Anti-cancer activity of biosynthesized silver nanoparticles using Avicennia marina against A549 lung cancer cells through ROS/mitochondrial damages

The biosynthesized Ag NPs was synthesized by using marine mangrove plant extract Avicennia marina. The synthesized Ag NPs was confirmed by various physiochemical characterization including UV-spectrometer and XRD analysis. In addition, the shape and of the synthesized Ag NPs was morphologically identified by SEM initially and TEM finally. After confirmation, the anti-cancer property of synthesized Ag NPs was confirmed at 50 µg/mL concentration against A549 lung cancer cells by MTT assay. Further, the ability to stimulate the ROS generation and mitochondrial membrane at the IC₅₀ concentration of Ag NPs was confirmed by fluorescence microscopy using DCFH-DA and rhodamine 123 dyes respectively. Finally, the result was concluded that the synthesized Ag NPs has improved anti-cancer activity against A549 cells at lowest concentration.     

Biosynthesized silver nanoparticles using Caulerpa taxifolia against A549 lung cancer cell line through cytotoxicity effect/morphological damage

The Caulerpa taxifolia is excellent marine green algae, which produced enormous bioactive compounds with more biological activities. Also, it is an excellent source for synthesis of Ag NPs with increased bioactivity against various infections. In our study, the marine algae marine algae Caulerpa taxifolia mediated Ag NPs was synthesized effectively. The synthesized Ag NPs was characterized well using UV-spectrometer and X-ray powder diffraction (XRD) and confirmed as synthesized particle was Ag NPs. The available structure of the Ag NPs was morphologically identified by scanning electron microscope (SEM), and exact minimum size, polydispersive spherical shape of the entire Ag NPs structure was confirmed by Transmission electron microscope (TEM). Further, the anti-cancer efficiency of biosynthesized Ag NPs against A549 lung cancer cells was found at 40 µg/mL concentration by cytotoxicity experiment. In addition, the phase contrast images of the result were supported the Ag NPs, which damaged the A549 morphologically clearly. Finally, florescence microscopic images were effectively proved the anti-cancerous effect against A549 lung cancer cells due to the condensed morphology of increased death cells. All the confirmed in-vitro results were clearly stated that the Caulerpa taxifolia mediated Ag NPs has superior anti-cancer agent against A549 lung cancer cells.     

Bioengineered silver nanoparticles using Curvularia pallescens and its fungicidal activity against Cladosporium fulvum

Microorganisms based biosynthesis of nanomaterials has triggered significant attention, due to their great potential as vast source of the production of biocompatible nanoparticles (NPs). Such biosynthesized functional nanomaterials can be used for various biomedical applications. The present study investigates the green synthesis of silver nanoparticles (Ag NPs) using the fungus Curvularia pallescens (C. pallescens) which is isolated from cereals. The C. pallescens cell filtrate was used for the reduction of AgNO₃ to Ag NPs. To the best of our knowledge C. pallescens is utilized first time for the preparation of Ag NPs. Several alkaloids and proteins present in the phytopathogenic fungus C. pallescens were mainly responsible for the formation of highly crystalline Ag NPs. The as-synthesized Ag NPs were characterized by using UV–Visible spectroscopy, X-ray diffraction and transmission electron microscopy (TEM). The TEM micrographs have revealed that spherical shaped Ag NPs with polydisperse in size were obtained. These results have clearly suggested that the biomolecules secreted by C. pallescens are mainly responsible for the formation and stabilization of nanoparticles. Furthermore, the antifungal activity of the as-prepared Ag NPs was tested against Cladosporium fulvum, which is the major cause of a serious plant disease, known as tomato leaf mold. The synthesized Ag NPs displayed excellent fungicidal activity against the tested fungal pathogen. The extreme zone of reduction occurred at 50 μL, whereas, an increase in the reduction activity is observed with increasing the concentration of Ag NPs. These encouraging results can be further exploited by employing the as synthesized Ag NPs against various pathogenic fungi in order to ascertain their spectrum of fungicidal activity.     

