周 贊, 蔡夢(mèng)思, 劉 虞, 孫燕燕,羅來高

1 杭州師范大學(xué)生命與環(huán)境科學(xué)學(xué)院, 杭州 310036 2 東營(yíng)市林業(yè)局, 東營(yíng) 257091 3 滁州學(xué)院生物與食品工程學(xué)院,滁州 239000

紅耳滑龜和中華草龜幼體體溫與運(yùn)動(dòng)的熱依賴性、補(bǔ)償生長(zhǎng)及免疫力

周 贊1, 蔡夢(mèng)思1, 劉 虞1, 孫燕燕2,*,羅來高3

1 杭州師范大學(xué)生命與環(huán)境科學(xué)學(xué)院, 杭州 310036 2 東營(yíng)市林業(yè)局, 東營(yíng) 257091 3 滁州學(xué)院生物與食品工程學(xué)院,滁州 239000

研究了紅耳滑龜(Trachemysscriptaelegans)和中華草龜(Chinemysreevesii)幼體的體溫與運(yùn)動(dòng)表現(xiàn)的熱依賴性、補(bǔ)償生長(zhǎng)能力及免疫能力。結(jié)果顯示：紅耳滑龜能顯著影響草龜?shù)捏w溫調(diào)節(jié)行為,具有較寬的體溫調(diào)節(jié)范圍和高低溫耐受幅,并具有較強(qiáng)的運(yùn)動(dòng)能力和天然免疫能力,表現(xiàn)出較為穩(wěn)定的補(bǔ)償生長(zhǎng)機(jī)制。在調(diào)溫環(huán)境中,與紅耳滑龜混合飼養(yǎng)的中華草龜平均體溫較分開飼養(yǎng)時(shí)明顯降低,且兩種飼養(yǎng)條件下的紅耳滑龜體溫調(diào)節(jié)的變異系數(shù)要明顯大于中華草龜。紅耳滑龜?shù)呐R界致死高低溫分別為41.2 ℃和3.6 ℃,分別高于和低于中華草龜?shù)呐R界致死高低溫(37.2 ℃和4.5 ℃)。分別測(cè)量?jī)煞N龜在體溫為20、28、36 ℃下的運(yùn)動(dòng)表現(xiàn),結(jié)果顯示兩種龜?shù)募灿嗡俸统掷m(xù)最大運(yùn)動(dòng)距離隨體溫的升高而增加；相同溫度下紅耳滑龜?shù)倪\(yùn)動(dòng)表現(xiàn)優(yōu)于中華草龜。兩種龜饑餓后均存在補(bǔ)償生長(zhǎng)現(xiàn)象,饑餓5d的兩種龜體重變化率不存在差異,饑餓10d的紅耳滑龜體重變化率顯著大于中華草龜,饑餓15d的中華草龜體重變化率顯著大于紅耳滑龜。3種饑餓處理下體重的變異系數(shù)中華草龜大于紅耳滑龜。兩種龜對(duì)植物凝集素均表現(xiàn)出明顯的免疫反應(yīng)性,紅耳滑龜對(duì)植物凝集素作出的免疫反應(yīng)較快,在6h就到達(dá)最大反應(yīng)。上述結(jié)果綜合表明,紅耳滑龜作為入侵種能快速適應(yīng)環(huán)境并威脅土著龜類,與其運(yùn)動(dòng)和生理機(jī)制上的潛在優(yōu)勢(shì)密切相關(guān)。

紅耳滑龜；中華草龜；入侵；體溫和運(yùn)動(dòng)；補(bǔ)償生長(zhǎng)；免疫能力

外來生物入侵已成為威脅人類生態(tài)環(huán)境安全的重大社會(huì)問題,是當(dāng)今世界三大環(huán)境難題之一,如何防治外來生物入侵,已成為政府和民眾關(guān)注的焦點(diǎn),也是學(xué)術(shù)界著力探討的重要課題之一。我國(guó)深受外來生物入侵之害,每年造成的經(jīng)濟(jì)損失不可估量。據(jù)統(tǒng)計(jì),我國(guó)的外來入侵生物已遍及34個(gè)省級(jí)行政區(qū),在700多種外來入侵種中,至少有352種入侵植物、267種入侵動(dòng)物和134種入侵微生物[1]。入侵形勢(shì)十分嚴(yán)峻,解決入侵問題刻不容緩。

