Optimization of soil viability testing for Ceratocystis lukuohia

Gabriela Benito and Marc Hughes, Pacific Cooperative Studies Unit, University of Hawaii at Manoa, Hilo, HI, Gabriela Benito, Marc Hughes and Flint Hughes, Institute of Pacific Islands Forestry, United States Department of Agriculture-Forest Service, Hilo, HI, Lisa Keith, Daniel K. Inouye U.S Pacific Basin-Agricultural Research Center, United States Department of Agriculture- Agricultural Research Station, Hilo, HI, Bruce Mackey, Agricultural Research Service, West Regional Research Center, United States Department of Agriculture, Albany, CA

Background/Question/Methods
Metrosideros polymorpha (ʻŌhiʻa) is Hawaiʻi’s keystone native tree, providing habitat and resources for multitudes of native flora and fauna; its forests are integral to maintaining Hawaiʻi’s healthy watersheds. In 2014 a new fungal pathogen, Ceratocystis lukuohia, was discovered in Hawaiʻi and identified as causing rapid tree wilt of ʻŌhiʻa trees, ultimately resulting in their death. This disease is locally referred to as Rapid ʻŌhiʻa Death (ROD). Previous field soil baiting attempts for the selective isolation of Ceratocystis lukuohia have been problematic, producing mixed results due to rapid degradation of carrot baits by non-target contaminants (e.g., bacteria and non-Ceratocystis fungi). We developed new methods to optimize detection of viable Ceratocystis lukuohia in field soils that included adding antibiotics to carrot baits, air drying soils to reduce soil moisture, and mechanical separation by soil particulate size.
Results/Conclusions
Results showed that application of streptomycin and ampicillin antibiotics effectively delayed carrot degradation throughout the 30-day observation period. Air drying soils for approximately 25 hours further reduced carrot bait degradation, presumably through elimination of anaerobic soil bacteria populations. Lastly, mechanical sieving partitioned soils into aggregate sizes that expressed varying levels of viability; 2000-micron and 500-micron soil aggregate sizes proved best for detection of viable C. lukuohia. Results confirmed that, in combination, these methods effectively detect viable Ceratocystis inoculum in soils, adding an important new diagnostic tool to combat Ceratocystis spread. We plan to use these techniques to confidently test for presence of viable Ceratocystis inoculum in soils of high priority ROD management areas.