In vitro assessment of the bacterial stress response and resistance evolution during multidrug-resistant bacterial invasion of the Xenopus tropicalis intestinal tract under typical stresses.

The intestinal microbiome might be both a sink and source of resistance genes (RGs). To investigate the impact of environmental stress on the disturbance of exogenous multidrug-resistant bacteria (mARB) within the indigenous microbiome and proliferation of RGs, an intestinal conjugative system was established to simulate the invasion of mARB into the intestinal microbiota in vitro. Oxytetracycline (OTC) and heavy metals (Zn, Cu, Pb), commonly encountered in aquaculture, were selected as typical stresses for investigation. Adenosine 5'-triphosphate (ATP), hydroxyl radical (OH·-) and extracellular polymeric substance (EPS) were measured to investigate their influence on the acceptance of RGs by intestinal bacteria. The results showed that the transfer and diffusion of RGs under typical combined stressors were greater than those under a single stressor. Combined effect of OTC and heavy metals (Zn, Cu) significantly increased the activity and extracellular EPS content of bacteria in the intestinal conjugative system, increasing intI3 and RG abundance. OTC induced a notable inhibitory response in Citrobacter and exerted the proportion of Citrobacter and Carnobacterium in microbiota. The introduction of stressors stimulates the proliferation and dissemination of RGs within the intestinal environment. These results enhance our comprehension of the typical stresses effect on the RGs dispersal in the intestine.

Assessment of biodegradation kinetics and mass transfer aspects in attached growth bioreactor for effective treatment of brilliant green dye from wastewater.

In this study, Bacillus licheniformis immobilized with low-density polyethylene (LDPE) was employed to degrade Brilliant Green (BG) dye from wastewater in a packed bed bioreactor (PBBR). Bacterial growth and extracellular polymeric substance (EPS) secretion were also assessed under different concentrations of BG dye. The impacts of external mass transfer resistance on BG biodegradation were also evaluated at different flow rates (0.3-1.2 L/h). A new mass transfer correlation [Formula: see text] was proposed to study the mass transfer aspects in attached-growth bioreactor. The intermediates, namely 3- dimethylamino phenol, benzoic acid, 1-4 benzenediol, and acetaldehyde were identified during the biodegradation of BG and, subsequently degradation pathway was proposed. Han - Levenspiel kinetics parameters µmax and Ks were found to be 0.185 per day and 115 mg/L, respectively. The new insight into mass transfer and kinetics support the design of efficiently attached growth bioreactor to treat a wide range of pollutants.

Soluble Extracellular Polymeric Substances Produced by Parachlorella kessleri and Chlorella vulgaris: Biochemical Characterization and Assessment of Their Cadmium and Lead Sorption Abilities.

In the present study, the potential of lead and cadmium removal by the extracellular polymeric substances (EPS) produced from Parachlorella kessleri and Chlorella vulgaris were investigated. Carbohydrates were the dominant components of EPS from both analyzed species. The contents of reducing sugars, uronic acids, and amino acids were higher in EPS synthesized by C. vulgaris than in EPS from P. kessleri. The analysis of the monosaccharide composition showed the presence of rhamnose, mannose and galactose in the EPS obtained from both species. The ICP-OES (inductively coupled plasma optical emission spectrometry) analyses demonstrated that C. vulgaris EPS showed higher sorption capacity in comparison to P. kessleri EPS. The sorption capacity of C. vulgaris EPS increased with the increase in the amount of metal ions. P. kessleri EPS had a maximum sorption capacity in the presence of 100 mg/L of metal ions. The FTIR analysis demonstrated that the carboxyl, hydroxyl, and carbonyl groups of EPS play a key role in the interactions with metal ions. The present study showed C. vulgaris EPS can be used as a biosorbent in bioremediation processes due to its biochemical composition, the presence of significant amounts of negatively charged uronic acids, and higher sorption capacity.

Extracellular Polymeric Substances Produced by the Thermophilic Cyanobacterium Gloeocapsa gelatinosa: Characterization and Assessment of Their Antioxidant and Metal-Chelating Activities.

