1. HEAVY METALS

2. Bioremediation of mercury contaminated soil

3. Forms of mercury:
1. Free Hg(II),
2. Particle-bound Hg(II), and
3. Hg(0).
Hg(II) and Hgp have atomospheric residence times on the order of weeks
Hg(0) can be retained approx. 0.5 to 2 years in the global atmospheric pool.

4. Toxicity
Hg(0) us readily absorbed through the lungs and passively diffuses through other biological tissues, but has a relatively short residence time.
It neither accumulates nor reacts with most biochemicals.

5. Toxicity
Hg(II) is highly reactive, but does not freely diffuse through cellular membranes and is inefficiently absorbed through the intestines.
Studies have reported that 80 to 85% of ingested doses are excreted in feces with the rest being eliminated over a period of days or months through urine.
As Hg(II) is processed and filtered by the body, it is likely to cause damage to the kidneys, liver, and gastrointestinal tract.

6. Toxicity
Methylmercury is lethal at much lower doses than Hg(0) and Hg(II) because it is reactive yet highly mobile. It is efficiently absorbed from the gastrointestinal tract with an estimated uptake of 95% in humans.
It is readily transported through blood vessel walls and into tissues.

7. Why are we concerned about them?
Mercury and mercurial compounds are nonnutritive heavy metals, which are hazardous to all biological organisms.

8. Basis for bioremediation
Bacteria have developed resistance to colonize mercury-contaminated environment: an operon of mercury resistant (mer) genes encoding transporters and enzymes for biochemical dotoxification. e.g. Pseudomonas strain K-62.
Some plants demonstrate moderate level of Hg tolerance. However, studies have showed high levels of Hg-tolerance and Hg-detoxification in transgenic plants.

9. Mechanisms

10. A. The mer operon contains a series of five or more genes that collectively confer Hg resistance to bacteria.
MerA and merB encode the enzymes Hg reductase and organomercurial lyase, respectively.
MerT and merP code for Hg(II) transport proteins, used by bacteria to shuttle Hg to the cytoplasm for enzymatic Hg detoxification.
MerD and merR encode proteins that regulate gene expression of the mer operon.

11. B. Organomercurial lyase (merB) detoxifies organic Hg by catalyzing the cleavage of the C-Hg bond
C. Mercuric reductase (merA) reduces Hg(II) to volatile Hg(0).

12. Hg(II) is the predominant form in soil, usually as stable HgS
Hg(II) is the predominant form in soil, usually as stable HgS. Hg(II) may be reduced to volatile Hg(0) abiotically or by mer+ bacteria, or methylated, primarily by anaerobic bacteria.

13. Mer genes -transgenic plants may help extract Hg from polluted sites either by translocating to their shoots or evaporating as Hg(0).

14. A. wild-type Arabidopsis.
B. merA transformed Arabidopsis
The growth media was spiked with 50 mM Hg(II)

15. Tobacco plants about 2 weeks from seeds
Tobacco plants about 2 weeks from seeds. The growth media was spiked with 50 mM Hg(II)

16. Yellow-poplar, germinated on selective medium for about 2 weeks The growth media was spiked with 50 mM Hg(II)

17. Arabidopsis seeds, about 4 wks on medium containing organomercurial salts:
a. 1mM methlmercury;
b. 1mM phenylmercuric acetate (PMA)

18. Hg(0) volatilization for wild-type and merA transformed Arabidopsis plantlet

19. Hg(0) volatilization for wild-type and merA transformed yellow-poplar plantlet

20. Hg(II) reduction in wild-type and merA-transformed tobacco plants

21. Example:

22. The magic market site in Trenton, New Jersey: A case study

23. Site description:
An old abandoned urban industrial site that has been designated as a “Superfund” site.
Typical of many of the so-called brownfield sites in NJ and across the US
Industrial uses including Gould Inc., a battery manufacturer.
Toxic lead was left around the old buildings

24. Problem
Neighborhood is left to cop with the industrial contamination left behind when the companies moved away.

25. Solution
Researchers from Rutgers Univ. devised and created a company, Phytotech, to implement its remediation.

26. Approaches
Starting with a 200 x 30-ft plot of freshly tilled earth, they sowed the tract with seeds of the Indian mustard plant that grows quickly and has the ability to accumulate lead from the soil in which it is planted.

