非洲白鳳蝶的多態型，左上方框內是雄性，線的上方是各種斑蝶與細蝶，線的下方是各型雌性白鳳蝶 photo from mimeticbutterflies.org

標題：Independent evolution of sexual dimorphism and female-limited mimicry in swallowtail butterflies (Papilio dardanus and Papilio phorcas)


摘要

Several species of swallowtail butterflies (genus Papilio) are Batesian mimics that express multiple mimetic female forms, while the males are monomorphic and nonmimetic. The evolution of such sex-limited mimicry may involve sexual dimorphism arising first and mimicry subsequently. Such a stepwise scenario through a nonmimetic, sexually dimorphic stage has been proposed for two closely related sexually dimorphic species: Papilio phorcas, a nonmimetic species with two female forms, and Papilio dardanus, a female-limited polymorphic mimetic species. Their close relationship indicates that female-limited polymorphism could be a shared derived character of the two species. Here, we present a phylogenomic analysis of the dardanus group using 3964 nuclear loci and whole mitochondrial genomes, showing that they are not sister species and thus that the sexually dimorphic state has arisen independently in the two species. Nonhomology of the female polymorphism in both species is supported by population genetic analysis of engrailed, the presumed mimicry switch locus in P. dardanus. McDonald-Kreitman tests performed on SNPs in engrailed showed the signature of balancing selection in a polymorphic population of P. dardanus, but not in monomorphic populations, nor in the nonmimetic P. phorcas. Hence, the wing polymorphism does not balance polymorphisms in engrailed in P. phorcas. Equally, unlike in P. dardanus, none of the SNPs in P. phorcas engrailed were associated with either female morph. We conclude that sexual dimorphism due to female polymorphism evolved independently in both species from monomorphic, nonmimetic states. While sexual selection may drive male-female dimorphism in nonmimetic species, in mimetic Papilios, natural selection for protection from predators in females is an alternative route to sexual dimorphism.

福翠鳳蝶 (Papilio phocas) 雄性 (上)與雌性 (下)， 另有一型與雄性相似

鳳蝶的雌性多態性與限性擬態的演化起源已經吸引科學家討論好久，但始終還沒找到一個定論。究竟那是個數個物種的共有衍徵？抑或是個別物種的獨有衍徵呢？或許可以從非洲白鳳蝶 (Papilio dardanus) 的例子中看出端倪。

福翠鳳蝶 (Papilio phocas)與非洲白鳳蝶同屬白鳳蝶種群 (dardanus species-group)，有些觀點認為這兩種可能是最近緣的種類。福翠鳳蝶同樣是只有雌性多態型，但不同的是其不參與擬態。也就是說，如果系統發生學的分析支持這兩個物種是姊妹群，那麼就支持雌性多態性的單一起源。

這個研究以基因體層級大規模的分析，以3964個基因與全粒腺體序列推測這兩者間的親緣關係。結果顯示兩者並未如預期的是最近的姊妹群。另外，此團隊也對控制擬態斑紋的engrailed基因的單核苷酸多態性 (SNP, Single Nucleotide Polymorphism)做分析，在白鳳蝶的多態型族群中偵測到平衡選擇 (balancing selection)的特徵，而在單態型與福翠鳳蝶的族群中都沒有。在非擬態的福翠鳳蝶的SNP分析中，engrailed基因也與雌性多態無關。根據以上資料可以推論，這兩個物種的雌性多態性是起源於單態且非擬態的狀態獨立演化而來，進而形成性雙型性。非擬態的物種的性雙型性可能是由性擇推動，而擬態物種的性雙型性來自天擇作用於雌性，讓這兩個看似相似的現象有著截然不同的機制。

圖片來自本文

原文標題：Identification of doublesex alleles associated with the female-limited Batesian mimicry polymorphism in Papilio memnon (open access)


摘要

The female-limited Batesian mimicry polymorphism in Papilio butterflies is an intriguing system for investigating the mechanism of maintenance of genetic polymorphisms. In Papilio polytes, an autosomal region encompassing the sex-determinant gene doublesex controls female-limited mimicry polymorphism. In the closely related species P. memnon, which also exhibits female-limited Batesian mimicry polymorphism, we identified two allelic sequences of the doublesex gene that corresponded exactly with the mimetic and non-mimetic female phenotypes. Thus, the genetic basis of the mimicry polymorphism in P. memnon is similar to that in P. polytes. However, the mimetic and non-mimetic alleles of the two species were not identical, and the divergence of alleles occurred independently in P. memnon and P. polytes. Different mutation-selection processes may have resulted in the convergent patterns of mimicry polymorphism in these Papilio butterflies.

