Lysins represent a novel class of anti-infectives derived from bacteriophage. Lysins are bacterial cell-wall hydrolytic enzymes that selectively and rapidly kill (≥3 log c.f.u. in 30 min) specific Gram-positive bacteria providing a targeted therapeutic approach with minimal impact on unrelated commensal flora. The potential for bacterial resistance to lysins is considered low due to targeting of highly conserved peptidoglycan components. Through cutting-edge genetic engineering, lysins can be assembled into large libraries of anti-infective agents tailored to any bacterium of interest including drug-resistant Gram-positive pathogens such as meticillin- and vancomycin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus faecalis and Enterococcus faecium, and penicillin-resistant Streptococcus pneumoniae. Lysins can eliminate bacteria systemically and topically from mucosal surfaces and biofilms, as evidenced by experimental models of sepsis, endocarditis, pneumonia, meningitis, and nasopharyngeal, skin and vaginal decolonization. Furthermore, lysins can act synergistically with antibiotics and, in the process, resensitize bacteria to non-susceptible antibiotics. Clinical trials are being prepared to assess the safety and pharmacokinetic properties of lysins in humans.