Table_3_Exposure to Crude Oil and Chemical Dispersant May Impact Marine Microbial Biofilm Composition and Steel Corrosion.XLSX

The release of hydrocarbons and chemical dispersant in marine environments may disrupt benthic ecosystems, including artificial reefs, formed by historic steel shipwrecks, and their associated organisms. Experiments were performed to determine the impacts of crude oil, dispersed crude oil, and dispersant on the community structure and function of microorganisms in seawater (SW) and biofilms formed on carbon steel, a common ship hull construction material. Steel corrosion was also monitored to illustrate how oil spills may impact preservation of steel shipwrecks. Microcosms were filled with seawater (SW) and incubated at 4°C. Carbon steel disks (CSDs) were placed in each tank, and tanks were amended with crude oil and/or dispersant or no treatment. SW and CSD biofilms were sampled biweekly for genetic analysis using Illumina sequencing of 16S ribosomal RNA gene amplicons. Predicted and sequenced bacterial metagenomes were analyzed to examine impacts of oil and dispersant on metabolic function. Gammaproteobacteria, Alphaproteobacteria, and Flavobacteriia dominated SW and biofilms. Bacterial community structure differed significantly between treatments for SW and biofilms. OTUs affiliated with known (Pseudomonas) and potential (Marinomonas) hydrocarbon-degraders were roughly twice as abundant in biofilms treated with oil and dispersed oil, and steel corrosion of CSDs in these treatments was higher compared to control and dispersant treatments. OTUs affiliated with the Rhodobacteraceae family (biofilm formers and potential oil degraders) were less abundant in the dispersant treatment compared to other treatments in biofilm and SW samples, but OTUs affiliated with the Pseudoalteromonas genus (biofilm formers and proposed hydrocarbon degraders) were more abundant in dispersant-treated biofilms. Overall, functional gene analyses revealed a decrease in genes (predicted using PICRUSt and observed in sequenced metagenomes) associated with hydrocarbon degradation in dispersant-treated biofilms. This study indicates that exposure to oil and dispersant could disrupt the composition and metabolic function of biofilms colonizing metal hulls, as well as corrosion processes, potentially compromising shipwrecks as ecological and historical resources.

海洋环境中烃类与化学分散剂的泄漏，可能会破坏由历史性钢制沉船形成的人工鱼礁（artificial reefs）及其相关生物组成的底栖生态系统（benthic ecosystems）。本研究开展实验，以明确原油、经分散剂乳化的原油与单一化学分散剂，对海水（seawater, SW）以及作为常见船体建造材料的碳钢表面形成的生物膜中微生物群落结构与功能的影响。同时监测钢材腐蚀情况，以阐明石油泄漏可能如何影响钢制沉船的保存状态。实验微宇宙体系（microcosms）中注入海水（SW），并置于4℃条件下培养。每个培养罐中放置碳钢圆盘（carbon steel disks, CSDs），随后分别施加原油处理、原油-分散剂复合处理、单一分散剂处理，或不做任何处理（对照组）。每隔两周采集海水与碳钢圆盘表面生物膜样本，通过16S核糖体RNA基因扩增子（16S ribosomal RNA gene amplicons）的Illumina测序技术开展遗传分析。对预测得到与实测获得的细菌宏基因组（bacterial metagenomes）进行分析，以检验油类与分散剂对微生物代谢功能的影响。γ-变形菌纲（Gammaproteobacteria）、α-变形菌纲（Alphaproteobacteria）与黄杆菌纲（Flavobacteriia）在海水与生物膜样本中占据优势类群。不同处理组的海水与生物膜样本的细菌群落结构均存在显著差异。隶属于已知烃类降解菌假单胞菌属（Pseudomonas）与潜在烃类降解菌海单胞菌属（Marinomonas）的操作分类单元（Operational Taxonomic Units, OTUs），在原油处理组与经分散剂乳化原油处理组的生物膜样本中的丰度约为对照组的两倍；且该两组处理中碳钢圆盘的腐蚀程度显著高于对照组与单一分散剂处理组。在生物膜与海水样本中，隶属于红杆菌科（Rhodobacteraceae，兼具生物膜形成能力与潜在原油降解能力）的操作分类单元在单一分散剂处理组中的丰度均低于其他处理组；但隶属于假交替单胞菌属（Pseudoalteromonas，兼具生物膜形成能力与被提议的烃类降解能力）的操作分类单元在经分散剂处理的生物膜样本中丰度更高。总体而言，功能基因分析结果显示，经分散剂处理的生物膜中，与烃类降解相关的基因（通过PICRUSt功能预测工具预测且在实测宏基因组中得到验证）的丰度出现下降。本研究表明，暴露于油类与化学分散剂环境中，可能会破坏金属船体表面定殖生物膜的群落组成与代谢功能，同时影响腐蚀过程，进而可能损害作为生态与历史资源的钢制沉船。