Retrotransposon-mediated disruption of a chitin synthase gene confers insect resistance to Bacillus thuringiensis Vip3Aa toxin.

The vegetative insecticidal protein Vip3Aa from Bacillus thuringiensis (Bt) has been produced by transgenic crops to counter pest resistance to the widely used crystalline (Cry) insecticidal proteins from Bt. To proactively manage pest resistance, there is an urgent need to better understand the genetic basis of resistance to Vip3Aa, which has been largely unknown. We discovered that retrotransposon-mediated alternative splicing of a midgut-specific chitin synthase gene was associated with 5,560-fold resistance to Vip3Aa in a laboratory-selected strain of the fall armyworm, a globally important crop pest. The same mutation in this gene was also detected in a field population. Knockout of this gene via CRISPR/Cas9 caused high levels of resistance to Vip3Aa in fall armyworm and 2 other lepidopteran pests. The insights provided by these results could help to advance monitoring and management of pest resistance to Vip3Aa.

Downregulation of a transcription factor associated with resistance to Bt toxin Vip3Aa in the invasive fall armyworm.

Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) have revolutionized control of some major pests. However, more than 25 cases of field-evolved practical resistance have reduced the efficacy of transgenic crops producing crystalline (Cry) Bt proteins, spurring adoption of alternatives including crops producing the Bt vegetative insecticidal protein Vip3Aa. Although practical resistance to Vip3Aa has not been reported yet, better understanding of the genetic basis of resistance to Vip3Aa is urgently needed to proactively monitor, delay, and counter pest resistance. This is especially important for fall armyworm (Spodoptera frugiperda), which has evolved practical resistance to Cry proteins and is one of the world's most damaging pests. Here, we report the identification of an association between downregulation of the transcription factor gene SfMyb and resistance to Vip3Aa in S. frugiperda. Results from a genome-wide association study, fine-scale mapping, and RNA-Seq identified this gene as a compelling candidate for contributing to the 206-fold resistance to Vip3Aa in a laboratory-selected strain. Experimental reduction of SfMyb expression in a susceptible strain using RNA interference (RNAi) or CRISPR/Cas9 gene editing decreased susceptibility to Vip3Aa, confirming that reduced expression of this gene can cause resistance to Vip3Aa. Relative to the wild-type promoter for SfMyb, the promoter in the resistant strain has deletions and lower activity. Data from yeast one-hybrid assays, genomics, RNA-Seq, RNAi, and proteomics identified genes that are strong candidates for mediating the effects of SfMyb on Vip3Aa resistance. The results reported here may facilitate progress in understanding and managing pest resistance to Vip3Aa.

Escalation by duplication: Milkweed bug trumps Monarch butterfly.

The iconic Monarch butterfly is probably the best-known example of chemical defence against predation, as pictures of vomiting naive blue jays in countless textbooks vividly illustrate. Larvae of the butterfly take up toxic cardiac glycosides from their milkweed hostplants and carry them over to the adult stage. These compounds (cardiotonic steroids, including cardenolides and bufadienolides) inhibit the animal transmembrane sodium-potassium ATPase (Na,K-ATPase), but the Monarch enzyme resists this inhibition thanks to amino acid substitutions in its catalytic alpha-subunit. Some birds also have substitutions and can feast on cardiac glycoside-sequestering insects with impunity. A flurry of recent work has shown how the alpha-subunit gene has been duplicated multiple times in separate insect lineages specializing in cardiac glycoside-producing plants. In this issue of Molecular Ecology, Herbertz et al. toss the beta-subunit into the mix, by expressing all nine combinations of three alpha- and three beta-subunits of the milkweed bug Na,K-ATPase and testing their response to a cardenolide from the hostplant. The findings suggest that the diversification and subfunctionalization of genes allow milkweed bugs to balance trade-offs between resistance towards sequestered host plant toxins that protect the bugs from predators, and physiological costs in terms of Na,K-ATPase activity.

