Bacillus cereus: Epidemiology, Virulence Factors, and Host-Pathogen Interactions

Bacillus cereus is an important human pathogen, and new findings have expanded our understanding of how this bacterium causes disease. B. cereus Hemolysin BL (HBL) and nonhemolytic enterotoxin (NHE) induce membrane pore formation, leading to activation of the NLRP3 inflammasome, systemic inflammation, and death. Lipopolysaccharide-induced tumor necrosis factor (TNF)-α factor (LITAF) and cell death-inducing P53 target 1 (CDIP1) are bona fide mammalian surface receptors of HBL. These newly identified toxin receptors and the NLRP3 inflammasome represent unique targets for potential future therapies against severe B. cereus infections. The toxin-producing bacterium Bacillus cereus is an important and neglected human pathogen and a common cause of food poisoning. Several toxins have been implicated in disease, including the pore-forming toxins hemolysin BL (HBL) and nonhemolytic enterotoxin (NHE). Recent work revealed that HBL binds to the mammalian surface receptors LITAF and CDIP1 and that both HBL and NHE induce potassium efflux and activate the NLRP3 inflammasome, leading to pyroptosis. These mammalian receptors, in part, contribute to inflammation and pathology. Other putative virulence factors of B. cereus include cytotoxin K, cereulide, metalloproteases, sphingomyelinase, and phospholipases. In this review, we highlight the latest progress in our understanding of B. cereus biology, epidemiology, and pathogenesis, and discuss potential new directions for research in this field.S.M.M. is supported by the Australian National University and the National Health and Medical Research Council of Australia under Project Grants (APP1141504, APP1146864, APP1162103 and APP1163358) and the R.D. Wright Career Development Fellowship (APP1162025). D.E.T. and S.M.M. are supported by Therapeutic Innovation Australia. D.E.T and A.M. are supported by The Gretel and Gordon Bootes Medical Research Foundation. A.M. is supported by a John Curtin School of Medical Research International PhD scholarshi

Bacillus cereus non-haemolytic enterotoxin activates the NLRP3 inflammasome

Inflammasomes are important for host defence against pathogens and homeostasis with commensal microbes. Here, we show non-haemolytic enterotoxin (NHE) from the neglected human foodborne pathogen Bacillus cereus is an activator of the NLRP3 inflammasome and pyroptosis. NHE is a non-redundant toxin to haemolysin BL (HBL) despite having a similar mechanism of action. Via a putative transmembrane region, subunit C of NHE initiates binding to the plasma membrane, leading to the recruitment of subunit B and subunit A, thus forming a tripartite lytic pore that is permissive to efflux of potassium. NHE mediates killing of cells from multiple lineages and hosts, highlighting a versatile functional repertoire in different host species. These data indicate that NHE and HBL operate synergistically to induce inflammation and show that multiple virulence factors from the same pathogen with conserved function and mechanism of action can be exploited for sensing by a single inflammasome

Microbial activators of the inflammasome

Innate immune recognition of microbial components serves as a cornerstone in mediating an effective immune response. Innate immune sensors and inflammasomes detect both intracellular and extracellular microorganisms. The inflammasome is an intracellular signalling complex comprising of a sensor, an adaptor protein ASC (known as apoptosis-associated speck-like protein containing a caspase activation and recruitment domain) and the cysteine protease caspase-1. Inflammasomes regulate secretion of the pro-inflammatory cytokines, IL-1β and IL-18, and induction of a cell death pathway known as pyroptosis. Certain intracellular bacteria require cytosolic access to activate the inflammasome, however, how extracellular bacteria are sensed by the inflammasome in the cytoplasm remains largely unclear. To understand the innate immune recognition of extracellular bacteria, a panel of clinically important intracellular and extracellular bacteria were analysed. This analysis led to the identification of an unknown secreted factor from the foodborne bacterium Bacillus cereus that activated the inflammasome without gaining cytosolic access. The tripartite enterotoxin called haemolysin BL (HBL) was identified as the novel activator of the NLRP3 inflammasome. I further identified another tripartite toxin called non-haemolytic enterotoxin (NHE), which also activated NLRP3 to induce inflammation and cell death. Mechanistically, both multi-component toxins assembled in a specific and linear order on the mammalian plasma membrane to form a lytic pore, which induces potassium efflux. Remarkably, HBL and NHE operated synergistically to drive inflammation in a mouse model of B. cereus infection. Administration of a small molecule NLRP3 inhibitor MCC950 inhibited inflammation induced by HBL and NHE in vivo and rescued mice from B. cereus-induced lethality. These data showcase the ability of a single inflammasome sensor to detect structurally and functionally similar toxins from the same bacterium. Overall, this research identified novel activators of the inflammasome and their molecular mechanisms in activating inflammation and cell death responses. Understanding the molecular basis of host-pathogen interactions has the potential to contribute therapies which target microbial virulence factors and/or the immune system in the treatment of infectious diseases

