Soil stabilisation for DNA metabarcoding of plants and fungi. Implications for sampling at remote locations or via third-parties

Storage of soil samples prior to metagenomic analysis presents a problem. If field sites are remote or if samples are collected by third parties, transport to analytical laboratories may take several days or even weeks. The bulk of such samples and requirement for later homogenisation precludes the convenient use of a stabilisation buffer, so samples are usually cooled or frozen during transit. There has been limited testing of the most appropriate storage methods for later study of soil organisms by eDNA approaches. Here we tested a range of storage methods on two contrasting soils, comparing these methods to the control of freezing at -80 °C, followed by freeze-drying. To our knowledge, this is the first study to examine the effect of storage conditions on eukaryote DNA in soil, including both viable organisms (fungi) and DNA contained within dying/dead tissues (plants). For fungi, the best storage regimes (closest to the control) were storage at 4 °C (for up to 14 d) or active air-drying at room temperature. The worst treatments involved initial freezing, followed by thawing which led to significant later spoilage. The key spoilage organisms were identified as Metarhizium carneum and Mortierella spp., with a general increase in saprotrophic fungi and reduced abundances of mycorrhizal/biotrophic fungi. Plant data showed a similar pattern, but with greater variability in community structure, especially in the freeze-thaw treatments, probably due to stochastic variation in substrates for fungal decomposition, algal proliferation and some seed germination. In the absence of freeze drying facilities, samples should be shipped refrigerated, but not frozen if there is any risk of thawing

Deep seam and minesoil carbon sequestration potential of the South Wales Coalfield, UK

Combustion of coal for energy generation has been a significant contributor to increased concentrations of atmospheric carbon dioxide. It is of interest to evaluate the potential of former coalfields for mitigating these increases by carbon sequestration and to compare different options to achieving this end. Here, carbon sequestration in residual coal seams and through reclamation of spoil tips is compared, and their carbon dioxide storage potential in the South Wales Coalfield estimated. Coal seam sequestration estimates come from an established methodology and consider the total unmined coal resource below 500 m deep with potential for carbon sequestration. The most likely effective deep seam storage capacity is 104.9 Mt carbon dioxide, taking account of reservoir conditions and engineering factors. Whilst many spoil tips in South Wales have been reclaimed, the focus has not been on carbon sequestration potential. Estimates of minesoil restoration sequestration capacity were based on a survey of restored minesoil and vegetation carbon stocks, mainly on sites 20–30 years after restoration; data from this survey were then extrapolated to the coalfield as a whole. Minesoil storage is estimated at 1.5 or 2.5 Mt (+2.2 Mt in tree biomass) carbon dioxide based on average grassland or woodland measurements, respectively; modelled data predicted equilibrium values of 2.9 and 2.6 Mt carbon dioxide respectively in grassland or woodland minesoils. If all sites achieved close to the maximum capacity in their land use class, minesoil storage capacity would increase to 2.1 or 3.9 Mt carbon dioxide, respectively. Combining the best woodland minesoil and standing biomass values, sequestration capacity increases to 7.2 Mt carbon dioxide. The wider social, economic, environmental and regulatory constraints to achieving this sequestration for each approach are discussed. Coal seam sequestration has a much higher capacity but sequestration in mine sites is less costly and has fewer regulatory constraints. Findings indicate a significant combined potential for carbon sequestration in the South Wales Coalfield and highlight challenges in achieving this potential. On a global scale, ex-coalfield sequestration could contribute to broader efforts to mitigate emissions

Maternal versus artificial rearing shapes the rumen microbiome having minor long-term physiological implications

Increasing productivity is a key target in ruminant science which requires better understanding of the rumen microbiota. This study investigated how maternal versus artificial rearing shapes the rumen microbiota using 24 sets of triplet lambs. Lambs within each sibling set were randomly assigned to natural rearing on the ewe (NN); ewe colostrum for 24 h followed by artificial milk feeding (NA); and colostrum alternative and artificial milk feeding (AA). Maternal colostrum feeding enhanced VFA production at weaning but not thereafter. At weaning, lambs reared on milk replacer had no rumen protozoa and lower microbial diversity, whereas natural rearing accelerated the rumen microbial development and facilitated the transition to solid diet. Differences in the rumen prokaryotic communities disappear later in life when all lambs were grouped on the same pasture up to 23 weeks of age. However, NN animals retained higher fungal diversity and abundances of Piromyces, Feramyces and Diplodiniinae protozoa as well as higher feed digestibility (+4%) and animal growth (+6.5%) during the grazing period. Nevertheless, no correlations were found between rumen microbiota and productive outcomes. These findings suggest that the early life nutritional intervention determine the initial rumen microbial community, but the persistence of these effects later in life is weak.</p

Vegetation and edaphic factors influence rapid establishment of distinct fungal communities on former coal-spoil sites

