In natural habitats, bacteria spend most of their time in some form of growth arrest. Little is known about deleterious mutations in such stages, and consequently there is limited understanding of what evolutionary events occur. In a deleterious mutation accumulation experiment in prolonged stationary phase of Escherichia coli, about 0.03 slightly deleterious mutations were observed per genome per day. This is over an order of magnitude higher than extrapolations from fast-growing cells, but in line with inferences from observations in adaptive stationary phase mutation experiments. These findings may affect understanding of bacterial evolution and the emergence of bacterial pathogenicity.