BackgroundPuberty is characterized by significant physical, hormonal, and psychological changes, which may be especially challenging for individuals with autism spectrum disorder (ASD). Although the etiology of ASD remains uncertain, studies suggest imbalances in hormones, such as testosterone, may modulate the autism phenotype. While differences in fetal and postnatal testosterone have been reported, there is limited literature regarding testosterone variations during adolescence in ASD. We investigated morning salivary testosterone levels in youth with ASD and typical development (TD) to explore hypothesized differences, expecting elevated hormonal levels in ASD compared to TD.MethodsYouth with ASD (n = 140) and TD (n = 104), ages 10 to 13 years, were enrolled as part of a longitudinal study on pubertal development. Pubertal stage was determined by gold standard physical examination, and salivary testosterone was collected in the morning immediately upon waking and 30 min after waking and averaged across 3 days. Diagnostic (ASD/TD) and sex (male/female) differences, as well as interactions with age and puberty, were examined using robust linear mixed effect models.ResultsYouth with ASD showed significantly elevated testosterone concentrations compared to same-age TD peers. After the inclusion of natural cubic splines to account for nonlinearity in age, a significant age-by-sex interaction emerged with distinct developmental slopes for males and females. At younger ages, females had higher testosterone, until about 11.5 years of age, when levels began to plateau, while male testosterone concentrations continued to rapidly increase and surpass females. As expected, more advanced pubertal development was associated with elevated testosterone. In contrast, no significant effect of parent-reported social communication symptoms was observed.LimitationsLimitations include an unequal sex distribution, non-representative sample (e.g., cognition and race/ethnicity), and inability to examine afternoon/evening testosterone due to detection limits.ConclusionsTestosterone may play a unique role in the presentation of ASD, especially during periods of dynamic hormonal changes including puberty. Inherent developmental (age, puberty) and sex-based (male, female) factors play a more prominent role in changes in testosterone levels during adolescence. Even so, future research is warranted to determine the differential expression and impact of exposure to excess testosterone during the pubertal transition for youth with ASD.