High molecular/low acetylated chitosans reduce adhesion of Campylobacter jejuni to host cells by blocking JlpA.

Infections caused by Campylobacter spp. are a major cause of severe enteritis worldwide. Multifactorial prevention strategies are necessary to reduce the prevalence of Campylobacter. In particular, antiadhesive strategies with specific inhibitors of early host-pathogen interaction are promising approaches to reduce the bacterial load. An in vitro flow cytometric adhesion assay was established to study the influence of carbohydrates on the adhesion of C. jejuni to Caco-2 cells. Chitosans with a high degree of polymerization and low degree of acetylation were identified as potent antiadhesive compounds, exerting significant reduction of C. jejuni adhesion to Caco-2 cells at non-toxic concentrations. Antiadhesive and also anti-invasive effects were verified by confocal laser scanning microscopy. For target identification, C. jejuni adhesins FlpA and JlpA were expressed in Escherichia coli ArcticExpress, and the influence of chitosan on binding to fibronectin and HSP90α, respectively, was investigated. While no effects on FlpA binding were found, a strong inhibition of JlpA-HSP90α binding was observed. To simulate real-life conditions, chicken meat was inoculated with C. jejuni, treated with antiadhesive chitosan, and the bacterial load was quantified. A strong reduction of C. jejuni load was observed. Atomic force microscopy revealed morphological changes of C. jejuni after 2 h of chitosan treatment, indicating disturbance of the cell wall and sacculi formation by electrostatic interaction of positively charged chitosan with the negatively charged cell surface. In conclusion, our data indicate promising antiadhesive and anti-invasive potential of high molecular weight, strongly de-acetylated chitosans for reducing C. jejuni load in livestock and food production. KEY POINTS: • Antiadhesive effects of chitosan with high DP/low DA against C. jejuni to host cells • Specific targeting of JlpA/Hsp90α interaction by chitosan • Meat treatment with chitosan reduces C. jejuni load.

Host-pathogen interaction: Enterobacter cloacae exerts different adhesion and invasion capacities against different host cell types.

New antibiotics are urgently needed due to the huge increase of multidrug-resistant bacteria. The underexplored gram-negative bacterium Enterobacter cloacae is known to cause severe urinary tract and lung infections (UTIs). The pathogenicity of E. cloacae in UTI has only been studied at the bioinformatic level, but until now not within systematic in vitro investigations. The present study assesses different human cell lines for monitoring the early steps of host-pathogen interaction regarding bacterial adhesion to and invasion into different host cells by flow cytometric adhesion assay, classical cell counting assay, gentamicin invasion assay, and confocal laser scanning microscopy. To our knowledge, this is the first report in which E. cloacae has been investigated for its interaction with human bladder, kidney, skin, and lung cell lines under in vitro conditions. Data indicate that E. cloacae exerts strong adhesion to urinary tract (bladder and kidney) and lung cells, a finding which correlates with the clinical relevance of the bacterium for induction of urinary tract and lung infections. Furthermore, E. cloacae ATCC 13047 barely adheres to skin cells (A-431) and shows no relevant interaction with intestinal cells (Caco-2, HT-29), even in the presence of mucin (HT29 MTX). In contrast, invasion assays and confocal laser scanning microscopy demonstrate that E. cloacae internalizes in all tested host cells, but to a different extent. Especially, bladder and kidney cells are being invaded to the highest extent. Defective mutants of fimH and fimA abolished the adhesion of E. cloacae to T24 cells, while csgA deletion had no influence on adhesion. These results indicate that E. cloacae has different pattern for adhesion and invasion depending on the target tissue, which again correlates with the clinical relevance of the pathogen. For detailed investigation of the early host-pathogen interaction T24 bladder cells comprise a suitable assay system for evaluation the bacterial adhesion and invasion.

Campylobacter sp.: Pathogenicity factors and prevention methods-new molecular targets for innovative antivirulence drugs?

Infections caused by bacterial species from the genus Campylobacter are one of the four main causes of strong diarrheal enteritis worldwide. Campylobacteriosis, a typical food-borne disease, can range from mild symptoms to fatal illness. About 550 million people worldwide suffer from campylobacteriosis and lethality is about 33 million p.a. This review summarizes the state of the current knowledge on Campylobacter with focus on its specific virulence factors. Using this knowledge, multifactorial prevention strategies can be implemented to reduce the prevalence of Campylobacter in the food chain. In particular, antiadhesive strategies with specific adhesion inhibitors seem to be a promising concept for reducing Campylobacter bacterial load in poultry production. Antivirulence compounds against bacterial adhesion to and/or invasion into the host cells can open new fields for innovative antibacterial agents. Influencing chemotaxis, biofilm formation, quorum sensing, secretion systems, or toxins by specific inhibitors can help to reduce virulence of the bacterium. In addition, the unusual glycosylation of the bacterium, being a prerequisite for effective phase variation and adaption to different hosts, is yet an unexplored target for combating Campylobacter sp. Plant extracts are widely used remedies in developing countries to combat infections with Campylobacter. Therefore, the present review summarizes the use of natural products against the bacterium in an attempt to stimulate innovative research concepts on the manifold still open questions behind Campylobacter towards improved treatment and sanitation of animal vectors, treatment of infected patients, and new strategies for prevention. KEY POINTS: • Campylobacter sp. is a main cause of strong enteritis worldwide. • Main virulence factors: cytolethal distending toxin, adhesion proteins, invasion machinery. • Strong need for development of antivirulence compounds.