The impact of varying sizes of silver nanoparticles on the induction of cellular damage in Klebsiella pneumoniae involving diverse mechanisms

Ramesa Shafi Bhat; Mona Alonazi; Abeer Mohammad Aldbass; Sooad Al-Daihan

doi:10.1515/gps-2024-0162

Article Open Access

The impact of varying sizes of silver nanoparticles on the induction of cellular damage in Klebsiella pneumoniae involving diverse mechanisms

Ramesa Shafi Bhat , Mona Alonazi , Abeer Mohammad Aldbass and Sooad Al-Daihan

Published/Copyright: November 15, 2024

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From the journal Green Processing and Synthesis Volume 13 Issue 1

Abstract

Silver nanoparticles (AgNPs) are extensively studied as potent antibacterial agents targeting antibiotic-resistant pathogens. Cellular damage induced through various mechanisms that can affect multiple cell components like the outer membrane, enzymes, and proteins is closely linked to their chemical and morphological characteristics. We investigated the impact of AgNPs’ size on their antibacterial effectiveness using two differently sized nanoparticles: silver nanoparticle-Citrus limon (AgCL) with an average size of 21 nm and silver nanoparticle-Citrus sinensis (AgCS) with an average size of 13 nm, derived from C. limon and C. sinensis through environmentally friendly methods. The study evaluated their antibacterial effects by assessing morphology changes via scanning electron microscopy, metabolic alterations using Fourier transform infrared (FT-IR) spectroscopy, and oxidative stress responses through biochemical markers in Klebsiella pneumoniae cells exposed to AgNPs. The results showed that both AgCL and AgCS exhibited remarkable antibacterial activity, evidenced by inhibition zones of 14 ± 1.5 and 16 ± 1.0 mm, respectively. Morphological changes in K. pneumoniae cells treated with AgNPs were size dependent, with notable alterations noted. FT-IR spectroscopy revealed size and concentration-dependent biochemical changes, particularly in shifts in functional groups involved in the fluidity of cell wall lipid, and protein structure. Exposure to AgNPs led to increased oxidative stress markers like lipid peroxides and reduced levels of enzymatic and non-enzymatic antioxidants, more prominently observed with smaller AgCS nanoparticles (13 nm). AgNPs induce oxidative stress and morphological changes in K. pneumoniae strains, with smaller nanoparticles demonstrating greater efficacy. These findings underscore the importance of nanoparticle size in optimizing the antibacterial properties against pathogens.

Keywords: silver nanoparticles; K. pneumonia ; size; FT-IR spectroscopy; oxidative stress; morphology

1 Introduction

Metallic nanoparticles are at the top of the discussion due to the rising incidence of microbial infections and antibiotic resistance [1]. Numerous studies have highlighted metallic nanoparticles for antimicrobial effects against bacterial infections [2]. These particles induce oxidative stress, protein dysfunction, and damage to microbial cells’ membranes and DNA, leading to cell death [3,4]. Metallic nanoparticles occupy a unique size, typically less than 100 nm in at least one dimension with a high specific surface area and a greater number of surface atoms [5,6]. Compared to other metallic nanoparticles, silver has gained interest in recent decades [7,8,9]. These nanoparticles typically consist of 20–15,000 silver atoms and generally have diameters smaller than 100 nm [10,11]. Due to their high surface area-to-volume ratio, they exhibit significant antimicrobial activity, even at low concentrations [12]. Moreover, they are cost-effective and have demonstrated low cytotoxicity and minimal immunological response [13]. These characteristics make silver nanoparticles (AgNPs) highly promising for a range of biomedical applications. They are utilized in drug delivery systems, medical imaging, and molecular diagnostics. Additionally, AgNPs find application in therapeutic areas such as surgical mesh, artificial joint replacements, wound dressings, and formulations to enhance wound healing [14,15,16,17].

