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Title: A comparison of methods used to unveil the genetic and metabolic pool in the built environment

Abstract

BACKGROUND:A majority of indoor residential microbes originate from humans, pets, and outdoor air and are not adapted to the built environment (BE). Consequently, a large portion of the microbes identified by DNA-based methods are either dead or metabolically inactive. Although many exceptions have been noted, the ribosomal RNA fraction of the sample is more likely to represent either viable or metabolically active cells. We examined methodological variations in sample processing using a defined, mock BE microbial community to better understand the scope of technique-based vs. biological-based differences in both ribosomal transcript (rRNA) and gene (DNA) sequence community analysis. Based on in vitro tests, a protocol was adopted for the analysis of the genetic and metabolic pool (DNA vs. rRNA) of air and surface microbiomes within a residential setting. RESULTS:We observed differences in DNA/RNA co-extraction efficiency for individual microbes, but overall, a greater recovery of rRNA using FastPrep (> 50%). Samples stored with various preservation methods at - 80°C experienced a rapid decline in nucleic acid recovery starting within the first week, although post-extraction rRNA had no significant degradation when treated with RNAStable. We recommend that co-extraction samples be processed as quickly as possible after collection. The in vivo analysis revealedmore » significant differences in the two components (genetic and metabolic pool) in terms of taxonomy, community structure, and microbial association networks. Rare taxa present in the genetic pool showed higher metabolic potential (RNA:DNA ratio), whereas commonly detected taxa of outdoor origins based on DNA sequencing, especially taxa of the Sphingomonadales order, were present in lower relative abundances in the viable community. CONCLUSIONS:Although methodological variations in sample preparations are high, large differences between the DNA and RNA fractions of the total microbial community demonstrate that direct examination of rRNA isolated from a residential BE microbiome has the potential to identify the more likely viable or active portion of the microbial community. In an environment that has primarily dead and metabolically inactive cells, we suggest that the rRNA fraction of BE samples is capable of providing a more ecologically relevant insight into the factors that drive indoor microbial community dynamics.« less

Authors:
 [1];  [2];  [3];  [4];  [2];  [2];  [1]
  1. Univ. of California, Berkeley, CA (United States). Dept. of Environmental Science, Policy, and Management; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Genomics and Systems Biology Division
  2. City Univ. of Hong Kong (China)
  3. Univ. of California, Berkeley, CA (United States). Dept. of Environmental Science, Policy, and Management; Univ. of California, San Francisco, CA (United States). Dept. of Physical Therapy and Rehabilitation Science
  4. Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Genomics and Systems Biology Division
Publication Date:
Research Org.:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC); National Science Foundation (NSF)
OSTI Identifier:
1559142
Grant/Contract Number:  
AC02-05CH11231
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Microbiome
Additional Journal Information:
Journal Volume: 6; Journal Issue: 1; Journal ID: ISSN 2049-2618
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; DNA; RNA; Indoor microbiome; Surface; Air; Sample storage; RNAStable; Extraction kit

Citation Formats

Gomez-Silvan, Cinta, Leung, Marcus H. Y., Grue, Katherine A., Kaur, Randeep, Tong, Xinzhao, Lee, Patrick K. H., and Andersen, Gary L. A comparison of methods used to unveil the genetic and metabolic pool in the built environment. United States: N. p., 2018. Web. doi:10.1186/s40168-018-0453-0.
Gomez-Silvan, Cinta, Leung, Marcus H. Y., Grue, Katherine A., Kaur, Randeep, Tong, Xinzhao, Lee, Patrick K. H., & Andersen, Gary L. A comparison of methods used to unveil the genetic and metabolic pool in the built environment. United States. doi:10.1186/s40168-018-0453-0.
Gomez-Silvan, Cinta, Leung, Marcus H. Y., Grue, Katherine A., Kaur, Randeep, Tong, Xinzhao, Lee, Patrick K. H., and Andersen, Gary L. Mon . "A comparison of methods used to unveil the genetic and metabolic pool in the built environment". United States. doi:10.1186/s40168-018-0453-0. https://www.osti.gov/servlets/purl/1559142.
@article{osti_1559142,
title = {A comparison of methods used to unveil the genetic and metabolic pool in the built environment},
author = {Gomez-Silvan, Cinta and Leung, Marcus H. Y. and Grue, Katherine A. and Kaur, Randeep and Tong, Xinzhao and Lee, Patrick K. H. and Andersen, Gary L.},
abstractNote = {BACKGROUND:A majority of indoor residential microbes originate from humans, pets, and outdoor air and are not adapted to the built environment (BE). Consequently, a large portion of the microbes identified by DNA-based methods are either dead or metabolically inactive. Although many exceptions have been noted, the ribosomal RNA fraction of the sample is more likely to represent either viable or metabolically active cells. We examined methodological variations in sample processing using a defined, mock BE microbial community to better understand the scope of technique-based vs. biological-based differences in both ribosomal transcript (rRNA) and gene (DNA) sequence community analysis. Based on in vitro tests, a protocol was adopted for the analysis of the genetic and metabolic pool (DNA vs. rRNA) of air and surface microbiomes within a residential setting. RESULTS:We observed differences in DNA/RNA co-extraction efficiency for individual microbes, but overall, a greater recovery of rRNA using FastPrep (> 50%). Samples stored with various preservation methods at - 80°C experienced a rapid decline in nucleic acid recovery starting within the first week, although post-extraction rRNA had no significant degradation when treated with RNAStable. We recommend that co-extraction samples be processed as quickly as possible after collection. The in vivo analysis revealed significant differences in the two components (genetic and metabolic pool) in terms of taxonomy, community structure, and microbial association networks. Rare taxa present in the genetic pool showed higher metabolic potential (RNA:DNA ratio), whereas commonly detected taxa of outdoor origins based on DNA sequencing, especially taxa of the Sphingomonadales order, were present in lower relative abundances in the viable community. CONCLUSIONS:Although methodological variations in sample preparations are high, large differences between the DNA and RNA fractions of the total microbial community demonstrate that direct examination of rRNA isolated from a residential BE microbiome has the potential to identify the more likely viable or active portion of the microbial community. In an environment that has primarily dead and metabolically inactive cells, we suggest that the rRNA fraction of BE samples is capable of providing a more ecologically relevant insight into the factors that drive indoor microbial community dynamics.},
doi = {10.1186/s40168-018-0453-0},
journal = {Microbiome},
issn = {2049-2618},
number = 1,
volume = 6,
place = {United States},
year = {2018},
month = {4}
}

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Figures / Tables:

Fig. 1 Fig. 1: Schematic chart of in vitro workflow organized by sequence of tasks involved in sampling and extraction. Multiple stages of the in vitro sampling and extraction processes (types of swabs, and surfaces, sample storage prior to extraction, extraction method, and nucleic acid preservation) were tested for the optimal methodsmore » in terms of nucleic acid recovery« less

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