Imaging microorganisms and pollutants at the microscale

荧光显微镜是研究自然环境中微生物而无需培养的有力工具。微生物细胞内的核酸可以用特异性结合靶生物的荧光染料染色。该技术可以对天然样品中的污染物降解微生物进行染色，以进行鉴定、定位和计数。微生物降解可以分解和解毒多环芳烃（PAHs）。多环芳烃是存在于原油或焦油等有机液体中的致癌污染物，也存在于英国受工业活动影响的许多土壤或沉积物中。多环芳烃的一个有趣的特性是它们在紫外光下的自发荧光，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微尺度成像的多环芳烃丰富的补丁在土壤和沉积物及其相关的微生物群落。为此，标准将来自识别多环芳烃丰富的补丁在土壤或沉积物中的荧光显微镜。一旦明确的区别多环芳烃丰富的补丁和其他天然土壤或沉积物成分的荧光之间已经实现，一个合适的方法将得出染色和图像相关的微生物使用荧光原位杂交（FISH），传统的落射荧光显微镜和共聚焦激光扫描显微镜。衍生的方法可用于详细研究，并在微观尺度上的微生物和污染物，如多环芳烃在自然环境中的共存。微尺度研究有助于揭示污染土壤和沉积物中多环芳烃微生物降解和衰减的确切机制。这种认识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.