Although dense animal communities at hydrothermal vents and cold seeps rely on symbioses with chemoautotrophic bacteria [1, 2], knowledge of the mechanisms underlying these chemosynthetic symbioses is still fragmentary because of the difficulty in culturing the symbionts and the hosts in the laboratory. Deep-sea Calyptogena clams harbor thioautotrophic bacterial symbionts in their gill epithelial cells [1, 2]. They have vestigial digestive tracts and nutritionally depend on their symbionts [3], which are vertically transmitted via eggs [4]. To clarify the symbionts' metabolic roles in the symbiosis and adaptations to intracellular conditions, we present the complete genome sequence of the symbiont of Calyptogena okutanii. The genome is a circular chromosome of 1,022,154 bp with 31.6% guanine + cytosine (G + C) content, and is the smallest reported genome in autotrophic bacteria. It encodes 939 protein-coding genes, including those for thioautotrophy and for the syntheses of almost all amino acids and various cofactors. However, transporters for these substances to the host cell are apparently absent. Genes that are unnecessary for an intracellular lifestyle, as well as some essential genes (e.g., ftsZ for cytokinesis), appear to have been lost from the symbiont genome. Reductive evolution of the genome might be ongoing in the vertically transmitted Calyptogena symbionts.
The genus Symbiodinium is the commonly observed symbiotic dinoflagellate (zooxanthellae) that forms mutual associations with various marine invertebrates. Numerous studies have revealed that the genus is comprised of a group of diverse taxa, and information on the phylogenetic relationships among the genus’ members is increasing. In this study, small subunit (SSU) ribosomal RNA (ssrRNA) gene sequences were determined for 15 more Symbiodinium strains from 12 relatively unstudied host taxa (Indo‐Pacific tridacnids, cardiids, sponge, and soft coral), 1 hitherto unreported free‐living Symbiodinium strain, and 4 other Symbiodinium strains from four other host taxa (Indo‐Pacific zoanthid, foraminifer, jellyfish, and mid‐Pacific hard coral). Their respective phylogenetic positions were inferred, and strains that are either closely related to or distinct from previously reported Symbiodinium taxa were revealed. The cultured Symbiodinium strains isolated from individuals of six species of tridacnids and three species of cardiids all had identical ssrRNA gene sequences, are closely related to S. microadriaticum Freudenthal, and are indistinguishable from the RFLP Type A strain previously reported. However, the ssrRNA gene sequences of clam symbionts that were obtained via gene cloning were different from those of the cultured isolates and represent strains that are close to the RFLP Type C strains. The Symbiodinium‐like dinoflagellate from the Indo‐Pacific sponge Haliclona koremella De Laubenfels is distinct from any of the Symbiodinium taxa studied and may be similar to the symbiont previously isolated from the stony coral Montipora patula Quelch. The isolates from the soft coral Sarcophyton glaucum Quoy et Gaimard and from the zoanthid Zoanthus sp. are both very closely related to S. pilosum Trench et Blank. The free‐living Symbiodinium isolate is very closely related to the symbiont isolated from the Indo‐Pacific foraminifer Amphisorus hemprichii Ehrenberg, which in turn is distinct from the Red Sea strain isolated from a similar host. Theisolate from Cassiopeia sp. is different from S. microadriaticum F., the type species harbored by Cassiopeia xamachana Bigelow, and is instead very closely related to S. pulchrorum Trench isolated from a sea anemone. The symbiont from the stony coral M. verrucosa Lamarck is a sister taxon to the symbionts isolated from the foraminifera Marginopora kudakajimensis Gudmundsson and Sorites orbiculus Forskål. These data suggest that polymorphic symbioses extend from cnidarians to some bivalve, foraminifer, and jellyfish host species.
Components of the proteinaceous cement secreted by barnacles have yet to be studied because of their insolubility. We solubilized and characterized the proteins of secondary cement, which is produced when the barnacle is detached from the substratum, in Megabalanus rosa. The cement was fractionated, according to its solubility in aqueous formic acid, into a soluble fraction, SF1 (21%); a fraction soluble after reduction, SF2 (37%); and a fraction insoluble after reduction, IF (42%). Analysis of the SF1 and SF2 by sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed that they contained three polypeptides (SF1-60 k, -57 k, -47 k) and one polypeptide (SF2-60 k), respectively. The amino acid compositions of these polypeptides were similar and their N-terminal amino acid sequences were identical. These polypeptides had an unusual amino acid composition, rich in Ser, Thr, Ala, and Gly, like the tube cement of a marine polychaete, Phragmatopoma californica. The IF, solubilized in aqueous formic acid after cleavage with cyanogen bromide, was shown by SDS-PAGE to contain eight fragment peptides (CB-peptides). N-terminal amino acid sequences of the CB-peptides were also determined. We conclude that the barnacle cement is composed of at least two types of protein: highly hydroxylated protein in the SF1 and SF2 and insoluble protein in the IF. The SDS-PAGE pattern of CB-peptides from the secondary cement was identical to that of the primary cement produced while the barnacle is attached to a substratum. In addition, immunoblot analysis, using a polyclonal antibody against one of the CB-peptides from the secondary cement, also cross-reacted with a CNBr-fragment peptide of the primary cement. These results indicate that the primary and secondary cements are similar in protein composition.
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