Transitory microbial habitat in the hyperarid Atacama Desert.

Traces of life are nearly ubiquitous on Earth. However, a central unresolved question is whether these traces always indicate an active microbial community or whether, in extreme environments, such as hyperarid deserts, they instead reflect just dormant or dead cells. Although microbial biomass and diversity decrease with increasing aridity in the Atacama Desert, we provide multiple lines of evidence for the presence of an at times metabolically active, microbial community in one of the driest places on Earth. We base this observation on four major lines of evidence: (i) a physico-chemical characterization of the soil habitability after an exceptional rain event, (ii) identified biomolecules indicative of potentially active cells [e.g., presence of ATP, phospholipid fatty acids (PLFAs), metabolites, and enzymatic activity], (iii) measurements of in situ replication rates of genomes of uncultivated bacteria reconstructed from selected samples, and (iv) microbial community patterns specific to soil parameters and depths. We infer that the microbial populations have undergone selection and adaptation in response to their specific soil microenvironment and in particular to the degree of aridity. Collectively, our results highlight that even the hyperarid Atacama Desert can provide a habitable environment for microorganisms that allows them to become metabolically active following an episodic increase in moisture and that once it decreases, so does the activity of the microbiota. These results have implications for the prospect of life on other planets such as Mars, which has transitioned from an earlier wetter environment to today's extreme hyperaridity.

Biochar Reduces Zinc and Cadmium but not Copper and Lead Leaching on a Former Sewage Field.

The leaching of trace metals from anthropogenically contaminated sites poses the risk of groundwater pollution. Biochar has recently been proposed as a soil additive to reduce trace-metal concentrations in the soil solution and to increase water retention, thus reducing drainage. However, field studies on the effects of biochar addition on trace-metal leaching are scarce. Therefore, we added 0, 1, 2.5, and 5 g 100 g of biochar derived from giant miscanthus ( × J.M. Greef & Deuter ex Hodk. & Renvoize) to soil contaminated by former wastewater irrigation and examined water retention and cumulative leaching of Zn, Cd, Cu, and Pb in a 2-yr field study. Cumulative trace-metal leaching was determined by self-integrating accumulators (SIAs) based on ion-exchange resins and compared with data calculated from mean concentrations in the soil solution collected with tension lysimeter plates and groundwater recharge rate. The highest rate of biochar addition increased water retention and thus reduced the amount of drainage water. Mean cumulative Zn and Cd fluxes decreased due to both reduced concentrations in the soil solution and reduced drainage. Although Cu and Pb concentrations in the soil solution increased with biochar addition, the reduced drainage resulted in similar fluxes in the biochar and the control treatment. The cumulative Zn, Cd, and Cu fluxes determined with SIAs were in the same range as the calculated values, while SIA-based Pb fluxes were much higher than those calculated. Since the suction plates excluded colloids, the high SIA-based Pb fluxes indicate colloidal transport and reveal the importance to elucidate the colloidal pathway for risk assessment.

Transport of pharmaceuticals in columns of a wastewater-irrigated Mexican clay soil.

The irrigation or agricultural land with wastewater is increasingly practiced in many parts of the world as a consequence of growing populations and urbanization. The risks emerging from pharmaceuticals that are contained in wastewater for soils and groundwater have hardly been investigated. We studied leaching and effects of naproxen, ibuprofen, bezafibrate, diclofenac, gemfibrocil, clarithromycin, trimethoprim, clindamycin, erythromycin, and metoprolol in a soil column experiment simulating an irrigation event with 8.6 cm of wastewater containing 20 microg L(-1) or 2000 microg L(-1) of each compound or of erythromycin alone. The leached fraction of applied pharmaceuticals ranged from 0.1 +/- 0.1% (clarithromycin, 2000 microg L(-1)) to 130 +/- 41% (naproxen, 20 microg L(-1)) and tended to increase with decreasing K(d) or K(oc). Naproxen transport was similar to that of the tracer chloride. Ibuprofen was also hardly retarded (R = 1.20 +/- 0.18), but showed a higher degradation rate of 0.02 +/- 0.004 h(-1) (2000 microg L(-1)) than naproxen. The transport of a pulse of 2000 microg L(-1) of bezafibrate could be described with a retardation factor of 1.5 and a degradation rate of 0.033 h(-1). The application of erythromycin alone or of a cocktail of all pharmaceuticals significantly increased soil CO2 emissions by 50% 1 d after the application. There is a considerable risk that pharmaceuticals are leached to groundwater during wastewater irrigation.