Cyanobacteria, particularly thermophilic strains, represent an important potential source of EPSs, harboring structural complexity that predicts diverse and specific bioactive potential. The thermophilic cyanobacteria Gloeocapsa gelatinosa, isolated from a natural hot source in Ain Echfa (Tunisia), was cultivated in a cylindrical reactor, and the production of biomass and EPSs was investigated. Results revealed that the strain is amongst the most efficient EPSs producers (0.89 g L-1) and that EPSs production was not correlated with the growth phase. EPSs were sulfated heteropolysaccharides containing carbohydrates (70%) based on nine different monosaccharides, mainly mannose (22%), and with the presence of two uronic acids. EPSs were formed by two polymers moieties with a molecular weight of 598.3 ± 7.2 and 67.2 ± 4.4 kDa. They are thermostable in temperatures exceeding 100 °C and have an anionic nature (zeta potential of -40 ± 2 mV). Atomic force microscopy showed that EPSs formed multimodal lumps with 88 nm maximum height. EPSs presented high water holding capacity (70.29 ± 2.36%) and solubility index (97.43 ± 1.24%), and a strong bivalent metal sorption capacity especially for Cu2+ (91.20 ± 1.25%) and Fe2+ (75.51 ± 0.71%). The antioxidant activity of G. gelatinosa EPSs was investigated using four methods: the ß-carotene-bleaching activity, DPPH assays, iron-reducing activity, and metal-chelating activity. EPS has shown high potential as free radicals' scavenger, with an IC50 on DPPH (0.2 g L-1) three-fold lower than ascorbic acid (0.6 g L -1) and as a metal chelating activity (IC50 = 0.4 g L-1) significantly lower than EDTA. The obtained results allow further exploration of the thermophilic G. gelatinosa for several biotechnological and industrial applications.

Impact of novel deflocculant ZnO/Chitosan nanocomposite film in disperser pretreatment enhancing energy efficient anaerobic digestion: Parameter assessment and cost exploration.

This paper proposed to interpret the novel method of extracellular polymeric substance (EPS) removal in advance to sludge disintegration to enrich bioenergy generation. The sludge has been subjected to deflocculation using Zinc oxide/Chitosan nanocomposite film (ZCNF) and achieved 98.97% of solubilization which enhance the solubilization of organics. The obtained result revealed that higher solubilization efficiency of 23.3% was attained at an optimal specific energy of 2186 kJ/kg TS and disintegration duration of 30 min. The deflocculated sludge showed 8.2% higher solubilization than the flocculated sludge emancipates organics in the form of 1.64 g/L of SCOD thereby enhancing the methane generation. The deflocculated sludge produces methane of 230 mL/g COD attained overall solid reduction of 55.5% however, flocculated and control sludge produces only 182.25 mL/g COD and 142.8 mL/g COD of methane. Based on the energy, mass and cost analysis, the deflocculated sludge saved 94.1% of energy than the control and obtained the net cost of 5.59 $/t which is comparatively higher than the flocculated and control sludge.

Sludge disintegration and anaerobic digestion enhancement by alkaline-thermal pretreatment: Economic evaluation and microbial population analysis.

Alkaline-thermal pretreatment was examined for waste activated sludge (WAS) disintegration and subsequent anaerobic digestion (AD). Pretreatment at 60 °C was estimated to provide better economic benefits than higher temperature conditions. The maximum methane yield of 215.6 mL/g COD was achieved when WAS was pretreated at 60 °C and pH 10 for 24 h, which was 46.6% higher than untreated WAS. The pretreatment condition also provided the maximum net savings. The degree of sludge disintegration, considering both loosely bound-extracellular polymeric substance and soluble COD, would be a better indicator to predict anaerobic digestibility than the solubilization rate that considers soluble COD alone. Microbial analysis implied that pretreatment facilitated the growth of hydrolytic bacteria, phyla Bacteroidetes and Firmicutes. In addition, sludge pretreatment enhanced the growth of both acetoclastic and hydrogenotrophic methanogens, genera Methanosaeta and Methanobacterium. The mild AT-PT would be useful to enhance the digestion performance and economic benefit of WAS digestion.

New methods for quantification of Fenton's reagent addition based on aged sludge indicators to improve filterability.

Fenton oxidation can effectively improve the dewaterability of aged sludge. Quantification of the addition of optimal reagents is central to the conditioning and dewatering of aged sludge. Improving the accuracy of quantification is significant to promote cost effectiveness. The effects of reagent addition and the mechanism governing the improved filterability of the aged sludge need to be understood uniformly. In this study, the optimal reagent additions have been determined using the response surface method (RSM) for five out of the eight aged sludges that were investigated. The physicochemical characteristics of eight aged sludges, including the extracellular polymer substance, undissolved organic matter, and suspension structure network, were investigated. Meanwhile, a comprehensive correlation analysis of critical indicators was conducted to investigate the interactions among the properties of the aged sludge. The effects of these interactions on the conditioning and filtration processes were examined, and a unified understanding of the combination of factors affecting the optimal reagent addition was obtained. The key factors were aggregate size, dewatering extent, yield stress, and organic substance content. Based on these results, a new reagent addition quantification method was developed along with an empirical model of the relationship between physicochemical properties and the economically optimal reagent addition.

