Detoxification Genes Research Articles

Transcriptomic analysis reveals global and temporal transcription changes during Candida glabrata adaptation to an oxidative environment

The ability to survive host-elicited oxidative stress is critical for microbial pathogens to cause infection. The human fungal pathogen C.glabrata can tolerate high levels of oxidative stress and proliferate inside phagocytes. Previous studies had successfully identified a transcription response to oxidative stress including induction of a core set of detoxification genes. However, the findings only represent an early snapshot of a highly dynamic process lacking temporal resolution. Here, we compare the transcriptome of C. glabrata at various points after exposure to hydrogen peroxide in order to study its adaptation to an oxidative environment. Our results reveal global and temporal gene expression changes during an immediate response; up-regulating genes related to peroxide detoxification, while down-regulating genes essential for growth. As cells adapt to the oxidative environment, a dramatic transcriptome reprogramming occurred to restore key cellular functions, protein homeostasis and biosynthesis of trehalose, carbohydrate, fatty acid and ergosterol. Interestingly, biofilm and drug transporter genes as well as many genes implicated in virulence, were induced during the adaptation stage. Our finding, therefore, suggests a role of oxidative stress adaptation in promoting virulence and drug resistance traits of C. glabrata during infection.

Detoxification and reproductive system-related gene expression following exposure to Cu(OH)2 nanopesticide in water flea (Daphnia magna Straus 1820).

The extensive use of copper-based nanopesticides in agriculture has led to their release into the aquatic environment and causes a potential risk to aquatic biota. However, there is a lack of knowledge regarding the possible toxic effect of these nanopesticides on non-target aquatic organisms including invertebrates. Therefore, in this study, effects of commonly used copper-based nanopesticide "Kocide 3000" on gene expression related to detoxification (cyp360a8, gst, P-gp, and hr96) and reproductive system (cut, cyp314, dmrt93, and vtg) in Daphnia magna was investigated through an acute toxicity test. In general, exposure to the nanopesticide caused significant down-regulation of detoxification genes after 24h and then significant up-regulation after 48h. Exposure to the nanopesticide, however, significantly induced cut expression after 24h. Moreover, dmrt93 and vtg genes were up-regulated after 48h exposure to the nanopesticide. On the other hand, the expression of dmrt93 and vtg down-regulated at high concentration of Cu(OH)2 nanopesticide (1.5ppm) after 96h. The results of this study provide first evidence into the crucial role of genes related to detoxification and reproductive system in response to Cu(OH)2 nanopesticide. The use of physiological, biochemical bioassays, as well as gene expression, can help explain the toxic effect of copper-based nanopesticides and provide more insight into the exact mechanism of toxicity in non-target aquatic organisms.

Mediator subunit MDT-15/MED15 and Nuclear Receptor HIZR-1/HNF4 cooperate to regulate toxic metal stress responses in Caenorhabditis elegans.

Zinc is essential for cellular functions as it is a catalytic and structural component of many proteins. In contrast, cadmium is not required in biological systems and is toxic. Zinc and cadmium levels are closely monitored and regulated as their excess causes cell stress. To maintain homeostasis, organisms induce metal detoxification gene programs through stress responsive transcriptional regulatory complexes. In Caenorhabditis elegans, the MDT-15 subunit of the evolutionarily conserved Mediator transcriptional coregulator is required to induce genes upon exposure to excess zinc and cadmium. However, the regulatory partners of MDT-15 in this response, its role in cellular and physiological stress adaptation, and the putative role for mammalian MED15 in the metal stress responses remain unknown. Here, we show that MDT-15 interacts physically and functionally with the Nuclear Hormone Receptor HIZR-1 to promote molecular, cellular, and organismal adaptation to cadmium and excess zinc. Using gain- and loss-of-function mutants and qRT-PCR and reporter analysis, we find that mdt-15 and hizr-1 cooperate to induce zinc and cadmium responsive genes. Moreover, the two proteins interact physically in yeast-two-hybrid assays and this interaction is enhanced by the addition of zinc or cadmium, the former a known ligand of HIZR-1. Functionally, mdt-15 and hizr-1 mutants show defective storage of excess zinc in the gut and are hypersensitive to zinc-induced reductions in egg-laying. Furthermore, mdt-15 but not hizr-1 mutants are hypersensitive to cadmium-induced reductions in egg-laying, suggesting potential divergence of regulatory pathways. Lastly, mammalian MDT-15 orthologs bind genomic regulatory regions of metallothionein and zinc transporter genes in a cadmium and zinc-stimulated fashion, and human MED15 is required to induce a metallothionein gene in lung adenocarcinoma cells exposed to cadmium. Collectively, our data show that mdt-15 and hizr-1 cooperate to regulate cadmium detoxification and zinc storage and that this mechanism is at least partially conserved in mammals.

