Analysis of the role of iron uptake mechanisms and addition of iron-doped apatite nanoparticles in phage infections in Staphylococcus aureus and Mycobacterium smegmatis
Rost, Linda Christina
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Antibiotic resistance has become a significant global public health issue, and phage therapy could serve as an adjuvant to traditional antibiotics. Phages are viruses that kill bacteria and produce more phages. Iron-doped apatite nanoparticles (IDANPs) have been shown to increase phage killing of bacteria. However, the mechanism of JB and Yodasoda infection of bacteria, and mechanism by which IDANPs increase phage infections, is unknown. Based on the iron composition of the IDANP, as well as extensive literature describing Staphylococcus aureus having aggressive iron uptake systems, it was hypothesized that IDANPs may affect such systems, and thereby be involved in the subsequent increase of phage-mediated bacterial death. In this work, the relationship between bacterial exposure to iron and subsequent phage infectivity was established, and IDANP effect on plaque size was determined. S. aureus cells were grown in various iron treatments, plaque assays were performed. There was a strong, positive relationship between iron treatments and plaque counts. The plaque counts were 29% higher in the 0.0004g/L iron treatment, 34% higher in 0.0008g/L, 60% higher in 0.0016 g/L and 82% higher in 0.0032g/L. When S. aureus and M. smegmatis cells were treated with IDANPs, plaque sizes were significantly larger, which may indicate increased infection in adjacent cells. Plaque sizes from IDANP-treated cells continued to increase in size as plates were incubated over 24, 48 and 96 hours. Plaque sizes also increased in size in the control cells in some time frames. S. aureus cells were also grown in 0.0016g/L iron treatment and treated with IDANPs, and there was a 65% increase in plaque counts. In higher iron treatments, it was difficult to achieve a lawn of cells to perform plaque assays. Cell growth was measured by performing serial dilutions and determining CFU/mL. There was no significant difference between cells grown in M9 minimal media or treated with IDANPs. Cells were also grown in the different iron treatments over three hours, with or without IDANPs. Less growth was observed in high iron treatments, but the differences were not significant. Cell growth was relatively slower in high iron levels in overnight treatments, and the results were significantly different. These data can be used to elucidate the relationship of iron uptake and phage killing, and therefore allow conjectures as to whether or not iron uptake mechanisms may be involved in the IDANP effect. Further research in this field can provide opportunities to develop reliable alternatives to antibiotics to treat bacterial infections.