Development of new G media for culture of Flavobacterium columnare and comparison with other media

• Three pairs of broth and agar media for culturing Flavobacterium columnare were developed.
• Within 24 h the bacterial culture on agar plates was confirmed and > 3 × 108 CFU/ml cells in broths were achieved.
• The clumps were eliminated so the colony counts were more accurate and reliable for all research purposes.

Each component of existing agar and broth media was compared based on observations of Flavobacterium columnare growth. Newly formulated G media resulted in the fastest proliferation of F. columnare cells in broth and yielded fully developed colonies on inoculated agar. In the formulation of the G media, consideration was given to factors such as Sodium chloride and antibiotics, which affect bacterial growth. Ingredients were added or subtracted from existing media, with the resulting formulation of 3 broths (Gb1, Gb2, Gb3) and 3 agars (Ga1, Ga2, Ga3). F. columnare culture on G agar media could be confirmed within 24 h for disease diagnostics. Because of the rapid growth of the bacterium, colony counts were performed after 24 h, avoiding the merging of adjacent colonies at 48 h. In G broth, the absorbance reached 0.3–0.5 within 24 h, indicating ≥ 3 × 108 CFU/ml in broth. In addition, no clumps formed in G broth, which ensured a uniform distribution of F. columnare cells compared to existing broths. The G media are superior for F. columnare culture because of the rapid uniform growth of the bacterium compared to existing media.Statement of relevanceThe newly developed G media significantly support faster growth of F. columnare in broth and on agar surface than that with the existing media.The formulations of the broth media are focused on the elimination of bacterial cell clumping and quick cell proliferation, and to maximize the colony development of F. columnare cultured on agar surface when the antibiotic of neomycin presented.The ingredients and their concentrations were thoroughly investigated and the reasons that new media are better than existing media are explained and discussed.