Several mechanisms behind the antibacterial effects of AgNPs have been proposed, but the exact mechanism is not fully understood. The continuous release of silver ions from AgNPs is the primary mechanism behind its antimicrobial activity [18,19]. These silver ions possess electrostatic attraction and a high affinity for sulfur-containing proteins, enabling them to bind to the bacterial cell wall and cytoplasmic membrane, increasing membrane permeability, and leading to disruption of the bacterial envelope [20,21]. Free silver ions can deactivate respiratory enzymes, triggering the production of reactive oxygen species (ROS) and disrupting adenosine triphosphate production inside the bacterial cells [22]. Sulfur and phosphorus are essential components of DNA, and the interaction between silver ions and these elements can interfere with DNA replication and cell division, potentially leading to microbial death [22]. Furthermore, silver ions can also inhibit protein synthesis by denaturing ribosomes in the bacterial cytoplasm [10]. The diameter of nanoparticles is a critical factor in influencing nanoparticle uptake by bacteria cells as smaller nanoparticles interact quickly and exhibit stronger antibacterial effects [23]. A higher surface area-to-mass ratio enhances their ability to penetrate bacterial membranes, thereby exerting potent antibiotic activity [24]. Smaller nanoparticles produce more radicals to combat infections by accelerating the dissolution rate of metallic ions from nanoparticles upon interaction with bacterial surfaces [23,24].

Multidrug-resistant strains are human pathogens responsible for infections like urinary tract infections, pneumonia, bloodstream infections, bacteremia, and sepsis [25,26]. Klebsiella pneumoniae, the second deadliest Gram-negative bacterium responsible for community and hospital-acquired bacteremia, has seen a significant increase in antibiotic resistance in recent years [27,28]. This trend poses a serious public health threat, as common infections like urinary tract infections become less responsive to antibiotic treatments. Consequently, managing infections caused by these pathogens has become increasingly difficult. AgNPs are in high demand for their applications in infectious diseases due to advanced drug delivery systems The unique properties of AgNP-based therapeutics and diagnostics offer promising prospects in overcoming challenges associated with acquired drug resistance in bacterial infections. Their specific size enables them to target resistant pathogenic strains effectively [29].

This research aimed to investigate the antibacterial mechanism of AgNPs mainly due to their particle size. Two differently sized Ag-NPs, produced using environmentally friendly methods were studied for their antibacterial effectiveness against the K. pneumoniae strain. This was assessed through the antibacterial activity followed by analyzing changes in cell morphology, composition, and biochemical modifications of K. pneumoniae cells in response to AgNPs of varying sizes.

2 Materials and methods

2.1 AgNPs

This study investigated two different sizes of AgNPs, with average diameters of 13 and 21 nm. The AgNPs were synthesized under optimized conditions per the methodology detailed in our recently published work [30]. Briefly, AgNPs were synthesized by gradually adding fresh juice of two different citrus fruit peels (Citrus sinensis and Citrus limon) into silver nitrate solution, maintaining a final silver nitrate concentration of 1 mM with constant stirring at room temperature. The color changes in solutions indicated the successful formation of AgNPs. Characterization of both types of nanoparticles included UV–Vis spectroscopy, zeta sizer analysis, zeta potential, transmission electron microscopy, scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy [30]. The 13-nm nanoparticles, denoted as silver nanoparticle-C. sinensis (AgCS), were synthesized using C. sinensis fruit peels, whereas the 21 nm nanoparticles, labeled as Silver Nanoparticle-C. limon (AgCL), were synthesized using C. limon fruit peels.

2.2 Antibacterial

2.2.1 K. pneumoniae culture

The gram-negative strain of K. pneumoniae was obtained from King Khalid Hospital in Riyadh, Saudi Arabia. The strain was revived on nutrient agar plates before use.

2.2.2 Anti-K. pneumoniae activity

The antibacterial activity against K. pneumoniae of AgCS and AgCL (1,000 µg·mL⁻¹) was evaluated using the disk diffusion method using 6 mm paper disks. Briefly, 20 µL of K. pneumoniae culture (10⁶ CFU·mL⁻¹) was evenly spread on sterile nutrient agar plates using a sterile cotton swab. After allowing the plates to dry for a few minutes, 50 µL of each test solution was applied to paper disks placed on the agar. Plates were incubated at 37°C for 18 h and the inhibitory zone (in mm) was measured [31].

