Cross-pollination in seed-blended refuge and selection for Vip3A resistance in a lepidopteran pest as detected by genomic monitoring.

The evolution of pest resistance to management tools reduces productivity and results in economic losses in agricultural systems. To slow its emergence and spread, monitoring and prevention practices are implemented in resistance management programs. Recent work suggests that genomic approaches can identify signs of emerging resistance to aid in resistance management. Here, we empirically examined the sensitivity of genomic monitoring for resistance management in transgenic Bt crops, a globally important agricultural innovation. Whole genome resequencing of wild North American Helicoverpa zea collected from non-expressing refuge and plants expressing Cry1Ab confirmed that resistance-associated signatures of selection were detectable after a single generation of exposure. Upon demonstrating its sensitivity, we applied genomic monitoring to wild H. zea that survived Vip3A exposure resulting from cross-pollination of refuge plants in seed-blended plots. Refuge seed interplanted with transgenic seed exposed H. zea to sublethal doses of Vip3A protein in corn ears and was associated with allele frequency divergence across the genome. Some of the greatest allele frequency divergence occurred in genomic regions adjacent to a previously described candidate gene for Vip3A resistance. Our work highlights the power of genomic monitoring to sensitively detect heritable changes associated with field exposure to Bt toxins and suggests that seed-blended refuge will likely hasten the evolution of resistance to Vip3A in lepidopteran pests.

Rapid Adaptation and Interspecific Introgression in the North American Crop Pest Helicoverpa zea.

Insect crop pests threaten global food security. This threat is amplified through the spread of nonnative species and through adaptation of native pests to control measures. Adaptations such as pesticide resistance can result from selection on variation within a population, or through gene flow from another population. We investigate these processes in an economically important noctuid crop pest, Helicoverpa zea, which has evolved resistance to a wide range of pesticides. Its sister species Helicoverpa armigera, first detected as an invasive species in Brazil in 2013, introduced the pyrethroid-resistance gene CYP337B3 to South American H. zea via adaptive introgression. To understand whether this could contribute to pesticide resistance in North America, we sequenced 237 H. zea genomes across 10 sample sites. We report H. armigera introgression into the North American H. zea population. Two individuals sampled in Texas in 2019 carry H. armigera haplotypes in a 4 Mbp region containing CYP337B3. Next, we identify signatures of selection in the panmictic population of nonadmixed H. zea, identifying a selective sweep at a second cytochrome P450 gene: CYP333B3. We estimate that its derived allele conferred a ∼5% fitness advantage and show that this estimate explains independently observed rare nonsynonymous CYP333B3 mutations approaching fixation over a ∼20-year period. We also detect putative signatures of selection at a kinesin gene associated with Bt resistance. Overall, we document two mechanisms of rapid adaptation: the introduction of fitness-enhancing alleles through interspecific introgression, and selection on intraspecific variation.

Evolutionary rescue in resistance to pesticides.

Evolutionary rescue occurs when the genetic evolution of adaptation saves a population from decline or extinction after environmental change. The evolution of resistance to pesticides is a special scenario of abrupt environmental change, where rescue occurs under (very) strong selection for one or a few de novo resistance mutations of large effect. Here, a population genetic model of evolutionary rescue with density-dependent population change is developed, with a focus on deriving results that are important to resistance management. Massive stochastic simulations are used to generate observations, which are accurately predicted using analytical approximations. Key results include the probability density function for the time to resistance and the probability of population extinction. The distribution of resistance times shows a lag period, a narrow peak and a long tail. Surprisingly, the mean time to resistance can increase with the strength of selection because, if a mutation does not occur early on, then its emergence is delayed by the pesticide reducing the population size. The probability of population extinction shows a sharp transition, in that when extinction is possible, it is also highly likely. Consequently, population suppression and (local) eradication can be theoretically achievable goals, as novel strategies to delay resistance evolution.

A relaxin receptor gene RpGPCR41 is involved in the resistance of Rhopalosiphum padi to pyrethroids.

