1. Please state project objectives and what work was done this quarter to address them: Objective 1: Screen FANA antisense oligonucleotide targeting CLas for efficacy in a field trial.During this quarter, we conducted field trials to assess the efficacy of FANA antisense oligonucleotides (ASOs) targeting CLas in infected citrus trees. We designed and synthesized FANA ASOs complementary to two essential CLas genes: the CLas NAD-dependent DNA Ligase gene (LigA) and the CLas DNA B-Helicase gene. Additionally, a scramble sequence FANA ASO was used as a negative control. Treatments were applied to 10-year-old, CLas-infected ‘Hamlin’ trees in 1-acre plots, with each treatment replicated three times in a randomized complete block design. The FANAs were delivered via microinjection, following the dosages determined in previous greenhouse assays.We collected data from the third replicate of the experiment, which encompassed the fall of 2022 and the spring of 2023. In the fall of 2022, CLas infection declined significantly after seven days in trees treated with LigA-FANA and antibiotics. At the 45-days after treament, CLas infection was observed in FANA ASO and antibiotic treatments but not in the Control-FANA treated trees. After 90 days, the lowest CLas infections were observed in LigA-FANA treated trees and antibiotic-treated trees. Preliminary data from spring of 2023 indicate that CLas infection significantly declined in antibiotic-treated trees from 0 to 60 days following application. Objective 2: Evaluate FANA antisense oligonucleotide targeting CLas to reduce vector transmission.We performed acquisition and inoculation assays to evaluate the impact of FANA ASOs on CLas transmission by the Asian citrus psyllid (ACP), the vector of citrus greening disease. In the acquisition assay, ACP nymphs were allowed to feed on FANA-treated infected citrus trees. In the inoculation assay, ACP collected from treated trees were transferred to uninfected citrus seedlings for inoculation feeding, and the development of CLas infection in the plants was monitored.In the fall of 2022, a reduction in CLas acquisition by ACP adults and their offspring feeding on FANA ASO-treated trees was observed. The lowest CLas infection rates were found in ACP feeding on antibiotic-treated trees, followed by LigA-FANA treated trees. Similar trends were observed in the spring of 2023, with reduced CLas acquisition by ACP adults feeding on Helicase-B-FANA treated trees and antibiotic-treated trees. This experiment was replicated in spring 2023 and data will be reported in a future report. 2. Please state what work is anticipated for next quarter: Next quarter, we plan to continue field trials to further assess the efficacy of FANA ASOs targeting CLas. A fourth and final applications of treatments will be applied to trees during this quarter. We will focus on analyzing the long-term effects of the treatments and evaluating the sustainability of the observed reductions in CLas infection rates. We will collect additional data on CLas titer, ACP transmission, and plant infection rates, extending the analysis timeline. Moreover, we aim to perform statistical comparisons between treatments to determine significant differences and evaluate treatment efficacy. 3. Please state budget status (underspend or overspend, and why): Spending is currently on track. We requested to use salary savings on the project to purchase additional treatments and conduct two full years of treatments.