It is under current debate whether mere intestinal carriage of the opportunistic pathogen PA does have any immunopathological impact in health and disease [
20,
29]. In a previous survey, we were able to show that following peroral challenge of clinically uncompromised microbiota-depleted mice with an identical clinical isolate, MDR PA was not only able to stably colonize the intestinal tract at high loads, but also to induce intestinal as well as even extra-intestinal (i.e. splenic) pro-inflammatory immune responses [
29]. Hence, mere intestinal carriage of MDR Gram-negative opportunistic pathogens such as PA by an antibiotic-pretreated, but otherwise asymptomatic individual might result in immunopathological sequelae that may even be aggravated by other comorbidities (and vice versa). This prompted us to unravel the interplay of MDR PA with the commensal intestinal microbiota and host immunity during chronic inflammation of the large intestinal tract here. Following peroral challenge, approximately two-thirds of TLR4 deficient IL10
−/−, but only up to 25% of IL10
−/− counterparts harbored MDR PA in their intestine 2 weeks following peroral challenge. Given that the intestinal microbiota composition determines host resistance against invading (opportunistic) pathogens [
28], we surveyed the intestinal gut microbiota before peroral challenge of mice of either genotype. Cultural analyses revealed that naive, conventionally colonized TLR4 deficient IL10
−/− mice harbored up to two orders of magnitude higher fecal enterobacteria as compared to IL10
−/− controls. In our previous studies we were able to demonstrate that elevated intestinal loads of enterobacterial commensals such as
Escherichia coli were able to override physiological colonization resistance and facilitated murine infection with the enteropathogen
Campylobacter jejuni [
30‐
32]. One also needs to take into consideration that intestinal inflammation per se might facilitate pathogenic colonization [
20,
25,
32,
33]. Before PA challenge histopathological severities of chronic colitis were comparable in either mice, however, and were not further aggravated upon PA challenge until the end of the study. The TLR4 dependent colonization properties of MDR PA observed here are supported by our very recent survey where we could show that healthy microbiota-depleted TLR4 deficient IL10
−/− mice harbored higher PA burdens in their gastrointestinal tract as compared to microbiota-depleted IL10
−/− counterparts [
34]. The PA induced intestinal and splenic pro-inflammatory immune responses do not follow a clear-cut pattern when comparing TLR4 deficient IL10
−/− mice and IL10
−/− controls, however. On one hand, besides higher intestinal opportunistic pathogenic loads, only MDR PA colonized TLR4 deficient IL10
−/− mice displayed increased numbers of innate as well as of adaptive immune cell populations such as macrophages and monocytes as well as of T lymphocytes in their large intestinal mucosa and lamina propria at day 14 p.i. These increases in immune cells were accompanied by more pronounced intestinal secretion of pro-inflammatory cytokines such as IFN-γ and TNF in colon and MLN, respectively. Of note, splenic TNF concentrations were lower in TLR4 deficient IL10
−/− mice as compared to IL10
−/− counterparts at day 14 p.i., which might be explained by more pronounced recruitment of leukocytes to the intestinal tract. One might argue that the more distinct immune responses observed in TLR4 deficient IL10
−/− mice were merely due to better gastrointestinal colonization properties of MDR PA as compared to IL10
−/− control mice. One needs to take into consideration, however, that an (opportunistic) pathogen does not necessarily need to be permanently abundant within the gastrointestinal tract in order to induce immunopathological responses [
22,
35‐
37]. It is rather the initial hit by the bacterial stimulus tipping the balance towards pro-inflammatory immune responses during (opportunistic) pathogen–host interactions [
35]. In fact, despite only sporadic colonization, IL10
−/−, but not TLR4 deficient IL10
−/− mice, on the other side, displayed increased numbers of apoptotic colonic epithelial cells that were paralleled by increased secretion of NO and IFN-γ in colon and MLN, respectively, whereas elevated IL-12p70 levels could be measured in extra-intestinal compartments like the spleen. In support, our very recent study revealed that TLR4 mediated PA induced inflammatory responses in otherwise healthy secondary abiotic IL10
−/− mice without colitis [
34]. Lipid A constitutes a core moiety of LPS derived from Gram-negative bacteria including
Pseudomonas and has been shown to activate NFκB signaling via TLR4 leading to increased pro-inflammatory cytokine expression [
38]. In turn, innate immune cells are recruited to the infection site and further accelerate host immune responses in order to combat the invading (opportunistic) pathogen [
3]. The impact of TLR4 dependent immunopathological sequelae of PA infection is even more intriguing for the following reasons. Firstly, mice are up to 1000 times more resistant to TLR4 ligands such as LPS than humans, and secondly, IL10 gene deficiency renders mice more sensitive to LPS action [
39].