Search Results (1,668)

Nitrate (NO₃⁻) serves as a pivotal molecule with dual functions in nutrient supply and signaling during plant growth and development. Precise monitoring of its spatiotemporal dynamics in planta is therefore essential for dissecting the regulatory mechanisms underlying plant nitrogen metabolism. However, conventional nitrate detection methods suffer from inherent limitations, including destructive sampling, insufficient spatiotemporal resolution, and an inability to achieve real-time whole-plant monitoring. Here, we report a genetically encoded nitrate biosensor, designated NitNRCL1, constructed using a split firefly luciferase complementation system. Functional validation in both prokaryotic and eukaryotic systems demonstrates that NitNRCL1 responds to changes in nitrate availability and generates stable chemiluminescent signals in bacteria and diverse plant species. Importantly, NitNRCL1 enables non-invasive, real-time, and whole-plant monitoring of nitrate levels in living plants. Using NitNRCL1, we successfully imaged the spatiotemporal dynamics of nitrate signaling in Arabidopsis thaliana. Collectively, our findings establish NitNRCL1 as a robust and novel tool for investigating nitrate transport, signaling, and metabolic pathways in plants. This biosensor advances our mechanistic understanding of plant nitrate biology and provides a technical foundation for breeding nitrogen-use-efficient crops and developing precision fertilization strategies. Full article

Objectives: This study evaluates the research landscape of the cyanobacterium Spirulina (recently reclassified as Limnospira), a strategic resource in the nutraceutical, pharmaceutical, and functional food industries. The central objective is to transition from the traditional “superfood” narrative to a structured analysis of its modern therapeutic potential as reflected in current scientific literature. This study employs bibliometric analysis to highlight research trends and thematic directions in Spirulina-related studies, rather than to experimentally validate therapeutic effects. Methods: The investigation employed an exploratory bibliometric analysis of 996 peer-reviewed articles indexed in the Web of Science (2016–2025). Using VOSviewer software, we mapped keyword co-occurrence networks, international collaborations, and institutional clusters to identify dominant thematic directions and emerging research frontiers in biotechnology and medicine. Results: Bibliometric mapping illustrates research trends and thematic associations reported in the scientific literature centered on pathophysiological mechanisms, particularly oxidative stress, inflammation, and hepatoprotection. While often referred to as “microalgae”, Spirulina is biologically a photosynthetic prokaryote with a unique lipid profile characterized by high gamma-linolenic acid (GLA) content, although clinical evidence remains heterogeneous. The analysis highlights a robust regional research hub in the Middle East and North Africa, led by Egypt and Saudi Arabia, in contrast to fragmented inter-continental collaboration. Conclusions: The steady upward trend in publications confirms expanding academic interest in Spirulina as a functional ingredient. However, this study underscores a persistent gap between in vitro bioactivity and standardized clinical validation. These findings provide a roadmap for future biotechnological developments, emphasizing the need for more rigorous, multi-center clinical trials to bridge the “superfood” perception with evidence-based therapeutic applications. Full article

Given the rapid mutation and high transmissibility of coronaviruses, especially SARS-CoV-2, comparative genomic studies are crucial for understanding viral evolution, transmission dynamics, and therapeutic development. In prior work, we analyzed and compared the spectral distribution patterns of various k-mer subsets across 920 genome sequences, spanning from primates to prokaryotes. This revealed an evolutionary mechanism in genome sequences, indicating the presence of both CG and TA-specific selection modes. In the present study, we further investigate the specific selection modes in coronavirus genomic sequences by examining the intrinsic distribution rules of 32 XYi 6-mer subset spectra. Our results show that coronavirus genomes exhibit only the CG-specific selection mode, with no evidence of TA-specific selection. Using the CG-specific selection mode, we identified CG1 6-mers as the fundamental subset underlying coronavirus genome evolution. To validate the CG1 subset, we constructed phylogenetic relationships for a set of coronaviruses and SARS-CoV-2 variant genomes. Comparative analysis confirmed that the resulting phylogenetic relationships align more closely with established knowledge. This study thus provides a theoretical framework for inferring phylogenetic relationships at the whole-genome level. Full article

