The actinomycete Kitasatospora sp. SeTe27, subjected to adaptive laboratory evolution (ALE) in the presence of selenite, varies its cellular morphology, redox stability, and tolerance to the toxic oxyanion.

The effects of oxyanions selenite (SeO32-) in soils are of high concern in ecotoxicology and microbiology as they can react with mineral particles and microorganisms. This study investigated the evolution of the actinomycete Kitasatospora sp. SeTe27 in response to selenite. To this aim, we used the Adaptive Laboratory Evolution (ALE) technique, an experimental approach that mimics natural evolution and enhances microbial fitness for specific growth conditions. The original strain (wild type; WT) isolated from uncontaminated soil gave us a unique model system as it has never encountered the oxidative damage generated by the prooxidant nature of selenite. The WT strain exhibited a good basal level of selenite tolerance, although its growth and oxyanion removal capacity were limited compared to other environmental isolates. Based on these premises, the WT and the ALE strains, the latter isolated at the end of the laboratory evolution procedure, were compared. While both bacterial strains had similar fatty acid profiles, only WT cells exhibited hyphae aggregation and extensively produced membrane-like vesicles when grown in the presence of selenite (challenged conditions). Conversely, ALE selenite-grown cells showed morphological adaptation responses similar to the WT strain under unchallenged conditions, demonstrating the ALE strain improved resilience against selenite toxicity. Whole-genome sequencing revealed specific missense mutations in genes associated with anion transport and primary and secondary metabolisms in the ALE variant. These results were interpreted to show that some energy-demanding processes are attenuated in the ALE strain, prioritizing selenite bioprocessing to guarantee cell survival in the presence of selenite. The present study indicates some crucial points for adapting Kitasatospora sp. SeTe27 to selenite oxidative stress to best deal with selenium pollution. Moreover, the importance of exploring non-conventional bacterial genera, like Kitasatospora, for biotechnological applications is emphasized.

From micro to macro: Physical-chemical characterization of wheat starch-based films modified with PEG200, sodium citrate, or citric acid.

Needing to extend the shelf-life of packaged food and the evolving consumer demands led researchers to seek innovative, eco-friendly, and biocompatible packaging solutions. Starch is among the most promising natural and renewable alternatives to non-degradable plastics. Here, we deeply study the structural features of starch films modified by adding citric acid (CA) or sodium citrate (SC) as a cross-linker and polyethylene glycol 200 (PEG200) as a plasticizer and obtained through solvent casting. The substances' influence on starch films was evaluated through Attenuated Total Reflection Fourier Transform Infrared (ATR-FTIR) and Solid-state Nuclear Magnetic Resonance (ss-NMR) spectroscopies. Films' macroscopic properties, such as swelling index, solubility, thermo-mechanical features, and moisture absorption, were also assessed to foresee potential applications. Proper amounts of CA, CS, and PEG200 improve film properties and inhibit starch chains' retrogradation and recrystallization. Besides, the chemical neighbourhood of nuclei observed through ss-NMR significantly changed alongside the polymer chains' mobility. The latter result indicates a different polymer chain structural organization that could justify the film's higher resistance to thermal degradation and elongation at the break. This methodological approach is effective in predicting the macroscopic behaviour of a polymeric material and could be helpful for the application of such products in food preservation.

Impact of Biogenic and Chemogenic Selenium Nanoparticles on Model Eukaryotic Lipid Membranes.

Microbial nanotechnology is an expanding research area devoted to producing biogenic metal and metalloid nanomaterials (NMs) using microorganisms. Often, biogenic NMs are explored as antimicrobial, anticancer, or antioxidant agents. Yet, most studies focus on their applications rather than the underlying mechanism of action or toxicity. Here, we evaluate the toxicity of our well-characterized biogenic selenium nanoparticles (bSeNPs) produced by the Stenotrophomonas maltophilia strain SeITE02 against the model yeast Saccharomyces cerevisiae comparing it with chemogenic SeNPs (cSeNPs). Knowing from previous studies that the biogenic extract contained bSeNPs in an organic material (OM) and supported here by Fourier transform infrared spectroscopy, we removed and incubated it with cSeNPs (cSeNPs_OM) to assess its influence on the toxicity of these formulations. Specifically, we focused on the first stages of the eukaryotic cell exposure to these samples─i.e., their interaction with the cell lipid membrane, which was mimicked by preparing vesicles from yeast polar lipid extract or phosphatidylcholine lipids. Fluidity changes derived from biogenic and chemogenic samples revealed that the bSeNP extract mediated the overall rigidification of lipid vesicles, while cSeNPs showed negligible effects. The OM and cSeNPs_OM induced similar modifications to the bSeNP extract, reiterating the need to consider the OM influence on the physical-chemical and biological properties of bSeNP extracts.

