Genomics technology for assessing soil pollution

Transcription and metabolite analysis is a powerful way to reveal physiological shifts in response to environmental pollution. Recent studies on earthworms, including one in BMC Biology, show that the type of pollution and its availability for uptake by organisms can differentially affect transcription and metabolism

Avoidance tests with the oribatid mite Oppia nitens (Acari: Oribatida) in cadmium-spiked natural soils

Avoidance behavior can be a useful parameter for assessing the ability of organisms to escape from pollutants in their environment. For soil evaluation, a variety of invertebrates is used including the oribatid mite Oppia nitens. Here, we tested the avoidance behavior of O. nitens using a two-chamber test and an escape test with exposures to different cadmium concentrations of up to 800 mg kg−1 dry LUFA 2.2 soil for 2, 4, and 6 days, and up to 7 weeks. With the two-chamber method, the oribatid mites had the choice between clean and polluted soils, whereas they were allowed to escape from a box with polluted soil to clean containers without soil with the escape method. Avoidance of cadmium was observed after 2 days in both tests and the net response of the mites in the two-chamber test increased with increasing cadmium exposure concentrations. Mite responses varied through time, especially with the escape method; with the avoidance behavior becoming more variable and overall non-significant with longer test durations. This is the first study investigating the escape test simultaneously with long-term avoidance of cadmium by O. nitens. This mite species is a promising species for avoidance testing in soil ecotoxicology, but more experiments are needed to evaluate the factors that influence its responses in laboratory tests and the consequences for its distribution in contaminated ecosystems

Transcriptomics reveals extensive inducible biotransformation in the soil-dwelling invertebrate Folsomia candida exposed to phenanthrene

Background: Polycyclic aromatic hydrocarbons are common pollutants in soil, have negative effects on soil ecosystems, and are potentially carcinogenic. The Springtail (Collembola) Folsomia candida is often used as an indicator species for soil toxicity. Here we report a toxicogenomic study that translates the ecological effects of the polycyclic aromatic hydrocarbon phenanthrene in soil to the early transcriptomic responses in Folsomia candida. Results: Microarrays wereused to examine two different exposure concentrations of phenanthrene, namely the EC10 (24.95 mg kg-1 soil) and EC50 (45.80 mg kg-1 soil) on reproduction of this springtail, which evoked 405 and 251 differentially expressed transcripts, respectively. Fifty transcripts were differential in response to either concentration. Many transcripts encoding xenobiotic detoxification and biotransformation enzymes (phases I, II, and III) were upregulated in response to either concentration. Furthermore, indications of general and oxidative stress were found in response to phenanthrene. Chitin metabolism appeared to be disrupted particularly at the low concentration, and protein translation appeared suppressed at the high concentration of phenanthrene; most likely in order to reallocate energy budgets for the detoxification process. Finally, an immune response was evoked especially in response to the high effect concentration, which was also described in a previous transcriptomic study using the same effect concentration (EC50) of cadmium. Conclusion: Our study provides new insights in the molecular mode of action of the important polluting class of polycyclic aromatic hydrocarbons in soil animals. Furthermore, we present a fast, sensitive, and specific soil toxicity test which enhances traditional tests and may help to improve current environmental risk assessments and monitoring of potentially polluted sites. © 2009 Nota et al; licensee BioMed Central Ltd

Functional and compositional responses in soil microbial communities along two metal pollution gradients : does the level of historical pollution affect resistance against secondary stress?

We examined how the exposure to secondary stressors affected the functional and compositional responses of microbial communities along two metal pollution gradients in Polish forests and whether responses were influenced by the level of metal pollution. Basal respiration rate and community composition, as determined by 16S rRNA gene-based denaturing gradient analysis, were studied in laboratory-incubated microcosms, containing soil samples taken from different locations in the field gradients, and subjected to secondary stress factors (arsenic, salt, benzo[a]pyrene or flooding). Soils adapted to higher metal contamination levels were more resistant to arsenic and salt stress compared to less polluted soils, indicating that functional and compositional responses to these stresses were related to the level of historical pollution in the long-term contaminated forest soils. However, community composition and functioning in soils along the two gradients were resistant to benzo[a]pyrene and flooding stress. Changes in respiration were significantly related to changes in community composition. Knowledge on the functional gene capabilities prior to adding the secondary stressor benefitted understanding the functional responses toward additional stressors. Our study highlights that microbial communities selected for metal resistance in the field might also become more resistant against some secondary stress factors; however, the type of stress and the level of historical pollution play a decisive role in community-level responses toward secondary stressors

