Gut-microbiota-targeted diets modulate human immune status.

Diet modulates the gut microbiome, which in turn can impact the immune system. Here, we determined how two microbiota-targeted dietary interventions, plant-based fiber and fermented foods, influence the human microbiome and immune system in healthy adults. Using a 17-week randomized, prospective study (n = 18/arm) combined with -omics measurements of microbiome and host, including extensive immune profiling, we found diet-specific effects. The high-fiber diet increased microbiome-encoded glycan-degrading carbohydrate active enzymes (CAZymes) despite stable microbial community diversity. Although cytokine response score (primary outcome) was unchanged, three distinct immunological trajectories in high-fiber consumers corresponded to baseline microbiota diversity. Alternatively, the high-fermented-food diet steadily increased microbiota diversity and decreased inflammatory markers. The data highlight how coupling dietary interventions to deep and longitudinal immune and microbiome profiling can provide individualized and population-wide insight. Fermented foods may be valuable in countering the decreased microbiome diversity and increased inflammation pervasive in industrialized society.

De novo genome assembly and functional insights of the first commercial pink Auricularia cornea.

A pioneering pink cultivar of Auricularia cornea, first commercially cultivated in 2022, lacks genomic data, hindering research in genetic breeding, gene discovery, and product development. Here, we report the de novo assembly of the pink A. cornea Fen-A1 genome and provide a detailed functional annotation. The genome is 73.17 Mb in size, contains 86 scaffolds (N50 âˆ¼ 5.49 Mb), 59.09% GC content and encodes 19,120 predicted genes with a BUSCO completeness of 92.60%. Comparative genomic analysis reveals the phylogenetic relatedness of Fen-A1 and remarkable gene family dynamics. Putative genes were found mapped to 3 antibiotic-related, 36 light-dependent and 25 terpene metabolites. In addition, 789 CAZymes genes were classified, revealing the dynamics of quality loss due to postharvest refrigeration. Overall, our work is the first report on a pink A. cornea genome and provides a comprehensive insight into its complex functions.

Investigating diversity and similarity between CBM13 modules and ricin-B lectin domains using sequence similarity networks.

BACKGROUND: The CBM13 family comprises carbohydrate-binding modules that occur mainly in enzymes and in several ricin-B lectins. The ricin-B lectin domain resembles the CBM13 module to a large extent. Historically, ricin-B lectins and CBM13 proteins were considered completely distinct, despite their structural and functional similarities. RESULTS: In this data mining study, we investigate structural and functional similarities of these intertwined protein groups. Because of the high structural and functional similarities, and differences in nomenclature usage in several databases, confusion can arise. First, we demonstrate how public protein databases use different nomenclature systems to describe CBM13 modules and putative ricin-B lectin domains. We suggest the introduction of a novel CBM13 domain identifier, as well as the extension of CAZy cross-references in UniProt to guard the distinction between CAZy and non-CAZy entries in public databases. Since similar problems may occur with other lectin families and CBM families, we suggest the introduction of novel CBM InterPro domain identifiers to all existing CBM families. Second, we investigated phylogenetic, nomenclatural and structural similarities between putative ricin-B lectin domains and CBM13 modules, making use of sequence similarity networks. We concluded that the ricin-B/CBM13 superfamily may be larger than initially thought and that several putative ricin-B lectin domains may display CAZyme functionalities, although biochemical proof remains to be delivered. CONCLUSIONS: Ricin-B lectin domains and CBM13 modules are associated groups of proteins whose database semantics are currently biased towards ricin-B lectins. Revision of the CAZy cross-reference in UniProt and introduction of a dedicated CBM13 domain identifier in InterPro may resolve this issue. In addition, our analyses show that several proteins with putative ricin-B lectin domains show very strong structural similarity to CBM13 modules. Therefore ricin-B lectin domains and CBM13 modules could be considered distant members of a larger ricin-B/CBM13 superfamily.

Genome sequencing of Elaeocarpus spp. stem blight pathogen Pseudocryphonectria elaeocarpicola reveals potential adaptations to colonize woody bark.

