Lignin concentrations in phloem and outer bark are not associated with resistance to mountain pine beetle among high elevation pines.

A key component in understanding plant-insect interactions is the nature of host defenses. Research on defense traits among Pinus species has focused on specialized metabolites and axial resin ducts, but the role of lignin in defense within diverse systems is unclear. We investigated lignin levels in the outer bark and phloem of P. longaeva, P. balfouriana, and P. flexilis; tree species growing at high elevations in the western United States known to differ in susceptibility to mountain pine beetle (Dendroctonus ponderosae; MPB). Pinus longaeva and P. balfouriana are attacked by MPB less frequently than P. flexilis, and MPB brood production in P. longaeva is limited. Because greater lignification of feeding tissues has been shown to provide defense against bark beetles in related genera, such as Picea, we hypothesized that P. longaeva and P. balfouriana would have greater lignin concentrations than P. flexilis. Contrary to expectations, we found that the more MPB-susceptible P. flexilis had greater phloem lignin levels than the less susceptible P. longaeva and P. balfouriana. No differences in outer bark lignin levels among the species were found. We conclude that lignification in Pinus phloem and outer bark is likely not adaptive as a physical defense against MPB.

Prey removal in cotton crops next to woodland reveals periodic diurnal and nocturnal invertebrate predation gradients from the crop edge by birds and bats.

Factors influencing the efficacy of insectivorous vertebrates in providing natural pest control services inside crops at increasing distances from the crop edge are poorly understood. We investigated the identity of vertebrate predators (birds and bats) and removal of sentinel prey (mealworms and beetles) from experimental feeding trays in cotton crops using prey removal trials, camera traps and observations. More prey was removed during the day than at night, but prey removal was variable at the crop edge and dependent on the month (reflecting crop growth and cover) and time of day. Overall, the predation of mealworms and beetles was 1-times and 13-times greater during the day than night, respectively, with predation on mealworms 3-5 times greater during the day than night at the crop edge compared to 95 m inside the crop. Camera traps identified many insectivorous birds and bats over crops near the feeding trays, but there was no evidence of bats or small passerines removing experimental prey. A predation gradient from the crop edge was evident, but only in some months. This corresponded to the foraging preferences of open-space generalist predators (magpies) in low crop cover versus the shrubby habitat preferred by small passerines, likely facilitating foraging away from the crop edge later in the season. Our results are in line with Optimal Foraging Theory and suggest that predators trade-off foraging behaviour with predation risk at different distances from the crop edge and levels of crop cover. Understanding the optimal farm configuration to support insectivorous bird and bat populations can assist farmers to make informed decisions regarding in-crop natural pest control and maximise the predation services provided by farm biodiversity.

Selection for high levels of resistance to double-stranded RNA (dsRNA) in Colorado potato beetle (Leptinotarsa decemlineata Say) using non-transgenic foliar delivery.

Insecticidal double-stranded RNAs (dsRNAs) silence expression of vital genes by activating the RNA interference (RNAi) mechanism in insect cells. Despite high commercial interest in insecticidal dsRNA, information on resistance to dsRNA is scarce, particularly for dsRNA products with non-transgenic delivery (ex. foliar/topical application) nearing regulatory review. We report the development of the CEAS 300 population of Colorado potato beetle (Leptinotarsa decemlineata Say) (Coleoptera: Chrysomelidae) with > 11,100-fold resistance to a dsRNA targeting the V-ATPase subunit A gene after nine episodes of selection using non-transgenic delivery by foliar coating. Resistance was associated with lack of target gene down-regulation in CEAS 300 larvae and cross-resistance to another dsRNA target (COPI ß; Coatomer subunit beta). In contrast, CEAS 300 larvae showed very low (~ 4-fold) reduced susceptibility to the Cry3Aa insecticidal protein from Bacillus thuringiensis. Resistance to dsRNA in CEAS 300 is transmitted as an autosomal recessive trait and is polygenic. These data represent the first documented case of resistance in an insect pest with high pesticide resistance potential using dsRNA delivered through non-transgenic techniques. Information on the genetics of resistance and availability of dsRNA-resistant L. decemlineata guide the design of resistance management tools and allow research to identify resistance alleles and estimate resistance risks.

