Conclusion
Our study on the Callicarpa genus in the Bonin Islands provides crucial insights into phylogenetic relationships, divergence patterns, and speciation drivers. The ancestral ecotype ST of C. subpubescens was over twofold older than the other species/ecotypes, followed by ecotype SD, species P and G, ecotype S, and finally ecotype SG, all diverging simultaneously. The concurrent diversification of species/ecotypes adapted to different environments suggests a connection to island aridification. Shifts from tall forests to lower forests with increased forest edges and an increase in dry environments likely triggered rapid phenotypic and genetic changes for adaptation, as observed in other organisms. Future research will focus on identifying genes associated with drought adaptation. Cryptic species were found within C. subpubescens , with ecotypes SD and ST considered distinct species based on leaf morphology, flowering phenology, and genetic divergence patterns. Although genetically distinct, ecotypes S and SG show no significant phenotypic differences and can be treated as the same species. Ecotype SH, presumed to be a hybrid between ecotypes ST and SG and found only on Hahajima Island, exhibited different characteristics from its parent ecotypes. Long-distance seed dispersal events likely contributed to the presence of the same ecotypes in different island groups. Certain birds, such as the brown-eared bulbul, Japanese wood pigeon, and jungle crow, are potential seed dispersers, with estimated dispersal distances aligning with the longer distances between islands. However, genetic differentiation among the same ecotypes in different island groups suggests rare long-distance dispersal. Our study provides a comprehensive understanding of phylogenetic relationships, divergence patterns, and speciation processes in Callicarpa in the Bonin Islands. Moreover, it emphasizes the importance of hybridization, adaptive radiation in response to changing environments, and long-distance seed dispersal in shaping the evolutionary history of plants in oceanic islands.
ACKNOWLEDGEMENTS
The authors are grateful to K. Hayama for assistance in the field survey; Drs. H. Kudo, M. Yasugi, A. Tezuka for their experimental support; Drs. K. Uchiyama, A. Izuno, J.R.P. Worth and H. Mori for their valuable advice in SNP analysis. We also thank Metropolis of Tokyo, the Ministry of the Environmental Government of Japan, and Forestry Agency of Japan for allowing this study. This research was conducted using the Ogasawara Field Research Station of Tokyo Metropolitan University. This work was funded by Grants-in-Aid for Science Research from the Japanese Society for Promotion of Science (JP26290073, JP15K07203, JP21K05694), the Environment Research and Technology Development Fund of the Ministry of the Environment, Japan (4-1402, 4MF-2202).
REFERENCES
Abobatta, W. F. (2021) Fruit orchards under climate change conditions: adaptation strategies and management. Journal of Applied Biotechnology & Bioengineering, 8 (3), 99-102. doi:10.15406/jabb.2021.08.00260
Aedo, C., Medina, L. & Fernández-Albert, M. (2013) Species richness and endemicity in the Spanish vascular flora. Nordic Journal of Botany, 31 (4), 478-488. doi:https://doi.org/10.1111/j.1756-1051.2012.00009.x
Alexander, D. H., Novembre, J. & Lange, K. (2009) Fast model-based estimation of ancestry in unrelated individuals. Genome Research, 19 (9), 1655-1664. doi:10.1101/gr.094052.109
Ando, H., Setsuko, S., Horikoshi, K., Suzuki, H., Umehara, S., Yamasaki, M., Hanya, G., Inoue‐Murayama, M. & Isagi, Y. (2016) Seasonal and inter‐island variation in the foraging strategy of the critically endangered Red‐headed Wood Pigeon Columba janthina nitens in disturbed island habitats derived from high‐throughput sequencing. Ibis, 158 (2), 291-304
Boyle, W. A. & Conway, C. J. (2007) Why migrate? A test of the evolutionary precursor hypothesis. Am Nat, 169 (3), 344-359. doi:10.1086/511335
Carlquist, S., Baldwin, B. G. & Carr, G. D. (2003) Tarweeds & silverswords: evolution of the Madiinae (Asteraceae) . St. Louis: Missouri Botanical Garden Press.
Catchen, J., Hohenlohe, P. A., Bassham, S., Amores, A. & Cresko, W. A. (2013) Stacks: an analysis tool set for population genomics.Molecular Ecology, 22 (11), 3124-3140
Catchen, J. M., Amores, A., Hohenlohe, P., Cresko, W. & Postlethwait, J. H. (2011) Stacks: building and genotyping loci de novo from short-read sequences. G3: Genes| genomes| genetics, 1 (3), 171-182
Dickman, E. E., Pennington, L. K., Franks, S. J. & Sexton, J. P. (2019) Evidence for adaptive responses to historic drought across a native plant species range. Evol Appl, 12 (8), 1569-1582. doi:10.1111/eva.12803
Drummond A, J. (2007) BEAST : Bayesian evolutionary analysis by sampling trees. BMC Evolutionary Biology, 7 , 214
Dunning Jr, J. B. (2007) CRC handbook of avian body masses : CRC press.
