Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota
Studies involving gut microbiome analysis play an increasing role in the evaluation of health and disease in humans and animals alike. Fecal sampling methods for DNA preservation in laboratory, clinical, and field settings can greatly influence inferences of microbial composition and diversity, but...
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ftdoajarticles:oai:doaj.org/article:7496063c58274b06ab3e23eb1bdfb510 2024-01-07T09:42:37+01:00 Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota Katti R. Horng Holly H. Ganz Jonathan A. Eisen Stanley L. Marks 2018-05-01T00:00:00Z https://doi.org/10.7717/peerj.4827 https://doaj.org/article/7496063c58274b06ab3e23eb1bdfb510 EN eng PeerJ Inc. https://peerj.com/articles/4827.pdf https://peerj.com/articles/4827/ https://doaj.org/toc/2167-8359 doi:10.7717/peerj.4827 2167-8359 https://doaj.org/article/7496063c58274b06ab3e23eb1bdfb510 PeerJ, Vol 6, p e4827 (2018) Fecal preservation Microbiome Canine 16S rRNA Gut microbiota Sample storage Medicine R Biology (General) QH301-705.5 article 2018 ftdoajarticles https://doi.org/10.7717/peerj.4827 2023-12-10T01:50:14Z Studies involving gut microbiome analysis play an increasing role in the evaluation of health and disease in humans and animals alike. Fecal sampling methods for DNA preservation in laboratory, clinical, and field settings can greatly influence inferences of microbial composition and diversity, but are often inconsistent and under-investigated between studies. Many laboratories have utilized either temperature control or preservation buffers for optimization of DNA preservation, but few studies have evaluated the effects of combining both methods to preserve fecal microbiota. To determine the optimal method for fecal DNA preservation, we collected fecal samples from one canine donor and stored aliquots in RNAlater, 70% ethanol, 50:50 glycerol:PBS, or without buffer at 25 °C, 4 °C, and −80 °C. Fecal DNA was extracted, quantified, and 16S rRNA gene analysis performed on Days 0, 7, 14, and 56 to evaluate changes in DNA concentration, purity, and bacterial diversity and composition over time. We detected overall effects on bacterial community of storage buffer (F-value = 6.87, DF = 3, P < 0.001), storage temperature (F-value=1.77, DF = 3, P = 0.037), and duration of sample storage (F-value = 3.68, DF = 3, P < 0.001). Changes in bacterial composition were observed in samples stored in −80 °C without buffer, a commonly used method for fecal DNA storage, suggesting that simply freezing samples may be suboptimal for bacterial analysis. Fecal preservation with 70% ethanol and RNAlater closely resembled that of fresh samples, though RNAlater yielded significantly lower DNA concentrations (DF = 8.57, P < 0.001). Although bacterial composition varied with temperature and buffer storage, 70% ethanol was the best method for preserving bacterial DNA in canine feces, yielding the highest DNA concentration and minimal changes in bacterial diversity and composition. The differences observed between samples highlight the need to consider optimized post-collection methods in microbiome research. Article in Journal/Newspaper Canis lupus Directory of Open Access Journals: DOAJ Articles PeerJ 6 e4827 |
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ftdoajarticles |
language |
English |
topic |
Fecal preservation Microbiome Canine 16S rRNA Gut microbiota Sample storage Medicine R Biology (General) QH301-705.5 |
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Fecal preservation Microbiome Canine 16S rRNA Gut microbiota Sample storage Medicine R Biology (General) QH301-705.5 Katti R. Horng Holly H. Ganz Jonathan A. Eisen Stanley L. Marks Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
topic_facet |
Fecal preservation Microbiome Canine 16S rRNA Gut microbiota Sample storage Medicine R Biology (General) QH301-705.5 |
description |
Studies involving gut microbiome analysis play an increasing role in the evaluation of health and disease in humans and animals alike. Fecal sampling methods for DNA preservation in laboratory, clinical, and field settings can greatly influence inferences of microbial composition and diversity, but are often inconsistent and under-investigated between studies. Many laboratories have utilized either temperature control or preservation buffers for optimization of DNA preservation, but few studies have evaluated the effects of combining both methods to preserve fecal microbiota. To determine the optimal method for fecal DNA preservation, we collected fecal samples from one canine donor and stored aliquots in RNAlater, 70% ethanol, 50:50 glycerol:PBS, or without buffer at 25 °C, 4 °C, and −80 °C. Fecal DNA was extracted, quantified, and 16S rRNA gene analysis performed on Days 0, 7, 14, and 56 to evaluate changes in DNA concentration, purity, and bacterial diversity and composition over time. We detected overall effects on bacterial community of storage buffer (F-value = 6.87, DF = 3, P < 0.001), storage temperature (F-value=1.77, DF = 3, P = 0.037), and duration of sample storage (F-value = 3.68, DF = 3, P < 0.001). Changes in bacterial composition were observed in samples stored in −80 °C without buffer, a commonly used method for fecal DNA storage, suggesting that simply freezing samples may be suboptimal for bacterial analysis. Fecal preservation with 70% ethanol and RNAlater closely resembled that of fresh samples, though RNAlater yielded significantly lower DNA concentrations (DF = 8.57, P < 0.001). Although bacterial composition varied with temperature and buffer storage, 70% ethanol was the best method for preserving bacterial DNA in canine feces, yielding the highest DNA concentration and minimal changes in bacterial diversity and composition. The differences observed between samples highlight the need to consider optimized post-collection methods in microbiome research. |
format |
Article in Journal/Newspaper |
author |
Katti R. Horng Holly H. Ganz Jonathan A. Eisen Stanley L. Marks |
author_facet |
Katti R. Horng Holly H. Ganz Jonathan A. Eisen Stanley L. Marks |
author_sort |
Katti R. Horng |
title |
Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
title_short |
Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
title_full |
Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
title_fullStr |
Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
title_full_unstemmed |
Effects of preservation method on canine (Canis lupus familiaris) fecal microbiota |
title_sort |
effects of preservation method on canine (canis lupus familiaris) fecal microbiota |
publisher |
PeerJ Inc. |
publishDate |
2018 |
url |
https://doi.org/10.7717/peerj.4827 https://doaj.org/article/7496063c58274b06ab3e23eb1bdfb510 |
genre |
Canis lupus |
genre_facet |
Canis lupus |
op_source |
PeerJ, Vol 6, p e4827 (2018) |
op_relation |
https://peerj.com/articles/4827.pdf https://peerj.com/articles/4827/ https://doaj.org/toc/2167-8359 doi:10.7717/peerj.4827 2167-8359 https://doaj.org/article/7496063c58274b06ab3e23eb1bdfb510 |
op_doi |
https://doi.org/10.7717/peerj.4827 |
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PeerJ |
container_volume |
6 |
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e4827 |
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1787423648285458432 |