Insecticide-resistant pollen beetles (Meligethes aeneus F) found in Danish oilseed rape (Brassica napus L) fields

The pollen beetle is the most important pest in Danish oilseed rape fields. In 2001, we screened a broad range of pollen beetle populations for pyrethroid and dimethoate resistance. A standard dip-test was used to test insecticide resistance in 18 populations collected from oilseed winter and spring rape fields. The beetles were treated with four different insecticides: the pyrethroids tau-fluvalinate, lambda-cyhalothrin and esfenvalerate, and the organophosphate dimethoate. The results show that up to 99% of the pollen beetles survived Danish standard doses of pyrethroids and up to 36% of the beetles survived standard doses of dimethoate.     

Occurrence of insecticide resistant pollen beetles (Meligethes aeneus F.) in Danish oilseed rape (Brassica napus L.) crops

The pollen beetle is the most important pest in Danish oilseed rape crops and it is essential that farmers are able to control this pest, especially in years when the economic damage threshold will be exceeded. About seven years ago, Danish oilseed rape growers began reporting that the pyrethroids seemed to be losing effectiveness towards pollen beetles. In 2001 18 populations collected from winter and spring oilseed rape fields were tested and very high level of insecticide resistance was found. In 2003 100 populations of pollen beetles were tested, using a FAO dip-test, for resistance to fenitrothion, lambda-cyhalothrin and tau-fluvalinate. It can be concluded that many Danish populations of pollen beetles are resistant to the pyrethroid lambda-cyhalothrin and to a lesser extent to the pyrethroid tau-fluvalinate. No resistance to the organophosphate fenitrothion was found.     

Cytochrome P450 mediated pyrethroid resistance in European populations of Meligethes aeneus (Coleoptera: Nitidulidae)

Pollen beetle, Meligethes aeneus (Coleoptera: Nitidulidae) is a major pest on several million hectares in European winter oilseed rape cultivation. Synthetic pyrethroids have been successfully used for many years to keep them under economic damage thresholds. Recently wide-spread resistance development to pyrethroids in pollen beetle populations was described in many European countries, including Germany, France, Poland, Denmark and others. Resistance monitoring is conducted by incubating beetles for 24 h in glass vials coated with different concentrations of lambda-cyhalothrin. Using such an assay format we were able to show cross-resistance to other pyrethroids, such as deltamethrin, cypermethrin, and to a somewhat lower extent bifenthrin, etofenprox and tau-fluvalinate. Here we also investigated in more detail in 27 different populations the biochemical mechanism of pyrethroid resistance. Synergism experiments revealed a high synergistic potential for piperonyl butoxide in vivo, whereas other compounds such as S,S,S-tributylphosphorotrithioate and diethylmaleimide failed to suppress pyrethroid resistance. Incubating microsomal fractions of pollen beetle with deltamethrin and subsequent LC–MS/MS analysis revealed 4-OH-deltamethrin as the major metabolite. Metabolite formation in vitro and pyrethroid resistance in vivo is correlated and inhibition trials with piperonyl butoxide, tebuconazole and aminobenzotriazole suggest the involvement of cytochrome P450′s. Furthermore we were able to show cross-resistance to tau-fluvalinate which is supported by the competitive inhibition of 4-OH-deltamethrin formation by increasing concentrations of tau-fluvalinate in microsomal hydroxylation assays. Although we provided clear experimental evidence for an oxidative mechanism of resistance in numerous populations, other mechanisms might be involved based on the data discussed.     

Pyrethroid resistance monitoring in European populations of pollen beetle (Meligethes spp.): a coordinated approach through the Insecticide Resistance Action Committee (IRAC)

BACKGROUND: Pollen beetle (Meligethes spp.) is a major pest of European oilseed rape crops. Its resistance to pyrethroid insecticides has been recorded in samples of beetles collected in Europe since at least 1999, and problems with the control of the beetle in the field have been widely reported. In 2007, a Pollen Beetle Working Group was formed through the Insecticide Resistance Action Committee (IRAC) in order to coordinate efforts for surveying pyrethroid resistance development. RESULTS: The results of the first 3 years of the pollen beetle pyrethroid susceptibility survey using a laboratory test are presented in this paper. Resistant beetle samples were collected from 20 of the 21 countries surveyed, with a general trend of increasing frequency and spread of resistant samples in European oilseed‐rape‐growing regions. CONCLUSION: Pyrethroid‐resistant beetles dominate in Western and Central Europe and are becoming established in the North and East, the main oilseed‐rape‐growing areas of Europe. The development and spread of pyrethroid‐resistant pollen beetles highlights the need for effective management strategies for oilseed rape insect pests. Copyright © 2011 Society of Chemical Industry     

