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This is the first report of Bactra bactrana (Kennel, 1901) (Lepidoptera: Tortricidae) attacking a major solanaceous crop, sweet pepper Capsicum annuum L. The infestation was detected in two greenhouses at the area of Tympaki (Southern Crete, Greece). The moth larvae caused typical symptoms of a fruit borer with numerous small holes on the surface of the peppers and extensive damage on the inside of the fruit as a result of the feeding activity. Unknown factors facilitated this major shift in host range since B. bactrana is typically a stem borer of sedges. In addition, the pest status of B. bactrana is currently under question, as in both cases the infestations by the moth were associated with significant yield losses. B. bactrana was moderately controlled with chemicals registered for Lepidoptera management in sweet pepper due to the boring nature of the infestation. Some comparative taxonomic notes are provided to facilitate accurate pest discrimination of related Bactra species. Finally, biological attributes of the species are summarized and are discussed from pest control and ecological perspectives. Because Bactra species have been used in augmentative releases for the control of sage, the implications of our findings on the release of biocontrol agents are placed in perspective. [ABSTRACT FROM AUTHOR]

In autumn 2007, an allium leafminer, Phytomyza gymnostoma Loew (Diptera: Agromyzidae), was recorded for the first time in non-commercial leek crops in the areas of Elassona and Katerini in Central Greece. Leek plants were severely damaged by the leafminer larvae infestation. Typical feeding symptoms were observed, caused by the mining behaviour of larvae within inner and outer leaves of leek plants, resulting in the formation of descending galleries. Pest control was performed locally using commercial insecticides with unknown efficacy mainly due to ignorance of the species biology. [ABSTRACT FROM AUTHOR]

In 2004 the banded greenhouse thrips, Hercinothrips femoralis (Reuter) (Thysanoptera: Thripidae), was recorded for the first time in greenhouse-grown organic banana in the area of Sitia (northeastern Crete). Banana fruits were severely damaged by the thrips infestation and a typical smoky-red discoloration of the fruit was observed. Pest control was poor using either high-pressure washing with potassium fatty acids, or commercially available thrips predators Neoseiulus cucumeris (Oudemans) (Acari: Phytoseiidae) and Orius laevigatus (Fieber) (Hemiptera: Anthocoridae). During 2005 H. femoralis was also found causing severe damage in conventional banana plantations in Arvi, the main banana-growing area of Crete. [ABSTRACT FROM AUTHOR]

The tansy leaf beetle Galeruca tanaceti (L.) is recorded for the first time in Crete as a pest of oregano Origanum vulgare (L.). Oregano is an important herb extensively cultivated both in Crete and mainland Greece. The absence of serious pests due to the production of phenolic compounds and essential oils in the plant tissue has promoted the organic cultivation of oregano. G. tanaceti is a pest of herbaceous plants that has developed appropriate behavioral and biochemical strategies to circumvent host plant defences. In addition, the tansy leaf beetle has very few natural enemies and may pose a serious threat to the organic oregano crops, unless extensive studies of the biology and integrated control of the pest are undertaken. [ABSTRACT FROM AUTHOR]

The European grapevine moth Lobesia botrana (Denis & Shiffermüller 1776) is an economically important pest of the vine-growing areas worldwide. Chemical insecticides have been used for its control; however, its resistance status is largely unknown in many regions. We monitored the susceptibility of several L. botrana populations from Greece and Turkey. In addition, based on RNAseq transcriptome analysis, we identified and phylogenetically classify the cytochrome P450 genes of L. botrana , as well as analysed target site sequences and looked for the presence of known resistance mutations. Resistance against chlorantraniliprole, alpha-cypermethrin, spinetoram, etofenprox, and acetamiprid was very low (below 2.5-fold in all cases, compared to a reference strain from Greece) in all populations from Greece that were included in the study. However, resistance against indoxacarb (4-30-fold), spinosad (5-59-fold), and deltamethrin (18-30 fold) was detected in the L. botrana populations from Turkey, compared to a reference population from Turkey. De novo transcriptome assembly and manual annotation, and subsequent PCR-based analysis of insecticide target sequences (i.e. voltage-gated sodium channel - VGSC: target of pyrethroids and oxadiazines; nicotinic acetylcholine receptor subunit a6 - nAChR_ α 6: target of spinosad; ryanodine receptor - RyR: target of diamides; glutamate-gated chloride channel - GluCl: target of avermectins and; acetylcholinesterase - AChE: target of organophosphates) showed the absence of known resistance mutations in all specimens from both countries. Finally, the L. botrana CYPome (116 genes) was manually analysed and phylogenetically characterised, to provide resources for future studies that will aim the analysis of metabolic resistance.