The complete life cycle of a Cretaceous beetle parasitoid.

Life cycles of parasites, particularly those with complex life histories and developmental pathways, are rarely preserved as fossils in total.1 The evidence is almost universally biased toward incomplete perspectives derived from a single sex or life stage.2,3 Here, we report a piece of Cretaceous Burmese amber that contains 28 males, a larviform female, and two longipede larvae of the wedge-shaped beetle Paleoripiphorus, and its potential cockroach host. Collectively, this fossil represents the complete series of free-living stages (except of the last larval instar) for a 99-million-year-old parasitoid insect from Myanmar (Figure 1 and Supplemental Information). The wedge-shaped beetles (Ripiphoridae) are of special interest among parasitoids because of their obligatory, protelean development in larvae of cockroaches, beetles, bees and wasps.4.

A new Bacillus thuringiensis protein for Western corn rootworm control.

The Western corn rootworm (WCR) Diabrotica virgifera virgifera LeConte is one of the most economically important insect pests in North America. Since 2003, transgenic maize expressing WCR-active proteins from Bacillus thuringiensis (Bt) have been widely adopted as the main approach to controlling WCR in the U.S. However, the emergence of field resistance to the Bt proteins in current commercial products has been documented in recent years, highlighting the need to develop additional tools for controlling this devasting pest. Here we report the discovery of Vpb4Da2 (initially assigned as Vip4Da2), a new insecticidal protein highly selective against WCR, through high-throughput genome sequencing of a Bt strain sourced from grain dust samples collected in the eastern and central regions of the US. Vpb4Da2 contains a sequence and domain signature distinct from families of other WCR-active proteins. Under field conditions, transgenic maize expressing Vpb4Da2 demonstrates commercial-level (at or below NIS 0.25) root protection against WCR, and reduces WCR beetle emergence by ≥ 97%. Our studies also conclude that Vpb4Da2 controls WCR populations that are resistant to WCR-active transgenic maize expressing Cry3Bb1, Cry34Ab1/Cry35Ab1 (reassigned as Gpp34Ab1/Tpp35Ab1), or DvSnf7 RNA. Based on these findings, Vpb4Da2 represents a valuable new tool for protecting maize against WCR.

Evaluation of reference genes for quantitative real-time PCR normalization in the scarab beetle Holotrichia oblita.

Quantitative real-time polymerase chain reaction (qPT-PCR) is commonly used to analyze gene expression, however, the accuracy of the normalized results is affected by the expression stability of reference genes. Holotrichia oblita (Coleoptera: Scarabaeidae) causes serious damage to crops. Reliable reference genes in H. oblita are needed for qRT-PCR analysis. Therefore, we evaluated 13 reference genes under biotic and abiotic conditions. RefFinder provided a comprehensive stability ranking, and geNorm suggested the optimal number of reference genes for normalization. RPL13a and RPL18 were the most suitable reference genes for developmental stages, tissues, and temperature treatments; RPL13a and RPS3 were the most suitable for pesticide and photoperiod treatments; RPS18 and RPL18 were the most suitable for the two sexes. We validated the normalized results using odorant-binding protein genes as target genes in different tissues. Compared with the selected suitable reference genes, the expression of OBP1 in antennae, abdomen, and wings, and OBP2 in antennae and wings were overestimated due to the instability of ACTb. These results identified several reliable reference genes in H. oblita for normalization, and are valuable for future molecular studies.

Satellitome Analysis in the Ladybird Beetle Hippodamia variegata (Coleoptera, Coccinellidae).

