View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Current Biology 22, 237–241, February 7, 2012 ª2012 Elsevier Ltd All rights reserved DOI 10.1016/j.cub.2011.12.017 Report Lead Transformation to Pyromorphite by Fungi Young Joon Rhee,1 Stephen Hillier,2 Depositions of these secondary minerals were also detected and Geoffrey Michael Gadd1,* inside fractured hyphae of P. javanicus (Figure 1F) and on 1Division of Molecular Microbiology, College of Life Sciences, the lead shot surface (Figures 1G and 1H). University of Dundee, Dundee DD1 5EH, Scotland, UK 2The James Hutton Institute, Craigiebuckler, Energy-Dispersive X-Ray Analysis and X-Ray Mapping Aberdeen AB15 8QH, Scotland, UK Energy-dispersive X-ray analysis (EDXA) revealed differences in elemental composition between the secondary lead minerals produced in biotic and abiotic treatments. In the abiotic Summary control, lead, carbon, and oxygen appeared as the main elements (Figure 2A), whereas phosphorus and chlorine Lead (Pb) is a serious environmental pollutant in all its together with lead, carbon, and oxygen appeared only in chemical forms [1]. Attempts have been made to immobilize the secondary minerals produced in the presence of lead in soil as the mineral pyromorphite using phosphate P. javanicus and M. anisopliae (Figures 2B and 2C). amendments (e.g., rock phosphate, phosphoric acid, and Some common lead minerals that contain phosphorus are apatite [2–5]), although our work has demonstrated that pyromorphite (Pb5[PO4]3X [X = F, Cl or OH]), plumbogummite soil fungi are able to transform pyromorphite into lead (PbAl3[PO4]2[OH]5), and tsumebite (Pb2Cu[OH]3[PO4]). oxalate [6, 7]. Lead metal, an important structural and indus- Plumbogummite and tsumebite contain other metals (Al, Cu), trial material, is subject to weathering, and soil contami- which can be easily detected in EDXA. The absence of nation also occurs through hunting and shooting [8, 9]. aluminum and copper in the EDXA spectra suggest that Although fungi are increasingly appreciated as geologic the mineral formed on fungal-exposed lead surfaces was agents [10–12], there is a distinct lack of knowledge about chloropyromorphite. X-ray mapping was able to confirm not their involvement in lead geochemistry. We examined the only the presence of phosphorus and chlorine but also the influence of fungal activity on lead metal and discovered exact location within the secondary minerals (Figure 3). Lead that metallic lead can be transformed into chloropyromor- (Figure 3C), phosphorus (Figure 3D), chlorine (Figure 3E), and phite, the most stable lead mineral that exists. This is of oxygen (Figure 3F) were all identically localized within the geochemical significance, not only regarding lead fate and secondary minerals. cycling in the environment but also in relation to the phos- phate cycle and linked with microbial transformations of X-Ray Powder Diffraction Analysis inorganic and organic phosphorus. This paper provides X-ray powder diffraction analysis (XRPD) revealed that the the first report of mycogenic chloropyromorphite formation secondary minerals on the surfaces of lead shot from abiotic from metallic lead and highlights the significance of this control plates included minium (Pb3O4) and hydrocerussite phenomenon as a biotic component of lead biogeochem- (Pb3[CO3]2[OH]2) together with a minor component of litharge istry, with additional consequences for microbial survival (PbO) after 2 months incubation (Figure 4A); additionally, in lead-contaminated environments and bioremedial treat- cerussite (PbCO3) is present after 3 months incubation (Fig- ments for Pb-contaminated land. ure 4B). However, lead shot taken from plates inoculated with P. javanicus showed evidence of chloropyromorphite Results (Pb5[PO4]3Cl) formation after incubation for 1 month, the mineral assemblage also including cerussite, hydrocerussite, Formation of Mycogenic Lead Minerals minor litharge, and minium (Figure 4C). After 3 months incuba- Lead shot was incubated in the absence and presence of tion, chloropyromorphite was much more prominent in the test fungi isolated from a former lead mining area near assemblage (Figure 4D). M. anisopliae showed cerussite Wanlockhead, Scotland, UK. Formation of corrosion products with minor litharge and hydrocerussite after incubation for around the lead shot was observed in all biotic treatments. 