Neoproterozoic origin and multiple transitions to macroscopic growth in green seaweeds.

The Neoproterozoic Era records the transition from a largely bacterial to a predominantly eukaryotic phototrophic world, creating the foundation for the complex benthic ecosystems that have sustained Metazoa from the Ediacaran Period onward. This study focuses on the evolutionary origins of green seaweeds, which play an important ecological role in the benthos of modern sunlit oceans and likely played a crucial part in the evolution of early animals by structuring benthic habitats and providing novel niches. By applying a phylogenomic approach, we resolve deep relationships of the core Chlorophyta (Ulvophyceae or green seaweeds, and freshwater or terrestrial Chlorophyceae and Trebouxiophyceae) and unveil a rapid radiation of Chlorophyceae and the principal lineages of the Ulvophyceae late in the Neoproterozoic Era. Our time-calibrated tree points to an origin and early diversification of green seaweeds in the late Tonian and Cryogenian periods, an interval marked by two global glaciations with strong consequent changes in the amount of available marine benthic habitat. We hypothesize that unicellular and simple multicellular ancestors of green seaweeds survived these extreme climate events in isolated refugia, and diversified in benthic environments that became increasingly available as ice retreated. An increased supply of nutrients and biotic interactions, such as grazing pressure, likely triggered the independent evolution of macroscopic growth via different strategies, including true multicellularity, and multiple types of giant-celled forms.

Historical biogeographical analysis of the Udoteaceae (Bryopsidales, Chlorophyta) elucidates origins of high species diversity in the Central Indo-Pacific, Western Indian Ocean and Greater Caribbean regions.

There is a growing interest in elucidating the biogeographical processes underlying biodiversity patterns of seaweeds, with recent studies largely focusing on red and brown macroalgae. This study focuses on the siphonous green algal family Udoteaceae, which is diverse and globally distributed in tropical to warm-temperate seas, and includes species that form important components of tropical reefs. We explored the historical processes that have shaped current biodiversity patterns in the family by analyzing a comprehensive dataset of 568 specimens sampled across its geographical range, and including 45 species, corresponding to 59% of the known diversity. Historical biogeographical analysis was based on a three-locus time-calibrated phylogeny, and probabilistic modeling of geographical range evolution. Many species were found to have restricted ranges, indicative of low dispersal capacity. Our analysis points toward a Western Tethys origin and early diversification of the Udoteaceae in the Triassic period. Three centers of diversity were identified, which are, in order of highest species richness, the Central Indo-Pacific, the Western Indian Ocean, and the Greater Caribbean. Different drivers have likely played a role in shaping these diversity centres. Species richness in the Central Indo-Pacific likely resulted from speciation within the region, as well as recolonization from neighbouring regions, and overlap of some wider ranged species, corroborating the "biodiversity feedback" model. Species richness in the Western Indian Ocean can be explained by ancient and more recent diversification within the region, and dispersal from the Central Indo-Pacific. The Greater Caribbean region was colonized more recently, followed by diversification within the region.

Plastid phylogenomics of the Sansevieria Clade of Dracaena (Asparagaceae) resolves a recent radiation.

Best known as low maintenance houseplants, sansevierias are a diverse group of flowering plants native to Africa, Madagascar, the Arabian Peninsula, and the Indian subcontinent. Traditionally recognised as a distinct genus, Sansevieria was recently merged with the larger genus Dracaena based on molecular phylogenetic data. Within the Sansevieria Clade of Dracaena, taxonomic uncertainties remain despite attempts to unravel the relationships between the species. To investigate the evolutionary relationships, morphological evolution and biogeographical history in the group, we aim to reconstruct a robust dated phylogenetic hypothesis. Using genome skimming, a chloroplast genome (cpDNA) dataset and a nuclear ribosomal (nrDNA) dataset were generated. The sampling included representatives of all sections and informal groups previously described in Sansevieria based on morphology. Analysis of the cpDNA dataset using a maximum likelihood approach resulted in a well-supported phylogeny. The time-calibrated phylogeny indicated a recent radiation with five main clades emerging in the Pliocene. Two strongly supported clades align with previously defined groups, i.e., Sansevieria section Dracomima, characterised by the Dracomima-type inflorescence, and the Zeylanica informal group, native to the Indian subcontinent. Other previously defined groups were shown to be polyphyletic; a result of convergent evolution of the identifying characters. Switches between flat and cylindrical leaves occurred multiple times in the evolution of the Sansevieria Clade. Similarly, the Cephalantha-type inflorescence has originated multiple times from an ancestor with a Sansevieria-type inflorescence. Analysis of the nrDNA dataset resulted in a phylogenetic hypothesis with low resolution, yet it supported the same two groups confirmed by the cpDNA dataset. This study furthers our understanding of the evolution of the Sansevieria Clade, which will benefit taxonomic and applied research, and aid conservation efforts.