Hippodamia variegata is one of the most commercialized ladybirds used for the biological control of aphid pest species in many economically important crops. This species is the first Coccinellidae whose satellitome has been studied by applying new sequencing technologies and bioinformatics tools. We found that 47% of the H. variegata genome is composed of repeated sequences. We identified 30 satellite DNA (satDNA) families with a median intragenomic divergence of 5.75% and A+T content between 45.6% and 74.7%. This species shows satDNA families with highly variable sizes although the most common size is 100-200 bp. However, we highlight the existence of a satDNA family with a repeat unit of 2 kb, the largest repeat unit described in Coleoptera. PCR amplifications for fluorescence in situ hybridization (FISH) probe generation were performed for the four most abundant satDNA families. FISH with the most abundant satDNA family as a probe shows its pericentromeric location on all chromosomes. This location is coincident with the heterochromatin revealed by C-banding and DAPI staining, also analyzed in this work. Hybridization signals for other satDNA families were located only on certain bivalents and the X chromosome. These satDNAs could be very useful as chromosomal markers due to their reduced location.

Modelling the effect of temperature on the biology and demographic parameters of the African coffee white stem borer, Monochamus leuconotus (Pascoe) (Coleoptera: Cerambycidae).

The African coffee white stem borer Monochamus leuconotus (Pascoe) (Coleoptera: Cerambycidae) is a destructive insect pest of Arabica coffee trees in African highlands. Our study aims to provide information on the pest biology as influenced by temperature, determine thermal thresholds, and provide life table parameters for M. leuconotus reared in the laboratory. The life cycle of M. leuconotus was studied at seven constant temperatures in the range 15-35 °C, with 80 ± 5% RH and a photoperiod of L:D 12:12. Linear and nonlinear models were fitted to laboratory data to describe the impact of temperature on M. leuconotus development, mortality, fecundity and senescence. The complete life cycle was obtained between 18 and 30 °C, with the egg incubation period ranging 10.8-29.2 days. The development time was longest for the larva, with 194.2 days at 30 °C and 543.1 days at 18 °C. The minimum temperature threshold (Tmin) was estimated at 10.7, 10.0 and 11.5 °C, for egg, larva and pupa, respectively. The maximum temperature threshold (Tmax) was estimated at 37.4, 40.6 and 40.0 °C for egg, larva and pupa, respectively. The optimum temperature for immature stage survival was estimated between 23.0 and 23.9 °C. The highest fecundity was 97.8 eggs per female at 23 °C. Simulated life table parameters showed the highest net reproductive rate (Ro) of 11.8 daughters per female at 26 °C and maximal intrinsic rate of increase (rm) between 26 and 28 °C, with a value of 0.008. Our results will help understanding M. leuconotus biology as influenced by temperature and may be used to predict the distribution and infestation risk under climate warming for this critical coffee pest.

Temperature and altitude modulate feeding attributes of Mexican beetle, Zygogramma bicolorata Pallister on Parthenium hysterophorus.

Zygogramma bicolorata Pallister (Coleoptera: Chrysomelidae) is an effective biocontrol agent of Parthenium hysterophorus L. which is an alien invasive herbaceous weed with a pan-tropical distribution. The present study aimed to assess the effects of temperature and altitude on feeding attributes (consumption rate, conversion efficiency and growth rate) of adults from the wild populations of Z. bicolorata inhabiting India and Nepal. Results revealed that adults inhabiting areas of low temperature (24°C ‒ 25°C) and high altitude (415 m ‒1400 m) were large and had higher food consumption rates. In contrast, those inhabiting areas of high temperature (34°C ‒ 36°C) and low altitude (81 m ‒ 229 m) were smaller and had higher food utilization efficiencies. In all the eco-climatic regions, females were larger than males and had higher feeding attributes than their counterparts. Temperature between 27°C and 30°C was found optimal for Z. bicolorata adults to convert and utilize the food biomass to body mass. Above the optimal temperature the feeding attributes decreased. Present results suggest that there exists a possibility for decrease in body size, and thereby weed biocontrol efficiency of Z. bicolorata adults with an increase in temperature due to global climate change.

Varietal susceptibility of maize to larger grain borer, Prostephanus truncatus (Horn) (Coleoptera; Bostrichidae), based on grain physicochemical parameters.

