Formation of biologically influenced palladium microstructures by Desulfovibrio desulfuricans and Desulfovibrio ferrophilus IS5.

A range of Desulfovibrio spp. can reduce metal ions to form metallic nanoparticles that remain attached to their surfaces. The bioreduction of palladium (Pd) has been given considerable attention due to its extensive use in areas of catalysis and electronics and other technological domains. In this study we report, for the first time, evidence for Pd(II) reduction by the highly corrosive Desulfovibrio ferrophilus IS5 strain to form surface attached Pd nanoparticles, as well as rapid formation of Pd(0) coated microbial nanowires. These filaments reached up to 8 µm in length and led to the formation of a tightly bound group of interconnected cells with enhanced ability to attach to a low carbon steel surface. Moreover, when supplied with high concentrations of Pd (≥ 100 mmol Pd(II) g-1 dry cells), both Desulfovibrio desulfuricans and D. ferrophilus IS5 formed bacteria/Pd hybrid porous microstructures comprising millions of cells. These three-dimensional structures reached up to 3 mm in diameter with a dose of 1200 mmol Pd(II) g-1 dry cells. Under suitable hydrodynamic conditions during reduction, two-dimensional nanosheets of Pd metal were formed that were up to several cm in length. Lower dosing of Pd(II) for promoting rapid synthesis of metal coated nanowires and enhanced attachment of cells onto metal surfaces could improve the efficiency of various biotechnological applications such as microbial fuel cells. Formation of biologically stimulated Pd microstructures could lead to a novel way to produce metal scaffolds or nanosheets for a wide variety of applications.

Inhibition of Sulfate Reduction and Cell Division by Desulfovibrio desulfuricans Coated in Palladium Metal.

The growth of sulfate-reducing bacteria (SRB) and associated hydrogen sulfide production can be problematic in a range of industries such that inhibition strategies are needed. A range of SRB can reduce metal ions, a strategy that has been utilized for bioremediation, metal recovery, and synthesis of precious metal catalysts. In some instances, the metal remains bound to the cell surface, and the impact of this coating on bacterial cell division and metabolism has not previously been reported. In this study, Desulfovibrio desulfuricans cells (1g dry weight) enabled the reduction of up to 1500 mmol (157.5 g) palladium (Pd) ions, resulting in cells being coated in approximately 1 µm of metal. Thickly coated cells were no longer able to metabolize or divide, ultimately leading to the death of the population. Increasing Pd coating led to prolonged inhibition of sulfate reduction, which ceased completely after cells had been coated with 1200 mmol Pd g-1 dry cells. Less Pd nanoparticle coating permitted cells to carry out sulfate reduction and divide, allowing the population to recover over time as surface-associated Pd diminished. Overcoming inhibition in this way was more rapid using lactate as the electron donor, compared to formate. When using formate as an electron donor, preferential Pd(II) reduction took place in the presence of 100 mM sulfate. The inhibition of important metabolic pathways using a biologically enabled casing in metal highlights a new mechanism for the development of microbial control strategies. IMPORTANCE Microbial reduction of sulfate to hydrogen sulfide is highly undesirable in several industrial settings. Some sulfate-reducing bacteria are also able to transform metal ions in their environment into metal phases that remain attached to their outer cell surface. This study demonstrates the remarkable extent to which Desulfovibrio desulfuricans can be coated with locally generated metal nanoparticles, with individual cells carrying more than 100 times their mass of palladium metal. Moreover, it reveals the effect of metal coating on metabolism and replication for a wide range of metal loadings, with bacteria unable to reduce sulfate to sulfide beyond a specific threshold. These findings present a foundation for a novel means of modulating the activity of sulfate-reducing bacteria.

The Critical Role of Environmental Synergies in the Creation of Bionanohybrid Microbes.

