Development of the First Cytochrome Oxidase I Barcode and Evidence for a Single Haplotype Associated with the Recent United States Invasion of the Pasture Mealybug Heliococcus summervillei (Pseudococcidae, Hemiptera)

Heliococcus summervillei is an emerging invasive mealybug that causes severe dieback in grasses in pastures and turfgrass landscapes. It is widespread in Australia and has recently been detected across the Caribbean, Mexico, and the United States. Accurate identification of mealybugs is challenging due to cryptic morphology, overlapping diagnostic characters, and limited taxonomic expertise and literature, which makes molecular tools essential for regulatory diagnostics and management. We developed the first Cytochrome Oxidase I (COI) barcode for H. summervillei and used it to examine mitochondrial variation across available populations. COI sequences reveal approximately a 10.2% mitochondrial split between the Type A and Type B variants. Phylogenetic, haplotype network, and genetic distance analyses show that all invasive range populations share one haplotype associated with a recent invasion in the United States, Australia, Pakistan, and the Caribbean, whereas the Barbados lineage contains two closely related haplotypes that represent a historically stable mitochondrial variant. Together, these results establish the first COI reference library for H. summervillei, clarify mitochondrial lineage structure, and provide a practical barcode tool that enables rapid identification of invasive populations and supports timely regulatory and pest management responses. Recognizing mitochondrial variants also establishes a framework for resolving lineage-specific biological and management traits and strengthens reconstruction of introduction pathways central to regulatory decision-making and limiting further spread.