Identification of a TonB-Dependent Receptor Involved in Lanthanide Switch by the Characterization of Laboratory-Adapted Methylosinus trichosporium OB3b.

Two methanol dehydrogenases (MDHs), MxaFI and XoxF, have been characterized in methylotrophic and methanotrophic bacteria. MxaFI contains a calcium ion in its active site, whereas XoxF contains a lanthanide ion. Importantly, the expression of MxaFI and XoxF is inversely regulated by lanthanide bioavailability, i.e., the "lanthanide switch." To reveal the genetic and environmental factors affecting the lanthanide switch, we focused on two Methylosinus trichosporium OB3b mutants isolated during routine cultivation. In these mutants, MxaF was constitutively expressed, but lanthanide-dependent XoxF1 was not, even in the presence of 25 µM cerium ions, which is sufficient for XoxF expression in the wild type. Genotyping showed that both mutants harbored a loss-of-function mutation in the CQW49_RS02145 gene, which encodes a TonB-dependent receptor. Gene disruption and complementation experiments demonstrated that CQW49_RS02145 was required for XoxF1 expression in the presence of 25 µM cerium ions. Phylogenetic analysis indicated that CQW49_RS02145 was homologous to the Methylorubrum extorquens AM1 lanthanide transporter gene (lutH). These findings suggest that CQW49_RS02145 is involved in lanthanide uptake across the outer membrane. Furthermore, we demonstrated that supplementation with cerium and glycerol caused severe growth arrest in the wild type. CQW49_RS02145 underwent adaptive laboratory evolution in the presence of cerium and glycerol ions, resulting in a mutation that partially mitigated the growth arrest. This finding implies that loss-of-function mutations in CQW49_RS02145 can be attributed to residual glycerol from the frozen stock. IMPORTANCE Lanthanides are widely used in many industrial applications, including catalysts, magnets, and polishing. Recently, lanthanide-dependent metabolism was characterized in methane-utilizing bacteria. Despite the global demand for lanthanides, few studies have investigated the mechanism of lanthanide uptake by these bacteria. In this study, we identify a lanthanide transporter in Methylosinus trichosporium OB3b and indicate the potential interaction between intracellular lanthanide and glycerol. Understanding the genetic and environmental factors affecting lanthanide uptake should not only help improve the use of lanthanides for the bioconversion of methane into valuable products like methanol but also be of value for developing biomining to extract lanthanides under neutral conditions.

Targeting microbial biofilms: by Arctium lappa l. synthesised biocompatible CeO2-NPs encapsulated in nano-chitosan.

This study is planned to synthesise new biocompatible, nano antimicrobial formulation against biofilm producing strains. Aqueous root extract of Arctium lappa l. was used to synthesise ceria nanoparticles (CeO2-NPs). The synthesised nanoparticles were encapsulated with nano-chitosan by sol-gel method and characterised using standard techniques. Gas chromatography-mass spectrometer of Arctium lappa l. revealed the presence of ethanol, acetone, 1- propanol, 2-methylethane, 1,1-di-ethoxy, 1-Butanol, and oleic acid acted as reducing and surface stabilising agents for tailoring morphology of CeO2-NPs. Erythrocyte integrity after treatment with synthesised nanomaterials was evaluated by spectrophotometer measurement of haemoglobin release having biocompatibility. Scanning electron microscopy revealed the formation of mono dispersed beads shaped particles with mean particle size of 26.2 nm. X-ray diffractometry revealed cubic crystalline structure having size of 28.0 nm. After encapsulation by nano-chitosan, the size of CeO2-NPs enhances to 48.8 nm making average coverage of about 22.6 nm. The synthesised nanomaterials were found effective to disrupt biofilm of S. aureus and P. aeruginosa. Interestingly, encapsulated CeO2-NPs revealed powerful antibacterial and biofilm disruption activity examined by fluorescent live/dead staining using confocal laser scanning microscopy. The superior antibacterial activities exposed by encapsulated CeO2-NPs lead to the conclusion that they could be useful for controlling biofilm producing multidrug resistance pathogens.

Uptake, translocation and impact of green synthesized nanoceria on growth and antioxidant enzymes activity of Solanum lycopersicum L.

