Revolutionizing healthcare: Harnessing nano biotechnology with zinc oxide nanoparticles to combat biofilm and bacterial infections-A short review.

A crucial pathogenic mechanism in many bacterial diseases is the ability to create biofilms. Biofilms are suspected to play a role in over 80 % of microbial illnesses in humans. In light of the critical requirement for efficient management of bacterial infections, researchers have explored alternative techniques for treating bacterial disorders. One of the most promising ways to address this issue is through the development of long-lasting coatings with antibacterial properties. In recent years, antibacterial treatments based on metallic nanoparticles (NPs) have emerged as an effective strategy in the fight over bacterial drug resistance. Zinc oxide nanoparticles (ZnO-NPs) are the basis of a new composite coating material. This article begins with a brief overview of the mechanisms that underlie bacterial resistance to antimicrobial drugs. A detailed examination of the properties of metallic nanoparticles (NPs) and their potential use as antibacterial drugs for curing drug-sensitive and resistant bacteria follows. Furthermore, we assess metal nanoparticles (NPs) as powerful agents to fight against antibiotic-resistant bacteria and the growth of biofilm, and we look into their potential toxicological effects for the development of future medicines.

Comprehensive Analysis of Bioactive Compounds in Wild Ganoderma applanatum Mushroom from Kerala, South India: Insights into Dietary Nutritional, Mineral, Antimicrobial, and Antioxidant Activities.

The present study focused on the mushroom Ganoderma, which has been used in Eastern countries for centuries as a food and medicinal source. Specifically, the fruiting bodies of Ganoderma applanatum from the Kerala Forest Research Institute in Thirussur, Kerala, India, were analyzed for their nutritional and medicinal properties. The methanolic extracts of G. applanatum were used to examine secondary metabolites and proximate profiles, revealing the presence of various phytochemicals such as terpenoids, phenolics, glycosides, alkaloids, flavonoids, and saponins. Further analysis revealed the presence of significant amounts of calcium, sodium, phosphorus, and manganese. The compounds were characterized using chromatographic analysis, FTIR, and GC-MS, which revealed potential therapeutic compounds with C-H and C-O bonds in the amide group, ß-glycosides, and C-C/C-O vibrations of phenolic substances. Mushroom extract at a concentration of 100 µg mL-1 exhibited potent antimicrobial activity against various pathogens. This study suggests that G. applanatum has a rich biochemical composition and pharmacological potential, making it a promising candidate for drug development and traditional medicine, and contributes valuable insights into its diverse therapeutic applications.

Intracellular Dynamics-Resolved Label-Free Scattering Reveals Real-Time Metabolism of Single Bacteria.

Nanoscopic investigation of bacterial cells is essential to reveal their physiological status, impacting all cellular functions. Currently, this requires labeled probes or targeted staining procedures. Herein, we report a new bacterial feature, intracellular dynamics-resolved Rayleigh scattering (IDRS), that visualizes spatiotemporal cytoplasmic transitions in unlabeled bacteria and characterizes their real-time physiological status in 10 s. From single-bacterium IDRS signals, we discovered unique spatial patterns and their multiple transitions in Gram-negative and Gram-positive bacteria. The magnitude of IDRS signal variation highly correlated with the metabolic status of bacteria, differentiating persistent subpopulations. This is also the first report demonstrating distinct real-time metabolic conditions of unlabeled drug-resistant bacteria that are exposed to different doses of antibiotics. Our strategy opens up a way to simultaneously trace in situ metabolic and antibiotic resistance statuses, which can be applied in single-cell level control of bacterial metabolism and efficacy with a heterogeneous nature.

Friends against the Foe: Synergistic Photothermal and Photodynamic Therapy against Bacterial Infections.

