PURPOSE: The purpose of this in vitro study was to assess the antimicrobial effect of sub 10-nm AgNPs in maxillofacial silicone against Staphylococcus aureus, Candida albicans, and mixed species biofilms containing both and to test the effectiveness of different AgNP concentrations against all 3 biofilms in vitro.
MATERIAL AND METHODS: Silicone disks (M511; Technovent Ltd) containing 0.0% (control), 0.1%, and 0.5% AgNPs were fabricated and treated with S. aureus, C. albicans, and mixed species strains of both in 24-well culture plates containing appropriate media. Each well received a 0.1-mL aliquot of the standardized suspension of microorganisms. The plates were incubated for 21 consecutive days, and colony-forming units per milliliter (CFU/mL) were measured on the first, third, fifth, seventh, fifteenth, and twenty-first day with the Miles and Misra method. Data were analyzed by 2-way ANOVA and the paired t test to evaluate the relationship between AgNP concentration, microbial strain, and time (α=.05). Mean CFU/mL differences for each time and for each biofilm category were assessed by repeated measure ANOVA.
RESULTS: AgNPs decreased the mean CFU/mL in both concentrations compared with the control. The 0.1% concentration showed sustained efficacy throughout the test, while the 0.5% concentration had high efficacy initially with a gradual decrease. However, the results were inconsistent for the mixed biofilm. The paired sample t test at day 3 and 15 and day 3 and 21 showed statistically significantly different results (P
RESULTS: Planktonic S. Typhi cells were cultured using standard nutrient broth whereas biofilm cells were cultured in a stressful environment using high shearing-force and bile to mimic the gallbladder. Sequencing libraries were prepared from S. Typhi planktonic cells and mature biofilm cells using the Illumina HiSeq 2500 platform, and the transcriptome data obtained were processed using Cufflinks bioinformatics suite of programs to investigate differential gene expression between the two phenotypes. A total of 35 up-regulated and 29 down-regulated genes were identified. The identities of the differentially expressed genes were confirmed using NCBI BLAST and their functions were analyzed. The results showed that the genes associated with metabolic processes and biofilm regulations were down-regulated while those associated with the membrane matrix and antibiotic resistance were highly up-regulated.
CONCLUSIONS: It is proposed that the biofilm phenotype of S. Typhi allows the bacteria to increase production of the membrane matrix in order to serve as a physical shield and to adhere to surfaces, and enter an energy conservation state in response to the stressful environment. Conversely, the planktonic phenotype allows the bacteria to produce flagella and increase metabolic activity to enable the bacteria to migrate and form new colonies of infection. This data provide a basis for further studies to uncover the mechanism of biofilm formation in S. Typhi and to discover novel genes or pathways associated with the development of the typhoid carrier state.