Biosynthesis of silver nanoparticles by the endophytic fungus Epicoccum nigrum and their activity against pathogenic fungi

There is an enormous interest in developing safe, cost-effective and environmentally friendly technologies for nano-materials synthesis. In the present study, extracellular biosynthesis of silver nanoparticles was achieved by Epicoccum nigrum, an endophytic fungus isolated from the cambium of Phellodendron amurense. The reduction of the silver ions was monitored by UV–visible spectrophotometry, and the characterization of the Ag NPs was carried out by X-ray diffraction and transmission electron microscopy. The synthesized Ag NPs were exceptionally stable. It was found that an alkaline pH favored the formation of Ag NPs and elevated temperature accelerated the reduction process. Furthermore, the antifungal activity of the Ag NPs was assessed using a microdilution method. The biosynthesized Ag NPs showed considerable activity against the pathogenic fungi. The current research opens a new path for the green synthesis of Ag NPs and the process is easy to scale up for biomedical applications.     

The system approach to the identification of aflatoxigenic fungi in foodstuffs and feedstuffs

Aspergillus flavus, A parasiticus, A nomius, A tamarii andA pseudotamarii are important microorganisms capable of producing aflatoxins and further mycotoxins. Aflatoxigenic Aspergillus species are morphologically similar species belonging to the Aspergillus section Flavi. The aflatoxigenic fungal strains were isolated from foods (cereals, pulses, oilseeds, dried fruit, spices), soil, air and water. Mycological analyses are based on valid standards and recommendations of the International Commission for Food Mycology (ICFM). The identification of isolated aflatoxigenic fungi in foodstuffs and feedstuffs can be proved by using classical mycological cultivation methods, diagnostic nutrient media, chemotaxonomy and molecular biological methods (PCR). The system approach to the identification of aflatoxigenic fungi combines these four methods. Thirty strains of the aflatoxigenic fungi were tested.     

Silver nanoparticles synthesized by Nigrospora oryzae showed antifungal activity

In this investigation, an alternate green-route based on myco-synthesised silver nanoparticles (Ag NPs) was evaluated to control plant disease to reduce the usage of synthetic chemicals. Here, we described biologically synthesised Ag NPs using the corn grain contaminant, Nigrospora oryzae, and were well-characterised by UV–visible spectrophotometer, X-ray powder diffraction (XRD), transmission electron microscopy (TEM), Energy Dispersive Spectroscopy (EDS) and particle size analyzer. The pathogenic behaviour of the Fusarium spp. were checked on Giza 86 and Giza 90 cultivars under greenhouse conditions. F. moniliforme and F. oxysporum exhibited high pathogenecity against Giza 90 and Giza 86 cultivars respectively. The antifungal activity of biosynthesised Ag NPs was evaluated against eight species of Fusaria causing damping-off of cotton seedlings. In vitro treatments with different concentrations of Ag NPs were achieved on Czapek Dox agar and Potato dextrose agar plates. Fungal growth was drastically retarded from 25 to 200 ppm of Ag NPs interaction. The antifungal activity of Ag NPs against the Fusarium spp. was clearly proven.     

Differently Environment Stable Bio-Silver Nanoparticles: Study on Their Optical Enhancing and Antibacterial Properties

