How sulfur species can accelerate the biological immobilization of the toxic selenium oxyanions and promote stable hexagonal Se0 formation.

Toxic selenium oxyanions and sulfur species are often jointly present in contaminated waters and soils. This study investigated the effect on kinetics and resulting products for bio-reduction of selenium oxyanions in the presence of biologically produced sulfur resulting from bio-oxidation of sulfide in (bio)gas-desulfurization (bio-S0) and of sulfate. Selenite and selenate (~2 mmol L-1) bio-reduction was studied in batch up to 28 days at 30 oC and pH 7 using lactic acid and a sulfate-reducing sludge, 'Emmtec'. Bio-S0 addition increased the selenite removal rate, but initially slightly decreased selenate reduction rates. Selenite reacted with biologically generated sulfide resulting in selenium-sulfur, which upon further bio-reduction creates a sulfur bio-reduction cycle. Sulfate addition increased the bio-reduction rate for both selenite and sulfate. Bio-S0 or sulfate promoted hexagonal selenium formation, whereas without these, mostly amorphous Se0 resulted. With another inoculum, 'Eerbeek', bio-S0 accelerated the selenite reduction rate less than for 'Emmtec' because of lower sulfur and higher selenite bio-reduction rates. Bio-S0 addition increased the selenate reduction rate slightly and accelerated hexagonal selenium formation. Hexagonal selenium formation is advantageous because it facilitates separation and recovery and is less mobile and toxic than amorphous Se0. Insights into the interaction between selenium and sulfur bio-reduction are valuable for understanding environmental pathways and considerations regarding remediation and recovery.

Autogenerative high pressure digestion: anaerobic digestion and biogas upgrading in a single step reactor system.

Conventional anaerobic digestion is a widely applied technology to produce biogas from organic wastes and residues. The biogas calorific value depends on the CH, content which generally ranges between 55 and 65%. Biogas upgrading to so-called 'green gas', with natural gas quality, generally proceeds with add-on technologies, applicable only for biogas flows > 100 m3/h. In the concept of autogenerative high pressure digestion (AHPD), methanogenic biomass builds up pressure inside the reactor. Since CO2 has a higher solubility than CH4, it will proportion more to the liquid phase at higher pressures. Therefore, AHPD biogas is characterised by a high CH4 content, reaching equilibrium values between 90 and 95% at a pressure of 3-90 bar. In addition, also H2S and NH3 are theoretically more soluble in the bulk liquid than CO2. Moreover, the water content of the already compressed biogas is calculated to have a dew point <--10 degrees C. Ideally, high-quality biogas can be directly used for electricity and heat generation, or injected in a local natural gas distribution net. In the present study, using sodium acetate as substrate and anaerobic granular sludge as inoculum, batch-fed reactors showed a pressure increase up to 90 bars, the maximum allowable value for our used reactors. However, the specific methanogenic activity (SMA) of the sludge decreased on average by 30% compared to digestion at ambient pressure (1 bar). Other results show no effect of pressure exposure on the SMA assessed under atmospheric conditions. These first results show that the proposed AHPD process is a highly promising technology for anaerobic digestion and biogas upgrading in a single step reactor system.

High-rate biological selenate reduction in a sequencing batch reactor for recovery of hexagonal selenium.

Recovery of selenium (Se) from wastewater provides a solution for both securing Se supply and preventing Se pollution. Here, we developed a high-rate process for biological selenate reduction to elemental selenium. Distinctive from other studies, we aimed for a process with selenate as the main biological electron sink, with minimal formation of methane or sulfide. A sequencing batch reactor, fed with an influent containing 120 mgSe L-1 selenate and ethanol as electron donor and carbon source, was operated for 495 days. The high rates (419 ± 17 mgSe L-1 day-1) were recorded between day 446 and day 495 for a hydraulic retention time of 6 h. The maximum conversion efficiency of selenate amounted to 96% with a volumetric conversion rate of 444 mgSe L-1 day-1, which is 6 times higher than the rates reported in the literature thus far. At the end of the experiment, a highly enriched selenate reducing biomass had developed, with a specific activity of 856 ± 26 mgSe-1day-1gbiomass-1, which was nearly 1000-fold higher than that of the inoculum. No evidence was found for the formation of methane, sulfide, or volatile reduced selenium compounds like dimethyl-selenide or H2Se, revealing a high selectivity. Ethanol was incompletely oxidized to acetate. The produced elemental selenium partially accumulated in the reactor as pure (≥80% Se of the total mixture of biomass sludge flocs and flaky aggregates, and ~100% of the specific flaky aggregates) selenium black hexagonal needles, with cluster sizes between 20 and 200 µm. The new process may serve as the basis for a high-rate technology to remove and recover pure selenium from wastewater or process streams with high selectivity.