Peptide deformylase (PDF) belongs to a conserved enzyme family critical for N-terminal methionine excision (NME), an essential protein maturation process in prokaryotes and eukaryotic organelles (chloroplasts, mitochondria). To explore the potential functions of OsPDFs in Oryza sativa, this study employed bioinformatics approaches and experimental validation to systematically identify and analyze the OsPDF gene family. Three OsPDF genes (OsPDF1A, OsPDF1B, OsPDF1B2) were identified in rice. These genes are exclusively distributed on chromosome 1. The biophysical properties of these proteins showed that OsPDF1A and OsPDF1B are alkaline proteins, while OsPDF1B2 is acidic, and all are hydrophilic with moderate thermostability potential. Synteny analysis revealed closer evolutionary relationships between Oryza sativa and the monocot Triticum aestivum than with dicots, reflecting conserved PDF function in gramineous plants. Analysis of cis-acting elements in the 2000 bp upstream region of OsPDF gene promoters revealed numerous elements associated with abiotic stress response and hormone regulation. Furthermore, quantitative real-time PCR (qRT-PCR) data supported these findings, indicating that OsPDF1A and OsPDF1B were upregulated under low-temperature stress, and all three OsPDF genes were transcriptionally activated by heat, salt and UV-B stresses, indicating their active involvement in rice growth, development, and abiotic stress tolerance. In summary, OsPDFs exhibit significant functions in rice’s stress adaptation, growth, and development. This study not only enhances our understanding of the OsPDF gene family’s genomic, evolutionary, and functional characteristics, but also provides new perspectives and foundational data for further exploring their regulatory mechanisms in protein maturation and abiotic stress responses, as well as their potential applications in rice stress tolerance breeding. Full article

Nile tilapia (Oreochromis niloticus) is a major aquaculture species worldwide, yet bacterial infections remain a critical constraint to production sustainability. Although pathogen-associated dysbiosis has been widely described, most studies have focused exclusively on bacterial communities, overlooking the multi-kingdom nature of the intestinal microbiota. This study evaluated the impact of experimental Providencia vermicola infection on both prokaryotic and microeukaryotic intestinal communities under controlled conditions. Using 16S (V3–V4) and 18S (V9) rRNA amplicon sequencing, we compared healthy and infected fish and assessed taxonomic, structural, and predicted functional changes. Infection was associated with significant compositional shifts, including increased relative abundances of Bacteroidota and Proteobacteria and decreased relative abundances of Fusobacteriota and Patescibacteria. Concomitantly, microeukaryotic groups such as Protalveolata, Nematozoa, and Phragmoplastophyta were significantly reduced. Functional prediction revealed metabolic pathway reconfiguration consistent with infection-associated ecological disturbance. Together, these results suggest that the pathogen challenge is associated with coordinated changes in the intestinal microbiota as an integrated system across multiple microbial kingdoms rather than as isolated bacterial shifts. This study supports ecosystem-level interpretations of dysbiosis and highlights the importance of incorporating cross-domain analyses into health assessment strategies in aquaculture species. Full article

Heat shock proteins 70 (HSP 70) are molecular chaperonins ubiquitously expressed in both prokaryotes and eukaryotes and are involved in the modulation and exacerbation of the immune response. The present study aimed to assess the immunohistochemical expression of HSP70 and apoptosis markers, such as TUNEL and Caspase-3, in 17 cases of cutaneous lupus erythematosus (CLE) in dogs to determine whether HSP70 expression correlates with cell apoptosis and to highlight possible involvement of HSP70 in the pathogenesis of CLE. The results revealed positive HSP70 expression in epidermal and inflammatory cells across all cases, with a significant correlation between HSP70 expression score and TUNEL-positive cells but not with Caspase-3-positive cells. This correlation could indicate a possible role for HSP70 in cell death via a caspase-independent apoptotic mechanism or other programmed cell death mechanisms, such as pyroptosis or necroptosis. The precise mechanisms by which HSP70 acts in this specific pathological context remain incompletely understood, but the results of this study provide important information for future investigations into autoimmune skin disease in dogs. Full article

