Development of DNA Aptamers as Therapeutic Tools Against Pseudomonas Aeruginosa

Antibiotic resistance is posing a serious threat to human health. Therefore, innovative therapeutic tools are needed to efficiently treat resistant bacteria. Aptamers are small synthetic oligonucleotides which show high specificity and affinity for a designated target. They evolve from random DNA oligonucleotide pools by means of an in vitro selection process called Systemic Evolution of Ligands by EXponential enrichment (SELEX). Aptamers have multiple applications in the biotechnological field and can be considered as biological drugs to target resistant microorganisms. In this work, we selected DNA aptamers to target the hemophore of the haem assimilation system (HasA) from Pseudomonas aeruginosa. The selection was performed with a DNA library with a random region of 40 nucleotides, and with HasA, which was produced in house. Twelve rounds of SELEX were performed, and the selection was monitored using qPCR and melting curve analysis. From round 1 to 12, the main fluorescence signal peak at 70°C, typical of the library, and then gradually shifted to 84°C. Selected oligonucleotides were sequenced by means of Next-Generation Sequencing, and one single sequence was found to be enriched at 75% in the pool. The most abundant candidates are currently being tested for their binding affinity and specificity. Meanwhile, our group is working on the aptamer selection towards the receptor of the haem assimilation system (HasR). Since iron is an essential micronutrient for P. aeruginosa, as for many bacteria, targeting and potentially blocking elements of the bacterium’s iron acquisition mechanisms, such as HasR and HasA, can put the bacterium at a significant metabolic disadvantage.