Charles Robin

Identification of candidate genes for fenvalerate resistance in Helicoverpa armigera using cDNA-AFLP

Publisher: Wiley

Date: 08-07-2008

DOI: 10.1111/J.1365-2583.2008.00809.X

Abstract: A cDNA- lified fragment length polymorphisms approach was undertaken to screen for candidate genes associated with fenvalerate resistance in the AN02 strain of Helicoverpa armigera. Larvae and adults of this strain manifest approximately 50-fold resistance, which is suppressible by piperonyl butoxide and controlled by the semidominant gene RFen1 previously mapped to AFLP Linkage Group 13. Two cytochrome P450s (CYP337B1 and CYP4S1), one carboxylesterase-like protein and one glutathione transferase were found to be constitutively upregulated in resistant insects. Mapping of these potential detoxification genes showed that one of them, the novel P450 CYP337B1, was tightly linked to the resistance locus. This suggests that the RFen1(R) allele has a cis-acting effect on CYP337B1 expression, and possible trans-acting effects on expression of other genes.

The spread of resistance to imidacloprid is restricted by thermotolerance in natural populations of Drosophila melanogaster

Publisher: Springer Science and Business Media LLC

Date: 18-03-2019

DOI: 10.1038/S41559-019-0837-Y

Abstract: Imidacloprid, the world's most used insecticide, has caused considerable controversy due to harmful effects on non-pest species and increasing evidence showing that insecticides have become the primary selective force in many insect species. The genetic response to insecticides is heterogeneous across populations and environments, leading to more complex patterns of genetic variation than previously thought. This motivated the investigation of imidacloprid resistance at different temperatures in natural populations of Drosophila melanogaster originating from four climate extremes replicated across two continents. Population and quantitative genomic analysis, supported by functional tests, have revealed a mixed genetic architecture to resistance involving major genes (Paramyosin and Nicotinic-Acetylcholine Receptor Alpha 3) and polygenes with a major trade-off with thermotolerance. Reduced genetic differentiation at resistance-associated loci indicated enhanced gene flow at these loci. Resistance alleles showed stronger evidence of positive selection in temperate populations compared to tropical populations in which chromosomal inversions In(2 L)t, In(3 R)Mo and In(3 R)Payne harbour susceptibility alleles. Polygenic architecture and ecological factors should be considered when developing sustainable management strategies for both pest and beneficial insects.

Cisandtrans-acting variants contribute to survivorship in a naïveDrosophila melanogasterpopulation exposed to ryanoid insecticides

Publisher: Cold Spring Harbor Laboratory

Date: 20-12-2018

DOI: 10.1101/502161

Abstract: Insecticide resistance is a paradigm of microevolution and insecticides are responsible for the strongest cases of recent selection in the genome of Drosophila melanogaster . Here we use a naïve population and a novel insecticide class to examine the ab initio genetic architecture of a potential selective response. Genome wide association studies of chlorantraniliprole susceptibility reveal variation in a gene of major effect, Stretchin Myosin light chain kinase ( Strn-Mlck ), which we validate with linkage mapping and transgenic manipulation of gene expression. We propose that allelic variation in Strn-Mlck alters sensitivity to the calcium depletion attributable to chlorantraniliprole’s mode of action. Genome-wide association studies also reveal a network of genes involved in neuromuscular biology. In contrast, phenotype to transcriptome associations identify differences in constitutive levels of multiple transcripts regulated by cnc, the homologue of mammalian Nrf2. This suggests that genetic variation acts in trans to regulate multiple metabolic enzymes in this pathway. The most outstanding association is with the transcription level of Cyp12d1 which is also affected in cis by copy number variation. Transgenic overexpression of Cyp12d1 reduces susceptibility to both chlorantraniliprole and the closely related insecticide cyantraniliprole. This systems genetics study reveals multiple allelic variants segregating at intermediate frequency in a population that is completely naïve to this new insecticide chemistry and it adumbrates a selective response among natural populations to these chemicals. Around the world insecticides are being deregistered and banned, as their environmental costs are deemed too great or their efficacy against pest insects is reduced through the evolution of insecticide resistance. With the introduction of replacement insecticides comes the responsibility to assess the way new insecticides perturb various levels of biological systems from insect physiology to ecosystems. We used a systems genetics approach to identify genetic variants affecting survivorship of Drosophila melanogaster exposed to chlorantraniliprole. The study population was completely naïve to this insecticide chemistry and yet we find associations with variants in neuromuscular genes and co-regulated detoxification genes. We predict that these variants will increase in populations of this ‘sentinel species’ as these insecticides are applied in the environment.

Identifying candidate detoxification genes in the ecdysteroid kinase-like (EcKL) and cytochrome P450 gene families inDrosophila melanogasterby integrating evolutionary and transcriptomic data

Publisher: Cold Spring Harbor Laboratory

Date: 19-02-2020

DOI: 10.1101/2020.02.17.951962

Abstract: The capacity to detoxify toxic compounds is essential for adaptation to the ecological niches of many organisms, especially insects. However, detoxification in insects is often viewed through the lens of mammalian detoxification research, even though the organ and enzyme systems involved have erged for over half a billion years. Phosphorylation is a non-canonical phase II detoxification reaction that, among animals, occurs near exclusively in insects, but the enzymes responsible have never been cloned or otherwise identified. We propose the hypothesis that members of the arthropod-specific ecdysteroid kinase-like (EcKL) gene family encode detoxicative kinases. To test this hypothesis, we annotated the EcKL gene family in 12 species of Drosophila and explored their evolution within the genus. Many ancestral EcKL clades are evolutionarily unstable and have experienced repeated gene gain and loss events, while others are conserved as single copy orthologs. Leveraging multiple published gene expression datasets from D. melanogaster , and using the cytochrome P450s—a canonical detoxification family—as a test case, we demonstrate relationships between xenobiotic induction, detoxification tissue-enriched expression and evolutionary instability in the EcKLs and the P450s. We also found previously unreported genomic and transcriptomic variation in a number of EcKLs and P450s associated with toxic stress phenotypes using a targeted phenome-wide association study (PheWAS) approach. Lastly, we devised a systematic method for identifying candidate detoxification genes in large gene families that is concordant with experimentally determined functions of P450 genes in D. melanogaster . Applying this method to the EcKLs suggested a significant proportion of these genes play roles in detoxification, and that the EcKLs may constitute a detoxification gene family in insects. Additionally, we estimate that between 11–16 uncharacterised D. melanogaster P450s are strong detoxification candidates. The poorly characterised ecdysteroid kinase-like (EcKL) gene family is hypothesised to encode enzymes responsible for detoxification by phosphorylation in insects. An integrative ‘detoxification score’ method accurately categorises the known functions of a canonical detoxification family, the cytochrome P450s, and suggests many EcKLs are also involved in detoxification. A targeted phenome-wide association study finds novel associations between EcKL/P450 variation and a number of toxic stress phenotypes, such as two unlinked EcKL paralogs that are both associated with developmental methylmercury resistance.

Positional cloning of a cyromazine resistance gene in Drosophila melanogaster

Publisher: Wiley

Date: 31-03-2006

DOI: 10.1111/J.1365-2583.2006.00622.X

Abstract: Cyromazine is an effective insecticide used to control dipteran insects. Its precise mode of action is yet to be determined, although it has been suggested that it interferes with the hormone system, sclerotization of the cuticle, or nucleic acid metabolism. To understand the way in which cyromazine acts, we have positionally cloned a cyromazine resistance gene from Drosophila melanogaster. Six cyromazine resistance alleles had previously been generated by ethyl methanasulphonate treatment. Two of these failed to complement each other and here we identify them as having independent non-sense mutations in CG32743, which is an ortholog of Smg1 of worms and mammals and encodes a phosphatidylinositol kinase-like kinase (PIKK). RNAi experiments confirm that cyromazine resistance can be achieved by knocking down CG32743. These are the first cyromazine resistant mutations identified at the nucleotide level. In mammals Smg1 phosphorylates P53 in response to DNA damage. This finding supports the hypothesis that cyromazine interferes with nucleic acid metabolism.

