Tao Yunzhe, Feng Yongwei, Peng Yu, Wang Xiang, Meng Xiangchuan, Xu Youjun, Han Xiaowan, Zhang Qingyang, Hu Hai-Yu
State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College Beijing 100050 China
School of Pharmaceutical Engineering, Key Laboratory of Structure-Based Drug Design & Discovery (Ministry of Education), Shenyang Pharmaceutical University Shenyang 110016 China.
Chem Sci. 2025 May 31. doi: 10.1039/d5sc01586c.
Cell wall-deficient bacteria (CWDB) are key contributors to antimicrobial resistance (AMR), enabling persistent infections by evading antibiotics through their transition to L-form states. Therefore, molecular tools for detecting L-form conversion and AMR mechanisms are crucial for developing novel strategies against bacterial infections. Herein, we present the development of small-sized, peptidoglycan-specific fluorogenic probes employing a two-step bioorthogonal strategy that enables real-time visualization of CWDB formation. Tz-FL-S rapidly reacts with the novel d-alanine derivative TCO-d-Ala at a rate of (2.61 ± 0.07) × 10 M s, resulting in a 4.9-fold increase in fluorescence intensity. This platform exhibited excellent labeling of peptidoglycan in both Gram-positive and Gram-negative bacteria (signal-to-noise ratio: 15 to 305), effectively capturing the transition from N-form to L-form. Furthermore, we investigated the impact of 14 kinds of antibiotics on L-form conversion and found 13 of them induced CWDB. Besides, we explored the relationship between L-form conversion and AMR. This research enhances our understanding of bacterial adaptations and resistance mechanisms, paving the way for innovative strategies to combat drug-resistant infections.
细胞壁缺陷细菌（CWDB）是导致抗菌药物耐药性（AMR）的关键因素，通过转变为L型状态躲避抗生素，从而引发持续性感染。因此，用于检测L型转变和AMR机制的分子工具对于开发对抗细菌感染的新策略至关重要。在此，我们展示了一种采用两步生物正交策略开发的小型肽聚糖特异性荧光探针，该探针能够实时可视化CWDB的形成。Tz-FL-S与新型d-丙氨酸衍生物TCO-d-Ala以(2.61 ± 0.07) × 10 M s的速率快速反应，导致荧光强度增加4.9倍。该平台在革兰氏阳性菌和革兰氏阴性菌中均表现出对肽聚糖的出色标记（信噪比：15至305），有效捕捉了从N型到L型的转变。此外，我们研究了14种抗生素对L型转变的影响，发现其中13种会诱导CWDB。此外，我们还探索了L型转变与AMR之间的关系。这项研究增进了我们对细菌适应性和耐药机制的理解，为对抗耐药感染的创新策略铺平了道路。