Effect of ibuprofen on extracellular polymeric substances (EPS) production and composition, and assessment of microbial structure by quantitative image analysis.

A Sequencing Batch Reactor (SBR) with activated sludge was operated with synthetic wastewater containing ibuprofen (IBU) to investigate the biomass stress-responses under long-term IBU exposure. There were 3 different phases: phase I as the control without IBU for 56 days, phase II (40 days), and phase III (60 days) containing IBU at 10 and 5 mg L-1 each. The overall performance of the SBR as well as the extracellular polymeric substances (EPS) in terms of polysaccharides, proteins, and humic acid substances were estimated. Morphological parameters of microbial aggregates in the presence of IBU (phase II and phase III) were assessed by quantitative image analysis (QIA). Removal efficiencies of chemical oxygen demand (COD) and ammonium (NH4+) were significantly reduced by IBU. Loosely bound EPS (LB-EPS) decreased during phase II and phase III, and tightly bound EPS (TB-EPS) was slightly higher in phase II than phase I. TB-EPS proteins were greater in phase II, perhaps to protect microbial cells from IBU exposure. These findings provided insight into both activated sludge stress-responses and EPS composition under long-term IBU exposure. Spearman correlation showed that EPS and morphological parameters significantly affected sludge settleability and flocculation. QIA also proved to be a powerful technique in investigating dysfunctions in activated sludge under IBU exposure.

Integrated review of resource recovery on aerobic granular sludge systems: Possibilities and challenges for the application of the biorefinery concept.

Aerobic Granular Sludge (AGS) is a biological treatment technology that has been extensively studied in the last decade. The possibility of resource recovery has always been highlighted in these systems, but real-scale applications are still scarce. Therefore, this paper aimed to present a systematic review of resources recovery such as water, energy, chemicals, raw materials, and nutrients from AGS systems, also analyzing aspects of engineering and economic viability. In the solid phase, sludge application in agriculture is an interesting possibility. However, the biosolids' metal concentration (the granules have high adsorption capacity due to the high concentration of extracellular polymeric substances, EPS) may be an issue. Another possibility is the recovery of Polyhydroxyalkanoates (PHAs) and Alginate-like exopolymers (bio-ALE) in the solid phase, emphasizing the last one, which has already been made in some Wastewater Treatment Plants (WWTPs), named and patented as Kaumera® process. The Operational Expenditure (OPEX) can be reduced by 50% in the WWTP when recovery of ALE is made. The ALE recovery reduced sludge yield by up to 35%, less CO2 emissions, and energy saving. Finally, the discharged sludge can also be evaluated to be used for energetic purposes via anaerobic digestion (AD) or combustion. However, the AD route has faced difficulties due to the low biodegradability of aerobic granules.

Optimization of biochemical sulfide potential (BSP) assay for anaerobic biodegradability assessment.

The anaerobic biodegradability assessment (biodegradation extent and kinetics) of organic wastes is critical for optimum design and evaluating treatment efficiencies for anaerobic treatment technologies. The biochemical sulfide potential (BSP) assay has previously demonstrated the advantages of its time efficiency and measurement accuracy for biologically assessing substrate degradability, while its application is limited by undefined operational parameters. In this study, the BSP assay was further optimized through a systematic investigation of a critical parameter, inoculum-to-substrate ratio (ISR), and the applicable kinetic model to unravel the potential use of BSP assays for anaerobic waste treatment. Under two series of experimental scenarios, the common ISR ranges of 0.5-4.0 (based on the traditional BMP assay) and extreme ISRs (as low as 0.1) were studied, in which the advantage of a BSP assay on extreme ISRs was highlighted. Meanwhile, the underlying cause and mechanism for biodegradability discrepancies under different ISRs (0.1-6.0) were further investigated. The extracellular polymeric substance (EPS) characterization of residual organics and the two-substrate first-order hydrolysis model analyses revealed that the hydrolysis process of slowly-biodegradable organics fraction was hindered under improper ISR conditions. Furthermore, the Cone model was evaluated as more appropriate for biodegradation kinetics analysis in BSP assays among the five common kinetic models (i.e., Exponential, Fitzhugh, Cone, Transference, and modified Gompertz models). Overall, the results provide fundamental guidance on designing consistent BSP assays and put a step forward in standardizing the BSP assay for anaerobic biodegradability assessments.

