Identification of Effectors From The Barley Powdery Mildew Pathogen, Blumeria graminis f. sp. hordei, by HIGS, VIGS, and T3S

Obligate biotrophic fungi must regulate host immunity to establish haustorial feeding structures. These pathogens secrete a variety of effector proteins believed to play essential roles in this process.  The barley powdery mildew fungus, Blumeria graminis f. sp. hordei, produces a large number of proteins specifically expressed in and predicted to be secreted from haustoria.  Fifty of these Blumeria Effector Candidates (BECs) were screened by Host Induced Gene Silencing (HIGS), and eight were identified that appear to contribute to infection.  These eight were further characterized by delivery via the bacterial type III secretion (T3S) system and by Barley stripe mosaic virus (BSMV) induced gene silencing.  We fused each of the coding sequences to the 5’ end of the avrBs2 effector gene from Xanthomonas campestris, which encodes the T3S signal.  When delivered into barley by a strain of X. campestris that otherwise elicits no visible plant response, one of the candidates, BEC1019, suppresses a defense-associated cell death elicited by another, co-inoculated, Xanthomonas species, X. oryzae.  In addition, BSMV-VIGS of BEC1019 resulted in much less sporulation and fungal growth.  Thus, BEC1019 may function to suppress defense and enhance virulence during the development of powdery mildew in barley.  BEC1019 homologs were detected in 57 other diverse fungi, including obligate plant pathogens, necrotrophic animal pathogens, and free-living non-pathogens.  T3S delivery of a BEC1019 ortholog from the human pathogen Candida albicans, also suppresses X. oryzae induced cell death in barley, suggesting that BEC1019 may represent a large family of conserved fungal virulence factors.