Multidrug-resistant (MDR) bacteria are rapidly emerging, coupled with the failure of current antibiotic therapy; thus, new alternatives for effectively treating infections caused by MDR bacteria are required. Hyperthermia-mediated photothermal therapy (PTT) and reactive oxygen species (ROS)-mediated photodynamic therapy (PDT) have attracted extensive attention as antibacterial therapies owing to advantages such as low invasiveness, low toxicity, and low likelihood of causing bacterial resistance. However, both strategies have notable drawbacks, including the high temperature requirements of PTT and the weak ability of PDT-derived ROS to penetrate target cells. To overcome these limitations, a combination of PTT and PDT has been used against MDR bacteria. In this review, we discuss the unique benefits and limitations of PTT and PDT against MDR bacteria. The mechanisms underlying the synergistic effects of the PTT-PDT combination are also discussed. Furthermore, we introduced advancements in antibacterial methods using nano-based PTT and PDT agents to treat infections caused by MDR bacteria. Finally, we highlight the existing challenges and future perspectives of synergistic PTT-PDT combination therapy against infections caused by MDR bacteria. We believe that this review will encourage synergistic PTT- and PDT-based antibacterial research and can be referenced for future clinical applications.

Characterization of Increased Extracellular Vesicle-Mediated Tigecycline Resistance in Acinetobacter baumannii.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is the most detrimental pathogen that causes hospital-acquired infections. Tigecycline (TIG) is currently used as a potent antibiotic for treating CRAB infections; however, its overuse substantially induces the development of resistant isolates. Some molecular aspects of the resistance mechanisms of AB to TIG have been reported, but they are expected to be far more complicated and diverse than what has been characterized thus far. In this study, we identified bacterial extracellular vesicles (EVs), which are nano-sized lipid-bilayered spherical structures, as mediators of TIG resistance. Using laboratory-made TIG-resistant AB (TIG-R AB), we demonstrated that TIG-R AB produced more EVs than control TIG-susceptible AB (TIG-S AB). Transfer analysis of TIG-R AB-derived EVs treated with proteinase or DNase to recipient TIG-S AB showed that TIG-R EV proteins are major factors in TIG resistance transfer. Additional transfer spectrum analysis demonstrated that EV-mediated TIG resistance was selectively transferred to Escherichia coli, Salmonella typhimurium, and Proteus mirabilis. However, this action was not observed in Klebsiella pneumonia and Staphylococcus aureus. Finally, we showed that EVs are more likely to induce TIG resistance than antibiotics. Our data provide direct evidence that EVs are potent cell-derived components with a high, selective occurrence of TIG resistance in neighboring bacterial cells.

Micrococcus luteus-derived extracellular vesicles attenuate neutrophilic asthma by regulating miRNAs in airway epithelial cells.

Bacterial extracellular vesicles (EVs) have been shown to regulate various pulmonary diseases, but their functions in asthma remain uncertain. To demonstrate the clinical significance of Micrococcus luteus-derived EVs (MlEVs) in asthma, we enrolled 45 asthmatic patients (20 patients with neutrophilic asthma [NA], 25 patients with eosinophilic asthma [EA]) and 40 healthy controls (HCs). When the prevalence of IgG1 and IgG4 specific to MlEVs was evaluated in serum by ELISA, lower levels of MlEV-specific IgG4 (but not IgG1) were noted in asthmatic patients than in HCs. Among asthmatic patients, significantly lower levels of MIEV-specific IgG4 were noted in patients with NA than in those with EA. Moreover, there was a positive correlation between serum MlEV-specific IgG4 levels and FEV1 (%) values. In asthmatic C57BL/6 mice, MlEVs significantly attenuated neutrophilic airway inflammation by reducing the production of IL-1ß and IL-17 in bronchoalveolar lavage fluid as well as the number of group 3 innate lymphoid cells (ILC3s) in lung tissues. To clarify the functional mechanism of MlEVs in NA, the effect of MlEVs on airway epithelial cells (AECs) and immune cells was investigated ex vivo. According to microarray analysis, MlEVs upregulated hsa-miR-4517 expression in AECs. Moreover, this miRNA could suppress IL-1ß production by monocytes, resulting in the inhibition of ILC3 activation and neutrophil recruitment. These findings suggest that MlEVs could be a novel therapeutic agent for managing unresolved NA by regulating miRNA expression in AECs.

A Nanocomposite with Extracellular Vesicles from Lactobacillus paracasei as a Bioinspired Nanoantibiotic Targeting Staphylococcus aureus.

