Pyrethroid Resistance In House Flies Research Articles

Resistance to pyrethroid insecticides in house flies, Musca domestica L., (Diptera: Muscidae) collected from urban areas in Punjab, Pakistan.

House flies are one of the major public health pests in urban settings. People usually use insecticides containing pyrethroids for the management of house flies; however, there is a lack of information on pyrethroid resistance in house flies from urban areas. In the present study, resistance to four pyrethroids (beta-cyfluthrin, deltamethrin, permethrin, transfluthrin) was assessed in house flies collected from urban areas of Punjab, Pakistan. Significant levels of resistance to all the pyrethroids were found in different strains of house flies. The resistance ratios (RRs) at the median lethal dose (LD50) level were in the range of 5.25- to 11.02-fold for beta-cyfluthrin, 7.22- to 19.31-fold for deltamethrin, 5.36- to 16.04-fold for permethrin, and 9.05- to 35.50-fold for transfluthrin. Pairwise comparison of the log LD50s revealed a highly significant correlation (p<0.01) between deltamethrin and permethrin, suggesting the possibility of a cross-resistance mechanism. The results revealed the occurrence of pyrethroid resistance in house flies from urban areas of Punjab. Regular resistance monitoring surveys and integrated approaches for the management of house flies are needed to retain the efficacy of these insecticides for a longer period of time.

Multiple cytochrome P450s overexpressed in pyrethroid resistant house flies (Musca domestica)

Cytochrome P450 monooxygenases-mediated detoxification is a major mechanism of pyrethroid resistance in house flies. Constitutive overexpression of P450 genes is the main cause for this type of insecticide resistance. In this study, three new P450 genes (CYP6A40, CYP6D8 and CYP6G4) over-expressed in a pyrethroid resistance strain (BJD) were identified. Developmental expression analysis showed that CYP6A40 and CYP6D8 were expressed in all life stages, while CYP6G4 was not transcribed in eggs or larvae. Permethrin resistance and expression level of cytochrome P450s in six house fly populations (the BJD strain and five field-derived house fly populations collected across China) were determined. Compared with the susceptible TJS strain, all the six populations exhibited different levels of resistance to permethrin. Piperonyl butoxide (PBO) reduced permethrin resistance level up to 109-fold, indicating that cytochrome P450-mediated detoxification is responsible for pyrethroid resistance in Chinese house flies. Significant overexpression of more than one of the eight P450 genes (CYP6A5v1, CYP6A5v2, CYP6A36, CYP6A40, CYP6D1, CYP6D3, CYP6D8 and CYP6G4) in the six pyrethroid resistant house fly populations was observed. Differential expression patterns of P450s over-represented in the resistance populations suggested evolutionary plasticity of P450-mediated pyrethroid resistance. CYP6G4 and CYP6D1, commonly over-expressed in resistant populations, were hypothesized to be the important P450s contributing to pyrethroid resistance of Chinese house flies.

Investigating the potential of selected natural compounds to increase the potency of pyrethrum against houseflies Muscadomestica (Diptera: Muscidae)

A study was undertaken to determine the efficacy of seven natural compounds compared with piperonyl butoxide (PBO) in synergising pyrethrum, with the intention of formulating an effective natural synergist with pyrethrum for use in the organic crop market. Discriminating dose bioassays showed PBO to be significantly more effective at synergising pyrethrum in houseflies than the seven natural compounds tested, causing 100% mortality in insecticide-susceptible WHO and resistant 381zb strains of housefly. The most effective natural synergists against WHO houseflies were dillapiole oil, grapefruit oil and parsley seed oil, with 59, 50 and 41% mortality respectively, compared with 18% mortality with unsynergised pyrethrum. Against 381zb houseflies, the most effective natural synergists were parsley seed oil and dillapiole oil. Esterase inhibition by the natural compounds and PBO in vitro showed no correlation with pyrethrum synergism in vivo, whereas the inhibition of oxidases in vitro more closely correlated with pyrethrum synergism in vivo. Dillapiole oil and parsley seed oil showed the greatest potential as pyrethrum synergists. PBO remained the most effective synergist, possibly owing to its surfactant properties, enhancing penetration of pyrethrins. The results suggest the involvement of oxidases in pyrethroid resistance in houseflies, with the efficacy of synergists showing a high correlation with inhibition of oxidases.

Genetics and mechanisms of permethrin resistance in the YPER strain of house fly

A strain of house fly was collected from the Third Yumenoshima Island in Japan and selected with the pyrethroid insecticide permethrin. Resistance to permethrin evolved to >18,400-fold in the selected (YPER) strain, and the mechanisms and genetics of resistance in this strain were examined. Permethrin resistance was decreased by pretreatment with piperonyl butoxide or 2-propynyl 2,3,6-trichlorophenyl ether, but not S, S, S-tributylphosphorotrithioate or diethylmaleate. The level of total cytochromes P450 was 2.7-fold increased, and the level of cytochrome b 5 was 1.5-fold increased, in YPER compared to the susceptible CS strain. These results suggest P450 monooxygenases, but not hydrolases or glutathione S-transferase, are a mechanism of resistance in the YPER strain. Analysis of the para-homologous sodium channel α-subunit gene in YPER indicates this strain has the super-kdr allele. Permethrin resistance in the YPER strain was inherited as a multigenic and incompletely recessive trait. A factorial analysis of resistance in the YPER strain indicated the relative contribution to resistance by the different autosomes was 2>3>5⩾1. Comparison of YPER with three other strains of house fly having high levels of permethrin resistance (LPR, NG98, and ALHF) indicates the genetic basis of this trait is variable between populations. This indicates there may be a greater difficulty for the development of diagnostic tools that could be used reliably over large areas to monitor pyrethroid resistance in house flies than was previously thought.