Brilliantia kiribatiensis, a new genus and species of Cladophorales (Chlorophyta) from the remote coral reefs of the Southern Line Islands, Pacific Ocean.

The marine green alga Brilliantia kiribatiensis gen. et sp. nov. is described from samples collected from the coral reefs of the Southern Line Islands, Republic of Kiribati, Pacific Ocean. Phylogenetic analysis of sequences of the large- and small-subunit rDNA and the rDNA internal transcribed spacer region revealed that Brilliantia is a member of the Boodleaceae (Cladophorales), containing the genera Apjohnia, Boodlea, Cladophoropsis, Chamaedoris, Phyllodictyon, and Struvea. Within this clade it formed a distinct lineage, sister to Struvea elegans, but more distantly related to the bona fide Struvea species (including the type S. plumosa). Brilliantia differs from the other genera by having a very simple architecture forming upright, unbranched, single-celled filaments attached to the substratum by a rhizoidal mat. Cell division occurs by segregative cell division only at the onset of reproduction. Based on current sample collection, B. kiribatiensis seems to be largely restricted to the Southern Line Islands, although it was also observed on neighboring islands, including Orona Atoll in the Phoenix Islands of Kiribati, and the Rangiroa and Takapoto Atolls in the Tuamotus of French Polynesia. This discovery highlights the likeliness that there is still much biodiversity yet to be discovered from these remote and pristine reefs of the central Pacific.

Facilitating population genomics of non-model organisms through optimized experimental design for reduced representation sequencing.

BACKGROUND: Genome-wide data are invaluable to characterize differentiation and adaptation of natural populations. Reduced representation sequencing (RRS) subsamples a genome repeatedly across many individuals. However, RRS requires careful optimization and fine-tuning to deliver high marker density while being cost-efficient. The number of genomic fragments created through restriction enzyme digestion and the sequencing library setup must match to achieve sufficient sequencing coverage per locus. Here, we present a workflow based on published information and computational and experimental procedures to investigate and streamline the applicability of RRS. RESULTS: In an iterative process genome size estimates, restriction enzymes and size selection windows were tested and scaled in six classes of Antarctic animals (Ostracoda, Malacostraca, Bivalvia, Asteroidea, Actinopterygii, Aves). Achieving high marker density would be expensive in amphipods, the malacostracan target taxon, due to the large genome size. We propose alternative approaches such as mitogenome or target capture sequencing for this group. Pilot libraries were sequenced for all other target taxa. Ostracods, bivalves, sea stars, and fish showed overall good coverage and marker numbers for downstream population genomic analyses. In contrast, the bird test library produced low coverage and few polymorphic loci, likely due to degraded DNA. CONCLUSIONS: Prior testing and optimization are important to identify which groups are amenable for RRS and where alternative methods may currently offer better cost-benefit ratios. The steps outlined here are easy to follow for other non-model taxa with little genomic resources, thus stimulating efficient resource use for the many pressing research questions in molecular ecology.

Systematics and Biogeography of the Red Algal Genus Yonagunia (Halymeniaceae, Rhodophyta) from the Indo-Pacific Including the Description of Two New Species from Taiwan.

