Transcriptional profile and differential fitness in a specialist milkweed insect across host plants varying in toxicity

Interactions between plants and herbivorous insects have been models for theories of specialization and co‐evolution for over a century. Phytochemicals govern many aspects of these interactions and have fostered the evolution of adaptations by insects to tolerate or even specialize on plant defensive chemistry. While genomic approaches are providing new insights into the genes and mechanisms insect specialists employ to tolerate plant secondary metabolites, open questions remain about the evolution and conservation of insect counterdefences, how insects respond to the diversity defences mounted by their host plants, and the costs and benefits of resistance and tolerance to plant defences in natural ecological communities. Using a milkweed‐specialist aphid (Aphis nerii) model, we test the effects of host plant species with increased toxicity, likely driven primarily by increased secondary metabolites, on aphid life history traits and whole‐body gene expression. We show that more toxic plant species have a negative effect on aphid development and lifetime fecundity. When feeding on more toxic host plants with higher levels of secondary metabolites, aphids regulate a narrow, targeted set of genes, including those involved in canonical detoxification processes (e.g., cytochrome P450s, hydrolases, UDP‐glucuronosyltransferases and ABC transporters). These results indicate that A. nerii marshal a variety of metabolic detoxification mechanisms to circumvent milkweed toxicity and facilitate host plant specialization, yet, despite these detoxification mechanisms, aphids experience reduced fitness when feeding on more toxic host plants. Disentangling how specialist insects respond to challenging host plants is a pivotal step in understanding the evolution of specialized diet breadths.     

Genetic differentiation associated with host plants and geography among six widespread species of South American Blepharoneura fruit flies (Tephritidae)

Tropical herbivorous insects are astonishingly diverse, and many are highly host‐specific. Much evidence suggests that herbivorous insect diversity is a function of host plant diversity; yet, the diversity of some lineages exceeds the diversity of plants. Although most species of herbivorous fruit flies in the Neotropical genus Blepharoneura are strongly host‐specific (they deposit their eggs in a single host plant species and flower sex), some species are collected from multiple hosts or flowers and these may represent examples of lineages that are diversifying via changes in host use. Here, we investigate patterns of diversification within six geographically widespread Blepharoneura species that have been collected and reared from at least two host plant species or host plant parts. We use microsatellites to (1) test for evidence of local genetic differentiation associated with different sympatric hosts (different plant species or flower sexes) and (2) examine geographic patterns of genetic differentiation across multiple South American collection sites. In four of the six fly species, we find evidence of local genetic differences between flies collected from different hosts. All six species show evidence of geographic structure, with consistent differences between flies collected in the Guiana Shield and flies collected in Amazonia. Continent‐wide analyses reveal – in all but one instance – that genetically differentiated flies collected in sympatry from different host species or different sex flowers are not one another's closest relatives, indicating that genetic differences often arise in allopatry before, or at least coincident with, the evolution of novel host use.     

Evidence for ecological speciation via a host shift in the holly leaf miner,Phytomyza glabricola (Diptera: Agromyzidae)

Evolutionary radiations have been well documented in plants and insects, and natural selection may often underly these radiations. If radiations are adaptive, the diversity of species could be due to ecological speciation in these lineages. Agromyzid flies exhibit patterns of repeated host‐associated radiations. We investigated whether host‐associated population divergence and evidence of divergent selection exist in the leaf miner Phytomyza glabricola on its sympatric host plants, the holly species, Ilex coriacea and I. glabra. Using AFLPs and nuclear sequence data, we found substantial genetic divergence between host‐associated populations of these flies throughout their geographic range. Genome scans using the AFLP data identified 13 loci under divergent selection, consistent with processes of ecological speciation. EF‐1α data suggest that I. glabra is the original host of P. glabricola and that I. coriacea is the novel host, but the AFLP data are ambiguous with regard to directionality of the host shift.     

