Genome-Wide Sequencing and an Open Reading Frame Analysis of Dichlorodiphenyltrichloroethane (DDT) Susceptible (91-C) and Resistant (91-R) Drosophila melanogaster Laboratory Populations

Abstract

The Drosophila melanogaster 91-R and 91-C strains are of common origin, however, 91-R has been intensely selected for dichlorodiphenyltrichloroethane (DDT) resistance over six decades while 91-C has been maintained as the non-selected control strain. These fly strains represent a unique genetic resource to understand the accumulation and fixation of mutations under laboratory conditions over decades of pesticide selection. Considerable research has been done to investigate the differential expression of genes associated with the highly DDT resistant strain 91-R, however, with the advent of whole genome sequencing we can now begin to develop an in depth understanding of the genomic changes associated with this intense decades-long xenobiotic selection pressure. Here we present the first whole genome sequencing analysis of the 91-R and 91-C fly strains to identify genome-wide structural changes within the open reading frames. Between-strain changes in allele frequencies revealed a higher percent of new alleles going to fixation for the 91-R strain, as compared to 91-C (P<0.0001). These results suggest that resistance to DDT in the 91-R laboratory strain could potentially be due primarily to new mutations, as well as being polygenic rather than the result of a few major mutations, two hypotheses that remain to be tested.

Comparative CYP‐omic analysis between the DDT‐susceptible and ‐resistant Drosophila melanogaster strains 91‐C and 91‐R

“…P450s in strains 91-C and 91-R As previously reported by Schmidt et al 32 and Harrop et al, 61 Cyp6g1 was tandemly duplicated in the D. melanogaster genome and the duplication event of Cyp6g1 is responsible for overexpression and increased resistance to DDT. We used the mapping data of whole-genome sequencing generated by Steele et al 48 to determine the CNV for differentially expressed P450s in 91-C and 91-R. Among 12 differentially expressed P450s, high variation in sequencing coverage between 91-C and 91-R was found in Cyp6g1 and Cyp6g2 (Table S3), supporting the theory that there may be CNV for these two P450s.…”

“…and Harrop et al ., Cyp6g1 was tandemly duplicated in the D. melanogaster genome and the duplication event of Cyp6g1 is responsible for overexpression and increased resistance to DDT. We used the mapping data of whole‐genome sequencing generated by Steele et al . to determine the CNV for differentially expressed P450s in 91‐C and 91‐R .…”

“…Illumina genome sequencing data sets (SRA accession number SRP041176) from 91-C and 91-R were generated in order to determine the presence of known or unique allelic variants putatively affecting the evolution of DDT resistance. 48 To identify functional polymorphisms in the cis-regulatory ARE and Nrf2 transcription factor binding sites that regulate the transcription of P450s with upstream AREs, sequences were searched for within approximately 300-bp upstream gene regions obtained from FlyBase.org. Genome data sets of 91-C and 91-R were mapped to the upstream regulatory regions of significantly up-and down-regulated P450s using the 'Map Reads to Reference' tool from CLC genomic workbench (Qiagen, Valencia, CA, USA).…”

“…[44][45][46] These two strains were derived from a known common origin, but subsequent differences in the DDT selection schemes have provided a valuable resource for comparative analysis within the context of a model for studying xenobiotic resistance mechanisms on a genomic-wide scale. Indeed, these two strains formed the basis of a comparative genomic study that identified genome regions with significant variance in nucleotide fixation putatively attributable to the effects of differential selection, which included one region encoding Cyp4g1 47,48 and ATP-binding cassette transporters (ABC transporters) open reading frames. 47,49 Previously, Brandt et al showed that the P450 inhibitor piperonyl butoxide (PBO) drastically reduces (but does not eliminate) DDT resistance in D. melanogaster, providing further evidence that differential expression of P450 genes is a component of the DDT resistance phenotype.…”

Selective Sweep Analysis in the Genomes of the 91-R and 91-C Drosophila melanogaster Strains Reveals Few of the ‘Usual Suspects’ in Dichlorodiphenyltrichloroethane (DDT) Resistance

“…Dr. Ranjan Ganguly of the University of Tennessee-Knoxville provided the DDT resistant and susceptible Drosophila strains, respectively 91-R and 91-C [ 85 ]. For detailed description of fly line maintenance, re-sequencing, and data filtering, please see Steele et al [ 86 ].…”