Responses of Detoxification Enzymes and Genes to Nicosulfuron Stress in Two Genotypes of Sweet Corn Differing in Nicosulfuron Tolerance

Nicosulfuron, a sulfonylurea herbicide is registered for use on field, controns many annual and perennial grass weeds. The accumulation of nicosulfuron will increase toxicity and induce unfavorable oxidative stress in plants. Acetolactate synthase (ALS, EC 2.2.1.6) and glutathione transferases (GSTs, E. C.2.5.1.18), which participate in detoxification of xenobiotics and limit oxidative damages of cellular macromolecules, are important groups of cytoprotective enzymes. This study aims to investigate the toxic effect of nicosulfuron on the detoxification enzyme activities and gene expressions in maize seedling. Specifically, Nicosulfuron-tolerant sweet inbred line ‘JP233’ and nicosulfuron-sensitive sweet inbred line ‘Z2H4’ are subjected to 80mg kg–1 nicosulfuron treatment when the fourth leaves are fully developed, and the resulting effects are compared to those processed by water. After nicosulfuron treatment, it is found that ALS and GST enzymes activities of Z2H4 are significantly lower than those of JP233. Compared to Z2H4, nicosulfuron treatment increases the expression levels of GST1, ALS1 and ALS2 genes in JP233. These results suggest that the increased transcription level of these detoxifying enzymes might play a vital role in reducing the toxicity of nicosulfuron and the oxidative stress induced by nicosulfuron in maize seedlings. The research will improve our understanding of the function of maize detoxification enzymes and genes in herbicide metabolism.

The Genome of the Blind Soil-Dwelling and Ancestrally Wingless Dipluran Campodea augens: A Key Reference Hexapod for Studying the Emergence of Insect Innovations.

The dipluran two-pronged bristletail Campodea augens is a blind ancestrally wingless hexapod with the remarkable capacity to regenerate lost body appendages such as its long antennae. As sister group to Insecta (sensu stricto), Diplura are key to understanding the early evolution of hexapods and the origin and evolution of insects. Here we report the 1.2-Gb draft genome of C. augens and results from comparative genomic analyses with other arthropods. In C. augens, we uncovered the largest chemosensory gene repertoire of ionotropic receptors in the animal kingdom, a massive expansion that might compensate for the loss of vision. We found a paucity of photoreceptor genes mirroring at the genomic level the secondary loss of an ancestral external photoreceptor organ. Expansions of detoxification and carbohydrate metabolism gene families might reflect adaptations for foraging behavior, and duplicated apoptotic genes might underlie its high regenerative potential. The C. augens genome represents one of the key references for studying the emergence of genomic innovations in insects, the most diverse animal group, and opens up novel opportunities to study the under-explored biology of diplurans.

Consequences of a short time exposure to a sublethal dose of Flupyradifurone (Sivanto) pesticide early in life on survival and immunity in the honeybee (Apis mellifera)

Dramatic losses of pollinating insects have become of global concern, as they threaten not only key ecosystem services but also human food production. Recent research provided evidence that interactions between ecological stressors are drivers of declining pollinator health and responsible for observed population collapses. We used the honeybee Apis mellifera and conducted a series of experiments to test for long-term effects of a single short exposure to the agricultural pesticide flupyradifurone to a second environmental stressor later in life. To do this, we exposed individuals during their larval development or early adulthood to sublethal dosages of flupyradifurone (0.025 μg for larvae and 0.645 μg for imagos), either pure or as part of an agricultural formulation (Sivanto). We afterwards exposed bees to a second ecological stressor infecting individuals with 10,000 spores of the fungal gut parasite Nosema ceranae. We found that pesticide exposures significantly reduced survival of bees and altered the expression of several immune and detoxification genes. The ability of bees to respond to these latter effects differed significantly between colonies, offering opportunities to breed bees with elevated levels of pesticide tolerance in the future. We conclude that short episodes of sublethal pesticide exposures during development are sufficient to trigger effects later in life and could therefore contribute to the widespread declines in bee health.

