Effect And Mechanism Of The Drilosphere On Soil DDT Degradation Pathway

As a typical synthetic organochlorine compound pesticide, 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane(DDT) was widely used in the 20 th century. Due to its persistent stability, high bioaccumulation, strong toxicity and long-range transport, the degradation and transformation of DDT is not only the focus but also an obstacle of research. Moreover, the fresh inputs of DDT from dicofol and antifouling paints, soil DDT contamination continues to be a serious environmental issue. Based on our previous early research that Amynthas robustus Perrier(A. robustus) could significantly enhance the degradation of soil DDT through unclear mechanism, we explored the effect and mechanism of drilosphere on soil DDT degradation by establishing soil column and microcosm incubation experiments. Molecular technologies, including terminal restriction fragment length polymorphism(T-RFLP), stable isotope probing(SIP) and high-thoughout sequencing(HTS), were applied to study the influence and mechanism of A. robustus and drilosphere on DDT degradation pathway. This study will provide theory support in developing new technologies on bioremediation of DDT contaminated soil. The main results and conclusions are as follows:(1) Via the soil column incubation with or without A. robustus in sterile or natural soils, we found that both indigenous microorganisms and A. robustus palys important roles in soil DDT degradation. The amount of DDT residual in sterile soil was steady within the first 6 weeks(higher than 95% of initial quantity), while exhibiting considerable decrease in natural soil during the whole incubation period. Moreover, both sterile and natural soil with A. robustus showed significant reduction, with only 75%, 50%, and 25% left at the 4th, 8th, and 14 th week, respectively. These results indicate that biotic degradation is the prime approach to remove soil DDT and A. robustus has direct effect on soil DDT degradaton.(2) By comparing the concentrations of DDT and its metabolites(DDTs) in CK soil(CK), bulk soil(Bulk), burrow lining(Burrow), cast(Cast), and gut content(Gut), we found that drilosphere formed by A. robustus is the “hotspot” in DDT degradation. A. robustus would not only form burrow lining to facilitate the DDT dehydrochlorination into DDE, but more importantly, utilize a different degradation pathway by reductively dechlorinating DDT/DDE to DDD/DDMU through gut digestion.(3) Moreover, by analyzing the chemical and biological properties of non-drilosphere and drilosphere matrixes, we further discovered that A. robustus would improve soil quality to accelerate DDT degradation. The p H of drilosphere matrixes increased along with incubation; the final p H of Cast reached 5.85. Concentrations of organic carbon, water soluble organic carbon, and hot-water soluble organic carbon in drilosphere matrixes were significantly increased compared with non-drilosphere matrixes. In addition, humin, microbial biomass carbon and nitrogen also increased with incubation time. Further canonical correspondence analysis(CCA) of soil properties and DDTs revealed that the higher p H and metabolism activity of organic substrate in drilosphere were the most important factors in driving DDTs microbial degradation.(4) In order to elucidate the microbial mechanism of the DDTs degradation enhancement by A. robustus, non-drilosphere and drilospher matrixes were applied in microcosm incubation. The result showed that the initial species and relative abundance of bacteria community are quite different: CK and Bulk matrixes have Chloroflexi as the dominant species, while Proteobacteria is the main species in Burrow, Cast and Gut. However, all matrixes were tended to have unclassified Sporolactobacillaceae as the predominant species as the incubation went on. With in contrast to CK, Bulk, Burrow, and Cast, the microflora in A. robustus gut showed remarkable increment in the Shannon diversity index, Pielou’s evenness index, Species richness index, and Simpson index during incubation. The CCA analysis indicates that the gut bacteria in A. robustus could promote DDTs degradation, while Simpson and microbial community composition are the main factors to affect DDT degradation in soil.(5) Eleven functional bacteria were identified by using DNA-SIP combined HTS participating in DDT ring-opening degradation, and mainly from Actinobacteria, Acidobacteria, Firmicutes, and Proteobacteria. The dominant degraders of 13C-DDT from each matrix are different: CK have Streptomyces,Bacillus, and unclassified Bacillaceae; Bulk have Dermacoccus, unclassified Bacillales, and other Bacillaceae; Burrow employs Dermacoccus,Brevibacterium, and unclassified Bacillales; Cast possess Dermacoccus,unclassified Xanthomonadaceae, and unclassified Bacillales; and Streptacidiphilus and Candidatus Solibacter in Gut. All these data indicate that the functional bacteria in non-drilosphere and drilosphere matrixes are quite different, and the dominant DDTs ring cleavage degraders are from Actinobacteria. A. robustus stimulates the functional bacteria Dermacoccus to improve the degradation of soil DDTs.Therefore, A. robustus significantly improved soil p H and metabolism activity of organic carbon of bulk soil by forming the drilosphere, changed the microbial community structure, and increased the relative abundances of functional bacteria, thus improving DDT degradation rate and affecting the degradation pathway of DDT degradation in soil.