Who is hiding behind these square scales? or The mystery of the origin of square shell plates in testate amoebae

Contributed by Anush Kosakyan

 

It would be hard to find a testate amoeba lover who does not know the genus Quadrulella. These vase-shaped species are a very unique group within family Hyalospheniidae (Arcellinida) since they are capable of secreting their own square or rectangular shell scales (or plates). These scales are characterisstic only for this genus, while most other members of the family are predators that build their shell by using mainly recycled plates taken or scavenged from other testate amoebae (i.e. kleptosquamy, Lahr et al. 2015).

***

My story started in the summer of 2013, when Edward Mitchell returned from his holiday in South Africa with a bag of Sphagnum samples. The samples were full of Quadrulella shells, including species that had never been seen since their original description back in 1957. And that was it….”the right time” for single-cell barcoding of this group. From this material, we obtained very interesting results that have been published recently (Kosakyan et al., 2016).

The most surprising result was that organisms presenting square shell plates appeared not to be monophyletic: two different groups of taxa with square scales (genus Quadrulella and newly established genus Mrabella) were indeed placed far from each other on the mtCOI-based phylogenetic tree. We thought of two possible evolutionary scenarios to explain this situation. These two hypotheses depend on whether the square-shaped plates in an organism have an autogenous or exogenous origin. This is a relevant aspect because hyalospheniids are known to engage in the behaviour of kleptosquamy, or recycling of shell plates produced by other organisms (Lahr et al., 2015).

Assuming that all amoebae that present square-shaped plates in their shells have produced them autogenously, the phylogenetic hypothesis presented here indicates homoplasy:

1) either the ability to produce square plates is an ancestral character that has been lost in a number of lineages (reversion); or

2) this character has evolved more than once independently (convergence).

We have assumed that convergence is the most parsimonious alternative (see the detailed discussion in the paper for further detail), with only two steps required – one event in genus Quadrulella and another independent event in genus Mrabella. Although we should not forget another unrelated group of organisms that are capable of producing square plates: the shelled amoeba Paraquadrulla is capable of producing calcareous square plates (the genus has not been sequenced yet, but while it is almost certainly an Arcellinid it is most likely unrelated to hyalosphenids or at best branches in a basal position to the whole group).

Alternatively, considering the fact that majority of hyalosphenids have the ability to scavenge plates from prey and use them to make the shell (kleptosquamy), a possible scenario is that Mrabella is in fact using scales from preyed Quadrulella, or scavenging these scales from the environment to construct the shell. And indeed, this was our first guess when we saw the description of Nebela galeata (current name Gibbocarina galeata) from Africa by Gauthier-Lièvre (1957). It is almost identical to Q. subcarinata in general shape and dimensions of the test (L=180-200, B=98-144, A=31-41 μm) (see Figure). Gauthier-Lièvre, 1957 (in her Fig. 10A) showed that square plates typical for Quadrulella can be integrated in the shell of N. galeata.  This is very easy to explain since Gauthier-Lièvre (1957) documented other Quadrulella species from the same locality where she found N. galeata and Q. subcarinata. Thus, other Quadrulella species could have provided the square plates use by these two species.

 fig10

Legend: A- Line drawing of Gibbocarina (Nebela) galeata from Congo by Gauthier-Lievre, 1957. B and C– Scanning electron microscopy and light microscopy micrographs of Mrabella subcarinata from South Africa. Scale bars =50 µm (in A) and 20 µm (in B). Modified from Kosakyan et al. 2016.

 

However,  things are not that easy in our case for two reasons:

1) Mrabella would specifically select these plates, a trait that is not known for any other hyalosphenid presenting kleptosquamy; these species generally use a mixture of plates from different origins to build their shell (see for instance: Apodera vas in Fig. 69 in Meisterfeld 2002);

2) if not specifically selecting, then Quadrulella would have to be the most abundant prey organism, or Quadrulella plates would have to be the most abundant in the environment, both options are likely not true since Quadrulella tends to be present in low abundance in comparison to Euglypha (although Euglypha was not abundant in our sample where we found Mrabella subcarinata), a genus of filose testate amoebae which is generally abundant and produces oval and ornamented siliceous plates.

