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Summary Tailored nanoparticles offer a novel approach to fight antibiotic‐resistant microorganisms. We analysed biogenic selenium nanoparticles (SeNPs) of bacterial origin to determine their antimicrobial activity against selected pathogens in their planktonic and biofilm states. SeNPs synthesized by Gram‐negative Stenotrophomonas maltophilia [Sm‐SeNPs(−)] and Gram‐positive Bacillus mycoides [Bm‐SeNPs(+)] 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 SeNPs were able to inhibit biofilm formation and also to disaggregate the mature glycocalyx in both P. aeruginosa and Candida spp. The Sm‐SeNPs(−) and Bm‐SeNPs(+) both achieved much stronger antimicrobial effects than synthetic selenium nanoparticles (Ch‐SeNPs). Dendritic cells and fibroblasts exposed to Sm‐SeNPs(−), Bm‐SeNPs(+) and Ch‐SeNPs 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 SeNPs 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. Biogenic SeNPs exhibited a higher antimicrobial and biofilm eradicating activity than chemically synthesized SeNPs. Moreover, they did not show toxicity against fibroblasts and dendritic cells.

Accurate identification and typing of microbes are crucial steps in gaining an awareness of the biological heterogeneity and reliability of microbial material within any proprietary or public collection. Paenibacillus polymyxa is a bacterial species of great agricultural and industrial importance due to its plant growth-promoting activities and production of several relevant secondary metabolites. In recent years, matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) has been widely used as an alternative rapid tool for identifying, typing, and differentiating closely related strains. In this study, we investigated the diversity of three P. polymyxa strains. The mass spectra of ATCC 842 T , DSM 292, and DSM 365 were obtained, analysed, and compared to select discriminant peaks using ClinProTools software and generate classification models. MALDI-TOF MS analysis showed inconsistent results in identifying DSM 292 and DSM 365 as belonging to P. polimixa species, and comparative analysis of mass spectra revealed the presence of highly discriminatory biomarkers among the three strains. 16S rRNA sequencing and Average Nucleotide Identity (ANI) confirmed the discrepancies found in the proteomic analysis. The case study presented here suggests the enormous potential of the proteomic-based approach, combined with statistical tools, to predict and explore differences between closely related strains in large microbial datasets.

A Gram-negative bacterial strain, namely Sel9 , was isolated from a sequencing batch reactor for the selection of a polyhydroxyalkanoate (PHA)-storing microbial biomass, fed with volatile fatty acids. 16S rRNA gene sequence and core genome analyses performed with maximum likelihood method evidenced that Sel9 belongs to the genus Thauera with the highest phylogenetic relatedness with Thauera butanivorans DSM 2080 (98.99%) and Thauera linaloolentis DSM 12138 (98.49%). Digital DNA-DNA hybridization and Average Nucleotide Identity (OrthoANI) values between strain Sel9 and the closest taxon, T. butanivorans DSM 2080 , were 53.20 and 93.71%, respectively, which were below the cut-off values for species delineation. The predominant cell fatty acids were summed feature 3 (C16:1 ω6c/C16:1 ω7c), C16:0 and summed feature 8 (C18:1 ω6c/C18:1 ω7c). Phosphatidylethanolamine and phosphatidylglycerol were the main polar lipids in the cell. Genome mining detected nine biosynthetic gene clusters, including ectoine, pyrroloquinoline quinone (PQQ)-redox and a genus-rare nonribosomal peptide synthetase (NRPS) gene cluster, plus the acyclic terpene utilization pathway predicting growth on linalool. The combination of phylogenetic, chemotaxonomic and phenotypic features led to consider strain Sel9 as a representative of a novel species within the genus Thauera. Therefore, given its remarkable ability to store carbon sources, for the type strain Sel9 (=LMG 33225 =BAC RE RB 2381 =VUCC 376 ) the name of Thauera carbonocopians sp. nov. is here proposed. Eventually, this study represents the first comprehensive investigation of biosynthetic gene clusters and the comparative genomics analysis of PHA metabolism within the genus Thauera.