Tolerance, Adaptation, and Cell Response Elicited by Micromonospora sp. Facing Tellurite Toxicity: A Biological and Physical-Chemical Characterization.

The intense use of tellurium (Te) in industrial applications, along with the improper disposal of Te-derivatives, is causing their accumulation in the environment, where oxyanion tellurite (TeO32-) is the most soluble, bioavailable, and toxic Te-species. On the other hand, tellurium is a rare metalloid element whose natural supply will end shortly with possible economic and technological effects. Thus, Te-containing waste represents the source from which Te should be recycled and recovered. Among the explored strategies, the microbial TeO32- biotransformation into less toxic Te-species is the most appropriate concerning the circular economy. Actinomycetes are ideal candidates in environmental biotechnology. However, their exploration in TeO32- biotransformation is scarce due to limited knowledge regarding oxyanion microbial processing. Here, this gap was filled by investigating the cell tolerance, adaptation, and response to TeO32- of a Micromonospora strain isolated from a metal(loid)-rich environment. To this aim, an integrated biological, physical-chemical, and statistical approach combining physiological and biochemical assays with confocal or scanning electron (SEM) microscopy and Fourier-transform infrared spectroscopy in attenuated total reflectance mode (ATR-FTIR) was designed. Micromonospora cells exposed to TeO32- under different physiological states revealed a series of striking cell responses, such as cell morphology changes, extracellular polymeric substance production, cell membrane damages and modifications, oxidative stress burst, protein aggregation and phosphorylation, and superoxide dismutase induction. These results highlight this Micromonospora strain as an asset for biotechnological purposes.

Differentiation among dairy products by combination of fast field cycling NMR relaxometry data and chemometrics.

A set of commercial milk and Sicilian cheeses was analysed by a combination of fast field cycling (FFC) nuclear magnetic resonance (NMR) relaxometry and chemometrics. The NMR dispersion (NMRD) curves were successfully analysed with a mathematical model applied on Parmigiano-Reggiano (PR) cheese. Regression parameters were led back to the molecular components of cheeses (water trapped in casein micelles, proteins and fats) and milk samples (water belonging to hydration shells around dispersed colloidal particles of different sizes and bulk water). The application of chemometric analysis on relaxometric data enabled differentiating milk from cheeses and revealing differences within the two sample groups of either cheeses or milk samples. Marked differences among cheeses were evidenced by statistical analysis of the sole quadrupolar peaks parameters, suggesting that these contain information on the nature of the milk used during cheese production. Hence, combination of FFC NMR and chemometrics represents a powerful tool to investigate alterations in dairy products.

Untargeted Metabolomics Investigation on Selenite Reduction to Elemental Selenium by Bacillus mycoides SeITE01.

