以粒線體序列探討臺灣水鹿地理親緣

Phylogeography of Formosan sambar deer inferred from polymorphism of mitochondrial DNA sequences

張郁琦(Yu-Chi Chang)

指導教授 ： 朱有田 ； 姜延年

國立臺灣大學/生物資源暨農學院/動物科學技術學研究所/碩士(2014年)

https://doi.org/10.6342/NTU.2014.10883

摘要

水鹿是亞洲鹿科動物中體型最大的水鹿，其七個亞種廣泛分布於亞洲不同地區，但在尼泊爾、印度、台灣地區之外的原生族群，都被認為已瀕臨滅絕的危機。臺灣水鹿身為臺灣野外最大的草食動物，是對棲地減少與氣候變遷敏感的指標性物種。同時，臺灣水鹿在東南亞高海拔地區的野生族群，由於氣候分明，導致其表現出特有的季節性海拔遷徙行為。關於臺灣水鹿的族群遺傳結構，以及臺灣水鹿與其他水鹿亞種的親緣關係，目前尚未有人進行深入之研究。因此，本研究欲使用水鹿的粒線體D-loop序列與粒線體全長序列作為分子標記來（1）探討臺灣水鹿與其他水鹿亞種的親緣關係；（2）分析臺灣水鹿的族群遺傳結構與親緣地理關係；（3）推測台灣水鹿的歷史族群數量變動趨勢。
本研究分析779個臺灣水鹿樣本，樣本分別來自雪霸國家公園、太魯閣國家公園、玉山國家公園、丹大野生動物重要棲息環境、大武山自然保留區、宜蘭南澳大濁水北溪與台東內本鹿古道。結果由379條D-loo序列中得到19種D-loop之基因單套型，其中4個基因單套型的核苷酸取代特徵與其他15個單套型明顯不同，因此在演化親緣樹上可分為陶塞雪霸類群與中央山脈主要類群，boostrap value高於94%。陶塞雪霸類群的基因單套型來自雪霸國家公園、太魯閣陶塞溪下游與丹大坂邊天長，而中央山脈主要類群的基因單套型則主要分布於太魯閣以南的各個樣區，但不包含太魯閣陶塞溪下游。此兩大遺傳類群根據粒線體全長序列所計算的分歧時間為37,250∼74,500年，座落於沃姆冰河期。在水鹿亞種的親緣關係樹上，臺灣水鹿形成一獨立遺傳類群，與海南島水鹿分歧時間為125,375∼250,750年前，與印度水鹿分歧時間為293,625∼587,250年前。
.利用SAMOVA、遺傳分化指數FST與網狀支序分析法分析臺灣水鹿族群遺傳結構，整合其結果將臺灣水鹿分為SP、NTS、CLNK、DDCT、NYS、SY、與DWTT七個族群後，發現SP與其他六個族群之間有最高的FST值，次高者出現在NTS與CLNK之間。這些突出的FST值表示陶塞雪霸地區族群與其他水鹿族群間分化程度極高。相反地，DDCT、NYS、SY、與DWTT四個族群之間FST值相對較低，顯示南部族群間基因交流順暢。結合地理資訊的網狀支序分析結果，暗示陶塞雪霸類群與中央山脈主要類群間的分歧可能是長距離拓殖或過去棲地破碎化所造成。然而，由於由於明顯分化的兩大遺傳類群間並沒有發現中間基因單套型，因此推測在距離隔離之外，還存在更嚴苛的因素限制了兩大遺傳類群間的基因交流。
以中性檢測分析、核苷酸錯位分布與貝氏天際線圖進一步分析臺灣水鹿的歷史族群數量變動趨勢，在中央山脈主要類群檢定的Fu’s Fs數值為負值且顯著（P < 0.001），核苷酸錯位分布也呈現單峰，因此支持該類群曾經歷族群擴張，但陶塞雪霸類群的中性檢測分析數值則皆為正值且不顯著，核苷酸錯位分布呈現半單峰，因此不支持。貝氏天際線圖顯示中央山脈主要類群約於10,000年前開始族群數量的成長，且在7,500年至今有明顯擴張現象。陶塞雪霸類群的族群擴增則較晚，約於2,000前族群數量開始增加，明顯的擴張出現於1,000年前至今。此外，陶塞雪霸類群在歷史上的有效族群數量也相對較小。
綜合上述結果，本研究定義臺灣水鹿粒線體序列的遺傳特徵，初步揭示了三種水鹿亞種間的親緣關係，並推測水鹿約在里斯冰河時期由亞洲大陸遷徙至台灣本島，然後在之後的間冰期分化成台灣亞種。台灣水鹿的族群遺傳結構有明顯分化，但主要原因可能不只有長距離的隔離。兩大遺傳類群在歷史族群數量變動上的差異，反映了低緯度地區較早的回暖，是南部水鹿族群領先於陶塞雪霸族群擴增的原因。考慮到冰河期氣候變遷對地形與植被分布的影響，食物與遷徙範圍的限制可能也使得陶塞雪霸類群水鹿族群擴增較慢。高海拔棲地嚴苛的環境不但影響了陶塞雪霸地區水鹿的歷史族群數量變動，也是阻礙了兩大遺傳類群間水鹿族群基因交流的潛在因素。

