Primary process of speciation in marine lakes of Palau Islands:

1. Primary process of speciation in marine lakes of Palau Islands: The case of the striped silverside, Atherinomorus endrachtensis (Atherinidae, Atheriniformes, Teleostei) Ryo Gotoh, N. Hanzawa Graduate School of Science and Engineering, Yamagata University Yamagata University

2. Background Terrestrial organisms Many study of geographical isolation have conducted. It is easy to set up geographical isolation models, such as oceanic islands. Ex) Darwin’s finch on Garapagos Islands Marine organisms ・Ocean is connected in the world ・Gene flow is occurring at the large scale Ex) Speciation caused by formation of the Isthmus of Panama ・This vicariant event had occurred several million years ago Primary process of speciation of marine organisms caused by geographical isolation is unclear We focused on marine lakes on Palau Islands

3. The formation procedure of marine lakes ・Several marine lakeshave anoxic layer (meromictic lakes) ・Toxic substances (ex. H2S) accumulate in anoxic layer ・Organisms could not migrate between marine lake and lagoon in the present Palau Islands and marine lakes ・Palau Islands are located in the West Pacific ・There are approximately 300 islands ・The limestone islands called “Rock Islands” are in the central and south parts of Palau ・There are approximately 70 marine lakes in Palau Islands

4. Organisms inhabiting marine lakes have been isolated since the lakes were formed “Geographical isolation model for marine organisms” Comparing genetic structures of lagoon populations and marine lake populations, we show primary process of speciation of marine organisms caused by geographical isolation.

5. Materials & Methods Atherinomorus endrachtensis (Atherinidae, Atheriniformes, Teleostei) Distribution Western Central Pacific: Philippines and Micronesia to northern Australia and the Solomon Islands. Biology Found in shallow coastal waters We conducted population genetic analyses based on mt DNA cyt b gene (1141bp) and control region (877bp)

6. KBB(23) Palau Islands NTOS(14) Sulawesi HLOS(15) JFOS (20) Mecherchar Island ONG (26) Ongael Island JFL (20) MEC3 Red letter shows marine lakes Bule letter shows lagoon (24) １０ｋｍ

7. Results & Discussion

8. Genetic diversity Cyt b Control region N H h π N H h π JFL204 0.284 0.026 2580.590 0.120 244 0.308 0.047 MEC3 241 0.0000.000 ONG 262 0.148 0.013 263 0.151 0.026 JFOS 2060.737 0.089 3317 0.936 0.335 2321 0.988 0.642 KBB 239 0.688 0.140 1313 1.000 0.680 NTOS146 0.747 0.099 HLOS154 0.371 0.035 1612 0.967 0.519 N: number of individuals, H: number of haplotypes, h: haplotype diversity π: nucletide diversity

9. lagoon Marine lake Genetic diversity of marine lake populations is much lower than that of lagoon populations Cyt b Control region π 0.5 π 0.5 0.5 0.5 h h

10. Palau Gene flow occurs at large scale Palau-Sulawesi =1,200km The relationships among marine lake populationsare unclear. Marine lake MEC3 JFL ONG Lagoon KBB JFOS NTOS HLOS Sulawesi Statistical parsimony network based on cyt b gene

11. JFL MEC3 ONG Lagoon Marine lake KBB MEC3 JFOS JFL NTOS ONG HLOS Sulawesi Statistical parsimony network based on control region

12. The ONG population ２４ １ １ ・One major haplotype was found ・The major haplotype was also found in lagoon population Founder effect

13. Founder effect Genetic drift Fixation Isolation

14. The ONG population ２４ １ １ ・One major haplotype was found ・The major haplotype was also found in lagoon population ・Two endemic haplotypes were found

15. CT repeat ５⇔４ The JFL and MEC3 populations ３ １６ ２０ ２ ・One major haplotype and several minor haplotypes were found ・The minor haplotypes was endemic in each marine lake and the haplotypes were derived from the major haplotype ・The number of times of CT repeat found in the JFL and the MEC3 population differs from lagoon population This strongly suggests that the minor haplotypes were emerged in marine lakes

16. Lagoon population has a large number of haplotypes It is unlikely that same haplotype was fixed in different marine lake populations by chance. However, the JFL and MEC3 population have the same major haplotype. JFL MEC3 We consider “stepwise formation hypothesis”, focusing on the facts that the two marine lakes are in the same island and differ in depth (JFL: 30m, MEC3: 20m)

17. Haplotype was fixed by founder effect Fixation index Fst 0.059(P<0.05) Migration from lagoon to JFL Migration from JFL to MEC3 Each marine lake population is isolated by anoxic layer Stepwise formation hypothesis Slightly genetic differentiation has already occurred between the JFL and the MEC3 population

18. A schematic showing the primary process of speciation in marine lake populations New mutation Founder effct Accumulated Population growth JFL population Colonized? Population growth MEC3 population Founder effct Population growth Ancestral lagoon population ONG population present ~12,000 years ago

19. Conclusion ・Genetic structure between lagoon and marine lake populations are obviouslydifferent ・Genetic structure among marine lake populations are also different ・Each marine lake population has endemic haplotypes Marine lakes are excellent model for geographical isolation of marine organisms

20. Thank you very for your attention!!