Pinoresinol diglucoside (PDG), one of the major lignans isolated from E. ulmoides Oliver bark, has various pharmacological functions, including antihypertension and prevention of osteoporosis. However, the glycosyltransferase-encoding gene (GT) involved in regulating the glycosylation of pinoresinol to form PDG has not been reported in E. ulmoides. In this study, we screened and cloned the EuGT8 gene from E. ulmoides based on our transcriptome data. The expression pattern of the EuGT8 gene exhibited a strong positive correlation with dynamic changes in the PDG contents in three different organs of E. ulmoides. The expression level of the EuGT8 gene and PDG content were significantly decreased in asODN-EuGT8-treated shoot tips in comparison with the control group. Prokaryotic expression of the EuGT8 gene revealed that the purified EuGT8 protein could catalyze the conversion of pinoresinol into PDG. In addition, we performed transcriptional and metabolomic analyses to compare the differences between transgenic Arabidopsis and WT plants. A total of 1799 DEGs and 294 DEMs were identified in transgenic and WT plants. KEGG enrichment analysis showed that the DEGs were mainly enriched in phenylpropanoid biosynthesis, secondary metabolite biosynthesis, and starch/sucrose metabolism pathways. The DEMs were mainly enriched in ABC transporters, aminoacyl-tRNA biosynthesis, biosynthesis of amino acids, phenylpropanoid biosynthesis, and flavone and flavonol biosynthesis pathways. Correlation analysis between DEGs and DEMs identified a total of 231 DEGs associated with 38 DEMs, which were mainly distributed in multiple metabolic pathways. This finding provides both theoretical insights and genetic resources for breeding high-PDG E. ulmoides varieties, facilitating marker-assisted selection (MAS) and promoting sustainable E. ulmoides production in Guizhou. Full article

Chlorinated paraffins (CPs) are widely used, structurally complex mixtures of chlorinated alkanes whose ecological risks in aquatic ecosystems have raised increasing concern. However, the toxic effects and molecular mechanisms of CPs on primary aquatic producers remain poorly understood. In this study, we used the eukaryotic green algae Chlorella sp. and the prokaryotic cyanobacterium Microcystis aeruginosa (M. aeruginosa) as test organisms to systematically investigate the effects of CPs with different carbon chain lengths, namely short-chain CPs (SCCPs), medium-chain CPs (MCCPs), and long-chain CPs (LCCPs), on algal growth, photosynthetic pigment content, antioxidant systems, cellular ultrastructure, and the underlying molecular responses. Our results showed that CPs toxicity to algae is significantly dependent on both CPs carbon-chain length and algal species. Exposure to 1.0 mg/L SCCPs for 96 h produced a growth inhibition of Chlorella sp. of 14.45%. CPs’ exposure significantly altered algal Chl-a content and elicited antioxidant defense responses, and affected the synthesis and extracellular release of MC-RR and MC-LR in M. aeruginosa. Ultrastructural observations revealed cell surface wrinkling and deformation in both Chlorella sp. and M. aeruginosa. Chlorella sp. additionally exhibited thylakoid disintegration and plasmolysis. Transcriptomic analysis indicated that CPs with different chain lengths significantly downregulated genes in Chlorella sp. associated with DNA replication and mismatch repair, suggesting impairment of replication initiation and elongation and compromised genome stability. Concurrently, genes encoding photosynthetic antenna proteins and carbon fixation were upregulated. In M. aeruginosa, CPs exposure markedly disturbed energy metabolism pathways, including glycolysis/gluconeogenesis and oxidative phosphorylation, which were generally downregulated. This study provides a comparative assessment of CPs’ toxicity between the eukaryotic algae Chlorella sp. and the prokaryotic algae M. aeruginosa, revealing that toxicity is co-determined by carbon chain length and algal species. Additionally, it provides critical toxicological data and establishes a theoretical foundation for the scientific assessment of the aquatic ecological risks posed by CPs with different carbon chain lengths. Full article

The application of nitrification inhibitors (Nis) with nitrogen fertilizers is increasingly used as a management strategy to improve nitrogen use efficiency in crop production systems. To evaluate the effects of Ni dicyandiamide (DCD) and 1,2,4-triazole (TZ) on the rhizosphere microbiome and strawberry yield (Fragaria × ananassa Duch.), a two-year field experiment was conducted with three treatments: unfertilized control (C), mineral nitrogen fertilizer (N) applied in two doses (40 + 40 kg N ha⁻¹ year⁻¹), and a single nitrogen application (80 kg N ha⁻¹ year⁻¹) combined with nitrification inhibitors (N + Ni). Soil microbiota were assessed using cultivation-based methods and metabarcoding of 16S rRNA and ITS2 regions. Total bacterial counts on complex media increased from 5.85 to 6.15 log CFU g⁻¹ in the N treatment, while remaining 5.89 in N + Ni. Microscopic fungi increased in fertilized treatments during spring but decreased in July of the second year. Microbial community composition differed among treatments, although sampling time explained a larger proportion of variability than fertilization. Relative abundance of Gemmatimonas decreased under N + Ni, whereas Nitrososphaera increased. Fungal Shannon diversity decreased in N + Ni, while prokaryotic diversity did not differ significantly. Despite similar levels of mineral nitrogen measured before harvest, strawberry yield increased significantly in the N + Ni treatment in the second year, reaching 109% higher values than the control and 80% higher than the N treatment. This may indicate that the fertilization regime including nitrification inhibitors influenced nitrogen availability earlier in the growing season. Full article

