Archamoebae: The Apogee (or Nadir) of Amoebozoan Evolution

Mastigamoeba aspera, the type species of the mastigamoebids. Photo by Josef Brief.

There's just one group of amoebozoans left for me to cover: the Archamoebae. Among Amoebozoa, the Archamoebae are easily distinguishable by one significant feature - they lack mitochondria. Mitochondria are also absent in Breviata (which was initially identified as an archamoeba as a result), but Breviata has double basal bodies attached to the cilium unlike the single basal body of Archamoebae (Walker et al., 2006). Members of the Archamoebae are either freshwater amoeboflagellates or non-ciliate animal endosymbionts. Because the Archamoebae are primarily defined by a character absence some authors have suggested that their monophyly is suspect, but molecular analyses support their recognition. The lack of mitochondria also lead to Archamoebae being one of the four groups of protists (along with the Diplomonadida, Microsporidia and Parabasalia) that were grouped together as the "Archezoa", and suggested to have diverged from other eukaryotes prior to the origin of mitochondria. The archezoan hypothesis began to fall from favour in the latter half of the 1990s as relationships were proposed between various 'archezoans' and specific groups of mitochondriate protists, such as between Archamoebae and other amoebozoans. Putative mitochondrion-derived organelles such as mitosomes and/or hydrogenosomes have now been identified from most Archamoebae (Walker et al., 2001; Gill et al., 2007).

Molecular data supports the division of Archamoebae into two clades, the Mastigamoebida and Pelobiontida (Cavalier-Smith et al., 2004). Production of pseudopodia in Mastigamoebida is eruptive (Walker et al., 2001), which you may recall is not the usual condition for amoebozoans (other than Archamoebae, eruptive pseudopodium is also found in Leptomyxida). Free-living mastigamoebids are ciliate for at least part of their life cycle, and movement is generally by means of the cilium or by gliding rather than by pseudopodium production. Mastigamoebids may or may not be multinucleate, and the cilium is usually connected to the nucleus. Taxonomy of the free-living amoeboflagellate mastigamoebids is in something of a state of flux - three genera have been distinguished, Mastigamoeba, Phreatamoeba and Mastigella, but opinions differ as to whether the latter two should be distinguished from the first. Phreatamoeba balamuthi has been placed as a separate genus because of its more complex life-cycle alternating between amoeboid and amoeboflagellate stages, with the amoeboid stage predominant, but the flagellate stage is otherwise not distinguishable from Mastigamoeba. Mastigella has been distinguished based on the absence of a connection between the cilium and the nucleus, but the presence or absence of such a connection can be difficult to distinguish in taxa where the connection is very fine (Walker et al., 2001). Part of the problem lies in the genus Mastigamoeba itself - Mastigamoeba contains about forty species of which some are very distinct from each other but many (including the type species) are insufficiently studied. When more of the Mastigamoeba species are studied it may lead to the genus' subdivision.


The giant micro-aerobic amoeboid Pelomyxa palustris, with the larger cell nearly three millimetres in length. The flecks of green inside it are endosymbiotic algae. Photo from here.

The Pelobiontida contain three distinct genera, Pelomyxa, Mastigina and Entamoeba, united by molecular data. Mastigina and Pelomyxa are both free-living amoeboflagellates. Mastigina setosa has one or a few nuclei and a single long cilium; however, movement is by pseudopodia while the cilium contributes little if any propulsion. Pelomyxa palustris is a gigantic amoeboid, up to five millimetres long and often with hundreds of nuclei in a single cell. It possesses small cilia, but it almost goes without saying that they do not contribute to moving the cell's massive bulk. Cytoplasmic movement in Pelomyxa has been described as "fountain streaming" - a wave of cytoplasm moves across the dorsal surface of the cell and spills over the front. Like Trichosphaerium, Pelomyxa can reproduce by budding off a piece of the cell with a few nuclei (Hickson, 1909) - indeed, at one point it was thought that Pelomyxa never underwent mitosis, an error that lead to its brief elevation to the status of an independent phylum, Caryoblastea.

Both of the two archamoebaen clades have given rise to non-ciliate lineages - the Endolimacidae (Endolimax and Endamoeba) among mastigamoebids, and Entamoeba among pelobionts. Both of these taxa are animal endosymbionts, living inside the gut of their host (the name Endolimax means "inner slug") - Endolimax and Entamoeba inhabit vertebrates (including humans), while Endamoeba is found in cockroaches (including termites). Endolimacidae are generally innocuous, but a few species of Entamoeba can cause great trouble for their hosts - among humans, E. histolytica causes dysentery, while E. gingivalis lives in the mouth and can cause gum disease.


Endamoeba blattae, an inhabitant of the digestive system of cockroaches. Photo from here.

Even with the downfall of the idea that Archamoebae are among the most archaic of eukaryotes (which kind of makes the name of clade a bit misleading, but we're stuck with it now), the relationships of this group are still interesting. Archamoebae possess a distinct conical arrangement of microtubules at the base of the cilium; in turn, this cone sits on top of the nucleus like a Vietnamese farmer's hat. The presence of a similar structure in many slime moulds lead Cavalier-Smith to unite Archamoebae and Mycetozoa in a clade called Conosa (or Conosea, depending on which paper you're reading and what rank Cavalier-Smith felt like putting it at at the time). Molecular data generally supports placing the two close together, but not always as an exclusive clade - often various examples of the taxa described in the last post muscle their way in. Many of these other amoebozoans, such as Phalansterium and Multicilia, possess similar (but not identical) microtubular cones, and Cavalier-Smith (2009) recently extended the Conosa to include these taxa as well. This extended Conosa is supported by most molecular analyses, but it has to be noted that all of the taxa involved show elevated rates of evolution - Pelomyxa and Trichosphaerium, in particular, show rates going through the roof - and the possibility of long-branch attraction cannot be entirely ruled out. If I may be allowed a somewhat strained analogy, it's a bit like when an election is held in a culturally diverse area between candidates of various backgrounds and both sexes, and all the winning candidates end up belonging to one particular subgroup. It's entirely possible that this was the valid result, and nothing untoward occurred in the ballot-counting process, but still, you can't help wondering.

And with that, I reach the end of the Amoebozoa (at least for now). If you want to look back on the other amoebozoan posts:

General Amoebozoa: Putting the Formless in Formation.
Breviata anathema: TAFKAMI; TAFKAMI Walks.
Tubulinea: The Paragons of Amoeboids; Amoeba: Much Wierder than You Think.
Discosea: Keeping a Low Profile.
Mycetozoa: The Diversity of Slime Moulds.
Various Amoebozoa incertae sedis ('Variosea'): Amoebozoan Oddments.

Now if only someone would do the same for Heterolobosea...

REFERENCES

Cavalier-Smith, T. 2009. Megaphylogeny, cell body plans, adaptive zones: causes and timing of eukaryote basal radiations. Journal of Eukaryotic Microbiology 56 (1): 26-33.

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40 (1): 21-48.

Gill, E. E., S. Diaz-Triviño, M. J. Barberà, J. D. Silberman, A. Stechmann, D. Gaston, I. Tamas & A. J. Roger. 2007. Novel mitochondrion-related organelles in the anaerobic amoeba Mastigamoeba balamuthi. Molecular Microbiology 66 (6): 1306–1320

Hickson, S. J. 1909. The Lobosa. In A Treatise on Zoology pt. 1. Introduction and Protozoa, first fascicle (R. Lankester, ed.) Adam & Charles Black: London.

Walker, G., A. G. B. Simpson, V. Edgcomb, M. L. Sogin & D. J. Patterson. 2001. Ultrastructural identities of Mastigamoeba punctachora, Mastigamoeba simplex and Mastigella commutans and assessment of hypotheses of relatedness of the pelobionts (Protista). European Journal of Protistology 37 (1): 25-49.