Background Biosurfactants are surface-active compounds with environmental and industrial applications. These molecules show higher biocompatibility, stability and efficiency compared to synthetic surfactants. On the other hand, biosurfactants are not cost-competitive to their chemical counterparts. Cost effective technology such as the use of low-cost substrates is a promising approach aimed at reducing the production cost. This study aimed to evaluate the biosurfactant production and activity by the novel strain Rhodococcus sp. SP1d by using different growth substrates. Therefore, to exploit the biosurfactant synthesized by SP1d for environmental applications, the effect of this compound on the bacteria biofilm formation was evaluated. Eventually, for a possible bioremediation application, the biosurfactant properties and its chemical characteristics were investigated using diesel as source of carbon. Results Rhodococcus sp. SP1d evidence the highest similarity to Rhodococcus globerulus DSM 43954 T and the ability to biosynthesize surfactants using a wide range of substrates such as exhausted vegetable oil, mineral oil, butter, n-hexadecane, and diesel. The maximum production of crude biosurfactant after 10 days of incubation was reached on n-hexadecane and diesel with a final yield of 2.38 ± 0.51 and 1.86 ± 0.31 g L − 1 respectively. Biosurfactants produced by SP1d enhanced the biofilm production of P. protegens MP12. Moreover, the results showed the ability of SP1d to produce biosurfactants on diesel even when grown at 10 and 18 °C. The biosurfactant activity was maintained over a wide range of NaCl concentration, pH, and temperature. A concentration of 1000 mg L − 1 of the crude biosurfactant showed an emulsification activity of 55% towards both xylene and olive oil and a reduction of 25.0 mN m − 1 of surface tension of water. Eventually, nuclear magnetic resonance spectroscopy indicated that the biosurfactant is formed by trehalolipids. Conclusions The use of low-cost substrates such as exhausted oils and waste butter reduce both the costs of biosurfactant synthesis and the environmental pollution due to the inappropriate disposal of these residues. High production yields, stability and emulsification properties using diesel and n-hexadecane as substrates, make the biosurfactant produced by SP1d a sustainable biocompound for bioremediation purpose. Eventually, the purified biosurfactant improved the biofilm formation of the fungal antagonistic strain P. protegens MP12, and thus seem to be exploitable to increase the adherence and colonization of plant surfaces by this antagonistic strain and possibly enhance antifungal activity.

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 (SeO 3 2− ) and tellurite (TeO 3 2− ) to their respective elemental forms (Se 0 and Te 0 ) in aerobic conditions, with generation of Se- and Te-nanoparticles (Se- and TeNPs). The isolate could bioconvert 2 mM SeO 3 2− and 0.5 mM TeO 3 2− to the corresponding Se 0 and Te 0 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 SeO 3 2− and TeO 3 2− 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 SeO 3 2− bioreduction, while TeO 3 2− 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.

A greenhouse pot experiment was carried out to evaluate the efficiency of arsenic phytoextraction by the fern Pteris vittata growing in arsenic-contaminated soil, with or without the addition of selected rhizobacteria isolated from the polluted site. The bacterial strains were selected for arsenic resistance, the ability to reduce arsenate to arsenite, and the ability to promote plant growth. P. vittata plants were cultivated for 4 months in a contaminated substrate consisting of arsenopyrite cinders and mature compost. Four different experimental conditions were tested: (i) non-inoculated plants; (ii) plants inoculated with the siderophore-producing and arsenate-reducing bacteria Pseudomonas sp. P1III2 and Delftia sp. P2III5 (A); (iii) plants inoculated with the siderophore and indoleacetic acid-producing bacteria Bacillus sp. MPV12, Variovorax sp. P4III4, and Pseudoxanthomonas sp. P4V6 (B), and (iv) plants inoculated with all five bacterial strains (AB). The presence of growth-promoting rhizobacteria increased plant biomass by up to 45% and increased As removal efficiency from 13% without bacteria to 35% in the presence of the mixed inoculum. Molecular analysis confirmed the persistence of the introduced bacterial strains in the soil and resulted in a significant impact on the structure of the bacterial community.

