Karen J. Marsh

The detoxification limitation hypothesis : Where did it come from and where is it going?

The detoxification limitation hypothesis is firmly entrenched in the literature to explain various aspects of the interaction between herbivores and plant toxins. These include explanations for the existence of specialist and generalist herbivores and for the prevalence of each of these. The hypothesis suggests that the ability of mammalian herbivores to eliminate plant secondary metabolites (PSMs) largely determines which plants, and how much, they can eat. The value of the hypothesis is that it provides a clear framework for understanding how plant toxins might limit diet breadth. Thus, it is surprising, given its popularity, that there are few studies that provide experimental support either for or against the detoxification limitation hypothesis. There are two likely reasons for this. First, Freeland and Janzen did not formally propose the hypothesis, although it is implicit in their paper. Second, it is a difficult hypothesis to test, requiring an understanding of the metabolic pathways that lead to toxin elimination. Recent attempts to test the hypothesis appear promising. Results suggest that herbivores can recognize mounting saturation of a detoxification pathway and adjust their feeding accordingly to avoid intoxication. One strategy they use is to ingest a food containing a toxin that is metabolized by a different pathway. This demonstrates that careful selection of food plants is a key to existing in a chemically complex environment. As more studies characterize the detoxification products of PSMs, we will better understand how widespread this phenomenon is.

Conflicting demands on detoxification pathways influence how common brushtail possums choose their diets

Most herbivores eat more and survive better when they have access to a variety of foods. One explanation involves the detoxification of plant secondary metabolites (PSMs). By feeding from a variety of plants that contain different classes of PSMs, animals can use multiple detoxification pathways and presumably consume more food. Although popular, this theory is difficult to test because it requires knowledge of the detoxification pathways of each PSM in the diet. We established that common brushtail possums (Trichosurus vulpecula) use various combinations of oxidation, hydrolysis, and conjugation with glucuronic acid (GA) or glycine to detoxify six PSMs. Compared to their ingestion of a single PSM, possums ate more when offered a choice between two diets containing PSMs that require apparently independent detoxification pathways (benzoate and 1,8-cineole, benzoate and p-cymene, benzoate and orcinol, benzoate and salicin, or orcinol and 1,8-cineole). However, possums still did not eat as much of these diets as they did of a basal diet free of PSMs. This suggests that detoxification pathways are never independent, but are separated instead by degrees. In contrast, possums offered a choice of two PSMs that require competing detoxification pathways (1,8-cineole and p-cymene, 1,8-cineole and salicin, or orcinol and salicin) ate no more than when offered diets containing one of the compounds. There was an exception: even though both rutin and orcinol are detoxified via conjugation with GA, the feeding behavior of possums did not suggest competition for detoxification pathways. This implies that the supply of GA is not limiting. This study provides the first convincing evidence that herbivorous mammals can eat more by selecting mixed diets with a diversity of PSMs that make full use of their detoxification potential. It also emphasizes that other behavioral and physiological factors, such as transient food aversions, influence feeding behavior.

Differential susceptibility to Eucalyptus secondary compounds explains feeding by the common ringtail (Pseudocheirus peregrinus) and common brushtail possum (Trichosurus vulpecula)

The effect of two plant secondary metabolites, tannins and formylated phloroglucinol compounds (FPCs), on the intake of Eucalyptus foliage by common ringtail (Pseudocheirus peregrinus) and common brushtail possums (Trichosurus vulpecula) was studied. We manipulated the amount of tannin that was free to bind with protein by coating foliage with polyethylene glycol 4000 (PEG) and relied on natural intraspecific variation in FPC concentrations. In contrast to ringtail possums, brushtail possums showed a greater tolerance to FPCs and ate more foliage when it was coated with PEG, suggesting that tannins limited their food intake. Brushtails detected the effects of tannins through immediate oral sensations rather than through systemic effects. Ringtail possums appeared highly tolerant of foliar tannins yet susceptible to low concentrations of FPCs. We could not detect any interaction between tannins and FPCs that affected the intake of Eucalyptus foliage by either species of possum. Although ringtail and brushtail possums are widely regarded as specialist and generalist folivores, respectively, their differential susceptibility to co-occurring secondary metabolites suggests greater complexity. Each possum species appears to be a specialist in its own right, which leads to a partitioning of available foliage. Brushtails avoid tannins and ringtails avoid FPCs.