No effect of selected engineered nanomaterials on reproduction and survival of the springtail Folsomia candida

Although the number of studies on engineered nanomaterial (NM) toxicity to soil invertebrates is increasing, only a few studies have reported toxicity of NMs to soil dwelling model species, such as the invertebrate Folsomia candida. The main objective of this study was to determine the toxicity of five different engineered NMs (WCCo, CuO, Fe2O3, organic pigment and MWCNTs) for the springtail F. candida. Copper(ii), cobalt and iron chlorides were taken as positive controls. A standardized OECD test was used to measure effects on reproduction and survival, and toxicity was related to metal concentrations in soil and pore water. None of the NMs exerted adverse effects on springtail reproduction and survival at concentrations up to 6400 mg per kg dry soil, whereas the Cu, Co and Fe chlorides resulted in 50% decline in springtail reproduction at 981, 469 and 569 mg metal ion per kg dry soil, respectively. The absence of toxicity of the NMs could partly be explained by the low porewater metal concentrations, suggesting low solubility or slow solubilisation. The fate of engineered NMs in soil is rather complex but needs better understanding to facilitate predicting exposure of soil organisms

Recombinational micro-evolution of functionally different metallothionein promoter alleles from Orchesella cincta

<p>Abstract</p> <p>Background</p> <p>Metallothionein (<it>mt</it>) transcription is elevated in heavy metal tolerant field populations of <it>Orchesella cincta </it>(Collembola). This suggests that natural selection acts on transcriptional regulation of <it>mt </it>in springtails at sites where cadmium (Cd) levels in soil reach toxic values This study investigates the nature and the evolutionary origin of polymorphisms in the metallothionein promoter (<it>pmt</it>) and their functional significance for <it>mt </it>expression.</p> <p>Results</p> <p>We sequenced approximately 1600 bp upstream the <it>mt </it>coding region by genome walking. Nine <it>pmt </it>alleles were discovered in NW-European populations. They differ in the number of some indels, consensus transcription factor binding sites and core promoter elements. Extensive recombination events between some of the alleles can be inferred from the alignment. A deviation from neutral expectations was detected in a cadmium tolerant population, pointing towards balancing selection on some promoter stretches. Luciferase constructs were made from the most abundant alleles, and responses to Cd, paraquat (oxidative stress inducer) and moulting hormone were studied in cell lines. By using paraquat we were able to dissect the effect of oxidative stress from the Cd specific effect, and extensive differences in <it>mt </it>induction levels between these two stressors were observed.</p> <p>Conclusion</p> <p>The <it>pmt </it>alleles evolved by a number of recombination events, and exhibited differential inducibilities by Cd, paraquat and molting hormone. In a tolerant population from a metal contaminated site, promoter allele frequencies differed significantly from a reference site and nucleotide polymorphisms in some promoter stretches deviated from neutral expectations, revealing a signature of balancing selection. Our results suggest that the structural differences in the <it>Orchesella cincta </it>metallothionein promoter alleles contribute to the metallothionein -over-expresser phenotype in cadmium tolerant populations.</p

Resilience of soil microbial communities to metals and additional stressors : DNA-based approaches for assessing "Stress-on-Stress" responses

International audienceMany microbial ecology studies have demonstrated profound changes in community composition caused by environmental pollution, as well as adaptation processes allowing survival of microbes in polluted ecosystems. Soil microbial communities in polluted areas with a long-term history of contamination have been shown to maintain their function by developing metal-tolerance mechanisms. In the present work, we review recent experiments, with specific emphasis on studies that have been conducted in polluted areas with a long-term history of contamination that also applied DNA-based approaches. We evaluate how the "costs" of adaptation to metals affect the responses of metal-tolerant communities to other stress factors ("stress-on-stress"). We discuss recent studies on the stability of microbial communities, in terms of resistance and resilience to additional stressors, focusing on metal pollution as the initial stress, and discuss possible factors influencing the functional and structural stability of microbial communities towards secondary stressors. There is increasing evidence that the history of environmental conditions and disturbance regimes play central roles in responses of microbial communities towards secondary stressors

Molecular mechanisms of zinc toxicity in the potworm Enchytraeus crypticus, analysed by high-throughput gene expression profiling

