In vivo antimicrobial activity of the hybrid peptide H4: a follow-up study Ammar Almaaytah,1 Qosay Albalas,2 Karem H Alzoubi3 1Department of Pharmaceutical Technology, Faculty of Pharmacy, Jordan University of Science and Technology, Irbid, Jordan; 2Department of Medicinal Chemistry and Pharmacognosy, Jordan University of Science and Technology, Irbid, Jordan; 3Department of Clinical Pharmacy, Jordan University of Science and Technology, Irbid, Jordan Background: The consistent upsurge in antimicrobial resistance globally is threatening the world population with the prospect of facing the post-antibiotic era. Dry pipelines and a drastic decrease of antimicrobial drug development accompany this rise in antimicrobial resistance. Governments and health authorities are calling for the development of novel classes of antimicrobial agents that would tackle this problem. Antimicrobial peptides represent a promising group of molecules for antimicrobial drug development due to their potency and rapid mode of killing. However, several obstacles, such as high mammalian cell toxicity and lack of target selectivity, have challenged the development of such agents. Methods: We have recently designed a novel hybrid peptide named H4 that exhibits potent antimicrobial activity and low toxicity in vitro. In order to confirm the potential therapeutic efficacy and safety of the peptide, we evaluated the in vivo activity and toxicity of H4 against Staphylococcus aureus peritonitis mice model. Results: Our results indicate that H4 is highly potent in eradicating bacterial infections in vivo with an effective dose50 value of 4.55±0.89 mg/kg. Additionally, the acute systemic toxicity results indicate that the peptide exhibits a high therapeutic index with no significant negative effects on the function of major body organs. Conclusion: H4 is a novel hybrid peptide with great potential for antimicrobial drug development. Keywords: antimicrobial peptides, hybrid peptides, in vivo activity, hybridization, antimicrobial resistance, drug development