Linkages of volatile fatty acids and polyhexamethylene guanidine stress during sludge fermentation: Metagenomic insights of microbial metabolic traits and adaptation

The massive use of polyhexamethylene guanidine (PHMG), as a typical bactericidal agent, raised environmental concerns to the public. This work comprehensively revealed the hormesis effects of PHMG occurred in waste activated sludge (WAS) on the generation of volatile fatty acids (VFAs) during anaerobic fermentation. The low level of PHMG (100 mg/g TSS) significantly promoted the VFAs generation (1283 mg COD/L, compared with 337 mg COD/L in the control) via synchronously facilitating the solubilization, hydrolysis, and acidification steps but inhibiting methanogenesis. Metagenomic analysis showed that the functional anaerobe (i.e., Bacteroides, Macellibacteroide and Parabacteroide) and corresponding genetic expressions responsible for extracellular hydrolysis (i.e., clpP), membrane transport (i.e., ffh and gspF), intracellular substrates metabolism (i.e., ald and paaF) and VFAs biosynthesis (i.e., ACACA and FASN) were enhanced in the optimal presence of PHMG. Moreover, the anaerobic species could respond and adapt to low PHMG stimuli via quorum sensing (i.e., cqsA, rpfC and rpfG), and thus maintain the high microbial metabolic activities. However, they were unable to tolerate the toxicity of excessive PHMG, resulting in the extremely low VFAs production. This work enlightened the effects of emerging pollutants on WAS fermentation at the genetic levels, and provided guidance on the WAS treatment and resource recovery.     

Enzymatic pretreatment mitigates the dissemination of antibiotic resistance genes via regulating microbial populations and gene expressions during food waste fermentation

Food waste (FW) has been recognized as essential reservoir for resource recovery via anaerobic fermentation, which could also bring the potential risk of antibiotic resistance genes (ARGs) dissemination. Although the structural deficiency of FW could be stimulated by enzymatic pretreatment to enhance fermentation efficiency, the influences of enzymatic pretreatment on ARGs fate and microbial metabolic pathways involved in ARGs dissemination have rarely been reported. This work proved that enzymatic pretreatment could effectively decrease the total abundance of ARGs (reduced by 13.8%-24.5%) during long-term FW fermentation. It was found that enzymatic pretreatment significantly reduced the ARGs belonging to the efflux pump, which might be ascribed to its ability to increase membrane permeability. Furthermore, enzymatic pretreatment was in favor of reducing microbial diversity and various potential ARGs host (e.g., Methanosarcina, Clostridium, Prevotella, Parabacteroides). Also, this pretreatment remarkably up-regulated the genetic expressions involved in ABC transporter (e.g., eryF and mntA) and down-regulated the genetic expressions that participated in DNA replication, two-component systems (e.g., uphA and cckA), and quorum sensing (e.g., rpfF and lsrG), thereby decreasing ARGs transmission. This study would expand the insight of the influences of pretreatment method on ARGs fate during FW fermentation, and offer practical guidance on the sustainable management of FW.     

Simultaneous determination of cefamandole and cefamandole nafate in human plasma and urine by high-performance liquid chromatography with column switching

A high-performance liquid chromatographic method with column switching has been developed for the simultaneous determination of cefamandole and cefamandole nafate in plasma and urine. The plasma and urine samples were injected onto a precolumn packed with Corasil RP C18 (37-50 microns) after simple dilution with an internal standard solution in 0.05 M phosphoric acid. Polar plasma and urine components were washed out using 0.05 M phosphoric acid. After valve switching, the concentrated drugs were desorbed in back-flush mode and separated by a reversed-phase C8 column with methanol-5 mM tetrabutylammonium bromide (45:55, v/v) as the mobile phase. The method showed excellent precision with good sensitivity and speed, and a detection limit of 0.5 microgram/ml. The total analysis time per sample was less than 30 min, and the mean coefficients of variation for intra- and inter-assay were both less than 4.9%. The method has been successfully applied to plasma and urine samples for human volunteers after intravenous injection of cefamandole nafate.     

