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Silver nanomaterials have been produced by chemical or biological methods requiring room to higher temperatures resulting in poor stable particles. Only a couple of reports use low temperature for their synthesis but, in these cases, nanoparticles aggregated in a short time. We hypothesized that psychrophilic bacteria could be found that produce stable nanoparticles at low temperature with antimicrobial activity that can be of use in biomedicine against pathogenic bacteria. Isolation of several bacteria from Antarctica was performed to obtain different psychrophilic bacteria, and their capability to extracellularly synthesize silver nanoparticles tested. Different conditions, including presence of sodium chloride and a low (4 °C) and a medium (30 °C) temperature, were used and the structure of the nanoparticles physicochemically characterized. The antimicrobial activity of freshly prepared and aged nanoparticles was determined against Gram-negative and Gram-positive bacteria. Nanoparticles were synthesized by all four psychrophilic bacteria at 4 °C and 30 °C. NaCl highly decreased the synthesis of nanosilver particles. Depending on the bacteria and temperature used, nanoparticles were spherical with sizes 5.0–11.1 nm, and stable after several months of incubation under light. The most stable nanoparticles were those kept at 4 °C and the highest detected activity was against Gram-positive bacteria. [ABSTRACT FROM AUTHOR]