Cascading effects of N input on tritrophic (plant–aphid–parasitoid) interactions

Because N is frequently the most limiting mineral macronutrient for plants in terrestrial ecosystems, modulating N input may have ecological consequences through trophic levels. Thus, in agro‐ecosystems, the success of natural enemies may depend not only from their herbivorous hosts but also from the host plant whose qualities may be modulated by N input. We manipulated foliar N concentrations by providing to Camelina sativa plants three different nitrogen rates (control, optimal, and excessive). We examined how the altered host‐plant nutritional quality influenced the performances of two aphid species, the generalist green peach aphid, Myzus persicae, and the specialist cabbage aphid, Brevicoryne brassicae, and their common parasitoid Diaeretiella rapae. Both N inputs led to increased N concentrations in the plants but induced contrasted concentrations within aphid bodies depending on the species. Compared to the control, plant biomass increased when receiving the optimal N treatment but decreased under the excessive treatment. Performances of M. persicae improved under the optimal treatment compared to the control and excessive treatments whereas B. brassicae parameters declined following the excessive N treatment. In no‐choice trials, emergence rates of D. rapae developing in M. persicae were higher on both optimum and excessive N treatments, whereas they remained stable whatever the treatment when developing in B. brassicae. Size of emerging D. rapae females was positively affected by the treatment only when it developed in M. persicae on the excessive N treatment. This work showed that contrary to an optimal N treatment, when N was delivered in excess, plant suitability was reduced and consequently affected negatively aphid parameters. Surprisingly, these negative effects resulted in no or positive consequences on parasitoid parameters, suggesting a buffered effect at the third trophic level. Host N content, host suitability, and dietary specialization appear to be major factors explaining the functioning of our studied system.     

Oviposition site preference and developmental performance of a gall‐inducing leafhopper on galled and non‐galled host plants

Oviposition preferences of herbivorous insects affect offspring performance. Both positive and negative links between oviposition preference and offspring performance have been reported for many species. A gall‐inducing leafhopper, Cicadulina bipunctata Melichar (Hemiptera: Cicadellidae), feeds on various Poaceae plants and induces galls of enhanced nutritional value for their offspring. Although gall induction by C. bipunctata improves nymphal performance, the oviposition preference of females between galled and non‐galled host plants is still unclear. In this paper, the nymphal performance and oviposition and feeding‐site preference of C. bipunctata were investigated using galled wheat, Triticum aestivum L., and non‐galled barley, Hordeum vulgare L., as host plants. The survival rate of C. bipunctata on wheat was significantly higher than on barley. In the choice test, significantly more eggs were laid into barley, whereas the number of eggs deposited on both hosts was not significantly different in the no‐choice test. The number of settling individuals per leaf area was not significantly different between wheat and barley, suggesting no clear preference for oviposition between these plants. Experience as a nymph with a growing host did not affect oviposition preference as adult female. The inconsistent correspondence between offspring performance and oviposition preference of C. bipunctata may reflect the high mobility of nymphs and/or differences in leaf area between host plants. The results indicate that the previous finding that oviposition preference and offspring performance are not always positively correlated in herbivorous insects is applicable to gall‐inducing insects.     

Insect oviposition preference between Epichloë‐symbiotic and Epichloë‐free grasses does not necessarily reflect larval performance

Variation in plant communities is likely to modulate the feeding and oviposition behavior of herbivorous insects, and plant‐associated microbes are largely ignored in this context. Here, we take into account that insects feeding on grasses commonly encounter systemic and vertically transmitted (via seeds) fungal Epichloë endophytes, which are regarded as defensive grass mutualists. Defensive mutualism is primarily attributable to alkaloids of fungal origin. To study the effects of Epichloë on insect behavior and performance, we selected wild tall fescue (Festuca arundinacea) and red fescue (Festuca rubra) as grass–endophyte models. The plants used either harbored the systemic endophyte (E+) or were endophyte‐free (E?). As a model herbivore, we selected the Coenonympha hero butterfly feeding on grasses as larvae. We examined both oviposition and feeding preferences of the herbivore as well as larval performance in relation to the presence of Epichloë endophytes in the plants. Our findings did not clearly support the female's oviposition preference to reflect the performance of her offspring. First, the preference responses depended greatly on the grass–endophyte symbiotum. In F. arundinacea, C. hero females preferred E+ individuals in oviposition‐choice tests, whereas in F. rubra, the endophytes may decrease exploitation, as both C. hero adults and larvae preferred E? grasses. Second, the endophytes had no effect on larval performance. Overall, F. arundinacea was an inferior host for C. hero larvae. However, the attraction of C. hero females to E+ may not be maladaptive if these plants constitute a favorable oviposition substrate for reasons other than the plants' nutritional quality. For example, rougher surface of E+ plant may physically facilitate the attachment of eggs, or the plants offer greater protection from natural enemies. Our results highlight the importance of considering the preference of herbivorous insects in studies involving the endophyte‐symbiotic grasses as host plants.     

