圈養無尾熊採食偏好與腸道發酵特性之評估

Abstract

本研究以臺北市立動物園圈養之無尾熊研究對象，探討一年四季所採食之桉樹葉成分的變化，對無尾熊採食偏好與其糞便菌相的影響。透過菌相的監測並以體外模式模擬無尾熊的後腸發酵，以了解無尾熊的腸道發酵特性狀況。 第一部分試驗採用6公6母共12隻無尾熊，採集一年度內(2021年5月至2022年3月)四個季節的各種新鮮桉樹葉，記錄無尾熊的採食評分並進行各桉樹葉之成分分析，同時進行無尾熊糞便菌相與發酵能力評估。成分分析的結果顯示，主食桉中的大葉桉（Eucalyptus.robusta）、細葉桉（E.tereticornis）、赤桉（E. camaldulensis）、脂桉（E.microcorys）與副食桉中的樹膠桉（E.viminalis）與白桉（E.alba）的採食評分及化學組成，在涼季暖季之間有顯著差異（p<0.05），在涼、暖季呈現不同的特性。但副食桉中的檸檬桉（E.citriodora）、小果灰桉（E.microcarpa）、斑桉（E.punctata）與玫瑰桉（E.rhodantha），於四季間成分變動性不大，且無尾熊並沒有在任何季節特別偏好此四種桉樹葉。雄性無尾熊偏好夏季的大葉桉與廣葉桉（E.amplifolia），秋季的大葉桉、脂桉、白桉，冬季的細葉桉、白桉。雌性無尾熊偏好夏季的大葉桉，秋季的細葉桉與樹膠桉。雄性無尾熊對同種桉樹葉在不同季節間的偏好性比起雌性來得較明顯。夏季的大葉桉與廣葉桉、秋季的樹膠桉與冬季細葉桉、白桉為無尾熊顯著偏好的五種葉子，分析桉樹葉中之水分、中洗纖維（Neutral detergent fiber，NDF）、粗蛋白（Crude protein，CP）、總酚（Total phenols，TP）、單寧酸（Tannic acid，TA）與可溶性碳水化合物（Water soluble carbohydrates，WSC）等成分後，採食營養成分如CP、WSC顯著較高者，無尾熊的採食評分也顯著較高，但抗營養因子TP、TA卻不一定較低，而在無尾熊不偏好的葉子如夏季的脂桉，出現WSC高但TP、TA卻低的現象，可見無尾熊偏好的葉子並不一定同時符合高營養成分且低抗營養因子的條件。 糞便細菌計數的結果顯示，各種細菌的比例在雄性與雌性無尾熊之間並無顯著差異，一年四季中無尾熊糞便菌相整體而言都是穩定的。年齡與性別也影響也不大，菌相的變化趨勢不如成分分析在涼季、暖季變換時有顯著的分別，即使葉片組成分有差異，無尾熊可能仍可透過靈活的採食策略來使腸道的環境維持穩定。 第二部分試驗以體外模式模擬無尾熊後腸發酵，分析後腸發酵特性並對照新鮮糞便中揮發性脂肪酸的組成。體外發酵的揮發性脂肪酸分析結果顯示，乳酸的比例在夏季顯著高於冬季，乙酸是冬季顯著高於夏季，丙酸是冬季顯著高於夏季與秋季，丁酸則是在秋季顯著高於冬季與春季。而揮發性脂肪酸結果也發現涼季與暖季呈現不同的特徵。雖然四季之間比例有些浮動，但是[乳酸+乙酸]與[丙酸+丁酸]的比例大致介於7:3至6:4，發酵狀況維持穩定。糞便揮發性脂肪酸分析結果顯示，性別與季節的不同不影響揮發性脂肪酸的組成，無尾熊即使在季節間對於桉樹葉產生不同的採食偏好，腸道發酵的環境還是能保持穩定。 綜上所述，無尾熊在四季的採食選擇的因素不同，而不同性別間選擇標準也不同，雖然無尾熊對於葉子的選擇標準不一，從發酵產物的角度來看，無尾熊可能透過其適應力能在面對眾多品種桉樹葉之下，權衡營養攝取與次級代謝物的危害，穩定其腸道菌相以及後腸發酵產生的揮發性脂肪酸的比例。

This study attempted to investigate the effects of variation in the composition of eucalyptus leaves throughout the year on the intake preferences and fecal flora. The flora assay and in vitro intestinal fermentation simulation were applied to evaluate the fermentation characteristics and of the koalas. In the first part of the experiment, twelve koalas (six males and six females) were involved in the experiment. Over the four seasons, fresh eucalyptus leaves were collected and analyzed.Dietary scores of koalas were recorded, the fecal flora and fermentation ability were examinated.The results showed significant differences (p<0.05) in dietary scores and chemical composition between the cool (winter and spring) and warm (summer and autumn) seasons for the main intake eucalyptus species (E.robusta, E.tereticornis, E.camaldulensis and E.microcorys) and two of subordinary intake species (E.viminalis and E.alba). However the other four subordinary intake species(E. microcarpa, E. citriodora, E. punctata, E. rhodantha) had no significant differences whether in dietary scores or chemical composition among four seasons. Male koalas preferred to intake E. robusta and E. amplifolia in summer, E.robusta, E.microcorys and E. alba in autumn, E.tereticornis and E.alba in winter. Female koalas preferred to intake E. robusta in summer, and E.tereticornis and E.viminalis in autumn. Males exhibited a greater preference for the same species of eucalyptus leaves between seasons than females. The chemical composition of five preferring intake eucalyptus species included E. robusta and E. amplifolia in summer, E. viminalis in autumn, E. tereticornis and E. alba in winter were assayed. The assay results of water content, neutral detergent fiber (NDF), crude protein (CP), total phenols (TP), tannic acid (TA), water soluble carbohydrates (WSC) indicated that the leaves preferred by koalas were not necessarily higher in nutrients (CP, WSC) and lower in anti-nutritional factors (NDF, TP, TA). Fecal bacteria count result showed no significant differences in the ratios of different types of microbe between whether genders or ages. Although eucalyptus leaves' composition varies from cool to warm season, the stable microflora indicated that koalas adopt a flexible dietary strategy to maintain a stable intestinal environment. In the second part of the experiment, in vitro hindgut fermentation was applied to simulate hindgut fermentation of koalas for fermentation characteristics evaluation and comparing the composition of VFAs in fresh feces. Results of in vitro fermentation showed different characteristics between the cool and warm seasons, the proportion of lactate was significantly higher in summer than in winter, acetate was significantly higher in winter than in summer, propionate was significantly higher in winter than in summer and autumn, and butyrate was significantly higher in autumn than in winter and spring. However, the ratio of [lactic acid + acetic acid] to [propionic acid + butyric acid] maintains a steady state varying from 7:3 to 6:4. Coupled with the results of fresh fecal VFA composition that showed no significant differences between whether genders or ages, while there are preferences for various species of eucalyptus leaves, the environment for intestinal fermentation is still stabilized. Although the selection criteria of eucalyptus leave varied among the four seasons, the strong adaptability of eucalyptus koalas was able to balance nutritional intake with the risk of secondary metabolites and stabilize their intestinal microflora and the proportion of volatile fatty acids produced by hindgut fermentation in the face of many species of eucalyptus leaves.