Prevalence of novel RNA viruses: Pheidole megacephala viruses (PmegV) in the invasive big-headed ant, Pheidole megacephala on Oʻahu island, Hawaiʻi and Successful RNA virus transmission using alginate hydrogels as a delivery system for Pheidole megacephala virus 3 (Picornaviridae: Picornavirales) to target invasive ants, Pheidole megacephala, and resulting mortality.

Abstract

Invasive ants pose a significant threat to global biodiversity, particularly in ecologically sensitive island ecosystems such as Hawaiʻi. Pheidole megacephala (Fabricius), the African big-headed ant, is one of the world’s worst invasive species and has become dominant in disturbed environments, where it disrupts food webs, supports pest outbreaks, and alters soil ecosystems. Conventional control methods offer limited long-term success and pose risks to non-target species, highlighting the need for sustainable, targeted alternatives. Biological control using RNA viruses represents a promising solution, with Polycipiviridae viruses demonstrating host specificity and pathogenic potential. In Hawaiʻi, where P. megacephala dominates urban and disturbed habitats in the absence of natural enemies, three novel positive-sense single-stranded RNA viruses, Pmeg V1, V2, and V3 were recently discovered, opening new avenues for biological control. This study first conducted an island-wide survey of P. megacephala colonies across Oʻahu (n=25), revealing that 56% of colonies harbored at least one virus, with Pmeg V2 was most prevalent (52%), followed by Pmeg V1 (36%) and Pmeg V3 (12%). Due to its low natural prevalence, Pmeg V3 was selected for experimental delivery to assess its infectivity and effects on colony health. We next evaluated the efficacy of an alginate hydrogels system for introducing Pmeg V3 to ten uninfected P. megacephala colonies. Reverse transcription-polymerase chain reaction (RT-PCR) confirmed successful viral transmission and tagged RT-PCR verified replication within worker ants. While Pmeg V3 infections reached queens and brood by Day 17, replication was limited to workers. Kaplan-Meier survival analyses showed significantly reduced survival in four of ten treated colonies. These findings support the potential of alginate hydrogels as an effective viral delivery system and establish foundational evidence for developing virus-based biological control strategies against invasive P. megacephala populations.