2.2.3 Minimum inhibitory concentration (MIC) and minimum bactericidal concentrations (MBC)

The broth microdilution method was used to determine the MIC and MBC values of AgNPs. AgCS and AgCL were serially diluted in nutrient broth to achieve concentrations ranging from 3.90 to 2,000 µg·mL⁻¹ AgNPs against K. pneumoniae cultures adjusted to approximately 10⁶ CFU·mL⁻¹. The plate was incubated at 37°C for 26 h to assess the turbidity. MIC was defined as the lowest AgNP concentration inhibiting K. pneumoniae growth, while MBC was defined as the concentration required to eliminate 99.9% of K. pneumoniae, indicated by no visible growth [32].

2.3 The growth kinetics, morphological, and biochemical changes in K. pneumoniae under AgNP stress

2.3.1 Growth curve

The growth curve of K. pneumoniae was studied under AgNP stress using two different-sized AgNPs (AgCS and AgCL) in sterilized nutrient broth. K. pneumoniae cells were incubated with 10 and 20 μg·mL⁻¹ of nanoparticles separately at 37°C in a shaker incubator for 24 h. The growth curve was monitored and compared with normal K. pneumoniae growth taken as control. Optical density (OD) at 600 nm was noted to assess the growth curve.

2.3.2 Morphological changes by SEM

SEM was used to examine the morphological changes induced by AgNP exposure in K. pneumoniae cells treated for 24 h. AgCS- and AgCL-treated cells from a 10 μg·mL⁻¹ batch were harvested for SEM to examine morphological alterations.

2.3.3 Biochemical changes by Fourier transform infrared (FT-IR) spectroscopy

FT-IR spectroscopy was utilized to identify the functional groups involved in biochemical changes, focusing on compositional and metabolic alterations in K. pneumoniae cells exposed to AgNPs, and comparing them to control cultures. FT-IR analysis was carried out on K. pneumoniae cells before and after AgCS and AgCL treatment each having a concentration of 10 and 20 μg·mL⁻¹.

K. pneumoniae cells treated with 10 and 20 μg·mL⁻¹ concentrations of AgCS and AgCL were initially monitored for growth curves. Due to insufficient cell counts in the 20 μg·mL⁻¹ treatment batch, we proceeded with experiments investigating stress markers using the 10 μg·mL⁻¹ concentrations of AgCS and AgCL.

2.4 Biochemical changes through stress markers in K. pneumoniae under AgNP treatment

K. pneumoniae cells incubated with 10 μg·mL⁻¹ of AgCS and AgCL in nutrient broth were harvested after 24 h. The cells were centrifuged at 10,000 × g for 15 min and washed with ice-cold saline. Cells were sonicated and centrifuged at 10,000 × g for 15 min to get the clear supernatant for measurement of the following stress markers.

2.4.1 Lipid peroxides

Lipid peroxides were determined using the thiobarbituric acid reactive substances assay, which quantifies malondialdehyde and other lipid peroxidation products [33].

2.4.2 Reduced glutathione (GSH)

Reduced GSH levels were measured by using 5,5′-dithiobis(2-nitrobenzoic acid), to produce a yellow-colored product upon reacting with sulfhydryl groups [34].

2.4.3 Glutathione S-transferase (GST) activity

GST was evaluated by its catalytic reaction involving GSH, a substrate for GST, and 1-chloro-2,4-dinitrobenzene [35].

2.4.4 Ascorbic acid

Vitamin C levels were estimated by using the Folin–Ciocalteu reagent, resulting in a blue-colored complex [36].

2.4.5 Total crude protein

The total protein content was estimated using the Bradford method with bovine serum albumin as a standard [37].

2.5 Statistical analysis

The results are expressed as the mean ± standard deviation, with each experiment conducted three times.

