Functional roles of nematodes associated with the walnut twig beetle and eastern black walnut in the inland northwest

Beetles (Scolytinae) form intimate associations with a taxonomically and functionally diverse suite of nematodes that are phytopathogens, fungal feeders, and entomoparasites. Despite their ubiquity, the ecological significance of nematodes in the lifecycles of economically important bark and ambrosia beetle species (Curculionidae: Scolytinae) and associated plant diseases remains largely unexplored. Thousand cankers disease (TCD) is caused by the walnut twig beetle (WTB, Pityophthorus juglandis Blackman) and the fungus Geosmithia morbida (Kolařík, Freeland, Utley & Tisserat; Ascoymycota: Hypocreales) and causes foliar senescence, progressive crown dieback, and mortality in black walnut (Juglans nigra L.) throughout western North America. In this study, nematodes recovered from P. juglandis and J. nigra in Idaho (ID) and Washington (WA) were identified morphologically and by constructing multilocus phylogenies to infer taxonomic relationships to taxa for which molecular data were available. We conducted assays to determine the extent to which nematodes feed and reproduce on G. morbida and other fungi commonly found in galleries of P. juglandis. Inoculation experiments were conducted to determine the effect of nematodes on the area of subdermal necrotic lesions (cankers) caused by G. morbida in branches of mature J. nigra and stems of seedlings. The phoretic nematode Bursaphlenhus juglandis (Ryss, Parker, Alvarez-Ortega, Nadeler & Subbotin) was frequently found under elytra of WTB in all locations, and a free-living nematode (Panagrolaimus sp.) was also widespread and found in the bark of mature trees. Both B. juglandis and Panagrolaimus sp. reduced the size of cankers caused by G. morbida in seedlings and branches of mature trees, respectively. However, these species may play opposite roles as disease synergists and antagonists based on the observation that exudates and/or microbiota associated with Panagrolaimus sp., but not B. juglandis destroyed G. morbida colonies in culture. Furthermore, B. juglandis contributed to foliar symptoms in seedlings inoculated with G. morbida. An entomoparasitic nematode (Aphelenchoididae), most closely resembling an Ektaphelenchus sp., was also found in the haemocoel of WTB. Infection rates were positively related to beetle population sizes as inferred from emergence rates. Ditylenchus sp. was also found in incubated walnut wood in WA and Rhabtidolaimus sp. was phoretic on P. juglandis and found in incubated walnut wood in WA and ID. The community of nematodes in J. nigra in WA and ID differed substantially from what has been observed associated with J. nigra in its native range.