diatoms (Bacillariophyta). I. Origin of the group and assessment of the monophyly of its major divisions

The diatoms are one of the best characterised algal groups. Despite this, little is known of the evolution of the group from the earliest cell to the myriad of taxa known today. Relationships among taxa at the family or generic level have been recognised in some diatoms. However, relationships at higher taxonomic levels are poorly understood and have often been strongly influenced by the first appearances of key taxa in the fossil record. An independent assessment of relationships among the diatoms at these higher taxonomic levels has been made using rRNA sequence data to infer phylogenetic relationships. In this paper we present an analysis of 18S rRNA data from several chosen centric, araphid and raphid pennate taxa. The phylogenetic inferences from these 18S rRNA sequences are supported by evidence from the fossil record and evidence from ontogenetic data. Ribosomal RNA data indicate that both the centric and araphid pennate lineages may not be monophyletic.     

Three extant genera of freshwater thalassiosiroid diatoms from Middle Eocene sediments in northern Canada

The evolutionary history of diatoms is only constrained partially by the fossil record. The timing of several key events, such as initial colonization of freshwater habitats by marine taxa, remains poorly resolved. Numerous specimens of the genera Cyclotella, Discostella, and Puncticulata (Ochrophyta: Thalassiosirales) have been recovered in Middle Eocene lacustrine sediments from the Giraffe Pipe locality in the Northwest Territories, Canada. These diatoms extend the fossil record of the family Stephanodiscaceae to at least 40 million years before present (Ma) and thus provide a new evolutionary milepost for the thalassiosiroid diatoms, an important clade of centric diatoms with global representation in both marine and freshwater environments. The quality of the fossil material enables detailed investigations of areolae, fultoportulae, and rimoportulae, revealing direct morphological affinities with a number of extant taxa. These observations extend the antiquity of several characters of phylogenetic importance within the thalassiosiroid diatoms, including the fultoportula, and imply that the entire lineage is considerably older than prior constraints from the fossil record, as suggested independently by several recent molecular phylogenies.     

Auxosporulation in Paralia guyana MacGillivary (Bacillariophyta) and Possible New Insights into the Habit of the Earliest Diatoms

Background

Diatoms are one of the most ecologically important aquatic micro-eukaryotes. As a group unambiguously recognized as diatoms, they seem to have appeared relatively recently with a limited record of putative remains from oldest sediments. In contrast, molecular clock estimates for the earliest possible emergence of diatoms suggest a considerably older date. Depending on the analysis, Paralia and Leptocylindrus have been recovered within the basal molecular divergences of diatoms. Thus these genera may be in the position to inform on characters that the earliest diatoms possessed.

Findings

Here we present auxospore development and structure of initial and post-auxospore cells in a representative of the ancient non-polar centric genus Paralia. Their initial frustules showed unusual, but not unprecedented, spore-like morphology. Similarly, initial frustules of Leptocylindrus have been long considered resting spores and a unique peculiarity of this genus. However, even though spore-like in appearance, initial cells of Paralia readily resumed mitotic divisions. In addition, Paralia post-auxospore cells underwent several rounds of mitoses in a multi-step process of building a typical, “perfect” vegetative valve. This degree of heteromorphy immediately post-auxosporulation is thus far unknown among the diatoms.

Implications

A spore-related origin of diatoms has already been considered, most recently in the form of the “multiplate diploid cyst” hypothesis. Our discovery that the initial cells in some of the most ancient diatom lineages are structurally spore-like is consistent with that hypothesis because the earliest diatoms may be expected to look somewhat similar to their ancestors. We speculate that because the earliest diatoms may have appeared less diatom-like and more spore-like, they could have gone unrecognized as such in the Triassic/Jurassic sediments. If correct, diatoms may indeed be much older than the fossil record indicates, and possibly more in line with some molecular clock predictions.     