3 Results

3.1 Antimicrobial activity

Disk diffusion tests were performed to assess the antibacterial effectiveness of AgCl and AgCS at a concentration of 1,000 μg·mL⁻¹ against K. pneumoniae, as depicted in Figure 1. The antibacterial efficacy was evaluated through inhibition zone diameters, MIC, and MBC shown in Tables 1 and 2. The K. pneumoniae strains showed sensitivity to 50 μL disks of AgCL and AgCS, yielding inhibition zones of 14 ± 1.5 mm and 16 ± 1.0 mm, respectively (Table 1). Additionally, AgCL and AgCS exhibited bacteriostatic effects against K. pneumoniae showing MIC of 15.6 and 7.81 μg·mL⁻¹, and MBC of 62.5 and 31.25 μg·mL⁻¹, respectively (Table 2). Gentamicin and Penicillin (10 µg each on 6-mm disks, Condalab) served as the standard antibiotics. The K. pneumoniae strain exhibited sensitivity to the gentamicin disk, demonstrating an IZ of 18 mm, whereas it showed resistance to the penicillin disk with no observable inhibition.

Figure 1

Agar plates the antibacterial activity of (a) AgNPs and (b) standard antibiotic against K. pneumonia.

Table 1

Inhibitory activity against K. pneumoniae by disk diffusion method

Strain	Zone of inhibition (mm)
Strain	AgCL	AgCS	Std1	Std 2
K. pneumonia	14 ± 1.5	16 ± 1.0	18	Resistant

Standard antibiotic (Std) 1 – gentamycin (10 µg); 2 – pencilline (10 μg).

Table 2

The MIC and MBC values of AgNPs against K. pneumonia

Strain	MIC (µg·mL⁻¹)		MBC (µg·mL⁻¹)
K. pneumonia	AgCL	AgCS	AgCL	AgCS
K. pneumonia	15.6	7.81	62.5	31.25

3.2 K. pneumoniae growth curve, metabolic, and structural changes under AgNP stress

K. pneumoniae strains were evaluated for their sensitivity to AgCl and AgCS at 10 and 20 μg·mL⁻¹. Figure 2 illustrates the bacterial growth rates under varying conditions with AgNPs. During the growth curve experiments, the suppression of bacterial growth was observed to correlate with nanoparticle size, demonstrating significantly stronger inhibition with AgCS (13 nm) than with AgCL (21 nm). Additionally, concentration-dependent inhibition was observed, as treatment with 20 μg·mL⁻¹ showed more pronounced growth suppression than 10 μg·mL⁻¹ compared to control.

Figure 2

Growth curves of K. pneumoniae incubated with AgCL and AgCS for 24 h (1 = 10 µg·mL⁻¹; 2 = 20 µg·mL⁻¹).

SEM analyses investigate size-dependent morphological changes induced by 10 μg·mL⁻¹ of AgCL and AgCS on K. pneumoniae cells (Figure 3). K. pneumoniae cells grown in normal media were used as controls for comparison. Cells treated with AgCl and AgCS exhibited distinct characteristics. Specifically, Figure 3a (control or untreated) shows regular, smooth rod-shaped K. pneumoniae cells; Figure 3b (10 μg·mL⁻¹ AgCL-treated) illustrates slight irregularities and cell aggregation; and Figure 3c (10 μg·mL⁻¹ AgCS-treated) demonstrates cell rupture, aggregation, and destruction.

Figure 3

SEM micrograph showing the K. pneumoniae cells treated with 10 μg·mL⁻¹ AgNPs: (a) control; (b) AgCL treated; and (c) AgCS treated.