Bacillus mycoides SeITE01 is an environmental isolate that transforms the oxyanion selenite ( SeO 3 2 - ) into the less bioavailable elemental selenium (Se0) forming biogenic selenium nanoparticles (Bio-SeNPs). In the present study, the reduction of sodium selenite (Na2SeO3) by SeITE01 strain and the effect of SeO 3 2 - exposure on the bacterial cells was examined through untargeted metabolomics. A time-course approach was used to monitor both cell pellet and cell free spent medium (referred as intracellular and extracellular, respectively) metabolites in SeITE01 cells treated or not with SeO 3 2 - . The results show substantial biochemical changes in SeITE01 cells when exposed to SeO 3 2 - . The initial uptake of SeO 3 2 - by SeITE01 cells (3h after inoculation) shows both an increase in intracellular levels of 4-hydroxybenzoate and indole-3-acetic acid, and an extracellular accumulation of guanosine, which are metabolites involved in general stress response adapting strategies. Proactive and defensive mechanisms against SeO 3 2 - are observed between the end of lag (12h) and beginning of exponential (18h) phases. Glutathione and N-acetyl-L-cysteine are thiol compounds that would be mainly involved in Painter-type reaction for the reduction and detoxification of SeO 3 2 - to Se0. In these growth stages, thiol metabolites perform a dual role, both acting against the toxic and harmful presence of the oxyanion and as substrate or reducing sources to scavenge ROS production. Moreover, detection of the amino acids L-threonine and ornithine suggests changes in membrane lipids. Starting from stationary phase (24 and 48h), metabolites related to the formation and release of SeNPs in the extracellular environment begin to be observed. 5-hydroxyindole acetate, D-[+]-glucosamine, 4-methyl-2-oxo pentanoic acid, and ethanolamine phosphate may represent signaling strategies following SeNPs release from the cytoplasmic compartment, with consequent damage to SeITE01 cell membranes. This is also accompanied by intracellular accumulation of trans-4-hydroxyproline and L-proline, which likely represent osmoprotectant activity. The identification of these metabolites suggests the activation of signaling strategies that would protect the bacterial cells from SeO 3 2 - toxicity while it is converting into SeNPs.

Biogenic Selenium Nanoparticles: A Fine Characterization to Unveil Their Thermodynamic Stability.

Among the plethora of available metal(loid) nanomaterials (NMs), those containing selenium are interesting from an applicative perspective, due to their high biocompatibility. Microorganisms capable of coping with toxic Se-oxyanions generate mostly Se nanoparticles (SeNPs), representing an ideal and green alternative over the chemogenic synthesis to obtain thermodynamically stable NMs. However, their structural characterization, in terms of biomolecules and interactions stabilizing the biogenic colloidal solution, is still a black hole that impairs the exploitation of biogenic SeNP full potential. Here, spherical and thermodynamically stable SeNPs were produced by a metal(loid) tolerant Micrococcus sp. Structural characterization obtained by Scanning Electron Microscopy (SEM) revealed that these SeNPs were surrounded by an organic material that contributed the most to their electrosteric stabilization, as indicated by Zeta (ζ) potential measurements. Proteins were strongly adsorbed on the SeNP surface, while lipids, polysaccharides, and nucleic acids more loosely interacted with SeNMs as highlighted by Fourier Transform Infrared Spectroscopy (FTIR) and overall supported by multivariate statistical analysis. Nevertheless, all these contributors were fundamental to maintain SeNPs stable, as, upon washing, the NM-containing extract showed the arising of aggregated SeNPs alongside Se nanorods (SeNRs). Besides, Density Functional Theory (DFT) calculation unveiled how thiol-containing molecules appeared to play a role in SeO32- bioreduction, stress oxidative response, and SeNP stabilization.

Processing of Metals and Metalloids by Actinobacteria: Cell Resistance Mechanisms and Synthesis of Metal(loid)-Based Nanostructures.

Metal(loid)s have a dual biological role as micronutrients and stress agents. A few geochemical and natural processes can cause their release in the environment, although most metal-contaminated sites derive from anthropogenic activities. Actinobacteria include high GC bacteria that inhabit a wide range of terrestrial and aquatic ecological niches, where they play essential roles in recycling or transforming organic and inorganic substances. The metal(loid) tolerance and/or resistance of several members of this phylum rely on mechanisms such as biosorption and extracellular sequestration by siderophores and extracellular polymeric substances (EPS), bioaccumulation, biotransformation, and metal efflux processes, which overall contribute to maintaining metal homeostasis. Considering the bioprocessing potential of metal(loid)s by Actinobacteria, the development of bioremediation strategies to reclaim metal-contaminated environments has gained scientific and economic interests. Moreover, the ability of Actinobacteria to produce nanoscale materials with intriguing physical-chemical and biological properties emphasizes the technological value of these biotic approaches. Given these premises, this review summarizes the strategies used by Actinobacteria to cope with metal(loid) toxicity and their undoubted role in bioremediation and bionanotechnology fields.

Volatile Compounds of Lemon and Grapefruit IntegroPectin.

