Epigallocatechin-3-gallate inhibits bacterial virulence and invasion of host cells

Abstract

Increasing antibiotic resistance and beneficial effects of host microbiota has motivated the search for anti-infective agents that attenuate bacterial virulence rather than growth. For example, we discovered that specific flavonoids such as baicalein and quercetin from traditional medicinal plant extracts could attenuate Salmonella enterica serovar Typhimurium type III protein secretion and invasion of host cells. Here, we show epigallocatechin-3-gallate from green tea extracts also inhibits the activity of S. Typhimurium type III protein effectors and significantly reduces bacterial invasion into host cells. These results reveal additional dietary plant metabolites that can attenuate bacterial virulence and infection of host cells.

Figure 1. Tea extracts inhibit SPI-1 T3SS

(A) Scheme for SopE2-CPG2-HA:Glu-CyFur reporter system. SopE2-CPG2-HA (SPI-1 T3SS) adopts enzymatic activity of carboxypeptidase G2 (CPG2) that when fused to the C-terminus of SopE2, a known S. Typhimurium T3SS bacterial effector, can be secreted and used for monitoring type III protein secretion via cleavage of fluorogenic substrates (Glu-CyFur). (B) Structures of catechin, gallic acid, catechin gallate, and epicatechin gallate. (C) Dose-dependent effect of EGCG, epicatechin gallate, chrysin, INP0007, and baicalein on the levels of SPI-1 T3SS secreted proteins (SipA, SipB, SopB, SipC, and SipD) and flagella components in S. Typhimurium growth media.

Figure 2. Epigallocatechin gallate (EGCG) inhibits SPI-1 T3SS

(A) Dose-dependent activity of EGCG on SPI-1 T3SS (SopE2-CPG2-HA) reporter in S. Typhimurium growth media and cell lysate. (B) Western blot analysis of SopE2-CPG2-HA levels in cell lysate. (C) IC₅₀ value of EGCG measured with the SopE2-CPG2-HA reporter fluorescence assay. Mean ± s.d., n = 3. (D) S. Typhimurim growth curve with 100 µM of EGCG.

Figure 3. EGCG inhibits SPI-1 T3SS-mediated bacterial invasion of epithelial cells

(A) Flow cytometry analysis of EGCG (100 μM) on S. Typhimurium invasion of HeLa cells judged by anti-S. Typhimurium antibody staining. MOI = 10, 30 minute infection. Experiment was done in triplicate and similar results were seen in two independent runs. (B) Quantification of the invasion studies and values are normalized to those of DMSO treated. (C) Immunofluorescence analysis of intracellular S. Typhimurium in HeLa cells with 100 μM EGCG. Scale bar = 10 μm.