FEMS Microbiology Ecology, 91, 2015 doi: 10.1093/femsec/fiu007 Advance Access Publication Date: 5 December 2014 Research Article
RESEARCH ARTICLE
Adaptation of soil microbial community structure and function to chronic metal contamination at an abandoned Pb-Zn mine ´ 1,# , Fernando Blanco1 , Tim Urich2 Lur Epelde1,∗,# , Anders Lanzen and Carlos Garbisu1 1
NEIKER-Tecnalia, Department of Ecology and Natural Resources, Soil Microbial Ecology Group, c/ Berreaga 1, E-48160 Derio, Spain and 2 Department of Ecogenomics and Systems Biology, University of Vienna, Althanstrasse 14, 1090 Vienna, Austria ∗ Corresponding author: NEIKER-Tecnalia, Department of Ecology and Natural Resources, c/ Berreaga 1, E-48160 Derio, Spain. Tel: +34-94-403-43-00;
Fax: +34-94-403-43-10; E-mail: [email protected] Shared first-authorship. One Sentence Summary: Our results provide an example of microbial communities in long-term metal contaminated environments, in terms of composition, diversity and function of highly and differentially transcribed genes. Editor: Cindy Nakatsu
#
ABSTRACT Toxicity of metals released from mine tailings may cause severe damage to ecosystems. A diversity of microorganisms, however, have successfully adapted to such sites. In this study, our objective was to advance the understanding of the indigenous microbial communities of mining-impacted soils. To this end, a metatranscriptomic approach was used to study a heavily metal-contaminated site along a metal concentration gradient (up to 3220 000 and 97 000 mg kg−1 of Cd, Pb and Zn, respectively) resulting from previous mining. Metal concentration, soil pH and amount of clay were the most important factors determining the structure of soil microbial communities. Interestingly, evenness of the microbial communities, but not its richness, increased with contamination level. Taxa with high metabolic plasticity like Ktedonobacteria and Chloroflexi were found with higher relative abundance in more contaminated samples. However, several taxa belonging to the phyla Actinobacteria and Acidobacteria followed opposite trends in relation to metal pollution. Besides, functional transcripts related to transposition or transfer of genetic material and membrane transport, potentially involved in metal resistance mechanisms, had a higher expression in more contaminated samples. Our results provide an insight into microbial communities in long-term metal-contaminated environments and how they contrast to nearby sites with lower contamination. Key words: evenness; metal resistance; metatranscriptomics; mine soil
INTRODUCTION The surface of the Earth is affected by mining operations with an area of 240 000 square kilometers (Furrer et al., 2002),
constituting one of the main sources of metal pollution (Wei and Zhou 2008). Metal extraction activities also contribute to the degradation of the surrounding environment by generating
Received: 25 June 2014; Accepted: 24 October 2014 C FEMS 2014. All rights reserved. For permissions, please e-mail: [email protected]
1
2
FEMS Microbiology Ecology, 2015, Vol. 91, No. 1
vast amounts of solid waste, frequently piled up in the vicinity of mining operations (Li 2006). Toxicity of metals and metalloids released from mine tailings can be detrimental to human health (Valko, Morris and Cronin et al., 2005) and other life, in˚ cluding microorganisms (Ba˚ ath 1989). Metal toxicity can occur through (i) displacement of essential metals from their native binding sites and (ii) ligand interactions, triggering cell membrane damage, alterations of enzyme specificity, disruption of cellular functions, modifications of DNA structure, etc. (Bruins, Kapil and Oehme 2000). Microorganisms, including fungi, are known to play key roles in soil and other ecosystems (Staley et al., 1997) and can influence the composition of plants and other larger organisms (Bardgett et al., 2005). Strong selective pressure from metalcontaminated environments on exposed microorganisms can thus have serious consequences for ecosystem function. However, microorganisms have been exposed and adapted to metal contamination since early history (Bruins, Kapil and Oehme 2000; Valls and de Lorenzo 2002). This has resulted in the evolution of several metal resistance mechanisms, such as extracellular and intracellular sequestration, exclusion by permeability barriers, enzymatic detoxification, reduction in sensitivity of cellular targets, efflux pumps, etc. (Nies 2003; Hobman, Yamamoto and Oshima 2007). Such resistance mechanisms form the basis for the use of microorganisms in bioremediation approaches. In addition to readily adapted organisms, these resistance systems may be carried by plasmids or transposons and can therefore be transferred very efficiently to other community members (Top et al., 1990; Sørensen et al., 2005). Unfortunately, little is known about the direct and indirect effects of metals on the composition, richness and evenness of microbial communities, including their ability for adaptation to metal contamination. To date, limited culture-independent molecular surveys have been reported describing the microbial communities of mining-impacted soils. However, recent developments have enabled for sequencing of the composite genomes (‘metagenomics’) or transcribed genes (‘metatranscriptomics’) of entire microbial communities in situ, particularly thanks to improved sequencing technology. In this study, our objective was to take advantage of this and use a metatranscriptomic approach to study the entire microbial community present in metal-contaminated environments, as well as possible biogeochemical roles of individual groups of organisms. To this end, we chose to study a contamination gradient at an abandoned Pb-Zn mine as an example of a heavily metalcontaminated site.
