Lu Jian-Jiang, Zhang Hao, Li Weiyi, Yi Jun-Bo, Sun Fei-Yun, Zhao Yi-Wei, Feng Liang, Li Zhuo, Dong Wen-Yi
School of Civil and Environmental Engineering, Harbin Institute of Technology (Shenzhen), Shenzhen 518055, China.
Shenzhen Research Institute, The Chinese University of Hong Kong, Shenzhen 518000, China.
Water Res. 2022 Nov 1;226:119243. doi: 10.1016/j.watres.2022.119243. Epub 2022 Oct 13.
Aerobic methane oxidation coupled with denitrification (AME-D) executed in membrane biofilm bioreactors (MBfRs) provides a high promise for simultaneously mitigating methane (CH) emissions and removing nitrate in wastewater. However, systematically experimental investigation on how oxygen partial pressure affects the development and characteristics of counter-diffusional biofilm, as well as its spatial stratification profiles, and the cooperative interaction of the biofilm microbes, is still absent. In this study, we combined Optical Coherence Tomography (OCT) with Confocal Laser Scanning Microscopy (CLSM) to in-situ characterize the development of counter-diffusion biofilm in the MBfR for the first time. It was revealed that oxygen partial pressure onto the MBfR was capable of manipulating biofilm thickness and spatial stratification, and then managing the distribution of functional microbes. With the optimized oxygen partial pressure of 5.5 psig (25% oxygen content), the manipulated counter-diffusional biofilm in the AME-D process obtained the highest denitrification efficiency, due mainly to that this biofilm had the proper dynamic balance between the aerobic-layer and anoxic-layer where suitable O gradient and sufficient aerobic methanotrophs were achieved in aerobic-layer to favor methane oxidation, and complete O depletion and accessible organic sources were kept to avoid constraining denitrification activity in anoxic-layer. By using metagenome analysis and Fluorescence in situ hybridization (FISH) staining, the spatial distribution of the functional microbes within counter-diffused biofilm was successfully evidenced, and Rhodocyclaceae, one typical aerobic denitrifier, was found to survive and gradually enriched in the aerobic layer and played a key role in denitrification aerobically. This in-situ biofilm visualization and characterization evidenced directly for the first time the cooperative path of denitrification for AME-D in the counter-diffused biofilm, which involved aerobic methanotrophs, heterotrophic aerobic denitrifiers, and heterotrophic anoxic denitrifiers.
在膜生物膜反应器（MBfR）中进行的好氧甲烷氧化耦合反硝化（AME-D）为同时减少甲烷（CH）排放和去除废水中的硝酸盐提供了很大的前景。然而，关于氧分压如何影响反向扩散生物膜的发育和特性、其空间分层剖面以及生物膜微生物的协同相互作用，仍缺乏系统的实验研究。在本研究中，我们首次将光学相干断层扫描（OCT）与共聚焦激光扫描显微镜（CLSM）相结合，对MBfR中反向扩散生物膜的发育进行原位表征。结果表明，MBfR上的氧分压能够控制生物膜的厚度和空间分层，进而管理功能微生物的分布。在优化的5.5 psig（氧含量25%）氧分压下，AME-D过程中被操纵的反向扩散生物膜获得了最高的反硝化效率，这主要是因为该生物膜在好氧层和缺氧层之间具有适当的动态平衡，其中好氧层实现了合适的氧梯度和充足的好氧甲烷氧化菌以促进甲烷氧化，同时保持完全的氧消耗和可利用的有机源以避免限制缺氧层中的反硝化活性。通过宏基因组分析和荧光原位杂交（FISH）染色，成功证明了反向扩散生物膜内功能微生物的空间分布，并且发现一种典型的好氧反硝化菌红环菌科在好氧层中存活并逐渐富集，在好氧反硝化中起关键作用。这种原位生物膜可视化和表征首次直接证明了反向扩散生物膜中AME-D反硝化的协同途径，该途径涉及好氧甲烷氧化菌、异养好氧反硝化菌和异养缺氧反硝化菌。
