Massively Parallel RNA Sequencing Identifies a Complex Immune Gene Repertoire in the lophotrochozoan Mytilus edulis

The marine mussel Mytilus edulis and its closely related sister species are distributed world-wide and play an important role in coastal ecology and economy. The diversification in different species and their hybrids, broad ecological distribution, as well as the filter feeding mode of life has made this genus an attractive model to investigate physiological and molecular adaptations and responses to various biotic and abiotic environmental factors. In the present study we investigated the immune system of Mytilus, which may contribute to the ecological plasticity of this species. We generated a large Mytilus transcriptome database from different tissues of immune challenged and stress treated individuals from the Baltic Sea using 454 pyrosequencing. Phylogenetic comparison of orthologous groups of 23 species demonstrated the basal position of lophotrochozoans within protostomes. The investigation of immune related transcripts revealed a complex repertoire of innate recognition receptors and downstream pathway members including transcripts for 27 toll-like receptors and 524 C1q domain containing transcripts. NOD-like receptors on the other hand were absent. We also found evidence for sophisticated TNF, autophagy and apoptosis systems as well as for cytokines. Gill tissue and hemocytes showed highest expression of putative immune related contigs and are promising tissues for further functional studies. Our results partly contrast with findings of a less complex immune repertoire in ecdysozoan and other lophotrochozoan protostomes. We show that bivalves are interesting candidates to investigate the evolution of the immune system from basal metazoans to deuterostomes and protostomes and provide a basis for future molecular work directed to immune system functioning in Mytilus

Transmigration of polymorphnuclear neutrophils and monocytes through the human blood-cerebrospinal fluid barrier after bacterial infection in vitro

Background: Bacterial invasion through the blood-cerebrospinal fluid barrier (BCSFB) during bacterial meningitis causes secretion of proinflammatory cytokines/chemokines followed by the recruitment of leukocytes into the CNS. In this study, we analyzed the cellular and molecular mechanisms of polymorphonuclear neutrophil (PMN) and monocyte transepithelial transmigration (TM) across the BCSFB after bacterial infection. Methods: Using an inverted transwell filter system of human choroid plexus papilloma cells (HIBCPP), we studied leukocyte TM rates, the migration route by immunofluorescence, transmission electron microscopy and focused ion beam/scanning electron microscopy, the secretion of cytokines/chemokines by cytokine bead array and posttranslational modification of the signal regulatory protein (SIRP) α via western blot. Results: PMNs showed a significantly increased TM across HIBCPP after infection with wild-type Neisseria meningitidis (MC58). In contrast, a significantly decreased monocyte transmigration rate after bacterial infection of HIBCPP could be observed. Interestingly, in co-culture experiments with PMNs and monocytes, TM of monocytes was significantly enhanced. Analysis of paracellular permeability and transepithelial electrical resistance confirmed an intact barrier function during leukocyte TM. With the help of the different imaging techniques we could provide evidence for para- as well as for transcellular migrating leukocytes. Further analysis of secreted cytokines/chemokines showed a distinct pattern after stimulation and transmigration of PMNs and monocytes. Moreover, the transmembrane glycoprotein SIRPα was deglycosylated in monocytes, but not in PMNs, after bacterial infection. Conclusions: Our findings demonstrate that PMNs and monoctyes differentially migrate in a human BCSFB model after bacterial infection. Cytokines and chemokines as well as transmembrane proteins such as SIRPα may be involved in this process

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 4: gonad and pyloric caeca mass and indices

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 2b: seawater-coelomic fluid PO2 gradients with slower warming

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 2: accumulation of anaerobic end products during heat stress under fully aerobic water conditions

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 4: PO2 gradients between seawater and coelomic fluid at acutely altered temperatures

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 3: A. rubens (fed vs. unfed) oxygen diffusion gradients

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Response of the sea star Asterias rubens to heat stress and ocean acidification: long-term and short-term experimental data

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations

Experiment on the response of the sea star Asterias rubens to heat stress and ocean acidification: experiment 1: A. rubens feeding rates

Robust estimates of marine species vulnerability to ongoing climate change require realistic stressor experiments. Here, we subjected an important coastal predator, the sea star Asterias rubens, to projected warming and ocean acidification over an annual seasonal cycle. Warming and, less so, acidification, had strongly season-specific impacts on animal energy budgets. Specifically, simulated future summer temperatures caused >95% sea star mortality, reduced feeding rate and body mass loss. Additional acute experiments demonstrated that respiratory oxygen flux was preferentially directed to support high summer metabolism at the expense of feeding-related processes. Using 15 years of field temperature data and end of century warming projections, we estimate that potentially lethal summer heat waves will occur in 20% of future years. Our study demonstrates the importance of assessing stress responses along seasonal thermal cycles and the high selective force that future summer heat waves likely can exert on coastal marine animal populations