High-throughput combination assay for studying biofilm formation of uropathogenic Escherichia coli.

Uropathogenic Escherichia coli, the most common cause for urinary tract infections, forms biofilm enhancing its antibiotic resistance. To assess the effects of compounds on biofilm formation of uropathogenic Escherichia coli UMN026 strain, a high-throughput combination assay using resazurin followed by crystal violet staining was optimized for 384-well microplate. Optimized assay parameters included, for example, resazurin and crystal violet concentrations, and incubation time for readouts. For the assay validation, quality parameters Z' factor, coefficient of variation, signal-to-noise, and signal-to-background were calculated. Microplate uniformity, signal variability, edge well effects, and fold shift were also assessed. Finally, a screening with known antibacterial compounds was conducted to evaluate the assay performance. The best conditions found were achieved by using 12 µg/mL resazurin for 150 min and 0.023% crystal violet. This assay was able to detect compounds displaying antibiofilm activity against UMN026 strain at sub-inhibitory concentrations, in terms of metabolic activity and/or biomass.

Theoretical Evaluation of Fluorinated Resazurin Derivatives for In Vivo Applications.

Primarily owing to the pronounced fluorescence exhibited by its reduced form, resazurin (also known as alamarBlue®) is widely employed as a redox sensor to assess cell viability in in vitrostudies. In an effort to broaden its applicability for in vivo studies, molecular adjustments are necessary to align optical properties with the near-infrared imaging window while preserving redox properties. This study delves into the theoretical characterisation of a set of fluorinated resazurin derivatives proposed by Kachur et al., 2015 examining the influence of fluorination on structural and electrochemical properties. Assuming that the conductor-like polarisable continuum model mimics the solvent effect, the density functional level of theory combining M06-2X/6-311G* was used to calculate the redox potentials. Furthermore, (TD-)DFT calculations were performed with PBE0/def2-TZVP to evaluate nucleophilic characteristics, transition states for fluorination, relative energies, and fluorescence spectra. With the aim of exploring the potential of resazurin fluorinated derivatives as redox sensors tailored for in vivo applications, acid-base properties and partition coefficients were calculated. The theoretical characterisation has demonstrated its potential for designing novel molecules based on fundamental principles.

[Establishment and Evaluation of a Resazurin-Based Microdilution Assay for Microbial Sensitivity Test of Neisseria gonorrhoeae]. / æ·‹ç— å¥ˆç‘ŸèŒè¯æ•è¯•éªŒåˆƒå¤©é’å¾®é‡ç¨€é‡Šæ³•çš„å»ºç«‹ä¸Žè¯„ä»·.

Objective: To establish and evaluate a microbial sensitivity test method for Neisseria gonorrhoeae based on resazurin coloration. Methods: Based on the broth microdilution method, resazurin was added as a live bacteria indicator. WHO G, a WHO gonococcal reference strain, was used to optimize the incubation time for resazurin-stained bacteria and the color change was visually observed to obtain the results. Agar dilution method (the gold standard) and resazurin-based microdilution assay were used to determine the minimum inhibitory concentration (MIC) of azithromycin, ceftriaxone, and spectinomycin for 3 reference strains and 32 isolates of Neisseria gonorrhoeae. The results were analyzed based on essential agreement (EA), which reflected the consistency of the MIC values, category agreement (CA), which reflected the consistency in the determination of drug resistance, intermediary, and sensitivity, very major error (VME), which reflected false sensitivity, and major error (ME), which reflected pseudo drug resistance, to evaluate the accuracy of resazurin-based microdilution assay as a microbial sensitivity test of of Neisseria gonorrhoeae. CA and EA rates≥90% and VME and ME rates≤3% were found to be the acceptable performance rates. Results: The results obtained 6 hours after resazurin was added were consistent with those of the agar dilution method and the resazurin-based microdilution assay was established accordingly based on this parameter. The EA of resazurin-based microdilution assay for measuring the MIC results of azithromycin, ceftriaxone, and spectinomycin was 97.1%, 91.5%, and 94.3%, respectively, and the CA was 88.6%, 94.3%, and 94.3%, respectively. The VME was 0% for all three antibiotics, while the ME was 11.4%, 5.7%, and 5.7%, respectively. Conclusion: The resazurin-based microdilution assay established in this study showed good agreement with agar dilution method for measuring the MIC of antibiotics against Neisseria gonorrhoeae. Moreover, the sensitivity results of this method were highly reliable and could be easily obtained through naked eye observation. Nonetheless, the results of drug resistance should be treated with caution and the optimization of parameters should be continued.