Additionally, there are a number of other described species in genus Quadrulella that differ from the tear-shaped morphology of Quadrulella s.str. These also present similar shapes to other genera: Quadrulella vas, Q. constricta Apodera vas, Q. lageniformis Padaungiella lageniformis, Q. tubulata P. tubulata. These “mirror” species could either be a result of convergent evolution or alternatively represent cases of the “classical” hyalosphenids (A. vas, Padaungiella ssp.) that live in environments where euglyphids are rare but Quadrulella are abundant enough to provide material for building their shells. We have suggested that at this point these Quadrulella species must be treated as incertae sedis, and their sequencing will certainly illuminate the conundrum of the evolution of square-shaped plates.

References

Gauther-Lievre L. 1957.  Additions aux Nebela d’Afrique. Bull. Soc. Hist. Nat. l’Afrique du Nord 48, 494-523.

Kosakyan, A., Lahr, D. J. G., Mulot, M., Meisterfeld, R., Mitchell, E. A. D. and Lara, E. 2016. Phylogenetic reconstruction based on COI reshuffles the taxonomy of hyalosphenid shelled (testate) amoebae and reveals the convoluted evolution of shell plate shapes. Cladistics, 32: 606-623. doi:10.1111/cla.12167

 

What’s in a name? Something (completely different) to be said about taxonomic nomenclature

By Edward A. D. Mitchell,

Laboratory of Soil Biodiversity, University of Neuchâtel, Switzerland

With the advent of high throughput sequencing, estimates of global diversity are being totally revised as well.  For us protistologists – arguably much more importantly – so is the picture of how diversity is distributed among the different branches of the tree of life. The image that emerges is one that shows a huge unknown diversity among protists, at all levels, from major groups (i.e. “environmental clades”) and within known groups (i.e. from more or less divergent groups to complexes of cryptic and pseudo-cryptic species). This is fascinating and to say the least mind-boggling and a much welcome development for making a case about the need to study protists more intensively. It is indeed impossible today to ignore this diversity and the many functional roles that protists play in all ecosystems.

But this unknown diversity also calls for a massive investment in taxonomy. And as we all know there are only few active protist taxonomists. We therefore need to train a new generation of taxonomists to meet the huge challenge of keeping up with the novel discoveries resulting from molecular studies. And indeed, combining molecular and traditional microscopy approaches is the key to doing this job properly.

But there is also an often overlooked but important aspect of taxonomy, besides the critically important fact that descriptions need to be done correctly in order not to make a huge mess of nomenclature: naming a species. Choosing an appropriate name is indeed not always easy. Should we name a species after an esteemed colleague, the geographical location where the species was found, a morphological feature of the species, or should we try to find a name that also allows non-specialists to relate to the species – thus providing an excellent opportunity to increase the impact of the finding and making a broader audience aware of the sheer existence of our beloved amoebae?

I firmly believe that witty names are useful. They make us happy, allow many lively discussions to take place among colleagues and are much appreciated by journalists always keen to report on “something completely different” (Monty Python, 1971).

The choice of names tells a lot about the personality and cultural references of the author. Here are three recent examples among testate amoebae:

Padaungiella Lara & Todorov 2012.

This genus of hyalosphenid testate amoebae is characterised by an elongated neck (Kosakyan et al., 2012). The idea for this name came to me while riding my bicycle (a great source of inspiration!). I knew about the existence of some African tribes that used metal necklaces to elongate the necks of woman (allegedly, if the woman cheated on her man he would remove the necklaces and she would die due to cervical spine injury). I did a bit of research on this and found out that there were two unrelated tribes using such necklaces, one in Africa, with independent circular necklaces piled on top of each other and one in Asia (Padaung – https://en.wikipedia.org/wiki/Kayan_people_(Myanmar)), with a single spiral necklace. The latter corresponded much better to the shape of the amoeba shell and hence we decided to choose this name.

At the time of writing this post there were 1’110 hits for Padaungiella in Google.

padaungiella

Padaungiella wailesii (left, picture by E. Mitchell) and a Padaung woman (right, source: http://www.chiangdao.com/chiangmai/karenlongneck.htm)

Nebela gimlii Singer & Lara 2015.