Amoebozoan Oddments

Arachnula, a branched marine amoeboid of uncertain affinities. Photo by D. Patterson et al.

If I am to continue with the Cavalier-Smith et al. (2004) classification of Amoebozoa, the next class I reach would be the Variosea. However, Cavalier-Smith et al.'s Variosea was the weakest of the classes they recognised. The characters it was based upon (usually single cilia or centrosomes, no fruiting bodies, non-eruptive pseudopodia) are almost certainly plesiomorphic for Amoebozoa (or possibly the clade of Amoebozoa excluding Breviata), and even the name 'Variosea' was coined to refer to the diverse morphologies the class covered. Molecular studies have not indicated variosean monophyly, though the majority of 'varioseans' may fall into a paraphyletic series below Archamoebae and Mycetozoa. The Smirnov et al. (2005) classification contained no grouping comparative to Variosea: they simply left the 'varioseans' as "Amoebozoa incertae sedis". Still, this is as good a place as any to introduce the 'varioseans', plus a couple of other amoebozoan taxa that don't fall under the aegis of any of the other groupings I've referred to. In no particular order, these are the 'Varipodida', Centramoebida (or Acanthopodida), Stereomyxa, Corallomyxa, Phalansterium, Trichosphaerium and Multicilia.

The name 'Varipodida' was introduced by Cavalier-Smith et al. (2004) for a grouping of the non-ciliate genera Gephyramoeba and Filamoeba (though as it turns out 'Gephyramoeba' was misidentified; the varipodidan has been redescribed as Acramoeba, while true Gephyramoeba is a member of Leptomyxida and hence tubulinean; Smirnov et al., 2008). Subsequent studies have been equivocal about the monophyly of such a grouping. Generally, the two genera are close to the clade containing Archamoebae and Mycetozoa (more on that clade in the next post), either just outside it or just within it. Filamoeba is a flattened, fan-shaped amoeboid which produces extremely slender, spine-like subpseudopodia that look superficially like filose pseudopodia (hence the name). However, unlike true filose pseudopodia as found in Rhizaria, the subpseudopodia of Filamoeba do not function in movement. Acramoeba has branched cells, with the branches producing slender subpseudopodia like those of Filamoeba. It's also worth mentioning Arachnula, which is similar to Acramoeba but larger and multinucleate. Similar slender subpseudopodia are also produced by dictyostelian slime moulds, adding further support to a relationship between Varipodida and Mycetozoa.


Balamuthia mandrillaris. Photo from here.

The Centramoebida include the flattened soil-living amoebae Acanthamoeba and Balamuthia. Acanthamoeba is similar to Filamoeba in producing slender subpseudopodia (there was a picture in this post), but Balamuthia does not. Both genera can be pathogenic to humans, though that is probably not their main mode of life - Acanthamoeba has been connected to eye infections, while both Acanthamoeba and Balamuthia can cause meningitis. The really interesting thing about the centramoebids, however, is that unlike any of the amoebozoans I've covered in the last few posts, they possess a centrosome. The centrosome is an organelle that is involved in controlling the activity of the microtubules that run through the cytoplasm. The centrosome (or, specifically, the centrioles within the centrosome) is also the organelle responsible for the production of cilia. Up until now, I've been looking at non-ciliate amoebozoans, but the common ancestor of the Amoebozoa would have been ciliate and some amoebozoans remain so. Just how many times cilia have been lost in the Amoebozoa is unknown - at least five, probably more (Nikolaev et al., 2006). Centramoebids have lost the cilium and the centrioles, but they still retain the evidence of their former presence in the centrosome.


Corallomyxa, though without a species name I'm not certain whether this is supposed to be true Corallomyxa or the rhizarian Filoreta tenera, originally misidentified as a Corallomyxa. Photo by David Patterson.

A similar centrosome to that of Centramoebida is also found in the marine genera Stereomyxa and Corallomyxa, and the three taxa may form a clade (Cavalier-Smith et al., 2004). Stereomyxa and Corallomyxa have slender branched, reticulate pseudopodia, making them look at first glance a bit like a small plasmodial slime mould (though they differ in lacking fruiting bodies). A recent study suggesting that Corallomyxa, at least, may be rhizarian rather than amoebozoan has turned out to be based on another misidentification (Bass et al., 2009).


Trichosphaerium in the naked phase. Photo by David Patterson.

Trichosphaerium is perhaps the most bizarre of all the amoebozoans - and not merely because of its ability to happily chow down on plastic. Trichosphaerium is a relatively large, multinucleate marine amoeboid that lives enclosed in a membranous test. The test is pierced by numerous openings, and the amoeboid extends its pseudopodia through those openings for feeding and locomotion (this differs from the situation in Pellita because the openings are permanent, rather than each individual pseudopodium forcing its way out through the covering). The life cycle of Trichosphaerium alternates between asexual and sexual generations; in the asexual generation, the test has a covering of sharp spicules, but in the sexual generation it is smooth. Reproduction in the asexual stage is by division, but division is generally unequal - a relatively small piece of cytoplasm containing a few nuclei is pinched off to become a separate cell, and the nuclear membrane does not break down during division. In the sexual generation, asexual division may occur as in the asexual generation while the cell is growing, but when the cell finishes growing it forms a cyst. Within the cyst, the cell divides rapidly so that each individual nucleus is contained within its own individual biciliate cell; when the cyst breaks open, these uninucleate cells fuse to give rise to the next generation (Hickson, 1909). Under crowded conditions, normal multinucleate cells have also been recorded fusing to give rise to gigantic amoeboid cells up to three millimetres in diameter, with over a thousand nuclei, that break apart into more normal multinucleate within the course of a week.


Phalansterium - drawing of a colony and of an individual cell embedded in its matrix. Image by Stuart Hedley & David Patterson.

Phalansterium is another highly unusual amoebozoan, for the simple reason that it is not an amoeboid (Cavalier-Smith et al., 2004). It's not even an amoeboflagellate. At one time, Phalansterium was united with the choanoflagellates, which it resembles in having a collar around the base of its single cilium and by living in colonies; however, in Phalansterium the collar is a single undivided fold while in choanoflagellates it is divided into microvilli, while Phalansterium has only a single centriole at the base of its cilium to choanoflagellates' two. Molecular analysis firmly plonks Phalansterium among the amoebozoans. Cavalier-Smith has made some pretty big calls in relation to unassuming little Phalansterium, claiming it is the closest known organism to the probable ancestral morphology for all eukaryotes. This, however, is based on his assumption that the unikont morphology is ancestral.


Drawing of Multicilia instructa. Image by Won Je Lee.

Finally, Multicilia is a marine amoebozoan with a covering of numerous radially-arranged cilia. Mikrjukov & Mylnikov (1998) found that movement in Multicilia was by irregular, uncoordinated beating of the flagella, causing the cell to roll over the substrate without any obvious organisation into front or back or up or down. Short pseudopodia are extended to capture Multicilia's favoured food - other amoebozoans. Generally cells are roughly globular, but in certain unfavourable conditions large branched cells can develop. Cavalier-Smith et al. (2004) suggested that Multicilia was related to the Flabellinea due to the presence of glycostyles in the cell coat, but Nikolaev et al. (2008) placed it closer to other ciliate amoebozoans.

REFERENCES

Bass, D., E. E.-Y. Chao, S. Nikolaev, A. Yabuki, K. Ishida, C. Berney, U. Pakzad, C. Wylezich & T. Cavalier-Smith. 2009. Phylogeny of novel naked filose and reticulose Cercozoa: Granofilosea cl. n. and Proteomyxidea revised. Protist 160 (1): 75-109.