Maize (Zea mays L) is one of main nutrients sources for humans and animals worldwide. In Africa, storage of maize ensures food resources availability throughout the year. However, it often suffers losses exceeding 20% due to insects such as the larger grain borer, Prostephanus truncatus (Horn) (Coleoptera; Bostrichidae), major pest of stored maize in the tropical countries. This study aims to select resistant varieties to reduce maize storage losses and explain the physicochemical parameters role in grains susceptibility. In the first study, maize grains were artificially infested under no-choice method with insects. Susceptibility parameters such as weight loss, grain damage, number of emerged insects, median development time and susceptibility index varied significantly through maize varieties. Dobie susceptibility index (SI) was assessed as a major indicator of resistance. The most resistant varieties were Early-Thaï, DMR-ES and Tzee-Yellow. Conversely, Synth-9243, Obatampa and Synth-C varieties were susceptible. SWAN, Across-Pool and Tzee-White were classified as moderately resistant varieties. The insect reproductive potential was significantly different in the nine maize varieties and Early-Thaï, DMR-ES and Tzee-Yellow varieties were the least favourable host. To assess the relationship between grains physicochemical characteristics and varietal susceptibility, moisture, total phenolics, palmitic acid, proteins, amylose, density and grain hardness were evaluated according to standardized methods. Palmitic acid, SI, insects emerged and grain damage were significantly and positively correlated with each other, and negatively correlated with grains hardness, phenolics and amylose contents. Maize susceptibility index was significantly and negatively correlated to amylose, and phenolics contents and positively correlated to palmitic acid content. This study identified three resistant maize varieties to P. tuncatus and revealed that the major factors involved in this resistance were hardness, phenolic and amylose contents of grains.

Combining Experimental Evolution and Genomics to Understand How Seed Beetles Adapt to a Marginal Host Plant.

Genes that affect adaptive traits have been identified, but our knowledge of the genetic basis of adaptation in a more general sense (across multiple traits) remains limited. We combined population-genomic analyses of evolve-and-resequence experiments, genome-wide association mapping of performance traits, and analyses of gene expression to fill this knowledge gap and shed light on the genomics of adaptation to a marginal host (lentil) by the seed beetle Callosobruchus maculatus. Using population-genomic approaches, we detected modest parallelism in allele frequency change across replicate lines during adaptation to lentil. Mapping populations derived from each lentil-adapted line revealed a polygenic basis for two host-specific performance traits (weight and development time), which had low to modest heritabilities. We found less evidence of parallelism in genotype-phenotype associations across these lines than in allele frequency changes during the experiments. Differential gene expression caused by differences in recent evolutionary history exceeded that caused by immediate rearing host. Together, the three genomic datasets suggest that genes affecting traits other than weight and development time are likely to be the main causes of parallel evolution and that detoxification genes (especially cytochrome P450s and beta-glucosidase) could be especially important for colonization of lentil by C. maculatus.

Spatio-temporal variation of Cerambycidae-host tree interaction networks.

Despite its high ecological importance, the commensal interactions at community level are poorly studied. In tropical dry forests (TDF) there is a great diversity of species adapted to the high seasonality that characterizes them; however, little is known regarding how the spatial and temporal availability of resources generates changes in the pattern of commensal interactions. We experimentally studied changes in the diversity, composition, and pattern of interactions in spatio-temporal associations between the saproxylophagous beetles and their host trees in a TDF in Morelos, Mexico. A total of 65 host tree species were selected, from which 16 wood sections were obtained per species. These sections were exposed in the field to allow oviposition by the cerambycids under four different (spatio-temporal) treatments. We analyzed the network structure and generated indices at species level (i.e., specialization, species strength, and effective partners) and those related to physical characteristics of the wood (hardness and degradation rate) and the cerambycids (body size). In total, 1,323 individuals of 57 species of cerambycids emerged. Our results showed that, independently of the space and time, the network presented a nested and modular structure, with a high specialization degree and a high turnover of cerambycid species and their interactions. In general, we found that the cerambycids are mostly associated with softwood species with a lower decomposition rate of wood, as well as with the most abundant host species. The commensalistic interactions between the cerambycids and their host trees are highly specialized but are not spatio-temporally static. The high turnover in the interactions is caused by the emergence patterns of cerambycids, which seem to restrict their use to certain species. The knowledge of the spatio-temporal variation in Cerambycidae-host tree interactions allows us to predict how environmental and structural changes in the habitat can modify the species ensemble, and therefore its interactions.

