Hybrid zones have been widely highlighted for their interest in understanding evolutionary processes. It is generally accepted that hybrid zones can be maintained in a balance between dispersal and selection. However, the selective forces can either be endogenous (i.e., genetic incompatibilities between parental taxa) or exogenous (i.e., parental taxa are adapted to different environments).

In this study, we evaluated these alternatives and determined the maintenance of a narrow hybrid zone between parapatric distributed Oxytropis diversifolia and O. leptophylla in Nei Mongol, China. For 507 individuals sampled from two populations in the hybrid zone, 12 O. diversifolia populations and five O. leptophylla populations, we measured leaf-morphological characteristics, quantified genetic structure using 11 nuclear microsatellite loci and five chloroplast DNA intergenic regions, collected micro- and macrohabitat data, and conducted geographical cline analysis.

We found that the two species differed in leaf morphology, and putative hybrids showed either intermediacy or a bias to O. diversifolia. Parental taxa formed two genetically distinct clusters, while populations in the hybrid zone consisted of both parental forms and various admixed individuals, exhibiting a bimodal pattern. The hybrid zone was coupled to ecological transitions of both microhabitat (i.e., the slope) and macroclimatic conditions. However, the genetic clines were significantly narrower than the environmental cline.

Our results indicate that endogenous selection can be primarily responsible for maintaining the hybrid zone, while local adaptation accounts for the position of the zone. We further suggest the probable outcome of hybridization could be introgression.

1. Morphology: 

Firstly, six leaf-morphological traits were measured by digital caliper for all 507 individuals of 19 populations: early leaf length, early leaf width, mature leaf length, mature leaf width, petiole length of early leaf, petiole length of mature leaf.

Secondly, we quantified the trichome density on leaf upper surface with an Olympus SZ61 stereo microscope (Olympus, Tokyo, Japan), and the number of trichomes on a 1mm×1mm surface was counted. In total we quantified 211 individuals of all the 19 populations.

2. Microsatellite genotyping:

11 nuclear microsatellite loci (N745892, N145635, N2724893, N2717495, N178451, N161850, N49251, N350553, N935993, N2528349, and N2697375) were scored across all 507 individuals according to the methods described by Wang et al. (2018).

Wang, H., Yang, H., Liu, P. L., Su, C., Xiao. L., & Chang, Z. Y. (2018). Isolation and characterization of microsatellite loci from Oxytropis diversifolia (Fabaceae). Applications in Plant Sciences, 6, e1168. https://doi.org/10.1002/aps3.1168

3. Microhabitat

For each individual sampled, a 40 cm × 40 cm square plot was surveyed, slope (by geologic compass), the percent of total vegetation cover, the percent of rocky ground (rock diameter > 0.2 cm), and the percent of bare ground (sand and soil) were measured. The latter three were scored on pictures by eye, and always done by the same experimenter (Hui WANG). In total, microhabitat data for 477 individuals in 18 populations were collected.

4. Macroclimate

In addition to our field sampling localities (19 records), occurrence records of the two parental species were collected from the Chinese Virtual Herbarium (CVH, http://www.cvh.ac.cn), the National Specimen Information Infrastructure (NSII, http://www.nsii.org.cn), and the Global Biodiversity Information Facility (GBIF, http://www.gbif.org). After removing wrong identifications, duplicate records and observations, and records without georeferences, in total we got 35 additional records for O. leptophylla. Nineteen contemporary bioclimatic variables of the time period 1970 – 2000 at 30-second resolution were downloaded from WorldClim (http://www.worldclim.org, Fick & Hijmans, 2017). To conduct PCA, the entire environmental space was built on variable values associated with 10,000 random points sampled from the whole study area.

Fick, S. E., & Hijmans, R. J. (2017). WorldClim 2: new 1‐km spatial resolution climate surfaces for global land areas. International journal of climatology, 37, 4302–4315. https://doi.org/10.1002/joc.5086

1. Morphology and 3. Microhabitat: integrated in one csv file, and can be opened in R.

2. Microsatellite genotyping: txt file can be opened in GENEPOP 4.7.

4. Macroclimate: rds file can be opened in R.