Megaherbivore browsers vs. tannins: is being big enough?

Megaherbivores have been of particular interest to scientists because of the physiological and ecological challenges associated with their extreme body size. Yet, one question that has seldom been explored is how browsing megaherbivores cope with plant secondary metabolites (PSMs), such as tannins, found in their food. It is possible that the sheer body size of these megaherbivores allows them to ingest tannins with no deleterious effects. However, it is plausible that megaherbivores must rely on other mechanisms to cope with PSMs, such as the production of salivary tannin-binding proteins. Thus, we aimed to determine whether megaherbivore browsers produce tannin-binding proteins to further reduce the consequences of ingesting a tannin-rich diet. Using a series of laboratory assays, we explored whether elephants, black rhinoceros, and giraffe had tannin-binding proteins in their saliva. We tested for the presence of proline-rich proteins in the saliva using two different approaches: (1) SDS-PAGE using Laemmli's (Laemmli, Nature 227:680-685, 1970) destaining method, and (2) comparative SDS-PAGE gels using Beeley et al.'s (Beeley et al. Electrophoresis 12:493-499, 1991) method for staining and destaining to probe for proline-rich proteins. Then, to test for the tannin-binding affinity of their saliva, we performed an inhibition assay. We did not observe proline-rich proteins in any of the megaherbivore species, but they did have other protein(s) in their saliva that have a high tannin-binding affinity. Our results highlight that, despite their large body sizes, and their abilities to tolerate low-quality food, browsing megaherbivores have likely evolved tannin-binding proteins as a way of coping with the negative effects of tannins.