Fungal communities as dual indicators of river biodiversity and water quality assessment.

Given their ecological importance, bioindicators are used for the assessment of the health of river ecosystems. This study explored the fungal compositions and the potential of fungal taxa as bioindicators for indicating the water quality of the Mekong River, as the use of fungal indicators of the Mekong River was not previously well characterized. The Mekong River exhibited dynamic variations in both physicochemical/hydrochemical properties and fungal communities according to seasons and locations. The results revealed the dominance of alkaline earth metal ions and weak acids in the water. The magnesium-bicarbonate water type was found in the dry season, but the water became the chloride-calcium type or mixed type of magnesium-bicarbonate and chloride-calcium in the rainy season at downstream sites. Fungal composition analysis revealed the dominance of Chytridiomycota in the dry season and intermediate periods, and Ascomycota and Basidiomycota in the rainy season. The fungal communities were influenced by stochastic and deterministic assembly processes, mainly homogeneous selection, heterogeneous selection, and dispersal limitation. The extent of environmental filtering implied that some fungal taxa were affected by environmental conditions, suggesting the possibility of identifying certain fungal taxa suitable for being bioindicators of water quality. Subsequently, machine learning with recursive feature elimination identified specific fungal bins mostly consisting of Agaricomycetes (mainly Polyporales, Agaricales, and Auriculariales), Dothideomycetes (mainly Pleosporales), Saccharomycetes (mainly Saccharomycetales), Chytridiomycota, and Rozellomycota as bioindicators that could predict ambient and irrigation water quality with high selectivity and sensitivity. These results thus promote the use of fungal indicators to assess the health of the river.

Interplay of xenobiotic-degrading and antibiotic-resistant microorganisms among the microbiome found in the air, handrail, and floor of the subway station.

Investigating the quality of the subway environment, especially regarding antibiotic resistance genes (ARGs) and xenobiotics, conveys ecological and health impacts. In this study, compositions and relations of microorganisms harboring ARGs and xenobiotic degradation and metabolism genes (XDGs) in the Sukhumvit subway station (MRT-SKV) in Bangkok was assessed by analyzing the taxonomic and genetic diversity of the microbiome in the air and on the surfaces of floor and handrail. The major bacteria in the MRT-SKV (including Moraxella, which was abundant in the bioaerosol and handrail samples, and Staphylococcus, which was abundant in the bioaerosol samples) were found to contain both ARGs and XDGs. The co-abundance correlation network revealed notable relationships among bacteria harboring antibiotic resistance genes (ARGs) and xenobiotic degradation genes (XDGs). Significant associations were observed between ARGs linked to glycopeptide and fluoroquinolone resistance and genes associated with benzoate, styrene, and atrazine degradation pathways, as well as between ARGs related to cephamycin, cephalosporin, and MLS resistance and XDGs associated with the cytochrome P450-dependent drug metabolism pathway. These correlations suggested that selective pressure exerted by certain xenobiotics and antibiotics can simultaneously affect both ARGs and XDGs in the environment and should favor correlations and co-survival among ARG- and XDG-containing bacteria in the environments. The correlations may occur via shared mechanisms of resistance to both xenobiotics and antibiotics. Finally, different correlation pairs were seen in different niches (air, handrail, floor) of the subway environment or different geolocations. Thus, the relationship between ARG and XDG pairs most likely depends on the unique characteristics of the niches and on the prominent types of xenobiotics and antibiotics in the subway environment. The results indicated that interactions and connections between microbial communities can impact how they function. These microorganisms can have profound effects on accumulation of xenobiotics and ARGs in the MRT-SKV.

Elucidating potential bioindicators from insights in the diversity and assembly processes of prokaryotic and eukaryotic communities in the Mekong River.

