Molecular phylogenetics and species delimitation of recently recorded Spilomelinae moths from Saudi Arabia

Approximately 4,200 described species constitute the subfamily Spilomelinae (Lepidoptera: Crambidae), yet molecular phylogenetic relationships among Arabian Peninsula populations remain poorly resolved. The present research employs mitochondrial cytochrome c oxidase subunit I (COI) and nuclear internal transcribed spacer 2 (ITS2) markers to evaluate species boundaries and phylogenetic affinities of Spilomelinae moths recently documented from Saudi Arabia. Specimens were collected from four biogeographically distinct zones across the kingdom between January 2022 and October 2023. DNA was extracted from thoracic tissue of 48 pinned specimens representing eight putative species. Maximum likelihood and Bayesian inference analyses recovered well-supported clades corresponding to morphologically defined species, with mean interspecific COI divergence ranging from 2.1% to 11.8%. The Automatic Barcode Gap Discovery method confirmed species delimitation for all taxa examined. The ITS2 phylogeny was largely concordant with the COI topology, although reduced resolution was observed among closely related Spoladea species. Two genetically distinct lineages within Hymenia recurvalis suggest cryptic diversity warranting further investigation. Saudi Arabian populations showed closest phylogenetic affinity to East African and South Asian conspecifics, supporting a multi-directional colonization pattern across the Arabian landmass. These molecular findings complement existing morphological taxonomy and provide a framework for integrative species identification in the region.