Pesticide resistance in arthropods: Ecology matters too.

Pesticide resistance development is an example of rapid contemporary evolution that poses immense challenges for agriculture. It typically evolves due to the strong directional selection that pesticide treatments exert on herbivorous arthropods. However, recent research suggests that some species are more prone to evolve pesticide resistance than others due to their evolutionary history and standing genetic variation. Generalist species might develop pesticide resistance especially rapidly due to pre-adaptation to handle a wide array of plant allelochemicals. Moreover, research has shown that adaptation to novel host plants could lead to increased pesticide resistance. Exploring such cross-resistance between host plant range evolution and pesticide resistance development from an ecological perspective is needed to understand its causes and consequences better. Much research has, however, been devoted to the molecular mechanisms underlying pesticide resistance while both the ecological contexts that could facilitate resistance evolution and the ecological consequences of cross-resistance have been under-studied. Here, we take an eco-evolutionary approach and discuss circumstances that may facilitate cross-resistance in arthropods and the consequences cross-resistance may have for plant-arthropod interactions in both target and non-target species and species interactions. Furthermore, we suggest future research avenues and practical implications of an increased ecological understanding of pesticide resistance evolution.

Resistance to RNA interference by plant-derived double-stranded RNAs but not plant-derived short interfering RNAs in Helicoverpa armigera.

Plant-mediated RNA interference (RNAi) has emerged as a promising technology for pest control through expression of double-stranded RNAs (dsRNAs) targeted against essential insect genes. However, little is known about the underlying molecular mechanisms and whether long dsRNA or short interfering RNAs (siRNAs) are the effective triggers of the RNAi response. Here we generated transplastomic and nuclear transgenic tobacco plants expressing dsRNA against the Helicoverpa armigera ATPaseH gene. We showed that expression of long dsRNA of HaATPaseH was at least three orders of magnitude higher in transplastomic plants than in transgenic plants. HaATPaseH-derived siRNAs are absent from transplastomic plants, while they are abundant in transgenic plants. Feeding transgenic plants to H. armigera larvae reduced gene expression of HaATPaseH and delayed growth. Surprisingly, no effect of transplastomic plants on insect growth was observed, despite efficient dsRNA expression in plastids. Furthermore, we found that dsRNA ingested by H. armigera feeding on transplastomic plants was rapidly degraded in the intestinal fluid. In contrast, siRNAs are relatively stable in the digestive system. These results suggest that plant-derived siRNAs may be more effective triggers of RNAi in Lepidoptera than dsRNAs, which will aid the optimization of the strategies for plant-mediated RNAi to pest control.

Mechanisms of Resistance to Insecticidal Proteins from Bacillus thuringiensis.

Insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are used in sprayable formulations or produced in transgenic crops as the most successful alternatives to synthetic pesticides. The most relevant threat to sustainability of Bt insecticidal proteins (toxins) is the evolution of resistance in target pests. To date, high-level resistance to Bt sprays has been limited to one species in the field and another in commercial greenhouses. In contrast, there are currently seven lepidopteran and one coleopteran species that have evolved practical resistance to transgenic plants producing insecticidal Bt proteins. In this article, we present a review of the current knowledge on mechanisms of resistance to Bt toxins, with emphasis on key resistance genes and field-evolved resistance, to support improvement of Bt technology and its sustainability.

Direct evidence for a new mode of plant defense against insects via a novel polygalacturonase-inhibiting protein expression strategy.

