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Deep sequencing of pyrethroid-resistant bed bugs reveals multiple mechanisms of resistance within a single population.

Adelman ZN, Kilcullen KA, Koganemaru R, Anderson MA, Anderson TD, Miller DM - PLoS ONE (2011)

Bottom Line: Using LD(50) bioassays, we determined that resistance ratios for Richmond strain bed bugs were ∼5200-fold to the insecticide deltamethrin.Following assembly, analysis of newly identified gene transcripts in both Harlan (susceptible) and Richmond (resistant) bed bugs revealed several candidate cytochrome P450 and carboxylesterase genes which were significantly over-expressed in the resistant strain, consistent with the idea of increased metabolic resistance.These data will accelerate efforts to understand the biochemical basis for insecticide resistance in bed bugs, and provide molecular markers to assist in the surveillance of metabolic resistance.

View Article: PubMed Central - PubMed

Affiliation: Fralin Life Science Institute and Department of Entomology, Virginia Tech, Blacksburg, Virginia, United States of America. zachadel@vt.edu

ABSTRACT
A frightening resurgence of bed bug infestations has occurred over the last 10 years in the U.S. and current chemical methods have been inadequate for controlling this pest due to widespread insecticide resistance. Little is known about the mechanisms of resistance present in U.S. bed bug populations, making it extremely difficult to develop intelligent strategies for their control. We have identified bed bugs collected in Richmond, VA which exhibit both kdr-type (L925I) and metabolic resistance to pyrethroid insecticides. Using LD(50) bioassays, we determined that resistance ratios for Richmond strain bed bugs were ∼5200-fold to the insecticide deltamethrin. To identify metabolic genes potentially involved in the detoxification of pyrethroids, we performed deep-sequencing of the adult bed bug transcriptome, obtaining more than 2.5 million reads on the 454 titanium platform. Following assembly, analysis of newly identified gene transcripts in both Harlan (susceptible) and Richmond (resistant) bed bugs revealed several candidate cytochrome P450 and carboxylesterase genes which were significantly over-expressed in the resistant strain, consistent with the idea of increased metabolic resistance. These data will accelerate efforts to understand the biochemical basis for insecticide resistance in bed bugs, and provide molecular markers to assist in the surveillance of metabolic resistance.

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Phylogenetic comparison of bed bug GST genes with other insects.Glutathione S-transferase (GST) genes of the bed bug, Cimex lectularius (pink circle), compared with published sequences from the pea aphid Acyrthosiphon pisum (gray diamond), honey bee, Apis mellifera (green circle), and the malaria mosquito, Anopheles gambiae (blue square). Predicted protein sequences were aligned using ClustalW; tree construction was performed using the Neighbor-joining method in MEGA 4.0. All branches with less than 50% bootstrap support were collapsed, support greater than 80% is indicated (*). Classification is based on Ranson et al [27].
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pone-0026228-g003: Phylogenetic comparison of bed bug GST genes with other insects.Glutathione S-transferase (GST) genes of the bed bug, Cimex lectularius (pink circle), compared with published sequences from the pea aphid Acyrthosiphon pisum (gray diamond), honey bee, Apis mellifera (green circle), and the malaria mosquito, Anopheles gambiae (blue square). Predicted protein sequences were aligned using ClustalW; tree construction was performed using the Neighbor-joining method in MEGA 4.0. All branches with less than 50% bootstrap support were collapsed, support greater than 80% is indicated (*). Classification is based on Ranson et al [27].

Mentions: Ten contigs encoding full-length glutathione S-transferase ORFs were identified and aligned with those of other insect species (Fig. 3). Surprisingly, only a single member of the delta-epsilon class was identified. This contrasts with 22 members from the malaria mosquito and 8 from pea aphid. Similar to both the pea aphid and honey bee [19], [20], bed bugs have undergone a lineage-specific expansion of the sigma class GSTs, the significance of which remains unknown (Fig. 3). We also identified at least 24 contigs corresponding to bed bug carboxylesterase (CE) genes. Lower coverage of these genes combined with much higher variability from other insect sequences made assignment to specific clades more challenging. A preliminary phylogeny is shown in Fig. S1, but we stress that higher coverage sequencing, as well as additional sequences from more closely related hemipterans will be required to properly establish the relationship of these sequences with those of other insects. Of note are at least seven bed bug CE contigs whose ORF is predicted to encode a signal peptide [21], and thus may be important in the metabolism of pyrethroid insecticides.