自從控制非洲的白鳳蝶 (Papilio dardanus) 的雌性限性擬態/多態性翅紋的基因座發現後，來到2014年，控制玉帶鳳蝶雌性限性擬態/多態性的超基因 doublesex 的序列與機制也被發掘出來 (可參考這篇文章與這個新聞) 。那麼有著類似斑紋變化，同樣是經典的擬態案例的大鳳蝶呢？

同樣是京都大學的曾田貞滋教授團隊與師大生科系的林仲平教授合作的研究，在這篇文章中，他們發現控制大鳳蝶的超基因與玉帶鳳蝶相似，都位於doublesex這個基因座上，也有表現擬態的顯性 (H) 與隱性 (h)。然而，雖然大鳳蝶與玉帶鳳蝶是同一個亞屬，但兩種目的基因的序列並不相同，而且系統發生學的分析顯示兩者間的基因來源是獨立演化而來。

系統發生學分析顯示玉帶鳳蝶與大鳳蝶的親緣關係

以系統發生學的方式分析doublesex基因序列的關係，顯示大鳳蝶與玉帶鳳蝶間的基因來源是獨立演化而來

控制表型的基因在種間是獨立演化而來其實是滿普遍的現象，但為何這個研究要特地分析兩種的序列以證明其獨立演化來源呢？因為以毒蝶來說，控制斑紋的超基因是會藉由雜交而在不同族群間交換，如果把以系統發生學的方式分析毒蝶的超基因，會發現每個支系混雜不同的族群，而在這篇文章中可以發現兩者的分化顯著，顯示其推動基因演化的機制與毒蝶有所差異。

圖片來源：https://reefbuilders.com/2014/11/14/survival-reef-part-1-masquerading-false-colours/#

標題：Temporal dynamics of the mimetic allele frequency at the doublesex locus, which controls polymorphic Batesian mimicry in Papilio memnon butterflies (open access)

摘要

Tracking allele frequencies is essential for understanding how polymorphisms of adaptive traits are maintained. In Papilio memnon butterflies, which exhibit a female-limited Batesian mimicry polymorphism (wing-pattern polymorphism), two alleles at the doublesex (dsx) locus correspond to mimetic and non-mimetic forms in females; males carry both dsx alleles but display only the non-mimetic form. This polymorphism is thought to be maintained by a negative frequency-dependent selection. By tracking dsx allele frequencies in both sexes at a Taiwanese site over four years, we found that the mimetic allele persists at intermediate frequencies even when the unpalatable model papilionid butterflies (Pachliopta and Atrophaneura species) were very rare or absent. The rates of male mate choice did not differ between the two female forms; neither did insemination number nor age composition, suggesting equivalent reproductive performance of the two forms over time. Our results characterised the temporal dynamics of the mimetic allele frequency in the field for the first time and give insights into underlying processes involved in the persistence of the female-limited Batesian mimicry polymorphism.

大鳳蝶是貝氏擬態的教科書經典案例，在亞洲對這個自家後院就看到的物種自不用說，而西方透過自華萊士時代的紀錄與研究，幾乎全世界都對這個例子的多態性、雌雄二態性與擬態間的關係與來源十分感興趣。這篇文章是京都大學的曾田貞滋教授 (Teiji Sota)及他的學生，與師大生科的林仲平教授合作，以台灣的大鳳蝶為材料，進行長達四年 (2013-2016) 的野外觀察與基因體研究的一部份成果。