Pyrethroid resistance and thiacloprid baseline susceptibility of European populations of Meligethes aeneus (Coleoptera: Nitidulidae) collected in winter oilseed rape

BACKGROUND: Pollen beetle, Meligethes aeneus F. (Coleoptera: Nitidulidae), is a major pest in European winter oilseed rape. Recently, control failures with pyrethroid insecticides commonly used to control this pest have been reported in many European countries. For resistance management purposes, the neonicotinoid insecticide thiacloprid was widely introduced as a new mode of action for pollen beetle control. RESULTS: A number of pollen beetle populations collected in Germany, France, Austria, Great Britain, Sweden, Denmark, Finland, Poland, Czech Republic and Ukraine were tested for pyrethroid resistance using lambda‐cyhalothrin‐coated glass vials (adult vial test). Most of the populations tested exhibited substantial levels of resistance to lambda‐cyhalothrin, and resistance ratios ranged from < 10 to > 2000. A similar resistance monitoring bioassay for the neonicotinoid insecticide thiacloprid was developed and validated by assessing baseline susceptibility data for 88 European pollen beetle populations. A variation of less than fivefold in response to thiacloprid was detected. The thiacloprid adult vial bioassay is based on glass vials coated with an oil‐dispersion‐based formulation of thiacloprid, resulting in a much better bioavailability compared with technical material. Analytical measurements revealed a > 56 and 28 day stability of thiacloprid and lambda‐cyhalothrin in coated glass vials at room temperature, respectively. No cross‐resistance between thiacloprid and lambda‐cyhalothrin based on log‐dose probit–mortality data was detected. CONCLUSION: Pyrethroid resistance in many European populations of M. aeneus was confirmed, whereas all populations are susceptible to thiacloprid when tested in a newly designed and validated monitoring bioassay based on glass vials coated with oil‐dispersion‐formulated thiacloprid. Based on the homogeneous results, it is concluded that thiacloprid could be an important chemical tool for pollen beetle resistance management strategies in European winter oilseed rape. Copyright © 2011 Society of Chemical Industry     

Pyrethroid sensitivity monitoring in Germany of oilseed rape pest insects other than pollen beetle

Pyrethroid resistant pollen beetles ( Meligethes aeneus ) are widely distributed in Germany and other European countries. Other insect pests on oilseed rape are also exposed to pyrethroids which often have more than one application per season. In spite of this scarce information about the sensitivity of other oilseed rape pest insects is available. To address this monitoring of pyrethroid sensitivity of different oilseed rape pest insects has been carried out since 2005. Laboratory experiments were conducted using an adult-vial-test design with samples collected in different areas of Germany.
Most samples of Ceutorhynchus napi , C. assimilis , C. pallidactylus , Dasineura brassicae , Phyllotreta spp. and Psylliodes chrysocephala tested from 2005–2007 showed 100% mortality at 0.003 and 0.015 µg/cm² lambda-cyhalothrin used as reference test pyrethroid. Some samples especially of C. pallidactylus , C. napi and C. assimilis showed a lower mortality with alive and healthy beetles at 0.015 µg/cm² lambda-cyhalothrin. For all samples tested from 2005–2007 100% mortality was found only at higher rates of 0.0375 or 0.075 µg/cm² lambda-cyhalothrin.
To date, no clear pyrethroid resistance has been detected in other oil seed rape pest insect populations in Germany.     

Ecological approaches to the control of pollen beetles in oilseed rape

At a time of increasing demand for rapeseed oil for biofuel and food use and as increasing areas are grown, the risk of pollen beetle resistance to pyrethroids presents a significant threat to the sustainability of the oilseed rape crop and to farm incomes. Measures are urgently required to reduce the use of insecticides against pollen beetles, to preserve activity of the limited armoury of insecticides and minimise environmental pollution. In this paper, the status of pollen beetle resistance to pyrethroid insecticides and current control methods are presented from a UK perspective. Three ecological approaches to the control of pollen beetles that are based on research into their behaviour and ecology and that of their natural enemies are highlighted: use of monitoring, trap cropping and conservation biological control. These approaches have the potential to significantly reduce insecticide use against pollen beetles by helping to identify when spray thresholds have been breached, reducing pest incidence in the crop and increasing populations of natural enemies, respectively.     