Drivers for spatio-temporal distribution patterns of overall planktonic prokaryotes and eukaryotes in riverine ecosystems are generally not fully understood. This study employed amplicon metabarcoding to investigate the distributions and assembly mechanisms of bacterial and eukaryotic communities in the Mekong River. The prevailing bacteria taxa were found to be Betaproteobacteria, Actinobacteria, and Bacteroidetes, while the dominant eukaryotic organisms were cryptophytes, chlorophytes, and diatoms. The community assemblages were influenced by a combination of stochastic and deterministic processes. Drift (DR) and dispersal limitation (DL), signifying the stochastic mechanism, were the main processes shaping the overall prokaryotic and eukaryotic communities. However, homogeneous selection (HoS), indicating deterministic mechanism, played a major role in the assembly process of core prokaryotic communities, especially in the wet season. In contrast, the core eukaryotic communities including Opisthokonta, Sar, and Chlorophyta were dominated by stochastic processes. The significance of HoS within prokaryotic communities was also found to exhibit a decreasing trend from the upstream sampling sites (Chiang Saen and Chiang Khan, Nong Khai) towards the downstream sites (Mukdahan, and Khong Chiam) of the Mekong River. The environmental gradients resulting from the site-specific variations and the gradual decrease in elevation along the river may have a potential influence on the role of HoS in community assembly. Crucial environmental factors that shape the phylogenetic structure within distinct bins of the core prokaryotic communities including water depth, temperature, chloride, sodium, and sulphate were identified, as inferred by their correlation with the beta Net Relatedness Index (betaNRI) during the wet season. Overall, these findings enhance understanding of the complex mechanisms governing the spatio-temporal dynamics of prokaryotic and eukaryotic communities in the Mekong River. Finally, insights gained from this study could provide information on further use of specific core bacteria as microbial-based bioindicators that are effective for the assessment and conservation of the Mekong River ecosystem.

AirDNA sampler: An efficient and simple device enabling high-yield, high-quality airborne environment DNA for metagenomic applications.

Analyzing temporal and spatial distributions of airborne particles of biological origins is vital for the assessment and monitoring of air quality, especially with regard to public health, environmental ecology, and atmospheric chemistry. However, the analysis is frequently impeded by the low levels of biomass in the air, especially with metagenomic DNA analysis to explore diversity and composition of living organisms and their components in the air. To obtain sufficient amounts of metagenomic DNA from bioaerosols, researchers usually need a long sampling time with an expensive high-volume air sampler. This work shows the utilization of an air sampling device containing an economical, high-volume portable ventilation fan in combination with customized multi-sheet filter holders to effectively obtain high yields of genomic DNA in a relatively short time. The device, named 'AirDNA' sampler, performed better than other commercial air samplers, including MD8 Airport and Coriolis compact air samplers. Using the AirDNA sampler, an average DNA yield of 40.49 ng (12.47-23.24 ng at 95% CI) was obtained in only 1 hour of air sampling with a 0.85 probability of obtaining ≥10 ng of genomic DNA. The genomic DNA obtained by the AirDNA system is of suitable quantity and quality to be further used for amplicon metabarcoding sequencing of 16S, 18S, and cytochrome c oxidase I (COI) regions, indicating that it can be used to detect various prokaryotes and eukaryotes. Our results showed the effectiveness of our AirDNA sampling apparatus with a simple setup and affordable devices to obtain metagenomic DNA for short-term or long-term spatiotemporal analysis. The technique is well suited for monitoring air in built environments, especially monitoring bioaerosols for health purposes and for fine-scale spatiotemporal environmental studies.

Beneficial bacterial-Auricularia cornea interactions fostering growth enhancement identified from microbiota present in spent mushroom substrate.