Assessment of yeast physiology during industrial-scale brewing practices using the redox-sensitive dye resazurin.

Beer refermentation in bottles is an industrial process utilized by breweries where yeast and fermentable extract are added to green beer. The beer is refermented for a minimum of 2 weeks before distribution, with the physiological state of the yeast a critical factor for successful refermentation. Ideally, fresh yeast that is propagated from a dedicated propagation plant should be used for refermentation in bottles. Here, we explored the applicability of the fluorescent and redox-sensitive dye, resazurin, to assess cellular metabolism in yeast and its ability to differentiate between growth stages. We applied this assay, with other markers of yeast physiology, to evaluate yeast quality during a full-scale industrial propagation. Resazurin allowed the discrimination between the different growth phases in yeast and afforded a more in-depth understanding of yeast metabolism during propagation. This assay can be used to optimize the yeast propagation process and cropping time to improve beer quality.

A New Electron Shuttling Pathway Mediated by Lipophilic Phenoxazine via the Interaction with Periplasmic and Inner Membrane Proteins of Shewanella oneidensis MR-1.

Although it has been established that electron mediators substantially promote extracellular electron transfer (EET), electron shuttling pathways are not fully understood. Here, a new electron shuttling pathway was found in the EET process by Shewanella oneidensis MR-1 with resazurin, a lipophilic electron mediator. With resazurin, the genes encoding outer-membrane cytochromes (mtrCBA and omcA) were downregulated. Although cytochrome deletion substantially reduced biocurrent generation to 1-12% of that of wild-type (WT) cells, the presence of resazurin restored biocurrent generation to 168 µA·cm-2 (ΔmtrA/omcA/mtrC), nearly equivalent to that of WT cells (194 µA·cm-2), indicating that resazurin-mediated electron transfer was not dependent on the Mtr pathway. Biocurrent generation by resazurin was much lower in ΔcymA and ΔmtrA/omcA/mtrC/fccA/cctA mutants (4 and 6 µA·cm-2) than in WT cells, indicating a key role of FccA, CctA, and CymA in this process. The effectiveness of resazurin in EET of Mtr cytochrome mutants is also supported by cyclic voltammetry, resazurin reduction kinetics, and in situ c-type cytochrome spectroscopy results. The findings demonstrated that low molecular weight, lipophilic electron acceptors, such as phenoxazine and phenazine, may facilitate electron transfer directly from periplasmic and inner membrane proteins, thus providing new insight into the roles of exogenous electron mediators in electron shuttling in natural and engineered biogeochemical systems.

A downstream process control tool based on the redox dye resazurin for rapid and accurate measurement of microbial metabolic activity.

Many sophisticated water treatment plants need a reliable, fast, and economical microbial load detection method. We refined a colorimetric assay using the redox dye resazurin to assess viable microorganisms. Here, we have used a mixed bacterial suspension of significant multi-drug-resistant coliform bacteria isolated from hospital wastewater and constructed a resazurin reduction calibration curve which could accurately predict the level of microbial contamination. The number of viable microorganisms was calculated from calibration curve in terms of log colony forming units (CFU) per milliliter. Ultrasonication disinfection of bacterial suspension for a duration of 50 min measured by resazurin assay depicted a reduction of 16.94%, 26.48%, and 37.69% at 410 W, 580 W, and 700 W, respectively. A synergistic effect of the combined methods of ultrasonication and heat disinfection treatments on raw wastewater and secondary wastewater effluent was observed and was also evaluated using both resazurin assay and standard plate count method. For raw wastewater, about 1.8 log reduction was observed for ultrasonication alone and 4 log CFU/mL reduction for thermosonication. In the secondary wastewater effluent, a reduction of 2.9 and 3.2 log CFU/mL was recorded for ultrasonication and thermosonication respectively. Resazurin microbial viability test results were highly comparable with conventional colony plate count for all treatment procedures, suggesting its appropriateness for quick and reliable wastewater sample microbial viability monitoring.

Improving droplet microfluidic systems for studying single bacteria growth.

