Study species and sampling
Callicarpa parvifolia grows in sunny dry dwarf scrub on rocky
ground in the Chichijima Islands (Toyoda, 2003), with a flowering peak
in July (Table 1), whereas C. glabra grows in the understory of
dry scrub in the Chichijima Islands (Toyoda, 2014) and has its flowering
peak in August. These species are classified as “endangered” and
“critically endangered” in the Red List of Threatened Species of Japan
(Ministry of the Environment Government of Japan, 2020), respectively.
In contrast, C. subpubescens is not listed as a threatened
species and is widely distributed in the Bonin and Volcano Islands.Callicarpa subpubescens exhibits different ecotypes, each with
distinct habitats and some with different flowering peaks. For example,
the Chichijima Islands’ ecotype (S) inhabits the forest edge of mesic
forests, with peak flowering in June. The Hahajima Islands have four
ecotypes: the glabrescent ecotype (SG), the tall ecotype (ST), the dwarf
ecotype (SD), and the hybrid ecotype (SH; previously called the
intermediate ecotype M; Setsuko et al. 2023). The ecotype SG inhabits
the understory of mesic forests, with a flowering peak in July. The
ecotype ST forms the canopy of tall mesic forests, with a flowering peak
in October. The ecotype SD forms the canopy of dry scrub, with two
flowering peaks in August and November. The ecotype SH forms the canopy
of mesic scrub (cloud forests) or inhabits the forest edge of mesic
forests, with a flowering peak in July (Setsuko et al., 2023; Sugai et
al., 2019). The Mukojima Islands have two ecotypes (STm and Sm), which
are genetically close to the ecotype ST in the Hahajima Islands and the
ecotype S in the Chichijima Islands (Sugai et al. 2019; also refer to
the Results section). However, substantial forest areas have been lost
due to feral goats (Shimizu, 2003), making it challenging to determine
their original habitats. Flowering surveys on the Mukojima Islands were
conducted only once in July 2010, as the islands are currently
uninhabited, with no ocean liner services, and distant from inhabited
Chichijima Island.
To cover species and ecotypes of each island in the Bonin Islands, leaf
samples were collected from 94 individuals across 14 populations for DNA
extraction (Table 1; Fig. S1). These samples included two populations in
the Mukojima Islands, six populations from the Chichijima Islands, and
six populations from the Hahajima Islands. As outgroups, one individual
each of Callicarpa japonica and Callicarpa mollis , which
grow in Kyoto Prefecture, Japan, was also collected (Fig. S1). Silica
gel was used to immediately dry leaf samples used for DNA extraction.
Leaves were also sampled for phenotypic measurements from the same
individuals used for DNA extraction. As leaf morphology varies within
individuals depending on sunlight exposure, leaves were collected from
the sunlit upper canopy. However, owing to time constraints, sunlit
leaves could not be collected from the SGi population.