Excoffier, L., Marchi, N., Marques, D. A., Matthey-Doret, R., Gouy, A. & Sousa, V. C. (2021) fastsimcoal2: demographic inference under complex evolutionary scenarios. Bioinformatics, 37 (24), 4882-4885
Franks, S. J., Kane, N. C., O’Hara, N. B., Tittes, S. & Rest, J. S. (2016) Rapid genome-wide evolution in Brassica rapa populations following drought revealed by sequencing of ancestral and descendant gene pools. Molecular Ecology, 25 (15), 3622-3631. doi:10.1111/mec.13615
Gillespie, R. G., Howarth, F. G. & Roderick, G. K. (2001) Adaptive radiation. In Levin SA (Ed.), Encyclopedia of biodiversity Vol. 1, pp. 25-44. New York: Academic Press.
Givnish, T. (Ed.) (1997) Adaptive radiation and molecular systematics: issues and approaches . Cambridge: Cambridge University Press.
Givnish, T. J., Millam, K. C., Mast, A. R., Paterson, T. B., Theim, T. J., Hipp, A. L., Henss, J. M., Smith, J. F., Wood, K. R. & Sytsma, K. J. (2009) Origin, adaptive radiation and diversification of the Hawaiian lobeliads (Asterales: Campanulaceae). Proceedings of the Royal Society B: Biological Sciences, 276 (1656), 407-416
Gossmann, T. I., Keightley, P. D. & Eyre-Walker, A. (2012) The effect of variation in the effective population size on the rate of adaptive molecular evolution in eukaryotes. Genome Biology and Evolution, 4 (5), 658-667
Grant, P. R. (1986) Ecology and Evolution of Darwin’s Finches (Princeton Science Library Edition) (REV - Revised ed.). Prinston: Princeton University Press.
Grant, P. R. (1998) Evolution on islands . Oxford: Oxford University Press.
Grant, P. R. & Grant, B. R. (2006) Evolution of Character Displacement in Darwin’s Finches. Science, 313 (5784), 224-226. doi:doi:10.1126/science.1128374
Grant, P. R. & Grant, B. R. (2007) How and why species multiply: the radiation of Darwin’s finches . Princeton: Princeton University Press.
Higuchi, Y. (1984) List of birds in the Ogasawara Islands, including Iwo Islands and Minamitori-shima. Strix, 3 , 73-87
Huson, D. H. & Bryant, D. (2006) Application of phylogenetic networks in evolutionary studies. Molecular Biology and Evolution, 23 (2), 254-267. doi:10.1093/molbev/msj030
Ito, M. & Ono, M. (1990) Allozyme diversity and the evolution ofCrepidiastrum (Compositae) on the Bonin (Ogasawara) Islands.The botanical magazine = Shokubutsu-gaku-zasshi, 103 (4), 449-459. doi:10.1007/BF02491263
Ito, M., Soejima, A. & Ono, M. (1997) Allozyme diversity ofPittosporum (Pittosporaceae) on the Bonin (Ogasawara) Islands.Journal of Plant Research, 110 (4), 455-462
Johnson, S. E., Hamann, E. & Franks, S. J. (2022) Rapid, parallel evolution of field mustard (Brassica rapa) under experimental drought.Evolution, 76 (2), 262-274. doi:10.1111/evo.14413
Johnson, S. E., Tittes, S. & Franks, S. J. (2023) Rapid, nonparallel genomic evolution of Brassica rapa (field mustard) under experimental drought. Journal of Evolutionary Biology, 36 (3), 550-562. doi:10.1111/jeb.14152
Kadereit, J. W. & Abbott, R. J. (2021) Plant speciation in the Quaternary. Plant Ecology & Diversity, 14 (3-4), 105-142. doi:10.1080/17550874.2021.2012849
Kagawa, K. & Takimoto, G. (2018) Hybridization can promote adaptive radiation by means of transgressive segregation. Ecology Letters, 21 (2), 264-274. doi:https://doi.org/10.1111/ele.12891
Kopelman, N. M., Mayzel, J., Jakobsson, M., Rosenberg, N. A. & Mayrose, I. (2015) Clumpak: a program for identifying clustering modes and packaging population structure inferences across K. Molecular Ecology Resources, 15 (5), 1179-1191