Background Selenite (SeO 3 2− ) oxyanion shows severe toxicity to biota. Different bacterial strains exist that are capable of reducing SeO 3 2− to non-toxic elemental selenium (Se 0 ), 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 . Results 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 SeO 3 2− within 12 and 24 hours, respectively. The resultant Se 0 aggregated to form nanoparticles and the amount of Se 0 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 SeO 3 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 SeO 3 2− reductase capability, revealing that enzymatic activity was mainly associated with the membrane fraction. Reduction of SeO 3 2− was also detected in the supernatant of bacterial cultures upon NADH addition. Conclusions 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 Se 0 precipitation and SeNPs assembly are suggested. SeO 3 2− is proposed to be enzimatically reduced to Se 0 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 SeO 3 2− reduction. Formation of SeNPs seems to be the result of an Ostwald ripening mechanism.

(i) In this study we propose new protocols for quantifying the concentration of carbohydrates, proteins, and lipids present in capping layers of biogenic selenium nanoparticles. We applied them to compare BioSeNPs produced by five different bacterial strains. (ii) A hypothesis to describe capping layer composition of the bacterial SeNPs is suggested: some biomolecules are bound more strongly than others to the core metalloid matrix, so that the diverse capping layer components differentially contribute to the overall structural characteristics of the nanostructures. (iii) The application of the approach here in combining quantification of cap‐associated biomolecules with the measurement of structural integrity related parameters, can give the biogenic nanomaterial field useful information to construct a data bank on biogenic nanomaterials. Summary Biogenic metal/metalloid nanoparticles of microbial origin retain a functional biomolecular capping layer that confers structural stability. Little is known about the composition of such capping material. In this study, selenium nanoparticles (SeNPs) synthesized by five different bacterial strains underwent comparative analysis with newly proposed protocols for quantifying the concentration of carbohydrates, proteins and lipids present in capping layers. SeNPs were therefore treated with two different detergents to remove portions of the surrounding caps in order to assess the resulting effects. Capping material quantification was carried out along with the measure of parameters such as hydrodynamic diameter, polydispersity and surface charge. SeNPs from the five strains showed differences in their distinct biomolecule ratios. On the other hand, structural changes in the nanoparticles induced by detergents did not correlate with the amounts of capping matrix removed. Thus, the present investigation suggests a hypothesis to describe capping layer composition of the bacterial SeNPs: some biomolecules are bound more strongly than others to the core metalloid matrix, so that the diverse capping layer components differentially contribute to the overall structural characteristics of the nanoparticles. Furthermore, the application of the approach here in combining quantification of cap‐associated biomolecules with the measurement of structural integrity‐related parameters can give the biogenic nanomaterial field useful information to construct a data bank on biogenically synthesized nanostructures.

PFASs (perfluoroalkyl and polyfluoroalkyl substances) are highly fluorinated, aliphatic, synthetic compounds with high thermal and chemical stability as well as unique amphiphilic properties which make them ingredients in a range of industrial processes. PFASs have attracted consideration due to their persistence, toxicity and bioaccumulation tendency in the environment. Recently, attention has begun to be addressed to shorter-chain PFASs, such as perfluorohexane sulfonate [PFHxS], apparently less toxic to and more easily eliminated from lab animals. However, short-chain PFASs represent end-products from the transformation of fluorotelomers whose biotic breakdown reactions have not been identified to date. This means that such emergent pollutants will tend to accumulate and persist in ecosystems. Since we are just learning about the interaction between short-chain PFASs and microorganisms, this study reports on the response to PFHxS of two Pseudomonas sp. strains isolated from environmental matrices contaminated by PFASs. The PFHxS bioaccumulation potential of these strains was unveiled by exploiting different physiological conditions as either axenic or mixed cultures under alkanothrofic settings. Moreover, electron microscopy revealed nonorthodox features of the bacterial cells, as a consequence of the stress caused by both organic solvents and PFHxS in the culturing substrate.

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.