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Aims: To characterize cellulolytic, hydrogen-producing clostridia on a comparable basis., Methods and Results: H(2) production from cellulose by six mesophilic clostridia was characterized in standardized batch experiments using MN301 cellulose, Avicel and cellobiose. Daily H(2) production, substrate degradation, biomass production and the end-point distribution of soluble fermentation products varied with species and substrates. All species produced a significant amount of H(2) from cellobiose, with Clostridium acetobutylicum achieving the highest H(2) yield of 2.3 mol H(2) mol(-1) hexose, but it did not degrade cellulose. Clostridium cellulolyticum and Clostridium populeti catalysed the highest H(2) production from cellulose, with yields of 1.7 and 1.6 mol H(2 )mol(-1) hexose from MN301 and 1.6 and 1.4 mol H(2) mol(-1) hexose from Avicel, respectively. These species also achieved 25-100% higher H(2) production rates from cellulose than the other species., Conclusions: These cellulolytic, hydrogen-producing clostridia varied in H(2) production, with Cl. cellulolyticum and Cl. populeti achieving the highest H(2) yields and cellulose degradation., Significance and Impact of the Study: The fermentation of cellulosic materials presents a means of H(2) production from renewable resources. This standardized comparison provides a quantitative baseline for improving H(2) production from cellulose through medium and process optimization and metabolic engineering.

Microbial fuel cells (MFCs) harness the electrochemical activity of certain microbes for the production of electricity from reduced compounds. Characterizations of MFC anode biofilms have collectively shown very diverse microbial communities, raising ecological questions about competition and community succession within these anode-reducing communities. Three sets of triplicate, two-chamber MFCs inoculated with anaerobic sludge and differing in energy sources (acetate, lactate, and glucose) were operated to explore these questions. Based on 16S rDNA-targeted denaturing gradient gel electrophoresis (DGGE), all anode communities contained sequences closely affiliated with Geobacter sulfurreducens (>99% similarity) and an uncultured bacterium clone in the Bacteroidetes class (99% similarity). Various other Geobacter-like sequences were also enriched in most of the anode biofilms. While the anode communities in replicate reactors for each substrate generally converged to a reproducible community, there were some variations in the relative distribution of these putative anode-reducing Geobacter-like strains. Firmicutes were found only in glucose-fed MFCs, presumably serving the roles of converting complex carbon into simple molecules and scavenging oxygen. The maximum current density in these systems was negatively correlated with internal resistance variations among replicate reactors and, likely, was only minimally affected by anode community differences in these two-chamber MFCs with high internal resistance.

Two different microbial fuel cell (MFC) configurations were investigated for electricity production from ethanol and methanol: a two-chambered, aqueous-cathode MFC; and a single-chamber direct-air cathode MFC. Electricity was generated in the two-chamber system at a maximum power density typical of this system (40+/-2 mW/m2) and a Coulombic efficiency (CE) ranging from 42% to 61% using ethanol. When bacteria were transferred into a single-chamber MFC known to produce higher power densities with different substrates, the maximum power density increased to 488+/-12 mW/m2 (CE = 10%) with ethanol. The voltage generated exhibited saturation kinetics as a function of ethanol concentration in the two-chambered MFC, with a half-saturation constant (Ks) of 4.86 mM. Methanol was also examined as a possible substrate, but it did not result in appreciable electricity generation. Analysis of the anode biofilm and suspension from a two-chamber MFC with ethanol using 16S rDNA-based techniques indicated that bacteria with sequences similar to Proteobacterium Core-1 (33.3% of clone library sequences), Azoarcus sp. (17.4%), and Desulfuromonas sp. M76 (15.9%) were significant members of the anode chamber community. These results indicate that ethanol can be used for sustained electricity generation at room temperature using bacteria on the anode in a MFC.

