Antibiotic Treatment Reduces Wing Size and Alters Wing Shape in a Neotropical Drosophila (Diptera: Drosophilidae)

Antibiotics are widely used in studies of insect–microorganism interactions, particularly to investigate symbiotic associations. However, antibiotic exposure may also induce unexpected physiological and developmental effects. Tetracycline is often used to remove Wolbachia from different hosts, allowing comparisons between infected and uninfected lines. This treatment, however, may also affect the host’s microbiota, physiology, and gene expression. In this study, we evaluated the effects of tetracycline on wing size and shape in 339 males of Drosophila prosaltans in three different generations. An original isofemale line was divided into sublines exposed to four generations of antibiotic treatment, followed by recovery generations in which the microbiota was reestablished, including reinoculation with feces from untreated males. Geometric morphometric analyses revealed a significant reduction in wing size in all treated sublines, which persisted after microbiota recovery. All sublines showed changes in wing shape after four generations of antibiotic treatment, and two returned to the original shape after recovery. These results suggest that the effects of tetracycline can persist for several generations, likely due to changes in the gut microbiota and gene expression. Such long-lasting effects may have broader phenotypic consequences and should be considered when interpreting studies that use antibiotics in host–symbiont interaction research. Graphical Abstract Tetracycline treatment eliminated Wolbachia and reduced wing size while modifying wing shape, effects that were not fully reversed after microbiota recovery. Morphological responses varied among sublines, indicating that antibiotic effects persist even after bacterial recolonization.