Copper (Cu) is known to accumulate in agricultural soils receiving urban waste products as fertilizers. We here report the use of the leucine incorporation technique to determine pollution-induced community tolerance (Leu-PICT) to Cu in a long-term agricultural field trial. A significantly increased bacterial community tolerance to Cu was observed for soils amended with organic waste fertilizers and was positively correlated with total soil Cu. However, metal speciation and whole-cell bacterial biosensor analysis demonstrated that the observed PICT responses could be explained entirely by Cu speciation and bioavailability artifacts during Leu-PICT detection. Hence, the agricultural application of urban wastes (sewage sludge or composted municipal waste) simulating more than 100 years of use did not result in sufficient accumulation of Cu to select for Cu resistance. Our findings also have implications for previously published PICT field studies and demonstrate that stringent PICT detection criteria are needed for field identification of specific toxicants.