Journal of microbiology and biotechnology最新文献

The microbiome significantly influences immune dysfunction and gut dysbiosis in patients with ankylosing spondylitis (AS). This study focuses on defining the distinct microbial characteristics within AS and biomarkers associated with disease activity. 44 patients with AS and 50 healthy controls (HC) were recruited. 16S rRNA sequencing was conducted to assess the microbiome of stool samples. The Ankylosing Spondylitis Disease Activity Score with C-reactive protein (ASDAS-CRP) was calculated for all AS patients; scores ≥ 2.1 indicated high disease activity, while < 2.1 indicated low disease activity. Similar alpha diversity profiles were maintained in both AS and HC cohorts, whereas significant differences were identified in beta diversity. The compositional prevalence of Proteobacteria, particularly Gammaproteobacteria, and Enterobacterales, including Escherichia spp., in the AS group was significantly increased. On the other hand, beneficial taxa, including Firmicutes, Clostridia, Clostridiales, Lachnospiraceae, Ruminococcaceae, and Faecalibacterium, were highly abundant in the HC group. Among patients with AS, alpha diversity decreased in the high disease activity group compared to the low disease activity group, while beta diversity did not differ significantly. Moreover, Coprobacter spp. abundance positively correlated with the Bath Ankylosing Spondylitis Disease Activity Index (p = 0.032) and the ASDAS-CRP (p = 0.023). Patients with AS exhibit distinct gut microbiota profiles, with increased Proteobacteria and decreased beneficial taxa such as Firmicutes. Greater disease activity is accompanied by reduced alpha diversity, while Coprobacter spp. abundance correlates with disease activity, suggesting its potential as a biomarker.

Amicyanin is a type 1 Cu protein that mediates electron transfer between methylamine dehydrogenase and cytochrome c-551i in Paracoccus denitrificans. In this study, a Ser mutation was introduced at either Asn47 or Asn54 located in the Cu-binding ligand loop containing His53 to determine their role in amicyanin functionality. Their spectral and redox properties, and protein stability according to temperature variance and oxidative stress were investigated. N47S amicyanin indicated similar redox potential and stability to native amicyanin. The reaction kinetic of N47S amicyanin toward methylamine dehydrogenase exhibited a similar electron transfer rate, but immensely improved binding affinity compared to native amicyanin. For N54S amicyanin, it attained a more positive redox potential and greatly reduced stability. N54S amicyanin also altered the reaction kinetics with increased electron transfer and decreased binding affinity. Combined with these results, computational simulations of the N47S and N54S mutations suggest that the Ser substitution at Asn54 alters the geometry of the Cu active site by changing the surrounding H-bond pattern. On the other hand, although N47S did not affect the active site, it can be deduced that the position of Asn47 in the loop is significantly altered to influence the interaction of amicyanin with MADH. Hence, we conclude that Asn47 takes charge of the amicyanin affinity for MADH while Asn54 regulates the electron transfer by altering the redox midpoint potential of the active site.

Exosomes are nanosized vesicles released from multivesicular bodies into the extracellular space. Lactic acid bacteria-derived exosomes have emerged as promising next-generation bioactive agents with beneficial effects on skin health and immune regulation. In this study, we aimed to investigate the anti-inflammatory and skin-lightening activities of exosomes derived from Latilactobacillus sakei isolated from the flowers of Aster koraiensis (BK-5 exosomes). Experiments were performed using lipopolysaccharide-stimulated RAW 264.7 macrophages and α-melanocyte-stimulating hormone-induced B16F10 melanoma cells. BK-5 exosomes exhibited no cytotoxicity in either cell line and effectively suppressed the production of proinflammatory cytokines, including interleukin (IL)-6, IL-1β, tumor necrosis factor-α, and prostaglandin E₂, as well as the expression of inducible nitric oxide synthase and cyclooxygenase-2. Moreover, BK-5 exosomes reduced the expression of melanogenesis-related proteins, including tyrosinase, tyrosinase-related protein (TRP)-1, TRP-2, and microphthalmia-associated transcription factor in a dose-dependent manner. Furthermore, BK-5 exosomes inhibited the phosphorylation of nuclear factor kappa-light-chain-enhancer of activated B cells and inhibitor of κB alpha, as well as the phosphorylation of mitogen-activated protein kinase pathway components, including extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38. Collectively, these findings confirm that BK-5 exosomes effectively attenuate inflammatory responses in RAW 264.7 macrophages through suppression of multiple signaling pathways and inhibit melanin biosynthesis in B16F10 cells, thereby demonstrating their potential as multifunctional bioactive materials applicable to pharmaceutical and cosmetic fields.

