Biogenic iron-silver nanoparticles inhibit bacterial biofilm formation due to Ag release as determined by a novel phycoerythrin-based assay.

Silver nanoparticles (Ag-NPs) can be considered as a cost-effective alternative to antibiotics. In the presence of Fe(III)-citrate and Ag, Klebsiella oxytoca DSM 29614 produces biogenic Ag-NPs embedded in its peculiar exopolysaccharide (EPS). K. oxytoca DSM 29614 was cultivated in a defined growth medium-containing citrate (as sole carbon source) and supplemented with Ag and either low or high Fe(III) concentration. As inferred from elemental analysis, transmission and scanning electron microscopy, Fourier transform infrared spectrometry and dynamic light scattering, Ag-EPS NPs were produced in both conditions and contained also Fe. The production yield of high-Fe/Ag-EPS NPs was 12 times higher than the production yield of low-Fe/Ag-EPS NPs, confirming the stimulatory effect of iron. However, relative Ag content and Ag ion release were higher in low-Fe/Ag-EPS NPs than in high-Fe/Ag-EPS NPs, as revealed by emission-excitation spectra by luminescent spectrometry using a novel ad hoc established phycoerythrin fluorescence-based assay. Interestingly, high and low-Fe/Ag-EPS NPs showed different and growth medium-dependent minimal inhibitory concentrations against Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 15442. In addition, low-Fe/Ag-EPS NPs exert inhibition of staphylococcal and pseudomonal biofilm formation, while high-Fe/Ag-EPS NPs inhibits staphylococcal biofilm formation only. Altogether, these results, highlighting the different capability of Ag release, support the idea that Fe/Ag-EPS NPs produced by K. oxytoca DSM 29614 can be considered as promising candidates in the development of specific antibacterial and anti-biofilm agents.Key points • Klebsiella oxytoca DSM 29614 produces bimetal nanoparticles containing Fe and Ag.• Fe concentration in growth medium affects nanoparticle yield and composition.• Phycoerythrin fluorescence-based assay was developed to determine Agrelease.• Antimicrobial efficacy of bimetal nanoparticle parallels Agions release.