An HS-SPME GC-MS analysis of the volatile compounds adsorbed at the outer surface of lemon and grapefruit pectins obtained via the hydrodynamic cavitation of industrial waste streams of lemon and grapefruit peels in water suggests important new findings en route to understanding the powerful and broad biological activity of these new pectic materials. In agreement with the ultralow degree of esterification of these pectins, the high amount of highly bioactive α-terpineol and terpinen-4-ol points to limonene (and linalool) decomposition catalyzed by residual citric acid in the citrus waste peel residue of the juice industrial production.

A combined physical-chemical and microbiological approach to unveil the fabrication, provenance, and state of conservation of the Kinkarakawa-gami art.

Kinkarakawa-gami wallpapers are unique works of art produced in Japan between 1870 and 1905 and exported in European countries, although only few examples are nowadays present in Europe. So far, neither the wallpapers nor the composing materials have been characterised, limiting the effective conservation-restoration of these artefacts accounting also for the potential deteriogen effects of microorganisms populating them. In the present study, four Kinkarakawa-gami wallpapers were analysed combining physical-chemical and microbiological approaches to obtain information regarding the artefacts' manufacture, composition, dating, and their microbial community. The validity of these methodologies was verified through a fine in blind statistical analysis, which allowed to identify trends and similarities within these important artefacts. The evidence gathered indicated that these wallpapers were generated between 1885 and 1889, during the so-called industrial production period. A wide range of organic (proteinaceous binders, natural waxes, pigments, and vegetable lacquers) and inorganic (tin foil and pigments) substances were used for the artefacts' manufacture, contributing to their overall complexity, which also reflects on the identification of a heterogeneous microbiota, often found in Eastern environmental matrices. Nevertheless, whether microorganisms inhabiting these wallpapers determined a detrimental or protective effect is not fully elucidated yet, thus constituting an aspect worth to be explored to deepen the knowledge needed for the conservation of Kinkarakawa-gami over time.

A New Water-Soluble Bactericidal Agent for the Treatment of Infections Caused by Gram-Positive and Gram-Negative Bacterial Strains.

Grapefruit and lemon pectin obtained from the respective waste citrus peels via hydrodynamic cavitation in water only are powerful, broad-scope antimicrobials against Gram-negative and -positive bacteria. Dubbed IntegroPectin, these pectic polymers functionalized with citrus flavonoids and terpenes show superior antimicrobial activity when compared to commercial citrus pectin. Similar to commercial pectin, lemon IntegroPectin determined ca. 3-log reduction in Staphylococcus aureus cells, while an enhanced activity of commercial citrus pectin was detected in the case of Pseudomonas aeruginosa cells with a minimal bactericidal concentration (MBC) of 15 mg mL-1. Although grapefruit and lemon IntegroPectin share equal MBC in the case of P. aeruginosa cells, grapefruit IntegroPectin shows boosted activity upon exposure of S. aureus cells with a 40 mg mL-1 biopolymer concentration affording complete killing of the bacterial cells. Insights into the mechanism of action of these biocompatible antimicrobials and their effect on bacterial cells, at the morphological level, were obtained indirectly through Fourier Transform Infrared spectroscopy and directly through scanning electron microscopy. In the era of antimicrobial resistance, these results are of great societal and sanitary relevance since citrus IntegroPectin biomaterials are also devoid of cytotoxic activity, as already shown for lemon IntegroPectin, opening the route to the development of new medical treatments of polymicrobial infections unlikely to develop drug resistance.

Biotechnology of Rhodococcus for the production of valuable compounds.

