Resistance, heteroresistance, and fitness costs drive colistin treatment failure during <i>Acinetobacter baumannii</i> pneumonia.

Abstract

<i>Acinetobacter baumannii</i> is an <i>Enterococcus</i>, <i>Staphylococcus</i>, <i>Klebsiella</i>, <i>Acinetobacter</i>, <i>Pseudomonas</i>, and <i>Enterobacter</i> species (ESKAPE) pathogen linked to healthcare-associated diseases. Due to evolved resistance, last-resort antibiotics such as the lipooligosaccharide (LOS)-targeting colistin are increasingly used to treat multidrug-resistant isolates. To track the evolution of colistin resistance within a host, we performed sequential oropharyngeal infections in immunocompetent or immune-depleted mice in the presence of inhaled colistin. Both resistant and heteroresistant <i>A. baumannii</i> strains emerged with <i>pmrB</i> mutations that efficiently competed with the susceptible parent in the presence of colistin. These <i>pmrB</i> mutants had a fitness cost in the absence of colistin treatment but retained their ability to colonize the host. In contrast, LOS-deficient <i>A. baumannii</i> mutants removed the target of colistin, but such mutants were unable to colonize the lung. The two pathogenic <i>pmrB</i> mutants showed clear evidence of LOS modification, which was linked to increased transcription of LOS modification enzymes, including the product of the cryptic <i>eptA</i> gene. Spontaneous insertion mutations that caused hyperexpression of <i>eptA</i> allowed the heteroresistant mutant to develop clinically significant colistin resistance. Insertion mutations upstream of the <i>eptA</i> gene or those disrupting <i>hns</i>, which encodes a small histone-like protein, resulted in increased <i>eptA</i> transcript, linking expression of this protein to clinically significant resistance. A resistant variant derived from the heteroresistant parent was stable in the absence of drug, but continued passaging selected for colistin-resensitized pseudorevertants that were largely due to disruption of the LOS modification enzymes. Therefore, colistin heteroresistance is an early stage in the stepwise acquisition of stable colistin resistance in <i>A. baumannii</i>.