The quagga mussel genome and the evolution of freshwater tolerance

Abstract

Freshwater dreissenid mussels evolved from marine ancestors during the Miocene ∼30 million years ago and today include some of the most successful and destructive invasive species of freshwater environments. Here, we sequenced the genome of the quagga mussel Dreissena rostriformis to identify adaptations involved in embryonic osmoregulation. We provide evidence that a lophotrochozoan-specific aquaporin water channel, a vacuolar ATPase subunit and a sodium/hydrogen exchanger are involved in osmoregulation throughout early cleavage, during which time large intercellular fluid-filled 'cleavage cavities' repeatedly form, coalesce and collapse, expelling excess water to the exterior. Independent expansions of aquaporins coinciding with at least five freshwater colonization events confirm their role in freshwater adaptation. Repeated aquaporin expansions and the evolution of membrane-bound fluid-filled osmoregulatory structures in diverse freshwater taxa point to a fundamental principle guiding the evolution of freshwater tolerance and provide a framework for future species control efforts.

Keywords: Dreissena; aquaporin; genome; osmoregulation; quagga.

Figure 1

The quagga mussel, D. rostriformis. (a) Quagga mussels form dense aggregations connected with strong byssal threads. Aggregates are often associated with other benthic species such as sponges. (b) Illustration of a single quagga mussel demonstrating the distinct banding pattern of the shell and the dense clump of byssus threads that enables them to adhere to both natural and manufactured substrates. (c) Global distribution of the quagga mussel highlighting native (blue) and colonized (red) habitats. (d) Condensed phylogeny of Bivalvia based on a supermatrix composed of 47 molluscan taxa covering 1,377 orthogroups. The quagga mussel is positioned amongst the Imparidentia. Species with sequenced genomes are marked with a *. All nodes possess Shimodaira–Hasegawa (SH) test support values equal to 1. Color figures are available at DNARES online.

Figure 2

Dreissenid COI phylogeny and 16S alignment. (a) The sequence in red is the COI from the genome sequenced here, sequences with ‘type’ in the name were obtained by Therriault et al. and the remaining sequences were obtained from the BoLD database. Dreissena stankovici and D. presbensis are likely to represent a single species called D. carinata (Dunker, 1853). SH test support values are indicated. (b) Multiple sequence alignment of 16S rRNA. The 16S rRNA from the genome sequenced here is named Dro 16S, sequences with ‘type’ in the name were obtained by Therriault et al. and the remaining sequences were downloaded from NCBI. The box highlights a motif identified by Therriault et al. as diagnostic for discerning bugensis (CCGG) from other D. rostriformis clades (CCAG). Color figures are available at DNARES online.

Figure 3

Embryonically expressed osmoregulatory genes. (a) Heat map of expression of candidate osmoregulatory genes highly expressed during embryogenesis prior to the free-swimming stage. Three genes (highlighted in bold) are highly expressed in Dreissena but not in similar stages of the marine oyster Cr. gigas (see Supplementary Material SM 6.2). (b) Phylogenetic tree of aquaporins with emphasis on classical lophotrochoaquaporins (red) with other classes (green—aquaglyceroporins, yellow—unorthodox, pink—EGLPs, white—undescribed annelid/brachiopod clade, light blue—aquaamoniaporins, blue—classical aquaporins) collapsed. Note the independent expansions associated with the freshwater lineages Dreissena, Corbicula, Limnoperna, the unionid mussels and the annelid leech Helobdella. * indicates a clade of long branch freshwater gastropod sequences previously annotated as malacoglyceroporins. ** indicates an expanded clade of marine brachiopod sequences. SH support values under 0.8 are not shown. (c) Peptide logo of the highly charged lophotrochoaquaporin loop D with occupancy and amino acid position indicated on the x-axis, respectively. (d) Predicted structure of the Dreissena lophotrochoaquaporin Dro.75921 loop D (magenta) wrapped to Bos taurus AQP1 (PDB: 1j4n.1) showing the predicted salt bridge formed between Arg-291 and Asp-293. Color figures are available at DNARES online.

Figure 4

Cleavage cavities in developing Dreissena embryos. (a–d) Formation of cleavage cavities during the first embryonic cleavage under ambient conditions. (e–h) First and second embryonic cleavages under high salinity demonstrating the lack of cleavage cavity formation. (i–l) Formation of cleavage cavities during the first embryonic cleavage under low salinity. (m–p) Failed first cleavage leading to rupture of the fertilization envelope and the extrusion of an amoeboid projection with a large vacuole. * indicates cleavage cavities, N indicates nuclei, v indicates intracellular vacuoles and arrows indicate fertilization envelope ruptures. Color figures are available at DNARES online.