Development of a Robust and Quantitative High-Throughput Screening Method for Antibiotic Production in Bacterial...

Area of Science:

• Microbiology
• Drug Discovery
• Biotechnology

Background:

• The rise of multidrug-resistant bacteria necessitates novel antibiotics.
• Antibiotic discovery has slowed, with few new mechanisms reaching the market.
• Current screening methods for natural product antibiotics are slow and error-prone.

Purpose of the Study:

• To develop a rapid, robust, and quantitative high-throughput liquid culture screening method for bacterial antibiotic production.
• To identify novel antibiotic-producing bacteria from large libraries.
• To adapt the method for screening both monocultures and cocultures.

Main Methods:

• A high-throughput liquid culture screening assay was developed and optimized.
• Over 260 bacterial species were screened in monoculture for antibiotic activity against *Staphylococcus aureus* and *Escherichia coli*.
• Bacteria not producing antibiotics in monoculture were screened in coculture using an adapted method.
• Antibiotic-producing isolates were identified via 16S rRNA sequencing.

Main Results:

• 38% of monocultures and 14% of cocultures inhibited *S. aureus* growth.
• 34% of monocultures and 30% of cocultures inhibited *E. coli* growth.
• Strong antibiotic producers (≥30% growth inhibition) were identified in 8% (monoculture) and 4% (coculture) of screenings against *S. aureus*, and 4% (monoculture) and 30% (coculture) against *E. coli*.
• Sequencing identified *Pseudomonas*, *Serratia*, and *Bacillus* as dominant antibiotic-producing genera, with novel producers *Herbaspirillum* and *Kluyvera* also identified.

Conclusions:

• The developed high-throughput liquid culture method is effective for discovering novel antibiotic-producing bacteria.
• Coculture screening significantly increases the detection rate of antibiotic-producing bacteria.
• This method accelerates the discovery of natural product antibiotics, crucial for combating antimicrobial resistance.