School of Environmental Sciences, University of Guelph, Guelph, Canada.
G360 Institute for Groundwater Research, University of Guelph, Guelph, Canada.
Int J Phytoremediation. 2021;23(8):846-856. doi: 10.1080/15226514.2020.1860901. Epub 2021 Jan 4.
Improved knowledge of the ecology of contaminant-degrading organisms is paramount for effective assessment and remediation of aromatic hydrocarbon-impacted sites. DNA stable isotope probing was used herein to identify autochthonous degraders in rhizosphere soil from a hybrid poplar phytoremediation system incubated under semi-field-simulated conditions. High-throughput sequencing of bacterial 16S and fungal internal transcribed spacer (ITS) genes in metagenomic samples separated according to nucleic acid buoyant density was used to identify putative toluene degraders. Degrader bacteria were found mainly within the Actinobacteria and Proteobacteria phyla and classified predominantly as , , , , , , and organisms. and fungi were also found to assimilate toluene, while several strains of the fungal poplar endophyte were indirectly implicated as potential degraders. Finally, PICRUSt2 predictive taxonomic functional modeling of 16S genes was performed to validate successful isolation of stable isotope-labeled DNA in density-resolved samples. Four unique sequences, classified within the , , or families, or within the Sphingobacteriales order were absent from PICRUSt2-generated models and represent potentially novel putative toluene-degrading species. This study illustrates the power of combining stable isotope amendment with advanced metagenomic and bioinformatic techniques to link biodegradation activity with unisolated microorganisms. This study used emerging molecular biological techniques to identify known and new organisms implicated in aromatic hydrocarbon biodegradation from a field-scale phytoremediation system, including organisms with phyto-specific relevance and having potential for downstream applications (amendment or monitoring) in future and existing systems. Additional novelty in this study comes from the use of taxonomic functional modeling approaches for validation of stable isotope probing techniques. This study provides a basis for expanding existing reference databases of known aromatic hydrocarbon degraders from field-applicable sources and offers technological improvements for future site assessment and management purposes.
提高对污染物降解生物的生态学认识，对于有效评估和修复芳烃污染场地至关重要。本研究采用 DNA 稳定同位素示踪法，在半现场模拟条件下，从杂交杨植物修复系统的根际土壤中鉴定出土著降解菌。根据核酸浮力密度分离的宏基因组样本中细菌 16S 和真菌内部转录间隔区（ITS）基因的高通量测序，用于鉴定可能的甲苯降解菌。降解菌主要存在于放线菌门和变形菌门，主要分类为 、 、 、 、 、 和 生物。 和 真菌也被发现能够同化甲苯，而几种杨树内生真菌 的菌株则被间接牵连为潜在的降解菌。最后，通过对 16S 基因进行 PICRUSt2 预测分类功能建模，验证了在密度分辨样本中成功分离稳定同位素标记 DNA。从 PICRUSt2 生成的模型中缺失了四个独特的序列，这些序列分类在 、 或 科或在鞘氨醇杆菌目中，代表可能是新型潜在的甲苯降解物种。本研究结合稳定同位素添加与先进的宏基因组和生物信息学技术，将生物降解活性与未分离的微生物联系起来，说明了其强大功能。本研究利用新兴的分子生物学技术，从现场规模的植物修复系统中鉴定出与芳烃生物降解有关的已知和新的生物，包括与植物特异性相关且具有未来和现有系统中（如添加或监测）下游应用潜力的生物。本研究的另一个新颖之处在于，采用分类功能建模方法验证稳定同位素示踪技术。本研究为从现场应用来源扩展已知芳烃降解菌的现有参考数据库提供了基础，并为未来的场地评估和管理目的提供了技术改进。