BACKGROUND: Elaeocarpus spp. stem blight, caused by Pseudocryphonectria elaeocarpicola, is a destructive disease, which will significantly reduce the productivity and longevity of Elaeocarpus spp. plants, especially in the Guangdong Province of China. However, few information is available for P. elaeocarpicola. To unravel the potential adaptation mechanism of stem adaptation, the whole genome of P. elaeocarpicola was sequenced by using the DNBSEQ and PacBio platforms. RESULTS: P. elaeocarpicola harbors 44.49 Mb genome with 10,894 predicted coding genes. Genome analysis revealed that the P. elaeocarpicola genome encodes a plethora of pathogenicity-related genes. Analysis of carbohydrate-active enzymes (CAZymes) revealed a rich variety of enzymes participated in plant cell wall degradation, which could effectively degrade cellulose, hemicellulose and xyloglucans in the plant cell wall and promote the invasion of the host plant. There are 213 CAZyme families found in P. elaeocarpicola, among which glycoside hydrolase (GH) family has the largest number, far exceeding other tested fungi by 53%. Besides, P. elaeocarpicola has twice as many genes encoding chitin and cellulose degradation as Cryphonectria parasitica, which belong to the same family. The predicted typical secreted proteins of P. elaeocarpicola are numerous and functional, including many known virulence effector factors, indicating that P. elaeocarpicola has great potential to secrete virulence effectors to promote pathogenicity on host plants. AntiSMASH revealed that the genome encoded 61 secondary metabolic gene clusters including 86 secondary metabolic core genes which was much higher than C. parasitica (49). Among them, two gene cluster of P. elaeocarpicola, cluster12 and cluster52 showed 100% similarity with the mycotoxins synthesis clusters from Aspergillus steynii and Alternaria alternata, respectively. In addition, we annotated cytochrome P450 related enzymes, transporters, and transcription factors in P. elaeocarpicola, which are important virulence determinants of pathogenic fungi. CONCLUSIONS: Taken together, our study represents the first genome assembly for P. elaeocarpicola and reveals the key virulence factors in the pathogenic process of P. elaeocarpicola, which will promote our understanding of its pathogenic mechanism. The acquired knowledge lays a foundation for further exploration of molecular interactions with the host and provide target for management strategies in future research.

Comparative genomic analysis of Planctomycetota potential for polysaccharide degradation identifies biotechnologically relevant microbes.

BACKGROUND: Members of the Planctomycetota phylum harbour an outstanding potential for carbohydrate degradation given the abundance and diversity of carbohydrate-active enzymes (CAZymes) encoded in their genomes. However, mainly members of the Planctomycetia class have been characterised up to now, and little is known about the degrading capacities of the other Planctomycetota. Here, we present a comprehensive comparative analysis of all available planctomycetotal genome representatives and detail encoded carbohydrolytic potential across phylogenetic groups and different habitats. RESULTS: Our in-depth characterisation of the available planctomycetotal genomic resources increases our knowledge of the carbohydrolytic capacities of Planctomycetota. We show that this single phylum encompasses a wide variety of the currently known CAZyme diversity assigned to glycoside hydrolase families and that many members encode a versatile enzymatic machinery towards complex carbohydrate degradation, including lignocellulose. We highlight members of the Isosphaerales, Pirellulales, Sedimentisphaerales and Tepidisphaerales orders as having the highest encoded hydrolytic potential of the Planctomycetota. Furthermore, members of a yet uncultivated group affiliated to the Phycisphaerales order could represent an interesting source of novel lytic polysaccharide monooxygenases to boost lignocellulose degradation. Surprisingly, many Planctomycetota from anaerobic digestion reactors encode CAZymes targeting algal polysaccharides - this opens new perspectives for algal biomass valorisation in biogas processes. CONCLUSIONS: Our study provides a new perspective on planctomycetotal carbohydrolytic potential, highlighting distinct phylogenetic groups which could provide a wealth of diverse, potentially novel CAZymes of industrial interest.

A bioinformatic workflow for in silico secretome prediction with the lignocellulose degrading ascomycete fungus Parascedosporium putredinis NO1.

