Zhifeng Fu

It is highly demanded to establish effective and sensitive method for detecting pathogenic bacteria because of their high morbidity. The conventional molecular recognition-based methods rely on heterogeneous mode, for which the passive diffusion of bacterial cells and signal tracer results in low transport efficiency and unsatisfied sensitivity. Herein, Fe3O4/polydopamine Janus micromotors were prepared and conjugated with bacteriophage acting as capture agent for bacteria and urease acting as propulsion fuel for Janus micromotors. With a bubble-driven approach, this strategy significantly accelerated transport and improved enrichment of bacterial cells onto the Janus micromotors, achieving a capture efficiency over 98%. By using MoS2/Pt conjugated with polymyxin B as the signal tracer, sensitive colorimetry of Pseudomonas aeruginosa was achieved with a dynamic range of 1.0 × 102 to 1.0 × 107 cfu mL–1. The whole procedure including incubation, washing, color development, and signal detection can be accomplished within 17 min. Usage of bacteriophage as capture agent led to satisfied selectivity for the target bacteria even in complex matrixes. The results for detecting Pseudomonas aeruginosa in artificial cerebrospinal fluid and contaminated milk demonstrated its reliability for real application. This work provides an ideal technical tool for dynamic enrichment and sensitive detection of pathogens based on functional self-propulsed micro/nanomotors.

A pH-responsive smart hydrogel dressing was developed for controlled antimicrobial therapy and accelerated wound healing.

Pseudomonas aeruginosa (P. aeruginosa) has long been one of the leading opportunistic pathogens responsible for serious healthcare-associated infective diseases. Development of rapid and accurate detection techniques for this pathogen is highly desired to achieve efficient prevention and control. This study developed a fluorescent detection technique targeting this pathogen by using recombinant phage tail fiber protein Gp47 as the recognition element. This protein was identified from a phage PAB1 isolated from hospital wastewater through genomic sequencing and bioinformatic analysis. It was then produced in its functional form via recombinant expression and purification. Gp47 exhibited broad-spectrum binding capability toward P. aeruginosa strains, recognizing all 30 clinical isolates tested, while completely lacking lytic activity. Thus, it effectively overcame the key limitations of the whole phage, including the narrow host range and tendency to lyse target bacteria. A sandwich method was constructed to detect P. aeruginosa by using Gp47 conjugated magnetic beads as capture carriers and fluorescein isothiocyanate labeled antimicrobial peptide as signal reporter. The method responded to P. aeruginosa linearly from 3.2 × 10² to 3.2 × 10⁶ CFU/mL, with a limit of detection of 109 CFU/mL and ideal specificity. It has been employed for detecting P. aeruginosa in complex matrices such as physiological saline, human urine, and milk powder with acceptable recoveries ranging from 71.23% to 110.87%. This work provides a high-performance recognition element for P. aeruginosa and helps develop valuable diagnostic tools for infective diseases.

Cryptococcus neoformans (C. neoformans) infections pose a critical threat to global health, with high mortality rates observed among immunocompromised individuals. Rapid, cost-effective diagnostics are essential for preventive treatment and reducing mortality in resource-scarce settings, yet the development of selective fluorescent probes for C. neoformans remains challenging. The Prp8 intein, a unique self-splicing protein in C. neoformans, represents an ideal target for developing fluorescent probes and antifungal drugs. In this study, a series of fluorescent probes integrating Prp8 intein inhibitors with an environment-sensitive fluorophore was developed for selective covalent binding to the Prp8 intein. Among these probes, H2 enables specific detection of C. neoformans with minimal interference from Candida species. A dual-staining approach combining capsule staining with intracellular Prp8 intein labeling significantly improves the accuracy of C. neoformans detection. Probe H2 was also successfully applied for fluorescence imaging of fungal infections in a Galleria mellonella model, demonstrating potential for assessing fungal infections in vivo. Furthermore, the fluorescence intensity of the probe exhibited a dose-dependent relationship with Prp8 inhibitor activity, enabling rapid screening of intein-splicing inhibitors within 2 h, thereby improving efficiency and reducing assay time.