在眾多的入侵種中,以紅耳滑龜(Trachemysscriptaelegans; 又名巴西龜、紅耳彩龜、翠龜)造成的危害較為嚴(yán)重。作為世界最危險(xiǎn)的100個(gè)入侵種之一,紅耳滑龜對(duì)入侵地生態(tài)系統(tǒng)造成不可逆的影響,歐洲在1997年立法禁止進(jìn)口紅耳滑龜,試圖從源頭上控制該種龜?shù)娜肭治：Γ蛔鳛樵a(chǎn)地的美國(guó)更是深受其害,迫使政府早在1975年就立法禁止紅耳滑龜交易[2- 3]。在中國(guó)臺(tái)灣的基隆河,紅耳滑龜作為優(yōu)勢(shì)種大量繁衍,大肆捕食河道內(nèi)的小型魚、貝及蛙類的卵和蝌蚪,嚴(yán)重破壞流域內(nèi)的生態(tài)平衡。調(diào)查發(fā)現(xiàn),紅耳滑龜占據(jù)該流域內(nèi)30%—40%的食物和空間資源,土著龜種幾乎消失[4]。紅耳滑龜?shù)娜肭诌€帶來了大量的致病菌,85%室內(nèi)培育個(gè)體攜帶副傷寒類沙門氏桿菌,并通過龜糞在水中傳播擴(kuò)散,感染水域附近的鳥獸,美國(guó)每年大約100—300萬副傷寒類沙門氏桿菌感染病例中的14%是由紅耳滑龜所引起的[5- 6]。紅耳滑龜還能與土著龜種雜交,對(duì)入侵地龜類遺傳結(jié)構(gòu)造成不可恢復(fù)的污染[7]。

已有研究顯示,紅耳滑龜表現(xiàn)如此巨大的危害與其環(huán)境適應(yīng)能力強(qiáng)有關(guān)。在歐洲地區(qū),該種龜具有較強(qiáng)的熱資源爭(zhēng)奪能力,能不斷侵占土著種-地中海擬水龜(Mauremysleprosa)的熱資源,提高自身曬背調(diào)溫機(jī)會(huì)[8]。在相同條件下,紅耳滑龜?shù)姆砟芰褪澄锵芰Ρ鹊刂泻M水龜強(qiáng)[9- 10]。加之圓形的體型構(gòu)造使其具有較高的熱惰性,能防止熱量過快散失[11]。因此紅耳滑龜自引入歐洲后,就得以快速適應(yīng)、繁衍。紅耳滑龜在污染較為嚴(yán)重的環(huán)境中表現(xiàn)為異常的適應(yīng)力,能通過體內(nèi)產(chǎn)生較多的SOD來清除環(huán)境中活性氧自由基,使機(jī)體免受活性氧傷害,土著龜-中華條頸龜(Mauremyssinensis)則因承受不了相應(yīng)的污染程度而被淘汰[12]。此外,紅耳滑龜具有較強(qiáng)的溫度耐受能力[13],這也是其擴(kuò)散如此廣泛的原因之一。

由此可見,與土著龜類相比,紅耳滑龜在入侵過程中對(duì)入侵地環(huán)境存在適應(yīng)優(yōu)勢(shì),其生理和行為指標(biāo)均表現(xiàn)出一定程度的優(yōu)越性,但該種龜較土著龜?shù)降妆憩F(xiàn)出多大程度的競(jìng)爭(zhēng)優(yōu)勢(shì)仍缺少足夠的數(shù)據(jù)。本研究以入侵種紅耳滑龜和國(guó)內(nèi)土著種-中華草龜(Chinemysreevesii)為對(duì)象,比較兩種龜在體溫調(diào)節(jié)和運(yùn)動(dòng)表現(xiàn)的熱依賴性、饑餓后補(bǔ)償生長(zhǎng)以及對(duì)外源蛋白免疫能力方面的差異,探討入侵龜對(duì)土著龜造成威脅的內(nèi)在機(jī)制。

環(huán)境溫度影響外溫動(dòng)物的體溫,進(jìn)而影響其生理反應(yīng)和行為表現(xiàn)[14-15]。動(dòng)物長(zhǎng)期暴露在極端溫度下可能導(dǎo)致死亡[16],上、下極限臨界溫度[臨界最高溫度(CTMax)和臨界溫度最低(CTMin)]被定義為動(dòng)物不能翻身或無正常反應(yīng)時(shí)的最高和最低耐受溫度[17-18]。盡管外溫動(dòng)物可在廣泛的溫度下存活,但它們的生理反應(yīng)和行為表現(xiàn)往往在適度較高的體溫下更活躍,因此許多外溫動(dòng)物在條件允許的情況下會(huì)試圖保持相對(duì)較高和穩(wěn)定的體溫[14,15,19-20]。

