Data_Sheet_1_Genetic Analysis of Citrobacter sp.86 Reveals Involvement of Corrinoids in Chlordecone and Lindane Biotransformations.docx

Chlordecone (Kepone®) and γ-hexachlorocyclohexane (γ-HCH or lindane) have been used for decades in the French West Indies (FWI) resulting in long-term soil and water pollution. In a previous work, we have identified a new Citrobacter species (sp.86) that is able to transform chlordecone into numerous products under anaerobic conditions. No homologs to known reductive dehalogenases or other candidate genes were found in the genome sequence of Citrobacter sp.86. However, a complete anaerobic pathway for cobalamin biosynthesis was identified. In this study, we investigated whether cobalamin or intermediates of cobalamin biosynthesis was required for chlordecone microbiological transformation. For this purpose, we constructed a set of four Citrobacter sp.86 mutant strains defective in several genes belonging to the anaerobic cobalamin biosynthesis pathway. We monitored chlordecone and its transformation products (TPs) during long-term incubation in liquid cultures under anaerobic conditions. Chlordecone TPs were detected in the case of cobalamin-producing Citrobacter sp.86 wild-type strain but also in the case of mutants able to produce corrinoids devoid of lower ligand. In contrast, mutants unable to insert the cobalt atom in precorrin-2 did not induce any transformation of chlordecone. In addition, it was found that lindane, previously shown to be anaerobically transformed by Citrobacter freundii without evidence of a mechanism, was also degraded in the presence of the wild-type strain of Citrobacter sp.86. The lindane degradation abilities of the various Citrobacter sp.86 mutant strains paralleled chlordecone transformation. The present study shows the involvement of cobalt-containing corrinoids in the microbial degradation of chlorinated compounds with different chemical structures. Their increased production in contaminated environments could accelerate the decontamination processes.

十氯酮（Chlordecone，Kepone®）与γ-六氯环己烷（γ-hexachlorocyclohexane，简称γ-HCH或林丹）已在法属西印度群岛（French West Indies, FWI）使用数十年，造成了长期的土壤与水体污染。前期研究中，我们鉴定出一株新型柠檬酸杆菌菌株（Citrobacter sp.86），其可在厌氧条件下将十氯酮转化为多种产物。在Citrobacter sp.86的基因组序列中，未发现已知还原性脱卤酶（reductive dehalogenases）或其他候选基因的同源序列，但完整存在钴胺素（cobalamin）厌氧生物合成途径。本研究旨在探究钴胺素或其生物合成中间体是否为十氯酮微生物转化所必需。为此，我们构建了4株Citrobacter sp.86突变菌株，分别缺失厌氧钴胺素生物合成途径中的多个关键基因。我们在厌氧液体培养基中开展长期孵育实验，监测十氯酮及其转化产物（transformation products, TPs）的动态变化。结果显示，能够产生钴胺素的Citrobacter sp.86野生型菌株中可检测到十氯酮转化产物；仅能合成缺少下部配体的钴啉化合物（corrinoids）的突变菌株，同样可实现该转化。与之相反，无法在预咕啉-2（precorrin-2）中插入钴原子的突变菌株，则未表现出任何十氯酮转化活性。此外，我们发现此前被报道可由弗氏柠檬酸杆菌（Citrobacter freundii）厌氧转化但机制未明的林丹，在Citrobacter sp.86野生型菌株存在时同样可被降解。各Citrobacter sp.86突变菌株的林丹降解能力与其十氯酮转化能力呈现一致趋势。本研究证实，含钴的钴啉化合物参与了不同化学结构氯化有机物的微生物降解过程；在污染环境中增强这类化合物的生成，或可加速污染修复进程。