27. A “Northwest Community Improvement Association” was established
A “Northwest Community Improvement Association” was established. From the collaboration among private industries, government agencies and local residents, the City of Trenton expanded its involvement in neighborhood remediation and successfully garnered an EPA grant for $200,000 to provide funds for the next step.

28. Mustard plants uptake and store Pb in their leaves rather than roots
Mustard plants uptake and store Pb in their leaves rather than roots. Their roots typically reach about 20 in into the ground, which is well below 6-8 in depth where lead contamination usually resides.
The plants reach maturity in about 6 weeks. The researchers were able to grow and harvest several cycles in the garden during the spring and summer of 1996.

29. Lead levels in the soil and in the plants were tested between each planting.
Later in the year, they planted pumpkins on the site and harvested pumpkins for Halloween. All these were done very publicly involving Trenton and environmental agency officials and community members.

30. Result
At the time harvesting pumpkin (Oct. 22, 1996), Lead concentrations in the soil have been reduced to established safety levels in 75% of the area they planted mustard plants earlier in the year.

31. Phytostabilization--A superfund site
location: in the Black Hills of western South Dakota
A 18-m stretch of Whitewood Creek was contaminated with arsenic and cadmium (surface and groundwater)
Cause: 130 years of gold mining activity

32. Remediation:
The university of Iowa planted 3100 hybrid poplar trees on one acre of the site to a depth of 1.6 m in old mine tailing along Whitewood Creek in April 1991.
A commercial N/P/K fertilizer was used to ensure vigorous early growth of the cuttings
Roots formed along the entire length of the cutting in the soil, and a root mass was established to prevent leachate generation and percolation to groundwater

33. Concentration mg/kg
Concentration of As, Cd, Pb, and Zn in subsuface soil at various soil depths.

34. Remediation:
As was the element of concern according to risk assessments at the site, but cadmium was monitored because of its potential for uptake and translocation to leaves, thus creating pathway of exposure at the site.
At the end of the first growing season, the trees had grown to four feet in height. There were evidence of phytoxicity of the tailings in the field and in laboratory pot studies, but vegetation was growing.

35. Remediation:
Poplar leaves in the field did not take up very much Cd or As (1.2 and 27 mg/kg dry weight).
These concentrations are below most limits (e.g. EPA 503 for biosolids) established for filed application of municipal sewage sludge or compost.
Results indicated that uptake of metals by the poplar trees was not a serious concern relative to published guidelines and native vegetation at the site.

36. There was a weak relationship between the total digestible As concentration in soils at the site and the concentration that was monitored in the stem tissue of young poplar trees.
There was a correlation between Zn and Cd concentration in leaves of poplar trees.

39. In general, relationships between total digestible metal in soil (or chelate extractable fractions) and the concentration of metals found in leaves were not statistically significant.

40. At the Whitewood Creek site, South Dakota, approximately 150 poplar trees were still living in 1995 from a total of trees planted between 1991 and 1993.
Overall survival rates were very low due to possible phytotoxicity, harsh climate, and animal browse.
The site is rather remote, and there was not one to maintain it on a daily basis.
Fencing was need to prevent animal browse (deer and cattle) that was very heavy at times.
Severe ice storm killed many of the trees.

41. Conclusion
It was difficulty to establish vegetation at the Whitewood Creek site given multiple stresses on the trees.
At a second site established in 1994 at an abandoned smelter in Dearing, Kansas, the addition of aged manure and straw allowed success in establishing a phytostabilization system with greater than 50% survival of the trees.

42. Processes influencing free metal ion activities in soil solution of a phytostabilization system

43. What would you do different if you were in charge of remediating this site?
Do you think phytostabilization is the most viable strategy for this site? Any other feasible remediation strategy ?