The tomato leafminer, Tuta absoluta, is an invasive crop pest that has evolved resistance to many of the insecticides used for its control. To facilitate the investigation of the underpinning mechanisms of resistance in this species we generated a contiguous genome assembly using long-read sequencing data. We leveraged this genomic resource to investigate the genetic basis of resistance to the diamide insecticide chlorantraniliprole in Spanish strains of T. absoluta that exhibit high levels of resistance to this insecticide. Transcriptomic analyses revealed that, in these strains, resistance is not associated with previously reported target-site mutations in the diamide target-site, the ryanodine receptor, but rather is associated with the marked overexpression (20- to >100-fold) of a gene encoding a UDP-glycosyltransferase (UGT). Functional expression of this UGT, UGT34A23, via ectopic expression in Drosophila melanogaster demonstrated that it confers strong and significant resistance in vivo. The genomic resources generated in this study provide a powerful resource for further research on T. absoluta. Our findings on the mechanisms underpinning resistance to chlorantraniliprole will inform the development of sustainable management strategies for this important pest., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The pepper fruit fly Atherigona orientalis (Schiner 1968) (Diptera: Muscidae) is a cosmopolitan tropical pest which has been recently recorded in several European countries. The biology of the pest has been primarily associated not only with decomposing fruits and vegetables, but even vertebrate and invertebrate carrion, dung and faeces. Relatively recently, A. orientalis has been reported as a primary pest of pepper fruits as well. In this short communication, we report, for the first time in Greece and, to the best of our knowledge, in Europe, cases of pepper fruit fly damage to pepper fruits in commercial greenhouse crops (in Crete in 2022). In this direction, possible implications and concerns regarding the occurrence of this pest in Crete are discussed.

Resistance to pyrethroid insecticides has evolved in Bactrocera oleae populations in Greece, threatening the efficacy of control interventions based on this insecticide class. Here we report the collection of populations from Crete, with resistance levels reaching up to 132-folds, compared to susceptible laboratory strains and show that pyrethroid resistance is substantially suppressed by the PBO synergist, suggesting the involvement of detoxification enzymes. To identify specific candidate genes implicated in resistance, we performed comparative transcriptomic analysis, between the pyrethroid resistant populations from Crete and the susceptible laboratory strains, using both whole bodies and Malpighian tubules. Several genes were found differentially transcribed between resistant and susceptible flies in each comparison, with P450s being among the most highly over-expressed detoxification genes in pyrethroid resistant populations. Four of the over-expressed P450s (Cyp6A61, Cyp6G6, Cyp4P6 and Cyp6G28) were recombinantly expressed in Escherichia coli and in vitro metabolism assays revealed that CYP6A61 is capable of metabolizing alpha-cypermethrin, while CYP6G6, CYP4P6 and CYP6G28 are capable of metabolizing deltamethrin. No metabolism of neonicotinoid insecticides was recorded. We further silenced CYP6G6 in vivo, via RNAi, which led to a small, but significant increase in deltamethrin toxicity. The study provides valuable information towards the development of molecular diagnostics and evidence-based insecticide resistance management strategies., Competing Interests: Declaration of Competing Interest The authors have no competing interests to declare., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Insecticide resistance has developed in several populations of the whitefly Bemisia tabaci worldwide and threatens to compromise the efficacy of chemical control. The molecular mechanisms underpinning resistance have been characterized and markers associated with the trait have been identified, allowing the development of diagnostics for individual insects., Results: TaqMan and Droplet Digital PCR (ddPCR) assays were developed and validated, in individual and pooled whitefly samples, respectively, for the following target-site mutations: the acetylcholinesterase (ace1) F331W mutation conferring organophosphate-resistance; the voltage-gated sodium channel (vgsc) mutations L925I and T929V conferring pyrethroid-resistance; and the acetyl-CoA carboxylase (acc) A2083V mutation conferring ketoenol-resistance. The ddPCR's limit of detection (LoD) was <0.2% (i.e. detection of one heterozygote whitefly in a pool of 249 wild-type individuals). The assays were applied in 11 B. tabaci field populations from four locations in Crete, Greece. The F331W mutation was detected to be fixed or close to fixation in eight of 11 B. tabaci populations, and at lower frequency in the remaining ones. The pyrethroid-resistance mutations were detected at very high frequencies. The A2083V spiromesifen resistance mutation was detected in eight of 11 populations (frequencies = 6.16-89.56%). Spiromesifen phenotypic resistance monitoring showed that the populations tested had variable levels of resistance, ranging from full susceptibility to high resistance. A strong spiromesifen-resistance phenotype-genotype (A2083V) correlation (r s  = -0.839, P = 0.002) was observed confirming the ddPCR diagnostic value., Conclusion: The ddPCR diagnostics developed in this study are a valuable tool to support evidence-based rational use of insecticides and resistance management strategies. © 2022 Society of Chemical Industry., (© 2022 Society of Chemical Industry.)

