Dual-ROS-scavenging and dual-lingering nanozyme-based eye drops alleviate dry eye disease.

Efficiently removing excess reactive oxygen species (ROS) generated by various factors on the ocular surface is a promising strategy for preventing the development of dry eye disease (DED). The currently available eye drops for DED treatment are palliative, short-lived and frequently administered due to the short precorneal residence time. Here, we developed nanozyme-based eye drops for DED by exploiting borate-mediated dynamic covalent complexation between n-FeZIF-8 nanozymes (n-Z(Fe)) and poly(vinyl alcohol) (PVA) to overcome these problems. The resultant formulation (PBnZ), which has dual-ROS scavenging abilities and prolonged corneal retention can effectively reduce oxidative stress, thereby providing an excellent preventive effect to alleviate DED. In vitro and in vivo experiments revealed that PBnZ could eliminate excess ROS through both its multienzyme-like activity and the ROS-scavenging activity of borate bonds. The positively charged nanozyme-based eye drops displayed a longer precorneal residence time due to physical adhesion and the dynamic borate bonds between phenyboronic acid and PVA or o-diol with mucin. The in vivo results showed that eye drops could effectively alleviate DED. These dual-function PBnZ nanozyme-based eye drops can provide insights into the development of novel treatment strategies for DED and other ROS-mediated inflammatory diseases and a rationale for the application of nanomaterials in clinical settings.

Construction of lymph nodes-targeting tumor vaccines by using the principle of DNA base complementary pairing to enhance anti-tumor cellular immune response.

Tumor vaccines, a crucial immunotherapy, have gained growing interest because of their unique capability to initiate precise anti-tumor immune responses and establish enduring immune memory. Injected tumor vaccines passively diffuse to the adjacent draining lymph nodes, where the residing antigen-presenting cells capture and present tumor antigens to T cells. This process represents the initial phase of the immune response to the tumor vaccines and constitutes a pivotal determinant of their effectiveness. Nevertheless, the granularity paradox, arising from the different requirements between the passive targeting delivery of tumor vaccines to lymph nodes and the uptake by antigen-presenting cells, diminishes the efficacy of lymph node-targeting tumor vaccines. This study addressed this challenge by employing a vaccine formulation with a tunable, controlled particle size. Manganese dioxide (MnO2) nanoparticles were synthesized, loaded with ovalbumin (OVA), and modified with A50 or T20 DNA single strands to obtain MnO2/OVA/A50 and MnO2/OVA/T20, respectively. Administering the vaccines sequentially, upon reaching the lymph nodes, the two vaccines converge and simultaneously aggregate into MnO2/OVA/A50-T20 particles through base pairing. This process enhances both vaccine uptake and antigen delivery. In vitro and in vivo studies demonstrated that, the combined vaccine, comprising MnO2/OVA/A50 and MnO2/OVA/T20, exhibited robust immunization effects and remarkable anti-tumor efficacy in the melanoma animal models. The strategy of controlling tumor vaccine size and consequently improving tumor antigen presentation efficiency and vaccine efficacy via the DNA base-pairing principle, provides novel concepts for the development of efficient tumor vaccines.

Human T-cell activation with Toxoplasma gondii antigens loaded in maltodextrin nanoparticles.

Toxoplasmosis is the most prevalent parasitic zoonosis worldwide, causing ocular and neurological diseases. No vaccine has been approved for human use. We evaluated the response of peripheral blood mononuclear cells (PBMCs) to a novel construct of Toxoplasma gondii total antigen in maltodextrin nanoparticles (NP/TE) in individuals with varying infectious statuses (uninfected, chronic asymptomatic, or ocular toxoplasmosis). We analyzed the concentration of IFN-γ after NP/TE ex vivo stimulation using ELISA and the immunophenotypes of CD4+ and CD8+ cell populations using flow cytometry. In addition, serotyping of individuals with toxoplasmosis was performed by ELISA using GRA6-derived polypeptides. Low doses of NP/TE stimulation (0.9 µg NP/0.3 µg TE) achieved IFN-γ-specific production in previously exposed human PBMCs without significant differences in the infecting serotype. Increased IFN-γ expression in CD4+ effector memory cell subsets was found in patients with ocular toxoplasmosis with NP/TE but not with TE alone. This is the first study to show how T-cell subsets respond to ex vivo stimulation with a vaccine candidate for human toxoplasmosis, providing crucial insights for future clinical trials.

