Accurate identification of three cryptic species of rodents of the genus Calomys using RAPD-PCR and mitDNA RFLP markers

GONZÁLEZ ITTIG, R.E.; CHIAPPERO, M.B.; PROVENSAL, C.; GARDENAL, C.N.

BIOCHEMICAL SYSTEMATICS AND ECOLOGY

PERGAMON-ELSEVIER SCIENCE LTD

Lugar: Amsterdam; Año: 2002 vol. 30 p. 425 - 432

0305-1978

Calomys musculinus, Calomys lauchaandCalomys venustusare cryptic species with overlapping distribution ranges.C. musculinusis the natural reservoir of Junin virus (Arenaviridae), the etiological agent of Argentine Hemorrhagic Fever. In epidemiological studies it is very important to unequivocally identify the species of individuals collected in the field in order to test virus infection. The purpose of this work was to describe molecular markers allowing a prompt and clear characterization of individuals of the three species. We studied the D-loop region of mitochondrial DNA by restriction fragment length polymorphism (RFLP). This region was amplified by PCR and the product was digested with nine restriction endonucleases (RE). Eight recognize 4 bp restriction sites (TaqαI, Tsp509 I, AciI, RsaI, AluI, NlaIII, HaeIII and MseI) and one recognized a 6 bp sequence (AseI). Two of them (AciI and HaeIII) did not distinguish any of the three species. AluI did not discriminate between C. musculinus and C. laucha, but clearly distinguished both from C. venustus. TaqI did not distinguish C.laucha from C. venustus, but differentiated both species from C. musculinus. MseI distinguished the three species, but some of the polymorphisms of C. musculinus are very similar to C. laucha?s restriction pattern. The enzyme NlaIII distinguished the three species, but it is highly polymorphic within species. The enzymes Tsp509 I, RsaI and AseI clearly discriminated the three species, and patterns obtained with the three of them are recommended for reliable identification of individuals collected in thefield. The same DNA samples were used to obtain Random Amplified Polymorphic DNAs (RAPDs) patterns. Several bands produced with primers A02, A06, A08, A09, B09 and OPA02 are species specific and could also be used for identification