Purpose: The purpose of this study was to evaluate how phage resistance influences the growth properties of Escherichia coli in a model system.
Methods: Bacteriophage insensitive mutants (BIMs) of Escherichia coli B (ATCC 11303) were produced by incubating suspensions of the bacteria with 10-fold dilutions of an initial 1010 PFU/ml stock of phage T4. Six colonies of BIMs (each isolated from different 10-fold phage dilutions) were tested for altered biochemical properties using the VITEK-2 system, susceptibility to antibiotics including extended spectrum beta-lactamases (ESBLs), aminoglycosides (kanamycin) and second generation cephalosporins. Also, growth curves in TSB were conducted for all BIMs.
Results: Compared to wildtype, all BIMs lost the ability to ferment glucose, and were positive for lactate and succinate alkalinisation, suggesting that amino acids in the growth medium were being degraded, resulting in acid stress within the bacterial cytoplasm. A primary mechanism of phage T4 resistance in E. coli is to mutate its primary receptors lipopolysaccharide and OmpC, which plays a major role in allowing entry of antibiotics into the bacterial cell. All BIMS were sensitive to ESBLs and cephalosporins but displayed increased MICs for kanamycin (four µg/ml compared to two µg/ml for the wildtype (one BIM had a MIC of eight µg/ml). Growth curves indicated that all BIMs except for one had longer exponential phases than the wild type, and all BIMs had higher ODs (0.85 to 0.9) at 600 nm than the wild type (0.7). Doubling times for the BIMs ranged from 33 to 40 mins in comparison to the wildtype (44 min).
Significance: These results indicate that phage induced mutations may influence fitness of E. coli in food, including resistance to antimicrobials that enter the cell through porins.