The Lymantria dispar IPLB-Ld652Y Cell Line Transcriptome Comprises Diverse Virus-Associated Transcripts

The enhanced viral susceptibility of the gypsy moth (Lymantria dispar)-derived IPLB-Ld652Y cell line has made it a popular in vitro system for studying virus-related phenomena in the Lepidoptera. Using both single-pass EST sequencing and 454-based pyrosequencing, a transcriptomic library of 14,368 putatively unique transcripts (PUTs) was produced comprising 8,476,050 high-quality, informative bases. The gene content of the IPLB-Ld652Y transcriptome was broadly assessed via comparison with the NCBI non-redundant protein database, and more detailed functional annotation was inferred by comparison to the Swiss-Prot subset of UniProtKB. In addition to L. dispar cellular transcripts, a diverse array of both RNA and DNA virus-associated transcripts was identified within the dataset, suggestive of a high level of viral expression and activity in IPLB-Ld652Y cells. These sequence resources will provide a sound basis for developing testable experimental hypotheses by insect virologists, and suggest a number of avenues for potential research

The Parasitic Wasp, Cotesia congregata (Say), Consists of Two Incipient Species Isolated by Asymmetric Reproductive Incompatibility and Hybrid Inability to Overcome Host Defenses

Parasitic wasps are highly diverse and play a major role in suppression of herbivorous insect pest populations. Several previously identified species of parasitic wasps have been found to be complexes of cryptic species resulting from adaptations to specific hosts or host foodplants. Cotesia congregata (Say) (Hymenoptera: Braconidae), which has long served as a model system for host-parasitoid interactions, can be used for investigating the process of diversification among sympatric populations that differ in host and host foodplant usage. Two incipient species of C. congregata have been identified in the USA mid-Atlantic region, “MsT wasps” originate from Manduca sexta (L.) (Lepidoptera: Sphingidae) on tobacco and “CcC wasps” originate from Ceratomia catalpae (Boisduval) (Lepidoptera: Sphingidae) on catalpa. Both wasp sources can develop in either host species. Hybrids resulting from MsT♂xCcC♀ crosses are fertile, whereas hybrids from CcC♂xMsT♀ crosses are typically sterile. In this study, we compared relative expression in vivo of seven C. congregata bracovirus (CcBV) genes among MsT and CcC parental and hybrid crosses. Also, we established hybrid crosses between MsT and CcC wasps and four additional host foodplant sources of C. congregata. Patterns of relative expression in vivo of MsT and CcC CcBV genes differed; a few were not expressed in hosts parasitized by CcC wasps. Overall, relative expression of CcBV genes from MsT and CcC wasps did not differ with respect to the host species parasitized. Low or absent expression of CcBV genes was found in hosts parasitized by sterile hybrids. For the most part, the other four host-foodplant wasp sources were reproductively compatible with either MsT or CcC wasps and hybrid crosses with the alternative wasp source were asymmetrically sterile. Crosses involving CcC males or MsT females produced sterile hybrids that lacked mature ovaries. Cumulatively, results indicate that C. congregata is composed of two sympatric incipient species that can utilize multiple host species rather than several host-associated races or cryptic species

De novo formation of an aggregation pheromone precursor by an isoprenyl diphosphate synthase-related terpene synthase in the harlequin bug.

Insects use a diverse array of specialized terpene metabolites as pheromones in intraspecific interactions. In contrast to plants and microbes, which employ enzymes called terpene synthases (TPSs) to synthesize terpene metabolites, limited information from few species is available about the enzymatic mechanisms underlying terpene pheromone biosynthesis in insects. Several stink bugs (Hemiptera: Pentatomidae), among them severe agricultural pests, release 15-carbon sesquiterpenes with a bisabolene skeleton as sex or aggregation pheromones. The harlequin bug, Murgantia histrionica, a specialist pest of crucifers, uses two stereoisomers of 10,11-epoxy-1-bisabolen-3-ol as a male-released aggregation pheromone called murgantiol. We show that MhTPS (MhIDS-1), an enzyme unrelated to plant and microbial TPSs but with similarity to trans-isoprenyl diphosphate synthases (IDS) of the core terpene biosynthetic pathway, catalyzes the formation of (1S,6S,7R)-1,10-bisaboladien-1-ol (sesquipiperitol) as a terpene intermediate in murgantiol biosynthesis. Sesquipiperitol, a so-far-unknown compound in animals, also occurs in plants, indicating convergent evolution in the biosynthesis of this sesquiterpene. RNAi-mediated knockdown of MhTPS mRNA confirmed the role of MhTPS in murgantiol biosynthesis. MhTPS expression is highly specific to tissues lining the cuticle of the abdominal sternites of mature males. Phylogenetic analysis suggests that MhTPS is derived from a trans-IDS progenitor and diverged from bona fide trans-IDS proteins including MhIDS-2, which functions as an (E,E)-farnesyl diphosphate (FPP) synthase. Structure-guided mutagenesis revealed several residues critical to MhTPS and MhFPPS activity. The emergence of an IDS-like protein with TPS activity in M. histrionica demonstrates that de novo terpene biosynthesis evolved in the Hemiptera in an adaptation for intraspecific communication

Comparative genomics of mutualistic viruses of Glyptapanteles parasitic wasps

Comparative genome analysis of two endosymbiotic polydnaviruses from Glyptapanteles parasitic wasps reveals new insights into the evolutionary arms race between host and parasite