2 Influence of mixed host populations on success of the parasitic dinoflagellate Amoebophrya

Proper taxon sampling is one of the greatest challenges to understanding phylogenetic relationships, perhaps as important as choice of optimality criterion or data type. This has been demonstrated in diatoms where centric diatoms may either be strongly supported as monophyletic or paraphyletic when analyzing SSU rDNA sequences using the same optimality criterion. The effect of ingroup and outgroup taxon sampling on relationships of diatoms is explored for diatoms as a whole and for the order Thalassiosirales. In the latter case, SSU rDNA and rbcL sequence data result in phylogenetic relationships that appear to be strongly incongruent with morphology and broadly incongruent with the fossil record. For example, Cyclotella stelligera Cleve & Grunow behaves like a rogue taxon, jumping from place to place throughout the tree. Morphological data place C. stelligera near the base of the freshwater group as sister to the extinct genus Mesodictyon Theriot and Bradbury, suggesting that it is an old, long branch that might be expected to "misbehave" in poorly sampled trees. Cyclotella stelligera and C. bodanica Grunow delimit the diameter of morphological diversity in Cyclotella , so increased sampling of intermediate taxa will be critical to resolving this part of the tree. Morphology is sampled for a much greater number of taxa and many transitional states of putative synapomorphies seem to suggest a robust morphological hypothesis. The Thalassiosirales are unstable with regards to taxon sampling in the genetic data, suggesting that perhaps the morphological hypothesis is (for now) preferable.     

Distance and Character-Based Evaluation of the V4 Region of the 18S rRNA Gene for the Identification of Diatoms (Bacillariophyceae)

DNA barcoding is a molecular tool that exploits a unique DNA sequence of a standardized gene or non-coding region for the species identification of unknown individuals. The investigation into a suitable barcode for diatoms is ongoing and there are several promising candidates including mitochondrial, plastidial and nuclear markers. We analyzed 272 sequences from 76 diatoms species in the orders Thalassiosirales, Lithodesmiales and Cymatosirales, using distance and character based approaches, to assess the applicability of a DNA barcode based on the hypervariable V4 region of the nuclear 18S rRNA gene. We show that the proposed V4 barcode separated ca. 97% of all centric diatom taxa tested using a threshold p-distance of 0.02 and that many problem pairs were further separated using a character based approach. The reliability of amplification, extensive reference library and variability seen in the V4 region make it the most promising candidate to date for a barcode marker for diatoms particularly when combined with DNA character analysis.     

Mio-pliocene faunal exchanges and african biogeography: the record of fossil bovids

The development of the Ethiopian biogeographic realm since the late Miocene is here explored with the presentation and review of fossil evidence from eastern Africa. Prostrepsiceros cf. vinayaki and an unknown species of possible caprin affinity are described from the hominid-bearing Asa Koma and Kuseralee Members (～5.7 and ～5.2 Ma) of the Middle Awash, Ethiopia. The Middle Awash Prostrepsiceros cf. vinayaki constitutes the first record of this taxon from Africa, previously known from the Siwaliks and Arabia. The possible caprin joins a number of isolated records of caprin or caprin-like taxa recorded, but poorly understood, from the late Neogene of Africa. The identification of these two taxa from the Middle Awash prompts an overdue review of fossil bovids from the sub-Saharan African record that demonstrate Eurasian affinities, including the reduncin Kobus porrecticornis, and species of Tragoportax. The fossil bovid record provides evidence for greater biological continuity between Africa and Eurasia in the late Miocene and earliest Pliocene than is found later in time. In contrast, the early Pliocene (after 5 Ma) saw the loss of any significant proportions of Eurasian-related taxa, and the continental dominance of African-endemic taxa and lineages, a pattern that continues today.     

1 Taxon sampling and inferences about diatom phylogeny

Proper taxon sampling is one of the greatest challenges to understanding phylogenetic relationships, perhaps as important as choice of optimality criterion or data type. This has been demonstrated in diatoms where centric diatoms may either be strongly supported as monophyletic or paraphyletic when analyzing SSU rDNA sequences using the same optimality criterion. The effect of ingroup and outgroup taxon sampling on relationships of diatoms is explored for diatoms as a whole and for the order Thalassiosirales. In the latter case, SSU rDNA and rbcL sequence data result in phylogenetic relationships that appear to be strongly incongruent with morphology and broadly incongruent with the fossil record. For example, Cyclotella stelligera Cleve & Grunow behaves like a rogue taxon, jumping from place to place throughout the tree. Morphological data place C. stelligera near the base of the freshwater group as sister to the extinct genus Mesodictyon Theriot and Bradbury, suggesting that it is an old, long branch that might be expected to “misbehave” in poorly sampled trees. Cyclotella stelligera and C. bodanica Grunow delimit the diameter of morphological diversity in Cyclotella, so increased sampling of intermediate taxa will be critical to resolving this part of the tree. Morphology is sampled for a much greater number of taxa and many transitional states of putative synapomorphies seem to suggest a robust morphological hypothesis. The Thalassiosirales are unstable with regards to taxon sampling in the genetic data, suggesting that perhaps the morphological hypothesis is (for now) preferable.     

