This project contains two objectives: 1) Control HLB by optimization of application of SA and its analogs. We are testing the control effect of SA and its analogs, e.g., ASM, Imidacloprid, DL-2-aminobutyric, 2,6-dichloro-isonicotinic acid, and 2,1,3 Benzothiadiazole via trunk injection in field trial. Oxytetracycline is used as a positive control, whereas water was used as a negative control. SA, Acibenzolar-S-methyl (ASM), benzo (1,2,3) thiadiazole-7-cabothionic acid S-methyl ester (BTH), and 2,6-dichloroisonicotinic acid (INA) have also been applied twice onto selected trees by foliar spray in November, 2015 during fall flush, arch 2016 during spring flush, and February 2017 during spring flush. In addition, three field trials for different compounds including SA are being conducted. Materials were applied once onto selected trees by foliar spray in September, 2016 during late summer-fall flush, were applied to selected trees by soil drench in September, 2016 during late summer-fall flush, in early March and June 2017. Trunk injection in August and September, 2016 during summer and late summer-fall flush. Trunk injection of SA showed significant control effect against HLB. The data for trunk injection has been collected and a manuscript has been submitted for publication. HLB disease severity,disease incidence surveys and Las titers were conducted before spray treatment in October, 2015 and at 6 months after the 1st application in April, 2016 and April 2017. SA analogs resulted in increased fruit yield in 2016, but not in 2017 probably due to hurricane damage and also slowed down the progress of Las titers compared to control. To compare the effect of suppressing SA hydroxylase, we also screened multiple SecA inhibitors which suppress the secretion of important virulence factors. Two effective SecA inhibitors have been tested in vitro. At least one SecA inhibitor has been shown to be specific against Las, but not E. coli. We are also investigating the possibility of modifying pathway of citrus to produce more SA in citrus using CRISPR. As experiment scheduled, SA, ASM, BTH and INA were applied to selected trees by foliar spray in March 2018 during spring flush. Admire, SA, ASM, BTH and INA were applied to selected trees by soil drench in March 2018 during spring flush. SA, ASM, BTH and INA were applied to selected trees by trunk injection from March to April 2018 during spring flush. HLB disease severity surveys and Las titer assays will conducted for treatments in April 2018 at 24 months after 1st application of soil drench or trunk injection and at 30 months after 1st application of foliar spray. 2) Control HLB using a combination of SA, SA analogs or SA hydroxylase inhibitors. The SA hydroxylase protein is being expressed in E.coli and purified. Several inhibitors identified using structure based design are being tested for their inhibitory effect against SA hydroxyalse. To further identify SA hydroxylase inhibitors or SA analogs that are not degraded by SA hydroxylase, we have expressed SA hydroxylase in tobacco and Arabidopsis. Overexpression of SA hydroxylase decreased HR induced by Pseudomonas spp, indicating that SA hydroxylase degrades SA. We have qualified SA with HPLC and conducted SAR related genes expression analysis. We have identified multiple SA analogs and tested whether they can be degraded by SA hydroxylase. 4 SahA inhibitors were trunk-injected during fall flush. Las titers and HLB disease severity of the treated trees are being tested periodically. One manuscript entitled: ‘Candidatus Liberibacter asiaticus’ Encodes a Functional Salicylic Acid (SA) Hydroxylase That Degrades SA to Suppress Plant Defenses” has been published by MPMI. We have completed the project. Application of plant defense inducers including SA have positive effect on HLB diseased trees at the early stage of HLB infection, but have no effect on HLB diseased trees at the late stage of infection. One manuscript entitled: “Developing citrus Huanglongbing management strategies based on the severity of symptoms in HLB-endemic citrus-producing regions” has been accepted for publication by Phytopathology.