Background: Myzus persicae has evolved resistance to various insecticides in Greece. Here we examine the effectiveness of the insecticide flupyradifurone against aphid clones collected from tobacco and peach in Greece during 2017-2020. Furthermore, we monitored the frequency of the neonicotinoid resistance mutation R81T in the sampled clones, and the association between the responses to flupyradifurone and acetamiprid., Results: Of 43 clones tested with flupyradifurone, 6.977%, 60.465% and 32.558% showed low (10-14), moderate (19-89) and high (104-1914) resistance factor (RF) values, respectively. Resistance was higher in clones from peach than from tobacco with 42.308% and 17.647% of clones (respectively) failing into the high RF category (median RF values 67.5 and 36.4 for clones from peach and tobacco, respectively). Acetamiprid resistance was detected in clones collected in 2019-2020, in line with our previous study in Greece. The analysis of the whole dataset (54 clones collected during 2017-2020) revealed that all tobacco clones had RF < 7.5, whereas 55.263%, 18.421% and 26.316% of the peach clones exhibited low (<12), moderate (20-48) and high (100-145) RF values, respectively. A significant but moderate association between flupyradifurone and acetamiprid responses was detected (r = 0.513, P < 0.001). The R81T mutation was detected in aphids from peach (5.6% and 32.6% as homozygotes and heterozygotes, respectively) and in one aphid specimen (heterozygote) from tobacco. R81T was partially associated with the resistance to both insecticides, but many highly resistant clones did not possess the mutation, indicating the possible operation of one or more alternative underlying resistance mechanisms., Conclusions: The use of flupyradifurone and acetamiprid in IPM/IRM should be based on further ongoing susceptibility monitoring. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

Background: Tuta absoluta is a devastating pest in tomato production areas worldwide. After its first introduction to Turkey in 2009, it quickly became the major pest of tomato-growing areas. Although some biocontrol agents have been used, especially in greenhouses, the main control of T. absoluta relies heavily on chemical insecticides. However, failure in chemical control has often been reported due to resistance development. In this study, we investigated (i) the population structure of 22 T. absoluta populations across Turkey by analysing haplotypes, based on the cytochrome oxidase subunit I gene; (ii) the efficacy of three registered insecticides from different classes (metaflumizone, chlorantraniliprole and spinosad) in real field-greenhouse conditions; and (iii) the geographic distribution of target-site mutations associated with insecticide resistance., Results: The efficacy of spinosad was higher than that of chlorantraniliprole and metaflumizone in the greenhouse trials, as documented by the mortality rates obtained, up to 14 days post application. Known resistance mutations in ryanodine receptors (RyR) (i.e. the I4790M/K and G4946E), nicotinic acetylcholine receptors (G275E), acetylcholinesterases (A201S) and voltage-gated sodium channels (F1845Y and V1848I) were found at various frequencies across the populations genotyped. The I4790K diamide resistance mutation in the RyR has been reported for the first time in T. absoluta populations. Although a total of eight haplotypes were found, the overall mean genetic distance was lower than 0.001, indicating the high genetic homogeneity among Turkish T. absoluta populations., Conclusion: The results will contribute to design area-wide resistance management programs in T. absoluta control in Turkey. However, more monitoring studies are needed to implement evidence-based insecticide resistance management strategies in the frame of integrated pest management. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

Cacoecimorpha pronubana (Hübner) (Lepidoptera, Tortricidae) is a highly polyphagous pest of a wide range of crop and ornamental plants. It is of Mediterranean origin and widespread in European and Mediterranean Plant Protection Organization (EPPO) region. For the first time, infestations of Hylotelephium spectabile (Boreau) Ohba (syn.: Sedum spectabile Boreau) (Saxifragales, Crassulaceae) ornamental plants by C. pronubana larvae, in private gardens in urban area of Drama, Greece, were found. Species identification was conducted based on morphology of female genitalia. In addition, due to reports on occurrence of cryptic C. pronubana species within Europe, DNA barcoding was carried out to determine the molecular status of the pest. This communication reports a new host of C. pronubana and places the Greek pest population along with European species clade.