Dose Dependence of Hypermethylation of Gene Promoters in Blood Leukocytes in Humans Occupationally Exposed to External γ-Radiation

Abstract—A study of promoter hypermethylation of cell cycle (RASSF1A, p16/INK4A, p14/ARF, p53, АТМ), xenobiotic detoxification (GSTP1), antioxidant defense (SOD3), and estrogen receptor (ESR1) genes in blood leukocytes of 88 nuclear industry workers from the Mayak facility (53–86 years old at sampling) exposed to prolonged external γ-radiation was carried out. Accumulated doses to red bone marrow (RBM) ranged from 0.2 to 2.95 Gy. The time between the end of work with radiation and blood sampling was on average 15.95 ± 1.22 years. Fifty unirradiated residents of Ozyorsk matched by age to the exposed group were enrolled as a control group. Promoter hypermethylation was analyzed using methylation-sensitive polymerase chain reaction (PCR) assay. The trend towards dependence of promoter hypermethylation of p14/ARF and RASSFA genes on age rather than radiation exposure (logistic regression: p = 0.072) confirms previous results. The frequency of individuals with promoter methylation of at least one of the following six genes: p16/INK4A, GSTP1, р53, АТМ,SOD3,ESR1, in irradiated group was significantly higher than in the control group (62.5 versus 34%, respectively, p = 0.001, OR = 3.24, 95% CI 1.56–6.69). A significantly elevated frequency of individuals with hypermethylated СpG islands in GSTP1 and SOD3 promoters was revealed among exposed subjects compared to the control group (p = 0.014 and p = 0.021, respectively). A statistically significant association between the cumulative dose and the number of methylated promoters was revealed (r = 0.304, р = 3.2 × 10–4). Therefore, the dose-dependent hypermethylation of CpG islands in gene promoters that is revealed in blood leukocytes long term after external exposure to γ-radiation was found.

Effects of a Four-Week High-Dosage Zinc Oxide Supplemented Diet on Commensal Escherichia coli of Weaned Pigs.

Strategies to reduce economic losses associated with post-weaning diarrhea in pig farming include high-level dietary zinc oxide supplementation. However, excessive usage of zinc oxide in the pig production sector was found to be associated with accumulation of multidrug resistant bacteria in these animals, presenting an environmental burden through contaminated manure. Here we report on zinc tolerance among a random selection of intestinal Escherichia coli comprising of different antibiotic resistance phenotypes and sampling sites isolated during a controlled feeding trial from 16 weaned piglets: In total, 179 isolates from “pigs fed with high zinc concentrations” (high zinc group, [HZG]: n = 99) and a corresponding “control group” ([CG]: n = 80) were investigated with regard to zinc tolerance, antimicrobial- and biocide susceptibilities by determining minimum inhibitory concentrations (MICs). In addition, in silico whole genome screening (WGSc) for antibiotic resistance genes (ARGs) as well as biocide- and heavy metal tolerance genes was performed using an in-house BLAST-based pipeline. Overall, porcine E. coli isolates showed three different ZnCl2 MICs: 128 μg/ml (HZG, 2%; CG, 6%), 256 μg/ml (HZG, 64%; CG, 91%) and 512 μg/ml ZnCl2 (HZG, 34%, CG, 3%), a unimodal distribution most likely reflecting natural differences in zinc tolerance associated with different genetic lineages. However, a selective impact of the zinc-rich supplemented diet seems to be reasonable, since the linear mixed regression model revealed a statistically significant association between “higher” ZnCl2 MICs and isolates representing the HZG as well as “lower ZnCl2 MICs” with isolates of the CG (p = 0.005). None of the zinc chloride MICs was associated with a particular antibiotic-, heavy metal- or biocide- tolerance/resistance phenotype. Isolates expressing the 512 μg/ml MIC were either positive for ARGs conferring resistance to aminoglycosides, tetracycline and sulfamethoxazole-trimethoprim, or harbored no ARGs at all. Moreover, WGSc revealed a ubiquitous presence of zinc homeostasis and – detoxification genes, including zitB, zntA, and pit. In conclusion, we provide evidence that zinc-rich supplementation of pig feed selects for more zinc tolerant E. coli, including isolates harboring ARGs and biocide- and heavy metal tolerance genes – a putative selective advantage considering substances and antibiotics currently used in industrial pork production systems.

