The Phenomenon of Functional Retention of Incorporated Chloroplasts in Sea Slugs (Sacoglossa, Heterobranchia, Mollusca) and Evolutionary Adaptations

Abstract

The phenomenon of retention of functional chloroplasts from their food algae in sacoglossan sea slugs - also called kleptoplasty - is a challenge for research, still leaving many questions unsolved, e. g. concerning differences between various sacoglossan species in their capacities of retention of functional chloroplasts. To investigate these differences and potential influencing factors in more detail, this thesis combined photobiological investigations with cell biological, ecological and behavioral analyses. Photosynthetic activity, i. e. the ongoing functioning of incorporated chloroplasts within the slug cells, was analyzed with a Pulse Amplitude Modulated Fluorometer (PAM). The analyses included several chloroplast-incorporating sea slug species, especially most of the few which are known as “top-performers” of long-term functional retention of chloroplasts: <em>Elysia timida, Elysia crispata (mangrove type </em>and<em> reef type), Elysia viridis </em>and<em> Plakobranchus ocellatus</em>. As further comparative species, <em>Bosellia mimetica, Thuridilla hopei </em>and<em> Placida dendritica</em> were included. Overall, relevant differences between species-specific spectra of photosynthetic capacities in various sacoglossan species could be confirmed and also considerable variation within. For <em>P. ocellatus</em>, PAM measurements of functional chloroplast retention for over seven months were reported, the longest time period documented up to now. Feeding experiments indicated that in addition to its known food algae <em>Codium fragile</em>, <em>E. viridis</em> also fed on and could incorporate chloroplasts from <em>Flabellia petiolata</em>, with even partly better capacities of chloroplast retention than with <em>C. fragile/vermilara</em>. Photobehavioral analyses, testing a former hypothesis that chloroplast retention potentially implies stronger phototactic behavior in sea slugs, in which the non-sacoglossan sea slug species <em>Cratena peregrina </em>and<em> Flabellina affinis</em> were additionally included as comparison without incorporation of chloroplasts, indicated a different coherence of photobehavior, as some sea slug species without chloroplasts or with rather fast digestion of chloroplasts reacted stronger positively phototactic than species with long-term kleptoplast retention. For <em>E. timida</em>, nevertheless a positive phototactic behavior could be observed, which might be connected to special adaptations in this species. With PAM-measurements could be demonstrated e. g. the efficiency to regulate fluorescence F emission of incorporated chloroplasts by varying opening and closing positions of the parapodial lobes in <em>E. timida</em>.<br /> Furthermore, effects of temperature on capacities of long-term photosynthetic activity were indicated by experimental trials under controlled laboratory conditions. The advantages of the laboratory culture system with <em>E. timida</em> as a model organism that could be successfully established could be revealed. In trials within the laboratory culture system, the capacity of <em>E. timida</em> to acquire kleptoplasts from another chloroplast donor – the alga <em>Acetabularia peniculus</em> – with similar retention capacities compared to their food alga <em>Acetabularia acetabulum</em>, could be demonstrated. On a cell biological level, indices for factors concerning special adaptations in relation to incorporation of chloroplasts were elucidated with transmission electron microscopy (TEM). The very first uptake of chloroplasts from the food alga <em>A. acetabulum</em> in juvenile <em>E. timida</em> could be illustrated with TEM. Furthermore, ecological parameters in the natural environment, especially concerning light conditions, could be demonstrated to affect photosynthetic activity of incorporated chloroplasts, which constituted the first demonstration of this kind. Two sea slug species, <em>E. timida </em>and<em> E. crispata mangrove type</em>, were investigated underwater with a Diving PAM Fluorometer in their natural habitat in France and in Florida, respectively, concerning kleptoplast photosynthetic activity and combined environmental and behavioral parameters. These represent to the current knowledge the first photosynthetic measurements of incorporated chloroplasts in sacoglossan sea slugs in their natural environment published so far.