Final Activity Report Summary - EU'S IMO-TRAIN (The European Union's "Infection models beyond cell culture" training) Despite significant research progress infectious diseases are still the major burden for human health, even in industrialised countries. Diseases such as urinary tract infections, which are usually not life-threatening, yet they affect the life quality and productivity of the individual and cause increased health care costs, are still prevalent. Due to the aging of the population and the successive increased use of medical services, some infections spread readily and novel diseases emerge. Disease is, in such circumstances, commonly caused by micro-organisms of low virulence potential, i.e. prevalent commensal organisms of humans. To analyse such infections with a complex interplay between the host and the microbe, disease mechanisms in the full biological context using global expression analysis, advanced bioinformatic analysis and in vivo infection models were addressed. In this early stage research training on infection models, the actual spectrum of infectious diseases relevant for European health including global challenges was studied. The relevant human pathogens, the gram-negative bacteria Escherichia coli, helicobacter pylori and salmonella enterica, the gram-positive bacteria streptococcus pneumoniae and the protozoa plasmodium falciparum were investigated. Through a recently developed technique of real-time imaging of bacterial proliferation using constitutively expressed green fluorescent protein we could investigate kidney infection with the urinary tract pathogen Escherichia coli in real time in live animals. Studies of bacterial proliferation demonstrated the contribution of virulence factors, such as haemolysin, and tissue responses, such as a novel innate vascular response to the mucosal infection in the live animal. On the other hand, streptococcus pneumoniae (pneumococcus) was an organism with low virulence potential, but also a major cause of community-acquired pneumonia. As part of the innate immune response, neutrophils produced Neutrophil extracellular traps (NETs) composed of Deoxyribonucleic acid (DNA) and DNA-binding proteins that bound, disarmed and killed bacteria. Pneumococci were not killed in NETs, but could degrade them with the help of a surface-located DNase and could counteract the action of neutrophils with an antiphagocytic capsule. Another organism with low virulence potential causing cancer was h. pylori. A consistent association between h. pylori pathogenicity-associated factor dupA and induction of the proinflammatory cytokine IL-8 in vitro, the development of Duodenal ulcer (DU) and Gastric cancer (GC) across the investigated ethnic groups was not present. Infections with bacteria of low virulence potential can be often correlated with the capacity of the microbes to form multicellular aggregates commonly called biofilms. Global expression analysis revealed feed-forward loop regulation of biofilm formation by the global regulator CsrA in E. coli. Structural components of biofilms, as well as enzymes producing the secondary messenger cyclic di-GMP, were negatively regulated by CsrA. Biofilm formation was also considered a colonisation factor. In vitro, the probiotic strain escherichia coli Nissle 1917 produced the biofilm-matrix component cellulose. Our in vivo and in vitro studies showed that cellulose was required for adherence in vitro to the epithelial cell line and in vivo in the mouse illeal loop model. Cellulose production of Nissle 1917 prevented induction of the proinflammatory cytokine IL-8. Cyclic di-GMP was a novel secondary messenger in bacteria. We could show that c-di-GMP affected the invasion of s. typhimurium in epithelial cell lines. In addition, the pathogen plasmodium falciparum caused human malaria. Repetitive interspersed family (RIF) genes, the largest multi-copy gene family of p. falciparum protected the parasite from the immune defences of the host. Combined phylogenetic and function shift analyses showed that the RIFIN proteins could be subdivided into two major groups that we named A- and B-RIFIN proteins.
