Noa Shenkar

An updated 18S rRNA phylogeny of tunicates based on mixture and secondary structure models.

BackgroundTunicates have been recently revealed to be the closest living relatives of vertebrates. Yet, with more than 2500 described species, details of their evolutionary history are still obscure. From a molecular point of view, tunicate phylogenetic relationships have been mostly studied based on analyses of 18S rRNA sequences, which indicate several major clades at odds with the traditional class-level arrangements. Nonetheless, substantial uncertainty remains about the phylogenetic relationships and taxonomic status of key groups such as the Aplousobranchia, Appendicularia, and Thaliacea.ResultsThirty new complete 18S rRNA sequences were acquired from previously unsampled tunicate species, with special focus on groups presenting high evolutionary rate. The updated 18S rRNA dataset has been aligned with respect to the constraint on homology imposed by the rRNA secondary structure. A probabilistic framework of phylogenetic reconstruction was adopted to accommodate the particular evolutionary dynamics of this ribosomal marker. Detailed Bayesian analyses were conducted under the non-parametric CAT mixture model accounting for site-specific heterogeneity of the evolutionary process, and under RNA-specific doublet models accommodating the occurrence of compensatory substitutions in stem regions. Our results support the division of tunicates into three major clades: 1) Phlebobranchia + Thaliacea + Aplousobranchia, 2) Appendicularia, and 3) Stolidobranchia, but the position of Appendicularia could not be firmly resolved. Our study additionally reveals that most Aplousobranchia evolve at extremely high rates involving changes in secondary structure of their 18S rRNA, with the exception of the family Clavelinidae, which appears to be slowly evolving. This extreme rate heterogeneity precluded resolving with certainty the exact phylogenetic placement of Aplousobranchia. Finally, the best fitting secondary-structure and CAT-mixture models suggest a sister-group relationship between Salpida and Pyrosomatida within Thaliacea.ConclusionAn updated phylogenetic framework for tunicates is provided based on phylogenetic analyses using the most realistic evolutionary models currently available for ribosomal molecules and an unprecedented taxonomic sampling. Detailed analyses of the 18S rRNA gene allowed a clear definition of the major tunicate groups and revealed contrasting evolutionary dynamics among major lineages. The resolving power of this gene nevertheless appears limited within the clades composed of Phlebobranchia + Thaliacea + Aplousobranchia and Pyuridae + Styelidae, which were delineated as spots of low resolution. These limitations underline the need to develop new nuclear markers in order to further resolve the phylogeny of this keystone group in chordate evolution.

Global Diversity of Ascidiacea

The class Ascidiacea presents fundamental opportunities for research in the fields of development, evolution, ecology, natural products and more. This review provides a comprehensive overview of the current knowledge regarding the global biodiversity of the class Ascidiacea, focusing in their taxonomy, main regions of biodiversity, and distribution patterns. Based on analysis of the literature and the species registered in the online World Register of Marine Species, we assembled a list of 2815 described species. The highest number of species and families is found in the order Aplousobranchia. Didemnidae and Styelidae families have the highest number of species with more than 500 within each group. Sixty percent of described species are colonial. Species richness is highest in tropical regions, where colonial species predominate. In higher latitudes solitary species gradually contribute more to the total species richness. We emphasize the strong association between species richness and sampling efforts, and discuss the risks of invasive species. Our inventory is certainly incomplete as the ascidian fauna in many areas around the world is relatively poorly known, and many new species continue to be discovered and described each year.

Tunicate mitogenomics and phylogenetics: peculiarities of the Herdmania momus mitochondrial genome and support for the new chordate phylogeny

BackgroundTunicates represent a key metazoan group as the sister-group of vertebrates within chordates. The six complete mitochondrial genomes available so far for tunicates have revealed distinctive features. Extensive gene rearrangements and particularly high evolutionary rates have been evidenced with regard to other chordates. This peculiar evolutionary dynamics has hampered the reconstruction of tunicate phylogenetic relationships within chordates based on mitogenomic data.ResultsIn order to further understand the atypical evolutionary dynamics of the mitochondrial genome of tunicates, we determined the complete sequence of the solitary ascidian Herdmania momus. This genome from a stolidobranch ascidian presents the typical tunicate gene content with 13 protein-coding genes, 2 rRNAs and 24 tRNAs which are all encoded on the same strand. However, it also presents a novel gene arrangement, highlighting the extreme plasticity of gene order observed in tunicate mitochondrial genomes. Probabilistic phylogenetic inferences were conducted on the concatenation of the 13 mitochondrial protein-coding genes from representatives of major metazoan phyla. We show that whereas standard homogeneous amino acid models support an artefactual sister position of tunicates relative to all other bilaterians, the CAT and CAT+BP site- and time-heterogeneous mixture models place tunicates as the sister-group of vertebrates within monophyletic chordates. Moreover, the reference phylogeny indicates that tunicate mitochondrial genomes have experienced a drastic acceleration in their evolutionary rate that equally affects protein-coding and ribosomal-RNA genes.ConclusionThis is the first mitogenomic study supporting the new chordate phylogeny revealed by recent phylogenomic analyses. It illustrates the beneficial effects of an increased taxon sampling coupled with the use of more realistic amino acid substitution models for the reconstruction of animal phylogeny.

