Collection, isolation, in vitro culture, and laboratory transmission of Hirsutella eleutheratorum (Hypocreales: Ophiocordycipitaceae) from coffee berry borer on Hawai'i Island.

The insect pathogenic fungus Hirsutella eleutheratorum was first reported as a pathogen of coffee berry borer (CBB) Hypothenemus hampei in Colombia in 1993. A similar CBB pathogen identified as Hirsutella sp. was reported also from Colombia in 2007; attempts at isolation and in vitro culture of this fungus were unsuccessful. During 2016 and 2017 on the island of Hawai'i, extensive sampling of CBB populations was conducted in coffee fields treated with Beauveria bassiana-based biopesticides and in untreated fields. Among the samples collected from two high-elevation sites in the district of South Kona were rare findings of adult foundress CBB infected with a species of Hirsutella fitting the description of H. eleutheratorum. Prevalence of the pathogen was, in all cases, very low (<1%), having no significant impact on pest populations, even under conditions supporting epizootics of B. bassiana. The fungus was readily isolated from freshly-killed CBB and cultured on potato dextrose agar (PDA). Molecular characterization identified the fungus as a member of the Hirsutella citriformis clade, which includes species recently placed in the genus Ophiocordyceps. Adult CBB exposed to fungus-killed beetles or to PDA cultures of the fungus succumbed to infection within 10-14 days. Under high-humidity laboratory conditions, the fungus emerged from the killed host and produced long, conidia-bearing synnemata characteristic of the species. To our knowledge, this is the first record of H. eleutheratorum from CBB in Hawai'i and the first account of isolation, in vitro culture, genetic characterization, host-to-host transfer, and culture-to-host transfer of this fungal pathogen.