Using population genetic theory and DNA sequences for species detection and identification in asexual organisms

Abstract

Background: It is widely agreed that species are fundamental units of biology, but there is little agreement on a definition of species or on an operational criterion for delimiting species that is applicable to all organisms.

Methodology/principal findings: We focus on asexual eukaryotes as the simplest case for investigating species and speciation. We describe a model of speciation in asexual organisms based on basic principles of population and evolutionary genetics. The resulting species are independently evolving populations as described by the evolutionary species concept or the general lineage species concept. Based on this model, we describe a procedure for using gene sequences from small samples of individuals to assign them to the same or different species. Using this method of species delimitation, we demonstrate the existence of species as independent evolutionary units in seven groups of invertebrates, fungi, and protists that reproduce asexually most or all of the time.

Conclusions/significance: This wide evolutionary sampling establishes the general existence of species and speciation in asexual organisms. The method is well suited for measuring species diversity when phenotypic data are insufficient to distinguish species, or are not available, as in DNA barcoding and environmental sequencing. We argue that it is also widely applicable to sexual organisms.

Figure 1. Speciation as seen in phylogenetic trees of asexual organisms descended from a single asexual founder.

(A) A single species is an inclusive population that is limited to some number of individuals N by the carrying capacity of the environment and will show extinction of lineages due to stochastic processes. The effective population size N_e is a smaller number reflecting the fact that different individuals are producing different numbers of offspring. This results in clades (open bars) and singlets (circle) separated by transient gaps with an average depth of 2N_e generations. (B) A tree in which two lineages have been physically separated, e.g. by distance, or have undergone divergent selection for adaptation to different niches (or both) long enough to complete lineage sorting and become reciprocally monophyletic. The populations form two clusters separated by a gap that becomes deeper over time until it is much deeper than 2N_e generations.

Figure 2. Using phylogenetic analysis and the 4× rule to identify species.

If phylogenetic analysis shows that small samples from two populations are reciprocally monophyletic, and if the mean sequence difference between them is more than four times θ = 2N_eμ estimated from the within-sample variation, then the samples came from different species.

Figure 3. Bdelloid rotifer: Maximum Likelihood tree made with PAUP using the procedure described by Hall .

The tree was rooted with clones of Rotaria, because several analyses support a basal position for this genus. Pairwise distances were corrected for multiple hits using a general time reversible model with site-specific substitution rates for the first, second, and third codon positions. Colored circles and triangles respectively indicate clades and singlets that were identified as species using the procedure described in the text. Pending formal species descriptions, these species are given temporary names consisting of the abbreviated genus and a number. Adi = Adineta; Hab = Habrotrocha; Sce = Scepanotrocha; Abr = Abrochtha; Mac = Macrotrachela; Rot = Rotaria; Pha = Philodina. Species labeled McBS1 and SnoF1 were not identified and are named after their sources (McBeth Spring and Snowy Range site F). Dashed lines were supported by <50% of 1000 neighbor-joining bootstrap replicas, each with 10 random-addition replicates.

Figure 4. Bdelloid rotifers: frequency distribution of uncorrected pairwise sequence differences, in bins of 1%.

The differences within species are not corrected for multiple hits, which is appropriate because all are less than 10%. The differences between species are corrected for multiple hits using the TVM+I+G model selected by ModelTest .