Deep Sequencing of Mixed Total DNA without Barcodes Allows Efficient Assembly of Highly Plastic Ascidian Mitochondrial Genomes

Ascidians or sea squirts form a diverse group within chordates, which includes a few thousand members of marine sessile filter-feeding animals. Their mitochondrial genomes are characterized by particularly high evolutionary rates and rampant gene rearrangements. This extreme variability complicates standard polymerase chain reaction (PCR) based techniques for molecular characterization studies, and consequently only a few complete Ascidian mitochondrial genome sequences are available. Using the standard PCR and Sanger sequencing approach, we produced the mitochondrial genome of Ascidiella aspersa only after a great effort. In contrast, we produced five additional mitogenomes (Botrylloides aff. leachii, Halocynthia spinosa, Polycarpa mytiligera, Pyura gangelion, and Rhodosoma turcicum) with a novel strategy, consisting in sequencing the pooled total DNA samples of these five species using one Illumina HiSeq 2000 flow cell lane. Each mitogenome was efficiently assembled in a single contig using de novo transcriptome assembly, as de novo genome assembly generally performed poorly for this task. Each of the new six mitogenomes presents a different and novel gene order, showing that no syntenic block has been conserved at the ordinal level (in Stolidobranchia and in Phlebobranchia). Phylogenetic analyses support the paraphyly of both Ascidiacea and Phlebobranchia, with Thaliacea nested inside Phlebobranchia, although the deepest nodes of the Phlebobranchia–Thaliacea clade are not well resolved. The strategy described here thus provides a cost-effective approach to obtain complete mitogenomes characterized by a highly plastic gene order and a fast nucleotide/amino acid substitution rate.

The solitary ascidian Herdmania momus: native (Red Sea) versus non-indigenous (Mediterranean) populations

During 2005 monthly samples of the solitary ascidian Herdmania momus were collected along the Mediterranean and Red Sea coasts of Israel, in order to investigate possible differences in life history strategies of the two populations. The samples were preserved, dissected, and measurements were made of length, total weight, gonad weight, oocyte diameter and the occurrence of symbionts was recorded. Additionally, field surveys showed that in the Mediterranean H. momus exclusively inhabit artificial substrates, and are common at greater depths than in Eilat (Red Sea). Individuals of H. momus in Eilat reproduced year round. Although individuals from the Mediterranean were significantly larger than individuals collected in Eilat their gonad indices and oocyte diameter measurements indicate that they have a short reproductive season. Copepods were found in 50% of the samples from both sites, while a pontoniine shrimp was found in 14% of the samples from Eilat only. The marked differences between the native vs. non-indigenous populations of H. momus are attributed to differential food availability, water temperature, currents and wave exposure. The increasing evidence of negative effects of non-indigenous ascidians on natural fauna from other regions in the world emphasizes the need for additional research regarding the ecology of ascidians along the coasts of Israel.

Non-indigenous ascidians (Chordata: Tunicata) along the Mediterranean coast of Israel

One of the most extensively documented marine bioinvasion phenomena is the invasion of Red Sea species to the Mediterranean Sea through the Suez Canal. The present study reports the occurrence of seven non-indigenous ascidian species along the Mediterranean coast of Israel: Ecteinascidia thurstoni, Ascidia cannelata, Phallusia nigra, Rhodosoma turcicum, Symplegma brakenhielmi, Microcosmus exasperatus and Herdmania momus . Five of these species (excluding P. nigra , and R. turcicum ) probably reached the Mediterranean via the Suez Canal, since they have an extra-tropical Indo-Pacific distribution and a restricted distribution in the eastern Mediterranean. This is the first record of E. thurstoni in the Mediterranean. The accumulating evidence for the negative impact of non-indigenous ascidians on local species and habitats raises the necessity for long-term studies and monitoring of this group.

