Phylogeographic and population genetic characteristics of a Korean endemic freshwater fish, Zacco koreanus

김유림 상지대학교 일반대학원 2021 국내석사

The formation of major river systems in the Korean Peninsula, including the Han River, Nakdong River, Geum River, and East-flowing river was influenced by the interplay between geological and glacial activities, which may lead to geographical isolation among freshwater fish populations in nature that were once connected physically. Limited gene flow among populations owing to the physical barrier, along with divergent natural selection in response to different local environments may in concert drive allopatric speciation through the build-up of genetic and ecological divergences, giving rise to approximately 67 Korean endemic freshwater fish species. Zacco koreanus (family Cyprinidae, genus Zacco), a Korean endemic freshwater fish, is widely distributed in most of the upper- and midstreams of rivers, and is suggested to represent three ecotypes according to distinct morphological characteristics in relation to their geographical distribution. In this study, I aimed to determine the phylogenetic relationships among four geographically separated river basins [Han River (HR; number of populations, N=10), East-flowing river (ER; N=4), Geum River (GR; N=2) and Nakdong River (NR; N=7)], and also to evaluate the population genetic structure among the four major river basins, using mitochondrial DNA COI (cytochrome oxidase I) and CR (control region). I found three distinct clades from 23 populations comprising 538 individuals analyzed; the HR formed a monophyletic lineage with the ER, and the GR and NR formed distinct clades, respectively. The NR populations represented a divergent lineage with a mean of 2.9% (2.6~3.4%) in genetic distances from the other lineages. Nevertheless, some populations (e.g. ODHR, TBNR, WAHR and etc.) were admixed with individuals of different lineages. Inter-population pairwise FST showed high levels of genetic differentiation with a range of FST= −0.01~0.96 (mean FST= 0.54), suggesting a generally low degree of gene flow occurring among/within populations/rivers, most likely due to geographic isolation acting as a physical barrier. I found a number of cases, where Z. koreanus individuals that had been originated from different clades of nonnative populations existed in certain riverine populations, which is most likely to result from artificial translocation by humans. In conclusion, I highlight that the NR populations represent their own distinct lineage, suggesting the possibility of cryptic or a species complex evolved in Z. koreanus. A further research is necessary to be performed to re-evaluate the taxonomic position of the NR populations by performing an integrated study of genomics and ecology. The possible adverse effects of human-induced hybridization or introgression between individuals from genetically different lineages on Z. koreanus and the resulted impacts upon the existing ecosystems are also needed to be studied. 한반도의 주요 강·하천은 지각변동과 빙하기의 영향으로 지리적인 격리가 뚜렷해지게 되었으며, 이는 개체군 간에 제한된 유전적확산과 서로 다른 환경에 대한 자연선택으로 인해 이소적 종분화를 일으켜 약 67종의 한국 고유담수어종의 분화를 야기했다. 한국 고유담수어종인 참갈겨니(Zacco koranus)는 잉어과 피라미속에 속하며, 대부분 하천의 중·상류에 널리 분포하고, 지리적 분포와 관련된 뚜렷한 형태적 특성에 따라 3개의 생태형을 나타낸다고 보고되었다. 본 연구에서는 미토콘드리아 DNA COI (cytochrome oxidase I)과 CR (control region) 부위를 분자마커로 이용하여, 지리적으로 격리된 수계[한강(HR; N, 개체군 수=10), 동해유입하천(ER; N=4), 금강(GR; N=2), 낙동강(NR; N=7)]에 분포하는 참갈겨니 개체군 간의 계통유연관계를 이해하고, 수계 간의 유전적 구조를 파악하였다. 총 538개체로부터 75개의 단상형이 확인되었으며, 한강수계의 개체군은 다른 수계 개체군보다 다양성이 높게 확인되었다(단상형풍부도; P=0.015). 수계 내의 개체군간 유전적 변이보다 수계 간 유전적 구조의 차이가 더 중대하게 존재함을 확인하였으며, 유전적 분화도(FST)는 -0.01~0.96 범위로 높게 나타나 개체군 및 하천 간에 지리적 장벽으로 인해 유전자 확산이 제한적임을 확인하였다. 이러한 양상은 국립공원내에서도 확인되었으며, 같은 국립공원내에 지리적으로 매우 가깝게 위치한 하천일지라도(6~20 km) 수계가 다름에 따라 상당한 유전적 차이(FST=0.00~0.92)를 나타내었다. 또한, 계통발생학적 분석을 통해서 비교적 명확하게 구분되는 3개의 계통이 지리적으로 격리된 서로 다른 하천에서 진화하였음이 관찰되었다. HR과 ER은 단계통군을 형성하였으며, GR과 NR은 각각 별개의 계통을 형성하였다. 특히, 낙동강 개체군은 다른 계통과 평균 2.9% (2.6~3.4%)의 유전적 거리를 가지고 있는 분화된 계통을 나타내었다. 이정도의 유전적 거리 수준을 고려하면 낙동강 수계 개체군들은 참갈겨니 내 잠재종 가능성을 시사한다. 계통이 수계 별로 명확히 구분되었음에도 불구하고 일부 집단은 수계 간 계통이 혼재되어 있음을 확인하였으며, 이는 인간에 의한 인위적인 이입의 영향으로 추정된다. 본 연구결과를 바탕으로 낙동강수계의 참갈겨니를 대상으로 유전체 및 생태학적 연구를 통합적으로 수행함으로써 분류학적 위치를 재평가할 필요가 있을 것으로 사료된다. 또한, 최근 인간에 의한 무분별한 이입으로 인한 서로 다른 수계의 고유 계통간 교잡, 유전적 오염 현상 등에 의한 참갈겨니의 생태에 미치는 효과 및 생태계에 미칠 부정적 영향도 중요한 향 후 연구주제로 생각된다.

