The dangers of not thinking about the relics


No, I’m not talking about Saint Erik’s skull from his beheading in 1160, which (as it turns out) might just be real. A hot topic in microbiology is a different kind of relic.

Amazing developments in microbial ecology in recent years have really opened up a new window to understanding the diversity of microbial communities and uncultivated organisms that are hanging out around us. As we continue to develop more affordable sequencing methods (and if you happen to sequence things, please take a few minutes to fill out a survey we posted earlier), the availability of sequence data has absolutely exploded.
Microbes play an important role in the environment, and linking specific lineages to what is actually happening involves figuring out who is metabolically active. Some studies previously been assumed that metagenomic data sets represent DNA from the living cells that drive biogeochemical processes, however, this is not the case.
A recent preprint posted by Paul Carini and colleagues, highlights the problems that arise due to what has been termed ‘relic DNA’, which refers to DNA that from dead cells. While this paper has not yet been peer reviewed, it was posted on bioRxiv, which is a cool way to make findings immediately available, and get feedback before submission. It’s been shown previously that extracellular DNA can hang out in the environment for multiple years, depending on specific environmental parameters.
I think their “one sentence” summary gets to the point pretty quickly…
 “Soils can harbor substantial amounts of DNA from dead microbial cells; this, ‘relic’ DNA inflates estimates of microbial diversity and obscures assessments of community structure.”
Samples were collected and divided into two subsamples, one of which had propidium monoazide (PMA) (which intercalates into free DNA / DNA in dead cells) added. When the sample with PMA is exposed to light, it covalently binds and modifies the DNA, inhibiting amplification by PCR. Basically, this study compared the total bacterial / fungal sequences obtained using qPCR in PMA treated and untreated samples. They were able to compare richness estimates, overall community composition, and taxon abundances between the two subsamples.
In total they analyzed 31 soils, and they found that this analysis revealed that about 14% of the taxa originally identified in the sequences were only in the relic DNA. In 24 of the samples, the richness decreased a ton more, by up to 55% even! How long has that DNA been there, and where did it come from in the first 31Fig1place? The authors suggest that it could represent microbes that used to live in the environment but no longer are alive due to changes in conditions; alternatively they could have died after transport from a different environment.
The big ‘take home’ messages were:

  • Actual microbial diversity in soil is lower than what is usually reported
  • Relic DNA might make it hard to really see any changes in seasonal changes of microbial communities or effects due to stuff like climate change.
  • This is alllll problemenatic when trying to figure stuff out about metagenomes.31Fig2

Removing this relic DNA before sequencing might give us a better / different window on the microbial diversity found in soils. This is most likely a problem in other environments, such as deep sea sediments or maybe in studies focues on host-microbiota systems. While the issue of potential ‘relic DNA’ has been discussed previously, this paper is a nice study clearly demonstrating that we still have a long way to go to understand complex microbial communities and that we might have less of the uncultured majority to culture than we thought.
This article was also highlighted here, and I came across a fun conversation on twitter too.
Carini P, Marsden PJ, Leff JW, Morgan EE, Strickland MS, Fierer N. Relic DNA is abundant in soil and obscures estimates of soil microbial diversity. bioRxiv. 2016 Jan 1:043372.

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