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Background  A deep divergence of mitochondrial DNA is common in species delimitated by morphological traits. Several hypotheses can explain such variations, such as cryptic species, introgression, allopatric divergence and ancestral lineage. The black-throated tit harbors several deeply divergent mitochondrial lineages. Two lineages with 5% divergence, but having a high level of gene flow, have been detected in its subspecies,  A. C. concinnus  and  A. c. talifuenses . In this study, we conducted a genetic analysis at a contact zone of these two lineages to identify whether these lineages either reflect a high level of intraspecific variation in mitochondrial loci or represent incipient speciation. Mitochondrial ND2 and 11 microsatellite loci were used to conduct phylogenetic and population structure analyses.    Results  ND2 haplotypes actually diverged into two groups within subspecies  A. c. talifuenses ; however, they formed a non-sister relationship when including all available GenBank ND2 sequences. Analyses of microsatellite data indicated no existing population structure and showed a pattern of isolation by distance. Individuals sampled at the contact zone were almost identified as F2 hybrids.    Conclusions  Isolation for 2.4 Ma, as suggested by a previous study, appeared to be insufficient to develop robust reproductive barriers. Reproductive barriers were weak, or even absent between the divergent lineages, highlighting that incipient speciation was unlikely to be the case. Considering the results from previous studies, the divergent lineages may be better explained by secondary contact after allopatric isolation because of Pleistocene climate changes, but other hypotheses cannot be definitively ruled out because of the lack of representative samples from the other distribution region and its relatives. Considering that divergence in morphology was hardly observed and the particular split in genetics, the two subspecies might be better merged. The current findings also highlight the likely contribution of male-biased dispersal to male-biased gene flow among mitochondrial lineages; more efforts are needed to illustrate the evolutionary history of the black-throated tit.    Electronic supplementary material  The online version of this article (doi: 10.1186/s12862-017-1114-9) contains supplementary material, which is available to authorized users.

bmc evolutionary biology

The absence of reproductive isolation between non-sister and deeply diverged mitochondrial lineages of the black-throated tit (Aegithalos concinnus) revealed by a multilocus genetic analysis in a contact zone