Evaluation of semiochemical based push-pull strategy for population suppression of ambrosia beetle vectors of laurel wilt disease in avocado.

Ambrosia beetles (Coleoptera: Curculionidae: Scolytinae and Platypodinae) bore into tree xylem to complete their life cycle, feeding on symbiotic fungi. Ambrosia beetles are a threat to avocado where they have been found to vector a symbiotic fungus, Raffaelea lauricola, the causal agent of the laurel wilt disease. We assessed the repellency of methyl salicylate and verbenone to two putative laurel wilt vectors in avocado, Xyleborus volvulus (Fabricius) and Xyleborus bispinatus (Eichhoff), under laboratory conditions. Then, we tested the same two chemicals released from SPLAT flowable matrix with and without low-dose ethanol dispensers for manipulation of ambrosia beetle populations occurring in commercial avocado. The potential active space of repellents was assessed by quantifying beetle catch on traps placed 'close' (~5-10 cm) and 'far' (~1-1.5 m) away from repellent dispensers. Ambrosia beetles collected on traps associated with all in-field treatments were identified to species to assess beetle diversity and community variation. Xyleborus volvulus was not repelled by methyl salicylate (MeSA) or verbenone in laboratory assays, while X. bispinatus was repelled by MeSA but not verbenone. Ambrosia beetle trap catches were reduced in the field more when plots were treated with verbenone dispensers (SPLAT) co-deployed with low-dose ethanol dispensers than when treated with verbenone alone. Beetle diversity was highest on traps deployed with low-dose ethanol lures. The repellent treatments and ethanol lures significantly altered the species composition of beetles captured in experiment plots. Our results indicate that verbenone co-deployed with ethanol lures holds potential for manipulating ambrosia beetle vectors via push-pull management in avocado. This tactic could discourage immigration and/or population establishment of ambrosia beetles in commercial avocado and function as an additional tool for management programs of laurel wilt.

A second VrPGIP1 allele is associated with bruchid resistance (Callosobruchus spp.) in wild mungbean (Vigna radiata var. sublobata) accession ACC41.

Two bruchid species, azuki bean weevil (Callosobruchus chinensis L.) and cowpea weevil (Callosobruchus maculatus F.), are the most important insect pests of mungbean [Vigna radiata (L.) Wilczek] after harvest. Improving bruchid resistance is a major goal for mungbean breeders. Bruchid resistance in mungbean is controlled by a single major locus, Br. The tightly linked VrPGIP1 and VrPGIP2, which encode polygalacturonase-inhibiting proteins (PGIPs), are the candidate genes at the Br locus associated with bruchid resistance. One VrPGIP1 resistance allele and two VrPGIP2 resistance alleles have been identified. In this study, we fine-mapped the bruchid-resistance genes in wild mungbean (V. radiata var. sublobata) accession ACC41 using the F2 population (574 individuals) derived from the 'Kamphaeng Saen 2' (susceptible) × ACC41 (resistant) cross. A QTL analysis indicated that the resistance to the azuki bean weevil and cowpea weevil in ACC41 is controlled by a major QTL (qBr5.1) and a minor QTL (qBr5.2), which are only 0.3 cM apart. qBr5.1 and qBr5.2 accounted for about 82% and 2% of the resistance variation in the F2 population, respectively. qBr5.1 was mapped to a 237.35-kb region on mungbean chromosome 5 containing eight annotated genes, including VrPGIP1 and VrPGIP2. An examination of the ACC41 VrPGIP1 and VrPGIP2 sequences revealed a new allele for VrPGIP1 (i.e., VrPGIP1-2). Compared with the wild-type sequence, VrPGIP1-2 has five SNPs, of which four cause amino acid changes (residues 125, 129, 188, and 336). A protein sequence analysis indicated that residues 125 and 129 in VrPGIP1-2 are in a ß-sheet B1 region, whereas residues 188 and 336 are in a C10-helix region and at the end of the C-terminal region, respectively. Because the ß-sheet B1 region is important for interactions with polygalacturonase (PG), residues 125 and 129 in VrPGIP1-2 likely contribute to bruchid resistance by inhibiting PG. Our results imply that VrPGIP1-2 is associated with the bruchid resistance of wild mungbean accession ACC41. This new resistance allele may be useful for breeding mungbean varieties exhibiting durable bruchid resistance.