Heavy metals immobilization and bioavailability in multi-metal contaminated soil under ryegrass cultivation as affected by ZnO and MnO2 nanoparticle-modified biochar.

Pollution by heavy metals (HMs) has become a global problem for agriculture and the environment. In this study, the effects of pristine biochar and biochar modified with manganese dioxide (BC@MnO2) and zinc oxide (BC@ZnO) nanoparticles on the immobilization and bioavailability of Pb, Cd, Zn, and Ni in soil under ryegrass (Lolium perenne L.) cultivation were investigated. The results of SEM-EDX, FTIR, and XRD showed that ZnO and MnO2 nanoparticles were successfully loaded onto biochar. The results showed that BC, BC@MnO2 and BC@ZnO treatments significantly increased shoots and roots dry weight of ryegrass compared to the control. The maximum dry weight of root and shoot (1.365 g pot-1 and 4.163 g pot-1, respectively) was reached at 1% BC@MnO2. The HMs uptake by ryegrass roots and shoots decreased significantly after addition of amendments. The lowest Pb, Cd, Zn and Ni uptake in the plant shoot (13.176, 24.92, 32.407, and 53.88 µg pot-1, respectively) was obtained in the 1% BC@MnO2 treatment. Modified biochar was more successful in reducing HMs uptake by ryegrass and improving plant growth than pristine biochar and can therefore be used as an efficient and cost effective amendment for the remediation of HMs contaminated soils. The lowest HMs translocation (TF) and bioconcentration factors were related to the 1% BC@MnO2 treatment. Therefore, BC@MnO2 was the most successful treatment for HMs immobilization in soil. Also, a comparison of the TF values of plant showed that ryegrass had a good ability to accumulate all studied HMs in its roots, and it is a suitable plant for HMs phytostabilization.

Radiotherapy-sensitized cancer immunotherapy via cGAS-STING immune pathway by activatable nanocascade reaction.

Radiotherapy-induced immune activation holds great promise for optimizing cancer treatment efficacy. Here, we describe a clinically used radiosensitizer hafnium oxide (HfO2) that was core coated with a MnO2 shell followed by a glucose oxidase (GOx) doping nanoplatform (HfO2@MnO2@GOx, HMG) to trigger ferroptosis adjuvant effects by glutathione depletion and reactive oxygen species production. This ferroptosis cascade potentiation further sensitized radiotherapy by enhancing DNA damage in 4T1 breast cancer tumor cells. The combination of HMG nanoparticles and radiotherapy effectively activated the damaged DNA and Mn2+-mediated cGAS-STING immune pathway in vitro and in vivo. This process had significant inhibitory effects on cancer progression and initiating an anticancer systemic immune response to prevent distant tumor recurrence and achieve long-lasting tumor suppression of both primary and distant tumors. Furthermore, the as-prepared HMG nanoparticles "turned on" spectral computed tomography (CT)/magnetic resonance dual-modality imaging signals, and demonstrated favorable contrast enhancement capabilities activated by under the GSH tumor microenvironment. This result highlighted the potential of nanoparticles as a theranostic nanoplatform for achieving molecular imaging guided tumor radiotherapy sensitization induced by synergistic immunotherapy.

Comprehensive dry eye therapy: overcoming ocular surface barrier and combating inflammation, oxidation, and mitochondrial damage.