Rhopalosiphum padi is a global pest that poses a significant threat to wheat crops and has developed resistance to various insecticides. G protein-coupled receptors (GPCRs), known for their crucial role in signaling and biological processes across insect species, have recently gained attention as a potential target for insecticides. GPCR has the potential to contribute to insect resistance through the regulation of P450 gene expression. However, GPCRs in R. padi remained unexplored until this study. We identified a total of 102 GPCRs in R. padi, including 81 receptors from family A, 10 receptors from family B, 8 receptors from family C, and 3 receptors from family D. Among these GPCR genes, 16 were up-regulated in both lambda-cyhalothrin and bifenthrin-resistant strains of R. padi (LC-R and BIF-R). A relaxin receptor gene, RpGPCR41, showed the highest up-regulated expression in both the resistant strains, with a significant increase of 14.3-fold and 22.7-fold compared to the susceptible strain (SS). RNA interference (RNAi) experiments targeting the relaxin receptor significantly increase the mortality of R. padi when exposed to the LC50 concentration of lambda-cyhalothrin and bifenthrin. The expression levels of five P450 genes (RpCYP6CY8, RpCYP6DC1, RpCYP380B1, RpCYP4CH2, and RpCYP4C1) were significantly down-regulated following knockdown of RpGPCR41 in LC-R and BIF-R strains. Our results highlight the involvement of GPCR gene overexpression in the resistance of R. padi to pyrethroids, providing valuable insights into the mechanisms underlying aphid resistance and a potential target for aphid control.

Population genomic insights into invasion success in a polyphagous agricultural pest, Halyomorpha halys.

Invasive species are increasingly threatening ecosystems and agriculture by rapidly expanding their range and adapting to environmental and human-imposed selective pressures. The genomic mechanisms that underlie such rapid changes remain unclear, especially for agriculturally important pests. Here, we used genome-wide polymorphisms derived from native, invasive, and intercepted samples and populations of the brown marmorated stink bug (BMSB), Halyomorpha halys, to gain insights into population genomics processes that have promoted the successful global invasion of this polyphagous pest. Our analysis demonstrated that BMSB exhibits spatial structure but admixture rates are high among introduced populations, resulting in similar levels of genomic diversity across native and introduced populations. These spatial genomic patterns suggest a complex invasion scenario, potentially with multiple bridgehead events, posing a challenge for accurately assigning BMSB incursions to their source using reduced-representation genomic data. By associating allele frequencies with the invasion status of BMSB populations, we found significantly differentiated single nucleotide polymorphisms (SNPs) located in close proximity to genes for insecticide resistance and olfaction. Comparing variations in allele frequencies among populations for outlier SNPs suggests that BMSB invasion success has probably evolved from standing genetic variation. In addition to being a major nuisance of households, BMSB has caused significant economic losses to agriculture in recent years and continues to expand its range. Despite no record of BMSB insecticide resistance to date, our results show high capacity for potential evolution of such traits, highlighting the need for future sustainable and targeted management strategies.

Global genomic signature reveals the evolution of fall armyworm in the Eastern hemisphere.

The major plant pest fall armyworm (FAW), Spodoptera frugiperda, is native to the Americas and has colonized Africa and Asia within the Eastern hemisphere since 2016, causing severe damage to multiple agricultural crop species. However, the genetic origin of these invasive populations requires more in-depth exploration. We analysed genetic variation across the genomes of 280 FAW individuals from both the Eastern hemisphere and the Americas. The global range-wide genetic structure of FAW shows that the FAW in America has experienced deep differentiation, largely consistent with the Z-chromosomal Tpi haplotypes commonly used to differentiate 'corn-strain' and 'rice-strain' populations. The invasive populations from Africa and Asia are different from the American ones and have a relatively homogeneous population structure, consistent with the common origin and recent spreading from Africa to Asia. Our analyses suggest that north- and central American 'corn-strain' FAW are the most likely sources of the invasion into the Eastern hemisphere. Furthermore, evidence based on genomic, transcriptomic and mitochondrial haplotype network analyses indicates an earlier, independent introduction of FAW into Africa, with subsequent migration into the recent invasive population.

Incomplete reproductive barriers and genomic differentiation impact the spread of resistance mutations between green- and red-colour morphs of a cosmopolitan mite pest.

