Search Results (168)

For the first time, hydrogels based on silica- and titanium-poly(ethylene glycol) have been used for immobilization of Gram-negative bacteria (Escherichia coli MG1655) and Gram-positive bacteria (Rhodococcus qingshengii X5) in a one-step sol–gel synthesis. Vibrational spectroscopy and thermogravimetric analysis have confirmed the formation of amorphous hybrid structures with a predominance of organic components and metal-oxide grids. Encapsulation efficiencies were 72–77% for Si-PEG-based hydrogel and 50–54% for Ti-PEG. Antimicrobial activity tests revealed that Si-PEG was non-toxic, while Ti-PEG reduced cell viability by 50%. For the first time, an analysis of the morphological properties of immobilized bacterial cells revealed the formation of a thin Si-PEG-based hydrogel shell around each cell and a thick polymer layer on the bacterial surface when encapsulated within Ti-PEG-based hydrogels. The catalytic activity of the biocatalysts, as measured by the ATP content, remained at 84–93% for Si-PEG-based hydrogel, and decreased to 5% for Ti-PEG-based hydrogel. Biocatalysts based on encapsulated bacteria in a Si-PEG-based hydrogel demonstrate high sensitivity and stability. Si-PEG-based hydrogel exhibits high biocompatibility, making it suitable for the effective encapsulation of various bacterial types with a “cell-in-shell” structure. Full article

Arsenic (As) toxicity drives the evolution of resistance mechanisms in environmental microorganisms. Bacteria of the Bacillus genus are frequently identified in isolates from arsenic-contaminated sites, highlighting the importance of understanding the molecular mechanisms related to this bacterial genus. Bacillus subtilis, a soil microorganism and Gram-positive model paradigm, employs multiple strategies to counteract As toxicity, including biosorption, redox transformation, active efflux, and inducible genetic regulation. This review provides a comprehensive analysis of the physiological and molecular mechanisms involved in arsenic response in B. subtilis and related species, focusing on the ars and ase operons. The ars operon, located within the mobile SKIN element, encodes a reductase (ArsC), an Acr3-type efflux pump (ArsB), a carbon–arsenic lyase (ArsI/YqcK), and a transcriptional repressor (ArsR), all co-regulated in response to arsenic. In turn, the ase operon contributes to resistance via an ArsB-type efflux system (AseA) and its own regulatory protein (AseR) but lacks an arsenate reductase. Additionally, genes such as aioAB, arrAB, and arsD are discussed, along with evidence for extracellular detoxification and cell surface immobilization of As. Studies on environmental Bacillus species are examined, pointing out the evolutionary implications of As resistance and the biotechnological potential for remediation of contaminated sites. Full article

Growing consumer awareness of the link between diet and health has increased interest in functional foods, including fermented juices. Grapefruit juice has potential health-promoting properties, but its bitter taste limits its acceptance by consumers. This study aimed to develop a fermentation process for debittering grapefruit juice at natural pH using Lacticaseibacillus rhamnosus and naringinase. Grapefruit juice was fermented with Lactic. rhamnosus using free naringinase and naringinase immobilized on carob gum and chitosan supports at 30 ± 0.2 °C for 72 h. Naringin concentration, bacterial cell count, total phenol content, organic acids, carbohydrates, antioxidant activity, and pH were analyzed. Naringinase immobilized on carob gum demonstrated the highest efficiency, hydrolyzing over 42% of naringin after 24 h (from 418.20 to 241.19 μg/mL). The free enzyme reduced the naringin concentration to 155.28 μg/mL after 48 h. The highest Lactic. rhamnosus cell count (2.05 × 10⁹ CFU/mL) was achieved with the free enzyme. Total phenol content decreased from 42.24 to 16.58 mg GAE/100 mL when using naringinase immobilized on chitosan. The combined use of naringinase and Lactic. rhamnosus enables the development of an integrated process that improves consumer acceptance with potential applications in the functional beverage industry. Full article