Background: The tomato leafminer, Tuta absoluta, is an economically important pest of tomatoes in Europe, Africa, Asia and South America. In the UK this species is controlled using an integrated pest management (IPM) programme which incorporates the insecticides spinosad and chlorantraniliprole. In response to UK grower concerns of loss of efficacy of these compounds at certain sites, insecticide bioassays were performed on five populations collected from four commercial glasshouses and potential mechanisms of resistance investigated., Results: We observed high levels of resistance to spinosad in four of the strains, and in two of these tolerance to chlorantraniliprole. Selection of one of these strains with chlorantraniliprole rapidly resulted in a line exhibiting potent resistance to this compound. Sequencing of messenger RNA encoding the nicotinic acetylcholine receptor (nAChR) α6 subunit, target of spinosad, revealed Taα6 transcripts in the spinosad-resistant strains that lack exon 4 and encode a highly truncated protein, or contain a triplet deletion in the predicted first transmembrane domain resulting in the loss of a highly conserved amino acid. Sequencing of the ryanodine receptor gene, encoding the target of diamide insecticides, of the chlorantraniliprole-selected line revealed an amino acid substitution (G4903V) that has been previously linked to diamide resistance in populations of T. absoluta in the Mediterranean and South America., Conclusion: Taken together our results reveal emerging resistance in UK populations of T. absoluta to two of the most important insecticides used as part of IPM, with significant implications for the control of this species in the UK. © 2019 Society of Chemical Industry., (© 2019 Society of Chemical Industry.)

Α reduction of pyrethroid efficacy has been recently recorded in Bactrocera oleae, the most destructive insect of olives. The resistance levels of field populations collected from Crete-Greece scaled up to 22-folds, compared to reference laboratory strains. Sequence analysis of the IIS4-IIS6 region of para sodium channel gene in a large number of resistant flies indicated that resistance may not be associated with target site mutations, in line with previous studies in other Tephritidae species. We analyzed the transcriptomic differences between two resistant populations versus an almost susceptible field population and two laboratory strains. A large number of genes was found to be significantly differentially transcribed across the pairwise comparisons. Interestingly, gene set analysis revealed that genes of the 'electron carrier activity' GO group were enriched in one specific comparison, which might suggest a P450-mediated resistance mechanism. The up-regulation of several transcripts encoding detoxification enzymes was qPCR validated, focusing on transcripts coding for P450s. Of note, the expression of contig00436 and contig02103, encoding CYP6 P450s, was significantly higher in all resistant populations, compared to susceptible ones. These results suggest that an increase in the amount of the CYP6 P450s might be an important mechanism of pyrethroid resistance in B. oleae., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Indoxacarb is an important active ingredient extensively used for the control of Tuta absoluta, a major tomato pest, playing a particular role in insecticide resistance management schemes., Results: Reduced susceptibility to indoxacarb was identified (1794-fold resistance) through toxicological bioassays in a field population from Greece and evolved rapidly to resistance after short laboratory selection. Combined bioassays with synergists and biochemical analysis suggested only a partial involvement of detoxification enzymes in the resistant phenotype. To investigate the role of target-site resistance, segment 6 of domain IV of the sodium channel in T. absoluta was cloned and the sequences compared between susceptible and indoxacarb-resistant T. absoluta insects. The presence of the F1845Y and the V1848I indoxacarb resistance mutations was detected and was strongly associated with the phenotype. These amino acid substitutions correspond to recently characterised indoxacarb resistance mutations in diamondback moth (Plutella xylostella). Robust and accurate PCR-RFLP assays were subsequently developed and successfully validated for detecting both indoxacarb resistance mutations in field T. absoluta populations., Conclusion: The identification of indoxacarb resistance mutations and the development of diagnostic tools will allow early detection of indoxacarb resistance, facilitating implementation of appropriate resistance management strategies, thus delaying the spread of resistance. © 2016 Society of Chemical Industry., (© 2016 Society of Chemical Industry.)