Super-resolution microscopy has transformed biological imaging by enabling nanoscale visualization of cellular structures beyond the diffraction limit. However, its effective application in highly dense molecular environments still poses challenges. This is the case for 3D PhotoActivated Localization Microscopy (PALM) achieved through astigmatism in bacterial cells. The limited volume of a single bacterium highly increases the probability of the intensity profiles emitted by single chromophores to overlap, thus strongly decreasing the number of localizations, leading to dramatic undersampling. Dual-color 3D super-resolution in Escherichia coli is achieved through a combination of PALM with Expansion Microscopy (Ex-PALM). PALM provides high specificity through photoactivable (PA) fusion proteins and high localization precision, while ExM physically expands the specimen and separate densely packed molecules. This hybrid approach enables dual-color 3D single-molecule localization with about 3 nm spatial resolution, thus allowing one to measure distances down to the molecular scale. This is achieved by optimizing ExM protocols in bacteria to achieve a 4-fold isotropic expansion, by minimizing both chromatic aberrations and signal crosstalk, and by improving single-molecule sensitivity through highly selective inclined illumination. The method is applied to measure the spatial distribution of HisF and HisH proteins, involved in E. coli histidine biosynthesis. By tagging each protein with a photoactivable fluorescent protein, Ex-PALM reveals that after being synthetized, they co-localize in the bacterial volume with an average 3D distance of 19 nm. By combining labeling specificity with Ex-PALM, an effective method is developed for studying molecular organization in prokaryotes and in high-density samples in general, such as cell organelles or molecular condensates, with broad applications in microbiology, synthetic biology, and cellular biophysics. Full article

The combined effects of organic pollutants and vertical soil gradients on microbial community assembly in long-term contaminated sites remain insufficiently understood. In this study, high-throughput sequencing was employed to characterize prokaryotic communities across depth-resolved soil profiles at a contaminated site in Tianjin, China. Microbial diversity, taxonomic composition, and predicted functional traits varied significantly with soil depth and pollutant distribution. Surface soils exhibited higher richness and diversity, with Shannon, Sobs, and PD indices decreasing with depth (p = 0.020, p = 0.002, and p < 0.001, respectively). Redundancy analysis showed that the first two axes explained 89.91% of the total variance, indicating strong associations between microbial community structure and environmental variables. Community differentiation was related to pollutant type, with aromatic hydrocarbons more strongly linked to surface assemblages and chlorinated compounds associated with deeper horizons. Although the overall abundance of predicted metabolic genes decreased with depth, the distribution of major functional categories, including pathways related to organic matter degradation, remained comparatively stable. Co-occurrence network analysis revealed a progressive decline in network connectivity and complexity along the vertical gradient, with the number of edges decreasing from 853 (L1) to 447 (L3) and average degree decreasing from 16.404 to 9.122. These findings highlight depth-related environmental filtering as a key mechanism structuring microbial communities under long-term organic contamination and provide a scientific basis for optimizing depth-specific in situ bioremediation strategies, such as targeting aromatic hydrocarbon degradation in surface soils and chlorinated compound remediation in deeper layers. Full article

To identify chitinase genes from the genome of the mycoparasitic Cladosporium sp. strain SYC23, bioinformatical analyses and real-time quantitative PCR (RT-qPCR) were employed to screen mycoparasitism-associated genes at 12, 24, 48, and 72 h post-induction with Aecidium pourthiaea rust spores. A total of eight chitinase genes were identified from SYC23 via bioinformatics analysis and designated CaChi34, CaChi40, CaChi45, CaChi67, CaChi82, CaChi92, CaChi93, and CaChi286 based on sequence and phylogenetic analyses. Analysis of the chitinase protein sequence characteristics revealed molecular weights ranging from 33.86 to 286.03 kDa and theoretical isoelectric points from 4.48 to 7.7. All CaChi genes contained the conserved GH18 domain, and promoter analysis showed that each harbored MYB-binding sites and pathogen-responsive elements. Mycoparasitism-related sequence clustering analysis indicated that the chitinase sequences of SYC23 shared the closest phylogenetic relationship with those from Trichoderma sp. RT-qPCR results following rust spore induction showed that five CaChi genes reached their highest expression levels at 24 h post-induction, CaChi45 was most highly expressed at 72 h post-induction, CaChi93 was continuously upregulated, and CaChi82 was continuously downregulated throughout the induction period. His-tagged recombinant CaChi93 protein was purified from E. coli and characterized. The results demonstrate that the enzymatic activity of CaChi93 was 0.929 U/mg, with optimal reaction conditions at 65 °C and pH 7. Treatment of A. pourthiaea rust spores with the recombinant CaChi93 chitinase confirmed that CaChi93 could effectively dissolve rust spore walls. In conclusion, this study confirms that the mycoparasitic Cladosporium sp. strain SYC23 can secrete chitinase to degrade the rust spore wall and induce spore death, thereby providing novel gene resources and a theoretical basis for the biological control of A. pourthiaea. Full article

