Insight into the Covalently Oriented Immobilization of Antibodies on Gold Nanoparticle Probes to Improve Sensitivity in the Colorimetric Detection of Listeria monocytogenes.

In this work, the covalently oriented conjugation of monoclonal Listeria monocytogenes antibody (mAb-Lis) to amino-terminated oligo(ethylene glycol)-capped gold nanoparticles (NH2-TEG-AuNPs) was studied. After NH2-TEG-AuNPs were synthesized, the amino-terminated ligands on the particles were then linked to the carboxyl groups in the mAb-Lis through EDC/NHS chemistry. By maintaining the pH of the solution at ∼5, the Fc region of the antibody could preferably attach to the particle surface, providing a specific Fab region that was available for binding with the target pathogen. The resulting mAb-NH-TEG-AuNPs could act as a colorimetric probe for L. monocytogenes based on a particular antigen-antibody interaction, which resulted in a drastic aggregation of particles. This caused the color of the colloidal solution to transition from red-pink to purple or even gray depending on the pathogen concentration. To perform quantitative analysis, the absorbance ratio of A650/A534 was monitored as a function of L. monocytogenes concentration using a spectrophotometer. The detection limit was very low at 11 CFU/mL. Furthermore, a lateral flow strip (LFS) was fabricated as a portable device for onsite utilization. LFS detection could be completed by the naked eye and by a smartphone. The detection limit of LFS was estimated to be 103 CFU/mL. Our methods exhibited a substantial improvement in sensitivity compared to that of previous studies on immuno-based nanoparticles. The assay could be completed in 15 min, and no cross reactivity by any pathogen was found. Hence, the designed AuNPs exhibit great promise for use in monitoring L. monocytogenes for food safety and in other applications.

Disinfection efficiency test for contaminated surgical mask by using Ozone generator.

BACKGROUND: Ozone (O3) is an effective disinfectant agent that leaves no harmful residues. Due to the global health crisis caused by the COVID-19 pandemic, surgical masks are in high demand, with some needing to be reused in certain regions. This study aims to evaluate the effects of O3 for pathogen disinfection on reused surgical masks in various conditions. METHODS: O3 generators, a modified PZ 2-4 for Air (2000 mg O3/L) and a modified PZ 7 -2HO for Air (500 mg O3/L), were used together with 1.063 m3 (0.68 × 0.68 × 2.3 m) and 0.456 m3 (0.68 × 0.68 × 1.15 m) acrylic boxes as well as a room-sized 56 m3 (4 × 4 × 3.5 m) box to provide 3 conditions for the disinfection of masks contaminated with enveloped RNA virus (105 FFU/mL), bacteria (103 CFU/mL) and fungi (102 spores/mL). RESULTS: The virucidal effects were 82.99% and 81.70% after 15 min of treatment with 2000 mg/L O3 at 1.063 m3 and 500 mg/L O3 at 0.456 m3, respectively. The viral killing effect was increased over time and reached more than 95% after 2 h of incubation in both conditions. By using 2000 mg/L O3 in a 1.063 m3 box, the growth of bacteria and fungi was found to be completely inhibited on surgical masks after 30 min and 2 h of treatment, respectively. Using a lower-dose O3 generator at 500 mg O3/L in 0.456 m3 provided lower efficiency, although the difference was not significant. Using O3 at 2000 mg O3/L or 500 mg O3/L in a 56 m3 room is efficient for the disinfection of all pathogens on the surface of reused surgical masks. CONCLUSIONS: This study provided the conditions for using O3 (500-2000 mg/L) to reduce pathogens and disinfect contaminated surgical masks, which might be applied to reduce the inappropriate usage of reused surgical masks.

Inhibition of Bacterial Efflux Pumps by Crude Extracts and Essential Oil from Myristica fragrans Houtt. (Nutmeg) Seeds against Methicillin-Resistant Staphylococcus aureus.

