Enteroaggregative Escherichia coli (EAEC) is an important diarrheal pathogen and a cause of both acute and chronic diarrhea. It is a common cause of pediatric bacterial diarrhea in developing countries. Despite its discovery in 1987, the intestinal tropism of the pathogen remains unknown. Cell lines used to study EAEC adherence include the HEp-2, T-84, and Caco-2 lines, but they exhibit abnormal metabolism and large variations in gene expression. Animal models either do not faithfully manifest human clinical symptoms or are cumbersome and expensive. Using human intestinal enteroids derived from all four segments of the human intestine, we find that EAEC demonstrates aggregative adherence to duodenal and ileal enteroids, with donor-driven differences driving a sheet-like and layered pattern. This contrasts with the colon, where segment-specific tropisms yielded a mesh-like adherence pattern dominated by interconnecting filaments. Very little to no aggregative adherence to jejunal enteroids was observed, regardless of the strain or donor, in contrast to a strong duodenal association across all donors and strains. These unique patterns of intestinal segment- or donor-specific adherence, but not the overall numbers of associated bacteria, were dependent on the major subunit protein of aggregative adherence fimbriae II (AafA), implying that the morphology of adherent clusters and the overall intestinal cell association of EAEC occur by different mechanisms. Our results suggest that we must give serious consideration to inter- and intrapatient variations in what is arguably the first step in pathogenesis, that of adherence, when considering the clinical manifestation of these infections.

The enteric bacterium and intracellular human pathogen Shigella causes hundreds of millions of cases of the diarrheal disease shigellosis per year worldwide. Shigella is acquired by ingestion of contaminated food or water; upon reaching the colon, the bacteria invade colonic epithelial cells, replicate intracellularly, spread to adjacent cells, and provoke an intense inflammatory response. There is no animal model that faithfully recapitulates human disease; thus, cultured cells have been used to model Shigella pathogenesis. However, the use of transformed cells in culture does not provide the same environment to the bacteria as the normal human intestinal epithelium. Recent advances in tissue culture now enable the cultivation of human intestinal enteroids (HIEs), which are derived from human intestinal stem cells, grown ex vivo, and then differentiated into "mini-intestines." Here, we demonstrate that HIEs can be used to model Shigella pathogenesis. We show that Shigella flexneri invades polarized HIE monolayers preferentially via the basolateral surface. After S. flexneri invades HIE monolayers, S. flexneri replicates within HIE cells and forms actin tails. S. flexneri also increases the expression of HIE proinflammatory signals and the amino acid transporter SLC7A5. Finally, we demonstrate that disruption of HIE tight junctions enables S. flexneri invasion via the apical surface.

Infection with the respiratory syncytial virus (RSV) early in life is essentially guaranteed and can lead to severe disease. Most RSV studies have involved either of two historic RSV/A strains infecting one of two cell lines, HEp-2 or A549 cells.

The translocation of bacteria across the intestinal epithelium of immunocompromised patients can lead to bacteremia and life-threatening sepsis. Extraintestinal pathogenic (ExPEC), so named because this pathotype infects tissues distal to the intestinal tract, is a frequent cause of such infections, is often multidrug resistant, and chronically colonizes a sizable portion of the healthy population. Although several virulence factors and their roles in pathogenesis are well described for ExPEC strains that cause urinary tract infections and meningitis, they have not been linked to translocation through intestinal barriers, a fundamentally distant yet important clinical phenomenon. Using untransformed human intestinal enteroids and transformed Caco-2 cells, we report that ExPEC strain CP9 binds to and invades the intestinal epithelium. ExPEC harboring a deletion of the gene encoding the mannose-binding type 1 pilus tip protein FimH demonstrated reduced binding and invasion compared to strains lacking known virulence factors. Furthermore, in a murine model of chemotherapy-induced translocation, ExPEC lacking colonized at levels comparable to that of the wild type but demonstrated a statistically significant reduction in translocation to the kidneys, spleen, and lungs. Collectively, this study indicates that FimH is important for ExPEC translocation, suggesting that the type 1 pilus is a therapeutic target for the prevention of this process. Our study also highlights the use of human intestinal enteroids in the study of enteric diseases.

The human gastrointestinal mucosal surface consists of a eukaryotic epithelium, a prokaryotic microbiota, and a carbohydrate-rich interface that separates them. In the gastrointestinal tract, the interaction of bacteriophages (phages) and their prokaryotic hosts influences the health of the mammalian host, especially colonization with invasive pathobionts. Antibiotics may be used, but they also kill protective commensals. Here, we report a novel phage whose lytic cycle is enhanced in intestinal environments. The tail fiber gene, whose protein product binds human heparan sulfated proteoglycans and localizes the phage to the epithelial cell surface, positions it near its bacterial host, a type of locational targeting mechanism. This finding offers the prospect of developing mucosal targeting phage to selectively remove invasive pathobiont species from mucosal surfaces. IMPORTANCE Invasive pathobionts or microbes capable of causing disease can reside deep within the mucosal epithelium of our gastrointestinal tract. Targeted effective antibacterial therapies are needed to combat these disease-causing organisms, many of which may be multidrug resistant. Here, we isolated a lytic bacteriophage (phage) that can localize to the epithelial surface by binding heparan sulfated glycans, positioning it near its host, Escherichia coli. This targeted therapy can be used to selectively remove invasive pathobionts from the gastrointestinal tract, preventing the development of disease.