Development and evaluation of pupal color-based genetic sexing strains in Anastrepha obliqua (Diptera: Tephritidae)

Anastrepha obliqua, a neotropical pest widely distributed in the Americas, attacks mango and other tropical fruits. In Mexico, it is controlled through integrated pest management, using the Sterile Insect Technique (SIT) as a main component. The applicability of SIT is significantly improved with the use of genetic sexing strains (GSS) that allow the possibility to release exclusively sterile males, the primary component of the technique. This study reports the isolation and characterization of two pupal mutations: black pupae (bp) and sphere pupae (sp), allowing for the first time the development of a genetic sexing system based on pupal color in this species. Inheritance analyses from reciprocal crosses between wild-type and mutant individuals showed F₂ phenotypic segregation consistent with a recessive Mendelian inheritance pattern, and linkage analysis indicated that the bp and sp loci are in separate chromosomes. Using the bp mutation, two GSS were developed through gamma irradiation [T(Y;bp⁺)/bp-22](GSS-22) and ethyl methanesulfonate treatment [T(Y;bp⁺)/bp-354](GSS-354). Both GSS exhibited sex-specific pupal differentiation but displayed a high frequency of recombinants. Despite an initial reduction in biological fitness, GSS-22 demonstrated greater genetic stability and a lower frequency of recombinants than GSS-354. Discrepancies between cytogenetic and genomic data, particularly regarding the localization of the gene responsible for the black pupae phenotype, underscore the need to integrate polytene chromosome and genomic analyses to characterize these translocations and improve GSS stability precisely. These results represent a breakthrough in the creation of genetic tools for the management of A. obliqua control.