Genome-wide characterization of hemocyanin gene family and adaptive evolution of heme oxygenase across four penaeid shrimp species

Penaeid shrimp aquaculture faces threats from diseases and stress, with hemocyanin plays vital roles in innate immunity and oxygen transport. This study systematically characterized the hemocyanin gene family in four Penaeid shrimp using comparative genomics and evolutionary analysis. OrthoFinder identified 18,916 gene families, with 7,957 shared across species. Phylogenetic analysis revealed Marsupenaeus japonicus diverged first (≈119 Mya), followed by Litopenaeus vannamei splitting from Fenneropenaeus chinensis and Penaeus monodon (≈68.5 Mya). Hemocyanin phylogenetic trees matched genome-wide trees, suggesting early duplication before speciation. Hemocyanin genes (11~18 per species) were more abundant than phenoloxidase genes (4~7 per species). A total of 586 genes in M. japonicus were under positive selection, including a truncated gene Heme oxygenase (HO) associated with structural divergence. The HO gene shows a conserved and ubiquitous expression pattern across four penaeid shrimp species, with high expression levels in immune- and respiratory-related tissues, implying its important roles in immune defense and stress tolerance. This study provides a comprehensive genomic and evolutionary characterization of the hemocyanin gene family in penaeid shrimp, shedding light on their molecular evolution and potential functional divergence, which lays a foundation for further exploring the roles of hemocyanins in immunity, stress resistance, and genetic improvement of cultured shrimp.