Exposure to AgNPs caused specific changes in FT-IR spectra of K. pneumoniae cells, as depicted in Figure 4 and Table 3. Typically, FT-IR spectra of bacteria exhibit four distinct regions: Region I (3,000–2,800 cm⁻¹) reflects cell membrane fatty acids, Region II (1,700–1,500 cm⁻¹) includes the amide I (1,650 cm⁻¹) and amide II (1,550 cm⁻¹) bands of proteins and peptides, Region III (1,500–1,200 cm⁻¹) corresponds to fatty acids, proteins, and phosphate-carrying molecules, and Region IV (1,200–900 cm⁻¹) shows absorption bands characteristic of polysaccharides or carbohydrates in microbial cell walls. Our findings indicated shifts in peak positions across nearly all these regions, suggesting alterations in the biochemical composition of cells following exposure to AgNPs. Figure 4a depicts the FT-IR spectra of K. pneumoniae treated with AgCL and AgCS at 10 and 20 μg·mL⁻¹ concentrations. Figure 4b compares the impact of different sizes of AgCL and AgCS on these spectra. The effect of varying concentrations of AgCL and AgCS on the FT-IR spectra of K. pneumoniae is shown in Figure 4. Remarkable shifts in FT-IR spectra were observed in a manner dependent on both size and concentration.

Figure 4

FT-IR spectra of K. pneumoniae treated with different AgNPs. (a) The FT-IR spectra of K. pneumoniae treated with AgNPs of various sizes. (b) The FT-IR spectra of K. pneumoniae treated with AgNPs at different concentrations N1 = AgCL; N2 = AgCS.

Table 3

FT-IR spectra showing observed peaks and probable functional groups in AgNP-treated K. pneumoniae

	Wave number				Functional group
Control	AgCL treated		AgCS treated
	10 μg·mL⁻¹	20 μg·mL⁻¹	10 μg·mL⁻¹	20 μg·mL⁻¹
Region I (3,800–2,800 cm ⁻¹)
	3,780.48	3,701.28	3,772.76	3,780.48	O–H stretch, free hydroxyl alcohols, phenols
3,394.72	3,356.14		3,402.43		N–H stretch primary, secondary amines, amides
	3,302.13	3,368.36	3,309.98	3,363.86	O–H stretch (Alcohols),
	3,209.56		3,209.55	3,248.13	O–H stretch carboxylic acids
3,070.68		3,163.26			C–H stretch, aromatics
3,032.10	3,024.38	3,024.28		3,024.38	C–H stretch alkanes
2,939.10	2,916.37	2,916.37	2,916.37		H–C═O: C–H stretch aldehydes
Region II (2,800–1,500 cm ⁻¹)
2,376.30	2,360.87	2,368.59	2,368.59	2,368.59	C(triple bond) N stretch nitriles
			2,152.56	2,217.13	–C(triple bond)C– stretch alkynes
1,944.25	1,944.25	1,944.25	1,944.25	1,944.25	C═O stretch ketones, saturated aliphatic
	1,867.09	1874.81	1874.81		C═O stretch aldehydes, saturated aliphatic
1,651.07	1,797.66	1,797.66	1,797.66	1,597.06	–C═C– stretch alkenes; C═O aldehyde
	1,507.06	1,597.06	1,597.06	1,527.62	N–O asymmetric stretch nitro compounds
Region III (1,500–1,200 cm ⁻¹)
	1,469.76	1,496.76	1,496.76		C–H bend alkanes
1,450.47	1,450.47	1,450.47	1,450.47		N–H bend (Nitro compounds),
1,404.18	1,404.18	1,404.18	1,404.18	1,404.18	N–O asymmetric stretch nitro compounds
1,242.46					C–N stretch aliphatic amines
Region IV (1,200–900 cm ⁻¹)
	1,149.57	1,149.57			C–O stretch alcohols, carboxylic acids, esters
1,072.42	1,072.42			1,080.14	═C–H bend (Alkenes)
	1,026.13	1,026.13	1,026.13	1,033.85	C═C–H bend (Alkenes) (Pectin)
	910.40	910.40	910.40	918.12	═C–H bend (Benzene), C–C stretch (fluorides)
		840.96			C–N stretch (amines)
748.38	756.10	756.10	756.10	756.10	C–N stretch (amines)
694.37	694.37	694.37	694.37		═C–H bend (chlorides)
624.94	617.22				═C–H bend (bromides)
532.35	540.07	540.07	532.35	532.35	C–C stretch (iodide)

3.3 Biochemical changes through stress markers in K. pneumoniae under AgNP treatment

K. pneumoniae cells treated with 10 μg·mL⁻¹ of AgCL and AgCS were evaluated for stress markers, as illustrated in Figure 5. Both types of AgNPs increased lipid peroxide levels, while a decrease in GSH, ascorbic acid, GST activity, and total protein levels was observed upon exposure. AgCS exhibited stronger effects on these stress markers than AgCL, as indicated in Figure 5.

