Table 1. Examples of human GI organoid cultures for studying host–microbe and host–pathogen interactions.

Microbe(s)/metabolites	Organoid	Method	Main findings	References
Clostridium difficile	Proximal colon organoids	Microinjection	C. difficile reduces MUC2 production and not MUC1, but is not capable of altering host mucus oligosaccharide composition.	[56]
	iPSC derived intestinal organoids, small intestine	Microinjection	C. difficile persistence and toxin production disrupts the epithelial paracellular barrier function.	[57]
Enterohemorrhagic Escherichia coli (EHEC)	Proximal colon organoids	Dissociation –Transwell monolayers	MUC2 and protocadherin 24 (PCDH24) are targeted by EHEC at early stages of infection. EHEC reduces colonic mucus and affects the brush border cytoskeleton in the absence of commensal bacteria.	[58]
Enteroaggregative E. coli (EAEC)	Enteroids	Dissociation – human intestinal monolayers	Differences in the intestinal segments as well as in donors/hosts contribute to unique patters of adherence and infection.	[59]
Enterotoxigenic and Enteropathogenic E. coli	Enteroids, co-cultured with human macrophages	Dissociation –Transwell monolayers	Macrophages enhance barrier function and maturity of enteroid monolayers. Macrophage and enteroid-derived cell co-ordinated response to infections.	[46]
E. coli ECOR2 and K-12 MG1655	Organoids — immature intestinal epithelium	Microinjection	Microbial colonisation of HIOs leads to hypoxia driven responses, increased antimicrobial peptide production and maturation of the mucus layer, and improved barrier function.	[60]
Salmonella Typhimurium	iPSC derived intestinal organoids, small intestine	Microinjection	HIOs effectively model aspects of S. Typhimurium-intestinal epithelium interactions. S. Typhimurium stimulation alters the gene expression patterns in HIOs.	[61]
	iPSC derived intestinal organoids, small intestine	Dissociation –Transwell monolayers	S. Typhimurium targets human-specific pathways by inducing host transcriptional changes (cytoskeletal rearrangement, polarized cytokine release, and hampering host immune defense system).	[62]
	iPSC derived intestinal organoids, colon and ileum	Microinjection	The IL-22 pathway facilitates control of microbial infection of the human intestinal epithelium, involving enhanced phagolysosomal fusion.	[63]
	Apical-out vs basal-out enteroids	Infection via media	Bacteria can induce actin ruffles to invade the human IECs and preferentially invade apical surfaces.	[45]
Listeria monocytogenes	Apical-out vs basal-out enteroids	Infection via media	L. monocytogenes invades the human IECs via attachment to basal receptors.	[45]
Human noroviruses (HuNoVs)	iPSC derived and IBD patient derived intestinal organoids, small intestine	Dissociation monolayers	Bile is required for strain-dependent HuNoV replication. Lack of appropriate histoblood group antigen expression in intestinal cells restricts virus replication.	[64]
Shiga toxin (Stx) producing E. coli O157:H7 and commensal E. Coli	iPSC derived intestinal organoids	Microinjection	Commensal E. coli remained within the lumen and did not cause damage. O157:H7 induced loss of actin and epithelial integrity and increased reactive oxygen species production. Both commensal and O157:H7 up-regulated genes associated with gastrointestinal maturation. O157:H7 up-regulated inflammatory responses and resulted in recruitment of human neutrophils.	[65]
Enteroviruses (Echovirus 11 (E11), Enterovirus 71 (EV71)) and Coxsackievirus B (CVB),	Stem cell-derived organoids from the small intestine		Virus-specific activation of antiviral and inflammatory signalling pathways in response to infection. Enteroviruses infect specific cell populations in the human intestine.	[66]
Helicobacter pylori	Gastric organoids	Microinjection	H. pylori induces inflammatory response. IL8 expression was substantially higher in gland-type organoids than in pit-type organoids.	[67]
Butyrate	Foetal small intestinal organoids		Butyrate affects cytokine responses in epithelial cells and enhances maturation markers and RA production.	[68]
Indoleacrylic Acid (IA) produced by Peptostreptococcus Species	Colonoids		IA promotes intestinal epithelial barrier function and mitigates inflammatory responses. IBD patient microbiota show diminished capability to utilise mucins and metabolise tryptophan.	[69]