Bacteria belonging to Rhodococcus genus represent ideal candidates for microbial biotechnology applications because of their metabolic versatility, ability to degrade a wide range of organic compounds, and resistance to various stress conditions, such as metal toxicity, desiccation, and high concentration of organic solvents. Rhodococcus spp. strains have also peculiar biosynthetic activities that contribute to their strong persistence in harsh and contaminated environments and provide them a competitive advantage over other microorganisms. This review is focused on the metabolic features of Rhodococcus genus and their potential use in biotechnology strategies for the production of compounds with environmental, industrial, and medical relevance such as biosurfactants, bioflocculants, carotenoids, triacylglycerols, polyhydroxyalkanoate, siderophores, antimicrobials, and metal-based nanostructures. These biosynthetic capacities can also be exploited to obtain high value-added products from low-cost substrates (industrial wastes and contaminants), offering the possibility to efficiently recover valuable resources and providing possible waste disposal solutions. Rhodococcus spp. strains have also recently been pointed out as a source of novel bioactive molecules highlighting the need to extend the knowledge on biosynthetic capacities of members of this genus and their potential utilization in the framework of bioeconomy. KEY POINTS: • Rhodococcus possesses promising biosynthetic and bioconversion capacities. • Rhodococcus bioconversion capacities can provide waste disposal solutions. • Rhodococcus bioproducts have environmental, industrial, and medical relevance. Graphical abstract.

Influence of Bacterial Physiology on Processing of Selenite, Biogenesis of Nanomaterials and Their Thermodynamic Stability.

We explored how Ochrobactrum sp. MPV1 can convert up to 2.5 mM selenite within 120 h, surviving the challenge posed by high oxyanion concentrations. The data show that thiol-based biotic chemical reaction(s) occur upon bacterial exposure to low selenite concentrations, whereas enzymatic systems account for oxyanion removal when 2 mM oxyanion is exceeded. The selenite bioprocessing produces selenium nanomaterials, whose size and morphology depend on the bacterial physiology. Selenium nanoparticles were always produced by MPV1 cells, featuring an average diameter ranging between 90 and 140 nm, which we conclude constitutes the thermodynamic stability range for these nanostructures. Alternatively, selenium nanorods were observed for bacterial cells exposed to high selenite concentration or under controlled metabolism. Biogenic nanomaterials were enclosed by an organic material in part composed of amphiphilic biomolecules, which could form nanosized structures independently. Bacterial physiology influences the surface charge characterizing the organic material, suggesting its diverse biomolecular composition and its involvement in the tuning of the nanomaterial morphology. Finally, the organic material is in thermodynamic equilibrium with nanomaterials and responsible for their electrosteric stabilization, as changes in the temperature slightly influence the stability of biogenic compared to chemogenic nanomaterials.

Mesoporous Silica-Based Materials with Bactericidal Properties.

Bacterial infections are the main cause of chronic infections and even mortality. In fact, due to extensive use of antibiotics and, then, emergence of antibiotic resistance, treatment of such infections by conventional antibiotics has become a major concern worldwide. One of the promising strategies to treat infection diseases is the use of nanomaterials. Among them, mesoporous silica materials (MSMs) have attracted burgeoning attention due to high surface area, tunable pore/particle size, and easy surface functionalization. This review discusses how one can exploit capacities of MSMs to design and fabricate multifunctional/controllable drug delivery systems (DDSs) to combat bacterial infections. At first, the emergency of bacterial and biofilm resistance toward conventional antimicrobials is described and then how nanoparticles exert their toxic effects upon pathogenic cells is discussed. Next, the main aspects of MSMs (e.g., physicochemical properties, multifunctionality, and biosafety) which one should consider in the design of MSM-based DDSs against bacterial infections are introduced. Finally, a comprehensive analysis of all the papers published dealing with the use of MSMs for delivery of antibacterial chemicals (antimicrobial agents functionalized/adsorbed on mesoporous silica (MS), MS-loaded with antimicrobial agents, gated MS-loaded with antimicrobial agents, MS with metal-based nanoparticles, and MS-loaded with metal ions) is provided.

Aerobic Growth of Rhodococcus aetherivorans BCP1 Using Selected Naphthenic Acids as the Sole Carbon and Energy Sources.

