A possible endophytic symbiotic bacterium of endemic species Androsace brevis (Primulaceae)

Background and Aims – Androsace brevis (Hegetschw.) Ces. is a narrow endemic plant living on ridges in a restricted area of the Southern Alps in Lombardy (Italy) and Switzerland. It prefers acid soils with low nitrogen content and blooms immediately after the snowmelt. Androsace brevis is proposed as a model species to study the effects of climate change on the web of interactions in mountain ecosystems (plant-arthropod, plant-microorganisms). During the sequencing of A. brevis genome carried out in a preliminary work, a significant amount of DNA belonging to a prokaryotic organism and not compatible with environmental contamination was detected. Our aim is to identify and characterise this microorganism. Methods – To evaluate the presence and diffusion of the bacterium, specific PCR primers were designed and tested on A. brevis individuals belonging to eight different natural populations. To isolate the bacterium from plant tissues, different growth media and conditions were tested. Bioinformatics analyses were performed to classify the bacterium and to identify protein functional domains trying to understand its relationship with the plant. Results – The bacterial genome was de novo assembled and identified as belonging to the Beijerinkiaceae family, Rhizobiales order. Nor the genus neither the species could be identified: at present there is no perfect match with any bacterial genome in public sequence databases. The Beijerinckiaceae family includes bacteria living in the phyllosphere, often methylotrophs or methanotrophs sharing nitrogen fixation capability and promoting plant growth. PCR assays confirmed the presence of the bacterium in all samples tested. However, so far, any attempt of isolating the bacterium has been unsuccessful. The in-silico analysis of the predicted functional domains suggests a likely symbiotic relationship with the plant, and preliminary microscope observations confirm the presence of endophytic bacteria inside plant tissues (leaves). Conclusions – The molecular and functional characterization of this microorganism could help to clarify the ecology of A. brevis, and to reduce the current lack of knowledge about high-altitude plant-bacteria interactions