Wannes Dermauw

The genome of Tetranychus urticae reveals herbivorous pest adaptations

The spider mite Tetranychus urticae is a cosmopolitan agricultural pest with an extensive host plant range and an extreme record of pesticide resistance. Here we present the completely sequenced and annotated spider mite genome, representing the first complete chelicerate genome. At 90 megabases T. urticae has the smallest sequenced arthropod genome. Compared with other arthropods, the spider mite genome shows unique changes in the hormonal environment and organization of the Hox complex, and also reveals evolutionary innovation of silk production. We find strong signatures of polyphagy and detoxification in gene families associated with feeding on different hosts and in new gene families acquired by lateral gene transfer. Deep transcriptome analysis of mites feeding on different plants shows how this pest responds to a changing host environment. The T. urticae genome thus offers new insights into arthropod evolution and plant–herbivore interactions, and provides unique opportunities for developing novel plant protection strategies.

The ABC gene family in arthropods: Comparative genomics and role in insecticide transport and resistance

About a 100 years ago, the Drosophila white mutant marked the birth of Drosophila genetics. The white gene turned out to encode the first well studied ABC transporter in arthropods. The ABC gene family is now recognized as one of the largest transporter families in all kingdoms of life. The majority of ABC proteins function as primary-active transporters that bind and hydrolyze ATP while transporting a large diversity of substrates across lipid membranes. Although extremely well studied in vertebrates for their role in drug resistance, less is known about the role of this family in the transport of endogenous and exogenous substances in arthropods. The ABC families of five insect species, a crustacean and a chelicerate have been annotated in some detail. We conducted a thorough phylogenetic analysis of the seven arthropod and human ABC protein subfamilies, to infer orthologous relationships that might suggest conserved function. Most orthologous relationships were found in the ABCB half transporter, ABCD, ABCE and ABCF subfamilies, but specific expansions within species and lineages are frequently observed and discussed. We next surveyed the role of ABC transporters in the transport of xenobiotics/plant allelochemicals and their involvement in insecticide resistance. The involvement of ABC transporters in xenobiotic resistance in arthropods is historically not well documented, but an increasing number of studies using unbiased differential gene expression analysis now points to their importance. We give an overview of methods that can be used to link ABC transporters to resistance. ABC proteins have also recently been implicated in the mode of action and resistance to Bt toxins in Lepidoptera. Given the enormous interest in Bt toxicology in transgenic crops, such findings will provide an impetus to further reveal the role of ABC transporters in arthropods.

A link between host plant adaptation and pesticide resistance in the polyphagous spider mite Tetranychus urticae

Plants produce a wide range of allelochemicals to defend against herbivore attack, and generalist herbivores have evolved mechanisms to avoid, sequester, or detoxify a broad spectrum of natural defense compounds. Successful arthropod pests have also developed resistance to diverse classes of pesticides and this adaptation is of critical importance to agriculture. To test whether mechanisms to overcome plant defenses predispose the development of pesticide resistance, we examined adaptation of the generalist two-spotted spider mite, Tetranychus urticae, to host plant transfer and pesticides. T. urticae is an extreme polyphagous pest with more than 1,100 documented hosts and has an extraordinary ability to develop pesticide resistance. When mites from a pesticide-susceptible strain propagated on bean were adapted to a challenging host (tomato), transcriptional responses increased over time with ∼7.5% of genes differentially expressed after five generations. Whereas many genes with altered expression belonged to known detoxification families (like P450 monooxygenases), new gene families not previously associated with detoxification in other herbivores showed a striking response, including ring-splitting dioxygenase genes acquired by horizontal gene transfer. Strikingly, transcriptional profiles of tomato-adapted mites resembled those of multipesticide-resistant strains, and adaptation to tomato decreased the susceptibility to unrelated pesticide classes. Our findings suggest key roles for both an expanded environmental response gene repertoire and transcriptional regulation in the life history of generalist herbivores. They also support a model whereby selection for the ability to mount a broad response to the diverse defense chemistry of plants predisposes the evolution of pesticide resistance in generalists.

