An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

Fopius arisanus (Sonan) is an important parasitoid of Tephritid fruit flies for at least two reasons. First, it is the one of only three opiine parasitoids known to infect the host during the egg stage1. Second, it has a wide range of potential fruit fly hosts. Perhaps due to its life history, F. arisanus has been a successfully used for biological control of fruit flies in multiple tropical regions2-4. One impediment to the wide use of F. arisanus for fruit fly control is that it is difficult to establish a stable laboratory colony5-9. Despite this difficulty, in the 1990s USDA researchers developed a reliable method to maintain laboratory populations of F. arisanus10-12. There is significant interest in F. arisanus biology13,14, especially regarding its ability to colonize a wide variety of Tephritid hosts14-17; interest is especially driven by the alarming spread of Bactrocera fruit fly pests to new continents in the last decade18. Further research on F. arisanus and additional deployments of this species as a biological control agent will benefit from optimizations and improvements of rearing methods. In this protocol and associated video article we describe an optimized method for rearing F. arisanus based on a previously described approach12. The method we describe here allows rearing of F. arisanus in a small scale without the use of fruit, using materials available in tropical regions around the world and with relatively low manual labor requirements

Functional genomics and microbiome profiling of the Asian longhorned beetle (\u3ci\u3eAnoplophora glabripennis\u3c/i\u3e) reveal insights into the digestive physiology and nutritional ecology of wood feeding beetles

Background: Wood-feeding beetles harbor an ecologically rich and taxonomically diverse assemblage of gut microbes that appear to promote survival in woody tissue, which is devoid of nitrogen and essential nutrients. Nevertheless, the contributions of these apparent symbionts to digestive physiology and nutritional ecology remain uncharacterized in most beetle lineages. Results: Through parallel transcriptome profiling of beetle- and microbial- derived mRNAs, we demonstrate that the midgut microbiome of the Asian longhorned beetle (Anoplophora glabripennis), a member of the beetle family Cerambycidae, is enriched in biosynthetic pathways for the synthesis of essential amino acids, vitamins, and sterols. Consequently, the midgut microbiome of A. glabripennis can provide essential nutrients that the beetle cannot obtain from its woody diet or synthesize itself. The beetle gut microbiota also produce their own suite of transcripts that can enhance lignin degradation, degrade hemicellulose, and ferment xylose and wood sugars. An abundance of cellulases from several glycoside hydrolase families are expressed endogenously by A. glabripennis, as well as transcripts that allow the beetle to convert microbe-synthesized essential amino acids into non-essential amino acids. A. glabripennis and its gut microbes likely collaborate to digest carbohydrates and convert released sugars and amino acid intermediates into essential nutrients otherwise lacking from their woody host plants. Conclusions: The nutritional provisioning capabilities of the A. glabripennis gut microbiome may contribute to the beetles’ unusually broad host range. The presence of some of the same microbes in the guts of other Cerambycidae and other wood-feeding beetles suggests that partnerships with microbes may be a facilitator of evolutionary radiations in beetles, as in certain other groups of insects, allowing access to novel food sources through enhanced nutritional provisioning

\u3ci\u3eWheat streak mosaic virus\u3c/i\u3e alters the transcriptome of its vector, wheat curl mite (\u3ci\u3eAceria tosichella Keifer\u3c/i\u3e), to enhance mite development and population expansion

Wheat streak mosaic virus (WSMV; genus Tritimovirus; family Potyviridae) is an economically important wheat virus that is transmitted by the wheat curl mite (WCM; Aceria tosichella Keifer) in a persistent manner. Virus–vector coevolution may potentially influence vector gene expression to prolong viral association and thus increase virus transmission efficiency and spread. To understand the transcriptomic responses of WCM to WSMV, RNA sequencing was performed to assemble and analyse transcriptomes of WSMV viruliferous and aviruliferous mites. Among 7291 de novo-assembled unigenes, 1020 were differentially expressed between viruliferous and aviruliferous WCMs using edgeR at a false discovery rate ≤0.05. Differentially expressed unigenes were enriched for 108 gene ontology terms, with the majority of the unigenes showing downregulation in viruliferous mites in comparison to only a few unigenes that were upregulated. Protein family and metabolic pathway enrichment analyses revealed that most downregulated unigenes encoded enzymes and proteins linked to stress response, immunity and development. Mechanistically, these predicted changes in mite physiology induced by viral association could be suggestive of pathways needed for promoting virus–vector interactions. Overall, our data suggest that transcriptional changes in viruliferous mites facilitate prolonged viral association and alter WCM development to expedite population expansion, both of which could enhance viral transmission

Wheat streak mosaic virus alters the transcriptome of its vector, wheat curl mite (Aceria tosichella Keifer), to enhance mite development and population expansion

