Control of fire blight (Erwinia amylovora) on apple trees with trunk-injected plant resistance inducers and antibiotics and assessment of induction of pathogenesis-related protein genes

Abstract

Management of fire blight is complicated by limitations on use of antibiotics in agriculture, antibiotic resistance development, and limited efficacy of alternative control agents. Even though successful in control, preventive antibiotic sprays also affect non-target bacteria, aiding the selection for resistance which could ultimately be transferred to the pathogen Erwinia amylovora. Trunk injection is a target-precise pesticide delivery method that utilizes tree xylem to distribute injected compounds. Trunk injection could decrease antibiotic usage in the open environment and increase the effectiveness of compounds in fire blight control. In field experiments, after 1-2 apple tree injections of either streptomycin, potassium phosphites (PH), or acibenzolar-S-methyl (ASM), significant reduction of blossom and shoot blight symptoms was observed compared to water injected control trees. Overall disease suppression with streptomycin was lower than typically observed following spray applications to flowers. Trunk injection of oxytetracycline resulted in excellent control of shoot blight severity, suggesting that injection is a superior delivery method for this antibiotic. Injection of both ASM and PH resulted in the significant induction of PR-1, PR-2, and PR-8 protein genes in apple leaves indicating induction of systemic acquired resistance (SAR) under field conditions. The time separating SAR induction and fire blight symptom suppression indicated that various defensive compounds within the SAR response were synthesized and accumulated in the canopy. ASM and PH suppressed fire blight even after cessation of induced gene expression. With the development of injectable formulations and optimization of doses and injection schedules, the injection of protective compounds could serve as an effective option for fire blight control.

Keywords: PR genes; SAR induction; acibenzolar-S-methyl; antibiotics; apple disease control; fire blight; potassium phosphites; trunk injection.

FIGURE 1

Control of blossom and shoot blight incidence in 2012 (A,B) and in 2013 (C,D) after 1–2 trunk injections of ‘Gala’ apple trees with protective compounds. WIC, water injected control; ASM, acibenzolar-S-methyl; PH, potassium salts of phosphorous acid (potassium phosphites); SS, streptomycin sulfate; NINIC, non-injected non-inoculated control. (A,C) Blossom blight incidence means across the three time points within one treatment followed by different letters are significantly different (t-test, 2012: p < 0.1, 2013: p < 0.05). (B) Shoot blight incidence means between treatments within one time point followed by different letters are significantly different (t-test, p < 0.1). (D) Shoot blight incidence means across the three time points within one treatment followed by different letters are significantly different (t-test, p < 0.05). Error bars represent SEM.

FIGURE 2

Shoot blight control in 2012 (A) and 2013 (B) after single injection of ‘Jonathan’ apple trees with oxytetracycline per each year. WIC, water injected control, OTC, oxytetracycline hydrochloride. Shoot blight severity means between treatments within one time point followed by different letters are significantly different (2012: Tukey’s HSD test, 2013: t-test, p < 0.05). Error bars represent SEM.

FIGURE 3

Relative expression of PR-1, PR-2, and PR-8 genes in ‘Gala’ apple leaves tested by qRT-PCR. Samples were collected in 2012 (A) and 2013 (B,C) following trunk injection/s of different compounds. PR, pathogenesis related genes; ASM, acibenzolar-S-methyl; PH, potassium salts of phosphorous acid (potassium phosphites); WIC, water injected control; NINIC, non-injected non-inoculated control. Mean gene expression followed by an asterisk (*) is significant relative to WIC or NINIC and normalized to actin gene (Pair Wise Fixed Reallocation Randomization test, α = 0.05). PR gene expression in (C) is shown in relation to NINIC since it was not significant in relation to WIC. Error bars represent SEM.