BACKGROUND: Dry Eye Disease (DED) is a prevalent multifactorial ocular disease characterized by a vicious cycle of inflammation, oxidative stress, and mitochondrial dysfunction on the ocular surface, all of which lead to DED deterioration and impair the patients' quality of life and social functioning. Currently, anti-inflammatory drugs have shown promising efficacy in treating DED; however, such drugs are associated with side effects. The bioavailability of ocular drugs is less than 5% owing to factors such as rapid tear turnover and the presence of the corneal barrier. This calls for investigations to overcome these challenges associated with ocular drug administration. RESULTS: A novel hierarchical action liposome nanosystem (PHP-DPS@INS) was developed in this study. In terms of delivery, PHP-DPS@INS nanoparticles (NPs) overcame the ocular surface transport barrier by adopting the strategy of "ocular surface electrostatic adhesion-lysosomal site-directed escape". In terms of therapy, PHP-DPS@INS achieved mitochondrial targeting and antioxidant effects through SS-31 peptide, and exerted an anti-inflammatory effect by loading insulin to reduce mitochondrial inflammatory metabolites. Ultimately, the synergistic action of "anti-inflammation-antioxidation-mitochondrial function restoration" breaks the vicious cycle associated with DED. The PHP-DPS@INS demonstrated remarkable cellular uptake, lysosomal escape, and mitochondrial targeting in vitro. Targeted metabolomics analysis revealed that PHP-DPS@INS effectively normalized the elevated level of mitochondrial proinflammatory metabolite fumarate in an in vitro hypertonic model of DED, thereby reducing the levels of key inflammatory factors (IL-1ß, IL-6, and TNF-α). Additionally, PHP-DPS@INS strongly inhibited reactive oxygen species (ROS) production and facilitated mitochondrial structural repair. In vivo, the PHP-DPS@INS treatment significantly enhanced the adhesion duration and corneal permeability of the ocular surface in DED mice, thereby improving insulin bioavailability. It also restored tear secretion, suppressed ocular surface damage, and reduced inflammation in DED mice. Moreover, it demonstrated favorable safety profiles both in vitro and in vivo. CONCLUSION: In summary, this study successfully developed a comprehensive DED management nanosystem that overcame the ocular surface transmission barrier and disrupted the vicious cycle that lead to dry eye pathogenesis. Additionally, it pioneered the regulation of mitochondrial metabolites as an anti-inflammatory treatment for ocular conditions, presenting a safe, efficient, and innovative therapeutic strategy for DED and other inflammatory diseases.

In vitro assessment of the effect of magnetic fields on efficacy of biosynthesized selenium nanoparticles by Alborzia kermanshahica.

Cyanobacteria represent a rich resource of a wide array of unique bioactive compounds that are proving to be potent sources of anticancer drugs. Selenium nanoparticles (SeNPs) have shown an increasing potential as major therapeutic platforms and led to the production of higher levels of ROS that can present desirable anticancer properties. Chitosan-SeNPs have also presented antitumor properties against hepatic cancer cell lines, especially the Cht-NP (Chitosan-NPs), promoting ROS generation and mitochondria dysfunction. It is proposed that magnetic fields can add new dimensions to nanoparticle applications. Hence, in this study, the biosynthesis of SeNPs using Alborzia kermanshahica and chitosan (CS) as stabilizers has been developed. The SeNPs synthesis was performed at different cyanobacterial cultivation conditions, including control (without magnetic field) and magnetic fields of 30 mT and 60 mT. The SeNPs were characterized by uv-visible spectroscopy, Fourier-transform infrared spectroscopy (FT-IR), Dynamic light scattering (DLS), zeta potential, and TEM. In addition, the antibacterial activity, inhibition of bacterial growth, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC), as well as the antifungal activity and cytotoxicity of SeNPs, were performed. The results of uv-visible spectrometry, DLS, and zeta potential showed that 60 mT had the highest value regarding the adsorption, size, and stabilization in compared to the control. FTIR spectroscopy results showed consistent spectra, but the increased intensity of peaks indicates an increase in bond number after exposure to 30 mT and 60 mT. The results of the antibacterial activity and the inhibition zone diameter of synthesized nanoparticles showed that Staphylococcus aureus was more sensitive to nanoparticles produced under 60 mT. Se-NPs produced by Alborzia kermanshahica cultured under a 60 mT magnetic field exhibit potent antimicrobial and anticancer properties, making them a promising natural agent for use in the pharmaceutical and biomedical industries.

