We investigated the capacity of CLP-producing Pseudomonas strains and their crude CLP extracts to control rice blast caused by Magnaporthe oryzae, both in a direct manner and via induced systemic resistance (ISR). In planta biocontrol assays showed that lokisin-, white line inducing principle (WLIP)-, entolysin- and N3-producing strains successfully induced resistance to M. oryzae VT5M1. Furthermore, crude extracts of lokisin, WLIP and entolysin gave similar ISR results when tested in planta. In contrast, a xantholysin-producing strain and crude extracts of N3, xantholysin and orfamide did not induce resistance against the rice blast disease. The role of WLIP in triggering ISR was further confirmed by using WLIP-deficient mutants. The severity of rice blast disease was significantly reduced when M. oryzae spores were pre-treated with crude extracts of N3, lokisin, WLIP, entolysin or orfamide prior to inoculation. In vitromicroscopic assays further revealed the capacity of crude N3, lokisin, WLIP, entolysin, xantholysin and orfamide to significantly inhibit appressoria formation by M. oryzae. In addition, the lokisin and WLIP biosynthetic gene clusters in the producing strains are described. In short, our study demonstrates the biological activity of structurally diverse CLPs in the control of the rice blast disease caused by M. oryzae. Furthermore, we provide insight into the non-ribosomal peptide synthetase genes encoding the WLIP and lokisin biosynthetic machineries.
Induce systemic resistance capacity of crude CLP extracts against rice blast caused by M. oryzae. (A) ISR potential of crude WLIP, lokisin, entolysin, orfamide, and xantholysin to M. oryzae on rice. For each treatment, potting soil was mixed with 25 μg/ml of each CLP prior to experimental set-up. Subsequently, roots of rice seedlings were dipped in 25 μg/ml solution of each CLP for 10 min before planting. Data of one experiment made up of three biological replicates, consisting of 12 plants each are shown. The experiment was repeated with comparable results. Different letters indicate statistically significant differences among treatments (ANOVA followed by a Tukey’s test: n = 36; α = 0.05). (B) Representative disease symptoms on second youngest leaf at 6 days post-inoculation with M. oryzae. Scale bar represents 1 cm. DC, disease control; HC, healthy control; Lok, lokisin; Ent, entolysin; Xan, xantholysin; Orf, orfamide; BTH, S-methyl 1,2,3-benzothiadiazole-7-carbothioate.