This work reports the study of the frescoes from the Casa de Fresco dos Sanches Baena in Vila Viçosa (Southeast Portugal) to allow their material characterisation, to identify the different microorganism populations and to assess their role in the deterioration of these paintings. The Casa de Fresco dos Sanches Baena is a small semi-underground building constructed in a Palace garden over a well and used as a cool refreshing place by the owners. The frescoes that cover the ceilings (composed of 4 panels) present rich mythological scenes and have other decorative elements, which make them an unusual example of this art form. Unfortunately, due to partial abandonment the paintings are in an advanced state of degradation exhibiting partial detachment of paint layers and mortars, salt efflorescences and abundant biological colonisations (Fig.1).