Cucurbit[8]uril-based supramolecular theranostics.

Different from most of the conventional platforms with dissatisfactory theranostic capabilities, supramolecular nanotheranostic systems have unparalleled advantages via the artful combination of supramolecular chemistry and nanotechnology. Benefiting from the tunable stimuli-responsiveness and compatible hierarchical organization, host-guest interactions have developed into the most popular mainstay for constructing supramolecular nanoplatforms. Characterized by the strong and diverse complexation property, cucurbit[8]uril (CB[8]) shows great potential as important building blocks for supramolecular theranostic systems. In this review, we summarize the recent progress of CB[8]-based supramolecular theranostics regarding the design, manufacture and theranostic mechanism. Meanwhile, the current limitations and corresponding reasonable solutions as well as the potential future development are also discussed.

Important Factors Affecting Induction of Cell Death, Oxidative Stress and DNA Damage by Nano- and Microplastic Particles In Vitro.

We have described the influence of selected factors that increase the toxicity of nanoplastics (NPs) and microplastics (MPs) with regard to cell viability, various types of cell death, reactive oxygen species (ROS) induction, and genotoxicity. These factors include plastic particle size (NPs/MPs), zeta potential, exposure time, concentration, functionalization, and the influence of environmental factors and cell type. Studies have unequivocally shown that smaller plastic particles are more cytotoxic, penetrate cells more easily, increase ROS formation, and induce oxidative damage to proteins, lipids, and DNA. The toxic effects also increase with concentration and incubation time. NPs with positive zeta potential are also more toxic than those with a negative zeta potential because the cells are negatively charged, inducing stronger interactions. The deleterious effects of NPs and MPs are increased by functionalization with anionic or carboxyl groups, due to greater interaction with cell membrane components. Cationic NPs/MPs are particularly toxic due to their greater cellular uptake and/or their effects on cells and lysosomal membranes. The effects of polystyrene (PS) vary from one cell type to another, and normal cells are more sensitive to NPs than cancerous ones. The toxicity of NPs/MPs can be enhanced by environmental factors, including UV radiation, as they cause the particles to shrink and change their shape, which is a particularly important consideration when working with environmentally-changed NPs/MPs. In summary, the cytotoxicity, oxidative properties, and genotoxicity of plastic particles depends on their concentration, duration of action, and cell type. Also, NPs/MPs with a smaller diameter and positive zeta potential, and those exposed to UV and functionalized with amino groups, demonstrate higher toxicity than larger, non-functionalized and environmentally-unchanged particles with a negative zeta potential.

Thermoresponsive Core-cross-linked Nanoparticles from HA-b-ELP Diblock Copolymers.

Stabilization against the dilution-dependent disassembly of self-assembled nanoparticles is a requirement for in vivo application. Herein, we propose a simple and biocompatible cross-linking reaction for the stabilization of a series of nanoparticles formed by the self-assembly of amphiphilic HA-b-ELP block copolymers, through the alkylation of methionine residues from the ELP block with diglycidyl ether compounds. The core-cross-linked nanoparticles retain their colloidal properties, with a spherical core-shell morphology, while maintaining thermoresponsive behavior. As such, instead of a reversible disassembly when non-cross-linked, a reversible swelling of nanoparticles' core and increase of hydrodynamic diameter are observed with lowering of the temperature.

Sortase A-Based Post-translational Modifications on Encapsulin Nanocompartments.