We investigated re-establishment of fungal communities on eight former colliery sites in South Wales following revegetation 22–27 y earlier. Regraded bare coal-spoil was seeded to sheep-grazed grasslands, with saplings planted into coal-spoil for woodlands. Metabarcoding (28S rRNA, D1 region) of soil fungal populations showed that woodland and grassland habitats were clearly divergent but edaphic variables only weakly affected fungal community structure. Root-associated basidiomycetes dominated all habitats, with ectomycorrhizal fungi more abundant in woodlands and Clavariaceae/Hygrophoraceae (‘CHEG’ fungi) in grasslands. The composition of coal-spoil grassland communities resembled that of a typical upland grassland site, suggesting that propagule immigration was not a limiting factor. However, fungal biomass (ergosterol) was 3-fold lower, reflecting high bulk density and poor structure. Re-establishment of fungal communities in coal-spoil soils represents an important barometer of restoration success. From a fungal conservation perspective, such sites represent important refugia for waxcap fungi subject to habitat loss from agricultural intensificatio

Teaching word recognition to children with severe learning difficulties: an exploratory comparison of teaching methods

Background: Some children with severe learning difficulties fail to begin word recognition. For these children there is a need for an effective and appropriate pedagogy. However, conflicting advice can be found regarding this derived from teaching approaches that are not based on a shared understanding of how reading develops or the skills that the non-reader needs to master. Purpose: In this research, three techniques for teaching word recognition in this context are described and compared: (1) the handle technique, (2) morphing method and (3) word alone. It also discusses whether it is appropriate for such small-scale research to influence pedagogy. Programme description: The handle technique uses an abstract mnemonic cue used to teach word recognition, and previous research indicates it is more successful than the presentation of words alone. The morphing method transforms a word into a photographic picture and a previous study suggested that it might also be more effective that presenting words alone. Sample: Six children between 11 and 13 years of age were selected. The criterion for selection was being unable to recognise any words from the British Ability Scales Reading Test. All the children attended a school for children with severe learning difficulties. Design and methods: A three-condition related design was used. The order in which the conditions were presented was counterbalanced and each child was taught 12 words, four words in each experimental condition. The children encountered each of the three methods and overall each word was taught via each method. Within conditions (teaching methods), the presentation of words was randomised. The number of words that the children could read (without cues) before each session was recorded, following the presentation of the uncued words in a random order. The difference in the number of words recognised between the three conditions was considered using a non-parametric statistical analysis. Results: The results suggest that the handle approach might be a more effective method of teaching word recognition. Conclusion: Research in this area is necessarily small in scale. However, it is ongoing and cumulative, and can give insights into potentially beneficial changes in classroom practice

Soil stabilisation for DNA metabarcoding of plants and fungi. Implications for sampling at remote locations or via third-parties

Storage of soil samples prior to metagenomic analysis presents a problem. If field sites are remote or if samples are collected by third parties, transport to analytical laboratories may take several days or even weeks. The bulk of such samples and requirement for later homogenisation precludes the convenient use of a stabilisation buffer, so samples are usually cooled or frozen during transit. There has been limited testing of the most appropriate storage methods for later study of soil organisms by eDNA approaches. Here we tested a range of storage methods on two contrasting soils, comparing these methods to the control of freezing at -80 °C, followed by freeze-drying. To our knowledge, this is the first study to examine the effect of storage conditions on eukaryote DNA in soil, including both viable organisms (fungi) and DNA contained within dying/dead tissues (plants). For fungi, the best storage regimes (closest to the control) were storage at 4 °C (for up to 14 d) or active air-drying at room temperature. The worst treatments involved initial freezing, followed by thawing which led to significant later spoilage. The key spoilage organisms were identified as Metarhizium carneum and Mortierella spp., with a general increase in saprotrophic fungi and reduced abundances of mycorrhizal/biotrophic fungi. Plant data showed a similar pattern, but with greater variability in community structure, especially in the freeze-thaw treatments, probably due to stochastic variation in substrates for fungal decomposition, algal proliferation and some seed germination. In the absence of freeze drying facilities, samples should be shipped refrigerated, but not frozen if there is any risk of thawing.</jats:p

Soil stabilisatizion for DNA metabarcoding of plants and fungi. Implications for sampling at remote locations or via third-parties

AbstractStorage of soil samples prior to metagenomic analysis presents a problem. If field sites are remote or if samples are collected by third parties, transport to analytical laboratories may take several days or even weeks. The bulk of such samples and requirement for later homogenisation precludes the convenient use of a stabilisation buffer, so samples are usually cooled or frozen during transit. There has been limited testing of the most appropriate storage methods for later study of soil organisms by eDNA approaches. Here we tested a range of storage methods on two contrasting soils, comparing these methods to the control of freezing at −80°C followed by freeze-drying. To our knowledge this is the first study to examine the effect of storage conditions on eukaryote DNA in soil, including both viable organisms (fungi) and DNA contained within dying/dead tissues (plants). For fungi, the best storage regimes (closest to the control) were storage a 4°C (for up to 14 d) or active air-drying at room temperature. The worst treatments involved initial freezing followed by thawing which led to significant later spoilage. The key spoilage organisms were identified as Metarhizium carneum and Mortierella spp., with a general increase in saprotrophic fungi and reduced abundances of mycorrhizal/biotrophic fungi. Plant data showed a similar pattern but with greater variability in community structure especially in the freeze-thaw treatments, probably due to stochastic variation in substrates for fungal decomposition, algal proliferation and some seed germination. In the absence of freeze drying facilities, samples should be shipped refrigerated but not frozen if there is any risk of thawing.</jats:p