Comparative phylogeographic inference
The eDNA extracted from water scoops revealed the large-scale
phylogeographic structure of B. toni . Two distinct lineages were
found in the northern and southern regions of Hokkaido, which were not
separated by obvious geographic barriers. The observed pattern differed
from that of other aquatic species reported in Hokkaido. BesidesB. toni , two other species of primary freshwater fishes are also
known to have colonized in Hokkaido from Sakhalin across the Soya
Strait: the pond minnow Rhynchocypris percnurus sachalinensis and
eight-barbel loach Lefua nikkonis (13 , 33 ). The
lineages of R. p. sachalinensis are clearly differentiated on the
east and west sides of the Hidaka Mountains at the center of Hokkaido
(34 ). The Japanese crayfish Cambaroides japonicus(35 ) also exhibits the similar phylogeographic pattern. The
earliest-branched lineage of L. nikkonis (belongs to the
Nemacheilidae family, like B. toni ) is distributed along the Sea
of Okhotsk (northeastern Hokkaido), while the late-branched lineage has
wide-range distribution without either north–south or east–west
divided structure (33 ). No prior study on aquatic organisms has
demonstrated clear north–south phylogeographic vicariance in Hokkaido
without current geographic
barriers.
Based on the estimated divergence times, we concluded that B.
toni immigrated multiple times to Hokkaido via land bridges during the
ice age. The north–south phylogeographic structure of this species
possibly arose from the long time lag (over one million years) between
the independent colonization of Hokkaido by the ancestral populations of
the two clades. The phylogeography of this species is a potentially
important indicator of the historical footprints in regions of Hokkaido
and the past connection between Hokkaido and Sakhalin. Through future
studies on this species in Sakhalin and other Far East regions, we could
understand the distribution expansion of B. toni and other
species from the continent to the islands. Our findings highlight thatB. toni , which has been overlooked as a common species, is a
valuable source of biogeographic information on past geology and
environments.