Two independent duplications forming the Cyp307a genes in Drosophila

Publisher: Elsevier BV

Date: 10-2007

DOI: 10.1016/J.IBMB.2007.05.017

Abstract: The conserved relationship between orthologs of many cytochrome P450 genes involved in ecdysone synthesis is not reflected in the evolution of the Drosophila Cyp307a genes. In Drosophila melanogaster Cyp307a1 (spook) and Cyp307a2 (spookier) both play essential roles in ecdysone synthesis and may possess biochemically redundant functions. Using phylogenetic analyses we show that the Drosophila Cyp307a genes were formed from two independent duplication events depicting a complicated evolutionary scenario. An initial duplication, from a Cyp307a2 ancestral gene produced the Cyp307a1 gene that has been maintained only in the Sophophoran subgenus. A second duplication in the Drosophila subgenus formed an additional paralog, Cyp307a3. Microsynteny is conserved for Cyp307a2 throughout the Drosophila species, but is not conserved between Cyp307a1 and Cyp307a3. These are located in different genomic positions in the Sophophora and Drosophila subgenera, respectively. Cyp307a3 appears to encode a functional gene product and is expressed in a different spatial and temporal manner to Cyp307a1. This suggests some level of functional ergence between the Cyp307a paralogs in different Drosophila species.

Gene duplication in the major insecticide target site, Rdl , in Drosophila melanogaster

Publisher: Proceedings of the National Academy of Sciences

Date: 19-08-2013

DOI: 10.1073/PNAS.1311341110

Abstract: The Resistance to Dieldrin gene, Rdl , encodes a GABA-gated chloride channel subunit that is targeted by cyclodiene and phenylpyrazole insecticides. The gene was first characterized in Drosophila melanogaster by genetic mapping of resistance to the cyclodiene dieldrin. The 4,000-fold resistance observed was due to a single amino acid replacement, Ala 301 to Ser. The equivalent change was subsequently identified in Rdl orthologs of a large range of resistant insect species. Here, we report identification of a duplication at the Rdl locus in D. melanogaster . The 113-kb duplication contains one WT copy of Rdl and a second copy with two point mutations: an Ala 301 to Ser resistance mutation and Met 360 to Ile replacement. In iduals with this duplication exhibit intermediate dieldrin resistance compared with single copy Ser 301 homozygotes, reduced temperature sensitivity, and altered RNA editing associated with the resistant allele. Ectopic recombination between Roo transposable elements is involved in generating this genomic rearrangement. The duplication phenotypes were confirmed by construction of a transgenic, artificial duplication integrating the 55.7-kb Rdl locus with a Ser 301 change into an Ala 301 background. Gene duplications can contribute significantly to the evolution of insecticide resistance, most commonly by increasing the amount of gene product produced. Here however, duplication of the Rdl target site creates permanent heterozygosity, providing unique potential for adaptive mutations to accrue in one copy, without abolishing the endogenous role of an essential gene.

Gene identification and proteomic analysis of the esterases of the cotton bollworm, Helicoverpa armigera

Publisher: Elsevier BV

Date: 2010

DOI: 10.1016/J.IBMB.2009.12.002

Abstract: Some of the resistance of Helicoverpa armigera to conventional insecticides such as organophosphates and synthetic pyrethroids appears to be due to metabolic detoxification by carboxylesterases. To investigate the H. armigera carboxyl/cholinesterases, we created a data set of 39 putative paralogous H. armigera carboxyl/cholinesterase sequences from cDNA libraries and other sources. Phylogenetic analysis revealed a close relationship between these sequences and 70 carboxyl/cholinesterases from the recently sequenced genome of the silkworm, Bombyx mori, including several conserved clades of non-catalytic proteins. A juvenile hormone esterase candidate from H. armigera was identified, and B. mori orthologues were proposed for 31% of the sequences examined, however low similarity was found between lepidopteran sequences and esterases previously associated with insecticide resistance from other insect orders. A proteomic analysis of larval esterases then enabled us to match seven of the H. armigera carboxyl/cholinesterase sequences to specific esterase isozymes. All identified sequences were predicted to encode catalytically active carboxylesterases, including six proteins with N-terminal signal peptides and N-glycans, with two also containing C-terminal signals for glycosylphosphatidylinositol anchor attachment. Five of these sequences were matched to zones of activity on native PAGE at relative mobility values previously associated with insecticide resistance in this species.

Adaptation and conservation insights from the koala genome

Publisher: Springer Science and Business Media LLC

Date: 02-07-2018

DOI: 10.1038/S41588-018-0153-5

The spread of resistance to imidacloprid is restricted by thermotolerance in natural populations of Drosophila melanogaster

Publisher: Springer Science and Business Media LLC

Date: 18-03-2019

DOI: 10.1038/S41559-019-0837-Y

Abstract: Imidacloprid, the world's most used insecticide, has caused considerable controversy due to harmful effects on non-pest species and increasing evidence showing that insecticides have become the primary selective force in many insect species. The genetic response to insecticides is heterogeneous across populations and environments, leading to more complex patterns of genetic variation than previously thought. This motivated the investigation of imidacloprid resistance at different temperatures in natural populations of Drosophila melanogaster originating from four climate extremes replicated across two continents. Population and quantitative genomic analysis, supported by functional tests, have revealed a mixed genetic architecture to resistance involving major genes (Paramyosin and Nicotinic-Acetylcholine Receptor Alpha 3) and polygenes with a major trade-off with thermotolerance. Reduced genetic differentiation at resistance-associated loci indicated enhanced gene flow at these loci. Resistance alleles showed stronger evidence of positive selection in temperate populations compared to tropical populations in which chromosomal inversions In(2 L)t, In(3 R)Mo and In(3 R)Payne harbour susceptibility alleles. Polygenic architecture and ecological factors should be considered when developing sustainable management strategies for both pest and beneficial insects.

Molecular cloning of an α-esterase gene cluster on chromosome 3R of Drosophila melanogaster

Publisher: Elsevier BV

Date: 03-1996

DOI: 10.1016/0965-1748(95)00081-X

Abstract: All or part of the alpha-esterase gene cluster in Drosophila melanogaster has been isolated by screening a YAC clone that spans cytological region 84D3-10 with consensus carboxyl/cholinesterase oligonucleotides. The cluster encompasses 11 putative esterase genes within 65 kb of genomic DNA and is one of the largest clusters of related protein-coding genes yet reported in Drosophila. The cluster must include the gene encoding the major alpha-esterase isozyme, EST9, which has previously been mapped to 84D3-5. It probably also includes the genes encoding the EST23, MCE and ALI esterases that have previously been mapped to 84D3-E2. The latter three are homologs of genes involved in organophosphate insecticide resistance in the sheep blowfly, Lucilia cuprina and the housefly, Musca domestica. Sequencing of one of the putative esterase genes in the Drosophila cluster, alpha E1, shows that it would encode features characteristic of an active carboxyl/cholinesterase, including the so-called catalytic triad, the nucleophilic elbow and oxyanion hole. It also shows that the closest relative of alpha E1 amongst previously published esterase sequences is ESTB1, which confers organophosphate resistance in Culex mosquitoes. We argue that we have cloned the D. melanogaster version of a major cluster of esterase genes which have variously mutated to confer organophosphate resistance in erse Diptera.