補(bǔ)償生長(zhǎng)是動(dòng)物遭受饑餓后表現(xiàn)出的一種有趣的生理反應(yīng)。這種反應(yīng)不僅具有生理生態(tài)學(xué)意義,而且對(duì)農(nóng)業(yè)生產(chǎn)和資源管理具有潛在的重要性。如果饑餓動(dòng)物在得到充足食物后表現(xiàn)出快速生長(zhǎng)沖刺并且最終趕上或超過他們一直在足夠食物條件下的體重,這種補(bǔ)償生長(zhǎng)叫做完全補(bǔ)償生長(zhǎng)或過量補(bǔ)償生長(zhǎng)[21-22]；如果饑餓動(dòng)物雖然表現(xiàn)出高生長(zhǎng)速率但最終沒有趕上穩(wěn)定足量飼養(yǎng)個(gè)體的體重,那么這種補(bǔ)償生長(zhǎng)叫做部分補(bǔ)償生長(zhǎng)[23]. 雖然已有多種動(dòng)物的補(bǔ)償生長(zhǎng)被研究,例如馴養(yǎng)動(dòng)物,魚類,軟體動(dòng)物,甲殼動(dòng)物等[22,24- 26],但對(duì)龜類補(bǔ)償生長(zhǎng)的研究很少,對(duì)中華鱉(Pelodiscussinensis) 幼體補(bǔ)償生長(zhǎng)反應(yīng)研究,表明中華鱉幼體在饑餓脅迫下首先利用脂肪作為主要能源以維持生存,只有完全的食物剝奪才可以誘發(fā)其部分補(bǔ)償生長(zhǎng)反應(yīng),而部分食物剝奪則不能誘發(fā)此反應(yīng)[27]。

1 材料和方法

1.1 實(shí)驗(yàn)動(dòng)物

實(shí)驗(yàn)用3月齡及6月齡紅耳滑龜和中華草龜幼體購(gòu)自杭州吳山花鳥市場(chǎng),其中3月齡龜用于測(cè)定體溫調(diào)節(jié)、運(yùn)動(dòng)表現(xiàn)及溫度耐受能力,6月齡龜用于測(cè)定補(bǔ)償生長(zhǎng)能力和免疫能力。所有動(dòng)物帶回實(shí)驗(yàn)室后,測(cè)定其形態(tài)指標(biāo),平均分至塑料整理箱中至少適應(yīng)1周。隨后進(jìn)行常規(guī)飼養(yǎng),提供足量食物,并在水體中添加鈣粉和抗生素,補(bǔ)充因?qū)嶒?yàn)期間無法正常曬背而導(dǎo)致的鈣質(zhì)缺失以及減少致病菌的傷害。

3月齡紅耳滑龜平均體重為(7.6±0.4) g(1SD)[N=20,范圍6.8—8.5 g；平均背甲長(zhǎng)×平均背甲寬為(32.3±0.8) ×(31.6±0.7) mm(1SD),背甲長(zhǎng)范圍：30.4—33.7 mm,背甲寬：30.1—32.7 mm]。中華草龜平均體重為(5.4±0.7)(1SD)[N=20,范圍為3.9—7.1 g；平均背甲長(zhǎng)×平均背甲寬為(28.5±1.2) ×(22.9±1.4) mm(1SD),背甲長(zhǎng)范圍：26.4—31.4 mm,背甲寬：19.8—25.5 mm]。

6月齡紅耳滑龜平均體重為(25.8±3.6) g(1SD)[N=33,范圍19.1—31.2 g；平均背甲長(zhǎng)×平均背甲寬為(52.0±2.3) ×(48.0± 1.6) mm(1SD),背甲長(zhǎng)范圍：47.3—56.8 mm,背甲寬：44.8—51.0 mm],中華草龜平均體重為(30.2±3.0) g(1SD)[N=33,范圍24.2—35.5 g；均背甲長(zhǎng)×平均背甲寬為(55.4±2.1) ×(42.9±1.8) mm(1SD),背甲長(zhǎng)范圍：51.0—49.7 mm,背甲寬：40.1—47.0 mm]。

1.2 體溫測(cè)定

體溫測(cè)定前,先將兩種龜分開飼養(yǎng)在長(zhǎng)×寬×高為47 cm×37 cm ×27 cm的整理箱中,用魚缸石鋪滿箱底的3/4,在其上規(guī)則的放置石棉瓦,為小龜提供掩所；將整理箱移入恒溫室內(nèi),調(diào)節(jié)室內(nèi)最低溫度為16oC,并在整理箱一側(cè)正上方懸掛一只150 W陶瓷加熱器,在箱內(nèi)形成16—60℃的溫度梯度。實(shí)驗(yàn)開始時(shí),將幼龜從箱子的低溫端釋放,每個(gè)箱子放置20只龜,提供足量的食物并及時(shí)換水。幼龜適應(yīng)2—3d后,用點(diǎn)溫計(jì)(精度為0.1℃)于每天16: 00測(cè)定動(dòng)物的泄殖腔溫度,每只龜至少測(cè)定2d。兩種龜分開飼養(yǎng)測(cè)定體溫結(jié)束后,將其以1∶1隨機(jī)混養(yǎng)在兩個(gè)相同的整理箱中,在相同的溫度環(huán)境中適應(yīng)2d后,用上述相同方式測(cè)定混養(yǎng)后兩種龜?shù)捏w溫。

