Endospore-forming bacteria in the genera Bacillus and Clostridium play pivot roles in food hygiene, medical sterilization, disease control, astrobiology, and so on. Sporulation is the process that a vegetative cell turns to an endospore in times of harsh environment to protect itself from being killed. When favorable condition arises, endospore will germinate, followed by outgrowth and eventually return to vegetative cell. Germination is affected by various factors including sporulation condition, duration and temperature of heat shock, choice of germinant, inoculation size of germination, germination temperature, and many others.

This thesis is composed by two parts. One is on microbiology and the other is on microfluidics. The first part consists of three independent projects (Project 1-3), each of which respectively investigated the effect of oxygen, heavy water, or static magnetic field on germination. Specifically, germination of Bacillus atrophaeus endospores were triggered in the presence or absence of these factors and monitored using terbium - dipicolinic acid fluorescence assay, phase contrast microscopy, conductance measurement, and optical density measurement. Culturability of endospore germinated with or without these external factors was also tested. We firstly reported that oxygen or heavy water had an inhibitory effect on germination, while static magnetic field didn’t had such effect.

The second part also consisted of three independent projects (Project 4-6), all of which used microfluidic technology to study germination or to manipulate endospores. In Project 4, we designed a novel digital microfluidic system to monitor germination process by conductometry. Droplet containing endospores was actuated and merged with droplet containing germinant. Germination was tracked by measuring the conductivity of the mixed droplet with minimized sample consumption. In Project 5, we proposed a dielectrophoresis microsystem to obtain high-purity endospore suspension. Under proper voltage and operation frequency, vegetative cells and endospores underwent positive dielectrophoresis and negative dielectrophoresis respectively and could hence be separated. In Project 6, we proposed another dielectrophoresis microsystem which was able to study the germination characteristics in the absence of chemical communications. Germinating endospores were fixed in a microchannel by dielectrophoresis while germination byproducts, which may be employed as medium of chemical communications, were continuously removed by flushing flow. The feasibility of systems proposed in Project 5 and 6 are based on numerical simulation.

This thesis contributed new information on both endospore physiology (science aspect) and microfluidics (engineering aspect).

桿菌屬和梭菌屬具有形成芽孢的能力。它們在食品衛生，醫學殺菌，疾病控制， 和天體生物學等領域都扮演著重要的角色。在惡劣的環境下，它們的營養細胞可 以形成芽孢，以保護自己不被殺死。在合適的情況下，芽孢會萌發，生長，最終 變成營養細胞。孢萌發過收到很多因素影響，諸如形成芽孢時的環境，芽孢萌發 前預熱激活的時長，萌發劑的選擇，芽孢萌發時的溫度，芽孢萌發時芽孢懸濁液 的濃度等等。

本論文由兩部分組成，第一部分是關於微生物學，第二部分是關於微流控技術。 第一部分由三個獨立的項目組成（項目 1-3）。三個項目分別研究了氧氣，重水 和恆定磁場對萎縮芽孢桿菌芽孢萌發過程的影響。具體來說，芽孢在施加或不施 加以上三個外界因素的條件下萌發。我們使用鋱-2,6-吡啶二羧酸（Tb-DPA）熒 光法，相差顯微鏡法，電導測定法和光密度測量法，來監測萌發過程。同時，我 們對在這些外界因素下萌發的芽孢的可培養性做了測試。我們首次報導了氧氣和 重水對萌發過程有抑製作用，而恆定磁場對萌發過程沒有影響。

第二部分也由三個相互獨立的子項目組成（項目 4-6）。這三個子項目均是通過 微流控的平台來研究萌發或者操控芽孢。具體來說，在第 4 個項目裡，我們創新 地在數字微流體芯片上，用電導測定法來監測芽孢萌發過程。我們驅動並混合了 含有芽孢的液滴和含有萌發劑的液滴，並用混合液滴電導率隨時間的變化來表徵 萌發過程。在第 5 個項目裡，我們提出了一種基於介電電泳的方法來獲取高純度 梭菌芽孢懸濁液。在適當的電壓和操作頻率下，營養細胞和芽孢分別受到正介電 電泳力和負介電電泳力的作用而被分離。在第 6 個項目裡，我們提出了一種基於 介電電泳的方法來研究芽孢在沒有化學通信情況下的萌發特點。我們在微流控管 道中用介電電泳力來固定正在萌發的芽孢，同時在入口處不斷注入流體，使得萌 發過程中所有的化學產物（這些產物可能被用作化學通信的媒介）可以被不斷地 沖刷走。我們用數值模擬的方法，對項目 5 和 6 中系統的可行性做了一個初步的 評估。

本論文從芽孢生理學（科學）和微流控技術（工程）兩方面，為細菌芽孢的研究 做出了新的貢獻。