Two-Phase Bactericidal Mechanism of Silver Nanoparticles against Burkholderia pseudomallei

Abstract

Silver nanoparticles (AgNPs) have a strong antimicrobial activity against a variety of pathogenic bacteria. The killing mechanism of AgNPs involves direct physical membrane destruction and subsequent molecular damage from both AgNPs and released Ag+. Burkholderia pseudomallei is the causative agent of melioidosis, an endemic infectious disease primarily found in northern Australia and Southeast Asia. B. pseudomallei is intrinsically resistant to most common antibiotics. In this study, the antimicrobial activity and mechanism of AgNPs (10–20 nm) against B. pseudomallei were investigated. The MIC and MBC for nine B. pseudomallei strains ranged from 32–48 μg/mL and 96–128 μg/mL, respectively. Concentrations of AgNPs less than 256 μg/mL were not toxic to human red blood cells. AgNPs exhibited a two-phase mechanism: cell death induction and ROS induction. The first phase was a rapid killing step within 5 min, causing the direct damage of the cytoplasmic membrane of the bacterial cells, as observed by a time-kill assay and fluorescence microscopy. During the period of 5–30 min, the cell surface charge was rapidly neutralized from -8.73 and -7.74 to 2.85 and 2.94 mV in two isolates of B. pseudomallei, as revealed by zeta potential measurement. Energy-dispersive X-ray (EDX) spectroscopy showed the silver element deposited on the bacterial membrane, and TEM micrographs of the AgNP-treated B. pseudomallei cells showed severe membrane damage and cytosolic leakage at 1/5 MIC and cell bursting at MBC. During the killing effect the released Ag+ from AgNPs was only 3.9% from the starting AgNPs concentration as observed with ICP-OES experiment. In the second phase, the ROS induction occurred 1–4 hr after the AgNP treatment. Altogether, we provide direct kinetic evidence of the AgNPs killing mechanism, by which cell death is separable from the ROS induction and AgNPs mainly contributes in the killing action. AgNPs may be considered a potential candidate to develop a novel alternative agent for melioidosis treatment with fast action.

Controlled Synthesis of Triangular Silver Nanoplates by Gelatin–Chitosan Mixture and the Influence of Their Shape on Antibacterial Activity

“…This process would cause membrane damage and consequently the cellular content leaks out, resulting in bacterial death [17][18][19]. Another antibacterial mechanism of Ag nanoparticles was described in several works, related to the generation of a high level of reactive oxygen species (ROS) and free radical species [20][21][22]. Furthermore, the surrounding media of AgNPs colloid is also related to their antibacterial activities.…”

Silver Nanoparticles Synthesized with Rumex hymenosepalus: A Strategy to Combat Early Mortality Syndrome (EMS) in a Cultivated White Shrimp

“…Journal of Nanomaterials was exposed to a high concentration of NPs (100 μg/mL) for 24 h. Subsequently, it was fixed in glutaraldehyde for its preparation and observation by electron microscopy. coli bacteria observed by TEM and show that cells contain additional membranous structures [42]; in Figure 8(b), we observe these additional structures. Yin et al have described using SEM AgNPs located on the cytoplasmic membrane and cell wall [70], similar results we observed in Figure 8(b).…”

“…Killing rate was determined by plotting the total number of viable cells as log10 CFU/mL versus time. Bactericidal activities have been defined as to 3 − log 10 decrease in CFU/mL (99.9% kill) [42].…”

Combating the melioidosis pathogen using antibiotics in combination with silver nanoparticles

“…The MICs and MBCs of the antibiotics (CAZ, IMI, MER, and GENT) and AgNPs were determined by the plate-counting method measured by serial dilutions, as previously described (37). Brie y, a range of concentrations of AgNPs (4-64 µg/mL) and antibiotics (0.25-1024 µg/mL) were prepared in a 96-well plate by serial dilution.…”