Diamide insecticides are used widely against lepidopteran pests, acting as potent activators of insect Ryanodine Receptors (RyRs) and thus inducing muscle contraction and eventually death. However, resistant phenotypes have recently evolved in the field, associated with the emergence of target site resistance mutations (G4946E/V and I4790M). We investigated the frequency of the mutations found in a resistant population of Tuta absoluta from Greece (G4946V ~79% and I4790M ~21%) and the associated diamide resistance profile: there are very high levels of resistance against chlorantraniliprole (9329-fold) and flubendiamide (4969-fold), but moderate levels against cyantraniliprole (191-fold). To further investigate functionally the contribution of each mutation in the resistant phenotype, we used CRISPR/Cas9 to generate genome modified Drosophila carrying alternative allele combinations, and performed toxicity bioassays against all three diamides. Genome modified flies bearing the G4946V mutation exhibited high resistance ratios to flubendiamide (91.3-fold) and chlorantraniliprole (194.7-fold) when compared to cyantraniliprole (5.4-fold). Flies naturally wildtype for the I4790M mutation were moderately resistant to flubendiamide (15.3-fold) but significantly less resistant to chlorantraniliprole (7.5-fold), and cyantraniliprole (2.3-fold). These findings provide in vivo functional genetic confirmation for the role and relative contribution of RyR mutations in diamide resistance and suggest that the mutations confer subtle differences on the relative binding affinities of the three diamides at an overlapping binding site on the RyR protein., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Background: The cotton whitefly Bemisia tabaci (Gennadius) is among the most important pests of numerous crops and a vector of more than 100 plant viruses, causing significant crop losses worldwide. Managing this pest as well as inhibiting the transmission of major viruses such as tomato yellow leaf curl virus (TYLCV) are of utmost importance for sustainable yields. The efficacy against both whitefly and virus transmission of the novel systemic butenolide insecticide flupyradifurone was investigated in this study., Results: The inhibition of TYLCV transmission by flupyradifurone was compared to that by thiamethoxam, a neonicotinoid insecticide reported to inhibit virus transmission. The experiment was performed under high virus pressure conditions (10 viruliferous insects per plant for 48 h) using a fully characterized field strain of B. tabaci. The insecticides were foliarly applied at recommended label rates under greenhouse conditions. Flupyradifurone suppressed virus transmission by 85% while levels of suppression after thiamethoxam treatments were just 25% and significantly lower. In untreated control plots, 100% of plants were infected by TYLCV. The observed difference in the potential to suppress virus transmission is linked to a strong knockdown effect as well as prolonged feeding inhibition in flupyradifurone treatments., Conclusion: Flupyradifurone is shown to be an extremely useful, fast-acting, new chemical tool in integrated crop management offering simultaneous control of whiteflies and strong suppression of viral infections via its rapid knockdown action and good residual activity. © 2017 Society of Chemical Industry., (© 2017 Society of Chemical Industry.)

The development of insecticide resistance in insect pests of crops is a growing threat to sustainable food production, and strategies that slow the development of resistance are therefore urgently required. The insecticide synergist piperonyl butoxide (PBO) inhibits certain insect detoxification systems and so may delay the evolution of metabolic resistance. In the current study we characterized resistance development in the silverleaf whitefly, Bemisia tabaci, after selection with either a neonicotinoid (thiacloprid) or pyrethroid (alpha-cypermethrin) insecticide alone or in combination with PBO. Resistance development was significantly suppressed (> 60%) in the line selected with alpha-cypermethrin + PBO compared to the line selected with alpha-cypermethrin alone. RNA sequencing (RNAseq) analyses revealed an increase in frequency of a knock-down resistance mutation but no differentially expressed genes were identified that could explain the sensitivity shift. No significant difference was observed in the level of resistance between the thiacloprid and thiacloprid + PBO selected lines, and RNA sequencing (RNAseq) analyses revealed that the cytochrome P450 monooxygenase CYP6CM1, known to metabolize neonicotinoids, was significantly upregulated (>10-fold) in both lines. The findings of this study demonstrate that PBO used in combination with certain insecticides can suppress the development of resistance in a laboratory setting; however, the mechanism by which PBO supresses resistance development remains unclear., (© 2016 The Royal Entomological Society.)