First molecular genotyping of voltage gated sodium channel alleles in Culex quinquefasciatus populations in Malaysia

A nationwide investigation was performed to detect the presence of 1014 mutation(s) in voltage gated sodium channel (kdr) gene of Culex quinquefasciatus from 14 residential areas across 13 states and a federal territory in Malaysia. Molecular genotyping of kdr mutation was performed via a modified three tubes allele-specific-polymerase chain reaction (AS-PCR) and direct sequencing of kdr gene. Based on the results of AS-PCR, homozygous susceptible (SS) genotype was found in nine out of 14 populations with 38 individuals from a total sample size of 140. Heterozygous (RS) genotype was most predominant (99 individuals) and distributed across all study sites. Homozygous resistance (RR) genotype was detected in Perak (one individual) and Selangor (two individuals). The resistance kdr allele frequencies ranged from 0.1 to 0.55, with the highest being detected in Cx. quinquefasciatus population from Selangor. This study has documented the first field-evolved instance of 1014F mutation in Malaysian mosquitoes and the findings of this study could be utilized in the implementation of strategic measures in vector control programs in Malaysia.     

Current status of resistance in pollen beetle (Meligethes aeneus F.) to selected active substances of insecticides in Poland

Chemical plant protection is still an indispensable method in effective oilseed rape protection against the pollen beetle ( Meligethes aeneus F.) in Poland. This species is able to develop strong resistance against active substances used in synthetic insecticides. The phenomenon of pollen beetle resistance is currently found in many European countries. Therefore the objective of the study was to determine the present susceptibility level of Polish populations of pollen beetle to the main groups of insecticides recommended in Poland for pollen beetle control. The research showed high resistance of most tested Polish pollen beetle populations to many active substances of all chemical groups of insecticides applied in Poland.     

Incidence and spread of knockdown resistance (kdr) in German Colorado potato beetle (Leptinotarsa decemlineata Say) populations

The Colorado beetle, Leptinotarsa decemlineata (Say), is one of the most important pests in many potato‐producing regions. Colorado beetle infestations are normally kept under economic damage thresholds by applying insecticides such as pyrethroids. Pyrethroids are known to act on voltage‐gated sodium channels and have been used for several decades to control L. decemlineata. Their continuous and widespread use has resulted in the development of resistance, which is often linked to a L1014F target‐site mutation in the voltage‐gated sodium channel, known as knockdown resistance (kdr). Since pyrethroids are used in many potato‐growing regions in Germany, more than 140 L. decemlineata samples were collected and analysed for the presence of the kdr allele by pyrosequencing diagnostics. Results showed that kdr is present in many German L. decemlineata populations, but even its homozygous presence does not substantially compromise the efficacy of recommended label rates of pyrethroids, as demonstrated by bioassays and crossing experiments. The implications of these findings for resistance management are briefly discussed.     

Incidence of pyrethroid‐resistant oilseed rape pests in Germany

BACKGROUND: Failures in pollen beetle control using pyrethroids since 2005 indicated pyrethroid resistance in Germany. Therefore, resistance monitoring using bioassays was established in Germany for oilseed rape pest insects. RESULTS: The spread and intensity of pyrethroid resistance of Meligethes aeneus increased from 2005 onwards, with no sensitive samples left in any region of Germany in 2011. Sensitivity also declined for the newly introduced actives bifenthrin, etofenprox (both class‐I pyrethroids) and tau‐fluvalinate; all three claimed to be less affected by resistance, although there was no clear cross‐resistance to lambda‐cyhalothrin (class‐II pyrethroid). In the German region with the longest tradition and high intensity of oilseed rape production, pyrethroid resistance of Psylliodes chrysocephala and Ceutorhynchus obstrictus, with resistance factors of up to 81 and 140 respectively, was detected. CONCLUSION: The intensive use of only one mode of action for many years is risky, because even pest insects with a low intrinsic resistance risk may develop resistance. Therefore, resistance strategies need to include several control options for pest insects needing regular treatments. Copyright © 2012 Society of Chemical Industry     

Frequency detection of pyrethroid resistance allele in Anopheles sinensis populations by real-time PCR amplification of specific allele (rtPASA)