Figure 5

The oxidative stress markers in K. pneumoniae under AgNP (10 μg·mL⁻¹) treatment. (a) Lipid peroxidation (MDA), (b) reduced GSH, (c) glutathione S-transferase activity, (d) ascorbic acid, and (e) total crude protein.

4 Discussion

Numerous studies have documented the antibacterial properties of AgNPs, but their intricate mechanism is not fully clear. This study investigates the impact of two different-sized AgNPs on the growth, biochemical profiles, and morphological changes related to oxidative stress in penicillin-resistant strains of K. pneumoniae. K. pneumoniae is a prevalent antibiotic-resistant bacterium that poses a substantial global public health threat. Given the challenges posed by antibiotic resistance, identifying novel therapeutic agents is imperative for effectively combating K. pneumoniae infections.

In this study, AgNPs of two different sizes were evaluated against K. pneumoniae: AgCL with an average diameter of 21 nm and AgCS with an average diameter of 13 nm. The results demonstrated that AgCS (13 nm) exhibited superior antibacterial activity compared to AgCL, suggesting an inverse relationship between particle size and bactericidal efficacy (Tables 1 and 2). This highlights the enhanced effectiveness of smaller-sized AgNPs in exerting antibacterial effects [38]. Smaller nanoparticles possess larger surface areas, facilitating increased release of Ag ions and improved interaction with bacterial cell membranes [39]. The antibacterial activity in this study was assessed using the disk diffusion method, which is particularly suitable for evaluating the antibacterial efficacy of Ag ions due to their ability to diffuse freely through agar matrices [40] (Figure 1). Cellular uptake and toxic level of AgNPs against bacteria mainly depend on their size [41]. The growth curves of K. pneumoniae treated with AgCS and AgCL reveal the lowest maximum OD and the longest lag time compared to untreated K. pneumoniae (Figure 2). Additionally, when bacteria are exposed to higher concentrations of NPs, the initial OD decreases, and this decline continues consistently over time, contrasting with lower nanoparticle concentrations. These findings underscore the dual impact of AgNP size and concentration on bacterial toxicity.

Our primary focus was the biochemical and morphological changes induced in K. pneumoniae cells with AgNP treatment. SEM was utilized to observe the surface morphology of K. pneumoniae, a widely used technique for visualizing bacterial structures [42]. Untreated cells exhibited a clear cell wall structure without any discernible alterations, whereas cells treated with AgCS and AgCL displayed distinct changes (Figure 3). AgCS-treated K. pneumoniae cells showed more irregular shapes, along with cell shrinkage, aggregation, and lysis compared to AgCL treated. These findings indicate that AgNPs induce size-dependent morphological changes by damaging bacterial membranes and causing leakage of cytoplasmic material.

Our SEM observations were further supported by FT-IR spectroscopic analysis of treated cells, (Table 3 and Figure 4) which revealed significant shifts across all four spectral regions (Regions I–IV). These shifts confirmed the morphological changes observed in SEM. More band shifts were observed with AgCS compared to AgCL mainly in the 3,400–2,900 cm⁻¹ range confirming the membrane alterations to damage the outer cell membrane to enable the entry of AgNPs into the cells (Figure 4a). FT-IR spectroscopy allows for rapid characterization of biochemical structural alterations in bacteria exposed to nanoparticles [43]. The technique identifies distinct regions for fatty acids, proteins peptides, carbohydrates, and fingerprints [44]. FT-IR is notably faster than conventional methods, applicable across various bacteria, and specific in differentiating chemical effects on intracellular levels [45]. Therefore, it facilitates clear discrimination between chemically exposed bacteria and controls, aiding in the rapid assessment of nanoparticle toxicity levels. Our results suggest several potential antimicrobial mechanisms. AgCS and AgCL may alter the fluidity of cell wall lipids by interacting with –CH groups of the membrane, as evidenced by shifts in the 3,400–2,900 cm⁻¹ range. Additionally, binding with amino acids in proteins and enzymes, resulting in changes to protein structures, was detected by shifts in the 1,500–2,000 cm⁻¹ range. Moreover, catalyzed oxidation and generation of ROS by AgCS and AgCL could lead to alterations in polysaccharide structures, indicated by shifts in the 900–1,200 cm⁻¹ range (Figure 4b, c).

