Six mosquito species in the Anopheles Hyrcanus group are known as vectors responsible for transmitting vivax malaria in South Korea. In this study, seasonal dynamics of Anopheles Hyrcanus group species and knockdown resistance (kdr) mutations in malaria-endemic sites, Paju and Gimpo, were monitored over two years. In August 2023, all six species were observed simultaneously in one of the Paju collecting sites, and kdr mutations were newly identified in all species except Anopheles kleini. Although Anopheles pullus was revealed as a relatively resistant species among five species populations without kdr mutation via bioassays, there were no critical differences in the voltage-sensitive sodium channel sequence. These findings suggest variability in pyrethroid resistance mechanisms among Anopheles Hyrcanus Group species.

To elucidate the mechanism of pyrethroid resistance in Helicoverpa armigera, the study explored three possibilities based on deltamethrin as a model pyrethroid; 1) the existence of mutations in the target site of deltamethrin, 2) the existence of variation at the genomic level between insecticide-susceptible and resistant strains, 3) differences in gene expression patterns between the strains. Based on these hypotheses, three levels of resistant strains and a susceptible strain as well as nine Korean field populations were used. As results, 1) any point mutations were not detected in sodium channel gene. 2) based on newly set Korean reference genome (GCA_026262555.1), approximately 3,369,837 and 1,032,689 variants (SNPs and Indels) were revealed from genome and ORFs, respectively. However, any specific variants were not found to be highly correlated with the level of insecticide resistance. 3) based on DEG analysis, some of detoxification enzyme genes were differently expressed particularly cytochrome P450 genes. Therefore, H. armigera possibly acquires deltamethrin resistance through a combination of actions, including over-expression of various detoxification enzymes such as CYP3 subfamilies and cuticular proteins.

Imported 313 cases of Dengue fever and 16 cases of Zika virus disease were reported in the ROK during 2016. Aedesalbopictus has been reported as a major vector mosquito for Dengue and Zika virus. Various class of pesticides havebeen used locally different pesticide class. To investigate the pyrethroid and organophosphate pesticide resistance, genomicDNA was used for Allele-specific PCR (AS-PCR) genotyping of the acethylcholine esterase to detect Gly119Ser mutationsand kdr gene to detect Phe1534Cys mutations. Detoxification enzyme activities were assessed using Ae. albopictus fromYeosu and Jeju. Activities of four detoxification enzymes eg., glutathione S-transferase (GST), Non specific esterases (α-naphthylacetate and β-naphthyl acetate), and cytochrome C oxidase were determined for each Ae. albopictus strain. In our studies,Ae. albopictus showed only susceptible sequences for AchE and kdr. Activity of cytochrome P-450 and non specific esterasewas higher in a field population than a laboratory strain, except for GST. This study might be helpful to understandthe insecticidal susceptibility and resistant for effective vector mosquito control program.

Aedes albopictus is an important vector for yellow fever, dengue fever, chikungunya fever and Zika virus. This mosquitohas been exposed to organophosphates and pyrethroids in Republic of Korea for a long time. Using the direct contactmortality bioassay, susceptibility strain and two field populations of Ae. albopictus from Busan, and Damyang. Insecticidalproperty were indicated to RR ratio values (Resistant ratio to strain of susceptibility) of LC50 (Midian Lethal concentration)to organophosphats and pyrethroids. Genomic DNA was used for Allele-specific PCR (AS-PCR) genotyping of the acethylcholineesterase to detect Gly119Ser mutations and kdr gene to detect Phe1534Cys mutations. Detoxification enzyme activitiesof Ae. albopictus from Busan and Damyang were assessed using microplate enzyme activity assays. Activities of fourdetoxification enzymes eg., glutathione S-transferase(GST), Non specific esterases (α-naphthyl acetate and β-naphthyl acetate),and cytochrome C oxidase were determined for each Ae. albopictus strain. This study might suggest that Ae. albopictuscontrol programs should be prepared for the management of organophosphate and pyrethroid insecticide resistance.

