Shatters, Robert G

Abstract Delivery of therapeutic biomolecules into plant vascular tissues remains a challenge in management of vector-borne plant pathogens. The symbiont concept uses reprogrammed Agrobacterium tumefaciens galls (called symbionts) to produce biomolecules while remaining connected to host vasculature. We evaluated whether symbionts expressing antimicrobial peptides (AMPs) suppress ‘ Candidatus Liberibacter solanacearum’ (CLso), the causal agent of potato zebra chip disease. Symbionts were engineered to express a Streptococcus mutans bacteriocin associated with bacterial membrane disruption (Blp-Sm), or an AMP isolated from finger lime and associated with resistance to citrus greening disease (MaSAMP). Effects of AMP-producing symbionts on CLso titers, infection incidence, pathogen movement, and disease symptoms were evaluated in tomato and potato. In tomato, neither AMP significantly reduced CLso titers or infection incidence. However, in potato, AMP-producing symbionts reduced CLso accumulation and movement from CLso-inoculated source shoots into non-inoculated sink shoots connected through underground tubers. Blp-Sm produced the strongest reduction in CLso accumulation and infection incidence in sink tissues. In separate assays where symbionts were established directly on potato seed tubers, MaSAMP significantly reduced CLso titers in stems and tubers and reduced zebra chip symptoms in tubers, despite no reduction of CLso titers in terminal leaves. These findings demonstrate that AMP-producing symbionts suppress vascular pathogen accumulation and movement within plants and highlight the symbiont concept as a potential platform for managing diseases caused by vascular-restricted pathogens. Further, they show the potato-CLso system is a promising infection model to both refine and improve symbiont technology, and to test additional AMPs for potency against related pathogens.

AbstractCandidatus Liberibacter asiaticus (CLas) bacterium has been strongly implicated as the causative agent of huanglongbing (HLB), or citrus greening, which is currently the most devastating citrus disease worldwide. HLB is transmitted by the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), in a persistent manner. We used quantitative-polymerase chain reaction (PCR) to detect CLas in dissected organs of individual D. citri adults infected with HLB in the laboratory or collected from field-infected citrus trees in South Florida. The proportion of infected (CLas-positive) dissected organs was 47–70% for the salivary glands, 72–80% for the alimentary canal, and 79–97.5% for the rest of the insect body. Statistical analysis indicated that, in both field- and laboratory-infected D. citri, the proportion of infected salivary glands was significantly lower than that of other parts in the insect body. With field-collected psyllids, the relative copy number of CLas genomes, compared with psyllid genomic DNA in each sample, was significantly higher in both the salivary gland and alimentary canal compared with that in the rest of the insect body for both males and females. These results provide the first PCR confirmation of CLas in the alimentary canal and salivary glands of D. citri and strongly suggest that the salivary glands constitute an important transmission barrier to CLas in the psyllid vector. Our results also suggest that CLas may replicate or accumulate in both the alimentary canal and salivary glands of D. citri.