Histological validation of ultrasound-guided neurography in early nerve regeneration.

Ultrasound-guided near-nerve neurography is a new tool that can be used to assess nerve regeneration before reinnervation occurs. In this study, ultrasound-guided near-nerve measurements were validated against axon diameter counts in rabbits during a 15-week regeneration period after a crush lesion of their peroneal nerve. The course of the nerve was determined ultrasonically, and the active near-nerve needle electrode was maneuvered just next to the nerve under ultrasound guidance. Measured action potentials were compared with axon diameter counts from histological sections of these same nerves. A moderate to good positive correlation was found, which reached a maximum of 0.7 at a cut-off of 3 microm, corresponding to the minimal size of the myelinated axons. Our results suggest that, following a similar validation study in humans, ultrasound-guided near-nerve neurography may be clinically useful when early evaluation of nerve activity is needed.

Metabolic interactions in methanogenic and sulfate-reducing bioreactors.

In environments where the amount of electron acceptors is insufficient for complete breakdown of organic matter, methane is formed as the major reduced end product. In such methanogenic environments organic acids are degraded by syntrophic consortia of acetogenic bacteria and methanogenic archaea. Hydrogen consumption by methanogens is essential for acetogenic bacteria to convert organic acids to acetate and hydrogen. Several syntrophic cocultures growing on propionate and butyrate have been described. These syntrophic fatty acid-degrading consortia are affected by the presence of sulfate. When sulfate is present sulfate-reducing bacteria compete with methanogenic archaea for hydrogen and acetate, and with acetogenic bacteria for propionate and butyrate. Sulfate-reducing bacteria easily outcompete methanogens for hydrogen, but the presence of acetate as carbon source may influence the outcome of the competition. By contrast, acetoclastic methanogens can compete reasonably well with acetate-degrading sulfate reducers. Sulfate-reducing bacteria grow much faster on propionate and butyrate than syntrophic consortia.

Effect of chronic light-dark shift stress on the immune response of the rat.

During a period of 35 weeks the night-day pattern of inbred Brown Norway female rats was changed weekly by alternating the light-dark (L-D) rhythm. After a period of 2 months, in a number of the animals, the cellular immune response was measured by means of Concanavalin A stimulation of peripheral blood (Con A) and a Popliteal Lymph Node Assay (PLNA). Serum corticosterone, plasma free fatty acids and peripheral leucocytes were determined as well. Seven months thereafter the remaining animals were sacrificed after which adrenal gland weight and spleen weight were established. Additionally, blood glucose and corticosterone were measured (corticosterone in vitro activity as well as the serum level). Both Con A and PLNA showed a significantly decreased immune response in the L-D shift stress group. Adrenal cortical activity measured in vitro as well as in vivo did not show any significant changes, neither at 2 months nor at 9 months. Therefore, the observed immunosuppressive effect of chronic light-dark shift stress can not be explained by an increased adrenal cortical activity. Other possible explanations for the effect of the light-dark shift stress on the immune response are discussed.

A microchip implant system as a method to determine body temperature of terminally ill rats and mice.