Lamichhaney, S., Han, F., Berglund, J., Wang, C., Almén, M. S., Webster, M. T., Grant, B. R., Grant, P. R. & Andersson, L. (2016) A beak size locus in Darwin’s finches facilitated character displacement during a drought. Science, 352 (6284), 470-474. doi:doi:10.1126/science.aad8786
Marijon, P., Chikhi, R. & Varré, J.-S. (2020) yacrd and fpa: upstream tools for long-read genome assembly. Bioinformatics, 36 (12), 3894–3896
Martinson, D. G., Pisias, N. G., Hays, J. D., Imbrie, J., Moore, T. C. & Shackleton, N. J. (1987) Age dating and the orbital theory of the ice ages: Development of a high-resolution 0 to 300,000-year chronostratigraphy. Quaternary Research, 27 (1), 1-29. doi:https://doi.org/10.1016/0033-5894(87)90046-9
McDade, L. A. (1992) Hybrids and phylogenetic systematics II. The impact of hybrids on cladistic analysis. Evolution, 46 (5), 1329-1346
Milligan, B. G. (1992) Plant DNA isolation. Molecular genetic analysis of populations: a practical approach. , 59-88
Ministry of the Environment Government of Japan (2020) Red List. Retrieved fromhttps://www.env.go.jp/content/900515981.pdf
Nishimura, O., Hara, Y. & Kuraku, S. (2017) gVolante for standardizing completeness assessment of genome and transcriptome assemblies.Bioinformatics, 33 (22), 3635-3637. doi:10.1093/bioinformatics/btx445
Nylander, J. (2002) MrModeltest v1. 0b. Program distributed by the author. Department of Systematic Zoology, Uppsala University
Olson, M. E., Soriano, D., Rosell, J. A., Anfodillo, T., Donoghue, M. J., Edwards, E. J., León-Gómez, C., Dawson, T., Camarero Martínez, J. J., Castorena, M., Echeverría, A., Espinosa, C. I., Fajardo, A., Gazol, A., Isnard, S., Lima, R. S., Marcati, C. R. & Méndez-Alonzo, R. (2018) Plant height and hydraulic vulnerability to drought and cold.Proceedings of the National Academy of Sciences of the United States of America, 115 (29), 7551-7556. doi:10.1073/pnas.1721728115
Ono, M. (1991) The Flora of the Bonin (Ogasawara) Islands. Aliso: A Journal of Systematic and Floristic Botany, 13 (1), 95-105
Ono, M., Kobayashi, S. & Kawakubo, N. (1986) Present situation of endangered plant species in the Bonin Islands. Ogasawara Research, 12 , 1-32
Peterson, B. K., Weber, J. N., Kay, E. H., Fisher, H. S. & Hoekstra, H. E. (2012) Double digest RADseq: an inexpensive method for de novo SNP discovery and genotyping in model and non-model species. PloS One, 7 (5), e37135
Porter, D. M. (1978) Endemism and evolution in Galapagos Islands vascular plant. Plants and islands. , 225-258
Puritz, J. B., Hollenbeck, C. M. & Gold, J. R. (2014) dDocent: a RADseq, variant-calling pipeline designed for population genomics of non-model organisms. PeerJ, 2 , e431
Rochette, N. C., Rivera‐Colón, A. G. & Catchen, J. M. (2019) Stacks 2: Analytical methods for paired‐end sequencing improve RADseq‐based population genomics. Molecular Ecology, 28 (21), 4737-4754
Ronquist, F., Teslenko, M., van der Mark, P., Ayres, D. L., Darling, A., Höhna, S., Larget, B., Liu, L., Suchard, M. A. & Huelsenbeck, J. P. (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology, 61 (3), 539-542. doi:10.1093/sysbio/sys029
Ruan, J. & Li, H. (2020) Fast and accurate long-read assembly with wtdbg2. Nat Methods, 17 (2), 155-158. doi:10.1038/s41592-019-0669-3
Rundle, H. D. & Nosil, P. (2005) Ecological speciation. Ecology Letters, 8 (3), 336-352
Schluter, D. (2000) The ecology of adaptive radiation . Oxford, UK: Oxford Universiity Press.