關鍵字

臺灣水鹿 ； 粒線體 ； 地理親緣

並列摘要

Sambar deer (Rusa unicolor) is the largest Asian deer, and most of its seven subspecies are under extinction due to human overexploitation and hunting pressure, except for the wild population in India, Nepal, and Taiwan. As the largest wild herbivore in Taiwan, which is the indicator sensitive to habitat loss and climate change in history, Formosan sambar deer (Rusa unicolor swinhoii) presently hold the only stable native populations in Southeast Asia high altitude region. The wild Formosan sambar deer populations distributed in one of the highest altitude sambar deer habitats which featuring season migration, however, the patterns of its population genetic structure and phylogenetic relationship with other subspecies had yet been revealed in previous studies. The aims of this study are (1) to investigate the phylogenetic relationship among wild Formosan sambar deer and other subspecies based on diversity of mitochondrial DNA; (2) to assess the genetic structure and phylogeography of Formosan sambar deer populations; (3) to infer the population demographic events-shaped phylogeographical pattern of Formosan sambar deer.
Seven hundred and seventy-nine Formosan sambar deer samples from Shei-pa National Park, Taroko National Park, Danda Wildlife Refuge, Yushan National Park, Da-Wu Shan Natural Reserve, Lupnik Trail, and Big Chosui River Trail were obtained. Nineteen haplotypes were obtained from three hundred and seventy-nine D-loop sequences of Formosan sambar deer. Distinct nucleotide substitution characterizes was crucially found in four haplotypes, and Formosan sambar deer were consequently split into two major clade, Taosai Shei-pa clade and Central Mountain Major clade supported by high bootstrap value (94%) statistically. Taosai Shei-pa clade included individuals from Shei-pa National Park, Taroko Taosai mid-down stream, and Danda Panpien-Tianchang. The Central Mountain Major clade contrarily included individuals from Taroko National Park (all regions except Taosai mid-down stream) and most other southern sampled regions. The divergence time between two major clade was 37,250~74,500 based on complete mitochondrial genome sequence, during Wurm glacial period. In the phylogenetic relationship of sambar subspecies, Formosan sambar deer forms a unique clade in phylogenetic trees, and the divergence time of Formosan sambar deer from Hainan sambar deer and Indian sambar deer were 125,375~250,750 and 293,625~587,250 years ago, respectively.
Population structure of Formosan sambar deer was assessed based on SAMOVA, FST calculation, and nested clade analysis combined with geographic distribution information. Based on the result, Formosan sambar populations were therefore divided in to seven groups: SP, NTS, CLNK, DDCT, NYS, SYS, and DWTT. The highest FST value appeared between SP and other six group, and the FST value between NTS and CLNK came seventh. These outstanding FST value indicated the evident differentiation of population in Taosai and Shei-Pa. In contrast, the relatively low FST value appeared between DCT, NYS, SYS, and DWTT group, revealing the smooth gene flow between these southern populations. The nested clade analysis integrated geography information suggested that long-distance colonization or past fragmentation caused the splitting between Taosai Shei-pa clade and Central Mountain Major clade. However, absent the middle haplotypes linked two clade, the other factors that severely restricted the gene flow between two major clades rather than distance isolation may have been existed.
The neutrality test, mismatch distribution, and Bayesian skyline plot were applied to study demographical history. The significant native value of Fu’s Fs test (P < 0.001) and uni-model mismatch distribution supported population expansion inference in Central Mountain Major clade, yet it did not support population expansion in Taosai Shei-pa clade. The Bayesian skyline plot revealed that the growth of population size of Central Mountain Major clade began at 10,000 years ago, and obviously expanded since 7,500 years before present. Slow expansion of Taosai Shei-pa clade began relatively later at 2,000 years ago, and the evident growth occurred since 1,000 before present. Furthermore, the relatively smaller estimated population size of Taosai Shei-pa clade is also revealed.
In conclusion, the genetic characteristics of Formosan sambar deer were defined, and preliminarily revealed phylogenetic relationship of 3 sambar subspecies. It was inferred that the migration of Formosan sambar deer from continent to Taiwan island occurred in the Riss ice age, and the division between two subspecies had taken place in the following interglacial period. The population structure of Formosan sambar deer showed evident differentiation, which may not completely caused by long distance isolation.

並列關鍵字

Formosan sambar deer ； Mitochondrial DNA ； Phylogeography

參考文獻

Avise, J. C., J. E. Neigel, and J. Arnold. 1984. Demographic influences on mitochondrial DNA lineage survivorship in animal populations. J. Mol. Evol., 20(2): 99-105.

Avise, J. C., J. Amold., R. M. Schneider, E. Bermingham, T. Lamb, J. E. Lamb, and N. C. Saunders. 1987. Intraspecific phylogeography: the mitochondrial DNA bridge between population genetics and systematics. Annu. Rev. Ecol. Syst. 18:489-522.

Avise, J. C., and K. Wollenberg. 1997. Phylogenetics and the origin of species. Proc. Natl. Acad. Sci. U.S.A. 94(15): 7748-7755.

Avise, J. C. 1998. The history and purview of phylogeography: a personal reflection. Molecular Ecology, 7(4), 371-379.

Avise, J. C. 2004. What is the field of biogeography, and where is it going. Taxon. 53(4):893-898.