This species was named after Gimli (http://lotr.wikia.com/wiki/Gimli), a dwarf in Tolkien’s Lord of the Rings saga (Singer et al., 2015). This dwarf wanders in forests during the saga, something dwarfs are not supposed to do much. This species being the smallest of the species complex and being found in forested bogs the name seemed appropriate. Newspapers as far as Austria wrote about this. http://www.krone.at/wissen/amoebenart-nach-figur-aus-herr-der-ringe-benannt-in-torfmoor-entdeckt-story-497678.

At the time of writing this post there were 188 hits for “Nebela gimlii” in Google.

Arcella gandalfi Féres, Porfírio-Sousa, Ribeiro, Rocha, Sterza, Souza, Soares & Lahr 2016.

This large Arcella species bears striking resemblance to Gandalf’s hat and thus logically was named after this even more famous character of the Lord of the Rings (Féres et al., 2016; Tolkien, 1954) (http://lotr.wikia.com/wiki/Gandalf).

arcella-gandalfii

Arcella gandalfi (left, from Féres et al., 2016) and Gandalf, as played by Ian McKellen in The Lord of the Rings triology, (right, source: http://lotr.wikia.com/wiki/Gandalf).

The media picked up on this story even more and this illustrates again how a well-chosen name can significantly contribute to making our field of research more visible.

Arcella gandalfi” has 31’600 hits on Google at the time of writing this post. To this date, news agencies in more than 15 countries reported on this, including: Brazil, USA, Germany, France, Russia, India, Mexico, Spain, Turkey, Argentina, Hungary, Indonesia, Croatia, South Korea, Ukraine.

Gandalf clearly wins! Well done fellows!

Who’s next? There’s no end to the fun!

References

Féres, J.C., Porfírio-Sousa, A.L., Ribeiro, G.M., Rocha, G.M., Sterza, J.M., Souza, M.B.G., Soares, C.E.A., Lahr, D.J.G., 2016. Morphological and Morphometric Description of a Novel Shelled Amoeba Arcella gandalfi sp. nov. (Amoebozoa: Arcellinida) from Brazilian Continental Waters Acta Protozool. 55(4).

Kosakyan, A., Heger, T.J., Leander, B.S., Todorov, M., Mitchell, E.A.D., Lara, E., 2012. COI Barcoding of Nebelid Testate Amoebae (Amoebozoa: Arcellinida): Extensive Cryptic Diversity and Redefinition of the Hyalospheniidae Schultze. Protist 163, 415-434.

Monty Python, 1971. And Now for Something Completely Different, in: MacNaughton, I. (Ed.), Monty Python’s Flying Circus. Columbia Pictures, United Kingdom, p. 95 minutes.

Singer, D., Kosakyan, A., Pillonel, A., Mitchell, E.A.D., Lara, E., 2015. Eight species in the Nebela collaris complex: Nebela gimlii (Arcellinida, Hyalospheniidae), a new species described from a Swiss raised bog. European Journal of Protistology 51, 79-85.

Tolkien, J.R.R., 1954. The Fellowship of the Ring, The Lord of the Rings, Boston. Ballantine Books, New York.

A new approach to testate amoeba paleoecology: reconstructing past environmental conditions from morpholological traits