Pesticide resistance represents a clear and trackable case of adaptive evolution with a strong societal impact. Understanding the factors associated with the evolution and spread of resistance is imperative to develop sustainable crop management strategies. The two-spotted spider mite Tetranychus urticae, a major crop pest with worldwide distribution and a polyphagous lifestyle, has evolved resistance to most classes of pesticides. Tetranychus urticae exists as either a green- or a red-coloured morph. However, the extent of genetic divergence and reproductive compatibility vary across populations of these colour morphs, complicating their taxonomic resolution at the species level. Here, we studied patterns of genetic differentiation and barriers to gene flow within and between morphs of T. urticae in order to understand the factors that influence the spread of resistance mutations across its populations. We derived multiple iso-female lines from Tetranychus populations collected from agricultural crops. We generated genomic and morphological data, characterized their bacterial communities and performed controlled crosses. Despite morphological similarities, we found large genomic differentiation between the morphs. This pattern was reflected in the incomplete, but strong postzygotic incompatibility in crosses between colour morphs, while crosses within morphs from different geographical locations were largely compatible. In addition, our results suggest recent/on-going gene flow between green-coloured T. urticae and T. turkestani. By screening the sequences of 10 resistance genes, we found evidence for multiple independent origins and for single evolutionary origins of target-site resistance mutations. Our results indicate that target-site mutations mostly evolve independently in populations on different geographical locations, and that these mutations can spread due to incomplete barriers to gene flow within and between populations.

Inhibition of ecdysone receptor (DcEcR) and ultraspiracle (DcUSP) expression in Diaphorina citri increased susceptibility to pesticides.

Diaphorina citri Kuwayama is of great concern because of its ability to transmit devastating citrus greening illness (Huanglongbing). One strategy for controlling HLB may involve limiting the spread of D. citri. Insecticides using dsRNA target genes may be a useful option to control D. citri. The ecdysone receptor (EcR) and ultraspiracle (USP) are crucial for the growth and reproduction of insects. This study identified the genes for D. citri ecdysone receptor (DcEcR) and ultraspiracle (DcUSP). According to the qPCR data, DcUSP peaked at the 5th-instar nymph stage, while DcEcR peaked at the adult stage. Females expressed DcEcR and DcUSP at much higher levels than males. RNAi was used to examine DcEcR and DcUSP function. The findings demonstrated that inhibition of DcEcR and DcUSP delayed nymph development and decreased survival and eclosion rates. dsEcR caused adults to develop deformed wings, and dsUSP caused nymphs to wither and die. Female adult ovaries developed slowly, and the females laid fewer eggs. Additionally, DcEcR and DcUSP were inhibited, increasing D. citri susceptibility to pesticides. These findings suggest that DcEcR and DcUSP are critical for D. citri development, growth, and reproduction and may serve as potential targets for D. citri management.

Inferring Adaptive Introgression Using Hidden Markov Models.

Adaptive introgression-the flow of adaptive genetic variation between species or populations-has attracted significant interest in recent years and it has been implicated in a number of cases of adaptation, from pesticide resistance and immunity, to local adaptation. Despite this, methods for identification of adaptive introgression from population genomic data are lacking. Here, we present Ancestry_HMM-S, a hidden Markov model-based method for identifying genes undergoing adaptive introgression and quantifying the strength of selection acting on them. Through extensive validation, we show that this method performs well on moderately sized data sets for realistic population and selection parameters. We apply Ancestry_HMM-S to a data set of an admixed Drosophila melanogaster population from South Africa and we identify 17 loci which show signatures of adaptive introgression, four of which have previously been shown to confer resistance to insecticides. Ancestry_HMM-S provides a powerful method for inferring adaptive introgression in data sets that are typically collected when studying admixed populations. This method will enable powerful insights into the genetic consequences of admixture across diverse populations. Ancestry_HMM-S can be downloaded from https://github.com/jesvedberg/Ancestry_HMM-S/.

Studies of an insecticidal inhibitor of acetyl-CoA carboxylase in the nematode C. elegans.

