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Female to male sex reversal was achieved in an emerging agricultural insect pest,  Drosophila suzukii , by creating a temperature-sensitive point mutation in the sex-determination gene,  transformer-2  ( tra-2 ), using CRISPR/Cas9 (clustered regularly interspaced palindromic repeats/CRISPR-associated) homology-directed repair gene-editing.  Ds-tra-2    ts2   mutants developed as normal fertile XX and XY adults at permissive temperatures below 20 °C, but at higher restrictive temperatures (26 to 29 °C) chromosomal XX females developed as sterile intersexuals with a predominant male phenotype, while XY males developed with normal morphology, but were sterile. The temperature-dependent function of the Ds-TRA-2 ts2  protein was also evident by the up- and down-regulation of female-specific  Ds-Yolk protein 1  ( Ds-Yp1 ) gene expression by temperature shifts during adulthood. This study confirmed the temperature-dependent function of a gene-edited mutation and provides a new method for the more general creation of conditional mutations for functional genomic analysis in insects, and other organisms. Furthermore, it provides a temperature-dependent system for creating sterile male populations useful for enhancing the efficacy of biologically-based programs, such as the sterile insect technique (SIT), to control  D. suzukii  and other insect pest species of agricultural and medical importance.

Temperature-dependent sex-reversal by a transformer-2 gene-edited mutation in the spotted wing drosophila, Drosophila suzukii