0 mm; (2) dark brown lesions of 1.0 to <4.0 mm; (3) black lesions of 4.0 to <7.0 mm; (4) black lesions of ≥7.0 mm in diameter that coalesce with one another; and (5) mostly coalesced black lesions covering more than 70% of the surface (or fully rotted) [25]. The Fusarium isolate pathogenic to ginseng roots was grown on CLA and PDA and identified based on the mycological characteristics referred to the descriptions of the Fusarium Laboratory Manual [24].

For molecular identification of the Fusarium isolate, genomic DNA was extracted from the mycelia of the pure fungal culture obtained by single spore isolation using PrepMan Ultra Sample Preparation Reagent (Applied Biosystems, Foster Everolimus in vivo City, CA, USA) [26]. The translation elongation factor-1α gene (EF-1α) was amplified through polymerase

chain reaction using primers EF1/EF2, and nucleotide sequences were generated using BigDye terminator version 3.1 cycle sequencing kits (Applied Biosystems) and registered in GenBank as GenBank Accession No. KC478361. Molecular identification of the pathogen was accomplished by BLAST analysis of the gene sequences by comparing sequence similarities to others registered in GenBank. To select antifungal bacteria against the Fusarium pathogen causing ginseng root rot, 392 bacteria were isolated from diseased ginseng roots and from mountain-, wetland-, and field-soils of various crops. For the dual culture tests, bacteria were grown in nutrient broth for 2 d, and 10 μL bacterial suspensions were spotted on Trichostatin A supplier three sections of the PDA. A mycelial plug (5 mm diameter) of the pathogen culture taken with a 5-mm-diameter cork-borer from the margin of a 7-d-old colony on the PDA was placed in the center of another PDA spotted with bacterial suspensions. After

1 wk of incubation, Cyclic nucleotide phosphodiesterase the pathogen mycelial growth of bacterial colonies (relative to the untreated control) was measured to determine the antifungal activity of the bacterial isolates. Three replications were used for each treatment. One bacterial isolate (isolate B2-5) out of 392 that showed a strong antifungal activity was selected and identified based on Gram staining, bacterial morphology, carbon source assimilation, and 16S ribosomal RNA (rRNA) gene sequencing analysis. Gram staining of the bacterial cells was conducted following the Laboratory Guide for Identification of Plant Pathogenic Bacteria [27]. The bacterial morphology was examined under a transmission electron microscope (JEM-1010, JEOL Ltd., Tokyo, Japan) operating at an accelerating voltage of 80 kV after negative staining with 1.0% uranyl acetate. Carbon source assimilation of the bacterial isolate was examined in the Biolog GN test kit (Biolog Inc., Hayward, CA, USA). For 16S rRNA gene sequencing analysis, the bacterial isolate was cultured on BHI agar at 28°C for 2 d, and its genomic DNA was extracted from the colony using a FastDNA spin kit (MP Biomedicals, Santa Ana, CA, USA).