Complex dynamic bacterial-fungal interactions play key roles during mushroom growth, ranging from mutualism to antagonism. These interactions convey a large influence on mushroom's mycelial and fruiting body formation during mushroom cultivation. In this study, high-throughput amplicon sequencing was conducted to investigate the structure of bacterial communities in spent mushroom substrates obtained from cultivation of two different groups of Auricularia cornea with (A) high yield and (B) low yield of fruiting body production. It was found that species richness and diversity of microbiota in group (A) samples were significantly higher than in group (B) samples. Among the identified 765 bacterial OTUs, 5 bacterial species found to exhibit high differential abundance between group (A) and group (B) were Pseudonocardia mangrovi, Luteimonas composti, Paracoccus pantotrophus, Sphingobium jiangsuense, and Microvirga massiliensis. The co-cultivation with selected bacterial strains showed that A. cornea TBRC 12900 co-cultivated with P. mangrovi TBRC-BCC 42794 promoted a high level of mycelial growth. Proteomics analysis was performed to elucidate the biological activities involved in the mutualistic association between A. cornea TBRC 12900 and P. mangrovi TBRC-BCC 42794. After co-cultivation of A. cornea TBRC 12900 and P. mangrovi TBRC-BCC 42794, 1,616 proteins were detected including 578 proteins of A. cornea origin and 1,038 proteins of P. mangrovi origin. Functional analysis and PPI network construction revealed that the high level of mycelial growth in the co-culture condition most likely resulted from concerted actions of (a) carbohydrate-active enzymes including hydrolases, glycosyltransferases, and carbohydrate esterases important for carbohydrate metabolism and cell wall generation/remodeling, (b) peptidases including cysteine-, metallo-, and serine-peptidases, (c) transporters including the ABC-type transporter superfamily, the FAT transporter family, and the VGP family, and (d) proteins with proposed roles in formation of metabolites that can act as growth-promoting molecules or those normally contain antimicrobial activity (e.g., indoles, terpenes, ß-lactones, lanthipeptides, iturins, and ectoines). The findings will provide novel insights into bacterial-fungal interactions during mycelial growth and fruiting body formation. Our results can be utilized for the selection of growth-promoting bacteria to improve the cultivation process of A. cornea with a high production yield, thus conveying potentially high socio-economic impact to mushroom agriculture.

Efficient multiplex CRISPR/Cpf1 (Cas12a) genome editing system in Aspergillus aculeatus TBRC 277.

CRISPR/Cas technology is a versatile tool for genome engineering in many organisms, including filamentous fungi. Cpf1 is a multi-domain protein of class 2 (type V) RNA-guided CRISPR/Cas endonuclease, and is an alternative platform with distinct features when compared to Cas9. However, application of this technology in filamentous fungi is limited. Here, we present a single CRISPR/Cpf1 plasmid system in Aspergillus aculeatus strain TBRC 277, an industrially relevant cell factory. We first evaluated the functionality of three Cpf1 orthologs from Acidaminococcus sp. BV3L6 (AsCpf1), Francisella tularensis subsp. novicida U112 (FnCpf1), and Lachnospiraceae bacterium (LbCpf1), in RNA-guided site-specific DNA cleavage at the pksP locus. FnCpf1 showed the highest editing efficiency (93 %) among the three Cpf1s. It was further investigated for its ability to delete a 1.7 kb and a 0.5 kb from pksP and pyrG genes, respectively, using two protospacers targeting these gene loci in a single crRNA array. Lastly, simultaneous editing of three sites within TBRC 277 genome was performed using three guide sequences targeting these two genes as well as an additional gene, kusA, which resulted in combined editing efficiency of 40 %. The editing of the NHEJ pathway by targeting kusA to generate a NHEJ-deficient strain of A. aculeatus TBRC 277 improved gene targeting efficiency and yielded more precise gene-editing than that of using wild-type strain. This promising genome-editing system can be used for strain improvement in industrial applications such as production of valuable bioproducts.

Development of a CRISPR/Cpf1 system for targeted gene disruption in Aspergillus aculeatus TBRC 277.