Anopheles sinensis is an important vector for Plasmodium vivax and thus has been targeted with pyrethroids in Republic of Korea. Using the direct contact mortality bioassay, two field populations of An. sinensis from Ganghwa-gun and Goyang-si were characterized for their resistance to pyrethroids with RR ratio values (Resistant ratio to imidacloprid) of 125.6 to 203.8 folds and 80.0 to 120 folds, respectively. Genomic DNA was used for Allele-specific PCR (AS-PCR) genotyping of the sodium channel genes to detect L1014S mutations. The homozygous susceptible Leu/Leu genotype in Ganghwa-gun and Goyang-si was 5.0% and 11.3% and the resistance genotypes were 95.0% and 88.7%, respectively. The homozygous Phe/Phe resistance genotype was the most prevalent as 35.4% in Ganghwa-gun and 44.0% in Goyang-si. Hence, this study suggests that malaria vector control programs should be prepared for the management of pyrethroid insecticide resistance.

Anopheles sinensis is an important vector for Plasmodium vivax and thus has been targeted with pyrethroids in Republic of Korea. Using the direct contact mortality bioassay, two field populations of An. sinensis from Paju-si and Yeoncheon-gun were characterized for their resistance to pyrethroids with RR ratio values (Resistant ratio to imidacloprid) of 96.8 to 167.2 folds and 34.2 to 98.4 folds, respectively. Genomic DNA was used for Allele-specific PCR (AS-PCR) genotyping of the sodium channel genes to detect L1014S mutations. The homozygous susceptible Leu/Leu genotype in Paju-si and Yeoncheon-gun was 3.3% and 37.1% and the resistance genotypes were 96.7% and 62.9%, respectively. The homozygous Phe/Phe resistance genotype was the most prevalent as 45.0% in Paju-si and 31.5% in Yeoncheon-gun. Hence, this study suggests that malaria vector control programs should be prepared for the management of pyrethroid insecticide resistance.

The cotton bollworm, Helicoverpa armigera (Hübner) is a serious agricultural pest which has evolved resistance against many chemical classes of insecticides. This species has evolved resistance to the synthetic pyrethroids across its native range and is becoming a truly global pest, after establishing in Brazil and having been recently recorded in North America. A chimeric cytochrome P450 gene, CYP337B3, has been shown to detoxify to fenvalerate and cypermethrin. The CYP337B3 gene has now been detected in different populations around the world, and is even found in South America. This gene is likely to have arisen independently in different geographic locations, probably through selection on pre-existing diversity, and there is ongoing movement of these alleles around the world. The alleles found in Brazil are those most commonly found in Asia, suggesting a potential origin for the incursion into the Americas. This information should be taken into account when devising control strategies for this invasive pest, both in its native range and in the Americas

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.

The green peach aphid (Myzus persicae) is a cosmopolitan pest of agricultural and horticultural crops and causes serious economic damages. M. persica has rapidly developed resistance to a wide variety of insecticides, including pyrethroids. Target site insensitivity mechanism mediated by two mutations (L1014F and M918T) on the para-type voltage-sensitive sodium channel (vssc) is mainly responsible for pyrethroid resistance. To predict the vssc resistance allele frequency, quantitative sequencing (QS) protocol was established. Frequency prediction equations generated from the plots of signal ratios and amplification critical time showed a high correlation coefficient (r2>0.993), indicating its high accuracy in prediction. QS results revealed that the kdr-type L1014F mutation is only present in Pyeongchang strain. No field strains of M. persicae possessed the super-kdr type M918T mutation. However, a novel M918L mutation was found by genotyping approach. The allele frequencies of M918L and L1014F were 0% to 53% in populations examined, and the level of M918L mutation frequency was closely related with pyrethroid resistance. Therefore, QS-based detection of M918L mutation frequency should faciltate the monitoring of pyrethroid resistance in the field.

Two point mutations (V419L and L925I) in the voltage-sensitive sodium channel (VSSC) α-subunit gene have been identified in deltamethrin-resistant bedbugs. To predict resistance allele frequencies of sodium channel mutations (V419L and L925I) in bedbugs at a population level, we developed quantitative sequencing (QS) protocol. The signal ratios between resistant and susceptible nucleotides were generated from sequencing chromatogram and plotted against the corresponding resistance allele frequencies. Linear regression coefficients of the plots were close to 1 (r2 = 0.9928 and 0.9998), suggesting that the signal ratios are reliable correlated with the resistance allele frequencies. To enable on-site monitoring of pyrethroid resistance in bed bugs, residual contact vial (RCV) bioassay method was established and used to determine median lethal concentration (LC50) values to deltamethrin for various bed bug strains. Resistance allele frequencies in these bedbug strains predicted by QS were correlated well with the RCV bioassay results, confirming the roles of two mutations in pyrethroid resistance. Taken together, employment of QS in conjunction with RCV bioassay should greatly facilitate resistance monitoring of bedbugs in the field.