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40 (1): 21-48.

Hickson, S. J. 1909. The Lobosa. In A Treatise on Zoology pt. 1. Introduction and Protozoa, first fascicle (R. Lankester, ed.) Adam & Charles Black: London.

Mikrjukov, K. A., & A. P. Mylnikov 1998. The fine structure of a carnivorous multiflagellar protist, Multicilia marina Cienkowski, 1881 (Flagellata incertae sedis). European Journal of Protistology 34: 391-401.

Nikolaev, S. I., C. Berny, N. B. Petrov, A. P. Mylnikov, J. F. Fahrni & J. Pawlowski. 2006. Phylogenetic position of Multicilia marina and the evolution of Amoebozoa. International Journal of Systematic and Evolutionary Microbiology 56: 1449-1458.

Smirnov, A. V., E. S. Nassonova & T. Cavalier-Smith. 2008. Correct identification of species makes the amoebozoan rRNA tree congruent with morphology for the order Leptomyxida Page 1987; with description of Acramoeba dendroida n. g., n. sp., originally misidentified as ‘Gephyramoeba sp.’ European Journal of Protistology 44 (1): 35-44.

Smirnov, A. V., E. S. Nassonova, E. Chao & T. Cavalier-Smith. 2007. Phylogeny, evolution, and taxonomy of vannellid amoebae. Protist 158 (3): 295-324.

Discosea: Keeping a Low Profile

Various views of Vannella devonica, a representative of the Vannellidae. Photos by A. Smirnov.

Having covered the Tubulinea, it's time to move on to some of the less familiar Amoebozoa. At this point, however, the competing classifications of Cavalier-Smith et al. (2004) and Smirnov et al. (2005) begin to part ways - at least superficially. Cavalier-Smith et al.'s class Discosea is roughly comparable to Smirnov et al.'s class Flabellinea, but the two are not identical. Rather, Flabellinea is a subgroup of Discosea, the latter also including a few taxa that Smirnov et al. listed as Amoebozoa incertae sedis. As such, I'll use Discosea for the larger group and Flabellinea for the smaller group. Monophyly of the Flabellinea has been recovered in a number of analyses (including both the studies just cited), monophyly of the Discosea is more doubtful. Kudryavtsev et al. (2005) found a monophyletic Discosea, but all other molecular analyses including non-flabellinean 'Discosea' show 'Discosea' as polyphyletic. The morphological characteristics of Discosea cited by Cavalier-Smith et al. (2004) ("highly flattened, often discoid amoebae that move slowly by a leading lamellipodium", and a tendency to secrete some sort of thickened protective covering or membrane) are not unique to members of the group. Nor is the other feature cited by Smirnov et al. (2005) as distinguishing Flabellinea from Tubulinea, that cytoplasmic flow in pseudopodia does not form a single central axis. Discoseans produce only a single anterior pseupodium when moving (which may or may not produce subpseudopodia), and no discosean has an adhesive uroid.

As well as the Flabellinea, Cavalier-Smith et al.'s Discosea included the Cochliopodiidae and Thecamoebida. Later analysis (Kudryavtsev et al., 2005) suggested the polyphyly of Thecamoebida, and the discosean component was restricted to the genus Dermamoeba (nevertheless, because Dermamoeba and Thecamoeba are ultrastructurally similar, I may as well cover both in this post). Subsequent studies have continued to indicate the polyphyly of 'Thecamoebida', but Pawlowski (2008) refers to unpublished data that might restore monophyly to the group. Cavalier-Smith et al. (2004) also suggested that the multiciliate Multicilia was a discosean, but Nikolaev et al. (2006) have since shown otherwise (Multicilia will feature in the next Amoebozoa post; with its removal, Discosea becomes an entirely non-ciliate group). Finally, though its authors refrained from saying so, it seems entirely possible that, if the Discosea should be monophyletic, the recently described distinctive genus Pellita (Smirnov & Kudryavtsev, 2005) may be discosean.


Thecamoeba striata. I suspect that the front of the cell is towards the lower left. Photo by Keisotyo.

A general characteristic of Amoebozoa that I have not yet had cause to mention is the presence of a glycocalyx, a protective covering of proteins and polysaccharides that lies outside but is connected to the membrane of the cell. The nature of the glycocalyx has often been used in differentiating amoebozoan taxa in the past, but recent studies suggest that it may be more variable than previously thought (e.g. Smirnov et al., 2007), so glycocalyx-based features should probably be treated with caution. Both 'Thecamoebida' and Flabellinea have particularly well-developed glycocalyces. In 'Thecamoebida', the glycocalyx is amorphous (probably the ancestral condition for Amoebozoa) but extremely thick. Thecamoebids on the move are generally oval or oblong in shape without anterior subpseudopodia. Thecamoeba has the dorsal surface of the cell shaped into longitudinal folds or wrinkles, while Dermamoeba has no dorsal folds or wrinkles (Smirnov & Goodkov, 1999).


Paramoeba aestuarina. The large dark spot is the "parasome" (or the endosymbiont Perkinsela amoebae, however you want to put it). Image from here.

In Flabellinea, the glycocalyx is usually differentiated into a covering of column-like glycostyles, though a number of flabellineans have lost the glycostyle coat (Smirnov et al., 2007). Flabellinea are divided into three families - Vanellidae, Paramoebidae and Vexilliferidae, with the latter two more closely related to each other than to Vanellidae. The basic form of Flabellinea is broad and fan-shaped; Vanellidae have a smooth leading edge without subpseudopodia, while Paramoebidae and Vexilliferidae produce subpseudopodia - short and blunt in Paramoebidae, long and slender in Vexilliferidae. Paramoebids were also distinguished in the past by their possession of a distinctive organelle known as the parasome. However, this distinction has been reworked in recent years - not because of doubts about the reality of the parasome, but because the parasome is now regarded as a separate organism in its own right, an endosymbiont rather than an organelle of paramoebids.


Diagram of Pellita, showing how the subpseudopodia extend through the thick cell coat. Figure from Smirnov & Kudryavtsev (2005).

The recently discovered Pellita (Smirnov & Kudryavtsev, 2005) resembles Flabellinea in possessing a cell covering of glycostyles. This covering is particularly thick in Pellita, and almost resembles a test rather than a coat. So thick is Pellita's covering that the normal means of amoebozoan movement and feeding via the projection of pseudopodia are not possible for it. Instead, Pellita produces short subpseudopodia with a covering of basic cell membrane only that muscle their way between the glycostyles until they project outside the coat. These subpseudopodia engulf individual bacteria if feeding, while for movement subpseudopodia produced near the leading edge of the cell adhere to the substrate and the cell rolls forward over the top of them.


Cochliopodium. Photos from here.

Finally, the Cochliopodiidae also possess an external covering, but in their case it is a rigid coat that is entirely separate from the cell membrane. In the genus Cochliopodium the coat consists of carbohydrate scales, while in the genera Gocevia and Paragocevia the coat is filamentous. The cochliopodiid coat differs from the test of Arcellinida in that it is restricted to the dorsal surface of the cell, not surrounding it as in arcellinidans. Also, the coat is divided between the daughter cells when the cochliopodiid divides; in Arcellinida, one daughter cell keeps the test while the other has to make an entire new test from scratch.

REFERENCES

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40 (1): 21-48.

Kudryavtsev, A., D. Bernhard, M. Schlegel, E. E-Y. Chao & T. Cavalier-Smith. 2005. 18S ribosomal RNA gene sequences of Cochliopodium (Himatismenida) and the phylogeny of Amoebozoa. Protist 156: 215-224.