Wheat streak mosaic virus (WSMV; genus Tritimovirus; family Potyviridae) is an economically important wheat virus that is transmitted by the wheat curl mite (WCM; Aceria tosichella Keifer) in a persistent manner. Virus–vector coevolution may potentially influence vector gene expression to prolong viral association and thus increase virus transmission efficiency and spread. To understand the transcriptomic responses of WCM to WSMV, RNA sequencing was performed to assemble and analyse transcriptomes of WSMV viruliferous and aviruliferous mites. Among 7291 de novo-assembled unigenes, 1020 were differentially expressed between viruliferous and aviruliferous WCMs using edgeR at a false discovery rate 0.05. Differentially expressed unigenes were enriched for 108 gene ontology terms, with the majority of the unigenes showing downregulation in viruliferous mites in comparison to only a few unigenes that were upregulated. Protein family and metabolic pathway enrichment analyses revealed that most downregulated unigenes encoded enzymes and proteins linked to stress response, immunity and development. Mechanistically, these predicted changes in mite physiology induced by viral association could be suggestive of pathways needed for promoting virus–vector interactions. Overall, our data suggest that transcriptional changes in viruliferous mites facilitate prolonged viral association and alter WCM development to expedite population expansion, both of which could enhance viral transmission

Divergent Switchgrass Cultivars Modify Cereal Aphid Transcriptomes

Schizaphis graminum Rondani (Hemiptera: Aphididae) and Sipha flava Forbes (Hemiptera: Aphididae) are two common pests of bioenergy grasses. Despite the fact that they are both considered generalists, they differ in their ability to colonize Panicum virgatum cultivars. For example, S. flava colonizes both P. virgatum cv. Summer and P. virgatum cv. Kanlow whereas S. graminum can only colonize Summer. To study the molecular responses of these aphids to these two switchgrass cultivars, we generated de novo transcriptome assemblies and compared the expression profiles of aphids feeding on both cultivars to profiles associated with feeding on a highly susceptible sorghum host and a starvation treatment. Transcriptome assemblies yielded 8,428 and 8,866 high-quality unigenes for S. graminum and S. flava, respectively. Overall, S. graminum responded strongly to all three treatments after 12 h with an upregulation of unigenes coding for detoxification enzymes while major transcriptional changes were not observed in S. flava until 24 h. Additionally, while the two aphids responded to the switchgrass feeding treatment by downregulating unigenes linked to growth and development, their responses to Summer and Kanlow diverged significantly. Schizaphis graminum upregulated more unigenes coding for stress-responsive enzymes in the Summer treatment compared to S. flava; however, many of these unigenes were actually downregulated in the Kanlow treatment. In contrast, S. flava appeared capable of overcoming host defenses by upregulating a larger number of unigenes coding for detoxification enzymes in the Kanlow treatment. Overall, these findings are consistent with previous studies on the interactions of these two cereal aphids to divergent switchgrass hosts

Metagenomic Profiling Reveals Lignocellulose Degrading System in a Microbial Community Associated with a Wood-Feeding Beetle

The Asian longhorned beetle (Anoplophora glabripennis) is an invasive, wood-boring pest that thrives in the heartwood of deciduous tree species. A large impediment faced by A. glabripennis as it feeds on woody tissue is lignin, a highly recalcitrant biopolymer that reduces access to sugars and other nutrients locked in cellulose and hemicellulose. We previously demonstrated that lignin, cellulose, and hemicellulose are actively deconstructed in the beetle gut and that the gut harbors an assemblage of microbes hypothesized to make significant contributions to these processes. While lignin degrading mechanisms have been well characterized in pure cultures of white rot basidiomycetes, little is known about such processes in microbial communities associated with wood-feeding insects. The goals of this study were to develop a taxonomic and functional profile of a gut community derived from an invasive population of larval A. glabripennis collected from infested host trees and to identify genes that could be relevant for the digestion of woody tissue and nutrient acquisition. To accomplish this goal, we taxonomically and functionally characterized the A. glabripennis midgut microbiota through amplicon and shotgun metagenome sequencing and conducted a large-scale comparison with the metagenomes from a variety of other herbivoreassociated communities. This analysis distinguished the A. glabripennis larval gut metagenome from the gut communities of other herbivores, including previously sequenced termite hindgut metagenomes. Genes encoding enzymes were identified in the A. glabripennis gut metagenome that could have key roles in woody tissue digestion including candidate lignin degrading genes (laccases, dye-decolorizing peroxidases, novel peroxidases and β- etherases), 36 families of glycoside hydrolases (such as cellulases and xylanases), and genes that could facilitate nutrient recovery, essential nutrient synthesis, and detoxification. This community could serve as a reservoir of novel enzymes to enhance industrial cellulosic biofuels production or targets for novel control methods for this invasive and highly destructive insect