Generally, limited research is extended in studying stability and applicational properties of silver nanoparticles (Ag NPs) synthesized by adopting ‘green chemistry’ protocol. In this work, we report on the synthesis of stable Ag NPs using plant-derived materials such as leaf extract of Neem (Azadirachta indica) and biopolymer pectin from apple peel. In addition, the applicational properties of Ag NPs such as surface-enhanced Raman scattering (SERS) and antibacterial efficiencies were also investigated. As-synthesized nanoparticles (NPs) were characterized using various instrumentation techniques. Both the plant materials (leaf extract and biopolymer) favored the synthesis of well-defined NPs capped with biomaterials. The NPs were spherical in shape with an average particle size between 14-27 nm. These bio-NPs exhibited colloidal stability in most of the suspended solutions such as water, electrolyte solutions (NaCl; NaNO₃), biological solution (bovine serum albumin), and in different pH solutions (pH 7; 9) for a reasonable time period of 120 hrs. Both the bio-NPs were observed to be SERS active through displaying intrinsic SERS signals of the Raman probe molecule (Nile blue A). The NPs were effective against the Escherichia coli bacterium when tested in nutrient broth and agar medium. Scanning and high-resolution transmission electron microscopy (SEM and HRTEM) images confirmed cellular membrane damage of nanoparticle treated E. coli cells. These environmental friendly template Ag NPs can be used as an antimicrobial agent and also for SERS based analytical applications.     

Complete green synthesis of silver-nanoparticles applying seed-borne Penicillium duclauxii

Seed-borne fungus Penicillium duclauxii was examined in this study to investigate its capability of synthesizing silver nanoparticles (Ag-NPs). In vitro experiments were conducted using corn-grain contaminating fungal isolate. Ag-NPs detection and characterization were assayed by the aid of spectroscopic techniques. Spectroscopy (energy dispersive), X-ray diffraction, transmission electron-microscope and optical absorption dimensions were employed.Ag-NPs with biosynthesized were used to test invitro against Bipolaris sorghicola; the cause of target leaf spot disease on sorghum plants. The myco-synthesis of Ag NPs using Penicillium duclauxii was proved in this study. Moreover, Bipolaris sorghicola was successfully inhibited by such Ag NPs in vitro.     

Comparison of Antimicrobial Properties of Silver Nanoparticles Synthesized from Selected Bacteria

Green silver nanoparticle (AgNP) biosynthesis is facilitated by the enzyme mediated reduction of Ag ions by plants, fungi and bacteria. The antimicrobial activity of green AgNPs is useful to overcome the challenge of antimicrobial resistance. Antimicrobial properties of biosynthesized AgNPs depend on multiple factors including culture conditions and the microbial source. The antimicrobial activity of AgNPs biosynthesized by Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923 and Acinetobacter baumannii (confirmed clinical isolate) were investigated in this study. Biosynthesis conditions (AgNO₃ concentration, pH, incubation temperature and incubation time) were optimized to obtain the maximum AgNP yield. Presence of AgNPs was confirmed by observing a characteristic UV–Visible absorbance peak in 420–435 nm range. AgNP biosynthesis was optimal at 0.4 g/L AgNO₃ concentration under alkaline conditions at 60–70 °C. The biosynthesized AgNPs showed higher stability compared to chemogenized AgNPs in the presence of electrolytes. AgNPs synthesized by P. aeruginosa were the most stable while NPs of S. aureus were the least stable. AgNPs synthesized by P. aeruginosa and S. aureus showed good antimicrobial potential against E. coli, P. aeruginosa, S. aureus, MRSA and Candida albicans. AgNPs synthesized by S. aureus had greater antimicrobial activity. The antimicrobial activity of NPs may vary depending on the size and the morphology of NPs.     

Characterization of Aspergillus flavus strains from Brazilian Brazil nuts and cashew by RAPD and ribosomal DNA analysis

Aims:  The aim of this study was to determine the genetic variability in Aspergillus flavus populations from Brazil nut and cashew and develop a polymerase chain reaction (PCR) detection method.
Methods and Results:  Chomatography analysis of 48 isolates identified 36 as aflatoxigenic (75%). One hundred and forty-one DNA bands were generated with 11 random amplified polymorphic DNA (RAPD) primers and analysed via unweighted pair group analysis, using arithmetic means (UPGMA). Isolates grouped according to host, with differentiation of those from A. occidentale also according to geographical origin. Aspergillus flavus -specific PCR primers ASPITSF2 and ASPITSR3 were designed from ribosomal DNA internal transcribed spacers (ITS 1 and 2), and an internal amplification control was developed, to prevent false negative results. Specificity to only A. flavus was confirmed against DNA from additional aspergilli and other fungi.
Conclusions:  RAPD-based characterization differentiated isolates according to plant host. The PCR primer pair developed showed specificity to A. flavus , with a detection limit of 10 fg.
Significance and Impact of the Study:  Genetic variability observed in A. flavus isolates from two Brazilian agroecosystems suggested reproductive isolation. The PCR detection method developed for A. flavus represents progress towards multiplex PCR detection of aflatoxigenic and nonaflatoxigenic strains in Hazard Analysis Critical Control Point systems.     