這個研究的關鍵在於：首先，貝氏擬態的前提是以負向頻率(negative frequency-dependent selection)為選汰機制，意思也就是在已知的環境中，擬態者的數量不能過多，否則天敵遇到大部份的個體都能吃的話，這個擬態斑紋就可能失效。然而像斑紋高度多態性又牽涉雌雄二態的大鳳蝶，控制擬態的超基因 (supergene) 的族群基因頻率在環境中，如何在擬態與非擬態型中取得平衡就是個很有趣的問題了。



這個團隊執行了幾個部份的調查與實驗。(1) 他們在台灣花蓮以穿越線調查蝴蝶的狀況，並且分析其年齡、身體尺寸、天敵攻擊的咬痕，與交配的次數；(2) 分析大鳳蝶與其擬態模型紅紋鳳蝶 (Pachliopta aristolochiae)、大紅紋鳳蝶 (Atrophaneura polyeuctes) 與台灣麝香鳳蝶 (Atrophaneura febanus) 在這四年中的族群數量 (2) 設計行為的實驗測試大鳳蝶雄性對不同表型的雌性是否具有交配偏好；(3) 野外採集的個體帶回至實驗室進行分生的分析，將其基因型統計是否符合哈溫定律。主要的結果支持兩性的基因頻度的調查皆符合哈溫定律，而雄性無論對擬態或非擬態型的雌性都沒有特殊的擇偶偏好。在四年的觀察與採集中，doublesex的基因頻度維持在中等程度，並未因擬態模型的數量變動偵測到顯著變化，也未發現明顯的負向頻度選汰。也就是說，擬態模型在環境中的頻度與雄性無明顯擇偶偏好，都對doublesex的基因頻度沒有顯著的影響, 而這可能是大鳳蝶能夠維持斑紋多態性的原因。

===
以下是對這個研究的一些觀點：

1. 這個研究的個體僅從花蓮取得，不知是否足以推及解釋台灣其他區域的族群。
2.  一般來說，大鳳蝶的擬態對象認為是紅紋鳳蝶 (Pachliopta aristolochiae)，而這個研究則認為不曾被紀錄是擬態對象的大紅紋鳳蝶 (Atrophaneura polyeuctes) 與台灣麝香鳳蝶 (Atrophaneura febanus) 也同時是其擬態模型。然而，紅紋鳳蝶與另兩種蝴蝶的棲地類型有所差異，根據這個研究的資料，在花蓮的研究地點紀錄的紅紋鳳蝶個體僅有1隻，遠低於大紅紋的21隻，有可能其觀測與採集地點並非天擇作用於擬態斑紋的主要活躍區域，因此擬態者的擬態模型與補食者的動物相的差異，可能影響基因頻率的觀察結果，而非單純的「維持在中等程度進而維持族群中翅紋的多態性」。若是未來能加入更廣泛區域的資料，能讓相關研究更加完整。
3. 雖然大鳳蝶的擬態關係常被用來解釋擬態現象，但畢竟是以人類的視覺角度來觀察，而且大鳳蝶翅紋多態性的變化程度遠比想象中的高，如果一點點差異就會造成擬態效果的變化，那麼所謂「擬態型」的個體與其基因頻度或許要再評估。

<書籍資訊>
だましのテクニックの進化 ―昆虫の擬態の不思議
単行本（ソフトカバー）: 212ページ 
出版社: オーム社 (2015/10/26) 
言語: 日本語 
ISBN-10: 4274505820 
ISBN-13: 978-4274505829 
発売日： 2015/10/26

<作者> 藤原 晴彥
東京大學生命科學研究系教授，以昆蟲為研究對象，其研究領域為反轉錄子的轉移、端粒酶的應用，昆蟲翅發生激素的建構機制，以及昆蟲體表擬態斑紋的形成的分子機制。

<內容簡介>
從擬態的歷史與昆蟲間的關係談起，昆蟲所呈現的擬態圖案，像是枯葉蝶、竹節蟲以及玉帶鳳蝶，綜合近年來的研究，以分子生物學的角度來討論斑紋是如何產生，現實上分子機制又是如何控制斑紋的表現，在演化上又扮演哪些重要的角色。書中也延伸說到瓢蟲的假死的神經控制，以及農藥對於這些神經系統的影響，在農業生態系中與害蟲間的關係。

photo from Papilio dardanus (wiki)

標題：Characterising the phenotypic diversity of Papilio dardanus wing patterns using an extensive museum collection.