A proline repeat polymorphism of the frost gene of drosophila melanogaster showing clinal variation but not associated with cold resistance

Publisher: Wiley

Date: 19-06-2012

DOI: 10.1111/J.1365-2583.2012.01149.X

Abstract: Genetic polymorphisms underlying adaptive shifts in thermal responses are poorly known even though studies are providing a detailed understanding of these responses at the cellular and physiological levels. The Frost gene of Drosophila melanogaster is a prime candidate for thermal adaptation it is up-regulated under cold stress and knockdown of this gene influences cold resistance. Here we describe an amino-acid INDEL polymorphism in proline repeat number in the structural component of this gene. The two main repeats, accounting for more than 90% of alleles in eastern Australia, show a strong clinal pattern the 6P allele was at a high frequency in tropical locations, and the 10P allele was common in temperate populations. However, the frequency of these alleles was not associated with three different assays of cold resistance. Adult transcription level of Frost was also unrelated to cold resistance as measured through post chill coma mobility. The functional significance of the proline repeat polymorphism therefore remains unclear despite its clinal pattern. The data also demonstrate the feasibility of using Roche/454 sequencing for establishing clinal patterns.

An Adaptive Allelic Series Featuring Complex Gene Rearrangements

Publisher: Public Library of Science (PLoS)

Date: 20-10-2011

DOI: 10.1371/JOURNAL.PGEN.1002347

Extracellular endonucleases in the midgut of Myzus persicae may limit the efficacy of orally delivered RNAi

Publisher: Springer Science and Business Media LLC

Date: 15-08-2019

DOI: 10.1038/S41598-019-47357-4

Abstract: Myzus persicae is a major pest of many crops including canola and Brassica vegetables, partly because it vectors plant viruses. Previously it has been reported that double-stranded RNA delivered to aphids by injection, artificial diet or transgenic plants has knocked down target genes and caused phenotypic effects. While these studies suggest that RNA interference (RNAi) might be used to suppress aphid populations, none have shown effects sufficient for field control. The current study analyses the efficacy of dsRNA directed against previously reported gene-targets on Green peach aphid ( Myzus persicae ) strains. No silencing effect was observed when dsRNA was delivered in artificial diet with or without transfection reagents. dsRNA produced in planta also failed to induce significant RNAi in M . persicae . Transcriptome analyses of the midgut suggested other potential targets including the Ferritin heavy chain transcripts, but they also could not be knocked down with dsRNA. Here we show that dsRNA is rapidly degraded by midgut secretions of Myzus persicae . Analysis of the transcriptome of the M . persicae midgut revealed that an ortholog of RNases from other insects was abundant.

GWAlpha: Genome-wide estimation of additive effects (alpha) based on trait quantile distribution from pool-sequencing experiments

Publisher: Oxford University Press (OUP)

Date: 21-12-2017

DOI: 10.1093/BIOINFORMATICS/BTW805

Abstract: Sequencing pools of in iduals (Pool-Seq) is a cost-effective way to gain insight into the genetics of complex traits, but as yet no parametric method has been developed to both test for genetic effects and estimate their magnitude. Here, we propose GWAlpha, a flexible method to obtain parametric estimates of genetic effects genome-wide from Pool-Seq experiments. We showed that GWAlpha powerfully replicates the results of Genome-Wide Association Studies (GWAS) from model organisms. We perform simulation studies that illustrate the effect on power of s le size and number of pools and test the method on different experimental data. GWAlpha is implemented in python, designed to run on Linux operating system and tested on Mac OS. It is freely available at flevel/GWAlpha. Supplementary data are available at Bioinformatics online.

Genomic and Transcriptomic Associations Identify a New Insecticide Resistance Phenotype for the Selective Sweep at the Cyp6g1 Locus of Drosophila melanogaster

Publisher: Oxford University Press (OUP)

Date: 08-2016

DOI: 10.1534/G3.116.031054

Abstract: Scans of the Drosophila melanogaster genome have identified organophosphate resistance loci among those with the most pronounced signature of positive selection. In this study, the molecular basis of resistance to the organophosphate insecticide azinphos-methyl was investigated using the Drosophila Genetic Reference Panel, and genome-wide association. Recently released full transcriptome data were used to extend the utility of the Drosophila Genetic Reference Panel resource beyond traditional genome-wide association studies to allow systems genetics analyses of phenotypes. We found that both genomic and transcriptomic associations independently identified Cyp6g1, a gene involved in resistance to DDT and neonicotinoid insecticides, as the top candidate for azinphos-methyl resistance. This was verified by transgenically overexpressing Cyp6g1 using natural regulatory elements from a resistant allele, resulting in a 6.5-fold increase in resistance. We also identified four novel candidate genes associated with azinphos-methyl resistance, all of which are involved in either regulation of fat storage, or nervous system development. In Cyp6g1, we find a demonstrable resistance locus, a verification that transcriptome data can be used to identify variants associated with insecticide resistance, and an overlap between peaks of a genome-wide association study, and a genome-wide selective sweep analysis.

Developmental expression and gene/enzyme identifications in the alpha esterase gene cluster of Drosophila melanogaster

Publisher: Wiley

Date: 15-09-2003

DOI: 10.1046/J.1365-2583.2003.00430.X

Abstract: Here we show how the 10 genes of the alpha esterase cluster of Drosophila melanogaster have erged substantially in their expression profiles. Together with previously described sequence ergence this suggests substantial functional ersification. By peptide mass fingerprinting and in vitro gene expression we have also shown that two of the genes encode the isozymes EST9 (formerly ESTC) and EST23. EST9 is the major 'alpha staining' esterase in zymograms of gut tissues in feeding stages while orthologues of EST23 confer resistance to organophosphorus insecticides in other higher Diptera. The results for EST9 and EST23 concur with previous suggestions that the products of the alpha esterase cluster function in digestion and detoxification of xenobiotic esters. However, many of the other genes in the cluster show developmental or tissue-specific expression that seems inconsistent with such roles. Furthermore, there is generally poor correspondence between the mRNA expression patterns of the remaining eight genes and isozymes previously characterized by standard techniques of electrophoresis and staining, suggesting that the alpha cluster might only account for a small minority of the esterase isozyme profile.

Universal primers for fluorescent labelling of PCR fragments-an efficient and cost-effective approach to genotyping by fluorescence

Publisher: Wiley

Date: 24-01-2012

DOI: 10.1111/J.1755-0998.2011.03104.X

Abstract: Directly labelling locus-specific primers for microsatellite analysis is expensive and a common limitation to small-budget molecular ecology projects. More cost-effective end-labelling of PCR products can be achieved through a three primer PCR approach, involving a fluorescently labelled universal primer in combination with modified locus-specific primers with 5' universal primer sequence tails. This technique has been widely used but has been limited largely due to a lack of available universal primers suitable for co- lifying large numbers of size overlapping loci and without requiring locus-specific PCR conditions to be modified. In this study, we report a suite of four high-performance universal primers that can be employed in a three primer PCR approach for efficient and cost-effective fluorescent end-labelling of PCR fragments. Amplification efficiency is maximized owing to high universal primer Tm values (approximately 60+ °C) that enhance primer versatility and enable higher annealing temperatures to be employed compared with commonly used universal primers such as M13. We demonstrate that these universal primers can be combined with multiple fluorophores to co- lify multiple loci efficiently via multiplex PCR. This method provides a level of multiplexing and PCR efficiency similar to microsatellite fluorescent detection assays using directly labelled primers while dramatically reducing project costs. Primer performance is tested using several alternative PCR strategies that involve both single and multiple fluorophores in single and multiplex PCR across a wide range of taxa.