BACKGROUND: CRISPR-Cas genome editing technologies have revolutionized biotechnological research particularly in functional genomics and synthetic biology. As an alternative to the most studied and well-developed CRISPR/Cas9, a new class 2 (type V) CRISPR-Cas system called Cpf1 has emerged as another versatile platform for precision genome modification in a wide range of organisms including filamentous fungi. RESULTS: In this study, we developed AMA1-based single CRISPR/Cpf1 expression vector that targets pyrG gene in Aspergillus aculeatus TBRC 277, a wild type filamentous fungus and potential enzyme-producing cell factory. The results showed that the Cpf1 codon optimized from Francisella tularensis subsp. novicida U112, FnCpf1, works efficiently to facilitate RNA-guided site-specific DNA cleavage. Specifically, we set up three different guide crRNAs targeting pyrG gene and demonstrated that FnCpf1 was able to induce site-specific double-strand breaks (DSBs) followed by an endogenous non-homologous end-joining (NHEJ) DNA repair pathway which caused insertions or deletions (indels) at these site-specific loci. CONCLUSIONS: The use of FnCpf1 as an alternative class II (type V) nuclease was reported for the first time in A. aculeatus TBRC 277 species. The CRISPR/Cpf1 system developed in this study highlights the feasibility of CRISPR/Cpf1 technology and could be envisioned to further increase the utility of the CRISPR/Cpf1 in facilitating strain improvements as well as functional genomics of filamentous fungi.

Uncovering multi-faceted taxonomic and functional diversity of soil bacteriomes in tropical Southeast Asian countries.

Environmental microbiomes encompass massive biodiversity and genetic information with a wide-ranging potential for industrial and agricultural applications. Knowledge of the relationship between microbiomes and environmental factors is crucial for translating that information into practical uses. In this study, the integrated data of Southeast Asian soil bacteriomes were used as models to assess the variation in taxonomic and functional diversity of bacterial communities. Our results demonstrated that there were differences in soil bacteriomes across different geographic locality with different soil characteristics: soil class and pH level. Such differences were observed in taxonomic diversity, interspecific association patterns, and functional diversity of soil bacteriomes. The bacterial-mediated biogeochemical cycles of nitrogen, sulfur, carbon, and phosphorus illustrated the functional relationship of soil bacteriome and soil characteristics, as well as an influence from bacterial interspecific interaction. The insights from this study reveal the importance of microbiome data integration for future microbiome research.

gcType: a high-quality type strain genome database for microbial phylogenetic and functional research.

Taxonomic and functional research of microorganisms has increasingly relied upon genome-based data and methods. As the depository of the Global Catalogue of Microorganisms (GCM) 10K prokaryotic type strain sequencing project, Global Catalogue of Type Strain (gcType) has published 1049 type strain genomes sequenced by the GCM 10K project which are preserved in global culture collections with a valid published status. Additionally, the information provided through gcType includes >12 000 publicly available type strain genome sequences from GenBank incorporated using quality control criteria and standard data annotation pipelines to form a high-quality reference database. This database integrates type strain sequences with their phenotypic information to facilitate phenotypic and genotypic analyses. Multiple formats of cross-genome searches and interactive interfaces have allowed extensive exploration of the database's resources. In this study, we describe web-based data analysis pipelines for genomic analyses and genome-based taxonomy, which could serve as a one-stop platform for the identification of prokaryotic species. The number of type strain genomes that are published will continue to increase as the GCM 10K project increases its collaboration with culture collections worldwide. Data of this project is shared with the International Nucleotide Sequence Database Collaboration. Access to gcType is free at http://gctype.wdcm.org/.

Mating-type switching and mating-type gene array expression in the methylotrophic yeast Ogataea thermomethanolica TBRC656.

The methylotrophic yeast, Ogataea thermomethanolica TBRC656, is an attractive host organism for heterologous protein production owing to the availability of protein expression vectors and a genome-editing tool. In this study, we focused on mating-type switching and gene expression in order to elucidate its sexual life cycle and establish genetic approaches applicable for the strain. A putative mating-type gene cluster was identified in TBRC656 that is syntenic to the cluster in Ogataea parapolymorpha DL-1 (previously named Hansenula polymorpha). Like DL-1, TBRC656 possesses two mating loci, namely MATa and MATα, and also shows flip-flop mating-type switching. Interestingly, unlike any other methylotrophic yeast, TBRC656 robustly switched mating type during late growth in rich medium (YPD). Under nutrient depletion, mating-type switching was observed within one hour. Transcription from both MATa and MATα mating loci was detected during growth in YPD, and possibly induced upon nitrogen depletion. Gene expression from MATα was detected as a single co-transcript from a three-gene array (α2-α1-a1S). Deletion of a putative a1S ORF at the MATα locus had no observed effect on mating-type switching but demonstrated significant effect on mating-type gene expression at both MATa and MATα loci.