Microbial fuel cells (MFCs) convert biodegradable materials into electricity, potentially contributing to an array of renewable energy production strategies tailored for specific applications. Since there are no known microorganisms that can both metabolize cellulose and transfer electrons to solid extracellular substrates, the conversion of cellulosic biomass to electricity requires a syntrophic microbial community that uses an insoluble electron donor (cellulose) and electron acceptor (anode). Electricity was generated from cellulose in an MFC using a defined coculture of the cellulolytic fermenter Clostridium cellulolyticum and the electrochemically active Geobacter sulfurreducens. In fed-batch tests using two-chamber MFCs with ferricyanide as the catholyte, the coculture achieved maximum power densities of 143 mW/ m2 (anode area) and 59.2 mW/m2 from 1 g/L carboxymethyl cellulose (CMC) and MN301 cellulose, respectively. Neither pure culture alone produced electricity from these substrates. The coculture increased CMC degradation from 42% to 64% compared to a pure C. cellulolyticum culture. COD removal using CMC and MN301 was 38 and 27%, respectively, with corresponding Coulombic efficiencies of 47 and 39%. Hydrogen, acetate, and ethanol were the main residual metabolites of the binary culture. Cellulose conversion to electricity was also demonstrated using an uncharacterized mixed culture from activated sludge with an aerobic aqueous cathode.

The purpose of these experiments was to determine the potential estrogenic, androgenic, and progestagenic activity of two cyclic siloxanes, octamethylcyclotetrasiloxane (D4) and decamethylcyclopentasiloxane (D5). Receptor-binding experiments and a luciferase reporter gene assay were used to determine if the materials were able to bind and activate either the estrogen receptors (ERs) or progesterone receptors (PRs)-alpha or beta. The rat uterotrophic assay (RUA) for estrogenic activity and the Hershberger assay for androgenic activity were utilized as the in vivo assays. For the ER-binding studies, D4 was shown to bind to ERalpha but not to ERbeta. D5 did not bind to either of the two receptors. D4 activated the reporter gene at 10 microM, while D5 was considered negative in the estrogen reporter gene assay. Neither material was a ligand for the PRs. Both the RUA and Hershberger assays were conducted using whole-body inhalation of the two materials for 16 h/day. D4 resulted in a small but significant increase in both wet and blotted uterine weight as well as increases in both luminal and glandular epithelial cell height in both Sprague Dawley and Fischer 344 rats. D5 was negative in both rat strains, indicating that D5 does not possess estrogenic activity. Neither material possessed any significant antiestrogenic activity. Both materials were negative in the Hershberger assay indicating that neither material possesses any significant androgenic activity. Our studies have shown that D4 exhibits a low affinity for ERalpha in vitro and a weakly estrogenic response in vivo.

Microbial fuel cells (MFCs) are not yet commercialized but they show great promise as a method of water treatment and as power sources for environmental sensors. The power produced by these systems is currently limited, primarily by high internal (ohmic) resistance. However, improvements in the system architecture will soon result in power generation that is dependent on the capabilities of the microorganisms. The bacterial communities that develop in these systems show great diversity, ranging from primarily delta-Proteobacteria that predominate in sediment MFCs to communities composed of alpha-, beta-, gamma- or delta-Proteobacteria, Firmicutes and uncharacterized clones in other types of MFCs. Much remains to be discovered about the physiology of these bacteria capable of exocellular electron transfer, collectively defined as a community of "exoelectrogens". Here, we review the microbial communities found in MFCs and the prospects for this emerging bioenergy technology.