In a series of experiments, Klebsiella pneumoniae was inoculated intratracheally into rats and mice, and the temperature of the animals was recorded twice daily using microchip transponders. Transponders are interrogated by radio frequencies and were implanted either subcutaneously or intraperitoneally. The microchip temperatures were compared with rectal temperatures taken at the same time. The purpose of the experiments was (a) to investigate the practicability and reliability of the ELAMS for temperature recording; (b) to compare values given by subcutaneously or intraperitoneally implanted transponders with rectal temperatures; and (c) to determine a 'temperature-cut-off point' as an alternative for 'death of the animal' as an end point for the experiment. The results showed that the ELAMS was easy to operate and no important drawbacks in the use of the system were observed. The temperatures generated by the transponders implanted subcutaneously and intraperitoneally did not differ significantly from rectal temperatures. In two out of three experiments on rats, it was shown that when the temperatures reached values below 36 degrees C, the median survival time of the animals was 24 h. In the one experiment on mice the same median survival time was observed at 36 degrees C. In one experiment using rats, however, the disease was so acute that death occurred before any temperature drop was seen. The results show that when a 36 degrees C cut-off point is used instead of the time of death in this particular animal model, the statistical analysis was not altered, but that it would spare animals further suffering for approximately 24 h. The argument that measuring body temperature is a laborious job and stressful to the animals is overcome when the ELAMS system is used.

Modelling the competition between sulphate reducers and methanogens in a thermophilic methanol-fed bioreactor.

Sulphate can be removed from wastewater by means of biological anaerobic reduction to sulphide. The reduction requires the presence of a substrate that can serve as an electron donor. Methanol a suitable electron donor for sulphate reduction under thermophilic conditions. In an anaerobic system containing methanol and sulphate, acetogenic bacteria (AB) and methanogenic archaea (MA) compete with sulphate reducing bacteria (SRB) for methanol or its degradation intermediates. Previously obtained results indicate that at 65 degrees C SRB and MA mainly compete for the intermediate hydrogen instead of methanol. For efficient use of methanol as electron donor for sulphate reduction it is important that for the treatment of sulphate wastewater in an anaerobic reactor SRB out-compete MA. The mechanisms that determine the outcome of the competition are, however, not well understood. This paper describes a model based on growth kinetics of methanol-oxidising AB, and hydrogen-consuming SRB and MA, that can describe the competition between SRB and MA in a methanol-fed bioreactor. We present the model and its calibration using experimental data, and we discuss its shortcomings and suggest possible improvements.

Methanol conversion in high-rate anaerobic reactors.

An overview on methanol conversion in high-rate anaerobic reactors is presented, with the focus on technological as well as microbiological aspects. The simple C1-compound methanol can be degraded anaerobically in a complex way, in which methanogens, sulfate reducing bacteria and homoacetogens interact cooperatively or competitively at substrate level. This interaction has large technological implications as it determines the final product of methanol mineralization, methane or carbon dioxide. The degradation route of methanol may be entirely different when environmental conditions change. Direct methanogenesis from methanol seems the predominant mineralization route under mesophilic conditions both in the absence and the presence of sulfate. Under thermophilic conditions methanol oxidation to carbon dioxide and hydrogen appears to play an important role. The UASB technology for mesophilic digestion of methanolic waste has presently reached full-scale maturity. The potential of methanol as feedstock for anaerobic processes is discussed.

New developments in reactor and process technology for sulfate reduction.

Sulfate reduction has been regarded in the past as an unwanted process in anaerobic treatment of sulfate-rich wastewaters. Research efforts were primarily focused on H2S toxicity, competition between sulfidogenic and methanogenic microorganisms and suppression of sulfidogenesis. More recently, the potential sulfidogenesis for treating a wide range of wastestreams contaminated with oxidized sulfurous compounds and/or heavy metals was also appreciated. Heavy metals can be removed by the formation and subsequent precipitation of poorly soluble metal sulfides. Basically two approaches can be distinguished in wastewater treatment: passive treatment using low-cost technologies and active treatment in newly developed bioreactors. Both strategies are discussed.

Competition for H2 between sulfate reducers, methanogens and homoacetogens in a gas-lift reactor.

