Search Results (294)

Ganoderma mushrooms produce bioactive metabolites with therapeutic potential, yet their tissue-level distribution is not well characterized. This study quantified triterpenoids, β-glucans, and phenolics across six anatomical sections of fruiting bodies from five wild Ganoderma species. Twenty-six triterpenoids were identified. Laccate species showed thicker context tissue enriched in ganoderic and lucidenic acids, resembling the chemotype of G. lucidum. Matte species displayed greater triterpenoid diversity, including elfvingic, applanoxidic, and ganoderenic analogues. Maximum contents reached 3.5% triterpenoids, 34.3% β-glucans, 20.8 mg TE·g⁻¹ (ABTS), 175.2 µmol Fe²⁺·g⁻¹ (FRAP), and 23.5 mg GAE·g⁻¹ phenolics. Triterpenoids and phenolics were highest in outer cap tissues, while β-glucans predominated in context layers. These patterns reflect functional tissue roles and developmental variation. The tissue distribution of metabolites in wild Ganoderma presented here identifies surface tissues as a major source of triterpenoids and phenolics, and internal tissues as a source of β-glucans. These traits represent selection targets for extraction and selective breeding to produce strains with thicker context tissue and higher triterpenoid and β-glucan yields. Full article

Avibacterium paragallinarum (Av. paragallinarum), the causative agent of infectious coryza, imposes substantial economic burdens on the poultry industry by inducing growth retardation in broilers and reducing egg production in laying hens by up to 40%. Disease control is hindered by the limited efficacy of available vaccines and the increasing prevalence of antibiotic resistance—challenges that are exacerbated by the pathogen’s capacity to form biofilms, which facilitate bacterial persistence and enhance drug tolerance. To systematically elucidate the genetic determinants underlying biofilm formation in Av. Paragallinarum, we constructed a high-density random mutant library using mini-Tn5 transposon mutagenesis, comprising 3106 individual mutants. Phenotypic screening via crystal violet staining identified 188 mutants displaying altered biofilm-forming capacity relative to the wild-type strain, including 172 with enhanced and 16 with reduced biofilm formation. Sequencing of transposon insertion sites in these mutants revealed 105 disrupted genes involved in diverse biological pathways, including amino acid metabolism, quorum sensing, and transmembrane transport. A representative subset of eight mutants was selected for detailed phenotypic characterization. Their biofilm phenotypes were consistent with the initial screening results; certain mutants exhibited markedly enhanced biofilm formation (e.g., Tn-2206), whereas others, including Tn-1504, Tn-2428, and Tn-2859, showed significant reductions in biofilm production. Notably, these three biofilm-deficient mutants—harboring disruptions in a TonB-dependent receptor (Tn-1504), a GntP family permease (Tn-2428), and a hypothetical protein (Tn-2859)—displayed drastically attenuated virulence in vitro. Compared with the wild-type strain, these mutants exhibited reductions in cytotoxicity (up to 66.38%), cell adhesion (up to 50.68%), and invasive capacity, while maintaining normal growth kinetics. These findings indicate that the identified genes may play crucial roles in biofilm-associated virulence and highlight Tn-1504, Tn-2428, and Tn-2859 as promising candidates for the development of live attenuated vaccines. Collectively, this study provides a comprehensive genetic foundation for the rational design of novel anti-biofilm strategies against Av. paragallinarum. Full article

Fish host complex intestinal bacterial communities that contribute to a wide range of functions, from nutrient assimilation to modulation of the immune system. Understanding how environmental and host-related factors shape the fish gut microbiota is essential for advancing sustainable aquaculture practices. This study compared the intestinal microbiota of gilthead sea bream (Sparus aurata) between wild and aquaculture populations in western Greece using 16S rRNA gene amplicon sequencing targeting the V3–V4 region, combined with culture-based methods. The analysis was based on a 97% similarity threshold and included 141 gastrointestinal samples of fish collected at two aquaculture facilities and two wild fisheries, representing two different growth phases (150 g and 300 g body weight). High-throughput sequencing data revealed a clear separation of gut microbial communities according to origin (wild vs. aquaculture), geographic location, and body growth phase, with most wild fish groups exhibiting higher microbial diversity than their farmed counterparts, except for group MES_150 which showed similar or lower values. The gut microbiota was dominated by Pseudomonadota (53%), Bacillota (29%), Actinomycetota (7%), Deinococcota (5%), and Bacteroidota (4%). A shared core microbiome, comprising Psychrobacter, Staphylococcus, Geobacillus, Aeromonas, Enterobacter, Pantoea, Bacillus, and Acinetobacter, was detected across all populations. Wild fish were enriched in Psychrobacter, Aeromonas, and Photobacterium, while aquaculture fish displayed higher abundances of Vibrio, Allomeiothermus, and Staphylococcus. Network analysis revealed mostly mutually exclusive interactions in both groups but distinct patterns of co-occurrence, driven mainly by Paenibacillus, Enterobacter, and Staphylococcus in wild samples, and by Vibrio, Aeromonas, and Pseudomonas in farmed fish. Culture-based assays demonstrated greater diversity in wild fish, dominated by Pseudomonas, Staphylococcus, and Vibrio strains, in contrast to the frequent occurrence of Staphylococcus and Psychrobacter in aquaculture samples. The findings suggest that aquaculture practices significantly alter gut microbial community structure and reduce diversity, with potential implications for fish health and disease resistance. The identified core and differentially abundant taxa provide candidates for probiotic development to improve aquaculture sustainability. Full article