Nikolaev, S. I., C. Berny, N. B. Petrov, A. P. Mylnikov, J. F. Fahrni & J. Pawlowski. 2006. Phylogenetic position of Multicilia marina and the evolution of Amoebozoa. International Journal of Systematic and Evolutionary Microbiology 56: 1449-1458.

Pawlowski, J. 2008. The twilight of Sarcodina: a molecular perspective on the polyphyletic origin of amoeboid protists. Protistology 5 (4): 281-302.

Smirnov, A. V., & A. V. Goodkov. 1999. An illustrated list of basic morphotypes of Gymnamoebia (Rhizopoda, Lobosea). Protistology 1: 20-29.

Smirnov, A. V., & A. A. Kudryavtsev. 2005. Pellitidae n. fam. (Lobosea, Gymnamoebia) – a new family, accommodating two amoebae with an unusual cell coat and an original mode of locomotion, Pellita catalonica n.g., n.sp. and Pellita digitata comb. nov. European Journal of Protistology 41 (4): 257-267.

Smirnov, A., E. Nassonova, C. Berney, J. Fahrni, I. Bolivar & J. Pawlowski. 2005. Molecular phylogeny and classification of the lobose amoebae. Protist 156: 129-142.

Smirnov, A. V., E. S. Nassonova, E. Chao & T. Cavalier-Smith. 2007. Phylogeny, evolution, and taxonomy of vannellid amoebae. Protist 158 (3): 295-324.

Tubulinea: The Paragons of Amoeboids

The basal tubulinean Echinamoeba. If you look very closely at the lower end of the cell, you can see the filaments of the adhesive uroid. Photo by David Patterson et al.

Due to popular demand (does two count as popular?), I'm continuing with the Amoebozoa series (previous installments are here and here). The two major classifications of Amoebozoa that have been published in recent years are those of Cavalier-Smith et al. (2004) and Smirnov et al. (2005). While at first glance the two systems appear quite different, there are few real significant differences between them. Mostly it's a matter of different names being used for similar concepts, plus the Cavalier-Smith et al. classification assigns positions to a number of taxa that the more conservative Smirnov et al. classification is content to list as Amoebozoa incertae sedis. The Cavalier-Smith et al. classification divides amoebozoans between seven 'classes', which offers a good basis for dividing my posts. One of Cavalier-Smith et al.'s classes currently includes a single species, the strange (and not necessarily amoebozoan) Breviata anathema, which has previously been covered here and here, while the two classes of their infraphylum Mycetozoa (slime moulds) include the first three entries for this post here. So that leaves four classes that I haven't yet covered in detail. In the coming posts, I'll start with the Tubulinea, move to the Discosea, waft through the 'Variosea' and finish with the Archamoebae. And if that seems like a lot to you, just be glad I didn't choose to do my amoeboid series on Foraminifera - we could have been here well into the next century.

The "Tubulinea" of Smirnov et al. (2005) are the same grouping as the "Lobosea" of Cavalier-Smith et al. (2004). I prefer to use the name Tubulinea because Lobosea has been used in the past for a much larger grouping, effectively all amoebozoans except Mycetozoa and (sometimes) Archamoebae. Also, the name Tubulinea refers to one of the characteristic features of this group, the production of tubular rather than flattened pseudopodia, with cytoplasmic flow within the pseudopodia or entire cell along a single axis. All Tubulinea lack cilia at all stages of the life cycle. The Tubulinea include the Tubulinida (Amoeboidea of Cavalier-Smith et al.), Arcellinida, Copromyxidae, Leptomyxida and Echinamoeba, and both the papers cited above produced identical phylogenies for this class.


Leptomyxa, type genus of the Leptomyxida. Note the branched morphology. Don't ask me to tell you which way this one's going - I think that might be the uroid towards the bottom right, but I'm not sure. Photo by David Patterson et al.

Echinamoeba forms the basalmost clade within the Tubulinea, together with the species 'Hartmannella' vermiformis (erroneous comments on the relationships of the family Hartmannellidae in Cavalier-Smith et al., 2004, are due to the use of H. vermiformis to represent Hartmannella; it wasn't until later that Smirnov et al., 2005, showed that H. vermiformis is not closely related to other Hartmannella species. The normal cell shape of Echinamoeba is "acanthopodian" - flattened with short, spinelike subpseudopodia (Smirnov & Goodkov, 1999); it only produces a more typically tubulinean cylindrical, monopodial form under particular conditions. The form it takes in these conditions is one produced by many amoeboids called the 'limax' form. Limax is a genus of slugs, and a microscopic slug is exactly what this form looks like. 'Hartmannella' vermiformis, on the other hand, is habitually worm-like, and has gained a certain notoriety as an unwitting vector for bacteria causing respiratory diseases in humans, particularly Legionnaire's disease (Brieland et al., 1997).

The Leptomyxida are the next group to branch off. The four genera of leptomyxidans - Leptomyxa, Rhizamoeba, Flabellula and Paraflabellula - resemble Echinamoeba by normally being flattened, and only adopting a tubular limax-like form occasionally. The normal form of Leptomyxa is reticulate, with a anastomosing net of pseudopodia. In the other three genera, the uroid (the trailing end of the moving cell) is adhesive, so when the cell is moving the posterior end is drawn out into smeared streaks. Rhizamoeba is monopodial, while the other two genera are fan-shaped. Paraflabellula produces short subpseudopodia from the anterior edge of the cell, Flabellula doesn't.


The fruiting body of Copromyxa arborescens, which grows up to 2.5 mm in height. Figure from Nesom & Olive (1972).

The Copromyxidae were placed by Cavalier-Smith et al. (2004) among the Tubulinea on the basis of their morphology (but were not represented in the molecular analysis), but were not even mentioned by Smirnov et al. (2005). Copromyxids include two little-studied genera, Copromyxa and Copromyxella. Cavalier-Smith et al. suggested that they are closer to Arcellinida and Tubulinida than other Tubulinea as these three groups are habitually rather than only intermittently tubular. Copromyxids differ from other Tubulinea in having a slime-mould type life cycle (I overlooked them in my earlier slime mould post). Life for copromyxids really is a pile of crap - their chosen habitat is animal dung. Fruiting bodies are produced by previously separate cells aggregating together to form a mound. Newly-arriving cells clamber over their confederates to reach the top of the pile, and the eventual result is a small, vaguely tree-like fruiting body (Bonner, 1982). Copromyxids are very similar in appearance to the non-amoebozoan acrasid slime moulds, and many earlier references combine the two.

The Arcellinida are the most speciose subgroup of Tubulinea (at least, as far as we know). Arcellinida are the testate Amoebozoa - they possess are hardened test of either secreted proteinaceous material or agglutinated mineral grains. The test is roughly vase-shaped, with a single opening through which the organism extends its pseudopodia. Phylogenetic studies (Nikolaev et al., 2005) confirm that the testate Amoebozoa form a monophyletic group (which, as I noted earlier, forms an interesting contrast to the polyphyletic testate Rhizaria), but the same cannot be said for proteinaceous- versus agglutinated-test formers, with lineages apparently switching between the two a number of times.


Nebela tubulosa, a member of the Arcellinida. Photo by Antonio Guillen.

Finally, the Tubulinida includes Amoeba itself and its nearest and dearest (such as Chaos, Saccamoeba and true Hartmannella). Tubulinida differ from Echinamoeba and Leptomyxida in being permanently tubular, never flattened. The cell may move limax-wise as a single pseudopodium (Saccamoeba, Hartmannella, sometimes Amoeba) or may form multiple pseudopodia (Chaos, other times Amoeba). Both Cavalier-Smith et al. and Smirnov et al. placed Chaos and Amoeba closest to one another, and indeed phylogenetically mixed together. As the only difference between the two seems to be the number of nuclei (one in Amoeba, more in Chaos) it would perhaps not be surprising if one or the other, or both, turned out to be polyphyletic.

