This study systematically elucidated the microbial community succession and functional gene dynamics during the postharvest spoilage process of Stropharia rugosoannulata by integrating physiological and biochemical indicators with metagenomic analysis. The experimental results demonstrated that as storage time extended, the activities of antioxidant enzymes (superoxide dismutase, peroxidase) in S. rugosoannulata significantly declined, while the content of membrane lipid peroxidation product malondialdehyde increased, leading to compromised cell membrane integrity and creating favorable conditions for microbial colonization. Metagenomic analysis revealed that during the spoilage phase (post-harvest day 14), the relative abundance of Pseudomonadota increased to 85.7%, with Pseudomonas replacing Ewingella as the absolutely dominant microbial population. Further functional gene analysis showed that the post-harvest day 14 exhibited significant enrichment of glycosyltransferases (GT0, GT1, GT2, GT4) and carbohydrate-binding modules (CBM10, CBM16, CBM50), along with pectinase (GH78), chitinase (GH19), and polysaccharide-modifying enzymes (CE4, CE11). This indicated a metabolic shift towards cell wall synthesis and substrate recognition. In contrast, the post-harvest day 7, prior to fruiting body softening, demonstrated high expression of glycoside hydrolases (GH1, GH2, GH4, GH94) and carbohydrate esterase CE8, focusing on the degradation of cellulose and starch. These findings, for the first time from a molecular ecology perspective, clarify that the essence of postharvest spoilage in S. rugosoannulata is a quality deterioration process driven by a Pseudomonas-dominated microbial community. The study provided a basis for the development of targeted antibacterial preservation strategies.

Sanghuangporus baumii polysaccharides (SBP) are recognized for their valuable pharmacological activities, driving increasing interest in their medicinal potential. However, the biosynthetic pathway of SBP remains incompletely characterized. Phosphoglucose isomerase (PGI), a key enzyme in carbohydrate metabolism, catalyzes the reversible isomerization between glucose-6-phosphate (G-6-P) and fructose-6-phosphate (F-6-P) and is hypothesized to regulate polysaccharide biosynthesis in this fungus. In this study, the pgi gene from S. baumii (sbpgi) was cloned and created sbpgi-silenced mutants using RNA interference (RNAi) to investigate its function. Silencing sbpgi resulted in an approximately 20% reduction in mycelial biomass but concurrently enhanced the production of exopolysaccharide (EPS) and intracellular polysaccharide (IPS) by approximately 2.0-fold and 1.9-fold after 9 days, respectively. Furthermore, suppression of sbpgi expression markedly decreased the content of cell wall β-1,3-glucan (by ~23%) while increasing chitin deposition by about 1.7-fold, leading to alterations in cell wall architecture, including thickness, and changes in stress tolerance. Transcriptional analysis revealed that sbpgi silencing significantly upregulated the expression of key genes in the polysaccharide biosynthetic pathway, including ugpg and pmm, highlighting the critical regulatory role of sbpgi in polysaccharide production. Our findings provide a foundation for metabolic engineering strategies to develop high-yielding strains for the industrial production of SBP.

Plastics are indispensable in modern society, but their increasing production and disposal pose serious environmental challenges, including pollution and the depletion of non-renewable resources. Poly(ethylene furanoate) (PEF), a bio-based polyester composed of ethylene glycol and 2,5-furandicarboxylic acid (FDCA), is attracting attention as a sustainable alternative to poly(ethylene terephthalate) (PET). In this study, we developed a fully biotechnological upcycling system for PEF. Our approach involved enzymatic depolymerization of PEF to release FDCA, followed by microbial conversion of FDCA into polyhydroxyalkanoate (PHA), a biodegradable polyester. From soil samples enriched with FDCA as the sole carbon source, we isolated two bacterial strains: Pseudomonas sp. S8-1 and Caballeronia sp. S8-5. These strains produced medium-chain-length and short-chain-length PHAs, respectively, in defined medium containing FDCA. For enzymatic depolymerization, we employed the thermostable ICCG variant (F243I/D238C/S283C/Y127G) of leaf-branch compost cutinase, known for its high PET-degrading activity. The depolymerization of PET by this enzyme was enhanced by the addition of calcium carbonate (CaCO3) powder to suppress acidification. Furthermore, the enzyme retained high activity even after partial purification by heat treatment at 60°C and efficiently depolymerized PEF as well. Finally, the PEF degradation solution was successfully utilized as a carbon source for PHA production by strain S8-5. These results demonstrate a proof-of-concept biorecycling system for PEF and represent a first step toward sustainable plastic management.