Naphthenic acids (NAs) are an important group of toxic organic compounds naturally occurring in hydrocarbon deposits. This work shows that Rhodococcus aetherivorans BCP1 cells not only utilize a mixture of eight different NAs (8XNAs) for growth but they are also capable of marked degradation of two model NAs, cyclohexanecarboxylic acid (CHCA) and cyclopentanecarboxylic acid (CPCA) when supplied at concentrations from 50 to 500 mgL-1. The growth curves of BCP1 on 8XNAs, CHCA, and CPCA showed an initial lag phase not present in growth on glucose, which presumably was related to the toxic effects of NAs on the cell membrane permeability. BCP1 cell adaptation responses that allowed survival on NAs included changes in cell morphology, production of intracellular bodies and changes in fatty acid composition. Transmission electron microscopy (TEM) analysis of BCP1 cells grown on CHCA or CPCA showed a slight reduction in the cell size, the production of EPS-like material and intracellular electron-transparent and electron-dense inclusion bodies. The electron-transparent inclusions increased in the amount and size in NA-grown BCP1 cells under nitrogen limiting conditions and contained storage lipids as suggested by cell staining with the lipophilic Nile Blue A dye. Lipidomic analyses revealed significant changes with increases of methyl-branched (MBFA) and polyunsaturated fatty acids (PUFA) examining the fatty acid composition of NAs-growing BCP1 cells. PUFA biosynthesis is not usual in bacteria and, together with MBFA, can influence structural and functional processes with resulting effects on cell vitality. Finally, through the use of RT (Reverse Transcription)-qPCR, a gene cluster (chcpca) was found to be transcriptionally induced during the growth on CHCA and CPCA. Based on the expression and bioinformatics results, the predicted products of the chcpca gene cluster are proposed to be involved in aerobic NA degradation in R. aetherivorans BCP1. This study provides first insights into the genetic and metabolic mechanisms allowing a Rhodococcus strain to aerobically degrade NAs.

Assembly, growth and conductive properties of tellurium nanorods produced by Rhodococcus aetherivorans BCP1.

Tellurite (TeO32-) is a hazardous and toxic oxyanion for living organisms. However, several microorganisms can bioconvert TeO32- into the less toxic form of elemental tellurium (Te0). Here, Rhodococcus aetherivorans BCP1 resting (non-growing) cells showed the proficiency to produce tellurium-based nanoparticles (NPs) and nanorods (NRs) through the bioconversion of TeO32-, depending on the oxyanion initial concentration and time of cellular incubation. Te-nanostructures initially appeared in the cytoplasm of BCP1 cells as spherical NPs, which, as the exposure time increased, were converted into NRs. This observation suggested the existence of an intracellular mechanism of TeNRs assembly and growth that resembled the chemical surfactant-assisted process for NRs synthesis. The TeNRs produced by the BCP1 strain showed an average length (>700 nm) almost doubled compared to those observed in other studies. Further, the biogenic TeNRs displayed a regular single-crystalline structure typically obtained for those chemically synthesized. The chemical-physical characterization of the biogenic TeNRs reflected their thermodynamic stability that is likely derived from amphiphilic biomolecules present in the organic layer surrounding the NRs. Finally, the biogenic TeNRs extract showed good electrical conductivity. Thus, these findings support the suitability of this strain as eco-friendly biocatalyst to produce high quality tellurium-based nanomaterials exploitable for technological purposes.

Stability of biogenic metal(loid) nanomaterials related to the colloidal stabilization theory of chemical nanostructures.

In the last 15 years, the exploitation of biological systems (i.e. plants, bacteria, mycelial fungi, yeasts, and algae) to produce metal(loid) (Me)-based nanomaterials has been evaluated as eco-friendly and a cost-effective alternative to the chemical synthesis processes. Although the biological mechanisms of biogenic Me-nanomaterial (Bio-Me-nanomaterials) production are not yet completely elucidated, a key advantage of such bio-nanostructures over those chemically synthesized is related to their natural thermodynamic stability, with several studies ascribed to the presence of an organic layer surrounding these Bio-Me-nanostructures. Different macromolecules (e.g. proteins, peptides, lipids, DNA, and polysaccharides) or secondary metabolites (e.g. flavonoids, terpenoids, glycosides, organic acids, and alkaloids) naturally produced by organisms have been indicated as main contributors to the stabilization of Bio-Me-nanostructures. Nevertheless, the chemical-physical mechanisms behind the ability of these molecules in providing stability to Bio-Me-nanomaterials are unknown. In this context, transposing the stabilization theory of chemically synthesized Me-nanomaterials (Ch-Me-nanomaterials) to biogenic materials can be used towards a better comprehension of macromolecules and secondary metabolites role as stabilizing agents of Bio-Me-nanomaterials. According to this theory, nanomaterials are generally featured by high thermodynamic instability in suspension, due to their high surface area and surface energy. This feature leads to the necessity to stabilize chemical nanostructures, even during or directly after their synthesis, through the development of (i) electrostatic, (ii) steric, or (iii) electrosteric interactions occurring between molecules and nanomaterials in suspension. Based on these three mechanisms, this review is focused on parallels between the stabilization of biogenic or chemical nanomaterials, suggesting which chemical-physical mechanisms may be involved in the natural stability of Bio-Me-nanomaterials. As a result, macromolecules such as DNA, polyphosphates and proteins may electrostatically interact with Bio-Me-nanomaterials in suspension through their charged moieties, showing the same properties of counterions in Ch-Me-nanostructure suspensions. Since several biomolecules (e.g. neutral lipids, nonionic biosurfactants, polysaccharides, and secondary metabolites) produced by metal(loid)-grown organisms can develop similar steric hindrance as compared to nonionic amphiphilic surfactants and block co-polymers generally used to sterically stabilize Ch-Me-nanomaterials. These biomolecules, most likely, are involved in the development of steric stabilization, because of their bulky structures. Finally, charged lipids and polysaccharides, ionic biosurfactants or proteins with amphiphilic properties can exert a dual effect (i.e. electrostatic and steric repulsion interactions) in the contest of Bio-Me-nanomaterials, leading to the high degree of stability observed.

