Identification of sequestered chloroplasts in photosynthetic and non-photosynthetic sacoglossan sea slugs (Mollusca, Gastropoda).

BACKGROUND: Sacoglossan sea slugs are well known for their unique ability among metazoans to incorporate functional chloroplasts (kleptoplasty) in digestive glandular cells, enabling the slugs to use these as energy source when starved for weeks and months. However, members assigned to the shelled Oxynoacea and Limapontioidea (often with dorsal processes) are in general not able to keep the incorporated chloroplasts functional. Since obviously no algal genes are present within three (out of six known) species with chloroplast retention of several months, other factors enabling functional kleptoplasty have to be considered. Certainly, the origin of the chloroplasts is important, however, food source of most of the about 300 described species is not known so far. Therefore, a deduction of specific algal food source as a factor to perform functional kleptoplasty was still missing.

RESULTS: We investigated the food sources of 26 sacoglossan species, freshly collected from the field, by applying the chloroplast marker genes tufA and rbcL and compared our results with literature data of species known for their retention capability. For the majority of the investigated species, especially for the genus Thuridilla, we were able to identify food sources for the first time. Furthermore, published data based on feeding observations were confirmed and enlarged by the molecular methods. We also found that certain chloroplasts are most likely essential for establishing functional kleptoplasty.

CONCLUSIONS: Applying DNA-Barcoding appeared to be very efficient and allowed a detailed insight into sacoglossan food sources. We favor rbcL for future analyses, but tufA might be used additionally in ambiguous cases. We narrowed down the algal species that seem to be essential for long-term-functional photosynthesis: Halimeda, Caulerpa, Penicillus, Avrainvillea, Acetabularia and Vaucheria. None of these were found in Thuridilla, the only plakobranchoidean genus without long-term retention forms. The chloroplast type, however, does not solely determine functional kleptoplasty; members of no-retention genera, such as Cylindrobulla or Volvatella, feed on the same algae as e.g., the long-term-retention forms Plakobranchus ocellatus or Elysia crispata, respectively. Evolutionary benefits of functional kleptoplasty are still questionable, since a polyphagous life style would render slugs more independent of specific food sources and their abundance.