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Biogeography and Phylogeny of Aigarchaeota, a Novel Phylum of Archaea Gisele Braga Goertz University of Nevada, Las Vegas

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Biogeography and Phylogeny of Aigarchaeota, a Novel Phylum of Archaea Gisele Braga Goertz University of Nevada, Las Vegas McNair Poster Presentations McNair Scholars Institute 2013 Biogeography and Phylogeny of Aigarchaeota, A Novel Phylum of Archaea Gisele Braga Goertz University of Nevada, Las Vegas Brian P. Hedlund University of Nevada, Las Vegas, [email protected] Follow this and additional works at: http://digitalscholarship.unlv.edu/mcnair_posters Part of the Genetics Commons Repository Citation Goertz, G. B., Hedlund, B. P. (2013). Biogeography and Phylogeny of Aigarchaeota, A Novel Phylum of Archaea. Available at: http://digitalscholarship.unlv.edu/mcnair_posters/33 This Poster is brought to you for free and open access by the McNair Scholars Institute at Digital Scholarship@UNLV. It has been accepted for inclusion in McNair Poster Presentations by an authorized administrator of Digital Scholarship@UNLV. For more information, please contact [email protected]. Biogeography and Phylogeny of Aigarchaeota, A Novel Phylum of Archaea Gisele Braga Goertz, McNair Scholar, Biological Sciences Major Dr. Brian P. Hedlund, Faculty Mentor, School of Life Sciences • ‘Aigarchaeota’ gene sequences mined from GenBank and out-group sequences were aligned in mothur using a 50,000 character SILVA reference alignment capable of aligning sequences from all three domains of life. • Alignments were manually curated in BioEdit version 7.1.3.0, trimmed, then saved as Phylogenetic Infer- ence Package (PHYLIP) version 4-compatible file. GROUPS • A distance matrix was computed using “Dnadist”, then used in the “Neighbor” program, which clusters lineages and constructs a tree. 1A ABSTRACT • Sequences were subsequently evaluated at the species (>97%), genus (>95%), family (>90%), order 1B (>85%), and phylum (>80%) level. 1C 1D ‘Aigarchaeota’ is a candidate phylum of Archaea known only by 16S rRNA gene fragments from cultiva- 1E tion-independent microbial surveys and a single composite genome from Candidatus ‘Caldiarchaeum sub- PRIMER EVALUATION • ‘Aigarchaeota’-specific qRT-PCR primers were designed by three different programs the SP-Designer, 2A terraneum’, an inhabitant of a subterranean gold mine in Japan. Gene sequences associated with ‘Aigarchaeo- 2B ARB, and AlleleID. ta’ have been found in a variety of geothermal habits, however a comprehensive analysis of the phylogeny 3 and distribution of ‘Aigarchaeota’ has not yet been done. Public databases were mined for 16S rRNA gene • Primers were classified as forward or reverse and submitted to the Ribosomal Database Project (RDP) 4 sequences related to known ‘Aigarchaeota’ and a combination of approaches were used to rigorously define Probe Match tool to eliminate primers that were non- ‘Aigarchaeota’ specific. 5 the phylogenetic boundaries of the phylum, investigate its distribution, and design potential ‘Aigarchaeota’- • Primers that did match well with ‘Aigarchaeota’ were reviewed in RDP to determine which genus-level 6 specific primers. Approximately 300 16S rRNA genes and gene fragments affiliated with ‘Aigarchaeota’ were groups were associated with each of the individual primers. 7 identified, phylogenic analyses suggested that ‘Aigarchaeota’ belonged to at least three family- to order-level • Primers were analyzed using OligoCalc to ensure appropriate GC content (~50%), melting temperature 8 groups and at least 13 genus-level groups, and supported the proposed relationship between ‘Aigarchaeota’, (60°C), and absence of hairpin formation and potential self-annealing sites. Thaumarchaeota, Crenarchaeota, and Korarchaeota in the so-called ‘TACK’ superphylum. A global distri- RESULTS Figure 3. Global distribution of ‘Aigarchaeota’ sequences by genus-level groups. bution was suggested with genus-level group habitat differentiation. Some groups were predominantly ter- restrial (1A, 2B, 3, 4 & 5) while other groups were mostly found in Marine habitats (6-8). Fifteen strong ‘Ai- CONCLUSION garchaeota’-specific primer candidates for the polymerase chain reaction (PCR) amplification of 16S rRNA genes were designed for further analyses. This study revealed, for the first time, the phylogenetic diversity and distribution of the novel phylum ‘Ai- garchaeota’. Currently available 16S rRNA gene sequences from global studies suggest that the phylum con- INTRODUCTION sists of at least 13 genus-level groups, and that ‘Aigarchaeota’ is situated firmly in the ‘TACK’ superphylum, but exists as a separate, unique phylum that does not possess a phylum-level relationship with the Miscella- Despite our knowledge of the diversity of life on earth there are major lineages of microbial life that have neous Cren Group (see Figure 1). Genus-level Group 2A was detected in both terrestrial and marine environ- never been studied that are referred to as “biological dark matter” or “microbial dark matter” (Marcy et al., ments. In contrast, Groups 1A, 2B, 3, 4 & 5 and Groups 6-8 are predominant in terrestrial and marine habitats 2007; Rinke, et