Protein-based encapsulin nanocompartments, known for their well-defined structures and versatile functionalities, present promising opportunities in the fields of biotechnology and nanomedicine. In this investigation, we effectively developed a sortase A-mediated protein ligation system in Escherichia coli to site-specifically attach target proteins to encapsulin, both internally and on its surfaces without any further in vitro steps. We explored the potential applications of fusing sortase enzyme and a protease for post-translational ligation of encapsulin to a green fluorescent protein and anti-CD3 scFv. Our results demonstrated that this system could attach other proteins to the nanoparticles' exterior surfaces without adversely affecting their folding and assembly processes. Additionally, this system enabled the attachment of proteins inside encapsulins which varied shapes and sizes of the nanoparticles due to cargo overload. This research developed an alternative enzymatic ligation method for engineering encapsulin nanoparticles to facilitate the conjugation process.

Overcoming thermostability challenges in mRNA-lipid nanoparticle systems with piperidine-based ionizable lipids.

Lipid nanoparticles (LNPs) have emerged as promising platforms for efficient in vivo mRNA delivery owing to advancements in ionizable lipids. However, maintaining the thermostability of mRNA/LNP systems remains challenging. While the importance of only a small amount of lipid impurities on mRNA inactivation is clear, a fundamental solution has not yet been proposed. In this study, we investigate an approach to limit the generation of aldehyde impurities that react with mRNA nucleosides through the chemical engineering of lipids. We demonstrated that piperidine-based lipids improve the long-term storage stability of mRNA/LNPs at refrigeration temperature as a liquid formulation. High-performance liquid chromatography analysis and additional lipid synthesis revealed that amine moieties of ionizable lipids play a vital role in limiting reactive aldehyde generation, mRNA-lipid adduct formation, and loss of mRNA function during mRNA/LNP storage. These findings highlight the importance of lipid design and help enhance the shelf-life of mRNA/LNP systems.

Autologous patient-derived exhausted nano T-cells exploit tumor immune evasion to engage an effective cancer therapy.

BACKGROUND: Active targeting by surface-modified nanoplatforms enables a more precise and elevated accumulation of nanoparticles within the tumor, thereby enhancing drug delivery and efficacy for a successful cancer treatment. However, surface functionalization involves complex procedures that increase costs and timelines, presenting challenges for clinical implementation. Biomimetic nanoparticles (BNPs) have emerged as unique drug delivery platforms that overcome the limitations of actively targeted nanoparticles. Nevertheless, BNPs coated with unmodified cells show reduced functionalities such as specific tumor targeting, decreasing the therapeutic efficacy. Those challenges can be overcome by engineering non-patient-derived cells for BNP coating, but these are complex and cost-effective approaches that hinder their wider clinical application. Here we present an immune-driven strategy to improve nanotherapeutic delivery to tumors. Our unique perspective harnesses T-cell exhaustion and tumor immune evasion to develop a groundbreaking new class of BNPs crafted from exhausted T-cells (NExT) of triple-negative breast cancer (TNBC) patients by specific culture methods without sophisticated engineering. METHODS: NExT were generated by coating PLGA (poly(lactic-co-glycolic acid)) nanoparticles with TNBC-derived T-cells exhausted in vitro by acute activation. Physicochemical characterization of NExT was made by dynamic light scattering, electrophoretic light scattering and transmission electron microscopy, and preservation and orientation of immune checkpoint receptors by flow cytometry. The efficacy of chemotherapy-loaded NExT was assessed in TNBC cell lines in vitro. In vivo toxicity was made in CD1 mice. Biodistribution and therapeutic activity of NExT were determined in cell-line- and autologous patient-derived xenografts in immunodeficient mice. RESULTS: We report a cost-effective approach with a good performance that provides NExT naturally endowed with immune checkpoint receptors (PD1, LAG3, TIM3), augmenting specific tumor targeting by engaging cognate ligands, enhancing the therapeutic efficacy of chemotherapy, and disrupting the PD1/PDL1 axis in an immunotherapy-like way. Autologous patient-derived NExT revealed exceptional intratumor accumulation, heightened chemotherapeutic index and efficiency, and targeted the tumor stroma in a PDL1+ patient-derived xenograft model of triple-negative breast cancer. CONCLUSIONS: These advantages underline the potential of autologous patient-derived NExT to revolutionize tailored adoptive cancer nanotherapy and chemoimmunotherapy, which endorses their widespread clinical application of autologous patient-derived NExT.

