title: Community fingerprinting in a sequencing world.
abstract: Despite decreasing costs, generating large-scale, well-replicated and multivariate microbial ecology investigations with sequencing remains an expensive and time-consuming option. As a result, many microbial ecology investigations continue to suffer from a lack of appropriate replication. We evaluated two fingerprinting approaches - terminal restriction fragment length polymorphism (T-RFLP) and automated ribosomal intergenic spacer analysis (ARISA) against 454 pyrosequencing, by applying them to 225 polar soil samples from East Antarctica and the high Arctic. By incorporating local and global spatial scales into the dataset, our aim was to determine whether various approaches differed in their ability and hence utility, to identify ecological patterns. Through the reduction in the 454 sequencing data to the most dominant OTUs, we revealed that a surprisingly small proportion of abundant OTUs (< 0.25%) was driving the biological patterns observed. Overall, ARISA and T-RFLP had a similar capacity as sequencing to separate samples according to distance at a local scale, and to correlate environmental variables with microbial community structure. Pyrosequencing had a greater resolution at the global scale but all methods were capable of significantly differentiating the polar sites. We conclude fingerprinting remains a legitimate approach to generating large datasets as well as a cost-effective rapid method to identify samples for elucidating taxonomic information or diversity estimates with sequencing methods.
authors.affiliation: School of Biotechnology and Biomolecular Sciences, UNSW Australia, Randwick, NSW, Australia.
authors.forename: Josie
authors.initials: J
authors.lastname: van Dorst
authors.forename: Andrew
authors.initials: A
authors.lastname: Bissett
authors.forename: Anne S
authors.initials: AS
authors.lastname: Palmer
authors.forename: Mark
authors.initials: M
authors.lastname: Brown
authors.forename: Ian
authors.initials: I
authors.lastname: Snape
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Stark
authors.forename: Ben
authors.initials: B
authors.lastname: Raymond
authors.forename: John
authors.initials: J
authors.lastname: McKinlay
authors.forename: Mukan
authors.initials: M
authors.lastname: Ji
authors.forename: Tristrom
authors.initials: T
authors.lastname: Winsley
authors.forename: Belinda C
authors.initials: BC
authors.lastname: Ferrari
mesh_terms: D000864:Antarctic Regions; D001110:Arctic Regions; D001419:Bacteria / Q000145:classification / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D058889:Molecular Typing; D010802:Phylogeny; D012150:Polymorphism, Restriction Fragment Length; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology
chemical_list: D004269:DNA, Bacterial; D021903:DNA, Ribosomal Spacer
keywords: ARISA; Antarctica; Arctic; T-RFLP; microbial ecology; polar soils