Insect ryanodine receptors (RyR) are the molecular target-site for the recently introduced diamide insecticides. Diamides are particularly active on Lepidoptera pests, including tomato leafminer, Tuta absoluta (Lepidoptera: Gelechiidae). High levels of diamide resistance were recently described in some European populations of T. absoluta, however, the mechanisms of resistance remained unknown. In this study the molecular basis of diamide resistance was investigated in a diamide resistant strain from Italy (IT-GELA-SD4), and additional resistant field populations collected in Greece, Spain and Brazil. The genetics of resistance was investigated by reciprocally crossing strain IT-GELA-SD4 with a susceptible strain and revealed an autosomal incompletely recessive mode of inheritance. To investigate the possible role of target-site mutations as known from diamondback moth (Plutella xylostella), we sequenced respective domains of the RyR gene of T. absoluta. Genotyping of individuals of IT-GELA-SD4 and field-collected strains showing different levels of diamide resistance revealed the presence of G4903E and I4746M RyR target-site mutations. These amino acid substitutions correspond to those recently described for diamide resistant diamondback moth, i.e. G4946E and I4790M. We also detected two novel mutations, G4903V and I4746T, in some of the resistant T. absoluta strains. Radioligand binding studies with thoracic membrane preparations of the IT-GELA-SD4 strain provided functional evidence that these mutations alter the affinity of the RyR to diamides. In combination with previous work on P. xylostella our study highlights the importance of position G4903 (G4946 in P. xylostella) of the insect RyR in defining sensitivity to diamides. The discovery of diamide resistance mutations in T. absoluta populations of diverse geographic origin has serious implications for the efficacy of diamides under applied conditions. The implementation of appropriate resistance management strategies is strongly advised to delay the further spread of resistance., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: Bemisia tabaci is one of the most damaging agricultural pests world-wide. Although its control is based on insecticides, B. tabaci has developed resistance against almost all classes of insecticides, including neonicotinoids., Results: We employed an RNA-seq approach to generate genome wide expression data and identify genes associated with neonicotinoid resistance in Mediterranean (MED) B. tabaci (Q1 biotype). Twelve libraries from insecticide resistant and susceptible whitefly populations were sequenced on an Illumina Next-generation sequencing platform, and genomic sequence information of approximately 73 Gbp was generated. A reference transcriptome was built by de novo assembly and functionally annotated. A total of 146 P450s, 18 GSTs and 23 CCEs enzymes (unigenes) potentially involved in the detoxification of xenobiotics were identified, along with 78 contigs encoding putative target proteins of six different insecticide classes. Ten unigenes encoding nicotinic Acetylcholine Receptors (nAChR), the target of neoinicotinoids, were identified and phylogenetically classified. No nAChR polymorphism potentially related with the resistant phenotypes, was observed among the studied strains. DE analysis revealed that among the 550 differentially (logFC > 1) over-transcribed unigenes, 52 detoxification enzymes were over expressed including unigenes with orthologues in P450s, GSTs, CCE and UDP-glucuronosyltransferases. Eight P450 unigenes belonging to clades CYP2, CYP3 and CYP4 were highly up-regulated (logFC > 2) including CYP6CM1, a gene already known to confer imidacloprid resistance in B. tabaci. Using quantitative qPCRs, a larger screening of field MED B. tabaci from Crete with known neonicotinoid phenotype was performed to associate expression levels of P450s with resistance levels. Expression levels of five P450s, including CYP6CM1, were found associated with neonicotinoid resistance. However, a significant correlation was found only in CYP303 and CYP6CX3, with imidacloprid and acetamiprid respectively., Conclusion: Our work has generated new toxicological data and genomic resources which will significantly enrich the available dataset and substantially facilitate the molecular studies in MED B. tabaci. No evidence of target site neonicotinoid resistance has been found. Eight P450 unigenes, including CYP6CM1, were found significantly over-expressed in resistant B. tabaci. This study suggests at least two novel P450s (CYP303 and CYP6CX3) as candidates for their functional characterization as detoxification mechanisms of neonicotinoid resistance in B. tabaci.

Cotton whitefly, Bemisia tabaci (Genn.) (Homoptera: Aleyrodidae) is a major sucking pest in many agricultural and horticultural cropping systems globally. The frequent use of insecticides of different mode of action classes resulted in populations resisting treatments used to keep numbers under economic damage thresholds. Recently it was shown that resistance to neonicotinoids such as imidacloprid is linked to the over-expression of CYP6CM1, a cytochrome P450 monooxygenase detoxifying imidacloprid and other neonicotinoid insecticides when recombinantly expressed in insect cells. However over-expression of CYP6CM1 is also known to confer cross-resistance to pymetrozine, an insecticide not belonging to the chemical class of neonicotinoids. In addition we were able to demonstrate by LC-MS/MS analysis the metabolisation of pyriproxyfen by recombinantly expressed CYP6CM1. Based on our results CYP6CM1 is one of the most versatile detoxification enzymes yet identified in a pest of agricultural importance, as it detoxifies a diverse range of chemical classes used to control whiteflies. Therefore we developed a field-diagnostic antibody-based lateral flow assay which detects CYP6CM1 protein at levels providing resistance to neonicotinoids and other insecticides. The ELISA based test kit can be used as a diagnostic tool to support resistance management strategies based on the alternation of different modes of action of insecticides., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Flonicamid is a novel systemic insecticide that acts as a feeding blocker with potential use against whiteflies within IPM control tactics. Flonicamid efficacy against Bemisia tabaci Mediterranean populations from Crete was examined, as well as side effects on selected beneficials used extensively in current IPM schemes., Results: Low variability in adulticide activity was detected (

