Widespread Occurrence of Two Carbon Fixation Pathways in Tubeworm Endosymbionts: Lessons from Hydrothermal Vent Associated Tubeworms from the Mediterranean Sea

Abstract

Vestimentiferan tubeworms (siboglinid polychetes) of the genus Lamellibrachia are common members of cold seep faunal communities and have also been found at sedimented hydrothermal vent sites in the Pacific. As they lack a digestive system, they are nourished by chemoautotrophic bacterial endosymbionts growing in a specialized tissue called the trophosome. Here we present the results of investigations of tubeworms and endosymbionts from a shallow hydrothermal vent field in the Western Mediterranean Sea. The tubeworms, which are the first reported vent-associated tubeworms outside the Pacific, are identified as Lamellibrachia anaximandri using mitochondrial ribosomal and cytochrome oxidase I (COI) gene sequences. They harbor a single gammaproteobacterial endosymbiont. Carbon isotopic data, as well as the analysis of genes involved in carbon and sulfur metabolism indicate a sulfide-oxidizing chemoautotrophic endosymbiont. The detection of a hydrogenase gene fragment suggests the potential for hydrogen oxidation as alternative energy source. Surprisingly, the endosymbiont harbors genes for two different carbon fixation pathways, the Calvin-Benson-Bassham (CBB) cycle as well as the reductive tricarboxylic acid (rTCA) cycle, as has been reported for the endosymbiont of the vent tubeworm Riftia pachyptila. In addition to RubisCO genes we detected ATP citrate lyase (ACL – the key enzyme of the rTCA cycle) type II gene sequences using newly designed primer sets. Comparative investigations with additional tubeworm species (Lamellibrachia luymesi, Lamellibrachia sp. 1, Lamellibrachia sp. 2, Escarpia laminata, Seepiophila jonesi) from multiple cold seep sites in the Gulf of Mexico revealed the presence of acl genes in these species as well. Thus, our study suggests that the presence of two different carbon fixation pathways, the CBB cycle and the rTCA cycle, is not restricted to the Riftia endosymbiont, but rather might be common in vestimentiferan tubeworm endosymbionts, regardless of the habitat.

Dual symbiosis with co‐occurring sulfur‐oxidizing symbionts in vestimentiferan tubeworms from a Mediterranean hydrothermal vent

“…Phylogenetic analyses resolved seven highly supported vestimentiferan symbiont clusters: six clusters that contained LES symbiont sequences, and one cluster that only contained sequences from hydrothermal vent habitats (Figs and B). Our analyses therefore confirmed the previously described ‘hydrothermal vent’ symbiont cluster, three ‘cold seep’ symbiont clusters (Cl‐1, 3 and 4) and a Mediterranean symbiont cluster (Cl‐2) that was recently identified by Thiel and colleagues (). Furthermore, we detected at least two additional symbiont clusters (Cl‐5 and Cl‐6) in our analyses, and these exclusively contained symbionts from Lamellibrachia species (Fig.…”

“…Until now, vestimentiferan Tws were widely believed to harbour a single sulfur‐oxidizing symbiont phylotype in their trophosome (reviewed by Bright and Lallier, ). Although a few molecular studies indicated the possibility of multiple symbionts in some vestimentiferan species, conclusive evidence was arguably still missing (Naganuma et al ., 1997; 2005; Elsaied et al ., ; Kimura et al ., ; Chao et al ., ; Duperron et al ., ; Thiel et al ., ). In all of our CARD‐FISH analyses, specific probes matching the two phylotypes found in our 16S rRNA clone libraries bound separate symbiont cells, providing evidence that these phylotypes represent two distinct coexisting symbionts.…”

“…The second group, which includes the genera Lamellibrachia , Escarpia , Paraescarpia and Seepiophila , is found mainly at cold seeps around the world. However, exceptions exist, as some ‘cold seep’ species such as Escarpia spicata , Lamellibrachia barhami , Lamellibrachia satsuma and Lamellibrachia anaximandri have been found at both vents and seeps (reviewed by Bright and Lallier, ; Thiel et al ., ). In addition, E. spicata has been described from a whale fall, and some Lamellibrachia Tws were found colonizing shipwrecks (Dando et al ., ; Feldman et al ., ; Hughes and Crawford, ; Gambi et al ., ).…”

“…In addition, Thiel and colleagues recently reported two different 16S rRNA phylotypes in clone libraries of E. laminata and Lamellibrachia sp. 2 from the Gulf of Mexico (Thiel et al ., ). The presence of more than one symbiont type co‐occurring in vestimentiferan Tws has therefore been hypothesized on a number of occasions, but has not yet been conclusively shown.…”

A self-potential investigation of submarine massive sulfides: Palinuro Seamount, Tyrrhenian Sea

“…Diffuse seepage of low-temperature hydrothermal fluids has been detected at Palinuro (Lupton et al, 2011;Thiel et al, 2012), but no active chimneys have been discovered.…”