The utilization of biomimetic materials that merge functional nanoparticles (NPs) with a cell-derived nanosized membrane is a state-of-the-art approach to harnessing cellular properties for biomedical applications. However, the development of biocompatible and species-selective biomimetic agents against hazardous pathogens threatening human health is still in its early stages. Herein, we report the synthesis and functional analysis of a novel nanoplatform in which a PEGylated MoS2-ZnO (MZ) nanocomposite was cloaked with a generally regarded as safe (GRAS)-grade Lactobacillus paracasei-derived extracellular vesicle (LPEV) for MZ-LPEV nanocomposite and evaluated its activity against Staphylococcus aureus. The MZ nanocomposite was characterized via X-ray diffraction, transmission electron microscopy, and X-ray photoelectron spectroscopy. The coating of MZ with LPEV was confirmed through nanoparticle tracking analysis and zeta potential measurements. MZ-LPEV exhibited 5- to 20-fold higher antibacterial activity than that of ZO NPs and MZ nanocomposite against S. aureus. Reactive oxygen species (ROS) production and bacterial membrane disruption were confirmed as antibacterial mechanisms of MZ-LPEV. Finally, MZ-LPEV exhibited enhanced biocompatibility and selectivity for S. aureus. All our results showed that LPEV could be utilized for developing synergistic nanoantibiotics against S. aureus.

Photo-Stimuli-Responsive CuS Nanomaterials as Cutting-Edge Platform Materials for Antibacterial Applications.

Photo-stimuli-responsive therapeutic nanomaterials have gained widespread attention as frontline materials for biomedical applications. The photoactivation strategies are classified as single-modality (based on either reactive oxygen species (ROS)-based photodynamic therapy (PDT), hyperthermia-based photothermal therapy (PTT)), or dual-modality (which combines PDT and PTT). Due to its minimal invasiveness, phototherapy has been extensively applied as an efficient therapeutic platform for many diseases, including skin cancers. However, extensive implementation of phototherapy to address the emergence of multidrug-resistant (MDR) bacterial infections remains challenging. This review focuses on copper sulfide (CuS) nanomaterials as efficient and cost-effective PDT and PTT therapeutic nanomaterials with antibacterial activity. The features and merits of CuS nanomaterials as therapeutics are compared to those of other nanomaterials. Control of the dimensions and morphological complexity of CuS nanomaterials through judicious synthesis is then introduced. Both the in vitro antibacterial activity and the in vivo therapeutic effect of CuS nanomaterials and derivative nanocomposites composed of 2D nanomaterials, polymers, metals, metal oxides, and proteins are described in detail. Finally, the perspective of photo-stimuli-responsive CuS nanomaterials for future clinical antibacterial applications is highlighted. This review illustrates that CuS nanomaterials are highly effective, low-toxic, and environmentally friendly antibacterial agents or platform nanomaterials for combatting MDR bacterial infections.

Membrane Vesicles Derived from Gut Microbiota and Probiotics: Cutting-Edge Therapeutic Approaches for Multidrug-Resistant Superbugs Linked to Neurological Anomalies.

Multidrug-resistant (MDR) superbugs can breach the blood-brain barrier (BBB), leading to a continuous barrage of pro-inflammatory modulators and induction of severe infection-related pathologies, including meningitis and brain abscess. Both broad-spectrum or species-specific antibiotics (ß-lactamase inhibitors, polymyxins, vancomycin, meropenem, plazomicin, and sarecycline) and biocompatible poly (lactic-co-glycolic acid) (PLGA) nanoparticles have been used to treat these infections. However, new therapeutic platforms with a broad impact that do not exert off-target deleterious effects are needed. Membrane vesicles or extracellular vesicles (EVs) are lipid bilayer-enclosed particles with therapeutic potential owing to their ability to circumvent BBB constraints. Bacteria-derived EVs (bEVs) from gut microbiota are efficient transporters that can penetrate the central nervous system. In fact, bEVs can be remodeled via surface modification and CRISPR/Cas editing and, thus, represent a novel platform for conferring protection against infections breaching the BBB. Here, we discuss the latest scientific research related to gut microbiota- and probiotic-derived bEVs, and their therapeutic modifications, in terms of regulating neurotransmitters and inhibiting quorum sensing, for the treatment of neurodegenerative diseases, such as Parkinson's and Alzheimer's diseases. We also emphasize the benefits of probiotic-derived bEVs to human health and propose a novel direction for the development of innovative heterologous expression systems to combat BBB-crossing pathogens.