The fate of silver nanoparticles in riverbank filtration systems - The role of biological components and flow velocity.

Riverbank filtration is a natural process that may ensure the cleaning of surface water for producing drinking water. For silver nanoparticles (AgNP), physico-chemical interaction with sediment surfaces is one major retention mechanism. However, the effect of flow velocity and the importance of biological retention, such as AgNP attachment to biomass, are not well understood, yet. We investigated AgNP (c = 0.6 mg L-1) transport at different spatial and temporal scales in pristine and previously pond water-aged sediment columns. Transport of AgNP under near-natural conditions was studied in a long-term riverbank filtration experiment over the course of one month with changing flow scenarios (i.e. transport at 0.7 m d-1, stagnation, and remobilization at 1.7 m d-1). To elucidate retention processes, we conducted small-scale lab column experiments at low (0.2 m d-1) and high (0.7 m d-1) flow rate using pristine and aged sediments. Overall, AgNP accumulated in the upper centimeters of the sediment both in lab and outdoor experiments. In the lab study, retention of AgNP by attachment to biological components was very effective under high and low flow rate with nearly complete NP accumulation in the upper 2 mm. When organic material was absent, abiotic filtration mechanisms led to NP retention in the upper 5 to 7 cm of the column. In the long-term study, AgNP were transported up to a depth of 25 cm. For the pristine sediment in the lab study and the outdoor experiments only erratic particle breakthrough was detected in a depth of 15 cm. We conclude that physico-chemical interactions of AgNP with sediment surfaces are efficient in retaining AgNP. The presence of organic material provides additional retention sites which increase the filtration capacity of the system. Nevertheless, erratic breakthrough events might transport NP into deeper sediment layers.

Dynamics and functional relevance of ammonia-oxidizing archaea in two agricultural soils.

Crucial steps in geochemical cycles are in many cases performed by more than one group of microorganisms, but the significance of this functional redundancy with respect to ecosystem functioning is poorly understood. Ammonia-oxidizing archaea (AOA) and their bacterial counterparts (AOB) are a perfect system to address this question: although performing the same transformation step, they belong to well-separated phylogenetic groups. Using pig manure amended with different concentrations of sulfadiazine (SDZ), an antibiotic that is frequently used in veterinary medicine, it was possible to affect AOB and AOA to different degrees. Addition of manure stimulated growth of AOB in both soils and, interestingly, also growth of AOA was considerably stimulated in one of the soils. The antibiotic treatments decreased the manure effect notably on AOB, whereas AOA were affected to a lower extent. Model calculations concerning the respective proportions of AOA and AOB in ammonia oxidation indicate a substantial contribution of AOA in one of the soils that further increased under the influence of SDZ, hence indicating functional redundancy between AOA and AOB.

Piggery manure used for soil fertilization is a reservoir for transferable antibiotic resistance plasmids.

In this study, the prevalence and types of transferable antibiotic resistance plasmids in piggery manure were investigated. Samples from manure storage tanks of 15 farms in Germany were analysed, representing diverse sizes of herds, meat or piglet production. Antibiotic resistance plasmids from manure bacteria were captured in gfp-tagged rifampicin-resistant Escherichia coli and characterized. The occurrence of plasmid types was also detected in total community DNA by PCR and hybridization. A total of 228 transconjugants were captured from 15 manures using selective media supplemented with amoxicillin, sulfadiazine or tetracycline. The restriction patterns of 81 plasmids representing different antibiotic resistance patterns or different samples clustered into seven groups. Replicon probing revealed that 28 of the plasmids belonged to IncN, one to IncW, 13 to IncP-1 and 19 to the recently discovered pHHV216-like plasmids. The amoxicillin resistance gene bla-TEM was detected on 44 plasmids, and sulphonamide resistance genes sul1, sul2 and/or sul3 on 68 plasmids. Hybridization of replicon-specific sequences amplified from community DNA revealed that IncP-1 and pHHV216-like plasmids were detected in all manures, while IncN and IncW ones were less frequent. This study showed that 'field-scale' piggery manure is a reservoir of broad-host range plasmids conferring multiple antibiotic resistance genes.

