Shifting the genomic gold standard for the prokaryotic species definition

Abstract

DNA-DNA hybridization (DDH) has been used for nearly 50 years as the gold standard for prokaryotic species circumscriptions at the genomic level. It has been the only taxonomic method that offered a numerical and relatively stable species boundary, and its use has had a paramount influence on how the current classification has been constructed. However, now, in the era of genomics, DDH appears to be an outdated method for classification that needs to be substituted. The average nucleotide identity (ANI) between two genomes seems the most promising method since it mirrors DDH closely. Here we examine the work package JSpecies as a user-friendly, biologist-oriented interface to calculate ANI and the correlation of the tetranucleotide signatures between pairwise genomic comparisons. The results agreed with the use of ANI to substitute DDH, with a narrowed boundary that could be set at approximately 95-96%. In addition, the JSpecies package implemented the tetranucleotide signature correlation index, an alignment-free parameter that generally correlates with ANI and that can be of help in deciding when a given pair of organisms should be classified in the same species. Moreover, for taxonomic purposes, the analyses can be produced by simply randomly sequencing at least 20% of the genome of the query strains rather than obtaining their full sequence.

Fig. 1.

Plotted results of ANIb versus ANIm. The triangles show those values that correspond to what taxonomists consider as “true” species according to the DDH values traditionally applied and that have previously been classified. Inset shows the regression lines of the pairwise comparisons of ANIb or ANIm values with their corresponding percentage of aligned stretches (percentage of nucleotides included in the study).

Fig. 2.

Plotted values of DDH versus ANIm. Triangles show values that correspond to what taxonomists consider as “true” species according to the DDH values traditionally applied and that have previously been classified. Squares indicate values that appear to be in the transition zone.

Fig. 3.

Plotted values of TETRA versus ANIm. Triangles show those values that correspond to what taxonomists consider as “true” species according to the DDH values traditionally applied and that have previously been classified. Squares indicate values that appear to be in the transition zone. Note that this is an enlarged (zoomed) portion of the graphic, and values <80% ANI and 0.90 TETRA have been skipped.