Application of quantitative IR spectral analysis of bacterial cells to acetone-butanol-ethanol fermentation monitoring

Clostridium acetobutylicum ATCC 824 was grown under three different acetone-butanol-ethanol (ABE) fermentation conditions: (1) strictly anaerobic conditions with vegetative inoculum; (2) semi-anaerobic conditions with vegetative inoculum; and (3) strictly anaerobic conditions with spore inoculum. Semi-anaerobic fermentation with vegetative inoculum and strictly anaerobic fermentation with spore inoculum produced solvents at high level. Strictly anaerobic fermentation with vegetative inoculum showed an “acid crash”, i.e. produced mainly acids and did not switch to predominant solvent production. The content of carbohydrates, nucleic acids, proteins and lipids in Clostridium cells during the fermentation were evaluated from the mid-IR spectra. The content of nucleic acids decreased with process time, and the lipid content increased, corresponding to ceasing growth and formation of the toxic fermentation products. It was shown that the physiological states of either solvent production or acid crash are reflected in the microbial biomass composition, which can be assessed by IR spectroscopy.     

A review of ARGs in WWTPs: Sources,stressors and elimination

Antibiotic resistance genes (ARGs) in aquatic environments, which seriously endanger human health and ecological safety, have become a worldwide concern due to their easy diffusion and proliferation. Wastewater treatment plants (WWTPs), which receive resistant bacteria and ARGs from a wide variety of sources (i.e., livestock farms, hospitals, antibiotic manufactures, and households), are regarded as important emission sources of aquatic ARGs. This review presents a quantitative profile of the majority sources of ARGs in the influent of WWTPs and discusses the potential factors that affect the concentration distribution of ARGs. Specifically, a noteworthy existence of ARGs, which ranged from 1E + 05 to 1E + 11 copies/mL, was detected in livestock breeding wastewater, and household wastewater (caused by the unlimited utilization of antibiotics) was determined to be the predominant contributor of ARGs in WWTPs. We summarized the selective pressure on ARGs and determined the positive correlation of the concentration of ARGs and the existence of many containments, including antibiotics, heavy metals (Zn and Cu were frequently reported), quaternary ammonium compounds, etc. In the last section, physical, chemical, and biological treatments for the removal of ARGs and their effluent in WWTPs are discussed and prospective future studies are summarized.     

Effectively Converting Cane Molasses into 2,3-Butanediol Using Clostridium ljungdahlii by an Integrated Fermentation and Membrane Separation Process

Firstly, 2,3-butanediol (2,3-BDO) is a chemical platform used in several applications. However, the pathogenic nature of its producers and the expensive feedstocks used limit its scale production. In this study, cane molasses was used for 2,3-BDO production by a nonpathogenic Clostridium ljungdahlii. It was found that cane molasses alone, without the addition of other ingredients, was favorable for use as the culture medium for 2,3-BDO production. Compared with the control (i.e., the modified DSMZ 879 medium), the differential genes are mainly involved in the pathways of carbohydrate metabolism, membrane transport, and amino acid metabolism in the case of the cane molasses alone. However, when cane molasses alone was used, cell growth was significantly inhibited by KCl in cane molasses. Similarly, a high concentration of sugars (i.e., above 35 g/L) can inhibit cell growth and 2,3-BDO production. More seriously, 2,3-BDO production was inhibited by itself. As a result, cane molasses alone with an initial 35 g/L total sugars was suitable for 2,3-BDO production in batch culture. Finally, an integrated fermentation and membrane separation process was developed to maintain high 2,3-BDO productivity of 0.46 g·L⁻¹·h⁻¹. Meanwhile, the varied fouling mechanism indicated that the fermentation properties changed significantly, especially for the cell properties. Therefore, the integrated fermentation and membrane separation process was favorable for 2,3-BDO production by C. ljungdahlii using cane molasses.     

