1a. Objectives (from AD-416)
Culture ZC Liberibacter bacterium and confirm its pathogenicity
1b. Approach (from AD-416)
The initial goal is to establish an in planta culture of Liberibacter. For in vitro culture, standard procedure for cultivation of fastidious prokaryotes will be followed. Liber A medium will be used as a starting point for cultivation. Growth of Liberibacter will be monitored by standard microbiological methods and by PCR detection. If Liberibacter is detected by PCR, various cultivation conditions and media compositions will be optimized for bacterial growth. Bacteria will be triple-cloned to obtain pure cultures. Transmission electron microscopy will be used for morphological characterization. Bacteriophages will also be explored for their relationships to pathogenicity.
3. Progress Report
This Reimbursable Agreement supports Objective 1.A of the parent project. The research goal was to study the biology of a newly found invasive bacterium, “Ca. L. solanacearum”, associated with potato zebra chip disease (ZCD). Beginning in August, 2010, potato plants with typical ZCD symptoms were found in fields in Southern California. Symptoms on affected plants included stunting, chlorosis, swollen internodes, proliferation of axillary buds and aerial tubers, browning of the vascular system in below-ground portions of stems, and leaf scorching. Using symptomatic potato shoots as scions, grafted tomato plants appeared to be able to host “Ca. L. solanacearum” as evidenced by polymerase chain reaction (PCR) detection of the bacterium. No potato scions were able to develop normal growth on grafted tomato. Instead, grafted potato shoots survived for only 1-3 weeks. Tomato plants grafted with ZCD scions developed marginal purpling of leaf margins(necrosis in a severe case) and stunting of new growth. “Ca. L. solanacearum” also was detected by PCR in non-symptomatic leaves. Overall, the graft-inoculated tomato plants did not succumb to “Ca. L. solanacearum” infection. These results suggest that the bacterium associated with ZCD can be maintained in tomato as an experimental host.