Targeted Suppression of Salmonella enterica Serotype Enteritidis via siRNA-Functionalized Graphene Oxide Nanocarriers Against 16S rRNA Gene

Abstract

Antimicrobial resistance in Salmonella enterica serotype Enteritidis demands new, gene-targeted therapeutics. We engineered a graphene oxide (GO) nanocarrier to deliver small interfering RNA (siRNA) directed against the essential 16S rRNA gene and evaluated its antibacterial efficacy in vitro. GO was synthesized via a modified Hummers’ method, activated with EDC/NHS, and covalently conjugated to anti-16S siRNA; physicochemical features were verified by UV–Vis spectroscopy (π–π* ≈230 nm; n–π* ≈300 nm). Efficient bacterial uptake GO–siRNA was confirmed by fluorescence microscopy. Functional activity was quantified by colony-forming unit (CFU) assays and RT-qPCR. Relative to untreated, GO-only, and siRNA-only controls, GO–siRNA reduced viable CFUs by >70% after 24 h (one-way ANOVA with Tukey’s post hoc, p<0.01) and decreased 16S rRNA levels by ~80% (ΔΔCt), p<0.001. Control groups showed no significant effects, indicating that therapeutic benefit required co-delivery. Results were reproducible across three independent experiments (mean ± SD). Together, these data establish a direct link between target knockdown and growth inhibition and validate GO as a non-viral vector for bacterial gene silencing. This RNA interference–based nanoplatform couples the sequence specificity of siRNA with the delivery capacity of GO, offering a non-antibiotic modality against S. Enteritidis and a potential route to address multidrug resistance. Future work will assess in vivo performance, biosafety, dosing, and durability of suppression, and will explore broader applicability to additional conserved bacterial targets.