Triple action of FeCl3-assisted hydrothermal treatment of digested sludge for deep dewatering.

In this study, a FeCl3-assisted hydrothermal treatment (HTT) process under mild conditions (90 °C-130 °C) was developed for deep dewatering of anaerobically digested sludge. HTT of sludge at 90 °C-130 °C with 4%-6% Fe3+ ions loading based on total sludge solids followed by mechanical dewatering reduced sludge water content from 82% to 38%-53% and sludge weight by 62%-72%. The treatment increased the flowability of sludge through reduction of apparent viscosity and disintegration of colloidal forces between sludge particles. This study unveiled that FeCl3-assisted HTT process had three mechanisms for improving sludge dewaterability and flowability. The treatment hydrolysed sludge flocs in the presence of Lewis acid FeCl3 and high temperature (90-130 °C). Fe3+ ions also improved dewaterability through the formation of double electric layers and neutralisation of surface negative charges, leading to flocculation of sludge flocs. More importantly, the hydrolysed sludge components produced during HTT process acted as reducing agents and led to in-situ generation of iron oxyhydroxide nanoparticles through reduction-oxidation reactions, further enhancing flocculation/co-precipitation of sludge flocs. The treatment reduced EPS content and changed conformational structures of EPS proteins by breaking down hydrogen bond-maintaining α-helix which led to a loose EPS protein structure and enhanced hydrophobicity and flocculability. Furthermore, the FeCl3-assisted treatment promoted immobilisation of the majority of heavy metals in the sludge matrix through co-precipitation/complexation reactions with iron species and organic/inorganic matters. This indicates that the FeCl3-assisted treatment reduced direct toxicity/bioavailability of the majority of heavy metals and the treated sludge may be suitable for land application. Overall, this study provides new insights into mechanism of FeCl3-assisted HTT process for dewaterability of anaerobically digested sludge and immobilisation of heavy metals.

Valorisation of keratin waste: Controlled pretreatment enhances enzymatic production of antioxidant peptides.

Conversion of keratin waste to value-added products not only reduces waste volumes but also creates new revenue streams for the animal production industry. In the present study, combination of alkaline pretreatment of cattle hair with enzymatic hydrolysis was studied to produce keratin hydrolysates with relatively high antioxidant activities. Firstly, the effect of pretreatment conditions at a high solid/liquid mass ratio of 1:2 with different NaOH loadings and temperatures was studied. Increasing NaOH concentration from 1.0% to 2.5% and temperature from room temperature to 110 °C increased hair hydrolysis by keratinase and protein recovery in hydrolysates. Mild pretreatment with 1.5% NaOH at 70 °C for 30 min led to a protein recovery of 30% in the enzymatic hydrolysate. The resulting hydrolysate showed a high antioxidant activity, scavenging 69% of the ABTS radical with a low EC50 of 0.8 mg/mL. Severe pretreatment with 2.5% NaOH at 110 °C for 30 min resulted in a higher protein recovery of 45%, but a lower ABTS radical scavenging activity of 56% and a higher EC50 of 1.3 mg/mL. The reduced antioxidant activity was attributed to the reduced proportion of small peptides (<3 kDa) and the increased extent of amino acid chemical modification. This study demonstrated that controlling alkali pretreatment conditions could lead to the production of enzymatic hydrolysates with higher antioxidant activities for potential value-adding applications. The information generated from this study will aid scale-up and commercialisation of processes with optimised antioxidant peptide production.

Effect of ferrous iron loading on dewaterability, heavy metal removal and bacterial community of digested sludge by Acidithiobacillus ferrooxidans.

