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Abstract <p>The rise of multidrug-resistant (MDR) pathogens, particularly extended-spectrum β-lactamase (ESBL)-producing <i>Klebsiella pneumoniae</i>, highlights the urgent need for alternative antimicrobial agents. This study reports the biogenic synthesis of zinc oxide Nanoparticles (ZnO NPs) using the endophytic bacterium <i>Bacillus amyloliquefaciens</i> NWR-14 and evaluates their antibacterial, antibiofilm, antioxidant, and cytotoxic properties. Characterization using UV–Vis spectroscopy, X-ray diffraction (XRD), Fourier-transform infrared (FTIR) spectroscopy, field emission scanning electron microscopy (FE-SEM), high-resolution transmission electron microscopy (HR-TEM), dynamic light scattering (DLS), and atomic force microscopy (AFM) confirmed a hexagonal wurtzite structure with an average particle size of 20–30&#xa0;nm. The ZnO NPs exhibited significant antibacterial activity against ESBL-producing <i>K. pneumoniae</i>, producing a mean zone of inhibition of 17.33 ± 0.57&#xa0;mm compared to 20.66 ± 1.15&#xa0;mm for ceftazidime–avibactam (<i>p</i> &lt; 0.05, one-way ANOVA, <i>n</i> = 3). The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were 25&#xa0;µg/mL and 100&#xa0;µg/mL, respectively (<i>p</i> &lt; 0.05). SEM analysis revealed extensive bacterial cell damage, including cell wall disruption and lysis. The ZnO NPs also demonstrated dose-dependent antibiofilm activity, achieving 81.21% inhibition at 60&#xa0;µg/mL (<i>p</i> &lt; 0.05) compared with untreated controls. Antioxidant assays indicated moderate free radical scavenging activity, while cytotoxicity studies on L929 fibroblast cells showed over 80% cell viability at 25&#xa0;µg/mL. These findings suggest that biogenic ZnO NPs synthesized using <i>B. amyloliquefaciens</i> NWR-14 present an eco-friendly and effective strategy to combat MDR pathogens and biofilm-related infections, with promising potential in biomedical and environmental applications.</p> Highlights <p>• Endophyte mediated green synthesis of ZnO NPs was achieved using <i>Bacillus amyloliquefaciens</i> NWR-14.</p> <p>• Physicochemical analyses confirmed crystalline wurtzite ZnO NPs of 20–30 nm size.</p> <p>• Biogenic ZnO NPs showed potent antibacterial activity against ESBL producing <i>Klebsiella pneumoniae</i>.</p> <p>• MIC and MBC values were 25 and 100 µg/mL, respectively.• ZnO NPs inhibited biofilm formation by more than 80 % at 60 µg/mL.</p> <p>• Cytotoxicity assays indicated good biocompatibility at low concentrations.</p> Graphical Abstract <p></p>

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Biogenic ZnO Nanoparticles from Bacillus amyloliquefaciens NWR-14 for Combating ESBL-Producing Klebsiella pneumoniae

  • 期刊论文

  • 发表日期：

    2026年2月10日

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  • Sijo Asokan,
  • Teena Jacob,
  • Tijo Cherian,
  • Afaf A AlSosowaa,
  • Faheem Q Al-Mojahid,
  • Smitha Vijayan

摘要

Abstract

The rise of multidrug-resistant (MDR) pathogens, particularly extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae, highlights the urgent need for alternative antimicrobial agents. This study reports the biogenic synthesis of zinc oxide Nanoparticles (ZnO NPs) using the endophytic bacterium Bacillus amyloliquefaciens NWR-14 and evaluates their antibacterial, antibiofilm, antioxidant, and cytotoxic properties. Characterization using UV–Vis spectroscopy, X-ray diffraction (XRD), Fourier-transform infrared (FTIR) spectroscopy, field emission scanning electron microscopy (FE-SEM), high-resolution transmission electron microscopy (HR-TEM), dynamic light scattering (DLS), and atomic force microscopy (AFM) confirmed a hexagonal wurtzite structure with an average particle size of 20–30 nm. The ZnO NPs exhibited significant antibacterial activity against ESBL-producing K. pneumoniae, producing a mean zone of inhibition of 17.33 ± 0.57 mm compared to 20.66 ± 1.15 mm for ceftazidime–avibactam (p < 0.05, one-way ANOVA, n = 3). The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were 25 µg/mL and 100 µg/mL, respectively (p < 0.05). SEM analysis revealed extensive bacterial cell damage, including cell wall disruption and lysis. The ZnO NPs also demonstrated dose-dependent antibiofilm activity, achieving 81.21% inhibition at 60 µg/mL (p < 0.05) compared with untreated controls. Antioxidant assays indicated moderate free radical scavenging activity, while cytotoxicity studies on L929 fibroblast cells showed over 80% cell viability at 25 µg/mL. These findings suggest that biogenic ZnO NPs synthesized using B. amyloliquefaciens NWR-14 present an eco-friendly and effective strategy to combat MDR pathogens and biofilm-related infections, with promising potential in biomedical and environmental applications.

Highlights

• Endophyte mediated green synthesis of ZnO NPs was achieved using Bacillus amyloliquefaciens NWR-14.

• Physicochemical analyses confirmed crystalline wurtzite ZnO NPs of 20–30 nm size.

• Biogenic ZnO NPs showed potent antibacterial activity against ESBL producing Klebsiella pneumoniae.

• MIC and MBC values were 25 and 100 µg/mL, respectively.• ZnO NPs inhibited biofilm formation by more than 80 % at 60 µg/mL.

• Cytotoxicity assays indicated good biocompatibility at low concentrations.

Graphical Abstract