Marine-derived filamentous fungi are a rich source of structurally diverse and biologically active natural products. However, many biosynthetic gene clusters (BGCs) in fungi remain silent under standard conditions. In this study, we employed a metabolic shunting strategy to disrupt azaphilone biosynthesis in the marine-derived fungus Penicillium sclerotiorum E23Y-1A by deleting the pathway-specific regulator gene A00667. HPLC analysis revealed the emergence of new metabolite peaks in the mutant strain Δ667 compared to the wild type. Subsequent purification yielded seven compounds: the mutant produced two novel meroterpenoids sclerotilins A and B (1 and 2) along with the known steroids ergosta-5,7,22-trien-3β-ol (3) and cerevisterol (4), while the wild type yielded the known steroid (22E)-5α,8α-epidioxyergosta-6,22-dien-3β-ol (5) and two azaphilones geumsanol G (6) and 5-chloro-3-[(1E,3R,4R,5S)-3,4-dihydroxy-3,5-dimethyl-1-hepten-1-yl]-1,7,8,8a-tetrahydro-7,8-dihydroxy-7-methyl-(7R,8R,8aS)-6H-2-benzopyran-6-one (7). Bioactivity assays showed that compound 6 exhibited moderate antimicrobial activity against Staphylococcus aureus, and compound 3 displayed moderate cytotoxicity against five human cancer cell lines. These results demonstrate that A00667 is essential for azaphilone biosynthesis and that its disruption leads to the production of structurally distinct natural products, highlighting the potential of pathway engineering to redirect fungal metabolism to yield novel natural products. Full article

This study provides an updated overview of Anthophila (wild bees and honey bees) diversity and conservation status in Algeria, explicitly distinguishing between the managed honey bee (Apis mellifera) and native wild Anthophila species. Using a systematic PRISMA-based literature analysis, more than 179 bee species have been documented across Mediterranean and semi-arid ecosystems, confirming their irreplaceable contribution to ecosystem resilience and crop pollination and beekeeping systems. The majority of Algeria’s Anthophila diversity is represented by endemic and native wild bees that sustain natural ecosystems. However, they are under growing human-caused (anthropogenic) pressures in the Anthropocene, including pressure from habitat loss and fragmentation, agricultural intensification, widespread pesticide use, and climate change. In addition, pathogenic threats such as Varroa destructor, Nosema, and associated viruses are well documented in honey bees, while evidence for their presence and impact in wild bees in Algeria remains very limited. These stressors not only weaken specialist species but also accelerate biotic homogenization dominated by A. mellifera. Recent genomic research on native honey bee populations has revealed adaptive signatures linked to immunity and social behavior, offering new opportunities for innovative conservation strategies based on molecular and genetic tools. Such insights highlight the value of preserving local strains, which may hold key traits for resilience under changing environmental conditions. To safeguard Anthophila biodiversity, this study underscores the urgent need for Algeria to implement proven conservation strategies, including habitat restoration initiatives and Anthophila-friendly farming approaches, which are common internationally but remain largely unaddressed at the national scale. By integrating cutting-edge genetic research, ecological restoration, and sustainable innovation, Algeria, with its diverse habitats and largely unexplored Anthophila fauna, holds considerable potential for advancing biodiversity conservation strategies that also support food security. However, this potential can only be realized through further in-depth research and comprehensive species inventories. Full article

