Determination, mechanism and monitoring of knockdown resistance in permethrin-resistant human head lice, Pediculus humanus capitis

J. Marshall Clark 한국응용곤충학회 2009 Journal of Asia-Pacific Entomology Vol.12 No.1

Permethrin resistance has been reported worldwide and clinical failures to commercial pediculicides containing permethrin have likewise occurred. Permethrin resistance in head lice populations from the U.S. is widespread but is not yet uniform and the level of resistance is relatively low (∼4–8 fold). Permethrinresistant lice are cross-resistant to pyrethrins, PBO-synergized pyrethrins and to DDT. Nix®, when applied to human hair tufts following manufacturer's instructions, did not provide 100% control when assessed by the hair tuft bioassay in conjunction with the in vitro rearing system. Resistance to permethrin is due to knockdown resistance (kdr), which is the result of three point mutations within the α-subunit gene of the voltage-gated sodium channel that causes amino acid substitutions, leading to nerve insensitivity. A three-tiered resistance monitoring system has been established based on molecular resistance detection techniques. Quantitative sequencing (QS) has been developed to predict the kdr allele frequency in head lice at a population level. The speed, simplicity and accuracy of QS made it an ideal candidate for a routine primary resistance monitoring tool to screen a large number of louse populations as an alternative to conventional bioassay. As a secondary monitoring method, real-time PASA (rtPASA) has been devised for a more precise determination of low resistance allele frequencies. To obtain more detailed information on resistance allele zygosity, as well as allele frequency, serial invasive signal amplification reaction (SISAR) has been developed as an individual genotyping method. Our approach of using three tiers of molecular resistance detection should facilitate large-scale routine resistance monitoring of permethrin resistance in head lice using field-collected samples. Permethrin resistance has been reported worldwide and clinical failures to commercial pediculicides containing permethrin have likewise occurred. Permethrin resistance in head lice populations from the U.S. is widespread but is not yet uniform and the level of resistance is relatively low (∼4–8 fold). Permethrinresistant lice are cross-resistant to pyrethrins, PBO-synergized pyrethrins and to DDT. Nix®, when applied to human hair tufts following manufacturer's instructions, did not provide 100% control when assessed by the hair tuft bioassay in conjunction with the in vitro rearing system. Resistance to permethrin is due to knockdown resistance (kdr), which is the result of three point mutations within the α-subunit gene of the voltage-gated sodium channel that causes amino acid substitutions, leading to nerve insensitivity. A three-tiered resistance monitoring system has been established based on molecular resistance detection techniques. Quantitative sequencing (QS) has been developed to predict the kdr allele frequency in head lice at a population level. The speed, simplicity and accuracy of QS made it an ideal candidate for a routine primary resistance monitoring tool to screen a large number of louse populations as an alternative to conventional bioassay. As a secondary monitoring method, real-time PASA (rtPASA) has been devised for a more precise determination of low resistance allele frequencies. To obtain more detailed information on resistance allele zygosity, as well as allele frequency, serial invasive signal amplification reaction (SISAR) has been developed as an individual genotyping method. Our approach of using three tiers of molecular resistance detection should facilitate large-scale routine resistance monitoring of permethrin resistance in head lice using field-collected samples.

A point mutation in a glutamate-gated chloride channel confers abamectin resistance in the two-spotted spider mite, Tetranychus urticae Koch

Deok Ho Kwon,Kyong Sup Yoon,J. Marshall Clark,Si Hyeock Lee 한국응용곤충학회 2010 한국응용곤충학회 학술대회논문집 Vol.2010 No.05

The molecular mechanisms and genetics of abamectin resistance mediated by target site insensitivity in the two-spotted spider mite, Tetranychus urticae, were investigated by comparing two isogenic AbaS and AbaR strains. Cloning and sequencing of full-length cDNA fragments of GABA-gated chloride channel genes revealed no polymorphisms between the two strains. However, sequence comparison of the full-length cDNA fragment of a T. urticae glutamate-gated chloride channel gene (TuGluCl) identified a G323D point mutation as being tentatively related with abamectin resistance. In individual F2 progenies obtained by backcrossing, the G323D genotype was confirmed to correlate with abamectin resistance. Bioassays using progeny from reciprocal crossings revealed that the abamectin resistance trait due to TuGluCl insensitivity is incompletely recessive.

