Alla Splichalova

Cross-talk of human gut with bifidobacteria.

The gut constitutes a prominent part of the immune system. Its commensal microflora plays an important role in defense and in tolerance to diet allergens. Disturbances in immune regulations may lead to food allergy. Among commensal bacteria, bifidobacteria are able to induce mechanisms of immune tolerance. Comprehension of their mutual cross-talk with the host is necessary for understanding their role in the diet and in food supplements.

Modulation of natural immunity in the gut by Escherichia coli strain Nissle 1917

The beneficial effect of probiotic Escherichia coli strain Nissle 1917 (EcN) suggests the gut epithelium plays a basic role in immune interactions with bacteria. Contrary to other commensal strains of Escherichia coli, EcN profoundly modulates the gut barrier to elevate its resistance to microbial pathogens. The present review documents the properties of EcN that have led to the protection of gnotobiotic pigs against lethal enteric infections. This effect could be important in light of the growing number of acquired deficiencies that paralyze gut immunity in humans.

Interference of Bifidobacterium choerinum or Escherichia coli Nissle 1917 with Salmonella Typhimurium in gnotobiotic piglets correlates with cytokine patterns in blood and intestine

The colonization, translocation and protective effect of two intestinal bacteria – PR4 (pig commensal strain of Bifidobacterium choerinum) or EcN (probiotic Escherichia coli strain Nissle 1917) – against subsequent infection with a virulent LT2 strain of Salmonella enterica serovar Typhimurium were studied in gnotobiotic pigs after oral association. The clinical state of experimental animals correlated with bacterial translocation and levels of inflammatory cytokines [a chemokine, interleukin (IL)‐8, a proinflammatory cytokine, tumour necrosis factor (TNF)‐α and an anti‐inflammatory cytokine, IL‐10] in plasma and intestinal lavages. Gnotobiotic pigs orally mono‐associated with either PR4 or EcN thrived, and bacteria were not found in their blood. No significant inflammatory cytokine response was observed. Mono‐association with Salmonella caused devastating septicaemia characterized by high levels of IL‐10 and TNF‐α in plasma and TNF‐α in the intestine. Di‐associated gnotobiotic pigs were given PR4 or EcN for 24 h. Subsequently, they were infected orally with Salmonella and euthanized 24 h later. Pigs associated with bifidobacteria before Salmonella infection suffered from severe systemic infection and mounted similar cytokine responses as pigs infected with Salmonella alone. In contrast, EcN interfered with translocation of Salmonella into mesenteric lymph nodes and systemic circulation. Pigs pre‐associated with EcN thrived and their clinical condition correlated with the absence of IL‐10 in their plasma and a decrease of TNF‐α in plasma and ileum.

Systemic and Local Cytokine Response of Young Piglets to Oral Infection with Salmonella enterica Serotype Typhimurium

One-week-old breast-fed miniature piglets were orally infected either with virulent LT2 strain or with a non-virulent SF1591 rough mutant ofSalmonella Typhimurium for 1 d. Both micro-organisms were cultivated from mesenteric lymph nodes but not from the blood of infected piglets. Interleukins (IL) 1β, 8, 18, tumor necrosis factor α (TNF-α) and interferon γ (IFN-γ) were quantified by ELISA in plasma and washes of a terminal part of the small bowel. In plasma, cytokines were mostly missing in noninfected piglets and either missing or low in infected piglets. In the gut of non-infected piglets, IL-1β, IL-8 and IL-18 were detected whereas TNF-α and IFN-γ were mostly missing. IFN-γ levels highly increased (p < 0.05) after infection with nonvirulent salmonellae. The variability of cytokine levels in the gut of suckling piglets is discussed.

Lipopolysaccharide induces inflammatory cytokines in the pig amnion

Inflammatory mediators that are induced by gram-negative bacteria in the course of intrauterine infections threaten successful pregnancy. To compare the effect of two different routes of cytokine induction, bacterial lipopolysaccharide (LPS) was administered in vivo either into the cord vein or into the amniotic cavity of pig fetuses in the second half of gestation for 20 h and cytokines were detected in the amnion.Tumor necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8) were induced in the amniotic epithelium after intra-amniotic but not after intra-venous administration of LPS. The presence of IL-8 was confirmed by RT-PCR. In contrast, transforming growth factor-beta1 (TGF-beta1) was expressed constitutively and was found in all samples of the amniotic epithelium. Amniotic fluid contained only minute levels of TNF-alpha. IL-8 levels in amniotic fluid increased after the treatment with LPS and the highest IL-8 levels were found in dead LPS-treated fetuses.