The 2009 pandemic H1N1 (pdm09) virus is both zoonotic and reverse-zoonotic, transmitting from swine to humans and vice versa. During the early zoonotic phase, immediately after the species jump and before substantial antigenic drift had accumulated, recombinant vaccine strains bearing hemagglutinin (HA) and neuraminidase (NA) from early pdm09 viruses often replicated poorly in embryonated chicken eggs (ECEs), contributing to delays and shortages in vaccine supply. Developing seed strains that are more productive in ECEs while preserving antigenicity and minimizing mammalian pathogenic potential is therefore essential for future pandemic preparedness. Efficient egg replication requires a balanced activity between HA and NA and their coordinated interaction with the polymerase subunit PB2. To this end, we generated PR8-derived recombinants combining PB2 backbones with distinct polymerase activities with targeted HA and NA modifications and edits to segment-specific 3' and 5' noncoding regions (NCRs). Comparative analysis of viral titers, together with sequence-based predictions of mutation effects, identified genotypes that improved replication in eggs while minimizing antigenic variations and reducing markers associated with mammalian virulence. Although further enhancement of viral yield is still warranted, these results delineate practical design principles, favoring balanced tuning of HA-NA functions, PB2 compatibility, and NCR context over large receptor-shift mutations, for engineering influenza seed strains. This work provides actionable guidance to support vaccine development and strengthen One-Health-oriented pandemic preparedness.

Emerging fungal pathogens pose a significant threat to global public health. Despite the availability of antifungal agents, their clinical efficacy is increasingly challenged by the rise of fungicide-resistant strains. Therefore, identifying novel therapeutic targets and ensuring the safe application of antifungal agents are critical for advancing treatment strategies. Autophagy, a fundamental cellular process that maintains intracellular homeostasis by degrading and recycling dysfunctional proteins and organelles, is implicated in fungal pathogenicity. It indicates that inhibition of autophagy represents a promising approach for antifungal development. In this study, we evaluate the antifungal potential of autophagy inhibitors targeting the Autophagy-related protein 4 (ATG4)-mediated cleavage of Autophagy-related protein 8 (ATG8). Our findings demonstrate that ebselen and its analogs effectively inhibit ATG4 activity in Cryptococcus neoformans, Aspergillus fumigatus, and Aspergillus niger, exhibiting fungicidal activity against Cryptococcus and Candida species. These results provide valuable insights into novel antifungal development strategies, highlighting the therapeutic potential of autophagy inhibitors against diverse pathogenic fungi.

The rise of bacterial resistance presents a pressing public health challenge, necessitating innovative antimicrobial solutions. Based on our prior work demonstrating the activity of pyrazolo[3,4-d] pyrimidine-based analogues against human dihydrofolate reductase (DHFR), we hypothesized that novel derivatives could function as dual-action antibacterial agents. In this study, we therefore evaluated the capacity of these compounds to both inhibit bacterial DHFR and disrupt quorum sensing (QS)-mediated virulence. The compounds were screened for antibacterial activity against several bacterial strains and the anti-DHF activity was assayed. Anti-virulence potential was assessed in vitro and in vivo, while interactions with the QS targets were investigated using in silico study. Staphylococcus aureus and Pseudomonas aeruginosa were chosen as representative clinically important gram-negative and gram-positive pathogens, respectively, to evaluate the antibacterial and anti-virulence activities. The in vitro and in silico DHFR inhibition assays revealed potent antibacterial activity for the synthesized compounds against various bacteria. Among the most promising candidates, compound 7_e exhibited potent antibacterial activity at low minimum inhibitory concentrations (MICs) and demonstrated synergy with conventional antibiotics. The in vitro and in vivo evaluations showed promising anti-virulence activities of the synthesized compounds, particularly 7_a and 7_e, against P. aeruginosa and S. aureus, respectively. Dynamics simulations showed the strong binding affinity of 7_a and 7_e to LasI/R and AgrC QS targets in P. aeruginosa and S. aureus, respectively. In conclusion, our findings demonstrate that pyrazolo[3,4-d]pyrimidine analogues, especially 7_a and 7_e, can function as effective dual-action agents by inhibiting DHFR and suppressing QS-mediated virulence, thus representing a promising new class of anti-virulence therapeutics against priority bacterial pathogens.

