A rapid PCR-based test for species identification of two cryptic bats Pipistrellus pipistrellus and P. pygmaeus and its application on museum and dropping samples

The identification of two cryptic bat species of the genus Pipistrellus using a non-destructive and quick method of multiplex PCR and agarose gel electrophoresis is described. Two primer combinations were able to produce species-specific bands that identified reliably individuals that were previously identified by mtDNA sequencing. Robustness of the method was subsequently successfully tested on 16 randomly selected free-living animals from central Europe (tissue samples obtained from a 3 mm punch of wing-membrane) identified to species on the basis of echolocation calls. Nine out of 15 museum specimens and 100% of fresh faecal samples from seven individuals were also successfully identified by this method. The described method thus provides a good way to routinely distinguish two Pipistrellus species by using non-destructive sampling of living individuals or droppings, and will be used in field studies of their ecology.