Comparative transcriptomic analyses revealed genes and pathways responsive to heat stress in Diaphorina citri

The Asian citrus psyllid Diaphorina citri Kuwayama (Hemiptera: Liviidae) is a serious pest that feeds on plant phloem sap of citrus trees, and transmits Candidatus Liberibacter asiaticus, a bacterium that induces the destructive disease called Huanglongbing. Increasing evidence suggests that high temperatures could affect various biological traits, including size, longevity, mortality, behavior and metabolism of D. citri. However, the relevant mechanisms of heat stress remain unclear. In this study, a large set of transcriptomic data derived from D. citri adults were generated and differentially expressed genes (DEGs) after heat stress were identified by RNA sequencing. A total of 118, 399 unigenes were obtained, from which 37, 665 were mapped to sequences from at least one database. Seven hundreds and twenty-two unigenes were affected by high temperature of 40 °C for 4 h, in which 486 up-regulated and 236 down-regulated, and part of heat shock proteins, antioxidant and detoxification genes and cathepsins were identified as the DEGs. KEGG pathway enrichment analysis demonstrated that part of genes involved in protein synthesis and processing, metabolism, immunity, and signal transduction were differentially expressed under heat stress. Furthermore, the mRNA expression levels of 20 DEGs were confirmed by qRT-PCR, which verified the accuracy of high-throughput sequencing. Our results revealed that the response of D. citri adults to high temperatures is associated with a range of changes involved in various physiological and biochemical processes. Our data provide a basis for future research to improve our understanding on the molecular mechanism for heat responses in D. citri adults.

Differences in susceptibility to insecticides among color morphs of the Asian citrus psyllid

Insect color morphs are known to vary in detoxification enzyme activity associated with susceptibility to insecticides. Diaphorina citri occurs in three distinct color morphs, and little data are available addressing this question. In this study, susceptibility to nine insecticides was compared among orange/yellow, gray/brown and blue/green morphs of adult D. citri. Imidacloprid and lambda-cyhalothrin resulted in higher mortality in the orange/yellow morph than in the blue/green morph. The orange/yellow morph was significantly more susceptible to acetamiprid, bifenthrin and chlorfenapyr than the blue/green and gray/brown morphs. For clothianidin and chlorpyriphos, the orange/yellow morph was significantly more susceptible than the gray/brown morphs. Susceptibility to dinotefuran and thiamethoxam was not significantly different between the three color morphs of D. citri. Biochemical enzyme assays revealed that the esterase, glutathione S-transferase and cytochrome P450 activities were significantly higher in the gray/brown and blue/green morphs than in the orange/yellow morph; however, the activities of three detoxification enzymes were not significantly different between gray/brown and blue/green morphs. We analyzed the relative expression among the color morphs of six detoxification genes involved in insecticide resistance. GSTS1 and EST-6 were expressed at significantly higher levels in the blue/green morph than in the orange/yellow and gray/brown morphs. The expression levels of CYP4g15, CYP303A1, CYP4C62 and CYP6BD5 were significantly greater in the gray/brown and blue/green morphs than in the orange/yellow morph. These results suggest that insecticide susceptibility in D. citri is associated with color morphs. Detoxification enzyme activity and the relative expression of six detoxification genes may partially explain these differences. These results may facilitate further work to develop a management strategy for control of D. citri.