3-Amino-4-hydroxybenzoic acid (3,4-AHBA) is a non-proteinogenic aromatic compound that functions as a key biosynthetic precursor for diverse secondary metabolites with pharmaceutical and industrial value. Microbial production of 3,4-AHBA offers a sustainable alternative to petroleum-based chemical synthesis; however, metabolic complexity and trade-offs between growth and product formation constrain rational strain design. Here, genome-scale metabolic (GSM) modeling and flux balance analysis (FBA) were integrated with targeted genetic engineering to elucidate and enhance 3,4-AHBA production in Streptomyces thermoviolaceus. A genome-scale metabolic model was constructed and expanded by incorporating the nspH–nspI gene operon, which encodes the 3,4-AHBA biosynthetic pathway. In silico FBA predicted substantial rewiring of central carbon metabolism, with carbon flux redirected from glycolysis and the tricarboxylic acid cycle toward aspartate-derived intermediates and 3,4-AHBA synthesis, accompanied by reduced biomass-associated flux. Guided by these predictions, an engineered strain (St::NspHI) was developed and experimentally evaluated. Consistent with model predictions, the engineered strain exhibited lower growth rates and glucose uptake than the wild type, reflecting a metabolic burden. Nevertheless, 3,4-AHBA production was achieved exclusively in the engineered strain. Comparison of simulated and experimental fluxes revealed overestimation by FBA, likely due to secondary metabolism and incomplete genome annotation. Overall, GSM-guided design enables optimization of precursor production. Full article

While the biobased, fermentative production of succinate by Escherichia coli represents a sustainable alternative to its conventional synthesis from petroleum, this process requires substantial amounts of inorganic carbon (C_i) to support CO₂-fixing reactions in the reductive branch of the tricarboxylic acid (rTCA) cycle. Accordingly, intracellular C_i availability represents a potential limiting factor during E. coli succinate fermentations. Here, we first investigate the role and importance of E. coli’s native CO₂ concentrating mechanism (CCM)—comprising two carbonic anhydrases (CAs), Can and CynT—by comparing and contrasting the behaviors of wild-type E. coli and the engineered succinate-producing strain, KJ122. Deletion of can and cynT significantly impaired the aerobic growth of both strains under low CO₂ atmosphere and/or low pH, outcomes that were further exacerbated under anaerobic conditions for KJ122. During bioreactor fermentations, KJ122 Δcan ΔcynT further exhibited a prolonged lag phase (~48 h) and 44% reduced succinate production relative to KJ122 by 96 h. Next, the relative functions and performance of mechanistically diverse, heterologous CCM components were investigated by characterizing their ability to restore growth and/or succinate production. While the cyanobacterial bicarbonate transporter SbtA and the C_i transporter DabAB from Halothiobacillus neapolitanus each complemented growth at 0.05% CO₂ and pH 6.5–7.5, neither fully restored succinate production by KJ122 Δcan ΔcynT. Moreover, individual overexpression of sbtA, dabAB, or can in KJ122 rendered no additional improvements to succinate production. Collectively, while these results point to the critical importance of CA for supporting efficient fermentative succinate production by E. coli, they also suggest that this native CCM alone is sufficient for ensuring C_i acquisition at requisite levels under the conditions examined. Full article

Cypripedium shanxiense S. C. Chen has high ornamental value; it relies on specific habitats and fungi. Wild C. shanxiense populations need urgent conservation because they are declining rapidly. This study investigated three wild C. shanxiense populations under different canopy densities in the Changbai Mountains, analyzing habitat characteristics and plant morphology. Tissue isolation methods, molecular identification techniques, and metagenomic approaches were applied separately to purify root-colonizing fungi and to investigate the composition and functions of rhizosphere fungi, thereby revealing the diversity of root mycobiome in C. shanxiense. Results revealed that C. shanxiense achieved the best growth when the canopy density was 85.29%, and the lowest growth was under 96.13% canopy density. Soil phosphorus and potassium contents reached their highest levels under 69.33% canopy density, while soil nitrogen and organic matter contents peaked at 85.29%. Soil organic matter and available nitrogen constitute the core nutrient factors for the growth of C. shanxiense. A total of 16 fungal strains were mainly enriched in the roots, all belonging to Ascomycota. Including numerous growth-promoting fungi and pathogenic fungi. The rhizosphere fungi were mainly enriched with Basidiomycota and Ascomycota. Functional genes related to replication, recombination, and repair, and Glycoside Hydrolases. This study clarifies the optimal growth conditions of this species and the dominant rhizosphere and root fungi, providing a scientific basis for the ecological restoration and conservation of rare species. Full article

