Imaging microorganisms and pollutants at the microscale

荧光显微镜是研究自然环境中无需培养的微生物的有力工具。微生物细胞内的核酸可以用荧光染料染色，这种染料专门结合到目标生物上。这项技术可以对天然样品中的污染物降解微生物进行染色，以便对其进行鉴定、定位和计数。微生物降解可以分解和解毒多环芳烃(PAHs)。多环芳烃是一种致癌污染物，存在于原油或焦油等有机液体中，也存在于受工业活动影响的英国许多土壤或沉积物中。多环芳烃的一个有趣的性质是它们在紫外光下的自体荧光，这意味着它们可以用荧光显微镜观察到。因此，荧光显微镜可能适合于同时对土壤和沉积物中富含多环芳烃的斑块及其相关微生物群落进行微尺度成像。为此，将制定用荧光显微镜识别土壤或沉积物中富含多环芳烃斑块的标准。一旦明确区分了富含多环芳烃的斑块和其他自然土壤或沉积物成分的荧光，就会得出一种适当的方法，使用荧光原位杂交(FISH)、传统的荧光显微镜和共聚焦激光扫描显微镜对相关微生物进行染色和成像。该方法可用于在微尺度上详细研究微生物与多环芳烃等污染物在自然环境中的共存情况。对污染土壤和沉积物中多环芳烃的微生物分解和衰减的精确控制机制，可以通过微观研究来阐明。这种洞察力对于生物修复技术的设计和受影响生态系统的风险评估至关重要。

Fluorescence microscopy is a powerful tool for studying microorganisms in their natural environment without cultivation. The nucleic acids inside microbial cells can be stained with fluorescing dyes which specifically bind to targeted organisms. This technique can stain pollutant degrading microorganisms in natural samples for their identification, localisation and enumeration. Microbial degradation can break down and detoxify polycyclic aromatic hydrocarbons (PAHs). PAHs are carcinogenic pollutants present in organic liquids like crude oil or tar and also in numerous soils or sediments in the United Kingdom impacted by industrial activity. An interesting property of PAHs is their autofluorescence under ultraviolet light which means that they can be visualised with fluorescence microscopy. Fluorescence microscopy may thus be suitable for the simultaneous microscale imaging of PAH-rich patches in soils and sediments and their associated microbial communities. To this end criteria will be derived for the identification of PAH-rich patches in soil or sediment by fluorescence microscopy. Once the unambiguous distinction between the fluorescence of PAH-rich patches and other natural soil or sediment constituents has been achieved, a suitable methodology will be derived to stain and image the associated microorganisms using fluorescence in situ hybridisation (FISH), conventional epifluorescence microscopy and confocal laser scanning microscopy. The derived methodology could be used to study in detail and at the microscale the co-localisation of microorganisms and pollutants such as PAHs in the natural environment. A microscale investigation may shed light on the exact mechanisms controlling the microbial breakdown and attenuation of PAHs in polluted soils and sediments. Such insight is of fundamental importance for the design of bioremediation technologies and for the risk assessment for impacted ecosystems.