1.3 運(yùn)動(dòng)表現(xiàn)測(cè)定

根據(jù)體溫測(cè)定的結(jié)果,設(shè)置20、28和36℃ 3個(gè)實(shí)驗(yàn)處理溫度檢測(cè)兩種龜?shù)倪\(yùn)動(dòng)表現(xiàn)。實(shí)驗(yàn)開始前1 h,將動(dòng)物置于預(yù)設(shè)溫度的恒溫箱內(nèi)適應(yīng),以確保動(dòng)物保持恒定的目標(biāo)體溫；實(shí)驗(yàn)開始前10 min,在一長(zhǎng)×寬為150 cm ×5 cm的帶刻度泳道中注入相同溫度的水,保持水深10 cm,并用點(diǎn)溫計(jì)實(shí)時(shí)監(jiān)控水溫,使水溫上下變動(dòng)不超過0.5℃。實(shí)驗(yàn)開始時(shí),將龜逐個(gè)從恒溫箱內(nèi)移至泳道中,一人用毛筆強(qiáng)烈驅(qū)趕動(dòng)物(筆頭頻繁碰觸龜?shù)娜惯?但不推動(dòng)),另一人用Canon Legria HF20攝像機(jī)拍錄動(dòng)物在泳道內(nèi)的整個(gè)運(yùn)動(dòng)過程。實(shí)驗(yàn)結(jié)束后,用會(huì)聲會(huì)影軟件分析動(dòng)物的運(yùn)動(dòng)過程。用兩項(xiàng)指標(biāo)評(píng)價(jià)龜?shù)倪\(yùn)動(dòng)表現(xiàn)：(1)疾游速,指幼龜游過30 cm的最快速度；(2)最大持續(xù)運(yùn)動(dòng)距離,指幼龜持續(xù)游動(dòng)的最大距離。

1.4 耐受溫度測(cè)定

臨界致死高溫(CTMax)和臨界致死低溫(CTMin)測(cè)定在光照培養(yǎng)箱(寧波萊福)中進(jìn)行。實(shí)驗(yàn)開始前,將動(dòng)物放于20 cm×15 cm×8 cm的干燥塑料盒中,并移入溫度預(yù)置為28℃的培養(yǎng)箱內(nèi)適應(yīng)30 min。隨后,箱體溫度以每20 min遞增或遞減2℃的速率向上或向下調(diào)節(jié),達(dá)到培養(yǎng)箱的上下限溫度后,通過加冰和設(shè)置白熾燈來調(diào)節(jié)溫度。CTMax 和CTMin 終點(diǎn)的判定以動(dòng)物在強(qiáng)烈刺激下不作出反應(yīng)(不能眨眼、翻身和爬動(dòng))、移出培養(yǎng)箱后能逐漸恢復(fù)正?；顒?dòng)為標(biāo)準(zhǔn),動(dòng)物拿出后立即測(cè)量并記錄泄殖腔溫度。

1.5 補(bǔ)償生長(zhǎng)能力測(cè)定

將30只紅耳滑龜和30只中華草龜編號(hào)后隨機(jī)分組,每組10只,飼養(yǎng)于室溫(28±0.5)℃恒溫室內(nèi)。3組龜分別作饑餓5、10d和15d處理,期間只提供足量飲水,并及時(shí)換水；饑餓后,足量投飼龜糧15、10d和5d。此外,再選取每種龜各3只作為對(duì)照,不進(jìn)行饑餓處理,在相同的室溫條件下常規(guī)飼養(yǎng)。測(cè)量動(dòng)物實(shí)驗(yàn)起始和終末的體重[27],計(jì)算最后5d的體重變化率用以評(píng)價(jià)補(bǔ)償生長(zhǎng)能力,以每天每克體重變化的毫克數(shù)表示,體重變化率計(jì)算如下：體重變化率=(終末體重-初始體重)×初始體重-1×生長(zhǎng)時(shí)間-1×1000,體重單位為g,生長(zhǎng)時(shí)間單位為d,體重變化率單位為mg g-1d-1。

1.6 外源性蛋白誘導(dǎo)的免疫反應(yīng)測(cè)定

實(shí)驗(yàn)開始前,隨機(jī)選取6月齡紅耳滑龜和中華草龜各10只,擦干后在干燥整理箱中適應(yīng)1d,隨后測(cè)定其左、右后肢腳掌厚度。配制6 mg/mL經(jīng)磷酸緩沖液溶解的植物凝集素(PHA, phytohemagglutinin; Sigma),在動(dòng)物的左后肢注射20 μL,右后肢注射相同劑量的磷酸緩沖液作為對(duì)照,在注藥后3、6、9和18 h時(shí)測(cè)量龜腳掌的厚度。打藥后,動(dòng)物置于干燥整理箱中,允許其自由活動(dòng)。水腫程度(水腫率)用打藥前后腳掌的厚度變化值與腳掌原始厚度比值的百分?jǐn)?shù)表示。