Genomic innovations, transcriptional plasticity and gene loss underlying the evolution and divergence of two highly polyphagous and invasive Helicoverpa pest species

Publisher: Springer Science and Business Media LLC

Date: 31-07-2017

DOI: 10.1186/S12915-017-0402-6

Sublethal larval exposure to imidacloprid impacts adult behaviour in Drosophila melanogaster

Publisher: Wiley

Date: 19-11-2019

DOI: 10.1111/JEB.13555

Abstract: Pesticides are now chronically found in numerous ecosystems incurring widespread toxic effects on multiple organisms. For insects, the larvae are very exposed to pesticide pollution and the acute effect of insecticides on larvae has been characterized in a range of species. However, the carry-on effects in adults of sublethal exposure occurring in larvae are not well characterized. Here, we use a collection of strains of Drosophila melanogaster differing in their larval resistance to a commonly used insecticide, imidacloprid, and we test the effect of larval exposure on behavioural traits at the adult stage. Focusing on locomotor activity and on courtship and mating behaviour, we observed a significant carry-on effect of imidacloprid exposure. The heritability of activity traits measured in flies exposed to imidacloprid was higher than measured in controls and in these, courtship traits were genetically less correlated from mating success. Altogether, we did not observe a significant effect of the larval insecticide resistance status on adult behavioural traits, suggesting that selection for resistance in larvae does not involve repeatable behavioural changes in adults. This lack of correlation between larval resistance and adult behaviour also suggests that resistance at the larval stage does not necessarily result in increased behavioural resilience at a later life stage. These findings imply that selection for resistance in larvae as well as for behavioural resilience to sublethal exposure in adult will combine and impose a greater evolutionary constraint. Our conclusions further substantiate the need to encompass multiple trait measures and life stages in toxicological assays to properly assess the environmental impact of pesticides.

A comparison of Drosophila melanogaster detoxification gene induction responses for six insecticides, caffeine and phenobarbital

Publisher: Elsevier BV

Date: 12-2006

DOI: 10.1016/J.IBMB.2006.09.004

Abstract: Modifications of metabolic pathways are important in insecticide resistance evolution. Mutations leading to changes in expression levels or substrate specificities of cytochrome P450 (P450), glutathione-S-transferase (GST) and esterase genes have been linked to many cases of resistance with the responsible enzyme shown to utilize the insecticide as a substrate. Many studies show that the substrates of enzymes are capable of inducing the expression of those enzymes. We investigated if this was the case for insecticides and the enzymes responsible for their metabolism. The induction responses for P450s, GSTs and esterases to six different insecticides were investigated using a custom designed microarray in Drosophila melanogaster. Even though these gene families can all contribute to insecticide resistance, their induction responses when exposed to insecticides are minimal. The insecticides spinosad, diazinon, nitenpyram, lufenuron and dicyclanil did not induce any P450, GST or esterase gene expression after a short exposure to high lethal concentrations of insecticide. DDT elicited the low-level induction of one GST and one P450. These results are in contrast to induction responses we observed for the natural plant compound caffeine and the barbituate drug phenobarbital, both of which highly induced a number of P450 and GST genes under the same short exposure regime. Our results indicate that, under the insecticide exposure conditions we used, constitutive over-expression of metabolic genes play more of a role in insect survival than induction of members of these gene families.

The haplolethal genewupAof Drosophila exhibits potential as a target for an X-poisoning gene drive

Publisher: Cold Spring Harbor Laboratory

Date: 26-06-2023

DOI: 10.1101/2023.06.23.546292

Abstract: A synthetic gene drive that targets haplolethal genes on the X-chromosome can skew the sex ratio towards males. Like an ‘X-shredder’ strategy it does not involve ‘homing’ and that has advantages including the reduction in gene drive resistance allele formation. We examine this ‘X-poisoning’ strategy by targeting four of the 11 known X-linked haplolethal/haplosterile genes of Drosophila melanogaster with CRISPR/cas9. We find that targeting the wupA gene during spermatogenesis skews the sex ratio so fewer than 14% of progeny are daughters. That is unless we cross the mutagenic males to X ^ XY female flies that bear attached-X chromosomes, which reverses the inheritance of the poisoned X chromosome so that sons inherit it from their mother in which case only 2% of the progeny are sons. These sex ratio biases suggests that most of the CRISPR/cas9 mutants we induced in the wupA gene are haplolethal but some are recessive lethal. We have used simulations to model release strategies of a gene drive where an X-poisoning Y chromosome is introduced to invasive populations of the pest Drosophila suzukii . While the reproductive output of carriers of X-poisoning drives is less than that of X-shredding drives due to their post-zygotic mode of action, the threshold dynamics of an X-poisoning drive result in more predictable outcomes than X-shredders that tend to exhibit ‘chasing’ more often. Like D. melanogaster and D. suzukii , the wupA orthologs in Anopheles mosquitos are found on X chromosomes making it a viable target in multiple species.

Fighting fly genes

Publisher: Elsevier BV

Date: 02-2007

DOI: 10.1016/J.TIG.2006.12.005

Abstract: Fighting by organisms such as mice and Drosophila provides model systems for investigating the genetic basis of aggression. Recent experiments to dissect male aggressive behaviour in Drosophila melanogaster, using gene expression analysis of selected lines followed by mutant analysis, have identified new candidate genes associated with male aggression, including one strong candidate that encodes a cytochrome P450 enzyme. Here, we describe the study of aggressive behaviour in flies and explore the possibility that cytochrome P450 is involved in aggression.

Transcriptome analysis of Drosophila melanogaster third instar larval ring glands points to novel functions and uncovers a cytochrome p450 required for development

Publisher: Oxford University Press (OUP)

Date: 02-2017

DOI: 10.1534/G3.116.037333

Abstract: In Drosophila melanogaster larvae, the ring gland (RG) is a control center that orchestrates major developmental transitions. It is a composite organ, consisting of the prothoracic gland, the corpus allatum, and the corpora cardiaca, each of which synthesizes and secretes a different hormone. Until now, the RG’s broader developmental roles beyond endocrine secretion have not been explored. RNA sequencing and analysis of a new transcriptome resource from D. melanogaster wandering third instar larval RGs has provided a fascinating insight into the ersity of developmental signaling in this organ. We have found strong enrichment of expression of two gene pathways not previously associated with the RG: immune response and fatty acid metabolism. We have also uncovered strong expression for many uncharacterized genes. Additionally, RNA interference against RG-enriched cytochrome p450s Cyp6u1 and Cyp6g2 produced a lethal ecdysone deficiency and a juvenile hormone deficiency, respectively, flagging a critical role for these genes in hormone synthesis. This transcriptome provides a valuable new resource for investigation of roles played by the RG in governing insect development.

Conservation of copper-transporting P(IB)-type ATPase function

Publisher: Springer Science and Business Media LLC

Date: 07-04-2010

DOI: 10.1007/S10534-010-9332-2

Abstract: Copper-transporting P(IB)-type ATPases are highly conserved, and while unicellular eukaryotes and invertebrates have only one, a gene duplication has occurred during vertebrate evolution. Copper-induced trafficking of mammalian ATP7A and ATP7B from the trans-Golgi Network towards the plasma membrane is critical for their role in copper homeostasis. In polarized epithelial cells ATP7A and ATP7B traffic towards the basolateral and apical membranes respectively. We examined the localization and function of DmATP7, the single Drosophila melanogaster orthologue, in cultured D. melanogaster and mammalian cells to explore the conservation of P(IB)-type ATPase function. Comparative genomic analysis demonstrated motifs involved in basolateral targeting and retention of ATP7A were conserved in DmATP7, whereas ATP7B targeting motifs were not. DmATP7 expression was able to correct the copper hyper-accumulation phenotype of cultured fibroblasts from a Menkes disease patient expressing a null ATP7A allele. DmATP7 was able to transport copper to the cupro-enzyme tyrosinase and under elevated copper conditions DmATP7 was able to traffic towards the plasma membrane and efflux copper, essentially phenocopying ATP7A. When expressed in polarized Madin-Darby Canine Kidney cells, DmATP7 translocated towards the basolateral membrane when exposed to elevated copper, similar to ATP7A. These results demonstrate DmATP7 is able to functionally compensate for the absence of ATP7A, with important trafficking motifs conserved in these distantly related orthologues.