Variation in DNA Methylation Is Not Consistently Reflected by Sociality in Hymenoptera

Changes in gene regulation that underlie phenotypic evolution can be encoded directly in the DNA sequence or mediated by chromatin modifications such as DNA methylation. It has been hypothesized that the evolution of eusocial division of labor is associated with enhanced gene regulatory potential, which may include expansions in DNA methylation in the genomes of Hymenoptera (bees, ants, wasps, and sawflies). Recently, this hypothesis garnered support from analyses of a commonly used metric to estimate DNA methylation in silico, CpG content. Here, we test this hypothesis using direct, nucleotide-level measures of DNA methylation across nine species of Hymenoptera. In doing so, we generated new DNA methylomes for three species of interest, including one solitary and one facultatively eusocial halictid bee and a sawfly. We demonstrate that the strength of correlation between CpG content and DNA methylation varies widely among hymenopteran taxa, highlighting shortcomings in the utility of CpG content as a proxy for DNA methylation in comparative studies of taxa with sparse DNA methylomes. We observed strikingly high levels of DNA methylation in the sawfly relative to other investigated hymenopterans. Analyses of molecular evolution suggest the relatively distinct sawfly DNA methylome may be associated with positive selection on functional DNMT3 domains. Sawflies are an outgroup to all ants, bees, and wasps, and no sawfly species are eusocial. We find no evidence that either global expansions or variation within individual ortholog groups in DNA methylation are consistently associated with the evolution of social behavior

Wheat streak mosaic virus alters the transcriptome of its vector, wheat curl mite (Aceria tosichella Keifer), to enhance mite development and population expansion

Wheat streak mosaic virus (WSMV; genus Tritimovirus; family Potyviridae) is an economically important wheat virus that is transmitted by the wheat curl mite (WCM; Aceria tosichella Keifer) in a persistent manner. Virus–vector coevolution may potentially influence vector gene expression to prolong viral association and thus increase virus transmission efficiency and spread. To understand the transcriptomic responses of WCM to WSMV, RNA sequencing was performed to assemble and analyse transcriptomes of WSMV viruliferous and aviruliferous mites. Among 7291 de novo-assembled unigenes, 1020 were differentially expressed between viruliferous and aviruliferous WCMs using edgeR at a false discovery rate 0.05. Differentially expressed unigenes were enriched for 108 gene ontology terms, with the majority of the unigenes showing downregulation in viruliferous mites in comparison to only a few unigenes that were upregulated. Protein family and metabolic pathway enrichment analyses revealed that most downregulated unigenes encoded enzymes and proteins linked to stress response, immunity and development. Mechanistically, these predicted changes in mite physiology induced by viral association could be suggestive of pathways needed for promoting virus–vector interactions. Overall, our data suggest that transcriptional changes in viruliferous mites facilitate prolonged viral association and alter WCM development to expedite population expansion, both of which could enhance viral transmission

Divergent Switchgrass Cultivars Modify Cereal Aphid Transcriptomes

Schizaphis graminum Rondani (Hemiptera: Aphididae) and Sipha flava Forbes (Hemiptera: Aphididae) are two common pests of bioenergy grasses. Despite the fact that they are both considered generalists, they differ in their ability to colonize Panicum virgatum cultivars. For example, S. flava colonizes both P. virgatum cv. Summer and P. virgatum cv. Kanlow whereas S. graminum can only colonize Summer. To study the molecular responses of these aphids to these two switchgrass cultivars, we generated de novo transcriptome assemblies and compared the expression profiles of aphids feeding on both cultivars to profiles associated with feeding on a highly susceptible sorghum host and a starvation treatment. Transcriptome assemblies yielded 8,428 and 8,866 high-quality unigenes for S. graminum and S. flava, respectively. Overall, S. graminum responded strongly to all three treatments after 12 h with an upregulation of unigenes coding for detoxification enzymes while major transcriptional changes were not observed in S. flava until 24 h. Additionally, while the two aphids responded to the switchgrass feeding treatment by downregulating unigenes linked to growth and development, their responses to Summer and Kanlow diverged significantly. Schizaphis graminum upregulated more unigenes coding for stress-responsive enzymes in the Summer treatment compared to S. flava; however, many of these unigenes were actually downregulated in the Kanlow treatment. In contrast, S. flava appeared capable of overcoming host defenses by upregulating a larger number of unigenes coding for detoxification enzymes in the Kanlow treatment. Overall, these findings are consistent with previous studies on the interactions of these two cereal aphids to divergent switchgrass hosts