An influential meal: host plant dependent transcriptional variation in the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae)

BackgroundTo understand the genetic mechanisms of insect herbivory, the transcriptional response of insects feeding on different host plant species has to be studied. Here, we generated gene expression data of the generalist herbivore Spodoptera exigua (Hübner) feeding on three selected host plant species and a control (artificial diet). The host plant species used in this study –cabbage (Brassica oleracea), maize (Zea mays) and tobacco (Nicotiana tabacum)- are members of different plant families that each employ specific defence mechanisms and toxins.ResultsSpodoptera exigua larvae had a higher growth rate, indicator for herbivore success, when feeding on Z. mays compared to larvae feeding on B. oleracea or N. tabacum. Larvae feeding on the different host plant species showed divergent transcriptional responses. We identified shared and unique gene expression patterns dependent of the host plant species the larvae fed on. Unique gene expression patterns, containing uniquely upregulated transcripts including specific detoxification genes, were found for larvae feeding on either B. oleracea or N. tabacum. No diet-specific gene cluster was identified for larvae feeding on the host for which larvae showed optimal herbivore success, Z. mays, or artificial diet. In contrast, for larvae feeding on hosts for which they showed low herbivore success, specific diet-dependent gene clusters were identified. Functional annotation of these clusters indicates that S. exigua larvae deploy particular host plant-specific genes for digestion and detoxification.ConclusionsThe lack of a host plant-specific gene activity for larvae feeding on Z. mays and the artificial diet suggest a general and non-specific gene activity for host plants with optimal herbivore success. Whereas the finding of specific gene clusters containing particular digestion and detoxifying genes expressed in larvae feeding on B. oleracea and N. tabacum, with low herbivore success, imply a host plant-specific gene activity for larvae feeding on host plants with suboptimal herbivore success. This observation leads to the conclusion that a polyphagous herbivore is able to feed on a large variation of host plants due to the flexibility and diversity of genes involved in digestion and detoxification that are deployed in response to particular host plant species.

Multiple detoxification genes confer imidacloprid resistance to Sitobion avenae Fabricius

The English grain aphid, Sitobion avenae Fabricius, is one of the major wheat pests distributed worldwide. Insecticide application is typically effective for controlling S. avenae, However, S. avenae has developed resistance to multiple insecticides. Thus, it is crucial to elucidate resistance mechanisms in S. avenae. In this study, the mechanisms of resistance to imidacloprid in S. avenae and the risk of cross resistance to other insecticides were investigated. An imidacloprid resistant strain (SA-R) of S. avenae was established via continuous imidacloprid selection with 42.7-fold resistance compared with the susceptible population (SA-S). There was cross resistance in SA-R between imidacloprid and other insecticides tested (acetamiprid, abamectin, chlorpyrifos, and omethoate), with ratios ranging from 10.7- to 24.7-fold higher than SA-S. S, S, S-tributyl phosphorotrithioate (TPP), piperonyl butoxide (PBO), and diethyl maleate (DEM) had synergism with imidacloprid in the SA-R strain with 2.85-, 3.59-, and 2.14-fold reductions, respectively, in LC50 values. Additionally, the activities of cytochrome P450 (P450), glutathione-S-transferases (GST), and carboxylesterase (COE) were significantly higher by 2.69-, 1.42-, and 1.39-fold in the SA-R strain than in the SA-S strain. Based on the S. avenae transcriptome database, 11 P450, two COE, four GST, and two esterase (EST) genes that were significantly induced by imidacloprid were selected to compare gene expression levels between SA-R and SA-S. Fourteen detoxification genes of the 19 tested from the SA-R strain were overexpressed compared to those of the SA-S strain. Silencing of CYP6A14-1, CYP307A1, GST1-1-1, and COE2 by dsRNA feeding resulted in increased susceptibility of SA-R aphids to imidacloprid. These results provide evidence that detoxification genes including CYP6A14-1, CYP307A1, GST1-1-1, and COE2 contribute to resistance to imidacloprid in S. avenae, which was confirmed by RNA interference (RNAi) feeding.