A wide range of bacteria can synthesize surface-associated nanoparticles (SANs) through exogenous metal ions reacting with sulfide produced via cysteine metabolism, resulting in the emergence of a biological-nanoparticle hybrid (bionanohybrid). The attached nanoparticles may couple to extracellular electron transfer, facilitating de novo photoelectrochemical processes. While SAN-cell coupling in hybrid organisms is opening a range of biotechnological possibilities, observation of bionanohybrids in nature is not commonly reported and their lab-based behavior remains difficult to control. We describe the critical role environmental synergy (microbial growth stage, cell densities, cysteine, and exogenous metal concentrations) plays in controlling the form and occurrence of Escherichia coli and Moorella thermoacetica bionanohybrids. SAN development depends on an appropriate cell density to metal ratio, with too few cells resulting in nanoparticle suppression through cytotoxicity or inhibition of cysteine conversion, and with too many cells diluting the number and size of particles produced. This cell number is governed by the concentration of cysteine present, which acts to protect the cells from metal ion toxicity. Exposing cells to metal and cysteine during the lag phase leads to SAN development, whereas cells in the exponential growth phase predominantly produce dispersed nanoparticles. Applying these principles more broadly, E. coli is shown to biosynthesize composite Bi/Cu sulfide SANs, and Clostridioides difficile can be coaxed into a bionanohybrid lifestyle by fine-tuning the cysteine dosage. Bionanohybrids maintain a remarkable ability for binary fission and sustained growth, opening doors to the production of SANs tailored to specific technological functions. IMPORTANCE Some bacteria can produce nanoscale-sized particles, which remain attached to the surface of the organism. The surface association of these nanoparticles creates a new mode of interaction between the microbe's environment and its internal cellular function, giving rise to a new hybrid lifeform, a biological nanoparticle hybrid (bionanohybrid). These hybrid organisms gain new or enhanced biological functions, and thus their creation opens a wide range of biotechnological possibilities. Despite this potential, the fundamental controls on bionanohybrid formation and occurrence remain poorly constrained. In this study, Escherichia coli K-12, Moorella thermoacetica, and Clostridioides difficile were used to test the combined influences of the growth phase, cell density, cysteine dose, and metal concentration in determining single and composite metal sulfide surface-associated nanoparticle production. The significance of this study is that it defined the critical synergies controlling nanoparticle formation on bacterial cell surfaces, unlocking the potential for bionanohybrid applications in a range of organisms.

Particles and enzymes: Combining nanoscale zero valent iron and organochlorine respiring bacteria for the detoxification of chloroethane mixtures.

Nanoscale zero valent iron (nZVI) and organochlorine respiring bacteria (ORB) are two technologies used to detoxify chlorinated aliphatic hydrocarbons (CAHs). nZVI can rapidly detoxify high CAH concentrations, but is quickly oxidised and unable to degrade certain CAHs (e.g., 1,2-dichlorothane). In contrast, ORB can dechlorinate CAHs resistant to nZVI (e.g., 1,2-dichlorothane) but are inhibited by other CAHs of concern degradable by nZVI (e.g., chloroform and carbon tetrachloride). Combining the two was proposed as a unique treatment train to overcome each technology's shortcomings. In this study, this combined remedy was investigated using a mixture of 1,2-dichloroethane, degradable by ORB but not nZVI, and 1,1,2-trichloroethane, susceptible to both. Results indicated that nZVI rapidly dechlorinated 1,1,2-trichloroethane when supplied above 0.5 g/L, however ORB were inhibited and unable to dechlorinate 1,2-dichloroethane. pH increase and ionic species associated with nZVI did not significantly impact ORB, pinpointing Fe(0) particles as responsible for ORB inhibition. Below 0.05 g/L nZVI, ORB activity was stimulated. Results suggest that combining ORB and nZVI at appropriate doses can potentially treat a wider range of CAHs than each individual remedy. At field sites where nZVI was applied, it is likely that in situ nZVI concentrations were below the threshold of negative consequences.

The application of nitric oxide to control biofouling of membrane bioreactors.

A novel strategy to control membrane bioreactor (MBR) biofouling using the nitric oxide (NO) donor compound PROLI NONOate was examined. When the biofilm was pre-established on membranes at transmembrane pressure (TMP) of 88-90 kPa, backwashing of the membrane module with 80 µM PROLI NONOate for 45 min once daily for 37 days reduced the fouling resistance (Rf ) by 56%. Similarly, a daily, 1 h exposure of the membrane to 80 µM PROLI NONOate from the commencement of MBR operation for 85 days resulted in reduction of the TMP and Rf by 32.3% and 28.2%. The microbial community in the control MBR was observed to change from days 71 to 85, which correlates with the rapid TMP increase. Interestingly, NO-treated biofilms at 85 days had a higher similarity with the control biofilms at 71 days relative to the control biofilms at 85 days, indicating that the NO treatment delayed the development of biofilm bacterial community. Despite this difference, sequence analysis indicated that NO treatment did not result in a significant shift in the dominant fouling species. Confocal microscopy revealed that the biomass of biopolymers and microorganisms in biofilms were all reduced on the PROLI NONOate-treated membranes, where there were reductions of 37.7% for proteins and 66.7% for microbial cells, which correlates with the reduction in TMP. These results suggest that NO treatment could be a promising strategy to control biofouling in MBRs.