al., 2013). One of these so-called “microbial dark matter” groups was originally described respectively. Groups 1B and 1C were only found in terrestrial habitats, while Group 1D was only found in ma- as pSL4 and related gene sequences were later grouped under the name Hot Water Crenarchaeotic Group 1 rine habitats. Potential primers designed in this study range from genus-level group specific to a combination (HWCG 1) (Barns et al., 1996; Nunoura et al., 2005). A metagenomics study of a microbial mat community of groups allowing for the detection of all potential ‘Aigarchaeota’ defined in this study. It’s important to note led to the construction of a complete composite genome for a member of the pSL4/HWCG 1 lineage pro- that there were fewer numbers of marine studies (17) than terrestrial studies (54), thus making it difficult to posed as Candidatus ‘Caldiarchaeum subterraneum’ and further analysis of the genome led to the proposi- make solid inferences. tion of the phylum ‘Aigarchaeota’ (Nunoura et al., 2005; Nunoura et al., 2011). Genomic analysis of Ca. subterraneum revealed unique eukaryote-type features which led to the proposal REFERENCES of a superphylum level relationship called the ‘TACK’ superphylum (Guy and Ettema, 2011). The cultivation- Figure 1. Neighbor-Joining Tree showing 16S rRNA gene phylogenetic relationship between ‘Aigarchaeota’ independent studies suggest a world-wide distribution of ‘Aigarchaeota’ and dominance of 16S rRNA gene Groups 1A – 8 and archaeal outgroups, with unsupported nodes identified with boxes. Scale bar represents one Barns SM, Delwiche CF, Palmer JD, Pace NR (1996) Perspectives on archaeal diversity, thermophily and monophyly from environ- clone libraries from habitats such as a deep sea hydrothermal vent in Okinawa Trough at 35-60 °C and Great substitution per 100 nucleotides. mental rRNA sequences. Proc. Natl. Acad. Sci. USA 93:9188–9193 Boiling Spring at 79-87°C ( Nunoura et al., 2010; Cole et al., 2012). Cole at al 2012 reported that ‘Aigar- Cole JK, Peacock JP, Dodsworth JA, Williams AJ, Thompson DB, Dong H, Wu G, Hedlund BP (2012) Sediment microbial communi- chaeota’ 16S rRNA gene sequences had the highest relative abundance at Site A (~54%), with a general trend ties in Great Boiling Spring are controlled by temperature and distinct from water communities. ISME J. 7: 718–729. LEGEND (°C) of decreasing relative abundance down to ~5% at 62°C (Site E), suggesting ‘Aigarchaeota’ niche differentia- <15 tion along a thermal gradient. Guy L & Ettema TJG (2011). The archaeal ‘TACK’ superphylum and the origin of eukaryotes. 15-50 Trends Microbiol. 19: 580–587. The purpose of this study was to gather 16S rRNA gene sequences potentially belonging to‘Aigarchaeota’ , 50-80 and to use those sequences to address several goals: >80 Marcy Y, Ouverney C, Bik EM, Lösekann T, Ivanova N, Martin HG, Szeto E, Platt D, Hugenholtz P, Relman DA, Quake SR (2007) (i) Rigorously define the phylum Unpublished/not available Dissecting biological “dark matter” with single-cell genetic analysis of rare and uncultivated TM7 microbes from the human mouth. (ii) Gain insight into the phylogenetic and potential taxonomic structure of the phylum Proceedings of the National Academy of Science 104: 11889-11894. Print. (iii) Assess the distribution of ‘Aigarchaeota’ as a whole, along with individual groups of ‘Aigarchaeota’ in Number of Gene Sequences Reported Nunoura T, Hirayama H, Takami H, Oida H, Nishi S, Shimamura S, Suzuki Y, Inagaki F, Takai K, Nealson KH, Horikoshi K (2005) nature Genetic and functional properties of uncultivated thermophilic crenarchaeotes from a subsurface gold mine as revealed by analysis of (iv) Design ‘Aigarchaeota’- specific 16S rRNA gene primers that combined will target genus-level groups 1-5 genome fragments. Environ. Microbiol. 7:1967–1984. 6-10 Nunoura T, Oida H, Nakaseama M, Kosaka A, Ohkubo SB, Kikuchi T et al. (2010). Archaeal diversity and distribution along thermal 11-25 and geochemical gradients in hydrothermal sediments at the Yonaguni Knoll IV Hydrothermal Field in the Southern Okinawa Trough. Appl Environ Microbiol. 76: 1198–1211 26-50 Nunoura T, Takaki Y, Kakuta J, Nishi S, Sugahara J, Kazama H, Chee GJ, Hattori M, Kanai A, Atomi H, Takai K, Takami H (2011) In- METHODOLOGY 51-75 sights into the evolution of archaea and eukaryotic protein modifier systems revealed by the genome of a novel archaeal group. Nucleic Acids Research 39: 3204-3223. PHYLOGENETIC TREE CONSTRUCTION Figure 2. ‘Aigarchaeota’ sequences organized to represent global sample location, sampling temperature, number of Rinke C, Schwientek P, Sczyrba A, Ivanova N, Anderson IJ, Cheng JF, Darling A, Malfatti S, Swan BK, Gies EA, Dodsworth JA, Hedlund sequences per location, and associated genus-level groups. BP, Tsiamis G, Sievert SM, Liu WT, Eisen JA, Hallam SJ, Kyrpides NC, Stepanauskas R, Rubin EM, Hugenholtz P, Woyke T, (2013 • The 16S rRNA gene sequence of Ca. ‘C. subterraneum’ was submitted to BLAST, and the accession num- Insights into the phylogeny and coding potential of microbial dark matter.Nature 00: 1-7 bers of all gene sequences with greater than 80% identity were recorded..
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