ena-PROJECT-LaternulaKGI-24-04-2012-13:20:41:206-3
The immune transcriptome of the bivalve Laternula elliptica from the Antarctic Peninsula
The ecophysiological and molecular response to environmental stressors resulting from melting and calving of tide water glaciers in Western Antarctica was investigated in the bivalve Laternula elliptica, which is a key species in Antarctic coastal ecosystems. The transcriptomal data set of L. elliptica was generated using 454 technology (454 Life Sciences, Branford, CT, USA). To maximize the number of expressed transcripts and facilitate the identification of candidate genes involved in injury, immune system and general stress response, samples from two age groups and different stress treatments were used for transcriptome generation. Animals were classified as “large/older” with 7.2 ± 0.5 cm and “small/younger” with 4.8 ± 0.5 cm shell length. RNA was extracted separately from samples of digestive gland, gill and siphon (one small and one large animal respectively from the treatments “starvation/ acute injury” and “food/ acute injury”) as well as from hemocyte cells (4 small animals from treatment “starvation”, 5 small from “food”, 4 large from “starvation/ acute injury” and 4 small as well as 4 large from “food/ acute injury”).  Briefly, Laternula elliptica individuals were collected by divers in Potter Cove, King George Island, Antarctic Peninsula (62°14´S, 58°40´W), between November 2008 and February 2009 at 7-15 m depth. Animals were kept in tanks for 10 days in aerated, natural seawater without sediment at 1°C. Following this acclimation, individuals from both size/age groups were exposed to different stress treatments (starvation and injury). Half of the animals of each size group were starved by keeping the animals in filtered seawater (0.2 µm final pore size, “starvation”). The other half was kept under feeding conditions in unfiltered seawater which was additionally enriched with particulate nutrients (treatment “food”). The mixed diet consisted of live microalgae as well as “artificial detritus” i.e. freeze dried and ground macroalgae (Ascoseira mirabilis) and krill and red bloodworms obtained from Tetra (Tetra Delica; Melle, Germany). Microalgae were fished in Potter Cove and cultured in 2L-bottles at 1°C. Animals were fed every second day. After 21days 50% of the small and large animals from the starvation and food treatment were injured (valves of the animals were cracked in such a way that also the mantle was injured and the siphon was cut at two places). Two days after the injury event i.e. at day 23 after the initial start of the different nutrition treatments, hemocyte cells and tissues were sampled. Hemolymph fluid containing the hemocyte cells of each individual was collected. The hemocytes pellet and the different tissues were shock-frozen in liquid nitrogen. For transcriptome generation, an additional set of sequence data, generated from mantle tissue of  L. elliptica collected at Rothera Research Station, Adelaide Island (range of shell size between 50.1-83.5mm), was provided by M. Clark of the British Antarctic Survey (BAS; for details see Clark et al., 2010. Insights into shell deposition in the Antarctic bivalve Laternula elliptica: gene discovery in the mantle transcriptome using 454 pyrosequencing. Bmc Genomics 11).
The ecophysiological and molecular response to environmental stressors resulting from melting and calving of tide water glaciers in Western Antarctica was investigated in the bivalve Laternula elliptica, which is a key species in Antarctic coastal ecosystems. The transcriptomal data set of L. elliptica was generated using 454 technology (454 Life Sciences, Branford, CT, USA). To maximize the number of expressed transcripts and facilitate the identification of candidate genes involved in injury, immune system and general stress response, samples from two age groups and different stress treatments were used for transcriptome generation. Animals were classified as “large/older” with 7.2 ± 0.5 cm and “small/younger” with 4.8 ± 0.5 cm shell length. RNA was extracted separately from samples of digestive gland, gill and siphon (one small and one large animal respectively from the treatments “starvation/ acute injury” and “food/ acute injury”) as well as from hemocyte cells (4 small animals from treatment “starvation”, 5 small from “food”, 4 large from “starvation/ acute injury” and 4 small as well as 4 large from “food/ acute injury”).  Briefly, Laternula elliptica individuals were collected by divers in Potter Cove, King George Island, Antarctic Peninsula (62°14´S, 58°40´W), between November 2008 and February 2009 at 7-15 m depth. Animals were kept in tanks for 10 days in aerated, natural seawater without sediment at 1°C. Following this acclimation, individuals from both size/age groups were exposed to different stress treatments (starvation and injury). Half of the animals of each size group were starved by keeping the animals in filtered seawater (0.2 µm final pore size, “starvation”). The other half was kept under feeding conditions in unfiltered seawater which was additionally enriched with particulate nutrients (treatment “food”). The mixed diet consisted of live microalgae as well as “artificial detritus” i.e. freeze dried and ground macroalgae (Ascoseira mirabilis) and krill and red bloodworms obtained from Tetra (Tetra Delica; Melle, Germany). Microalgae were fished in Potter Cove and cultured in 2L-bottles at 1°C. Animals were fed every second day. After 21days 50% of the small and large animals from the starvation and food treatment were injured (valves of the animals were cracked in such a way that also the mantle was injured and the siphon was cut at two places). Two days after the injury event i.e. at day 23 after the initial start of the different nutrition treatments, hemocyte cells and tissues were sampled. Hemolymph fluid containing the hemocyte cells of each individual was collected. The hemocytes pellet and the different tissues were shock-frozen in liquid nitrogen. For transcriptome generation, an additional set of sequence data, generated from mantle tissue of  L. elliptica collected at Rothera Research Station, Adelaide Island (range of shell size between 50.1-83.5mm), was provided by M. Clark of the British Antarctic Survey (BAS; for details see Clark et al., 2010. Insights into shell deposition in the Antarctic bivalve Laternula elliptica: gene discovery in the mantle transcriptome using 454 pyrosequencing. Bmc Genomics 11).