Nitric oxide treatment for the control of reverse osmosis membrane biofouling.

Biofouling remains a key challenge for membrane-based water treatment systems. This study investigated the dispersal potential of the nitric oxide (NO) donor compound, PROLI NONOate, on single- and mixed-species biofilms formed by bacteria isolated from industrial membrane bioreactor and reverse osmosis (RO) membranes. The potential of PROLI NONOate to control RO membrane biofouling was also examined. Confocal microscopy revealed that PROLI NONOate exposure induced biofilm dispersal in all but two of the bacteria tested and successfully dispersed mixed-species biofilms. The addition of 40 µM PROLI NONOate at 24-h intervals to a laboratory-scale RO system led to a 92% reduction in the rate of biofouling (pressure rise over a given period) by a bacterial community cultured from an industrial RO membrane. Confocal microscopy and extracellular polymeric substances (EPS) extraction revealed that PROLI NONOate treatment led to a 48% reduction in polysaccharides, a 66% reduction in proteins, and a 29% reduction in microbial cells compared to the untreated control. A reduction in biofilm surface coverage (59% compared to 98%, treated compared to control) and average thickness (20 µm compared to 26 µm, treated compared to control) was also observed. The addition of PROLI NONOate led to a 22% increase in the time required for the RO module to reach its maximum transmembrane pressure (TMP), further indicating that NO treatment delayed fouling. Pyrosequencing analysis revealed that the NO treatment did not significantly alter the microbial community composition of the membrane biofilm. These results present strong evidence for the application of PROLI NONOate for prevention of RO biofouling.

Optimal dosing regimen of nitric oxide donor compounds for the reduction of Pseudomonas aeruginosa biofilm and isolates from wastewater membranes.

Membrane fouling by bacterial biofilms remains a key challenge for membrane-based water purification systems. Here, the optimal biofilm dispersal potential of three nitric oxide (NO) donor compounds, viz. sodium nitroprusside, 6-(2-hydroxy-1-methyl-2-nitrosohydrazino)-N-methyl-1-hexanamine (MAHMA NONOate) and 1-(hydroxy-NNO-azoxy)-L-proline, disodium salt, was investigated using Pseudomonas aeruginosa PAO1 as a model organism. Dispersal was quantitatively assessed by confocal microscopy [bacterial cells and the components of the extracellular polymeric substances (EPS) (polysaccharides and extracellular DNA)] and colony-forming unit counts. The three NO donor compounds had different optimal exposure times and concentrations, with MAHMA NONOate being the optimal NO donor compound. Biofilm dispersal correlated with a reduction in both bacterial cells and EPS. MAHMA NONOate also reduced single species biofilms formed by bacteria isolated from industrial membrane bioreactor and reverse osmosis membranes, as well as in isolates combined to generate mixed species biofilms. The data present strong evidence for the application of these NO donor compounds for prevention of biofouling in an industrial setting.

An assessment of the fate, behaviour and environmental risk associated with sunscreen TiO2 nanoparticles in UK field scenarios.

The fate of Ti was examined in an activated sludge plant serving over 200,000 people. These studies revealed a decrease of 30 to 3.2 µg/L of Ti < 0.45 µm from influent to effluent and a calculated Ti presence of 305 mg/kg DW in wasted sludge. Thus, using sludge as a fertiliser would result in a predicted deposition of up to 250 mg/m² of Ti to soil surfaces using a recommended maximal agricultural application rate. Given the major use of TiO2 in many industrial and domestic applications where loss to the sewer is possible, this measured Ti was presumed to have been largely TiO2, a proportion of which will be nanoparticle sized. To assess the behaviour of engineered nanoparticle (ENP) TiO2 in sewage and toxicology studies, Optisol (Oxonica Materials Ltd) and P25 (Evonik Industries AG), which are representative of forms used in sunscreen and cosmetic products, were used. These revealed a close association of TiO2 ENPs with activated sludge. Using commercial information on consumption, and removal rates for sewage treatment, predictions were made for river water concentrations for sunscreen TiO2 ENPs for the Anglian and Thames regions in Southern England. The highest predicted value from these exercises was 8.8 µg/L for the Thames region in which it was assumed that one in four people used the recommended application of sunscreen during a low flow (Q95) period. Ecotoxicological studies using potentially vulnerable species indicated that 1000 µg/L TiO2 ENP did not affect the viability of a mixed community of river bacteria in the presence of UV light. Direct exposure to TiO2 ENPs did not impair the immuno-effectiveness of earthworm coelomocyte cells at concentrations greatly above those predicted for sewage sludge.