Genomics of Body/Head Lice and Its Application

Si Hyeock Lee,Ju Hyeon Kim,J. Marshall Clark,Barry B. Pittendrigh 한국응용곤충학회 2014 한국응용곤충학회 학술대회논문집 Vol.2014 No.04

Both the body louse (Pediculus humanus humanus) and the head louse (P. humanus capitis) are obligatory human ectoparasites. The body louse is a serious public health threat because it transmits a variety of human diseases whereas the head lice causes one of the most prevalent human infestations, pediculosis. Recent genome analysis revealed that both body and head lice have one of the smallest insect genomes (~108 Mb). Comparison of transcriptional profiles uncovered that almost the same number of genes was annotated both in the head louse (10,770 genes) and the body louse (10,771 genes) among 10,775 protein-coding genes predicted from the body louse genome. Despite their similar genetic background, body and head lice have several differences in their biological features, such as choice of habitat on human host, body size and vector competence. Annotation of major detoxification genes revealed that they are dramatically reduced in human lice compared to other insects except for the honey bee and that, despite the overall reduction in number, human lice retain at least a minimum repertoire of genes known to confer metabolic or toxicokinetic resistance to insecticides, suggesting their high potential for resistance development. Comparison of insecticide target site gene sequences and transcription levels of detoxification genes enabled the identification of toxicodynamic and metabolic factors of insecticide resistance and further allowed the development of molecular markers for resistance detection. Transcriptional profiling during tolerance was used to identify ivermectinmetabolizing detoxification genes, indicating that such an approach may allow proactive resistance management. Comparison of genomes and transcriptomes between body and head lice suggested that vector competence difference is not attributed to the difference in the composition of immune related genes but rather to their transcriptional regulation and/or not-yet-identified epigenetic factors.

Cross-strain comparison of cypermethrin-induced cytochrome P450 transcription under different induction conditions in diamondback moth

Baek, Ji Hyeong,Clark, J. Marshall,Lee, Si Hyeock Elsevier 2010 Pesticide biochemistry and physiology Vol.96 No.1

<P><B>Abstract</B></P><P>Synergism with piperonyl butoxide indicated that enhanced P450 activity is likely associated with pyrethroid resistance in an isogenic cypermethrin-resistant (CR) strain of diamondback moth (DBM), <I>Plutella xylostella</I> (L.). Eleven P450 genes, including 8 novel ones, were cloned from DBM, and their basal and cypermethrin-induced transcription levels were compared between insecticide-susceptible (Sus) and CR strains. One P450 gene was determined to be constitutively overtranscribed in CR strain. To determine the conditions resulting in maximum levels of P450 gene induction by cypermethrin, several factors, including routes of insecticide administration (topical vs. leaf dip), exposure amounts and durations of exposure, were examined. In general, the leaf dip method resulted in greater levels of induction in a wider array of P450 genes. The conditions of ‘low sub-lethal amounts and short exposures’ to cypermethrin were more efficient in P450 induction than those of ‘high amounts (e.g., LD<SUB>50</SUB> or LC<SUB>50</SUB>) and long exposures’. Cross-strain comparisons revealed that 8 of 11 P450 genes were induced (1.5- to 2.2-fold) in CR larvae whereas only a single P450 gene was induced in Sus larvae under optimal induction conditions. This finding demonstrates that selective P450 gene induction by cypermethrin in addition to constitutive overexpression can confer metabolic resistance in the CR strain. Current study also provides a framework for future induction experiments for the transcriptional profiling of insect detoxification genes in response to insecticide exposure.</P>

Estimation of Knockdown Resistance in Diamondback Moth Using Real-time PASA

Deok Ho Kwon,J. Marshall Clark,Si Hyeock Lee 한국농약과학회 2003 한국농약과학회 학술발표대회 논문집 Vol.- No.-

Transcriptome profiling of the diamondback moth treated with sublethal doses of five different insecticides and characterization of commonly responding genes