Plant cell wall-associated polygalacturonase-inhibiting proteins (PGIPs) are widely distributed in the plant kingdom. They play a crucial role in plant defense against phytopathogens by inhibiting microbial polygalacturonases (PGs). PGs hydrolyze the cell wall polysaccharide pectin and are among the first enzymes to be secreted during plant infection. Recent studies demonstrated that herbivorous insects express their own PG multi-gene families, raising the question whether PGIPs also inhibit insect PGs and protect plants from herbivores. Preliminary evidence suggested that PGIPs may negatively influence larval growth of the leaf beetle Phaedon cochleariae (Coleoptera: Chrysomelidae) and identified BrPGIP3 from Chinese cabbage (Brassica rapa ssp. pekinensis) as a candidate. PGIPs are predominantly studied in planta because their heterologous expression in microbial systems is problematic and instability and aggregation of recombinant PGIPs has complicated in vitro inhibition assays. To minimize aggregate formation, we heterologously expressed BrPGIP3 fused to a glycosylphosphatidylinositol (GPI) membrane anchor, immobilizing it on the extracellular surface of insect cells. We demonstrated that BrPGIP3_GPI inhibited several P. cochleariae PGs in vitro, providing the first direct evidence of an interaction between a plant PGIP and an animal PG. Thus, plant PGIPs not only confer resistance against phytopathogens, but may also aid in defense against herbivorous beetles.

Genomic analysis of novel Yarrowia-like yeast symbionts associated with the carrion-feeding burying beetle Nicrophorus vespilloides.

BACKGROUND: Mutualistic interactions with microbes can help insects adapt to extreme environments and unusual diets. An intriguing example is the burying beetle Nicrophorus vespilloides, which feeds and reproduces on small vertebrate carcasses. Its fungal microbiome is dominated by yeasts that potentially facilitate carcass utilization by producing digestive enzymes, eliminating cadaver-associated toxic volatiles (that would otherwise attract competitors), and releasing antimicrobials to sanitize the microenvironment. Some of these yeasts are closely related to the biotechnologically important species Yarrowia lipolytica. RESULTS: To investigate the roles of these Yarrowia-like yeast (YLY) strains in more detail, we selected five strains from two different phylogenetic clades for third-generation sequencing and genome analysis. The first clade, represented by strain B02, has a 20-Mb genome containing ~ 6400 predicted protein-coding genes. The second clade, represented by strain C11, has a 25-Mb genome containing ~ 6300 predicted protein-coding genes, and extensive intraspecific variability within the ITS-D1/D2 rDNA region commonly used for species assignments. Phenotypic microarray analysis revealed that both YLY strains were able to utilize a diverse range of carbon and nitrogen sources (including microbial metabolites associated with putrefaction), and can grow in environments with extreme pH and salt concentrations. CONCLUSIONS: The genomic characterization of five yeast strains isolated from N. vespilloides resulted in the identification of strains potentially representing new YLY species. Given their abundance in the beetle hindgut, and dominant growth on beetle-prepared carcasses, the analysis of these strains has revealed the genetic basis of a potential symbiotic relationship between yeasts and burying beetles that facilitates carcass digestion and preservation.

Timing strains of the marine insect Clunio marinus diverged and persist with gene flow.

Genetic divergence of populations in the presence of gene flow is a central theme in speciation research. Theory predicts that divergence can happen with full range overlap - in sympatry - driven by ecological factors, but there are few empirical examples of how ecologically divergent selection can overcome gene flow and lead to reproductive isolation. In the marine midge Clunio marinus (Diptera: Chironomidae) reproduction is ecologically restricted to the time of the lowest tides, which is ensured through accurate control of development and adult emergence by circalunar and circadian clocks. As tidal regimes differ along the coastline, locally adapted timing strains of C. marinus are found in different sites across Europe. At the same time, ecologically suitable low tides occur at both full and new moon and twice a day, providing C. marinus with four nonoverlapping temporal niches at every geographic location. Along the coast of Brittany, which is characterized by a steep gradient in timing of the tides, we found an unusually large number of differentially adapted timing strains, and the first known instances of sympatric C. marinus strains occupying divergent temporal niches. Analysis of mitochondrial genotypes suggests that these timing strains originated from a single recent colonization event. Nuclear genotypes show strong gene flow, sympatric timing strains being the least differentiated. Even when sympatric strains exist in nonoverlapping temporal niches, timing adaptations do not result in genome-wide genetic divergence, suggesting timing adaptations are maintained by permanent ecological selection. This constitutes a model case for incipient ecological divergence with gene flow.