Genomic and transcriptomic analyses in Drosophila suggest that the ecdysteroid kinase-like (EcKL) gene family encodes the ‘detoxification-by-phosphorylation’ enzymes of insects

Publisher: Elsevier BV

Date: 08-2020

DOI: 10.1016/J.IBMB.2020.103429

Copy Number Variation and Transposable Elements Feature in Recent, Ongoing Adaptation at the Cyp6g1 Locus

Publisher: Public Library of Science (PLoS)

Date: 24-06-2010

DOI: 10.1371/JOURNAL.PGEN.1000998

Evolution of genes and genomes on the Drosophila phylogeny

Publisher: Springer Science and Business Media LLC

Date: 11-2007

DOI: 10.1038/NATURE06341

OfftargetFinder: A web tool for species-specific RNAi design

Publisher: Oxford University Press (OUP)

Date: 24-12-2016

DOI: 10.1093/BIOINFORMATICS/BTV747

Abstract: Motivation: RNA interference (RNAi) technology is being developed as a weapon for pest insect control. To maximize the specificity that such an approach affords we have developed a bioinformatic web tool that searches the ever-growing arthropod transcriptome databases so that pest-specific RNAi sequences can be identified. This will help technology developers finesse the design of RNAi sequences and suggests which non-target species should be assessed in the risk assessment process. Availability and implementation: rnai.specifly.org. Contact: crobin@unimelb.edu.au

Reconciliation of a gene network and species tree

Publisher: Elsevier BV

Date: 07-2019

DOI: 10.1016/J.JTBI.2019.04.001

Abstract: The phylogenetic trees of genes and the species which they belong to are similar, but distinct due to various evolutionary processes which affect genes but do not create new species. Reconciliations map the gene tree into the species tree, explaining the discrepancies by events including gene duplications and losses. However, when duplicate genes undergo recombination (a phenomenon known as paralog exchange, or non-allelic homologous recombination), the phylogeny of the genes becomes a network, not a tree. In this paper, we explore how to reconcile a gene network to a species tree with duplications and losses. We propose an extension of the lowest common ancestor (LCA) mapping which solves the problem for tree-child gene networks, show that a restricted version of the problem is polynomial-time solvable and bounds the optimal position of each gene node in the full problem, and show that the full problem is fixed-parameter tractable in the level of the gene network. This provides a formal foundation for the development of efficient algorithms to solve this problem.

Structural Variants and Selective Sweep Foci Contribute to Insecticide Resistance in theDrosophilaGenetic Reference Panel

Publisher: Oxford University Press (OUP)

Date: 11-2018

DOI: 10.1534/G3.118.200619

Abstract: Patterns of nucleotide polymorphism within populations of Drosophila melanogaster suggest that insecticides have been the selective agents driving the strongest recent bouts of positive selection. However, there is a need to explicitly link selective sweeps to the particular insecticide phenotypes that could plausibly account for the drastic selective responses that are observed in these non-target insects. Here, we screen the Drosophila Genetic Reference Panel with two common insecticides malathion (an organophosphate) and permethrin (a pyrethroid). Genome-wide association studies map survival on malathion to two of the largest sweeps in the D. melanogaster genome Ace and Cyp6g1. Malathion survivorship also correlates with lines which have high levels of Cyp12d1, Jheh1 and Jheh2 transcript abundance. Permethrin phenotypes map to the largest cluster of P450 genes in the Drosophila genome, however in contrast to a selective sweep driven by insecticide use, the derived allele seems to be associated with susceptibility. These results underscore previous findings that highlight the importance of structural variation to insecticide phenotypes: Cyp6g1 exhibits copy number variation and transposable element insertions, Cyp12d1 is tandemly duplicated, the Jheh loci are associated with a Bari1 transposable element insertion, and a Cyp6a17 deletion is associated with susceptibility.

Cis - and trans -acting variants contribute to survivorship in a naïve Drosophila melanogaster population exposed to ry

Publisher: Proceedings of the National Academy of Sciences

Date: 07-05-2019

DOI: 10.1073/PNAS.1821713116

Abstract: Around the world insecticides are being deregistered and banned, as their environmental costs are deemed too great or their efficacy against pest insects is reduced through the evolution of insecticide resistance. With the introduction of replacement insecticides comes the responsibility to assess the way new insecticides perturb various levels of biological systems, from insect physiology to ecosystems. We used a systems genetics approach to identify genetic variants affecting survivorship of Drosophila melanogaster exposed to chlorantraniliprole. The study population was completely naïve to this insecticide chemistry and yet we find associations with variants in neuromuscular genes and coregulated detoxification genes. We predict that these variants will increase in populations of this “sentinel species” as these insecticides are applied in the environment.

Smg1 Nonsense Mutations Do Not Abolish Nonsense-Mediated mRNA Decay in Drosophila melanogaster

Publisher: Oxford University Press (OUP)

Date: 09-2005

DOI: 10.1534/GENETICS.105.045674

Abstract: Smg1 is a key component of nonsense-mediated decay (NMD) in Caenorhabditis elegans and mammals. Here we report that two nonsense alleles of the ortholog of Smg1 do not affect NMD in Drosophila melanogaster.

Birth and Death of Genes and Functions in the β-Esterase Cluster of Drosophila

Publisher: Springer Science and Business Media LLC

Date: 18-06-2009

DOI: 10.1007/S00239-009-9236-3

Ecdysteroid kinase-like (EcKL) paralogs confer developmental tolerance to caffeine in Drosophila melanogaster

Publisher: Cold Spring Harbor Laboratory

Date: 16-09-2021

DOI: 10.1101/2021.09.15.460555

Abstract: A unique aspect of metabolic detoxification in insects compared to other animals is the presence of xenobiotic phosphorylation, about which little is currently understood. Our previous work raised the hypothesis that members of the taxonomically restricted ecdysteroid kinase-like (EcKL) gene family encode the enzymes responsible for xenobiotic phosphorylation in the model insect Drosophila melanogaster (Diptera: Ephydroidea)—however, candidate detoxification genes identified in the EcKL family have yet to be functionally validated. Here, we test the hypothesis that EcKL genes in the rapidly evolving Dro5 clade are involved in the detoxification of plant and fungal toxins in D. melanogaster . The mining and reanalysis of existing data indicated multiple Dro5 genes are transcriptionally induced by the plant alkaloid caffeine and that adult caffeine susceptibility is associated with a novel naturally occurring indel in CG31370 (Dro5-8) in the Drosophila Genetic Reference Panel (DGRP). CRISPR-Cas9 mutagenesis of five Dro5 EcKLs substantially decreased developmental tolerance of caffeine, while in idual overexpression of two of these genes— CG31300 (Dro5-1) and CG13659 (Dro5-7)—in detoxification-related tissues increased developmental tolerance. In addition, we found Dro5 loss-of-function animals also have decreased developmental tolerance of the fungal secondary metabolite kojic acid. Taken together, this work provides the first compelling functional evidence that EcKLs encode detoxification enzymes and suggests that EcKLs in the Dro5 clade are involved in the metabolism of multiple ecologically relevant toxins in D. melanogaster . We also propose a biochemical hypothesis for EcKL involvement in caffeine detoxification and highlight the many unknown aspects of caffeine metabolism in D. melanogaster and other insects. Phosphorylation is an under-characterised detoxification reaction in insects Dro5 EcKL genes are good detoxification candidate genes in Drosophila melanogaster Knockout and misexpression of some Dro5 genes modulated tolerance to caffeine Dro5 genes may also confer tolerance to the fungal toxin kojic acid Caffeine tolerance could be adaptive for Drosophila associating with Citrus fruits