The increasing availability of microbial genome sequences provides a reservoir of information for the identification of new microbial enzymes. Genes encoding proteins engaged in extracellular processes are of particular interest as these mediate the interactions microbes have with their environments. However, proteomic analysis of secretomes is challenging and often captures intracellular proteins released through cell death and lysis. Secretome prediction workflows from sequence data are commonly used to filter proteins identified through proteomics but are often simplified to a single step and are not evaluated bioinformatically for their effectiveness. Here, a workflow to predict a fungal secretome was designed and applied to the coding regions of the Parascedosporium putredinis NO1 genome. This ascomycete fungus is an exceptional lignocellulose degrader from which a new lignin-degrading enzyme has previously been identified. The 'secretome isolation' workflow is based on two strategies of localisation prediction and secretion prediction each utilising multiple available tools. The workflow produced three final secretomes with increasing levels of stringency. All three secretomes showed increases in functional annotations for extracellular processes and reductions in annotations for intracellular processes. Multiple sequences isolated as part of the secretome lacked any functional annotation and made exciting candidates for novel enzyme discovery.

Multifaceted roles of plant glycosyl hydrolases during pathogen infections: more to discover.

MAIN CONCLUSION: Carbohydrates are hydrolyzed by a family of carbohydrate-active enzymes (CAZymes) called glycosidases or glycosyl hydrolases. Here, we have summarized the roles of various plant defense glycosidases that possess different substrate specificities. We have also highlighted the open questions in this research field. Glycosidases or glycosyl hydrolases (GHs) are a family of carbohydrate-active enzymes (CAZymes) that hydrolyze glycosidic bonds in carbohydrates and glycoconjugates. Compared to those of all other sequenced organisms, plant genomes contain a remarkable diversity of glycosidases. Plant glycosidases exhibit activities on various substrates and have been shown to play important roles during pathogen infections. Plant glycosidases from different GH families have been shown to act upon pathogen components, host cell walls, host apoplastic sugars, host secondary metabolites, and host N-glycans to mediate immunity against invading pathogens. We could classify the activities of these plant defense GHs under eleven different mechanisms through which they operate during pathogen infections. Here, we have provided comprehensive information on the catalytic activities, GH family classification, subcellular localization, domain structure, functional roles, and microbial strategies to regulate the activities of defense-related plant GHs. We have also emphasized the research gaps and potential investigations needed to advance this topic of research.

Pontiella agarivorans sp. nov., a novel marine anaerobic bacterium capable of degrading macroalgal polysaccharides and fixing nitrogen.

Marine macroalgae produce abundant and diverse polysaccharides, which contribute substantially to the organic matter exported to the deep ocean. Microbial degradation of these polysaccharides plays an important role in the turnover of macroalgal biomass. Various members of the Planctomycetes-Verrucomicrobia-Chlamydia (PVC) superphylum are degraders of polysaccharides in widespread anoxic environments. In this study, we isolated a novel anaerobic bacterial strain NLcol2T from microbial mats on the surface of marine sediments offshore Santa Barbara, CA, USA. Based on 16S ribosomal RNA (rRNA) gene and phylogenomic analyses, strain NLcol2T represents a novel species within the Pontiella genus in the Kiritimatiellota phylum (within the PVC superphylum). Strain NLcol2T is able to utilize various monosaccharides, disaccharides, and macroalgal polysaccharides such as agar and É©-carrageenan. A near-complete genome also revealed an extensive metabolic capacity for anaerobic degradation of sulfated polysaccharides, as evidenced by 202 carbohydrate-active enzymes (CAZymes) and 165 sulfatases. Additionally, its ability of nitrogen fixation was confirmed by nitrogenase activity detected during growth on nitrogen-free medium, and the presence of nitrogenases (nifDKH) encoded in the genome. Based on the physiological and genomic analyses, this strain represents a new species of bacteria that may play an important role in the degradation of macroalgal polysaccharides and with relevance to the biogeochemical cycling of carbon, sulfur, and nitrogen in marine environments. Strain NLcol2T (= DSM 113125T = MCCC 1K08672T) is proposed to be the type strain of a novel species in the Pontiella genus, and the name Pontiella agarivorans sp. nov. is proposed.IMPORTANCEGrowth and intentional burial of marine macroalgae is being considered as a carbon dioxide reduction strategy but elicits concerns as to the fate and impacts of this macroalgal carbon in the ocean. Diverse heterotrophic microbial communities in the ocean specialize in these complex polymers such as carrageenan and fucoidan, for example, members of the Kiritimatiellota phylum. However, only four type strains within the phylum have been cultivated and characterized to date, and there is limited knowledge about the metabolic capabilities and functional roles of related organisms in the environment. The new isolate strain NLcol2T expands the known substrate range of this phylum and further reveals the ability to fix nitrogen during anaerobic growth on macroalgal polysaccharides, thereby informing the issue of macroalgal carbon disposal.