Population bulk segregant mapping uncovers resistance mutations and the mode of action of a chitin synthesis inhibitor in arthropods

Because of its importance to the arthropod exoskeleton, chitin biogenesis is an attractive target for pest control. This point is demonstrated by the economically important benzoylurea compounds that are in wide use as highly specific agents to control insect populations. Nevertheless, the target sites of compounds that inhibit chitin biogenesis have remained elusive, likely preventing the full exploitation of the underlying mode of action in pest management. Here, we show that the acaricide etoxazole inhibits chitin biogenesis in Tetranychus urticae (the two-spotted spider mite), an economically important pest. We then developed a population-level bulk segregant mapping method, based on high-throughput genome sequencing, to identify a locus for monogenic, recessive resistance to etoxazole in a field-collected population. As supported by additional genetic studies, including sequencing across multiple resistant strains and genetic complementation tests, we associated a nonsynonymous mutation in the major T. urticae chitin synthase (CHS1) with resistance. The change is in a C-terminal transmembrane domain of CHS1 in a highly conserved region that may serve a noncatalytic but essential function. Our finding of a target-site resistance mutation in CHS1 shows that at least one highly specific chitin biosynthesis inhibitor acts directly to inhibit chitin synthase. Our work also raises the possibility that other chitin biogenesis inhibitors, such as the benzoylurea compounds, may also act by inhibition of chitin synthases. More generally, our genetic mapping approach should be powerful for high-resolution mapping of simple traits (resistance or otherwise) in arthropods.

Genome sequence of the Asian Tiger mosquito, Aedes albopictus, reveals insights into its biology, genetics, and evolution

Significance Aedes albopictus is a highly adaptive species that thrives worldwide in tropical and temperate zones. From its origin in Asia, it has established itself on every continent except Antarctica. This expansion, coupled with its ability to vector the epidemic human diseases dengue and Chikungunya fevers, make it a significant global public health threat. A complete genome sequence and transcriptome data were obtained for the Ae. albopictus Foshan strain, a colony derived from mosquitoes from its historical origin. The large genome (1,967 Mb) comprises an abundance of repetitive DNA classes and expansions of the numbers of gene family members involved in insecticide resistance, diapause, sex determination, immunity, and olfaction. This large genome repertory and plasticity may contribute to its success as an invasive species. The Asian tiger mosquito, Aedes albopictus, is a highly successful invasive species that transmits a number of human viral diseases, including dengue and Chikungunya fevers. This species has a large genome with significant population-based size variation. The complete genome sequence was determined for the Foshan strain, an established laboratory colony derived from wild mosquitoes from southeastern China, a region within the historical range of the origin of the species. The genome comprises 1,967 Mb, the largest mosquito genome sequenced to date, and its size results principally from an abundance of repetitive DNA classes. In addition, expansions of the numbers of members in gene families involved in insecticide-resistance mechanisms, diapause, sex determination, immunity, and olfaction also contribute to the larger size. Portions of integrated flavivirus-like genomes support a shared evolutionary history of association of these viruses with their vector. The large genome repertory may contribute to the adaptability and success of Ae. albopictus as an invasive species.

The cys-loop ligand-gated ion channel gene family of Tetranychus urticae: implications for acaricide toxicology and a novel mutation associated with abamectin resistance

The cys-loop ligand-gated ion channel (cysLGIC) super family of Tetranychus urticae, the two-spotted spider mite, represents the largest arthropod cysLGIC super family described to date and the first characterised one within the group of chelicerates. Genome annotation, phylogenetic analysis and comparison of the cysLGIC subunits with their counterparts in insects reveals that the T. urticae genome encodes for a high number of glutamate- and histamine-gated chloride channel genes (GluCl and HisCl) compared to insects. Three orthologues of the insect γ-aminobutyric acid (GABA)-gated chloride channel gene Rdl were detected. Other cysLGIC groups, such as the nAChR subunits, are more conserved and have clear insect orthologues. Members of cysLGIC family mediate endogenous chemical neurotransmission and they are prime targets of insecticides. Implications for toxicology associated with the identity and specific features of T. urticae family members are discussed. We further reveal the accumulation of known and novel mutations in different GluCl channel subunits (Tu_GluCl1 and Tu_GluCl3) associated with abamectin resistance in T. urticae, and provide genetic evidence for their causality. Our study provides useful toxicological insights for the exploration of the T. urticae cysLGIC subunits as putative molecular targets for current and future chemical control strategies.

