Kamonchat Prommarit, Ajaraporn Sriboonlert, Passorn Wonnapinij
Fireflies (Lampyridae) are bioluminescent beetles with broad ecological roles across temperate and tropical ecosystems, occupying diverse habitats including forests, wetlands, grasslands, mangroves, and riverine systems. The subfamily Luciolinae is primarily distributed across Asia and the Indo-Pacific. Phylogenetic relationships among three closely related Luciolinae genera - Medeopteryx, Pteroptyx, and Trisinuata - remain unresolved using mitochondrial genome data alone. This study used nuclear genome data to resolve relationships among these genera and identify a lighter-weight nuclear marker panel for expanding taxon sampling. Draft genomes were reconstructed for fifteen firefly species, eight from the focal genera, and analyzed with five published firefly genomes. Using BUSCO and OrthoFinder, 1,011 nuclear single-copy orthologs (SCOs) were identified for phylogenomic inference. Discordance between concatenation- and coalescence-based phylogenies indicated incomplete lineage sorting (ILS). The coalescence-based phylogeny recoveredPteroptyxas monophyletic and sister to a (Medeopteryx,Trisinuata) clade, with Trisinuata nested within a non-monophyletic Medeopteryx; however, quartet support at the base of Pteroptyx, particularly at Pt. valida, was low.Filtering for compositional homogeneity, clock-likeness, and species-tree concordance yielded 103 SCOs with a significantly higher proportion of parsimony-informative sites than non-selected loci, retaining the backbone topology with higher gene concordance support at scored clades, while ILS-driven discordance at Pt. valida persists - confirming that the reduced panel retains phylogenetic resolving power for future taxon sampling. These findings demonstrate a practical framework for using nuclear SCOs to resolve close phylogenetic relationships within Luciolinae. Future work should expand taxon sampling - especially forTrisinuata - alongside long-read assemblies, for a more robust phylogenomic framework.