Consumption of gossypol increases fatty acid-amino acid conjugates in the cotton pests Helicoverpa armigera and Heliothis virescens.

Gossypol is a toxic sesquiterpene dimer produced by cotton plants which deters herbivory by insects and vertebrates. Two highly reactive aldehyde groups contribute to gossypol toxicity by cross-linking herbivore proteins. We identified another consequence of consuming gossypol in two insect pests of cotton: increased amounts of fatty acid-amino acid conjugates (FACs). Eight different FACs in the feces of larval Helicoverpa armigera and Heliothis virescens increased when larvae consumed artificial diet containing gossypol, but not a gossypol derivative lacking free aldehyde groups (SB-gossypol). FACs are produced by joining plant-derived fatty acids with amino acids of insect origin in the larval midgut tissue by an unknown conjugase, and translocated into the gut lumen by an unknown transporter. FACs are hydrolyzed back into fatty acids and amino acids by an aminoacylase (L-ACY-1) in the gut lumen. The equilibrium level of FACs in the lumen is determined by a balance between conjugation and hydrolysis, which may differ among species. When heterologously expressed, L-ACY-1 of H. armigera but not H. virescens was inhibited by gossypol; consistent with the excretion of more FACs in the feces by H. armigera. FACs are known to benefit the plant host by inducing anti-herbivore defensive responses, and have been hypothesized to benefit the herbivore by acting as a surfactant and increasing nitrogen uptake efficiency. Thus in addition to its direct toxic effects, gossypol may negatively impact insect nitrogen uptake efficiency and amplify the signal used by the plant to elicit release of volatile compounds that attract parasitoids.

Two calcium-binding chaperones from the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) involved in diapause.

Molecular chaperones are crucial for the correct folding of newly synthesized polypeptides, in particular, under stress conditions. Various studies have revealed the involvement of molecular chaperones, such as heat shock proteins, in diapause maintenance and starvation; however, the role of other chaperones in diapause and starvation relatively is unknown. In the current study, we identified two lectin-type chaperones with calcium affinity, a calreticulin (LdCrT) and a calnexin (LdCnX), that were present in the fat body of the Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae) during diapause. Both proteins possessed an N-globular domain, a P-arm domain, and a highly charged C-terminal domain, while an additional transmembrane domain was present in LdCnX. Phylogenetic analysis revealed distinction at the order level. Both genes were expressed in multiple tissues in larval and adult stages, and constitutively throughout development, though a starvation response was detected only for LdCrT. In females, diapause-related expression analysis in the whole body revealed an upregulation of both genes by post-diapause, but a downregulation by diapause only for LdCrT. By contrast, males revealed no alteration in their diapause-related expression pattern in the entire body for both genes. Fat body-specific expression analysis of both genes in relation to diapause revealed the same expression pattern with no alteration in females and downregulation in males by post-diapause. This study suggests that calcium-binding chaperones play similar and possibly gender-specific roles during diapause.

Silencing of an ABC transporter, but not a cadherin, decreases the susceptibility of Colorado potato beetle larvae to Bacillus thuringiensis ssp. tenebrionis Cry3Aa toxin.