Bacterial inclusion bodies (IBs) are still generally considered to be waste products of recombinant protein production, despite various studies that have challenged this conventional view in the last two decades, and have been proposed for use as immobilized enzymes in vivo for biocatalysis. Current advances in genetic and molecular biology make it possible to perform multienzymatic reactions or enzymatic cascades to synthesize valuable products. When cascades need cofactor regener tion, it is difficult to use “cheap” whole cells or their lysates, and “expensive” enzyme purification is required. The capture of enzymatic activity into active IBs (aIBs), well-separable protein aggregates from cell lysate, could represent a usable compromise between purified enzymes and cell lysates. It is shown here that the combination of two polyphosphate kinases (PPKs) in the form of aIBs leads to almost 10-fold ATP regeneration and 100% UTP utilization without degradation into adenosine or uridine. PPKs have been combined with N-acetylhexosamine 1-kinase and N-acetylglucosamine-1-phosphate uridyltransferase to produce valuable UDP-N-acetylglucosamine, but the described approach could be used in various multienzymatic syntheses to avoid enzyme purification and ensure nucleotide triphosphate regeneration. Full article

Nanoporous gold nanoparticles (np-AuNPs) combine inertness, a nanoscale structure, and a porous framework with high surface area, conductivity, and biocompatibility, making them ideal for biosensing, catalysis, fuel cells, and drug delivery. Their open pore structure and low-coordinated atoms enhance biomolecule capture and mass transfer, while their tunable size, pore volume, and ease of surface modification make them promising biosensor transducers. However, synthesizing colloidal np-AuNPs in a simple way with controllable size and scalability remains challenging. The existing approaches mostly rely on specialized equipment, complex setups, and expert knowledge, while still facing challenges in terms of scalability. In this study, we present a simple, seedless, wet-chemical synthesis of colloidal np-AuNPs via the co-reduction of Au/Ag alloys followed by dealloying. By adjusting the Au:Ag ratio, we produced np-AuNPs sized ~120–530 nm, which were immobilized on electrodes for detecting lipopolysaccharide (LPS), a toxic component of Gram-negative bacterial membranes. The LPS biosensor exhibited excellent sensitivity towards detecting wild-type LPS, with a low limit of detection (LOD) of 0.1244 ng/L. This work demonstrates the effective synthesis and application of np-AuNPs in LPS biosensing. Full article

Pseudomonas aeruginosa is a clinically significant pathogen with high antibiotic resistance, necessitating rapid and reliable diagnostic methods. In this study, we developed a whole-cell aptamer selection method for P. aeruginosa using an Eppendorf-tube-based SELEX system, where bacterial cells were directly incubated with an ssDNA library. This configuration enhanced the recovery of bound aptamers and overcame the cell quantity limitations often encountered in microtiter-plate-based SELEX. After 10 selection rounds, six aptamer candidates were obtained and evaluated for affinity. Molecular docking analysis revealed that aptamer T1 possessed the highest target selectivity. To demonstrate diagnostic applicability, aptamer T1 was integrated into a hybrid lateral flow immunoassay (LFIA), replacing the conventional detection antibody. In this format, the AuNP–aptamer complex bound to the target bacteria and was captured by a specific antibody immobilized on the test line. The LFIA achieved a visual detection limit of 2.34 × 10² CFU/mL within 15 min, showing high specificity and suitability for point-of-care applications. This study presents the first demonstration of an aptamer–antibody hybrid LFIA for bacterial detection and highlights the potential of aptamers as low-cost, rapidly synthesized recognition elements adaptable for the detection of other infectious agents. Full article

