Elimination of plant toxins by herbivorous woodrats: revisiting an explanation for dietary specialization in mammalian herbivores

Constraints on rates of detoxification and elimination of plant toxins are thought to be responsible for limiting dietary specialization in mammalian herbivores. This hypothesis, known as the detoxification limitations hypothesis, suggests that most mammalian herbivores are generalists to avoid overdosing on toxins from a single plant species. The hypothesis also predicts that the few mammalian specialists that exist should have adaptations for rapid detoxification and elimination of plant secondary compounds. We took a pharmacological approach to test whether specialists eliminate toxins from the bloodstream faster than generalists. We compared elimination rate and total exposure of alpha-pinene in closely related dietary specialist and generalist woodrats, Neotoma stephensi and N. albigula, respectively. Animals were orally gavaged with alpha-pinene, a plant secondary compound present in the natural diets of both woodrat species. We collected venous blood at 3, 6, 10, 15, and 20 min post-ingestion of alpha-pinene. Blood was analyzed for alpha-pinene concentration using gas chromatography. We found that specialist and generalist woodrats did not differ in elimination rates of alpha-pinene. However, specialists had lower exposure levels of alpha-pinene than generalists due to lower initial delivery of alpha-pinene to the general circulation. The levels of alpha-pinene detected in the bloodstream of specialists were 4.7-5.3x lower over all time intervals than generalists. Thus, specialists encounter a functionally lower dose of toxin than generalists. We suggest that the lower exposure level of specialist woodrats may be due to mechanisms in the gut that decrease toxin absorption. Regardless of mechanism, lower exposure to plant toxins may allow specialists to forage on diets with high toxin concentrations thereby facilitating dietary specialization.     

Diet breadth of mammalian herbivores: nutrient versus detoxification constraints

Two hypotheses, nutrient constraints and detoxification limitation, have been proposed to explain the lack of specialists among mammalian herbivores. The nutrient constraint hypothesis proposes that dietary specialization in mammalian herbivores is rare because no one plant can provide all requisite nutrients. The detoxification limitation hypothesis suggests that the mammalian detoxification system is incapable of detoxifying high doses of similar secondary compounds present in a diet of a single plant species. We experimentally tested these hypotheses by comparing the performance of specialist and generalist woodrats (Neotoma) on a variety of dietary challenges. Neotoma stephensi is a narrow dietary specialist with a single species, one-seeded juniper, Juniperus monosperma, comprising 85–95% of its diet. Compared with other plants available in the habitat, juniper is low in nitrogen and high in fiber, phenolics, and monoterpenes. The generalist woodrat, N. albigula, also consumes one-seeded juniper, but to a lesser degree. The nutrient constraint hypothesis was examined by feeding both species of woodrats a low-nitrogen, high-fiber diet similar to that found in juniper. We found no differences in body mass change, or apparent digestibility of dry matter or nitrogen between the two species of woodrats after 35 days on this diet. Moreover, both species were in positive nitrogen balance. We tested the detoxification limitation hypothesis by comparing the performance of the generalist and specialist on diets with and without juniper leaves, the preferred foliage of the specialist, as well as on diets with and without α-pinene, the predominant monoterpene in juniper. We found that on the juniper diet, compared with the specialist, the generalist consumed less juniper and lost more mass. Urine pH, a general indicator of overall detoxification processes, declined in both groups on the juniper diet. The generalist consumed half the toxin load of the specialist yet its urine pH was slightly lower. Moreover, the generalist consumed significantly less of the treatment with high concentrations of α-pinene compared to the control treatment, while the specialist consumed the same amount of food regardless of α-pinene concentration. For both groups, urine pH declined as levels of α-pinene in the diet increased. The generalist produced a significantly more acidic urine than the specialist on the treatment with the highest α-pinene concentration. Our results suggest that in this system, specialists detoxify plant secondary compounds differently than generalists and plant secondary compounds may be more important than low nutrient levels in maintaining dietary diversity in generalist herbivores. Received: 5 May 1999 / Accepted: 14 November 1999     

Differential hepatic gene expression of a dietary specialist (Neotoma stephensi) and generalist (Neotoma albigula) in response to juniper (Juniperus monosperma) ingestion