Protoparvoviruses are highly contagious pathogens that cause severe, often fatal diseases in both domestic and wild carnivores. Golden jackal (Canis aureus) populations have experienced expansion in recent years, increasingly occupying urban and peri-urban areas. Despite this, they remain largely overlooked in scientific research. This study aimed to detect and characterise Protoparvovirus carnivoran1 circulating in a golden jackal population in Croatia and to assess their role in the epidemiology of parvovirus infections in companion animals. Small intestines from 55 jackals hunted in 2024 and 2025 were tested for Protoparvovirus carnivoran1 using real-time PCR. Positive samples were found across all sampling sites, with an overall positivity rate of 40%. Based on characteristic amino acid residues within the VP2 protein, the viruses detected in jackals were classified as feline panleukopenia virus (FPV). Phylogenetic analysis of the VP2 protein demonstrated considerable genetic diversity among strains circulating in Croatia. Additionally, a distinct group was identified, shared exclusively by Croatian domestic cats and golden jackals. Amino acid analysis revealed the novel A91T mutation, found only in jackals, and the E411Q mutation, unique to Croatian FPV strains. Structural modelling of the VP2 protein indicates that the observed mutations are located on the protein surface, within the antibody-binding site. These findings highlight the potential role of wild carnivores in parvovirus epidemiology and underscore the importance of including them in future surveillance and research efforts. Full article

Recently acquired evidence indicates that bacteria can utilise yeasts as survival niches. This study investigated the presence of hidden, intracellular bacteria (endobacteria) within wild yeasts collected from natural ecosystems and evaluated whether biotechnological processes influenced these bacterial communities. We examined the microbiotas of 28 axenic cultures of wild yeasts; these were selected due to their potential brewing and biocontrol uses and were isolated from habitats associated with Quercus and Vitis. We also analysed the microbiotas present after these strains were used to ferment beer wort. Bacterial communities were characterised using 16S rRNA gene amplicon metagenomics. The results indicate that yeast strains and their endobacterial partners have coevolved, and their compositions are shaped by the environmental conditions. Substantial bacterial diversity was detected across strains in both axenic cultures and post-fermentation samples. The ecological origin of the yeast (oak- or grape-associated) did not significantly affect the endobacterial community structure. Across all samples, the dominant phyla were Proteobacteria, Actinobacteria, Firmicutes, and Cyanobacteria, with Proteobacteria representing over 90% of sequences. Most bacterial genera were shared between axenic and fermentation conditions. However, Escherichia and Comamonas predominated in axenic cultures, while Parvibaculum dominated after fermentation. These findings suggest that yeasts constitute stable microhabitats for bacterial communities, and their relative abundances can shift during fermentation processes. Full article

A growing body of evidence indicates that artificial manipulation of transcriptional regulation is a powerful approach to activate cryptic biosynthetic gene clusters (BGCs) of secondary metabolites (SMs) in fungi. In this study, one mutant strain MNP-2-OE::veA was constructed by overexpressing the global transcription regulator veA in an Arctic-derived strain Aspergillus sydowii MNP-2. Chemical investigation of the mutant OE::veA resulted in the isolation of one novel polyhydroxy anthraquinone (1) together with nine known metabolites (2–10), which were unambiguously characterized by various spectroscopic methods including 1D and 2D NMR and HR-ESI-MS as well as via comparison with literature data. Biosynthetically, compounds 1 and 10 as new arising chemicals were, respectively, formed by type II polyketide synthase (T2PK) and non-ribosomal peptide synthetase (NRPS), which were silent in the wild-type (WT) strain MNP-2. A bioassay showed that only compound 3 had weak inhibitory effect on human pathogen Candida albicans, with a MIC value of 64 ug/mL, and 4 displayed in vitro weak cytotoxic activity against HCT116 cells (IC₅₀ = 44.47 μM). These results indicate that overexpression of veA effectively awakened the cryptic BGCs in fungal strains and enhanced their structural diversity in natural products. Full article

Xylanases (EC 3.2.1.8) are value-added enzymes essential for biomass deconstruction and are widely used in the pulp and paper, food, feed, and biofuel sectors. This review provides a comprehensive analysis of the current state and future prospects of xylanase research and application. It begins by examining the structural diversity of xylan substrates and the corresponding classification of xylanase enzymes, their catalytic mechanisms, and methods for their functional study, such as inhibitor analysis. The discussion then covers the challenges and methods involved in the purification of xylanases from complex biological mixtures. While natural microbial sources (fungi and bacteria) remain important, the limitations of wild-type (WT) strains for industrial production are highlighted. The review assesses the most common recombinant production systems, including Escherichia coli, Bacillus subtilis, and Komagataella phaffii, comparing their advantages for high-yield enzyme production. Finally, the paper focuses on protein engineering strategies as powerful tools for enhancing key enzyme properties (thermostability, specific activity, and pH tolerance). By integrating fundamental knowledge with applied technological approaches, this review underscores the critical role of xylanases in industrial biotechnology and identifies future research directions for their optimization. Full article