Does anthropogenic nitrogen deposition induce phosphorus limitation in herbivorous insects?

Anthropogenic nitrogen deposition has shifted many ecosystems from nitrogen (N) limitation to phosphorus (P) limitation. Although well documented in plants, no study to date has explored whether N deposition exacerbates P limitation at higher trophic levels, or focused on the effects of induced plant P limitation on trophic interactions. Insect herbivores exhibit strict N : P homeostasis, and should therefore be very sensitive to variations in plant N : P stoichiometry and prone to experiencing deposition‐induced P limitation. In the current study, we investigated the effects of N deposition and P availability on a plant‐herbivorous insect system. Using common milkweed (Asclepias syriaca) and two of its specialist herbivores, the monarch caterpillar (Danaus plexippus) and milkweed aphid (Aphis asclepiadis) as our study system, we found that experimental N deposition caused P limitation in milkweed plants, but not in either insect species. However, the mechanisms for the lack of P limitation were different for each insect species. The body tissues of A. asclepiadis always exhibited higher N : P ratios than that of the host plant, suggesting that the N demand of this species exceeds P demand, even under high N deposition levels. For D. plexippus, P addition increased the production of latex, which is an important defense negatively affecting D. plexippus growth rate. As a result, we illustrate that P limitation of herbivores is not an inevitable consequence of anthropogenic N deposition in terrestrial systems. Rather, species‐specific demands for nutrients and the defensive responses of plants combine to determine the responses of herbivores to P availability under N deposition.     

Dietary specialization and the effects of plant species on potential multitrophic interactions of three species of nymphaline caterpillars

The diet breadth of insect herbivores influences their response to variation in plant quality, and these bitrophic interactions have implications for the higher‐level trophic interactions between herbivores and their natural enemies. In this comparative study, we examined the role of host plant species and plant secondary chemistry on the potential interactions between three species of nymphaline caterpillars and their natural enemies. The caterpillar species (all Lepidoptera: Nymphalidae) varied in their degree of specialization: the buckeye, Junonia coenia Hübner, is a specialist on plants that contain iridoid glycosides (IGs); the white peacock, Anartia jatrophae L., feeds on plants in five families, some of which contain IGs and some of which do not; and the painted lady, Vanessa cardui L., is a generalist, feeding on plants in at least 15 families. Each species was reared on leaves of an introduced host plant, Plantago lanceolata L. (Plantaginaceae), which produces two IGs, aucubin and catalpol, and on another plant species that is a common host plant. These alternate host plants were Plantago major L. (Plantaginaceae) for J. coenia, Bacopa monnieri (L.) Pennell (Plantaginaceae) for A. jatrophae, and Malva parviflora L. (Malvaceae) for V. cardui. We examined growth, sequestration, and immune response of these caterpillars on the different host plant species. Junonia coenia developed more rapidly and sequestered higher IG concentrations when reared on P. lanceolata, whereas both other species grew more slowly on P. lanceolata. Host plant did not influence immune response of J. coenia or A. jatrophae, whereas V. cardui immune response was weaker when reared on P. lanceolata. Junonia coenia was most efficient at IG sequestration and A. jatrophae was least efficient, when all three species were reared on P. lanceolata. These results indicate that diet breadth may play an important role in structuring tritrophic interactions, and this role should be further explored.     

