Genome and Transcriptome Analyses Facilitate Genetic Control of Wohlfahrtia magnifica, a Myiasis-Causing Flesh Fly.

ABSTRACT

Myiasis caused by Wohlfahrtia magnifica is a widespread parasitic infestation in mammals. The infested host suffers from damage as the developing larvae feed on its tissues. For the control of myiasis infestation, genetic methods have been shown to be effective and promising as an alternative to insecticides. Combining genome, isoform sequencing (Iso-Seq), and RNA sequencing (RNA-seq) data, we isolated and characterized two sex-determination genes, W. magnifica transformer (Wmtra) and W. magnifica transformer2 (Wmtra2), whose orthologs in a number of insect pests have been utilized to develop genetic control approaches. Wmtra transcripts are sex-specifically spliced; only the female transcript encodes a full-length functional protein, while the male transcript encodes a truncated and non-functional polypeptide due to the presence of the male-specific exon containing multiple in-frame stop codons. The existence of five predicted TRA/TRA2 binding sites in the male-specific exon and the surrounding intron of Wmtra, as well as the presence of an RNA-recognition motif in WmTRA2 may suggest the auto-regulation of Wmtra by its own protein interacting with WmTRA2. This results in the skipping of the male-specific exon and translation of the full-length functional protein only in females. Our comparative study in dipteran species showed that both the WmTRA and WmTRA2 proteins exhibit a high degree of similarity to their orthologs in the myiasis-causing blow flies. Additionally, transcriptome profiling performed between adult females and adult males reported 657 upregulated and 365 downregulated genes. Functional analysis showed that among upregulated genes those related to meiosis and mitosis Gene Ontology (GO) terms were enriched, while, among downregulated genes, those related to muscle cell development and aerobic metabolic processes were enriched. Among the female-biased gene set, we detected five candidate genes, vasa (vas), nanos (nanos), bicoid (bcd), Bicaudal C (BicC), and innexin5 (inx5). The promoters of these genes may be able to upregulate Cas9 expression in the germline in Cas9-based homing gene drive systems as established in some flies and mosquitoes. The isolation and characterization of these genes is an important step toward the development of genetic control programs against W. magnifica infestation.