Cerium oxide nanoparticles (nanoceria) were synthesized by a novel, simple green chemistry procedure using Elaeagnus angustifolia leaf extract as a reducing and capping agent. The crystalline nature of nanoceria was confirmed by XRD analysis. FTIR analysis revealed that phytochemicals are present on the surface of nanoceria. SEM and TEM images revealed that the nanoceria are well dispersed, spherical in shape with a particle size range in between 30 and 75 nm. Thereafter, the effects of various concentrations of cerium oxide (CeO2) and green synthesized nanoceria on growth and metabolism of Solanum lycopersicum (tomato) were investigated. The bio-accumulation of Ce in tomato seedlings was found to be dose dependent and the results showed that with the increase in exposure concentrations, the accumulation of Ce contents in both root and shoots augmented. However, unlike nanoceria treated seedlings, Ce contents in the roots with CeO2 treatments were negligible than that in the shoots at lower concentrations and this suggested the immobilization of Ce in CeO2 treatment at lower concentrations. Nanoceria at 500 and 1000 mg/L resulted in inhibitory effect on growth of test plant as compared to CeO2 component. The exposure of plants to nanoceria and CeO2 has resulted in significant reduction in pigment content, increased LP, EL and H2O2 content. The activities of antioxidant enzymes viz. SOD, CAT, APX and GPX were significantly up regulated on exposure of nanoceria and CeO2. It is concluded that plant exposure with nanoceria at concentrations of 20 and 100 mg/L were more beneficial for growth and metabolism of tomato plants than that of CeO2 at equivalent concentrations.

Dietary nano cerium oxide promotes growth, relieves ammonia nitrogen stress, and improves immunity in crab (Eriocheir sinensis).

Oxidative stress plays a crucial role in ammonia nitrogen toxicity. In this study, the beneficial effects of dietary nano cerium oxide (nano CeO2) as a potent antioxidant were examined in the Chinese mitten crab (Eriocheir sinensis). Crabs were fed a diet supplemented with 0, 0.2, 0.4, 0.8, 1.6, 3.2, 6.4, or 12.8 mg/kg nano CeO2 for 60 d. The optimum supplementation level of nano CeO2 that significantly increased weight gain rate and decreased feed coefficient was 0.8 mg/kg. This level also offered immune protection when crabs were kept under ammonia nitrogen stress and/or exposed to pathogen infection (Aeromonas hydrophila). Supplementation with 0.8 mg/kg of CeO2 (i) relieved pathological damage to the hepatopancreas; (ii) increased hemocyte counts, including total number of hemocytes, granulocytes, and hyalinocytes; (iii) decreased malondialdehyde content and increased antioxidant enzyme activities of superoxide dismutase and catalase in the hemolymph; (iv) increased the activities of lysozyme, acid phosphatase, and alkaline phosphatase in the hemolymph; and (v) increased gene and protein expression of cathepsin L in the hepatopancreas. Mortality increased when crabs were injected with bacteria under ammonia nitrogen stress, but dietary supplementation with 0.8 mg/kg nano CeO2 decreased the mortality rate. Thus, the results of this study suggested that dietary supplementation with nano CeO2 in crabs promoted growth and up-regulated immunity to bacterial infection under ammonia nitrogen stress.

Effect of cerium on growth and antioxidant metabolism of Lemna minor L.

An increasing input rate of rare earth elements in the environment is expected because of the intense extraction of such elements form their ores to face human technological needs. In this study Lemna minor L. plants were grown under laboratory conditions and treated with increasing concentrations of cerium (Ce) ions to investigate the effects on plant growth and antioxidant systems. The growth increased in plants treated with lower Ce concentrations and reduced in plants treated with higher concentrations, compared to control plants. In plants treated with higher Ce concentrations lower levels of chlorophyll and carotenoid and the appearance of chlorotic symptoms were also detected. Increased levels of hydrogen peroxide, antioxidant metabolites and antioxidant activity confirmed that higher Ce concentrations are toxic to L. minor. Ce concentration in plant tissues was also determined and detectable levels were found only in plants grown on Ce-supplemented media. The use of duckweed plants as a tool for biomonitoring of Ce in freshwater is discussed.

