4.1 Phylogeographical patterns driven by river isolation
Through the examination of the ribotypes and chlorotypes of 217
individuals from 10 populations of the Hainan-endemic herbaceousP. heterotricha in the south-central mountain system of Hainan
Island, low within-population genetic diversity and high genetic
differentiation (Fst ) among populations were revealed (Figure 1).
There was a significant phylogeographical structure and AMOVA showed
high genetic variation mainly from inter-regions (Table 4). Three clades
NW, SE and SW have evolved in prolonged isolation, as intimated by
bayesian analysis (Figure 3), which is supported by results of STRUCTURE
v.2.3.4. (Figure 4), and low gene flow (Nm ) among regions were
detected. Two main genetic barriers were identified based on Monmonierʼs
algorithm (Figure 5), which spatially concordant with the Changhua River
and Wanglou River in south-central mountains system of Hainan Island
(Figure 2, 3 and 4).
The average population genetic divergence detected in P.
heterotricha is high overall (GST = 0.801 and
0.765 for nrITS and cpDNA gene respectively), considering that genetic
differentiation among populations turns to be stronger whereGST > 0.25 (Buso et al. ,
1998). Primulina heterotricha grows widely in rocky streamsides
of forest valleys and humid or arid limestone and granites habitat,
which are widespread in south-central Hainan Island. Yet, there was a
slight IBD, although pollen or seed dispersal is very limited among most
populations (Nm << 1, Table 3) owing to the
discussed isolation effect of Changhua River, and the lack of adaptation
to long-range seed dispersal. Furthermore, the distribution of effective
pollinators (genus Glossamegilla ) is interrupted easily by rivers
and mountains, and it even shows altitudinal differences within the same
mountain system (Zhong et al. , 2014).
Geographic discontinuities are indeed a significant factor for
population differentiation through weakening or blocking gene flow
(Slatkin, 1985), and their impacts apparently have driven regional
genetic differentiation within P. heterotricha populations. These
results are in line with previous studies in other groups, showing that
the geographic barrier associated with the course of Changhua River may
have stimulated lineage diversification. Two examples are the multiple
intra-specific genetic lineages detected in the endemic speciesMetapetrocosmea peltata (Merr. et Chun) W. T. Wang (Gesneriaceae)
and genus Oreocharis (Gesneriaceae), which also presented a
strong phylogeographical population structures (Li et al. , 2020;
Ling et al. , 2020a).
The major role of mountains and valleys in the origins and evolution of
plants has been widely reported in the Andes (Antonelli et al., 2009;
Pennington et al. , 2010), Ghats (Robin et al. , 2015) and
Hengduan Mountain (Liu et al. , 2013). And similar results of
genetic structure associated with sharp geographical discontinuities
have been reported in many Asian plants and animals. Regarding other
families and phyla, Myricaria laxiflora (Franch.) P.Y. Zhang et
Y.J. Zhang (Tamaricaceae) in the Three Gorges mountain region (Liu et
al. , 2009), Taxus wallichiana Zucc. (Taxaceae) in the
Himalaya-Hengduan Mountains region (Liu et al. , 2013), and
several endemic montane birds from the Western Ghats (Robin et
al. , 2015), all showed nested genetic patterns across complex
mountain system isolated by valleys or rivers. The ensuing gene flow
limitation, genetic drift enhancement and possibly natural selection,
can likely have induced the genetic structure detected, that may in the
long term lead to reproductive isolation and speciation.
Our genetic data pinpoint the lower Changhua River as the stronger
gene-flow barrier between NW and SW clades (93% bootstrap) with minimumNm = 0.04 and 0.07 for nrDNA and cpDNA sequences, followed by a
secondary influence of the Wanglou River, separating SE and SW clades
with Nm = 0.10 and 0.07 for nrDNA and cpDNA sequences, then the
third influence of the upper Changhua River dividing NW and SE clades
with Nm = 0.40 and 0.41 for nrDNA and cpDNA sequences (Figure 5,
90% bootstrap). Notably, the Nansheng River splits nrITS clade SE into
two subclades (BS below 80%, Figure 5A). Besides, according to the
BARRIER analysis, the isolation magnitudes were proportional to the size
of the rivers; the lower Changhua River features the greatest impact,
possibly owing to its much larger width (1660 m, vs only 300 m of the
upper Changhua River and 160 m of the Wanglou River), and bigger
discharge volume.