Molecular phylogenetic analysis of green tide forming macroalga Ulva community structure and species diversity along the southern coasts of Korea

박혜진 상지대학교 일반대학원 2023 국내석사

In recent years, ‘macroalgal blooms’, or often called ‘seaweed tides’ frequently occur in coastal oceans worldwide, which is perhaps attributed to accelerating climate change as well as increasing human activity. “Green tide” events caused by the proliferation of green macroalgae (genus Ulva) not only damage the local economy but also disturb the coastal ecosystems, resulting in the loss of regional biodiversity. This phenomenon is of growing concerns because of its adverse ecological and economic outcomes. In Korean waters, green macroalgal blooms have been observed continuously throughout the year, particularly along the northeastern coastlines of Jeju Island since the 2000s. More recently, however, local green tide incidences can sometimes be seen in the southern and eastern coasts of the mainland (the South Sea and East Sea), near nutrient-rich environments, such as seashores and/or harbor ports largely affected by human-mediated nutrient loads. Which Ulva species primarily contribute to the local green tides in Korean waters, however, remain largely unexplored. Whether geographic and seasonal variation exists in the Ulva community structure and species diversity remain also unknown. Ulva species are notorious to be difficult for morphology-based species identification due to their high degree of phenotypic plasticity. In this study, in order to investigate the Ulva community structure on Jeju Island and the South Sea between 2019 and 2021 years, the chloroplast barcode tufA gene was sequenced and phylogenetically analyzed for a total of 966 specimens from 46 sites. In addition, nuclear 5s rDNA marker was further analyzed for 90 Ulva specimens to delimitate the species belonging to the LPP (Ulva linza-prolifera-procera) clade. The primary objective of this study was to assess the Ulva community structure, species composition, and seasonal variation in Jeju Island and in the South Sea based on molecular phylogenetic analyses. I found that there was a difference in the Ulva community structure between Jeju Island and the South Sea, and also observed that there was seasonal variation in species composition for both regions. A total of 9 Ulva species were found in Jeju Island, with Ulva ohnoi (43.7%) and Ulva pertusa (synonym, Ulva australis; 29.3%) being the predominant species throughout the year. U. ohnoi showed a sharp increase in relative frequency (59.2%) during autumn relative to U. pertusa. In the South Sea, U. pertusa (34.8%) and LPP sp 1. (; U. linza) (25.2%) were the most dominant species, with LPP sp 1. (; U. linza) absent during the autumn season. Ulva ohnoi was rarely found except for in autumn (43.1%). Overall, considerable spatio-temporal variation in species composition and distribution was observed between the two regions. The findings of this study will provide valuable information on molecular genetic data of the genus Ulva occurring on the coasts of Korea, which will assist in developing management strategy for mitigating the negative influences of green tide events on the Korean coast. Furthermore, this study provides valuable information on the genetic diversity of the genus Ulva in this region, which can serve as a foundation for future investigations on the taxonomy, ecology, and conservation of Ulva species in coastal ecosystems.