Identification of carbohydrate gene clusters obtained from in vitro fermentations as predictive biomarkers of prebiotic responses.

BACKGROUND: Prebiotic fibers are non-digestible substrates that modulate the gut microbiome by promoting expansion of microbes having the genetic and physiological potential to utilize those molecules. Although several prebiotic substrates have been consistently shown to provide health benefits in human clinical trials, responder and non-responder phenotypes are often reported. These observations had led to interest in identifying, a priori, prebiotic responders and non-responders as a basis for personalized nutrition. In this study, we conducted in vitro fecal enrichments and applied shotgun metagenomics and machine learning tools to identify microbial gene signatures from adult subjects that could be used to predict prebiotic responders and non-responders. RESULTS: Using short chain fatty acids as a targeted response, we identified genetic features, consisting of carbohydrate active enzymes, transcription factors and sugar transporters, from metagenomic sequencing of in vitro fermentations for three prebiotic substrates: xylooligosacharides, fructooligosacharides, and inulin. A machine learning approach was then used to select substrate-specific gene signatures as predictive features. These features were found to be predictive for XOS responders with respect to SCFA production in an in vivo trial. CONCLUSIONS: Our results confirm the bifidogenic effect of commonly used prebiotic substrates along with inter-individual microbial responses towards these substrates. We successfully trained classifiers for the prediction of prebiotic responders towards XOS and inulin with robust accuracy (≥ AUC 0.9) and demonstrated its utility in a human feeding trial. Overall, the findings from this study highlight the practical implementation of pre-intervention targeted profiling of individual microbiomes to stratify responders and non-responders.

Comparative genome analysis of the genus Marivirga and proposal of two novel marine species: Marivirga arenosa sp. nov., and Marivirga salinae sp. nov.

BACKGROUND: The phylum Bacteroidota represents a significant proportion of heterotrophic bacteria found in marine ecosystems. Members of the phylum Bacteroidota are actively involved in the degradation of biopolymers such as polysaccharides and proteins. Bacteroidota genomes exhibit a significant enrichment of various enzymes, including carbohydrate-active enzymes (CAZymes), carboxypeptidases, esterases, isomerases, peptidases, phosphatases, and sulfatases. The genus Marivirga, a member of the family Marivirgaceae within the phylum Bacteroidota, comprises six documented species. During a microbial diversity study, three novel Marivirga strains (BKB1-2 T, ABR2-2, and BDSF4-3 T) were isolated from the West Sea, Republic of Korea. RESULTS: To explore the taxonomic status and genomic characteristics of the novel isolates, we employed a polyphasic taxonomic approach, which included phylogenetic, chemotaxonomic and comprehensive genome analysis. The three isolates were Gram-stain-negative, aerobic, rod-shaped, moderately halophilic, and had a gliding motility. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values among the two isolates, BKB1-2 T and BDSF4-3 T, and the six reference strains were 70.5-76.5% for ANI and 18.1-25.7% for dDDH. Interestingly, the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the strains harbor genes for a comprehensive pathway for dissimilatory nitrate reduction to ammonium (DNRA), as well as other nitrogen pathways for the reduction of nitrite, nitric oxide, and nitrous oxide. Additionally, the antiSMASH analysis indicated that the strains contained three to eight biosynthetic gene clusters (BGCs) associated with the synthesis of secondary metabolites. Furthermore, the strains carried a high number of CAZyme ranging from 53 to 152, which was also demonstrated by an in vitro analysis of degradation of the polysaccharide cellulose, chitin, laminarin, starch, and xylan. Additionally, all the strains carried genes for the metabolism of heavy metals, and exhibited tolerance to heavy metals, with minimum inhibitory concentrations (MICs) in millimoles (mM) in ranges of Co2+ (3-6), Cu2+ (0.2-0.4), Ni2+ (3-5), Zn2+ (2-4), Mn2+ (20-50), and Hg2+ (0.3). CONCLUSIONS: Based on polyphasic taxonomic approach, the three isolated strains represent two novel species names Marivirga arenosa sp. nov. (BKB1-2 T = KCTC 82989 T = InaCC B1618T), and Marivirga salinae sp. nov. (BDSF4-3 T = KCTC 82973 T = InaCC B1619T).