Influence of CeO2 and ZnO nanoparticles on cucumber physiological markers and bioaccumulation of Ce and Zn: a life cycle study.

With the dramatic increase in nanotechnologies, it has become increasingly likely that food crops will be exposed to excess engineered nanoparticles (NPs). In this study, cucumber plants were grown to full maturity in soil amended with either CeO2 or ZnO NPs at concentrations of 0, 400, and 800 mg/kg. Chlorophyll and gas exchange were monitored, and physiological markers were recorded. Results showed that, at the concentrations tested, neither CeO2 nor ZnO NPs impacted cucumber plant growth, gas exchange, and chlorophyll content. However, at 800 mg/kg treatment, CeO2 NPs reduced the yield by 31.6% compared to the control (p ≤ 0.07). ICP-MS results showed that the high concentration treatments resulted in the bioaccumulation of Ce and Zn in the fruit (1.27 mg of Ce and 110 mg Zn per kg dry weight). µ-XRF images exhibited Ce in the leaf vein vasculature, suggesting that Ce moves between tissues with water flow during transpiration. To the authors' knowledge, this is the first holistic study focusing on the impacts of CeO2 and ZnO NPs in the life cycle of cucumber plants.

Cerium binding activity of pectins isolated from the seagrasses Zostera marina and Phyllospadix iwatensis.

Cerium binding activity of three different water soluble pectin compounds of different origin was studied in a batch sorption system. The Langmuir, Freundlich and BET sorption models were adopted to describe the binding reactions between metal ions and pectin molecules. The Langmuir model provided the best fit. Within the pH range from 4.0 to 6.0, the largest amount of the cerium ions was bound by pectin isolated from the seagrass Phylospadix iwatensis in comparison to pectin extracted from the seagrass Zostera marina and pectin obtained from citrus peel (commercial grade). The Langmuir constants were also highest for the pectin samples isolated from the seagrass P. iwatensis. The results obtained from this study suggest that pectin is a prospective source for the development of radioisotope-removing pharmaceuticals.

Interplay between Fe 3d and Ce 4f magnetism and Kondo interaction in CeFeAs(1-x)P(x)O probed by 75As and 31P NMR.

A detailed (31)P (I = 1/2) and (75)As (I = 3/2) NMR study on polycrystalline CeFeAs(1-x)P(x)O alloys is presented. The magnetism of CeFeAsO changes drastically upon P substitution on the As site. CeFePO is a heavy fermion system without long-range order whereas CeFeAsO exhibits an Fe 3d SDW type of ordering accompanied by a structural transition from tetragonal (TT) to orthorhombic (OT) structure. Furthermore, Ce 4f(1) orders antiferromagnetically (AFM) at low temperature. At the critical concentration where the Fe magnetism is diminished the Ce-Ce interaction changes to a ferromagnetic (FM) type of ordering. Three representative samples of the CeFeAs(1-x)P(x)O (x = 0.05, 0.3 and 0.9) series are systematically investigated. (1) For the x = 0.05 alloy a drastic change of the linewidth at 130 K indicates the AFM-SDW type of ordering of Fe and the structural change from the TT to the OT phase. The linewidth roughly measures the internal field in the ordered state and the transition is most likely first order. The small and nearly constant shift from (31)P and (75)As NMR suggests the presence of competing hyperfine interactions between the nuclear spins and the 4f and 3d ions of Ce and Fe. (2) For the x = 0.3 alloy, the evolution of the Fe-SDW type of order takes place at around 70 K corroborating the results of bulk measurement and µSR. Here we found evidence for phase separation of paramagnetic and magnetic SDW phases. (3) In contrast to the heavy fermion CeFePO for the x = 0.9 alloy a phase transition is found at 2 K. The field-dependent NMR shift gives evidence of FM ordering. Above the ordering the spin-lattice relaxation rate (31)(1/T(1)) shows unconventional, non-Korringa-like behaviour which indicates a complex interplay of Kondo and FM fluctuations.

Competitive inhibition and selectivity enhancement by Ca in the uptake of inorganic elements (Be, Na, Mg, K, Ca, Sc, Mn, Co, Zn, Se, Rb, Sr, Y, Zr, Ce, Pm, Gd, Hf) by carrot (Daucus carota cv. U.S. harumakigosun).

