Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/93008
標題: Profiling of Amino Acids Consumption of Bacteria by Flow-injection Analysis with Electrochemical Detection: Evaluation of Nutrition Requirements and Differentiation of Bacterial Species
以流動注入法結合電化學分析偵測不同菌種對 胺基酸消耗之研究:營養需求與菌種區別評估
作者: Wen-Hua Hsu
許文華
關鍵字: 胺基酸營養需求;胺基酸消耗;菌種區別;Amino acids requirements;Amino acids consumption;Bacterial species differentiation
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摘要: 
細菌使用胺基酸作為細胞中碳及氮分子來源與合成蛋白質,為生理代謝上之關鍵物質,而細菌依據種別與生化特性需求之不同有可能會選擇使用不同的胺基酸。本研究的目的為利用流動注入分析搭配電化學偵測胺基酸的濃度變化,評估不同菌種對於組成蛋白質的20個基本胺基酸的個別消耗能力,以建構不同菌種的胺基酸消耗圖譜(pattern),並以此圖譜作為區別細菌種別的依據。實驗使用8個菌種共28個菌株,分別為各3株的Staphylococcus aureus、S. hyicus、Streptococcus suis、Enterococcus faecalis、Escherichia coli、7株P. multocida、1株P. aeruginosa與5株不同血清型的Salmonella enterica。實驗先將細菌數量調整為約1x109 CFU/mL,並培養於含有單一胺基酸的液態培養基中18個小時,然後取上清液偵測胺基酸的濃度變化。結果顯示,不同菌種間的20個胺基酸消耗差異可形成特異性的胺基酸消耗譜。革蘭氏陽性菌相對於陰性菌消耗較多的glycine、proline與tyrosine,陰性菌則消耗較多asparagine與cysteine。其中S. aureus與S. hyicus除了proline外其他胺基酸的消耗情況非常類似,E. coli與Salmonella的胺基酸消耗需求較相近。P. aeruginosa相較於其他菌種在20個胺基酸上幾乎都有大量的消耗。然而大部分S. suis, E. faecalis與P. multocida對20個胺基酸都只有少量的消耗。以主成分分析與gene comparison模式分析發現可以根據種別間不同的胺基酸消耗差異區別細菌種別。五種血清型的Salmonella在20個胺基酸中有13個有消耗上的差異,此外P. multocida A型與D型、或有無產毒株在胺基酸的消耗程度上也有不同。總而言之,所實驗的菌種具有特徵性的胺基酸消耗譜,藉由胺基酸消耗譜有機會能應用於細菌種別甚至是病原菌的區別與鑑定,且此方法能快速地提供在營養貧乏環境下,細菌對胺基酸的營養需求與其代謝相關之重要資訊。這些結果將有助於開發新的細菌鑑別方法以及設計特殊需求的選擇性培養基。

Amino acids (AAs) are critical substances for nitrogen and carbon metabolism in bacteria. Different bacteria may have different ability to utilize AA depending on their biochemical requirements. Bacteria utilizing AA in a species-dependent manner have been reported. The aims of this study were to establish AA consumption profiles of various bacteria and to identify bacteria species by characteristic AA consumption pattern. Twenty-eight bacterial strains including 3 isolates of Staphylococcus aureus, S. hyicus, Streptococcus suis, Enterococcus faecalis, Escherichia coli, 7 isolates of P. multocida, 5 serovars of Salmonella entrica and 1 isolates of P. aeruginosa were analyzed for their ability to utilize 20 basic AAs. The bacteria were cultured in media containing 3 mM of each AA for 18 hours and the supernatant was analyzed for the remaining AA concentration by flow-injection analysis with electrochemical detection (FIA-EC). The results showed that most bacteria species have characteristic AAs consumption pattern. Gram positive bacteria tended to utilize glycine, tyrosine and proline more in contrast to gram negative bacteria utilizing more asparagine and cysteine. S. aureus exhibited similar pattern to S. hyicus (except for proline) while patterns of E. coli, and Salmonella serotypes were alike. P. aeruginosa consumed large quantities of almost all tested AAs but S. suis, E. faecalis and P. multocida barely used the 20 AAs. Principal component analysis and gene comparison model analysis revealed that bacterial species could be differentiated by their AA consumption. In addition, consumptions for 13 AAs by 5 serotypes of Salmonella entrica were statistically different. Consumptions of AA are also different between type A and type D or between toxigenic and nontoxigenic P. multocida. In conclusion, most of the tested bacteria have characteristic AA consumption pattern such that differentiation of bacteria species could be potentially made. Furthermore, our findings provided pivotal information regarding survival requirements of bacteria under mimimum-nutrional environment and information regarding AA metabolism important for development of markers for bacterial species identification and for design of functional culture medium.
URI: http://hdl.handle.net/11455/93008
其他識別: U0005-2811201416194530
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