Project: POCI-01-0145-FEDER-030431

Project name:	CLASInVivo - Development of NAM-CLASI-FIVH method: a new in vitro tool for an accurate analysis of diversity and spatial organization of gastric microbiota
Project code:	POCI-01-0145-FEDER-030431
Main Objective:	Reforçar a investigação, o desenvolvimento tecnológico e a inovação
Intervention Region:	Norte
Proposing institution/Lead promoter/Coordinating entity:	Faculdade de Engenharia da Universidade do Porto
Partner(s)/Co-promoter(s)/Participating institution(s):	Instituto de Patologia e Imunologia Molecular da Universidade do Porto
Date of approval:	2018-03-20
Start date:	2018-07-26
Completion date:	2022-07-25

Eligible Cost of the Project

Total Eligible Cost:	186.303,47 EUR
Eligible Cost in the University of Porto:	159.245,97 EUR
Faculdade de Engenharia da Universidade do Porto:	159.245,97 EUR

Total Financial Support

União Europeia - FEDER:	158.357,95 EUR
Orçamento de Estado:	27.945,52 EUR

Financial Support to the University of Porto

Total of the University of Porto:	159.245,97 EUR
União Europeia \| União Europeia - FEDER \| Faculdade de Engenharia da Universidade do Porto:	135.359,07 EUR
Nacional/Regional \| Orçamento de Estado \| Faculdade de Engenharia da Universidade do Porto:	23.886,90 EUR

Objectives, activities and expected/achieved results

Objetivos e Atividades Considering, the impact that NAMs have on FISH performance, our goal is to combine them with the two recent FISH procedures referred above in one technique designated by NAM-CLASI-FIVH; to our knowledge, there are no published results on this matter. First, to accomplish our goal, the NAMs probes used in NAM-CLASI-FIVH technique will be developed and optimized in conditions similar those found in stomach; then, the technique will be validated in gastric biopsies samples. Once this has been achieved, the developed protocol will be tested and adpated in a mouse model. Then, as a case study, we will analyse the microbial changes over time in a mouse model in terms of composition and spatial organization during H. pylori infection.
Resultados Esperados/Atingidos An EUB LNA/2'OMe probe conjugated to one fluorophore were tested against the species; to rank the fluorophores, 8 versions of EUB LNA/2'OMe probe, each conjugated to one of 8 different fluorophores were tested against E. coli. Strong variations on the signal intensity were found between species and between fluorochromes However, this methodology allowed the establishment of balanced species/fluorochromes pairs and provided an unambiguous separation of up to eight fluorophores Validation tests with different proportions of bacteria labelled with the different fluorophores showed that the frequency of each bacterium in the mixture is equivalent to the frequency of each species in the individual images from our spectral imaging-FISH. In fact, the separate spectral imaging combined with FISH it works since the frequency of each label type in individual images and mixtures are similar.

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