Treatment sequences of a phage and colistin combination trigger different evolution pathways of a multidrug-resistant...

To better understand the underlying mechanisms of treatment order of phage and antibiotic synergy (PAS), particularly in the context of planktonic bacterial populations.

Phage and antibiotic synergy (PAS) is often adopted clinically to address shortcomings of phage monotherapy. Building on a previously identified PAS combination, IME-AB2 phage and colistin, demonstrating sequential effects, we conducted a systematic investigation to elucidate mechanisms driving the phenotypic outcomes of three treatment orders (phage-first, simultaneous, and antibiotic-first) against a resistant Acinetobacter baumannii.

Our findings revealed phage pretreatment effectively decapsulated bacteria to promote colistin binding and bacterial killing, while colistin pretreatment altered bacterial morphology and intracellular machinery, reducing phage reproduction and hence the PAS level. For simultaneous treatment, colistin also reduced the phage reproduction rate, but phage-mediated bacterial decapsulation promoted colistin binding and outer membrane disruption, leading to significant PAS. Notably only the simultaneous treatment is capable of suppressing resistance evolvement. Genome sequencing analysis revealed that colistin-pretreated bacteria modified lipopolysaccharide/outer membrane structure by regulating the QseB/C signal transduction system to promote resistance. On the other hand, phage-pretreated bacteria acquired mutations in gene clusters related to polysaccharide synthesis, impairing phage adsorption at the cost of weakening biofilm formation ability.

Overall, this extended mechanistic study provided important insights into the impacts of treatment orders on PAS and resistance dynamics, suggesting the need for more fundamental studies to realize the full clinical potential of the PAS strategy.