A conservative approach for species delimitation based on multi-locus DNA sequences: a case study of the genusGiraffa(Mammalia, Cetartiodactyla)

Molecular data are now commonly used in taxonomy for delimiting cryptic species. In the case of giraffes, which were treated as a single species ( Giraffa camelopardalis ) during half of a century, several molecular studies have suggested a splitting into four to seven species, but the criteria applied for taxonomic delimitation were not fully described. In this study, we have analysed all multi-locus DNA sequences available for giraffes using multispecies coalescent (MSC: *BEAST, BPP and STACEY), population genetic (STRUCTURE, allelic networks, haplotype network and bootstrapping) and phylogenetic (MrBayes, PhyML, SuperTRI) methods to identify the number of species. Our results show that depending on the method chosen, different taxonomic hypotheses, recognizing from two to six species, can be considered for the genus Giraffa. Our results confirm that MSC methods can lead to taxonomic over-splitting, as they delimit geographic structure rather than species. The 3-species hypothesis, which recognizes G. camelopardalis sensu strico, G. giraffa , and G. tippelskirchi , is highly supported by phylogenetic analyses and also corroborated by most population genetic and MSC analyses. The three species show high levels of nucleotide divergence in both nuclear (0.35-0.51 %) and mitochondrial sequences (3-4 %), and they are characterised by 7 to 12 exclusive synapomorphies (ES) detected in nine of the 21 nuclear introns analysed for this study. By contrast, other putative species, such as G. peralta , G . reticulata , G. thornicrofti or G. tippelskirchi sensu stricto, do not exhibit any ES in nuclear genes. A robust mito-nuclear conflict was found for the position and monophyly of G. giraffa and G. tippelskirchi , which is explained firstly by a mitochondrial introgression from Masai giraffe to southeastern giraffe during the Pleistocene, and secondly, by gene flow mediated by male dispersal between southern populations (subspecies G.g. giraffa and G.g. angolensis ).