Jennifer G. Klutsch

Direction of interaction between mountain pine beetle ( Dendroctonus ponderosae ) and resource-sharing wood-boring beetles depends on plant parasite infection

AbstractPlant pathogens can have cascading consequences on insect herbivores, though whether they alter competition among resource-sharing insect herbivores is unknown. We experimentally tested whether the infection of a plant pathogen, the parasitic plant dwarf mistletoe (Arceuthobium americanum), on jack pine (Pinus banksiana) altered the competitive interactions among two groups of beetles sharing the same resources: wood-boring beetles (Coleoptera: Cerambycidae) and the invasive mountain pine beetle (Dendroctonus ponderosae) (Coleoptera: Curculionidae). We were particularly interested in identifying potential mechanisms governing the direction of interactions (from competition to facilitation) between the two beetle groups. At the lowest and highest disease severity, wood-boring beetles increased their consumption rate relative to feeding levels at moderate severity. The performance (brood production and feeding) of mountain pine beetle was negatively associated with wood-boring beetle feeding and disease severity when they were reared separately. However, when both wood-boring beetles and high severity of plant pathogen infection occurred together, mountain pine beetle escaped from competition and improved its performance (increased brood production and feeding). Species-specific responses to changes in tree defense compounds and quality of resources (available phloem) were likely mechanisms driving this change of interactions between the two beetle groups. This is the first study demonstrating that a parasitic plant can be an important force in mediating competition among resource-sharing subcortical insect herbivores.

Defence syndromes in lodgepole – whitebark pine ecosystems relate to degree of historical exposure to mountain pine beetles

Warming climate is allowing tree-killing bark beetles to expand their ranges and access naïve and semi-naïve conifers. Conifers respond to attack using complex mixtures of chemical defences that can impede beetle success, but beetles exploit some compounds for host location and communication. Outcomes of changing relationships will depend on concentrations and compositions of multiple host compounds, which are largely unknown. We analysed constitutive and induced chemistries of Dendroctonus ponderosaes primary historical host, Pinus contorta, and Pinus albicaulis, a high-elevation species whose encounters with this beetle are transitioning from intermittent to continuous. We quantified multiple classes of terpenes, phenolics, carbohydrates and minerals. Pinus contorta had higher constitutive allocation to, and generally stronger inducibility of, compounds that resist these beetle-fungal complexes. Pinus albicaulis contained higher proportions of specific monoterpenes that enhance pheromone communication, and lower induction of pheromone inhibitors. Induced P. contorta increased insecticidal and fungicidal compounds simultaneously, whereas P. albicaulis responses against these agents were inverse. Induced terpene accumulation was accompanied by decreased non-structural carbohydrates, primarily sugars, in P. contorta, but not P. albicaulis, which contained primarily starches. These results show some host species with continuous exposure to bark beetles have more thoroughly integrated defence syndromes than less-continuously exposed host species.

Weathering the storm: how lodgepole pine trees survive mountain pine beetle outbreaks

Recent mountain pine beetle outbreaks in western North America killed millions of lodgepole pine trees, leaving few survivors. However, the mechanism underlying the ability of trees to survive bark beetle outbreaks is unknown, but likely involve phytochemicals such as monoterpenes and fatty acids that can drive beetle aggregation and colonization on their hosts. Thus, we conducted a field survey of beetle-resistant lodgepole pine (Pinus contorta) trees to retrospectively deduce whether these phytochemicals underlie their survival by comparing their chemistry to that of non-attacked trees in the same stands. We also compared beetle attack characteristics between resistant and beetle-killed trees. Beetle-killed trees had more beetle attacks and longer ovipositional galleries than resistant trees, which also lacked the larval establishment found in beetle-killed trees. Resistant trees contained high amounts of toxic and attraction-inhibitive compounds and low amounts of pheromone-precursor and synergist compounds. During beetle host aggregation and colonization, these compounds likely served three critical roles in tree survival. First, low amounts of pheromone-precursor (α-pinene) and synergist (mycrene, terpinolene) compounds reduced or prevented beetles from attracting conspecifics to residual trees. Second, high amounts of 4-allyanisole further inhibited beetle attraction to its pheromone. Finally, high amounts of toxic limonene, 3-carene, 4-allyanisole, α-linolenic acid, and linoleic acid inhibited beetle gallery establishment and oviposition. We conclude that the variation of chemotypic expression of local plant populations can have profound ecological consequences including survival during insect outbreaks.