Exposure to AgCS and AgCL resulted in oxidative damage, indicated by elevated oxidative markers such as lipid peroxides in K. pneumoniae cells. Concurrently, there was a depletion observed in both enzymatic and non-enzymatic antioxidants, notably GSH and GST (Figure 5). GSH is predominantly consumed through GST-catalyzed reactions, which serve as a crucial detoxification mechanism in bacterial cells [46,47]. These responses underscore the defense strategies employed by K. pneumoniae to mitigate oxidative stress induced by exposure to AgCS and AgCL. The toxicity of Ag NPs to bacterial cells is primarily attributed to Ag ions. Ag ions disrupt the bacterial respiratory chain and bind to sulfhydryl groups within the cellular membrane and essential enzymes, thereby causing damage to the bacteria [48]. The size-dependent antibacterial activity in our results is due to the faster dissolution of smaller nanoparticles, leading to increased release of Ag+ ions. Another critical factor contributing to AgCS and AgCL’s antibacterial properties in our results is the generation of ROS, which induces oxidative stress and apoptosis in cells [49]. ROS production triggered by AgNPs operates through two main mechanisms. First, reactive groups like radicals or oxidants, often present on the nanoparticle surface, directly contribute to ROS formation. Second, AgNPs can deactivate ROS protection pathways, such as damaging GSH, a typical cellular antioxidant, thereby reducing scavenging activity against ROS [50].

5 Conclusions

AgNPs have been extensively studied as antibacterial agents against multidrug-resistant pathogens. We examined the impact of AgNPs of varying sizes on cellular damage in penicillin-resistant K. pneumoniae cells through various mechanisms. The efficacy of AgNPs against K. pneumoniae showed a strong correlation with their size and concentration. We examined morphological changes and modifications in functional groups resulting from nanoparticle treatments to evaluate their antibacterial efficacy. In cells exposed to AgNPs, we noted biochemical alterations such as reductions in GSH, GST, ascorbic acid, and total protein along with increases in lipid peroxidation. Due to the low cell count after treatment with 20 µg·mL⁻¹ AgNPs, we limited the investigation of stress markers to a 10 µg·mL⁻¹ concentration of AgNPs. Developing biocompatible silver-based agents to combat multidrug resistance could significantly aid in controlling harmful infections.

Acknowledgments

The authors appreciate the Researchers Supporting Project number (RSP 2024R237), King Saud University, Riyadh, Saudi Arabia.

Funding information: This research project was supported by Researchers Supporting Project number (RSP 2024R237), King Saud University, Riyadh, Saudi Arabia.
Author contributions: Ramesa Shafi Bhat: writing – original draft, writing – review and editing, methodology, formal analysis; visualization. Mona Alonazi: resources. Abeer M. Aldbass: formal analysis. Sooad Al-Daihan: writing – review and editing.
Conflict of interest: Authors state no conflict of interest.
Data availability statement: All data generated or analyzed during this study are included in this published article.

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Received: 2024-07-20

Accepted: 2024-10-08

Published Online: 2024-11-15

This work is licensed under the Creative Commons Attribution 4.0 International License.

Articles in the same Issue

https://doi.org/10.1515/gps-2024-0162

Keywords for this article

silver nanoparticles; K. pneumonia; size; FT-IR spectroscopy; oxidative stress; morphology

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