title: Local diversity of heathland Cercozoa explored by in-depth sequencing.
abstract: Cercozoa are abundant free-living soil protozoa and quantitatively important in soil food webs; yet, targeted high-throughput sequencing (HTS) has not yet been applied to this group. Here we describe the development of a targeted assay to explore Cercozoa using HTS, and we apply this assay to measure Cercozoan community response to drought in a Danish climate manipulation experiment (two sites exposed to artificial drought, two unexposed). Based on a comparison of the hypervariable regions of the 18S ribosomal DNA of 193 named Cercozoa, we concluded that the V4 region is the most suitable for group-specific diversity analysis. We then designed a set of highly specific primers (encompassing ~270 bp) for 454 sequencing. The primers captured all major cercozoan groups; and >95% of the obtained sequences were from Cercozoa. From 443 350 high-quality short reads (>300 bp), we recovered 1585 operational taxonomic units defined by >95% V4 sequence similarity. Taxonomic annotation by phylogeny enabled us to assign >95% of our reads to order level and ~85% to genus level despite the presence of a large, hitherto unknown diversity. Over 40% of the annotated sequences were assigned to Glissomonad genera, whereas the most common individually named genus was the euglyphid Trinema. Cercozoan diversity was largely resilient to drought, although we observed a community composition shift towards fewer testate amoebae.
authors.affiliation: Section of Terrestrial Ecology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.|Section for Genetics and Evolutionary Biology, Department of Biosciences, University of Oslo, Oslo, Norway.
authors.forename: Christoffer Bugge
authors.initials: CB
authors.lastname: Harder
authors.affiliation: Section of Terrestrial Ecology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Regin
authors.initials: R
authors.lastname: Rønn
authors.affiliation: Section of Microbiology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Asker
authors.initials: A
authors.lastname: Brejnrod
authors.affiliation: Department of Life Sciences, The Natural History Museum, Cromwell Road, London, UK.|Centre for Environment, Fisheries and Aquaculture Science (Cefas), The Nothe, Weymouth, Dorset, UK.
authors.forename: David
authors.initials: D
authors.lastname: Bass
authors.affiliation: Section of Microbiology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Waleed Abu
authors.initials: WA
authors.lastname: Al-Soud
authors.affiliation: Section of Terrestrial Ecology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Flemming
authors.initials: F
authors.lastname: Ekelund
mesh_terms: D044822:Biodiversity; D056907:Cercozoa / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D059014:High-Throughput Nucleotide Sequencing; D010802:Phylogeny; D012987:Soil / Q000469:parasitology*
chemical_list: D017931:DNA Primers; D004275:DNA, Ribosomal; D012987:Soil