Antimicrobial resistance remains a global threat with ~ 5 million deaths in 2019 alone and 10 million deaths projected every year by 2050. Current tools employed in the analysis of bacteria can be time inefficient, leading to delayed diagnosis and treatment. In this work, we develop a microfluidic setup capable of bacteria incubation and detection of growth in ~ 2 h. We fabricated polydimethylsiloxane (PDMS) microchips via soft lithography, enclosed microchannels by plasma bonding to glass, and utilized PDMS blocks for simplified connection of devices to a flow system. We generated uniform droplets enclosing zero, one or two bacteria within our devices, and incubated droplet-encapsulated bacteria with 100 × lower concentrations of a fluorescence probe of bacterial growth compared to prior work. We assessed bacterial growth via laser induced fluorescence after room temperature incubation for 2 h and obtained a range of signals corresponding to droplets with or without bacteria. Our devices allow for online droplet incubation, monitoring, detection, and tracking. Developing microfluidic chips for single bacteria studies will improve the analysis and treatment of antimicrobial resistance.

A New Method to Determine Antioxidant Activities of Biofilms Using a pH Indicator (Resazurin) Model System.

Biopolymeric films were prepared with gelatin, plasticizer, and three different types of antioxidants (ascorbic acid, phytic acid, and BHA) corresponding to different mechanisms in activity. The antioxidant activity of films was monitored for 14 storage days upon color changes using a pH indicator (resazurin). The instant antioxidant activity of films was measured by a DPPH free radical test. The system using resazurin was composed of an agar, an emulsifier, and soybean oil to simulate a highly oxidative oil-based food system (AES-R). Gelatin-based films (GBF) containing phytic acid showed higher tensile strength and energy to break than all other samples due to the increased intermolecular interactions between phytic acid and gelatin molecules. The oxygen barrier properties of GBF films containing ascorbic acid and phytic acid increased due to the increased polarity, while GBF films containing BHA showed increased oxygen permeability compared to the control. According to "a-value" (redness) of the AES-R system tested with films, films incorporating BHA showed the most retardation of lipid oxidation in the system. This retardation corresponds to 59.8% antioxidation activity at 14 days, compared with the control. Phytic acid-based films did not show antioxidant activity, whereas ascorbic acid-based GBFs accelerated the oxidation process due to its prooxidant activity. The comparison between the DPPH free radical test and the control showed that the ascorbic acid and BHA-based GBFs showed highly effective free radical scavenging behavior (71.7% and 41.7%, respectively). This novel method using a pH indicator system can potentially determine the antioxidation activity of biopolymer films and film-based samples in a food system.

Resazurin Reduction-Based Assays Revisited: Guidelines for Accurate Reporting of Relative Differences on Metabolic Status.

Cell viability and metabolic activity are ubiquitous parameters used in biochemistry, molecular biology, and biotechnological studies. Virtually all toxicology and pharmacological projects include at some point the evaluation of cell viability and/or metabolic activity. Among the methods used to address cell metabolic activity, resazurin reduction is probably the most common. At variance with resazurin, resorufin is intrinsically fluorescent, which simplifies its detection. Resazurin conversion to resorufin in the presence of cells is used as a reporter of metabolic activity of cells and can be detected by a simple fluorometric assay. UV-Vis absorbance is an alternative technique but is not as sensitive. In contrast to its wide empirical "black box" use, the chemical and cell biology fundamentals of the resazurin assay are underexplored. Resorufin is further converted to other species, which jeopardizes the linearity of the assays, and the interference of extracellular processes has to be accounted for when quantitative bioassays are aimed at. In this work, we revisit the fundamentals of metabolic activity assays based on the reduction of resazurin. Deviation to linearity both in calibration and kinetics, as well as the existence of competing reactions for resazurin and resorufin and their impact on the outcome of the assay, are addressed. In brief, fluorometric ratio assays using low resazurin concentrations obtained from data collected at short time intervals are proposed to ensure reliable conclusions.

Rapid ResaCeftazidime-Avibactam Enterobacterales NP Test: Rapid Detection of Ceftazidime-Avibactam Susceptibility in Enterobacterales.

