Aims: This article shows the effect of nanosecond pulsed electric field (nsPEF) on Escherichia coli, which could imply a durable change in protein expressions and then impacted the phenotype of surviving bacteria that might lead to increase pathogenicity. Methods and Results: The effects of nsPEF on E. coli viability and membrane permeabilization were investigated. One log(10) reduction in bacterial counts was achieved at field strength of 10 7 V m(-1) with a train of 500 successive pulses of 60 9 10(-9) s. Incubation of germs after treatment with propidium iodide showed that membrane permeabilization was reversible. Possible protein changes of surviving bacteria were checked to assess potential phenotypical changes using two-dimensional electrophoresis. In our study, after 40 generations, only UniProt #P39187 was up-regulated with P = 0.05 compared with the control and corresponded to the uncharacterized protein YtfJ. Antibiograms were used to check whether or not the pattern of cultivable bacteria after nsPEF deliveries changed. Conclusions: The results tend to show that nsPEFs are able to inactivate bacteria and have probably no serious impact in E. coli protein patterns. Significance and Impact of the Study: The use of nsPEF is a safe promising new nonthermal method for bacterial inactivation in the food processing and environmental industry.