Finally, I'd like to end this post on a bit of speculation. In the first Amoebozoa post, I described the ridiculously large genome of Tubulinida species (thanks to commentor George X for pointing out that the species I referred to as Amoeba dubia is now known as Polychaos dubium). I tried to find if any studies had been done on the detailed genetic structure of these species, to see if there was any clue as to just why Tubulinida have such enormous genomes, but I couldn't find any. Indeed, when searching on Amoeba in Google Scholar, I was struck by the dates shown for most of the results - a significant proportion dating back to the period from the 1940s to the 1960s. It looks like Amoeba proteus, so popular as a model organism in the early days of cell biology due to its large size making it easy to observe and manipulate, may have since fallen in popularity. I can guess at some reasons why that might be - I get the impression that Amoeba's rarity makes it tricky to find, that it is difficult to maintain in culture once you do find it, and I wonder if, in a time when electron microscopy has become almost routine, Amoeba is large enough that its size has become a positive disadvantage rather than advantage.

In the absence of much information about Amoeba's genome beyond its size, speculation becomes ill-founded. The large size of the genome doesn't necessarily indicate a proportionally large number of genes - it could be that Amoeba is carrying a particularly heavy load of non-coding DNA. Also, the previously-mentioned cyclic nature of the Amoeba genome, with the amount of DNA increasing and decreasing over the course of the division cycle by a factor of nearly three (Parfrey et al., 1998), suggests that Amoeba proteus is at least hexaploid. Again, it might not be - perhaps instead of the entire genome being replicated three times, a smaller number of chromosomes are replicated many times (as we know happens with such things as B chromosomes in animals). But, with all those caveats, I still can't help wondering if the cyclic Amoeba genome is related to its unusual success as an asexual organism. Let us assume that the entire genome is replicated in the cell cycle, and that it is random which of the resulting replicate chromosomes gets retained and which disposed of prior to division. The result would be that the effective mutation rate of Amoeba would probably be noticeably higher than in organisms with more straightforward genetic cycles. Could this be what has allowed Amoeba to survive for so long seemingly without the benefit of recombination?

REFERENCES

Bonner, J. T. 1982. Evolutionary strategies and developmental constraints in the cellular slime molds. American Naturalist 119 (4): 530-552.

Brieland, J. K., J. C. Fantone, D. G. Remick, M. LeGendre, M. McClain & N. C. Engleberg. 1997. The role of Legionella pneumophila-infected Hartmannella vermiformis as an infectious particle in a murine model of Legionnaire's disease. Infection and Immunity 65 (12): 5330-5333.

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40 (1): 21-48.

Nesom, M., & L. S. Olive. 1972. Copromyxa arborescens, a new cellular slime mold. Mycologia 64 (6): 1359-1362.

Nikolaev, S. I., E. A. D. Mitchell, N. B. Petrov, C. Berney, J. Fahrni & J. Pawlowski. 2005. The testate lobose amoebae (order Arcellinida Kent, 1880) finally find their home within Amoebozoa. Protist 156: 191-202.

Parfrey, L. W., D. J. G. Lahr & L. A. Katz. 2008. The dynamic nature of eukaryotic genomes. Molecular Biology and Evolution 25 (4): 787-794.

Smirnov, A. V., & A. V. Goodkov. 1999. An illustrated list of basic morphotypes of Gymnamoebia (Rhizopoda, Lobosea). Protistology 1: 20-29.

Smirnov, A., E. Nassonova, C. Berney, J. Fahrni, I. Bolivar & J. Pawlowski. 2005. Molecular phylogeny and classification of the lobose amoebae. Protist 156: 129-142.

Amoebozoan Classification: Putting the Formless in Formation

Chaos carolinense, the species generally regarded today as the main exemplar of the genus Chaos (see the note below). In case you were wondering, this individual is moving towards the right. Photo by David Patterson.

In my last post, I described some of the oddities of the well-known micro-organism Amoeba. In this post, I'll expand the field of view to look more generally at the clade of Amoebozoa. Study of amoeboids has certainly been going on for a long time - an amoeboid was among the few micro-organisms listed by Linnaeus (1758), under the name of Volvox chaos*. But how, you may be wondering, does one go about characterising a shapeless blob? And how does one distinguish one type of shapeless blob from another?

*Later raised by Linnaeus to the status of a separate genus, Chaos. The name Chaos is still in use for a genus very closely related to Amoeba (the main difference is that Chaos is multinucleate, while Amoeba has a single nucleus) but it pays not to look to carefully at the taxonomy. Debate about the identity of the original Volvox chaos raged down the years - whether it was the modern Chaos, the modern Amoeba, or something else entirely - but the debate was largely futile, because the original description cited by Linnaeus illustrates little more than a shapeless blob with a few dots over it. Many authors included the modern 'Chaos' in the genus Pelomyxa, another large multinucleate amoeboid, but Pelomyxa is an entirely different beast. King & Jahn's (1948) argument for recognition of the three genera Amoeba, Pelomyxa and Chaos with those names is in accord with the modern usage, but has the air more of arbitrary pragmatism rather than adherence to priority - the genus that is now Chaos needed a name, and the name Chaos was going begging. The modern usage is now well-established, and it wouldn't really benefit anyone to go stirring things up now.

If you are wondering exactly that, then I'm sorry to point out to you that you're under something of a misunderstanding about the nature of cellular structure. Not that I blame you, because it's an easy enough misunderstanding to develop. Most basic descriptions of intra-cellular structure might suggest that the interior of a cell is basically liquid (or at most jelly-like) with the nucleus and other organelles freely floating about like so many chunks of carrots and peas in a vegetable soup. But while the cytoplasm is fluid, it's not water. It's a complex mixture of all sorts of molecules - actin filaments, microtubules, etc. - almost more like an enormous bowl of noodles than a broth. It is the interactions between these molecules that give a cell its shape, and also that make it move. The movement and shape-changes of an amoeboid are not random, but follow a pattern. And different types of amoeboid will move according to a different pattern. The form and manner that the amoeboid adopts while moving is generally one of the first things to observe in its identification.



Jahn et al. (1974) divided amoeboid micro-organisms into two classes based on the mode of pseudopodium formation. In one class (as shown in the figure from Jahn et al., 1974, above), the cytoplasm was liquified and pushed forward by contraction, re-coagulating at the front of the resulting broad, lobose pseudopodium. In the second class (shown in the figure below), long filamentous pseudopodia were extended with each side of the pseudopodium moving against the other in an opposite direction.



The distinction between lobose and filose amoeboids has been reinforced with further study, though as it turns out both modes have evolved multiple times (filose pseudopodia more often than lobose pseudopodia). Filose pseudopodia are found among such organisms as the Rhizaria (including foraminifers and radiolarians), while Amoebozoa are characterised by lobose pseudopodia (in those amoebozoans that don't produce distinct pseudopodia, the entire cell moves in this way). Lobose pseudopodia are also found among the Heterolobosea, another group of micro-organisms not closely related to Amoebozoa (heteroloboseans include Naegleria, the causative agent for amoebic meningitis, and acrasid slime moulds). Nevertheless, pseudopodium formation differs between the two in that in amoebozoans, movement is smooth and continuous, while heteroloboseans produce pseudopodia eruptively, cycling between periods of extension and periods of "resting" (heteroloboseans also have a distinct mitochondrial structure from amoebozoans).


Acanthamoeba, a common amoebozoan in soil and fresh water, occasionally causing eye infections in humans. Acanthamoeba produce distinctive short, narrow subpseudopodia from the single flattened cell-wide pseudopodium, as seen in this photo by David Patterson.