Biosynthesized silver nanoparticles for inhibition of antibacterial resistance and biofilm formation of methicillin-resistant coagulase negative Staphylococci

The ability of a natural stabilizing and reducing agent on the synthesis of silver nanoparticles (Ag NPs) was explored using a rapid and single-pot biological reduction method using Nocardiopsis sp. GRG1 (KT235640) biomass. The UV–visible spectral analysis of Ag NPs was found to show a maximum absorption peak located at a wavelength position of ∼422 nm for initial conformation. The major peaks in the XRD pattern were found to be in excellent agreement with the standard values of metallic Ag NPs. No other peaks of impurity phases were observed. The morphology of Ag NPs was confirmed through TEM observation, demonstrating that the particle size distribution of Ag NPs entrenched in spherical particles is in a range between 20 and 50 nm. AFM analysis further supported the nanosized morphology of the synthesized Ag NPs and allowed quantifying the Ag NPs surface roughness. The synthesized Ag NPs showed significant antibacterial and antibiofilm activity against biofilm positive methicillin-resistant coagulase negative Staphylococci (MR-CoNS), which were isolated from urinary tract infection as determined by spectroscopic methods in the concentration range of 5–60 µg/ml. The inhibition of biofilm formation with coloring stain was morphologically imaged by confocal laser scanning microscopy (CLSM). Morphological alteration of treated bacteria was observed by SEM analysis. The results clearly indicate that these biologically synthesized Ag NPs could provide a safer alternative to conventional antibiofilm agents against uropathogen of MR-CoNS.     

A mycological and bacteriological survey on feed ingredients and mixed poultry feeds in Reunion island

A survey was carried out in Reunion island to obtain data on the occurrence of fungi, aflatoxigenic strains of Aspergillus flavus, aflatoxins, total aerobic bacteria and salmonellae of 150 samples of mixed poultry feeds and raw materials. These were collected at five farms over a 3-month period during the warm rainy season.White corn and Brazilian soybean meal seemed to present a better microbiological quality than yellow corn and US soybean meal.Mixed poultry feeds presented a high total mold count reflecting the mold flora of raw materials. The most frequent and abundant fungi were Aspergillus flavus, A. glaucus group, Fusarium spp., Penicillium spp., A. candidus, Mucor spp., A. restrictus, Scopulariopsis spp., Cladosporium spp. and A. versicolor. Of the 118 A. flavus strains screened, 42 (35.6%) were aflatoxigenic. Yellow corn samples were the most frequently contamined with aflatoxigenic strains (54.5%), followed by mixed feeds (44%).Of the 66 samples tested, 24 (36%) contained aflatoxins (traces to 22 ng/g). A good correlation seemed to exist between presence of at least one aflatoxigenic strain per sample and presence of aflatoxins.     

