The misuse of antibiotics is causing widespread antibiotic resistance, creating an urgent need for new treatment options such as nanoparticle-based therapies. This study aimed to compare silver nanoparticles (AgNPs) produced via green synthesis methods with those made through traditional chemical processes. Furthermore, the study investigated and contrasted the bacterial responses to these two types of AgNPs over a 21-day period of selection pressure using experimental evolution techniques. Analysis using scanning electron microscopy and transmission electron microscopy revealed a consistent, uniform morphology among the AgNPs produced via chemical methods. In contrast, AgNPs synthesized through green methods displayed an irregular morphology. Despite these morphological differences, all nanoparticles from both synthesis approaches were under 100 nm in diameter. These findings were further supported by the absorption spectrum data, which showed a maximum absorption peak between the 400 and 500 nm wavelength range. E. coli exposed to green synthesized AgNPs for 21 days adapted to their presence, exhibiting both enhanced resistance to the green synthesized AgNPs themselves and the development of cross-resistance to ionic silver, a pattern not observed in chemically synthesized AgNP-selected populations. Populations selected using chemical synthesized AgNPs did not develop increased resistance to either chemically or green synthesized AgNPs; however, they showed a slight increase in resistance to ionic silver. Genomics analysis identified polymorphism in genes in a green synthesized AgNP-resistant line including but not limited to the multidrug efflux transporter system (EmrAB), DUF4756 family protein (D1792_RS05680), putative zinc-binding protein YnfU/cold shock-like protein (ynfU/cspB) and imcF-related family protein (D1792_RS10035). Bacterial resistance to chemical AgNPs involves specific polymorphisms in key bacterial components like the RNA polymerase sigma factor (RpoE) and the EmrAB efflux pump. Collectively, the method used to synthesize the AgNPs influences their antibacterial efficacy and the likelihood of bacteria developing resistance. Understanding this interaction is vital for developing effective and resistance-controlled applications of AgNPs across medicine, environmental science, and industry.

The antimicrobial resistance crisis has led to the use of metals and bacteriophages as possible alternatives to antibiotics. Experimental studies have examined interactions between ionic/nano-silver and bacteriophages against multidrug-resistant bacteria. However, these approaches have often failed to examine whether silver affects the stability and infectivity of bacteriophages. Here, we utilized experimental evolution to evolve resistance to ionic silver in bacteriophage T7. High ionic silver concentrations that do not represent physiological exposure conditions were used to impose strong selective pressure. Evolution of ionic silver resistance in phage T7 was rapid, as evidenced by recovery of bacteriophage growth in E. coli following repeated exposures to ionic silver, enhanced infectivity of silver-selected populations relative to parallel control and ancestral populations under increasing ionic silver concentrations, and greater suppression of E. coli growth in standard medium. Furthermore, silver resistance evolved without loss of thermal or pH stability under the conditions tested. The genomic foundation of silver resistance was relatively simple, with positive and negative natural selection differentiating the silver-selected populations from the controls and ancestral populations across serial passages in silver. Support for replication-associated adaptation under ionic silver selection may be reflected in recurrent mutations identified in genes involved in transcription, DNA replication, and genome maintenance, including T7p07 (RNA polymerase), T7p10 (DNA ligase), and T7p29 (DNA polymerase I). These findings highlight the importance of evaluating phage –silver combination strategies within an evolutionary framework that accounts for the adaptive capacity of bacteriophages under silver selection.

Antimicrobial resistance (AMR) represents a critical global health challenge in the 21st century, with drug-resistant infections accounting for approximately 4.95 million deaths annually; moreover, forecasts suggest a significant increase in this toll by 2050. Traditional approaches to antibiotic development have demonstrated an inability to keep pace with bacterial evolution. Consequently, there has been a resurgence of scientific interest in alternative therapeutic strategies, specifically bacteriophage therapy and metal-based antimicrobials. This narrative review examines the molecular processes that help bacteria evolve resistance to both phages and metals, a topic often studied separately. It encompasses receptor modification, CRISPR–Cas adaptive immunity, abortive infection systems, efflux pump evolution, the rewiring of regulatory networks, and also examines the impact of dual selection on these mechanisms. We investigate the complex, combined evolutionary effects of exposure to phages and metals, both simultaneously and sequentially. This includes mutations that confer resistance to multiple stressors, the co-selection of resistance traits associated with mobile genetic elements, and the transfer of metal resistance genes between bacteria via phages. The outer membrane, metal transport systems, and efflux pumps are key cellular areas where phages and metals create overlapping and interacting selective pressures. Direct experimental evidence for combined phage–metal selection remains scarce, and we therefore distinguish throughout between direct phage–metal evidence, evidence transferred from phage–antibiotic and metal–antibiotic systems, and mechanistic hypotheses that remain to be tested. Understanding these complex evolutionary dynamics is essential for formulating combination therapeutic approaches that reduce the emergence of resistance, improve antimicrobial effectiveness, and address the growing challenge of multidrug-resistant pathogens in both clinical and environmental settings.