Ceftazidime-avibactam (CZA), a novel ß-lactam/ß-lactamase inhibitor combination, has good antibacterial activity against carbapenem-resistant Enterobacterales (CRE) producing class A and C and some class D carbapenemases, but in recent years, the emergence of CZA-resistant Enterobacterales bacteria is growing. Therefore, rapid, accurate, and timely detection of CZA is necessary for clinical anti-infection treatment. In this study, the rapid ResaCeftazidime-avibactam Enterobacterales NP test was developed; its principle is that metabolically active bacteria (CZA-resistant strains) can change resazurin-PrestoBlue, a viability colorant, from blue to purple or pink in the presence of CZA, whereas CZA-susceptible strains cannot. We used 178 Enterobacterales isolates to evaluate the performance of this test. This test allowed the susceptibility of Enterobacterales to CZA to be detected within 4.5 h with an overall performance of 96% category agreement (CA), 7% major errors (MEs), and 0% very major errors (VMEs). Performance for Escherichia coli included 100% CA and 0% MEs and VMEs. Performance for Klebsiella pneumoniae included 99% CA and 2% MEs and 0% VMEs. Performance for Enterobacter cloacae included 87% CA, 25% MEs, and 0% VMEs. Moreover, this test is both economical ($1.0106 per isolate) and convenient, as it only requires basic laboratory equipment. In a word, the rapid ResaCeftazidime-avibactam Enterobacterales NP test is rapid and feasible, which may provide certain backing for the rapid screening and timely treatment of CZA-resistant strains in the clinic.

Resazurin rapid screening for antibacterial activities of organic and inorganic nanoparticles: Potential, limitations and precautions.

Nanoparticles have been used as antibacterial agents in several products. To optimize their effectiveness, synthesis processes and particle modifications have been developed, creating the need for a rapid screening method to investigate their potencies. Owing to the opacity and insolubility of nanoparticles, a classical method to determine antibacterial activity-such as the minimum inhibitory concentration (MIC), which relies on turbidimetry-might not apply to them. In this study, we demonstrate the potential of a dye (resazurin)-based assay as an indicator of bacterial growth to rapidly screen the antibacterial activities of both organic and inorganic nanomaterials against both gram-negative (E. coli) and gram-positive (S. aureus) bacteria. The results indicate that the resazurin-based assay successfully determine the MIC of organic lipid nanocarriers, and several inorganic nanoparticles. However, the use of resazurin require a precaution for nanoparticles with photocatalytic properties, which may cause dye degradation at higher concentrations. In this study, resazurin bleaching was observed at approximately >50 mg/ml of TiO2. In summary, the modified MIC assay with resazurin can evaluate antibacterial activity of nanomaterials, whose turbidity interferer conventional MIC assay. This modification conserves an advantage of MICs assay which are simple and reliable. This would be useful for screening of antibacterial nanomaterials.

A novel yeast-based screening system for potential compounds that can alleviate human α-synuclein toxicity.

AIMS: This study aimed to establish a yeast-based screening system for potential compounds that can alleviate the toxicity of α-synuclein (α-syn), a neuropathological hallmark of Parkinson's disease, either inhibition of α-syn aggregation or promotion of ubiquitin-mediated degradation of α-syn. METHODS AND RESULTS: A powerful yeast-based screening assay using the rsp5A401E -mutant strain, which is hypersensitive to α-syn aggregation, was established by two-step gene replacement and further overexpressed the GFP-fused α-syn in the drug-sensitive yeast strain with a galactose-inducible multicopy plasmid. The rsp5A401E -mutant strain treated with baicalein, a known α-syn aggregation inhibitor, showed better α-syn toxicity alleviation than the same background wild type strain as accessed by comparison on the reduction kinetics of viable dye resazurin fluorometrically (λex 540/λem 590 nm). The rsp5A401E -mutant yeast-based assay system showed high sensitivity as it could detect as low as 3.13 µmol l-1 baicalein, the concentration that lower than previously report detected by the in vitro assay. CONCLUSIONS: Our yeast-based system has been effective for screening potential compounds that can alleviate α-syn toxicity with high sensitivity and specificity. SIGNIFICANCE AND IMPACT OF THE STUDY: Yeast-based assay system can be used to discover novel neuroprotective drug candidates which may be either efficiently suppress-α-syn aggregation or enhance ubiquitin-dependent degradation.

Screening and evaluating honokiol from Magnolia officinalis against Nocardia seriolae infection in largemouth bass (Micropterus Salmoides).

