title: Direct 16S rRNA-seq from bacterial communities: a PCR-independent approach to simultaneously assess microbial diversity and functional activity potential of each taxon.
abstract: The analysis of environmental microbial communities has largely relied on a PCR-dependent amplification of genes entailing species identity as 16S rRNA. This approach is susceptible to biases depending on the level of primer matching in different species. Moreover, possible yet-to-discover taxa whose rRNA could differ enough from known ones would not be revealed. DNA-based methods moreover do not provide information on the actual physiological relevance of each taxon within an environment and are affected by the variable number of rRNA operons in different genomes. To overcome these drawbacks we propose an approach of direct sequencing of 16S ribosomal RNA without any primer- or PCR-dependent step. The method was tested on a microbial community developing in an anammox bioreactor sampled at different time-points. A conventional PCR-based amplicon pyrosequencing was run in parallel. The community resulting from direct rRNA sequencing was highly consistent with the known biochemical processes operative in the reactor. As direct rRNA-seq is based not only on taxon abundance but also on physiological activity, no comparison between its results and those from PCR-based approaches can be applied. The novel principle is in this respect proposed not as an alternative but rather as a complementary methodology in microbial community studies.
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Riccardo
authors: R
authors: Rosselli
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Ottavia
authors: O
authors: Romoli
authors: Department of Biotechnology, University of Verona, Verona 37134, Italy.
authors: Nicola
authors: N
authors: Vitulo
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Alessandro
authors: A
authors: Vezzi
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Stefano
authors: S
authors: Campanaro
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Fabio
authors: F
authors: de Pascale
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Riccardo
authors: R
authors: Schiavon
authors: RWL Water/Eurotec WTT, Water Treatment Technologies, S.r.l., Padova, Italy.
authors: Maurizio
authors: M
authors: Tiarca
authors: RWL Water/Eurotec WTT, Water Treatment Technologies, S.r.l., Padova, Italy.
authors: Fabio
authors: F
authors: Poletto
authors: Department of Agronomy Animals, Food, Natural Resources and Environment, DAFNAE, University of Padova, Legnaro (Padova) Italy.
authors: Giuseppe
authors: G
authors: Concheri
authors: Department of Biology, University of Padova, Padova, Italy.
authors: Giorgio
authors: G
authors: Valle
authors: Department of Agronomy Animals, Food, Natural Resources and Environment, DAFNAE, University of Padova, Legnaro (Padova) Italy.
authors: Andrea
authors: A
authors: Squartini
mesh_terms: D001419:Bacteria / Q000145:classification / Q000235:genetics; D059013:Microbial Consortia / Q000235:genetics*; D009876:Operon; D012329:RNA, Bacterial / Q000235:genetics*; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D017423:Sequence Analysis, RNA / Q000379:methods*
chemical_list: D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S