Comparative analysis of bacterial communities associated with healthy and diseased corals in the Indonesian sea.

Coral reef ecosystems are impacted by climate change and human activities, such as increasing coastal development, overfishing, sewage and other pollutant discharge, and consequent eutrophication, which triggers increasing incidents of diseases and deterioration of corals worldwide. In this study, bacterial communities associated with four species of corals: Acropora aspera, Acropora formosa, Cyphastrea sp., and Isopora sp. in the healthy and disease stages with different diseases were compared using tagged 16S rRNA sequencing. In total, 59 bacterial phyla, 190 orders, and 307 genera were assigned in coral metagenomes where Proteobacteria and Firmicutes were pre-dominated followed by Bacteroidetes together with Actinobacteria, Fusobacteria, and Lentisphaerae as minor taxa. Principal Coordinates Analysis (PCoA) showed separated clustering of bacterial diversity in healthy and infected groups for individual coral species. Fusibacter was found as the major bacterial genus across all corals. The lower number of Fusibacter was found in A. aspera infected with white band disease and Isopora sp. with white plaque disease, but marked increases of Vibrio and Acrobacter, respectively, were observed. This was in contrast to A. formosa infected by a black band and Cyphastrea sp. infected by yellow blotch diseases which showed an increasing abundance of Fusibacter but a decrease in WH1-8 bacteria. Overall, infection was shown to result in disturbance in the complexity and structure of the associated bacterial microbiomes which can be relevant to the pathogenicity of the microbes associated with infected corals.

Designing cellulolytic enzyme systems for biorefinery: From nature to application.

Cellulolytic enzymes play a key role on conversion of lignocellulosic plant biomass to biofuels and biochemicals in sugar platform biorefineries. In this review, we survey composite carbohydrate-active enzymes (CAZymes) among groups of cellulolytic fungi and bacteria that exist under aerobic and anaerobic conditions. Recent advances in designing effective cellulase mixtures are described, starting from the most complex microbial consortium-based enzyme preparations, to single-origin enzymes derived from intensively studied cellulase producers such as Trichoderma reesei, Talaromyces cellulolyticus, and Penicellium funiculosum, and the simplest minimal enzyme systems comprising selected sets of mono-component enzymes tailor-made for specific lignocellulosic substrates. We provide a comprehensive update on studies in developing high-performance cellulases for biorefineries.

A novel sucrose-based expression system for heterologous proteins expression in thermotolerant methylotrophic yeast Ogataea thermomethanolica.

The thermotolerant methylotrophic yeast Ogataea thermomethanolica TBRC656 is a potential host for heterologous protein expression. In this study, a novel expression system was developed for O. thermomethanolica based on the maltase (mal) gene promoter from this organism. The OtMal promoter function was tested for expression of fungal enzymes as reporter genes. Measurement of xylanase reporter enzyme activity showed that the OtMal promoter was repressed during growth on glucose and was activated by sucrose. When sucrose was used as a carbon source, the OtMal promoter was approximately twice as strong as the constitutive OtGAP promoter. Comparison of the OtMal promoter with the methanol-inducible OtAOX promoter showed that OtMal promoter drove 1.2 and 1.7-fold higher expression of xylanase and phytase reporter, respectively, than OtAOX promoter under inducing conditions at 24 h. Our results indicated that this novel expression system could be useful for the production of heterologous proteins from sucrose in yeast O. thermomethanolica.

The global catalogue of microorganisms 10K type strain sequencing project: closing the genomic gaps for the validly published prokaryotic and fungi species.

