| Summary: | Apple tree canker infected by <i>Valsa mali</i> var. <i>mali</i> is a serious and widely distributed disease in China. <i>Saccharothrix yanglingensis</i> Hhs.015 is an endophytic actinomycete isolated from cucumber roots, and it has been proven that this strain is a promising biocontrol agent on apple tree canker in previous studies. The aim of this study was to elucidate the active ingredients in its metabolites. Two pentaene macrolides, WH01 and WH02, were isolated from strain Hhs.015, and their structures were elucidated based on the extensive spectroscopic analysis. WH01 and WH02 were identified as fungichromin and 1′-deoxyfungichromin, among which WH02 is a novel compound. These two compounds showed strong in vitro and in vivo antifungal activity against <i>V. mali</i>. By comparison of the structures of hyphae cells treated by pure compound and fermentation broth, it has been proven that pentaene macrolides are the main active ingredients in the metabolites of strain Hhs.015. This is the first report on the antifungal activity of fungichromin and its analogs on <i>V. mali</i>, and the 28-member pentaene macrolides were also firstly isolated from the genus of <i>Saccharothrix.</i>
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