Zinc (Zn) is known to be relatively toxic to some soil-living invertebrates including the ecologically important enchytraeid worms. To reveal the molecular mechanisms of zinc toxicity we assessed the gene expression profile of Enchytraeus crypticus (Enchytraeidae), exposed to the reproduction effect concentrations EC10 and EC50, over 4 consecutive days, using a high-throughput microarray (species customized). Three main mechanisms of toxicity to Zn were observed: 1) Zn trafficking (upregulation of zinc transporters, a defence response to regulate the cellular zinc level), 2) oxidative stress (variety of defence mechanisms, triggered by Reactive Oxygen Species (ROS)), and 3) effects on the nervous system (possibly the primary lesion explaining the avoidance behaviour and also why enchytraeids are relatively susceptible to Zn). The adverse outcome at the organism level (reproduction EC50) could be predicted based on gene expression (male gonad development, oocyte maturation), with Zn at the EC50 affecting processes related to higher stress levels. The gene expression response was time-dependent and reflected the cascade of events taking place over-time. The 1 to 4 days of exposure design was a good strategy as it captured the time for sequence of events towards zinc adverse outcomes in E. crypticus

Structural and functional variation in soil fungal communities associated with litter bags containing maize leaf

Soil fungi are key players in the degradation of recalcitrant organic matter in terrestrial ecosystems. To examine the organisms and genes responsible for complex organic matter degradation in soil, we tracked changes in fungal community composition and expressed genes in soil adjacent to mesh bags containing maize leaves undergoing decomposition. Using high-throughput sequencing approaches, changes in fungal community composition were determined by targeting 18S rRNA gene sequences, whereas community gene expression was examined via a metatranscriptomic approach. The majority of the 93 000 partial 18S rRNA gene sequences generated, were affiliated with the Ascomycota and Basidiomycota. Fungal diversity was at least 224 operational taxonomic units at the 97% similarity cutoff level. During litter degradation, the relative proportion of Basidiomycota increased, with a decrease in Ascomycota : Basidiomycota ratios over time. The most commonly detected decomposition-associated fungi included Agaricomycetes and Tremellales as well as unclassified Mucoromycotina. The majority of protein families found in the metatranscriptomic data were affiliated to fungal groups described to degrade plant-derived cellulose, such as Mucoraceae, Chaetomiaceae, Sordariaceae, Sebacinaceae, Tremellaceae, Psathyrellaceae and Schizophyllaceae. The combination of high-throughput rRNA gene-based and metatranscriptomic approaches provided perspectives into the organisms and genes involved in complex organic matter in soi

Reference genes for QRT-PCR tested under various stress conditions in Folsomia candida and Orchesella cincta (Insecta, Collembola)

<p>Abstract</p> <p>Background</p> <p>Genomic studies measuring transcriptional responses to changing environments and stress currently make their way into the field of evolutionary ecology and ecotoxicology. To investigate a small to medium number of genes or to confirm large scale microarray studies, Quantitative Reverse Transcriptase PCR (QRT-PCR) can achieve high accuracy of quantification when key standards, such as normalization, are carefully set. In this study, we validated potential reference genes for their use as endogenous controls under different chemical and physical stresses in two species of soil-living Collembola, <it>Folsomia candida </it>and <it>Orchesella cincta</it>. Treatments for <it>F. candida </it>were cadmium exposure, phenanthrene exposure, desiccation, heat shock and pH stress, and for <it>O. cincta </it>cadmium, desiccation, heat shock and starvation.</p> <p>Results</p> <p>Eight potential reference genes for <it>F. candida </it>and seven for <it>O. cincta </it>were ranked by their stability per stress factor using the programs geNorm and Normfinder. For <it>F. candida </it>the succinate dehydrogenase (<it>SDHA</it>) and eukaryotic transcription initiation factor 1A (<it>ETIF</it>) genes were found the most stable over the different treatments, while for <it>O. cincta</it>, the beta actin (<it>ACTb</it>) and tyrosine 3-monooxygenase (<it>YWHAZ</it>) genes were the most stable.</p> <p>Conclusion</p> <p>We present a panel of reference genes for two emerging ecological genomic model species tested under a variety of treatments. Within each species, different treatments resulted in differences in the top stable reference genes. Moreover, the two species differed in suitable reference genes even when exposed to similar stresses. This might be attributed to dissimilarity of physiology. It is vital to rigorously test a panel of reference genes for each species and treatment, in advance of relative quantification of QRT-PCR gene expression measurements.</p