Molecular characterization of the interaction between Helianthus annuus and Verticillium dahliae

The soil-borne fungus Verticillium dahliae Klebahn is the primary causal agent of Verticillium wilt in sunflower. The long viability of the resting structures, which can survive in the soil for more than 10-15 years, makes it more difficult to control this disease. To date, only limited information is available regarding the interaction between V. dahliae and sunflower, although such information is critical for a better understanding of plant defense mechanisms against this pathogen and for developing an effective strategy to manage Verticillium wilt sunflower. In this study, two highly aggressive (Vd1396-9 and Vd1398-21) and two weakly aggressive (Vs06-07 and Vs06-14) isolates of V. dahliae were used to inoculate susceptible (IS8048) and moderately resistant (IS6111) sunflower hybrids. A fusion protein of VdNEP (V. dahliae necrosis and ethylene-inducing protein) gene was produced, purified, and used as another treatment. The disease severity and symptoms caused by four Verticillium isolates and VdNEP were compared and induced plant defense responses were investigated in order to study the function of VdNEP involved in the sunflower- V. dahliae interaction. Our results indicated that VdNEP caused typical symptoms of Verticillium wilt in sunflower, including chlorosis, necrosis and vascular discoloration. The disease severity caused by VdNEP was similar to that caused by Verticillium isolate Vs06-14, which indicated that VdNEP not only acted as an elicitor but also as a pathogenicity factor involved in the disease development of Verticillium wilt. Furthermore, VdNEP induced the hypersensitive response, oxidative burst and the accumulation of fluorescent compounds in sunflower. Pathogenesis-related genes (Ha-PR-3, and Ha-PR-5), two defensin genes (Ha-PDF and Ha-CUA1) and genes encoding Ha-ACO, Ha-CHOX, Ha-GST and Ha-SCO were up-regulated by VdNEP, which indicated that multiple signaling pathways were involved. Two genes (Ha-PAL and Ha-NML1) were slightly inhibited after infiltration with VdNEP, which suggested that sunflower resistance against Verticillium wilt might be SA-independent. Interestingly, VdNEP enhanced the root fresh and dry weight in the moderately resistant sunflower hybrid but not in the susceptible one, indicating that it may play other roles in the interaction between sunflower and V. dahilae such as promoting plant growth.