Understanding the Role of Prevotella Genus in the Digestion of Lignocellulose and Other Substrates in Vietnamese Native Goats’ Rumen by Metagenomic Deep Sequencing

Bacteria in rumen play pivotal roles in the digestion of nutrients to support energy for the host. In this study, metagenomic deep sequencing of bacterial metagenome extracted from the goats’ rumen generated 48.66 GB of data with 3,411,867 contigs and 5,367,270 genes. The genes were mainly functionally annotated by Kyoto Encyclopedia of Genes and Genomes (KEGG) Carbohydrate-Active enZYmes (CAZy), and HMMER database, and taxonomically classified by MEGAN. As a result, 65,554 genes encoding for 30 enzymes/proteins related to lignocellulose conversion were exploited, in which nine enzymes were seen for the first time in goat rumen. Prevotella was the most abundant genus, contributing 30% hemicellulases and 36% enzymes/proteins for lignocellulose pretreatment, and supporting 98.8% of feruloyl esterases and 71.7% acetylxylan esterases. In addition, 18 of the 22 most lignocellulose digesting- potential contigs belonged to Prevotella. Besides, Prevotella possessed many genes coding for amylolytic enzymes. One gene encoding for endoxylanase was successfully expressed in E. coli. The recombinant enzyme had high Vmax, was tolerant to some salts and detergents, worked better at pH 5.5–6.5, temperature 40–50 °C, and was capable to be used in practices. Based on these findings, we confirm that Prevotella plays a pivotal role for hemicellulose digestion and significantly participates in starch, cellulose, hemicellulose, and pectin digestion in the goat rumen.

Isolation and Screening of Microorganisms for the Effective Pretreatment of Lignocellulosic Agricultural Wastes

Lignocellulosic waste is the most abundant biorenewable biomass on earth, and its hydrolysis releases highly valued reducing sugars. However, the presence of lignin in the biopolymeric structure makes it highly resistant to solubilization thereby hindering the hydrolysis of cellulose and hemicellulose. Microorganisms are known for their potential complex enzymes that play a dominant role in lignocellulose conversion. Therefore, the current study was designed to isolate and screen potential microorganisms for their selective delignification ability for the pretreatment of lignocellulosic biomass. An extensive isolation and screening procedure yielded 36 desired isolates (22 bacteria, 7 basidiomycete fungi, and 7 filamentous fungi). Submerged cultivation of these desired microorganisms revealed 4 bacteria and 10 fungi with potent lignocellulolytic enzyme activities. The potent isolates were identified as Pleurotus, Trichoderma, Talaromyces, Bacillus, and Chryseobacterium spp. confirmed by morphological and molecular identification. The efficiency of these strains was determined through enzyme activities, and the degraded substrates were analyzed through scanning electron microscopy (SEM) and X-ray diffraction (XRD). Among all isolated microbes, Pleurotus spp. were found to have high laccase activity. The cellulose-decomposing and selective delignification strains were subjected to solid-state fermentation (SSF). SSF of field waste corn stalks as a single-carbon source provides Pleurotus spp. better condition for the secretion of ligninolytic enzymes. These isolated ligninolytic enzymes producing microorganisms may be used for the effective pretreatment of lignocellulosic agricultural wastes for the production of high value-added natural products by fermentation.

Conversion of Lignocellulose for Bioethanol Production, Applied in Bio–Polyethylene Terephthalate

The increasing demand for petroleum-based polyethylene terephthalate (PET) grows population impacts daily. A greener and more sustainable raw material, lignocellulose, is a promising replacement of petroleum-based raw materials to convert into bio-PET. This paper reviews the recent development of lignocellulose conversion into bio-PET through bioethanol reaction pathways. This review addresses lignocellulose properties, bioethanol production processes, separation processes of bioethanol, and the production of bio–terephthalic acid and bio–polyethylene terephthalate. The article also discusses the current industries that manufacture alcohol-based raw materials for bio-PET or bio-PET products. In the future, the production of bio-PET from biomass will increase due to the scarcity of petroleum-based raw materials.

Effect of the method for the elimination of inhibitors present in Miscanthus giganteus hydrolysates on ethanol production effectiveness

The pretreatment of lignocellulosic material performed to improve substrate’s susceptibility to enzymatic hydrolysis is usually accompanied by reactions leading to the synthesis of compounds that inhibit the metabolic activity of microorganisms. Their toxicity is the main obstacle to the successful bioconversion of lignocellulosic hydrolysates. The identification of these inhibitors and the choice of the optimal detoxication method are crucial for the improving the efficiency of fermentation processes. Material rinsing with water after processing is a common detoxication practice. However, it generates material losses, thus affecting contents of saccharides in the fermentation medium, which may in turn trigger higher costs of lignocellulose conversion to ethanol and other products with a higher added value. A study was undertaken to determine the effect of selected methods for the detoxication of an enzymatic hydrolysate from Miscanthus giganteus on the fermentation efficiency of saccharide derivatives. The experiment conducted with Mucor rouxii DSM 1191 demonstrated the usability of the detoxication method based on the activated carbon. After 96-h fermentation of Miscanthus hydrolysates, the alcohol content in the post-reaction medium was higher by 14% than in the control experiment wherein the material was rinsed with water after pretreatment. The experiment carried out with Saccharomyces cerevisiae 7, NRRL 978 showed no positive impact of the alternative detoxication methods replacing material rinsing on the efficiency of ethanol synthesis. The highest concentration of this metabolite (2.04% (v/v)) was obtained in the experimental variant in which the mentioned operation was coupled with detoxication of hydrolysates using calcium hydroxide.

