Synergism of imipenem with fosfomycin associated with the active cell wall recycling and heteroresistance in Acinetobacter calcoaceticus-baumannii complex

The carbapenem-resistant Acinetobacter calcoaceticus-baumannii (ACB) complex has become an urgent threat worldwide. Here, we determined antibiotic combinations and the feasible synergistic mechanisms against three couples of ACB (A. baumannii (AB250 and A10), A. pittii (AP1 and AP23), and A. nosocomialis (AN4 and AN12)). Imipenem with fosfomycin, the most effective in the time-killing assay, exhibited synergism to all strains except AB250. MurA, a fosfomycin target encoding the first enzyme in the de novo cell wall synthesis, was observed with the wild-type form in all isolates. Fosfomycin did not upregulate murA, indicating the MurA-independent pathway (cell wall recycling) presenting in all strains. Fosfomycin more upregulated the recycling route in synergistic strain (A10) than non-synergistic strain (AB250). Imipenem in the combination dramatically downregulated the recycling route in A10 but not in AB250, demonstrating the additional effect of imipenem on the recycling route, possibly resulting in synergism by the agitation of cell wall metabolism. Moreover, heteroresistance to imipenem was observed in only AB250. Our results indicate that unexpected activity of imipenem on the active cell wall recycling concurrently with the presence of heteroresistance subpopulation to imipenem may lead to the synergism of imipenem and fosfomycin against the ACB isolates.