Molecular discrimination between Rana latastei and R. italica

The two endemic Italian brown frog species, Rana latastei and R. italica, have a very similar morphology: the diagnostic characters are the tympanum, larger in R. latastei and the nostril distance that is almost equal to the interorbital length in R. latastei, while is slightly greater in R. italica. The most important parameter in order to discriminate between the two species is their different areals: R. latastei is mostly widespread in the Po Valley, while R. italica inhabits almost only the Apennine rivers. The S1 satellite DNA from Palaearctic brown frogs displays a different structure in all the previously tested species, and this complete species- specificity was verified also for R. latastei and R. italica. Southern Blot of DNA samples from the two species, digested with five different restriction enzymes, displays different patterns of hybrid bands. Both species present two repetitive units S1a and S1b. The S1a repetitive unit has the same size but different restriction sites, while the S1b unit differs both in size and restriction sites. In conclusion, the S1 satellite DNA is a powerful molecular marker in order to discriminate the Palaearctic brown frog species.