We have studied the mode of action of the insecticide spirotetramat in the nematode Caenorhabditis elegans. A combination of symptomology, forward genetics and genome editing show that spirotetramat acts on acetyl-CoA carboxylase (ACC) in C. elegans, as it does in insects. We found C. elegans embryos exposed to spirotetramat show a cell division defect which closely resembles the phenotype of loss-of-function mutations in the gene pod-2, which encodes ACC. We then identified two mutations in the carboxyl transferase domain of pod-2 (ACC) which confer resistance and were confirmed using CRISPR/Cas9. One of these mutations substitutes an invertebrate-specific amino acid with one ubiquitous in other taxa; this residue may, therefore, be a determinant of the selectivity of spirotetramat for invertebrates. Such a mutation may also be the target of selection for resistance in the field. Our study is a further demonstration of the utility of C. elegans in studying bioactive chemicals.

Acaricide resistance status and identification of resistance mutations in populations of the two-spotted spider mite Tetranychus urticae from Ethiopia.

The intensive use of pesticides is a common practice for the management of the two-spotted spider mite, Tetranychus urticae, in greenhouses and field farms of Ethiopia. However, incidence of resistance and possible resistance mechanisms in T. urticae populations from Ethiopia have not yet been studied. Here, we assessed the toxicity of various acaricides-bifenazate, abamectin, emamectin benzoate, profenofos, fenbutatin oxide, fenpyroximate, amitraz and chlorfenapyr-on T. urticae populations sampled from six flower greenhouse farms, three strawberry greenhouse farms, one field-grown vegetable farm and two wild populations. In parallel, all populations were screened for known target-site mutations. All tested populations were fully susceptible to bifenazate, abamectin, emamectin benzoate and profenofos, but resistant against fenbutatin oxide and fenpyroximate. Four populations showed considerable levels of resistance against amitraz and one population was resistant to chlorfenapyr. Several target-site mutations were identified in the tested populations, including G119S, A201S, T280A, G328A and F331W/C/Y in acetylcholinesterase and the F1538I and L1024V mutation in the voltage-gated sodium channel. The F1538I mutation was found in eight out of 12 populations, whereas the L1024V mutation was only found in two populations. The H92R mutation in the PSST subunit of complex I and the I1017F mutation in chitin synthase 1 was detected in half of the tested populations. The G326E and I321T mutations in the glutamate-gated chloride channel 3 were also detected, but more rarely, whereas mitochondrial cytochrome b mutations were not detected. The current study revealed multiple resistance patterns in Ethiopian T. urticae populations and together with the wide presence of target-site mutations, calls for the wise use of acaricides in the management of T. urticae in Ethiopia.

Evolutionary epidemiology predicts the emergence of glyphosate resistance in a major agricultural weed.

The evolution of resistance to herbicides is a striking example of rapid, human-directed adaptation with major consequences for food production. Most studies of herbicide resistance are performed reactively and focus on post hoc determination of resistance mechanisms following the evolution of field resistance. If the evolution of resistance can be anticipated, however, pro-active management to slow or prevent resistance traits evolving can be advocated. We report a national-scale study that combines population monitoring, glyphosate sensitivity assays, quantitative genetics and epidemiological analyses to pro-actively identify the prerequisites for adaptive evolution (directional selection and heritable genetic variation) to the world's most widely used herbicide (glyphosate) in a major, economically damaging weed species, Alopecurus myosuroides. Results highlighted pronounced, heritable variability in glyphosate sensitivity amongst UK A. myosuroides populations. We demonstrated a direct epidemiological link between historical glyphosate selection and current population-level sensitivity, and show that current field populations respond to further glyphosate selection. This study provides a novel, pro-active assessment of adaptive potential for herbicide resistance, and provides compelling evidence of directional selection for glyphosate insensitivity in advance of reports of field resistance. The epidemiological approach developed can provide a basis for further pro-active study of resistance evolution across pesticide resistance disciplines.

Possibilities for rationally exploiting co-evolution in addressing resistance to insecticides, and beyond.

Certain biorational chemical agents used against insect pests impact essential stages or processes in insect life cycles when applied for pest management. Development of resistance to these agents, while involving maintenance of the natural role of the chemical agent, frequently requires the evolution of a new chemical structure by the resistant organism. When considering the process of resistance development, one could theoretically consider biorational structural determination rather than the less predictable or feasible generation of a novel replacement insecticide. At first consideration, this process might exclude toxicants such as typical pest control agents and rather be a phenomenon reserved principally for signalling processes such as are fulfilled by pheromones and other semiochemicals. However, because there is a unique co-evolutionary relationship between chemical defence and the physiology of the antagonistic organism, this process can be further explored for potential to overcome resistance to toxins. Given further consideration, newly evolved chemical defences may rationally provide options for new resistance-defeating chemistry. This review therefore discusses the potential for overcoming insecticide resistance through targeted application of this approach. Potential for use of a similar approach to counteract fungicide and herbicide resistance is also considered. Furthermore, the possible applications of this approach to address drug or pharmaceutic resistance are also considered.

