Molecular phylogeography relationships of freshwater crabs in East Asian islands

東亞島嶼(台灣、琉球和日本)所產的淡水蟹，有2科5屬，種類約有50種。其中澤蟹屬與清溪蟹屬為東亞島嶼淡水蟹的優勢屬，不產於亞洲大陸地區。利用16S rRNA的基因片段，比較東亞產淡水蟹之親緣關係，期能了解東亞優勢淡水蟹的起源。結果顯示，東亞的清溪蟹類群中，台灣的Candidiopotamon rathbunae與琉球的C. okinawense 和C. kumejimense有較近的親緣關係，與琉球的相近屬－琉球蟹屬的親緣關係則較遠。台灣的C. rathbunae，可見以中央山脈為中心，將其分為東、西兩大族群，分離年代為4.8 ± 0.8 mya，此年代接近台灣島露出海面的年代。西半群可再分成西北部、西部、西南部三群；東半群也可再分成南部、東南部、東部三群。東亞澤蟹屬可分為4大群，首先分離出日本本島群，其次分離出中琉球群，再分離出南琉球-北台灣群，最後是台灣澤蟹群。從類群分離的時間先後之親緣關係樹來看，東亞澤蟹屬的祖先，在Miocene之前就在日本本島繁衍，然後逐漸往南遷移。台灣產澤蟹屬的分群，部分分群的可信度雖未達50%，但若參照C. rathbunae的分群，亦可看出以中央山脈為界的兩大群，分離年代為3.2 ± 0.5 mya，此為中央山脈急速隆起的年代。中央山脈以西可分成南琉球-北台灣群、西部群、北部群與西南群；中央山脈以東又可分成南部群與東部群，但每群仍有部分種類延伸至其他地理區。從東亞島嶼淡水蟹的親緣關係樹，可發現澤蟹屬與清溪蟹類群各自成一單系群，兩者分化的年代，約為9.0 ± 1 mya，與亞洲大陸的淡水蟹在一千萬年前即分離，分離的原因可能是，沖繩海槽的陷落，此時台灣島尚未露出海面，因此，這些淡水蟹應該集中於日本本島區域，然後逐漸南下散佈。此結果支持在Miocene時，東亞島嶼與亞洲大陸連接在一起的假說 。Freshwater crabs in the East Asian islands have about fifty species, belonging to five genera. Among them, genera Geothelphusa and Candidiopotamon are dominant, couldn't find in the Asian continent. The phylogenetic relationship among freshwater crabs in the East Asian islands were studied using partial sequence data from mitochondrial large subunit 16S rRNA gene. Phylogenetic analysis based on sequence data of East Asian Candidiopotamon group revealed that C. rathbunae in Taiwan have closest relatives with C. okinawense and C. kumejimense in central Ryukyus. In Taiwan, C. rathbunae can be divided into two geographic groups by the Central Range. The divergence time between eastern and western groups can be estimated about 4.8 ± 0.8 mya, which can be correlated with the emergence time of Taiwan island when the Philippine Sea plate collided with continental Asian tectonic plate. Western groups can be subdivided into three subgroups (northwestern, western and southwestern groups), and Eastern groups also can be subdivided into three subgroups (southern, south eastern and eastern groups). East Asian Geothelphusa can be subdivided into 4 groups. The group of the main-islands of Japan was diverged firstly, the central Ryukyus group was diverged secondly, then the southern Ryukyus — northern Taiwanese group was diverged, the last was Taiwanese Geothelphusa. From the divergence time of every groups indicate that the ancestors of East Asian Geothelphusa have distributed all over main-islands of Japan since Miocene, then dispersed southward. Taiwanese Geothelphusa are similar to the distribution of C. rathbunae, also can be divided into two groups and the Central Range could isolate eastern and western groups. The divergence time is 3.2 ± 0.5 mya and it matches the rapid uplift of the Central Range. Based on the phylogenetic tree of the East Asian freshwater crabs, it is clear that both Geothelphusa and Candidiopotamon groups are monophyletic. The two groups were diverged in 9.0 ± 1 mya and the dating could be 10 mya compared with the Chinese freshwater crabs. Caused by the sinking of Okinawa Trough, the islands were separated from the continent. Meanwhile, Taiwan was still under the sea level. Freshwater crabs should be distributed all over main-islands of Japan, then dispersed southward. The results support the hypothesis that the East Asian islands was linked together with the Asian continent in Miocene.中文摘要………………………………………………………………… Ⅰ 英文摘要………………………………………………………………… Ⅱ 誌謝……………………………………………………………………… Ⅲ 目錄……………………………………………………………………… Ⅳ 表目錄..…………………………………………………………………. Ⅴ 圖目錄..…………………………………………………………………. Ⅵ 頁 壹、前言………………………………………………………………… 1 貳、材料與方法………………………………………………………… 9 一、標本的採集……………………………………………………… 9 二、分子生物學方法………………………………………………… 9 2.1萃取分離genomic DNA ……………………………………… 9 2.2 PCR ( Polymerase chain reaction )與DNA定序……………… 11 三、分子時鐘………………………………………………………… 12 四、數據分析………………………………………………………… 13 參、結果………………………………………………………………… 14 一、東亞島嶼清溪蟹類群之親緣關係……………………………… 14 二、東亞島嶼澤蟹屬之親緣關係…………………………………… 15 三、東亞島嶼淡水蟹之親緣關係…………………………………… 17 肆、討論………………………………………………………………… 19 一、東亞島嶼清溪蟹類群之親緣關係…………………………….. 19 二、東亞島嶼澤蟹屬之親緣關係…………………………………… 20 三、東亞島嶼淡水蟹之親緣關係…………………………………… 21 伍、結論………………………………………………………………… 25 陸、參考文獻…………………………………………………………… 2