Uncovering the lignin-degrading potential of Serratia quinivorans AORB19: insights from genomic analyses and alkaline lignin degradation.

BACKGROUND: Lignin is an intricate phenolic polymer found in plant cell walls that has tremendous potential for being converted into value-added products with the possibility of significantly increasing the economics of bio-refineries. Although lignin in nature is bio-degradable, its biocatalytic conversion is challenging due to its stable complex structure and recalcitrance. In this context, an understanding of strain's genomics, enzymes, and degradation pathways can provide a solution for breaking down lignin to unlock the full potential of lignin as a dominant valuable bioresource. A gammaproteobacterial strain AORB19 has been isolated previously from decomposed wood based on its high laccase production. This work then focused on the detailed genomic and functional characterization of this strain based on whole genome sequencing, the identification of lignin degradation products, and the strain's laccase production capabilities on various agro-industrial residues. RESULTS: Lignin degrading bacterial strain AORB19 was identified as Serratia quinivorans based on whole genome sequencing and core genome phylogeny. The strain comprised a total of 123 annotated CAZyme genes, including ten cellulases, four hemicellulases, five predicted carbohydrate esterase genes, and eight lignin-degrading enzyme genes. Strain AORB19 was also found to possess genes associated with metabolic pathways such as the ß-ketoadipate, gentisate, anthranilate, homogentisic, and phenylacetate CoA pathways. LC-UV analysis demonstrated the presence of p-hydroxybenzaldehyde and vanillin in the culture media which constitutes potent biosignatures indicating the strain's capability to degrade lignin. Finally, the study evaluated the laccase production of Serratia AORB19 grown with various industrial raw materials, with the highest activity detected on flax seed meal (257.71 U/L), followed by pea hull (230.11 U/L), canola meal (209.56 U/L), okara (187.67 U/L), and barley malt sprouts (169.27 U/L). CONCLUSIONS: The whole genome analysis of Serratia quinivorans AORB19, elucidated a repertoire of genes, pathways and enzymes vital for lignin degradation that widens the understanding of ligninolytic metabolism among bacterial lignin degraders. The LC-UV analysis of the lignin degradation products coupled with the ability of S. quinivorans AORB19 to produce laccase on diverse agro-industrial residues underscores its versatility and its potential to contribute to the economic viability of bio-refineries.

Carbohydrate-active enzymes involved in rice cell wall metabolism.

Plant cell walls are complex, multifunctional structures, built up of polysaccharides and proteins. The configuration and abundance of cell wall constituents determine cellular elongation and plant growth. The emphasis of this review is on rice, a staple crop with economic importance, serving as model for grasses/cereals. Recent advancements have contributed to a better understanding of the grass/cereal cell wall. This review brings together the current knowledge about the organisation and metabolism of the rice cell wall, and addresses gaps and missing information connected to the cell wall of rice and the enzymes involved. Several cell wall fractions, including cellulose, mixed-linkage glucans and glucuronoarabinoxylans, are well-understood in rice and other grasses/grains. Conversely, there are still open questions and missing links when it comes down to xyloglucans, glucomannans, pectin, lignin and arabinogalactan proteins. There is still a large and untapped potential to identify carbohydrate-active enzymes (CAZymes), to characterise their activity and to elucidate their involvement in the metabolism of the mentioned cell wall fractions. With this review, we demonstrate the current state and demarcate the research areas with potential for further investigations.

