Department of Soil Centre Science Soil Research

Gut passage of arsenic contaminated soil through the earthworm Lumbricus terrestris increases arsenic mobility Tom Sizmur | Michael Watts | John Charnock | Barbara Palumbo-Roe | Mark E. Hodson

Introduction In order to properly assess the risks that arsenic (As) poses to the environment a greater understanding of how soil biota influence the fate of As in soil is required. The bioavailability of As in soil is dependent on its mobility and speciation but little is known about how this is affected by the passage of soil through the earthworm gut. Earthworm casts are often produced on the surface of the soil and there is therefore potential for As to be leached out of the casts into water courses during periods of rainfall and overland flow. We carried out three experiments to determine the impact of passage through the gut of a UK native earthworm, Lumbricus terrestris L. on the mobility and speciation of As in soil (1130 mg kg-1 As) collected at Devon Great Consols, an abandoned Cu and As mine.

Mobility of As over time in earthworm casts (Experiment 1) 25

As / mg kg-1

L. terrestris were incubated in bags containing 500g of soil (five specimens per bag) for 7 days alongside earthworm-free bags containing 50g of soil. After incubation the earthworms were removed and their guts voided on filter paper. The filter papers were then sealed, moist in petri dishes for 0, 1, 7, 14, 28 or 56 days to simulate moist casts ageing in the soil environment. Bulk earthworminhabited soil and earthworm-free soil was treated likewise. After the ageing, casts and soils were dried. Air-dried soil/cast samples were extracted with ultra pure water. Arsenic concentration was determined in the supernatant.

Casts

20

Bulk

15

Control

10 5

There was a significantly (p<0.01) greater concentration of water soluble As in the casts compared to the bulk or control soil at all time points (Figure 1).

0 0

1

7

14

Days of ageing

28

56

Figure 1: Water soluble As in L. terrestris casts, bulk earthworm-inhabited soil and earthworm-free control soil after 7 days earthworm incubation and then ageing for 0, 1, 7, 14, 28 or 56 days (n = 5, error bars = standard errors).

L. terrestris were incubated in columns containing 900g of soil (two specimens per column) for 112 days alongside four earthworm-free columns of soil. After incubation the earthworms were removed from the soil and their guts voided on filter paper for 48 hours. Casts were collected and frozen. Station 16.5 at SRS Daresbury Laboratory, Warrington, UK was used to obtain As K-edge spectra of earthworm casts to compare with bulk earthworm inhabited soil and earthworm-free soil. The As in the solid phase of the soil was present as As(V) in the casts, bulk earthworm inhabited and control soils (Figure 2). There was no indication that the earthworms changed the bulk speciation of the As in the casts after 112 days of incubation.

Normalised absorbtion

Speciation of total As in earthworm casts (Experiment 2)

11850

As:glutathione, 1:3 Sodium arsenite Sodium arsenate Casts

Bulk Control

11864

11879

11894

11915

11947

Figure 2: XANES spectra of As K-edge of L. terrestris casts, bulk earthworm-inhabited soil and earthworm-free control soil compared with spectra from known standards.

11986

Energy / eV

Speciation of mobile As in earthworm casts (Experiment 3)

There was a significant (p<0.01) increase in the concentration of water soluble As(V) in the casts compared to bulk earthworm-inhabited and control soil (Figure 3).

10

As / mg kg-1

L. terrestris were incubated in boxes containing 1kg of soil (ten specimens per box) for 7 days alongside five earthworm-free boxes of soil. After incubation the earthworms were removed from the soil and their guts voided on filter paper for 48 hours. Casts were collected and dried along with bulk earthworm-inhabited soil and earthworm-free soil. Air-dried soil/cast samples were extracted with ultra pure water. Arsenate (AsV) and arsenite As(III) species of As were then quantitatively determined in the supernatants within 24 hours of extraction by HPLC-ICP-MS.

12

8

AsV

6

As III

4 2

Figure 3: Speciation and concentration of water soluble As in L. terrestris casts, bulk earthworminhabited soil and earthworm-free control soil (n = 5, error bars = standard errors)

0

Casts

Bulk

Control

Discussion and Conclusions

Acknowledgements • Tom Sizmur is supported by a BBSRC studentship, with CASE support from British Geological Survey –University Funding Initiative Contact information • Soil Research Centre, University of Reading, Whiteknights, RG6 6DW • Email: [email protected]

The earthworm gut is a pH-neutral environment containing enzymes capable of degrading organic matter. The passage of soil through the gut elevated the pH and degraded organic matter (data not shown). Passage through the earthworm gut did not increase the concentration of soluble As(III), but instead caused desorption of As(V) and the release of organically bound As(V) into solution resulting in no overall change in As redox state in the soil but an increase in As mobility.