(The) 'golden tide' seaweed Sargassum horneri in Korean coastal waters : identification of genetic origins of floating populations and genetic comparisons between floating and benthic populations

변서연 Sangji University 2019 국내석사

Recently, brown seaweeds Sargassum horneri blooming on the eastern coast of China drifted in large biomass with oceanic currents (called “golden tide”) and subsequently floated along the southern coastlines Korea including Jeju Island. These floating populations posed a large threat to both the local economy and coastal ecosystems. I aimed to track geographic or genetic origins of the floating S. horneri populations and to investigate evolutionary relationships and the genetic structure between floating (N=14) and Korean benthic (N=5) populations using two genetic markers, such as mitochondria (mt) DNA cox3 (cytochrome c oxidase subunit 3) and seven microsatellites. Results of genetic structure analysis with microsatellites clearly revealed two separate genetic clusters of each comprised of floating and benthic populations. High levels of inter-population differentiation were detected within Korean benthic samples. Given a shared mtDNA haplotype and oceanic circulation systems, the floating populations may have been originated from the southeastern coast of China (e.g. Zhoushan, Zhejjiang province). The floating populations from the same periods during a 2015-2018 year were genetically more different from one another than those from different periods, suggesting that the floating populations might be of multiple genetic sources within the geographic origin(s). However, my phylogeographic analysis showed that variation in mtDNA cox3 is not sufficiently enough to distinguish floating from Korean benthic populations. Therefore, I further analysed sequence polymorphisms in the whole organelle genomes of a Korean benthic sample (from Jindo Island in the South Sea) in comparison to Chinese sample’s genomes to trace genetic origins of floating biomass more precisely. Using mitochondrial and chloroplast genomes of S. horneri from Nanji Island, Zhejiang in China as a reference, I successfully mapped a mitochondrial genome of 34,620 bp and a chloroplast genome of 124,068 bp for the Korean benthic sample. I found that although the mt genome had a shorter length in size than the chloroplast genome, it had considerably more variable sites. MtDNA cox2 (cytochrome c oxidase subunit 2) gene showed the highest number of six Single Nucleotide Polymorphism (SNP) sites and an inter-genic region between nad7 (NADH dehydrogenase subunit 7) and trnP (proline tRNA) genes showed a 14 bp insertion/deletion mutation (indel). I am currently testing those ‘candidate’ genetic markers (an indel and SNPs) on the mt genome for discriminating between floating and Korean benthic samples. My thesis will inform management efforts including the development of “S. horneri blooming forecasting system”, which will assist in mitigating ecological and economic damages on the Korean coastal ecosystems in the future.

DNA 수선 유전자 APE1 저해제 발굴을 위한 Pharmacophore modeling 및 가상스크리닝

이인원 상지대학교 일반대학원 2017 국내석사

AP Endonuclease I(APE1)는 DNA에서 염기가 없는 부위를 인지하여 염기를 수리하는 DNA 염기 절단 수리 경로(BER)에 관여하는 효소이다. APE1은 정상세포 뿐만 아니라 종양세포에서도 염기 수선이 이루어지며 저해제를 통해 종양세포에서의 APE1 작용을 억제하여 항암치료를 위한 약물 타겟으로 연구되고 있다. 새로운 APE1 저해제를 개발함에 있어 전체 과정 중 저해제로서 활용 가능한 후보군을 선정하는 후보물질 탐색 단계가 매우 중요하다. 그래서 시간과 비용을 줄이고 가능성 있는 후보군을 선정하기 위하여 컴퓨팅 기법이 사용되고 있다. 본 연구에서는 APE1을 저해할 가능성이 높은 화합물들을 찾기 위하여 PDB에서 APE1을 포함하는 단백질 구조와 ZINC database에서 신약 개발 단계에서 약리학적 기능 및 생물학적 활성을 가지는 Leads-Now subset을 Docking과 Pharmacophore modeling 기법으로 연구를 실시하였다. Docking과 Pharmacophore modeling을 수행하여 얻은 binding affinity와 pharmacophore score 결과를 종합하여 각각의 결과에서 교집합에 해당하는 화합물들을 최종 후보물질로 선정하였다. Docking의 결과로 4,220,644개, Pharmacophore modeling의 결과로 3,718,602개가 확인되었다. 두 결과의 교집합은 pharmacophore score에 따라 6개로 그룹으로 나뉘었으며, pharmacophore score를 기준으로 하였을 때 pharmacophore score가 가장 높은 그룹이 APE1을 저해할 가능성이 있는 화합물들로 판단된다. Docking과 Pharmacophore 결과에서 상위 20,000개를 비교하여 교집합에 해당하는 25개의 화합물을 얻었다. 25개의 화합물들 중 pharmacophore score가 가장 높은 그룹에 속하는 14개의 화합물들이 APE1 저해제로써 가장 가능성이 높은 후보물질로 예측되어 14개의 화합물들을 최종 후보물질로 선정하였고, 14개의 화합물들이 APE1을 저해할 가능성이 높으며 실험할 가치가 있다고 판단되어 이후 관련 연구들에서 시작점이 될 수 있다고 생각된다.

