Nosiheptide, an archetypal member of thiopeptide antibiotics, arises from post-translational modifications of a ribosomally synthesized precursor peptide that contains an N-terminal leader peptide (LP) sequence and a C-terminal core peptide (CP) sequence. The precursor peptide of nosiheptide (NosM) is comprised of a leader peptide with 37 amino acids and a core peptide containing 13 amino acids. Nosiheptide exhibits potent activity against multidrug-resistant Gram-positive bacterial pathogens. Nosiheptide exhibited extremely potent activity against all contemporary MRSA strains tested including multiple drug-resistant clinical isolates, with MIC values ≤ 0.25 mg l(-1). Nosiheptide was also highly active against Enterococcus spp. and the contemporary hypervirulent BI/NAP1/027 strain of Clostridium difficile but was inactive against most Gram-negative strains tested. Time-kill analysis revealed nosiheptide to be rapidly bactericidal against MRSA in a concentration- and time-dependent manner, with a nearly 2-log kill noted at 6 h at 10 × MIC. Nosiheptide also demonstrated in vivo activity in a murine model of MRSA infection, and therefore represents a promising antibiotic for the treatment of serious infections caused by contemporary strains of MRSA. Nosiheptide also has significant anti-hepatitis B virus (anti-HBV) activity in cell culture.