Carpopeltis maillardii has been regarded as a widely distributed species in the Indo-Pacific region. In this study, we analyzed the genetic diversity of C. maillardii and related species collected from Taiwan and the Indian Ocean based on rbcL sequences, in order to elucidate species boundaries, diversity, and biogeographic patterns. Our analyses show that C. maillardii specimens are only distantly related to the genus Carpopeltis (type: C. phyllophora) but instead form a clade together with species of Yonagunia. We therefore propose the new combination Yonagunia maillardii comb. nov. In addition, two new species (Yonagunia palmata sp. nov. and Yonagunia taiwani-borealis sp. nov.) are described from Taiwan. The close relationship of Yonagunia to Grateloupia is corroborated by detailed observations of the female reproductive structures, which demonstrate that the development of auxiliary cell ampullae before and after diploidization is similar to that of Grateloupia sensu stricto. Namely, the ampullae are composed of only two orders of unbranched filaments in which only a few ampullar cells are incorporated into a basal fusion cell after diploidization of the auxiliary cell and the pericarp consists almost entirely of secondary medullary filaments. Of all Yonagunia species, Y. maillardii has the widest distribution in the Indo-Pacific, and can be identified in the field by its relatively thin, feathery, and highly branched morphology. Most other species, including those that occur in Taiwan, are seemingly more range-restricted. Our phylogenetic analyses resulted in a well-resolved phylogeny of Yonagunia, with an origin estimated in the Eocene-Oligocene, and diversification of species mainly in the Miocene.

Chlorocladiella gen. nov. (Pithophoraceae, Cladophorales, Chlorophyta), Including Four New Species From Various Freshwater Habitats in China.

Samples of Pithophoraceae, collected in diverse freshwater and damp terrestrial habitats from tropical and subtropical China, were characterized morphologically and ultrastructurally, and their phylogenetic position was determined based on nuclear ribosomal DNA sequences. Our phylogenetic analysis resolved a novel lineage of Pithophoraceae, sister to Aegagropilopsis. Based on our phylogenetic results, morphological observations and comparative rDNA ITS2 secondary structure analysis, we propose Chlorocladiella gen. nov., characterized by a well-developed system of prostrate filaments, and describe four new species, C. cochlea sp. nov., C. erecta sp. nov., C. medogensis sp. nov., and C. pisformis sp. nov. Two species were found growing on damp soil, which is an unusual habitat for cladophoralean green algae, indicating that the diversity of Cladophorales in terrestrial habitats may be greater than currently recognized.

Reassessment of the classification of Bryopsidales (Chlorophyta) based on chloroplast phylogenomic analyses.

The Bryopsidales is a morphologically diverse group of mainly marine green macroalgae characterized by a siphonous structure. The order is composed of three suborders - Ostreobineae, Bryopsidineae, and Halimedineae. While previous studies improved the higher-level classification of the order, the taxonomic placement of some genera in Bryopsidineae (Pseudobryopsis and Lambia) as well as the relationships between the families of Halimedineae remains uncertain. In this study, we re-assess the phylogeny of the order with datasets derived from chloroplast genomes, drastically increasing the taxon sampling by sequencing 32 new chloroplast genomes. The phylogenies presented here provided good support for the major lineages (suborders and most families) in Bryopsidales. In Bryopsidineae, Pseudobryopsis hainanensis was inferred as a distinct lineage from the three established families allowing us to establish the family Pseudobryopsidaceae. The Antarctic species Lambia antarctica was shown to be an early-branching lineage in the family Bryopsidaceae. In Halimedineae, we revealed several inconsistent phylogenetic positions of macroscopic taxa, and several entirely new lineages of microscopic species. A new classification scheme is proposed, which includes the merger of the families Pseudocodiaceae, Rhipiliaceae and Udoteaceae into a more broadly circumscribed Halimedaceae, and the establishment of tribes for the different lineages found therein. In addition, the deep-water genus Johnson-sea-linkia, currently placed in Rhipiliopsis, was reinstated based on our phylogeny.

Improving phylogenetic inference of core Chlorophyta using chloroplast sequences with strong phylogenetic signals and heterogeneous models.