1 month (Figure 4E) but along with minor chloropyromorphite, Microscopic examination showed widespread corrosion on whereas after 3 months incubation (Figure 4F), chloropyromor- lead shot surfaces as well as the presence of secondary phite was considerably more abundant. Metallic lead from mineral formations of various shapes together with fungal the lead shot substrate is also apparent in most traces. Note hyphae (Figure 1). Control lead shot, incubated in the absence that the peaks for some phases (minerals) are occasionally of the fungi, also showed some mineral deposition on the displaced from their ideal positions, and this is attributed to lead surface resulting from abiotic corrosion (Figures 1A and the nonideal geometry of the sample, i.e., spherical rather 1B). The secondary minerals formed on lead shot incubated than flat. Additionally, the peaks for chloropyromorphite are with Metarhizium anisopliae showed several different mor- notably broader than the other phases, indicating a very fine phologies, the most distinctive shapes observed being hexag- size or a cryptocrystalline nature of the phase. onal columns and small spheroids (Figures 1C and 1D). In Paecilomyces javanicus-treated lead shot, small spheroids Discussion were dominant in most of the samples (Figures 1E–1H). Most metallic lead contamination of the environment is caused by the use of lead shot in firearms [13, 14] and in fishing *Correspondence: [email protected] weights or jigs, although non-lead substitutes are now widely Current Biology Vol 22 No 3 238 Figure 1. Scanning Electron Microscopy Images of Lead Shot Surfaces Incubated in the Absence or Presence of Experimental Organisms Images were obtained using a Philips XL30 environ- mental scanning electron microscope (ESEM) operating at an accelerating voltage of 15 kV. (A and B) Surface of control lead shot showing some deterioration caused by abiotic effects (scale bars repre- sent 500 mm and 10 mm, respectively). (C and D) Secondary lead mineral and pyromorphite formation on lead shot surface after incubation with M. anisopliae (scale bars represent 5 mm and 2 mm, respectively). (E) Deposits of pyromorphite on the surface of lead shot incubated with P. javanicus. This image was taken after removing the fungal hyphae, which were covering the lead shot (scale bar represents 10 mm). Inset is a higher magnification image of the area indicated by the square (scale bar represents 1 mm). (F) Pyromorphite formation within broken hypha of P. javanicus (scale bar represents 5 mm). (G) Pyromorphite deposition on the surface of lead shot incubated with P. javanicus (scale bar represents 10 mm). The arrow indicates the area of sample shown in (H) at higher magnification. (H) Pyromorphite deposition on the surface of lead shot incubated with P. javanicus (detail of indicated area in G; scale bar represents 2 mm). All samples were incu- bated for 3 months at 25C. Typical images are shown from many similar examples. oxides are exposed to various pH conditions [1]. Despite lead being bound strongly to soil components such as soil colloids and humic substances, attempts have still been made to immobilize lead in situ by the formation of stable lead minerals that can withstand weath- ering processes or biological attack. In lead-contaminated soil, the lead con- taining mineral pyromorphite can form by interaction between mobile Pb species, phos- phate, and chloride. Chloropyromorphite (Pb5[PO4]3Cl), with a solubility product (Ksp) of 10284.4, is the most stable lead mineral in the Earth’s crust [16], and the various forms of pyromorphite (Pb5[PO4]3X [X = F, Cl and OH]) formation have been widely pro- posed as a remediation mechanism for the used in freshwater angling [15]. In one study, lead shot from sequestration and immobilization of contaminant Pb in soil ammunition was found to accumulate in a shooting range to [3, 16, 18, 19]. Soil amendment remediation techniques that amounts of the order of tens of thousands of kilograms per apply apatite or bone meal to encourage the formation of hectare per year [8]. Apart from such direct introduction of pyromorphite have received wide attention [20].The addition metallic lead into the environment, soil can also be contami- of phosphoric acid [2], apatite [3], and rock phosphate [4, 5] nated with lead from other sources such as industrial wastes has been used to immobilize lead in this way. All of these (i.e., paints, printing inks, lead water pipes, lead glazed procedures require a step that includes acidification of the pottery, battery casings, etc.), as well as residues from leaded soil. This allows the dissolution of the Pb and P source, which petrol [16]. Lead in all its forms is regarded as an important and increases the efficacy of pyromorphite formation but also potentially dangerous pollutant because of toxicological promotes leaching of other metals, which might cause addi- effects toward humans [1, 9, 17], and as with other toxic tional contamination [19]. In addition, some phosphate metals, various remedial techniques have been investigated amendment remediation techniques