Dietary specialization is thought to be rare in mammalian herbivores because of limitations of their detoxification system in processing large doses of a single type of plant secondary compound (PSC). Therefore, in order to specialize on a single species of plant, mammalian herbivores must have a highly efficient detoxification system for the particular types of PSCs they ingest. Using microarray technology, we looked at the expression of hepatic genes of a dietary specialist, Neotoma stephensi, and a sympatric generalist, Neotoma albigula, in response to diets containing different levels of one-seeded juniper (Juniperus monosperma). We found large between species differences in gene expression, as well as large within species differences when specialists fed a low juniper diet (25% juniper) were compared to specialists fed their ecologically relevant level of juniper (70% juniper). We also tested the hypothesis that the specialist relies on less costly phase I detoxification enzymes more than phase II compared to the generalist. Although we found that the specialist had higher cumulative as well as average expression of phase I versus phase II enzymes, the generalist had a similar pattern of expression for phase I versus phase II enzymes.     

Testing the diet-breadth trade-off hypothesis: differential regulation of novel plant secondary compounds by a specialist and a generalist herbivore

Specialist herbivores are predicted to have evolved biotransformation pathways that can process large doses of secondary compounds from the plant species on which they specialize. It is hypothesized that this physiological specialization results in a trade-off such that specialists may be limited in ability to ingest novel plant secondary compounds (PSCs). In contrast, the generalist foraging strategy requires that herbivores alternate consumption of plant species and PSC types to reduce the possibility of over-ingestion of any particular PSC. The ability to behaviorally regulate is a key component of this strategy. These ideas underpin the prediction that in the face of novel PSCs, generalists should be better able to maintain body mass and avoid toxic consequences compared to specialists. We explored these predictions by comparing the feeding behavior of two herbivorous rodents: a juniper specialist, Neotoma stephensi, and a generalist, Neotoma albigula, fed diets with increasing concentrations of phenolic resin extracted from the creosote bush (Larrea tridentata), which produces a suite of PSCs novel to both species. The specialist lost more mass than the generalist during the 15-day trial. In addition, although the specialist and generalist both regulated phenolic resin intake by reducing meal size while on the highest resin concentration (4%), the generalist began to regulate intake on the 2% diet. The ability of the generalist to regulate intake at a lower PSC concentration may be the source of the generalist’s performance advantage over the specialist. These data provide evidence for the hypothesis that the specialist’s foraging strategy may result in behavioral as well as physiological trade-offs in the ability to consume novel PSCs.     

Plant Secondary Compounds as Diuretics: An Overlooked Consequence

Plant secondary compounds are deterrents and toxins to a varietyof herbivores. The effect of secondary compounds on water balanceof herbivores is virtually unexplored, yet many secondary compoundsare renowned for their diuretic effects in humans and laboratoryrats. We review data from the ethnopharmocological literatureon plants with diuretic effects. We also present our data fromexperiments on water intake of specialist (Neotoma stephensi)and generalist woodrats (N. albigula) consuming plant secondarycompounds from their natural diet. We measured effects of dietarysecondary compounds on voluntary water consumption, urine volumeand urine osmolarity. Ingestion of secondary compounds increasedwater intake and urine output and decreased urine osmolarityin both species. However, the generalist was more impacted bydietary secondary compounds than the specialist. Our resultscombined with that from the literature suggest that diuresismay be a prevalent consequence of ingestion of secondary compounds.Many herbivores live in arid habitats with limited access tofree-standing water, thus an increase in the desire for watermay have profound consequences on foraging behavior and fitness.     

An oak (Quercus agrifolia) specialist (Neotoma macrotis) and a sympatric generalist (Neotoma lepida) show similar intakes and digestibilities of oak

Dietary specialization is thought to be rare in mammalian herbivores as a result of either a limitation in their detoxification system to metabolize higher doses of plant secondary compounds or deficiencies in nutrients present in a diet composed of a single species of plant. Neotoma macrotis is an oak specialist, whereas Neotoma lepida is a dietary generalist when sympatric with N. macrotis. We hypothesized that N. macrotis would have a higher tolerance for and digestibility of oak. We determined the two species' tolerances for oak by feeding them increasing concentrations of ground oak leaves until they could no longer maintain body mass. The highest concentration on which both species maintained body mass was 75% oak. There were no differences between the species in their abilities to digest dry matter, nitrogen, or fiber in the oak diets. The species' similar tolerances for oak were probably due to their similar abilities to digest and potentially assimilate the ground oak leaves.     