The Colorado potato beetle, Leptinotarsa decemlineata (Coleoptera: Chrysomelidae), is a major pest of potato plants worldwide and is notorious for its ability to develop resistance to insecticides. Cry3 toxins synthesized by Bacillus thuringiensis ssp. tenebrionis have been used successfully to manage this pest. Resistance to Cry toxins is a concerning problem for many insect pests; therefore, it is important to determine the mechanisms by which insects acquire resistance to these toxins. Cadherin-like and ABC transporter proteins have been implicated in the mode of action of Cry toxins as mutations in these genes render lepidopterans resistant to them; however, clear consensus does not exist on whether these proteins also play a role in Cry3 toxin activity and/or development of resistance in coleopterans. In the current study, we identified the L. decemlineata orthologues of the cadherin (LdCAD) and the ABCB transporter (LdABCB1) that have been implicated in the mode of action of Cry toxins in other coleopterans. Suppression of LdABCB1 via RNA interference reduced toxin-related larval mortality, whereas partial silencing of LdCAD did not. Our results suggest that the ABCB is involved in the mode of action of Cry3Aa toxins; however, no evidence was found to support the role of cadherin as a receptor of Cry3Aa in L. decemlineata.

Microbiome-assisted carrion preservation aids larval development in a burying beetle.

The ability to feed on a wide range of diets has enabled insects to diversify and colonize specialized niches. Carrion, for example, is highly susceptible to microbial decomposers, but is kept palatable several days after an animal's death by carrion-feeding insects. Here we show that the burying beetle Nicrophorus vespilloides preserves carrion by preventing the microbial succession associated with carrion decomposition, thus ensuring a high-quality resource for their developing larvae. Beetle-tended carcasses showed no signs of degradation and hosted a microbial community containing the beetles' gut microbiota, including the yeast Yarrowia In contrast, untended carcasses showed visual and olfactory signs of putrefaction, and their microbial community consisted of endogenous and soil-originating microbial decomposers. This regulation of the carcass' bacterial and fungal community and transcriptomic profile was associated with lower concentrations of putrescine and cadaverine (toxic polyamines associated with carcass putrefaction) and altered levels of proteases, lipases, and free amino acids. Beetle-tended carcasses develop a biofilm-like matrix housing the yeast, which, when experimentally removed, leads to reduced larval growth. Thus, tended carcasses hosted a mutualistic microbial community that promotes optimal larval development, likely through symbiont-mediated extraintestinal digestion and detoxification of carrion nutrients. The adaptive preservation of carrion coordinated by the beetles and their symbionts demonstrates a specialized resource-management strategy through which insects modify their habitats to enhance fitness.

An Insect Counteradaptation against Host Plant Defenses Evolved through Concerted Neofunctionalization.

Antagonistic chemical interactions between herbivorous insects and their host plants are often thought to coevolve in a stepwise process, with an evolutionary innovation on one side being countered by a corresponding advance on the other. Glucosinolate sulfatase (GSS) enzyme activity is essential for the Diamondback moth, Plutella xylostella, to overcome a highly diversified secondary metabolite-based host defense system in the Brassicales. GSS genes are located in an ancient cluster of arylsulfataselike genes, but the exact roles of gene copies and their evolutionary trajectories are unknown. Here, we combine a functional investigation of duplicated insect arylsulfatases with an analysis of associated nucleotide substitution patterns. We show that the Diamondback moth genome encodes three GSSs with distinct substrate spectra and distinct expression patterns in response to glucosinolates. Contrary to our expectations, early functional diversification of gene copies was not indicative of a coevolutionary arms race between host and herbivore. Instead, both copies of a duplicated arylsulfatase gene evolved concertedly in the context of an insect host shift to acquire novel detoxifying functions under positive selection, a pattern of duplicate gene retention that we call "concerted neofunctionalization."

How do toxins from Bacillus thuringiensis kill insects? An evolutionary perspective.

Three-domain Cry toxins from the bacterium Bacillus thuringiensis (Bt) are increasingly used in agriculture to replace chemical insecticides in pest control. Most chemical insecticides kill pest insects swiftly, but are also toxic to beneficial insects and other species in the agroecosystem. Cry toxins enjoy the advantages of high selectivity and the possibility of the application by sprays or transgenic plants. However, these benefits are offset by the limited host range and the evolution of resistance to Bt toxins by insect pests. Understanding how Bt toxins kill insects will help to understand the nature of both problems. The recent realization that ABC transporters play a central role in the killing mechanism will play an important role in devising solutions.

