FIGURE 2.

The high-throughput interbacterial competition platform. (A) Interbacterial competition assay. Cultured attacker Agrobacterium tumefaciens and recipient Escherichia coli were mixed and then spotted on the Agrobacterium Kill (AK) agar medium to allow interbacterial competition for 16 h at 25°C followed by recovery of mixed cultures, serially diluted, and then spread onto lysogeny broth (LB) plate supplemented with appropriate antibiotics to select for recipient cells. (B) High-throughput interbacterial competition screening platform. Recipient cells were grown and mixed with attacker A. tumefaciens in a 96-well plate. The bacterial mixture was dropped onto the AK agar medium competition surface using an automated pipetting system. The competition surface was made on a microplate lid. Recovery was performed using microplate replicator. The candidates are the strains that show multiple colonies grown after recovery as opposed to wild-type controls and most strains with no or few colonies. This high-throughput A. tumefaciens type VI secretion system (T6SS) killing platform enables ∼400 mutant screens per day. This figure was created with BioRender (https://biorender.com/).