Abstract:[Objective] Nisin, a bioactive antimicrobial peptide, inhibits many Gram-positive bacteria causing food spoilage and infection. In this study, cost-effective screening method for high-yield nisin strains was developed using high-throughput screening technology to provide a practical method for screening during the industrial production process. [Methods] Mutants (2511) were obtained by applying ultraviolet mutagenesis treatments to Lactococcus lactis ATCC 11454, followed by development of a 96-well plate high-throughput screening method using a Biomek FXP automatic workstation to select mutant strains for culture dilution. Diluted bacterial solutions were transferred to another 96-well plate with Micrococcus luteus grown to logarithmic phase, and an improved turbidimetric method was used to rapidly detect the biological activity of nisin and screen the mutants associated with the high-yield strains. High-yield strains were fermented by shake-flask fermentation to evaluate the screening method. [Results] Turbidimetric detection revealed nisin activity in the range between 10 and 25 IU/mL, with M. luteus biomass detected after a 2 h reaction with nisin. After two rounds of high-throughput screening, about 50 high-yield strains were obtained from the 2511 mutants, with 8 accurately detected by shake-flask fermentation. Results indicated that the nisin yield of the tested strains all increased, with one strain showing a 30% increase in production, indicating the efficacy of the high-throughput screening method. [Conclusion] Our high-throughput screening method was capable of screening high nisin-producing bacteria.
