OPTIMISATION OF BIOMARKERS FOR IMMUNOTOXICITY IN MUSSELS: A Combined Laboratory-in situ Experiment

Abstract

The development of new biological effect markers for immunotoxicity in mussels could enhance biomonitoring programmes. To reach the European Union's goal of good environmental status in the marine environment, new tools are needed to ensure the health of marine ecosystems. Previous research has shown that pollution could affect mussels' immune systems in in vivo and ex vivo experiments. However, biomarkers for immunotoxicity have never been tested in field experiments; in which mussels from a polluted environment have their immune systems induced and their response compared to that of a healthy immune system. Here, we show that there are differences in immune response between blue mussels from a polluted site and the reference site. M. edulis was collected from Gothenburg harbour, used as a proxy for a polluted site, and from a more pristine site in the archipelago on the west coast of Sweden. The mussels were then inoculated with either lipopolysaccharide (LPS) or artificial seawater. After 24 hours of LPS exposure, haemocyte viability, total haemocyte count and differential haemocyte count were evaluated using flow cytometry. After an additional 24 hours, the cell-free haemolymph and haemocytes were used in a lysozyme activity assay, and the gene expression in the gills and haemocytes was evaluated for immune-relevant and efflux genes. The species of the mussels was also identified through PCR of the Glu-5’ gene. The result showed an increase in total haemocyte count and granulocytes in the treated group from the polluted site compared to the treated group from the more pristine site (p=0.07). More dead cells in the mussels from Gothenburg harbour could be observed; however, this was not statistically significant. Differences in inter-individual variation in efflux genes between the sites were also observed. Downregulation of the antimicrobial peptide genes was statistically significant in the treated groups compared to control from both sites. The current thesis suggests these biomarkers as candidates for further optimisation.