摘要

The history of 20th Century evolutionary biology can be followed through the study of mimetic butterflies. From the initial findings of discontinuous polymorphism through the debates regarding the evolution of mimicry and the step-size of evolutionary change, to the studies on supergene evolution and molecular characterisation of butterfly genomes, mimetic butterflies have been at the heart of evolutionary thought for over 100 years. During this time, few species have received as much attention and in-depth study as Papilio dardanus. To assist all aspects of mimicry research, we present a complete data-derived overview of the extent of polymorphism within this species. Using historical samples permanently held by the NHM London, we document the extent of phenotypic variation and characterise the diversity present in each of the subspecies and how it varies across Africa. We also demonstrate an association between ‘‘imperfect’’ mimetic forms and the transitional race formed in the area where Eastern and Western African populations meet around Lake Victoria. We present a novel portal for access to this collection, www.mimeticbutterflies.org, allowing remote access to this unique repository. It is hoped that this online resource can act as a nucleus for the sharing and dissemination of other collections databases and imagery connected with mimetic butterflies.

非洲白鳳蝶的多態型，左上方框內是雄性，線的上方是各種斑蝶與細蝶，線的下方是各型雌性白鳳蝶 photo from mimeticbutterflies.org

非洲的白鳳蝶(Papilio dardanus)是知名的貝氏擬態的鳳蝶，其豐富的多態型各自擬態非洲各地的細蝶、斑蝶等不好吃的種類。白鳳蝶的擬態實際上只有雌性參與其中，而雄性只有一種型態，在多態型的雌性中，除了擬態各種不同的蝴蝶外，神奇的也存在雄性型的雌性。

1960年代，透過孟德爾式的遺傳學雜交試驗瞭解不同族群間的斑紋變化，而且結果顯示白鳳蝶的斑紋變化是來自一個超基因；到了2008年，基於現代的分生技術，終於找到超基因的位置。即使如此，雌性的型態實在太多了，除了上面這些看起來精準擬態的斑紋外，也有更多看起來是中間型的種類，因此雖然已經研究到基因體層級的情況下，基礎的斑紋與地理分佈還沒有整體性的調查過。

雌性各種不同的型與名稱 photo from Fig. 1

這篇透過英國倫敦自然史博物館(the Natural History Museum, London)的極大量鱗翅目館藏，蒐集標籤上的採集資訊，將採集的地點轉換成現代的地點並查出座標，把各個不同的斑紋分類後，再轉換為二維座標系統輸入地理資訊系統 (GIS)，把各個不同的斑紋分類後，就能每個型態的分佈呈現在現代的地圖上，並且透過採集點的連線，推估分佈範圍。

取樣密度與地點 photo from Fig. 3 各型的分佈範圍 photo from Fig. 6

結果就如上圖，把大部分的斑紋的分佈範圍推估出來，而那些看起來不完美擬態的斑紋分佈於維多利亞湖附近，支持這是西部與東部族群相遇的雜交區。

中間型的分佈區域 photo from Fig. 7

這分析的概念很簡單，但卻是瞭解一個擬態群的多樣性最快的方式，而且最重要的，這仰賴保存完整與大量的標本館藏。這樣的研究說明透過良好的想法與完整的分析，即使基礎的研究也能從中得到大量的資訊，也更凸顯標本蒐藏的重要性。

photo from K. Kunte

標題：A genetic mechanism for female-limited Batesian mimicry in Papilio butterfly.