Behavioural response to combined insecticide and temperature stress in natural populations of Drosophila melanogaster

Publisher: Wiley

Date: 17-03-2016

DOI: 10.1111/JEB.12844

Abstract: Insecticide resistance evolves extremely rapidly, providing an illuminating model for the study of adaptation. With climate change reshaping species distribution, pest and disease vector control needs rethinking to include the effects of environmental variation and insect stress physiology. Here, we assessed how both long-term adaptation of populations to temperature and immediate temperature variation affect the genetic architecture of DDT insecticide response in Drosophila melanogaster. Mortality assays and behavioural assays based on continuous activity monitoring were used to assess the interaction between DDT and temperature on three field-derived populations from climate extremes (Raleigh for warm temperate, Tasmania for cold oceanic and Queensland for hot tropical). The Raleigh population showed the highest mortality to DDT, whereas the Queensland population, epicentre for derived alleles of the resistance gene Cyp6g1, showed the lowest. Interaction between insecticide and temperature strongly affected mortality, particularly for the Tasmanian population. Activity profiles analysed using self-organizing maps show that the insecticide promoted an early response, whereas elevated temperature promoted a later response. These distinctive early or later activity phases revealed similar responses to temperature and DDT dose alone but with more or less genetic variance depending on the population. This change in genetic variance among populations suggests that selection particularly depleted genetic variance for DDT response in the Queensland population. Finally, despite similar (co)variation between traits in benign conditions, the genetic responses across population differed under stressful conditions. This showed how stress-responsive genetic variation only reveals itself in specific conditions and thereby escapes potential trade-offs in benign environments.

Phylogenomics of the ecdysteroid kinase-like (EcKL) gene family in insects highlights roles in both steroid hormone metabolism and detoxification

Publisher: Cold Spring Harbor Laboratory

Date: 07-2023

DOI: 10.1101/2023.06.29.546846

Abstract: The evolutionary dynamics of large gene families can offer important insights into the functions of their in idual members. While the ecdysteroid kinase-like (EcKL) gene family has previously been linked to the metabolism of both steroid moulting hormones and xenobiotic toxins, the functions of nearly all EcKL genes are unknown and there is little information on their evolution across all insects. Here, we perform comprehensive phylogenetic analyses on a manually annotated set of EcKL genes from 140 insect genomes, revealing the gene family is comprised of at least 13 subfamilies that differ in retention and stability. Our results show the only two genes known to encode ecdysteroid kinases belong to different subfamilies and therefore ecdysteroid metabolism functions must be spread throughout the EcKL family. We also provide comparative phylogenomic evidence that EcKLs are involved in detoxification across insects, with positive associations between family size and dietary chemical complexity, and also find similar evidence for the cytochrome P450 and glutathione S-transferase gene families. Unexpectedly, we find that the size of the clade containing a known ecdysteroid kinase is positively associated with host plant taxonomic ersity in Lepidoptera, possibly suggesting multiple functional shifts between hormone and xenobiotic metabolism. This work provides a robust framework for future functional studies into the EcKL gene family and opens promising new avenues for exploring the genomic basis of dietary adaptation in insects, including the classically studied co-evolution of butterflies with their host plants. The ecdysteroid kinase-like (EcKL) gene family has been linked to both steroid inactivation and detoxification in insects, but the functions of most of its members are unknown. Here, we study the evolutionary history of the EcKLs and showcase how phylogenetics can inform the functional characterisation of enzyme families. EcKL family size varies over an order of magnitude and is associated with chemically complex diets, implicating numerous genes in detoxification. We also find a surprising link between a steroid-metabolising clade and host plant ersity in butterflies and moths, suggesting new detoxification functions may evolve through an insect-plant ‘chemical arms-race’. This work advances numerous functional hypotheses for multiple EcKL clades and proposes a new classification system for this poorly characterised gene family.

Molecular Evolution of Glutathione S-Transferases in the Genus Drosophila

Publisher: Oxford University Press (OUP)

Date: 11-2007

DOI: 10.1534/GENETICS.107.075838

Abstract: As classical phase II detoxification enzymes, glutathione S-transferases (GSTs) have been implicated in insecticide resistance and may have evolved in response to toxins in the niche-defining feeding substrates of Drosophila species. We have annotated the GST genes of the 12 Drosophila species with recently sequenced genomes and analyzed their molecular evolution. Gene copy number variation is attributable mainly to unequal crossing-over events in the large δ and ε clusters. Within these gene clusters there are also GST genes with slowly erging orthologs. This implies that they have their own unique functions or have spatial/temporal expression patterns that impose significant selective constraints. Searches for positively selected sites within the GSTs identified G171K in GSTD1, a protein that has previously been shown to be capable of metabolizing the insecticide DDT. We find that the same radical substitution (G171K) in the substrate-binding domain has occurred at least three times in the Drosophila radiation. Homology-modeling places site 171 distant from the active site but adjacent to an alternative DDT-binding site. We propose that the parallel evolution observed at this site is an adaptive response to an environmental toxin and that sequencing of historical alleles suggests that this toxin was not a synthetic insecticide.

Acinetobacter baumannii K27 and K44 capsular polysaccharides have the same K unit but different structures due to the presence of distinct wzy genes in otherwise closely related K gene clusters.

Publisher: Oxford University Press (OUP)

Date: 27-12-2016

DOI: 10.1093/GLYCOB/CWV168

Abstract: Capsular polysaccharides (CPSs), from Acinetobacter baumannii isolates 1432, 4190 and NIPH 70, which have related gene content at the K locus, were examined, and the chemical structures established using 2D(1)H and(13)C NMR spectroscopy. The three isolates produce the same pentasaccharide repeat unit, which consists of 5-N-acetyl-7-N-[(S)-3-hydroxybutanoyl] (major) or 5,7-di-N-acetyl (minor) derivatives of 5,7-diamino-3,5,7,9-tetradeoxy-D-glycero-D-galacto-non-2-ulosonic (legionaminic) acid (Leg5Ac7R), D-galactose, N-acetyl-D-galactosamine and N-acetyl-D-glucosamine. However, the linkage between repeat units in NIPH 70 was different to that in 1432 and 4190, and this significantly alters the CPS structure. The KL27 gene cluster in 4190 and KL44 gene cluster in NIPH 70 are organized identically and contain lga genes for Leg5Ac7R synthesis, genes for the synthesis of the common sugars, as well as anitrA2 initiating transferase and four glycosyltransferases genes. They share high-level nucleotide sequence identity for corresponding genes, but differ in the wzy gene encoding the Wzy polymerase. The Wzy proteins, which have different lengths and share no similarity, would form the unrelated linkages in the K27 and K44 structures. The linkages formed by the four shared glycosyltransferases were predicted by comparison with gene clusters that synthesize related structures. These findings unambiguously identify the linkages formed by WzyK27 and WzyK44, and show that the presence of different wzy genes in otherwise closely related K gene clusters changes the structure of the CPS. This may affect its capacity as a protective barrier for A. baumannii.