The impact of zero-valent iron nanoparticles on a river water bacterial community.

Zero-valent iron (ZVI) nanoparticles are of interest because of their many potential biomedical and environmental applications. However, these particles have recently been reported to be cytotoxic to bacterial cells. The overall objective of this study was to determine the impact of 100mg/L ZVI nanoparticles on the diversity and structure of an indigenous river water bacterial community. Response during exposure for 36 days was determined by denaturing gel gradient electrophoresis (DGGE) analysis of bacterial 16S rRNA genes, amplified from extracted DNA, and viable and total cell abundances were determined by plate counting and fluorescent microscopy of DAPI-stained cells. Changes in river water chemistry were also monitored. Addition of ZVI nanoparticles led to a rapid decrease in oxidation-reduction potential (ORP) (+196 to -281 mV) and dissolved oxygen (DO) concentration (8.2-0.6 mg/L), both of which stabilized during the experiment. Interestingly, both viable and total bacterial cell abundances increased and pH decreased, characteristic of an active microbial community. Total community structure was visualized using rank-abundance plots fitted with linear regression models. The slopes of the regression models were used as a descriptive statistic of changes in evenness over time. Importantly, despite bacterial growth, addition of ZVI nanoparticles did not influence bacterial community structure.

Inhibition of biological TCE and sulphate reduction in the presence of iron nanoparticles.

Iron (Fe) nanoparticles are increasingly being employed for the remediation of Chlorinated Aliphatic Hydrocarbon (CAH) contaminated sites. However, these particles have recently been reported to be cytotoxic to bacterial cells, and may therefore have a negative impact on exposed microbial communities. The overall objective of this study was to investigate the impact of Fe nanoparticles on the biodegradation of CAHs by an indigenous dechlorinating bacterial community. Also, to determine the most appropriate combination and/or application of bimetallic (Ni/Fe) nanoparticles and dechlorinating bacteria for the remediation of CAH contaminated sites. Addition of Fe nanoparticles to groundwater collected from a CAH contaminated site in Derby, UK, led to a decrease in the oxidation-reduction potential (ORP) and an increase in pH. The biological degradation rate of TCE was observed to progressively decrease in the presence of increasing Fe nanoparticle concentrations; which ranged from 0.01 to 0.1 gL(-1), and cease completely at concentrations of 0.3 gL(-1) or above. Concentrations greater than 0.3 gL(-1) led to a decline in viable bacterial counts and the inhibition of biological sulphate reduction. The most appropriate means of combining bimetallic (Ni/Fe) nanoparticles and indigenous dechlorinating bacteria was to employ a two step process: initially stimulating the biodegradation of TCE using acetate, followed by the addition of bimetallic nanoparticles to degrade the remaining cis-1,2-DCE and VC.

Optimization of nano-scale nickel/iron particles for the reduction of high concentration chlorinated aliphatic hydrocarbon solutions.

The use of nano-scale particles as a means of environmental remediation still provides a comparatively novel approach for the treatment of contaminated waters. The current study compares the reactivity of micro-scale Fe, nano-scale Fe and nano-scale Ni/Fe (nickel/iron) particles specifically for dechlorination of solutions containing 350 mg L(-1) of TCE (concentration measured at a contaminated site in Derbyshire, UK). The results indicated that employing 1 g L(-1) of reactive material for dechlorination in the monometallic form (both micro- and nano-scale) exhibited very little reduction capability compared with the bimetallic Ni/Fe nano-scale particles, containing 28.9% Ni (in molar), which achieved complete dechlorination of the TCE in solution within 576 h. Experiments were also performed to determine the optimum bimetallic composition of the Ni/Fe particles for TCE reduction. This revealed that 3.2% Ni was the optimum Ni/Fe molar ratio for both maximum dehalogenation performance and minimum release of Ni into solution. Using particles of the most effective bimetallic composition, experiments were carried out to determine the concentration required for optimal TCE reduction. Over the range of nano-scale particle concentrations tested (0.1-9 g L(-1)), reduction rates of TCE increased with greater TCE:nano-scale particle ratios. However, a concentration range of 1-3 g L(-1) was selected as the most appropriate for site remediation, since more concentrated solutions demonstrated only small increases in rates of reaction. Finally, in order to test the long term performance and reactivity of the 3.2% Ni/Fe bimetallic nano-scale particles, weekly spikes of 350 mg L(-1) TCE were injected into a 3 g L(-1) nano-scale particle batch reactor. Results showed that the bimetallic nano-scale particles had the ability to reduce 1750 mg L(-1) TCE and remained active for at least 13 weeks.