1.7 數(shù)據(jù)分析

用Statistica 8.0和Sigmaplot 11.0軟件包進(jìn)行統(tǒng)計(jì)分析與作圖,所有數(shù)據(jù)經(jīng)正態(tài)性(Kolmogorov-Smirnov test)和方差均質(zhì)性檢測(cè)(F-max test)后進(jìn)行參數(shù)統(tǒng)計(jì)分析,不符合參數(shù)統(tǒng)計(jì)的數(shù)據(jù)經(jīng)自然兌數(shù)或三角函數(shù)轉(zhuǎn)換后進(jìn)行參數(shù)統(tǒng)計(jì)。用單因子方差分析(one-way ANOVA)檢測(cè)各項(xiàng)指標(biāo)的種間差異,用重復(fù)檢驗(yàn)方差分析(repeated measures ANOVA)檢測(cè)溫度對(duì)同一物種各項(xiàng)生理指標(biāo)的影響。實(shí)驗(yàn)或飼養(yǎng)過程中,死亡個(gè)體的數(shù)據(jù)不用于統(tǒng)計(jì)分析。所有描述性統(tǒng)計(jì)值均用平均值±標(biāo)準(zhǔn)離差表示,變異系數(shù)(coefficient of variation)用數(shù)據(jù)組的標(biāo)準(zhǔn)離差與平均值的比值乘上100來表示,顯著性水平設(shè)置為α=0.05。

2 結(jié)果

2.1 體溫調(diào)節(jié)

圖1 紅耳滑龜和中華草龜幼體的體溫調(diào)節(jié) Fig.1 Body temperature regulation of hatchling T. S. elegans and C. reevesii *P < 0.05; ns無顯著差異

兩種龜分開飼養(yǎng)時(shí),紅耳滑龜幼體平均調(diào)節(jié)體溫為26.5 ℃,最低和最高體溫分別為20.1 ℃和32.9 ℃,體溫調(diào)節(jié)的變異系數(shù)為11.1；中華草龜幼體平均調(diào)節(jié)體溫為28.4 ℃,最低和最高體溫分別為24.9 ℃和32.4 ℃,體溫調(diào)節(jié)的變異系數(shù)為8.3(圖1)。兩種幼龜混養(yǎng)時(shí),紅耳滑龜平均調(diào)節(jié)體溫為26.1 ℃,最低和最高體溫分別為21.8 ℃和32.9 ℃,體溫調(diào)節(jié)的變異系數(shù)為10.9；中華草龜平均調(diào)節(jié)體溫為26.5 ℃,最低和最高體溫分別為22.2 ℃和31.5 ℃,體溫調(diào)節(jié)的變異系數(shù)為9.2(圖1)。

單因子方差分析顯示,分開飼養(yǎng)時(shí),紅耳滑龜和中華草龜幼體體溫調(diào)節(jié)存在顯著的種間差異(F1,37=4.98,P= 0.032)；混養(yǎng)時(shí),龜?shù)捏w溫調(diào)節(jié)不存在種間差異(F1,37=0.22,P= 0.65)。混養(yǎng)顯著影響中華草龜體溫調(diào)節(jié)(F1,18=7.18,P= 0.015),但對(duì)紅耳滑龜體溫調(diào)節(jié)不存在影響(F1,19=0.24,P= 0.63)(圖1)。兩種條件下,紅耳滑龜體溫調(diào)節(jié)的變異系數(shù)均大于中華草龜。

2.2 運(yùn)動(dòng)能力的熱依賴性

單因子方差分析顯示,3個(gè)溫度下紅耳滑龜幼體的疾游速均顯著大于中華草龜幼體(P<0.0001)(圖2)。重復(fù)檢驗(yàn)方差分析顯示,兩種龜?shù)募灿嗡倬S著水溫的升高而增加(P<0.002),但中華草龜疾游速的增幅小于紅耳滑龜,且28℃和36℃下中華草龜?shù)募灿嗡俨淮嬖诓町?F1,15=0.29,P=0.60)。低溫和高溫下,紅耳滑龜?shù)某掷m(xù)最大運(yùn)動(dòng)距離顯著大于中華草龜(P<0.005),但28℃下,兩種龜?shù)某掷m(xù)最大運(yùn)動(dòng)距離不存在差異(P=0.51)(圖2)。重復(fù)檢驗(yàn)方差分析顯示,兩種龜?shù)某掷m(xù)最大運(yùn)動(dòng)距離隨水溫的升高而增加(P<0.003),但中華草龜?shù)脑龇∮诩t耳滑龜,且28℃和36℃下中華草龜?shù)某掷m(xù)最大運(yùn)動(dòng)距離不存在差異(F1,15=0.005,P=0.95)。

圖2 紅耳滑龜和中華草龜幼體運(yùn)動(dòng)表現(xiàn)的熱依賴性Fig.2 Thermal dependence of locomotor performance of hatchling T. S. elegans and C. reevesii

2.3 耐受溫度

圖3 紅耳滑龜和中華草龜幼體的臨界致死高溫與低溫 Fig.3 The critical thermal maximum (CTMax) and critical thermal minimum (CTMin) of hatchling T. S. elegans and C. reevesii

