Bacteriophage resistance in Lactococcus lactis ssp. lactis using antisense ribonucleic acid

Area of Science:

• Microbiology
• Molecular Biology
• Bacteriophage Research

Background:

• Bacteriophages pose a significant threat to Lactococcus lactis dairy starter cultures.
• Conserved genes within bacteriophages are potential targets for antimicrobial strategies.
• Antisense RNA technology offers a method to inhibit gene expression.

Purpose of the Study:

• To investigate the potential of antisense RNA to confer resistance in Lactococcus lactis against bacteriophage infection.
• To identify and characterize conserved bacteriophage genes suitable for antisense RNA targeting.
• To evaluate the efficacy of antisense RNA expression in inhibiting bacteriophage replication.

Main Methods:

• Identification and characterization of two conserved open reading frames (gp18C and gp24C) in L. lactis bacteriophages.
• Confirmation of gene presence using polymerase chain reaction (PCR).
• Construction of plasmids expressing antisense RNA targeting gp18C and gp24C under a Lactococcus lactis promoter.

Main Results:

• Antisense RNA expression reduced bacteriophage plaque formation efficiency by up to 50%, resulting in smaller, irregular plaques.
• Bacteriophage replication was severely inhibited in strains expressing antisense RNA.
• A significant decrease in bacteriophage numbers was observed within the first 3 hours of infection compared to control strains.

Conclusions:

• Antisense RNA targeting conserved bacteriophage genes is an effective strategy for developing phage-resistant Lactococcus lactis strains.
• This approach holds promise for protecting dairy starter cultures from bacteriophage contamination.
• Further research can explore optimizing antisense RNA constructs for enhanced phage resistance.