Population dynamics of zooxanthellae during a bacterial bleaching event

Each summer 80–90% of the colonies of Oculina patagonica undergo bleaching off the Mediterranean coast of Israel. To investigate fluctuations through a yearly bleaching cycle, monthly measurements of zooxanthella density, mitotic index and chlorophyll-a concentration were conducted. Results showed (1) a significant negative correlation between sea surface temperature (SST) and zooxanthella density; (2) both significantly lower zooxanthella mitotic index and higher chlorophyll-a per zooxanthella content during the bleaching season compared with the non-bleaching period; (3) prior to bleaching, a lag between the peak of zooxanthella density and chlorophyll-a concentration followed by a similar lag during recovery. Zooxanthella density declined significantly between March and May while chlorophyll-a concentration peaked in April, and then declined. Zooxanthella density increased significantly in November while chlorophyll-a concentration increased significantly in January. We conclude that during bacterial bleaching events, zooxanthellae are severely damaged. However, by the time of the following bleaching event the coral tissues regain their “normal” (pre-bleaching) zooxanthella population density.

Ascidian recruitment patterns on an artificial reef in Eilat (Red Sea)

Although ascidians are conspicuous members of the fouling community not much is known regarding their recruitment patterns in coral reefs. A 1-year study was carried out along the Red Sea coast of Israel to examine the effects of season and spatial distribution on ascidian recruitment to artificial marine structures. In general, autumn and spring were characterized by higher coverage with a significantly higher percentage of cover of Didemnum granulatum in autumn and higher numbers of Herdmania momus in spring. These species contributed the most to similarity between treatments consequently setting the pattern for each group (colonial and solitary). Halocynthia spinosa had significantly higher numbers during winter and Phallusia nigra was absent in spring and winter. H. momus showed a preference for horizontal surfaces. P. nigra and Ascidia cannelata showed a preference for floating units. It is concluded that the ascidian recruitment patterns are species-specific and vary between seasons, orientation and position on the substrata and in the water column.

Back to solitude: Solving the phylogenetic position of the Diazonidae using molecular and developmental characters.

The order Aplousobranchia (Chordata, Ascidiacea) contains approximately 1500 species distributed worldwide. Their phylogeny, however, remains unclear, with unresolved family relationships. While most Aplousobranchia are colonial, debates exist concerning the phylogenetic position of families such as the Diazonidae and Cionidae, which exhibit a solitary lifestyle and share morphological characteristics with both Aplousobranchia and Phlebobranchia orders. To clarify the phylogenetic position of the Diazonidae and Cionidae, we determined the complete mitochondrial sequence of the solitary diazonid Rhopalaea idoneta. The phylogenetic reconstruction based on the 13 mitochondrial protein coding genes strongly supports a positioning of Diazonidae well-nested within the Aplousobranchia rather than a positioning as a sister clade of the Aplousobranchia. In addition, we examined the regenerative ability of R. idoneta. Similar to colonial Aplousobranchia, R. idoneta was found to be able to completely regenerate its thorax. Ciona, also known to possess high regenerative abilities, is the Aplousobranchia sister clade rather than a member of the Phlebobranchia. Our results thus indicate that the colonial lifestyle was acquired in the Aplousobranchia, starting from a Ciona-like solitary ancestor and secondarily lost in Diazonidae representatives such as Rhopalaea. The solitary lifestyle of Rhopalaea is thus a derived characteristic rather than an ancestral trait.

Monitoring the magnitude of marine vessel infestation by non-indigenous ascidians in the Mediterranean

Invasive ascidians (Chordata, Tunicata) are dominant nuisance organisms. The current study investigated the role of marine vessels in their dispersal and introduction. An examination of 45 dry-docked marine vessels, comprising recreational, commercial, and military craft, in five Israeli shipyards along the Mediterranean coast, revealed non-indigenous ascidians (NIA) on every second vessel investigated. Military vessels featured the highest ascidian abundance and richness, potentially related to their maintenance routine. Niche areas on the vessels such as sea chests and the propeller exhibited the highest occurrence of ascidians. Overall, these findings provide strong evidence that marine vessels play an acute role in NIA introduction and dispersal, with military vessels and niche areas on all the vessels being more susceptible to serving as vectors. A discovery of a new introduced species during the surveys suggests that the monitoring of marine vessels can serve as an effective tool for the early detection of NIA.