摘要

In Batesian mimicry, animals avoid predation by resembling distasteful models. In the swallowtail butterfly Papilio polytes, only mimetic-form females resemble the unpalatable butterfly Pachliopta aristolochiae. A recent report showed that a single gene, doublesex (dsx), controls this mimicry; however, the detailed molecular mechanisms remain unclear. Here we determined two whole-genome sequences of P. polytes and a related species, Papilio xuthus, identifying a single ~130-kb autosomal inversion, including dsx, between mimetic (H-type) and non-mimetic (h-type) chromosomes in P. polytes. This inversion is associated with the mimicry-related locus H, as identified by linkage mapping. Knockdown experiments demonstrated that female-specific dsx isoforms expressed from the inverted H allele (dsx(H)) induce mimetic coloration patterns and simultaneously repress non-mimetic patterns. In contrast, dsx(h) does not alter mimetic patterns. We propose that dsx(H) switches the coloration of predetermined wing patterns and that female-limited polymorphism is tightly maintained by chromosomal inversion.

photo from Fig 3

基因knockdown後，不同處理的斑紋表現 photo from Fig. 5

左邊那隻是紅紋鳳蝶(Pachilopta aristolochiae)，右邊那隻是黃星鳳蝶(Papilio (Chilasa) epycides)

這看起來只是兩種不同蝴蝶雜交，但在擬態生物學裡這個異常的地方在於，紅紋鳳蝶是一種泛亞洲分佈，很多擬態者的擬態模型，而黃星鳳蝶則被認為是一種擬態斑蝶的擬態者

標題：Multicomponent deceptive signals reduce the speed at which predators learn that prey are profitable

摘要

Many prey use multicomponent deceptive signals to fool predators into mistaking them for inedible objects, toxic prey, or danger- ous animals. However, recent experiments have suggested that multicomponent deceptive signals are no more effective in deter- ring predators than single-component signals, making it difficult to understand how they have evolved. Here, we use an established experimental system in which naive domestic chicks are presented with models of snake-mimicking caterpillars to test the idea that multicomponent deceptive signals reduce the speed at which predators learn that prey are profitable. We presented chicks with a series of 4 trials in which they encountered a single type of caterpillar model. The type of model differed among our 4 experimental groups that were arranged in a 2 × 2 factorial design: models either possessed eyespots or did not and were in either the resting or defensive posture. Chicks’ responses to the same model prey were then retested following an extended 72-h retention period. Chicks rapidly attacked prey with no defensive traits and initially showed similar levels of wariness to prey with either 1 or 2 deceptive traits. However, chicks learned that single-trait caterpillars were profitable more quickly than 2-trait caterpillars and retained their learned responses better. This suggests that prey with multicomponent deceptive signals may have a selective advantage over prey with single-component deceptive signals when predators repeatedly encounter such prey.

這個研究的概念跟討論Battus philenor的是不是每個部分的顏色對於警戒都有貢獻的概念有點類似，就是說通常有警戒斑紋的物種，也會有其他的第二防禦機制，像是某種行為配上顏色，這樣可能對於嚇阻捕食者更有效率。但就演化上來說，如果警戒色或行為已經夠有效率的話，又為何需要花更多的時間與資源來發展第二防禦？這真的會比較有效嗎？

文中透過行為學的實驗來討論這個問題，捕食者使用的是出生七天內的小家雞，獵物則是用麵粉配上食用色素，做成像Papilio canadiensis或是P. glaucus的幼蟲樣，這又分成兩個行為的型態：休息姿勢與防禦姿勢，配上有眼睛與沒眼睛，就是一個2x2因子的實驗設計。結果顯示沒有任何防禦機制（沒眼睛+休息姿勢）的最快被攻擊，只有眼睛或只有防禦姿勢在一開始的測試中與沒有防禦機制相似，之後的攻擊延遲上升，而有兩種防禦機制（有眼睛+防禦姿勢）攻擊延遲最長。結果顯示有兩種防禦機制能有效的讓捕食者遲疑。

～個人觀感～
這類實驗通常能解釋某些行為學上的觀點，可是他們覺得這個樣子的幼蟲之所以有防禦的樣子，不是單純的因為眼斑，而是因為他們認為這是有關蛇的擬態（snake mimicry）（嗯........不像吧，難道加拿大的蛇長這樣嗎？）


另一個點是這個研究團隊通常用剛出生的小雞做實驗，為的是剛出生時沒有任何經驗，很善良天真，餵什麼吃什麼，而且很可愛（誤），可以避免某些因為經驗帶來的實驗誤差，也很容易看的到學習曲線。但相對來說，因為這個年齡算是未成年，也就是思想不成熟，腦類發育可能未完全，這樣考量的話，使用這個年紀的雞來做實驗得到的結果，是不是真能夠反映自然界的狀況呢？