Among amoebozoans except testate forms, archamoebae and slime moulds, Smirnov & Goodkov (1999) recognised nineteen "morphotypes" distinguished by their mode of movement - whether the amoeboid extends multiple pseudopodia, or moves as a single unit; the form of the uroid (the posterior end of the cell while the amoeboid is moving); whether the surface of the cell is ridged or smooth; and other such details. Though Smirnov & Goodkov (1999) explicitly established their morphotype distinctions as identification characters only, without necessarily indicating higher classification, molecular phylogenetic studies have indicated a rough (but not exact) correlation between locomotive mode and phylogeny (Smirnov et al., 2005). For instance, Tubulinea, the class of amoebozoans including Amoeba and Chaos, generally produce tubular or subcylindrical pseudopodia with cytoplasm streaming down a distinct single central axis. Members of another class, Flabellinea, have flattened pseudopodia without a single central axis.


Vannella simplex, a member of the Flabellinea. Note the single broad flat fan-shaped pseudopodium. Photo from here.

Not everything is movement, of course. Other features distinguishing amoebozoans include the texture and ornamentation (if any) of the outer cell surface; the shape, distribution and number of the nucleus/nuclei and other organelles; and the presence and nature of a protective test (interestingly, while the testate filose amoeboids do not appear to form a monophyletic group among the Rhizaria, the testate lobose amoebae do seem to be monophyletic among the Amoebozoa - Nikolaev et al., 2005). There is a good detailed online guide at Alexey Smirnov's website (and a hat-tip to Psi Wavefunction for pointing the site out to me). A number of the higher taxa among the Amoebozoa have become reasonably robust in the last few years - if you're not too completely sick of amoeboids, I may introduce you to a few over the next few posts.

REFERENCES

Jahn, T. L., E. C. Bovee & D. L. Griffith. 1974. Taxonomy and evolution of the Sarcodina: a reclassification. Taxon 23 (4): 483-496.

King, R. L., & T. L. Jahn. 1948. Concerning the genera of amebas. Science 107: 293-294.

Nikolaev, S. I., E. A. D. Mitchell, N. B. Petrov, C. Berney, J. Fahrni & J. Pawlowski. 2005. The testate lobose amoebae (order Arcellinida Kent, 1880) finally find their home within Amoebozoa. Protist 156: 191-202.

Smirnov, A. V., & A. V. Goodkov. 1999. An illustrated list of basic morphotypes of Gymnamoebia (Rhizopoda, Lobosea). Protistology 1: 20-29.

Smirnov, A., E. Nassonova, C. Berney, J. Fahrni, I. Bolivar & J. Pawlowski. 2005. Molecular phylogeny and classification of the lobose amoebae. Protist 156: 129-142.

Amoeba: Much Wierder than You Think

Amoeba proteus extending pseudopodia to feed on a hapless ciliate. Note how the pseudopodia completely surround the ciliate, cutting off any escape, before they close in on it. A fantastic photo by Wim van Egmond - you owe it to yourself to visit that link.

I have been challenged (or at least, I think I have been challenged) to write some posts on amoebozoans, the clade of eukaryotes that includes such organisms as Amoeba and most slime moulds. As amoebozoans are unequivocally neat organisms, I'm happy to take up the challenge, but I thought Id start by focusing on the most famous amoebozoan genus of all, Amoeba itself. There are about five or so species of Amoeba (at least that I'm aware of), but most of what I'm going to say in this post applies equally to all of them. I think I'm safe in claiming that Amoeba is not just the most famous amoebozoan, it's also the most famous of all unicellular eukaryotes. Almost all general biology textbooks will include two examples of 'protists', and one of them will always be Amoeba (the other will be either Euglena or Paramecium). The funny thing about this ubiquity of the Amoeba exemplar, however, is that as unicellular protists go, Amoeba is actually (a) apparently not that common, and (b) seriously wierd*.

*Euglena and Paramecium aren't that typical either.

What makes Amoeba so odd? For a start, Amoeba is amoeboid* (kind of by definition, really). This might not seem so unusual at a glance (many micro-organisms are amoeboid), but the thing is that Amoeba is always amoeboid. It never possesses cilia. Many (if not most) other amoeboid eukaryotes transform into amoeboflagellates or flagellates for at least part of their life-cycle, or possess flagellated gametes, while the majority of unicellular eukaryotes are permanently flagellated**. Even among amoebozoans, cilia are not that unusual; they're still present in Breviata, Multicilia, Phalansterium, Mastigamoebidae, Pelomyxa and many Mycetozoa, though cilia have been entirely lost among amoebozoans at least nine times (Cavalier-Smith et al., 2004).

*Simply for the sake of avoiding confusion, I prefer to avoid the common use of the name "amoeba" to refer to any organism with an Amoeba-like morphology.

**A brief explanation about the terms "cilium" and "flagellum". Originally, the term "cilium" was used for small hair-like locomotory structures, usually present in large numbers, while "flagellum" referred to larger whip-like structures of which a cell would usually only have one or a few. As our knowledge of unicellular diversity broadened, the boundary between the two became increasingly blurred, and fundamentally they're all the same structure. On the other hand, "flagella" in bacteria, though superficially resembling flagella in eukaryotes, are structurally very different (eukaryote flagella are organelles formed of membrane-bound microtubules, while bacterial flagella are formed of a single protein strand). As a result, recent authors have tended to restrict the term "flagellum" to bacteria, and expand the term "cilium" to cover all eukaryote locomotory structures (a replacement term "undulipodium" never caught on [thankfully]). However, terms such as "flagellate" are still pretty well entrenched in their old sense.


Amoeba 'radiosa', photo by David Patterson & Aimlee Laderman. Despite the use of the name, there is not really such a species as 'Amoeba radiosa'. Rather, the name is used to indicate amoebae that have become detached from the substrate and are free-floating in the water column, where they abandon their usual flattened form and adopt a form with slender pseudopodia radiating from a spherical centre. Once they come back into contact with a solid surface, they will return to their normal morphology.

The second unusual thing about Amoeba (which is perhaps not unconnected to the first thing) is its reproductive habits. Most people are aware that Amoeba reproduce by division. That happens to be the only way that Amoeba reproduce (Chapman-Andresen, 1971); they are (so far as anyone knows) entirely asexual. While asexual reproduction is normal for many organisms, exclusively asexual lineages are something of a rarity. Most asexually reproducing organisms have more aphid-like life cycles - they reproduce asexually as long as conditions are favourable for doing so, but convert to sexual reproduction when times get tough. Even bacteria, which mostly don't engage in sexual reproduction per se, are able to engage in processes such as conjugation that still allow for gene flow.

And the third wierd thing about Amoeba has to be its genetics. Amoeba genomes are simply huge - the largest genomes known to exist, in fact. We humans have a genome that clocks in at a little under three billion base pairs of DNA. Amoeba proteus, the best-known species of Amoeba, has a genome containing closer to three hundred billion base pairs. And even that effort pales in comparison to Amoeba dubia, which carries around a whopping six hundred and seventy billion base pairs. That's right - the difference in genome size alone between the two species is larger than the total genome size of any other organism! The actual genetic structure of Amoeba, however, appears to be little-known. The genome of A. proteus is divided between more than five hundred chromosomes, which is hardly surprising considering its size. By means unknown, however, this enormous genome can be reduced to nearly a third of its normal size over the course of cell division (Parfrey et al., 2008). Presumably the normally polyploid amoeba jettisons excess chromosomes prior to division then recreates them from the remainder afterwards.


