Illicium difengpi Kib and Kim, an endangered plant unique to karst areas in China, has evolved an extremely high tolerance to arid environments. To elucidate the molecular mechanisms of the response to drought stress in I. difengpi, physiological index determination and transcriptome sequencing experiments were conducted in biennial seedlings grown under different soil moisture conditions (70~80%, 40~50% and 10~20%). With increasing drought stress, the leaf chlorophyll content decreased, while the proline (Pro), soluble sugar (SS) and malondialdehyde (MDA) contents increased; superoxide dismutase (SOD) and peroxidase (POD) activities also increased. Transcriptome sequencing and pairwise comparisons of the treatments revealed 2489, 4451 and 753 differentially expressed genes (DEGs) in CK70~80 vs. XP40~50, CK70~80 vs. XP10~20 and XP40~50 vs. XP10~20, respectively. These DEGs were divided into seven clusters according to their expression trends, and the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment results of different clusters indicated that genes in the hormone signal transduction and osmotic regulation pathways were greatly activated under mild drought stress. When drought stress increased, the DEGs related to membrane system and protein modification and folding were all upregulated; simultaneously, chitin catabolism- and glycolysis/gluconeogenesis-related genes were continuously upregulated throughout drought stress, while the genes involved in photosynthesis were downregulated. Here, 244 transcription factors derived from 10 families were also identified. These results lay a foundation for further research on the adaptation of I. difengpi to arid environments in karst areas and the establishment of a core regulatory relationship in its drought resistance mechanism.

Castanopsis is one of the most common genus of trees in subtropical evergreen broad-leaved forests and tropical monsoon rainforests in China. Castanopsis hainanensis and Castanopsis wenchangensis are endemic to Hainan Island, but they were once confused as the same species due to very similar morphologies. In this study, nuclear microsatellite markers and chloroplast genomes were used to delimit C. hainanensis and C. wenchangensis. The allelic variations of nuclear microsatellites revealed that C. hainanensis and C. wenchangensis were highly genetically differentiated with very limited gene admixture. Both showed higher genetic diversity within populations and lower genetic diversity among populations, and neither had further population genetic structure. Furthermore, C. wenchangensis and C. hainanensis had very different chloroplast genomes. The independent genetic units, very limited gene admixture, different distribution ranges, and distinct habitats all suggest that C. wenchangensis and C. hainanensis are independent species, thus they should be treated as distinct conservation units.

Castanopsis chinensis (Spreng.) Hance is widespread in the subtropical forests of China. Castanopsis qiongbeiensis G.A. Fu and Castanopsis glabrifolia J. Q. Li & Li Chen are limited to the coastal beaches of Wenchang county in the northeast of Hainan Island, and have similar morphological characteristics to C. chinensis. It is supposed that C. qiongbeiensis and C. glabrifolia are closely related to C. chinensis. In the present study, the genetic differentiation, gene flow, and genetic relationship of C. chinensis, C. qiongbeiensis, and C. glabrifolia were investigated by using 15 nuclear microsatellite markers; a total of 308 individuals from 17 populations were sampled in the three species. The allelic variation of nuclear microsatellites revealed moderate but significant genetic differentiation (F_CT = 0.076) among C. chinensis, C. qiongbeiensis, and C. glabrifolia, and genetic differentiation between C. chinensis and C. glabrifolia was larger than that between C. chinensis and C. qiongbeiensis. Demographic simulations revealed unidirectional gene flow from C. chinensis to C. glabrifolia and C. qiongbeiensis, which highlight dispersal from mainland to island. The isolation effect of Qiongzhou Strait increased the genetic differentiation of species on both sides of the strait; however, the differentiation was diminished by gene flow that occurred during the historical period when Hainan Island was connected to mainland China. Our results supported the argument that C. glabrifolia should be considered an independent species and argued that C. qiongbeiensis should be regarded as an incipient species and independent conservation unit.

The cytoplasmic genome of one species may be replaced by that of another species without leaving any trace of past hybridization in its nuclear genome, which can thus confuse the inference of genealogical relationship and evolutionary history of many congeneric species. In this study, we used sequence variations of chloroplast DNA and restriction site-associated DNA to investigate gene exchange between Castanopsis fabri and Castanopsis lamontii, and to infer the divergence history of the two species by comparing different divergence models based on the joint allele frequency spectrum. We evaluated climatic niche similarity of the two species using climatic variables across their entire distribution range in subtropical China. Clear genetic differentiation was revealed between C. fabri and C. lamontii, and gene exchange between the two species was discovered as a consequence of secondary contact. The gene exchange rates were variable across the genome. Gene exchange could allow C. fabri to widen its habitat through pollen swamping and broaden its climatic niche, and the chloroplast genome of C. lamontii is captured by C. fabri during this process. These results further our understanding of the timing and contribution of gene exchange to species divergence in forests.