The effect of inactivating tannins on the intake of Eucalyptus foliage by a specialist Eucalyptus folivore (Pseudocheirus peregrinus) and a generalist herbivore (Trichosurus vulpecula)

The paucity of evidence on eucalypt browsing by common brushtail and common ringtail possums suggests that ringtails preferentially eat foliage from trees within the subgenus Monocalyptus. In contrast, brushtails eat less eucalypt foliage than do ringtails and prefer trees from the subgenus Symphyomyrtus. Trees from these subgenera differ in their defensive chemicals. Both contain tannins but it appears that only the symphyomyrts synthesise formylated phloroglucinol compounds (FPCs). We fed possums foliage from several individual Eucalyptus rossii and E. consideniana, both monocalypts, to avoid the confounding factor of FPCs, and examined the effects of blocking tannins by dipping foliage in polyethylene glycol (PEG). Ringtails and brushtails differed in their abilities to eat foliage from these eucalypts. The ringtails ate much more than did the brushtails and showed a small (about 10%) but significant increase in feeding in response to PEG. The brushtails were reluctant to eat foliage from either eucalypt species but doubled their intake when leaves were coated with PEG. Even so, they still did not eat enough to meet maintenance requirements for energy and nitrogen. Neither ringtails nor brushtails preferred foliage from any individual E. rossii tree, suggesting that all trees were equally defended. However, brushtails preferred foliage from some E. consideniana to others. Monocalypt tannins are clearly important barriers to feeding in brushtail possums, but further research with higher doses of PEG will confirm whether they are the only deterrent chemicals in monocalypt foliage. ZO y s o ly K. J a et

Behavioural contributions to the regulated intake of plant secondary metabolites in koalas

In a given period of time, herbivores often eat less as dietary plant secondary metabolite (PSM) concentrations increase. This reduction in total food intake is interpreted as a need for the herbivore to regulate PSM ingestion in order to avoid toxication. However, regulation of PSM ingestion involves more than the reduction of total intake; it involves an alteration of meal patterns, through a reduction in the number and/or the size of the meals eaten. Despite this, studies of how herbivores alter their meal patterns when offered varying concentrations of PSMs are rare. We investigated whether koalas adjust the number and/or the size of their meals when offered eucalypt foliage varying naturally in concentrations of formylated phloroglucinol compounds (FPCs), a group of PSMs that have previously been shown to inhibit total food intake. High FPC concentrations caused koalas to eat more slowly, eat shorter meals and eat less per meal, which resulted in a reduced total intake. Because increasing FPC concentrations did not cause koalas to alter the number of meals that they ate, clear individual differences between koalas were observed, where some consistently ate fewer larger meals and others ate many smaller meals. Thus, different feeding strategies may still achieve the same outcome of a regulated intake of PSMs. The changes observed match the meal patterns of other herbivores ingesting PSMs known to stimulate nausea and emetic pathways, supporting the idea that feedback signals from nausea are an important way that koalas avoid toxication when eating eucalypt foliage.

The effect of plant secondary metabolites on the interplay between the internal and external environments of marsupial folivores

Most woody plants contain a diverse array of plant secondary metabolites (PSMs) that deter vertebrate herbivores. However, mammalian folivores have evolved a complex of physiological and behavioural strategies to counter these compounds, leading to the development of an “evolutionary arms race”. Marsupial folivores are ideal models to investigate the role of PSMs in the interaction between the external foraging environment and the digestive physiology of mammalian herbivores, as we have a very strong understanding of the diversity and modes of action of PSMs in Eucalyptus, as well as the mechanisms by which animals overcome the effects of these compounds. Studies of marsupial folivores have benefited from the facts that: these herbivores subsist on relatively poor quality diets; they include feeding types from specialist species such as the koala, to generalists; and life history factors such as maternal investment in reproduction can be measured more easily than in eutherians. Here, we describe patterns of spatial variation in the types and distributions of plant secondary metabolites in Australian forests and discuss how this variation influences foraging behaviour, habitat selection and life history strategies in arboreal, folivorous marsupials. We also provide a summary of our understanding of the mechanisms by which marsupials detect and regulate their intake of toxic compounds. While our examples are drawn largely from studies of the interaction between marsupials and Eucalyptus, this knowledge is applicable to advancing our understanding of interactions in plant–mammal systems more broadly. We also identify and discuss key areas that should be the focus of future research.