Acidithiobacillus ferrooxidans ILS-2 was adapted in digested sludge and used to treat sludge for dewaterability improvement. Results showed that increasing ferrous iron loading increased sludge dewaterability, but the inoculation of the bioleaching strain had little effect on sludge dewaterability compared to controls without the strain. The total extracellular polymeric substances (EPS) contents of sludges with and without bioleaching treatment were similar except for bioleaching treatment at 10% ferrous iron loading (on sludge total solids) where total EPS was higher with bioleaching treatment. However, bioleaching treatment for 48 h had a notable effect on removal of heavy metals, such as Mn, Ni and Zn, especially at the high loadings of ferrous iron. In the presence of A. ferrooxidans, the removal of Ni, Mn and Zn reached 93%, 88% and 80%, respectively, at a ferrous iron loading of 21%. The sequencing of 16S rRNA genes indicated that increasing ferrous iron loadings to 15% and 21% increased the relative abundance of Acidithiobacillus, Acidocella (with A. ferrooxidans) and Carboxylicivirga (without A. ferrooxidans) but decreased the abundance of Pseudomonas and Acinetobacter after 48 h treatment. This study enhanced the understanding of the correlations between bioleaching treatment of digested sludge, sludge dewaterability, heavy metal removal and bacterial communities.

Microbial oil production from acidified glycerol pretreated sugarcane bagasse by Mortierella isabellina.

An integrated microbial oil production process consisting of acidified glycerol pretreatment of sugarcane bagasse, enzymatic hydrolysis, microbial oil production by Mortierella isabellina NRRL 1757 and oil recovery by hydrothermal liquefaction (HTL) of fungal biomass in fermentation broth was assessed in this study. Following pretreatment, the effect of residual pretreatment hydrolysate (containing glycerol) on enzymatic hydrolysis was firstly studied. The residual pretreatment hydrolysate (corresponding to 2.0-7.5% glycerol) improved glucan enzymatic digestibilities by 10-11% compared to the enzymatic hydrolysis in water (no buffer). Although residual pretreatment hydrolysate at 2.0-5.0% glycerol slightly inhibited the consumption of glucose in enzymatic hydrolysate by M. isabellina NRRL 1757, it did not affect microbial oil production due to the consumption of similar amounts of total carbon sources including glycerol. When the cultivation was scaled-up to a 1 L bioreactor, glucose was consumed more rapidly but glycerol assimilation was inhibited. Finally, HTL of fungal biomass in fermentation broth without any catalyst at 340 °C for 60 min efficiently recovered microbial oils from fungal biomass and achieved a bio-oil yield of 78.7% with fatty acids being the dominant oil components (∼89%). HTL also led to the hydrogenation of less saturated fatty acids (C18:2 and C18:3) to more saturated forms (C18:0 and C18:1).

Co-utilization of acidified glycerol pretreated-sugarcane bagasse for microbial oil production by a novel Rhodosporidium strain.

Acidified glycerol pretreatment is very effective to deconstruct lignocellulosics for producing glucose. Co-utilization of pretreated biomass and residual glycerol to bioproducts could reduce the costs associated with biomass wash and solvent recovery. In this study, a novel strain Rhodosporidium toruloides RP 15, isolated from sugarcane bagasse, was selected and tested for coconversion of pretreated biomass and residual glycerol to microbial oils. In the screening trails, Rh. toruloides RP 15 demonstrated the highest oil production capacity on glucose, xylose, and glycerol among the 10 strains. At the optimal C:N molar ratio of 140:1, this strain accumulated 56.7, 38.3, and 54.7% microbial oils based on dry cell biomass with 30 g/L glucose, xylose, and glycerol, respectively. Furthermore, sugarcane bagasse medium containing 32.6 g/L glucose from glycerol-pretreated bagasse and 23.4 g/L glycerol from pretreatment hydrolysate were used to produce microbial oils by Rh. toruloides RP 15. Under the preliminary conditions without pH control, this strain produced 7.7 g/L oil with an oil content of 59.8%, which was comparable or better than those achieved with a synthetic medium. In addition, this strain also produced 3.5 mg/L carotenoid as a by-product. It is expected that microbial oil production can be significantly improved through process optimization.

A genetic and metabolic approach to redirection of biochemical pathways of Clostridium butyricum for enhancing hydrogen production.