Phylogenetic relationships within the green algal phylum Chlorophyta have proven difficult to resolve. The core Chlorophyta include Chlorophyceae, Ulvophyceae, Trebouxiophyceae, Pedinophyceae and Chlorodendrophyceae, but the relationships among these classes remain unresolved and the monophyly of Ulvophyceae and Trebouxiophyceae are highly controversial. We analyzed a dataset of 101 green algal species and 73 protein-coding genes sampled from complete and partial chloroplast genomes, including six newly sequenced ulvophyte genomes (Blidingia minima NIES-1837, Ulothrix zonata, Halochlorococcum sp. NIES-1838, Scotinosphaera sp. NIES-154, Caulerpa brownii and Cephaleuros sp. HZ-2017). We applied the Tree Certainty (TC) score to quantify the level of incongruence between phylogenetic trees in chloroplast genomic datasets, and show that the conflicting phylogenetic trees of core Chlorophyta stem from the most GC-heterogeneous sites. With removing the most GC-heterogeneous sites, our chloroplast phylogenomic analyses using heterogeneous models consistently support monophyly of the Chlorophyceae and of the Trebouxiophyceae, but the Ulvophyceae was resolved as polyphyletic. Our analytical framework provides an efficient approach to reconstruct the optimal phylogenetic relationships by minimizing conflicting signals.

The endemic Cladophorales (Ulvophyceae) of ancient Lake Baikal represent a monophyletic group of very closely related but morphologically diverse species.

Lake Baikal, the oldest lake in the world, is home to spectacular biodiversity and extraordinary levels of endemism. While many of the animal species flocks from Lake Baikal are famous examples of evolutionary radiations, the lake also includes a wide diversity of endemic algae that are not well investigated with regards to molecular-biological taxonomy and phylogeny. The endemic taxa of the green algal order Cladophorales show a range of divergent morphologies that led to their classification in four genera in two families. We sequenced partial large- and small-subunit rDNA as well as the internal transcribed spacer region of 14 of the 16 described endemic taxa to clarify their phylogenetic relationships. One endemic morphospecies, Cladophora kusnetzowii, was shown to be conspecific with the widespread Aegagropila linnaei. All other endemic morphospecies formed a monophyletic group nested within the genus Rhizoclonium (Cladophoraceae), a very surprising result, in stark contrast to their morphological affinities. The Baikal clade represents a species flock of closely related taxa with very low genetic differentiation. Some of the morphospecies were congruent with lineages recovered in the phylogenies, but due to the low phylogenetic signal in the rDNA sequences the relationships within the Baikal clade were not all well resolved. The Baikal clade appears to represent a recent radiation, based on the low molecular divergence within the group, and it is hypothesized that the large morphological variation results from diversification in sympatry from a common ancestor in Lake Baikal.

Distinct genetic differentiation and species diversification within two marine nematodes with different habitat preference in Antarctic sediments.

BACKGROUND: Dispersal ability, population genetic structure and species divergence in marine nematodes are still poorly understood, especially in remote areas such as the Southern Ocean. We investigated genetic differentiation of species and populations of the free-living endobenthic nematode genera Sabatieria and Desmodora using nuclear 18S rDNA, internal transcribed spacer (ITS) rDNA, and mitochondrial cytochrome oxidase I (COI) gene sequences. Specimens were collected at continental shelf depths (200-500 m) near the Antarctic Peninsula, Scotia Arc and eastern side of the Weddell Sea. The two nematode genera co-occurred at all sampled locations, but with different vertical distribution in the sediment. A combination of phylogenetic (GMYC, Bayesian Inference, Maximum Likelihood) and population genetic (AMOVA) analyses were used for species delimitation and assessment of gene flow between sampling locations. RESULTS: Sequence analyses resulted in the delimitation of four divergent species lineages in Sabatieria, two of which could not be discriminated morphologically and most likely constitute cryptic species. Two species were recognised in Desmodora, one of which showed large intraspecific morphological variation. Both genera comprised species that were restricted to one side of the Weddell Sea and species that were widely spread across it. Population genetic structuring was highly significant and more pronounced in the deeper sediment-dwelling Sabatieria species, which are generally less prone to resuspension and passive dispersal in the water column than surface Desmodora species. CONCLUSIONS: Our results indicate that gene flow is restricted at large geographic distance in the Southern Ocean, which casts doubt on the efficiency of the Weddell gyre and Antarctic Circumpolar Current in facilitating circum-Antarctic nematode species distributions. We also show that genetic structuring and cryptic speciation can be very different in nematode species isolated from the same geographic area, but with different habitat preferences (surface versus deeper sediment layers).