Effects of class-specific, synthetic, and natural proteinase inhibitors on life-history traits of the cotton bollworm Helicoverpa armigera.

Herbivorous insects have more difficulty obtaining proteins from their food than do predators and parasites. The scarcity of proteins in their diet requires herbivores to feed voraciously, thus heavily damaging their host plants. Plants respond to herbivory by producing defense compounds, which reduce insect growth, retard development, and increase mortality. Herbivores use both pre- and postdigestive response mechanisms to detect and avoid plant defense compounds. Proteinase inhibitors (PIs) are one example of plant compounds produced as a direct defense against herbivory. Many insects can adapt to PIs when these are incorporated into artificial diets. However, little is known about the effect of PIs on diet choice and feeding behavior. We monitored the diet choice, life-history traits, and gut proteinase activity of Helicoverpa armigera larvae using diets supplemented with synthetic and natural PIs. In choice experiments, both neonates and fourth-instar larvae preferred the control diet over PI-supplemented diets, to varying degrees. Larvae that fed on PI-supplemented diets weighed less than those that fed on the control diet and produced smaller pupae. Trypsin-specific PIs had a stronger effect on mean larval weight than did other PIs. A reduction of trypsin activity but not of chymotrypsin activity was observed in larvae fed on PI-supplemented diets. Therefore, behavioral avoidance of feeding on plant parts high in PIs could be an adaptation to minimize the impact of this plant's defensive strategy.

Parasite-host specificity: A cross-infection study of the parasite Ophryocystis elektroscirrha.

Many parasites are constrained to only one or a few hosts, showing host specificity. It remains unclear why some parasites are specialists and other parasites are generalists. The parasite Ophryocystis elektroscirrha (OE) is a neogregarine protozoan thought to be restricted to monarch butterflies, Danaus plexippus (Nymphaliae) and D. gilippus. Recently, we found OE-like spores in other Lepidoptera, specifically in three noctuid moths: Helicoverpa armigera, H. assulta and H. punctigera, as well as another nymphalid, Parthenos sylvia. To our knowledge, this is the first report of OE-like parasite infections in species other than the genus Danaus. In sequencing 558 bp of 18S rRNA, we found the genetic similarity between OE from D. plexippus and OE-like parasite from the moths H. armigera and H. punctigera to be 95.2%. When we conducted cross-species infection experiments, we could not infect the moths with OE from D. plexippus, but OE-like parasite from H. armigera did infect D. plexippus and a closely related moth species Heliothis virescens. Interestingly, we did not find the OE-like parasite in the H. armigera population from Spain. Inter-population infection experiments with H. armigera demonstrated a higher sensitivity to OE-like infection in the population from Spain compared to the populations from Australia and China. These results suggest geographic variation in OE-like susceptibility and coevolution between parasite and host. Our findings give important new insights into the prevalence and host specificity of OE and OE-like parasites, and provide opportunities to study parasite transmission over spatial and temporal scales.

Genetic mapping of male pheromone response in the European corn borer identifies candidate genes regulating neurogenesis.

The sexual pheromone communication system of moths is a model system for studies of the evolution of reproductive isolation. Females emit a blend of volatile components that males detect at a distance. Species differences in female pheromone composition and male response directly reinforce reproductive isolation in nature, because even slight variations in the species-specific pheromone blend are usually rejected by the male. The mechanisms by which a new pheromone signal-response system could evolve are enigmatic, because any deviation from the optimally attractive blend should be selected against. Here we investigate the genetic mechanisms enabling a switch in male response. We used a quantitative trait locus-mapping approach to identify the genetic basis of male response in the two pheromone races of the European corn borer, Ostrinia nubilalis Male response to a 99:1 vs. a 3:97 ratio of the E and Z isomers of the female pheromone is governed by a single, sex-linked locus. We found that the chromosomal region most tightly linked to this locus contains genes involved in neurogenesis but, in accordance with an earlier study, does not contain the odorant receptors expressed in the male antenna that detect the pheromone. This finding implies that differences in the development of neuronal pathways conveying information from the antenna, not differences in pheromone detection by the odorant receptors, are primarily responsible for the behavioral response differences among the males in this system. Comparison with other moth species reveals a previously unexplored mechanism by which male pheromone response can change in evolution.