A Review of Immunomodulatory Reprogramming by Probiotics in Combating Chronic and Acute Diabetic Foot Ulcers (DFUs).

Diabetic foot ulcers (DFUs) are characterized by a lack of angiogenesis and distal limb diabetic neuropathy. This makes it possible for opportunistic pathogens to protect the biofilm-encased micro-communities, causing a delay in wound healing. The acute and chronic phases of DFU-associated infections are distinguished by the differential expression of innate proinflammatory cytokines and tumor necrosis factors (TNF-α and -ß). Efforts are being made to reduce the microbial bioburden of wounds by using therapies such as debridement, hyperbaric oxygen therapy, shock wave therapy, and empirical antibiotic treatment. However, the constant evolution of pathogens limits the effectiveness of these therapies. In the wound-healing process, continuous homeostasis and remodeling processes by commensal microbes undoubtedly provide a protective barrier against diverse pathogens. Among commensal microbes, probiotics are beneficial microbes that should be administered orally or topically to regulate gut-skin interaction and to activate inflammation and proinflammatory cytokine production. The goal of this review is to bridge the gap between the role of probiotics in managing the innate immune response and the function of proinflammatory mediators in diabetic wound healing. We also highlight probiotic encapsulation or nanoformulations with prebiotics and extracellular vesicles (EVs) as innovative ways to tackle target DFUs.

Bioconjugated Thymol-Zinc Oxide Nanocomposite as a Selective and Biocompatible Antibacterial Agent against Staphylococcus Species.

Owing to the rapid spread of antibiotic resistance among Staphylococcus species, effective and low-risk alternatives to antibiotics are being actively searched. Thymol (THO), the most abundant component of the oil extracted from thyme, can be considered as a natural antibacterial alternative. However, the low antibacterial activity and non-selectivity of THO limit its usage as a universal anti-Staphylococcus agent. Herein, we report the bioconjugation of THO with ZnO nanoparticle (ZO), which resulted in the TZ nanocomposite (NC), as a potent and selective antibacterial agent against Staphylococcus species, particularly S. epidermidis. The cell-free supernatant (CFS) of ATCC 25923 cultures was employed for the production of TZ NC. Successful production of TZ NC was confirmed via X-ray diffraction (XRD), Fourier-transform infrared (FT-IR) spectroscopy, and ultraviolet-visible (UV-Vis) studies. TZ NC had selective efficacy against Staphylococcus species, with MIC values 2-32-fold lower than THO. The antibacterial mechanisms of TZ NC are proposed to involve membrane rupture, suppression of biofilm formation, and modulation of new cell wall and protein-synthesis-associated cellular pathways. Its biocompatibility against HCT116 cells was also checked. Our findings suggest that the TZ nanocomposite could improve the selectivity and bactericidal activity of THO against target species.

A MoS2 based silver-doped ZnO nanocomposite and its antibacterial activity against ß-lactamase expressing Escherichia coli.

Multidrug-resistant (MDR) Gram-negative bacteria including Escherichia coli are increasingly resistant to current antibiotics. Among the strategies implemented to eradicate such MDR pathogens, approaches based on two-dimensional (2D) nanomaterials have received considerable attention. In particular, the excellent physicochemical properties of 2D molybdenum disulfide (MoS2) nanosheets, including a high surface area, good conductivity, and good surface retention, are advantageous for their use as bactericidal agents. Herein, we report the fabrication of a MoS2-based nanocomposite conjugated with silver-doped zinc oxide (AZM) as an effective antibacterial agent against E. coli species. The properties of AZM were characterized, and its antibacterial activity against MDR E. coli strains with different resistance types was evaluated. MoS2 was found to activate the antibacterial activity of AZM and provide enhanced selectivity against MDR E. coli strains expressing ß-lactamases. We proposed that membrane disruption of bacterial cell walls was the major cell death mechanism for MDR E. coli. Furthermore, surface charge perturbation could explain the differences in AZM activity against MDR E. coli strains expressing a ß-lactamase and a mobilized colistin resistance (mcr-1) gene product. Thus, a MoS2-based nanocomposite with a functional conjugation strategy could be a selective nano-antibacterial platform against infections caused by MDR E. coli with resistance against ß-lactam antibiotics.