An Effective and Promising Strategy for Plant Protection: Synthesis of L-Carvone-Based Thiazolinone-Hydrazone/Nanochitosan Complexes with Antifungal Activity and Sustained Releasing Performance.

The development of novel natural product-derived nano-pesticide systems with loading capacity and sustained releasing performance of bioactive compounds is considered an effective and promising plant protection strategy. In this work, 25 L-carvone-based thiazolinone-hydrazone compounds 4a~4y were synthesized by the multi-step modification of L-carvone and structurally confirmed. Compound 4h was found to show favorable and broad-spectrum antifungal activity through the in vitro antifungal activity evaluation of compounds 4a~4y against eight phytopathogenic fungi. Thus, it could serve as a leading compound for new antifungal agents in agriculture. Moreover, the L-carvone-based nanochitosan carrier 7 bearing the 1,3,4-thiadiazole-amide group was rationally designed for the loading and sustained releasing applications of compound 4h, synthesized, and characterized. It was proven that carrier 7 had good thermal stability below 200 °C, dispersed well in the aqueous phase to form numerous nanoparticles with a size of~20 nm, and exhibited an unconsolidated and multi-aperture micro-structure. Finally, L-carvone-based thiazolinone-hydrazone/nanochitosan complexes were fabricated and investigated for their sustained releasing behaviors. Among them, complex 7/4h-2 with a well-distributed, compact, and columnar micro-structure displayed the highest encapsulation efficiency and desirable sustained releasing property for compound 4h and thus showed great potential as an antifungal nano-pesticide for further studies.

The Influence of the Variable Wettability Characteristics of Layers on the Transport of Nanoparticles in the Context of Drug Delivery in Skin Structures.

The rapid development of nanotechnology has offered the possibility of creating nanosystems that can be used as drug carriers. The use of such carriers offers real opportunities for the development of non-invasive drug delivery through skin structures. However, in addition to the ability to create suitable nanocarriers, it is also necessary to know how they move through dermal layers. The human skin consists of layers with different wettability characteristics, which greatly complicates how introduced substances move through it. In this work, an experimental study of the diffusion process of nanoparticles through partitions with different wettability properties was carried out. Conventional diffusion tests using Franz chambers were used for this purpose. We quantified how the wettability of the barrier, the number of layers, and their mutual configuration affect the transport of nanoparticles. Based on the results, an analysis of the phenomena taking place, depending on the wettability of the partition, was carried out. A model relationship was also proposed to determine the effective diffusion coefficient, taking into account the influence of the wettability and porosity of the barrier.

High-Spatial-Resolution Benchtop X-ray Fluorescence Imaging through Bragg-Diffraction-Based Focusing with Bent Mosaic Graphite Crystals: A Simulation Study.

X-ray fluorescence imaging (XFI) can localize diagnostic or theranostic entities utilizing nanoparticle (NP)-based probes at high resolution in vivo, in vitro, and ex vivo. However, small-animal benchtop XFI systems demonstrating high spatial resolution (variable from sub-millimeter to millimeter range) in vivo are still limited to lighter elements (i.e., atomic number Z≤45). This study investigates the feasibility of focusing hard X-rays from solid-target tubes using ellipsoidal lens systems composed of mosaic graphite crystals with the aim of enabling high-resolution in vivo XFI applications with mid-Z (42≤Z≤64) elements. Monte Carlo simulations are performed to characterize the proposed focusing-optics concept and provide quantitative predictions of the XFI sensitivity, in silico tumor-bearing mice models loaded with palladium (Pd) and barium (Ba) NPs. Based on simulation results, the minimum detectable total mass of PdNPs per scan position is expected to be on the order of a few hundred nanograms under in vivo conform conditions. PdNP masses as low as 150 ng to 50 ng could be detectable with a resolution of 600 µm when imaging abdominal tumor lesions across a range of low-dose (0.8 µGy) to high-dose (8 µGy) exposure scenarios. The proposed focusing-optics concept presents a potential step toward realizing XFI with conventional X-ray tubes for high-resolution applications involving interesting NP formulations.

