Discovery of chemoautotrophic symbiosis in the giant shipworm Kuphus polythalamia (Bivalvia: Teredinidae) extends wooden-steps theory

Abstract

The "wooden-steps" hypothesis [Distel DL, et al. (2000) Nature 403:725-726] proposed that large chemosynthetic mussels found at deep-sea hydrothermal vents descend from much smaller species associated with sunken wood and other organic deposits, and that the endosymbionts of these progenitors made use of hydrogen sulfide from biogenic sources (e.g., decaying wood) rather than from vent fluids. Here, we show that wood has served not only as a stepping stone between habitats but also as a bridge between heterotrophic and chemoautotrophic symbiosis for the giant mud-boring bivalve Kuphus polythalamia This rare and enigmatic species, which achieves the greatest length of any extant bivalve, is the only described member of the wood-boring bivalve family Teredinidae (shipworms) that burrows in marine sediments rather than wood. We show that K. polythalamia harbors sulfur-oxidizing chemoautotrophic (thioautotrophic) bacteria instead of the cellulolytic symbionts that allow other shipworm species to consume wood as food. The characteristics of its symbionts, its phylogenetic position within Teredinidae, the reduction of its digestive system by comparison with other family members, and the loss of morphological features associated with wood digestion indicate that K. polythalamia is a chemoautotrophic bivalve descended from wood-feeding (xylotrophic) ancestors. This is an example in which a chemoautotrophic endosymbiosis arose by displacement of an ancestral heterotrophic symbiosis and a report of pure culture of a thioautotrophic endosymbiont.

Keywords: Teredinidae; chemoautotrophy; shipworm; symbiosis; thioautotrophy.

Fig. 1.

Comparative anatomy and life position of Kuphus polythalamia and Lyrodus pedicellatus. (A) Fresh specimen of K. polythalamia (PMS-1672Y) removed from its calcareous tube, (B) calcareous tube of K. polythalamia (PMS-1674K) removed from sediment, (C) diagram depicting the anatomy and life position of K. polythalamia in sediment, and (D) Inset from box in C depicting the anatomy and life position of the wood-feeding shipworm Lyrodus pedicellatus in wood. (Scale bars: A–C, 5.0 cm; D, 0.5 cm.) b, bacteria; c, cecum; g, gill; HS⁻, hydrogen sulfide; m, mouth; p, pallet; s, siphon; t, calcareous tube; v, valve (shell); vm, visceral mass. Movie S1 shows a specimen of K. polythalamia being removed from its tube and dissected.

Fig. 2.

Endosymbiont ultrastructure and localization. (A) Scanning electron micrograph of freeze-fractured gill; (B) Inset from A; (C) Inset from B; (D) Inset from C; (E) transmission electron micrograph (TEM) of endosymbionts in gill; (F) TEM of isolate 2141T. cs, carboxysomes; icm, intracytoplasmic membranes; im, inner membrane; om, outer membrane; s, symbionts; sg, sulfur globules; vm, vacuole membrane. (Scale bars: A, 500 µm; B, 100 µm; C, 10 µm; D, 2 µm; E and F, 0.5 µm.) (G–J) FISH images showing (G) bacterial probe Eub338 (Cy5, red); (H) symbiont probe Kp1 (Cy3, green); (I) nucleic acid stain (SYTOX, blue); and (J) merged images G–I. (Scale bars: G–J, 10 µm.) Negative controls are in SI Appendix, Fig. S1.

Fig. 3.

Endosymbiont and host phylogeny. (A) Bayesian inference subtree for symbionts of K. polythamia and closely related bacteria (excerpted from SI Appendix, Fig. S2) based on partial 16S rRNA sequences (*uncultivated environmental sample). (B) Bayesian inference tree for the bivalve family Teredinidae and related families within the order Myida based on partial 18S and 28S rRNA sequences, excerpted from previously published work (22). Reproduced from ref. , with permission from Elsevier. Numbers at nodes indicate posterior probabilities. Scale bars denote nucleotide substitutions per site. Arrow in B indicates proposed acquisition of xylotrophic symbionts.

Fig. 4.

Endosymbiont metagenome and isolate 2141T genome. (A) Metagenome read clusters plotted as a function of GC content, coverage, and read counts (2141T-like clusters are shown in red). Note that the majority of reads fall into two clusters with similar read counts, GC content, and coverage. (B) BLASTn alignment of the 2141T-like bacterial sequences from metagenomes of two individual specimens of K. polythalamia, specimen 2132W, sample 2249p (green), and specimen 2133X, sample 2110w (blue) to the genome of strain 2141T (purple). Color shades correspond to the identity range of 80–100% compared with the reference genome (2141T). The GC content of the genome of strain 2141T is plotted in black. See SI Appendix, Table S3 for additional specimen, sample, and metagenome data.