Novel family of terpene synthases evolved from trans-isoprenyl diphosphate synthases in a flea beetle.

Sesquiterpenes play important roles in insect communication, for example as pheromones. However, no sesquiterpene synthases, the enzymes involved in construction of the basic carbon skeleton, have been identified in insects to date. We investigated the biosynthesis of the sesquiterpene (6R,7S)-himachala-9,11-diene in the crucifer flea beetle Phyllotreta striolata, a compound previously identified as a male-produced aggregation pheromone in several Phyllotreta species. A (6R,7S)-himachala-9,11-diene-producing sesquiterpene synthase activity was detected in crude beetle protein extracts, but only when (Z,E)-farnesyl diphosphate [(Z,E)-FPP] was offered as a substrate. No sequences resembling sesquiterpene synthases from plants, fungi, or bacteria were found in the P. striolata transcriptome, but we identified nine divergent putative trans-isoprenyl diphosphate synthase (trans-IDS) transcripts. Four of these putative trans-IDSs exhibited terpene synthase (TPS) activity when heterologously expressed. Recombinant PsTPS1 converted (Z,E)-FPP to (6R,7S)-himachala-9,11-diene and other sesquiterpenes observed in beetle extracts. RNAi-mediated knockdown of PsTPS1 mRNA in P. striolata males led to reduced emission of aggregation pheromone, confirming a significant role of PsTPS1 in pheromone biosynthesis. Two expressed enzymes showed genuine IDS activity, with PsIDS1 synthesizing (E,E)-FPP, whereas PsIDS3 produced neryl diphosphate, (Z,Z)-FPP, and (Z,E)-FPP. In a phylogenetic analysis, the PsTPS enzymes and PsIDS3 were clearly separated from a clade of known coleopteran trans-IDS enzymes including PsIDS1 and PsIDS2. However, the exon-intron structures of IDS and TPS genes in P. striolata are conserved, suggesting that this TPS gene family evolved from trans-IDS ancestors.

Burying beetles regulate the microbiome of carcasses and use it to transmit a core microbiota to their offspring.

Necrophagous beetles utilize carrion, a highly nutritious resource that is susceptible to intense microbial competition, by treating it with antimicrobial anal and oral secretions. However, how this regulates the carcass microbiota remains unclear. Here, we show that carcasses prepared by the burying beetle Nicrophorus vespilloides undergo significant changes in their microbial communities subsequent to their burial and "preparation." Prepared carcasses hosted a microbial community that was more similar to that of beetles' anal and oral secretions than to the native carcass community or the surrounding soil, indicating that the beetles regulated the carcass microbiota. A core microbial community (Xanthomonadaceae, Enterococcaceae, Enterobacteriaceae and Yarrowia yeasts) was transmitted by the beetles to the larvae via the anal and oral secretions and the carcass surface. These core taxa proliferated on the carcass, indicating a growth conducive environment for these microbes when associated with beetles. However, total bacterial loads were higher on decomposing carcasses without beetles than on beetle-prepared carcasses, indicating that the beetles and/or their associated symbionts suppress the growth of competing microbes. Thus, apart from being a nutritional resource, the carcass provides a medium for vertical transmission of a tightly regulated symbiotic microbiota, whose activity on the carcass and in the larval gut may involve carcass preservation as well as digestion.