Tomaž Langerholc

Functional cell models of the gut and their applications in food microbiology — A review

Abstract Animal experimentation has a long tradition for risk assessment of new drugs before they reach the clinic. To reduce expensive animal experimentation, attempts have been made to build inexpensive and convenient intestinal functional cell models to study toxicity and bioavailability of new substances along with providing relevant models to study interactions between the host, pathogens and intestinal microflora. We review the available cell lines and models of the intestine and their potential uses. Tumor derived cell lines such as Caco-2, T84 and HT-29 are widely used despite many drawbacks, which are discussed with respect to complexity of the gut, where various cell types interact with commensal microbiota and gut-associated lymphoid tissue. To address this complexity, 3D models of human and animal gut represent a promising in vitro system to mimic in vivo situation without the use of transformed cell lines.

Anti-adhesion activity of thyme (Thymus vulgaris L.) extract, thyme post-distillation waste, and olive (Olea europea L.) leaf extract against Campylobacter jejuni on polystyrene and intestine epithelial cells.

BACKGROUND In order to survive in food-processing environments and cause disease, Campylobacter jejuni requires specific survival mechanisms, such as biofilms, which contribute to its transmission through the food chain to the human host and present a critical form of resistance to a wide variety of antimicrobials. RESULTS Phytochemical analysis of thyme ethanolic extract (TE), thyme post-hydrodistillation residue (TE-R), and olive leaf extract (OE) using high-performance liquid chromatography with photodiode array indicates that the major compounds in TE and TE-R are flavone glucuronides and rosmarinic acid derivatives, and in OE verbascoside, luteolin 7-O-glucoside and oleuroside. TE and TE-R reduced C. jejuni adhesion to abiotic surfaces by up to 30% at 0.2-12.5 µg mL(-1) , with TE-R showing a greater effect. OE from 3.125 to 200 µg mL(-1) reduced C. jejuni adhesion to polystyrene by 10-23%. On the other hand, C. jejuni adhesion to PSI cl1 cells was inhibited by almost 30% over a large concentration range of these extracts. CONCLUSION Our findings suggest that TE, the agro-food waste material TE-R, and the by-product OE represent sources of bioactive phytochemicals that are effective at low concentrations and can be used as therapeutic agents to prevent bacterial adhesion.

Occurrence and toxicity of musks and UV filters in the marine environment

Emerging chemical contaminants in the marine ecosystem represent a threat to the environment and also to human health due to insufficient knowledge about their toxicity and bioaccumulation in the food chain. Consequently, many of them are not regulated. In this review we focus on musks and organic UV filters. For both groups of compounds we describe occurrence in the marine environment, toxic effects identified so far and methods used to identify such effects. The final objective of this work is to identify gaps in the understanding of their toxicology.

Toxic effects of perfluorinated compounds at human cellular level and on a model vertebrate

This work aims at deepening the understanding of the mode of action of some of the most prominent perfluorinated compounds (PFCs) by detecting in a realistic way their effects. To this end, after adjusting the exposure media taking into account the biological model employed and the physico-chemical properties of PFCs, we evaluated the toxic effects of PFOA, PFOS and PFNA in a human macrophage cell line (TLT cells) and in zebrafish embryos. We performed such evaluation on individual compounds and mixtures. Acute toxicity was greater for PFOS in zebrafish; however, it was greater for PFNA in TLT cells. PFNA was also the compound producing the greatest levels of oxidative stress, both in zebrafish and TLT cells. Additionally, in both biological systems, it showed a much stronger effect on mixtures in comparison to the others PFCs tested in this work. Mixture studies in zebrafish showed that acute toxicity depended on the concentration and that the mixture was far more toxic than the individual compounds. This study highlights the importance of studying PFCs in realistic conditions on various biological models.