Feeding rates of a mammalian browser confirm the predictions of a 'foodscape' model of its habitat.

Adequate nutrition is a fundamental requirement for the maintenance and growth of populations, but complex interactions between nutrients and plant toxins make it difficult to link variation in plant quality to the ecology of wild herbivores. We asked whether a ‘foodscape’ model of habitat that uses near-infrared spectroscopy to describe the palatability of individual trees in the landscape, predicted the foraging decisions of a mammalian browser, the koala (Phascolarctos cinereus). Specifically, we considered four behavioural decision points at which nutritional quality may influence an animal’s decision. These were: which tree to enter, whether to feed from that tree, when to stop eating, and how long to remain in that tree. There were trends for koalas to feed in eucalypt trees that were more palatable than unvisited neighbouring conspecific trees, and than trees that they visited but did not eat. Koalas ate longer meals in more palatable trees, and stayed longer and spent more time feeding per visit to these trees. Using more traditional chemical analyses, we identified that an interaction between the concentrations of formylated phloroglucinol compounds (a group of plant secondary metabolites) and available N (an integrated measure of tannins, digestibility and N) influenced feeding. The study shows that foodscape models that combine spatial information with integrated measures of food quality are a powerful tool to predict the feeding behaviour of herbivores in a landscape.

A Metabolomic Approach to Identifying Chemical Mediators of Mammal–Plant Interactions

Different folivorous marsupials select their food from different subgenera of Eucalyptus, but the choices cannot be explained by known antifeedants, such as formylated phloroglucinol compounds or tannins, or by nutritional quality. Eucalypts contain a wide variety of plant secondary metabolites so it is difficult to use traditional methods to identify the chemicals that determine food selection. Therefore, we used a metabolomic approach in which we employed 1H nuclear magnetic resonance spectroscopy to compare chemical structures of representatives from the two subgenera and to identify chemicals that consistently differ between them. We found that dichloromethane extracts of leaves from most species in the subgenus Eucalyptus differ from those in Symphyomyrtus by the presence of free flavanones, having no substitution in Ring B. Although flavanoids are known to deter feeding by certain insects, their effects on marsupials have not been established and must be tested with controlled feeding studies.

Translating physiological signals to changes in feeding behaviour in mammals and the future effects of global climate change

Dr Olivier Bonnet and Dr Paulo Carvalho. BDM, KJM MDD and WJF are supported by grants from the Australian Research Council (DP140100228; DE120101263) and MDD also by the National Science Foundation (DEB 1342615).

Continuous monitoring of feeding by koalas highlights diurnal differences in tree preferences

Abstract Context. It is difficult to measure feeding rates for most wild nocturnal mammalian herbivores. Thus, although koalas are a popular species to study, we have a poor understanding of their activity patterns and feeding ecology. Researchers often assume that the trees that koalas occupy during the day indicate feeding preferences, but they may better reflect preferred resting sites. Aims. We recorded the activities of koalas, with an emphasis on their feeding, particularly when they fed, the trees that they fed from, the number of meals they consumed and the variability in these measurements. Methods. We continuously monitored eight koalas by audio- and radio-telemetry for 14 consecutive 24-h periods each. We followed two koalas at a time and recorded the trees they visited, when, where and how long they fed, and the size and nutritional composition of the trees in the landscape. Key results. Individual koalas varied in how many trees they visited, how many meals they ate and how long they spent feeding during each 24-h period. They preferred Eucalyptus globulus trees during the day, but fed mainly at night, with a preference for E. viminalis. The trees that koalas visited during the day were larger than those that they visited at night. Conclusions. The trees that koalas occupied during the day were poor indicators of their diet preferences, whereas the daily feeding activities of individual koalas varied widely. Implications. Predicting a koala’s diet from the trees it occupies during the day is fraught with error. Although the trees that koalas rest in are important in the species ecology for reasons other than feeding, we should refrain from using them to predict an animal’s diet. Because feeding activity is difficult to measure, it is probably best done indirectly by analysing leaf-cuticle fragments or waxes in faeces. The substantial day-to-day variation in koala activities also indicates that behavioural and physiological studies of koalas require long monitoring periods – a week or longer.