Microbial fuel cells (MFCs) represent a new method for treating animal wastewaters and simultaneously producing electricity. Preliminary tests using a two-chambered MFC with an aqueous cathode indicated that electricity could be generated from swine wastewater containing 8320 +/- 190 mg/L of soluble chemical oxygen demand (SCOD) (maximum power density of 45 mW/m2). More extensive tests with a single-chambered air cathode MFC produced a maximum power density with the animal wastewater of 261 mW/m2 (200 omega resistor), which was 79% larger than that previously obtained with the same system using domestic wastewater (146 +/- 8 mW/m2) due to the higher concentration of organic matter in the swine wastewater. Power generation as a function of substrate concentration was modeled according to saturation kinetics, with a maximum power density of P(max) = 225 mW/m2 (fixed 1000 omega resistor) and half-saturation concentration of K(s) = 1512 mg/L (total COD). Ammonia was removed from 198 +/- 1 to 34 +/- 1 mg/L (83% removal). In order to try to increase power output and overall treatment efficiency, diluted (1:10) wastewater was sonicated and autoclaved. This pretreated wastewater generated 16% more power after treatment (110 +/- 4 mW/m2) than before treatment (96 +/- 4 mW/m2). SCOD removal was increased from 88% to 92% by stirring diluted wastewater, although power output slightly decreased. These results demonstrate that animal wastewaters such as this swine wastewater can be used for power generation in MFCs while at the same time achieving wastewater treatment.

Bioremediation of perchlorate-contaminated groundwater can occur via bacterial reduction of perchlorate to chloride. Although perchlorate reduction has been demonstrated in bacterial pure cultures, little is known about the efficacy of using perchlorate-reducing bacteria as inoculants for bioremediation in the field. A pilot-scale, fixed-bed bioreactor containing plastic support medium was used to treat perchlorate-contaminated groundwater at a site in Southern California. The bioreactor was inoculated with a field-grown suspension of the perchlorate-respiring bacterium Dechlorosoma sp. strain KJ and fed groundwater containing indigenous bacteria and a carbon source amendment. Because the reactor was flushed weekly to remove accumulated biomass, only bacteria capable of growing in biofilms in the reactor were expected to survive. After 26 days of operation, perchlorate was not detected in bioreactor effluent. Perchlorate remained undetected by ion chromatography (detection limit 4 mug L(-1)) during 6 months of operation, after which the reactor was drained. Plastic medium was subsampled from top, middle, and bottom locations of the reactor for shipment on blue ice and storage at -80 degrees C prior to analysis. Microbial community DNA was extracted from successive washes of thawed biofilm material for PCR-based community profiling by 16S-23S ribosomal intergenic spacer analysis (RISA). No DNA sequences characteristic of strain KJ were recovered from any RISA bands. The most intense bands yielded DNA sequences with high similarities to Dechloromonas spp., a closely related but different genus of perchlorate-respiring bacteria. Additional sequences from RISA profiles indicated presence of representatives of the low G+C gram-positive bacteria and the Cytophaga-Flavobacterium-Bacteroides group. Confocal scanning laser microscopy and fluorescence in situ hybridization (FISH) were also used to examine biofilms using genus-specific 16S ribosomal RNA probes. FISH was more sensitive than RISA profiling in detecting possible survivors from the initial inoculum. FISH revealed that bacteria hybridizing to Dechlorosoma probes constituted <1% of all cells in the biofilms examined, except in the deepest portions where they represented 3-5%. Numbers of bacteria hybridizing to Dechloromonas probes decreased as biofilm depth increased, and they were most abundant at the biofilm surface (23% of all cells). These spatial distribution differences suggested persistence of low numbers of the inoculated strain Dechlorosoma sp. KJ in parts of the biofilm nearest to the plastic medium, concomitant with active colonization or growth by indigenous Dechloromonas spp. in the biofilm exterior. This study demonstrated the feasibility of post hoc analysis of frozen biofilms following completion of field remediation studies.

A protocol for simultaneously interrogating bacterial viability and identity using in situ, culture-independent methods is described. Viability is assayed using ethidium monoazide (EMA) staining of cells with compromised membranes, and identity is determined using fluorescent in situ hybridization (FISH). Experiments with planktonic cultures were used to demonstrate the compatibility of EMA staining and FISH after covalently bonding EMA to nucleic acids by photoreaction. Applications to biofilm samples showed that diffusion limitations in the biofilm matrix were not problematic and that effective discrimination of viable target cells within a mixed microbial community was possible.