Owing to target recognition hindrance, common metal-organic frameworks (MOFs) as nanocarriers lead to the dramatically decreased catalytic activity (by 90%) of immobilized enzymes. Herein, a dopamine-modulated enclosure strategy was proposed to successfully immobilize alkaline phosphatase (ALP) on phosphatase-like Ce-2-methylimidazole (2-MIm) MOFs for the formation of a Ce-2-MIm@ALP@polydopamine (PDA) biocomposite. Although the activity proportion (1.22%) of Ce-2-MIm to Ce-2-MIm@ALP@PDA was negligible, Ce-2-MIm MOFs facilitated 2.5-fold higher catalytic activity of immobilized ALP than free ALP. Note that shortened Ce-Ce bonds and increased Ce-OH groups were verified by X-ray absorption spectroscopy for the increased active Ce sites on the Ce-2-MIm support. Thereby, the phosphatase-like Ce-2-MIm possessed enhanced substrate affinity (K_m = 86.79 μM) 16.5 times superior to ordinary Ce-based nanozymes (K_m = 1428.57 μM). The experimental data and density functional theory calculations demonstrated that the flexible scaffold coating of PDA endowed Ce-2-MIm with improved oxygen vacancies for the preferable binding affinity and hydrolysis activity. The marked affinity effect of the Ce-2-MIm-based support imparted the dominant improvement of catalytic activity to fixed enzymes in the enzyme immobilization. For the proof-of-concept application on chemiluminescence (CL) catalysis, the prepared Ce-2-MIm@ALP@PDA is successfully employed for the detection of carbendazol (CAB), with a low detection limit of 56 pg/mL. This work highlights the merits of nanozymes as an efficient affinity-type support for enzyme assemblies, which exhibits great potential for considerable activity exaltation of enzymes even in an immobilization manner.

Multimodal probes can output multidimensional signals and meet the demand for discriminating highly similar enantiomers. Their diverse signal transducing mechanisms broaden the discrimination scope of enantiomers and ensure adaptability in various application scenarios. Herein, an achiral multimodal probe termed gold-cobalt nanoparticles (AuCoNPs) was synthesized, which showed outstanding enantioselectivity, Fenton-like activity, and localized surface plasmon resonance effects. It promoted the chemiluminescent (CL) signal of the luminol system and exhibited color and temperature (TP) response characteristics. The assay platform based on a multimodal probe can discriminate 11 kinds of enantiomers, showing an enantioselectivity of 4.29 for glutamic acid (Glu). For the CL mode, the stronger binding affinity of l-Glu to AuCoNPs reduced its adsorption onto the poly(vinyl alcohol)/sodium alginate hydrogel, resulting in a weaker CL signal compared to d-Glu. For the RGB mode, the aggregation state of AuCoNPs was affected by the affinity of Glu enantiomers to the probe, creating a distinguishable color gradient. This enables rapid visual discrimination for Glu enantiomers using smartphone RGB imaging within 3 min. For the TP mode, the aggregation state affected the photothermal conversion, producing different TP changes induced by the enantiomers. Density functional theory calculations, contact angle measurements, and zeta potential measurements were used to explore the discrimination mechanism. The multimodal probe shows promising practicality for chiral discrimination due to the combinational usage of multidimensional signals.

Traditional methods for detecting bacteria are usually limited by low efficiency, high cost, and resource dependency. This study developed a recyclable affinity chromatographic device using phage-derived cell wall binding domains (CBDs) as recognition ligands for point-of-care testing of bacteria. The device trapped nickel-crosslinked resin (NR) beads within two layers of PTFE sieve, enabling efficient conjugation of His-tagged CBD. With a horseradish peroxidase (HRP)-based colorimetry, Acinetobacter baumannii (A. baumannii) was detected by using CBD18 and polymyxin B-HRP conjugate as the recognition ligand and signal tracer, respectively. With the same protocol, CBD108 and vancomycin-HRP conjugates were used for the colorimetry of methicillin-resistant Staphylococcus aureus (MRSA). In this process, the affinity chromatographic device was eluted using an imidazole solution, enabling its recycling use. With the device, A. baumannii and MRSA were quantified within 30 min, with linear ranges of 2.4 × 10²-2.4 × 10⁸ CFU/mL and 2.1 × 10²-2.1 × 10⁸ CFU/mL, respectively. It exhibited high selectivity for the target bacteria due to the combinational usage of CBDs and antibiotics. The NR beads trapped in the device maintained a conjugation performance of no less than 93.6% after 20 run cycles. This study provides a rapid, sensitive, and recyclable technical tool for point-of-care testing of bacteria in resource-limited countries and institutes.