Genomic information is essential for taxonomic, phylogenetic, and functional studies to comprehensively decipher the characteristics of microorganisms, to explore microbiomes through metagenomics, and to answer fundamental questions of nature and human life. However, large gaps remain in the available genomic sequencing information published for bacterial and archaeal species, and the gaps are even larger for fungal type strains. The Global Catalogue of Microorganisms (GCM) leads an internationally coordinated effort to sequence type strains and close gaps in the genomic maps of microorganisms. Hence, the GCM aims to promote research by deep-mining genomic data.

CRISPR-Cas9 enabled targeted mutagenesis in the thermotolerant methylotrophic yeast Ogataea thermomethanolica.

Ogataea thermomethanolica TBRC656 is a thermotolerant methylotrophic yeast suitable for heterologous protein expression at various temperatures. However, the lack of efficient methods for targeted gene mutagenesis limits strain engineering in this yeast. In this study, we applied a CRISPR-Cas9-based tool for targeted gene mutagenesis in O. thermomethanolica. The putative unfolded protein response regulator OtHAC1, and the OtMAL1 (maltase) and OtMAL2 (maltose permease) genes involved with sucrose and maltose utilization were targeted for CRISPR-Cas9 mutagenesis. Plasmids were constructed for integrative and episomal expression of CRISPR-Cas9 elements in O. thermomethanolica in which Cas9 and gRNA are transcribed from the alcohol oxidase (AOX) promoter. The expression of these genome-editing elements is controlled by derepression with glycerol and gRNA are flanked by self-cleaving ribozymes. For integrative system, OtHAC1, OtMAL1 and OtMAL2 were disrupted at 63%, 97% and 93%, respectively. In addition, OtMAL1 was also disrupted with episomal system at 92%. These findings indicate that the CRISPR-Cas9 system described herein is thus applicable for studying gene function and strain engineering in yeast O. thermomethanolica.

Characterization of cellulolytic microbial consortium enriched on Napier grass using metagenomic approaches.

Energy grass is a promising substrate for production of biogas by anaerobic digestion. However, the conversion efficiency is limited by the enzymatically recalcitrant nature of cellulosic wastes. In this study, an active, structurally stable mesophilic lignocellulolytic degrading microbial consortium (Np-LMC) was constructed from forest compost soil microbiota by successive subcultivation on Napier grass under facultative anoxic conditions. According to tagged 16S rRNA gene amplicon sequencing, increasing abundance of facultative Proteobacteria was found in the middle of batch cycle which was then subsequently replaced by the cellulose degraders Firmicutes and Bacteroidetes along with decreasing CMCase, xylanase, and ß-glucanase activity profiles in the supernatant after 5 days of incubation. Anaerobic/facultative bacteria Dysgonomonas and Sedimentibacter and aerobic bacteria Comamonas were the major genera found in Np-LMC. The consortium was active on degradation of the native and delignified grass. Direct shotgun sequencing of the consortium metagenome revealed relatively high abundance of genes encoding for various lignocellulose degrading enzymes in 23 glycosyl hydrolase (GH) families compared to previously reported cellulolytic microbial communities in mammalian digestive tracts. Enzymes attacking cellulose and hemicellulose were dominated by GH2, 3, 5, 9, 10, 26, 28 and 43 in addition to a variety of carbohydrate esterases (CE) and auxiliary activities (AA), reflecting adaptation of the enzyme systems to the native herbaceous substrate. The consortium identified here represents the microcosm specifically bred on energy grass, with potential for enhancing degradation of fibrous substrates in bioenergy industry.

Simultaneous saccharification and viscosity reduction of cassava pulp using a multi-component starch- and cell-wall degrading enzyme for bioethanol production.