Chloramination for secondary disinfection of drinking water often promotes the growth of nitrifying bacteria in the distribution system due to the ammonia introduced by chloramine formation and decay. This study involved the application of molecular biology techniques to explore the types of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) present in several full-scale chloraminated systems. The results of AOB community characterization indicated the ubiquitous detection of representatives from the Nitrosomonas genus, with Nitrosospira constituting a negligible or small fraction of the AOB community in all but one sample. Cloning and sequencing demonstrated the presence of AOB representatives within the Nitrosomonas oligotropha cluster, a phylogenetic subgroup of AOB from which isolates demonstrate a high affinity for ammonia. For the NOB communities, Nitrospira were detected in most of the samples, while Nitrobacter were only detected in a few samples. These results provide insight into the types of AOB responsible for nitrification episodes in full-scale chloraminated systems, which should help direct future studies aimed at characterizing relevant AOB growth and inactivation properties. Furthermore, the detection of NOB in most of the samples suggests a need to evaluate the contribution of biological nitrite oxidation relative to chemical oxidation in these systems.

Nitrification in drinking water distribution systems is a common operational problem for many utilities that use chloramines for secondary disinfection. The diversity of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in the distribution systems of a pilot-scale chloraminated drinking water treatment system was characterized using terminal restriction fragment length polymorphism (T-RFLP) analysis and 16S rRNA gene (ribosomal DNA [rDNA]) cloning and sequencing. For ammonia oxidizers, 16S rDNA-targeted T-RFLP indicated the presence of Nitrosomonas in each of the distribution systems, with a considerably smaller peak attributable to Nitrosospira-like AOB. Sequences of AOB amplification products aligned within the Nitrosomonas oligotropha cluster and were closely related to N. oligotropha and Nitrosomonas ureae. The nitrite-oxidizing communities were comprised primarily of Nitrospira, although Nitrobacter was detected in some samples. These results suggest a possible selection of AOB related to N. oligotropha and N. ureae in chloraminated systems and demonstrate the presence of NOB, indicating a biological mechanism for nitrite loss that contributes to a reduction in nitrite-associated chloramine decay.

Aerated-anoxic processes operate under the principle that small additions of oxygen to an anoxic reactor induce simultaneous nitrification and denitrification. In these systems, ammonia oxidation in the anoxic zone can easily account for 30-50% of the total nitrification in the reactor, even though the dissolve oxygen concentration is usually below detection limit. To investigate whether the nitrification efficiency in aerated-anoxic processes was due to the presence of specialized ammonia-oxidizing bacteria (AOB), an analysis of the AOB population in an aerated-anoxic Orbal process and a conventional nitrogen removal process was carried out using phylogenetic analyses based on the ammonia monooxygenase A (amoA) gene. Terminal restriction fragment length polymorphism (TRFLP) analyses revealed that Nitrosospira-like organisms were one of the major contributors to ammonia oxidation in a full-scale aerated-anoxic Orbal reactor. However, the relative populations of Nitrosospira-like and Nitrosomonas-like AOB were not constant and appeared to have seasonal variability. Cloning and sequence comparison of amoA gene fragments demonstrated that most of the AOB in the aerated-anoxic Orbal process belonged to the Nitrosospira sp. and Nitrosomonas oligotropha lineages. The abundance of Nitrosospira-like organisms in aerated-anoxic reactors is significant, since this group of AOB has not been usually associated with nitrification in wastewater treatment plants.

Background: Recent basic science research has greatly added to our knowledge of pain mechanisms. Application of this knowledge to cancer pain syndromes has led to new and innovative approaches to cancer pain management., Methods: The mechanisms involved in the three main cancer pain syndromes (somatic, visceral, and neuropathic) are reviewed, and various therapeutic options are discussed., Results: Advances in knowledge in neurophysiology, neuroanatomy, and pharmacology have allowed a greater understanding of the peripheral and central mechanisms of pain. New drugs and interventional techniques based on this knowledge have improved the control of cancer pain., Conclusions: Understanding the neurophysiology of cancer pain promotes use of the most appropriate palliative measures for pain control.