Understanding speciation process of Korean endemic fat minnow (Rhynchocypris kumgangensis) and its close relative, Deogyu fat minnow (Rhynchocypris deogyuensis), based on genetic (genomic) and morphological analyses

황순영 상지대학교 일반대학원 2023 국내석사

Natural populations of freshwater fishes occurring in geographically isolated river drainages are restricted to migrate from one river/stream to another, often resulting in ecological/genetic divergence in response to diversifying selection, the formation of ecotypes and even allopatric speciation. However, evidence for the speciation through allopatric processes in freshwater fish systems remains still scarce. Kumkang fat minnow, Rhynchocypris kumgangensis is a Korean endemic coldwater fish that occurs only in the uppermost regions of stream or river. Rhynchocypris kumgangensis, which inhabits the Gucheondong Valley of the Deogyusan National Park, was recently classified as a new species, Deogyu fat minnow, Rhynchocypris deogyuensis in 2017 based on genetic and morphological analyses. However, whether they are separate species and what the level of their genetic and ecological (morphological) divergence is remain unclear. I here analyzed the population genetic structure of R. kumgangensis with respect to R. deogyuensis using mitochondrial DNA (mtDNA) cytochrome oxidase I (COI) and control region (CR) sequences and my newly developed eight microsatellite loci. Moreover, the level of genetic divergence was quantitatively assessed between the presumed two species at the whole mitogenome level. I further assessed quantitatively the morphology between R. kumgangensis and R. deogyuensis by analyzing metric (morphological) traits and also geometric morphometrics on body shape. I found that mtDNA and microsatellite diversities were considerably higher for 11 populations of R. kumgangensis than one population of R. deogyuensis. Rhynchocypris deogyuensis population harbored only a single haplotype and showed a very high level of inbreeding (FIS=0.25). The comparative analysis of the mitogenomes showed approximately 2.9% difference (pairwise identity = 97.1%) between R. kumgangensis and R. deogyuensis, supporting the species-level divergence, given the suggested >2-3% genetic distance for species identification in most animals. Only four of seven metric traits examined significantly differed between R. kumgangensis and R. deogyuensis. However, analysis of geometric morphometrics on body shape indicated significant morphological divergence between the two species. Lastly, the observed low values of both length-weight relationship and condition factor particularly for R. deogyuensis population are likely to be attributed to combined effects of the elevated level of inbreeding and depleted genetic diversity. Overall, our combined genetic, genomic and morphological analyses suggest the considerable divergence between R. kumgangensis and R. deogyuensis, supporting the hypothesis that they are indeed distinct species. The mechanisms underpinning how they speciate still need to be studied further in detail.

납자루아과(Pisces: Acheilognathinae) 담수어류 3종의 난과 치어 종 동정을 위한 분자마커 개발 및 숙주조개(작은말조개 : Unio douglasiae sinuolatus)에 대한 산란양상