單因子方差分析顯示,中華草龜和紅耳滑龜幼體的臨界致死高溫(F1,28=7.48,P= 0.011)和臨界致死低溫(F1,28=4.38,P= 0.045)存在明顯的種間差異。其中,紅耳滑龜平均臨界致死高溫和低溫分別為41.2℃和3.6℃,中華草龜則分別為37.2℃和4.5℃(圖3),紅耳滑龜?shù)母叩蜏啬褪苣芰γ黠@比中華草龜強(qiáng)。

2.4 饑餓后的補(bǔ)償生長(zhǎng)

單因子方差分析顯示,兩種幼龜?shù)酿囸I處理組與對(duì)照組相比,均存在明顯的補(bǔ)償生長(zhǎng)現(xiàn)象(P<0.041)。在補(bǔ)償生長(zhǎng)階段,饑餓5d處理的兩種龜,體重變化率不存在差異(F1,16=0.03,P=0.86),饑餓10d處理的紅耳滑龜體重變化率顯著大于中華草龜(F1,15=5.12,P=0.038),而饑餓15d處理的中華草龜體重變化率顯著大于紅耳滑龜(F1,16=8.29,P=0.011)(圖4)。3種饑餓處理下的中華草龜平均體重變化率為5.6 mg g-1d-1,變異系數(shù)為72.8；紅耳滑龜?shù)钠骄w重變化率為5.2 mg g-1d-1,變異系數(shù)為24.6,紅耳滑龜?shù)捏w重變化率更為穩(wěn)定。

2.5 植物凝集素誘導(dǎo)的水腫反應(yīng)

兩種幼龜注射植物凝集素后,左后肢均出現(xiàn)水腫現(xiàn)象。去除部分測(cè)量誤差導(dǎo)致的無效樣本,剩余12個(gè)樣本用于統(tǒng)計(jì)分析。單因子方差分析顯示,注藥后3h和6h,紅耳滑龜水腫程度明顯要比中華草龜強(qiáng)(P<0.0055),注藥后9h和18h,兩者水腫程度不存在差異(P> 0.075)(圖5)。重復(fù)檢驗(yàn)方差分析顯示,紅耳滑龜4個(gè)時(shí)間點(diǎn)的水腫程度存在顯著差異(F3,15=3.70,P=0.036),注藥后6h水腫達(dá)到最大程度,隨后開始消腫；中華草龜4個(gè)時(shí)間點(diǎn)的水腫程度也存在顯著差異(F3,15= 14.48,P=0.00011),隨時(shí)間延長(zhǎng),水腫程度逐漸加強(qiáng)(圖5)。

圖4 恢復(fù)生長(zhǎng)階段中華草龜和紅耳滑龜幼體的體重變化 Fig.4 Body mass changes of hatchling C. reevesii and T. S. elegans during the compensatory growth

圖5 植物凝集素誘導(dǎo)的紅耳滑龜和中華草龜幼體腳掌水腫反應(yīng) Fig.5 Phytohemagglutinin-induced swelling in the paws of hatchling C. reevesii and T. S. elegans

3 討論

本研究結(jié)果顯示,從單物種分開飼養(yǎng)到兩物種混合飼養(yǎng),紅耳滑龜幼體的平均調(diào)節(jié)體溫不發(fā)生漂移,而中華草龜幼體的平均調(diào)節(jié)體溫從28.4 ℃降至26.5 ℃,這可能與中華草龜近距離接觸熱源的機(jī)會(huì)下降有關(guān)。較多的體溫調(diào)節(jié)機(jī)會(huì)可以使爬行動(dòng)物保持最佳體溫,能提高其運(yùn)動(dòng)能力、食物消化能力及其它生理狀態(tài)。曬背機(jī)會(huì)的降低直接影響中華草龜?shù)钠骄{(diào)節(jié)體溫,進(jìn)而降低其生理、行為表現(xiàn),而紅耳滑龜混養(yǎng)前后平均調(diào)節(jié)體溫不存在差異,表明其調(diào)溫機(jī)會(huì)或調(diào)溫節(jié)律不發(fā)生改變?？梢?在混養(yǎng)環(huán)境中,紅耳滑龜對(duì)熱資源的競(jìng)爭(zhēng)擁有明顯的優(yōu)勢(shì)。這與Cadi和Joly的報(bào)道類似,在紅耳滑龜和歐洲澤龜(Emysorbicularis)混養(yǎng)的池塘中,前者占據(jù)的曬背場(chǎng)更多,導(dǎo)致后者的調(diào)溫機(jī)會(huì)明顯減少[28]。受紅耳滑龜?shù)挠绊?地中海擬水龜?shù)臅癖彻?jié)律發(fā)生明顯變化,該種龜往往避開與紅耳滑龜同一時(shí)間點(diǎn)曬背,這導(dǎo)致其曬背時(shí)間也明顯縮短[8]。

