In the present study, 77 strains of endophytic actinomycetes isolated from cabbage were screened in order to assess their biocontrol potential against Alternaria brassicicola on cabbage seedlings. In the first and second screening trials, cabbage seedlings pretreated with mycelial suspensions of each isolate were spray-inoculated with A. brassicicola. Strain MBCN152-1, which exhibited the best protection in screening trials and had no adverse effects on seedling growth, was selected for the greenhouse trial. In the greenhouse trial, cabbage seedlings, which had been grown in plug trays filled with soil mix containing spores of MBCN152-1 (1×10 8 spores g -1 of soil mix), were spray-inoculated with A. brassicicola and grown in greenhouse conditions. MBCN152-1 reduced disease incidence and significantly increased the number of viable seedlings. The efficacy of MBCN152-1 against damping-off caused by seed-borne A. brassicicola was then evaluated. Cabbage seeds, artificially infested with A. brassicicola, were sown in soil mix containing MBCN152-1 spores. The disease was completely suppressed when infested seeds were sown in a soil mix blended with MBCN152-1 at 1.5×10 7 spores g -1 of soil mix. These results strongly suggest that MBCN152-1 has the potential to control A. brassicicola on cabbage plug seedlings. MBCN152-1 was identified as a Streptomyces humidus-related species based on 16S rDNA sequencing. Scanning electron microscopy showed that the hyphae of MBCN152-1 multiplied on the surface of the seedlings and penetrated their epidermal cells. In conclusion, strain MBCN152-1 is a promising biocontrol agent against A. brassicicola on cabbage plug seedlings.

In October 2004, Pythium ultimum var. ultimum was isolated from rotten stems of cabbage plug seedlings in a commercial nursery in Mie Prefecture (Japan). The isolated fungus was then used to inoculate seedlings and subsequently reisolated from the seedlings with the damping-off disease, showing that P. ultimum var. ultimum is a new pathogen causing cabbage seedling disease.Key words Cabbage · Damping-off · Plug seedlings · Pythium ultimum var. ultimumIn October 2004, several plug seedlings of cabbage (Brassica oleracea L., cv. Miharu) at the two-to three-leaf stage had water-soaked rot on the stems around cotyledon axils in the greenhouse of a commercial nursery in Mie Prefecture (Japan). The diseased seedlings finally wilted. Colorless hyphae growing out from the lesions on hypocotyls were microscopically observed. A Pythium-like fungus ( Fig. 1a) was consistently isolated from the diseased hypocotyls with high frequency. Although damping-off of cabbage plug seedlings caused by Pythium megalacanthum de Bary, Pythium aphanidermatum (Edson) Fitzpatrick, and Pythium zingiberis Takahashi has been reported Abiko 1998, 2000), the Pythium sp. isolated from these diseased hypocotyls differed morphologically from those species. The purpose of this study was to identify the fungus causing this symptom, using morphological observation and a pathogenicity test.Hypocotyls excised from diseased seedlings were placed on 1.5% water agar plates and incubated at 25°C for 2-3 days. Hyphal tips from hyphae growing from the diseased hypocotyls were placed singly on potato sucrose agar (PSA) plates. Three representative isolates, designated as 1021-1, 1021-4, and 1027-1, were identified to species. These isolates, when grown on corn meal agar (CMA) supplemented with 0.1% fish powder at 25°C, formed white aerial mycelium. After a 1-week incubation, they produced oogonia, antheridia, and oopsores on the medium (Fig 1b). Oogonia were terminal and globose (17-28 µm in diameter) with a smooth surface. Antheridia were sac like, and >85% were monoclinous. The mean number of antheridia per oogonium was almost one. These morphological features coincided with those of Pythium ultimum Trow var. ultimum described by van der Plaats-Niterink (1981) (Table 1).To induce zoosporangia of isolates 1021-1, 1021-4, and 1027-1, pieces of autoclaved leaves of cabbage or bentgrass were inoculated with actively growing mycelial plugs (5 mm diameter) of the isolates at 25°C for 24 h. Then, four to six pieces of the leaves were soaked in 10 ml of petri solution [150 mg/l KH 2 PO 4 , 150 mg/l MgSO 4 , 60 mg/l KCl, and 400 mg/l Ca(NO 3 ) 2 ] in a petri dish (9 cm diameter) after the inocula were removed. No zoosporangium formed after incubation at 25°C within 7 days or at 5°C within 6 days followed by 24-48 h incubation at 25°C.The optimal temperature for mycelial growth on potato dextrose agar (PDA) of three isolates (1021-1, 1021-4, and 1027-1) was around 25°C. Growth rate at 25°C was 26-27 mm/24 h (Fig. 2). All isolates grew at 5°C, but not at 40°C.Based on ...

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