To investigate the level of pyrethroid resistance in Anopheles sinensis Wiedemann 1828 (Diptera: Culicidae), a major malaria vector in Korea, we cloned and sequenced the IIS4-6 transmembrane segments of the sodium channel gene that encompass the most widely known kdr mutation sites. Sequence analysis revealed the presence of the major Leu-Phe mutation and a minor Leu-Cys mutation at the same position in permethrin-resistant field populations of An. sinensis. To establish a routine method for monitoring resistance, we developed a simple and accurate real-time PCR amplification of specific allele (rtPASA) protocol for the estimation of resistance allele frequencies on a population basis. The kdr allele frequency of a field population predicted by the rtPASA method (60.8%) agreed well with that determined by individual genotyping (61.7%), demonstrating the reliability and accuracy of rtPASA in predicting resistance allele frequency. Using the rtPASA method, the kdr allele frequencies in several field populations of An. sinensis were determined to range from 25.0 to 96.6%, suggestive of widespread pyrethroid resistance in Korea.     

Different methods of monitoring susceptibility of oilseed rape beetles to insecticides

The resistance of oilseed rape pollen beetles to lambda-cyhalothrin has increased in Germany over the period 2002 to 2007. Oilseed rape pollen beetles are very good at dispersing, therefore field results should always be compared with those of laboratory experiments. The response of oilseed rape pollen beetles to active substances may not be the same as that to formulated products. Bioassays in which beetles are exposed to insecticides on either complete inflorescences or flower buds give different results. It is evident that samples of the genus Meligethes collected in the field can include several different species. Monitoring for insecticide resistance should be done using only the true oilseed rape pollen beetle.     

High resistance to pyrethroid insecticides in the Polish pollen beetle (<Emphasis Type="Italic">Meligethes aeneus F.</Emphasis>): the role of oxidative metabolism

Pollen beetle (Meligethes aeneus F.) (PB) causes very serious problems in oilseed rape (Brassica napus var. oleifera L.) cultivations in Poland and other European countries. Pyrethroid insecticides are the most frequently used type for controlling PB in Poland. This chemical group has been used in Poland since the beginning of the 1980’s and its strong selection pressure on PB resulted in some level of resistance of this pest to many active substances. The aim of this investigation was to analyze, with the use of synergists, the main mechanisms responsible for resistance of PB to pyrethroids. The results indicate the main resistance factor in Polish PB populations to be monooxygenase enzymes.     

Sodium channel mutations (T929I and F1534S) found in pyrethroid-resistant strains of the cigarette beetle,Lasioderma serricorne (Coleoptera: Anobiidae)

RNA-seq data analysis of cigarette beetle (Lasioderma serricorne) strains having different sensitivities to pyrethroids identified sodium channel mutations in strains showing pyrethroid resistance: the T929I and F1534S mutations. These results suggest that reduced sensitivity of the sodium channel confers the pyrethroid resistance of L. serricorne. Results also showed that the F1534S mutation mostly occurred concurrently with the T929I mutation. The functional relation between both mutations for pyrethroid resistance is discussed.     

Determination of knockdown resistance allele frequencies in global human head louse populations using the serial invasive signal amplification reaction

BACKGROUND: Pediculosis is the most prevalent parasitic infestation of humans. Resistance to pyrethrin‐ and pyrethroid‐based pediculicides is due to knockdown (kdr)‐type point mutations in the voltage‐sensitive sodium channel α‐subunit gene. Early detection of resistance is crucial for the selection of effective management strategies. RESULTS: Kdr allele frequencies of lice from 14 countries were determined using the serial invasive signal amplification reaction. Lice collected from Uruguay, the United Kingdom and Australia had kdr allele frequencies of 100%, while lice from Ecuador, Papua New Guinea, South Korea and Thailand had kdr allele frequencies of 0%. The remaining seven countries investigated, including seven US populations, two Argentinian populations and populations from Brazil, Denmark, Czech Republic, Egypt and Israel, displayed variable kdr allele frequencies, ranging from 11 to 97%. CONCLUSION: The newly developed and validated SISAR method is suitable for accurate monitoring of kdr allele frequencies in head lice. Proactive management is needed where kdr‐type resistance is not yet saturated. Based on sodium channel insensitivity and its occurrence in louse populations resistant to pyrethrin‐ and pyrethroid‐based pediculicides, the T917I mutation appears to be a key marker for resistance. Results from the Egyptian population, however, indicate that phenotypic resistance of lice with single or double mutations (M815I and/or L920F) should also be determined. Copyright © 2010 Society of Chemical Industry     