Background: Chicken infectious anemia virus (CIAV) is a globally significant immunosuppressive pathogen that causes substantial economic losses to the poultry industry, with particularly severe outbreaks in China in recent years. Given the limitations of existing vaccines, especially the residual virulence associated with live attenuated vaccines, there is an urgent need to develop novel, safer, and more effective vaccine strategies. Methods: In this study, the VP1 and VP2 genes of CIAV were cloned and expressed in Escherichia coli to develop a cost-effective subunit vaccine. Since VP1 primarily formed inclusion bodies, a “VP2-assisted co-refolding” strategy was employed. This involved denaturing VP1 and refolding it via gradient dialysis in the presence of soluble VP2, thereby leveraging VP2’s natural chaperone-like function to restore conformational epitopes. The refolded VP1/VP2 protein complexes, emulsified at different ratios, were used to immunize 3-day-old specific pathogen-free (SPF) chickens, followed by challenge with a virulent CIAV strain. Results: The vaccine formulation with a VP1:VP2 ratio of 1:1 provided the best protection, achieving 71.4% (5/7) protective efficacy, as evidenced by significantly reduced thymic atrophy and a higher thymus index. Conclusions: These findings validate the feasibility of using an economical prokaryotic expression system combined with a rational protein refolding strategy to produce a protective subunit vaccine candidate against CIAV, offering a promising alternative for disease control. Full article

Background: African swine fever virus (ASFV) causes a fatal hemorrhagic disease in domestic pigs and wild boars. While live attenuated vaccines (LAVs) provide protection, their use raises safety concerns. Therefore, the aim of the present study was to identify viral B-cell antigens associated with protection and to test their potential using highly immunogenic vaccine delivery platforms. Methods: We employed a microarray of 169 ASFV proteins expressed in a cell-free prokaryotic system to identify immunodominant antigens using sera from immune pigs. Six structural proteins were selected and formulated into AP205 virus-like particles (VLPs). Additionally, replication-defective vesicular stomatitis virus (VSV)-based vaccine candidates expressing glycosylated CD2v and EP153R proteins were generated. Three groups of specific pathogen-free pigs were immunized with either VLP- or VSV-based vaccines and challenged with the virulent ASFV Georgia 2007 strain. Control groups included pigs immunized with the attenuated ASFV Estonia 2014 strain and a naïve group. Results: Most vaccine candidates induced detectable antibody responses against target ASFV proteins. However, neither VLP- nor VSV-based vaccines provided protection, as clinical scores, hematology, cytokine responses, and viremia levels were similar to those in the negative control group. In contrast, only the ASFV Estonia 2014 strain elicited a robust T-cell response and protective immunity. Conclusions: These findings highlight the challenges in identifying protective B-cell antigens of ASFV and emphasize the pivotal role of cellular immunity in mediating protection. Full article

Lysin motif (LysM) domain-containing proteins are widespread in prokaryotes and eukaryotes, and play crucial roles in microbe-host interactions. In recent decades, a large number of LysM domain-containing proteins have been identified and confirmed to participate in various biological processes, including microbial growth, fungal pathogenesis, and recognition of pathogens by plant immune receptors. Emerging evidence has shown that some LysM domain-containing proteins in plant pathogenic fungi have evolved as key virulence factors. They manipulate host immune responses mainly by interfering with the plant’s perception of chitin, a core pathogen-associated molecular pattern (PAMP) of fungal cell walls. However, the functions of LysM domain-containing proteins in plant pathogenic fungi have not been systematically summarized. In this review, we discuss the latest advances in the structural characteristics, classification, and functional mechanisms of these proteins, as well as their applications in plant disease control. We also propose the current challenges and future research directions in this field. This review aims to deepen the understanding of the molecular mechanisms underlying plant-fungal interactions mediated by LysM domain-containing proteins and provide theoretical references for developing novel and environmentally friendly strategies for fungal disease management in agriculture. Full article