Health-care-associated infections contribute to increased mortality among hospitalized patients worldwide. The most common pathogens include fungi and antibiotic-resistant bacteria. Treatment and prevention of these infections with available antibiotics and hospital sanitation are challenging, and there are currently no approved vaccines for these pathogens. In this study, Yan et al. developed a protein-free vaccine that protects against infection with bacterial and fungal nosocomial pathogens. The vaccine consisted of a combination of three components: Al(OH)3, monophosphoryl lipid A and fungal mannan. The tripartite vaccine conferred protection against bloodstream infection and pneumonia in mice by methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecalis, extended-spectrum β-lactamase-expressing Escherichia coli, and carbapenem-resistant strains of Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. The vaccine was also protective against the fungal pathogens Rhizopus delemar and Candida albicans. Protection lasted up to 28 days after a single dose, activating innate, rather than adaptive, immunity. The vaccine induced epigenetic changes in macrophages that modulated phagocytosis and inflammatory response to infection.