Figure 3 Delta rank comparison of highly significant (p < 0.001) Cladocopium and Durusdinium GO categories. Higher delta ranks indicate upregulation and lower delta ranks indicate downregulation.
We were concerned that the variance in the number of symbiont counts per sample would skew the results of our functional analyses. This was primarily due to the fact that read counts for each symbiont covaried with the symbiont’s proportion in the host. While this effect was formally accounted for in our analyses by including the log10(total count) as a covariate in DESeq2 models, doubts remained as to whether this would not create additional artifacts. One reassuring argument is that the main gene expression response (to codominance) was not aligned with the minor-major axis (i.e. with variation in total counts) but rather happened in the middle of the total counts’ scale. Still, to further test for possible residual effects of total counts variation on our results, we resampled the counts across samples so that each sample would have similar counts. We reduced the counts to 30,000 per sample, 15,000 per sample, and 8,000 per sample, the latter being the lowest number of counts for a symbiont type observed in a sample. We repeated the DESeq2 and GO_MWU analyses for each resampled dataset and compared the GO delta ranks of each. Reassuringly, we observed similar results at the GO delta-ranks level across all resampling trials, indicating that our functional results were largely unaffected by differences in coverage across samples (Fig. S3).