Soil heavy-metal pollution is one of the most serious environmental issues in the world. There is an urgent need to develop feasible strategies for the remediation of polluted soil. Biochar has great potential to reduce heavy metal phytotoxicity and promote plant growth, but its mechanisms are still unclear. In this study, phosphoric acid and magnesium composite-modified tea branch biochar (PMB) was prepared and characterized. The effects of PMB at 5% addition on pakchoi growth, Cd/Pb accumulation and subcellular distribution in pakchoi, soil physicochemical characteristics and enzyme activities, Cd/Pb bioavailability, bacterial community structure, and diversity in Cd/Pb co-contaminated soils was investigated by a pot experiment. The results showed that PMB significantly alleviated the phytotoxicity of Cd and Pb. The application of PMB effectively increased the plant height and biomass and Cd and Pb proportion in the cell wall, while reducing Cd and Pb accumulation and their distribution in cytoplasm and organelles in pakchoi plants. PMB significantly improved the activities of urease, invertase, and catalase and reduced the available Cd and Pb contents in soil. Moreover, PMB changed the structure and diversity of the soil bacterial community. The relative abundance of several beneficial microbial phyla, including Acidobacteriota, Bacteroidota, Actinobacteriota, and Gemmatimonadota, increased by 13.81%, 19.02%, 68.09%, and 34.79%, respectively. The Shannon and Chao1 index also increased significantly. This study provides an effective strategy for simultaneous Cd and Pb immobilization in soil, promoting plant growth and inhibiting heavy metal accumulation in vegetables, which highlights the application of PMB in sustainable agro-ecosystems. Full article

Implant-associated infections (IAIs) are major complications in dental and orthopedic implants, potentially compromising osseointegration and eventually causing implant loosening or removal. Thus, early prevention of bacterial adhesion and biofilm formation is critical for successful long-term osseointegration. Polyetheretherketone (PEEK) exhibits excellent physicochemical properties and an elastic modulus similar to bone tissue, making it a promising material for dental and orthopedic implants. However, its inherent lack of antibacterial properties limits its ability to prevent IAIs. Herein, an antibacterial coating with controlled drug release and excellent biocompatibility is designed by immobilizing minocycline (Mino)-doped carboxymethyl chitosan (CMCS) onto the PEEK surface via a polydopamine (PDA)-mediated Michael addition and Schiff base reaction. The coating is characterized by SEM, XPS, water contact angle measurements, and in vitro Mino release assays. Antibacterial activity is evaluated using the zone of inhibition (ZOI), turbidity, and colony counting assays, while biocompatibility is assessed through a SEM analysis of cell morphology and CCK-8 assay. The results show that the Mino-modified coating is successfully fabricated on the PEEK surface, achieving sustained Mino release for up to 14 days. Among the three Mino concentrations, the PEEK-0.5Mino group demonstrates the best balance of antibacterial activity and biocompatibility, highlighting its potential for preventing IAIs in orthopedic and dental applications. Full article

Background/Objectives: Extracellular vesicles (EVs) are nanoscale, membrane-enclosed structures that play key roles in intercellular communication and biological regulation. Among them, Lactobacillus paracasei-derived EVs (Lp-EVs) have attracted attention for their anti-inflammatory and anti-aging properties, making them promising candidates for therapeutic and cosmetic use. However, methods for specific detection and quantitative evaluation of Lp-EVs are still limited. This study aims to develop a surface plasmon resonance (SPR)-based sensor system for the precise and selective detection of Lp-EVs. Methods: Anti-p40 antibodies were immobilized on gold thin films to construct an SPR sensing platform. The overexpression of the p40 protein on Lp-EVs was confirmed using flow cytometry and Western blotting. For functional evaluation, Lp-EVs were applied to an artificial skin membrane mounted on a Franz diffusion cell, followed by SPR-based quantification and fluorescence imaging to assess their skin penetration behavior. Results: The developed SPR sensor demonstrated high specificity and a detection limit of 0.12 µg/mL, with a linear response range from 0.1 to 0.375 µg/mL. It successfully discriminated Lp-EVs from other bacterial EVs. In the skin diffusion assay, Lp-EVs accumulated predominantly in the epidermal layer without penetrating into the dermis, likely due to their negative surface charge and interaction with the hydrophobic epidermal lipid matrix. Fluorescence imaging confirmed this epidermal confinement, which increased over 24 h. Conclusions: This study presents a sensitive and selective SPR-based platform for detecting Lp-EVs and demonstrates their potential for targeted epidermal delivery. These findings support the use of Lp-EVs in skin-focused therapeutic and cosmetic applications. Future studies will explore strategies such as microneedle-assisted delivery to enhance transdermal penetration and efficacy. Full article

