We have set up all of the experiments in the greenhouse and field needed to accomplish those objectives. We have also set up experiments in paralell with the field experiments. Given that citrate is the preferred carbon source for Liberibacter and that plants load citrate into phloem in order to acid mine insoluble phosphate from the soil, we expect foliar phosphate fertilization to reduce citrate in phloem so drastically, that it starves CLas.
We now know that foliarly-applied potassium phosphate will decrease phloem citrate levels in half after just one month of treatment compared to citrus saplings that were fed calcium phosphate to the roots. We are now analyzing samples to see whether citrate levels continue to decline after three months. We will keep this experiment going for the length of the project and will sample phloem citrate levels every three months.
The first greenhouse experiments on needed phosphate levels taught us the appropriate levels of potassium phosphate to spray on citrus trees in the field. In April 2019, a field trial commenced in a grove o 20-year-old infected trees near Lake Hamilton in Polk County. There were 10 replicates and four treatments in a randomized complete block design. We spray the trees with 0, 1x, 3x, and 9x the optimal level observed in the greenhouse every two months. Even at 1x, the plants are receiving enough P for flushing and fruit development. The plants are sprayed six times a year including after each flush. We sample these trees for phloem CLas levels every three months. The samples from the first nine months are now being analyzed. We expect to see CLas titer declines in the foliar phosphate-treated plants after one year of treatment.
A second field trial was established in August 2019 in the Immokalee area. That trial is the same design as that in Polk County. The idea was to have a trial on the ridge (Polk County) and flatwoods (Collier County) regions of citrus production. In both trials, the outcomes being measured are CLas titer in leaf midribs and leaf area index. Baseline samples were taken prior to the first spray and subsequent sampling will be done at 6, 12, and 18 months after the first spray. We have now sampled these trees for the first six months after treatment.
As we wait for field results, we are now testing the effect of a foliar potassium phosphate spray (compared to root-applied calcium phosphate) on CLas titer in graft-infected trees in the greenhouse. We are assaying those plants for CLas titer now and expect to have results in a week. This experiment will tell us whether foliar phosphate treatments can prevent CLas infection.
Given the speed with which foliar potassium phosphate can reduce organic hose plants acids levels in citrus phloem, we expect to see positive results in the field in reducing CLas titer in the first quarter of 2020. I am pleased to report that this project is working as planned so far. Foliar potassium phosphate does reduce citrate levels and the levels of other organic acids in phloem. As organic acids, particularly citrate, are the preferred carbon source for Liberibacter crescens, we expect this treatment to starve the pathogen.
We expect to observe CLas titer declines in these experiments over the next two months. If we do, a manuscript will be prepared very quickly in order to disseminate our results as quickly as possible. We will also ask CRDF for help in identifying more field locations for this work.
An extension article will follow shortly thereafter.
Our team (Triplett, Vincent, Killiny, and Wang) are working very well together and meet to discuss the project every two weeks.