Simulating the Degradation of Odor Precursors in Primary and Waste-Activated Sludge During Anaerobic Digestion

Degradation of known odor precursors in sludge during anaerobic digestion was systematically studied and simulated using the Anaerobic Digestion Model Number 1 (ADM1). The degradation of various protein fractions (particulate, soluble, and bound), volatile fatty acids (VFAs), lipids, and amino acids of primary sludge (PS) and waste-activated sludge (WAS) were monitored during anaerobic digestion. The degradation kinetic constants of the odor precursors namely, protein, lipid, and VFAs were determined. Relationships between degradations of protein fractions and volatile suspended solid were established; a strong relationship between bound protein, a major odor precursor, and volatile suspended solid degradation was found. No statistically significant difference in bound protein reduction was observed between PS and WAS. ADM1 was successfully used to simulate the lab scale continuous anaerobic digestion; model results with optimized parameters showed good agreement with the experimental data for methane production and several other sludge parameters including odor precursors such as lipids, VFAs, and proteins.     

Fermentation condition outweighed truffle species in affecting volatile organic compounds analyzed by chromatographic fingerprint system

The influences of fermentation conditions and truffle species (i.e., Tuber melanosporum, Tuber sinense, Tuber indicum, and Tuber aestivum) on the volatile organic compounds (VOCs) originated from truffle fermentation mycelia were studied by using chromatographic fingerprint system for the first time. Gas chromatography combined with statistical methods including similarity analysis and hierarchical cluster analysis was applied to develop chromatographic fingerprint system for truffle VOCs evaluation. Fermentation conditions affected the VOCs from truffle fermentation mycelia much more significantly than truffle species. This indicated that it is possible to adjust the aroma of truffle fermentation mycelia similar with the natural fruiting-body through the control of fermentation process.     

Application of the ‘Zero-Crossing’ Derivative Spectrophotometry to the Analysis of Mixtures of Cefoperazone and Cefamandole Nafate in Pure and Pharmaceutical Dosage Forms

Abstract

‘Zero - crossing’ first and second derivative spectrophotometry has been employed for the quantitation of mixtures of cefoperazone Na and cefamandole nafate. Beer's law holds for up to 44 ug/ml of each cephalosporin, in both derivative modes. The method has been applied to the recovery of these antibiotics in mixtures of injectable dosage forms.     

Biosynthesis of Polyhydroxyalkanoates (PHAs) by the Valorization of Biomass and Synthetic Waste

Synthetic pollutants are a looming threat to the entire ecosystem, including wildlife, the environment, and human health. Polyhydroxyalkanoates (PHAs) are natural biodegradable microbial polymers with a promising potential to replace synthetic plastics. This research is focused on devising a sustainable approach to produce PHAs by a new microbial strain using untreated synthetic plastics and lignocellulosic biomass. For experiments, 47 soil samples and 18 effluent samples were collected from various areas of Punjab, Pakistan. The samples were primarily screened for PHA detection on agar medium containing Nile blue A stain. The PHA positive bacterial isolates showed prominent orange–yellow fluorescence on irradiation with UV light. They were further screened for PHA estimation by submerged fermentation in the culture broth. Bacterial isolate 16a produced maximum PHA and was identified by 16S rRNA sequencing. It was identified as Stenotrophomonas maltophilia HA-16 ({"type":"entrez-nucleotide","attrs":{"text":"MN240936","term_id":"1712425297","term_text":"MN240936"}}MN240936), reported first time for PHA production. Basic fermentation parameters, such as incubation time, temperature, and pH were optimized for PHA production. Wood chips, cardboard cutouts, plastic bottle cutouts, shredded polystyrene cups, and plastic bags were optimized as alternative sustainable carbon sources for the production of PHAs. A vital finding of this study was the yield obtained by using plastic bags, i.e., 68.24 ± 0.27%. The effective use of plastic and lignocellulosic waste in the cultivation medium for the microbial production of PHA by a novel bacterial strain is discussed in the current study.     

