First Report of Verticillium Wilt Caused by Verticillium dahliae on Fava Bean in the United States

Fava bean, Vicia faba L., broad bean, is eaten in many cuisines, and is planted worldwide as a rotation crop for erosion control and soil improvement. In April 2014, fava bean plants were collected from a seed field in Santa Clara County, CA. Clusters of stunted and declining plants were reported before flowering with further decline after seed set. Plants showed symptoms of chlorosis, wilting, and death, with dark streaks in the vascular tissue of the roots. Ten percent of the plants were symptomatic, the field yielding 40% of the seed expected. Pieces (4 mm3) of streaked vascular tissue were excised from the roots and stems, surface sterilized in 0.6% NaOCl for 2 min, and placed onto one-half-strength acidified potato dextrose agar (APDA). Fungal colonies were subcultured onto NP-10 (Kabir et al. 2004) when verticillate conidiophores were observed on APDA. Colonies on NP-10 consisted of fine, hyaline hyphae with verticillate conidiophores producing hyaline conidia, measuring from 3.2 to 5.0 × 2.1 to 3.5 μm (4.0 × 2.8 μm average), in slimy masses. Microsclerotia formed after a week in culture, causing the center of the colony to turn a blackish brown. Morphological characteristics were consistent with those of Verticillium dahliae Kleb. (Hawksworth and Talboys 1970). The internal transcribed spacer region (ITS) of rDNA was amplified from one isolate using ITS1/ITS4 primers as described by White et al. (1990), and the amplicon sequenced (GenBank accession no. KX025142). BLAST analysis of the 521-bp amplicon showed 100% identity with a sequence of V. dahliae from cosmos in Italy (GQ130129). Pathogenicity of the California fava bean isolate of V. dahliae was determined by inoculating the clipped roots of 13-day-old seedlings (four to six leaves) of V. faba ‘Broad Windsor Fava’ with a spore suspension (106 spores/ml) for 15 min. Spores were harvested by flooding 2-week-old cultures of V. dahliae grown on APDA with sterile distilled water. Two isolates were tested on groups of five plants each. Roots of five healthy plants were clipped and dipped in water as the control treatment. Plants were then planted in a commercial potting mix in 10-cm-diameter pots and kept in a growth chamber at 20°C with a 12-h photoperiod. After 21 days, all inoculated plants were chlorotic and wilted with black streaks in the roots. Root and stem isolations were performed, and V. dahliae grew from vascular tissue from stem and roots of all inoculated plants on NP-10 media. The experiment was repeated with 30-day-old plants with similar results. No symptoms developed on control plants, and Verticillium spp. did not grow from isolated root or stem pieces from the noninoculated plants in either experiment. On the basis of morphological and ITS sequence information, the fungus was identified as V. dahliae. V. dahliae is an economically important pathogen with a wide host range worldwide including fava in Spain (Berbegal and Armengol 2009). The affected field in California had a history of tomato and tomatillo seed crops, as well as vegetables for consumption. To our knowledge, this is the first report of Verticillium wilt on fava bean in the United States. © The American Phytopathological Society.