Alpinia katsumadai Extracts Inhibit Adhesion and Invasion of Campylobacter jejuni in Animal and Human Foetal Small Intestine Cell Lines

Alpinia katsumadai is used in traditional Chinese medicine for abdominal distention, pain, and diarrhoea. Campylobacter jejuni is the most common cause of bacterial food‐borne diarrhoeal illnesses worldwide. Adhesion to gut epithelium is a prerequisite in its pathogenesis. The antimicrobial, cytotoxic, and anti‐adhesive activities of a chemically characterised extract (SEE) and its residual material of hydrodistillation (hdSEE‐R) from A. katsumadai seeds were evaluated against C. jejuni. Minimal inhibitory concentrations for SEE and hdSEE‐R were 0.5 mg/mL and 0.25 mg/mL, respectively, and there was no cytotoxic influence in the anti‐adhesion tests, as these were performed at much lower concentrations of these tested plant extracts. Adhesion of C. jejuni to pig (PSI) and human foetal (H4) small‐intestine cell lines was significantly decreased at lower concentrations (0.2 to 50 µg/mL). In the same concentration range, the invasiveness of C. jejuni in PSI cells was reduced by 45% to 65% when they were treated with SEE or hdSEE‐R. The hdSEE‐R represents a bioactive waste with a high phenolic content and an anti‐adhesive activity against C. jejuni and thus has the potential for use in pharmaceutical and food products. Copyright

Generation of Lactobacillus plantarum strains with improved potential to target gastrointestinal disorders related to sugar malabsorption

Malabsorption of dietary sugars is a common cause of gastrointestinal discomfort, affecting up to one in three people with debilitating symptoms, such as abdominal pain, osmotic diarrhoea, bloating and flatulence. Besides dietary interventions, it has been suggested that ingestion of lactobacilli may alleviate these symptoms. The objectives of this study were to generate strains with improved potential to ameliorate sugar malabsorption related gastrointestinal disorders. Initial selection was made from 183 natural isolates of lactic acid bacteria, on the basis of broad sugar fermentation ability, absence of gas production, gastrointestinal survival and susceptibility to important medical antimicrobials. Two strains of L. plantarum (KR6 and M5) exhibited favourable characteristics for all criteria, and were further optimised through random mutagenesis and selection approaches. Ultraviolet light (UV) exposure resulted in mutants characterized by better survival (for 1.9 log and 1.4 log) in gastrointestinal conditions. Subsequent exposure to ethyl methanesulfonate (EMS) provided mutants with greater tolerance to glucose induced catabolic repression. UV and UV-EMS mutants of L. plantarum M5 showed improved adhesion ability. As a result of this optimisation, L. plantarum MP2026 and L. plantarum MP2420 have been identified as promising candidates for probiotics, intended for alleviation of gastrointestinal discomfort originating from sugar malabsorption.

Cytokine production in vitro and in rat model of colitis in response to Lactobacillus plantarum LS/07

Over the past decade, it has become clear that specific probiotic lactobacilli are valuable in the prevention and treatment of infectious and inflammatory diseases of gastrointestinal tract but their successful application would benefit greatly from a better understanding of the mechanisms of individual strains. Hence, each probiotic strain should be characterized for their immune activity before being proposed for clinical applications. The aim of the study was to characterize the immunomodulatory activity of the strain Lactobacillus (L.) plantarum LS/07 in vitro using functional gut model and to study its anti-inflammatory potential in dextran sulphate sodium (DSS)-induced colitis in rats. We showed that L. plantarum LS/07 induced production of IL-10 in macrophages derived from blood monocytes as well as monocyte/macrophages cell line stimulated indirectly via enterocytes in vitro. In rat model of colitis, L. plantarum LS/07 attenuated the DSS-induced signs of inflammatory process in colon such as weight loss, diarrhoea, infiltration of inflammatory cells associated with decreased colon weight/length ratio, inhibited gut mucosa destruction and depletion of goblet cells. Moreover, the strain increased the concentration of anti-inflammatory cytokine IL-10 in mucosal tissue. In conclusion, the protective effects of L. plantarum LS/07 in the DSS-induced colitis model seem to be related to the stimulation of IL-10 and the restoration of goblet cells and indicate it as a good candidate to prevent and treat diseases associated with inflammation.

Basic microbiology : Instructions on Laboratory Exercises

Bridging the gap between the theoretical and the practical application of what students are learning is the first step towards understanding the central roles of microorganisms in nature and in our daily lives. Our goal for these instructions on laboratory exercises is to provide a bridge that helps our foreign students, who choose the Microbiology course (Erasmus exchange program), to integrate lectures with laboratory experience.