Enhanced Bacterial Wilt Resistance in Potato Through Expression of Arabidopsis EFR and Introgression of Quantitative Resistance from Solanum commersonii

Abstract

Bacterial wilt (BW) caused by Ralstonia solanacearum is responsible for substantial losses in cultivated potato (Solanum tuberosum) crops worldwide. Resistance genes have been identified in wild species; however, introduction of these through classical breeding has achieved only partial resistance, which has been linked to poor agronomic performance. The Arabidopsis thaliana (At) pattern recognition receptor elongation factor-Tu (EF-Tu) receptor (EFR) recognizes the bacterial pathogen-associated molecular pattern EF-Tu (and its derived peptide elf18) to confer anti-bacterial immunity. Previous work has shown that transfer of AtEFR into tomato confers increased resistance to R. solanacearum. Here, we evaluated whether the transgenic expression of AtEFR would similarly increase BW resistance in a commercial potato line (INIA Iporá), as well as in a breeding potato line (09509.6) in which quantitative resistance has been introgressed from the wild potato relative Solanum commersonii. Resistance to R. solanacearum was evaluated by damaged root inoculation under controlled conditions. Both INIA Iporá and 09509.6 potato lines expressing AtEFR showed greater resistance to R. solanacearum, with no detectable bacteria in tubers evaluated by multiplex-PCR and plate counting. Notably, AtEFR expression and the introgression of quantitative resistance from S. commersonii had a significant additive effect in 09509.6-AtEFR lines. These results show that the combination of heterologous expression of AtEFR with quantitative resistance introgressed from wild relatives is a promising strategy to develop BW resistance in potato.

Keywords: EFR; Ralstonia solanacearum; Solanum commersonii; Solanum tuberosum; bacterial wilt; pattern recognition receptor; quantitative resistance.

FIGURE 1

AtEFR potato plants are phenotypically similar to non-transformed plants. Representative 90-day-old plants grown from tubers in greenhouse. (A) INIA Iporá. (B) clone 09509.6. No differences were observed in controlled conditions between wild-type and transformed plants at phenotypic level.

FIGURE 2

AtEFR expression in potato confers elf18 responsiveness. (A) AtEFR gene detection by PCR of transgenic potato lines. (B) Protein expression of AtEFR detection by Western blot. Upper panel anti-HA blot; lower panel CBB membrane staining as loading control. (C) ROS production triggered by 100 nM elf18 in INIA Iporá and clone 09509.6 leaf discs measured as RLU over a period of 60 min. Results are average ± standard error (n = 16). Experiments were performed twice with similar results.

FIGURE 3

Transgenic expression of AtEFR in potato confers BW resistance. (A) BW progress curves on INIA Iporá (left) and clone 09509.6 (right) after soil inoculation with R. solanacearum strain UY031. The mean of two experiments are represented in each point. (B) AUDPC values for the average wilting score ± SD as means of two independent experiments. Data were pooled across trials of repeated experiments because no significant effects involving trials were found in the analyses of variance. Transformed AtEFR potato plants of both genotypes responded statistically different when compared to wild-type controls, as assayed by ANOVA (p < 0.001). Tukey’s HSD test as means with different letters are significantly different (p < 0.05).

FIGURE 4

Occurrence of latent infections is reduced in AtEFR potato lines. (A) Latent infection in stems. Bars indicate percentage of plant survival after inoculation (28 d.p.i.). The proportion of positive replicates by PCR to R. solanacearum are denoted in dark gray, while negative for bacterial presence is denoted in light gray. Data samples correspond to averages of two independent experiments (n = 4). (B) Latent infection in tubers. Bars indicate percentage of plant survival (90 d.p.i.), while the proportions of positive replicates by PCR to R. solanacearum are denoted in dark gray and negative tubers in light gray.