Rapid monoterpene induction promotes the susceptibility of a novel host pine to mountain pine beetle colonization but not to beetle-vectored fungi

Chemical induction can drive tree susceptibility to and host range expansions of attacking insects and fungi. Recently, mountain pine beetle (Dendroctonus ponderosae Hopkins; MPB) has expanded its host range from its historic host lodgepole pine (Pinus contorta var. latifolia Douglas ex Loudon) to jack pine (Pinus banksiana Lamb) in western Canada. Beetle success in jack pine forests likely depends upon the suitability of tree chemistry to MPB and its symbiotic phytopathogenic fungi. In particular, how rapid induced defenses of jack pine affect MPB colonization and the beetles symbionts is unknown. In the field, we characterized and compared differences in rapid induced phloem monoterpenes between lodgepole and jack pines in response to various densities of Grosmannia clavigera (Robinson-Jeffery and Davidson)-a MPB symbiotic fungus used to simulate beetle attack-inoculations. Overall, lodgepole pine had higher limonene and myrcene, but lower α-pinene, concentrations than jack pine. However, myrcene concentrations in jack pine increased with inoculation density, while that in lodgepole pine did not respond to density treatments. We compared the growth and reproduction of MPBs symbiotic fungi, G. clavigera, Ophiostoma montium (Rumford) von Arx and Leptographium longiclavatum Lee, Kim and Breuil, grown on media amended with myrcene, α-pinene and limonene at concentrations reflecting two induction levels from each pine species. Myrcene and α-pinene amendments inhibited the growth but stimulated the reproduction of G. clavigera, whereas limonene stimulated its growth while inhibiting its reproduction. However, the growth and reproduction of the other fungi were generally stimulated by monoterpene amendments. Overall, our results suggest that jack pine rapid induction could promote MPB aggregation due to high levels of α-pinene (pheromone precursor), a positive feedback of myrcene (pheromone synergist) and low levels of limonene (resistance). Jack pine is likely as susceptible to MPB-vectored fungi as lodgepole pine, indicating that jack pine induction will likely not adversely affect symbiont activities enough to inhibit the invasion of MPB into jack pine forests.

Fungal Volatiles Can Act as Carbon Sources and Semiochemicals to Mediate Interspecific Interactions Among Bark Beetle-Associated Fungal Symbionts.

Mountain pine beetle (Dendroctonus ponderosae) has killed millions of hectares of pine forests in western North America. Beetle success is dependent upon a community of symbiotic fungi comprised of Grosmannia clavigera, Ophiostoma montium, and Leptographium longiclavatum. Factors regulating the dynamics of this community during pine infection are largely unknown. However, fungal volatile organic compounds (FVOCs) help shape fungal interactions in model and agricultural systems and thus may be important drivers of interactions among bark beetle-associated fungi. We investigated whether FVOCs can mediate interspecific interactions among mountain pine beetle’s fungal symbionts by affecting fungal growth and reproduction. Headspace volatiles were collected and identified to determine species-specific volatile profiles. Interspecific effects of volatiles on fungal growth and conidia production were assessed by pairing physically-separated fungal cultures grown either on a carbon-poor or -rich substrate, inside a shared-headspace environment. Fungal VOC profiles differed by species and influenced the growth and/or conidia production of the other species. Further, our results showed that FVOCs can be used as carbon sources for fungi developing on carbon-poor substrates. This is the first report demonstrating that FVOCs can drive interactions among bark beetle fungal symbionts, and thus are important factors in beetle attack success.

A Native Parasitic Plant Systemically Induces Resistance in Jack Pine to a Fungal Symbiont of Invasive Mountain Pine Beetle

Conifer trees resist pest and pathogen attacks by complex defense responses involving different classes of defense compounds. However, it is unknown whether prior infection by biotrophic pathogens can lead to subsequent resistance to necrotrophic pathogens in conifers. We used the infection of jack pine, Pinus banksiana, by a common biotrophic pathogen dwarf mistletoe, Arceuthobium americanum, to investigate induced resistance to a necrotrophic fungus, Grosmannia clavigera, associated with the mountain pine beetle, Dendroctonus ponderosae. Dwarf mistletoe infection had a non-linear, systemic effect on monoterpene production, with increasing concentrations at moderate infection levels and decreasing concentrations at high infection levels. Inoculation with G. clavigera resulted in 33 times higher monoterpene concentrations and half the level of phenolics in the necrotic lesions compared to uninoculated control trees. Monoterpene production following dwarf mistletoe infection seemed to result in systemic induced resistance, as trees with moderate disease severity were most resistant to G. clavigera, as evident from shorter lesion lengths. Furthermore, trees with moderate disease severity had the highest systemic but lowest local induction of α-pinene after G. clavigera inoculation, suggesting a possible tradeoff between systemically- and locally-induced defenses. The opposing effects to inoculation by G. clavigera on monoterpene and phenolic levels may indicate the potential for biosynthetic tradeoffs by the tree between these two major defense classes. Our results demonstrate that interactions between a biotrophic parasitic plant and a necrotrophic fungus may impact mountain pine beetle establishment in novel jack pine forests through systemic effects mediated by the coordination of jack pine defense chemicals.