최희규 상지대학교 2019 국내석사

잉어과(Cyprinidae) 납자루아과(Acheilognathinae) 어류는 몸이 납작하고 체고가 높은 소형 담수어류로서 우리나라에는 2속 14종이 기록되어 있다. 납자루아과 어류는 담수산 석패과(Bivalvia: Unionidae) 패류의 아가미 안에 산란하는 독특한 생물학적 특징을 가지고 있으며, 이러한 독특한 번식 행동을 가진 담수어류 종은 생활사 초기단계가 숙주조개에 전적으로 의존되기 때문에 서식처 환경에 숙주조개 개체 수 감소 시 멸종위기 가능성이 높아진다. 최근 인간의 인위적인 간섭에 의한 오염 및 하천사업으로 인해 납자루아과가 숙주로 이용하는 석패과 패류의 개체수가 급격히 감소하고 있으며, 이로 인해 납자루아과의 개체수가 전 세계적으로 감소하고 있는 추세이다. 국내의 경우 납자루아과 담수어류에 대한 산란양상을 파악하기 위한 생태학적 연구가 진행된 바 있으나, 아직 유전학적인 방법을 이용한 숙주조개 속 난 및 치어에 대한 종 판별 연구는 수행되지 않았다. 따라서, 본 연구에서는 납자루아과 어류의 생태학적인 연구에 PCR 기반 RFLP (Restriction Fragment Length Polymorphism; 제한절편 길이 다형성) 분자기법을 활용하여 난 및 치어 대상 납자루아과 어류 3종의 동정을 좀 더 빠르고 정확하게 파악하고 납자루아과 어류의 종별 산란양상 및 번식생태 이해에 대한 기여가 목적이다. 연구를 위한 조사지역은 강원도 홍천 내촌천(HN), 덕치천(HD), 정선 골지천(JG) 및 조양강(JJ)을 대상으로 납자루아과 어류의 산란양상를 알아보고자 하였다. 연구대상 지역 내 납자루아과 어류는 내촌천과 골지천에서 1종(묵납자루; Acheilognathus signifer), 덕치천에서 3종(각시붕어; Rhodeus uyekii, 묵납자루; A. signifer, 줄납자루; A. yamatsutae), 조양강에서 2종(묵납자루, 줄납자루)으로 확인되었고, 네 집단에서 모두 동서하고 있는 숙주조개인 작은말조개(Unio douglasiae sinuolatus) 982개체를 함께 채집하였다. 제한절편 길이 다형성 분자마커를 이용하여 납자루아과 어류의 난 및 치어로 확인되는 작은말조개(N=163)에서 총 646개의 납자루아과 어류의 난 및 치어(묵납자루 454개체, 줄납자루 43개체, 각시붕어 149개체)를 확인하였다. 각 집단 간 숙주조개 크기에 따른 산란선호도를 조사하기 위해 난 및 치어가 확인된 숙주조개(mussels with eggs and/or fry)와 확인되지 않은 숙주조개(mussels without eggs and/or fry)의 각장(shell length), 각고(shell height) 및 각폭(shell width)의 평균 크기를 비교 분석하였다. 그 결과 3종의 납자루아과 어류가 동서하는 덕치천의 경우, 난 및 치어가 확인된 숙주조개가 확인되지 않은 숙주조개보다 각장(1.98mm), 각고(0.85mm) 및 각폭(0.73mm)의 크기가 통계적으로 유의하게 크게 나타났으며(Mann-Whitney, P = 0.002, P = 0.012, P = 0.009), 다른 세 개의 집단에서도 통계적으로 유의하지는 않았으나 유사한 경향의 결과가 관찰되었다. 이러한 결과는 본 연구 대상 납자루아과 어류 3종은 크기가 큰 작은말조개를 산란을 위한 숙주로서 선호 함을 의미하고, 이러한 조개 크기에 대한 선호도가 동서하는 종의 수가 많을수록 더 강화됨을 시사한다. 또한, 2종 이상의 납자루아과 어류가 서식하는 덕치천(HD)과 조양강(JJ)에서 묵납자루와 줄납자루 2종이 동일한 조개에 산란하는 것이 관찰되었다(N = 4). 이는 납자루아과 어류가 2종이상 동서할 때, 동일한 자원인 작은말조개를 자신의 산란숙주로 이용하기 위한 종간경쟁(interspecific competition)의 가능성을 시사한다. 기존 연구된 생태학적 연구에 유전학적인 방법을 추가하여 각 집단 간, 종 간 숙주조개 크기에 대한 산란양상을 보다 정확히 규명하였으며, 이러한 연구 결과를 바탕으로 향후 이들 자연개체군의 효과적인 유지, 관리 및 보전 방법 개발에 활용할 수 있는 과학적 기초 데이터를 제공하고자 한다. For freshwater bitterling fishes (Cyprinidae, Pisces), 14 different species have been described so far from South Korea. During a spawning season, females develop a long ovipositor that they use to lay eggs in the gill cavities of mussels through the mussels’exhalant siphon. This unique spawning behavior renders these fishes likely to become endangered as host mussel populations are declining. During recent years, because of increasing human impacts, freshwater mussels have significantly decreased in population size, and several species are now under threat of extinction globally. In this study, I investigated spawning patterns of three bitterling fishes (Pisces: Acheilognathinae), including Acheilognathus signifer, A. yamatsutae and Rhodeus uyekii, on the host mussels (Unio douglasiae sinuolatus). This study was conducted at four streams [Hongcheon Naechoncheon (HN); Deokchicheon (HD) from the North Han River basin; Jeongseon Goljicheon (JG) and Joyanggang (JJ) from the South Han River] in South Korea. Only A. signifer occurred at two sites of HN and JG, whereas A. signifier, A. yamatsutae and R. uyekii co-existed in HD and A. signifier and A. yamatsutae co-occurred at JJ. Among the sampled mussels (N = 982) from four populations, a total of 646 eggs/fry were observed within the mussels (N = 163; 16.6%) and identified as A. signifer (N = 454; 70.3%), A. yamatsutae (N = 43; 6.6%) and R. uyekii (N = 149; 23.1%). A species identification for the eggs and fry of these three species was done by my developed PCR-based RFLP (Restriction Fragment Length Polymorphism). To investigate the spawning preference of the Acheilognathinae fishes in relation to the size of the mussels, I estimated and compared length, height and width of the mussels between those with/without the eggs and/or fry. For HD where three species co-existed, the mussels having the eggs and fry were significantly larger in the length (Mann-Whitney U, P = 0.002; difference = 1.98 mm), height (P = 0.012; 0.85 mm) and width (P = 0.009; 0.73mm) than mussels not having them. However, this trend was not significant in the other three sites. These findings indicate that my study species of the Acheilognathinae fishes tend to prefer larger mussels as their spawning hosts, and this tendency increases with a number of coexisting bitterling fish species. In HD and JJ where more than two bitterling species occurred, I found mussels (N = 4) having the eggs/fry of both A. signifier and A. yamatsutae. These results suggest the possibility of interspecific competition between the Acheilognathinae fishes for utilizing the same resource of mussels for their spawning. My thesis helps to better understand the spawning patterns and reproductive ecology of the Acheilognathinae fishes, which will provide important information for future maintenance, management and conservation of these natural, but endangered species.