Recognition and Detoxification of the Insecticide DDT by Drosophila melanogaster Glutathione S-Transferase D1

Publisher: Elsevier BV

Date: 06-2010

DOI: 10.1016/J.JMB.2010.04.020

Abstract: GSTD1 is one of several insect glutathione S-transferases capable of metabolizing the insecticide DDT. Here we use crystallography and NMR to elucidate the binding of DDT and glutathione to GSTD1. The crystal structure of Drosophila melanogaster GSTD1 has been determined to 1.1 A resolution, which reveals that the enzyme adopts the canonical GST fold but with a partially occluded active site caused by the packing of a C-terminal helix against one wall of the binding site for substrates. This helix would need to unwind or be displaced to enable catalysis. When the C-terminal helix is removed from the model of the crystal structure, DDT can be computationally docked into the active site in an orientation favoring catalysis. Two-dimensional (1)H,(15)N heteronuclear single-quantum coherence NMR experiments of GSTD1 indicate that conformational changes occur upon glutathione and DDT binding and the residues that broaden upon DDT binding support the predicted binding site. We also show that the ancestral GSTD1 is likely to have possessed DDT dehydrochlorinase activity because both GSTD1 from D. melanogaster and its sibling species, Drosophila simulans, have this activity.

Population genomics provides insights into lineage divergence and local adaptation within the cotton bollworm

Publisher: Wiley

Date: 27-01-2022

DOI: 10.1111/1755-0998.13581

Abstract: The cotton bollworm Helicoverpa armigera is a cosmopolitan pest and its erse habitats plausibly contribute to the formation of erse lineages. Despite the significant threat it poses to economic crops worldwide, its evolutionary history and genetic basis of local adaptation are poorly understood. In this study, we de novo assembled a high‐quality chromosome‐level reference genome of H . a . armigera (contig N50 = 7.34 Mb), with 99.13% of the HaSCD2 assembly assigned to 31 chromosomes (Z‐chromosome + 30 autosomes). We constructed an ultradense variation map across 14 cotton bollworm populations and identified a novel lineage in northwestern China. Historical inference showed that effective population size changes coincided with global temperature fluctuation. We identified nine differentiated genes in the three H . armigera lineages ( H . a . armigera , H . a . conferta and the new northwestern Chinese lineage), of which per and clk genes are involved in circadian rhythm. Selective sweep analyses identified a series of Gene Ontology categories related to climate adaptation, feeding behaviour and insecticide tolerance. Our findings reveal fundamental knowledge of the local adaptation of different cotton bollworm lineages and will guide the formulation of cotton bollworm management measures at different scales.

Ecdysteroid kinase-like (EcKL) paralogs confer developmental tolerance to caffeine in Drosophila melanogaster

Publisher: Elsevier BV

Date: 2022

DOI: 10.1016/J.CRIS.2022.100030

The Molecular Evolution of Cytochrome P450 Genes within and between Drosophila Species

Publisher: Oxford University Press (OUP)

Date: 20-04-2014

DOI: 10.1093/GBE/EVU083

Population differentiation between Australian and ChineseHelicoverpa armigeraoccurs in distinct blocks on the Z-chromosome

Publisher: Cambridge University Press (CUP)

Date: 05-02-2018

DOI: 10.1017/S0007485318000081

Abstract: Over the last 40 years, many types of population genetic markers have been used to assess the population structure of the pest moth species Helicoverpa armigera . While this species is highly vagile, there is evidence of inter-continental population structure. Here, we examine Z-chromosome molecular markers within and between Chinese and Australian populations. Using 1352 polymorphic sites from 40 Z-linked loci, we compared two Chinese populations of moths separated by 700 km and found virtually no population structure ( n = 41 and n = 54, with % of variation discriminating between populations). The levels of nucleotide ersity within these populations were consistent with previous estimates from introns in Z-linked genes of Australian s les (π = 0.028 vs. 0.03). Furthermore, all loci surveyed in these Chinese populations showed a skew toward rare variants, with ten loci having a significant Tajima's D statistic, suggesting that this species could have undergone a population expansion. Eight of the 40 loci had been examined in a previous study of Australian moths, of which six revealed very little inter-continental population structure. However, the two markers associated with the Cyp303a1 locus that has previously been proposed to be a target of a selective sweep, exhibited allele structuring between countries. Using a separate dataset of 19 Australian and four Chinese moths, we scanned the molecular variation distributed across the entire Z-chromosome and found distinct blocks of differentiation that include the region containing Cyp303a1 . We recommend some of these loci join those associated with insecticide resistance to form a set of genes best suited to analyzing population structure in this global pest.

What can genetic association panels tell us about evolutionary processes in insects?

Publisher: Elsevier BV

Date: 02-2019

DOI: 10.1016/J.COIS.2018.12.004

Abstract: If we are to fully comprehend the evolution of insect ersity at a genomic level we need to understand how natural selection can alter genetically encoded characters within populations. Genetic association panels have the potential to be standard bearers in this endeavour. They enable the mapping of phenotypes to genotypes at unprecedented resolution while simultaneously providing population genomic s les that can be interrogated for the tell-tale signs of selection. Analyses of these panels promise to elucidate the entanglement of gene ontologies, pathways, developmental processes and evolutionary constraints, and inform how these are shaped by adaptation.

Mining insect genomes for functionally affiliated genes

Publisher: Elsevier BV

Date: 02-2019

DOI: 10.1016/J.COIS.2018.12.006

Abstract: Several hundred insect genome assemblies are already publicly available, and this total grows on a weekly basis. A major challenge now confronting insect science is how best to use genomic data to improve our understanding of insect biology. We consider a framework for genome analysis based on functional affiliation, that is, groups of genes involved in the same biological process or pathway, and explore how such an approach furthers our understanding of several aspects of insect phenotype. We anticipate that this approach will prove useful for future research across the breadth of insect studies, whatever organism or trait it involves.

Structural variants and selective sweep foci contribute to insecticide resistance in theDrosophila melanogastergenetic reference panel

Publisher: Cold Spring Harbor Laboratory

Date: 16-04-2018

DOI: 10.1101/301937

Abstract: Patterns of nucleotide polymorphism within populations of Drosophila melanogaster suggest that insecticides have been the selective agents driving the strongest recent bouts of positive selection. However, there is a need to explicitly link selective sweep loci to the particular insecticide phenotypes that could plausibly account for the drastic selective responses that are observed in these non-target insects. Here, we screen the Drosophila Genetic Reference Panel with two common insecticides malathion (an organophosphate) and permethrin (a pyrethroid). Genome wide association studies map ‘survival-on-malathion’ to two of the largest sweeps in the D. melanogaster genome Ace and Cyp6g1 . Malathion survivorship also correlates with lines which have high levels of Cyp12d1 and Jheh1 and Jheh2 transcript abundance. Permethrin phenotypes map to the largest cluster of P450 genes in the Drosophila genome, however in contrast to a selective sweep driven by insecticide use, the derived state seems to be associated with susceptibility. These results underscore previous findings that highlight the importance of structural variation to insecticide phenotypes: Cyp6g1 exhibits copy number variation and transposable element insertions, Cyp12d1 is tandemly duplicated, the Jheh loci are associated with a Bari1 transposable element insertion, and a Cyp6a17 deletion is associated with susceptibility.

Low doses of the organic insecticide spinosad trigger lysosomal defects, elevated ROS, lipid dysregulation, and neurodegeneration in flies

Publisher: eLife Sciences Publications, Ltd

Date: 22-02-2022

DOI: 10.7554/ELIFE.73812

Abstract: Large-scale insecticide application is a primary weapon in the control of insect pests in agriculture. However, a growing body of evidence indicates that it is contributing to the global decline in population sizes of many beneficial insect species. Spinosad emerged as an organic alternative to synthetic insecticides and is considered less harmful to beneficial insects, yet its mode of action remains unclear. Using Drosophila , we show that low doses of spinosad antagonize its neuronal target, the nicotinic acetylcholine receptor subunit alpha 6 (nAChRα6), reducing the cholinergic response. We show that the nAChRα6 receptors are transported to lysosomes that become enlarged and increase in number upon low doses of spinosad treatment. Lysosomal dysfunction is associated with mitochondrial stress and elevated levels of reactive oxygen species (ROS) in the central nervous system where nAChRα6 is broadly expressed. ROS disturb lipid storage in metabolic tissues in an nAChRα6-dependent manner. Spinosad toxicity is ameliorated with the antioxidant N-acetylcysteine amide. Chronic exposure of adult virgin females to low doses of spinosad leads to mitochondrial defects, severe neurodegeneration, and blindness. These deleterious effects of low-dose exposures warrant rigorous investigation of its impacts on beneficial insects.