Zombie-Ant Fungi Emerged from Non-manipulating, Beetle-Infecting Ancestors.

The manipulation of animal behavior by parasitic organisms is one of the most complex adaptations to have arisen via natural selection. Among the most impressive examples of behavioral manipulation are the zombie-ant fungi [1]. In this association, ants are controlled to leave the colony and perform a stereotyped death grip behavior, where they bite onto vegetation over foraging trails, before being killed for the post mortem fungal growth. Manipulation functions to provide a platform outside the nest, from which fungal parasites actively shoot out spores, targeting foraging ants because within colony transmission is prevented by strong social immunity exhibited by social insect societies [2, 3]. It is not clear how such complex examples of host manipulation arose. To address this, we performed a broad-scale phylogenetic reconstruction of the order Hypocreales, to which the zombie-ant fungi, Ophiocordyceps, belong. In order to understand the patterns of host association and host switching along the evolution of Ophiocordyceps, we performed ancestral character state reconstruction analysis. We found that zombie-ant fungi likely arose from an ancestor that infected beetle larvae residing in soil or decaying wood, similar to extant beetle-infecting Ophiocordyceps species. Surprisingly, the jump led to an extensive species radiation observed after the development of behavioral manipulation. We suggest that the jump from solitary beetle larva to ants within a colony exposed the fungus to the robust social immunity of ant societies.

Soybean plants expressing the Bacillus thuringiensis cry8-like gene show resistance to Holotrichia parallela.

BACKGROUND: Cry8-like from Bacillus thuringiensis (Bt) encodes an insecticidal crystal (Cry) protein. Holotrichia parallela (Coleoptera: Scarabaeoidae), commonly known as the dark black chafer, is a troublesome pest of soybean (Glycine max). To test whether cry8-like can confer resistance against H. parallela to soybean, we introduced cry8-like from the Bt strain HBF-18 into soybean cultivar Jinong 28. RESULTS: Quantitative reverse transcription-PCR analysis demonstrated that cry8-like was expressed most highly in soybean leaves. In addition, Southern blot assays revealed that one copy of the integrated fragment was present in the transformed plants. Eight independent cry8-like transgenic lines were subsequently fed on by H. parallela. Under H. parallela feeding stress, the survival rates of the non-transgenic plants were 92% lower than those of the transgenic plants. The mortality rate of H. parallela increased when the larvae fed on the roots of T1 transgenic soybean plants. Moreover, the surviving larvae were deformed, and their growth was inhibited. CONCLUSIONS: Collectively, our data suggest that transgenic soybean plants expressing the cry8-like gene are more resistant to H. parallela than non-transgenic plants and that transgenic expression of the cry8-like gene may represent a promising strategy for engineering pest tolerance. The events generated in this study could thus be utilized in soybean breeding programs.

Cabbage stem flea beetle's (Psylliodes chrysocephala L.) susceptibility to pyrethroids and tolerance to thiacloprid in the Czech Republic.

