Simpson Lab - Department of Biology, Dalhousie University, Canada

• The diversity and evolutionary affinities of 'excavates'
• (Extremely) halophilic protozoa
• The cytoskeleton of very deep-branching eukaryotes.

The diversity and evolutionary affinities of 'excavates'

A number of protist groups have a morphologically distinctive feeding groove. We refer to these groups (and their immediate relatives) as 'excavates', and have proposed that they are descended from a common ancestor, forming a clade called 'Excavata' (see Simpson, 2003 or Simpson and Roger, 2004 for an introduction and history). Excavata is now often treated as one of the 5-6 proposed 'super-groups' that would collectively encompass almost all of the known diversity of eukaryotes.

The excavate organisms are very important to understanding early eukaryote evolution because the assemblage includes most of the eukaryotic groups that, individually, have been championed as 'primitive' eukaryotes. This includes diplomonads (e.g. Giardia), parabasalids (e.g. Trichomonas), Euglenozoa (e.g. trypanosomes), and jakobids (e.g. Reclinomonas) - an obscure group with the most primitive-looking (bacterial-like) mitochondrial genomes known. If excavates are a monophyletic group, ALL of these individual claims for primitive status will be untenable, as will some of the most influential ideas about early eukaryotic cell evolution.

Determining whether excavates really do form a monophyletic group is one of the major questions in eukaryote evolution, and is currently unresolved. The deeper-level relationships amongst excavate groups are also incompletely understood. Through our own isolation efforts, and in collaboration with others, we are broadening the taxonomic sampling of the most poorly understood, yet phlyogenetically crucial excavate groups. This will provide a better basis for large-scale molecular phylogenetic studies, and for future genomic and cell biological work (see below). We also do in-depth transmission electron microscopy studies of some of these organisms. Recent work has seen us define a new deep branch of jakobids called Andalucia (see Lara, Chatzinotas and Simpson, 2006; Simpson, Perley and Lara, 2008), and several new relatives of diplomonads (Park et al, in press; Kolisko et al., in prep).

In collaboration with Andrew Roger's lab at Dalhousie University we have also conducted taxon-rich multi-gene phylogenetic analyses to understand the relationships amongst excavates, and to attempt to resolve the position of excavates relative to other eukaryotic supergroups (e.g. Simpson, Inagaki and Roger, 2006). This work is ongoing (Hampl et al. 2009).

More specific work on excavates:

We are also remain interested in the diversity and evolution of certain established groups within excavates, in particular Diplomonads and Euglenozoa. These are quite well-known groups whose internal phylogenies have been difficult to establish because of rapid evolution of SSUrRNA genes, and because of historical undersampling of free-living heterotrophic forms, in favour of photosynthetic or parasitic taxa.

We are specifically interested in the identity of the major clades of Euglenozoa, and how they are related, and also the relationships between free-living and parasitic kinetoplastids. See Simpson and Roger, 2004; Simpson et al. 2004, 2006 for (relatively) recent work on euglenozoan evolution.

We are also examining diplomonad relationships. In arecent study in collaboration with researchers from Charles University in Prague, we examined the relationships amongst typical diplomonads and the 'enteromonads' - see Kolisko et al. 2008.

(Extremely) Halophilic Protozoa

Microbiology textbooks give the impression that very hypersaline habitats (>25% salt) are exclusively inhabited by prokayotes, mostly Haloarchaea, and the alga Dunaliella. In reality there are a large number of heterotrophic protozoa that have been reported in samples from these environments - at least 30 morphologically distinguishable species by our conservative estimate (Park et al., 2009). These include ciliates, amoebae, and several groups of flagellates, some of which have unknown affinities within eukaryotes.

In collaboration with Prof. Byung Cheol Cho of Seoul National University, we have been charactering novel isolates of extremely halophilic (or borderline extremely halophilic) protozoa using electron microscopy and molecular phylogenetic techniques. We have identified several new genera (Park et al., 2006, 2009), and established that Pleurostomum, a flagellate of previously unknown affinities, is in fact a heterolobosean, whose closest relatives include the well-known amoeboflagellate Naegleria (Park et al., 2007).

A curious observation is that some of the protozoa that have been reported in the past from extremely hypersaline habitats are morphologically indistinguishable from species tht are found in marine habitats. In ongoing work we are trying to establish whether there are sharply defined halophilic or halotolerant molecularly-defined clades within these morphospecies.

Extremely hypersaline pond at the margin of the Salton Sea, California (A.G.B. Simpson, 2009)

The cytoskeleton of very deep-branching eukaryotes

In addition to the relationships amongst eukaryotes, we are interested in questions of deep-level cell evolution, for example, whether the eukaryotic supergroups contain homologous cytoskeletal elements, and whether we can know the basic cell structure of the common ancestors of eukaryotic super-groups, or even of all living eukaryotes.

Complementary to our work on excavates (see above), we are reconstructing the microtubular cytoskeletons of some poorly studied eukaryotes that represent very deep, independent branches in the phylogenetic tree of eukaryotes. At present we are examining Breviata, a flagellate that appears to be the deepest branch within the supergroup Amoebozoa (see Minge et al., 2009), and apusomonads, which are flagellates of very uncertain placement that are possibly most closely related to Amoebozoa and Opisthokonts (see Kim, Simpson and Graham, 2006). We are also examining Ancyromonas (Planomonas), which is possibly related to apusomonads, but otherwise of very uncertain placement. This work is being performed in close collaboration with Giselle Walker (University of Cambridge).