Cannabidiol-Loaded Solid Lipid Nanoparticles Ameliorate the Inhibition of Proinflammatory Cytokines and Free Radicals in an In Vitro Inflammation-Induced Cell Model.

Cannabidiol (CBD) is a non-psychoactive compound derived from Cannabis sativa. It has demonstrated promising effects in combating inflammation and holds potential as a treatment for the progression of chronic inflammation. However, the clinical application of CBD is limited due to its poor solubility and bioavailability. This study introduces an effective method for preparing CBD-loaded solid lipid nanoparticles (CBD-SLNs) using a combination of low-energy hot homogenization and ultrasonication. We enhanced this process by employing statistical optimization with response surface methodology (RSM). The optimized CBD-SLN formulation utilizes glyceryl monostearate as the primary lipid component of the nanocarrier. The CBD-SLN formulation is screened as a potential tool for managing chronic inflammation. Stable, uniformly dispersed spherical nanoparticles with a size of 123 nm, a surface charge of -32.1 mV, an encapsulation efficiency of 95.16%, and a drug loading of 2.36% were obtained. The CBD-SLNs exhibited sustained release properties, ensuring prolonged and controlled CBD delivery, which could potentially amplify its therapeutic effects. Additionally, we observed that CBD-SLNs significantly reduced both reactive oxygen and nitrogen species and proinflammatory cytokines in chondrocyte and macrophage cell lines, with these inhibitory effects being more pronounced than those of free CBD. In conclusion, CBD-SLNs demonstrated superiority over free CBD, highlighting its potential as an effective delivery system for CBD.

Green Synthesized Chitosan Nanoparticles for Controlling Multidrug-Resistant mecA- and blaZ-Positive Staphylococcus aureus and aadA1-Positive Escherichia coli.

Antimicrobial resistance has recently been considered an emerging catastrophe globally. The public health and environmental threats were aggravated by the injudicious use of antibiotics in animal farming, aquaculture, and croup fields, etc. Consequently, failure of antibiotic therapies is common because of the emergence of multidrug-resistant (MDR) bacteria in the environment. Thus, the reduction in antibiotic spillage in the environment could be an important step for overcoming this situation. Bear in mind, this research was focused on the green synthesis of chitosan nanoparticles (ChiNPs) using Citrus lemon (Assam lemon) extract as a cross-linker and application in controlling MDR bacteria to reduce the antibiotic spillage in that sector. For evaluating antibacterial activity, Staphylococcus aureus and Escherichia coli were isolated from environmental specimens, and their multidrug-resistant pattern were identified both phenotypically by disk diffusion and genotypically by detecting methicillin- (mecA), penicillin- (blaZ), and streptomycin (aadA1)-resistance encoding genes. The inhibitory zone's diameter was employed as a parameter for determining the antibacterial effect against MDR bacteria revealing 30 ± 0.4 mm, 34 ± 0.2 mm, and 36 ± 0.8 mm zones of inhibition against methicillin- (mecA) and penicillin (blaZ)-resistant S. aureus, and streptomycin (aadA1)-resistant E. coli, respectively. The minimum inhibitory concentration at 0.31 mg/mL and minimum bactericidal concentration at 0.62 mg/mL of yielded ChiNPs were used as the broad-spectrum application against MDR bacteria. Finally, the biocompatibility of ChiNPs was confirmed by showing a negligible decrease in BHK-21 cell viability at doses less than 2 MIC, suggesting their potential for future application in antibiotic-free farming practices.