Summary In this study, we investigated the tactic response of Pseudomonas putida G7, a representative soil bacterium, towards silver nanoparticles (AgNPs). The study integrated the characterization of surface area and size distribution of AgNPs, toxicity determinations, based on ATP production, and assessment of the repellent reaction by means of an inverted capillary assay (‘chemical‐in‐pond’ method), and changes in the motility behaviour determined by computer‐assisted motion analysis. Our data demonstrate, for the first time, that nanoparticles can elicit a negative tactic response in bacteria at low but environmentally relevant, sublethal concentrations. Data obtained by the chemical‐in‐pond method indicated that cells exposed to 0.1 mg l −1 of two AgNPs preparations, differing in particle size (maximum diameter ≤ 100 nm and ≤ 150 nm respectively), were repelled in the gradients created inside the capillaries. However, cells exposed to similar low concentration of AgNO 3 did not demonstrate any detectable repellent response, although it reduced cell viability by 20%, a decrease comparable to that caused by AgNPs. Computer analysis of swimming behaviour of cells exposed to AgNPs (0.2 mg l −1 ) revealed a significant increase in turning events, as compared with unexposed controls, which is characteristic of bacterial repellent response. Greater AgNPs concentrations (up to 100 mg l −1 ) also induced changes in the swimming behaviour, although they did not induce any detectable repellent response as determined by the chemical‐in‐pond assays. In contrast, AgNO 3 failed to induce the repellent swimming behaviour within the wide range of concentrations tested (0.001–100 mg l −1 ), and caused a significant inhibition of cell motility at a concentration above 0.1 mg l −1 . The evidence presented here suggests there are likely to be alternative mechanisms by which nano‐scale silver induces a repellent response, which is more direct than the toxic response of macro‐forms of silver, attributed to ion formation and exposure.