Chronic constipation is a common gastrointestinal disorder characterized by impaired intestinal motility and dysbiosis of the gut microbiota. This study evaluated the therapeutic potential of the probiotic yeast Saccharomyces cerevisiae var. boulardii (HO12) on loperamide-induced constipation in rats. We assessed fecal parameters, intestinal transit, histological alterations, neurotransmitter levels, and short-chain fatty acid (SCFA) production. HO12 administration effectively alleviated constipation symptoms in a dose-dependent manner. It significantly increased fecal number, weight, water content, and improved gastrointestinal transit, indicating enhanced intestinal motility. HO12 also restored serotonin levels and the expression of serotonin-related genes, key regulators of enteric neurotransmission. Furthermore, HO12 increased the production of acetic acid and total SCFAs, contributing to a favorable intestinal environment. Histological analysis demonstrated that HO12 restored mucosal thickness, crypt cell distribution, and interstitial cells of Cajal, thereby supporting intestinal functional recovery. In conclusion, our results demonstrate that HO12 alleviates loperamide-induced constipation by modulating intestinal neurotransmitters, enhancing intestinal motility, and increasing SCFA production. These findings suggest that S. boulardii HO12 has strong potential as a therapeutic probiotic for constipation management.

Severe Fever with Thrombocytopenia Syndrome virus (SFTSV) is a tick-borne pathogen associated with high mortality, particularly among the elderly, and poses a significant public health threat in East Asia's rural areas. Improved diagnostic approaches are crucial for effective disease management in resource-limited settings. In this study, we developed a portable Reverse Transcription Loop-Mediated Isothermal Amplification (RT-LAMP) platform for rapid and accurate SFTSV detection. Primers targeting the conserved L gene were designed using 158 sequences collected between 2019 and 2023, and a multiplex assay incorporating the β-actin gene as an internal control to verify amplification efficiency in alignment with clinical performance requirements. We assessed the assay's sensitivity and specificity, ultimately establishing the RT-LAMP platform in a portable, clinical-grade device. The RT-LAMP assay achieved a detection limit of 500 RNA copies per reaction and provided results within 20 min. It demonstrated high specificity without cross-reactivity with other pathogens. Performance comparisons with commercial qRT-PCR kits using clinical specimens confirmed the RT-LAMP platform's reliability. The newly developed RT-LAMP platform is a rapid, accurate, and cost-effective diagnostic tool for SFTSV with strong potential for point-of-care testing (POCT). Its portability and ease of use make it ideal for application in resource-constrained environments, thereby enhancing SFTSV surveillance and improving public health outcomes in affected regions.

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by progressive cartilage degradation, synovial inflammation, and impaired joint function. This study investigated the therapeutic potential of a botanical mixture, NWG11-02, comprising Elaeagnus angustifolia L. (oleaster) extract (OE) and Sophora japonica L. fruit extract (SJFE) in a monosodium iodoacetate (MIA)-induced OA rat model. Oral administration of the mixture for four weeks significantly improved weight-bearing function and preserved cartilage architecture, as evidenced by reduced Mankin scores and enhanced proteoglycan retention in toluidine blue-stained sections. Molecular analyses revealed that the mixture markedly enhanced the expression of cartilage anabolic markers (aggrecan and type II collagen) and reduced matrix-degrading enzymes (MMP-3 and MMP-13), while suppressing the expression of pro-inflammatory cytokines (COX-2, TNF-α, IL-1β, IL-6) at both mRNA and protein levels. These effects exhibited a dose-dependent pattern, with the medium-dose mixture (80 mg/kg) demonstrating superior efficacy compared to individual extracts. No systemic toxicity was observed during the treatment period. Collectively, these findings suggest that NWG11-02 exerts multi-faceted protective effects on joint tissues and may serve as a promising natural therapeutic strategy for OA management.

Saccharomyces cerevisiae and Cyberlindnera jadinii are widely utilized in the natural food seasoning industry as sources of flavor enhancing nucleotides such as inosine monophosphate (IMP) and guanosine monophosphate (GMP), which contribute to umami taste and support sodium reduction in food. However, wild type yeast strains produce GMP at levels that are inadequate for industrial scale applications, necessitating metabolic engineering strategies to increase production efficiency. This study employed a CRISPR-Cas9-based scarless genome engineering approach to enhance GMP biosynthesis in S. cerevisiae via promoter replacement. The key genes IMD3 and GUA1, responsible for converting IMP to GMP, were overexpressed to redirect purine flux toward GMP production. To address precursor limitations, ZWF1 and RKI1, involved in the pentose phosphate pathway, were also overexpressed. In parallel, the expression of STB5 and RAP1 was increased to enhance NADPH regeneration and relieve transcriptional bottlenecks. As a result, the final engineered strain SCJ-7 demonstrated a 1.77-fold increase in GMP titer and a 1.40-fold increase in GMP content during flask fermentation compared to the wild-type. In fed-batch fermentation, GMP titer was further improved by 27.6%. These findings demonstrate that combining metabolic flux enhancement with transcriptional regulation provides an effective and scalable strategy for boosting GMP production in S. cerevisiae, offering strong potential for industrial application in the food industry.