利用拉曼顯微光譜技術及成像法觀測紅球菌SD-74生物薄膜在原處的生長過程

Abstract

在自然界中絕大部分的細菌生活在生物薄膜（Biofilm）中，在生物薄膜中細菌被包埋在由胞外聚合物組成的基質中並貼附在表面上，最近的研究指出細菌以生物薄膜的形態生活可以提高其存活率例如產生極高的抗藥性等等，雖然生物薄膜的特殊結構及功能引起了各類科學上的興趣，在探討其在活體及在原處的化學特性仍是一大挑戰。 在本論文中我們使用拉曼顯微光譜技術及成像法來針對菌種紅球菌SD-74（Rhodococcus sp. SD-74）的生物薄膜及其生長過程做討論。拉曼光譜技術提供了許多優點於在原處的生物薄膜研究，包括非破壞性、不需要特殊標記及不受來自水的訊號干擾等。我們成功地獲得在生物薄膜生長過程的詳細分佈及組成改變資訊，我們發現隨著生物薄膜的生長在基質中的類胡蘿蔔素濃度極巨的增加，為了解釋這個類胡蘿蔔素累積的現象我們提出了一個假設：在此類胡蘿蔔伴演抗氧化的角色來對抗可能的氧化壓力，而我們所做的在不同光照條件的拉曼成像實驗結果支持這個假設。

Most bacteria in natural habitats live as biofilms, in which bacterial cells are embedded in matrices of extracellular polymeric substances adherent to a surface. Recent studies have revealed that the biofilm mode of living increases the survival rate of bacteria by e.g., acquiring extremely high antibiotic resistance. Although unique structures and functions of biofilms have attracted much attention in many disciplines of science, it remains challenging to study biofilms in vivo and in situ as well as with high chemical specificity. In the present study, we focus on the biofilms of the bacterial species of practical interest known as Rhodococcus sp. SD-74 and monitor the process of its biofilm development using Raman microspectroscopy and imaging. Because Raman spectroscopy is nondestructive and label-free, and is less hampered by the presence of water, Raman microspectroscopy and imaging are an ideal tool to study biofilms in situ. We successfully obtained detailed distribution and composition changes within R. sp. SD-74 biofilm during the development process. We found that the concentration of carotenoids in the biofilm matrix drastically increases as biofilm development proceeds. To interpret this phenomenon of carotenoid accumulation, we hypothesize that carotenoids act as an antioxidant in R. sp. SD-74 biofilms and counter possible oxidative stress. Raman imaging experiments performed under different light-illumination conditions support this hypothesis.