Fate of Streptococcus pyogenes and epithelial cells following internalization

Abstract

The fate of GAS and epithelial cells following internalization was determined in this study. HEp-2 cells harbouring intracellular bacteria were treated with antibiotics to kill extracellular adherent bacteria, washed, and the fate of bacteria and epithelial cells was assessed up to 24 h post-infection. In the absence of antibiotics, massive bacterial growth was apparent in the cell medium, accompanied by extensive cell death, suggesting that intracellular bacteria had multiplied and damaged the monolayer. Addition of the internalization inhibitor, cytochalasin D, either pre-or post-internalization prevented bacterial growth and cell injury; post-internalization treatment with chloramphenicol had the same effect. Analysis of three apoptotic markers in HEp-2 cells -chromatin condensation, DNA laddering and translocation of phosphatidylserine onto the cell-surface membrane -indicated that HEp-2 cells underwent apoptosis. Taken together, the data presented here support a model in which intenalized bacteria can induce their own externalization into the medium by a process that requires both an intact host-cell cytoskeleton and de novo synthesis of bacterial proteins. Concomitantly, intracellular and, apparently, extracellular free bacteria induce apoptosis through their cytotoxic activity, and release essential nutrients required for their growth.

Lactobacilli Reduce Cell Cytotoxicity Caused by Streptococcus pyogenes by Producing Lactic Acid That Degrades the Toxic Component Lipoteichoic Acid

“…S. pyogenes strain S165 did not invade the pharyngeal epithelial cell line FaDu, as determined by plating viable counts after killing extracellular bacteria with gentamicin (data not shown). This is in accordance with previous reports showing that GAS invasion of eukaryotic cells correlates with the isolation site (17,26,53) and that invasive blood isolates invade poorly (30). Taken together, these results indicate that lactic acid treatment, but not heat killing, abolishes the cytotoxic effect of GAS.…”

“…A pathogenicity feature of GAS is the ability to induce cytotoxicity in human cells (33, 37), which facilitates bacterial entry into deeper tissues of the body (2, 9). Invasion of host cells (17,26,53), as well as many virulence factors of GAS, such as the surface component lipoteichoic acid (LTA) (12, 19-22, 49, 51) and the toxins streptolysin O and streptolysin S (11,29,45), is reported to induce host cell damage. In addition, pathogeninduced apoptosis and necrosis lead to shedding of the cell surface complement regulator CD46 (14, 25).…”

SpyA, a C3-Like ADP-Ribosyltransferase, Contributes to Virulence in a Mouse Subcutaneous Model of Streptococcus pyogenes Infection

“…Although invasion of epithelial cells is usually assumed to represent increased virulence, the role of intracellular streptococci during infection remains enigmatic. A number of reports demonstrate that GAS can be found intracellularly within the tonsils of patients with recurrent tonsillitis (48,50) and that bacteria can resurge from host cells (38,47 (4,7,28,54), and that internalization prevents passage of bacteria through disrupted tight junctions into deeper tissue (12). Our data suggest that a reduction in intracellular bacteria in vitro may be associated with the decrease in chemokine production seen during in vivo infection with wild-type GAS and consequently increased virulence.…”

“…It has been demonstrated that GAS can enter epithelial cells (24,30,34,38,42,54), so we investigated whether or not SpyA affects bacterial entry into epithelial monolayers in vitro. HeLa cells were infected with wild-type or spyA mutant bacteria at a multiplicity of infection (MOI) of ϳ10:1, and association and internalization into these cells was measured.…”

“…An increase in the incidence of invasive infections beginning in the latter part of the 20th century has resulted in increased research into and identification of the organism's numerous virulence determinants. Though typically considered an extracellular pathogen, GAS have been shown to enter and survive intracellularly in macrophages (60), neutrophils (40,41), and epithelial cells (24,30,34,38,42,54).…”