Inflammasomes are cytosolic multiprotein complexes that initiate host defense against bacterial pathogens by activating caspase-1-dependent cytokine secretion and cell death. In mice, specific nucleotide-binding domain, leucine-rich repeat-containing family, apoptosis inhibitory proteins (NAIPs) activate the nucleotide-binding domain, leucine-rich repeat-containing family, CARD domain-containing protein 4 (NLRC4) inflammasome upon sensing components of the type III secretion system (T3SS) and flagellar apparatus. NAIP1 recognizes the T3SS needle protein, NAIP2 recognizes the T3SS inner rod protein, and NAIP5 and NAIP6 recognize flagellin. In contrast, humans encode a single functional NAIP, raising the question of whether human NAIP senses one or multiple bacterial ligands. Previous studies found that human NAIP detects both flagellin and the T3SS needle protein and suggested that the ability to detect both ligands was achieved by multiple isoforms encoded by the single human gene. Here, we show that human NAIP also senses the Typhimurium T3SS inner rod protein PrgJ and that T3SS inner rod proteins from multiple bacterial species are also detected. Furthermore, we show that a single human NAIP isoform is capable of sensing the T3SS inner rod, needle, and flagellin. Our findings indicate that, in contrast to murine NAIPs, promiscuous recognition of multiple bacterial ligands is conferred by a single human NAIP.

dCoxiella burnetii replicates within permissive host cells by employing a Dot/Icm type IV secretion system (T4SS) to translocate effector proteins that direct the formation of a parasitophorous vacuole. C57BL/6 mouse macrophages restrict the intracellular replication of the C. burnetii Nine Mile phase II (NMII) strain. However, eliminating Toll-like receptor 2 (TLR2) permits bacterial replication, indicating that the restriction of bacterial replication is immune mediated. Here, we examined whether additional innate immune pathways are employed by C57BL/6 macrophages to sense and restrict NMII replication. In addition to the known role of TLR2 in detecting and restricting NMII infection, we found that TLR4 also contributes to cytokine responses but is not required to restrict bacterial replication. Furthermore, the TLR signaling adaptors MyD88 and Trif are required for cytokine responses and restricting bacterial replication. The C. burnetii NMII T4SS translocates bacterial products into C57BL/6 macrophages. However, there was little evidence of cytosolic immune sensing of NMII, as there was a lack of inflammasome activation, T4SS-dependent cytokine responses, and robust type I interferon (IFN) production, and these pathways were not required to restrict bacterial replication. Instead, endogenous tumor necrosis factor (TNF) produced upon TLR sensing of C. burnetii NMII was required to control bacterial replication. Therefore, our findings indicate a primary role for TNF produced upon immune detection of C. burnetii NMII by TLRs, rather than cytosolic PRRs, in enabling C57BL/6 macrophages to restrict bacterial replication.T o initiate innate immune defense against bacterial pathogens, infected host cells utilize pattern recognition receptors (PRRs) to detect pathogen-associated molecular patterns (PAMPs) (1-3). Toll-like receptors (TLRs) located at the cell surface and within endosomes detect extracellular PAMPs such as bacterial lipoproteins and lipopolysaccharide (LPS) (4). Downstream of TLRs, the adaptor proteins MyD88 and Trif activate several signaling pathways, including NF-B, mitogen-activated protein kinases (MAPKs), and interferon (IFN) regulatory factor 3 (IRF3), which direct the expression of proinflammatory cytokines and other antimicrobial effectors (4). For intracellular bacterial pathogens, cytosolic PRRs, such as those of the nucleotide binding domain/ leucine-rich repeat (NLR) and RIG-I-like receptor (RLR) families, often are critical for host defense as they respond to PAMPs introduced into the host cell cytosol by bacterial pore-forming toxins or specialized secretion systems (5-8). In addition, cytosolic sensing can lead to the assembly of a multiprotein complex termed the inflammasome, which activates the host proteases caspase-1 and caspase-11, resulting in the release of IL-1 family cytokines and a form of cell death known as pyroptosis (9-16). These innate immune pathways collaborate to restrict intracellular bacterial infection through both cell-intrinsic and -extrinsic mechanisms (17)(18...

The alveolar epithelium is a central signal relay between infected and bystander myeloid cells that orchestrates antibacterial defense.

Early responses mounted by both tissue-resident and recruited innate immune cells are essential for host defense against bacterial pathogens. In particular, both neutrophils and Ly6Chi monocytes are rapidly recruited to sites of infection. While neutrophils and monocytes produce bactericidal molecules, such as reactive nitrogen and oxygen species, both cell types are also capable of synthesizing overlapping sets of cytokines important for host defense. Whether neutrophils and monocytes perform redundant or non-redundant functions in the generation of anti-microbial cytokine responses remains elusive. Here, we sought to define the contributions of neutrophils and Ly6Chi monocytes to cytokine production and host defense during pulmonary infection with Legionella pneumophila, responsible for the severe pneumonia Legionnaires’ disease. We found that both neutrophils and monocytes are critical for host defense against L. pneumophila. Both monocytes and neutrophils contribute to maximal IL-12 and IFNγ responses, and monocytes are also required for TNF production. Moreover, natural killer (NK) cells, NKT cells, and γδ T cells are sources of IFNγ, and monocytes direct IFNγ production by these cell types. Thus, neutrophils and monocytes cooperate in eliciting an optimal cytokine response that promotes effective control of bacterial infection.

Young age is a risk factor for prolonged colonization by common pathogens residing in their upper respiratory tract (URT). Why children present with more persistent colonization is unknown and there is relatively little insight into the host-pathogen interactions that contribute to persistent colonization. To identify factors permissive for persistent colonization during infancy, we utilized an infant mouse model of Streptococcus pneumoniae colonization in which clearance from the mucosal surface of the URT requires many weeks to months. Loss of a single bacterial factor, the pore-forming toxin pneumolysin (Ply), and loss of a single host factor, IL-1α, led to more persistent colonization. Exogenous administration of Ply promoted IL-1 responses and clearance, and intranasal treatment with IL-1α was sufficient to reduce colonization density. Major factors known to affect the duration of natural colonization include host age and pneumococcal capsular serotype. qRT-PCR analysis of the uninfected URT mucosa showed reduced baseline expression of genes involved in IL-1 signaling in infant compared to adult mice. In line with this observation, IL-1 signaling was important in initiating clearance in adult mice but had no effect on early colonization of infant mice. In contrast to the effect of age, isogenic constructs of different capsular serotype showed differences in colonization persistence but induced similar IL-1 responses. Altogether, this work underscores the importance of toxin-induced IL-1α responses in determining the outcome of colonization, clearance versus persistence. Our findings about IL-1 signaling as a function of host age may provide an explanation for the increased susceptibility and more prolonged colonization during early childhood.