Supramolecular organization of fucoxanthin–chlorophyll proteins in centric and pennate diatoms

Fucoxanthin–chlorophyll proteins (FCP) are the major light-harvesting proteins of diatom algae, a major contributor to marine carbon fixation. FCP complexes from representatives of centric (Cyclotella meneghiniana) and pennate (Phaeodactylum tricornutum) diatoms were prepared by sucrose gradient centrifugation and studied by means of electron microscopy followed by single particle analysis. The oligomeric FCP from a centric diatom were observed to take the form of unusual chain-like or circular shapes, a very unique supramolecular assembly for such antennas. The existence of the often disputed oligomeric form of FCP in pennate diatoms has been confirmed. Contrary to the centric diatom FCP, pennate diatom FCP oligomers are very similar to oligomeric antennas from related heterokont (Stramenopila) algae. Evolutionary aspects of the presence of novel light-harvesting protein arrangement in centric diatoms are discussed.     

THE EVOLUTION OF ELONGATE SHAPE IN DIATOMS1

Diatoms have been classified historically as either centric or pennate based on a number of features, cell outline foremost among them. The consensus among nearly every estimate of the diatom phylogeny is that the traditional pennate diatoms (Pennales) constitute a well‐supported clade, whereas centric diatoms do not. The problem with the centric–pennate classification was highlighted by some recent analyses concerning the phylogenetic position of Toxarium, whereby it was concluded that this “centric” diatom independently evolved several pennate‐like characters including an elongate, pennate‐like cell outline. We performed several phylogenetic analyses to test the hypothesis that Toxarium evolved its elongate shape independently from Pennales. First, we reanalyzed the original data set used to infer the phylogenetic position of Toxarium and found that a more thorough heuristic search was necessary to find the optimal tree. Second, we aligned 181 diatom and eight outgroup SSU rDNA sequences to maximize the juxtapositioning of similar primary and secondary structure of the 18S rRNA molecule over a much broader sampling of diatoms. We then performed a number of phylogenetic analyses purposely based on disparate sets of assumptions and found that none of these analyses supported the conclusion that Toxarium acquired its pennate‐like outline independently from Pennales. Our results suggest that elongate outline is congruent with SSU rDNA data and may be synapomorphic for a larger, more inclusive clade than the traditional Pennales.     

The species composition and distribution of diatom algae in sphagnum bogs in European Russia: The Polistovo-Lovatskii land tract

The species composition of diatom algae in the Polistovo-Lovatskii sphagnum tract (Rdeiskoye bog, Novgorod oblast) \are studied. The systematic and ecological-geographical analyses of the flora are performed. A total of 256 species and intraspecific taxa differing in ecological specificities and frequencies of occurrence in samples are revealed. A considerable number of centric diatoms is found. It is shown that primary and secondary diatom communities are formed in large sphagnum tracts. The horizontal heterogeneity of diatom communities in sphagnum bog is revealed. The species diversities of diatom algae in different bogs are compared.     

Centric diatom algae of the Selenga River and its delta branches

Different stages of the study of taxonomical composition of centric diatoms in the Selenga River are considered. A revision of the species composition and a general systematic list are made. New for the Selenga River diatom species are discovered. The role of small-sized centric diatoms in different years and vegetation periods is assessed. A comparison of the composition and the level of development of centric diatoms in 1958–1959 and 2002–2005 is made.     