Repurposing of Ciclopirox to Overcome the Limitations of Zidovudine (Azidothymidine) against Multidrug-Resistant Gram-Negative Bacteria.

Multidrug-resistant (MDR) Gram-negative bacteria are the top-priority pathogens to be eradicated. Drug repurposing (e.g., the use of non-antibiotics to treat bacterial infections) may be helpful to overcome the limitations of current antibiotics. Zidovudine (azidothymidine, AZT), a licensed oral antiviral agent, is a leading repurposed drug against MDR Gram-negative bacterial infections. However, the rapid emergence of bacterial resistance due to long-term exposure, overuse, or misuse limits its application, making it necessary to develop new alternatives. In this study, we investigated the efficacy of ciclopirox (CPX) as an alternative to AZT. The minimum inhibitory concentrations of AZT and CPX against MDR Gram-negative bacteria were determined; CPX appeared more active against ß-lactamase-producing Escherichia coli, whereas AZT displayed no selectivity for any antibiotic-resistant strain. Motility assays revealed that ß-lactamase-producing Escherichia coli strains were less motile in nature and more strongly affected by CPX than a parental strain. Resistance against CPX was not observed in E. coli even after 25 days of growth, whereas AZT resistance was observed in less than 2 days. Moreover, CPX effectively killed AZT-resistant strains with different resistance mechanisms. Our findings indicate that CPX may be utilized as an alternative or supplement to AZT-based medications to treat opportunistic Gram-negative bacterial infections.

Wash-Free Amperometric Escherichia coli Detection via Rapid and Specific Proteolytic Cleavage by Its Outer Membrane OmpT.

Various methods have been developed for the detection of Escherichia coli (E. coli); however, they are complex and time-consuming. OmpT─a cell membrane endopeptidase of E. coli─strongly embedded in the outer membrane of only E. coli, exposed to external solutions, with high proteolytic activity, could be a suitable target molecule for the rapid and straightforward detection of E. coli. Herein, a wash-free, sensitive, and selective amperometric method for E. coli detection, based on rapid and specific proteolytic cleavage by OmpT, has been reported. The method involved (i) rapid proteolytic cleavage of consecutive amino acids, after cleavage by OmpT, linked to an electrochemical species (4-aminophenol, AP), by leucine aminopeptidase (LAP, an exopeptidase), (ii) affinity binding of E. coli on an electrode, and (iii) electrochemical-enzymatic (EN) redox cycling. OmpT cleaved the intermediate peptide bond of a peptide substrate containing alanine-arginine-arginine-leucine-AP (-A-R-R-L-AP), forming R-L-AP, followed by the cleavage of two peptide bonds of R-L-AP sequentially by LAP, to liberate an electroactive AP. Affinity binding and EN redox cycling, in addition to rapid proteolytic cleavage by OmpT and LAP, enabled high electrochemical signal amplification. Two-sequential-cleavage was employed for the first time in protease-based detection. The calculated detection limit for E. coli cells in tap water (approximately 103 CFU/mL after 1 h incubation) was lower than those obtained without affinity binding and EN redox cycling. The detection method was highly selective to E. coli as OmpT is present in only E. coli. High sensitivity, selectivity, and the absence of wash steps make the developed detection method practically promising.

Wash-Free, Sandwich-Type Protein Detection Using Direct Electron Transfer and Catalytic Signal Amplification of Multiple Redox Labels.

Direct electron transfer (DET) between a redox label and an electrode has been used for sensitive and selective sandwich-type detection without a wash step. However, applying DET is still highly challenging in protein detection, and a single redox label per probe is insufficient to obtain a high electrochemical signal. Here, we report a wash-free, sandwich-type detection of thrombin using DET and catalytic signal amplification of multiple redox labels. The detection scheme is based on (i) the redox label-catalyzed oxidation of a reductant, (ii) the conjugation of multiple redox labels per probe using a poly-linker, (iii) the low nonspecific adsorption of the conjugated poly-linker due to uncharged, reduced redox labels, and (iv) a facile DET using long, flexible poly-linker and spacer DNA. Amine-reactive phenazine ethosulfate and NADH were used as the redox label and reductant, respectively. N3-terminated polylysine was used as the poly-linker for the conjugation between an aptamer probe and multiple redox labels. Approximately 11 redox labels per probe and rapid catalytic NADH oxidation enable high signal amplification. Thrombin in urine could be detected without a wash step with a detection limit of ∼50 pM, which is practically promising for point-of-care testing of proteins.

