Ultrastructural, Immunohistochemical, and Biochemical Variation Amongst Symbiodinium Isolates
Mazzillo Mays, Maria
Type of Degreedissertation
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Symbiodinium spp. reside intracellularly in a complex symbiosome (host and symbiont-derived) in a specific host-symbiont associations. Symbiodinium is a diverse genus with variation equivalent to that of other dinoflagellate orders. First, this investigation examines antigenic variation in the algal mucilage secretions at the host-symbiont interface. Cultured Symbiodinium from a variety of clades were labeled with one or the other of two antibodies that bind to components of the symbiont mucilage (PC3 antibody, developed using a clade B alga cultured from Aiptasia pallida; BF10 antibody, developed using a clade F alga cultured from Briareum sp.). PC3 antigen was found in cultured Symbiodinium from clades A and B, but not clades C, D, E and F. The correlation between labeling and clade may account for some of the specificity between host and symbiont in the field. Within clades A and B there was variation in the amount of label present. BF10 antigen was more specific and only found in cultures of the same cp23S-rDNA strain the antibody was created against. Since the mucilage forms the host-symbiont interface, variation in its molecular composition is likely to be the source of signals involved in recognition and specificity. Next, I examined variation in the ultrastructural morphology amongst 5 Symbiodinium isolates (2 from clade A, and 1 from clade B, 1 from clade C, and a free-living isolate from clade E) to create a baseline standard for future ultrastructural work and to determine if there is any ultrastructural basis for taxonomic description of species within this genus. Ultrastructural examinations were performed using High Pressure Rapid Freezing. All 5 isolates studied exhibited the standard dinokaryon nucleus, a peripheral multi-lobed chloroplastwith a single stalked pyrenoid surrounded by a starch cap, a dinoflagellate-type mitochondria with tubular cristae, and various other cellular inclusions. Additional small electron dense inclusions were typically positioned near the cell periphery and may be related to mucocysts sometimes seen in dinoflagellates. Some isolates (CCMP421 and Y109) had a few cells that exhibited large electron dense structures containing calcium oxalate crystals. Cell size amongst different isolates was significantly different and consistent with previous studies on the same or similar isolates. Examination of the ultrastructure of 5 isolates of Symbiodinium revealed that there is very little variation in morphological characters. Therefore, it is unlikely that isolates or species of Symbiodinium can be usefully differentiated on the basis of morphology alone. Finally, in order to continue examining Symbiodinium isolates for variation that may impact the association with a specific host, I investigated the variation of storage products (lipid droplets and starch granules) within 5 isolates. Amongst the 5 isolates examined, the two isolates from clade A contained the least amount of both starch and lipid. However, another small isolate from clade B contained some of the largest amounts of both starch and lipid. A clade F isolate also contained many storage products while the only free-living isolate examined, a clade E isolate, contained more lipid and starch than the clade A symbionts but less than the clade B and F isolates. This variation in storage products amongst isolates grown under the same culture regime reveals physiological differences that may relate to an individual isolate's suitability as a symbiotic partner under certain environmental conditions such as those arising from global climate change. Further biochemical examination of excreted exudate for the presence of carbohydrate and protein components found that in all 5 isolates, more carbohydrate compounds were secreted into the culture medium than protein. Examination of the composition of PC3 antigen, a component of the exudate of some Symbiodinium isolates, found that the antigen iscomposed, at least in part, of carbohydrate structures. The presence of carbohydrate components in the exudate compounds that are associated with the cell surface is consistent with the hypothesis that initiation and perhaps maintenance of the host-symbiont relationship is mediated by a glycan/lectin interaction.