The Balkan Green-veined White (Pieris napi balcana): did the barcoding approach resolve its taxonomic status?

To resolve taxonomic status and distribution of the enigmatic Pieris (napi) balcana Lorković, 1970 specimens from a P. napi aggregate were collected in the karst habitats of the western part of the Croatian Dinaric Alps. Wing pattern characters were studied on a large sample (n=323). Egg laying of balcana females on the larval host‐plant Arabis turrita was observed in spring at three localities, and butterflies have been reared on the same plant. DNA barcodes of 58 individuals (39 localities) were obtained and complemented by sequences of a fragment of the nuclear wingless (wg) gene. Tissue from fresh adult males was used for karyotyping (n=21). ‘Balcana’ shares the DNA barcode with the subalpine P. (napi) bryoniae, while balcana/bryoniae and napi haplotypes segregated into two distinct mitochondrial clusters. Barcoding results agree with the absence of reproductive isolation between balcana and bryoniae in former hybridisation studies, and both taxa have the same polymorphic karyotype, different from the napi karyotype. ‘Balcana’ and napi could not be distinguished by wg sequence data. At 26 localities, we found only bryoniae/balcana DNA barcodes, and P. (napi) napi was detected only in probably parapatric area. On Žumberak Mt. we discovered F1 hybrids with napi barcode and hybrid wing pattern.