Quantitative trait locus mapping and functional genomics of an organophosphate resistance trait in the western corn rootworm, Diabrotica virgifera virgifera.

The western corn rootworm, Diabrotica virgifera virgifera, is an insect pest of corn and population suppression with chemical insecticides is an important management tool. Traits conferring organophosphate insecticide resistance have increased in frequency amongst D. v. virgifera populations, resulting in the reduced efficacy in many corn-growing regions of the USA. We used comparative functional genomic and quantitative trait locus (QTL) mapping approaches to investigate the genetic basis of D. v. virgifera resistance to the organophosphate methyl-parathion. RNA from adult methyl-parathion resistant and susceptible adults was hybridized to 8331 microarray probes. The results predicted that 11 transcripts were significantly up-regulated in resistant phenotypes, with the most significant (fold increases ≥ 2.43) being an α-esterase-like transcript. Differential expression was validated only for the α-esterase (ST020027A20C03), with 11- to 13-fold greater expression in methyl-parathion resistant adults (P < 0.05). Progeny with a segregating methyl-parathion resistance trait were obtained from a reciprocal backcross design. QTL analyses of high-throughput single nucleotide polymorphism genotype data predicted involvement of a single genome interval. These data suggest that a specific carboyxesterase may function in field-evolved corn rootworm resistance to organophosphates, even though direct linkage between the QTL and this locus could not be established.

IDENTIFICATION OF PROTEASOME ALPHA6 SUBUNIT ASSOCIATED WITH DELTAMETHRIN RESISTANCE IN Drosophila melanogaster Kc CELLS.

Differential expression of the proteasome alpha6 (prosalpha6) was previously reported between Plutella xylostella strains that are resistant or susceptible to the pesticide deltamethrin (DM). This finding indicated that the prosalpha6 may be involved in DM resistance. In this article, qPCR analysis revealed that the prosalpha6 was also significantly upregulated in Drosophila Kc cells treated with DM. To better understand the contribution of prosalpha6 in DM resistance, RNA interference, heterologous expression, and a proteasome inhibitor (MG-132) were used. MG-132 was used to suppress proteasomal activity, and the dsRNA was designed to block the function of prosalpha6. The results indicated that both MG-132 and prosalpha6 knockdown decreased the cellular viability following DM treatment. Prosalpha6 was cloned and transfected into Drosophila Kc cells. The result showed that overexpression of prosalpha6 in Drosophila Kc cells conferred some protection against DM. Taken together, our results indicate that prosalpha6 is involved in Drosophila cells DM resistance.

Differential Gene Expression Analysis of the Epacromius coerulipes (Orthoptera: Acrididae) Transcriptome.

