Multidrug-resistant bacteria (MRB) and their biofilms, both of which develop high levels of drug tolerance, cause severe threats to global health. This study demonstrates that biocompatible fluorescent silicon-containing nanodots can be a multifunctional platform for simultaneously imaging and eliminating MRB and their biofilms. Ultrasmall epoxy group (oxirane)-functionalized organosilica nanodots (OSiNDs) with a high photoluminescence quantum yield of ≈31% are synthesized via a simple one-step hydrothermal treatment of an epoxy group-containing silane molecule, 3-glycidoxypropyltrimethoxysilane, and an organic dye, rose bengal. The resultant OSiNDs can be employed as a universal imaging reagent for visualizing various bacteria/biofilms, including MRB and their biofilms. Moreover, the epoxy group-terminated OSiNDs can be conjugated with amine-containing reagents only via the simple stirring of the mixtures at an elevated temperature (e.g., 60 °C) for several hours (e.g., 3 h) without the addition of activating reagents. The amine-containing antibiotic vancomycin (Van) can thus be easily conjugated with the OSiNDs, and the obtained OSiNDs-Van can successfully inhibit the growth of MRB and even eliminate their biofilms. Collectively, the present work may give new impetus to the development of novel antibacterial and anti-biofilm agents for overcoming the drug resistance of bacteria.