Summary: Tailored nanoparticles offer a novel approach to fight antibiotic‐resistant microorganisms. We analysed biogenic selenium nanoparticles (Se NP s) of bacterial origin to determine their antimicrobial activity against selected pathogens in their planktonic and biofilm states. Se NP s synthesized by Gram‐negative Stenotrophomonas maltophilia [Sm‐Se NP s(−)] and Gram‐positive Bacillus mycoides [Bm‐Se NP s(+)] were active at low minimum inhibitory concentrations against a number of clinical isolates of Pseudomonas aeruginosa but did not inhibit clinical isolates of the yeast species Candida albicans and C. parapsilosis . However, the Se NP s were able to inhibit biofilm formation and also to disaggregate the mature glycocalyx in both P. aeruginosa and Candida spp. The Sm‐Se NP s(−) and Bm‐Se NP s(+) both achieved much stronger antimicrobial effects than synthetic selenium nanoparticles (Ch‐Se NP s). Dendritic cells and fibroblasts exposed to Sm‐Se NP s(−), Bm‐Se NP s(+) and Ch‐Se NP s did not show any loss of cell viability, any increase in the release of reactive oxygen species or any significant increase in the secretion of pro‐inflammatory and immunostimulatory cytokines. Biogenic Se NP s therefore appear to be reliable candidates for safe medical applications, alone or in association with traditional antibiotics, to inhibit the growth of clinical isolates of P. aeruginosa or to facilitate the penetration of P. aeruginosa and Candida spp. biofilms by antimicrobial agents.

The present study deals with Se⁰- and Te⁰-based nanoparticles bio-synthesized by two selenite- and tellurite-reducing bacterial strains, namely Stenotrophomonas maltophilia SeITE02 and Ochrobactrum sp. MPV1, isolated from polluted sites. We evidenced that, by regulating culture conditions and exposure time to the selenite and tellurite oxyanions, differently sized zero-valent Se and Te nanoparticles were produced. The results revealed that these Se⁰ and Te⁰ nanoparticles possess antimicrobial and biofilm eradication activity against Escherichia coli JM109, Pseudomonas aeruginosa PAO1, and Staphylococcus aureus ATCC 25923. In particular, Se⁰ nanoparticles exhibited antimicrobial activity at quite low concentrations, below that of selenite. Toxic effects of both Se⁰ and Te⁰ nanoparticles can be related to the production of reactive oxygen species upon exposure of the bacterial cultures. Evidence so far achieved suggests that the antimicrobial activity seems to be strictly linked to the dimensions of the nanoparticles: indeed, the highest activity was shown by nanoparticles of smaller sizes. In particular, it is worth noting how the bacteria tested in biofilm mode responded to the treatment by Se⁰ and Te⁰ nanoparticles with a susceptibility similar to that observed in planktonic cultures. This suggests a possible exploitation of both Se⁰ and Te⁰ nanoparticles as efficacious antimicrobial agents with a remarkable biofilm eradication capacity.

Selenite (SeO₃ ²⁻) oxyanion shows severe toxicity to biota. Different bacterial strains exist that are capable of reducing SeO₃ ²⁻ to non-toxic elemental selenium (Se⁰), with the formation of Se nanoparticles (SeNPs). These SeNPs might be exploited for technological applications due to their physico-chemical and biological characteristics. The present paper discusses the reduction of selenite to SeNPs by a strain of Bacillus sp., SeITE01, isolated from the rhizosphere of the Se-hyperaccumulator legume Astragalus bisulcatus. Use of 16S rRNA and GyrB gene sequence analysis positioned SeITE01 phylogenetically close to B. mycoides. On agarized medium, this strain showed rhizoid growth whilst, in liquid cultures, it was capable of reducing 0.5 and 2.0 mM SeO₃ ²⁻ within 12 and 24 hours, respectively. The resultant Se⁰ aggregated to form nanoparticles and the amount of Se⁰ measured was equivalent to the amount of selenium originally added as selenite to the growth medium. A delay of more than 24 hours was observed between the depletion of SeO₃ ² and the detection of SeNPs. Nearly spherical-shaped SeNPs were mostly found in the extracellular environment whilst rarely in the cytoplasmic compartment. Size of SeNPs ranged from 50 to 400 nm in diameter, with dimensions greatly influenced by the incubation times. Different SeITE01 protein fractions were assayed for SeO₃ ²⁻ reductase capability, revealing that enzymatic activity was mainly associated with the membrane fraction. Reduction of SeO₃ ²⁻ was also detected in the supernatant of bacterial cultures upon NADH addition. The selenite reducing bacterial strain SeITE01 was attributed to the species Bacillus mycoides on the basis of phenotypic and molecular traits. Under aerobic conditions, the formation of SeNPs were observed both extracellularly or intracellullarly. Possible mechanisms of Se⁰ precipitation and SeNPs assembly are suggested. SeO₃ ²⁻ is proposed to be enzimatically reduced to Se⁰ through redox reactions by proteins released from bacterial cells. Sulfhydryl groups on peptides excreted outside the cells may also react directly with selenite. Furthermore, membrane reductases and the intracellular synthesis of low molecular weight thiols such as bacillithiols may also play a role in SeO₃ ²⁻ reduction. Formation of SeNPs seems to be the result of an Ostwald ripening mechanism.

Summary: In an effort to prevent the formation of pathogenic biofilms on hydroxyapatite ( HA )‐based clinical devices and surfaces, we present a study evaluating the antimicrobial efficacy of S pherical biogenic Se‐ N anostructures Em bedded in O rganic material (Bio Se‐ NEMO ‐S) produced by Bacillus mycoides Sel TE 01 in comparison with two different chemical selenium nanoparticle (Se NP ) classes. These nanomaterials have been studied as potential antimicrobials for eradication of established HA ‐grown biofilms, for preventing biofilm formation on HA ‐coated surfaces and for inhibition of planktonic cell growth of Pseudomonas aeruginosa NCTC 12934 and Staphylococcus aureus ATCC 25923. Bio Se‐ NEMO resulted more efficacious than those chemically produced in all tested scenarios. Bio Se‐ NEMO produced by B. mycoides Sel TE 01 after 6 or 24 h of Na ₂ SeO ₃ exposure show the same effective antibiofilm activity towards both P. aeruginosa and S. aureus strains at 0.078 mg ml ⁻¹ (Bio Se‐ NEMO ₆ ) and 0.3125 mg ml ⁻¹ (Bio Se‐ NEMO ₂₄ ). Meanwhile, chemically synthesized Se NP s at the highest tested concentration (2.5 mg ml ⁻¹ ) have moderate antimicrobial activity. The confocal laser scanning micrographs demonstrate that the majority of the P. aeruginosa and S. aureus cells exposed to biogenic Se NP s within the biofilm are killed or eradicated. Bio Se‐ NEMO therefore displayed good antimicrobial activity towards HA ‐grown biofilms and planktonic cells, becoming possible candidates as new antimicrobials.