Extreme differences in rates of molecular evolution of foraminifera revealed by comparison of ribosomal DNA sequences and the fossil record

Foraminifera have one of the best known fossil records among the unicellular eukaryotes. However, the origin and phylogenetic relationships of the extant foraminiferal lineages are poorly understood. To test the current paleontological hypotheses on evolution of foraminifera, we sequenced about 1,000 base pairs from the 3' end of the small subunit rRNA gene (SSU rDNA) in 22 species representing all major taxonomic groups. Phylogenies were derived using neighbor- joining, maximum-parsimony, and maximum-likelihood methods. All analyses confirm the monophyletic origin of foraminifera. Evolutionary relationships within foraminifera inferred from rDNA sequences, however, depend on the method of tree building and on the choice of analyzed sites. In particular, the position of planktonic foraminifera shows important variations. We have shown that these changes result from the extremely high rate of rDNA evolution in this group. By comparing the number of substitutions with the divergence times inferred from the fossil record, we have estimated that the rate of rDNA evolution in planktonic foraminifera is 50 to 100 times faster than in some benthic foraminifera. The use of the maximum-likelihood method and limitation of analyzed sites to the most conserved parts of the SSU rRNA molecule render molecular and paleontological data generally congruent.      

A time-calibrated multi-gene phylogeny of the diatom genus Pinnularia

Pinnularia is an ecologically important and species-rich genus of freshwater diatoms (Bacillariophyceae) showing considerable variation in frustule morphology. Interspecific evolutionary relationships were inferred for 36 Pinnularia taxa using a five-locus dataset. A range of fossil taxa, including newly discovered Middle Eocene forms of Pinnularia, was used to calibrate a relaxed molecular clock analysis and investigate temporal aspects of the genus' diversification. The multi-gene approach resulted in a well-resolved phylogeny of three major clades and several subclades that were frequently, but not universally, delimited by valve morphology. The genus Caloneis was not recovered as monophyletic, confirming that, as currently delimited, this genus is not evolutionarily meaningful and should be merged with Pinnularia. The Pinnularia-Caloneis complex is estimated to have diverged between the Upper Cretaceous and the early Eocene, implying a ghost range of at least 10 million year (Ma) in the fossil record.     

The complete nucleotide sequence of the small-subunit ribosomal RNA coding region for the cycadZamia pumila: Phylogenetic implications

Summary The DNA sequence of the small-subunit ribosomal RNA coding region for the cycadZamia pumila L. was determined. TheZamia smallsubunit rRNA was found to be 1813 nucleotides in length and approximately 92% identical to published angiosperm small-subunit rRNA sequences. Conserved regions interspersed with variable regions are observed corresponding to those found in other eukaryotic small-subunit sequences. Using representatives from protist, fungal, plant, and animal groups, a distance matrix was constructed of average nucleotide substitution rates for pairs of organisms. Phylogenetic trees were inferred from similarities between sequences. The sequence ofZamia represents the earliest divergence from the higher plant lineage reported to date for small-subunit rRNA data. Inferred phylogenies also support a monophyletic origin for the angiosperms consistent with studies citing phenotypic characters.     

Barcoding diatoms: evaluation of the V4 subregion on the 18S rRNA gene,including new primers and protocols

Diatoms are present in all types of water bodies and their species diversity is influenced greatly by environmental conditions. This means that diatom occurrence and abundances are suitable indicators of water quality. Furthermore, continuous screening of algal biodiversity can provide information about diversity changes in ecosystems. Thus, diatoms represent a desirable group for which to develop an easy to use, quick, efficient, and standardised organism identification tool to serve routine water quality assessments. Because conventional morphological identification of diatoms demands specialised in-depth knowledge, we have established standard laboratory procedures for DNA barcoding in diatoms. We (1) identified a short segment (about 400 bp) of the SSU (18S) rRNA gene which is applicable for the identification of diatom taxa, and (2) elaborated a routine protocol including standard primers for this group of microalgae. To test the universality of the primer binding sites and the discriminatory power of the proposed barcode region, 123 taxa, representing limnic diatom diversity, were included in the study and identified at species level. The effectiveness of the barcode was also scrutinised within a closely related species group, namely the Sellaphora pupula taxon complex and relatives.     

