The susceptibility of S. aureus and P. aeruginosa to CoLig NPs was determined in 96-well microtiter plates through the broth dilution method as previously described [47 (link)]. Different dilutions of CoLig NPs were incubated at 37 °C with bacterial suspensions (5 × 105 CFU/mL) in Nutrient broth. After 24 h, the optical density at 600 nm was measured (Ultramark Microplate Imaging System, Bio-Rad Laboratories S.r.l., Segrate, Italy) to obtain the minimum inhibitory concentration (MIC). As a control, Lig NPs were used.
The morphology of S. aureus and P. aeruginosa treated with CoLig NPs was examined using SEM (Merlin Zeiss, operating at 1 kV). Bacterial cultures were grown in NB overnight and then diluted to an OD600 = 0.01. The suspension was mixed with CoLig NPs to achieve a final concentration of 2.4 mg/mL and transferred to a 48-well plate containing silicon wafers. After 24 h at 37 °C, the liquid was drained, and the bacteria remaining on the wafers were fixed overnight in a 2% paraformaldehyde solution. Finally, the bacteria were dehydrated by incubating the wafers with increasing concentrations of ethanol for 1 h each (25, 50, 75, and 100%). The same process was repeated for untreated bacteria.
The morphology of S. aureus and P. aeruginosa treated with CoLig NPs was examined using SEM (Merlin Zeiss, operating at 1 kV). Bacterial cultures were grown in NB overnight and then diluted to an OD600 = 0.01. The suspension was mixed with CoLig NPs to achieve a final concentration of 2.4 mg/mL and transferred to a 48-well plate containing silicon wafers. After 24 h at 37 °C, the liquid was drained, and the bacteria remaining on the wafers were fixed overnight in a 2% paraformaldehyde solution. Finally, the bacteria were dehydrated by incubating the wafers with increasing concentrations of ethanol for 1 h each (25, 50, 75, and 100%). The same process was repeated for untreated bacteria.
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