Nocardiosis caused by Nocardia seriolae is a major threat to the aquaculture industry. Given that prolonged therapy administration can lead to a growth of antibiotic resistant strains, new antibacterial agents and alternative strategies are urgently needed. In this study, 80 medicinal plants were selected for antibacterial screening to obtain potent bioactive compounds against N. seriolae infection. The methanolic extracts of Magnolia officinalis exhibited the strongest antibacterial activity against N. seriolae with the minimal inhibitory concentration (MIC) of 12.5 µg/ml. Honokiol and magnolol as the main bioactive components of M. officinalis showed higher activity with the MIC value of 3.12 and 6.25 µg/ml, respectively. Sequentially, the evaluation of antibacterial activity of honokiol in vivo showed that honokiol had good biosafety, and could significantly reduce the bacterial load of nocardia-infected largemouth bass (p < .001). Furthermore, the survival rate of nocardia-infected fish fed with 100 mg/kg honokiol was obviously improved (p < .05). Collectively, these results suggest that medicinal plants represent a promising reservoir for discovering active components against Nocardia, and honokiol has great potential to be developed as therapeutic agents to control nocardiosis in aquaculture.

Study of antimycobacterial, cytotoxic, and mutagenic potential of polymeric nanoparticles of copper (II) complex.

This study aimed to encapsulate and characterise a potential anti-tuberculosis copper complex (CuCl2(INH)2.H2O:I1) into polymeric nanoparticles (PNs) of polymethacrylate copolymers (Eudragit®, Eu) developed by nanoprecipitation method. NE30D, S100 and, E100 polymers were tested. The physicochemical characterisations were performed by DLS, TEM, FTIR, encapsulation efficiency and, in vitro release studies. Encapsulation of I1 in PN-NE30D, PN-E100, and PN-S100 was 26.3%, 94.5%, 22.6%, respectively. The particle size and zeta potentials were 82.3 nm and -24.5 mV for PNs-NE30D, 304.4 nm and +18.7 mV for PNs-E100, and 517.9 nm and -6.9 mV for PNs-S100, respectively. All PDIs were under 0.5. The formulations showed an I1 controlled release at alkaline pH with 29.7% from PNs-NE30D, 7.9% from PNs-E100 and, 28.1% from PNs-S100 at 1 h incubation. PNs were stable for at least 3 months. Particularly, PNs-NE30D demonstrated moderate inhibition of M. tuberculosis and low cytotoxic activity. None of the PNs induced mutagenicity.

Development and Validation of an In Vitro Resazurin-Based Susceptibility Assay against Madurella mycetomatis.

We present an in vitro susceptibility assay for Madurella mycetomatis hyphae using resazurin for endpoint reading. Using this assay, reproducible MICs were obtained for amphotericin B, itraconazole, voriconazole, posaconazole, terbinafine, and micafungin. Results were comparable with those of a 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide salt (XTT)-based susceptibility assay. The lowest MICs were obtained for itraconazole and posaconazole (MIC50, 0.016 µg/ml) followed by voriconazole (MIC50, 0.063 µg/ml). Amphotericin B, micafungin, and terbinafine appeared much less effective.

Evaluation of antimicrobial susceptibility testing of Nocardia spp. isolates by broth microdilution with resazurin and spectrophotometry.

BACKGROUND: Nocardia species are ubiquitous in natural environments and can cause nocardiosis. In the present study, the use of Resazurin salt and Spectrophotometry were proposed as alternative methods to reduce subjectivity in the interpretation of susceptibility results to antimicrobials by the broth microdilution method for Nocardia spp. RESULTS: The susceptibility of Nocardia spp. isolates to Amikacin, Ciprofloxacin, Minocycline and Trimethoprim-Sulfamethoxazole was evaluated by Minimum Inhibitory Concentration (MIC) determinations by the broth microdilution method. To verify cellular growth, the colour-changing dye Resazurin was applied, the Optical Densities were measured on a spectrophotometer, and both were compared to Clinical and Laboratory Standards Institute (CLSI) Gold Standard method (visual MIC determination). Percentages of essential and categorical agreements and interpretative categorical errors were calculated within each method (intra-reading) and between them (inter-reading). The Gold Standard visual reading demonstrated 100% of essential and categorical intra-reading agreements for Amikacin, and there was no error when compared with the alternative methods. For Ciprofloxacin, the comparison between the Gold Standard and the Spectrophotometric reading showed 91.5% of essential agreement. In the categorical intra-reading analysis for Minocycline, there were 88.1 and 91.7% in the Gold Standard and in the Spectrophotometric readings, respectively, and 86.4% of concordance between them. High rates of categorical agreement were also observed on the Trimethoprim-Sulfamethoxazole analyses, with 93.7% for the Gold Standard, 84.9% for the Resazurin readings, and 80.5% between them. CONCLUSIONS: The alternative methods with Resazurin and Spectrophotometric readings showed high agreement rates with the Gold Standard.

Rapid phenotypic antimicrobial susceptibility testing using nanoliter arrays.

