First Report of Verticillium Wilt of Faba Bean Caused by Verticillium dahliae in Spain.

Faba bean (Vicia faba L.) crops in eastern-central Spain are usually grown in rotation or double cropped with artichoke (Cynara cardunculus L. var. scolymus (L.) Fiori). In this region, artichoke is grown annually and is severely affected by Verticillium dahliae Kleb. (1). During February of 2007, wilt symptoms were observed at harvesting time on faba bean fields located in Castellón Province (eastern-central Spain). Symptoms consisted of leaf yellowing, wilting, and gradual death of the leaves while stems generally remained green except for severely affected plants. The vascular tissue in the stems showed a tan-to-light brown discoloration and plants were stunted. For isolation, crown and stem sections (10 cm long) were surface disinfected for 1 min in 1.5% NaOCl and washed twice with sterile distilled water. The sections were cut longitudinally and small pieces of discolored vascular tissue were plated onto potato dextrose agar (PDA) amended with streptomycin sulfate (0.5 g liter^-1). Plates were incubated at 25°C in the dark. V. dahliae was consistently isolated and colonies transferred to PDA were identified on the basis of the presence of microsclerotia and conidiophore morphology. Identity of monoconidial isolates 3H, 4H, 7H, and 8H was confirmed by specific multiplex nested-PCR assays using primers NDf/NDr in the first PCR round and INTND2f/INTND3r/MCR2B in the second round (2). PCR markers amplified with these primers were originally developed for the detection and vegetative compatibility group (VCG) identification of V. dahliae isolates infecting artichoke plants. Isolates 3H, 4H, 7H, and 8H amplified the 688-bp and the 964-bp markers indicating that they belong to VCG2B. Recent studies identified VCG2B as the prevalent group in the population of V. dahliae affecting artichoke in Castellón Province (3). Pathogenicity of two selected isolates, 3H and 7H, was determined on faba bean (cv. Muchamiel) and artichoke seedlings (cv. Madrigal) at the two-true-leaf stage. Seedlings were inoculated by watering the roots with 25 ml of a conidial suspension (10⁶ conidia ml^-1) harvested from 3-week-old cultures grown on PDA. Ten replicates (each one in individual pots) for each isolate and plant species were used, with an equal number of control plants. Plants were maintained in a greenhouse at 23 to 25°C. Within 1 month of inoculation, symptoms developed on all inoculated plants as severe stunting, leaf necrosis, and wilting. The fungus was reisolated from vascular tissues of the crown area and the stems of inoculated seedlings, completing Koch's postulates. Symptoms were not visible in the control seedlings and V. dahliae was not isolated from them. To our knowledge, this is the first report of V. dahliae infecting faba bean in Spain. Verticillium wilt had been previously reported on V. faba in Greece (4). Verticillium wilt of faba bean may bear importance in the epidemiology of the disease in artichoke as an alternative host for inoculum increase and survival of V. dahliae under field conditions. References: (1) M. Berbegal et al. Plant Dis. 91:1131, 2007. (2) M. Collado-Romero et al. Online publication. doi:10.1111/j.1365-3059.2008.01981.x. Plant Pathol., 2008. (3) R. M. Jiménez-Díaz et al. Phytopathology 96:288, 2006. (4) E. K. Ligoxigakis and D. J. Vakalounakis. Plant Pathol. 43:755, 1994.