Sex-specific variation in basal innate immune gene and protein expression across mucosal and immune tissues of adult rainbow trout (Oncorhynchus mykiss)

Rainbow trout (Oncorhynchus mykiss) is a key aquaculture species in many countries, but the role of biological sex in shaping fish immune responses remains understudied. Characterizing sex-based immune variation is essential for optimizing disease management, vaccination strategies, and selective breeding programs. This study investigates the baseline innate immunity in sexually mature male and female rainbow trout (n=23/sex) by examining both transcript and protein expression in immune relevant tissues, including head kidney, spleen, foregut, midgut, and hindgut. RT-qPCR was used to quantify the expression of key innate immune genes and hormone receptors, while ELISA measured cytokines and immunoglobulins (IgT and IgM) in head kidney and spleen. Results revealed statistically significant sex- and tissue-specific differences at the transcript level, with males exhibiting elevated expression of interferons (ifn-1 and ifn-γ) inflammatory and regulatory cytokines (il1β, il10, tgf-β), and interferon-stimulated genes (pkr) particularly in gut-mucosal tissues. Androgen receptor transcripts (adr-a and adr-b) were also significantly higher in males, whereas estrogen receptor expression showed a complex, tissues- and isoform-specific pattern. In contrast, protein-level differences were limited with only IgT showing higher expression in the female head kidney. These findings demonstrate that sex is a significant determinant of baseline innate immunity in rainbow trout with male-biased transcript level responses and limited protein-level regulation, which highlights the need to consider sex as a biological variable in fish immunology and aquaculture applications, especially when facilities use only one sex for production.