Individuals with malaria exhibit increased morbidity and mortality when infected with Gram-negative (Gr-) bacteria. To explore this experimentally, we performed co-infection of mice with Plasmodium chabaudi and Citrobacter rodentium, an extracellular Gr- bacterial pathogen that infects the large intestine. While single infections are controlled effectively, co-infection results in enhanced virulence that is characterized by prolonged systemic bacterial persistence and high mortality. Mortality in co-infected mice is associated with disrupted iron metabolism, elevated levels of plasma heme, and increased mitochondrial reactive oxygen species (ROS) production by phagocytes. In addition, iron acquisition by the bacterium plays a key role in pathogenesis because co-infection with a mutant C. rodentium strain lacking a critical iron acquisition pathway does not cause mortality. These results indicate that disrupted iron metabolism may drive mortality during co-infection with C. rodentium and P. chabaudi by both altering host immune responses and facilitating bacterial persistence.

Owing to immature immune systems and impaired colonization resistance mediated by the microbiota, infants are more susceptible to enteric infections. Maternal antibodies can provide immunity, with maternal vaccination offering a protective strategy. We find that oral infection of adult females with the enteric pathogen Citrobacter rodentium protects dams and offspring against oral challenge. Parenteral immunization of dams with heat-inactivated C. rodentium reduces pathogen loads and mortality in offspring but not mothers. IgG, but not IgA or IgM, transferred through breast milk to the intestinal lumen of suckling offspring, coats the pathogen and reduces intestinal colonization. Protective IgG largely recognizes virulence factors encoded within the locus of enterocyte effacement (LEE) pathogenicity island, including the adhesin Intimin and T3SS filament EspA, which are major antigens conferring protection. Thus, pathogen-specific IgG in breast milk induced during maternal infection or immunization protects neonates against infection with an attaching and effacing pathogen.

The microbiota confers host protection by limiting the colonization of pathogenic bacteria in the gut, but the mechanisms by which pathogens overcome colonization resistance remain poorly understood. Using a high-density transposon screen in the enteric pathogen Citrobacter rodentium, we find that the bacterium requires amino acid biosynthesis pathways to colonize conventionally raised mice, but not germ-free or antibiotic-treated animals. These metabolic pathways are induced during infection by the presence of the gut microbiota. Reduced amounts of amino acids are found in the guts of conventionally raised mice compared with germ-free animals. Dietary administration of high protein increases amino acid levels in the gut and promotes pathogen colonization. Thus, the depletion of amino acids by the microbiota limits pathogen colonization, and in turn, the pathogen activates amino acid biosynthesis to expand in the presence of the microbiota.

Despite the accepted health benefits of consuming dietary fiber, little is known about the mechanisms by which fiber deprivation impacts the gut microbiota and alters disease risk. Using a gnotobiotic mouse model, in which animals were colonized with a synthetic human gut microbiota composed of fully sequenced commensal bacteria, we elucidated the functional interactions between dietary fiber, the gut microbiota, and the colonic mucus barrier, which serves as a primary defense against enteric pathogens. We show that during chronic or intermittent dietary fiber deficiency, the gut microbiota resorts to host-secreted mucus glycoproteins as a nutrient source, leading to erosion of the colonic mucus barrier. Dietary fiber deprivation, together with a fiber-deprived, mucus-eroding microbiota, promotes greater epithelial access and lethal colitis by the mucosal pathogen, Citrobacter rodentium. Our work reveals intricate pathways linking diet, the gut microbiome, and intestinal barrier dysfunction, which could be exploited to improve health using dietary therapeutics.

Monocytes play a crucial role in antimicrobial host defence, but the mechanisms by which they protect the host during intestinal infection remains poorly understood. Here we show that depletion of CCR2(+) monocytes results in impaired clearance of the intestinal pathogen Citrobacter rodentium. After infection, the de novo recruited CCR2(+) monocytes give rise to CD11c(+)CD11b(+)F4/80(+)CD103(-) intestinal macrophages (MPs) within the lamina propria. Unlike resident intestinal MPs, de novo differentiated MPs are phenotypically pro-inflammatory and produce robust amounts of IL-1β (interleukin-1β) through the non-canonical caspase-11 inflammasome. Intestinal MPs from infected mice elicit the activation of RORγt(+) group 3 innate lymphoid cells (ILC3) in an IL-1β-dependent manner. Deletion of IL-1β in blood monocytes blunts the production of IL-22 by ILC3 and increases the susceptibility to infection. Collectively, these studies highlight a critical role of de novo differentiated monocyte-derived intestinal MPs in ILC3-mediated host defence against intestinal infection.