Unlabelled: We compared the quantitative leakage between forearm and conventional IV regional anesthesia (IVRA). Forearm IVRA remains unpopular because of the theoretical risk of local anesthetic leakage through the interosseous vessels. IVRA was simulated on the forearm or arm during two separate, randomized sessions using a double tourniquet in 14 volunteers. A radiolabeled substance, DISIDA (99m Tc-disofenin) with a structure similar to lidocaine, was injected instead of local anesthetic. Volumes of 0.4 mL/kg (maximum 25 mL), were used for forearm IVRA and 0.6 mL/kg (maximum 45 mL) for conventional IVRA. A gamma camera recorded radioactivity levels in the limb distal to the tourniquet every 30 s while the tourniquet was inflated (25 min) and for 20 min postdeflation. The leakage of radiolabeled substance during inflation was similar in both groups, 6%+/-12% (mean +/- SD) from the forearm and 10%+/-20% from the upper arm. After deflation, mean loss of radioactivity was higher in conventional IVRA, 70%+/-7% vs 57%+/-11% and 82%+/-5% vs 69%+/-11% at 3 and 20 min, respectively (P < 0.001). We conclude that forearm IVRA results in tourniquet leakage comparable to conventional IVRA and is potentially safer because the required dose of local anesthetic is smaller., Implications: Using a tourniquet on the forearm for IV regional anesthesia does not increase the risk of drug leakage. This is potentially a safer technique compared with conventional IV regional anesthesia because a much smaller dose of local anesthetic is required.

Pain is one of the most distressing symptoms associated with cancer. Basic science research has provided much insight into the mechanisms of peripheral and central pain and the actions of new drugs. Despite these advances, pain accompanying malignancy can be difficult to treat. Pain most commonly presents when the tumor has invaded somatic,visceral, or neural structures. An understanding of pain mechanisms is essential when deciding on the appropriate treatment. New therapeutic options have been developed and will hopefully provide clinicians with tools to successfully alleviate cancer pain.

Increased ambient temperature affects apparent oral glucose tolerance to an extent which may have clinical implications for the diagnosis of impaired glucose tolerance and gestational diabetes. As a first step in order to better define the nature of this effect, we have examined, in a climate chamber, the effects of ambient temperature at four levels (20, 25, 30, and 35 degrees C) on glucose and insulin responses to a standard 75 g oral glucose tolerance test in seven non-diabetic male subjects. Plasma glucose responses to ambient temperature were compared with the responses of core (auditory canal) and skin temperatures. The 2-h plasma glucose was affected in a nonlinear manner by ambient temperature (5.4 +/- 0.2, 5.3 +/- 0.4, 6.5 +/- 0.3, 6.4 +/- 0.4 mmol/l at 20, 25, 30, and 35 degrees C, P = 0.015) with the effect localised between 25 and 30 degrees C (P = 0.012). Core temperature responded in a similar manner (36.6 +/- 0.1, 36.6 +/- 0.1, 36.9 +/- 0.1, 37.0 +/- 0.1, (P = 0.0005) with the effect localised 25 and 30 degrees C (P = 0.011). However skin temperature increased significantly with each 5 degrees C increase in ambient temperature (30.2 +/- 0.5, 33.0 +/- 0.5, 34.2 +/- 0.2, 35.2 +/- 0.2, P < or = 0.0001). We conclude that the acute effect of ambient temperature on apparent glucose tolerance is most likely due to redistribution of blood flow between cutaneous and visceral beds driven by changes in core temperature. The absence of temperature effects between the two lowest, and between the two highest temperatures, provides workable guidelines for the standardisation of conditions during oral glucose tolerance tests in circumstances where temperature may have clinically significant effects.