Molecular ecological investigations of the golden tide forming Sargassum horneri in northwestern Pacific: understanding the origins and mechanisms

변서연 상지대학교 일반대학원 2023 국내박사

Genetic structure and ecological characteristics among local populations of a Korean endangered freshwater fish, Brachymystax lenok tsinlingensis: implications for its effective conservation and restoration strategies : 한국 멸종위기 담수어종 열목어(Brachymystax lenok tsinlingensis) 집

장지은 상지대학교 일반대학원 2017 국내석사

During recent years, indigenous freshwater fishes have profoundly decreased in population size, and a number of species are now under threat of extinction globally. To mitigate this breakdown of endemic fish populations, the use of translocation of animals to natural habitats to which they are not native as a conservation tool is a central issue under debate in conservation biologists, but may be the ‘unavoidable’ only choice where management augmentation of the target population is not possible naturally in practice. The Manchurian trout or lenok, Brachymystax lenok tsinlingensis (family Salmonidae), is a cold freshwater fish endemic to Northeast Asia. South Korea lenok populations that comprise the southern range edge of this species have recently decreased strikingly in size primarily due to anthropogenic pressure such as habitat destruction, agricultural run-off, water pollution and possibly climate change, and are now becoming critically endangered. To recover the number of individuals of B. lenok tsinlingensis, stocking or translocation programs have routinely been conducted by local governments since 1980s in South Korea, but their source population was not recorded, so it remains unknown. Moreover, several local sites have been repopulated using stocking via the release of hatchery-reared or -bred fish or translocation of individuals from non-native habitats. However, whether and how these translocated populations have successfully been established in novel environmental conditions and have ecologically and/or genetically diverged from native populations remain untested. In my thesis, I assessed the current population status of B. lenok tsinlingensis in South Korea by estimating the levels of genetic diversity and genetic structure for six natural (Yanggu, Injae, Seorak, Bangtae, Hongcheon and Taebaek) and four restored (Pyeongchang, Chiak, Jeongseon and Bonghwa) populations using mitochondrial DNA (mtDNA) control region sequences and eight nuclear microsatellite loci. Using mtDNA control region sequences, I also performed intraspecific phylogenetic analysis to understand evolutionary history of this species in the Northeastern Asia. Moreover, I examined whether ecological divergence occurred among natural and restored populations by comparing aspects of body shape using geometric morphometric analysis. Levels of within-population genetic diversity were low suggesting past effective population sizes (Ne) have been small. Each population had one, or at most two, haplotypes within the mtDNA control region. Microsatellite allelic richness (AR) ranged from 2.38 to 4.13 (overall mean = 3.12). I also observed that South Korea lenok populations were significantly genetically isolated from one another, with private mtDNA haplotypes and microsatellite alleles, suggesting that limited gene flow has been occurring among populations. A large-scale phylogenetic analysis of Northeast Asian populations revealed that South Korean lineages are more closely related to those of China (Yellow River) than to those of North Korea and Russia. I found that five South Korea lenok populations (Bangtae, Hongcheon, Pyeongchang, Chiak and Bonghwa) analyzed are morphologically significantly differentiated one another based on my geometric morphometric analysis (canonical variate analysis; CVA, P < 0.001). By focusing on the source population of Hongcheon and the introduced population of Mt. Chiaksan National Park, I found that source and introduced populations display ecological divergence with respect to body shape and trophic level, which is perhaps due to changing environmental conditions between the two habitats during the last few decades. Overall, I suggest that future restoration efforts aimed at South Korean lenok populations should consider both the genetic and ecological (morphological) characteristics documented here, which should assist in accomplishing effective conservation strategies for this highly cherished species.

