Exploring the Links between Testate Amoeba Traits and Eutrophication in Lakes

There is considerable potential to apply traits-based approaches to the subfossil remains of shell-forming micro-organisms which preserve well in sediments and whose short generation times make it possible to achieve high-temporal resolution in palaeoecological studies. In this paper we review progress in applying traits-based approaches to freshwater testate amoebae (Arcellinida), a diverse group of protists which are abundant in lakes and are valuable palaeoecological indicators. Drawing on published studies from the last ~10 years, we describe the methodologies which have been applied to delimit testate amoeba (TA) traits and review the challenges associated with their measurement and interpretation. We also showcase the results of ongoing work in seven lakes (UK, Canada) which aims to (i) examine the character and causes of trait-based variability in palaeolimnological settings; (ii) apply novel biometric approaches to aid in trait delimitation; and (iii) explore the potential for combining phylogenetic with advanced morphometric approaches to better understand the ecological and evolutionary significance of TA traits. We applied geometric morphometric analysis to define test size and shape indices and summarise testate amoeba community dynamics along a temporal gradient of eutrophication in a large shallow lake in Scotland, UK. Cluster analysis of test size and shape indices yielded three assemblages, each dominated by a single shape: elongate, spherical and ovoid. When plotted stratigraphically, we observed increases in spherical tests, decreases in elongate tests and shrinking of test size coeval with eutrophication. Decreases in the elongate cluster may reflect benthic conditions with reduced oxygen levels, while increases in the spherical cluster are likely associated with an expanding macrophyte community that promoted pelagic and epibiotic life habits. Shrinking of test size may be a stress response to eutrophication and/or warming temperatures. Tracking community dynamics using test size and shape indices was found to be as effective as using traditional species-based approaches to summarize key palaeolimnological changes, with the added benefit of being free of taxonomic bias. The approach thus shows significant potential for future studies of aquatic community change in nutrient-impacted lakes. To further investigate the functional significance of the Arcellinida shape groups, we examined the phylogenetic signal of morphological traits in elongate Difflugia species which occur in eutrophic and mesotrophic lakes. Previous phylogenetic work has shown that whilst overall test morphology (e.g., spherical or elongate) is generally conserved in Arcellinida lineages, the taxonomic significance of other traits (e.g., size, ornamentation, mixotrophy/heterotrophy metabolism type) is not well understood. Our analyses revealed two clades which could be reliably separated by test size and the presence/absence of mixotrophy. This suggests that test size may reflect trophic level, with smaller taxa occupying lower trophic levels. In addition to having larger tests, elongate mixotrophic Difflugia are characterised by wide, flat bases and inflation of the lower part of the test. These morphological traits may provide additional space for endosymbionts and/or increased surface area to aid light transmission. Continued research into the ecological and evolutionary significance of morphological traits will serve to strengthen palaeoecological inferences, increasing the importance of lacustrine testate Arcellinida as environmental proxies.