Historical biogeography of the highly diverse brown seaweed Lobophora (Dictyotales, Phaeophyceae).

The tropical to warm-temperate marine brown macroalgal genus Lobophora (Dictyotales, Phaeophyceae) recently drew attention because of its striking regional diversity. In this study we reassess Lobophora global species diversity, and species distributions, and explore how historical factors have shaped current diversity patterns. We applied a series of algorithmic species delineation techniques on a global mitochondrial cox3 dataset of 598 specimens, resulting in an estimation of 98-121 species. This diversity by far exceeds traditional diversity estimates based on morphological data. A multi-locus time-calibrated species phylogeny using a relaxed molecular clock, along with DNA-confirmed species distribution data was used to analyse ancestral area distributions, dispersal-vicariance-founder events, and temporal patterns of diversification under different biogeographical models. The origin of Lobophora was estimated in the Upper Cretaceous (-75 to -60 MY), followed by gradual diversification until present. While most speciation events were inferred within marine realms, founder events also played a non-negligible role in Lobophora diversification. The Central Indo-Pacific showed the highest species diversity as a result of higher speciation events in this region. Most Lobophora species have small ranges limited to marine realms. Lobophora probably originated in the Tethys Sea and dispersed repeatedly in the Atlantic (including the Gulf of Mexico) and Pacific Oceans. The formation of the major historical marine barriers (Terminal Tethyan event, Isthmus of Panama, Benguela upwelling) did not act as important vicariance events. Long-distance dispersal presumably represented an important mode of speciation over evolutionary time-scales. The limited geographical ranges of most Lobophora species, however, vouch for the rarity of such events.

Molecular phylogeny of the Cladophoraceae (Cladophorales, Ulvophyceae), with the resurrection of Acrocladus Nägeli and Willeella Børgesen, and the description of Lurbica gen. nov. and Pseudorhizoclonium gen. nov.

The taxonomy of the Cladophoraceae, a large family of filamentous green algae, has been problematic for a long time due to morphological simplicity, parallel evolution, phenotypic plasticity, and unknown distribution ranges. Partial large subunit (LSU) rDNA sequences were generated for 362 isolates, and the analyses of a concatenated dataset consisting of unique LSU and small subunit (SSU) rDNA sequences of 95 specimens greatly clarified the phylogeny of the Cladophoraceae. The phylogenetic reconstructions showed that the three currently accepted genera Chaetomorpha, Cladophora, and Rhizoclonium are polyphyletic. The backbone of the phylogeny is robust and the relationships of the main lineages were inferred with high support, only the phylogenetic position of both Chaetomorpha melagonium and Cladophora rupestris could not be inferred unambiguously. There have been at least three independent switches between branched and unbranched morphologies within the Cladophoraceae. Freshwater environments have been colonized twice independently, namely by the freshwater Cladophora species as well as by several lineages of the Rhizoclonium riparium clade. In an effort to establish monophyletic genera, the genera Acrocladus and Willeella are resurrected and two new genera are described: Pseudorhizoclonium and Lurbica.

The chloroplast genomes of Bryopsis plumosa and Tydemania expeditiones (Bryopsidales, Chlorophyta): compact genomes and genes of bacterial origin.

