Using metagenomic analyses to estimate the consequences of enrichment bias for pathogen detection.

BACKGROUND

Enriching environmental samples to increase the probability of detection has been standard practice throughout the history of microbiology. However, by its very nature, the process of enrichment creates a biased sample that may have unintended consequences for surveillance or resolving a pathogenic outbreak. With the advent of next-generation sequencing and metagenomic approaches, the possibility now exists to quantify enrichment bias at an unprecedented taxonomic breadth.

FINDINGS

We investigated differences in taxonomic profiles of three enriched and unenriched tomato phyllosphere samples taken from three different tomato fields (n = 18). 16S rRNA gene meteganomes were created for each of the 18 samples using 454/Roche's pyrosequencing platform, resulting in a total of 165,259 sequences. Significantly different taxonomic profiles and abundances at a number of taxonomic levels were observed between the two treatments. Although as many as 28 putative Salmonella sequences were detected in enriched samples, there was no significant difference in the abundance of Salmonella between enriched and unenriched treatments.

CONCLUSIONS

Our results illustrate that the process of enriching greatly alters the taxonomic profile of an environmental sample beyond that of the target organism. We also found evidence suggesting that enrichment may not increase the probability of detecting a target. In conclusion, our results further emphasize the need to develop metagenomics as a validated culture independent method for pathogen detection.