The innate immune response elicited by Group A Streptococcus is highly variable among clinical isolates and correlates with the emm type

Abstract

Group A Streptococcus (GAS) infections remain a significant health care problem due to high morbidity and mortality associated with GAS diseases, along with their increasing worldwide prevalence. Macrophages play a key role in the control and clearance of GAS infections. Moreover, pro-inflammatory cytokines production and GAS persistence and invasion are related. In this study we investigated the correlation between the GAS clinical isolates genotypes, their known clinical history, and their ability to modulate innate immune response. We constituted a collection of 40 independent GAS isolates representative of the emm types currently prevalent in France and responsible for invasive (57.5%) and non-invasive (42.5%) clinical manifestations. We tested phagocytosis and survival in mouse bone marrow-derived macrophages and quantified the pro-inflammatory mediators (IL-6, TNF-α) and type I interferon (INF-β) production. Invasive emm89 isolates were more phagocytosed than their non-invasive counterparts, and emm89 isolates more than the other isolates. Regarding the survival, differences were observed depending on the isolate emm type, but not between invasive and non-invasive isolates within the same emm type. The level of inflammatory mediators produced was also emm type-dependent and mostly invasiveness status independent. Isolates of the emm1 type were able to induce the highest levels of both pro-inflammatory cytokines, whereas emm89 isolates induced the earliest production of IFN-β. Finally, even within emm types, there was a variability of the innate immune responses induced, but survival and inflammatory mediator production were not linked.

Figure 1. Phagocytosis and survival of GAS clinical isolates in BMDMs.

Cells were infected with GAS as described in Material and Methods. (A) Percentage of phagocytosis of all (black bars), invasive (black symbols) and non-invasive (white symbols) GAS strains of different emm types. The results are expressed as the percentage of bacterial CFUs recovered after 30 min post-antibiotics treatment relative to the initial inoculum. (B) Bacterial survival experiments were carried out as described in the Material and Methods and expressed as the percentage of phagocytosed bacteria that survived. (C) Intracellular bacteria were not killed by extracellular antibiotics. Cells were infected with GAS as described and after washing, either medium with antibiotics (ATB) (black) or medium alone (dash) was added to cells. The results represent the mean ± SD of three independent experiments, with significance levels indicated between given isolates from the same emm type or in between emm types (*p≤0.05; **p≤0.01; ***p≤0.005).

Figure 2. GAS clinical isolates induced pro-inflammatory mediator and IFN-β secretion by infected BMDMs.

Graphics represent IL-6, TNF-α and IFN-β quantification in the cell culture supernatant at T2h (A), T4h (B) and T6h (C) after infection by the different emm type isolates. The mean values immune mediator productions induced by all (black bars), invasive (black symbols) and non-invasive (white symbols) of isolates of each emm type are shown. To note, the scales are different, within the IL-6 data in panel A and, within the IFN-β data in panel C. The results represent the mean ± SD of 3 independent experiments; with significance levels indicated between emm1 and other emm types by stars above the corresponding black bar and within emm types by stars above a line overlapping the corresponding the black and white symbols (*p≤0.05; **p≤0.01; ***p≤0.005).