BACKGROUND: Species of Bryopsidales form ecologically important components of seaweed communities worldwide. These siphonous macroalgae are composed of a single giant tubular cell containing millions of nuclei and chloroplasts, and harbor diverse bacterial communities. Little is known about the diversity of chloroplast genomes (cpDNAs) in this group, and about the possible consequences of intracellular bacteria on genome composition of the host. We present the complete cpDNAs of Bryopsis plumosa and Tydemania expeditiones, as well as a re-annotated cpDNA of B. hypnoides, which was shown to contain a higher number of genes than originally published. Chloroplast genomic data were also used to evaluate phylogenetic hypotheses in the Chlorophyta, such as monophyly of the Ulvophyceae (the class in which the order Bryopsidales is currently classified). RESULTS: Both DNAs are circular and lack a large inverted repeat. The cpDNA of B. plumosa is 106,859 bp long and contains 115 unique genes. A 13 kb region was identified with several freestanding open reading frames (ORFs) of putative bacterial origin, including a large ORF (>8 kb) closely related to bacterial rhs-family genes. The cpDNA of T. expeditiones is 105,200 bp long and contains 125 unique genes. As in B. plumosa, several regions were identified with ORFs of possible bacterial origin, including genes involved in mobile functions (transposases, integrases, phage/plasmid DNA primases), and ORFs showing close similarity with bacterial DNA methyltransferases. The cpDNA of B. hypnoides differs from that of B. plumosa mainly in the presence of long intergenic spacers, and a large tRNA region. Chloroplast phylogenomic analyses were largely inconclusive with respect to monophyly of the Ulvophyceae, and the relationship of the Bryopsidales within the Chlorophyta. CONCLUSIONS: The cpDNAs of B. plumosa and T. expeditiones are amongst the smallest and most gene dense chloroplast genomes in the core Chlorophyta. The presence of bacterial genes, including genes typically found in mobile elements, suggest that these have been acquired through horizontal gene transfer, which may have been facilitated by the occurrence of obligate intracellular bacteria in these siphonous algae.

Exploration of nuclear DNA markers for population structure assessment in the desmid Micrasterias rotata (Zygnematophyceae, Streptophyta).

Freshwater green microalgae are diverse and widely distributed across the globe, yet the population structuring of these organisms is poorly understood. We assessed the degree of genetic diversity and differentiation of the desmid species, Micrasterias rotata. First, we compared the sequences of four nuclear regions (actin, gapC1, gapC2, and oee1) in 25 strains and selected the gapC1 and actin regions as the most appropriate markers for population structure assessment in this species. Population genetic structure was subsequently analyzed, based on seven populations from the Czech Republic and Ireland. Hudson's Snn statistics indicated that nearest-neighbor sequences occurred significantly more frequently within geographical populations than within the wider panmictic population. Moreover, Irish populations consistently showed higher genetic diversity than the Czech samples. These results are in accordance with the unbalanced distribution of alleles in many land plant species; however, the large genetic diversity in M. rotata differs from levels of genetic diversity found in most land plants.

Cryptic diversity in southern African kelp.

The southern coast of Africa is one of the few places in the world where water temperatures are predicted to cool in the future. This endemism-rich coastline is home to two sister species of kelps of the genus Ecklonia maxima and Ecklonia radiata, each associated with specific thermal niches, and occuring primarily on opposite sides of the southern tip of Africa. Historical distribution records indicate that E. maxima has recently shifted its distribution ~ 70 km eastward, to sites where only E. radiata was previously reported. The contact of sister species with contrasting thermal affinities and the occurrence of mixed morphologies raised the hypothesis that hybridization might be occurring in this contact zone. Here we describe the genetic structure of the genus Ecklonia along the southern coast of Africa and investigate potential hybridization and cryptic diversity using a combination of nuclear microsatellites and mitochondrial markers. We found that both species have geographically discrete genetic clusters, consistent with expected phylogeographic breaks along this coastline. In addition, depth-isolated populations were found to harbor unique genetic diversity, including a third Ecklonia lineage. Mito-nuclear discordance and high genetic divergence in the contact zones suggest multiple hybridization events between Ecklonia species. Discordance between morphological and molecular identification suggests the potential influence of abiotic factors leading to convergent phenotypes in the contact zones. Our results highlight an example of cryptic diversity and hybridization driven by contact between two closely related keystone species with contrasting thermal affinities.