Exposure of honey bee larvae to thiamethoxam and its interaction with Nosema ceranae infection in adult honey bees

During their lifetime honey bees (Apis mellifera) rarely experience optimal conditions. Sometimes, a simultaneous action of multiple stressors, natural and chemical, results in even greater effect than of any stressor alone. Therefore, integrative investigations of different factors affecting honey bees have to be carried out. In this study, adult honey bees exposed to thiamethoxam in larval and/or adult stage and infected with Nosema ceranae were examined. Newly emerged bees from colonies, non-treated or treated with thiamethoxam, were organized in six groups and kept in cages. Thiamethoxam treated bees were further exposed to either thiamethoxam or Nosema (groups TT and TN), or simultaneously to both (group TTN). Newly emerged bees from non-treated colonies were exposed to Nosema (group CN). From both, treated and non-treated colonies two groups were organized and further fed only with sugar solution (groups C and TC). Here, we present the expression profile of 19 genes in adult worker honey bees comprising those involved in immune, detoxification, development and apoptosis response. Results showed that gene expression patterns changed with time and depended on the treatment. In group TC at the time of emergence the majority of tested genes were downregulated, among which nine were significantly altered. The same gene pattern was observed on day six, where the only significantly upregulated gene was defensin-1. On day nine most of analyzed genes in all experimental groups showed upregulation compared to control group, where upregulation of antimicrobial peptide genes abaecin, defensin-1 and defensin-2 was significant in groups TT and TTN. On day 15 we observed a similar pattern of expression in groups TC and TT exposed to thiamethoxam only, where most of the detoxification genes were downregulated. Additionally RNA loads of Nosema and honey bee viruses were recorded. We detected a synergistic interaction of thiamethoxam and Nosema, reflected in lowest honey bee survival.

Identification of epigenetic mechanisms in paddy crop associated with lowering environmentally related cadmium risks to food safety

Cadmium (Cd) is a toxic metal that contributes to human diseases such as pediatric cancer and cardiovascular dysfunction. Epigenetic modification caused by Cd exposure is the major factor in etiology of environmentally-relevant diseases. However, the underlying epigenetic mechanism for Cd uptake and accumulation in food crops, particularly those growing in Cd-contaminated environments, is largely unknown. This study investigated uncharacterized regulatory mechanisms and biological functions of global DNA hypomethylation at CG sites that are associated with gene expression for Cd detoxification and accumulation in the food crop rice. Mutation of the CG maintenance enzyme OsMET1 confers rice tolerance to Cd exposure. Genome-wide analysis of OsMET1 loss of function mutant Osmet1 and its wild type shows numerous loci differentially methylated and upregulated genes for Cd detoxification, transport and accumulation. We functionally identified a new locus for a putative cadmium tolerance factor (here termed as OsCTF) and demonstrated that Cd-induced DNA demethylation is the drive of OsCTF expression. The 3′-UTR of OsCTF is the primary site of DNA and histone (H3K9me2) demethylation, which is associated with higher levels of OsCTF transcripts detected in the Osmet1 and Ossdg714 mutant lines. Mutation of OsCTF in rice led to hypersensitivity to Cd and the Osctf line accumulated more Cd, whereas transfer of OsCTF back to the Osctf mutant completely restored the normal phenotype. Our work unveiled an important epigenetic mechanism and will help develop breeding crops that contribute to food security and better human health.

MicroRNAs as Potential Biomarkers of Insecticide Exposure: A Review.

Insecticides are key weapons for the control of pests. Large scale use of insecticides is harmful to the ecosystem, which is made up of a wide range of species and environments. MicroRNAs (miRNAs) are a class of endogenous single-stranded noncoding small RNAs in length of 20-24 nucleotides (nt), which extensively regulate expression of genes at transcriptional and post-transcriptional levels. The current research on miRNA-induced insecticide resistance reveals that dysregulated miRNAs cause significant changes in detoxification genes, particularly cytochrome P450s. Meanwhile, insecticide-induced changes in miRNAs are related to the decline of honeybees and threatened the development of zebrafish and other animals. Additionally, miRNAs are involved in insecticide-induced cytotoxicity, and dysregulated miRNAs are associated with human occupational and environmental exposure to insecticides. Therefore, miRNAs are valuable novel biomarkers of insecticide exposure, and they are potential factors to explain the toxicological effects of insecticides.