Molecular mechanisms of inflammasome signaling

The inflammasome is a macromolecular protein complex that mediates proteolytic cleavage of pro-IL-1β and -IL-18 and induces cell death in the form of pyroptosis. Certain nucleotide-binding oligomerization domain-like receptors (NLRs), absent in melanoma 2 (AIM2)-like receptors (ALRs), or tripartite motif (TRIM) family receptors trigger the assembly of an inflammasome in response to pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs). Recent studies have revealed a multitude of host components and signals that are essential for controlling canonical and noncanonical inflammasome activation and pyroptosis. These include pore-forming gasdermin proteins, the never in mitosis A-related kinase 7 (NEK7), IFN-inducible proteins (IFIs), reactive oxygen species (ROS), autophagy, potassium efflux, mitochondrial perturbations, and microbial metabolites. Here, we provide a comprehensive overview of the molecular and signaling mechanisms that provide stringent regulation over the activation and effector functions of the inflammasome.This work was supported by The John Curtin School of Medical Research Ph.D. Scholarship from the Australian National University (to A.M.), a grant from the Gretel and Gordon Bootes Foundation (to S.M.M.), and the R. G. Menzies Early Career Fellowship from the National Health and Medical Research Council of Australia (to S.M.M.

Cytosolic Recognition of Microbes and Pathogens: Inflammasomes in Action

Infection is a dynamic biological process underpinned by a complex interplay between the pathogen and the host. Microbes from all domains of life, including bacteria, viruses, fungi, and protozoan parasites, have the capacity to cause infection. Infection is sensed by the host, which often leads to activation of the inflammasome, a cytosolic macromolecular signaling platform that mediates the release of the proinflammatory cytokines interleukin-1β (IL-1β) and IL-18 and cleavage of the pore-forming protein gasdermin D, leading to pyroptosis. Host-mediated sensing of the infection occurs when pathogens inject or carry pathogen-associated molecular patterns (PAMPs) into the cytoplasm or induce damage that causes cytosolic liberation of danger-associated molecular patterns (DAMPs) in the host cell. Recognition of PAMPs and DAMPs by inflammasome sensors, including NLRP1, NLRP3, NLRC4, NAIP, AIM2, and Pyrin, initiates a cascade of events that culminate in inflammation and cell death. However, pathogens can deploy virulence factors capable of minimizing or evading host detection. This review presents a comprehensive overview of the mechanisms of microbe-induced activation of the inflammasome and the functional consequences of inflammasome activation in infectious diseases. We also explore the microbial strategies used in the evasion of inflammasome sensing at the host-microbe interaction interface.A.M. is supported by a scholarship from The John Curtin School of Medical Research, The Australian National University. S.M.M. is supported by the Australian National University Futures Award, The Gretel and Gordon Bootes Medical Research Foundation, the National Health and Medical Research Council of Australia (under project grants APP1141504 and APP1146864), and the R. G. Menzies Early Career Fellowship (grant APP1091544)

A multicomponent toxin from Bacillus cereus incites inflammation and shapes host outcome via the NLRP3 inflammasome

Host recognition of microbial components is essential in mediating an effective immune response. Cytosolic bacteria must secure entry into the host cytoplasm to facilitate replication and, in doing so, liberate microbial ligands that activate cytosolic innate immune sensors and the inflammasome. Here, we identified a multicomponent enterotoxin, haemolysin BL (HBL), that engages activation of the inflammasome. This toxin is highly conserved among the human pathogen Bacillus cereus. The three subunits of HBL bind to the cell membrane in a linear order, forming a lytic pore and inducing activation of the NLRP3 inflammasome, secretion of interleukin-1β and interleukin-18, and pyroptosis. Mechanistically, the HBL-induced pore results in the efflux of potassium and triggers the activation of the NLRP3 inflammasome. Furthermore, HBL-producing B. cereus induces rapid inflammasome-mediated mortality. Pharmacological inhibition of the NLRP3 inflammasome using MCC950 prevents B. cereus-induced lethality. Overall, our results reveal that cytosolic sensing of a toxin is central to the innate immune recognition of infection. Therapeutic modulation of this pathway enhances host protection against deadly bacterial infections