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Title: Optimization of a Method To Quantify Soil Bacterial Abundance by Flow Cytometry

Abstract

Bacterial abundance is a fundamental metric for understanding the population dynamics of soil bacteria and their role in biogeochemical cycles. Despite its importance, methodological constraints hamper our ability to assess bacterial abundance in terrestrial environments. Here, we aimed to optimize the use of flow cytometry (FCM) to assay bacterial abundances in soil while providing a rigorous quantification of its limitations. Soil samples were spiked withEscherichia colito evaluate the levels of recovery efficiency among three extraction approaches. The optimized method added a surfactant (a tetrasodium pyrophosphate [TSP] buffer) to 0.1 g of soil, applied an intermediate degree of agitation through shaking, and used a Nycodenz density gradient to separate the cells from background debris. This procedure resulted in a high (average, 89%) level of cell recovery. Recovery efficiencies did not differ significantly among sites across an elevation gradient but were positively correlated with percent carbon in the soil samples. Estimated abundances were also highly repeatable between technical replicates. The method was applied to samples from two field studies and, in both cases, was sensitive enough to detect treatment and site differences in bacterial abundances. We conclude that FCM offers a fast and sensitive method to assay soil bacterial abundance from relativelymore » small amounts of soil. Further work is needed to assay differential biases of the method across a wider range of soil types.« less

Authors:
 [1];  [1];  [1];  [1];  [1]
  1. Univ. of California, Irvine, CA (United States)
Publication Date:
Research Org.:
Univ. of California, Irvine, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER); National Science Foundation (NSF)
OSTI Identifier:
1595495
Alternate Identifier(s):
OSTI ID: 1612541
Grant/Contract Number:  
SC0016410; DEB-1457160; MRI-1126749
Resource Type:
Published Article
Journal Name:
mSphere
Additional Journal Information:
Journal Volume: 4; Journal Issue: 5; Journal ID: ISSN 2379-5042
Publisher:
American Society for Microbiology
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 54 ENVIRONMENTAL SCIENCES; Microbiology; bacterial cell count; ecosystem types; extraction procedure; flow cytometry; soil

Citation Formats

Khalili, Banafshe, Weihe, Claudia, Kimball, Sarah, Schmidt, Katharina T., and Martiny, Jennifer B. H. Optimization of a Method To Quantify Soil Bacterial Abundance by Flow Cytometry. United States: N. p., 2019. Web. doi:10.1128/msphere.00435-19.
Khalili, Banafshe, Weihe, Claudia, Kimball, Sarah, Schmidt, Katharina T., & Martiny, Jennifer B. H. Optimization of a Method To Quantify Soil Bacterial Abundance by Flow Cytometry. United States. https://doi.org/10.1128/msphere.00435-19
Khalili, Banafshe, Weihe, Claudia, Kimball, Sarah, Schmidt, Katharina T., and Martiny, Jennifer B. H. Wed . "Optimization of a Method To Quantify Soil Bacterial Abundance by Flow Cytometry". United States. https://doi.org/10.1128/msphere.00435-19.
@article{osti_1595495,
title = {Optimization of a Method To Quantify Soil Bacterial Abundance by Flow Cytometry},
author = {Khalili, Banafshe and Weihe, Claudia and Kimball, Sarah and Schmidt, Katharina T. and Martiny, Jennifer B. H.},
abstractNote = {Bacterial abundance is a fundamental metric for understanding the population dynamics of soil bacteria and their role in biogeochemical cycles. Despite its importance, methodological constraints hamper our ability to assess bacterial abundance in terrestrial environments. Here, we aimed to optimize the use of flow cytometry (FCM) to assay bacterial abundances in soil while providing a rigorous quantification of its limitations. Soil samples were spiked withEscherichia colito evaluate the levels of recovery efficiency among three extraction approaches. The optimized method added a surfactant (a tetrasodium pyrophosphate [TSP] buffer) to 0.1 g of soil, applied an intermediate degree of agitation through shaking, and used a Nycodenz density gradient to separate the cells from background debris. This procedure resulted in a high (average, 89%) level of cell recovery. Recovery efficiencies did not differ significantly among sites across an elevation gradient but were positively correlated with percent carbon in the soil samples. Estimated abundances were also highly repeatable between technical replicates. The method was applied to samples from two field studies and, in both cases, was sensitive enough to detect treatment and site differences in bacterial abundances. We conclude that FCM offers a fast and sensitive method to assay soil bacterial abundance from relatively small amounts of soil. Further work is needed to assay differential biases of the method across a wider range of soil types.},
doi = {10.1128/msphere.00435-19},
journal = {mSphere},
number = 5,
volume = 4,
place = {United States},
year = {2019},
month = {10}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1128/msphere.00435-19

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Cited by: 20 works
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Evaluation of quantitative recovery of bacterial cells and DNA from different lake sediments by Nycodenz density gradient centrifugation
journal, March 2010


Flow cytometric analysis of bacteria- and virus-like particles in lake sediments
journal, March 2006


Recovery and quantification of bacterial cells associated with streambed sediments
journal, October 2008


Flow cytometric analysis of benthic prokaryotes attached to sediment particles
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  • Journal of Microbiological Methods, Vol. 79, Issue 2
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Past, present and future applications of flow cytometry in aquatic microbiology
journal, August 2010


Soil bacterial quantification approaches coupling with relative abundances reflecting the changes of taxa
journal, July 2017


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journal, March 2008


Flow Cytometric Assessment of Bacterial Abundance in Soils, Sediments and Sludge
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Works referencing / citing this record:

Identifying Mechanisms for Successful Ecological Restoration with Salvaged Topsoil in Coastal Sage Scrub Communities
journal, April 2020

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