Population genetic structure of seagrass, Zostera marina, on the southern coasts of Korea (Jeju Island, South Sea): the cornerstone for its future conservation and restoration plans : 한국 두 연안(제주도, 남해)에 서식하는 잘피 거머리말 집단의 유전적 구조: 미래 보전

김재환 상지대학교 일반대학원 2017 국내석사

Seagrasses provide numerous ecosystem services for coastal and estuarine environments such as nursery functions, erosion protection, pollution filtration, and carbon sequestration. Zostera marina (common name “eelgrass”) is one of the seagrass bed-forming species distributed widely in the northern hemisphere, including the Korean Peninsula. Recently, however, there has been a drastic decline in the population size of Z. marina worldwide, including Korea. We examined the current population status of this species on the southern coast of Korea by estimating the levels of genetic diversity and genetic structure of 10 geographically disconnected populations using eight nuclear microsatellite markers. The level of genetic diversity was found to be significantly lower for populations on Jeju Island [mean allelic richness (AR) = 2.15], which is located off the southernmost region of the Korean Peninsula, than for those in the South Sea (mean AR = 3.73), which is on the southern coast of the mainland. South Korea eelgrass populations were all substantially genetically divergent from one another, suggesting that limited contemporary gene flow has been taking place among populations. We also found weak but detectable temporal variation in genetic structure within a site over 10 years. In additional depth comparisons, statistically significant genetic differentiation was observed between shallow (or middle) and deep zones in each of the three sites tested. Depleted genetic diversity, small effective population sizes (Ne) and limited connectivity for populations on Jeju Island indicate that these populations may be vulnerable to local extinction under changing environmental conditions, especially given that Jeju Island is one of the fastest warming regions around the world. Overall, our work will inform conservation and restoration efforts, including transplantation for eelgrass populations at the southern tip of the Korean Peninsula, for this ecologically highly significant species.

Morphological evolution and genetic divergence in two Korean salmonid fish species