Permanent residents or temporary lodgers: characterizing intracellular bacterial communities in the siphonous green alga Bryopsis.

The ecological success of giant celled, siphonous green algae in coastal habitats has repeatedly been linked to endophytic bacteria living within the cytoplasm of the hosts. Yet, very little is known about the relative importance of evolutionary and ecological factors controlling the intracellular bacterial flora of these seaweeds. Using the marine alga Bryopsis (Bryopsidales, Chlorophyta) as a model, we explore the diversity of the intracellular bacterial communities and investigate whether their composition is controlled by ecological and biogeographic factors rather than the evolutionary history of the host. Using a combination of 16S rDNA clone libraries and denaturing gradient gel electrophoresis analyses, we show that Bryopsis harbours a mixture of relatively few but phylogenetically diverse bacterial species. Variation partitioning analyses show a strong impact of local environmental factors on the presence of Rickettsia and Mycoplasma in their association with Bryopsis. The presence of Flavobacteriaceae and Bacteroidetes, on the other hand, reflects a predominant imprint of host evolutionary history, suggesting that these bacteria are more specialized in their association. The results highlight the importance of interpreting the presence of individual bacterial phylotypes in the light of ecological and evolutionary principles such as phylogenetic niche conservatism to understand complex endobiotic communities and the parameters shaping them.

Extensive cryptic species diversity and fine-scale endemism in the marine red alga Portieria in the Philippines.

We investigated species diversity and distribution patterns of the marine red alga Portieria in the Philippine archipelago. Species boundaries were tested based on mitochondrial, plastid and nuclear encoded loci, using a general mixed Yule-coalescent (GMYC) model-based approach and a bayesian multilocus species delimitation method. The outcome of the GMYC analysis of the mitochondrial encoded cox2-3 dataset was highly congruent with the multilocus analysis. In stark contrast with the current morphology-based assumption that the genus includes a single, widely distributed species in the Indo-West Pacific (Portieria hornemannii), DNA-based species delimitation resulted in the recognition of 21 species within the Philippines. Species distributions were found to be highly structured with most species restricted to island groups within the archipelago. These extremely narrow species ranges and high levels of intra-archipelagic endemism contrast with the wide-held belief that marine organisms generally have large geographical ranges and that endemism is at most restricted to the archipelagic level. Our results indicate that speciation in the marine environment may occur at spatial scales smaller than 100 km, comparable with some terrestrial systems. Our finding of fine-scale endemism has important consequences for marine conservation and management.

Host specificity and coevolution of Flavobacteriaceae endosymbionts within the siphonous green seaweed Bryopsis.

The siphonous green seaweed Bryopsis harbors complex intracellular bacterial communities. Previous studies demonstrated that certain species form close, obligate associations with Flavobacteriaceae. A predominant imprint of host evolutionary history on the presence of these bacteria suggests a highly specialized association. In this study we elaborate on previous results by expanding the taxon sampling and testing for host-symbiont coevolution Therefore, we optimized a PCR protocol to directly and specifically amplify Flavobacteriaceae endosymbiont 16S rRNA gene sequences, which allowed us to screen a large number of algal samples without the need for cultivation or surface sterilization. We analyzed 146 Bryopsis samples, and 92 additional samples belonging to the Bryopsidales and other orders within the class Ulvophyceae. Results indicate that the Flavobacteriaceae endosymbionts are restricted to Bryopsis, and only occur within specific, warm-temperate and tropical clades of the genus. Statistical analyses (AMOVA) demonstrate a significant non-random host-symbiont association. Comparison of bacterial 16S rRNA and Bryopsis rbcL phylogenies, however, reveal complex host-symbiont evolutionary associations, whereby closely related hosts predominantly harbor genetically similar endosymbionts. Bacterial genotypes are rarely confined to a single Bryopsis species and most Bryopsis species harbored several Flavobacteriaceae, obscuring a clear pattern of coevolution.