Epacromius coerulipes (Ivanov) is one of the most widely distributed locusts. To date, the main methods to kill locusts still rely on chemical controls, which can result in the selection of locusts with resistance to chemical pesticides. Butene-fipronil is a new pesticide that was discovered by the structural modification of fipronil. This pesticide has been used to control various agricultural pests and has become an important pesticide product to control pests that exhibit resistance to other pesticides, including locusts. To extend its useful half-life, studies of the initiation and progression of resistance to this pesticide are needed. Herein, two E. coerulipes strains, a pesticide-sensitive (PS) and a pesticide-resistant (PR) strain, were chosen to undergo de novo assembly by paired-end transcriptome Illumina sequencing. Overall, 63,033 unigenes were detected; the average gene length was 772 bp and the N50 was 1,589 bp. Among these unigenes, ∼ 25,132 (39.87% of the total) could be identified as known proteins in bioinformatic databases from national centers. A comparison of the PR and PS strains revealed that 2,568 genes were differentially expressed, including 1,646 and 922 genes that were up- and down-regulated, respectively. According to the Gene Ontology (GO) database, among biological processes the metabolic process group was the largest group (6,900 genes, 22.47%) and contained a high frequency of differentially expressed genes (544 genes, 27.54%). According to the Clusters of Orthologous Groups (COG) categories, 28 genes, representing 2.98% of all genes, belonged to the group of genes involved in the biosynthesis, transportation, and catabolism of secondary metabolites. The differentially expressed genes that we identified are involved in 50 metabolic pathways. Among these pathways, the metabolism pathway was the most represented. After enrichment analysis of differential gene expression pathways, six pathways--ribosome; starch, and sucrose metabolism; ascorbate and aldarate metabolism; drug metabolism-cytochrome P450; metabolism of xenobiotics by cytochrome P450; and glutathione metabolism--showed a high degree of enrichment. Among these pathways, drug metabolism-cytochrome P450, metabolism of xenobiotics by cytochrome P450, and glutathione metabolism have been associated with pesticide metabolism. Furthermore, 316 unigenes in the E. coerulipes transcriptome encode detoxifying enzymes and 76 unigenes encode target proteins of pesticides. Among these genes, 23 genes that encode detoxifying enzymes in the resistance group were found to be up-regulated. The transcriptome sequencing results of E. coerulipes established a genomics database of E. coerulipes for the first time. This study also establishes a molecular basis for gene function analysis of E. coerulipes Moreover, it provides a theoretical resource for mechanistic studies on pesticide resistance through the screening and investigation of resistance genes.

Pyrethroid resistance in Phytoseiulus macropilis (Acari: Phytoseiidae): cross-resistance, stability and effect of synergists.

Phytoseiulus macropilis Banks (Acari: Phytoseiidae) is an effective predator of Tetranychus urticae Koch (Acari: Tetranychidae). The objectives of this research were to study the stability of fenpropathrin resistance and the cross-resistance relationships with different pyrethroids, and also to evaluate the effect of synergists [piperonyl butoxide (PBO), diethyl maleate (DEM) and S,S,S-tributyl phosphorotrithioate (DEF)] on fenpropathrin resistant and susceptible strains of this predaceous mite. The stability of fenpropathrin resistance was studied under laboratory conditions, using P. macropilis populations with initial frequencies of 75 and 50% of resistant mites. The percentages of fenpropathrin resistant mites were evaluated monthly for a period of up to 12 months. A trend toward decreased resistance frequencies was observed only during the first 3-4 months. After this initial decrease, the fenpropathrin resistance was shown to be stable, maintaining constant resistance frequencies (around 30%) until the end of the evaluation period. Toxicity tests carried out using fenpropathrin resistant and susceptible strains of P. macropilis indicated strong positive cross-resistance between fenpropathrin and the pyrethroids bifenthrin and deltamethrin. Bioassays with the synergists DEM, DEF and PBO were also performed. The maximum synergism ratio (SR = LC50 without synergist/LC50 with synergist) detected for the three evaluated synergists (PBO, DEM, DEF) was 5.86 (for DEF), indicating low influence of enzyme detoxification processes in fenpropathrin resistance.

Biotic interactions govern genetic adaptation to toxicants.

The genetic recovery of resistant populations released from pesticide exposure is accelerated by the presence of environmental stressors. By contrast, the relevance of environmental stressors for the spread of resistance during pesticide exposure has not been studied. Moreover, the consequences of interactions between different stressors have not been considered. Here we show that stress through intraspecific competition accelerates microevolution, because it enhances fitness differences between adapted and non-adapted individuals. By contrast, stress through interspecific competition or predation reduces intraspecific competition and thereby delays microevolution. This was demonstrated in mosquito populations (Culex quinquefasciatus) that were exposed to the pesticide chlorpyrifos. Non-selective predation through harvesting and interspecific competition with Daphnia magna delayed the selection for individuals carrying the ace-1(R) resistance allele. Under non-toxic conditions, susceptible individuals without ace-1(R) prevailed. Likewise, predation delayed the reverse adaptation of the populations to a non-toxic environment, while the effect of interspecific competition was not significant. Applying a simulation model, we further identified how microevolution is generally determined by the type and degree of competition and predation. We infer that interactions with other species-especially strong in ecosystems with high biodiversity-can delay the development of pesticide resistance.