Amoeba proteus on the move (towards the top left of the photo). Note the knobbly bit at the bottom right corner. This is the uroid, and represents the trailing end of the cell. The form of the uroid has turned out to be surprisingly useful in identifying amoeboids. Another photo by David Patterson.

One other feature of the Amoeba nucleus is worth mentioning. The nucleus contains a number of stellate aggregations of condensed helical structures just inside the nuclear envelope that, when first observed, were not unreasonably thought to represent condensed chromosomes. However, further study showed that the nuclear helices were composed of a mixture of proteins and RNA (not DNA) and seemed to be able to be transported out of the nucleus into the surrounding cytoplasm (Minassian & Bell, 1976). The helices disappear over the course of cell division, but are regenerated afterwards. The exact function of these helices is still unknown. Minassian & Bell (1976) seem to have suggested (in a rather cagy way that would have allowed for ready back-tracking if they turned out to be wrong, and which I may have easily misinterpreted) that they could be related to ribosome formation. Gągola et al. (2003), in contrast, note the attachment of actin filaments to the helices, and imply that they may play a role in cell motility (Amoeba with removed nuclei are unable to move*, while amoeboid animals cells can continue to move even without their nuclei).

*Removing the nucleus from an Amoeba is as simple as slicing it in half.

REFERENCES

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40 (1): 21-48.

Chapman-Andresen, C. 1971. Biology of the large amoebae. Annual Review of Microbiology 25: 27-48.

Gągola, M., W. Kłopocka, A. Grębecki & R. Makuch. 2003. Immunodetection and intracellular localization of caldesmon-like proteins in Amoeba proteus. Protoplasma 222: 75-83.

Minassian, I., & L. G. E. Bell. 1976. Studies on changes in the nuclear helices of Amoeba proteus during the cell cycle. J. Cell Sci. 20: 273-287.

Parfrey, L. W., D. J. G. Lahr & L. A. Katz. 2008. The dynamic nature of eukaryotic genomes. Molecular Biology and Evolution 25 (4): 787-794.

TAFKAMI Walks

Various individuals of the amoeboflagellate Breviata anathema. Note particularly the extended anterior and posterior pseudopodia on numbers 13, 14 and 18. Figure from Walker et al. (2006).

There's been a couple of really interesting things come through the pipeline lately. For this post, I'm not going to talk (yet) about yesterday's publication of an analysis of acanthodians that suggests that they are not a monophyletic grouping. If you want to know what that's all about, ask Adam Yates (and if you don't know what an acanthodian is, I've briefly discussed them previously). Today, I'm going to discuss another recent publication.

One of the first posts I wrote for this site (in fact, the sixth) was on an organism that I dubbed TAFKAMI, or The Amoeboid Formerly Known As Mastigamoeba invertens. This organism had been originally identified as 'Mastigamoeba invertens' when isolated in 1992, and was eventually properly described by Walker et al. (2006) as Breviata anathema* (the real Mastigamoeba invertens is known only from an undiagnostic description published in 1892, and, short of someone inventing a time machine so that they can look over its original describer's shoulder, will probably never be identifiable). Breviata is an amitochondriate, microaerobic amoeboid or amoeboflagellate (depending on life cycle stage). As explained in the previous post, Breviata has proven to be an obscenely difficult organism to place phylogenetically. Its position in phylogenetic analyses has been very unstable, and it jumps wildly about depending on the analysis parameters. The earliest division in eukaryotes appears to be between unikonts (animals, fungi and amoebozoans, which have a single flagellum with a single basal body) and bikonts (including plants, algae and excavates, with flagella in doublets or with double basal bodies), and it has not even been conclusively established whether Breviata is a unikont or a bikont. It has a single flagellum like a unikont, but two basal bodies attached to that flagellum like a bikont**, and sturdy branching filose pseudopodia like nothing else. Whatever its position, it seems likely that the divergence of Breviata from other eukaryotes happened not long after the the origin of crown eukaryotes in total.

*Tragically, Walker et al. (2006) gave no etymology for the new name. I have always wondered what exactly is so anathematic about Breviata anathema.

**Just to confuse matters, there are unikonts with double basal bodies, and bikonts with single flagella. However, bikonts with single flagella always retain two basal bodies. The anterior basal body in bikonts is always the younger of the two, and when the posterior basal body dies off the anterior body moves to the back and a new basal body grows in front of it. Those unikonts with two basal bodies still lack this distinctive growth pattern. Unfortunately, the flagellar growth pattern has not yet been studied for Breviata.

A new paper published by Minge et al. (2009) presents a new phylogenetic analysis incorporating Breviata anathema that draws on 17,283 nucleotide sites from no less than 78 genes (for contrast, the analysis of Breviata by Walker et al. used 1274 sites). The results of this analysis place Breviata with the amoebozoans, the clade including the majority of amoeboids with lobose pseudopodia. The support for this result is actually not too bad for this high a level of evolutionary divergence. Under certain analytical parameters, Breviata fell within other amoebozoans as sister to the other amitochondriate amoeboids Entamoeba and Mastigamoeba proper, but in the majority of cases it was the sister group to all other amoebozoans. This seems the more likely option as Breviata lacks certain sequence signatures (including a small insertion) characteristic of other Amoebozoa.

Sadly, as interesting as this result is, and as impressive as the amount of data used is, the analysis of Minge et al. (2009) suffers a fatal flaw. Though the analysis by Walker et al. did not give a conclusive result, the position they suggested to be most likely for Breviata was as sister to the Apusozoa. Apusozoans are a small group of flagellates with doubled flagella, and have been suggested to represent the basalmost divergence in the bikont lineage. As well as the double basal bodies, Apusozoa also produce filose pseudopodia like Breviata. Unfortunately, due to lack of data, the analysis by Minge et al. (2009) doesn't include a single apusozoan. While I'm personally sceptical of an apusozoan-Breviata relationship, I do think that without their inclusion the results of Minge et al. can't really be taken as conclusive.

Even if the phylogenetic results can't be entirely trusted, Minge et al. (2009) do have some interesting things to say. One of the interesting results from Walker et al. (2006) was the identification in Breviata of what appeared to be a hydrogenosome. Hydrogenosomes are hydrogen-processing organelles found in a number of anaerobic eukaryotes that have been shown to be altered mitochondria (Akhmanova et al., 1998). If Breviata did have a hydrogenosome, that would add to an increasing amount of evidence that all of the various 'amitochondriate' eukaryotes living today actually descend from ancestors that once had mitochondria (in contrast to previous opinions that they diverged from other eukaryotes prior to the origin of mitochondria). Among the genes possessed by Breviata, Minge et al. identify a number of genes derived from the pre-mitochondrial endosymbiont, confirming that Breviata's lack of mitochondria is a secondary feature.

Finally, there is the way Breviata moves. Amoeboids move, of course, by the extension of pseudopodia, but the exact method by which pseudopodia are produced can differ significantly between taxa. Indeed, in organisms with few permanent morphological features, the mode of pseudopodium formation has turned out to have a fair amount of phylogenetic significance (Smirnov et al., 2005). With its unique phylogenetic position, it seems only fitting that Breviata should have a unique mode of movement - it walks. A pseudopodium is protruded from the front of the cell and attached to the substrate. The rest of the cell body then rolls forward over the attached pseudopodium (like a tractor on treads, is Minge et al.'s analogy), until the pseudopodium is left trailing behind before being retracted and another pseudopod is extended from the front to repeat the process. No other organism has a mode of movement like Breviata - always twirling, twirling, twirling towards the future!

REFERENCES

Akhmanova, A., F. Voncken, T. van Alen, A. van Hoek, B. Boxma, G. Vogels, M. Veenhuis & J. H. P. Hackstein. 1998. A hydrogenosome with a genome. Nature 396: 527-528.