Xenobiotic metabolism of plant secondary compounds in juniper (Juniperus monosperma) by specialist and generalist woodrat herbivores, genus Neotoma

Mammalian herbivores routinely consume diets laden with often-toxic xenobiotics, yet the manner in which mammalian herbivores detoxify these plant secondary compounds (PSC) is largely unknown. Theory predicts that specialists rely more heavily on functionalization pathways whereas generalists rely on conjugation pathways to metabolize PSC in their diet. We took a pharmacological approach to determine how a specialist (Neotoma stephensi) of juniper foliage (Juniperus monosperma) and a generalist (N. albigula) may process the same dietary PSC. We investigated the xenobiotic metabolizing enzymes of the specialist and generalist on a control diet and a low (25%) juniper diet. We also examined enzyme activities in the specialist on a high (70%) juniper diet. We assayed for cytochrome P450 concentration and biotransformation activities of three specific cytochrome P450 isozymes (CYP1A, CYP2B, CYP3A), NAD(P)H:quinone oxidoreductase, glutathione conjugation, sulfation and glucuronidation. Results provide partial evidence for the hypothesis in that the specialist and generalist consuming juniper at a level similar to their natural diet, differ in the level of conjugation enzyme activity with generalists having higher activity overall than specialists.     

Ingestion of plant secondary compounds causes diuresis in desert herbivores

Plant secondary compounds are recognized deterrents and toxins to a variety of herbivores. The effect of secondary compounds on water balance of herbivores is virtually unexplored, yet secondary compounds could potentially cause a decrease in an animal's ability to maintain water balance. We investigated the effects of secondary compounds, alpha-pinene and creosote resin, on water balance in three species of herbivorous woodrats (Neotoma stephensi, N. albigula, N. lepida). In separate experiments, we measured the effect of these secondary compounds on voluntary water consumption, urine volume and urine osmolarity. In both experiments, water intake and urine volume increased and urine osmolarity decreased compared to controls. Water balance of specialist or experienced woodrats was less affected than generalists and woodrats with less prior experience with particular secondary compounds. Our results suggest that secondary compounds have diuretic-like effects on herbivores. Woodrats live in arid habitats with limited access to freestanding water; thus an increase in water requirements may have profound consequences on foraging behavior and fitness.     

Warmer ambient temperatures depress liver function in a mammalian herbivore

Diet selection in mammalian herbivores is thought to be mainly influenced by intrinsic factors such as nutrients and plant secondary compounds, yet extrinsic factors like ambient temperature may also play a role. In particular, warmer ambient temperatures could enhance the toxicity of plant defence compounds through decreased liver metabolism of herbivores. Temperature-dependent toxicity has been documented in pharmacology and agriculture science but not in wild mammalian herbivores. Here, we investigated how ambient temperature affects liver metabolism in the desert woodrat, Neotoma lepida. Woodrats (n = 21) were acclimated for 30 days to two ambient temperatures (cool = 21°C, warm = 29°C). In a second experiment, the temperature exposure was reduced to 3.5 h. After temperature treatments, animals were given a hypnotic agent and clearance time of the agent was estimated from the duration of the hypnotic state. The average clearance time of the agent in the long acclimation experiment was 45% longer for animals acclimated to 29°C compared with 21°C. Similarly, after the short exposure experiment, woodrats at 29°C had clearance times 26% longer compared with 21°C. Our results are consistent with the hypothesis that liver function is reduced at warmer environmental temperatures and may provide a physiological mechanism through which climate change affects herbivorous mammals.     

A specialist herbivore (Neotoma stephensi) absorbs fewer plant toxins than does a generalist (Neotoma albigula)

Detoxification capacity of enzymes in the liver is thought to be the primary factor governing dietary toxin intake by mammalian herbivores. Recently, toxin absorption in the gut was proposed as an alternative process that also influences toxin intake. We examined the role of the gut in regulating toxin absorption by quantifying excretion of a plant secondary compound in the feces. We hypothesized that specialists have a greater capacity to reduce intestinal absorption of toxins than do generalists. To test this hypothesis, we compared fecal excretion of alpha-pinene in specialist (Neotoma stephensi) and generalist (Neotoma albigula) woodrats. Alpha-pinene is the most abundant monoterpene in Juniperus monosperma, which occurs in the natural diet of both woodrat species. Woodrats were fed alpha-pinene in diets containing juniper foliage for 3 wk and, in a separate experiment, were given a single oral dose of alpha-pinene. Feces were collected from animals at the end of each experiment and analyzed for alpha-pinene concentration using gas chromatography. Both woodrat species excreted unchanged alpha-pinene in the feces. However, specialist woodrats excreted 40% more alpha-pinene per unit ingested from a juniper diet and excreted nearly four times a greater percentage of an oral dose of alpha-pinene compared with generalists.     