爬行動(dòng)物溫度耐受能力受外部因素和內(nèi)部因素的影響,包括馴化環(huán)境、地理、性別、年齡和生理狀態(tài)等[29- 35]。高低溫耐受能力影響動(dòng)物的地理分布范圍,低緯度種類往往具有較強(qiáng)的耐受高溫,高緯度種類則具有較強(qiáng)的耐受低溫[36,37]。紅耳滑龜?shù)呐R界致死高溫和低溫分別高于和低于中華草龜,其高低溫耐受幅比中華草龜寬,表明其潛在的分布范圍比中華草龜廣,有利于該種龜向高緯度和低緯度地區(qū)擴(kuò)散。單養(yǎng)和混養(yǎng)時(shí),紅耳滑龜體溫調(diào)節(jié)的變異系數(shù)均比中華草龜要大,表明紅耳滑龜?shù)捏w溫調(diào)節(jié)幅較寬,也進(jìn)一步證實(shí)其潛在的分布范圍要廣于中華草龜。

運(yùn)動(dòng)表現(xiàn)是影響爬行動(dòng)物適合度的重要特征,受眾多生理和生態(tài)因素影響[38]。爬行動(dòng)物疾跑(游)速常隨體溫升高而加快,當(dāng)體溫到達(dá)一定限度時(shí),又會(huì)隨體溫增加而減慢[39-41]。疾跑(游)速可以體現(xiàn)爬行動(dòng)物的最大運(yùn)動(dòng)潛能,持續(xù)最大運(yùn)動(dòng)距離則能體現(xiàn)其最大運(yùn)動(dòng)耐力[42],這兩項(xiàng)指標(biāo)直接與野外幼龜獲取食物資源和逃避天敵的能力有關(guān)[43]。本研究中,紅耳滑龜和中華草龜幼體運(yùn)動(dòng)能力模式與中華花龜(Ocadiasinensis)[43]和中華鱉(TrionyxSinensis)[44]幼體相似,均隨體溫的升高而加強(qiáng)。相同溫度下的中華草龜相比,紅耳滑龜具有更快的疾游速和更長(zhǎng)的持續(xù)最大運(yùn)動(dòng)距離,因此也就具有更佳的運(yùn)動(dòng)潛能,在野外競(jìng)爭(zhēng)食物和躲避天敵時(shí),也就比中華草龜具有更多的優(yōu)勢(shì)。

在自然環(huán)境中,食物分布存在空間和時(shí)間異質(zhì)性,動(dòng)物常常在生活周期的一定階段因食物短缺而受到饑餓脅迫[45]。耐饑餓能力高低是爬行動(dòng)物能否在自然環(huán)境中維持正常存活的主要因素之一,而饑餓攝食后的補(bǔ)償生長(zhǎng)能力高低則是爬行動(dòng)物迅速恢復(fù)饑餓前體能狀態(tài)的重要機(jī)制,強(qiáng)而穩(wěn)定的補(bǔ)償生長(zhǎng)能力有助于動(dòng)物在食物資源多變的環(huán)境中保持穩(wěn)定的生存狀態(tài)[46,47]。本研究中,盡管兩種龜均存在饑餓攝食后補(bǔ)償生長(zhǎng)現(xiàn)象,且3種饑餓處理下兩種龜?shù)钠骄w重變化率相近,說明兩種龜在長(zhǎng)期饑餓后都能通過補(bǔ)償生長(zhǎng)來彌補(bǔ)饑餓對(duì)體重的影響,但紅耳滑龜?shù)难a(bǔ)償生長(zhǎng)變異系數(shù)要明顯比中華草龜小,其在饑餓不同時(shí)間后所表現(xiàn)的補(bǔ)償生長(zhǎng)能力相同,即具有更穩(wěn)定的補(bǔ)償生長(zhǎng)能力,這有利于紅耳滑龜維持較穩(wěn)定的生存狀態(tài)。

脊椎動(dòng)物免疫能力的強(qiáng)弱是影響其環(huán)境適應(yīng)能力的重要因素,用外源性蛋白(如植物凝集素)誘導(dǎo)水腫反應(yīng)則是評(píng)價(jià)脊椎動(dòng)物免疫能力強(qiáng)弱的重要技術(shù),水腫反應(yīng)越強(qiáng)烈,表明機(jī)體抵御外源性蛋白作用的能力越強(qiáng),該技術(shù)目前已被廣泛應(yīng)用[48-49]。在自然環(huán)境中存在各種致病菌,爬行動(dòng)物在適應(yīng)環(huán)境或種間競(jìng)爭(zhēng)過程中,往往面臨各種不確定致病菌的影響,機(jī)體對(duì)抗致病菌的免疫應(yīng)答能力越強(qiáng),其存活幾率就越大[50]。本研究中,兩種龜首次注射植物凝集素,表現(xiàn)的是其天然免疫能力。兩種龜對(duì)植物凝集素均表現(xiàn)出明顯的免疫反應(yīng)性,但紅耳滑龜對(duì)植物凝集素作出的免疫反應(yīng)比中華草龜要快,表明該種對(duì)外源性蛋白作出的免疫應(yīng)答比中華草龜要迅速。因此,與土著種——中華草龜相比,紅耳滑龜快速的免疫應(yīng)答能力使其在適應(yīng)環(huán)境時(shí)能保持更強(qiáng)的優(yōu)勢(shì)。