장지은 상지대학교 일반대학원 2022 국내박사

‘진화생물학’은 세대 간에 일어나는 생물 개체군의 형태와 행동과 같은 표현형과 유전자형의 변화과정을 연구하는 학문으로서, 생물의 서식처 환경변화에 대한 적응 능력과 관련된 기작과 그 과정 및 양상을 이해하는 학문이다. 저명한 유전학자이자 진화생물학자인 Theodosius Dobzhansky가 그의 가장 유명한 에세이에서 언급했듯이 진화생물학은 모든 다양한 생물학 관련 학문의 기초가 된다. 최근에는 분자생물학 기술 발전에 힘입어 생태유전학, 집단/보전유전학, 진화유전체학, 생물정보학과 같은 다른 많은 학문 영역으로 확장되어가는 추세이다. 생물이 자신을 둘러싼 서식처 환경에 반응하여 어떻게 적응 형질(adaptive trait)이 진화하는가를 이해하는 것은 진화생물학 학문의 기초적인 매우 중요한 질문 중 하나로서 많은 과학자에 의해 연구되어 왔지만 많은 질문들이 아직 완전히 해결되지 못하였다. 적응 형질을 포함하는 표현형 진화는 종간 및 종내 다양성(예: 생태형)에 기여하는 진화 기작을 이해하는 것뿐만 아니라 동일종 내 개체군 간에서 발생하는 개체군 유전적, 표현형적 분화 및 분기과정을 추론하는데 중요하다. 또한 변화하는 환경에서 생물 개체군의 반응은 때때로 동시대의 시간규모에서 예상한 시간보다 훨씬 더 빠르게 발생할 수 있기 때문에 인위적 활동은 변화하는 환경에서 생물체의 표현형 또는 유전자형과 연관된 적응 형질의 진화를 촉진시키는 역할을 할 수 있다. 진화적 관점에서 서식지 환경에 대한 자연선택의 결과로 나타나는 담수어류에서 형태와 같은 적응 형질의 표현형적 특성 변화 및 개체군간 유전적 분화를 파악하고 이 형질과 연관된 유전자 또는 유전적 변이를 모색하는 것은 표현형 진화, 개체군 분기, 국지적 적응 및 종분화를 통합적으로 이해하기 위해 필수적이다. 본 연구에서는 다양한 표현형적 특성을 갖는 연어과 어류(family Salmonidae)를 대상종으로 하여 표현형(형태) 진화, 개체군 분기 및 적응과 관련된 중요한 진화적 질문들을 다루었다. 첫번째 장에서는 인위적 복원 사업에 의해 도입된 열목어(Manchurian trout, Brachymystax lenok tsinlingensis) 개체군을 원개체군과 비교 분석을 통하여 잠재적인 생태적 및 진화적 분화가 발생했는지 여부를 파악하였고, 실제로 지난 불과 수십 년 동안 변화된 서식지 환경에서 복원된 개체군이 표현형 적응의 결과로 형태와 영양지위에서 생태학적 분화가 발생한 것을 확인할 수 있었다. 두번째 장에서는 넓은 지리적 분포를 가진 열목어를 대상으로 종간 또는 종과 파생된 아종 및 개체군 사이의 유전적 및 형태학적 특성 차이를 비교하여 그들이 서식 환경에 어떻게 진화역사적으로 적응하였는지에 대한 분화 정도를 정량 평가하였다. 이 연구에서 한국 열목어를 아종으로 명명한 근거를 근연종과의 비교 분석을 통해서 기하형태적 및 유전(유전체)적 특성 차이를 구명하여 진화적 역사 및 분류학적 위치를 재조명하였다. 마지막으로 우리나라 자생 연어(chum salmon, Oncorhynchus keta) 개체군 보전 관리를 목적으로 오랜 기간 동안 시행되어온 산란장의 치어 방류 방법이 한반도 개체군 유전적 특성(유전적 다양성, 개체군 유전적 구조 등)에 미치는 영향을 파악하고자 하였다. 우리나라 연어 개체군은 다른 지역 개체군(일본, 러시아, 미국, 캐나다)과 비교 시 유전적 다양성이 낮으나, 종 수준 다양성에 대한 기여도가 가장 높고 고유한 유전적 특성을 가짐을 확인할 수 있었다. 또한, 우리나라 연어 개체군은 단일 유전자풀을 구성하는 비교적 혼합된 개체군 구조 특성을 가지고 있었으나 몇몇 개체군간 유전적 분화를 확인하였고, 이 결과는 향후 한반도 연어 개체군의 효과적인 보전 관리 방안 마련에 기여할 수 있을 것으로 기대한다. A major topic in the field of ‘evolutionary biology’ is involved with the processes, patterns and mechanisms underlying changes in phenotypes, such as the form and behaviour, and the associated genotypes, leading to differential fitness among living organisms over generations in nature. As a prominent geneticist and evolutionary biologist Theodosius Dobzhansky mentioned in his most famous essays, evolutionary biology serves as the basis of all diverse areas of biology. In recent years, there is an expanding tendency in new research fields in evolutionary biology, such as ecological genetics, population/conservation genetics, evolutionary genomics and bioinformatics. Mechanisms underlying how organisms become evolving adaptive traits in response to their surrounding habitat environments is one of the fundamental, but important questions in evolutionary biology. Therefore, the topic of the mechanisms driving adaptive evolution has been extensively studied by a number of researchers for long time periods, while the questions have not been fully resolved yet. Understanding how adaptive phenotypes evolve is important for not only understanding the evolutionary mechanisms contributing to inter- as well as intraspecific variability (i.e., ecotype), but also inferring the mechanisms causing differentiation and/or divergence among populations of a species. As population’s response to changing environments can sometimes occur much more rapidly than expected (at a contemporary timescale), anthropogenic activities can play a role as a facilitator in driving phenotypic or genetic adaptation of organisms responding to different environmental conditions. Determining ecological characteristics of adaptive traits that may evolve as a result of natural selection in response to changing environments and also searching for the potentially related genes and/or genetic variation are key steps to comprehend complex interactions in the environments where phenotypic evolution, genetic divergence, local adaptation and speciation may occur. In this PhD thesis, I chose salmonid fishes (family Salmonidae) as my study organisms for addressing some important evolutionary questions related to phenotypic (morphological) evolution, inter-population genetic divergence and adaptation. Salmonid fishes can be ideal to address how local adaptation and phenotypic evolution arise, as they evolve diverse forms of morphological and behavioural variants. The results of my thesis can provide a foundation for future study of ecology and evolution of indigenous freshwater fishes occurring in Korea. By using combined ecological and genetic analyses, my thesis aimed to address the questions as follows: 1) to investigate whether ecological/evolutionary differentiation has occurred in phenotypes between original (donor) and introduced (recipient) populations of the Manchurian trout (Brachymystax lenok tsinlingensis) by human-mediated translocations for conservation and restoration efforts. I found rapid ecological divergence in morphology and trophic niches in translocated populations responding to changing habitat conditions during only the last few decades; 2) to quantitatively assess the levels of genetic and morphological differentiation among closely related species, subspecies and populations using genetic/genomic and geometric morphometric analyses, in order to better understand how the Manchurian trout evolved in different regional habitats across Northeast Asia. The results of this study first provide crucial information on the evolutionary history and taxonomic status of the subspecies of Korean lenok; 3) to determine the effects of hatchery fish releasing management practices (for the purpose of increasing returning rates of wild salmon adults) on genetic characteristics, such as genetic diversity, differentiation and population structure of Korean indigenous chum salmons (Oncorhynchus keta). Overall, well admixed population structure as a single gene pool with some local-scale detectable structure was identified, which informs on the effective future management plans for this ecologically and economically valuable species. More importantly, a lower genetic diversity but higher contribution to the overall species-level diversity and also a unique genetic integrity of the Korean population highlights its evolutionary significance for species-level conservation.