Penicillin facilitates the entry of antisense constructs intoStreptococcus mutans

Abstract

Antisense oligonucleotides (AS-ODN) target genes in a sequence-specific manner inhibit gene function and have potential use as antimicrobial agents. Cell barriers, such as peptidoglycan, cell surface proteins and lipopolysaccharide membranes, prevent delivery of AS-ODN into the bacterial cell, limiting their use as an effective treatment option. The β-lactam antibiotic penicillin was examined for its ability to deliver phosphorothioate oligodeoxyribonucleotides (PS-ODNs) and γ(32) P-ODN into Streptococcus mutans OMZ175. Treatment of lag-phase S.mutans OMZ175 cells with penicillin and FBA (PS-ODN targeting the fructose-biphosphate aldolase gene), resulted in prolonged suppression of growth (>24h) and fba expression (656.9±194.4-fold decrease at 5h). Suppression of both cell growth and fba expression corresponded with a greater amount of γ(32) P-ODN becoming cell associated, with a maximum γ(32) P-ODN concentration per cell achieved 5h after penicillin treatment (6.50±1.39×10(8) molecules per CFU). This study confirms that for S.mutans OMZ175, the peptidoglycan layer acts as a major barrier preventing AS-ODN penetration and suggests that the use of agents such as penicillin that interfere with peptidoglycan integrity can significantly increase the uptake of PS-ODN by these cells.