Comparative study of antifungal activity of two preparations of green silver nanoparticles from Portulaca oleracea extract

The green silver nanoparticles (green AgNPs) exhibit an exceptional antimicrobial property against different microbes, including bacteria and fungi. The current study aimed to compare the antifungal activities of both the crude aqueous extract of Portulaca oleracea or different preparations of green AgNPs biosynthesized by mixing that aqueous extract with silver nitrate (AgNO₃). Two preparations of the green AgNPs were synthesized either by mixing the aqueous extract of P. oleracea with silver nitrate (AgNO₃) (normal AgNPs) or either irradiation of the AgNPs, previously prepared, under ⁶⁰Co γ-ray using chitosan (gamma-irradiated AgNPs). Characterization of different AgNPs were tested by Zeta potential analyzer, Ultraviolet (UV) Visible Spectroscopy, and Fourier-Transform Infrared (FTIR) spectrometry. Three different plant pathogenic fungi were tested, Curvularia spicifera, Macrophomina phaseolina, and Bipolaris sp. The antifungal activities were evaluated by Transmission Electron Microscope (TEM) for either the crude aqueous extract of P. oleracea at three doses (25%, 50%, and 100%) or the newly biosynthesized AgNPs, normal or gamma-irradiated. With a few exceptions, the comparative analysis revealed that the irradiated green AgNPs at all three concentrations showed a relatively stronger antifungal effect than the normal AgNPs against all the three selected fungal strains. UV–visible spectroscopy of both preparations showed surface plasmon resonance at 421 nm. TEM results showed that both AgNPs were aggregated and characterized by a unique spherical shape, however, the gamma-irradiated AgNPs were smaller than the non-irradiated AgNPs (0.007–0.026 µM vs. 0.009–0.086 µM). TEM photographs of the fungal strains treated with the two AgNPs preparations showed flaccid structures, condensed hyphae, and shrunken surface compared with control cells. The data suggested that the biosynthesized P. oleracea AgNPs have antifungal properties against C. spicifera, M. phaseolina, and Bipolaris sp. These AgNPs may be considered a fungicide to protect different plants against phytopathogenic fungi.     

Biosynthesis of silver nanoparticles using Malva parviflora and their antifungal activity

Cheeseweed mallow (Malva parviflora L.) was used to biosynthesize silver nanoparticles. The biosynthesized silver nanoparticles were classified by UV–vis Spectroscopy and Fourier-Transform Infrared Spectroscopy (FT-IR). The shape and size distribution were visualized by Transmission Electron Microscopy (TEM), Field Emission Scanning Electron Microscopy (FE-SEM), and Zeta potential analysis. The chemical composition of M. parviflora leaf extract was identified by Gas Chromatography and Mass Spectroscopy (GC/MS). Finally, in vitro antifungal assay was done to assess the potential of biosynthesized silver nanoparticles and crude leaf extract of M. parviflora for inhibiting the mycelial growth of phytopathogenic fungi. The UV–vis analysis manifests the formation of silver nanoparticles. FTIR analysis established that chemicals of the leaf extract stabilized the biosynthesized silver nanoparticles by binding with the free silver ions. The TEM, FE-SEM and zeta potential analyzer confirmed that the biosynthesized silver nanoparticles were mostly spherical with an average diameter of 50.6 nm. The biosynthesized silver nanoparticles and leaf extract of M. parviflora effectively mitigate the mycelial growth of Helminthosporium rostratum, Fusarium solani, Fusarium oxysporum, and Alternaria alternata. The maximum reduction in mycelial growth by biosynthesized nanoparticles was observed against H. rostratum (88.6%). Whereas, the leaf extract of M. parviflora was most effective against F. solani (65.3%). Thus, the biosynthesis of nanoparticle assisted by M. parviflora is a feasible and eco-friendly method for the synthesis of silver nanoparticles. Further the silver nanoparticles and leaf extract of M. parviflora could be explored for the development of the fungicide.     

