STSM of Eleonora Fusi (University of Milan) to Freie Universität Berlin, Germany
Effect of nutrients and nutraceuticals on the development of intestinal tract and immune response.

1. Aim of visit

The aim of this visit was to assess the effect of some nutrients, nutraceuticals and additives on the development of the intestinal tract and the immune response in young animals. The attention was focused on the role of zinc oxide in counteracting the early Lawsonia intracellularis infection in weaned piglets. For this purpose IPEC2 cells and porcine intestinal explants isolated from young animals were used. The role of zinc in the anti-inflammatory response and resistance to infection is well known. In particular, zinc oxide (ZnO) appears to have a strong protective effect in resisting intestinal diseases. Several studies conducted on piglets showed that dietary ZnO supplementation could prevent or reduce intestinal diseases, but further understanding of the role of dietary supplementation in preventing the early bacterial infection will offer alternative strategies in livestock management.

2. Description of the work carried out during the visit
The undifferentiated porcine intestinal epithelial cell line (IPEC2) derived from jejunum was used. Because IPEC2 are not immortalized, they represent an ideal model of normal intestinal epithelium Stimulation of the cells were performed using several concentrations of ZnO and different incubation periods. The ZnO concentration and the incubation time which showed the most effective effect were selected for the following part of the project.
In the second part of the project the infection design was performed. Enterisol® ileitis vaccine in reason of two infection doses was used. After the pre-treatment with ZnO the intestinal cells were infected. In order to evaluate the condition of cell culture cellular vitality (trypan blue) and cell counting were carried out. To evaluate the rate of infection the following analyses were performed: flow cytometry (CFDA Lawsonia intracellularis staining); in situ hybridization, qPCR.
The last part of STSM involved the preparation of intestinal explants isolated from weaned pigs. The use of intestinal explants showed the interaction between the intestinal microflora, host intestinal epithelium, and pathogen under physiological condition. On the intestinal explants now are being used the experimental assays described above. Moreover, the expression of selected cytokines and chemokine mediators will be evaluated.

3. Description of the main results obtained

The preliminary results obtained from the STSM project point to indicate that the rate of infection of Lawsonia intracellularis and the efficacies of ZnO pretreatment varied from assay to assay in relation to the cell condition and to the intestinal microflora present on the explants. Further additional experimental approach have been planned in order to investigate the composition of the beneficial microflora that could prevent the early infection and the influence that a dietary treatment such as ZnO could improve.

4. Conclusions
The STSM was successful because it has allowed the establishment of a physiological complex of in vitro models for nutritional studies. These models will be used to test other nutritional compounds and infective agents. The visit offers the chance to the fellow to learn and well trained new techniques, which were new for her. Moreover, new protocols were set up by the fellow. This will give her the possibility to apply these techniques in her home institution in future research.

5. Future perspectives
The collaboration with the host institution was very successful. Based on the successful collaboration during this short term scientific visit, the host and guest institutes are willing to continue the collaboration.
The continuation of this collaboration will be included the conclusion of the experimental plan by assessing of the expression selected cytokines and chemokine mediators in both in vitro models in order to understand the development of the immune response.

6. Projected publications/articles resulting or to result from the STSM
It is planned to disseminate the present and on coming results at least in a conference abstract (15th Meeting of the European Society of Veterinary and Comparative Nutrition, 14-16 September, 2011, Zaragoza, Spain). Possible publication in a peer-review journal such as “Archives of animal nutrition” will be also considered. The COST action Feed for Health FA0802 will be acknowledged in the resulting publications.

Eleonora Fusi STSM
Design & Programming: Zenlex