Molecular biogeography and host relations of a parasitoid fly

Successful geographic range expansion by parasites and parasitoids may also require host range expansion. Thus, the evolutionary advantages of host specialization may trade off against the ability to exploit new host species encountered in new geographic regions. Here, we use molecular techniques and confirmed host records to examine biogeography, population divergence, and host flexibility of the parasitoid fly, Ormia ochracea (Bigot). Gravid females of this fly find their cricket hosts acoustically by eavesdropping on male cricket calling songs; these songs vary greatly among the known host species of crickets. Using both nuclear and mitochondrial genetic markers, we (a) describe the geographical distribution and subdivision of genetic variation in O. ochracea from across the continental United States, the Mexican states of Sonora and Oaxaca, and populations introduced to Hawaii; (b) demonstrate that the distribution of genetic variation among fly populations is consistent with a single widespread species with regional host specialization, rather than locally differentiated cryptic species; (c) identify the more‐probable source populations for the flies introduced to the Hawaiian islands; (d) examine genetic variation and substructure within Hawaii; (e) show that among‐population geographic, genetic, and host song distances are all correlated; and (f) discuss specialization and lability in host‐finding behavior in light of the diversity of cricket songs serving as host cues in different geographically separate populations.     

Experimental warming increases herbivory by leaf‐chewing insects in an alpine plant community

Climate warming is predicted to affect species and trophic interactions worldwide, and alpine ecosystems are expected to be especially sensitive to changes. In this study, we used two ongoing climate warming (open‐top chambers) experiments at Finse, southern Norway, to examine whether warming had an effect on herbivory by leaf‐chewing insects in an alpine Dryas heath community. We recorded feeding marks on the most common vascular plant species in warmed and control plots at two experimental sites at different elevations and carried out a brief inventory of insect herbivores. Experimental warming increased herbivory on Dryas octopetala and Bistorta vivipara. Dryas octopetala also experienced increased herbivory at the lower and warmer site, indicating an overall positive effect of warming, whereas B. vivipara experienced an increased herbivory at the colder and higher site indicating a mixed effect of warming. The Lepidoptera Zygaena exulans and Sympistis nigrita were the two most common leaf‐chewing insects in the Dryas heath. Based on the observed patterns of herbivory, the insects life cycles and feeding preferences, we argue that Z. exulans is the most important herbivore on B. vivipara, and S. nigrita the most important herbivore on D. octopetala. We conclude that if the degree of insect herbivory increases in a warmer world, as suggested by this study and others, complex interactions between plants, insects, and site‐specific conditions make it hard to predict overall effects on plant communities.     

Are the metabolomic responses to folivory of closely related plant species linked to macroevolutionary and plant–folivore coevolutionary processes?

The debate whether the coevolution of plants and insects or macroevolutionary processes (phylogeny) is the main driver determining the arsenal of molecular defensive compounds of plants remains unresolved. Attacks by herbivorous insects affect not only the composition of defensive compounds in plants but also the entire metabolome. Metabolomes are the final products of genotypes and are constrained by macroevolutionary processes, so closely related species should have similar metabolomic compositions and may respond in similar ways to attacks by folivores. We analyzed the elemental compositions and metabolomes of needles from three closely related Pinus species with distant coevolutionary histories with the caterpillar of the processionary moth respond similarly to its attack. All pines had different metabolomes and metabolic responses to herbivorous attack. The metabolomic variation among the species and the responses to folivory reflected their macroevolutionary relationships, with P. pinaster having the most divergent metabolome. The concentrations of terpenes were in the attacked trees supporting the hypothesis that herbivores avoid plant individuals with higher concentrations. Our results suggest that macroevolutionary history plays important roles in the metabolomic responses of these pine species to folivory, but plant–insect coevolution probably constrains those responses. Combinations of different evolutionary factors and trade‐offs are likely responsible for the different responses of each species to folivory, which is not necessarily exclusively linked to plant–insect coevolution.     

Changes in phytophagous insect host ranges following the invasion of their community: Long‐term data for fruit flies

The invasion of an established community by new species can trigger changes in community structure. Invasions often occur in phytophagous insect communities, the dynamics of which are driven by the structure of the host assemblage and the presence of competitors. In this study, we investigated how a community established through successive invasions changed over time, taking the last invasion as the reference. The community included four generalist and four specialist species of Tephritidae fruit flies. We analyzed a long‐term database recording observed numbers of flies per fruit for each species on 36 host plants, over 18 years, from 1991 to 2009. Community structure before the last invasion by Bactrocera zonata in 2000 was described in relation to host plant phylogeny and resource availability. Changes in the host range of each species after the arrival of B. zonata were then documented by calculating diversity indices. The flies in the community occupied three types of niches defined on the basis of plant phylogeny (generalists, Solanaceae specialist, and Cucurbitaceae specialists). After the arrival of B. zonata, no change in the host range of specialist species was observed. However, the host ranges of two generalist species, Ceratitis quilicii and Ceratitis capitata, tended to shrink, as shown by the decreases in species richness and host plant α‐diversity. Our study shows increased host specialization by generalist phytophagous insects in the field following the arrival of an invasive species sharing part of their resources. These findings could be used to improve predictions of new interactions between invaders and recipient communities.     

