Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Erika L. Ware - Alabama State University
Co-Author(s): Atul Chaudhari, Shreekumar Pillai, and Shree Singh, Alabama State University, Montgomery, AL
Bacterial resistance to modern antibiotics is a serious concern and new strategies are needed to inhibit the infection from spreading. Metallic nanoparticles such as silver coated carbon nanotubes (AgCNTs) have demonstrated good antibacterial activity. Antimicrobial peptides (APs) have also shown efficacy against drug-resistant pathogens. However, the APs are unstable at different pH conditions, and may need to be combined with nanoparticles to be effective. In this study, we investigated the antibacterial effect of AgCNTs functionalized with a proprietary peptide TP226 (Therapeutic Peptides, Inc., fAgCNTs) under two different pH conditions (acidic pH=5 and alkaline pH=9) against a gram- negative bacterium, Salmonella Typhimurium and a gram-positive bacterium, Staphylococcus aureus. We hypothesized that the pH variations would regulate the gene expression profile in bacteria exposed to fAgCNTs. Studies included minimum inhibitory concentration assays and gene transcription assays carried out by qRT-PCR. The expression of ATP-binding cassette transporter genes (dpp, artP and livJ) in Salmonella Typhimurium and cell wall biosynthesis gene (ftsZ) in Staphylococcus aureus were investigated. The MIC assay showed that fAgCNTs were highly effective (MIC values lower than 0.9 µg/ml) as opposed to the plain AgCNTs (MIC value 62.5 µg/ml). Transcriptional analysis of the genes associated with transporter systems in Salmonella Typhimurium and cell wall biosynthesis in Staphylococcus aureus revealed that these genes were either up-regulated or down-regulated several folds (~3-4 folds) in fAgCNTs treated bacteria at alkaline pH compared to neutral pH. The expression of dppA was up regulated and expression of artP and livJ was down regulated at pH=9 in Salmonella Typhimurium exposed to fAgCNTs. The down-regulation of artP and livJ and up-regulation of dppA at pH=9 validates the better antibacterial activity of the fAgCNTs at pH=9. Similarly, the up regulation of cell division mediating gene ftsZ at pH=9 in Staphylococcus aureus indicated a strong stress response. Our results show that fAgCNTs are much more effective at alkaline pH compared to neutral pH. Currently, we are investigating the effects of fAgCNTs on the expression of several genes associated with cell division and amino acid biosynthesis in both the pathogens.
Funder Acknowledgement(s): This work was supported by NSF-REU (DBI-1358923) to Komal Vig (PI) and by NSFCREST (HRD-1241701) to Shree S. Singh (PI).
Faculty Advisor: Shreekumar Pillai, firstname.lastname@example.org
Role: My role in the lab was to make the media (LB Broth) of pH=7.4 and alkaline pH=9.1 media. After making the LB Broth, I grew the Salmonella and Staph bacteria in the different pH's overnight in a shaking incubator. Measuring the gene transcription changes of the functionalized silver carbon nanotubes in the bacteria, I had a control group and one with the sample of the funtionalized silver carbon nanotubes for each. After it grew, I conducted cDNA and RNA synthesis. Then, I finally tested the different genes of Salmonella and Staphylcoccus in the QRT-PCR, to see the gene expression regulation in regards to the control.