Detoxification of phoxim by a gut bacterium of Delia antiqua.

The presence of certain associated bacteria has been reported to increase pest resistance to pesticides, which poses a serious threat to food security and the environment. Researches on the above microbe-derived pesticide resistance would bring innovative approaches for pest management. Investigations into the phoxim resistance of Delia antiqua, one Liliaceae crop pests, revealed the contribution of a phoxim-degrading gut bacterium, D39, to this resistance. However, how the strain degraded phoxim was unknown. In this study, the role of D39 in phoxim degradation and resistance was first confirmed. DT, which had an identical taxonomy but lacked phoxim-degrading activity, was analyzed alongside D39 via comparative genomics to identify the potential phoxim degrading genes. In addition, degradation metabolites were identified, and a potential degradation pathway was proposed. Furthermore, the main gene responsible for degradation and the metabolites of phoxim were further validated via prokaryotic expression. The results showed that D39 contributed to resistance in D. antiqua larva by degrading phoxim. Phoxim was degraded by an enzyme encoded by the novel gene phoD in D39 to O,O-diethyl hydrogen phosphorothioate and 2-hydroxyimino-2-phenylacetonitrile. Finally, downstream products were metabolized in the tricarboxylic acid cycle. Further analysis via prokaryotic expression of phoD confirmed its degradation activity. The mechanisms through which gut microbes promote pesticide resistance are elucidated in this study. These results could aid in the development of innovative pest control methods. In addition, this information could also be used to identify microbial agents that could be applied for the remediation of pesticide contamination.

Relationship between acaricide resistance and acetylcholinesterase gene polymorphisms in the cattle tick Rhipicephalus microplus.

In this study, we aimed to develop a comprehensive methodology for identifying amino acid polymorphisms in acetylcholinesterase transcript 2 (AChE2) in acaricide-resistant Rhipicephalus microplus ticks. This included assessing AChE2 expression levels through qPCR and conducting 3D modeling to evaluate the interaction between acaricides and AChE2 using docking techniques. The study produced significant results, demonstrating that acaricide-resistant R. microplus ticks exhibit significantly higher levels of AChE expression than susceptible reference ticks. In terms of amino acid sequence, we identified 9 radical amino acid substitutions in AChE2 from acaricide-resistant ticks, when compared to the gene sequence of the susceptible reference strain. To further understand the implications of these substitutions, we utilized 3D acaricide-AChE2 docking modeling to examine the interaction between the acaricide and the AChE2 catalytic site. Our models suggest that these amino acid polymorphisms alter the configuration of the binding pocket, thereby contributing to differences in acaricide interactions and ultimately providing insights into the acaricide-resistance phenomenon in R. microplus.

Title: Relations entre la résistance aux acaricides et les polymorphismes du gène de l'acétylcholinestérase chez la tique du bétail Rhipicephalus microplus. Abstract: Notre étude vise à développer une méthodologie complète pour identifier les polymorphismes d'acides aminés dans le transcrit 2 de l'acétylcholinestérase (AChE2) chez les tiques Rhipicephalus microplus résistantes aux acaricides. Cela comprend l'évaluation des niveaux d'expression d'AChE2 via qPCR et la réalisation d'une modélisation 3D pour évaluer l'interaction entre les acaricides et l'AChE2 à l'aide de techniques d'amarrage moléculaire. L'étude a produit des résultats significatifs, démontrant que les tiques R. microplus résistantes aux acaricides présentent des niveaux d'expression d'AChE significativement plus élevés que les tiques sensibles de référence. En termes de séquence d'acides aminés, nous avons identifié 9 substitutions d'acides aminés dans AChE2 provenant de tiques résistantes aux acaricides par rapport à la séquence génétique de la souche sensible de référence. Pour mieux comprendre les implications de ces substitutions, nous avons utilisé la modélisation de l'amarrage acaricide-AChE2 pour examiner l'interaction entre l'acaricide et le site catalytique AChE2. Nos modèles suggèrent que ces polymorphismes d'acides aminés modifient la configuration de la poche de liaison, contribuant ainsi aux différences dans les interactions acaricides et fournissant finalement un aperçu du phénomène de résistance aux acaricides chez R. microplus.