Re-evaluation of the conformational structure of sulfadiazine species using NMR and ab initio DFT studies and its implication on sorption and degradation.

In the environment, the sorption and the degradation of organic pollutants are of increasing interest. The investigation of the chemical structures provides a basis for the development of a suitable binding model approach and for the mechanistic understanding of the chemical fate processes. The aim of this study was the identification of different species of the antibiotic compound sulfadiazine (SDZ) using (1)H and (13)C NMR experiments and ab initio density functional theory (DFT) calculations. In the neutral, aprotic solvent dimethylsulfoxide-d(6) (DMSO-d(6)), a new sulfadiazine structure containing an O-H-N hydrogen bond was identified. In the protic solvent water-d(2) and in dependence on pH and the position of the amidogen hydrogen atom nine possible SDZ conformations were analyzed and five structures were identified. Good conformity between theory and calculation of (1)H NMR was observed. Unfortunately, (13)C NMR is not sensitive enough for comparison and differentiation. In order to verify the identified structures, additional NBO/NLMO (natural localized molecular orbital) analyses were conducted (calculation of net atomic charges, bond polarity, atomic valence, and electron delocalization). Finally, conformation optimizations were performed in order to investigate the stability of the SDZ species. We showed that SDZ contains no S=O double bond, but that it has two S-O single bonds. Surprisingly, negative charges were observed at the pyrimidine nitrogen atom. With these results, the known structure of SDZ was revised. Studies of the geometrical structure and the torsion angles showed that SDZ is very flexible and can be easily fitted to the sorbent. These observations would explain the strong sorbance and hence the rapid formation of non-extractable residues in the environment because SDZ acts as a strong ligand. These results show that that the sulfonamide hydrogen is important for the biological activity but the pyrimidine nitrogen and the sulfonamide oxygen is responsible for the sorbance in environment.

Is colloid-facilitated phosphorus leaching triggered by phosphorus accumulation in sandy soils?

The leaching of colloidal phosphorus (P(coll)) contributes to P losses from agricultural soils. In an irrigation experiment with undisturbed soil columns, we investigated whether the accumulation of P in soils due to excess P additions enhances the leaching of colloids and P(coll) from sandy soils. Furthermore, we hypothesized that large concentrations of P(coll) occur at the onset of leaching events and that P(coll) mobilized from topsoils is retained in subsoils. Soil columns of different P saturation and depth (0-25 and 0-40 cm) were collected at a former disposal site for liquid manure and at the Thyrow fertilization experiment in northeastern Germany. Concentrations of total dissolved P, P(coll), Fe(coll), Al(coll), optical density, zeta potential, pH, and electrical conductivity of the leachates were determined. Colloidal P concentrations ranged from 0.46 to 10 micromol L(-1) and contributed between 1 and 37% to total P leaching. Large P(coll) concentrations leached from the P-rich soil of the manure disposal site were rather related to a large P-content of colloids than to the mobilization of additional colloids. Concentrations of colloids and P(coll) in leachates from P-poor and P-rich columns from Thyrow did not differ significantly. In contrast, accumulation of P in the Werbellin and the Thyrow soil consistently increased dissolved P concentrations to maximum values as high as 300 micromol L(-1). We observed no first-flush of colloids and P(coll) at the beginning of the leaching event. Concentrations of P(coll) leached from 40-cm soil columns were not smaller than those leached from 25-cm columns. Our results illustrate that an accumulation of P in sandy soils does not necessarily lead to an enhanced leaching of colloids and P(coll), because a multitude of factors independent from the P status of soils control the mobility of colloids. In contrast, P accumulation generally increases dissolved P concentrations in noncalcareous soils due to the saturation of the P sorption capacity. This indicates that leaching of dissolved P might be a more widespread environmental problem in areas with P-saturated sandy soils than leaching of P(coll).

Retention and remobilization mechanisms of environmentally aged silver nanoparticles in an artificial riverbank filtration system.

