home .Featured Microbiome Methods Q & A – submit your questions …

Microbiome Methods Q & A – submit your questions …

I get a lot of emails from people asking for advice on some aspect of the methods used in microbiome studies.  In most cases, I probably do not know the ideal answer.  In addition, even when I do know the answer it would be useful to the community to have this posted publicly rather than in a private email.  So I am going to refer people to this post in the future and suggest they ask their question as a comment and then I can alert people to the question via social media, etc.

So – if you have any questions about microbiome methods – fire away.

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4 thoughts on “Microbiome Methods Q & A – submit your questions …

  1. What are the major differences between and among mehods, and is there any reasonable way to compare results from studies that used different methods?

    1. Well, I am not sure which methods you specifically are referring to here but some general comments:

      1. If you want to compared results between studies, the best thing to do is to use comparable / identical methods. This is one of the reasons there have been efforts to develop standards for microbiome studies. See for example work on this from NIST https://www.nist.gov/programs-projects/microbiome-community-measurements or work from the Earth Microbiome project http://www.earthmicrobiome.org/protocols-and-standards/.

      2. Differences in every step in a microbiome study – from design, to sampling, to data collection, to data analysis can make it difficult to make comparisons between the results of a study. This is one of the reasons why it is helpful for people to release their raw data – this would allow at least in theory people to compare data use the same analysis rather than comparing results from different analyses.

      3. As for major differences I think some of the biggest factors for DNA based microbiome studies are sample storage, DNA extraction, and DNA amplification protocols (if amplification is used). For example, if the data you are looking at involved PCR amplification and sequencing (e.g., rRNA gene amplicon studies) then differences in PCR primers and PCR conditions can lead to big differences that make it difficult to compare studies. For DNA based studies, differences in DNA extraction and storage protocols can lead to significant differences in inferences about microbial communities. This issue is discussed in some of the comments on this post: https://www.microbe.net/2015/01/31/best-practices-for-sample-storage-prior-to-microbiome-dna-analysis-freeze-buffer-process/.

  2. I was completely sold on using a mock microbial community to help with downstream sequence/data analyses (I have used the one which was available from BEI Resources, Manassas, VA, USA; https://www.beiresources.org/Catalog/otherProducts/HM-783D.aspx. It’s no longer available, although it’s not clear on the website why). However, after reading this article I’m no longer quite so convinced: https://www.biorxiv.org/content/early/2017/07/03/155358. So how is the best way to add a ‘community control’, and how would this differ from ‘spike-ins’?

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