02088nas a2200253   4500008004100000245008500041210006900126300001300195490000700208520131900215653001001534653003101544653001901575653001401594100001601608700001601624700002201640700001401662700001901676700001701695700001701712700001801729856008701747       2015                        eng d00aScraping the bottom of the barrel: are rare high throughput sequences artifacts?0 aScraping the bottom of the barrel are rare high throughput seque  a221 -2250 v133 a<p>Metabarcoding data generated using next-generation sequencing (NGS) technologies are overwhelmed with rare taxa and skewed in Operational Taxonomic Unit (OTU) frequencies comprised of few dominant taxa. Low frequency OTUs comprise a rare biosphere of singleton and doubleton OTUs, which may include many artifacts. We present an in-depth analysis of global singletons across sixteen NGS libraries representing different ribosomal RNA gene regions, NGS technologies and chemistries. Our data indicate that many singletons (average of 38 % across gene regions) are likely artifacts or potential artifacts, but a large fraction can be assigned to lower taxonomic levels with very high bootstrap support (&sim;32 % of sequences to genus with &ge;90 % bootstrap cutoff). Further, many singletons clustered into rare OTUs from other datasets highlighting their overlap across datasets or the poor performance of clustering algorithms. These data emphasize a need for caution when discarding rare sequence data en masse: such practices may result in throwing the baby out with the bathwater, and underestimating the biodiversity. Yet, the rare sequences are unlikely to greatly affect ecological metrics. As a result, it may be prudent to err on the side of caution and omit rare OTUs prior to downstream analyses.</p>10afungi10aHigh-throughput sequencing10aRare biosphere10aSingleton1 aBrown, S.P.1 aVeach, A.M.1 aRigdon-Huss, A.R.1 aGrond, K.1 aLickteig, S.K.1 aLothamer, K.1 aOliver, A.K.1 aJumpponen, A.  uhttps://www.sciencedirect.com/science/article/abs/pii/S1754504814001068?via%3Dihub