Jianxiong Dou,1,* Weilong Shang,1,* Huagang Peng,1 Yi Yang,1 Juan Chen,2 Yifan Rao,3 Li Tan,1 Zhen Hu,1 Yuting Wang,1 Xiaonan Huang,1 Yuhua Yang,1 Jianghong Wu,1 Qiwen Hu,1 Chuan Xiao,1 Xiancai Rao1,3 

1Department of Microbiology, College of Basic Medical Sciences, Army Medical University, Key Laboratory of Microbial Engineering Under the Educational Committee in Chongqing, Chongqing, 400038, People’s Republic of China; 2Department of Pharmacy, Xinqiao Hospital, Army Medical University, Chongqing, 400037, People’s Republic of China; 3Department of Emergency Medicine, Xinqiao Hospital, Army Medical University, Chongqing, 400037, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Xiancai Rao, Email xcrao@tmmu.edu.cn; raoxiancai@126.com Chuan Xiao, Email xiaochuanapple@tmmu.edu.cn

Introduction: Methicillin-resistant Staphylococcus aureus (MRSA), a notorious multidrug-resistant (MDR) pathogen, frequently resides and proliferates within macrophages, contributing to refractory and recurrent infections. Conventional antibiotics exhibit limited efficacy against intracellular MRSA due to poor cellular penetration.
Methods: Vancomycin (VAN) was encapsulated into membrane vesicles (ΔagrAMVs) derived from the attenuated S. aureus strain RN4220ΔagrA, generating VAN-loaded nanoparticles (ΔagrAMV-VAN). In vitro and in vivo experiments were performed to test the efficacy of ΔagrAMV-VAN in intracellular MRSA clearance.
Results: ΔagrAMV-VAN demonstrated sustained VAN release and efficient extracellular MRSA eradication. Moreover, macrophages actively internalized ΔagrAMV-VAN, leading to VAN accumulation in intracellular compartments and M1 macrophage polarization, which increased MRSA killing. In vivo animal experiments revealed that ΔagrAMV-VAN was safe and effectively eliminated intracellular MRSA in abdominal infections.
Conclusion: Our findings propose a nanotherapeutic strategy that uses bacterial-derived vesicles for targeted antibiotic delivery, overcoming the intrinsic limitations of conventional therapies against intracellular MDR pathogens.

Keywords: MRSA, ΔagrAMV-VAN nanoparticles, macrophage activation, intracellular infections, intracellularly bacterial clearance