Antibiotic resistance is a major global health concern that requires action across all sectors of society. In particular, to allow conservative and effective use of antibiotics clinical settings require better diagnostic tools that provide rapid determination of antimicrobial susceptibility. We present a method for rapid and scalable antimicrobial susceptibility testing using stationary nanoliter droplet arrays that is capable of delivering results in approximately half the time of conventional methods, allowing its results to be used the same working day. In addition, we present an algorithm for automated data analysis and a multiplexing system promoting practicality and translatability for clinical settings. We test the efficacy of our approach on numerous clinical isolates and demonstrate a 2-d reduction in diagnostic time when testing bacteria isolated directly from urine samples.

Drug-induced reactive oxygen species-mediated inhibitory effect on growth of Trypanosoma evansi in axenic culture system.

Trypanosoma evansi, an extracellular haemoflagellate, has a wide range of hosts receptive and susceptible to infection, in which it revealed highly inconsistent clinical effects. Drugs used for the treatment of trypanosomosis have been utilized for more than five decades and have several problems like local and systemic toxicity. In the present investigation, imatinib and sorafenib were selected as drugs as they are reported to have the potential to cause reactive oxygen species (ROS)-mediated effect in cancer cells. Both have also been reported to have potential against T. brucei, T. cruzi and Leishmania donovani. To date, imatinib and sorafenib have not evaluated for their growth inhibitory effect against T. evansi. Imatinib and sorafenib showed significant (p < 0.001) inhibition on parasite growth and multiplication with IC50 (50% inhibitory concentration) values 6.12 µM and 0.33 µM respectively against T. evansi. Both the drug molecules demonstrated for the generation of ROS in T. evansi and were found up to 65% increased level of ROS as compared with negative control in the axenic culture system. Furthermore, different concentrations of imatinib and sorafenib were found non-toxic on horse peripheral blood mononuclear cells and Vero cell lines. Also, in conclusion, our results demonstrated that imatinib- and sorafenib-induced generation of ROS contributed inhibitory effect on the growth of Trypanosoma evansi in an axenic culture system.

Optimization of a High-Throughput 384-Well Plate-Based Screening Platform with Staphylococcus aureus ATCC 25923 and Pseudomonas aeruginosa ATCC 15442 Biofilms.

In recent years, bacterial infections have become a main concern following the spread of antimicrobial resistance. In addition, bacterial biofilms are known for their high tolerance to antimicrobials and they are regarded as a main cause of recalcitrant infections in humans. Many efforts have been deployed in order to find new antibacterial therapeutic options and the high-throughput screening (HTS) of large libraries of compounds is one of the utilized strategies. However, HTS efforts for anti-biofilm discovery remain uncommon. Here, we miniaturized a 96-well plate (96WP) screening platform, into a 384-well plate (384WP) format, based on a sequential viability and biomass measurements for the assessment of anti-biofilm activity. During the assay optimization process, different parameters were evaluated while using Staphylococcus aureus and Pseudomonas aeruginosa as the bacterial models. We compared the performance of the optimized 384WP platform to our previously established 96WP-based platform by carrying out a pilot screening of 100 compounds, followed by the screening of a library of 2000 compounds to identify new repurposed anti-biofilm agents. Our results show that the optimized 384WP platform is well-suited for screening purposes, allowing for the rapid screening of a higher number of compounds in a run in a reliable manner.

Screening sediment samples used as anolytes in microbial fuel cells for microbial electron transfer activity using DREAM assay.

OBJECTIVE: The dye reduction-based electron-transfer activity monitoring (DREAM) assay was employed to screen sediment and wastewater samples functioning as anolytes in a microbial fuel cell (MFC) for their microbial electron transfer activity. RESULTS: Electron transfer to redox dyes from microbial activity reduced the dyes and the resulting extent of reduction was measured as DREAM assay coefficient. Methylene blue was decolourised, while resazurin underwent florigenic change from blue to pink to colourless upon formation of resorufin and dihydroxyresorufin. DREAM assay coefficient conformed to power density obtained in the MFC. A correlation was observed between chemical oxygen demand of the sample and the DREAM coefficient (+ 0.934) and also between DREAM coefficient and power density generated (+ 0.976). Highest DREAM coefficient and power density was observed for activated sludge. CONCLUSIONS: The DREAM assay is a rapid, sensitive and low-cost method to assess microbial electron transfer activity for inocula used as anolytes in a MFC.