High Throughput ANI Analysis of 90K Prokaryotic Genomes Reveals Clear Species Boundaries

Overview

Journal Nat Commun

Specialty Biology

Date 2018 Dec 4

PMID 30504855

Citations 2056

Authors

Chirag Jain

Luis M Rodriguez-R

Adam M Phillippy

Konstantinos T Konstantinidis

Srinivas Aluru

Affiliations

Soon will be listed here.

Abstract

A fundamental question in microbiology is whether there is continuum of genetic diversity among genomes, or clear species boundaries prevail instead. Whole-genome similarity metrics such as Average Nucleotide Identity (ANI) help address this question by facilitating high resolution taxonomic analysis of thousands of genomes from diverse phylogenetic lineages. To scale to available genomes and beyond, we present FastANI, a new method to estimate ANI using alignment-free approximate sequence mapping. FastANI is accurate for both finished and draft genomes, and is up to three orders of magnitude faster compared to alignment-based approaches. We leverage FastANI to compute pairwise ANI values among all prokaryotic genomes available in the NCBI database. Our results reveal clear genetic discontinuity, with 99.8% of the total 8 billion genome pairs analyzed conforming to >95% intra-species and <83% inter-species ANI values. This discontinuity is manifested with or without the most frequently sequenced species, and is robust to historic additions in the genome databases.

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References

Wilmes P, Simmons S, Denef V, Banfield J . The dynamic genetic repertoire of microbial communities. FEMS Microbiol Rev. 2008; 33(1):109-32. PMC: 2704941. DOI: 10.1111/j.1574-6976.2008.00144.x. View

Berlin K, Koren S, Chin C, Drake J, Landolin J, Phillippy A . Assembling large genomes with single-molecule sequencing and locality-sensitive hashing. Nat Biotechnol. 2015; 33(6):623-30. DOI: 10.1038/nbt.3238. View

Mende D, Sunagawa S, Zeller G, Bork P . Accurate and universal delineation of prokaryotic species. Nat Methods. 2013; 10(9):881-4. DOI: 10.1038/nmeth.2575. View

Olm M, Brown C, Brooks B, Banfield J . dRep: a tool for fast and accurate genomic comparisons that enables improved genome recovery from metagenomes through de-replication. ISME J. 2017; 11(12):2864-2868. PMC: 5702732. DOI: 10.1038/ismej.2017.126. View

Edgar R . Search and clustering orders of magnitude faster than BLAST. Bioinformatics. 2010; 26(19):2460-1. DOI: 10.1093/bioinformatics/btq461. View

Yarza P, Yilmaz P, Pruesse E, Glockner F, Ludwig W, Schleifer K . Uniting the classification of cultured and uncultured bacteria and archaea using 16S rRNA gene sequences. Nat Rev Microbiol. 2014; 12(9):635-45. DOI: 10.1038/nrmicro3330. View

Koren S, Walenz B, Berlin K, Miller J, Bergman N, Phillippy A . Canu: scalable and accurate long-read assembly via adaptive -mer weighting and repeat separation. Genome Res. 2017; 27(5):722-736. PMC: 5411767. DOI: 10.1101/gr.215087.116. View

Konstantinidis K, Ramette A, Tiedje J . Toward a more robust assessment of intraspecies diversity, using fewer genetic markers. Appl Environ Microbiol. 2006; 72(11):7286-93. PMC: 1636164. DOI: 10.1128/AEM.01398-06. View

Cohan F . Bacterial species and speciation. Syst Biol. 2002; 50(4):513-24. DOI: 10.1080/10635150118398. View

10.

Richter M, Rossello-Mora R . Shifting the genomic gold standard for the prokaryotic species definition. Proc Natl Acad Sci U S A. 2009; 106(45):19126-31. PMC: 2776425. DOI: 10.1073/pnas.0906412106. View

11.

Konstantinidis K, Tiedje J . Genomic insights that advance the species definition for prokaryotes. Proc Natl Acad Sci U S A. 2005; 102(7):2567-72. PMC: 549018. DOI: 10.1073/pnas.0409727102. View

12.

Luo C, Walk S, Gordon D, Feldgarden M, Tiedje J, Konstantinidis K . Genome sequencing of environmental Escherichia coli expands understanding of the ecology and speciation of the model bacterial species. Proc Natl Acad Sci U S A. 2011; 108(17):7200-5. PMC: 3084108. DOI: 10.1073/pnas.1015622108. View

13.

Lee I, Kim Y, Park S, Chun J . OrthoANI: An improved algorithm and software for calculating average nucleotide identity. Int J Syst Evol Microbiol. 2015; 66(2):1100-1103. DOI: 10.1099/ijsem.0.000760. View

14.

Ivanova N, Sorokin A, Anderson I, Galleron N, Candelon B, Kapatral V . Genome sequence of Bacillus cereus and comparative analysis with Bacillus anthracis. Nature. 2003; 423(6935):87-91. DOI: 10.1038/nature01582. View

15.

Straub T, Zhaxybayeva O . A null model for microbial diversification. Proc Natl Acad Sci U S A. 2017; 114(27):E5414-E5423. PMC: 5502593. DOI: 10.1073/pnas.1619993114. View

16.

Kurtz S, Phillippy A, Delcher A, Smoot M, Shumway M, Antonescu C . Versatile and open software for comparing large genomes. Genome Biol. 2004; 5(2):R12. PMC: 395750. DOI: 10.1186/gb-2004-5-2-r12. View

17.

Varghese N, Mukherjee S, Ivanova N, Konstantinidis K, Mavrommatis K, Kyrpides N . Microbial species delineation using whole genome sequences. Nucleic Acids Res. 2015; 43(14):6761-71. PMC: 4538840. DOI: 10.1093/nar/gkv657. View

18.

Kim M, Oh H, Park S, Chun J . Towards a taxonomic coherence between average nucleotide identity and 16S rRNA gene sequence similarity for species demarcation of prokaryotes. Int J Syst Evol Microbiol. 2014; 64(Pt 2):346-351. DOI: 10.1099/ijs.0.059774-0. View

19.

Guy L, Kultima J, Andersson S . genoPlotR: comparative gene and genome visualization in R. Bioinformatics. 2010; 26(18):2334-5. PMC: 2935412. DOI: 10.1093/bioinformatics/btq413. View

20.

Shapiro B, Friedman J, Cordero O, Preheim S, Timberlake S, Szabo G . Population genomics of early events in the ecological differentiation of bacteria. Science. 2012; 336(6077):48-51. PMC: 3337212. DOI: 10.1126/science.1218198. View