The Na⁺/K⁺/2Cl⁻ cotransporter (NKCC) gene encodes a critical membrane transporter involved in cellular ion homeostasis and plays a pivotal role in osmoregulation and salinity adaptation in aquatic organisms. This study identified and validated SNP markers in the NKCC gene associated with low-salinity tolerance in Scylla paramamosain. Four SNPs (g.196C>A, g.8374T>A, g.8385T>A and g.91143T>A) were screened and genotyped in low-salinity tolerant and intolerant groups. Association analysis revealed that mutant genotypes at all four sites were significantly enriched in the tolerant group (p <0.05), with the values of odds ratios (OR) greater than 1. The tolerant group exhibited significantly higher genetic diversity than the intolerant group. Haplotype analysis showed the wild CTTT haplotype dominated in the intolerant group, while mutant-containing haplotypes were significantly elevated in the tolerant group. A positive correlation was observed between the mutant and NKCC expression. Functional validation by qRT-PCR demonstrated that mutant allele carriers exhibited significantly higher NKCC mRNA expression levels than the wild-type carriers. Moreover, the expression level of homozygous mutations is significantly higher than that of heterozygous mutations. These validated SNPs could provide effective molecular markers for marker-assisted selection breeding of low-salinity tolerant S. paramamosain strains, offering important theoretical and practical implications for sustainable aquaculture development. Full article

Ganoderma sichuanense is a widely used medicinal and edible fungus. Genomic studies have revealed substantial genetic variation among its different strains, indicating that a genetic transformation system optimized for one genotype may not be effective in others. However, no study has systematically evaluated the efficiency of a genetic transformation system across diverse genotypes, which has potentially limited functional genetic studies in this species. In this study, we first evaluated eight wild and cultivated monokaryotic strains with different genotypes based on their hygromycin B resistance and green fluorescent protein (GFP) expression efficiency. Three strains (CCMJ1500101, CCMJ1509001, and CCMJ1507802) were identified as capable of stable foreign gene expression, achieving transformation efficiencies of 20.0–66.7% via PEG-mediated protoplast transformation. Subsequently, a CRISPR/Cas9 system incorporating seven key elements to enhance editing efficiency was constructed and applied to these three strains using the ura3 gene as a test target. Gene editing efficiencies varied significantly among genotypes, ranging from 14.3% to 75.0%, confirming the system’s high efficacy and genotype dependence. Importantly, to rigorously assess the robustness and versatility of the established transformation platform, we further validated its broad applicability in the best-performing strain, CCMJ1500101, by successfully editing five functional genes involved in growth, development, and metabolism. Notably, gene inversion events were detected for the first time in edited transformants of Ganoderma, providing new clues for understanding non-homologous end joining (NHEJ) repair in this species. This study establishes a robust dual-sgRNA CRISPR/Cas9 platform for G. sichuanense and provides valuable strain resources to facilitate future gene functional studies and genetic improvement. Full article

Porcine circovirus types 2 (PCV2) and 3 (PCV3) are major pathogens affecting swine health and productivity, yet important gaps remain in understanding their evolution and circulation in Europe, particularly within wild boar populations that may serve as reservoirs. This study examined the genetic diversity and evolutionary dynamics of PCV2 and PCV3 in Portugal, drawing on viral genomes obtained from domestic pigs and wild boars to explore transmission patterns, spillover events and the contribution of recombination to viral emergence. We identified two PCV2 genotypes (PCV2a and PCV2d) and two PCV3 genotypes (PCV3-2a and PCV3-3g) circulating in Portuguese swine. Phylogeographic reconstruction revealed multiple introductions of both PCV2 and PCV3 from China into Europe, followed by regional diversification and subsequent spread within European wild boar populations. Evidence of bidirectional viral exchange between domestic pigs and wild boars was also observed. Recombination played a notable role in PCV2 evolution, with consistent signals detected among PCV2a sequences and indications that the PCV2h genotype likely originated from a recombinant event involving a Portuguese PCV2a strain and a Chinese PCV2d strain. By contrast, no recombination was detected in PCV3, suggesting that its evolution is primarily mutation-driven. Overall, these findings highlight the complex evolutionary history of swine circoviruses in Europe and underscore the importance of continuous genomic surveillance in both domestic and wild hosts. The study reinforces the value of a One Health approach for monitoring and controlling emerging circoviruses with implications for animal health and livestock production. Full article