Molecular identification of temperate Cricetidae and Muridae rodent species using fecal samples collected in a natural habitat

Molecular species identification from biological material collected at field sites has become an established ecological tool. However, extracting and amplifying DNA from degraded field samples, such as prey remains and feces that have been exposed to the elements, remains a challenge and costly. We collected 115 fecal samples of unknown small mammals, resembling fecal droppings of voles and mice (i.e., Cricetidae and Muridae), from a salt marsh in The Netherlands. We modified a previously published protocol into a relatively low-cost method with a PCR success of 95%. We demonstrate that species identification is possible for both Cricetidae and Muridae species using fecal samples of unknown age deposited in the field. For 90 samples, sequences of the variable control region in the mitochondrial genome were obtained and compared to published DNA sequences of small mammals occurring in north European salt marshes. A single sample, probably environmentally contaminated, appeared as Sus scrofa (n = 1). We positively identified house mouse Mus musculus, being the positive control (n = 1), and common vole Microtus arvalis (n = 88). In 81 sequences of 251 nt without ambiguous bases, ten haplotypes were present. These haplotypes, representing the central lineage of the western subspecies M. arvalis arvalis, were separated by 20 mutations from published control region haplotypes of the western European lineages sampled in France. Unlike earlier studies of cytochrome b variation in coastal European populations, we did not find indications of recent purging of genetic variation in our study area.