The lethal effects of inorganic acid on phoE+ Escherichia coli strains, grown at neutral pHo, were enhanced by chloramphenicol, apparently because some organisms acquire acid tolerance (habituate) during challenge and chloramphenicol stops this. Phosphate (and/or polyphosphate) present during challenge prevented killing and damage by acid to outer membranes, DNA and cellular enzymes but did not prevent acid pHo enhancing novobiocin activity. To reverse acid effects, phosphate must interact with or cross the outer membrane but need not enter the cytoplasm; it is probable that it competes with H+ (or protonated anions) for passage through the PhoE pore. Phosphate also prevented induction of beta-galactosidase in a strain with the cadA promoter fused to lacZ. Four unc mutants showed essentially normal acid sensitivity and habituation; the same was true for strains with lesions in fur, oxyR, katF, phoP, cadA and hycB. In contrast, deletion of rpoH led to slightly increased acid sensitivity for cells grown at pHo 7.0, although habituation was relatively normal.