Discrimination, crypticity, and incipient taxa in entamoeba.

AMOEBAE and amoeboid protists have transitioned from studies based on single morphological traits (pseudopodia; Levine et al. 1980), to single gene (subunit rRNA -SSU rRNA; Adl et al. 2005; Burki and Pawlowski 2006; Pawlowski and Burki 2009; Stensvold et al. 2010, 2011), and to multigene (Bapteste et al. 2002; Pawlowski and Burki 2009) phylogenies at the supertaxon, supergroup, class, genus, or species level. Single gene analyses of metabolic traits (e.g. alcohol dehydrogenase adhe) contribute to conflictive phylogenetic depictions, due to horizontal acquisition (horizontal gene transfer -HGT) of genes from prokaryotes and unicellular eukaryotes (Andersson 2005; Andersson et al. 2006; Bapteste and Boucher 2009; Espinosa et al. 2001, 2009; Paz-y-Mino C. and Espinosa 2010). In On the Origin of Species (1859), Charles Darwin questioned whether the species concept reflected natural groups, “we shall have to treat species in the same manner as those naturalists treat genera, who admit that genera are merely artificial combinations made for convenience… we shall at least be freed from the vain search for the undiscovered and undiscoverable essence of the term species”. Persistent difficulties in resolving clear lineages have questioned the species concept to describe diverging populations of prokaryotes (Bacteria and Archaea, Pace 2006; Doolittle and Zhaxybayeva 2009) or unicellular eukaryotes (protistan polyphyletic groups, Finlay 2004). Although it is widely accepted that value of multiple integrated approaches would render ‘holistic taxonomies’ (Finlay 2004; Pawlowski and Burki 2009), most phylogenetic studies are still based on single-gene or morphological traits. Such methodologies conceal natural lineages, which are considered ‘cryptic’ (Mallet 2010). Stebbins (1950) defined cryptic species as “… population systems, which were believed to belong to the same species until genetic evidence showed the existence of isolating mechanisms separating them”. In protists, unresolved phylogenies generate heated debates to unravel ‘cryptic’ morphospecies (populations of unicellular eukaryotes with similar physiological properties and mating incompatibilities; Caron et al. 2009; Finlay 2004). Phylogenetic analyses would benefit from the integration of physiological, ecological, and behavioral studies. Most phylogenies of the Entamoeba lineage are based on SSU-rRNA analyses (Stensvold et al. 2010, 2011). Kin discrimination has been suggested as a behavioral mechanism in social microbes to resolve ‘cryptic’ varieties, given that individuals in a population signal to ‘like’ individuals (Kalla et al. 2011; Perez-Ponce de Leon and Nadler 2010; Saez and Lozano 2005). In single or multicellular organisms behavior toward kin differs from interactions with non-kin (Hamilton 1964; Queller et al. 2003; Wilson 2000). Altruistic behavior increases toward members of the same group, a phenomenon known as ‘the green beard effect’ (Dawkins 1976). Discrimination at the unicellular level has been detected in bacteria (Myxococcus xanthus; Vos and Velicer 2009) and protists (Dictyostelium, Polysphondylium violaceum; Kalla et al. 2011; Li and Purugganan 2011; Mehdiabadi et al. 2006; Ostrowski et al. 2008; Queller et al. 2003). The Entamoeba lineage is an ideal model to combine discrimination in a non-social ameba with morphological, multi-gene and ecological studies in attempting to resolve phylogenetic varieties considered ‘cryptic’. Here we demonstrate that two strains of Entamoeba invadens (IP-1 and VK-1:NS) aggregate only with members of their own population suggesting they distinguish members of the same strain based on behavioral and chemical signals. Discrimination could help resolve conflictive branching and crypticity among Entamoeba varieties and demonstrate the evolutionary history of this lineage. We discuss how gradual genetic changes combined with selective pressures that gave strain members the ability to discriminate between alike and non-alike-individuals drove the diversification of the Entamoeba lineage and contrast this analysis with proposals invoking ‘common design’ (independent major taxa emergence with no common ancestry; Nelson 1996) and ‘special Entamoeba creation’ (Sherwin 2009), which are unfounded.