Exploring the Role of Bacterial Extracellular Polymeric Substances for Sustainable Development in Agriculture.

The incessant need to increase crop yields has led to the development of many chemical fertilizers containing NPK (nitrogen-phosphorous-potassium) which can degrade soil health in the long term. In addition, these fertilizers are often leached into nearby water bodies causing algal bloom and eutrophication. Bacterial secondary metabolites exuded into the extracellular space, termed extracellular polymeric substances (EPS) have gained commercial significance because of their biodegradability, non-toxicity, and renewability. In many habitats, bacterial communities faced with adversity will adhere together by production of EPS which also serves to bond them to surfaces. Typically, hygroscopic, EPS retain moisture in desiccating conditions and modulate nutrient exchange. Many plant growth-promoting bacteria (PGPR) combat harsh environmental conditions like salinity, drought, and attack of pathogens by producing EPS. The adhesive nature of EPS promotes soil aggregation and restores moisture thus combating soil erosion and promoting soil fertility. In addition, these molecules play vital roles in maintaining symbiosis and nitrogen fixation thus enhancing sustainability. Thus, along with other commercial applications, EPS show promising avenues for improving agricultural productivity thus helping to address land scarcity as well as minimizing environmental pollution.

Optimization of a real-time high-throughput assay for assessment of Streptococcus mutans metabolism and screening of antibacterial dental adhesives.

OBJECTIVE: The present work shows the optimization of a high-throughput bioluminescence assay to assess the metabolism of intact Streptococcus mutans biofilms and its utility as a screening method for nanofilled antibacterial dental materials. METHODS: The assay was optimized by monitoring changes in bioluminescence mediated by variation of the experimental parameters investigated (growth media and sucrose concentration, inoculum:D-Luciferin ratio, dilution factor, inoculum volume, luminescence wavelength, replicate and luciferase metabolic activity). Confocal microscopy was then used to demonstrate the impact of biofilm growth conditions on the 3-D distribution of extracellular polymeric substance (EPS) within Streptococcus mutans biofilms and its implications as confounding factors in high-throughput studies (HTS). RESULTS: Relative Luminescence Unit (RLU) values from the HTS optimization were analyzed by multivariate ANOVA (α = 0.05) and coefficients of variation, whereas data from 3-D structural parameters and RLU values of biofilms grown on experimental antibacterial dental adhesive resins were analyzed using General Linear Models and Student-Newman-Keuls post hoc tests (α = 0.05). Confocal microscopy demonstrated that biofilm growth conditions significantly influenced the quantity and distribution of EPS within the 3-D structures of the biofilms. An optimized HTS bioluminescence assay was developed and its applicability as a screening method in dentistry was demonstrated using nanofilled experimental antibacterial dental adhesive resins. SIGNIFICANCE: The present study is anticipated to positively impact the direction of future biofilm research in dentistry, because it offers fundamental information for the design of metabolic-based assays, increases the current levels of standardization and reproducibility while offering a tool to decrease intra-study variability.

Characterization and assessment of barnacle larval settlement-inducing activity of extracellular polymeric substances isolated from marine biofilm bacteria.

Extracellular polymeric substances (EPSs) are the hydrated gelatinous matrix produced by microorganisms for attachment in a biofilm environment. In this study, the compositional variation between EPSs of three marine biofilm bacteria (Pseudoalteromonas shioyasakiensis, Vibrio harveyi and Planomicrobium sp.) were analysed by GC-MS, 1H NMR, FT-IR and XRD and SEM. The ecological significance of exopolymers was assessed in vivo using marine model organism barnacle larvae for their settlement-inducing activity. Chemical analysis revealed the presence of glycan fucosylated oligosaccharides, tetraose, trisaccharides, iso-B-Pentasaccharides, sialyllactose, oligomannose, galacto-N-biose, difucosyl-para-lacto-N-neohexaose, 3'-sialyl N-acetyllactosamine and isoglobotriaose-ß-N(Acetyl)-Propargyl in all extracted EPSs. Bioassay results indicated that treatment of the barnacle larvae with EPSs from three bacterial strains enhanced settlement on substrates. In conclusion, this study highlighted the role of water-soluble EPSs in the invertebrate larval settlement on artificial materials.