Reported values for growth kinetic parameters show an order in competitivity of heterotrophic sulfate reducing bacteria>methanogens>homoacetogens for the substrate hydrogen. This order suggests that methanogens can succesfully compete with consortia of heterotrophic SRB and homoacetogens when H2/CO2 is present as sole substrate. However, we found in experiments using gas-lift reactors inoculated with anaerobic sludge and fed with H2/CO2 and sulfate, that heterotrophic sulfate reduction rapidly and completely outcompeted methanogenesis, whereas a low amount of acetate was formed. Thus, in disagreement with the above competitivity order, hydrogen is more readily consumed by homoacetogenesis than by methanogenesis, indicating that the competition is not kinetically determined. The superior settling velocity of sulfidogenic-acetogenic sludge compared to that of methanogenic sludge suggests that the former sludge is better retained, which can explain the predominance of sulfate reduction/homoacetogenesis over methanogenesis.

Rabbits as a model for research into craniofacial distraction osteogenesis.

Various factors affect the choice of the appropriate animal for craniofacial research. We have evaluated the rabbit as a suitable animal for research on craniofacial distraction osteogenesis. We describe our experience with housing and handling them, surgical and experimental protocols, and compare them with other animals. We introduce, and describe the use of, a continuous hydraulic distractor on the nasal bones of the rabbit. Fifty-two skeletally mature New Zealand White rabbits were used. Forty-two of the 52 operations were uneventful. Ten of the fifty-two developed complications, of which two were animal-related, and the other eight distractor-related. During the experiments the animals stayed healthy, and the distraction procedures were well tolerated. Rabbits are excellent for use in biological research on craniofacial distraction osteogenesis. Specifically, their nasal bones are easily accessible, the size and shape of the nasal bones allow various commercially available as well as custom-made distractors to be attached to the bones easily, their care and housing are relatively simple and inexpensive, and harvesting of tissue for further analyses is no problem because their skulls are of a manageable size and shape compared with other laboratory animals.

Performance of a thermophilic sulfate and sulfite reducing high rate anaerobic reactor fed with methanol.

Thermophilic sulfate and sulfite reduction was studied in lab-scale Expanded Granular Sludge Bed (EGSB) reactors operated at 65 degrees C and pH 7.5 with methanol as the sole carbon and energy source for the sulfate- and sulfite-reducing bacteria. At a hydraulic retention time (HRT) of 10 h, maximum sulfite and sulfate elimination rates of 5.5 g SO3(2-) L(-1) day(-1) (100% elimination) and 5.7 g SO4(2-) L(-1) day(-1) (55% elimination) were achieved, resulting in an effluent sulfide concentration of approximately 1800 mg S L(-1). Sulfate elimination was limited by the sulfide concentration, as stripping of H2S from the reactor with nitrogen gas was found to increase the sulfate elimination rate to 9.9 g SO4(2-) L(-1) day(-1) (100% elimination). At a HRT of 3 h, maximum achievable sulfite and sulfate elimination rates were even 18 g SO3(2-) L(-1) day(-1) (100% elimination) and 11 g SO4(2-) L(-1) day(-1) (50% elimination). At a HRT of 3 h, the elimination rate was limited by the biomass retention of the system. 5.5 +/- 1.8% of the consumed methanol was converted to acetate, which was not further degraded by sulfate reducing bacteria present in the sludge. The acetotrophic activity of the sludge could not be stimulated by cultivating the sludge for 30 days under methanol-limiting conditions. Omitting cobalt as trace element from the influent resulted in a lower acetate production rate, but it also led to a lower sulfate reduction rate. Sulfate degradation in the reactor could be described by zeroth order kinetics down to a threshold concentration of 0.05 g L(-1), while methanol degradation followed Michaelis-Menten kinetics with a Km of 0.037 g COD L(-1).

Thermophilic sulfate reduction and methanogenesis with methanol in a high rate anaerobic reactor.

