Surfing the vegetal pole in a small population: extracellular vertical transmission of an 'intracellular' deep-sea clam symbiont

Abstract

Symbiont transmission is a key event for understanding the processes underlying symbiotic associations and their evolution. However, our understanding of the mechanisms of symbiont transmission remains still fragmentary. The deep-sea clam Calyptogena okutanii harbours obligate sulfur-oxidizing intracellular symbiotic bacteria in the gill epithelial cells. In this study, we determined the localization of their symbiont associating with the spawned eggs, and the population size of the symbiont transmitted via the eggs. We show that the symbionts are located on the outer surface of the egg plasma membrane at the vegetal pole, and that each egg carries approximately 400 symbiont cells, each of which contains close to 10 genomic copies. The very small population size of the symbiont transmitted via the eggs might narrow the bottleneck and increase genetic drift, while polyploidy and its transient extracellular lifestyle might slow the rate of genome reduction. Additionally, the extracellular localization of the symbiont on the egg surface may increase the chance of symbiont exchange. This new type of extracellular transovarial transmission provides insights into complex interactions between the host and symbiont, development of both host and symbiont, as well as the population dynamics underlying genetic drift and genome evolution in microorganisms.

Effects of a long-term rearing system for deep-sea vesicomyid clams on host survival and endosymbiont retention

“…In the gill bacteriocytes of freshly collected clams, we detected numerous symbiont cells in which electrontransparent vesicles were frequently observed (81% of the 63 symbiont cells counted), as previously noted [4] (Fig. 3a).…”

“…However, the symbiont of P. okutanii was recently found to exhibit polyploidy [4]. Because the genomic copy number of the gill symbionts of P. okutanii under starvation conditions is unknown, quantification of rRNA genes via qPCR may not necessarily lead to reliable results regarding symbiont abundance.…”

“…For gill tissue sections, the fixed gills were prepared as described previously [4]. Briefly, pieces of the gills were post-fixed with 2.0% osmium tetroxide dissolved in filtered artificial seawater (FASW) for 2 h at 4 °C.…”

“…These species harbor sulfur-oxidizing intracellular symbiotic bacteria (Gram-negative gammaproteobacteria) in their gill epithelial cells [2], and the clams depend on the symbionts for nutrition [3]. Recently, the robust and unique associations between Phreagena clams and their symbionts were revealed through genome sequencing of symbionts [2] and by studying the transmission mode of the symbionts [4].…”

“…We sampled the clams from the three tanks at certain intervals, and observed the abundance of the symbiotic bacteria in the gill epithelia by in situ hybridization that targeted symbiont 16S rRNA. Furthermore, we observed the sulfur globules-assumed to be sites of elemental sulfur storage [4,7,8]-in the symbiont cells and investigated the expression of one of the key enzymes potentially responsible for sulfur metabolism in the symbiont. After presenting the results below, we discuss the issues that need to be addressed to achieve more stable rearing of Phreagena clams.…”