Establishing typical values for hemocyte mortality in individual California mussels, Mytilus californianus

The purposes of this study were to: (1) establish FACS as a method to analyze hemocytes in M. californianus, (2) evaluate whether hemolymph can be repeatedly drawn from an individual mussel without adverse effects, and if so, whether the time interval between repeated samples affects hemocyte mortality or organism survival, and (3) describe the typical intra- and inter-individual variability in hemocyte mortality across repeated samples in mussels not exposed to typical abiotic stressors. To assess whether individuals’ hemolymph could be repeatedly sampled (without killing the animal), and to establish baseline hemocyte mortality values in unperturbed mussels, hemolymph was sampled from the same individuals at time zero and three subsequent timepoints, with varying intervals between subsequent samples: 24, 48, 72 or 168 h. Each of these four different sampling intervals defined a separate experimental Group (n = 14-15 mussels per Group). We evaluated each individual's hemocyte mortality (using FACS), and then compared these individual-group samples to pooled-group samples that we created (of four individuals each). We assessed total hemocyte mortality, and mortality of the granulocyte and hyalinocyte populations as well. The attached excel file contains columns for the Mussel ID, the specific Group that the mussel was a part of, and the Sampling Timepoint (#1-4). There are two columns for each the hyalinocyte and granulocyte data: 1) the overall percentage of hyalinocytes or granulocytes relative to the total hemocyte population, and 2) the percentage of dead hyalinocytes or granulocytes relative to their respective cell type. There is one column for the overall percentage of dead hemocytes. The pooled samples that were obtained for each Group's sampling timepoint are indicated in MusselID column as "pooled_..." with an identifying number linked to that Group. There were 2 pooled samples for each Group's sample timepoint (i.e., 8 pooled samples per Group). The mussel morphometric data (shell height, width, length, and body mass) are also included for each individual