The Antimicrobial Properties of Silver Nanoparticles in Bacillus subtilis Are Mediated by Released Ag+ Ions

The superior antimicrobial properties of silver nanoparticles (Ag NPs) are well-documented, but the exact mechanisms underlying Ag-NP microbial toxicity remain the subject of intense debate. Here, we show that Ag-NP concentrations as low as 10 ppm exert significant toxicity against Bacillus subtilis, a beneficial bacterium ubiquitous in the soil. Growth arrest and chromosomal DNA degradation were observed, and flow cytometric quantification of propidium iodide (PI) staining also revealed that Ag-NP concentrations of 25 ppm and above increased membrane permeability. RedoxSensor content analysis and P_hag-GFP expression analysis further indicated that reductase activity and cytosolic protein expression decreased in B. subtilis cells treated with 10–50 ppm of Ag NPs. We conducted X-ray absorption near-edge structure (XANES) and extended X-ray absorption fine structure (EXAFS) analyses to directly clarify the valence and fine structure of Ag atoms in B. subtilis cells placed in contact with Ag NPs. The results confirmed the Ag species in Ag NP-treated B. subtilis cells as Ag₂O, indicating that Ag-NP toxicity is likely mediated by released Ag⁺ ions from Ag NPs, which penetrate bacterial cells and are subsequently oxidized intracellularly to Ag₂O. These findings provide conclusive evidence for the role of Ag⁺ ions in Ag-NP microbial toxicity, and suggest that the impact of inappropriately disposed Ag NPs to soil and water ecosystems may warrant further investigation.     

Revision von Stylothalamia columnaris Le Maitre 1935 (Sphinctozoa,Porifera) aus dem Lias von Marokko

A revision of the original material ofStromatomorpha californica var.columnaris Le Maitre 1935 from the Pliensbachian of Morocco and a comparison withStylothalamia dehmi Ott 1967 from the Carnian of Eastern Alps confirm v.Hillebrandts supposition that the two species could be identical. Kurzfassung: Eine Revision des Original-Materials vonStromatomorpha californica var.columnaris le Maitre 1935 aus dem Pliensbachien von Marokko und ein Vergleich mitStylothalamia dehmi Ott 1967 aus dem Kam der Ost-Alpen bestätigen die Vermutung v.Hillebrandts (1971), daß beide Arten ident sind.     

Detection of aflatoxigenic fungi in feeds using the PCR method

The possibility of using the polymerase chain reaction (PCR) to speed up and specify the detection of aflatoxigenic fungi isolated from feed was investigated. The method, applied to 2 genes encoding the biosynthesis of aflatoxins (apa-2 and ver-1), was optimized on two collection cultures (Aspergillus flavus CCM F-108 and A. parasiticus CCM F-550). The specificity of the optimized PCR method was proved on collection cultures of different kinds of fungi. Fifty feed samples out of which 18 showed positive findings of aflatoxigenic fungi on an Aspergillus Flavus and Parasiticus Agar (AFPA) medium were tested. Isolated strains of Aspergillus strains were verified using the PCR method; its reaction products were detected in 1% agarose gel by electrophoresis. The results almost exclusively matched those gained from the AFPA medium.     

Synthesis of silver nanoparticles using plant derived 4-N-methyl benzoic acid and evaluation of antimicrobial,antioxidant and antitumor activity

The present study is to investigate the antitumor, antioxidant and antibacterial potential of silver nanoparticles (Ag NPs) synthesized from a phenolic derivative 4-N-methyl benzoic acid, isolated from a medicinal plant (Memecylon umbellatum Burm F). The Bio-inspired nanoparticles (NPs) were analyzed by using UV–vis spectroscopy, FTIR, HRTEM, Zeta potential and XRD techniques. The UV–vis spectroscopy study at the band of 430 nm confirmed the nanoparticles formation. HRTEM report showed that the AgNPs synthesized were in the size range 7–23 nm. The harvested nanoparticles were subjected to anti-bacterial assay and a dose dependent inhibitory action was observed against the tested human pathogens. Among the tested bacteria, Acinetobacter baumannii was found to be highly sensitive to AgNPs (diameter of zone of inhibition was 31 mm). Further, the silver nanoparticles exhibited a good anti-tumor activity against the breast cancer cell line (MCF 7) with an IC₅₀ value of 42.19 µg/mL. As the present study confirmed a good antibacterial, antioxidant and antitumor activity in the nanoparticles synthesized using 4-N-methyl benzoic acid derived from a medicinal plant, the product can be further tested to formulate a good lead compound for biomedical applications.     