Clostridium butyricum, a well known H(2) producing bacterium, produces lactate, butyrate, acetate, ethanol, and CO(2) as its main by-products from glucose. The conversion of pyruvate to lactate, butyrate and ethanol involves oxidation of NADH. It was hypothesized that the NADH could be increased if the formation of these by-products could be eliminated, resulting in enhancing H(2) yield. Herein, this study aimed to establish a genetic and metabolic approach for enhancing H(2) yield via redirection of metabolic pathways of a C. butyricum strain. The ethanol formation pathway was blocked by disruption of aad (encoding aldehyde-alcohol dehydrogenase) using a ClosTron plasmid. Although elimination of ethanol formation alone did not increase hydrogen production, the resulting aad-deficient mutant showed approximately 20% enhanced performance in hydrogen production with the addition of sodium acetate. This work demonstrated the possibility of improving hydrogen yield by eliminating the unfavorable by-products ethanol and lactate.

Genetic manipulation of butyrate formation pathways in Clostridium butyricum.

Clostridium butyricum is one of the commonly used species for fermentative hydrogen production. While producing H2, it can produce acids (lactic, acetic and butyric acids) and CO2, as well as a small amount of ethanol. It has been proposed that elimination of competing pathways, such as the butyrate formation pathway, should increase H2 yields in Clostridium species. However, the application of this strategy has been hindered by the unavailability of genetic tools for these organisms. In this study, we successfully transferred a plasmid (pMTL007) to C. butyricum by inter-specific conjugation with Escherichia coli and disrupted hbd, the gene encoding ß-hydroxybutyryl-CoA dehydrogenase in C. butyricum. Fermentation data showed that inactivation of hbd in C. butyricum eliminated the butyrate formation pathway, resulting in a significant increase in ethanol production and an obvious decrease in H2 yield compared with the wild type strain. However, under low partial pressure of H2, the hbd-deficient strain showed increased H2 production with the simultaneous decrease of ethanol production, indicating that H2 production by C. butyricum may compete for NADH with the ethanol formation pathway. Together with the discovery of a potential bifurcating hydrogenase, this study extends our understanding of the mechanism of H2 production by C. butyricum.

Metabolic flux network and analysis of fermentative hydrogen production.

Fermentative hydrogen production (FHP) has received a great R & D interest in recent decades, as it offers a potential means of producing H2 from a variety of renewable resources, even wastewater via a low energy continuous process. Various extracellular metabolites including ethanol, acetate, butyrate and lactate can be produced during the fermentation, building a complex metabolic network of the FHP. Except for the recognition of its complexity, the metabolic flux network has not been well understood. Studies on biochemical reactions and metabolic flux network associated with the FHP in anaerobic fermentation system have only been drawn attention in recent years. This review summarizes the biochemical reactions taking place in the metabolic network of FHP. We discuss how the key operation factors influence metabolism in the FHP process. Recently developed and applied technologies for metabolic flux analysis have been described. Future studies on the metabolic network to enhance fermentative hydrogen production by strict anaerobes are recommended. It is expected that this review can provide useful information in terms of fundamental knowledge and update technology for scientists and research engineers in the field of biological hydrogen production.

A bacteriophage-related chimeric marine virus infecting abalone.

Marine viruses shape microbial communities with the most genetic diversity in the sea by multiple genetic exchanges and infect multiple marine organisms. Here we provide proof from experimental infection that abalone shriveling syndrome-associated virus (AbSV) can cause abalone shriveling syndrome. This malady produces histological necrosis and abnormally modified macromolecules (hemocyanin and ferritin). The AbSV genome is a 34.952-kilobase circular double-stranded DNA, containing putative genes with similarity to bacteriophages, eukaryotic viruses, bacteria and endosymbionts. Of the 28 predicted open reading frames (ORFs), eight ORF-encoded proteins have identifiable functional homologues. The 4 ORF products correspond to a predicted terminase large subunit and an endonuclease in bacteriophage, and both an integrase and an exonuclease from bacteria. The other four proteins are homologous to an endosymbiont-derived helicase, primase, single-stranded binding (SSB) protein, and thymidylate kinase, individually. Additionally, AbSV exhibits a common gene arrangement similar to the majority of bacteriophages. Unique to AbSV, the viral genome also contains genes associated with bacterial outer membrane proteins and may lack the structural protein-encoding ORFs. Genomic characterization of AbSV indicates that it may represent a transitional form of microbial evolution from viruses to bacteria.