Riverbank filtration systems are important structures that ensure the cleaning of infiltrating surface water for drinking water production. In our study, we investigated the potential risk for a breakthrough of environmentally aged silver nanoparticles (Ag NP) through these systems. Additionally, we identified factors leading to the remobilization of Ag NP accumulated in surficial sediment layers in order to gain insights into remobilization mechanisms. We conducted column experiments with Ag NP in an outdoor pilot plant consisting of water-saturated sediment columns mimicking a riverbank filtration system. The NP had previously been aged in river water, soil extract, and ultrapure water, respectively. We investigated the depth-dependent breakthrough and retention of NP. In subsequent batch experiments, we studied the processes responsible for a remobilization of Ag NP retained in the upper 10 cm of the sediments, induced by ionic strength reduction, natural organic matter (NOM), and mechanical forces. We determined the amount of remobilized Ag by ICP-MS and differentiated between particulate and ionic Ag after remobilization using GFAAS. The presence of Ag-containing heteroaggregates was investigated by combining filtration with single-particle ICP-MS. Single and erratic Ag breakthrough events were mainly found in 30 cm depth and Ag NP were accumulated in the upper 20 cm of the columns. Soil-aged Ag NP showed the lowest retention of only 54%. Remobilization was induced by the reduction of ionic strength and the presence of NOM in combination with mechanical forces. The presence of calcium in the aging- as well as the remobilizing media reduced the remobilization potential. Silver NP were mainly remobilized as heteroaggregates with natural colloids, while dissolution played a minor role. Our study indicates that the breakthrough potential of Ag NP in riverbank filtration systems is generally low, but the aging in soil increases their mobility. Remobilization processes are associated to co-mobilization with natural colloids.

Contrasting effects of biochar on N2O emission and N uptake at different N fertilizer levels on a temperate sandy loam.

Biochar has been frequently suggested as an amendment to improve soil quality and mitigate climate change. To investigate the optimal management of nitrogen (N) fertilization, we examined the combined effect of biochar and N fertilizer on plant N uptake and N2O emissions in a cereal rotation system in a randomized two-factorial field experiment on a sandy loam soil in Brandenburg, Germany. The biochar treatment received 10Mgha-1 wood-derived biochar in September 2012. Four levels of N fertilizer, corresponding to 0, 50%, 100%, 130% of the recommended fertilizer level, were applied in winter wheat (Triticum aestivum L.)) and winter rye (Secale cereal L.) in 2013 and 2014 followed by the catch crop oil radish (Raphanus sativus L. var. oleiformis). Biomass and N uptake of winter wheat and winter rye were significantly affected by the level of N fertilizer but not by biochar. For N uptake of oil radish an interaction effect was observed for biochar and N fertilizer. Without applied fertilizer, 39% higher N uptake was found in the presence of biochar, accompanied by higher soil NH4+ content and elevated cumulative CO2 emissions. At 130% of the recommended fertilizer level, 16% lower N uptake and lower cumulative N2O emissions were found in the biochar-mediated treatment. No significant change in abundance of microbial groups and nosZ gene were observed. Our results highlight that biochar can have a greenhouse gas mitigation effect at high levels of N supply and may stimulate nutrient uptake when no N is supplied.

Insecticide dissipation after repeated field application to a Northern Thailand Ultisol.

Nontarget effects of pesticides may occur when the active ingredients have a long persistence in the environment. The half-lives of six insecticides commonly used in Thai fruit orchards under tropical field conditions were determined. A mixture of endosulfan-alpha and -beta, chlorpyrifos, malathion, dimethoate, and mevinphos was applied five times in 10-day intervals onto an Ultisol (lychee plantation ground-covered with grass vegetation, northwestern Thailand). On days 1, 3, 5, 7, and 10 after each application, composite samples of the topsoil (0-10 cm) were collected and exhaustively extracted. Fitting a first-order model to the datasets revealed rapid initial dissipation [half-lives from 2.2 +/- 0.4 (malathion) to 5.4 +/- 1.3 days (chlorpyrifos)]. Volatilization appeared to be a major process of pesticide dissipation, especially for malathion and mevinphos. Because 8% of the applied endosulfan-alpha and -beta had been converted to the sulfate metabolite within 1 day after the first application, also microbial degradation contributed significantly to pesticide dissipation. Nevertheless, no trend in half-lives over the five application cycles could be observed, indicating that accelerated microbial degradation did not occur for these insecticides following the five applications. Precipitation and soil moisture were key parameters of dissipation, but dissipation processes were too complex to be generalized for all substances studied. Despite their short half-lives, all pesticides except mevinphos accumulated in soil (up to 656%; endosulfan-alpha), and this accumulation correlated significantly with the hydrophobicity of the substances (r = 0.88). This was interpreted as an aging process, and it was concluded that pesticide aging must be considered relevant also in tropical environments, where it has received very limited attention so far.