Evaluation and Functional Characterization of a Biosurfactant Produced by Lactobacillus plantarum CFR 2194

The study details the investigations on the ability of Lactobacillus plantarum CFR 2194, an isolate from kanjika, a rice-based ayurvedic fermented product, to produce biosurfactant. Surfactant production, as a function of fermentation time, indicates that the maximum production occurred at 72 h under stationary conditions. Isolation, partial purification, and characterization of the biosurfactant produced have been carried out, and Fourier transform infrared spectroscopy (FTIR) spectra demonstrated that biosurfactants were constituted by protein and polysaccharide fractions, i.e., possessed the structure typical of glycoprotein, which is affected by the medium composition and the phase of growth of the biosurfactant-synthesizing strain. Critical micelle concentration (cmc) of the biosurfactant was found to be 6 g l^?1. The emulsification index (EI), emulsification activity (EA), and emulsion stability (ES) values of the biosurfactant have confirmed its emulsification property. Aqueous fractions of the produced biosurfactant exhibited a significant antimicrobial activity against the food-borne pathogenic species: Escherichia coli ATCC 31705, E. coli MTCC 108, Salmonella typhi, Yersinia enterocolitica MTCC 859, and Staphylococcus aureus F 722. More importantly, the biosurfactant from L. plantarum showed antiadhesive property against above food-borne pathogens. The results thus indicate the potential for developing strategies to prevent microbial colonization of food contact surfaces and health-care prosthesis using these biosurfactants.     

The use of an automatic on-line system for monitoring penicillin cultivation in a bubble-colum loop reactor

Penicillium chrysogenum is cultivated by a fed-batch mode to produce penicillin V. During fermentation, the concentrations of the medium components must be held at predetermined levels, which will change during fermentation, e.g., in the growth phase the concentration of the carbon sources and the nitrogen sources (urea and ammonium) must be high enough to maximize biomass production, whereas in the production stage these sources should be limited. To achieve optimal substrate concentrations, continuous measurement of various components in the fermentation broth is necessary. This is done by using a sterilizable ultrafiltation sampling probe and an air-segmented automatic flow analysis system to determine reducing sugars, dissolved organic carbon, ammonium, urea, sulfate, phosphate and penicillin V concentrations; spectrophotometric and gas-sensing electrodes are used in order to guarantee dependable results throughout the 290-h fermentation process, the analysis system is automatically cleaned and calibrated, and blanks are determined. The results are stored and evaluated by computer.     

Effect of Salt Addition and Fermentation Time on Phenolics,Microbial Dynamics,Volatile Organic Compounds,and Sensory Properties of the PDO Table Olives of Gaeta (Italy)

‘Oliva di Gaeta’ is almost certainly the most important and well-known PDO denomination for table olives in Italy. Their production is based on a specific two-stage trade preparation called the ‘Itrana’ method. In this work, we investigated how variations in the duration of the initial water fermentation (i.e., 15 and 30 days) and the salt concentration (i.e., 6% and 8% NaCl) influence the chemical features, microbial dynamics, polyphenols, volatile organic compounds, and sensory features of ‘Oliva di Gaeta’. The time of the addition of salt did not affect the final concentration in the brine, but a longer initial water fermentation (before salt addition) led to lower pH values. The bacterial count constantly increased until the salt addition (i.e., either 15 or 30 days), while the yeast population peaked on day 30. Generally, the two different salt concentrations did not affect the count of microorganisms at the end of fermentation, with the only exception being a higher lactic acid bacteria count for the treatment with 6% salt added at 30 days. At commercial maturity, the crucial bitter tastant oleuropein was not completely removed from the drupes, and differences in salt concentration and the length of the first-stage water fermentation did not influence its content at the end of olive curing. Richer volatile profiles of olives were detected with higher-salt treatments, while the combination of low salt and early saline treatment provided a more distinct profile. Longer initial water fermentation caused a small increase in some phenolic compounds (e.g., iso-verbascoside, verbascoside, and hydroxytyrosol-glucoside). A panel test indicated that salt application at 30 days resulted in a more “Sour” and “Bitter” taste, irrespective of the salt concentration. The low salt concentration coupled with the late saline treatment resulted in more “Fruity” notes, probably due to the higher production of esters by lactobacilli. The slightly bitter perception of the olives was consistent with the partial removal of oleuropein. Our work revealed the characteristics of the ‘Itrana’ method and that the variation in salt concentration and its time of application changes parameters ranging from the microbial dynamics to the sensory profile. Specifically, our data indicate that 6% NaCl coupled with a longer initial water fermentation is the most different condition: it is less effective in blocking microbial growth but, at the same time, is more potent in altering the nutritional (e.g., polyphenols) and sensorial qualities (e.g., bitterness and fruitiness) of ‘Oliva di Gaeta’.     