The ecology of the planktonic diatom Cyclotella and its implications for global environmental change studies

The fossil record of diatoms in lake sediments can be used to assess the effects of climate variability on lake ecosystems if ecological relationships between diatom community structure and environmental parameters are well understood. Cyclotella sensu lato taxa are a key group of diatoms that are frequently dominant members of phytoplankton communities in low‐ to moderate‐productivity lakes. Their relative abundances have fluctuated significantly in palaeolimnological records spanning over a century in arctic, alpine, boreal and temperate lakes. This suggests that these species are sensitive to environmental change and may serve as early indicators of ecosystem effects of global change. Yet patterns of change in Cyclotella species are not synchronous or unidirectional across, or even within, regions, raising the question of how to interpret these widespread changes in diatom community structure. We suggest that the path forward in resolving seemingly disparate records is to identify clearly the autecology of Cyclotella species, notably the role of nutrients, dissolved organic carbon and light, coupled with better consideration of both the mechanisms controlling lake thermal stratification processes and the resulting effects of changing lake thermal regimes on light and nutrients. Here we begin by reviewing the literature on the resource requirements of common Cyclotella taxa, illustrating that many studies reveal the importance of light, nitrogen, phosphorus, and interactions among these resources in controlling relative abundances. We then discuss how these resource requirements can be linked to shifts in limnological processes driven by environmental change, including climate‐driven change in lakewater temperature, thermal stratification and nutrient loading, as well as acidification‐driven shifts in nutrients and water clarity. We examine three case studies, each involving two lakes from the same region that have disparate trends in the relative abundances of the same species, and illustrate how the mechanisms by which these species abundances are changing can be deciphered. Ultimately, changes in resource availability and water clarity are key factors leading to shifts in Cyclotella abundances. Tighter integration of the autecology of this important group of diatoms with environmental change and subsequent alterations in limnological processes will improve interpretations of palaeolimnological records, and clarify the drivers of seemingly disparate patterns in fossil records showing widespread and rapid changes across the northern hemisphere.     

Seasonal changes on planktonic diatom communities along an inshore-offshore gradient in the Gulf of Gabes (Tunisia)

We investigated the composition, biomass and cell size dynamics of the marine planktonic diatom species along a coastal-open sea gradient in relation to the hydrological characteristics, during four oceanographic cruises in July 2005, May–June 2006, September 2006 and March 2007 in the Gulf of Gabes. The study of the marine planktonic diatoms throughout the sampling period showed the presence of 40 different species belonging to 22 pennate and 18 centric diatom species. Centric diatoms were more abundant than the pennate ones; 56% and 44% of total diatom abundance, respectively. Diatoms were very abundant, representing about 60% of the total phytoplankton abundance, with an exception in July 2005 (19%) during which Dictyochophyceae were the most dominant group (41% of the total phytoplankton abundance). Diatoms, which were dominant in the coastal samples, mainly proliferate in the semi-mixing conditions (May–June 2006), whereas they declined in the offshore area, most likely due to silicate shortage. In this period, in spite of the high abundance of diatom planktonic cells, only diatom biomass was correlated with silica amount, proving that biomass was a better ecological bio-indicator than abundance. The results suggest that the marine planktonic diatoms taxa were generally adapted to specific hydrological structure. In fact, the dominance index of diatoms showed that the biodiversity of diatoms increased gradually along the coastal-open sea gradient except in May–June 2006 during which a slight decrease along the inshore towards offshore areas was observed. This index increase depended on a coastal-open sea distance during thermal stratification (July 2005 and September 2006) and mixing periods (March 2007).     

Comparison of molecular and paleontological data in diatoms suggests a major gap in the fossil record

Diatoms, unicellular eukaryotic algae with a siliceous skeleton, offer the rare advantage of displaying both an extensive fossil record and numerous extant species, thus providing the opportunity of confronting molecular and paleontological data in a protist group. A portion of the 28S ribosomal RNA was sequenced from 5 diatoms, the divergence times of which are well known. The nucleotide substitution rate was estimated in these unicellular eukaryotes and compared with the rate of multicellular eukaryotes, using a broad data base comprising metazoans and metaphytes. When using fossil record derived divergence times, our results show that the nucleotide substitution rate is about 5 times faster in diatoms than in chordates. But, when using the relative rate test, it is observed that, over a long time period, the nucleotide substitution rate may in fact have been slightly slower in diatoms than in chordates. For this contradiction, two possible explanations are proposed: (i) a failure of the relative rate test, (ii) a gap in the pre-Jurassic diatom fossil record. We have checked that our results concerning the relative rate test were valid. Thus, the second hypothesis, which implies pre-Jurassic diatom evolution, in fact already suggested by some non-molecular evidences, is favoured. Decoupling of morphological differentiation from genetic speciation also appears to have occurred and may account in part for the underestimation of the dates of recent cladogenesis events.     