We investigated the uptake of inorganic elements (Be, Na, Mg, K, Ca, Sc, Mn, Co, Zn, Se, Rb, Sr, Y, Zr, Ce, Pm, Gd, and Hf) and the effect of Ca on their uptake in carrots (Daucus carota cv. U.S. harumakigosun) by the radioactive multitracer technique. The experimental results suggested that Na, Mg, K, and Rb competed for the functional groups outside the cells in roots with Ca but not for the transporter-binding sites on the plasma membrane of the root cortex cells. In contrast, Y, Ce, Pm, and Gd competed with Ca for the transporters on the plasma membrane. The selectivity, which was defined as the value obtained by dividing the concentration ratio of an elemental pair, K/Na, Rb/Na, Be/Sr, and Mg/Sr, in the presence of 0.2 and 2 ppm Ca by that of the corresponding elemental pair in the absence of Ca in the solution was estimated. The selectivity of K and Rb in roots was increased in the presence of Ca. The selectivity of Be in roots was not affected, whereas the selectivity of Mg was increased by Ca. These observations suggest that the presence of Ca in the uptake solution enhances the selectivity in the uptake of metabolically important elements against unwanted elements.

Site-selective and hydrolytic two-strand scission of double-stranded DNA using Ce(IV)/EDTA and pseudo-complementary PNA.

By combining Ce(IV)/EDTA with two pseudo-complementary peptide nucleic acids (pcPNAs), both strands in double-stranded DNA were site-selectively hydrolyzed at the target site. Either plasmid DNA (4361 bp) or its linearized form was used as the substrate. When two pcPNAs invaded into the double-stranded DNA, only the designated portion in each of the two strands was free from Watson-Crick base pairing with the counterpart DNA or the pcPNA. Upon the treatment of this invasion complex with Ce(IV)/EDTA at 37 degrees C and pH 7.0, both of these single-stranded portions were selectively hydrolyzed at the designated site, resulting in the site-selective two-strand scission of the double-stranded DNA. Furthermore, the hydrolytic scission products were successfully connected with foreign double-stranded DNA by using ligase. The potential of these artificial systems for manipulation of huge DNA has been indicated.

Combination of S1 nuclease and PNA for site-selective hydrolysis of double-stranded DNA. Comparison with the site-selective hydrolysis using Ce(IV)/EDTA.

The potential of the combination of SI nuclease and pseudo-complementary PNA (pcPNA) for site-selective scission of double-stranded DNA has been investigated. Through strand invasion of two pcPNAs, single-stranded portions were formed in both strands of substrate DNA. In the initial stage of the enzymatic digestion, two scission fragments were obtained due to the hydrolysis at these two gap-like sites. On prolonged reactions, however, these products (as well as the substrate DNA) were further digested to smaller fragments. Under the conditions employed here, only Ce(IV)/EDTA is available for the preparation of desired fragments from double-stranded DNA.

Manipulation of double-stranded DNA by artificial restriction enzyme composed of Ce(IV)/EDTA and PNA.

Through the invasion of pseudo-complementary PNA (pePNA) to double-stranded DNA, gap-like structures were formed at predetermined sites in both strands of PBR322 plasmid DNA. These gap-like sites were selectively hydrolyzed by Ce(IV)/EDTA complex, and two designed fragments were obtained. Furthermore, the scission fragment by this artificial restriction enzyme was successfully ligated with foreign DNA.

The influence of age on the efficiency of delayed therapy with Ca-DTPA for cerium in rats.

The influence of age on the effectiveness of chelation therapy starting 24 h after cerium administration was studied in 2- and 6-week-old rats. 141Ce was administered IP, followed after 24 and 48 h by IP administration of trisodium calcium diethylenetriaminepentaacetate (Ca-DTPA). The whole body and organ retention of 141Ce was determined 6 days after its administration. The chelation therapy significantly reduced cerium retention in both age groups but was more efficient in older animals. This age-related effect was, however, less pronounced than after immediate treatment with DTPA observed in our previous experiments, indicating that age as one of the factors which influence the efficacy of chelation treatment is probably more important in conditions of early than in conditions of delayed therapy.