Broadly reactive aptamers targeting bacteria belonging to different genera using a sequential toggle cell-SELEX.

Area of Science:

• Microbiology
• Biotechnology
• Molecular Biology

Background:

• Conventional cell-SELEX typically isolates aptamers targeting a single bacterial species.
• Developing broad-spectrum antimicrobial agents remains a significant challenge in combating diverse bacterial infections.

Purpose of the Study:

• To introduce and validate a novel method, sequential toggle cell-SELEX (STC-SELEX), for isolating DNA aptamers with broad reactivity against multiple bacterial genera.
• To demonstrate the efficacy of STC-SELEX in generating aptamers that bind to a panel of phylogenetically diverse bacteria.

Main Methods:

• Employed a sequential selection process where the DNA library was exposed to different bacterial species (Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Citrobacter freundii, Bacillus subtilis, Staphylococcus epidermidis) in successive rounds.
• The selection process involved a toggling loop, where the library was repeatedly incubated with the first and last bacterial species used in the sequence.
• Isolated aptamers were characterized for their binding affinity and specificity.

Main Results:

• The STC-SELEX method successfully isolated single-stranded DNA aptamers with broad reactivity against the tested bacterial genera.
• The isolated aptamers exhibited dissociation constants in the nanomolar range (9.22–38.5 nM).
• These aptamers demonstrated specificity, showing no significant affinity for bacterial species not included in the selection process.

Conclusions:

• STC-SELEX is a viable strategy for isolating aptamers with broad affinity across different bacterial taxa.
• This approach holds promise for the development of novel diagnostic tools and broad-spectrum therapeutic agents against bacterial infections.