Background: Tuta absoluta (Meyrick), in only a few years, has become a serious threat to global tomato production. Depending on the cropping system and infestation pressure, T. absoluta control may rely heavily on insecticide applications. By means of a validated IRAC bioassay method, the toxicity of all insecticides registered for T. absoluta control in Greece has been estimated. A non-registered pyrethroid insecticide was also included in this study., Results: Low heterogeneity was detected in the populations tested with most insecticides. The LC50 ranged from 0.31 to 1.31 mg L(-1) for flubendiamide, from 0.12 to 0.53 mg L(-1) for chlorantraniliprole, from 0.03 to 0.12 mg L(-1) for emamectin benzoate, from 0.08 to 0.26 mg L(-1) for spinosad, from 31.8 to 159.5 mg L(-1) for metaflumizone, from 1.73 to 17.5 mg L(-1) for indoxacarb, from 530 to 2038 mg L(-1) for chlorpyriphos and finally from 475 to 794 mg L(-1) for cypermethrin. The variability of the LC50 values among the tested populations was low (RR under 5×), except for indoxacarb (RR = 10×). In the absence of a reference strain, comparisons with the recommended label rates were performed. Evidence of potential control failures was detected using probit analysis estimates for cypermethrin, chlorpyriphos and metaflumizone., Conclusions: For most registered insecticides, a solid set of baseline data has been presented that can be used in future resistance monitoring studies. The interaction of metaflumizone with T. absoluta has been discussed, and for chlorpyriphos it is suspected that the resistance level is underestimated with the present dataset. Finally, it has been demonstrated that the pyrethroid cypermethrin would provide insufficient control of the pest., (© 2012 Society of Chemical Industry.)

Background: Tuta absoluta(Meyrick) is one of the most serious pests of tomato recently introduced in the Mediterranean region. A novel bioassay method designed for the accurate determination of insecticide toxicity on T. absoluta (IRAC method No. 022) was validated by three different laboratories [Greece (NAGREF), Italy (UC) and Spain (UPCT)] on European populations., Results: The insecticides indoxacarb and chlorantraniliprole were used as reference products. The IRAC leaf dip method is easy to perform, producing repeatable, homogeneous responses. LC(50) values for indoxacarb ranged between 1.8 and 17.9 mg L(-1) (NAGREF), 0.93 and 10.8 mg L(-1) (UC) and 0.20 and 0.70 mg L(-1) (UPCT), resulting in a tenfold, 12-fold and fourfold difference between the least and most susceptible populations at each laboratory respectively. For chlorantraniliprole, LC(50) values ranged between 0.10 and 0.56 mg L(-1) (NAGREF), 0.23 and 1.34 mg L(-1) (UC) and 0.04 and 0.24 mg L(-1) (UPCT), resulting in a sixfold difference in all three cases. Overall, UPCT reported lower mean LC(50) to indoxacarb, while UC reported higher LC(50) to chlorantraniliprole., Conclusions: The new bioassay is reliable, providing a useful tool in the design of IRM strategies. Within each country/lab, the variability observed in the results for both indoxacarb and chlorantraniliprole can be attributed to natural variation. Future research is necessary to determine the extent to which it is possible to compare results among laboratories., (Copyright © 2012 Society of Chemical Industry.)

