Regulation of Sclerotinia sclerotiorum growth by the mycoviral replicase-associated protein

Hypovirulent mycoviruses offer a promising potential for developing sustainable bio-fungicides. However, the mechanisms through which they reduce fungal aggressiveness remain unclear, particularly regarding the viral factors involved and their modes of action. In this work, we investigated the hypovirulence mechanism by which Sclerotinia sclerotiorum hypovirulent-associated DNA virus 1 (SsHADV1) impaired the necrotrophic plant fungi Sclerotinia sclerotiorum. We first identified the replication-associated protein (REP) of SsHADV1 as the key factor of hypovirulence. We demonstrated that the transient expression in plants of the viral REP or its ectopic application directly restricts fungal growth and reduces oxalic acid production, which is an essential factor for fungal pathogenicity.  This anti-fungal function of REP does not involve the suppression of RNA silencing but seems to be dependent of its ATPase domain. To establish its biological application, we engineered Bacillus subtilis as an anti-fungal delivery platform.  Remarkably, soil pre-treatment with bacteria secreting REP significantly controlled basal stem rot of sunflower plants by Sclerotinia sclerotiorum.  Furthermore, it enhanced fungal susceptibility to sublethal concentration of commercial fungicide. This study introduces a novel source of anti-fungal agent and their mode of function with application possibility.