Myristicafragrans Houtt. (Nutmeg) is a widely known folk medicine across several parts of Asia, particularly used in antimicrobial treatment. Bacterial resistance involves the expression of efflux pump systems (chromosomal norA and mepA) in methicillin-resistant Staphylococcus aureus (MRSA). Crude extract (CE) and essential oil (EO) obtained from nutmeg were applied as efflux pump inhibitors (EPIs), thereby enhancing the antimicrobial activity of the drugs they were used in. The major substances in CE and EO, which function as EPIs, in a descending order of % peak area include elemicin, myristicin, methoxyeugenol, myristicin, and asarone. Here, we investigated whether the low amount of CE and EO used as EPIs was sufficient to sensitize MRSA killing using the antibiotic ciprofloxacin, which acts as an efflux system. Interestingly, synergy between ciprofloxacin and CE or EO revealed the most significant viability of MRSA, depending on norA and mepA, the latter being responsible for EPI function of EO. Therefore, CE and EO obtained from nutmeg can act as EPIs in combination with substances that act as efflux systems, thereby ensuring that the MRSA strain is susceptible to antibiotic treatment.

Influence of Gold Nanoparticles with Different Surface Charges on Localization and Monocyte Behavior.

The use of gold nanoparticles (AuNP) has been established in nanocarriers, diagnostics, and biosensors. Access to the targeted sites of these nanomaterials could directly involve the first line of defense, the innate immune system. Charges of nanomaterials play a critical role in a number of aspects such as stabilization, cellular uptake, modulation, and function of cells. Interactions and modulations of the charged nanomaterials against the innate immune system may occur even at very low concentration. To understand the effects of charges on monocyte behavior, in this study, the positively and negatively charged AuNP (AuNP+ve and AuNP-ve) of the similar size and shape on cytotoxicity, recognition, cellular behavior, and function were evaluated in vitro using U937 human monocyte cells as an innate immunity model. Both types of AuNP at various concentrations (0-5 nM) exhibited low toxicity. In addition, the cellular internalization of the AuNP+ve and AuNP-ve, as determined by TEM, occurred by different mechanisms, and the internalization had no effect on cellular destruction, as implied by the low levels of %LDH. Interestingly, the AuNP+ve recognition and internalization seemingly entered cells through receptor dependence and strongly affected cellular response to express both pro-inflammatory (IL-1ß) and anti-inflammatory (TGF-ß) cytokines, while the AuNP-ve stimulated TNF-α expression. Nevertheless, the AuNP-treated cells maintained normal function when exposed to planktonic bacteria. Thus, these results indicated that one part of the immune system interacted with different surface-charged AuNP, suggesting appropiate immunomodulation in biomedicine.

Prebiotic, Antipathogenic Bacteria and Hypocholesterolemia Properties of Fermented Rice Bran Extracts Derived from Black Rice and Germinated Brown Rice.

Rice bran is a rich source of health-promoting nutrition and bioactive compounds; nevertheless, the properties of rice brans depend on cultivars, ages, and preparation methods, drawing the potential of raw materials for health benefits. Therefore, this research aimed to investigate the health-promoting properties of fermented rice bran extracts from cultivar black rice (H7F) and germinated brown rice (G13F), focusing on their prebiotic, antipathogenic bacteria activity and safety demonstrated in vitro and in vivo study models, respectively. Here, the screening of metabolites' change after rice bran fermentation by ATR-FTIR spectra revealed specific peaks corresponding to the composited components of protein, carbohydrate, and lipid. Then, in the in vitro study, the prebiotic capability of H7F and G13F extracts was demonstrated by a growth-promoting effect on Lactobacillus delbrueckii subsp. lactis under specific acidic conditions. Furthermore, antipathogenic bacterial activity against Escherichia coli and Staphylococcus aureus was presented at 25 mg/mL of MIC values and 50 mg/mL of MBC of both fermented rice bran extracts, eliminating the bacteria by interfering with the biofilm formation. For safety, an acute and chronic toxicity study using Wistar rats was conducted, in which changes in the body and organ weights, histopathology of organs, blood chemistry, and hematological parameters were observed after H7F and G13F treatment. Desirably, they showed no toxicity, with a significant reduction in blood cholesterol levels in the chronic treatment of H7F and G13F. Conclusively, the overall results evidenced the health benefits of H7F and G13F related to their prebiotic and antipathogenic bacteria properties and hypocholesterolemia potential with a high level of safety. Therefore, the fermented rice bran extracts were demonstrated as potential materials for the further development of functional ingredients and health products.