Genome of the lepidopleurid chiton Hanleya hanleyi (Mollusca, Polyplacophora).

Mollusca is the second most species-rich phylum and includes animals as disparate as octopuses, clams, and chitons. Dozens of molluscan genomes are available, but only one representative of the subphylum Aculifera, the sister taxon to all other molluscs, has been sequenced to date, hindering comparative and evolutionary studies. To facilitate evolutionary studies across Mollusca, we sequenced the genome of a second aculiferan mollusc, the lepidopleurid chiton Hanleya hanleyi (Bean 1844), using a hybrid approach combining Oxford Nanopore and Illumina reads. After purging redundant haplotigs and removing contamination from this 1.3% heterozygous genome, we produced a 2.5 Gbp haploid assembly (>4X the size of the other chiton genome sequenced to date) with an N50 of 65.0 Kbp. Despite a fragmented assembly, the genome is rather complete (92.0% of BUSCOs detected; 79.4% complete plus 12.6% fragmented). Remarkably, the genome has the highest repeat content of any molluscan genome reported to date (>66%). Our gene annotation pipeline predicted 69,284 gene models (92.9% of BUSCOs detected; 81.8% complete plus 11.1% fragmented) of which 35,362 were supported by transcriptome and/or protein evidence. Phylogenomic analysis recovered Polyplacophora sister to all other sampled molluscs with maximal support. The Hanleya genome will be a valuable resource for studies of molluscan biology with diverse potential applications ranging from evolutionary and comparative genomics to molecular ecology.

Phylogenomics of Aplacophora (Mollusca, Aculifera) and a solenogaster without a foot.

Recent molecular phylogenetic investigations strongly supported the placement of the shell-less, worm-shaped aplacophoran molluscs (Solenogastres and Caudofoveata) and chitons (Polyplacophora) in a clade called Aculifera, which is the sister taxon of all other molluscs. Thus, understanding the evolutionary history of aculiferan molluscs is important for understanding early molluscan evolution. In particular, fundamental questions about evolutionary relationships within Aplacophora have long been unanswered. Here, we supplemented the paucity of available data with transcriptomes from 25 aculiferans and conducted phylogenomic analyses on datasets with up to 525 genes and 75 914 amino acid positions. Our results indicate that aplacophoran taxonomy requires revision as several traditionally recognized groups are non-monophyletic. Most notably, Cavibelonia, the solenogaster taxon defined by hollow sclerites, is polyphyletic, suggesting parallel evolution of hollow sclerites in multiple lineages. Moreover, we describe Apodomenia enigmatica sp. nov. , a bizarre new species that appears to be a morphological intermediate between Solenogastres and Caudofoveata. This animal is not a missing link, however; molecular and morphological studies show that it is a derived solenogaster that lacks a foot, mantle cavity and radula. Taken together, these results shed light on the evolutionary history of Aplacophora and reveal a surprising degree of morphological plasticity within the group.

Molecular phylogeny of Caudofoveata (Mollusca) challenges traditional views.

The shell-less, worm-shaped Caudofoveata (=Chaetodermomorpha) is one of the least known groups of molluscs. The taxon consists of 141 recognized species found from intertidal environments to the deep-sea where they live burrowing in sediment. Evolutionary relationships of the group have been debated, but few studies based on morphological or molecular data have investigated the phylogeny of the group. Here we use molecular phylogenetics to resolve relationships among and within families of Caudofoveata. Phylogenetic analyses were performed using selected mitochondrial and nuclear genes from species from all recognized families of Caudofoveata. In resulting trees and contrary to traditional views, Prochaetodermatidae forms the sister clade to a clade containing the other two currently recognized families, Chaetodermatidae and Limifossoridae. The monophyly of Prochaetodermatidae is highly supported, but Limifossoridae and Chaetodermatidae are not recovered as monophyletic. Most of the caudofoveate genera are also not recovered as monophyletic in our analyses. Thus results from our molecular data suggest that the current classification of Caudofoveata is in need of revision, and indicate evolutionary scenarios that differ from previously proposed hypotheses based on morphology.

Mitogenomics reveals phylogenetic relationships of caudofoveate aplacophoran molluscs.

The worm-shaped, shell-less aplacophoran molluscs Caudofoveata and Solenogastres have recently received attention as part of the clade Aculifera, but relationships within these two lineages are still largely unknown. Here, we use complete mitochondrial genomes to shed light on higher-level relationships within Caudofoveata. Mitochondrial genomes have been sequenced for many diverse molluscs, but only two mitochondrial genomes from aplacophoran molluscs (the caudofoveates Scutopus ventrolineatus and Chaetoderma nitidulum) are available to date. We sequenced and assembled complete or near complete mitochondrial genomes of five additional species of Caudofoveata (Falcidens acutargatus, Falcidens halanychi, Scutopus robustus, Psilodens balduri and Spathoderma clenchi) and one species of Solenogastres (Neomenia carinata) for comparison to available mitochondrial genomes of aculiferans. Comparison of mitochondrial gene order among different lineages revealed a highly conserved order of protein coding genes corresponding to the hypothesized ancestral gene order for Mollusca. Unique arrangements of tRNAs were found among lineages of aculiferan molluscs as well as among caudofoveate taxa. Phylogenetic analyses of amino acid sequences for the 13 protein-coding genes recovered a monophyletic Aplacophora. Within Caudofoveata, Chaetodermatidae, but not Limifossoridae, was recovered monophyletic. Taken together, our results suggest that mitochondrial genomes can serve as useful molecular markers for aculiferan phylogenetics, especially for more recent phylogenetic events.

Fast evolving 18S rRNA sequences from Solenogastres (Mollusca) resist standard PCR amplification and give new insights into mollusk substitution rate heterogeneity.

BACKGROUND: The 18S rRNA gene is one of the most important molecular markers, used in diverse applications such as molecular phylogenetic analyses and biodiversity screening. The Mollusca is the second largest phylum within the animal kingdom and mollusks show an outstanding high diversity in body plans and ecological adaptations. Although an enormous amount of 18S data is available for higher mollusks, data on some early branching lineages are still limited. Despite of some partial success in obtaining these data from Solenogastres, by some regarded to be the most "basal" mollusks, this taxon still remained problematic due to contamination with food organisms and general amplification difficulties. RESULTS: We report here the first authentic 18S genes of three Solenogastres species (Mollusca), each possessing a unique sequence composition with regions conspicuously rich in guanine and cytosine. For these GC-rich regions we calculated strong secondary structures. The observed high intra-molecular forces hamper standard amplification and appear to increase formation of chimerical sequences caused by contaminating foreign DNAs from potential prey organisms. In our analyses, contamination was avoided by using RNA as a template. Indication for contamination of previously published Solenogastres sequences is presented. Detailed phylogenetic analyses were conducted using RNA specific models that account for compensatory substitutions in stem regions. CONCLUSIONS: The extreme morphological diversity of mollusks is mirrored in the molecular 18S data and shows elevated substitution rates mainly in three higher taxa: true limpets (Patellogastropoda), Cephalopoda and Solenogastres. Our phylogenetic tree based on 123 species, including representatives of all mollusk classes, shows limited resolution at the class level but illustrates the pitfalls of artificial groupings formed due to shared biased sequence composition.