With the advances of sequencing tools, the fields of environmental microbiology and soil ecology have been transformed. Today, the unculturable majority of soil microbes can be sequenced. Although ...these tools give us tremendous power and open many doors to answer important questions, we must understand how sample processing may impact our results and interpretations. Here, we test the impacts of four soil storage methods on downstream amplicon metabarcoding and qPCR analyses for fungi and bacteria. We further investigate the impact of thaw time on extracted DNA to determine a safe length of time during which this can occur with minimal impact on study results. Overall, we find that storage using standard cold packs with subsequent storage at −20°C is little different than immediate storage in liquid nitrogen, suggesting that the historical and current method is adequate. We further find evidence that storage at room temperature or with aid of RNAlater can lead to changes in community composition and in the case of RNAlater, lower gene copies. We therefore advise against these storage methods for metabarcoding analyses. Finally, we show that over 1 month, DNA extract thaw time does not impact diversity or qPCR metrics. We hope that this work will help researchers working with soil bacteria and fungi make informed decisions about soil storage and transport to ensure repeatability and accuracy of results and interpretations.
Here, we test the impacts of four soil storage methods on downstream amplicon metabarcoding and qPCR analyses for fungi and bacteria and the impact of thaw time on extracted DNA to determine a safe length of time during which this can occur with minimal impact on study results. Overall, we find that storage using standard cold packs with subsequent storage at −20°C is little different than immediate storage in liquid nitrogen, suggesting that the historical and current method is adequate. We also show that over 1 month, DNA extract thaw time does not impact diversity or qPCR metrics.
Petroleum hydrocarbon (PHC) degradation slows significantly during the winter which substantially increases the time it takes to remediate soil in Arctic landfarms. The aim of this laboratory trial ...was to assess the potential of a meat and bonemeal (MBM) biochar to stimulate PHC degradation in contaminated soil collected from Iqaluit, Canada. Over 90 days, 3% (w/w) MBM biochar significantly increased F3- (equivalent nC16-C34) PHC degradation rate constants (k) in frozen soils when compared to the fertilizer (urea and monoammonium phosphate) control. Taking into consideration extensive variability within treatments and negative k values, this difference may not reflect significant remediation. Decreasing C17/Pr and C18/Ph ratios in the frozen soil suggest that this reduction is a result of microbial degradation rather than volatilization. Amendment type and application rate affected the immediate abiotic losses of F2 and F3-PHC in sterile soils, with the greatest losses occurring in compost-amended treatments in the first 24 h. In frozen soils, MBM biochar was found to increase liquid water content (θliquid) but not nutrient supply rates. Under frozen but not thawed conditions, genes for aromatic (C2,3O and nahAc) but not aliphatic (alkB) PHC degradation increased over time in both biochar-amended and control treatments but total viable PHC-degrading populations only increased in biochar-amended soils. Based on these results, it is possible that PHC degradation in biochar-amended soils is active and even enhanced under frozen conditions, but further investigation is required.
•Laboratory-scale PHC degradation was investigated under frozen and thawed conditions using biochar and compost amendments.•Under frozen conditions, F3-PHC degradation rate constants were higher in biochar-amended soils.•Liquid water content increased in frozen soils amended with biochar, but not nutrient supply rates.•Total PHC-degrading microbial populations were stimulated in frozen soils amended with biochar.•Biochar applications at 3% (w/w) have the potential to increase PHC remediation rates in frozen soils.
To integrate biochar as a practical and successful remedial amendment at northern landfarms, components of its formulation and application must be optimized for site-specific environmental ...conditions. Different biochar amendments were applied to petroleum hydrocarbon (PHC)-contaminated soil at two northern field sites (Iqaluit and Whitehorse) and in a laboratory study at -5°C to determine the effects of application method (injection or incorporation) and biochar type (wood, fishmeal, bonemeal, and/or compost) on PHC degradation and associated soil properties. Incorporation decreased F2 (equivalent C-C) and F3 (equivalent C-C) PHC concentrations in soil after 31 d, whereas injection did not decrease PHC concentrations until Day 334. Bonemeal-derived biochar selectively stimulated F3-PHC degradation in frozen soil over 90 d under controlled laboratory conditions. In the field, there was little difference in PHC degradation between biochar types and the fertilizer control. Incorporation also increased NO availability, and in field trials, all biochars increased NO availability relative to the fertilizer control, whereas the effects of biochars on NH and PO were variable. Aromatic functional gene abundance was enhanced when treatments were incorporated, compared with when injected. In field trials, 6% Zakus wood plus fertilizer inhibited aliphatic and aromatic gene abundance. Liquid water content increased in incorporated treatments, specifically those amended with fishmeal biochar. Incorporation was the most successful application method for these northern soils, and although biochar amendments are not clearly effective in reducing PHC concentrations, there is evidence to suggest it can beneficially influence soil properties and PHC degradation under specific environmental conditions.
Aphanomyces euteiches is a soil-borne pathogen that causes root rot in susceptible pulse crops. Although widespread across Canada, A. euteiches was not reported in Saskatchewan until 2012. The ...distribution of this pathogen across the Canadian prairies is unknown and it is unclear whether it is native to the region. The objective of this study was to conduct a survey of A. euteiches across the major agricultural soils of Saskatchewan, Canada to determine its distribution and to assess the relative importance of factors related to its distribution and possible spread across the province. Soil samples were collected from lentil and pea fields and their associated roadside ditches, as well as from native and tame rangelands. Soil samples were analyzed using a quantitative polymerase chain reaction (qPCR) assay to detect and quantify the presence of A. euteiches. In addition, root rot severity was assessed on affected pea and lentil plants, and the physical and chemical properties of the soils were measured. A. euteiches was present across all land use (i.e., annual cropland, roadside ditches, and rangeland) and soil types in Saskatchewan. A. euteiches abundance was highest in samples collected from annually cropped fields. Within annual cropland, A. euteiches abundance was higher in pea fields than lentil fields. Soil moisture, total and organic carbon, and total nitrogen were positively correlated to A. euteiches abundance in field and roadside soils. Based on this survey, A. euteiches is wide-spread across all the major agricultural regions of Saskatchewan and is not limited to specific soil or land use types. The frequent detection of A. euteiches in native rangeland indicates it is likely native to Saskatchewan. The significant relationship between A. euteiches gene abundance and disease severity in pea fields indicates that the qPCR assay may be a useful indicator for predicting the potential for certain agricultural soils to cause A. euteiches root rot in pea crops.
•euteiches is widespread across all soil and land use types in Saskatchewan.•Soil moisture, total C, and total N were correlated with A. euteiches abundance.•Frequent detection in rangeland indicates A. euteiches is likely native to region.
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