Aryl hydrocarbon receptor regulates the expression of LmGSTd7 and is associated with chlorpyrifos susceptibility in Locusta migratoria

The aryl hydrocarbon receptor (AhR) belongs to the bHLH-PAS (basic Helix-Loop-Helix-Period/ARNT/Single minded) family of transcription factors. AhR is a ligand-activated transcription factor, which participates in the sensing and transmitting stimuli of endogenous and exogenous chemicals, and subsequently activates the transcription of genes related to various physiological and detoxification functions. In this study, a single full-length LmAhR sequence was cloned and characterized. RNA interference (RNAi) and insecticide bioassays showed that LmAhR plays a vital role in chlorpyrifos susceptibility. To better identify aryl hydrocarbon receptor from locusta migratoria (LmAhR)-regulated genes involved in chlorpyrifos susceptibility, a comparative transcriptome analysis was performed using double-stranded (ds)GFP- and dsLmAhR-injected Locusta migratoria. Differential gene expression analysis identified 145 down-regulated and 67 up-regulated genes (P ≤ 0.05 and fold change ≥2) in dsLmAhR-knockdown insects. We selected 27 down-regulated genes and verified their expression levels using reverse transcription quantitative PCR. Finally, one glutathione S-transferase (GST) gene (LmGSTd7) was selected as a candidate detoxification gene and was further validated via RNAi and chlorpyrifos bioassays. Our data suggest that AhR is associated with chlorpyrifos susceptibility via the regulation of LmGSTd7 expression in L. migratoria. © 2019 Society of Chemical Industry.

Status of Insecticide Resistance and Its Mechanisms in Anopheles gambiae and Anopheles coluzzii Populations from Forest Settings in South Cameroon

A key factor affecting malaria vector control efforts in Cameroon is the rapid expansion of insecticide resistance in Anopheles gambiae s.l (An. gambiae) populations; however, mechanisms involved in insecticide resistance in forest mosquito populations are still not well documented yet. The present study was conducted to screen molecular mechanisms conferring insecticide resistance in An. gambiae s.l. populations from the South Cameroon forest region. WHO bioassays were conducted with F0 An. gambiae females aged three to four days from forest (Sangmelima, Nyabessan, and Mbandjock) and urban sites (Yaoundé (Bastos and Nkolondom)), against pyrethroids (permethrin 0.75% and deltamethrin 0.05%) and carbamates (bendiocarb 0.1%). Members of the An. Gambiae s.l. species complex were identified using molecular diagnostic tools. TaqMan assays were used to screen for target site mutations. The expression profiles of eight genes implicated in insecticide resistance were assessed using RT-qPCR. Cuticle hydrocarbon lipids were measured to assess their potential implication in insecticide resistance. Both An. Gambiae and An. coluzzii were detected. An. gambiae was highly prevalent in Sangmelima, Nyabessan, Mbandjock, and Nkolondom. An. coluzzii was the only species found in the Yaoundé city center (Bastos). Low mortality rate to both pyrethroids and bendiocarb was recorded in all sites. High frequency of L1014F allele (75.32–95.82%) and low frequencies of L1014S (1.71–23.05%) and N1575Y (5.28–12.87%) were recorded. The G119S mutation (14.22–35.5%) was detected for the first time in An. gambiae populations from Cameroon. This mutation was rather absent from An. coluzzii populations. The detoxification genes Cyp6m2, Cyp9k1, Cyp6p4, Cyp6z1, as well as Cyp4g16 which catalyzes epicuticular hydrocarbon biosynthesis, were found to be overexpressed in at least one population. The total cuticular hydrocarvbon content, a proxy of cuticular resistance, did not show a pattern associated with pyrethroid resistance in these populations. The rapid emergence of multiple resistance mechanisms in An. Gambiae s.l. population from the South Cameroon forest region is of big concern and could deeply affect the sustainability of insecticide-based interventions strategies in this region.

Transcriptomics of monarch butterflies (Danaus plexippus) reveals that toxic host plants alter expression of detoxification genes and down-regulate a small number of immune genes.