Abamectin is metabolized by CYP392A16, a cytochrome P450 associated with high levels of acaricide resistance in Tetranychus urticae

Abamectin is one of the most important insecticides worldwide. It is used against major agricultural pests and insects of public health importance, as well as against endoparasites in animal health. Abamectin has been used successfully for the control of the spider mite Tetranychus urticae, a major agricultural pest with global distribution, an extremely diverse host range, and a remarkable ability to develop resistance against insecticides including abamectin. Target site resistance mutations may explain a large part of resistance, although genetic evidence and transcriptomic data indicated that additional mechanisms may also be implicated in the abamectin resistant phenotype. To investigate a functional link between cytochrome P450-mediated metabolism and abamectin resistance, we recombinantly expressed three cytochrome P450s (CYP392A16, CYP392D8 and CYP392D10) that have been associated with high levels of abamectin resistance in a resistant T. urticae strain isolated from Greece. CYP392A16 was expressed predominately in its P450 form however, both CYP392D8 and CYP392D10 were expressed predominately as P420, despite optimization efforts on expression conditions. CYP392A16 catalyses the hydroxylation of abamectin (Kcat=0.54 pmol/min/pmol P450; Km=45.9 μM), resulting in a substantially less toxic compound as confirmed by bioassays with the partially purified metabolite. However, CYP392A16 did not metabolize hexythiazox, clofentezine and bifenthrin, active ingredients that also showed reduced toxicity in the abamectin resistant strain. Among a number of fluorescent and luminescent substrates screened, Luciferin-ME EGE was preferentially metabolized by CYP392A16, and it may be a potential diagnostic probe for metabolic resistance detection and monitoring.

A gene horizontally transferred from bacteria protects arthropods from host plant cyanide poisoning

Cyanogenic glucosides are among the most widespread defense chemicals of plants. Upon plant tissue disruption, these glucosides are hydrolyzed to a reactive hydroxynitrile that releases toxic hydrogen cyanide (HCN). Yet many mite and lepidopteran species can thrive on plants defended by cyanogenic glucosides. The nature of the enzyme known to detoxify HCN to β-cyanoalanine in arthropods has remained enigmatic. Here we identify this enzyme by transcriptome analysis and functional expression. Phylogenetic analysis showed that the gene is a member of the cysteine synthase family horizontally transferred from bacteria to phytophagous mites and Lepidoptera. The recombinant mite enzyme had both β-cyanoalanine synthase and cysteine synthase activity but enzyme kinetics showed that cyanide detoxification activity was strongly favored. Our results therefore suggest that an ancient horizontal transfer of a gene originally involved in sulfur amino acid biosynthesis in bacteria was co-opted by herbivorous arthropods to detoxify plant produced cyanide. DOI: http://dx.doi.org/10.7554/eLife.02365.001

The economic importance of acaricides in the control of phytophagous mites and an update on recent acaricide mode of action research

Acaricides are one of the cornerstones of an efficient control program for phytophagous mites. An analysis of the global acaricide market reveals that spider mites such as Tetranychus urticae, Panonychus citri and Panonychus ulmi are by far the most economically important species, representing more than 80% of the market. Other relevant mite groups are false spider mites (mainly Brevipalpus), rust and gall mites and tarsonemid mites. Acaricides are most frequently used in vegetables and fruits (74% of the market), including grape vines and citrus. However, their use is increasing in major crops where spider mites are becoming more important, such as soybean, cotton and corn. As revealed by a detailed case study of the Japanese market, major shifts in acaricide use are partially driven by resistance development and the commercial availability of compounds with novel mode of action. The importance of the latter cannot be underestimated, although some compounds are successfully used for more than 30 years. A review of recent developments in mode of action research is presented, as such knowledge is important for devising resistance management programs. This includes spirocyclic keto-enols as inhibitors of acetyl-CoA carboxylase, the carbazate bifenazate as a mitochondrial complex III inhibitor, a novel class of complex II inhibitors, and the mite growth inhibitors hexythiazox, clofentezine and etoxazole that interact with chitin synthase I.

Genotype to phenotype, the molecular and physiological dimensions of resistance in arthropods.

The recent accumulation of molecular studies on mutations in insects, ticks and mites conferring resistance to insecticides, acaricides and biopesticides is reviewed. Resistance is traditionally classified by physiological and biochemical criteria, such as target-site insensitivity and metabolic resistance. However, mutations are discrete molecular changes that differ in their intrinsic frequency, effects on gene dosage and fitness consequences. These attributes in turn impact the population genetics of resistance and resistance management strategies, thus calling for a molecular genetic classification. Mutations in structural genes remain the most abundantly described, mostly in genes coding for target proteins. These provide the most compelling examples of parallel mutations in response to selection. Mutations causing upregulation and downregulation of genes, both in cis (in the gene itself) and in trans (in regulatory processes) remain difficult to characterize precisely. Gene duplications and gene disruption are increasingly reported. Gene disruption appears prevalent in the case of multiple, hetero-oligomeric or redundant targets.