This project sought to evaluate the importance of skin temperature during heat acclimation, using an isothermal-strain model. Two groups of seven matched males, participated (1 h per day, 10 days) in one of two conditions: (i) temperate physical training (TEMP: 22.4 +/- 0.7 degrees C, relative humidity (r.h.) 41.0 +/- 0.9%); or (ii) combined physical training and heat acclimation (HEAT: 38.2 +/- 0.7 degrees C, r.h. 39.7 +/- 1.3%). Isothermal strain was induced in both groups by rapidly elevating rectal temperature by 1 degree C (cycling), then holding it constant by manipulating external work. Subjects completed two three-phase heat stress tests (39.8 +/- 0.1 degrees C, r.h. 38.6 +/- 1.2), consisting of 20 min rest, then 20 min cycling at each of 30% and 45% of peak power, before and after each regimen. While there was a difference of 4.2 degrees C in mean skin temperature between treatments, both regimens elicited a similar peripheral sudomotor increase, indicating a core temperature dependent adaptation. However, based on significant pre- vs. post-acclimation decreases in average auditory canal temperature (0.4 +/- 0.1 degree C), average forehead skin blood flow (26%), average perceived exertion (11%), and a 5% increase in average forehead sweat rate (0.1 +/- 0.04 mg cm-2 min-1), the HEAT regimen elicited a more complete acclimation. While elevation in core temperature is critical to acclimation, it also appears necessary to expose subjects to an external thermal stress. This observation has not been previously demonstrated under conditions of isothermal strain, and verifies the importance of skin temperature elevation in the acclimation process.

Comparative 16S rRNA sequencing was used to evaluate phylogenetic relationships among selected strains of ammonia- and nitrite-oxidizing bacteria. All characterized strains were shown to be affiliated with the proteobacteria. The study extended recent 16S rRNA-based studies of phylogenetic diversity among nitrifiers by the comparison of eight strains of the genus Nitrobacter and representatives of the genera Nitrospira and Nitrospina. The later genera were shown to be affiliated with the delta subdivision of the proteobacteria but did not share a specific relationship to each other or to other members of the delta subdivision. All characterized Nitrobacter strains constituted a closely related assemblage within the alpha subdivision of the proteobacteria. As previously observed, all ammonia-oxidizing genera except Nitrosococcus oceanus constitute a monophyletic assemblage within the beta subdivision of the proteobacteria. Errors in the 16S rRNA sequences for two strains previously deposited in the databases by other investigators (Nitrosolobus multiformis C-71 and Nitrospira briensis C-128) were corrected. Consideration of physiology and phylogenetic distribution suggested that nitrite-oxidizing bacteria of the alpha and gamma subdivisions are derived from immediate photosynthetic ancestry. Each nitrifier retains the general structural features of the specific ancestor's photosynthetic membrane complex. Thus, the nitrifiers, as a group, apparently are not derived from an ancestral nitrifying phenotype.

A survey of treatment agencies showed that alcohol misusers' families remain on the periphery of the treatment process. When the spouses or children did receive services, it was usually in the form of individual counseling. Despite their growing popularity, conjoint couples therapy, group couples therapy and family treatment were seldom used.

The effects of introducing the Team Approach to the management of the lower limb amputee has been assessed in a consecutive series of 233 patients over a five year period. During the first year, baseline data was collected and during the subsequent yearly phases the effects of introducing a physiotherapist co-ordinator, visiting prosthetist and medical officer from the local Artificial Limb and Appliance Centre (ALAC), and finally trained surgeons were studied. During the final phases of the study, the effects of changing team staff were monitored. The results have shown that only when the full Team Approach is adopted are the best results achieved, but that, once this approach is established, staff changes can be made without serious reductions in effectiveness. The study has shown that the team can reduce in-patient stay by 20 days; reduce the need for post-discharge physiotherapy by 94%; increase the proportion of patients discharged with a prosthesis more than fivefold and increase the effectiveness of long term rehabilitation threefold.