Minge, M. A., J. D. Silberman, R. J. S. Orr, T. Cavalier-Smith, K. Shalchian-Tabrizi, F. Burki, Å. Skjæveland & K. S. Jakobsen. 2009. Evolutionary position of breviate amoebae and the primary eukaryote divergence. Proceedings of the Royal Society of London B 276: 597-604.

Smirnov, A., E. Nassonova, C. Berney, J. Fahrni, I. Bolivar & J. Pawlowski. 2005. Molecular phylogeny and classification of the lobose amoebae. Protist 156: 129-142.

Walker, G., J. B. Dacks & T. M. Embley. 2006. Ultrastructural description of Breviata anathema, n. gen., n. sp., the organism previously studied as "Mastigamoeba invertens". Journal of Eukaryotic Microbiology 53 (2): 65-78.

The Diversity of Slime Moulds

How could you not love an organism that manages to combine both slime and mould? Slime moulds are saprobic organisms (i.e. they gain their nutrients by breaking down dead organic matter) that spend most of their life cycle feeding as separate amoeboid cells or disaggregated plasmodium. However, when conditions become right all the cells or plasmodium near each other will stream together to form a fungus-like fruiting body that releases spores, as shown in the diagram above borrowed from here. Because slime moulds thus resemble protozoa for part of their life cycle but fungi at other times, they were an early protagonist in the destruction of the idea that all organisms could be divided between plants and animals. Slime moulds, it turns out, are mostly not related to plants or animals. As our understanding of organismal phylogeny has progressed, it has become clear that not all slime moulds are even related to other slime moulds. Instead, the term has been used to cover a number of phylogenetically disparate organisms with little in common other than similar life cycles. However, the majority of references to slime moulds out there fail to mention this, focusing on only a small part of "slime mould" diversity, so I thought I'd give a brief overview of the full diversity of organisms with a slime mould-type life cycle.



1 - Myxogastrea: The plasmodial or acellular slime moulds, also known as Myxomycetes. This is the largest group of "slime moulds" - both in terms of number of species and the size reached by some species. While most other groups of slime moulds are fairly microscopic, myxogastreans reach sizes where they can easily be seen with the naked eye, at which point they are usually mistaken for fungi. During the feeding stage of their life cycle, myxogastreans form a plasmodium - a spreading mass that is not divided into individual cells, like threads of jelly or mucus (photo above from here). Recent phylogenetic analyses agree that myxogastreans belong to the Amoebozoa, the clade that includes more familiar amoeboids such as, well Amoeba (Cavalier-Smith et al., 2004). Indeed, the amoeboflagellate genus Hyperamoeba has been shown to represent a polyphyletic assortment of myxogastreans that have dropped the plasmodial habit.



2 - Dictyostelia: While myxogastreans may be the largest group of slime moulds, dictyostelians may be the most famous, because they take the standard coolness of the slime mould life cycle and turn the dial up to eleven. The photo above from here shows the various stages of the life cycle of the most famous dictyostelian, Dictyostelium discoideum. Dictyostelians are cellular slime moulds - while myxogastreans form a plasmodium, dictyostelians spend their nutritive phase as separate individual amoeboids. When the time comes for reproduction, the separate amoeboids swarm together to form a slug-shaped mass that actually moves as one, like something out of a Japanese cartoon. The dictyostelian slug crawls around until it finds a suitable location, at which point it extends outwards to form a sporangium on the end of a long thin stalk. The complexities of Dictyostelium's life cycle have made it a favoured study organism for such topics as kin selection, as researchers attempt to identify what cues incite slug formation, and why some individual amoeboids forming the sporangium stalk are seemingly willing to sacrifice their own reproductive potential in order to promote the reproduction of those cells forming the sporangium.

Phylogenetically, dictyostelians are also amoebozoans, closely related to myxogastreans. However, analyses are currently unable to resolve whether amoebozoan slime moulds share a single origin (forming a clade called Mycetozoa) or whether dictyostelians and myxogastreans independently originated from closely related but separate amoeboid ancestors.



3 - Protostelia: Three small families of slime moulds, the Protosteliidae, Cavosteliidae and Ceratiomyxidae, form a spreading nutritive phase similar to that of the Myxogastrea, and have often been regarded as closely related to the ultrastructurally similar myxogastreans. However, while myxogastreans form a truly acellular plasmodium, different protostelians form a pseudoplasmodium, with cells retaining their individual identity (Protosteliidae and Cavosteliidae), or a plasmodium that breaks up into individual cells before sporangium formation (Ceratiomyxidae). Ceratiomyxidae form small coral-like fruiting bodies - the photo above by Keisotyo shows a Ceratiomyxa species - while Protosteliidae form minute sporangia on slender stalks like dictyostelians. If mycetozoans form a single group, protostelians may represent a morphological connection between the cellular dictyostelians and the acellular myxogastreans. A relationship between protostelians and other mycetozoans was supported by Baldauf (1999), but the group remains little studied. The protostelians themselves are of doubtful monophyly, and some families may be closer to myxogastreans than others.

4 - Buddenbrockia: One parasitic animal was only recently identified as having a slime mould-like life cycle, with disassociated cells in its host aggregating together to give rise to a worm-like reproductive stage. The sordid details were covered in an earlier post.



5 - Acrasea: Acrasids are cellular slime moulds like dictyostelians, and indeed were once united with dictyostelians under the name of Acrasiomycetes. Like dictyostelians, acrasids live as individual amoeboids that aggregate together to form raised sporangia. The photo above from here shows the fruiting bodies of the best-known acrasid, Acrasis. However, acrasids are ultrastructurally distinct from dictyostelians, and are not even amoebozoans - rather, they belong to a protozoan group called Heterolobosea that also includes Naegleria, the organism that causes amoeboid meningitis, and belongs to the Excavata eukaryote superclade. Whether or not it was due to the mistaken assumption that acrasids were closely related to the intensely studied Dictyostelium, or whether it was due to the fact that acrasids seem to be most often found growing on animal poo, studies of acrasids are laughably rare, and only Acrasis rosea appears to have received any recent attention.

6 - Labyrinthulea: The slime nets are members of the Heterokonta, the clade also including brown and golden algae (among others), and have been covered before at this site - twice, in fact.

One protist group, the Phytomyxa or Plasmodiophoromycota, has often been included with the slime moulds due to its formation of a plasmodium for part of its life-cycle. However, phytomyxans, which are parasites of plant roots belonging to the Rhizaria (the eukaryote superclade including foraminiferans and radiolarians), do not seem to have an aggregative phase of the life-cycle comparable to other slime moulds. The best known phytomyxan is Plasmodiophora brassicae, the cause of club root in cabbages and other brassicas.



7 - Myxococcales: Finally comes a group that has never been regarded as slime moulds, but which has a very similar life cycle. The reason why Myxococcales have never been lumped with slime moulds is because they are not eukaryotes of any kind, but bacteria. Myxococcales are saprobic bacteria generally found in soil. They are capable of gliding motility, a form of movement by means other than flagella, though the exact mechanism remains little known. When nutrient supplies run low, some species of Myxococcales are capable of swarming together in a similar manner to cellular slime moulds and releasing dispersive spores. Myxococcales are therefore one of the few groups of bacteria to have developed multicellularity.

REFERENCES

Baldauf, S. L. 1999. A search for the origins of animals and fungi: comparing and combining molecular data. American Naturalist 154 (S4): 178-188.

Cavalier-Smith, T., E. E.-Y. Chao & B. Oates. 2004. Molecular phylogeny of Amoebozoa and the evolutionary significance of the unikont Phalansterium. European Journal of Protistology 40: 21-48.