Enhancing bovicin HC5 production in Streptococcus equinus HC5 through adaptive laboratory evolution under thermal...

Area of Science:

• Microbiology and Biotechnology
• Metabolic Engineering
• Bacteriocin Production

Background:

• Bovicin HC5, a bacteriocin from Streptococcus equinus HC5, has antimicrobial properties but suffers from low production yields.
• Low yields limit the practical application of bovicin HC5 in controlling pathogenic and spoilage microorganisms.

Purpose of the Study:

• To investigate the effect of temperature on S. equinus HC5 growth and bovicin HC5 production.
• To enhance bovicin HC5 production using adaptive laboratory evolution (ALE) under heat stress.
• To characterize the resulting high-producing variants.

Main Methods:

• Determined the optimal growth temperature for the wild-type S. equinus HC5 strain.
• Applied ALE for 400 generations at elevated temperatures (47°C and 48°C) to select for improved bovicin HC5 production.
• Analyzed bovicin HC5 yield, bvcA gene expression, thermal resistance, growth rate, biomass accumulation, membrane composition, and genomic mutations in selected variants.

Main Results:

• The optimal growth temperature for S. equinus HC5 was 42°C; growth ceased above 49°C.
• Two ALE-derived variants showed up to a 140% increase in bovicin HC5 production.
• The highest-producing variant (S. equinus HC5 40048) exhibited increased bvcA expression, enhanced thermal resistance, higher growth rate (1.33 h⁻¹), and greater biomass (OD₆₀₀nm = 4.03) at 48°C compared to the wild-type (0.98 h⁻¹, OD₆₀₀nm = 1.96).

Conclusions:

• Adaptive laboratory evolution is an effective strategy for enhancing bovicin HC5 production in S. equinus HC5.
• ALE-generated variants demonstrate improved thermal resistance and metabolic characteristics, linked to mutations in genes regulating protein modification, transcription, and transport.
• The study provides valuable insights for optimizing bacteriocin biosynthesis and developing S. equinus HC5 as a microbial production platform.