Crataegus bretschneideri (Rosaceae), a separate species endemic to Northeast of China

Crataegus bretschneideri is considered a variety of Crataegus pinnatifida, but it has also been proposed as a separate species. We used ITS and three cpDNA sequence fragments (psbA-trnH, trnG-trnS, trnH-trnK) to perform maximum parsimony, maximum likelihood, Bayesian interence and neighbor-joining molecular phylogenetic tree analysis with 108 individuals of 19 hawthorn (Crataegus) species and 4 outgroups (Malus). ITS, cpDNA and cpDNA-ITS molecular phylogenetic trees constructed using 4 methods have similar consistency respectively. In the cpDNA-ITS molecular phylogenetic tree, C. bretschneideri is an independent clade separated from Crataegus maximowiczii and C. pinnatifida, which provides a separate species condition. In the ITS (nuclear inheritance) molecular phylogenetic tree, C. bretschneideri and C. pinnatifida are closely related and are distantly related to C. maximowiczii. The cpDNA (maternal inheritance) molecular phylogenetic tree indicated that C. bretschneideri is closely related to C. maximowiczii but distantly related to C. pinnatifida. Furthermore, the 108 Crataegus individuals belonged to 15 chloroplast haplotypes, as determined using the DNASP analysis; C. bretschneideri and C. maximowiczii have the same chloroplast haplotype, H1; C. pinnatifida contains three haplotypes, H13-H15. Taken together with previous results of our laboratory, we confirm that C. bretschneideri is not a variety of C. pinnatifida, but a separate species that gradually developed from natural hybrid offspring in mixed forest areas with C. maximowiczii as the female parent and C. pinnatifida as the male parent