title: Capturing single cell genomes of active polysaccharide degraders: an unexpected contribution of Verrucomicrobia.
abstract: Microbial hydrolysis of polysaccharides is critical to ecosystem functioning and is of great interest in diverse biotechnological applications, such as biofuel production and bioremediation. Here we demonstrate the use of a new, efficient approach to recover genomes of active polysaccharide degraders from natural, complex microbial assemblages, using a combination of fluorescently labeled substrates, fluorescence-activated cell sorting, and single cell genomics. We employed this approach to analyze freshwater and coastal bacterioplankton for degraders of laminarin and xylan, two of the most abundant storage and structural polysaccharides in nature. Our results suggest that a few phylotypes of Verrucomicrobia make a considerable contribution to polysaccharide degradation, although they constituted only a minor fraction of the total microbial community. Genomic sequencing of five cells, representing the most predominant, polysaccharide-active Verrucomicrobia phylotype, revealed significant enrichment in genes encoding a wide spectrum of glycoside hydrolases, sulfatases, peptidases, carbohydrate lyases and esterases, confirming that these organisms were well equipped for the hydrolysis of diverse polysaccharides. Remarkably, this enrichment was on average higher than in the sequenced representatives of Bacteroidetes, which are frequently regarded as highly efficient biopolymer degraders. These findings shed light on the ecological roles of uncultured Verrucomicrobia and suggest specific taxa as promising bioprospecting targets. The employed method offers a powerful tool to rapidly identify and recover discrete genomes of active players in polysaccharide degradation, without the need for cultivation.
authors.affiliation: Bigelow Laboratory for Ocean Sciences, West Boothbay Harbor, Main, United States of America.
authors.forename: Manuel
authors.initials: M
authors.lastname: Martinez-Garcia
authors.forename: David M
authors.initials: DM
authors.lastname: Brazel
authors.forename: Brandon K
authors.initials: BK
authors.lastname: Swan
authors.forename: Carol
authors.initials: C
authors.lastname: Arnosti
authors.forename: Patrick S G
authors.initials: PS
authors.lastname: Chain
authors.forename: Krista G
authors.initials: KG
authors.lastname: Reitenga
authors.forename: Gary
authors.initials: G
authors.lastname: Xie
authors.forename: Nicole J
authors.initials: NJ
authors.lastname: Poulton
authors.forename: Monica
authors.initials: M
authors.lastname: Lluesma Gomez
authors.forename: Dashiell E D
authors.initials: DE
authors.lastname: Masland
authors.forename: Brian
authors.initials: B
authors.lastname: Thompson
authors.forename: Wendy K
authors.initials: WK
authors.lastname: Bellows
authors.forename: Kai
authors.initials: K
authors.lastname: Ziervogel
authors.forename: Chien-Chi
authors.initials: CC
authors.lastname: Lo
authors.forename: Sanaa
authors.initials: S
authors.lastname: Ahmed
authors.forename: Cheryl D
authors.initials: CD
authors.lastname: Gleasner
authors.forename: Chris J
authors.initials: CJ
authors.lastname: Detter
authors.forename: Ramunas
authors.initials: R
authors.lastname: Stepanauskas
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D044602:Cellulases / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence; D005456:Fluorescent Dyes / Q000378:metabolism; D005618:Fresh Water / Q000382:microbiology; D016680:Genome, Bacterial; D005936:Glucans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010802:Phylogeny; D011134:Polysaccharides / Q000378:metabolism*; D012623:Seawater / Q000382:microbiology; D017422:Sequence Analysis, DNA; D059010:Single-Cell Analysis; D061327:Verrucomicrobia / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D014871:Water Microbiology; D014990:Xylans / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D005936:Glucans; D011134:Polysaccharides; D014990:Xylans; C008247:laminaran; D044602:Cellulases