Yue Gao,Kyungmun Kim,Deok Ho Kwon,In Hong Jeong,J. Marshall Clark,Si Hyeock Lee 한국응용곤충학회 2017 한국응용곤충학회 학술대회논문집 Vol.2017 No.10

To identify genes that commonly respond to the treatment of different insecticides and are responsible for the toleranceenhancement, transcriptomic profiles of larvae treated with sublethal doses of the five insecticides were compared withthat of untreated control. A total of 117,181 transcripts with a mean length of 662 bp were generated by de novo assembly,of which 35,329 transcripts were annotated. Among them, 125, 143, 182, 215 and 149 transcripts were determined tobe up-regulated whereas 67, 45, 60, 60 and 38 genes were down-regulated following treatments with these five insecticides.The most notable examples of commonly responding over-transcribed genes were two cytochrome P450 genes and ninecuticular protein genes. In contrast, several genes composing the mitochondrial energy generation system were significantlydown-regulated in all treated larvae. Considering the distinct structure and mode of action of the five insecticides tested,the differentially expressed genes identified in this study appear to be involved in general chemical defense at the initialstage of intoxication. Their possible roles in the tolerance/resistance development were discussed.

Transcriptome profiling of the diamondback moth treated with sublethal doses of five different insecticides and characterization of commonly responding genes

Yue Gao,Kyungmun Kim,Deok Ho Kwon,In Hong Jeong,J. Marshall Clark,Si Hyeock Lee 한국농약과학회 2017 한국농약과학회 학술발표대회 논문집 Vol.2017 No.11

Pyrethroid Resistance Monitoring of the Common Bedbug, Cimex lectularius L., Based on Quantitative Sequencing and Residual Filter Contact Vial Bioassay

Keon-Mook Seong,Da-Young Lee,Kyong Sup Yoon,J. Marshall Clark,Si Hyeock Lee 한국농약과학회 2009 한국농약과학회 학술발표대회 논문집 Vol.2009 No.2

Molecular Mechanism and Detection of Pyrethroid Resistance in the Common Bed Bug, Cimex lectularius

Deok Ho Kwon,Keon Mook Seong,Kyong Sup Yoon,John Marshall Clark,Si Hyeock Lee 한국응용곤충학회 2014 한국응용곤충학회 학술대회논문집 Vol.2014 No.04

The common bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), is an ectoparasitic pest that feeds on humans as well as other mammals. We investigate that point mutations on the voltage-sensitive sodium channel are associated with the resistance to pyrethroids. Two point mutations (V419L and L925I) in the voltage-sensitive sodium channel (VSSC) α-subunit gene have been identified in deltamethrin-resistant bed bugs. L925I, located the intracellular loop between IIS4 and IIS5, has been previously found in a highly pyrethroid-resistant populations of whitefly. V419L, located in the IS6 transmembrane segment, is a novel mutation. To establish a population-based genotyping method as a molecular resistance monitoring tool, a quantitative sequencing (QS) protocol was developed. Frequency prediction equations were generated from the plots by linear regression, and the signal ratios were shown to highly correlate with resistance allele frequencies (r2 > 0.993). In addition to QS, the filter contact vial bioassay (FCVB) method was established and used to determine the baseline susceptibility and resistance of bed bugs to pyrethroids. A pyrethroid-resistant strain showed > 9375- and 6990-fold resistance to deltamethrin and λ-cyhalothrin, respectively. Resistance allele frequencies in different bed bug populations predicted by QS correlated well with the FCVB results, confirming the roles of the two mutations in pyrethroid resistance. Taken together, employment of QS in conjunction with FCVB method should greatly facilitate the detection and monitoring of pyrethroid resistant bed bugs in the field.

Identification and characterization of genes associated with insecticide tolerance induction in the western flower thrips, Frankliniella occidentalis

Yue Gao,Min Ju Kim,Ju Hyeon Kim,In Hong Jeong,J. Marshall Clark,Si Hyeock Lee 한국응용곤충학회 2020 한국응용곤충학회 학술대회논문집 Vol.2020 No.10