Forest refugia revisited: nSSRs and cpDNA sequences support historical isolation in a wide-spread African tree with high colonization capacity, Milicia excelsa (Moraceae)

Abstract

The impact of the Pleistocene climate oscillations on the structure of biodiversity in tropical regions remains poorly understood. In this study, the forest refuge theory is examined at the molecular level in Milicia excelsa, a dioecious tree with a continuous range throughout tropical Africa. Eight nuclear microsatellites (nSSRs) and two sequences and one microsatellite from chloroplast DNA (cpDNA) showed a deep divide between samples from Benin and those from Lower Guinea. This suggests that these populations were isolated in separate geographical regions, probably for several glacial cycles of the Pleistocene, and that the nuclear gene pools were not homogenized despite M. excelsa's wind-pollination syndrome. The divide could also be related to seed dispersal patterns, which should be largely determined by the migration behaviour of M. excelsa's main seed disperser, the frugivorous bat Eidolon helvum. Within Lower Guinea, a north-south divide, observed with both marker types despite weak genetic structure (nSSRs: F(ST) = 0.035, cpDNA: G(ST) = 0.506), suggested the existence of separate Pleistocene refugia in Cameroon and the Gabon/Congo region. We inferred a pollen-to-seed dispersal distance ratio of c.1.8, consistent with wide-ranging gene dispersal by both wind and bats. Simulations in an Approximate Bayesian Computation framework suggested low nSSR and cpDNA mutation rates, but imprecise estimates of other demographic parameters, probably due to a substantial gene flow between the Lower Guinean gene pools. The decline of genetic diversity detected in some Gabonese populations could be a consequence of the relatively recent establishment of a closed canopy forest, which could negatively affect M. excelsa's reproductive system.

Speciation slowing down in widespread and long-living tree taxa: insights from the tropical timber tree genus Milicia (Moraceae)

“…These included 550 individuals previously studied in Dainou et al (2010) and sampled from the Lower Guinea and Benin (Dahomey gap) and 46 collected from herbarium collections at the National Botanical Garden of Belgium. During the sampling work, collaborators did not pay attention to the taxonomic identity of individuals.…”

“…We applied the method as described in Dainou et al (2010) with some slight differences. Only the admixture model was run with an interaction parameter c ¼ 0, which means that no spatial information was included in the analysis.…”

“…psbA-trnH was amplified in a total volume of 25 ml with the Phusion polymerase (Finnzymes Espoo, Finland) following the manufacturer's protocol. PCR conditions for trnC-ycf6 were as described in Dainou et al (2010) and were also applied for At103. From the subsample of 260 individuals selected for sequencing, 195, 225 and 202 were successfully amplified for psbA-trnH, trnC-ycf6 and At103, respectively, with 172 individuals common to the three fragments.…”

“…Ofori et al (2003) tried to assess genetic differentiation between Milicia species via chloroplast markers, but their populations were assigned to species according to vegetation types because of difficulties in obtaining diagnostic morphological traits of the sampled individuals. Within M. excelsa, Dainou et al (2010) demonstrated the existence of different gene pools in the Lower Guinea (the rainforest from the Atlantic Ocean until the western side of the Oubangui-Congo River; Supplementary Figure S1), despite extensive gene dispersal (Bizoux et al, 2009), and suggested a relatively recent fragmentation during Pleistocene ice ages. Curiously, Dainou et al (2010) also showed that West African M. excelsa populations from Benin in the Dahomey gap (the savannah interval separating the Upper Guinean and Lower Guinean forest blocks; Supplementary Figure S1) strongly diverged from Central African samples, despite the fact that M. excelsa presents a continuous range from West to Central Africa.…”

“…Within M. excelsa, Dainou et al (2010) demonstrated the existence of different gene pools in the Lower Guinea (the rainforest from the Atlantic Ocean until the western side of the Oubangui-Congo River; Supplementary Figure S1), despite extensive gene dispersal (Bizoux et al, 2009), and suggested a relatively recent fragmentation during Pleistocene ice ages. Curiously, Dainou et al (2010) also showed that West African M. excelsa populations from Benin in the Dahomey gap (the savannah interval separating the Upper Guinean and Lower Guinean forest blocks; Supplementary Figure S1) strongly diverged from Central African samples, despite the fact that M. excelsa presents a continuous range from West to Central Africa. This pattern questions whether M. excelsa individuals from Benin are more closely related to West African M. regia or to Central African M. excelsa populations.…”