Polychlorinated biphenyls (PCBs) in soils of the Moscow region: concentrations and small-scale distribution along an urban-rural transect.

In soils of the Moscow region, we examined PCB concentrations in bulk samples and aggregate fractions. Topsoils under grassland and forest at five locations along a southeast-bound transect from Moscow and at a northeastern background location (grassland only) were analysed. We collected aggregates >1 cm and fractionated them into interior and exterior portions and also analysed the remaining soil without the aggregates >1 cm. The concentrations of the sum of 17 PCBs (sigma17PCBs) in 35 bulk soil samples ranged from 3.1 to 42 microg kg(-1). This was 48-61% of the sigma33PCBs determined in 23 selected samples. The congeners 138(+158), 101 and 52 were most abundant. All PCB concentrations and the degree of chlorination declined with increasing distance from Moscow. The PCBs were accumulated in the aggregate exterior (on average 146% of the sigma17PCBs in bulk soil). We conclude that the ecotoxicological risk of PCBs in soils may not be properly assessed with the conventional bulk soil analysis.

Initial Effects of Differently Treated Biogas Residues from Municipal and Industrial Wastes on Spring Barley Yield Formation.

Soil application of biogas residues (BGRs) is important for closing nutrient cycles. This study examined the efficiency and impact on yields and yield formation of solid-liquid separated residues from biodegradable municipal and industrial wastes (bio-waste) in comparison to complete BGRs, nitrification inhibitor, agricultural BGRs, mineral fertilizer and unfertilized plots as control. The experiment was set up as a randomized block design on silt loam Cambisol. Biogas residues from four biogas plants were evaluated. Plants per m², ears per plant, grains per ear and thousand grain weight (TGW) were measured at harvest. Fertilization with BGRs resulted in similar biomass yields compared with mineral fertilizer. Mineral fertilizer (71 dt/ha) and plots fertilized with liquid fraction (59-62 dt/ha) indicated a trend to higher yields than solid fraction or complete BGR due to its high ammonia content. Liquid fractions and fraction with nitrification inhibitor induced fewer plants per m² than corresponding solid and complete variants due to a potential phytotoxicity of high NH4-N concentration during germination. However, barley on plots fertilized with liquid fraction compensated the disadvantages at the beginning during the vegetation period and induced higher grain yields than solid fraction. This was attributable to a higher number of ears per plant and grains per ear. In conclusion, BGRs from biodegradable municipal and industrial wastes can be used for soil fertilization and replace considerable amounts of mineral fertilizer. Our study showed that direct application of the liquid fraction of BGR is the most suitable strategy to achieve highest grain yields. Nevertheless potential phytotoxicity of the high NH4-N concentration in the liquid fraction should be considered.

Colloidal and dissolved phosphorus in sandy soils as affected by phosphorus saturation.

Fertilization exceeding crop requirements causes an accumulation of phosphorus (P) in soils, which might increase concentrations of dissolved and colloidal P in drainage. We sampled soils classified as Typic Haplorthods from four fertilization experiments to test (i) whether increasing degrees of phosphorus saturation (DPS) increase concentrations of dissolved and colloidal P, and (ii) if critical DPS levels can be defined for P release from these soils. Oxalate-extractable concentrations of P, iron (Fe), and aluminum (Al) were quantified to characterize DPS. Turbidity, zeta potential, dissolved P, and colloidal P, Fe, Al, and carbon (C) concentrations were determined in water and KCl extracts. While concentrations of dissolved P decreased with increasing depth, concentrations of water-extractable colloidal P remained constant. In topsoils 28 +/- 17% and in subsoils 94 +/- 8% of water-extractable P was bound to colloids. Concentrations of dissolved P increased sharply for DPS > 0.1. Colloidal P concentrations increased with increasing DPS because of an additional mobilization of colloids and due to an increase of the colloids P contents. In addition to DPS, ionic strength and Ca(2+) affected the release of colloidal P. Hence, using KCl for extraction improved the relationship between DPS and colloidal P compared with water extraction. Accumulation of P in soils increases not only concentrations of dissolved P but also the risk of colloidal P mobilization. Leaching of colloidal P is potentially important for inputs of P into water bodies because colloidal P as the dominant water-extractable P fraction in subsoils was released from soils with relatively low DPS.