Resolving an ancient, rapid radiation in Saxifragales

Despite the prior use of approximately 9000 bp, deep-level relationships within the angiosperm clade, Saxifragales remain enigmatic, due to an ancient, rapid radiation (89.5 to 110 Ma based on the fossil record). To resolve these deep relationships, we constructed several new data sets: (1) 16 genes representing the three genomic compartments within plant cells (2 nuclear, 10 plastid, 4 mitochondrial; aligned, analyzed length = 21,460 bp) for 28 taxa; (2) the entire plastid inverted repeat (IR; 26,625 bp) for 17 taxa; (3) "total evidence" (50,845 bp) for both 17 and 28 taxa (the latter missing the IR). Bayesian and ML methods yielded identical topologies across partitions with most clades receiving high posterior probability (pp = 1.0) and bootstrap (95% to 100%) values, suggesting that with sufficient data, rapid radiations can be resolved. In contrast, parsimony analyses of different partitions yielded conflicting topologies, particularly with respect to the placement of Paeoniaceae, a clade characterized by a long branch. In agreement with published simulations, the addition of characters increased bootstrap support for the putatively erroneous placement of Paeoniaceae. Although having far fewer parsimony-informative sites, slowly evolving plastid genes provided higher resolution and support for deep-level relationships than rapidly evolving plastid genes, yielding a topology close to the Bayesian and ML total evidence tree. The plastid IR region may be an ideal source of slowly evolving genes for resolution of deep-level angiosperm divergences that date to 90 My or more. Rapidly evolving genes provided support for tip relationships not recovered with slowly evolving genes, indicating some complementarity. Age estimates using penalized likelihood with and without age constraints for the 28-taxon, total evidence data set are comparable to fossil dates, whereas estimates based on the 17-taxon data are much older than implied by the fossil record. Hence, sufficient taxon density, and not simply numerous base pairs, is important in reliably estimating ages. Age estimates indicate that the early diversification of Saxifragales occurred rapidly, over a time span as short as 6 million years. Between 25,000 and 50,000 bp were needed to resolve this radiation with high support values. Extrapolating from Saxifragales, a similar number of base pairs may be needed to resolve the many other deep-level radiations of comparable age in angiosperms.     

Identifying from recent sediment records the effects of nutrients and climate on diatom dynamics in Loch Leven

1. Changes in nutrients and climate have occurred over approximately the same timescales in many European lake catchments. Here, we attempt to interpret the sedimentary diatom record of a large shallow lake, Loch Leven, in relation to these pressures using information gained from analysis of long‐term data sets of water quality, climate and planktonic diatoms. 2. The core data indicate the enrichment of Loch Leven starting in c. 1800–1850, most likely from agricultural practices in the catchment, with a more marked phase since c. 1940–1950 caused by increased phosphorus inputs from sewage treatment works, land drainage and a woollen mill. 3. While the recent diatom plankton remains are dominated by taxa associated with nutrient‐rich conditions, an increase in Aulacoseira subarctica relative to Stephanodiscus taxa since the mid‐1980s suggests that reductions in external catchment sources of nutrients (since 1985) may have resulted in partial recovery. This observation accords well with the long‐term monitoring series of water chemistry and phytoplankton. 4. On a decadal‐centennial scale, the eutrophication signal in the sediment record outweighs any evidence of climate as a control on the diatom community. However, at an inter‐annual scale, while the diatom data exhibit high variability, there are several changes in species composition in the recent fossil record that may be attributed to climatic controls. 5. The study highlights the value of a palaeolimnological approach, particularly when coupled with long‐term data sets, for developing our understanding of environmental change at a range of temporal scales. The diatom record in the sediment can be used effectively to track recovery from eutrophication, but requires greater understanding of contemporary ecology to fully interpret climate impacts. 6. The study illustrates the complexity of ecosystem response to synchronous changes in nutrients and climate, and the difficulty of disentangling the effects of these multiple, interacting pressures.