Evolutionary irony: evidence that ‘defensive’ plant spines act as a proximate cue to attract a mammalian herbivore

Many plants produce structural defenses to deter feeding by herbivores. However, many previous studies testing whether spines are effective at defending against mammalian herbivores have produced equivocal results. These ambiguous results are hypothesized to be due to herbivore counter‐adaptations. We investigated potential counter‐adaptations in a population of white‐throated woodrats Neotoma albigua that specialize on cactus by investigating feeding behavior and preference for cacti varying in spinescence. Neotoma albigula exhibited a unique behavior of clipping cactus spines, which renders these defenses ineffective. Strikingly, these woodrats chose to collect spiny cacti over experimentally de‐spined cacti, demonstrating that spines act as a proximal cue that attracts woodrats. This attraction is likely due to the higher protein and lower fiber content of spiny cacti compared to naturally non‐spiny cacti. Thus, the ‘defensive’ spines of cacti are ineffective against a specialist herbivore and instead serve as an indicator of nutritional quality that promotes herbivory. Our results support the ‘rule‐of‐thumb’ hypothesis of foraging, which states that herbivores forage according to obvious visual cues that are indicative of nutritional content, rather than sampling nutrient composition of plants. We propose that specialist herbivores are unique systems in which to study other counter‐adaptations to structural defenses and ‘rule‐of‐thumb’ foraging strategies.     

Woodrat (<Emphasis Type="Italic">Neotoma</Emphasis>) herbivores maintain nitrogen balance on a low-nitrogen,high-phenolic forage, <Emphasis Type="Italic">Juniperus monosperma</Emphasis>

The acquisition of adequate quantities of nitrogen is a challenge for herbivorous vertebrates because many plants are in low nitrogen and contain secondary metabolites that reduce nitrogen digestibility. To investigate whether herbivores maintain nitrogen balance on plant diets low in nitrogen and high in secondary compounds, we studied the effect of juniper (Juniperus monosperma) ingestion on the nitrogen balance of two species of herbivorous woodrats (Neotoma stephensi and N. albigula). These woodrat species feed on the foliage of juniper: N. stephensi is a juniper specialist, whereas N. albigula is a generalist that incorporates some juniper in its diet. Based on the nitrogen contents of the natural diets of these woodrats, we predicted that the generalist would be in negative nitrogen balance on a juniper diet whereas the specialist would not be affected. We found that both species of woodrat had low-nitrogen requirements (334.2 mg N/kg^0.75/day) and that a diet of 50% juniper did not result in negative nitrogen balance for either species. However, excretion patterns of nitrogen were altered; on the 50% juniper diet, fecal nitrogen losses increased ~38% and urinary nitrogen losses were half that of the control diet. The results suggest that absorption and detoxification of juniper secondary compounds may be more important for restricting juniper intake by the generalist than nitrogen imbalance.     

Differential expression and activity of catechol-O-methyl transferase (COMT) in a generalist (Neotoma albigula) and juniper specialist (Neotoma stephensi) woodrat

Mammalian herbivores, particularly dietary specialists must have an efficient means to metabolize the high doses of plant secondary compounds they consume. We found previously that Neotoma stephensi, a juniper specialist, upregulated catechol-O-methyl transferase (COMT) mRNA almost seven fold in response to an ecologically relevant diet (70% juniper). To further investigate the relevance of this enzyme with respect to juniper metabolism, we compared the protein expression, activity and kinetics of the two forms of COMT, soluble (S-COMT) and membrane bound (MB-COMT), in the blood, kidneys and liver of N. stephensi on its natural juniper diet to that of N. stephensi fed an experimental diet of 70% juniper as well as a non-toxic control diet under laboratory conditions. In addition, we compared these results to that of Neotoma albigula, a generalist species, which consumes a diet of 25% juniper in the wild. The specialist consuming juniper under both field and laboratory conditions had increased S-COMT expression and activity in their livers and kidneys, and increased S-COMT activity in their blood compared to the specialist and generalist fed the control diet. The specialist showed expression and activity of S-COMT in their kidneys that was as high as or higher than that in their livers. The generalist had an elevated V_max for MB-COMT compared to the specialist that resulted in higher activity for MB-COMT than the specialist despite lower expression of MB-COMT in the generalist's livers and kidneys. This high activity MB-COMT may be in part responsible for differences in the behaviors of the generalist compared to the specialist. We conclude that S-COMT is important in the specialist's ability to consume high levels of juniper.     