Setsuko, S., Ohtani, M., Sugai, K., Nagamitsu, T., Kato, H. & Yoshimaru, H. (2017) Genetic variation of pantropical Terminalia catappa plants with sea-drifted seeds in the Bonin Islands: suggestions for transplantation guidelines. Plant Species Biology, 32 (1), 13-24. doi:https://doi.org/10.1111/1442-1984.12121
Setsuko, S., Sugai, K., TAMAKI, I., Hayama, K. & Kato, H. (2023) Local environmental adaptation contributes to the maintenance of ecotypes ofCallicarpa subpubescens (Lamiaceae), in spite of frequent hybridization and low pre- and post-mating barriers. Authorea . doi:10.22541/au.167481187.76575231/v2
Setsuko, S., Sugai, K., Tamaki, I., Takayama, K. & Kato, H. (2022) Contrasting genetic diversity between Planchonella obovata sensu lato (Sapotaceae) on old continental and young oceanic island populations in Japan. PloS One, 17 (9), e0273871. doi:10.1371/journal.pone.0273871
Setsuko, S., Sugai, K., Tamaki, I., Takayama, K., Kato, H. & Yoshimaru, H. (2020) Genetic diversity, structure, and demography of Pandanus boninensis (Pandanaceae) with sea drifted seeds, endemic to the Ogasawara Islands of Japan: Comparison between young and old islands.Molecular Ecology, 29 (6), 1050-1068
Shibazaki, F. & Hoshi, Y. (2006) Japanese Wood Pigeons (Columba janthina nitens ) as a seed disperser in the Ogasawara Islands, southern Japan. Strix, 24 , 171-176
Shimizu, Y. (1999) Natural History of the Ogasawara Islands.Forest Science (Shinrin Kagaku), 25 , 42-45
Shimizu, Y. (2003) The nature of Ogasawara and its conservation.GLOBAL ENVIRONMENTAL RESEARCH-ENGLISH EDITION-, 7 (1), 3-14
Shimizu, Y. & Tabata, H. (1991) Forest structure, composition, and distribution on a Pacific island, with reference to ecological release and speciation.
Simão, F. A., Waterhouse, R. M., Ioannidis, P., Kriventseva, E. V. & Zdobnov, E. M. (2015) BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics, 31 (19), 3210-3212. doi:10.1093/bioinformatics/btv351
Soejima, A., Nagamasu, H., Ito, M. & Ono, M. (1994) Allozyme diversity and the evolution of Symplocos (Symplocaceae) on the Bonin (Ogasawara) Islands. Journal of Plant Research, 107 , 221-227
Sugai, K., Mori, K., Murakami, N. & Kato, H. (2019) Strong genetic structure revealed by microsatellite variation in Callicarpaspecies endemic to the Bonin (Ogasawara) Islands. Journal of Plant Research, 132 (6), 759-775. doi:10.1007/s10265-019-01144-4
Sugai, K., Setsuko, S., Nagamitsu, T., Murakami, N., Kato, H. & Yoshimaru, H. (2013) Genetic differentiation in Elaeocarpus photiniifolia (Elaeocarpaceae) associated with geographic distribution and habitat variation in the Bonin (Ogasawara) Islands. J Plant Res, 126 (6), 763-774
Swofford, D. L. (2002) PAUP: phylogenetic analysis using parsimony (and other methods), version 4.0 beta. http://paup. csit. fsu. edu/
Tennekes, H. (2009) The simple science of flight: from insects to jumbo jets . Massachusetts, USA: MIT press.
Toyoda, T. (2003) Flora of Bonin Islands . Kanagawa, Japan: Aboc-sha.
Toyoda, T. (2014) The endemic plants of the Bonin Islands . Kanagawa, Japan: Woodspress.
Wagner, C. E., Keller, I., Wittwer, S., Selz, O. M., Mwaiko, S., Greuter, L., Sivasundar, A. & Seehausen, O. (2013) Genome‐wide RAD sequence data provide unprecedented resolution of species boundaries and relationships in the L ake Victoria cichlid adaptive radiation.Molecular Ecology, 22 (3), 787-798
Yoshikawa, T., Kawakami, K. & Masaki, T. (2019) Allometric scaling of seed retention time in seed dispersers and its application to estimation of seed dispersal potentials of theropod dinosaurs. Oikos, 128 (6), 836-844. doi:https://doi.org/10.1111/oik.05827
DATA ACCESSIBILITY AND BENEFIT-SHARING
Genotype data has been deposited at FigShare: https://figshare.com/account/articles/24356665. The PacBio Sequel raw reads are available at NCBI Sequence Reads Archive (DRA017215), and the reference genome sequence of C. subpubescence has been deposited at DDBJ/EMBL/GenBank under the accessions BTTA01000001–BTTA0106011.
AUTHOR CONTRIBUTIONS
SS, SN, IY and HK designed the research. SS, SN, KS and HK sampled materials. SN and AJN performed the laboratory work. TUI performed de novo genome assembly. SS and IT performed data analysis. All co-authors discussed the results. SS and IT wrote the paper.
Table 1 Population characteristics of the three Callicarpaspecies in the Bonin Islands examined in this study.