Contemporary evolution of host plant range expansion in an introduced herbivorous beetle Ophraella communa

Host range expansion of herbivorous insects is a key event in ecological speciation and insect pest management. However, the mechanistic processes are relatively unknown because it is difficult to observe the ongoing host range expansion in natural population. In this study, we focused on the ongoing host range expansion in introduced populations of the ragweed leaf beetle, Ophraella communa, to estimate the evolutionary process of host plant range expansion of a herbivorous insect. In the native range of North America, O. communa does not utilize Ambrosia trifida, as a host plant, but this plant is extensively utilized in the beetle's introduced range. Larval performance and adult preference experiments demonstrated that native O. communa beetles show better survival on host plant individuals from introduced plant populations than those from native plant populations and they also oviposit on the introduced plant, but not on the native plant. Introduced O. communa beetles showed significantly higher performance on and preference for both introduced and native A. trifida plants, when compared with native O. communa. These results indicate the contemporary evolution of host plant range expansion of introduced O. communa and suggest that the evolutionary change of both the host plant and the herbivorous insect involved in the host range expansion.     

Effects of quantitative and qualitative differences in volatiles from host‐ and non‐host‐infested maize on the attraction of the larval parasitoid Cotesia kariyai

Cotesia kariyai Watanabe (Hymenoptera: Braconidae) is a specialist larval parasitoid of Mythimna separata Walker (Lepidoptera: Noctuidae). Cotesia kariyai wasps use herbivore‐induced plant volatiles (HIPVs) to locate hosts. However, complex natural habitats are full of volatiles released by both herbivorous host‐ and non‐host‐infested plants at various levels of intensity. Therefore, the presence of non‐hosts may affect parasitoid decisions while foraging. Here, the host‐finding efficiency of naive C. kariyai from HIPVs influenced by host‐ and non‐host‐infested maize [Zea mays L. (Poaceae)] plants was investigated with a four‐arm olfactometer. Ostrinia furnacalis Guenée (Lepidoptera: Crambidae) was selected as a non‐host species. One unit (1 U) of host‐ or non‐host‐infested plant was prepared by infesting a potted plant with five host or seven non‐host larvae. In two‐choice bioassays, host‐infested plants fed upon by different numbers of larvae, and various units of host‐ and non‐host‐infested plants (infestation units; 1 U, 2 U, and 3 U) were arranged to examine the effects of differences in volatile quantity and quality on the olfactory responses of C. kariyai with the assumption that volatile quantity and quality changes with differences in numbers of insects and plants. Cotesia kariyai was found to perceive quantitative differences in volatiles from host‐infested plants, preferring larger quantities of volatiles from larger numbers of larvae or plants. Also, the parasitoids discriminated between healthy plants, host‐infested plants, and non‐host‐infested plants by recognising volatiles released from those plants. Cotesia kariyai showed a reduced preference for host‐induced volatiles, when larger numbers of non‐host‐infested plants were present. Therefore, quantitative and qualitative differences in volatiles from host‐ and non‐host‐infested plants appear to affect the decision of C. kariyai during host‐habitat searching in multiple tritrophic systems.     

Ecological adaptation and reproductive isolation in sympatry: genetic and phenotypic evidence for native host races of Rhagoletis pomonella

Ecological speciation with gene flow may be an important mode of diversification for phytophagous insects. The recent shift of Rhagoletis pomonella from its native host downy hawthorn (Crataegus mollis) to introduced apple (Malus domestica) in the northeastern United States is a classic example of sympatric host race formation. Here, we test whether R. pomonella has similarly formed host races on four native Crataegus species in the southern United States: western mayhaw (C. opaca), blueberry hawthorn (C. brachyacantha), southern red hawthorn (C. mollis var. texana) and green hawthorn (C. viridis). These four southern hosts differ from each other in their fruiting phenology and in the volatile compounds emitted from the surface of their fruits. These two traits form the basis of ecological reproductive isolation between downy hawthorn and apple flies in the north. We report evidence from microsatellite population surveys and eclosion studies supporting the existence of genetically differentiated and partially reproductively isolated host races of southern hawthorn flies. The results provide an example of host shifting and ecological divergence involving native plants and imply that speciation with gene flow may be commonly initiated in Rhagoletis when ecological opportunity presents itself.     