Aerobic denitrifying microorganisms, with their strong environmental adaptability, low dissolved oxygen concentration requirements, rapid growth rate, and high nitrogen removal efficiency, significantly compensate for the shortcomings of traditional aerobic chemolithoautotrophic nitrification and anaerobic heterotrophic denitrification models. The introduction of aerobic denitrifiers can effectively enhance the removal of nitrate nitrogen. However, directly inoculating aerobic denitrifiers into wastewater leads to issues such as easy loss of bacterial cells and difficulty in forming a dominant flora, thus preventing the long-term maintenance of their enhancing effect on denitrification performance. To address this problem, microbial immobilization technology has been introduced into the remediation process of nitrogen-polluted water bodies. This technology can maintain a high biomass concentration, provide a stable breeding ground for microorganisms, and effectively prevent the rapid loss of microorganisms. This article systematically reviews the current status of the isolation of aerobic denitrifying bacteria, key enzymes, and genes, as well as the application progress of aerobic denitrifying bacteria and their immobilization technology, aiming to provide solid theoretical support for the practical application of aerobic denitrification technology and promote its further development in the field of nitrogen pollution control. Full article

This study presents a proof-of-concept evaluation of optimized whole-cell biosensors designed for the real-time detection of crop infections. Genetically engineered luminescent bacterial strains were used to detect volatile organic compounds (VOCs) emitted by crops during spoilage. Key factors investigated include bacterial uniformity, nutrient supply, and temperature effects. The results demonstrated that lower temperatures (+4 °C) yielded higher sensor sensitivity and prolonged bacterial viability. A proof-of-concept evaluation was conducted in storage-like conditions, showing effective infection detection in potatoes. These findings underscore the potential of whole-cell-based biosensors for monitoring postharvest production in cold storage environments. Full article

It is known that the ciliate Paramecium cell surface including cilia is completely covered by high-molecular-mass GPI-anchored proteins named surface antigens (SAgs). However, their functions are not well understood. It was found that ciliate Paramecium caudatum reversibly changes its SAgs depending on the absence or presence of the endonuclear symbiotic bacterium Holospora obtusa in the macronucleus. Immunofluorescence microscopy with a monoclonal antibody produced SAg of the H. obtusa-free P. caudatum strain RB-1-labeled cell surface of the H. obtusa-free P. caudatum RB-1 cell but not the H. obtusa-bearing RB-1 cell. When this antibody was added to the living P. caudatum RB-1 cells, only H. obtusa-free cells were immobilized. An immunoblot with SAgs extracted from Paramecium via cold salt/ethanol treatment showed approximately 266-kDa SAgs in the extract from H. obtusa-free cells and 188 and 149-kDa SAgs in the extract from H. obtusa-bearing cells. H. obtusa-free RB-1 cells produced from H. obtusa-bearing cells via treatment with penicillin-G-potassium re-expressed 266-kDa SAg, while the 188 and 149-kDa SAgs disappeared. This phenotypic change in the SAgs was not induced by degrees of starvation or temperature shifts. These results definitively show that Paramecium SAgs have functions related to bacterial infection. Full article