Ingestion of juniper foliage reduces metabolic rates in woodrat (Neotoma) herbivores

Ingestion of plant secondary compounds by herbivores is predicted to increase resting or basal metabolic rates. We tested this hypothesis with two species of woodrat herbivores, Neotoma stephensi and Neotoma albigula, consuming diets of juniper (Juniperus monosperma), which is rich in plant secondary compounds. In nature, N. stephensi specializes on juniper, whereas N. albigula consumes a variety of plant species including juniper. We measured resting metabolic rates (RMR) of woodrats on control, 25% juniper and a treatment containing the maximum tolerable dose of juniper (50% juniper for N. albigula and 70% juniper for N. stephensi). Ingestion of a juniper diet resulted in decreased RMR in both species of woodrats. We propose several potential mechanisms for metabolic depression of Neotoma on juniper diets. Our novel results underscore the need for more studies utilizing plant-based diets to determine the general effect of plant secondary compounds on metabolic rates of herbivores.     

Gut microbes of mammalian herbivores facilitate intake of plant toxins

The foraging ecology of mammalian herbivores is strongly shaped by plant secondary compounds (PSCs) that defend plants against herbivory. Conventional wisdom holds that gut microbes facilitate the ingestion of toxic plants; however, this notion lacks empirical evidence. We investigated the gut microbiota of desert woodrats (Neotoma lepida), some populations of which specialise on highly toxic creosote bush (Larrea tridentata). Here, we demonstrate that gut microbes are crucial in allowing herbivores to consume toxic plants. Creosote toxins altered the population structure of the gut microbiome to facilitate an increase in abundance of genes that metabolise toxic compounds. In addition, woodrats were unable to consume creosote toxins after the microbiota was disrupted with antibiotics. Last, ingestion of toxins by naïve hosts was increased through microbial transplants from experienced donors. These results demonstrate that microbes can enhance the ability of hosts to consume PSCs and therefore expand the dietary niche breadth of mammalian herbivores.     

The dilemma of foraging herbivores: dealing with food and fear

For foraging herbivores, both food quality and predation risk vary across the landscape. Animals should avoid low-quality food patches in favour of high-quality ones, and seek safe patches while avoiding risky ones. Herbivores often face the foraging dilemma, however, of choosing between high-quality food in risky places or low-quality food in safe places. Here, we explore how and why the interaction between food quality and predation risk affects foraging decisions of mammalian herbivores, focusing on browsers confronting plant toxins in a landscape of fear. We draw together themes of plant–herbivore and predator–prey interactions, and the roles of animal ecophysiology, behaviour and personality. The response of herbivores to the dual costs of food and fear depends on the interplay of physiology and behaviour. We discuss detoxification physiology in dealing with plant toxins, and stress physiology associated with perceived predation risk. We argue that behaviour is the interface enabling herbivores to stay or quit food patches in response to their physiological tolerance to these risks. We hypothesise that generalist and specialist herbivores perceive the relative costs of plant defence and predation risk differently and intra-specifically, individuals with different personalities and physiologies should do so too, creating individualised landscapes of food and fear. We explore the ecological significance and emergent impacts of these individual-based foraging outcomes on populations and communities, and offer predictions that can be clearly tested. In doing so, we provide an integrated platform advancing herbivore foraging theory with food quality and predation risk at its core.     

Role of cytochrome P450 2B sequence variation and gene copy number in facilitating dietary specialization in mammalian herbivores

Theory postulates that dietary specialization in mammalian herbivores is enabled by a specialized set of liver enzymes that process the high concentrations of similar plant secondary metabolites (PSMs) in the diets of specialists. To investigate whether qualitative and quantitative differences in detoxification mechanisms distinguish dietary specialists from generalists, we compared the sequence diversity and gene copy number of detoxification enzymes in two woodrat species: a generalist, the white‐throated woodrat (Neotoma albigula) and a juniper specialist, Stephens’ woodrat (N. stephensi). We focused on enzymes in the cytochrome P450 subfamily 2B (CYP2B), because previous research suggests this subfamily plays a key role in the processing of PSMs. For both woodrat species, we obtained and sequenced CYP2B cDNA, generated CYP2B phylogenies, estimated CYP2B gene copy number and created a homology model of the active site. We found that the specialist possessed on average ~5 more CYP2B gene copies than the generalist, but the specialist's CYP2B sequences were less diverse. Phylogenetic analysis of putative CYP2B homologs resolved woodrat species as reciprocally monophyletic and suggested evolutionary convergence of distinct homologs on similar key amino acid residues in both species. Homology modelling of the CYP2B enzyme suggests that interspecific differences in substrate preference and function likely result from amino acid differences in the enzyme active site. The characteristics of CYP2B in the specialist, that is greater gene copy number coupled with less sequence variation, are consistent with specialization to a narrow range of dietary toxins.     