Herbivorous insects have evolved many mechanisms to overcome plant chemical defences, including detoxification and sequestration. Herbivores may also use toxic plants to reduce parasite infection. Plant toxins could directly interfere with parasites or could enhance endogenous immunity. Alternatively, plant toxins could favour down-regulation of endogenous immunity by providing an alternative (exogenous) defence against parasitism. However, studies on genomewide transcriptomic responses to plant defences and the interplay between plant toxicity and parasite infection remain rare. Monarch butterflies (Danaus plexippus) are specialist herbivores of milkweeds (Asclepias spp.), which contain toxic cardenolides. Monarchs have adapted to cardenolides through multiple resistance mechanisms and can sequester cardenolides to defend against bird predators. In addition, high-cardenolide milkweeds confer monarch resistance to a specialist protozoan parasite (Ophryocystis elektroscirrha). We used this system to study the interplay between the effects of plant toxicity and parasite infection on global gene expression. We compared transcriptional profiles between parasite-infected and uninfected monarch larvae reared on two milkweed species. Our results demonstrate that monarch differentially express several hundred genes when feeding on A.curassavica and A.incarnata, two species that differ substantially in cardenolide concentrations. These differentially expressed genes include genes within multiple families of canonical insect detoxification genes, suggesting that they play a role in monarch toxin resistance and sequestration. Interestingly, we found little transcriptional response to infection. However, parasite growth was reduced in monarchs reared on A.curassavica, and in these monarchs, several immune genes were down-regulated, consistent with the hypothesis that medicinal plants can reduce reliance on endogenous immunity.

A high throughput multi-locus insecticide resistance marker panel for tracking resistance emergence and spread in Anopheles gambiae

The spread of resistance to insecticides in disease-carrying mosquitoes poses a threat to the effectiveness of control programmes, which rely largely on insecticide-based interventions. Monitoring mosquito populations is essential, but obtaining phenotypic measurements of resistance is laborious and error-prone. High-throughput genotyping offers the prospect of quick and repeatable estimates of resistance, while also allowing resistance markers to be tracked and studied. To demonstrate the potential of highly-mulitplexed genotypic screening for measuring resistance-association of mutations and tracking their spread, we developed a panel of 28 known or putative resistance markers in the major malaria vector Anopheles gambiae, which we used to screen mosquitoes from a wide swathe of Sub-Saharan Africa (Burkina Faso, Ghana, Democratic Republic of Congo (DRC) and Kenya). We found resistance association in four markers, including a novel mutation in the detoxification gene Gste2 (Gste2-119V). We also identified a duplication in Gste2 combining a resistance-associated mutation with its wild-type counterpart, potentially alleviating the costs of resistance. Finally, we describe the distribution of the multiple origins of kdr resistance, finding unprecedented diversity in the DRC. This panel represents the first step towards a quantitative genotypic model of insecticide resistance that can be used to predict resistance status in An. gambiae.

Integrated analysis of physiological, transcriptomic and metabolomic responses and tolerance mechanism of nitrite exposure in Litopenaeus vannamei

Nitrite accumulation in aquatic environments is a potential risk factor that disrupts multiple physiological functions in aquatic animals. In this study, the physiology, transcriptome and metabolome of the control group (LV-C), nitrite-tolerance group (LV-NT) and nitrite-sensitive group (LV-NS) were investigated to identify the stress responses and mechanisms underlying the nitrite tolerance of Litopenaeus vannamei. After LV-NT and LV-NS were subjected to nitrite stress, the hemocyanin contents were significantly decreased, and hepatopancreas showed severe histological damage compared with LV-C. Likewise, the antioxidant enzymes were also significantly changed after nitrite exposure. The transcriptome data revealed differentially expressed genes associated with immune system, cytoskeleton remodeling and apoptosis in LV-NT and LV-NS. The combination of transcriptomic and metabolomic analysis revealed nitrite exposure disturbed metabolism processes in L. vannamei, including amino acid metabolism, nucleotide metabolism and lipid metabolism. The multiple comparative analysis implicated that higher nitrite tolerance of LV-NT than LV-NS may be attributed to enhanced hypoxia inducible factor-1α expression to regulate energy supply and gaseous exchange. Moreover, LV-NT showed higher antioxidative ability, detoxification gene expression and enhanced fatty acids contents after nitrite exposure in relative to LV-NS. Collectively, all these results will greatly provide new insights into the molecular mechanisms underlying the stress responses and tolerance of nitrite exposure in L. vannamei.