title: MetaGaAP: A Novel Pipeline to Estimate Community Composition and Abundance from Non-Model Sequence Data.
abstract: Next generation sequencing and bioinformatic approaches are increasingly used to quantify microorganisms within populations by analysis of 'meta-barcode' data. This approach relies on comparison of amplicon sequences of 'barcode' regions from a population with public-domain databases of reference sequences. However, for many organisms relevant 'barcode' regions may not have been identified and large databases of reference sequences may not be available. A workflow and software pipeline, 'MetaGaAP,' was developed to identify and quantify genotypes through four steps: shotgun sequencing and identification of polymorphisms in a metapopulation to identify custom 'barcode' regions of less than 30 polymorphisms within the span of a single 'read', amplification and sequencing of the 'barcode', generation of a custom database of polymorphisms, and quantitation of the relative abundance of genotypes. The pipeline and workflow were validated in a 'wild type'Alphabaculovirusisolate,Helicoverpa armigerasingle nucleopolyhedrovirus (HaSNPV-AC53) and a tissue-culture derived strain (HaSNPV-AC53-T2). The approach was validated by comparison of polymorphisms in amplicons and shotgun data, and by comparison of predicted dominant and co-dominant genotypes with Sanger sequences. The computational power required to generate and search the database effectively limits the number of polymorphisms that can be included in a barcode to 30 or less. The approach can be used in quantitative analysis of the ecology and pathology of non-model organisms.
authors.affiliation: School of Earth, Environmental and Biological Sciences, Queensland University of Technology, Brisbane City QLD 4000, Australia. chris.noune@connect.qut.edu.au.
authors.forename: Christopher
authors.initials: C
authors.lastname: Noune
authors.affiliation: School of Earth, Environmental and Biological Sciences, Queensland University of Technology, Brisbane City QLD 4000, Australia. caroline.hauxwell@qut.edu.au.
authors.forename: Caroline
authors.initials: C
authors.lastname: Hauxwell
keywords: HaSNPV-AC53; MetaGaAP; baculoviruses; bioinformatics; community analysis; meta-barcoding; metapopulation