Sulfate reduction outcompeted methanogenesis at 65 degrees C and pH 7.5 in methanol and sulfate-fed expanded granular sludge bed reactors operated at hydraulic retention times (HRT) of 14 and 3.5 h, both under methanol-limiting and methanol-overloading conditions. After 100 and 50 days for the reactors operated at 14 and 3.5 h, respectively, sulfide production accounted for 80% of the methanol-COD consumed by the sludge. The specific methanogenic activity on methanol of the sludge from a reactor operated at HRTs of down to 3.5 h for a period of 4 months gradually decreased from 0. 83 gCOD. gVSS(-1). day(-1) at the start to a value of less than 0.05 gCOD. gVSS(-1). day(-1), showing that the relative number of methanogens decreased and eventually became very low. By contrast, the increase of the specific sulfidogenic activity of sludge from 0. 22 gCOD. gVSS(-1). day(-1) to a final value of 1.05 gCOD. gVSS(-1). day(-1) showed that sulfate reducing bacteria were enriched. Methanol degradation by a methanogenic culture obtained from a reactor by serial dilution of the sludge was inhibited in the presence of vancomycin, indicating that methanogenesis directly from methanol was not important. H(2)/CO(2) and formate, but not acetate, were degraded to methane in the presence of vancomycin. These results indicated that methanol degradation to methane occurs via the intermediates H(2)/CO(2) and formate. The high and low specific methanogenic activity of sludge on H(2)/CO(2) and formate, respectively, indicated that the former substrate probably acts as the main electron donor for the methanogens during methanol degradation. As sulfate reduction in the sludge was also strongly supported by hydrogen, competition between sulfate reducing bacteria and methanogens in the sludge seemed to be mainly for this substrate. Sulfate elimination rates of up to 15 gSO(4)(2-)/L per day were achieved in the reactors. Biomass retention limited the sulfate elimination rate.

Cometabolic degradation of trichloroethylene by Pseudomonas cepacia G4 in a chemostat with toluene as the primary substrate.

Pseudomonas cepacia G4 is capable of cometabolic degradation of trichloroethylene (TCE) if the organism is grown on certain aromatic compounds. To obtain more insight into the kinetics of TCE degradation and the effect of TCE transformation products, we have investigated the simultaneous conversion of toluene and TCE in steady-state continuous culture. The organism was grown in a chemostat with toluene as the carbon and energy source at a range of volumetric TCE loading rates, up to 330 mumol/liter/h. The specific TCE degradation activity of the cells and the volumetric activity increased, but the efficiency of TCE conversion dropped when the TCE loading was elevated from 7 to 330 mumol/liter/h. At TCE loading rates of up to 145 mumol/liter/h, the specific toluene conversion rate and the molar growth yield of the cells were not affected by the presence of TCE. The response of the system to varying TCE loading rates was accurately described by a mathematical model based on Michaelis-Menten kinetics and competitive inhibition. A high load of 3,400 mumol of TCE per liter per h for 12 h caused inhibition of toluene and TCE conversion, but reduction of the TCE load to the original nontoxic level resulted in complete recovery of the system within 2 days. These results show that P. cepacia can stably and continuously degrade toluene and TCE simultaneously in a single-reactor system without biomass retention and that the organism is more resistant to high concentrations and shock loadings of TCE than Methylosinus trichosporium OB3b.

Adjuvant intraoperative photodynamic therapy diminishes the rate of local recurrence in a rat mammary tumour model.

The use of photodynamic therapy (PDT) as an adjunct to curative tumour resection was investigated in a tumour recurrence model, using rat mammary adenocarcinoma BN472. Tumours were inoculated subcutaneously in 60 animals and resected after 21 days of growth. Immediately after removal, the operation site was exposed to 320-450 nm light of 0.1 W cm-2 and 60 J cm-2 after photosensitisation with either Photofrin (5 mg kg-1 i.v. 48 h before illumination) or 5-aminolaevulinic acid (ALA) (2 mg ml-1 in drinking water for 9 days). Porphyrin concentrations were measured in tissue samples. After 28 days, animals treated with adjunctive PDT had a significantly longer tumour-free interval than controls (P < 0.01); median 25 days (Photofrin), 18 days (ALA), 14 days (controls). Moreover, in the PDT groups significantly fewer rats had lymph node metastasis. A prophyrin concentration ratio between tumour and mammary tissue of 2:1 was found after Photofrin and 4:1 after ALA. The results indicate that adjuvant intraoperative PDT may be a safe and effective method of destroying residual tumour, thereby preventing locoregional tumour recurrence.