From glycans to green biotechnology: exploring cell wall dynamics and phytobiota impact in plant glycopathology.

Filamentous plant pathogens, including fungi and oomycetes, pose significant threats to cultivated crops, impacting agricultural productivity, quality and sustainability. Traditionally, disease control heavily relied on fungicides, but concerns about their negative impacts motivated stakeholders and government agencies to seek alternative solutions. Biocontrol agents (BCAs) have been developed as promising alternatives to minimize fungicide use. However, BCAs often exhibit inconsistent performances, undermining their efficacy as plant protection alternatives. The eukaryotic cell wall of plants and filamentous pathogens contributes significantly to their interaction with the environment and competitors. This highly adaptable and modular carbohydrate armor serves as the primary interface for communication, and the intricate interplay within this compartment is often mediated by carbohydrate-active enzymes (CAZymes) responsible for cell wall degradation and remodeling. These processes play a crucial role in the pathogenesis of plant diseases and contribute significantly to establishing both beneficial and detrimental microbiota. This review explores the interplay between cell wall dynamics and glycan interactions in the phytobiome scenario, providing holistic insights for efficiently exploiting microbial traits potentially involved in plant disease mitigation. Within this framework, the incorporation of glycobiology-related functional traits into the resident phytobiome can significantly enhance the plant's resilience to biotic stresses. Therefore, in the rational engineering of future beneficial consortia, it is imperative to recognize and leverage the understanding of cell wall interactions and the role of the glycome as an essential tool for the effective management of plant diseases.

Functional characterization and structural insights of three cutinases from the ascomycete Fusarium verticillioides.

Cutinases are serine esterases that belong to the α/ß hydrolases superfamily. The natural substrates for these enzymes are cutin and suberin, components of the plant cuticle, the first barrier in the defense system against pathogen invasion. It is well-reported that plant pathogens produce cutinases to facilitate infection. Fusarium verticillioides, one important corn pathogens, is an ascomycete upon which its cutinases are poorly explored. Consequently, the objective of this study was to perform the biochemical characterization of three precursor cutinases (FvCut1, FvCut2, and FvCut3) from F. verticillioides and to obtain structural insights about them. The cutinases were produced in Escherichia coli and purified. FvCut1, FvCut2, and FvCut3 presented optimal temperatures of 20, 40, and 35 °C, and optimal pH of 9, 7, and 8, respectively. Some chemicals stimulated the enzymatic activity. The kinetic parameters revealed that FvCut1 has higher catalytic efficiency (Kcat/Km) in the p-nitrophenyl-butyrate (p-NPB) substrate. Nevertheless, the enzymes were not able to hydrolyze polyethylene terephthalate (PET). Furthermore, the three-dimensional models of these enzymes showed structural differences among them, mainly FvCut1, which presented a narrower opening cleft to access the catalytic site. Therefore, our study contributes to exploring the diversity of fungal cutinases and their potential biotechnological applications.

Genomic analysis of the class Phycisphaerae reveals a versatile group of complex carbon-degrading bacteria.

Bacteria of the phylum Planctomycetota have received much attention over the years due to their unique cell biology and potential for biotechnological application. Within the phylum, bacteria of the class Phycisphaerae have been found in a multitude of environmental datasets. However, only a few species have been brought into culture so far and even enrichments are scarce. Therefore, very little is known about their lifestyle, which has hindered efforts to estimate their environmental relevance. Here, we analysed all medium- and high-quality Phycisphaerae genomes represented in the genome taxonomy database to learn more about their physiology. We combined automatic and manual annotation efforts to provide a bird's eye view of their diverse energy metabolisms. Contrasting previous reports, we did not find indications for the presence of genes for anaerobic ammonium oxidation in any Phycisphaerae genome. Instead, we found that many members of this class are adapted to a facultative anaerobic or strictly fermentative lifestyle and may be specialized in the breakdown of carbon compounds produced by other organisms. Based on these findings, we provide a practical overview of organic carbon substrates predicted to be utilized by Phycisphaerae families.