This study introduces a one-pot colorimetric nucleic acid test (NAT) platform that integrates silver nanoparticle (AgNP)-based DNA isolation and colorimetric detection of bacterial genes. The NAT platform is comprised with purification and reaction units that enable cell lysis, DNA purification, loop-mediated isothermal amplification (LAMP), and colorimetric detection. In the purification unit, polyethyleneimine (PEI)-capped AgNPs were used as cell lysis agents because of their cell-disrupting and antimicrobial properties and were immobilized on a glass fiber membrane for DNA capture and isolation. The reaction unit enabled colorimetric detection of DNA amplicons, achieved by the synthesis of AgNPs on chromatography paper formed via the reduction of silver ions present on the paper, mediated by the use of sodium ascorbate, a reducing agent, present in the LAMP reagent, after the reaction. AgNPs were formed only in the presence of the target amplicons in the positive samples after reaction at 65 °C for 5 min. Bacterial DNA was efficiently extracted using this method, and Enterococcus faecium was detected with a detection limit of 10² CFU/mL. This platform is a promising alternative for rapid and cost-effective nucleic acid testing in resource-limited settings. Full article

Microalgal biotechnology is gaining attention due to its potential to produce pigments, lipids, biofuels, and value-added products. However, challenges persist in terms of the economic viability of microalgal lipid production in photobioreactors due to slow growth rates, expensive media, complex downstream processing, limited product yields, and contamination risks. Recent studies suggest that co-cultivating microalgae with bacteria can enhance the profitability of microalgal bioprocesses. Immobilizing bacteria offers advantages such as protection against shear forces, the prevention of overgrowth, and continuous product secretion. Previous work has shown that biopolymeric immobilization of Paenibacillus polymyxa enhances 2,3-butanediol production. In this study, a novel co-fermentation process was developed by exploiting the chemical crosstalk between a freshwater microalga Scenedesmus obliquus, also known as Tetradesmus obliquus, and an immobilized plant-growth-promoting bacterium, Paenibacillus polymyxa. This co-cultivation resulted in increased metabolite production, with a 1.5-fold increase in the bacterial 2,3-butanediol concentration and a 3-fold increase in the microalgal growth rates compared to these values in free-cell co-cultivation. Moreover, the co-culture with the immobilized bacterium exhibited a 5-fold increase in the photosynthetic pigments and a 3-fold increase in the microalgal lipid concentration compared to these values in free-cell co-cultivation. A fixed bed photobioreactor was further constructed, and the co-cultivation bioprocess was implemented to improve the bacterial 2,3-butanediol and microalgal lipid production. In conclusion, this study provides conclusive evidence for the potential of co-cultivation and biopolymeric immobilization techniques to enhance 2,3-butanediol and lipid production. Full article

The hydrothermal carbonization (HTC) of biomass presents a sustainable approach for waste management and production of value-added materials such as hydrochar, which holds promise as an adsorbent and support matrix for bacterial immobilization applied, e.g., for bioremediation processes of sites contaminated with phthalate ester plasticizers such as diethyl phthalate (DEP). In the present study, hydrochar was synthesized from vine shoots (VSs) biomass employing the following parameters during the HTC process: 260 °C for 30 min with a 1:10 (w/v) biomass-to-water ratio. The resulting vine shoots hydrochar (VSs-HC) was characterized for porosity, elemental composition, and structural properties using Scanning Electron Microscopy (SEM), Energy-Dispersive X-ray Spectroscopy (EDX), and Raman spectroscopy. Elemental analysis confirmed the presence of key elements in the VSs structure, elements essential for char formation during the HTC process. The VSs-HC exhibited a macroporous structure (>0.5 μm), facilitating diethyl phthalate (DEP) adsorption, bacterial adhesion, and biofilm formation. Adsorption studies showed that the VSs-HC achieved a 90% removal rate for 4 mM DEP within the first hour of contact. Furthermore, VS-HC was tested as a support matrix for a bacterial consortium (Pseudomonas spp. and Microbacterium sp.) known to degrade DEP. The immobilized bacterial consortium on VSs-HC demonstrated enhanced tolerance to DEP toxicity, degrading 76% of 8 mM DEP within 24 h, compared with 14% by planktonic cultures. This study highlights VSs-HC’s potential as a sustainable and cost-effective material for environmental bioremediation, offering enhanced bacterial cell viability, improved biofilm formation, and efficient plasticizer removal. These findings provide a pathway for mitigating environmental pollution through scalable and low-cost solutions. Full article