Detection of Latent Infections of Ralstonia solanacearum Race 3 Biovar 2 in Geranium

Ralstonia solanacearum race 3 biovar 2 is a regulated quarantine pathogen that infects solanaceous hosts such as potato as well as geranium, where it causes either bacterial wilt (also known as Southern Wilt) or a symptomless latent infection. Geranium growers and government regulators need reliable detection methods to identify infected plant material before it is exported. We previously found that R. solanacearum-infected geranium plants can shed millions of bacteria in effluent water that flows from pots. We tested a nondestructive sampling method wherein effluent water from infected plants grown under commercial conditions was both dilution plated and filter concentrated for real-time polymerase chain reaction (PCR). Under field conditions in Guatemala, effluent shedding of infected geranium plants was highly variable. Comprehensive growth chamber studies confirmed that latently infected and mildly symptomatic geranium plants often but not invariably shed detectable numbers of bacteria in their effluent. At the peak of bacterial shedding, just under 90% of infected plants shed detectable bacteria whereas, at the lowest point, 44% shed detectable numbers of pathogen cells. Bacterial shedding peaked several weeks after inoculation regardless of whether plants were symptomatic or latently infected. Bacterial stem population sizes did not correlate with either effluent population sizes or disease index rating. Finally, we found that the effluent from geranium plants grown in volcanic rock scoria medium contains inhibitors that reduce the effectiveness of real-time PCR detection methods.