至於他們用黏土做的手工獵物可以看這裡，不得不說這看起來根本是另一個什麼（雖然捏的是挺可愛），然後這些笨雞還真的學到了什麼（到底學到什麼啊？）

標題：Warning Signal Efficacy: Assessing the Effects of Color, Iridescence, and Time of Day in the Field


摘要

Warning coloration deters predators from attacking distasteful or toxic prey. Signal features that influence warning color effectiveness are not well understood, and in particular, we know very little about how effective short-wavelength and iridescent colors are as warning color elements in nature and how warning signal effectiveness changes throughout the day. We tested the effect of these factors on predation risk in nature using specimens of the distasteful pipevine swallowtail butterfly, Battus philenor. B. philenor adults display both iridescent blue and diffusely reflecting orange components in their warning signal. We painted B. philenor wings to create five different model types: all-black, only-iridescent-blue, only- orange, iridescent-blue-and-orange (intact signal), and matte-blue-and- orange. We placed 25 models in each of 14 replicate field sites for 72 h and checked for attacks three times each day. Model type affected the likelihood of attack; only-orange models were, the only model attacked significantly less than the all-black model. Iridescence did not enhance or decrease warning signal effectiveness in our experiment because matte- blue-and-orange models were attacked at the same rate as iridescent- blue-and-orange models. Time of day did not differentially affect model type. Video recordings of attacks revealed that insectivorous birds were responsible. The results of this experiment, when taken with previous work, indicate that the response to blue warning coloration is likely dependent on predator experience and context, but that iridescence per se does not affect warning signals in a natural context.

在讀行為學、生態學或是普通生物學的時候，念到擬態的部分會搭配一個很典型的例子，然後就會自然而然很少懷疑這些百年前的科學前所指定的擬態群，牠們的警戒訊號到底有沒有發揮作用。或是有時候會認知一個物種身上所有的斑紋，都對警戒訊號有貢獻，特別是那些有金屬色的物種們。

這個研究就在嘗試解決這個疑慮。Pipevine swallowtail (Battus philenor) 是一種廣泛分佈在北美洲的鳳蝶，幼生期的寄主植物就如俗名，是取食馬兜鈴類的植物，這種植物含有高量植物鹼，幼蟲取食後會儲存在體內，成體時會透過某種機制，像是被攻擊的時候ˇ噴放臭氣來抵抗捕食者，因此這個鳳蝶被認為是北美的線蛺蝶(Limenitis arthemis)與 大黃鳳蝶 (Papilio glaucus)雌性的擬態模型。這個鳳蝶的腹面有幾個明顯的大橘斑，還有閃亮的金屬藍色，而金屬色對於人類來說十分閃亮，甚至有可能反射人類看不到的偏極光或紫外光，而且金屬色在許多涉入擬態的物種都存在，只是之前的研究預設這樣顯眼的顏色必然對於警戒色有貢獻，那麼究竟有沒有在人類的感官之外的故事呢？

作者嘗試的用顏料操作斑紋，把斑紋塗成四種樣式：全黑、只有金屬藍、只有橘斑、無金屬的藍色+橘斑，加上一個對照組，然後把處理過的樣本放到野外，看看哪個部分對於警戒色有貢獻，以及記錄一天內不同的時間對於這些模型的攻擊率的差異。結果顯示會來攻擊的是食蟲性的鳥類，紅橘斑毫無疑問的是一個很重要的訊號，但金屬藍在處理間卻沒有任何的差異，也就是這個金屬色並未影響警戒訊號的有效性。一天內不同的時間對於被攻擊率沒有差異。

雖然沒有預想中的功用，但動物的體色也可能在性擇求偶這方面有所用處。也有很多其他區域的擬態群，牠們的金屬色也還沒被檢測過，雖然有的看起來就像這個例子，與擬態明顯的相關，這讓我們對於警戒色與擬態的功用有更多探索的空間，也提醒我們很多早期的自然學家體認的擬態群，他們想像中的故事可能跟自然界實際發生的劇情可能有很大的落差。