A metagenomic survey of the fecal microbiome of the African savanna elephant (Loxodonta africana).

The African savanna elephant (Loxodonta africana) is the largest terrestrial animal on Earth and is found primarily in Southern and Eastern Africa. It is a hindgut, colonic fermenter and subsists on a diet of raw plant materials found in its grazing area. In this study the bacterial, archaeal and fungal populations of seven African savanna elephant fecal metagenomes were first characterized using amplicon sequencing. On the genus level it was observed that the p-1088-a5 gut group in the bacteriome, Methanocorpusulum and Methanobrevibacter in the archaeome and Alternaria, Aurobasidium, Didymella and Preussia in the mycome, predominated. Subsequently, metagenomic shotgun sequencing was employed to identify possible functional pathways and carbohydrate-active enzymes (CAZymes). Carbohydrate catabolic pathways represented the main degradation pathways, and the fecal metagenome was enriched in the glycohydroside (GH) class of CAZymes. Additionally, the top GH families identified - GH43, GH2, GH13 and GH3 - are known to be associated with cellulytic, hemicellulytic and pectolytic activities. Finally, the CAZymes families identified in the African savanna elephant were compared with those found in the Asian elephant and it was demonstrated that there is a unique repository of CAZymes that could be leveraged in the biotechnological context such as the degradation of lignocellulose for the production of second-generation biofuels and energy.

Optimizing liquid fermentation for Wolfiporia cocos: gene expression and biosynthesis of pachymic acid and mycelial biomass.

Wolfiporia cocos, a versatile fungus acclaimed for its nutritional and therapeutic benefits in Traditional Chinese Medicine, holds immense potential for pharmaceutical and industrial applications. In this study, we aimed to optimize liquid fermentation techniques and culture medium composition to maximize mycelial biomass (MB) yield, pachymic acid (PA) concentration, and overall PA production. Additionally, we investigated the molecular basis of our findings by quantifying the expression levels of genes associated with PA and MB biosynthesis using quantitative real-time polymerase chain reaction. Under the optimized fermentation conditions, significant results were achieved, with maximum MB reaching 6.68 g l-1, PA content peaking at 1.25 mg g-1, and a total PA yield of 4.76 g l-1. Notably, among the four examined genes, squalene monooxygenase, exhibited enhanced expression at 0.06 ratio under the optimized conditions. Furthermore, within the realm of carbohydrate-active enzymes, the glycoside hydrolases 16 family displayed elevated expression levels at 21 ratios, particularly during MB production. This study enhances understanding of genetic mechanism governing MB and PA production in W. cocos, highlighting the roles of squalene monooxygenase and glycoside hydrolases 16 carbohydrate-active enzymes.

Recent expansion of metabolic versatility in Diplonema papillatum, the model species of a highly speciose group of marine eukaryotes.

BACKGROUND: Diplonemid flagellates are among the most abundant and species-rich of known marine microeukaryotes, colonizing all habitats, depths, and geographic regions of the world ocean. However, little is known about their genomes, biology, and ecological role. RESULTS: We present the first nuclear genome sequence from a diplonemid, the type species Diplonema papillatum. The ~ 280-Mb genome assembly contains about 32,000 protein-coding genes, likely co-transcribed in groups of up to 100. Gene clusters are separated by long repetitive regions that include numerous transposable elements, which also reside within introns. Analysis of gene-family evolution reveals that the last common diplonemid ancestor underwent considerable metabolic expansion. D. papillatum-specific gains of carbohydrate-degradation capability were apparently acquired via horizontal gene transfer. The predicted breakdown of polysaccharides including pectin and xylan is at odds with reports of peptides being the predominant carbon source of this organism. Secretome analysis together with feeding experiments suggest that D. papillatum is predatory, able to degrade cell walls of live microeukaryotes, macroalgae, and water plants, not only for protoplast feeding but also for metabolizing cell-wall carbohydrates as an energy source. The analysis of environmental barcode samples shows that D. papillatum is confined to temperate coastal waters, presumably acting in bioremediation of eutrophication. CONCLUSIONS: Nuclear genome information will allow systematic functional and cell-biology studies in D. papillatum. It will also serve as a reference for the highly diverse diplonemids and provide a point of comparison for studying gene complement evolution in the sister group of Kinetoplastida, including human-pathogenic taxa.