Background: A major problem of crop protection in Crete, Greece, is the control of Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae) with chemical insecticides owing to the rapid development of resistance. The aim of this study was to investigate the establishment of resistance and the underlying mechanisms to major insecticide classes with classical bioassays and known biochemical resistance markers., Results: During a 2005-2007 survey, 53 Q biotype populations were collected. Application history records showed extensive use of neonicotinoids, organophosphates, carbamates and pyrethroids. High resistance levels were identified in the majority of populations (>80%) for imidacloprid (RF: 38-1958x) and alpha-cypermethrin (RF: 30-600x). Low resistance levels (RF < 12) were observed for pirimiphos-methyl. A strong correlation between resistance to imidacloprid and the number of applications with neonicotinoids was observed. Significant correlations were observed between COE and P450-dependent monoxygenase activity with resistance to alpha-cypermethrin and imidacloprid respectively. A propoxur-based AChE diagnostic test indicated that iAChE was widespread in most populations. Resistance levels for alpha-cypermethrin were increased when compared with a previous survey (2002-2003). Differentiation of LC(50) values between localities was observed for imidacloprid only., Conclusion: Bemisia tabaci resistance evolved differently in each of the three insecticides studied. Imidacloprid resistance seems less established and less persistent than alpha-cypermethrin resistance. The low resistance levels for pirimiphos-methyl suggest absence of cross-resistance with other organophosphates or carbamates used., (2008 Society of Chemical Industry)

The two most damaging biotypes of Bemisia tabaci, B and Q, have both evolved strong resistance to the neonicotinoid insecticide imidacloprid. The major mechanism in all samples investigated so far appeared to be enhanced detoxification by cytochrome P450s monooxygenases (P450s). In this study, a polymerase chain reaction (PCR) technology using degenerate primers based on conserved P450 helix I and heme-binding regions was employed to identify P450 cDNA sequences in B. tabaci that might be involved in imidacloprid resistance. Eleven distinct P450 cDNA sequences were isolated and classified as members of the CYP4 or CYP6 families. The mRNA expression levels of all 11 genes were compared by real-time quantitative RT-PCR across nine B and Q field-derived strains of B. tabaci showing strong resistance, moderate resistance or susceptibility to imidacloprid. We found that constitutive over-expression (up to approximately 17-fold) of a single P450 gene, CYP6CM1, was tightly related to imidacloprid resistance in both the B and Q biotypes. Next, we identified three single-nucleotide polymorphic (SNP) markers in the intron region of CYP6CM1 that discriminate between the resistant and susceptible Q-biotype CYP6CM1 alleles (r-Q and s-Q, respectively), and used a heterogeneous strain to test for association between r-Q and resistance. While survivors of a low imidacloprid dose carried both the r-Q and s-Q alleles, approximately 95% of the survivors of a high imidacloprid dose carried only the r-Q allele. Together with previous evidence, the results reported here identify enhanced activity of P450s as the major mechanism of imidacloprid resistance in B. tabaci, and the CYP6CM1 gene as a leading target for DNA-based screening for resistance to imidacloprid and possibly other neonicotinoids in field populations.

The effects of the entomopathogenic fungus Lecanicillium longisporum (Zimmerman) Zare & Gams on three parameters of behaviour (feeding, reproduction and movement) of the green peach aphid Myzus persicae (Homoptera: Aphididae) were investigated in the laboratory. Visual analysis of video tapes established that honeydew excretion events of mycosed aphids gradually declined from 2 d post inoculation and reproduction rate was significantly reduced 2 d prior to death (which occurred on day 6); both parameters were stable in controls over the same period. A detailed comparison was made between mobility of aphids during infection with two isolates of L. longisporum, using image analysis of video recordings. Both isolates caused an increase in activity at the beginning of mycosis (during fungal germination and cuticle invasion) though the intensity and the duration of this behaviour varied with the isolate. The possibility that increased movement in early mycosis helps disseminate disease is discussed in the light of the observation that saprophytic surface growth occurs on living M. persicae as it does in at least some other Lecanicillium spp-insect interactions.

The resistance levels to alpha-cypermethrin, bifenthrin, pirimiphos-methyl, endosulfan and imidacloprid were determined in Bemisia tabaci (Gennadius) from Crete. Five B tabaci populations collected from greenhouse and outdoor crops were bioassayed and compared with a reference susceptible strain. Bemisia tabaci collected in a floriculture greenhouse exhibited the highest resistance against all insecticides: at LC50, resistance factors were 23-fold for bifenthrin, 80-fold for alpha-cypermethrin, 18-fold for pirimiphos-methyl, 58-fold for endosulfan and 730-fold for imidacloprid. A population collected on outdoor melons was more susceptible than the reference strain against all insecticides tested, suggesting the occurrence of local highly susceptible B tabaci populations in 'refugia'. In pairwise comparisons of resistance levels, correlation was observed between the LC50 values of the pyrethroid insecticides bifenthrin and alpha-cypermethrin.