Bacteria facing chalchogens: biogenic formation of Se and Te nanoparticles and evaluation of their antimicrobial potential

The chalcogens selenium and tellurium can be found as trace elements in the environment. Selenium and tellurium in their elemental form (i.e. Se0 and Te0) are insoluble in water and almost non-toxic, while the oxyanions selenite (SeO32-) and tellurite (TeO32-) are highly soluble in water and toxic to biological systems. The toxicity of these oxyanions has been ascribed to their oxidizing activity, which can interfere with fundamental cellular functions. Bacteria have developed different mechanisms for the transformation of metal oxyanions to non-toxic forms. In fact, a number of bacterial strains has revealed the capacity to reduce selenite to elemental selenium in different conditions, with formation of nanoparticles with various morphological characteristics, either inside the cells or extracellularly. Similarly, tellurite reduction to elemental tellurium nanoparticles has been reported in different cellular compartments within a wide range of bacterial species. This appears particularly worth of note since the recent boost of nanotechnology has attracted growing interest in the exploitation of the natural ability of biological systems to generate nanomaterial with well-defined properties both in the cytoplasm and outside the bacterial cells. Indeed, this can be considered as a green and ecofriendly alternative to the chemical and physical methods conventionally used to synthesize nanomaterials. In the present PhD work, the utilization of bacterial strains to produce selenium and tellurium nanoparticles along with the possible applications of these biogenic nanomaterials have been evaluated. In the first part of this dissertation, the mechanisms involved in the biogenic formation of nanoparticles by different microbial isolates have been analyzed. In particular, the reduction of selenite to elemental selenium nanoparticles has been studied in Stenotrophomonas maltophilia SeITE02, Bacillus mycoides SeITE01, while the reduction of both selenite and tellurite has been evaluated in Ochrobactrum sp. MPV1. On the basis of the evidences discussed in this thesis, the involvement of both thiolic compounds (glutathione for Stenotrophomonas maltophilia SeITE02 and Ochrobactrum sp. MPV1, bacillithiols for Bacillus mycoides SeITE01) and intracellular/extracellular enzymes can be singled out. On the other hand, tellurite reduction in Ochrobactrum sp. MPV1 may be attributed to the activity of an intracellular NADH-dependent enzyme, exhibiting a reducing mechanism different from that involved in selenite reduction. In the second part of the dissertation, biogenic nanomaterials produced by the microbial strains of interest are characterized in terms of physico-chemical parameters as well as for their biological activity. Biogenic SeNPs showed similar sizes (dependent on incubation time) and high stability (negative ζ-potential). In this respect, also biogenic TeNPs from Ochrobactrum sp. MPV1 revealed a high stability (positive ζ-potential). Moreover, the antimicrobial potential of these biogenic nanomaterials has been investigated against reference strains. In particular, Se0 nanoparticles exhibited antimicrobial activity at quite low concentrations. 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 that bacteria tested in biofilm mode of growth responded to the treatment by Se0 and Te0 nanoparticles with a susceptibility similar to that observed in planktonic cultures. Comparative tests were also performed with biogenic SeNPs extracted from two different bacterial strains, namely Stenotrophomonas maltophilia SeITE02 and Bacillus mycoides SeITE01. These biogenic nanoparticles showed a higher antimicrobial potential than exerted by the chemically synthesized ones. Finally, toxicity and induction of cytokine production by these nanomaterials were tested on human fibroblasts and dendritic cells, evidencing on the one hand no toxic effects, while on the other induction of cytokine production only at high concentrations (250 and 500 mg/L). All these results open the perspective of a possible exploitation of both Se0 and Te0 nanoparticles as efficacious antimicrobial agents with a remarkable biofilm eradication capacity.