Fungicidal and anti-aflatoxigenic effects of the essential oil of Cymbopogon citratus (DC.) Stapf. (lemongrass) against Aspergillus flavus Link. isolated from stored rice

AIMS: To develop a natural fungicide against aflatoxigenic fungi, to protect stored rice, using the essential oil of lemongrass. METHODS AND RESULTS: Aspergillus flavus Link. was isolated from stored rice and identified as an aflatoxigenic strain. Lemongrass oil was tested against A. flavus and the test oil was fungistatic and fungicidal against the test pathogen at 0.6 and 1.0 mg ml(-1), respectively. Aflatoxin production was completely inhibited at 0.1 mg ml(-1). The results obtained from the thin layer chromatographic bioassay and gas chromatography indicated citral a and b as the fungicidal constituents in lemongrass oil. During the fumigant toxicity assay of lemongrass oil, the sporulation and the mycelial growth of the test pathogen were inhibited at the concentrations of 2.80 and 3.46 mg ml(-1), respectively. CONCLUSION: Lemongrass oil could be used to manage aflatoxin formation and fungal growth of A. flavus in stored rice. SIGNIFICANCE AND IMPACT OF THE STUDY: Currently, fungicides are not used to control fungal pests or mycotoxin production on stored rice. Rice treated with the essential oil of lemongrass could be used to manage fungal pests as well as the insect pests in stored rice. The essential oil is chemically safe and acceptable to consumers, as synthetic chemical fungicides can cause adverse health effects to consumers.     

Antibacterial and antibiofilm efficacy of Ag NPs,Ni NPs and Al2O3 NPs singly and in combination against multidrug-resistant Klebsiella pneumoniae isolates

BackgroundAlthough traditional antibiotic therapy provided an effective approach to combat pathogenic bacteria, the long-term and widespread use of antibiotic results in the evolution of multidrug-resistant bacteria. Recent progress in nanotechnology offers an alternative opportunity to discover and develop novel antibacterial agents.MethodsA total of 51 K. pneumoniae strains were collected from several specimens of hospitalized patients and identified by two parallel methods (biochemical tests and Vitek-2 system). The antibiotic sensitivity of isolates was evaluated by disk diffusion antibiogram and Vitek-2 system. The biofilms formation ability of antibiotic-resistant strains was examined by microtiter plate and tube methods based on crystal violet staining. The molecular technique was used to determine key genes responsible for biofilms formation of clinical isolates. The antibacterial and antibiofilm activities of Ag NPs, Ni NPs, Al₂O₃ NPs singly (NPs) and in combination (cNPs) were investigated against selected strains using standard methods. Moreover, the cytotoxicity of NPs was evaluated on mouse neural crest-derived (Neuro-2A) cell line.ResultsThe results of bacterial studies revealed that more than 80 % of the isolates were resistant to commonly used antibiotics and about 95 % of them were able to form biofilms. Moreover, the presence of fimA and mrkA genes were determined in all biofilm-producing strains. The results of antibacterial and antibiofilm activities of NPs and cNPs demonstrated the lower MIC and MBEC values for Al₂O₃ NPs singly as well as for Ag/Ni cNPs and Ag/Al₂O₃ cNPs in combination, respectively. Overall, the inhibitory effects of cNPs were superior to NPs against all strains. Furthermore, the results of the checkerboard assays showed that Ag NPs act synergistically with two other NPs against multidrug-resistant Klebsiella pneumoniae (MDR-K. pneumoniae) isolates. The in vitro cytotoxicity assay revealed no significant toxicity of NPs against Neuro-2A cells.ConclusionIn the present study, the combination of Ag NPs, Ni NPs, and Al₂O₃ NPs were used against MDR-K. pneumoniae strains and antibacterial and antibiofilm activities were observed for Ag/Ni cNPs and Ag/Al₂O₃ cNPs.