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Body temperature, thermal dependence of locomotor performance,compensatory growth, and the immunity of hatchling Red-eared turtles and Chinese pond turtles

ZHOU Zan1, CAI Mengsi1, LIU Yu1, SUN Yanyan2,*, LUO Laigao3

1CollegeofLifeandEnvironmentalSciences,HangzhouNormalUniversity,Hangzhou310036,China2DongyingForestryBureau,Dongying257091,China3CollegeofBiologyandFoodEngineeringChuzhouUniversity,Chuzhou239000,China

Invasion by alien species has become a major problem that threatens the security of our ecological environment. It is one of the three major environmental problems in the world today. Preventing alien invasions has become a focus for both government and public sectors, and a focal topic within academic circles. Of the many of invasive species, the Red-eared turtle (Trachemysscriptaelegans) poses a particularly serious threat. As one of the 100 most dangerous invasive species in the world, Red-eared turtles have caused irreversible effects on the ecosystems they invade. Previous studies have shown that the Red-eared turtle is able to cause such extensive damage because of its ability to adapt to its environment. Some of their physiological and behavioral indicators show an advantage, but there is still insufficient data to explain their competitive advantage overall. This paper examines the body temperature, dependence of locomotion on temperature, compensatory growth, and the immunity of hatchling Red-eared turtles and Chinese pond turtles (Chinemysreevesii). We found that Red-eared turtles could significantly influence the thermoregulatory behavior of the Chinese pond turtle. The Red-eared turtle exhibited a wider range of body temperatures and thermal tolerance, better locomotor performance, and innate immunity in comparison to its counterpart; indicating a more stable compensatory growth mechanism. When exposed to a thermal gradient, Red-eared turtles that were bred together with Chinese pond turtles selected lower body temperatures and had larger coefficient of variation of body temperatures than those bred separately. The upper (CTMax) and lower (CTMin) limits of thermal tolerance were 41.2 ℃ and 3.6 ℃ for the Red-eared turtle, and 37.2 ℃ and 4.5 ℃ for the Chinese pond turtle, respectively. We therefore concluded that Red-eared turtles have an advantage in the thermal resource competition in a polyculture environment. The locomotor performance of the two species was measured when the body temperature was held at three different levels: 20, 28 ℃ and 36 ℃. Sprint speed and maximal distance increased with body temperature of both species and the locomotor performance of the Red-eared turtle was better than that of the Chinese pond turtle when compared at the same body temperature. Therefore, the Red-eared turtle exhibited better movement potential and likely has more advantages in the field during competition for food and predator evasion. Compensatory growth was observed in the two turtles following starvation. The change in body mass of the two species increased evenly under the five-day starvation treatment. Body mass increased more rapidly in the Red-eared turtle than in the Chinese pond turtle when subjected to the 10-day-long starvation treatment and more slowly when under the 15-day starvation treatment. The coefficient of variation for body mass was larger in the Chinese pond turtle than in the Red-eared turtle. Both species showed immunoreactivity to phytohemagglutinin; with the immunoreactivity of the Red-eared turtle taking six hours to reach its maximum level-significantly faster than that of the Chinese pond turtle. These results indicate that the Red-eared turtle can adapt to new environments quickly and therefore threaten the native turtles as an alien species. This adaption could be ascribed to the potential advantages in their locomotor and physiological characteristics.

Red-eared turtle; Chinese pond turtle; invasion; body temperature and locomotor performance; compensatory growth; immunity

浙江省重點(diǎn)科技創(chuàng)新團(tuán)隊(duì)項(xiàng)目(2010R50039)；杭州西溪濕地研究中心開放研究基金資助項(xiàng)目(WR005)；安徽省自然科學(xué)基金項(xiàng)目(1508085MC61)

2015- 02- 11;

日期:2016- 03- 03

10.5846/stxb201502110333

*通訊作者Corresponding author.E-mail: sunyanyan8866@163.com

周贊, 蔡夢(mèng)思, 劉虞, 孫燕燕,羅來高.紅耳滑龜和中華草龜幼體體溫與運(yùn)動(dòng)的熱依賴性、補(bǔ)償生長(zhǎng)及免疫力.生態(tài)學(xué)報(bào),2016,36(21):7014- 7022.

Zhou Z, Cai M S, Liu Y, Sun Y Y, Luo L G.Body temperature, thermal dependence of locomotor performance, compensatory growth, and the immunity of hatchling Red-eared turtles and Chinese pond turtles.Acta Ecologica Sinica,2016,36(21):7014- 7022.