Genetics of Pyrethroid Resistance in a Strain (ALHF) of House Flies (Diptera: Muscidae)

Five house fly lines were derived from crosses of the pyrethroid-resistant ALHF (wildtype) and the susceptible aabys (bearing recessive morphological markers on each of five autosomes) strains. Each line was homozygous for one mutant-type marker from aabys. The level of resistance to permethrin was measured for each line to determine the genetic linkage of pyrethroid resistance in ALHF. Permethrin resistance in ALHF was 6600-fold compared with that in aabys. Resistance in flies bearing a mutant-type marker on autosome 4 was similar to that in ALHF. Flies with mutant-type markers on autosomes 1 and 2 had relatively lower resistance than ALHF; flies with mutant-type markers on autosomes 3 and 5 had much lower levels of resistance. These results demonstrated that factors on autosomes 3 and 5 play very important roles in pyrethroid resistance, whereas factors on autosomes 1 and 2 may have relatively small roles in resistance. Piperonyl butoxide (PBO) increased toxicity of permethrin in strains with mutant-type markers on autosomes 3 and 4 similar to that in ALHF. Slightly decreased synergism ratios in strains with autosomes 1 and 2 mutant-type markers compared with ALHF indicated that factors on autosomes 1 and 2 might make a small contribution in P450 monooxygenase-mediated resistance. However, when the autosome 5 mutant-type marker was present, PBO did not substantially decrease resistance, suggesting that the factor(s) on autosome 5 plays the most important role in P450 monooxygenase-mediated resistance. The resistance ratios of permethrin + PBO in strains with mutant-type markers on autosomes 1, 2, and 5 were significantly lower than those in ALHF, suggesting that factors on autosomes 1, 2, and 5 might be involved in pyrethroid resistance mechanisms other than P450-mediated detoxication. Injection did not change levels of resistance in the house flies tested, revealing that decreased rate of cuticular penetration (pen) probably does not play an important role in pyrethroid resistance in ALHF. The interaction and regulation of different mechanisms and/or factors involved in pyrethroid resistance in house flies are discussed.

EB35 ジョージア産イエバエのピレスロイド抵抗性とチトクロム P450 (CYP6D1) の過剰発現

EB35 ジョージア産イエバエのピレスロイド抵抗性とチトクロム P450 (CYP6D1) の過剰発現

Overexpression of Cytochrome P450 CYP6D1 Is Associated with Monooxygenase-Mediated Pyrethroid Resistance in House Flies from Georgia

CYP6D1 is a house fly cytochrome P450 that catalyzes metabolism of pyrethroid insecticides. Overexpression of CYP6D1 is responsible for monooxygenase-mediated pyrethroid resistance in the LPR (Learn pyrethroid resistant) house fly strain, which was originally collected in New York State. To determine whether CYP6D1 is involved in monooxygenase-mediated pyrethroid resistance in other populations, we obtained two pyrethroid-resistant strains from Georgia and investigated the relationship between piperonyl butoxide suppressible permethrin resistance and CYP6D1 gene expression. The Georgia strain, collected from a poultry facility in about 1983, showed no PBO suppressible permethrin resistance and CYP6D1 was not overexpressed. The NG98-u strain, collected from a poultry facility in Georgia State in 1998, showed PBO suppressible resistance and CYP6D1 mRNA was overexpressed. Furthermore, the level of resistance and CYP6D1 mRNA increased after only two generations of permethrin selection. Overall, the synergistic ratios were well correlated with the expression of CYP6D1. CYP6D1 protein detected by immunoblotting was well correlated with CYP6D1 mRNA expression. These results suggest that increased expression of CYP6D1 in the resistant flies collected from Georgia in 1998 is due to increased transcription of the gene (as has been identified in LPR) and that CYP6D1-mediated pyrethroid resistance is found in house flies from both New York and Georgia.

CDNA and deduced protein sequence of Cyp6D1: the putative gene for a cytochrome p450 responsible for pyrethroid resistance in house fly

A microsomal cytochrome P450 from the house fly (Musca domestica), termed P450 lpr, is involved in P450 monooxygenase-mediated pyrethroid resistance and is expressed at 8-fold higher levels in the insecticide resistant LPR strain compared to a susceptible strain. An internal cDNA sequence was amplified by polymerase chain reaction (PCR) using degenerate primers based on known P450 lpr, polypeptide sequences, and the remainder of the sequence was amplified by single side-specific PCR. A 1.8 kb cDNA sequence was obtained from 3 overlapping PCR products, with an open reading frame encoding a P450 protein of 516 residues ( M r 59,182). This gene has been designated CYP6D1 within the P450 gene superfamily. CYP6D1 exhibits most similarity (28.2–29.8% positional identity) to butterfly CYP6B1, house fly CYP6A1 and Drosophila CYP6A2, and also exhibits comparable similarity (24.7% identity) to rat CYP3AI. The deduced protein sequence contains a hydrophobic N-terminal region and conserved sequences thought to be involved in heme-binding and electron donor-protein interactions. Comparison of CYP6D1 with its four most similar proteins (CYP6B1, CYP6A1, CYP6A2 and CYP3A1) reveals the presence of extensive stretches of residues in an alignment row in 2 possible substrate-binding regions. Three introns of 74, 66 and 64 bp, having 5′-GT and AG-3′ ends, split the CYP6D1 coding region in genomic DNA. Results indicate that CYP6D1 is likely the P450 lpr gene.