stg fimbrial operon from S. Typhi STH2370 contributes to association and cell disruption of epithelial and macrophage-like cells

Abstract

Background: Salmonella enterica serovar Typhi (S. Typhi) stg operon, encoding a chaperone/usher fimbria (CU), contributes to an increased adherence to human epithelial cells. However, one report suggests that the presence of the Stg fimbria impairs the monocyte--bacteria association, as deduced by the lower level of invasion to macrophage-like cells observed when the stg fimbrial cluster was overexpressed. Nevertheless, since other CU fimbrial structures increase the entry of S. Typhi into macrophages, and considering that transcriptomic analyses revealed that stg operon is indeed expressed in macrophages, we reassessed the role of the stg operon in the interaction between S. Typhi strain STH2370 and human cells, including macrophage-like cells and mononuclear cells directly taken from human peripheral blood.

Results: We compared S. Typhi STH2370 WT, a Chilean clinical strain, and the S. Typhi STH2370 Δstg mutant with respect to association and invasion using epithelial and macrophage-like cells. We observed that deletion of stg operon reduced the association and invasion of S. Typhi, in both cellular types. The presence of the cloned stg operon restored the WT phenotype in all the cases. Moreover, we compared Salmonella enterica sv. Typhimurium 14028s (S. Typhimurium, a serovar lacking stg operon) and S. Typhimurium heterologously expressing S. Typhi stg. We found that the latter presents an increased cell disruption of polarized epithelial cells and an increased association in both epithelial and macrophage-like cells.

Conclusions: S. Typhi stg operon encodes a functional adhesin that participates in the interaction bacteria-eukaryotic cells, including epithelial cells and macrophages-like cells. The phenotypes associated to stg operon include increased association and consequent invasion in bacteria-eukaryotic cells, and cell disruption.

Figure 1

Association and invasion of HEp-2 epithelial cells. The strains used include S. Typhi STH2370 WT (black), S. Typhi STH2370 ΔstgABCD::FRT (Δstg) (white), S. Typhi STH2370 ΔstgABCD::FRT/pSstg (Δstg/pSstg) (dark grey), and S. Typhi STH2370 ΔstgABCD::FRT/pSU19 (Δstg/pSU19) (light grey) (a); and S. Typhimurium 14028s WT (black), S. Typhimurium 14028s WT/pSstg (white), and S. Typhimurium 14028s/pSU19 (dark grey) (b). The figure shows values expressed as the mean ± standard deviation of three full biological replicates, each time in technical triplicate. *p < 0.05 (Student’s-test) compared with the WT in the corresponding group.

Figure 2

Cell permeability assay of S. Typhi STH2370 WT, and S. Typhimurium 14028s WT and derivatives, through H-T29 human cell line monolayers. The arrow indicates the time at which gentamicin was added. The experiments were performed in three full biological replicates, each time in technical triplicate. The values are expressed as the mean ± standard deviation of three independent experiments. *p < 0.005 (Student’s-test).

Figure 3

Association and invasion of monocytes U937 (a) or mononuclear cells directly extracted from human blood (b). The strains used include S. Typhi STH2370 WT (black), S. Typhi STH2370 ΔstgABCD::FRT (Δstg) (white), S. Typhi STH2370 ΔstgABCD::FRT/pSstg (Δstg/pSstg) (fark grey), and S. Typhi STH2370 ΔstgABCD::FRT/pSU19 (Δstg/pSU19) (light grey) (a); and S. Typhimurium 14028s WT (black), S. Typhimurium 14028s WT/pSstg (white), and S. Typhimurium 14028s/pSU19 (dark grey) (b). The figure shows values expressed as the mean ± standard deviation of three full biological replicates, each time in technical triplicate. *p < 0.05 (Student’s-test) compared with the WT in the corresponding group.