Root Uptake And Accumulation Of Heavy Metals And Rhizosphere Bacteria Community Of Hyperaccumulator Sedum Alfredii Hance

Heavy metal pollution is a widespread and important environmental concern. Migration and transformation of heavy metals in soil-plant system are associated with root uptake and the rhizosphere environment. Understanding the characteristics of metal uptake and accumulation by roots of hypcraccumulator, as well as the role of rhizosphere microbe involved in metal hyperaccumulation, will provide insight into the mechanisms of metal tolerance and accumulation and help to improve the practical application of hyperaccumulators.Sedum alfredii Hance is a Zn/Cd hyperaccumulator native to Pb/Zn rich regions of China.This species also exhibits strong ability to both tolerate and accumulate considerable amounts of Pb. The aim of this study was to obtain fundamental information and important role of its root in metal tolerate and accumulation, by investigation of Cd/Pb localization and speciation in roots of two ecotypes of S. alfredii using synchrotron radiation techonology. Meanwhile,16S rRNA gene sequencing was used to investigate the rhizobacteria community structure and explore key taxa associated with plant remediation in rhizosphere of S. alfredii planted in Cd or Pb contaminated soils as well as Cd-Pb cocontaminated soil. The main results include:1. Synchrotron radiation X-ray fluorescence (XRF) and X-ray absorption spectroscopy (XAS)analysis were used to compare the in vivo Cd distribution and speciation in root tissue-level and cell-level of hyperaccumulating ecotype (HE) and non-hyperaccumulating ecotype (NHE)of S. alfredii. The results showed that, Cd was mainly distribulated in the root meristem zone and the stele area of HE. For NHE, Cd was mainly localized on meristem zone with little Cd in stele, which indicated that HE has the ability to transport more Cd to its shoot through the xylem. In addition, large amont of Cd was distributed in the lateral root (including lateral root tip and lateral root primordia, etc.) and the stele around lateral root, which indicated that more Cd would be absorbed through the lateral root into the root xylem and then be transported and accumulated in shoot. Cadmium can be more quickly and efficiently absorbed into the roots through the lateral roots. In S. alfredii, the dominant chemical forms of Cd in root tissues were Cd2+, Cd-citrate, Cd-cell wall and Cd-GSH compounds. The proportions of Cd2+ and Cd-citrate in HE root were higher than that in NHE.2. The characteristics of in vivo Pb distribution and speciation in roots of accumulating ecotype (AE) and non-accumulating ecotype (NAE) of S. alfredii were investigated by XRF and XAS analysis. Preferential Pb accumulation in meristem zone and stele area was observed in both ecotypes. Root distribution patterns of P and Cl were similar with Pb.Although Pb distribution in meristem zone of AE root was much lower than that of NAE, its concentration in stele of AE root was much higher than that of NAE. It suggested that Pb was easier to load into the xylem of AE root and then translocated to the shoot, while it was remained in the root apex of NAE. In addition, more Pb was uptaken by lateral roots of AE and directly transport into the stele. The dominant chemical form of Pb in roots of both AE and NAE was similar to previously defined Pb-cell wall compounds.3. 16S rRNA gene amplicon sequencing was used to investigate the rhizospheric bacterial communities of HE and NHE S. alfredii planted in Cd contaminated soil. Both planting of two ecotypes of S. alfredii and elevated Cd levels significantly decreased bacterial alpha-diversity and altered bacterial community structure in soils. The HE rhizosphere harbored a unique bacterial community differing from those in its bulk soil and NHE counterparts.Several key taxa from Actinobacteria, Bacteroidetes, and TM7 were especially abundant in HE rhizospheres under high Cd stress. The Actinobacterial genus Streptomyces was responsible for the majority of the divergence of bacterial community composition between the HE rhizosphere and other soil samples. In the HE rhizosphere, the abundance of Streptomyces was 3.31-16.45 fold higher than that in other samples under high Cd stress.These results suggested that both the presence of the hyperaccumulator S. alfredii and Cd-exposure select for a specialized rhizosphere bacterial community during phytoextraction of Cd-contaminated soils, and that key taxa, such as the species (OTU8159) affiliated to genus Streptomyces , may play an important role in metal hyperaccumulation.4. Rhizosphere bacteria community structures of two ecotypes of S. alfredii in Pb contaminated soil were investigated using 16S rRNA gene amplicon sequencing. Similar with results obtain from Cd contaminated soil, both planting of two ecotypes and elevated Pb levels (especially moderate-Pb level) significantly decreased bacterial alpha-diversity and altered bacterial community structure in soils. The rhizobacteria community structure composition of AE was significantly different with bulk soils or NAE rhizosphere. Several key taxa from phyla Alphaproteobacteria (Asticcacaulis and Dongia), Betaproteobacteria{Dechloromonas and Rhizobacter), and Bacteroidetes (Flavobacterium) were especially abundant in AE rhizosphere, while phyla Firmicutes (Clostridium sensu stricto Paenibacillus), Deltaproteobacteria (Geobacter) and Gammaproteobacteria (Pseudomonas)were more abundant in NAE rhizosphere. The number of the specific OTUs in the rhizosphere of two ecotypes were decreased with the increase of Pb levels.5. The rhizosphere characteristics of S. alfredii were investigated 6 months after planting in weathered field soils contaminated with Cd, Zn, Pb, and Cu. In contrast with NHE, HE S.alfredii was more tolerant to the metals and accumulated higher levels of Cd and Zn. The HE was characterized by a unique rhizosphere including extensive root systems, reduced soil pH,higher metal bioavailability, and increased rhizomicrobial activity. The bioavailability of metals was significantly correlated with the HE's unique bacterial communities (P<0.005).The HE harbored abundant Streptomyces (9.43%, family Streptomycetaceae), Kribbella(1.08%, family Nocardioidaceae), and an unclassified genus (1.09%, family Nocardioidaceae)in its rhizosphere, a composition which differed from that of the NHE. PICRUSt analysis predicted high relative abundances of imputed functional profiles in the HE rhizosphere related to membrane transport and amino acid metabolism. This study revealed the rhizosphere characteristics, particularly the unique bacterial rhizobiome of a hyperaccumulator, which may provide a new approach to facilitating heavy metal phytoextraction.