An efficient bacterial consortium (designated BPA-1), comprising Bacillus subtilis SX-6, Pseudomonas sp. SX-10, and Georgenia sp. SY-1, was successfully constructed for the decolorization of the azo dye Congo Red (CR). BPA-1 exhibited significant thermotolerance and heavy metal resistance, achieving over 90% CR decolorization within 60 h at 47°C under co-stress conditions with Zn²⁺, Mn²⁺, and Pb²⁺ (50 mg/L each). The consortium demonstrated broad substrate specificity, effectively decolorizing 12 structurally diverse azo dyes. Enzymatic assays revealed the involvement of laccase, manganese peroxidase, lignin peroxidase, and azoreductase in CR biodegradation. Metabolic pathway analysis indicated a three-stage degradation mechanism: (1) Asymmetric cleavage of azo bonds (-N=N-) generated 4,4'-diazaldenylbiphenyl and 4-amino-1-naphthalenesulfonic acid (Intermediate II); (2) Deamination converted Intermediate II to 3,4-dihydroxy-1-naphthalenesulfonic acid, followed by desulfurization to form naphthalene-1,2,3,4-tetraol; (3) Complete mineralization of intermediates occurred through subsequent oxidative steps. Notably, 4,4'-diazaldenylbiphenyl was further transformed into 4,4'-diaminobiphenyl, confirming the consortium's capacity for multi-step detoxification.

Bacillus thuringiensis (Bt) has broad spectrum multipotent functionalities for pest and disease suppression, and growth promotion (PGP) of plants. Therefore, potency of 27 rice rhizospheric and 2 commercial Bt isolates was assessed for biocidal and PGP traits. Functionally rhizospheric Bts were broadly superior than commercial Bts. Virulence of the Bts varied against rice leaf folder (LF, Cnaphalocrocis medinalis) and stripe stem borer (SSB, Chilo suppresalis) larvae in laboratory, net house and field tests. Drosophila diet (DD) incorporation, cut leaf and field assays proved virulence of 5-9 Bt isolates against LF larvae with LC50s 1.99 - 6.31 x 108, 2.18 x 106 - 2.25 x 109 and 3.16 x 106 - 1.25 x 109 bacteria-spore-crystal (BSC)/ml, respectively, and TB261 was most (LC50s 2.18 x 106 - 3.98 x 108 BSC/ml) infective. DD and cut stem assays for SSB proved virulence of 5 and 6 Bts with LC50s 9.20 x 106 - 3.62 x 108 and 9.21 x 106- 3.24 x 108 BSC/ml, respectively, and maximum (LC50s 9.20 - 9.21 x 108 BSC/ml) infectivity of TB263. Eight Bts inhibited 1-4 out of 7 rice pathogens and 16 Bts antagonized 1-4 out of 9 entomopathogenic fungi. Biocidal principles of the Bts were cell wall/membrane hydrolyzing exoenzymes, toxin/inhibitors and crystal toxins. Furthermore, the Bts were also inhibited by 3 insecticides and 2 fungicides. The Bts possessed 1-4 PGP and phytostimulation traits also. The potent rhizospheric Bt can be prospected for overall improvement/sustenance of rice.

