Instituto de Biología Molecular y Celular de Rosario, Consejo Nacional de Investigaciones Científicas y Tecnológicas, Universidad Nacional de Rosario, Rosario, Santa Fe, Argentina.
J Antimicrob Chemother. 2022 Oct 28;77(11):3050-3063. doi: 10.1093/jac/dkac278.
The overprescription and misuse of classical antimicrobial compounds to treat gastrointestinal or systemic salmonellosis have been accelerating the surge of antibiotic-recalcitrant bacterial populations, posing a major public health challenge. Therefore, alternative therapeutic approaches to treat Salmonella infections are urgently required.
To identify and characterize actinobacterial secreted compounds with inhibitory properties against the Salmonella enterica PhoP/PhoQ signal transduction system, crucial for virulence regulation.
The methodology was based on a combination of the measurement of the activity of PhoP/PhoQ-dependent and -independent reporter genes and bioguided assays to screen for bioactive inhibitory metabolites present in culture supernatants obtained from a collection of actinobacterial isolates. Analogues of azomycin were used to analyse the functional groups required for the detected bioactivity and Salmonella mutants and complemented strains helped to dissect the azomycin mechanism of action. The tetrazolium dye colorimetric assay was used to investigate azomycin potential cytotoxicity on cultured macrophages. Salmonella intramacrophage replication capacity upon azomycin treatment was assessed using the gentamicin protection assay.
Sublethal concentrations of azomycin, a nitroheterocyclic compound naturally produced by Streptomyces eurocidicus, repressed the Salmonella PhoP/PhoQ system activity by targeting PhoP and inhibiting its transcriptional activity in a PhoQ- and aspartate phosphorylation-independent manner. Sublethal, non-cytotoxic concentrations of azomycin prevented Salmonella intramacrophage replication.
Azomycin selectively inhibits the activity of the Salmonella virulence regulator PhoP, a new activity described for this nitroheterocyclic compound that can be repurposed to develop novel anti-Salmonella therapeutic approaches.
过度开具和滥用经典抗菌化合物来治疗胃肠道或全身性沙门氏菌病，加速了抗生素耐药细菌群体的激增，对公共健康构成了重大挑战。因此，迫切需要替代治疗方法来治疗沙门氏菌感染。
鉴定和表征放线菌分泌的化合物，这些化合物具有抑制沙门氏菌 enterica PhoP/PhoQ 信号转导系统的活性，该系统对于毒力调节至关重要。
该方法基于测量 PhoP/PhoQ 依赖性和非依赖性报告基因的活性以及生物导向测定的组合，筛选来自放线菌分离物集合的培养上清液中存在的具有生物活性的抑制代谢物。使用氮杂霉素类似物分析检测到的生物活性所需的功能基团，沙门氏菌突变体和互补菌株有助于剖析氮杂霉素的作用机制。使用四唑染料比色法研究氮杂霉素对培养的巨噬细胞的潜在细胞毒性。使用庆大霉素保护测定法评估氮杂霉素处理后沙门氏菌在巨噬细胞内的复制能力。
氮杂霉素是一种天然产生于链霉菌 eurocidicus 的硝基杂环化合物，亚致死浓度的氮杂霉素通过靶向 PhoP 并以 PhoQ 和天冬氨酸磷酸化非依赖性方式抑制其转录活性来抑制沙门氏菌 PhoP/PhoQ 系统的活性。亚致死、非细胞毒性浓度的氮杂霉素可防止沙门氏菌在巨噬细胞内的复制。
氮杂霉素选择性抑制沙门氏菌毒力调节因子 PhoP 的活性，这是该硝基杂环化合物的新活性，可以重新用于开发新型抗沙门氏菌治疗方法。