The cabbage stem flea beetle (CSFB), Psylliodes chrysocephala (Coleoptera: Chrysomelidae), has recently become a major pest species in winter oilseed rape in the Czech Republic. The susceptibility of CSFB populations from two localities to six pyrethroids, two neonicotinoids, one organophosphate and one oxadiazine was evaluated in 2015-2018 in glass vial experiments. The susceptibility of CSFB to thiacloprid and thiamethoxam was evaluated in feeding experiment in 2017 and 2018. High susceptibility of CSFB populations to lambda-cyhalothrin, cypermethrin, esfenvalerate, tau-fluvalinate, etofenprox, deltamethrin, chlorpyrifos, indoxacarb and acetamiprid was observed in the glass vial experiments. The LC50 and LC90 data obtained for pyrethroids in these experiments in 2015 represent baseline for CSFB resistance monitoring to pyrethroids in the Czech Republic. High tolerance of CSFB to thiacloprid of CSFB was demonstrated in glass vial and the feeding experiment, too.

Relationship between the weights of seed beetles of the genus Megacerus Fåhraeus, 1839 (Coleoptera: Chrysomelidae: Bruchinae) and their host seeds of the family Convolvulaceae.

We studied seeds from a set of plant species from the Convolvulaceae family. Seeds collected from natural populations and infested with beetles of genus Megacerus were monitored until the beetle emergence. We analyze the relationship between body weight of beetles and seed weight of host plants, and its connection with between-species differences and sexual dimorphism. The results show that differences in the scaling of body weight of beetles are associated with sexual dimorphism. For the same species of beetle, the females tend to have heavier bodies than the males. Differences between host plants species in the weight of seeds are related to differences in the body weight Megacerus species, resulting in a distinctive pattern of seed infestation across hosts. Small-sized (lighter) species of beetles tended to infest small-sized (lighter) seed species and, correspondingly, heavier beetles species tended to do it in heavier seed species. Mechanisms of female oviposition preferences may be involved to generate that pattern. In general, the beetle weight showed an asymptotic relation with the host seed weight. The greater the weight of the seed, the greater the weight of adult beetle was. However, the proportion in weights reaches an asymptotic value probably because beetles reached the maximum possible weight for their species. We conclude that the process of specialization in the seed-beetle assemblage studied is influenced by intrinsic traits of the species involved in the interaction (beetles and seeds) and by differences between sexes in their sexual-allocation paths.

Functions of mountain pine beetle cytochromes P450 CYP6DJ1, CYP6BW1 and CYP6BW3 in the oxidation of pine monoterpenes and diterpene resin acids.

The mountain pine beetle (MPB; Dendroctonus ponderosae) is a forest insect pest that attacks several different pine (Pinus) species in its native range of distribution in western North America. MPB are exposed for most of their life cycle to the chemical defenses of their hosts. These defenses are dominated by oleoresin secretions containing mostly various monoterpenes and diterpene resin acids (DRAs). Cytochrome P450 enzymes (P450s) of the MPB are thought to be involved in the metabolism of at least some of these defense compounds. Here we describe the cloning and characterization of three MPB P450s, CYP6DJ1, CYP6BW1 and CYP6BW3, and their functions in the oxidation of various monoterpenes and diterpene resin acids. CYP6DJ1 oxidizes the monoterpenes (+)-(4R)-limonene, (-)-(4S)-limonene and terpinolene and produces (4R,8R)-limonene-8,9-epoxide, (4R,8S)-limonene-8,9-epoxide, (4S,8S)-limonene-8,9-epoxide, (4S,8R)-limonene-8,9-epoxide, perilla alcohol and several unidentified oxidized compounds. These products of CYP6DJ1 were also identified in extracts of MPB treated with the same monoterpenes. CYP6BW1 and CYP6BW3 both oxidize the DRAs abietic acid, dehydroabietic acid, neoabietic acid, levopimaric acid, palustric acid, and isopimaric acid, producing hydroxylated and epoxidized DRAs. CYP6DJ1, CYP6BW1 and CYP6BW3 appear to contribute to the metabolism of oleoresin terpenes as part of the MPB's ability to cope with host defenses.