In September 2011, a Phytophthora sp. was isolated from wasabi (Wasabia japonica Matsum.) grown commercially in hydroponic culture in a large production facility in southwest Michigan. Approximately 20% of the plants were affected, resulting in serious losses for the grower. Plants exhibited severe wilting and root and crown rot, with soft water-soaked lesions on the crown and dark lesions on the roots. Small pieces of root tissue with dark lesions were excised and plated onto potato dextrose agar and unclarified V8 agar plates amended with 25 ppm of benomyl, 100 ppm of ampicillin, 30 ppm of rifampicin, and 100 ppm of pentachloronitrobenzene. Isolates of a Phytophthora sp. were recovered from root tissue. Isolates produced sporangia abundantly in culture. Sporangia averaged 48 μm long × 34 μm wide and were ellipsoid to ovoid, occasionally obpyriform, and were nonpapillate and noncaducous. Distinct hyphal swellings were noted and chlamydospores were observed rarely in culture. The isolate used for inoculations did not produce oospores alone in culture but was able to produce oospores when paired with an A1 culture of P. capsici and incubated in the dark. Oospores were not observed when the isolate was paired with an A2 culture of P. capsici. No growth was observed at 35°C, and the isolate was identified as Phytophthora cryptogea based on morphological and physiological traits. Pathogen identity was further confirmed using PCR primers specific to P. cryptogea (1). In addition, a BLAST search was conducted using the nucleotide database collection in GenBank comparing our isolate against Phytophthora spp., with 99% sequence similarity to P. cryptogea in two sequenced genes, beta tubulin and cytochrome c oxidase 1 (2). Sequences for the isolate were deposited in the GenBank database under accession numbers JX041520 and JX041521. To fulfill Koch's postulates, six small, potted wasabi seedlings were inoculated by placing 3 g of 1-month-old infested millet (100 g of millet, 72 ml of distilled water, 0.08 g of asparagine, and 10 7-mm diameter V8 agar plugs with actively growing P. cryptogea) onto the soil surface of each pot under coconut coir mulch. Plants were watered heavily after soil infestation and as needed thereafter. Three control plants were inoculated with sterile millet seed. The experiment was repeated once. Wilting was observed within 5 and 7 days, respectively, in the first and second experiment. All six inoculated plants were severely wilted within 25 and 56 days, respectively, except for a single plant in the second experiment that never wilted. Root and crown rot was observed on wilted plants and dark lesions could be observed on root tissue. P. cryptogea was recovered from five of the six plants inoculated in each experiment. None of the control plants in either experiment displayed symptoms of wilting, and the pathogen was not recovered from these plants when pieces of root tissue were excised and plated onto amended V8 agar. To our knowledge, this is the first report of P. cryptogea causing crown and root rot of wasabi.
References: (1) D. Minerdi et al. Eur. J. Plant Pathol. 122:227, 2008. (2) L. M. Quesada-Ocampo et al. Phytopathology 101:1061, 2011.