Biocontrol of strawberry gray mold caused by Botrytis cinerea with the termite associated Streptomyces sp. sdu1201 and actinomycin D

Abstract

Strawberry gray mold caused by Botrytis cinerea is one of the most severe diseases in pre- and post-harvest periods. Although fungicides have been an effective way to control this disease, they can cause serious "3R" problems (Resistance, Resurgence and Residue). In this study, Streptomyces sp. sdu1201 isolated from the hindgut of the fungus-growing termite Odontotermes formosanus revealed significant antifungal activity against B. cinerea. Four compounds (1-4) were isolated from Streptomyces sp. sdu1201 and further identified as actinomycins by the HRMS and 1D NMR data. Among them, actinomycin D had the strongest inhibitory activity against B. cinerea with the EC₅₀ value of 7.65 μg mL^-1. The control effect of actinomycin D on strawberry gray mold was also tested on fruits and leaves in vitro, and its control efficiency on leaves was 78.77% at 3 d. Moreover, actinomycin D can also inhibit the polarized growth of germ tubes of B. cinerea. Therefore, Streptomyces sp. sdu1201 and actinomycin D have great potential to gray mold as biocontrol agents.

Keywords: Streptomyces; actinomycin D; antifungal activity; biocontrol; gray mold.

Figure 1

Phylogenetic tree of strain Streptomyces sp. sdu1201 based on 16S rRNA sequences by maximum-likelihood method.

Figure 2

HPLC-MS data of crude extracts of Streptomyces sp. sdu1201 cultured in different media.

Figure 3

Antifungal activity of Streptomyces sp. sdu1201. (A) Antifungal activity of the fermentation broth of Streptomyces sp. sdu1201 against six pathogenic fungi. (B) The inhibitory rate of the cell culture and supernatant of Streptomyces sp. sdu1201 against B. cinerea. Vertical bars represent the standard errors of the means. Different letters above the bars indicate statistically significant differences at p < 0.05.

Figure 4

Chemical structures of compounds 1–4.

Figure 5

Complete genome and actinomycin D BGC of Streptomyces sp. sdu1201. (A) Circular map of genome. The seven circles (outer to inner) represent forward genome size, strand coding sequences, reverse-strand coding sequences, repetitive sequences, transfer RNA (blue) and ribosomal RNA (purple), GC content, and GC skew. (B) Comparison of actinomycin D BGC from Streptomyces sp. sdu1201 (middle) with reference actinomycin D BGC from S. anulatus (top) and S. costaricanus ZS0073 (bottom).

Figure 6

Bioassay of fermentation broth against B. cinerea on detached strawberry fruits. (A) Symptoms on strawberry fruits in different treatments. (B) Lesion area on strawberry fruits in B. cinerea and S1201 + B. cinerea treatment at 2 d and 3 d. (C) Control efficiency of fermentation broth against B. cinerea at 2 d and 3 d. Vertical bars represent the standard errors of the means. Different letters above the bars indicate statistically significant differences at p < 0.05.

Figure 7

Bioassay of compound 2 against B. cinerea on detached strawberry leaves. (A) Symptoms on strawberry leaves in different treatments. (B) Lesion area on strawberry leaves in B. cinerea and compound 2 + B. cinerea treatment from 1 d to 4 d. (C) Control efficiency of compound 2 against B. cinerea ranging from 1 d to 4 d. Vertical bars represent the standard errors of the means. Different letters above the bars indicate statistically significant differences at p < 0.05.

Figure 8

Changes of conidia and germ tubes of B. cinerea after treated with different concentrations of compound 2 in 24 h. (A) Images for morphological changes of conidia and germ tubes in different treatments Bar = 100 μm. (B) Inhibitory effect of compound 2 against germ tube elongation of B. cinerea. Columns with different letters represent significant difference at p < 0.05.