In the biosynthesis of peptidoglycan (PG), murE protein (MurE) adds a diamino acid at position 3 of the peptide chain of peptidoglycan. The diamino acid that is added by MurE and makes cross-linkage with adjacent peptide chain differs depending on the bacterial species: Gram-negative bacteria add meso-diaminopimelic acid (DAP), while most Gram-positive bacteria add L-lysine (Lys). In this study, the murE gene of Levilactobacillus brevis that transfers Lys in PG synthesis was cloned into Escherichia coli that has DAP-type PG. The transformant cells harboring L. brevis murE showed reduction of colony forming units during cultivation, and were elongated or burst when murE was expressed. Amino acid analysis of solubilized PG revealed that the Lys/DAP ratio increased in the PG of the transformants. Interestingly, aspartic acid that is responsible for the formation of cross-linkages between Lys and other peptide chain in the PG of L. brevis also increased, suggesting that Lys-type PG with Asp cross-linkage was partially formed by the cloning of murE gene.

Superoxide dismutases (SODs) play crucial roles in protecting cells against oxidative stress by catalyzing the dismutation of superoxide radicals. In Aspergillus nidulans, five putative SOD genes have been predicted in the genome; however, their comparative expression profiles and physiological functions remain largely uncharacterized. In this study, we analyzed the expression levels of all five SOD genes at different growth stages and examined the oxidative stress sensitivity of corresponding gene-disrupted strains. We found that sodA exhibited high and constitutive expression across all growth stages, while sodB was predominantly expressed in conidia (asexual spores). Disruption mutants of sodA and sodB showed increased sensitivity to oxidative agents, confirming their functional importance. Subcellular fractionation and SOD activity assays revealed that SodA was localized in the cytoplasm, whereas SodB was primarily localized in mitochondria. These results highlight the growth stage-specific expression and distinct cellular roles of SodA and SodB in A. nidulans, providing novel insights into the oxidative stress defense system in filamentous fungi.

Methyl methacrylate (MMA), the primary raw material of acrylic resin, is an important polymeric material due to its increasing demand and ease of recycling. The most promising biosynthetic route for MMA involves the condensation of methanol with methacrylyl-CoA (MAA-CoA), an intermediate in the valine degradation pathway. The toxicity of MAA-CoA, poor stability and low activity of the heterologous pathway enzymes make this biosynthetic pathway less feasible. For enabling the evolutionary engineering of this pathway and its components (enzymes), we constructed a biosensor system in which the cellular level of key intermediate MAA-CoA can be evaluated in a high-throughput manner. With the aid of this MAA-CoA sensory system, we could establish the functional pathway from isobutyric acid to MAA-CoA. The sensor described in this paper should be valuable tool in the design-build-test-learn cycle for optimizing and breeding this MMA pathway.

Methionine gamma-lyase enzyme was isolated and purified from Mucor irregularis PQ344458 fungal isolates, that obtained from plant root, the isolates were identified through observation of their colony morphological features, implementation of PCR and DNA sequencing via sanger-chain termination approach, then data of DNA sequence alignment, phylogenetic tree, percent identity was generated. Through implementation of several stages that involved using of ion-exchange chromatography, gel-filtration chromatography, ammonium sulphate, enzyme isolation and purification stages were accomplished. The enzyme extract then, was analyzed for its protein content, specific activity and Impact of pH, temperature, inhibitors and activators on its kinetics. Additionally, MTT and DPPH radical scavenging assays were carried-out to reveal information about anti-cancer and anti-oxidant activities of methionine gamma-lyase enzyme. MTT assay results of %viable cells were 15% for HeLa cells and 6.6% for U937 cells at maximum concentration of the enzyme extract. Moreover, DPPH scavenging activity results were 82% at maximum concentration.

We investigated the effect of a starter culture on the fungal communities of commercial karebushi. Aspergillus pseudoglaucus was initially identified as the starter fungus. In karebushi samples from two manufacturers relying on naturally occurring molds, Aspergillus chevalieri was the dominant species, accompanied by Aspergillus montevidensis and Aspergillus sydowii, while A. pseudoglaucus was not detected. Among samples from six manufacturers that used the starter culture, A. pseudoglaucus was dominant in only three; in the remaining three, A. chevalieri predominated despite the starter being used. These results suggest that indigenous fungi, particularly A. chevalieri, present in the processing environment can outcompete the starter culture, influence the fungal community, and potentially contribute to the qualitative diversity of karebushi.