Important paper on how biases in DNA extraction can shape inferences from metagenomics (re spore formers)

A moderately new paper is out that is an excellent example of how biases in DNA extraction can have major impacts on inferences from culture independent DNA studies. The paper is in what I think is a generally non open access journal (Computational and Structural Biotechnology Journal) for fortunately has been made open access

Source: Under-detection of endospore-forming Firmicutes in metagenomic dataComput Struct Biotechnol J. 2015 Apr 25;13:299-306. doi: 10.1016/j.csbj.2015.04.002. eCollection 2015. Filippidou S, Junier T, Wunderlin T, Lo CC, Li PE, Chain PS, Junier P.

Fig. 2. Profile similarity hits for Spo0A and Gpr protein profiles in metagenomes from different origins. The color code identifying different environments is presented under the results. The genomes included in profile testing (see Fig. 1A) were also included in the analysis and are presented in white (endospore-formers) and gray (non-spore formers).


Microbial diversity studies based on metagenomic sequencing have greatly enhanced our knowledge of the microbial world. However, one caveat is the fact that not all microorganisms are equally well detected, questioning the universality of this approach. Firmicutes are known to be a dominant bacterial group. Several Firmicutes species are endospore formers and this property makes them hardy in potentially harsh conditions, and thus likely to be present in a wide variety of environments, even as residents and not functional players. While metagenomic libraries can be expected to contain endospore formers, endospores are known to be resilient to many traditional methods of DNA isolation and thus potentially undetectable. In this study we evaluated the representation of endospore-forming Firmicutes in 73 published metagenomic datasets using two molecular markers unique to this bacterial group (spo0A and gpr). Both markers were notably absent in well-known habitats of Firmicutes such as soil, with spo0A found only in three mammalian gut microbiomes. A tailored DNA extraction method resulted in the detection of a large diversity of endospore-formers in amplicon sequencing of the 16S rRNA and spo0A genes. However, shotgun classification was still poor with only a minor fraction of the community assigned to Firmicutes. Thus, removing a specific bias in a molecular workflow improves detection in amplicon sequencing, but it was insufficient to overcome the limitations for detecting endospore-forming Firmicutes in whole-genome metagenomics. In conclusion, this study highlights the importance of understanding the specific methodological biases that can contribute to improve the universality of metagenomic approaches.

I noticed this paper a few months ago and posted about it to Twitter:

But only now getting around to blogging about it. This is definitely worth a look.

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Jonathan Eisen

I am an evolutionary biologist and a Professor at U. C. Davis. My lab is in the UC Davis Genome Center and I hold appointments in the Department of Medical Microbiology and Immunology in the School of Medicine and the Department of Evolution and Ecology in the College of Biological Sciences. My research focuses on the origin of novelty (how new processes and functions originate). To study this I focus on sequencing and analyzing genomes of organisms, especially microbes and using phylogenomic analysis (see my lab site here which has more information on lab activities).  In addition to research, I am heavily involved in the Open Access publishing and Open Science movements.