Physical-Chemical Properties of Biogenic Selenium Nanostructures Produced by Stenotrophomonas maltophilia SeITE02 and Ochrobactrum sp. MPV1.

Stenotrophomonas maltophilia SeITE02 and Ochrobactrum sp. MPV1 were isolated from the rhizosphere soil of the selenium-hyperaccumulator legume Astragalus bisulcatus and waste material from a dumping site for roasted pyrites, respectively. Here, these bacterial strains were studied as cell factories to generate selenium-nanostructures (SeNS) under metabolically controlled growth conditions. Thus, a defined medium (DM) containing either glucose or pyruvate as carbon and energy source along with selenite () was tested to evaluate bacterial growth, oxyanion bioconversion and changes occurring in SeNS features with respect to those generated by these strains grown on rich media. Transmission electron microscopy (TEM) images show extra- or intra-cellular emergence of SeNS in SeITE02 or MPV1 respectively, revealing the presence of two distinct biological routes of SeNS biogenesis. Indeed, the stress exerted by upon SeITE02 cells triggered the production of membrane vesicles (MVs), which surrounded Se-nanoparticles (SeNPsSeITE02-G_e and SeNPsSeITE02-P_e with average diameter of 179 ± 56 and 208 ± 60 nm, respectively), as highlighted by TEM and scanning electron microscopy (SEM), strongly suggesting that MVs might play a crucial role in the excreting mechanism of the SeNPs in the extracellular environment. On the other hand, MPV1 strain biosynthesized intracellular inclusions likely containing hydrophobic storage compounds and SeNPs (123 ± 32 nm) under pyruvate conditioning, while the growth on glucose as the only source of carbon and energy led to the production of a mixed population of intracellular SeNPs (118 ± 36 nm) and nanorods (SeNRs; average length of 324 ± 89). SEM, fluorescence spectroscopy, and confocal laser scanning microscopy (CLSM) revealed that the biogenic SeNS were enclosed in an organic material containing proteins and amphiphilic molecules, possibly responsible for the high thermodynamic stability of these nanomaterials. Finally, the biogenic SeNS extracts were photoluminescent upon excitation ranging from 380 to 530 nm, whose degree of fluorescence emission (λem = 416-640 nm) was comparable to that from chemically synthesized SeNPs with L-cysteine (L-cys SeNPs). This study offers novel insights into the formation, localization, and release of biogenic SeNS generated by two different Gram-negative bacterial strains under aerobic and metabolically controlled growth conditions. The work strengthens the possibility of using these bacterial isolates as eco-friendly biocatalysts to produce high quality SeNS targeted to possible biomedical applications and other biotechnological purposes.

Biosynthesis of selenium-nanoparticles and -nanorods as a product of selenite bioconversion by the aerobic bacterium Rhodococcus aetherivorans BCP1.

