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You're Not Supposed to Be There

Contaminated kits and reagents may have affected the results of many microbiome analyses, according to a new BMC Biology paper from the University of Aberdeen's Alan Walker and colleagues.

The researchers report that DNA extraction kits and other laboratory reagents are often contaminated with soil and skin bacteria. They sent aliquots from serial dilutions of a pure culture of Salmonella bongori to three UK institutes for 16S rRNA gene sequence profiling. Only S. bongori was identified in the undiluted samples, but the diluted samples contained other bacteria as well. By the fifth serial dilution, contaminating bacteria were dominant.

Similarly, shotgun metagenomic analysis of undiluted and diluted S. bongori aliquots processed using four different kits also turned up contamination, and that contaminants became more dominant players as the samples were increasingly diluted.

Though the contaminating bacteria varied by site and kit, Walker and his colleagues note that many of them were soil or water-dwelling bacteria.

Additionally, they say that while contamination is an issue for all studies, it is particularly worrisome for studies examining rare microbes or ones studying low-biomass samples. They suggest that researchers include negative control samples in their studies.

"I think many people are still surprised that if you put nothing into your sequencing pipeline, you come out with something that looks like a well-ordered microbiome," co-author Nick Loman from the University of Birmingham tells Ed Yong at Not Exactly Rocket Science. "If you talk about the old guard, they'll say that contamination has been a problem since day one. But the generation of scientists who are now furiously engaged in microbiome research needs to relearn the lessons of the past. You've got to assume that your results might be explained by a technical factor unless you've ruled it out."

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