Polycyclic aromatic hydrocarbons (PAHs) in soils of the Moscow Region--concentrations, temporal trends, and small-scale distribution.

The knowledge of the environmental fate of polycyclic aromatic hydrocarbons (PAHs) is restricted to few climatic regions of the world almost excluding the Taiga. Our objectives were to (i) separate anthropogenic from background contributions to PAH concentrations and (ii) determine temporal trends in PAH concentrations during the last century including the change in distribution of PAHs in interior and exterior portions of aggregates in soils of the Moscow region. Along a southeast-bound transect from Moscow (windward in winter) and at a background location northeast of Moscow (leeward in winter), seven topsoil samples were collected in 1910-1954 and 35 in 1998-2003. We fractionated the soils in interior and exterior portions of aggregates > 10 mm and remaining soil without aggregates. The sum of 21 PAHs (sigma21PAHs) concentrations in recent bulk soil ranged from 59 to 1350 ng g(-1). The concentrations of all PAHs were lower outside than in Moscow. The range of the concentrations of the sigma21PAHs in archived soil samples (159-1280 ng g(-1)) was similar as in recent soils. In most recent and archived samples, naphthalene and phenanthrene, were most abundant. The concentrations of low-molecular-weight PAHs decreased during the last century at most sites; those of high-molecular-weight compounds increased. The sigma21PAHs concentrations were accumulated in the exterior of aggregates (109%) and depleted in the interior (95%) relative to the concentration in bulk soil (defined as 100%), which was similar to that in the soil without aggregates (99%). The differences between aggregate interior and exterior did not change during the last century. The dominance of naphthalene and phenanthrene is typical of remote regions. The urban influence on PAH concentrations in the last century was small.

Effects of different biochars and digestate on N2O fluxes under field conditions.

Field studies that have investigated the effects of char materials on the emission of nitrous oxide (N2O) are still scarce. Therefore, we conducted a field trial with bio- and hydrochars and measured N2O emissions for one whole year. It was hypothesised that the incorporation of chars reduces the emissions of N2O. Chars were produced by pyrolysis and hydrothermal carbonisation (HTC) using either maize silage or wood residues as feedstock. In addition, after production chars were post-treated with digestate in order to accelerate the ageing process of the chars. Chars and digestate were applied to the soil to raise the C content. Emissions of N2O were measured weekly and soil samples for inorganic nitrogen (N) and soil water-content were taken once a month. Additionally, the abundance of functional marker genes from denitrification (nosZ) was determined in October 2012 and in June 2013. The treatment with pure digestate emitted the most N2O compared to the control and char treatments. However, this was significant only in one case. There were no great differences between the char treatments due to high spatial variability and gene abundance of nosZ did not differ between treatments. Overall, emissions of N2O were relatively low. This was attributed to the heterogeneous distribution of the chars and the sandy soils that did not favour the production of N2O. To conclude, the emissions of N2O were mainly influenced by temperature and precipitation and to a minor extent by the type of char and post-treatment.

Diverse aadA gene cassettes on class 1 integrons introduced into soil via spread manure.

Class 1 integrons are assumed to play an important role in dissemination of antibiotic resistance genes. Class 1 integrons from clinical isolates often have 5'- and 3'-conserved segments flanking the variable region where gene cassettes integrate. Primers targeting the conserved segments were used to amplify the variable region from total community DNA of manure and soil. Integron gene cassette diversity in manure and manured soils was studied by cloning, restriction fragment length polymorphism (RFLP) analysis and sequencing. The analyzed group of integrons could be detected in manured soils but not in soil without a history of manure treatment. aadA gene cassettes encoding resistance to streptomycin and spectinomycin were frequently detected in the clone libraries. To elucidate whether "clinical" class 1 integron gene cassettes are typically introduced via manure into agricultural soil, a survey with a unique set of manure samples originating from 15 different pig-producing facilities was performed. aadA gene cassettes were detected in all field-scale manures, confirming that aadA genes localized on class 1 integrons are introduced via piggery manure into agricultural soils. In addition, class 1 integrons without gene cassettes were detected in total community DNA of many manures.