竞争性特异等位基因PCR检测山东甜菜夜蛾对高效氯氰菊酯的抗性

为明确山东地区甜菜夜蛾对高效氯氰菊酯抗性水平,采用竞争性特异等位基因PCR方法,检测甜菜夜蛾钠离子通道基因片段上L1014F的突变频率.结果表明,敏感种群个体中均未发现突变个体;田间章丘种群92％个体具有L1014F突变,其中64％为L1014F突变纯合子,28％为L1014F突变杂合子.经高效氯氰菊酯汰选5代后的抗性种群检测个体中均未发现敏感型纯合子.安丘种群的L1014F突变等位基因频率最高,为85％,泰安、滕州和滨州种群分别为50％、43.33％和30％,相关性分析表明甜菜夜蛾L1014F突变等位基因频率与对高效氯氰菊酯不敏感度呈线性正相关(Y=1.106x +0.079,R2 =0.93).     

Experiences in controlling resistant pollen beetle by type I ether pyrethroid Trebon 30 EC in Germany

Oil seed rape (OSR) is an important crop in Germany with a long growing history. Due to the prevalence of various springtime pests an average of two pyrethroid applications are made. Pyrethroid resistance in Germany was found locally in 2003, widespread distribution of resistance was observed in 2006, a year of very strong pest pressure. Field trials in 2005 and the following years and laboratory analysis showed good control of resistant pollen beetle ( Meligethes aeneus ) by type I ether pyrethroid etofenprox, the active substance of Trebon 30 EC. Etofenprox differs significantly from common pyrethroids, which are usually ester pyrethroids including an α-amino-group and halogen substitutions. Trebon 30 EC has comparable efficacy on all important OSR pests but it has superior efficacy on resistant pollen beetle compared to common pyrethroids. To cope with pyrethroid resistance of pollen beetle, three different modes of action (MOA)s are postulated, which may be type I pyrethroids, neonicotinoids and organophosphates (OPs). Because type I pyrethroids, such as Trebon 30 EC, perform well at low temperatures, have a broad spectrum of activity, are considered as not to be toxic to bees, and give a high level of control of resistant pollen beetle, they will have an important role in the insecticide spraying regime of OSR.     

Biochemical and molecular diagnosis of insecticide resistance conferred by esterase, MACE, kdr and super-kdr based mechanisms in Italian strains of the peach potato aphid, Myzus persicae (Sulzer)

In this paper we analysed the basis of insecticide resistance in 59 Italian strains of the peach potato aphid Myzus persicae using both molecular and biochemical assays. Our data as a whole clearly indicate that most M. persicae strains (76.3%) have high or extremely high production of an esterase enzyme which sequester and detoxify insecticides with esteric group. Kdr genotypes conferring resistance towards pyrethoids are present in 57.7% of the analysed populations. Moreover, 26.5% of the kdr positive strains possess also the M918T mutation conferring super-kdr phenotype. Strains with modified AChE (MACE) are not so numerous (27.1%), although they can be found almost everywhere in Italy. Considering all the strains analysed, both MACE and kdr phenotypes are associated with high levels of esterase activity. In Central–Southern regions, kdr and MACE resistance mechanisms resulted in linkage disequilibrium. Bioassays performed in order to evaluate the efficacy of a pyrethroid insecticide against a strain possessing a F979S mutation within its para-type sodium channel gene suggests that this amino acid substitution could affect the sodium channel responsivity to pyrethroids.     

Frequencies of the M918I mutation in the sodium channel of the diamondback moth in China, Thailand and Japan and its association with pyrethroid resistance

In addition to the allele frequencies of the L1014F and T929I mutations which are involved in nerve-insensitive resistance to a pyrethroid, those of the M918I mutation were examined using field strains obtained in China, Thailand, and Japan during 2009-2011. Results show that the resistance allele frequencies at the L1014F site were 89-100%, 97-100% and 65-85%, respectively, for strains in China, Thailand, and Japan. The respective allele frequencies at the T929I site were 86-100%, 70-97% and 58-84% for Chinese, Thai, and Japanese strains. With low frequencies up to 27%, M918I was found in Japan and China, but not in Thailand. The strain homozygous for the M918I and L1014F mutations was established and its resistance level to a pyrethroid was examined. The strain lacks a portion of the sodium channel gene corresponding to the 3′ portion of exon 18a, intron 18, and the 5′ portion of exon 18b. Nevertheless, the strain showed a similar level of resistance to that which was homozygous for the T929I and L1014F mutations.