Metagenomic analysis of carbohydrate-active enzymes and their contribution to marine sediment biodiversity.

Marine sediments constitute the world's most substantial long-term carbon repository. The microorganisms dwelling in these sediments mediate the transformation of fixed oceanic carbon, but their contribution to the carbon cycle is not fully understood. Previous culture-independent investigations into sedimentary microorganisms have underscored the significance of carbohydrates in the carbon cycle. In this study, we employ a metagenomic methodology to investigate the distribution and abundance of carbohydrate-active enzymes (CAZymes) in 37 marine sediments sites. These sediments exhibit varying oxygen availability and were isolated in diverse regions worldwide. Our comparative analysis is based on the metabolic potential for oxygen utilisation, derived from genes present in both oxic and anoxic environments. We found that extracellular CAZyme modules targeting the degradation of plant and algal detritus, necromass, and host glycans were abundant across all metagenomic samples. The analysis of these results indicates that the oxic/anoxic conditions not only influence the taxonomic composition of the microbial communities, but also affect the occurrence of CAZyme modules involved in the transformation of necromass, algae and plant detritus. To gain insight into the sediment microbial taxa, we reconstructed metagenome assembled genomes (MAG) and examined the presence of primary extracellular carbohydrate active enzyme (CAZyme) modules. Our findings reveal that the primary CAZyme modules and the CAZyme gene clusters discovered in our metagenomes were prevalent in the Bacteroidia, Gammaproteobacteria, and Alphaproteobacteria classes. We compared those MAGs to organisms from the same taxonomic classes found in soil, and we found that they were similar in its CAZyme repertoire, but the soil MAG contained a more abundant and diverse CAZyme content. Furthermore, the data indicate that abundant classes in our metagenomic samples, namely Alphaproteobacteria, Bacteroidia and Gammaproteobacteria, play a pivotal role in carbohydrate transformation within the initial few metres of the sediments.

Unique properties of a Dictyostelium discoideum carbohydrate-binding module expand our understanding of CBM-ligand interactions.

Deciphering how enzymes interact, modify, and recognize carbohydrates has long been a topic of interest in academic, pharmaceutical, and industrial research. Carbohydrate-binding modules (CBMs) are noncatalytic globular protein domains attached to carbohydrate-active enzymes that strengthen enzyme affinity to substrates and increase enzymatic efficiency via targeting and proximity effects. CBMs are considered auspicious for various biotechnological purposes in textile, food, and feed industries, representing valuable tools in basic science research and biomedicine. Here, we present the first crystallographic structure of a CBM8 family member (CBM8), DdCBM8, from the slime mold Dictyostelium discoideum, which was identified attached to an endo-ß-1,4-glucanase (glycoside hydrolase family 9). We show that the planar carbohydrate-binding site of DdCBM8, composed of aromatic residues, is similar to type A CBMs that are specific for crystalline (multichain) polysaccharides. Accordingly, pull-down assays indicated that DdCBM8 was able to bind insoluble forms of cellulose. However, affinity gel electrophoresis demonstrated that DdCBM8 also bound to soluble (single chain) polysaccharides, especially glucomannan, similar to type B CBMs, although it had no apparent affinity for oligosaccharides. Therefore, the structural characteristics and broad specificity of DdCBM8 represent exceptions to the canonical CBM classification. In addition, mutational analysis identified specific amino acid residues involved in ligand recognition, which are conserved throughout the CBM8 family. This advancement in the structural and functional characterization of CBMs contributes to our understanding of carbohydrate-active enzymes and protein-carbohydrate interactions, pushing forward protein engineering strategies and enhancing the potential biotechnological applications of glycoside hydrolase accessory modules.