A defined, fetal bovine serum (FBS)-free medium was developed and validated for the isolation, cultivation, and quantification of Renibacterium salmoninarum (Rs), the etiological agent of Bacterial Kidney Disease (BKD). By employing precise combinations of selected peptone compounds along with optimized titrations of cysteine and NaCl at 0.15%, a chemically defined medium was obtained that supports robust Rs growth in both liquid and solid media. This system ethically replaces the use of FBS while maintaining high levels of sustainability, reproducibility, and adaptability.
The new medium enabled stable and abundant growth of all evaluated clinical strains, which retained their transmissibility and pathogenicity. Bacteria could be easily transferred between liquid and solid media without loss of viability, demonstrating the versatility of the system. Subsequent infection of the SHK-1 cell line with these strains exhibited varying degrees of cytotoxicity, but no correlation with passage through the new medium, suggesting that these differences are attributable to minor genetic variations, later confirmed in Salmo salar in vivo assays.
Finally, it was confirmed that this medium does not interfere with programmed cell death mechanisms associated with infection (apoptosis), allowing its use in immunopathological studies without culture-induced artifacts. This platform represents an ethical and technical advancement for BKD diagnostics, with applications in both laboratory and field settings.