Deep sequencing of pyrethroid-resistant bed bugs reveals multiple mechanisms of resistance within a single population.

Adelman ZN, Kilcullen KA, Koganemaru R, Anderson MA, Anderson TD, Miller DM - PLoS ONE (2011)

Phylogenetic comparison of bed bug GST genes with other insects.Glutathione S-transferase (GST) genes of the bed bug, Cimex lectularius (pink circle), compared with published sequences from the pea aphid Acyrthosiphon pisum (gray diamond), honey bee, Apis mellifera (green circle), and the malaria mosquito, Anopheles gambiae (blue square). Predicted protein sequences were aligned using ClustalW; tree construction was performed using the Neighbor-joining method in MEGA 4.0. All branches with less than 50% bootstrap support were collapsed, support greater than 80% is indicated (*). Classification is based on Ranson et al [27].
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198472&req=5

pone-0026228-g003: Phylogenetic comparison of bed bug GST genes with other insects.Glutathione S-transferase (GST) genes of the bed bug, Cimex lectularius (pink circle), compared with published sequences from the pea aphid Acyrthosiphon pisum (gray diamond), honey bee, Apis mellifera (green circle), and the malaria mosquito, Anopheles gambiae (blue square). Predicted protein sequences were aligned using ClustalW; tree construction was performed using the Neighbor-joining method in MEGA 4.0. All branches with less than 50% bootstrap support were collapsed, support greater than 80% is indicated (*). Classification is based on Ranson et al [27].
Mentions: Ten contigs encoding full-length glutathione S-transferase ORFs were identified and aligned with those of other insect species (Fig. 3). Surprisingly, only a single member of the delta-epsilon class was identified. This contrasts with 22 members from the malaria mosquito and 8 from pea aphid. Similar to both the pea aphid and honey bee [19], [20], bed bugs have undergone a lineage-specific expansion of the sigma class GSTs, the significance of which remains unknown (Fig. 3). We also identified at least 24 contigs corresponding to bed bug carboxylesterase (CE) genes. Lower coverage of these genes combined with much higher variability from other insect sequences made assignment to specific clades more challenging. A preliminary phylogeny is shown in Fig. S1, but we stress that higher coverage sequencing, as well as additional sequences from more closely related hemipterans will be required to properly establish the relationship of these sequences with those of other insects. Of note are at least seven bed bug CE contigs whose ORF is predicted to encode a signal peptide [21], and thus may be important in the metabolism of pyrethroid insecticides.

Bottom Line: Using LD(50) bioassays, we determined that resistance ratios for Richmond strain bed bugs were ∼5200-fold to the insecticide deltamethrin.Following assembly, analysis of newly identified gene transcripts in both Harlan (susceptible) and Richmond (resistant) bed bugs revealed several candidate cytochrome P450 and carboxylesterase genes which were significantly over-expressed in the resistant strain, consistent with the idea of increased metabolic resistance.These data will accelerate efforts to understand the biochemical basis for insecticide resistance in bed bugs, and provide molecular markers to assist in the surveillance of metabolic resistance.

View Article: PubMed Central - PubMed

Affiliation: Fralin Life Science Institute and Department of Entomology, Virginia Tech, Blacksburg, Virginia, United States of America. zachadel@vt.edu

ABSTRACT
A frightening resurgence of bed bug infestations has occurred over the last 10 years in the U.S. and current chemical methods have been inadequate for controlling this pest due to widespread insecticide resistance. Little is known about the mechanisms of resistance present in U.S. bed bug populations, making it extremely difficult to develop intelligent strategies for their control. We have identified bed bugs collected in Richmond, VA which exhibit both kdr-type (L925I) and metabolic resistance to pyrethroid insecticides. Using LD(50) bioassays, we determined that resistance ratios for Richmond strain bed bugs were ∼5200-fold to the insecticide deltamethrin. To identify metabolic genes potentially involved in the detoxification of pyrethroids, we performed deep-sequencing of the adult bed bug transcriptome, obtaining more than 2.5 million reads on the 454 titanium platform. Following assembly, analysis of newly identified gene transcripts in both Harlan (susceptible) and Richmond (resistant) bed bugs revealed several candidate cytochrome P450 and carboxylesterase genes which were significantly over-expressed in the resistant strain, consistent with the idea of increased metabolic resistance. These data will accelerate efforts to understand the biochemical basis for insecticide resistance in bed bugs, and provide molecular markers to assist in the surveillance of metabolic resistance.

Show MeSH
Related in: MedlinePlus