Enhanced Biomass Yield of and Saccharification in Transgenic Tobacco Over-Expressing β-Glucosidase

Here, we report an increase in biomass yield and saccharification in transgenic tobacco plants (Nicotiana tabacum L.) overexpressing thermostable β-glucosidase from Thermotoga maritima, BglB, targeted to the chloroplasts and vacuoles. The transgenic tobacco plants showed phenotypic characteristics that were significantly different from those of the wild-type plants. The biomass yield and life cycle (from germination to flowering and harvest) of the transgenic tobacco plants overexpressing BglB were 52% higher and 36% shorter than those of the wild-type tobacco plants, respectively, indicating a change in the genome transcription levels in the transgenic tobacco plants. Saccharification in biomass samples from the transgenic tobacco plants was 92% higher than that in biomass samples from the wild-type tobacco plants. The transgenic tobacco plants required a total investment (US$/year) corresponding to 52.9% of that required for the wild-type tobacco plants, but the total biomass yield (kg/year) of the transgenic tobacco plants was 43% higher than that of the wild-type tobacco plants. This approach could be applied to other plants to increase biomass yields and overproduce β-glucosidase for lignocellulose conversion.

Complete Genome Sequence of Rhodococcus ruber R1, a Novel Strain Showing a Broad Catabolic Potential toward Lignin-Derived Aromatics

Rhodococcus ruber R1 was isolated from a pulp mill wastewater treatment plant because of its ability to use methoxylated aromatics as growth substrates. We report the 5.56-Mb genome sequence of strain R1, which can provide insights into the biodegradation of lignin-derived phenolic monomers and potentially support processes for lignocellulose conversion.

Coupling structural characterization with secretomic analysis reveals mechanism of the disruption of cross-linked structure in bamboo culms by Echinodontium taxodii

Background: Overcoming the biomass recalcitrance is essential for efficient utilization of lignocellulosic biomass in industrial bio-refining. White-rot fungi can overcome the biomass recalcitrance and accelerate the conversion of lignocellulose to biofuels via a large number of special extracellular lignocellulolytic enzymes. Previous studies try to dissect the function of extracellular enzymes on biomass resistant cross-linked structures by secretome analysis, but the bio-alteration of cross-linked structures is ignored usually. A deeper and detailed understanding of relationship between secretome and bio-alteration of cross-linked structure in lignocellulosic biomass is still lack. Results: As an efficient wood-decaying fungus, Echinodontium taxodii could improve the conversion efficiency of lignocellulose to biofuels. This study coupled comparative analysis of fungal secretomes and 2D HSQC NMR analysis of lignocellulose fractions, aiming to elucidate the role of extracellular enzymes from Echinodontium taxodii 2538 in the disruption of resistant cross-linked structure of bamboo culms. Carboxylesterases, alcohol oxidases and Class-II peroxidases showed importance in the cleavage of cross-linked structures, including ester and ether linkages of lignin-carbohydrate complexes (LCCs) and inter-unit linkages of lignin, which contributed to biomass resistance removal and cellulose exposure during the early stage of fungal decay. Moreover, the rapid oxidation of Cα-OH was found to contribute to the lignin bio-depolymerization. Conclusions: These findings revealed the detailed mechanisms of biomass recalcitrance reduction by fungal pretreatment, and provide insight into efficient strategy of lignocellulose conversion. It will advance the development in design of enzyme cocktail for efficient lignocellulose bio-refinery.

Hydrolysis Kinetics of Oil Palm Empty Fruit Bunch in Ionic Liquids and Cellulase Integrated System

Ionic liquids (ILs) are developing as potential solvents in lignocellulose solvation, which enables cellulase accessibility into the substrate. Nevertheless, ILs could result in enzyme deactivation because of the high polarity. Therefore, developing a system of ILs-compatible cellulase (IL-E) to promote lignocellulose conversion into sugars is a challenge in ILs applications. This study used an IL-E to attain high conversion yield of sugars from oil palm empty fruit bunch (EFB). Cellulase (Tr-Cel) from Trichoderma reesei was stable in the ILs, 1-ethyl-3-methyl imidazolium diethyl phosphate [EMIM]DEP and choline acetate [Cho]OAc. The inhibition and deactivation of cellulase were evaluated using the model substrate, carboxymethyl cellulose (CMC) and EFB as a lignocellulosic material to assess the hydrolytic activity. The enzyme kinetics revealed that [Cho]OAc acted as a noncompetitive inhibitor. Additionally, [EMIM]DEP may not be considered as an inhibitor as it increases the Vmax and does not significantly affect the KM. In both cases, the study proved that IL did not result in a severe loss of cellulase activity, which is a promising outcome for one-pot hydrolysis of lignocellulosic materials.