Dissecting the Insect Metabolic Machinery Using Twin Ion Mass Spectrometry: A Single P450 Enzyme Metabolizing the Insecticide Imidacloprid in Vivo

Publisher: American Chemical Society (ACS)

Date: 11-03-2014

DOI: 10.1021/AC404188G

Abstract: Insecticide resistance is one of the most prevalent ex les of anthropogenic genetic change, yet our understanding of metabolic-based resistance remains limited by the analytical challenges associated with rapidly tracking the in vivo metabolites of insecticides at nonlethal doses. Here, using twin ion mass spectrometry analysis of the extracts of whole Drosophila larvae and excreta, we show that (i) eight metabolites of the neonicotinoid insecticide, imidacloprid, can be detected when formed by susceptible larval genotypes and (ii) the specific overtranscription of a single gene product, Cyp6g1, associated with the metabolic resistance to neonicotinoids, results in a significant increase in the formation of three imidacloprid metabolites that are formed in C-H bond activation reactions that is, Cyp6g1 is directly linked to the enhanced metabolism of imidacloprid in vivo. These results establish a rapid and sensitive method for dissecting the metabolic machinery of insects by directly linking single gene products to insecticide metabolism.

Insights into DDT Resistance from the Drosophila melanogaster Genetic Reference Panel

Publisher: Oxford University Press (OUP)

Date: 21-09-2017

DOI: 10.1534/GENETICS.117.300310

Abstract: Insecticide resistance is considered a classic model of microevolution, where a strong selective agent is applied to a large natural population, resulting in a change in frequency of alleles that confer resistance. While many insecticide resistance variants have been characterized at the gene level, they are typically single genes of large effect identified in highly resistant pest species. In contrast, multiple variants have been implicated in DDT resistance in Drosophila melanogaster however, only the Cyp6g1 locus has previously been shown to be relevant to field populations. Here we use genome-wide association studies (GWAS) to identify DDT-associated polygenes and use selective sweep analyses to assess their adaptive significance. We identify and verify two candidate DDT resistance loci. A largely uncharacterized gene, CG10737, has a function in muscles that ameliorates the effects of DDT, while a putative detoxifying P450, Cyp6w1, shows compelling evidence of positive selection.

Rethinking the ecdysteroid source during Drosophila pupal–adult development

Publisher: Elsevier BV

Date: 2023

DOI: 10.1016/J.IBMB.2022.103891

High nucleotide diversity and limited linkage disequilibrium in Helicoverpa armigera facilitates the detection of a selective sweep

Publisher: Springer Science and Business Media LLC

Date: 15-07-2015

DOI: 10.1038/HDY.2015.53

Australian National University

Location: Australia

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University Of Technology Sydney

Location: Australia

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University Of Sydney

Location: Australia

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University Of Melbourne

Location: Australia

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Discovery Projects - Grant ID: DP190102512

Start Date: 03-2019

End Date: 12-2023

Amount: $552,000.00

Summary: Potential of gene drives to eliminate incursions of Drosophila suzukii. This project aims to test the efficacy and evolutionary stability of different types of gene drives, and model whether gene drives can be used to eliminate incursions of Drosophila suzukii into Australia. It is now possible to use genome editing technology to alter populations of organisms using ‘gene drives’. Multiple strategies have been conceived with a major distinction between those that aim to eliminate populations ver .... Potential of gene drives to eliminate incursions of Drosophila suzukii. This project aims to test the efficacy and evolutionary stability of different types of gene drives, and model whether gene drives can be used to eliminate incursions of Drosophila suzukii into Australia. It is now possible to use genome editing technology to alter populations of organisms using ‘gene drives’. Multiple strategies have been conceived with a major distinction between those that aim to eliminate populations versus those that aim to modify populations. This project will examine these strategies in two fly species, the model, Drosophila melanogaster and the devastating pest of horticulture, Drosophila suzukii. The project expects to assess a gene drive strategy to control the invasive pest that threatens the Australian soft-skinned fruit industries.
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Funder: Australian Research Council

Discovery Projects - Grant ID: DP0557497

Start Date: 2005

End Date: 12-2008

Amount: $280,000.00

Summary: Using comparative genomics to identify genes responsible for adaptation to environmental toxins. The US National Human Genome Research Institute has committed to sequencing the genomes of ten different Drosophila (fly) species. We will search these genomes, and two others that are already available, for genes that allow flies to cope with environmental toxins found in the plants upon which they feed and breed. These same genes have the potential to degrade many of the insecticides used to con .... Using comparative genomics to identify genes responsible for adaptation to environmental toxins. The US National Human Genome Research Institute has committed to sequencing the genomes of ten different Drosophila (fly) species. We will search these genomes, and two others that are already available, for genes that allow flies to cope with environmental toxins found in the plants upon which they feed and breed. These same genes have the potential to degrade many of the insecticides used to control insect pests. Hence, this research will contribute to ongoing efforts to minimize the threat to agriculture posed by the insecticide resistance that frequently evolves in pest species.
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Funder: Australian Research Council

Discovery Projects - Grant ID: DP0985013

Start Date: 03-2009

End Date: 12-2013

Amount: $780,000.00

Summary: Testing the DNA decay hypothesis of ecological specialization. Australia's biodiversity has been increasingly threatened by climate change and fragmentation from habitat loss. To conserve biodiversity we need to identify species most at risk of extinction. One way species avoid extinction is to evolve and adapt to changing conditions, however, it now appears that many species have a limited adaptive potential. Here we develop and test a new idea that helps to predict species most threatened b .... Testing the DNA decay hypothesis of ecological specialization. Australia's biodiversity has been increasingly threatened by climate change and fragmentation from habitat loss. To conserve biodiversity we need to identify species most at risk of extinction. One way species avoid extinction is to evolve and adapt to changing conditions, however, it now appears that many species have a limited adaptive potential. Here we develop and test a new idea that helps to predict species most threatened by climate change and other types of stresses. We also identify the sets of genes that are involved in adapting to dry/cold conditions and toxins. This information provides a rapid way of identifying species most at risk and least likely to adapt, and a new perspective on Australia's biodiversity.
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Funder: Australian Research Council

Discovery Projects - Grant ID: DP230101111

Start Date: 07-2023

End Date: 07-2026

Amount: $590,000.00

Summary: Using genetic Allee effects to manage invasive populations. An invasion can be started with only a small number of individuals, and it is very difficult to reliably detect these individuals. This project aims to develop new genetic technology that can send small founder populations extinct without affecting large populations. This technology removes the problem of having to detect small populations; these small populations will go extinct on their own, without the need for management interventio .... Using genetic Allee effects to manage invasive populations. An invasion can be started with only a small number of individuals, and it is very difficult to reliably detect these individuals. This project aims to develop new genetic technology that can send small founder populations extinct without affecting large populations. This technology removes the problem of having to detect small populations; these small populations will go extinct on their own, without the need for management intervention. This technology could be used to prevent establishment and spread of invasive species and agricultural pests. Through a combination of experimentation and modelling, the project develops this technology and assesses its use in applied problems ranging across environment, agriculture, and health.
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Funder: Australian Research Council