In this study, an efficient ethanol production process using simultaneous saccharification and viscosity reduction of raw cassava pulp with no prior high temperature pre-gelatinization/liquefaction step was developed using a crude starch- and cell wall-degrading enzyme preparation from Aspergillus aculeatus BCC17849. Proteomic analysis revealed that the enzyme comprised a complex mixture of endo- and exo-acting amylases, cellulases, xylanases, and pectina ses belonging to various glycosyl hydrolase families. Enzymatic hydrolysis efficiency was dependent on the initial solid loading in the reaction. Reduction in mixture viscosity was observed with a rapid decrease in complex viscosity from 3785 to 0.45 Pa s with the enzyme dosage of 2.19 mg/g on a dried weight basis within the first 2 h, which resulted from partial destruction of the plant cell wall fiber and degradation of the released starch granules by the enzymes as shown by scanning electron microscopy. Saccharification of cassava pulp at an initial solid of 16% (w/v) in a bench-scale bioreactor resulted in 736.4 mg glucose/g, which is equivalent to 82.92% glucose yield based on the total starch and glucan in the substrate, after 96 h at 40 °C. Simultaneous saccharification and fermentation of cassava pulp by Saccharomyces cerevisiae with the uncooked enzymatic process led to a final ethanol concentration of 6.98% w/v, equivalent to 96.7% theoretical yield based on the total starch and cellulose content. The results demonstrated potential of the enzyme for low-energy processing of cassava pulp in biofuel industry.

Characterization of cellulolytic enzyme system of Schizophyllum commune mutant and evaluation of its efficiency on biomass hydrolysis.

Schizophyllum commune is a basidiomycete equipped with an efficient cellulolytic enzyme system capable of growth on decaying woods. In this study, production of lignocellulose-degrading enzymes from S. commune mutant G-135 (SC-Cel) on various cellulosic substrates was examined. The highest cellulase activities including CMCase, FPase, and ß-glucosidase were obtained on Avicel-PH101 while a wider range of enzymes attacking non-cellulosic polysaccharides and lignin were found when grown on alkaline-pretreated biomass. Proteomic analysis of SC-Cel also revealed a complex enzyme system comprising seven glycosyl hydrolase families with an accessory carbohydrate esterase, polysaccharide lyase, and auxiliary redox enzymes. SC-Cel obtained on Avicel-PH101 effectively hydrolyzed all agricultural residues with the maximum glucan conversion of 98.0% using corn cobs with an enzyme dosage of 5 FPU/g-biomass. The work showed potential of SC-Cel on hydrolysis of various herbaceous biomass with enhanced efficiency by addition external ß-xylosidase.

Heterologous expression of Aspergillus aculeatus endo-polygalacturonase in Pichia pastoris by high cell density fermentation and its application in textile scouring.

BACKGROUND: Removal of non-cellulosic impurities from cotton fabric, known as scouring, by conventional alkaline treatment causes environmental problems and reduces physical strength of fabrics. In this study, an endo-polygalacturonase (EndoPG) from Aspergillus aculeatus produced in Pichia pastoris was evaluated for its efficiency as a bioscouring agent while most current bioscouring process has been performed using crude pectinase preparation. RESULTS: The recombinant EndoPG exhibited a specific activity of 1892.08 U/mg on citrus pectin under the optimal condition at 50 °C, pH 5.0 with a V max and K m of 65,451.35 µmol/min/mL and 15.14 mg/mL, respectively. A maximal activity of 2408.70 ± 26.50 U/mL in the culture supernatant was obtained by high cell density batch fermentation, equivalent to a 4.8 times greater yield than that from shake-flask culture. The recombinant enzyme was shown to be suitable for application as a bioscouring agent, in which the wettability of cotton fabric was increased by treatment with enzyme at 300 U/mL scouring solution at 40 °C, pH 5.0 for 1 h. The bio-scoured fabric has comparable wettability to that obtained by conventional chemical scouring, but has higher tensile strength. CONCLUSION: The work has demonstrated for the first time functions of A. aculeatus EndoPG on bioscouring in eco-textile processing. EndoPG alone was shown to possess effective scouring activity. High expression level and homogeneity could be achieved in bench-scale bioreactor.