Soil physicochemical and biological characterization For the soil physicochemical characterization, samples were sieved to
RESEARCH ARTICLE
Adaptation of soil microbial community structure and function to chronic metal contamination at an abandoned Pb-Zn mine ´ 1,# , Fernando Blanco1 , Tim Urich2 Lur Epelde1,∗,# , Anders Lanzen and Carlos Garbisu1 1
NEIKER-Tecnalia, Department of Ecology and Natural Resources, Soil Microbial Ecology Group, c/ Berreaga 1, E-48160 Derio, Spain and 2 Department of Ecogenomics and Systems Biology, University of Vienna, Althanstrasse 14, 1090 Vienna, Austria ∗ Corresponding author: NEIKER-Tecnalia, Department of Ecology and Natural Resources, c/ Berreaga 1, E-48160 Derio, Spain. Tel: +34-94-403-43-00;
Fax: +34-94-403-43-10; E-mail: [email protected] Shared first-authorship. One Sentence Summary: Our results provide an example of microbial communities in long-term metal contaminated environments, in terms of composition, diversity and function of highly and differentially transcribed genes. Editor: Cindy Nakatsu
#
ABSTRACT Toxicity of metals released from mine tailings may cause severe damage to ecosystems. A diversity of microorganisms, however, have successfully adapted to such sites. In this study, our objective was to advance the understanding of the indigenous microbial communities of mining-impacted soils. To this end, a metatranscriptomic approach was used to study a heavily metal-contaminated site along a metal concentration gradient (up to 3220 000 and 97 000 mg kg−1 of Cd, Pb and Zn, respectively) resulting from previous mining. Metal concentration, soil pH and amount of clay were the most important factors determining the structure of soil microbial communities. Interestingly, evenness of the microbial communities, but not its richness, increased with contamination level. Taxa with high metabolic plasticity like Ktedonobacteria and Chloroflexi were found with higher relative abundance in more contaminated samples. However, several taxa belonging to the phyla Actinobacteria and Acidobacteria followed opposite trends in relation to metal pollution. Besides, functional transcripts related to transposition or transfer of genetic material and membrane transport, potentially involved in metal resistance mechanisms, had a higher expression in more contaminated samples. Our results provide an insight into microbial communities in long-term metal-contaminated environments and how they contrast to nearby sites with lower contamination. Key words: evenness; metal resistance; metatranscriptomics; mine soil
INTRODUCTION The surface of the Earth is affected by mining operations with an area of 240 000 square kilometers (Furrer et al., 2002),
constituting one of the main sources of metal pollution (Wei and Zhou 2008). Metal extraction activities also contribute to the degradation of the surrounding environment by generating
Received: 25 June 2014; Accepted: 24 October 2014 C FEMS 2014. All rights reserved. For permissions, please e-mail: [email protected]
1
2
FEMS Microbiology Ecology, 2015, Vol. 91, No. 1
vast amounts of solid waste, frequently piled up in the vicinity of mining operations (Li 2006). Toxicity of metals and metalloids released from mine tailings can be detrimental to human health (Valko, Morris and Cronin et al., 2005) and other life, in˚ cluding microorganisms (Ba˚ ath 1989). Metal toxicity can occur through (i) displacement of essential metals from their native binding sites and (ii) ligand interactions, triggering cell membrane damage, alterations of enzyme specificity, disruption of cellular functions, modifications of DNA structure, etc. (Bruins, Kapil and Oehme 2000). Microorganisms, including fungi, are known to play key roles in soil and other ecosystems (Staley et al., 1997) and can influence the composition of plants and other larger organisms (Bardgett et al., 2005). Strong selective pressure from metalcontaminated environments on exposed microorganisms can thus have serious consequences for ecosystem function. However, microorganisms have been exposed and adapted to metal contamination since early history (Bruins, Kapil and Oehme 2000; Valls and de Lorenzo 2002). This has resulted in the evolution of several metal resistance mechanisms, such as extracellular and intracellular sequestration, exclusion by permeability barriers, enzymatic detoxification, reduction in sensitivity of cellular targets, efflux pumps, etc. (Nies 2003; Hobman, Yamamoto and Oshima 2007). Such resistance mechanisms form the basis for the use of microorganisms in bioremediation approaches. In addition to readily adapted organisms, these resistance systems may be carried by plasmids or transposons and can therefore be transferred very efficiently to other community members (Top et al., 1990; Sørensen et al., 2005). Unfortunately, little is known about the direct and indirect effects of metals on the composition, richness and evenness of microbial communities, including their ability for adaptation to metal contamination. To date, limited culture-independent molecular surveys have been reported describing the microbial communities of mining-impacted soils. However, recent developments have enabled for sequencing of the composite genomes (‘metagenomics’) or transcribed genes (‘metatranscriptomics’) of entire microbial communities in situ, particularly thanks to improved sequencing technology. In this study, our objective was to take advantage of this and use a metatranscriptomic approach to study the entire microbial community present in metal-contaminated environments, as well as possible biogeochemical roles of individual groups of organisms. To this end, we chose to study a contamination gradient at an abandoned Pb-Zn mine as an example of a heavily metalcontaminated site.
Soil physicochemical and biological characterization For the soil physicochemical characterization, samples were sieved to
