Screening of anti-virulence compounds inhibiting type Ⅲ secretion system of Salmonella Typhimurium

Abstract:

[Background] Salmonella enterica is a common foodborne enteric pathogen that infects humans and animals and causes food poisoning, typhoid fever and other diseases. In recent years, antibiotic abuse has aggravated the antibiotic resistance of S. enterica, and it is urgent to develop new anti-infectives. After the contact with the intestinal epithelium, S. enterica can deliver several effector proteins into the host cells through the type Ⅲ secretion system (T3SS). The effector proteins can manipulate many common cellular processes, including host immune responses, cytoskeletal dynamics, vesicle transport, and signaling pathways, which facilitate the infection in the host cells. T3SS, composed of more than 20 proteins, is a protein transport nanomachine ubiquitous in most Gram-negative bacterial pathogens and symbionts. Its structure and function are highly conserved in diverse pathogens. The SctV family protein, one of the most conserved components of T3SS, is located at the core part of T3SS and involved in energy supply and effector secretion of T3SS. The mutation of key residues in SctV results in the failure of S. enterica invasion. [Objective] To obtain the anti-virulence T3SS inhibitors with the SctV family proteins of Salmonella as the targets by virtual screening. [Methods] The in vitro interaction assay, secretion assay, bacterial growth curve assay, and bacterial invasion assay were combined to analyze and validate the inhibitory effect of candidate molecules. [Results] The compounds C4 (pneumocandin B0) and C5 (purpurin) significantly inhibited the secretion of T3SS effectors and further prevented S. Typhimurium from invading NCM460 cells. Instead of binding to the intracellular part of SctV, C4 and C5 employed other mechanisms to inhibit the virulence of S. Typhimurium. [Conclusion] How C4 inhibits the secretion of T3SS effectors remains unclear. C5 may down-regulate the expression of the regulator gene hilD to inhibit the secretion of T3SS effectors and further hinder the invasion of S. Typhimurium into NCM460 cells. This work provides new targets and ideas for the development of novel anti-infectives targeting T3SS. Moreover, it provides a theoretical basis for the optimization and modification of subsequent T3SS anti-virulence inhibitors.