High throughput ANI analysis of 90K prokaryotic genomes reveals clear species boundaries
Authors: Srinivas Aluru
Luis M Rodriguez-R
Adam M Phillippy
Chirag Jain
Konstantinos T Konstantinidis
Journal: Nature Communications (2018)
Discipline: Multidisciplinary
Keywords: microbiology; genetic diversity; genomes; species boundaries; Whole-genome similarity metrics
Affiliations: School of Computational Science and Engineering, Georgia Institute of Technology, Atlanta, GA, 30332, USA., National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, 20894, USA., School of Civil and Environmental Engineering, Georgia Institute of Technology, Atlanta, GA, 30332, USA., School of Biological Sciences, Georgia Institute of Technology, Atlanta, GA, 30332, USA., School of Civil and Environmental Engineering, Georgia Institute of Technology, Atlanta, GA, 30332, USA. ., School of Biological Sciences, Georgia Institute of Technology, Atlanta, GA, 30332, USA. ., School of Computational Science and Engineering, Georgia Institute of Technology, Atlanta, GA, 30332, USA. ., Institute for Data Engineering and Science, Georgia Institute of Technology, Atlanta, GA, 30332, USA. .

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 &gt;95% intra-species and &lt;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. 

Retracted: No

PubPeer comments:
 Readers should become aware of a recent preprint entitled  There is no evidence of a universal genetic boundary among microbial species that calls into question part of the conclusions of Jain et al., 2018 paper. From the Abstract:  A fundamental question in studying microbial diversity is whether there is a species boundary and if the boundary can be delineated by a universal genetic discontinuity. To address this question, Jain et al. computed the pairwise average nucleotide identity (ANI) of 91,761 microbial (bacterial and archaeal) genomes (the 90K genome dataset) and found that the ANI values from the 8 billion comparisons follow a strong bimodal distribution. The authors concluded that a clear genetic discontinuum and species boundary were evident from the unprecedented large-scale ANI analysis. As a result, the researchers advocated that an ANI of 95% can be used to accurately demarcate all currently named microbial species. **While the FastANI program described in the paper is useful, we argue that the paper s conclusion of a universal genetic boundary is questionable and resulted from the substantial biased sampling in genome sequencing. We also caution against being overly confident in using 95% ANI for microbial species delineation as the high benchmarks reported in the paper were inflated by using highly redundant genomes**. , 
 #1 A reply to the mentioned article (Re-evaluating the evidence for a universal genetic boundary among microbial species; https: doi.org 10.1038 s41467-021-24128-2) has been published: Reply to: u201cRe-evaluating the evidence for a universal genetic boundary among microbial speciesu201d ,