Xanthan gum production from cassava bagasse hydrolysate with Xanthomonas campestris using alternative sources of nitrogen

Cassava bagasse was hydrolyzed using HCl and the hydrolysate was used for the production of xanthan gum using a bacterial culture of Xanthomonas campestris. Cassava bagasse hydrolysate with an initial concentration of approx 20 g of glucose/L proved to be the best substrate concentration for xanthan gum production. Among the organic and inorganic nitrogen sources tested to supplement the medium—urea, yeast extract, peptone, potassium nitrate, and ammonium sulfate—potassium nitrate was most suitable. Ammonium sulfate was the least effective for xanthan gum production, and it affected sugar utilization by the bacterial culture. In media with an initial sugar concentration of 48.6 and 40.4 g/L, at the end of fermentation about 30 g/L of sugars was unused. Maximum xanthan gum (about 14 g/L) was produced when fermentation was carried out with a medium containing 19.8 g/L of initial reducing sugars supplemented with potassium nitrate and fermented for 72 h, and it remained almost the same until the end of fermentation (i.e., 96 h).     

Polyampholyte-tuned lyotrop lamellar liquid crystalline systems

The influence of a polyampholyte, i.e., poly(N,N′-diallyl-N,N′-dimethyl-altmaleamic carboxylate) (PalH), on the lamellar liquid crystalline (LC) system sodium dodecyl sulfate (SDS)/decanol/water was investigated by means of microdifferential scanning calorimetry, small-angle X-ray diffraction (SAXS), and cryo-scanning electron microscopy. After incorporating PalH into the lamellar liquid crystalline system, SAXS measurements show that three different LC phases exist: i.e., a swelling, slightly swelling, and non-swelling one. At pH 4, the positively charged polymer with an extended conformation can directly adsorb at the anionic head groups of the surfactant and more compact vesicles are formed at room temperature. At pH 9, the electrostatic interactions between the polyampholyte (in a more coiled conformation) and the sulfate head groups of the SDS are leveled off and incompact vesicles are formed at room temperature. That means in presence of the polyampholyte the morphology of the LC phase, i.e., the supramolecular vesicle structure, can be tuned by varying the pH and/or the temperature.

Effects of Salts Contained in Lignocellulose-Derived Sugar Streams on Microbial Lipid Production

This study aimed at developing low-cost, robust non-sterile fermentation processes for microbial lipid production from lignocellulose-derived sugars. Three representative oleaginous yeasts, Lipomyces tetrasporus (NRRL Y-11562), Rhodotorula toruloides (NRRL Y-1091), and Yarrowia lipolytica (NRRL YB-437), were tested for lipid production via non-sterile fermentation. Under optimal non-sterile conditions, all the tested strains had good performance on salt tolerance and lipid production. L. tetrasporus (NRRL Y-11562) gave the highest lipid titer of 12.79 g/L along with the depletion of both glucose and xylose, while Y. lipolytica (NRRL YB-437) showed the lowest lipid production and limited capability of xylose utilization. The key factors, including inoculation size, initial pH, and salt, all contributed to successful non-sterile fermentation. This study demonstrated that it is feasible to perform both sterile and non-sterile fermentation for lipid production using salt-containing lignocellulose-derived sugar streams.     

Biohydrogen Production Through Dark Fermentation by a Microbial Consortium Using Whey Permeate as Substrate