Specificity and seasonal prevalence of anther smut disease Microbotryum on sympatric Himalayan Silene species

Host sympatry provides opportunities for cross‐species disease transmission and compounded disease effects on host population and community structure. Using the Silene–Microbotryum interaction (the castrating anther smut disease), eleven Himalayan Silene species were assessed in regions of high host diversity to ascertain levels of pathogen specificity. We also investigated disease prevalence, seasonal dynamics of infection and flowering patterns in five co‐blooming Silene species. We identified several new Microbotryum lineages with varying degrees of specialization that is likely influenced by degrees of host divergence and ecological similarities (i.e. shared pollinator guilds). Affected species had 15%–40% of plants infected by anther smut. Flowering was seasonally overlapping among host species (except for the species pair S. asclepiadea and S. atrocastanea), but diseased flowering onset was earlier than healthy plants, leading to dramatic seasonal shifts in observed disease prevalence. Overlapping distributions and flowering provides opportunities for floral pathogen movement between host species, but host specialization may be constrained by the plant phylogenetic relatedness, adaptation to micro‐habitats and difference in pollinator/vector guilds.     

The paradoxical rarity of a fruit fly fungus attacking a broad range of hosts

Understanding the factors that determine the realized and potential distribution of a species requires knowledge of abiotic, physiological, limitations as well as ecological interactions. Fungi of the order Laboulbeniales specialize on arthropods and are typically thought to be highly specialized on a single species or closely related group of species. Because infections are almost exclusively transmitted through direct contact between the hosts, the host ecology, to a large extent, determines the distribution and occurrence of the fungus. We examined ~20,000 fruit flies (Diptera: Dacinae) collected in Malaysia, Sulawesi, Australia, and the Solomon Islands between 2017 and 2019 for fungal infections and found 197 infected flies across eight different Bactrocera species. Morphology and 1,363 bps of small subunit (18S) DNA sequences both support that the infections are from a single polyphagous fungal species Stigmatomyces dacinus—a known ectoparasite of these fruit flies. This leads to the question: why is S. dacinus rare, when its hosts are widespread and abundant? In addition, the hosts are all Bactrocera, a genus with ~480 species, but 37 Bactrocera species found sympatric with the hosts were never infected. Host‐selection does not appear to be phylogenetically correlated. These results suggest a hidden complexity in how different, but closely related, host species vary in their susceptibility, which somehow limits the abundance and dispersal capability of the fungus.     

Diadegma insulare development is altered by Plutella xylostella reared on water‐stressed host plants

Natural enemies of herbivores function in a multitrophic context, and their performance is directly or indirectly influenced by herbivores and their host plants. Very little is known about tritrophic interactions between host plants, pests and their parasitoids, particularly when the host plants are under any stress. Herbivores and their natural enemies’ response to plants under stress are diverse and variable. Therefore, in this study we investigated how diamondback moth, Plutella xylostella (L.), reared on water‐stressed host plants (Brassica napus L. and Sinapis alba L.) influenced the development of its larval parasitoid, Diadegma insulare (Cresson). No significant differences were observed in development of P. xylostella when reared on water‐stressed host plants. However, all results indicated that water stress had a strong effect on developmental parameters of D. insulare. Development of D. insulare was delayed when the parasitoid fed on P. xylostella, reared on stressed host plants. Egg to adult development of D. insulare was faster on non‐stressed B. napus than non‐stressed S. alba followed by stressed B. napus and S. alba. Female parasitoids were heavier on non‐stressed host plants than stressed counterparts. Furthermore, the parasitoid lived significantly longer on stressed B. napus. However, body size was not affected by water treatment. Most host plant parameters measured were significantly lower for water‐stressed than non‐stressed treatments. Results suggest that development of this important and effective P. xylostella parasitoid was influenced by both water stress and host plant species.