Biotechnological approaches for the conservation of Campanula sabatia De Not., a endangered Italian species

Campanula sabatia is one of the most vulnerable endemism of the Maritime and Ligurian Alps hot spot of biodiversity. According to IUCN categories, the species has been recently evaluated as vulnerable. 100 seeds from 9 populations have been collected and checked for their in vitro germination aptitude; after one month of culture, the final germination percentage (FGP) reached a mean of 79% with significant differences among populations. The mean germination time (MGT) was appreciable between a minimum of 17 to a maximum of 21 days. From the seedlings of each population, it was possible to clone and multiply the shoots onto an agarised Murashige and Skoog (MS) medium containing 1.33 μM of BA. Data on multiplication trials were strictly population dependent: mean values higher than 6 were observed in 6 out of 9 populations, the highest number of shoots/seedling/month (11) was obtained in populations 2, 4 and 8. The explants, cultured onto hormone-free MS medium, were able to develop a complete and functional root system after 25 days. The plants were recovered in the glasshouse and after acclimatization (25%) were used for in situ re-introduction and ex situ collection. AFLP markers were used to detect the genetic diversity within and among the populations, representative of the range of the species. For 83 C. sabatia samples, four combinations of selected primers generated a total of 100 polymorphic bands, that were used for the genetic distances analysis and the determination of allelic frequency. Nei‟s genetic distances ranged from a minimum of 0.063 to a maximum of 0.216. Nei’s gene diversity values varied between 0.0538 and 0.2194, with mean values of 0.1721 and 0.2434 at population and species level, espectively. The genetic differentiation coefficient among populations was 0.3018; the gene flow level was estimated on 1.1567. The scored data showed that C. sabatia has a remarkable genetic diversity, probably due to a high genetic exchange rate among populations. Conservation actions have been strongly suggested in relation to the genetic variability pattern.