Nowadays, hydrogen produced globally has been synthesized from fossil fuel with limited source. Therefore, research has been developed in order to explore biological H₂ production by dark fermentation. The purpose of this work was to evaluate the effect of initial pH and ferrous sulfate and ammonium sulfate concentrations on the production of biohydrogen by dark fermentation. The process was carried out in batch mode under anaerobic conditions, in the absence of light, and at standard room temperature and pressure. A microbial consortium provided by the effluent treatment plant of a local dairy company was inoculated into a synthetic medium supplemented with cheese whey permeate (20 g/L of lactose) as a carbon source. The influence of three variables was analyzed by a central composite design 2⁽³⁾, and the optimum results of hydrogen yield (4.13 mol H₂/mol lactose) and productivity (86.31 mmol H₂/L/day) were achieved at initial pH 7.0 and FeSO₄ and (NH₄)₂SO₄ concentrations of 0.6 and 1.5 g/L, respectively. Under these conditions, the kinetic parameters of fermentation were investigated by analyzing the profile of H₂ yield and productivity, metabolite concentrations, pH, and concentration of dissolved iron. In the kinetic analysis, the modified Gompertz equation described adequately the fermentative hydrogen production from cheese whey permeate (R ²?=?0.98). The profile of ethanol and volatile organic acids showed that lactic acid and butyric acid were the main metabolites produced, and the sum of both by-products corresponded to about 58 % of the total metabolites.     

Enzymatic Hydrolysis and Succinic Acid Fermentation from Steam-Exploded Corn Stalk at High Solid Concentration by Recombinant Escherichia coli

Steam-exploded corn stalk biomass was used as the substrate for succinic acid production via lignocellulose enzymatic hydrolysis and fermentation. Succinic acid fermentation was investigated in Escherichia coli strains overexpressing cyanobacterium Anabaena sp. 7120 ecaA gene encoding carbonic anhydrase (CA). For the washed steam-exploded corn stalk at 30 % substrate concentration, i.e., 30 % water-insoluble solids (WIS), enzymatic hydrolysis yielded 97.5 g/l glucose solution and a cellulose conversion of 73.6 %, thus a high succinic acid level up to 38.6 g/l. With the unwashed steam-exploded corn stalk, though a cellulose conversion of 71.2 % was obtained in hydrolysis at 30 % solid concentration (27.9 % WIS), its hydrolysate did not ferment at all, and the hydrolysate of 25 % solid loading containing 3.8 g/l acetic acid and 168.2 mg/l furfural exerted a strong inhibition on succinic acid production.     

Production of Acid-Stable and High-Maltose-Forming α-Amylase of Bacillus acidicola by Solid-State Fermentation and Immobilized Cells and Its Applicability in Baking

Among matrices used for immobilizing Bacillus acidicola cells [calcium alginate, chitosan + alginate, scotch brite, and polyurethane foam (PUF)], ??-amylase production was highest by PUF-immobilized cells (9.1?U?ml^?1), which is higher than free cells (7.2?U?ml^?1). The PUF-immobilized cells could be reused over seven cycles with sustained ??-amylase production. When three variables (moisture, starch, and ammonium sulfate), which significantly affected enzyme production in solid-state fermentation (SSF), were optimized using response surface methodology, 5.6-fold enhancement in enzyme production was attained. The enzyme production in SSF is 3.8-fold higher than that in submerged fermentation. The bread made by supplementing dough with ??-amylase of B. acidicola scored better than those with the xylanase of Bacillus halodurans and thermostable ??-amylase of Geobacillus thermoleovorans.     

Effect of graphitic carbon nitride powders on adsorption removal of antibiotic resistance genes from water

There is a growing need to eliminate antibiotic resistance genes (ARGs) in the environment and mitigate widespread antibiotic resistance. Graphitic carbon nitride (g-C₃N₄) was successfully synthesized via facile thermal polymerization approach and its potential for adsorption treatment of ARGs in water was examined. Batch adsorption experimental results revealed that g-C₃N₄ powders had robust adsorption activity for the gene ampC and ermB. Adsorption kinetics and isotherms were systematically investigated to explain the adsorption mechanism. The apparent adsorption equilibrium could be reached within 180 min. The adsorption process effectively removed ARGs (ampC and ermB) from water with 3.2 log and 4.2 log reductions, respectively. In addition, experimental data were analyzed by several models and simulated well with Langmuir isotherm and pseudo-second-order model. It indicated that adsorption process might be dominated by the chemical rate-limiting step. Moreover, the effects of temperature and pH on the removal of ARGs were conducted and the isoelectric point (IEP) was obtained. Finally, we have demonstrated that the g-C₃N₄ is a novel adsorbent and can be used as column packing to remove ARGs by filtration.