The wide anthropogenic use of selenium compounds represents the major source of selenium pollution worldwide, causing environmental issues and health concerns. Microbe-based strategies for metal removal/recovery have received increasing interest thanks to the association of the microbial ability to detoxify toxic metal/metalloid polluted environments with the production of nanomaterials. This study investigates the tolerance and the bioconversion of selenite (SeO32-) by the aerobically grown Actinomycete Rhodococcus aetherivorans BCP1 in association with its ability to produce selenium nanoparticles and nanorods (SeNPs and SeNRs). The BCP1 strain showed high tolerance towards SeO32- with a Minimal Inhibitory Concentration (MIC) of 500mM. The bioconversion of SeO32- was evaluated considering two different physiological states of the BCP1 strain, i.e. unconditioned and/or conditioned cells, which correspond to cells exposed for the first time or after re-inoculation in fresh medium to either 0.5 or 2mM of Na2SeO3, respectively. SeO32- bioconversion was higher for conditioned grown cells compared to the unconditioned ones. Selenium nanostructures appeared polydisperse and not aggregated, as detected by electron microscopy, being embedded in an organic coating likely responsible for their stability, as suggested by the physical-chemical characterization. The production of smaller and/or larger SeNPs was influenced by the initial concentration of provided precursor, which resulted in the growth of longer and/or shorter SeNRs, respectively. The strong ability to tolerate high SeO32- concentrations coupled with SeNP and SeNR biosynthesis highlights promising new applications of Rhodococcus aetherivorans BCP1 as cell factory to produce stable Se-nanostructures, whose suitability might be exploited for biotechnology purposes.

Ochrobactrum sp. MPV1 from a dump of roasted pyrites can be exploited as bacterial catalyst for the biogenesis of selenium and tellurium nanoparticles.

BACKGROUND: Bacteria have developed different mechanisms for the transformation of metalloid oxyanions to non-toxic chemical forms. A number of bacterial isolates so far obtained in axenic culture has shown the ability to bioreduce selenite and tellurite to the elemental state in different conditions along with the formation of nanoparticles-both inside and outside the cells-characterized by a variety of morphological features. This reductive process can be considered of major importance for two reasons: firstly, toxic and soluble (i.e. bioavailable) compounds such as selenite and tellurite are converted to a less toxic chemical forms (i.e. zero valent state); secondly, chalcogen nanoparticles have attracted great interest due to their photoelectric and semiconducting properties. In addition, their exploitation as antimicrobial agents is currently becoming an area of intensive research in medical sciences. RESULTS: In the present study, the bacterial strain Ochrobactrum sp. MPV1, isolated from a dump of roasted arsenopyrites as residues of a formerly sulfuric acid production near Scarlino (Tuscany, Italy) was analyzed for its capability of efficaciously bioreducing the chalcogen oxyanions selenite (SeO32-) and tellurite (TeO32-) to their respective elemental forms (Se0 and Te0) in aerobic conditions, with generation of Se- and Te-nanoparticles (Se- and TeNPs). The isolate could bioconvert 2 mM SeO32- and 0.5 mM TeO32- to the corresponding Se0 and Te0 in 48 and 120 h, respectively. The intracellular accumulation of nanomaterials was demonstrated through electron microscopy. Moreover, several analyses were performed to shed light on the mechanisms involved in SeO32- and TeO32- bioreduction to their elemental states. Results obtained suggested that these oxyanions are bioconverted through two different mechanisms in Ochrobactrum sp. MPV1. Glutathione (GSH) seemed to play a key role in SeO32- bioreduction, while TeO32- bioconversion could be ascribed to the catalytic activity of intracellular NADH-dependent oxidoreductases. The organic coating surrounding biogenic Se- and TeNPs was also characterized through Fourier-transform infrared spectroscopy. This analysis revealed interesting differences among the NPs produced by Ochrobactrum sp. MPV1 and suggested a possible different role of phospholipids and proteins in both biosynthesis and stabilization of such chalcogen-NPs. CONCLUSIONS: In conclusion, Ochrobactrum sp. MPV1 has demonstrated to be an ideal candidate for the bioconversion of toxic oxyanions such as selenite and tellurite to their respective elemental forms, producing intracellular Se- and TeNPs possibly exploitable in biomedical and industrial applications.