Cryptic species
Phenotypic and genetic differences, as well as diversification patterns,
were used to assess classifications as distinct species. First, the leaf
phenotypes of species P (C. parvifolia ) and G (C. glabra ),
as well as ecotype SD (C. subpubescens ), exhibited distinct and
distinguishable distributions in the PCA of leaf morphology (Fig. 5).
However, the ecotypes of C. subpubescens , excluding ecotype SD,
displayed PCA plots with less clarity in terms of separation.
Callicarpa parvifolia , endemic to the Chichijima Islands grows in
very dry and sunny rocky areas and is notably distinguishable from otherCallicarpa species and ecotypes owing to its small, rounded,
remarkably hairy, and thick leaves (Fig. S11). Similarly, C.
glabra , also endemic to the Chichijima Islands grows in the dry scrub
understory and is characterized by its small, slender, hairless, and
rather thick leaves. Phylogenetically, these two species form a sister
clade, each maintaining monophyly (Fig. 3b). Accordingly, the current
taxonomy of the two species appears reasonable.
Ecotype SD, found only in the Hahajima Islands, forms a canopy in dry
scrubs, with leaves that are small, round, hairy, and thick, similar toC. parvifolia (Fig. S11). However, it can be distinguished fromC. parvifolia by significantly lower leaf hair density and
thickness. Phylogenetically, ecotype SD and C. parvifolia were
located different clades (Fig. 3b). A parallel evolutionary process of
adaptation to dry environments likely resulted in a morphology closely
resembling that of C. parvifolia . Additionally, compared with
ecotype S from the Chichijima Islands, treated as C.
subpubescens , ecotype SD differs both in its leaf morphology and
flowering phenology, with prolonged flowering from summer to winter
(Table 1), warranting recognition as a new species.
Although ecotype ST in the Hahajima Islands could not be clearly
distinguished from other ecotypes of C. subpubescens except
ecotype SD in leaf morphology (Fig. 5), it can be distinguished from
other ecotypes using flowering phenology (the flowering season of
ecotype ST is autumn, whereas those of ecotypes S, SG, and SH are
summer; Table 1). Phylogenetically, we consider ecotype ST to be a
cryptic species, as it diverged from the ancestral species at an age
more than twofold greater than that of other genetic groups based on
population demography analysis (Fig. 4).
Both ecotype S in the Chichijima Islands and ecotype SG in the Hahajima
Islands flower in summer (Table 1). Ecotype S is identical to the
registered type specimen of C. subpubescens (specimen no.
K000674714), exhibiting fine, soft hairs on its leaves, whereas ecotype
SG has almost no hairs on its leaves. However, the number of hairs did
not differ compared with ecotype SG (Fig. S11), and the plot
distributions of ecotypes S and SG overlapped in PCA (Fig. 5). Despite
belonging to different genetic groups, no obvious phenotypic differences
were observed. Several examples exist in the Bonin Islands of the same
species differentiated genetically due to gene flow restriction caused
by the different island groups they inhabit (Setsuko et al., 2017;
Setsuko et al., 2022; Setsuko et al., 2020; Sugai et al., 2013).
Therefore, considering ecotype SG as homologous to ecotype S is
reasonable.
Ecotype SH, thought to be derived from a hybrid of ecotypes ST and SG,
is found only on Hahajima Island (Figs. 1 and 2a). Ecotypes SH and ST
possessed similar leaves, with ecotype SH occupying part of the broader
distribution of ecotype ST in the PCA plot (Fig. 5). However, the plot
distributions of ecotypes SH and SG in PCA did not overlap. As the
flowering season of ecotypes SG and SH is summer and that of ecotype ST
is autumn (Table 1), ecotypes SH and ST are distinguished by their
flowering periods, whereas ecotypes SH and SG are roughly distinguished
by their leaf morphologies. In naturally distributed individuals,
habitat information can aid taxonomic classification of ecotypes SG and
SH, as ecotype SH inhabits high-elevation cloud forests and forms the
forest canopy layer, whereas ecotype SG inhabits the understory of mesic
forests, except in the high-elevation areas of the Hahajima Island
(Setsuko et al., 2023). Based on these findings, we propose thatC. subpubescens can be divided into three species in addition to
one hybrid-derived taxon, rather than one species.
Concerning ecotypes in the Mukojima Islands, the flowering of STm and Sm
was investigated only once in July, with these ecotypes found to be in
the early and late stages of flowering, respectively. July marked the
beginning of flowering for ecotype ST in the Hahajima Islands, and the
end of flowering for ecotype S in the Chichijima Islands (Table 1).
Combining flowering information with the phylogenetic tree results from
this study, Sm was considered the same ecotype as S, whereas STm was
considered the same ecotype as ST. Determining whether ecotypes STm and
Sm in the Mukojima Islands can be considered identical to ecotypes ST
and S, respectively, necessitates a more thorough investigation into the
flowering periods of ecotypes STm and Sm. Furthermore, although only a
minimal number of hybrid individuals (two) were found in the Mukojima
Islands (Figs. 1 and 2), the extent to which they form hybrid zones,
akin to ecotype SH on Hahajima Island (Setsuko et al., 2023), remains
unclear. However, given the coexistence of lineages from different
origins on the same island, there is a possibility that this
codistribution contributes to hybrid speciation (Kagawa & Takimoto,
2018). Further research should include more comprehensive investigations
into hybridization in the Mukojima Islands.