Local and systemic occurrences of HMGB1 in gnotobiotic piglets infected with E. coli O55 are related to bacterial translocation and inflammatory cytokines

High mobility group box 1 (HMGB1), a nuclear protein, can be secreted by stimulated cells or released from damaged cells. It is recognized as a late mediator of sepsis, but its extracellular occurrence has primarily been studied on the systemic level. Acute and chronic diseases of the gastrointestinal tract, however, have usually been connected with immediate local cell damage. We present local and systemic findings of HMGB1 in Escherichia coli O55-caused infection, in relation to inflammatory cytokines, using a pig gnotobiotic infection model. High levels of HMGB1 were detected in the intestine of those piglets that suffered from infection (fever, anorexia, and diarrhea), as compared to their E. coli 055-infected counterparts that thrived. These local changes were also reflected at the systemic level and related to inflammatory cytokines. Based on our findings of high levels of HMGB1 in the intestinal content of the infection-suffering gnotobiotic piglets, its concurrent presence with inflammatory cytokines, and the published literature, we propose that the detection and analysis of HMGB1 levels in feces can be a non-invasive method of clinical evaluation of the severity of enteric infections.

Innate Immune Response in the Gut against Salmonella - review

Innate immunity is shaped by a complex of redundant and pleiotropic factors that ensure recognition, alert and suppression of pathogens. Innate immune responses in the gut are complicated by the requirement of parallel tolerance to commensal microflora predominating in cell numbers and species. In normal individuals, the intestinal mucosa together with relevant lymph nodes represents a robust barrier against systemic spread of non-typhoid Salmonella. Contemporary insights into these defense mechanisms are reviewed.

Effect of Bacterial Virulence on IL‐18 Expression in the Amnion Infected with Escherichia coli

Problem:  The upregulation of inflammatory substances threatens pregnancy. Interleukin‐18 (IL‐18) is elevated in women who miscarried. The purpose of this study was to develop a pig model of chorioamnionitis to study the effect of bacterial virulence on IL‐18 response in experimentally infected amnion.

Expression of inflammatory markers in pig amnion after intraamniotic infection with nonpathogenic or enteropathogenicEscherichia coli

The pig amnion wasin vivo intraamniotically infected withE. coli for 10 h at 80–85 d of gestation either with the nonpathogenic O86 strain or enteropathogenic O55 strain. TNF-α, IL-10, IL-1β and IFN-γ were determined in amniotic fluids by ELISA, the expression of cytokines and some other inflammatory markers was determined by immunohistochemistry. Intraamniotic infection induced high levels of TNF-α in amniotic fluids which correlated with bacterial virulence whereas IL-10 was induced only by O86. The IL-1β level did not increase significantly and was expressed in all infected membranes. IFN-γ was negligible or absent. TNF-α, IL-12p40, calprotectin, HSP65 and gp91phox were found by immunohistochemistry only in amnion membranes infected with the enteropathogenic strain O55.

A modified MacConkey agar for selective enumeration of necrotoxigenic E. coli O55 and probiotic E. coli Nissle 1917

An agar selective enumeration of necrotoxigenic Escherichia coli O55 (NTEC2) and probiotic E. coli Nissle 1917, using modified MacConkey agar, was developed to study bacterial interference between these E. coli strains in a gnotobiotic piglet model. Replacement of lactose with saccharose in the agar enables the direct visual enumeration of red colonies of E. coli O55 and yellow colonies of E. coli Nissle 1917 that are co-cultured in the same Petri dish. A total of 336 colonies (168 for each color) were subjected to strain-specific PCR identification with LNA probes. Sensitivity, specificity, and positive and negative predictive values were 96.43%, 95.83%, 95.86% and 96.41% respectively in E. coli O55, and 98.21%, 97.02%, 97.06% and 98.19% respectively in E. coli Nissle 1917. Color-based enumeration of both E. coli strains in colonic contents and mesenteric lymph nodes homogenates of gnotobiotic piglets demonstrated the applicability of this method for the gnotobiotic piglet model of enteric diseases.