A single vertebrate DNA virus protein disarms invertebrate immunity to RNA virus infection

Virus-host interactions drive a remarkable diversity of immune responses and countermeasures. We found that two RNA viruses with broad host ranges, vesicular stomatitis virus (VSV) and Sindbis virus (SINV), are completely restricted in their replication after entry into Lepidopteran cells. This restriction is overcome when cells are co-infected with vaccinia virus (VACV), a vertebrate DNA virus. Using RNAi screening, we show that Lepidopteran RNAi, Nuclear Factor-kappaB, and ubiquitin-proteasome pathways restrict RNA virus infection. Surprisingly, a highly conserved, uncharacterized VACV protein, A51R, can partially overcome this virus restriction. We show that A51R is also critical for VACV replication in vertebrate cells and for pathogenesis in mice. Interestingly, A51R colocalizes with, and stabilizes, host microtubules and also associates with ubiquitin. We show that A51R promotes viral protein stability, possibly by preventing ubiquitin-dependent targeting of viral proteins for destruction. Importantly, our studies reveal exciting new opportunities to study virus-host interactions in experimentally-tractable Lepidopteran systems

The Occurrence of Photorhabdus-Like Toxin Complexes in Bacillus thuringiensis

Recently, genomic sequencing of a Bacillus thuringiensis (Bt) isolate from our collection revealed the presence of an apparent operon encoding an insecticidal toxin complex (Tca) similar to that first described from the entomopathogen Photorhabdus luminescens. To determine whether these genes are widespread among Bt strains, we screened isolates from the collection for the presence of tccC, one of the genes needed for the expression of fully functional toxin complexes. Among 81 isolates chosen to represent commonly encountered biochemical phenotypes, 17 were found to possess a tccC. Phylogenetic analysis of the 81 isolates by multilocus sequence typing revealed that all the isolates possessing a tccC gene were restricted to two sequence types related to Bt varieties morrisoni, tenebrionis, israelensis and toumanoffi. Sequencing of the ∼17 kb tca operon from two isolates representing each of the two sequence types revealed >99% sequence identity. Optical mapping of DNA from Bt isolates representing each of the sequence types revealed nearly identical plasmids of ca. 333 and 338 kbp, respectively. Selected isolates were found to be toxic to gypsy moth larvae, but were not as effective as a commercial strain of Bt kurstaki. Some isolates were found to inhibit growth of Colorado potato beetle. Custom Taqman® relative quantitative real-time PCR assays for Tc-encoding Bt revealed both tcaA and tcaB genes were expressed within infected gypsy moth larvae

Gene content evolution in the arthropods

Arthropods comprise the largest and most diverse phylum on Earth and play vital roles in nearly every ecosystem. Their diversity stems in part from variations on a conserved body plan, resulting from and recorded in adaptive changes in the genome. Dissection of the genomic record of sequence change enables broad questions regarding genome evolution to be addressed, even across hyper-diverse taxa within arthropods. Using 76 whole genome sequences representing 21 orders spanning more than 500 million years of arthropod evolution, we document changes in gene and protein domain content and provide temporal and phylogenetic context for interpreting these innovations. We identify many novel gene families that arose early in the evolution of arthropods and during the diversification of insects into modern orders. We reveal unexpected variation in patterns of DNA methylation across arthropods and examples of gene family and protein domain evolution coincident with the appearance of notable phenotypic and physiological adaptations such as flight, metamorphosis, sociality, and chemoperception. These analyses demonstrate how large-scale comparative genomics can provide broad new insights into the genotype to phenotype map and generate testable hypotheses about the evolution of animal diversity

Brown marmorated stink bug, Halyomorpha halys (Stål), genome: putative underpinnings of polyphagy, insecticide resistance potential and biology of a top worldwide pest

Background Halyomorpha halys (Stål), the brown marmorated stink bug, is a highly invasive insect species due in part to its exceptionally high levels of polyphagy. This species is also a nuisance due to overwintering in human-made structures. It has caused significant agricultural losses in recent years along the Atlantic seaboard of North America and in continental Europe. Genomic resources will assist with determining the molecular basis for this species’ feeding and habitat traits, defining potential targets for pest management strategies. Results Analysis of the 1.15-Gb draft genome assembly has identified a wide variety of genetic elements underpinning the biological characteristics of this formidable pest species, encompassing the roles of sensory functions, digestion, immunity, detoxification and development, all of which likely support H. halys’ capacity for invasiveness. Many of the genes identified herein have potential for biomolecular pesticide applications. Conclusions Availability of the H. halys genome sequence will be useful for the development of environmentally friendly biomolecular pesticides to be applied in concert with more traditional, synthetic chemical-based controls

Brown marmorated stink bug, Halyomorpha halys (Stål), genome: putative underpinnings of polyphagy, insecticide resistance potential and biology of a top worldwide pest

Halyomorpha halys (Stål), the brown marmorated stink bug, is a highly invasive insect species due in part to its exceptionally high levels of polyphagy. This species is also a nuisance due to overwintering in human-made structures. It has caused significant agricultural losses in recent years along the Atlantic seaboard of North America and in continental Europe. Genomic resources will assist with determining the molecular basis for this species' feeding and habitat traits, defining potential targets for pest management strategies