Dwarf mistletoe infection in jack pine alters growth–defense relationships

Trees utilize a combination of chemical and anatomical defenses against a myriad of attacking organisms. However, persistent pathogen infection that alters resource acquisition may impact growth and defense relationships, which could have consequences for tree resistance. We characterized systemic chemical and anatomical changes in jack pine (Pinus banksiana) in response to infection by the parasitic plant dwarf mistletoe (Arceuthobium americanum) and identified how the growth-defense relationship is altered due to infection severity. Our study found that the growth and defense relationship in jack pine was altered due to infection and that chemical defenses in the phloem received a relatively higher priority than radial growth and anatomical defenses. Chemical defenses in the phloem had a non-linear relationship with infection severity with increasing concentrations of monoterpenes in trees with moderate infection and decreasing concentrations at high infection. In contrast, both radial growth and vertical resin duct production decreased with increasing infection severity. While constitutive resin duct counts and many monoterpene compound concentrations were positively correlated, this relationship was not maintained in infected trees. Furthermore, radial growth and basal area increment was positively correlated with resin duct production and monoterpene concentration in non-infected trees but had fewer relationships in severely infected trees. We conclude that while both chemical and anatomical defenses may be used as indicators for potential resistance to biotic stress in pines, changes in resource allocation patterns between these defenses after infection will likely have consequences on tree resistance to subsequent biotic attacks.

Anatomical defenses against bark beetles relate to degree of historical exposure between species and are allocated independently of chemical defenses within trees: Resin ducts of lodgepole and whitebark pine

Conifers possess chemical and anatomical defences against tree-killing bark beetles that feed in their phloem. Resins accumulating at attack sites can delay and entomb beetles while toxins reach lethal levels. Trees with high concentrations of metabolites active against bark beetle-microbial complexes, and more extensive resin ducts, achieve greater survival. It is unknown if and how conifers integrate chemical and anatomical components of defence or how these capabilities vary with historical exposure. We compared linkages between phloem chemistry and tree ring anatomy of two mountain pine beetle hosts. Lodgepole pine, a mid-elevation species, has had extensive, continual contact with this herbivore, whereas high-elevation whitebark pines have historically had intermittent exposure that is increasing with warming climate. Lodgepole pine had more and larger resin ducts. In both species, anatomical defences were positively related to tree growth and nutrients. Within-tree constitutive and induced concentrations of compounds bioactive against bark beetles and symbionts were largely unrelated to resin duct abundance and size. Fewer anatomical defences in the semi-naïve compared with the continually exposed host concurs with directional differences in chemical defences. Partially uncoupling chemical and morphological antiherbivore traits may enable trees to confront beetles with more diverse defence permutations that interact to resist attack.

Drought stress leads to systemic induced susceptibility to a nectrotrophic fungus associated with mountain pine beetle in Pinus banksiana seedlings

Conifers have complex defense responses to initial attacks by insects and pathogens that can have cascading effects on success of subsequent colonizers. However, drought can affect a plant’s ability to respond to biotic agents by potentially altering the resources needed for the energetically costly production of induced defense chemicals. We investigated the impact of reduced water on induced chemical defenses of jack pine (Pinus banksiana) seedlings from initial attack by biotic agents and resistance to subsequent challenge inoculation with a pathogenic fungal associate of mountain pine beetle (Dendroctonus ponderosae), Grosmannia clavigera. Applications of phytohormones (methyl salicylate and methyl jasmonate) and G. clavigera were used for initial induction of defenses. Monoterpene concentrations varied with initial induction from fungal and phytohormone application while watering treatment had no effect. Seedlings treated with G. clavigera and methyl jasmonate had the greatest monoterpene concentrations compared to the control and methyl salicylate-treated seedlings. However, the monoterpene response to the challenge inoculation varied with watering treatments, not with prior induction treatments, with lower monoterpene concentrations in fungal lesions on seedlings in the low to moderate watering treatments compared to normal watering treatment. Furthermore, prior induction from phytohormones resulted in systemic cross-induction of resistance to G. clavigera under normal watering treatment but susceptibility under low watering treatment. Seedlings stressed by low water conditions, which also had lower stomatal conductance than seedlings in the normal watering treatment, likely allocated resources to initial defense response but were left unable to acquire further resources for subsequent responses. Our results demonstrate that drought can affect interactions among tree-infesting organisms through systemic cross-induction of susceptibility.