Biomimetic Nanoparticles Coated with Bacterial Outer Membrane Vesicles as a New-Generation Platform for Biomedical Applications.

The biomedical field is currently reaping the benefits of research on biomimetic nanoparticles (NPs), which are synthetic nanoparticles fabricated with natural cellular materials for nature-inspired biomedical applications. These camouflage NPs are capable of retaining not only the physiochemical properties of synthetic nanoparticles but also the original biological functions of the cellular materials. Accordingly, NPs coated with cell-derived membrane components have achieved remarkable growth as prospective biomedical materials. Particularly, bacterial outer membrane vesicle (OMV), which is a cell membrane coating material for NPs, is regarded as an important molecule that can be employed in several biomedical applications, including immune response activation, cancer therapeutics, and treatment for bacterial infections with photothermal activity. The currently available cell membrane-coated NPs are summarized in this review. Furthermore, the general features of bacterial OMVs and several multifunctional NPs that could serve as inner core materials in the coating strategy are presented, and several methods that can be used to prepare OMV-coated NPs (OMV-NPs) and their characterization are highlighted. Finally, some perspectives of OMV-NPs in various biomedical applications for future potential breakthrough are discussed. This in-depth review, which includes potential challenges, will encourage researchers to fabricate innovative and improvised, new-generation biomimetic materials through future biomedical applications.

Bovine Serum Albumin-Immobilized Black Phosphorus-Based γ-Fe2O3 Nanocomposites: A Promising Biocompatible Nanoplatform.

The interactions between proteins and nanoparticles need to be fully characterized as the immobilization of proteins onto various nanoplatforms in the physiological system often results in the change of surface of the protein molecules to avoid any detrimental issues related to their biomedical applications. Hence, in this article, the successful low-temperature synthesis of a BP-based γ-Fe2O3 (IB) nanocomposite and its interactive behavior with bovine serum albumin (BSA)-a molecule with chemical similarity and high sequence identity to human serum albumin-are described. To confirm the formation of γ-Fe2O3 and the IB nanocomposite, X-ray diffraction, transmission electron microscopy, and X-ray photoelectron spectroscopy analyses of the materials were performed. Additionally, the physical interaction between BSA and the IB nanocomposite was confirmed via UV-Vis and photoluminescence spectral analyses. Finally, the biocompatibility of the BSA-immobilized IB nanocomposite was verified using an in vitro cytotoxicity assay with HCT-15 colon cancer cells. Our findings demonstrate that this newly developed nanocomposite has potential utility as a biocompatible nanoplatform for various biomedical applications.

A New Surface Charge Neutralizing Nano-Adjuvant to Potentiate Polymyxins in Killing Mcr-1 Mediated Drug-Resistant Escherichia coli.

Resistance to polymyxins when treating multidrug-resistant (MDR) Gram-negative bacterial infections limit therapeutic options. Here, we report the synthesis of a nickel (Ni) doped Zinc oxide (NZO) combined with black phosphorus (BP) (NZB) nanocomposite and its synergistic action with polymyxin B (PolB) against polymyxin-resistant Escherichia coli harboring mobilized colistin resistance (mcr-1) gene. NZB and PolB combination therapy expressed a specific and strong synergy against Mcr-1 expressing E. coli cells. The underlying mechanism of the synergy is the charge neutralization of the E. coli cell surface by NZB, resulting in a more feasible incorporation of PolB to E. coli. The synergistic concentration of NZB with PolB was proved biocompatible. Thus, the NZB is the first biocompatible nano-adjuvant to polymyxins against polymyxin-resistant E. coli cells, recognizing the physical status of bacteria instead of known adjuvants targeting cellular gene products. Therefore, NZB has the potential to revive polymyxins as leading last-resort antibiotics to combat polymyxin-resistant Gram-negative bacterial infections.