Plant secondary metabolites and vertebrate herbivores--from physiological regulation to ecosystem function

Plant secondary metabolites can constrain the diet of vertebrates and these effects can flow through to community dynamics. Recent studies have moved beyond attempting to correlate diet choice with secondary metabolite profiles and instead focus on mechanisms that animals use to detect toxins and to regulate their intake and absorption. These include molecularly determined taste specificity, serotonin-mediated learning and the control of toxin absorption by permeability-glycoproteins. Focus on the detoxification pathways employed by specialist and generalist herbivores has facilitated explicit tests of the long-standing hypothesis that detoxification rates limit feeding. Understanding the molecular basis of differences amongst species in their tolerance of plant secondary metabolites opens many opportunities for understanding the evolutionary history of interactions between vertebrates and their food plants.     

Evidence for local specialization in a generalist mammalian herbivore, Neotoma fuscipes

Herbivores with very plastic dietary requirements, or so-called generalist species, can include individuals that develop specialized feeding habits through their experience with local chemically-defended plants. Local specialization has important implications for understanding a variety of ecological and evolutionary dynamics. However, the extent to which individuals within a generalist species specialize on local plants and the consequences of such specialization remain poorly understood, especially in non-insect herbivores. To better understand this phenomenon, we determined the diet and food preferences of a generalist mammalian herbivore, the dusky-footed woodrat ( Neotoma fuscipes ), in two adjacent but distinct plant communities. Based on a combination of cafeteria trials and stable isotope analyses (δ¹³C and δ¹⁵N), our results indicate that woodrats display preferences for local plants and tend to avoid novel chemically-defended plants. Moreover, both methodologies support the conclusion that individual woodrats are dietary specialists restricting their diets to only a few (2-3) of the available plant species. In juniper woodland, woodrats prefer western juniper ( Juniperus occidentalis ), while less than one km away in mixed-coniferous forest, woodrats prefer incense cedar ( Calocedrus decurrens ). Both plants contain high levels of plant secondary compounds that require detoxification mechanisms within consumers. Therefore, preferences are likely indicative of underlying physiological adaptations that could promote further behavioral, physiological and ultimately genetic differences between woodrats in different habitats. This study provides additional evidence for local specialization and urges caution when using the term generalist to characterize feeding behaviors at the individual level.     

The tri-trophic interactions hypothesis: interactive effects of host plant quality, diet breadth and natural enemies on herbivores

Several influential hypotheses in plant-herbivore and herbivore-predator interactions consider the interactive effects of plant quality, herbivore diet breadth, and predation on herbivore performance. Yet individually and collectively, these hypotheses fail to address the simultaneous influence of all three factors. Here we review existing hypotheses, and propose the tri-trophic interactions (TTI) hypothesis to consolidate and integrate their predictions. The TTI hypothesis predicts that dietary specialist herbivores (as compared to generalists) should escape predators and be competitively dominant due to faster growth rates, and that such differences should be greater on low quality (as compared to high quality) host plants. To provide a preliminary test of these predictions, we conducted an empirical study comparing the effects of plant (Baccharis salicifolia) quality and predators between a specialist (Uroleucon macolai) and a generalist (Aphis gossypii) aphid herbivore. Consistent with predictions, these three factors interactively determine herbivore performance in ways not addressed by existing hypotheses. Compared to the specialist, the generalist was less fecund, competitively inferior, and more sensitive to low plant quality. Correspondingly, predator effects were contingent upon plant quality only for the generalist. Contrary to predictions, predator effects were weaker for the generalist and on low-quality plants, likely due to density-dependent benefits provided to the generalist by mutualist ants. Because the TTI hypothesis predicts the superior performance of specialists, mutualist ants may be critical to A. gossypii persistence under competition from U. macolai. In summary, the integrative nature of the TTI hypothesis offers novel insight into the determinants of plant-herbivore and herbivore-predator interactions and the coexistence of specialist and generalist herbivores.