Contributed by Edward A. D. Mitchell

Laboratory of Soil Biodiversity, University of Neuchâtel, Switzerland

Testate amoebae are well known to be good indicators of micro-environmental gradients and especially soil moisture, water table depth and pH. This has been known since the early 20th century (Harnisch, 1925). The predictable distribution patterns of many species not only systematically result in seeing these variables emerge as being significantly correlated to testate amoeba community data in ecological studies but also allow the development of inference models – so called “transfer functions” – based on testate amoeba community data to reconstruct (infer) these variables either from modern samples or more generally from subfossil communities extracted from peat deposits or lake sediments. As there are few proxies to reconstruct past hydrological changes, testate amoebae have become part of the standard toolbox of palaeoecologists (Charman, 2001; Mitchell et al., 2008). Indeed, developing and using such transfer function has been the main reason to study testate amoebae and indeed most papers dealing with testate amoebae published in the last couple of decades. These organisms were thus being used as tools using correlative approaches rather than considered as study subjects in their own right.
Simon van Bellen and colleagues have just published a paper in which they present a new approach for palaeo-environmental inference based on testate amoeba morphological traits rather than community data (van Bellen et al., 2017). They used nine different traits and related them to the depth to the water table (DWT), the variable most commonly inferred from testate amoebae. Their data set is from Tierra-del-Fuego, but could almost be from anywhere in the World where Sphagnum peatlands occur. The list of traits includes morphological traits such as biovolume, aperture position, test compression, aperture size and test composition, a binary physiological trait (mixotrophy vs. heterotrophy) and two phylogenetic traits (Arcellinida and Euglyphida – there were no Amphitremids in the data). Most traits showed a relationship to DWT. For example, Arcellinida show an almost perfect negative linear correlation to DWT (i.e. the propostion of Arcellinida decreases with increasing DWT, in other words, the wetter the habitat, the more Arcelinida dominate the community). By contrast Euglyphida show a positive correlation to DWT (so the drier it gets the more the testate amoeba community is dominated by Euglyphida).

vabellen

An extract from Figure 3 of van Bellen et al. 2017. Relationship between the community weighed means of Arcellinida (top) and Euglyphida (bottom) vs depth to water table (DWT, horizontal axis – left is wet, right is dry). The two phylogenetic groups clearly show opposite responses to the humidity gradient.

Interestingly this pattern corresponds to a ratio used in mineral soil, the LF index (Bonnet, 1976), where “L” stands for Lobosea (Arcellinida) and “F” stands for Filosa (Euglyphida). By analogy to MacArthur and Wilson’s concept of r (ruderal) and K (competitors) life history classification (MacArthur and Wilson, 1967), Euglyphida are considered as r strategists while Arcellinida are on average more K strategists. Euglyphids are on average smaller and thus more likely to feed on smaller prey such as bacteria. Being smaller also means that they may tolerate dry conditions better than the generally larger arcellinids that require a thicker water film to move.

The study of testate amoebae functional traits is clearly rapidly becoming a dynamic field of research within our community (Arrieira et al., 2015; Fournier et al., 2016; Fournier et al., 2015; Fournier et al., 2012; Jassey et al., 2016; Jassey et al., 2015; Lamentowicz et al., 2015; Marcisz et al., 2016; Marcisz et al., 2014). It will be interesting to follow the next developments in this area!

References

Arrieira, R.L., Schwind, L.T.F., Bonecker, C.C., Lansac-Toha, F.A., 2015. Use of functional diversity to assess determinant assembly processes of testate amoebae community. Aquatic Ecology 49, 561-571.

Bonnet, L., 1976. Le peuplement thécamoebien édaphique de la Côte-d’Ivoire. Sols de la région de Lamto. Protistologica 12, 539-554.

Charman, D.J., 2001. Biostratigraphic and palaeoenvironmental applications of testate amoebae. Quaternary Science Reviews 20, 1753-1764.

Fournier, B., Coffey, E.E.D., van der Knaap, W.O., Fernandez, L.D., Bobrov, A., Mitchell, E.A.D., 2016. A legacy of human-induced ecosystem changes: spatial processes drive the taxonomic and functional diversities of testate amoebae in Sphagnum peatlands of the Galapagos. Journal of Biogeography 43, 533-543.

Fournier, B., Lara, E., Jassey, V.E.J., Mitchell, E.A.D., 2015. Functional traits as a new approach for interpreting testate amoeba palaeo-records in peatlands and assessing the causes and consequences of past changes in species composition. Holocene 25, 1375-1383.

Fournier, B., Malysheva, E., Mazei, Y., Moretti, M., Mitchell, E.A.D., 2012. Toward the use of testate amoeba functional traits as indicator of floodplain restoration success. Eur. J. Soil Biol. 49, 85-91.

Harnisch, O., 1925. Studien zur Ökologie und Tiergeographie der Moore. Zoologisch Jahrbuch (Abteilung Systematik) 51, 1-166.

Jassey, V.E.J., Lamentowicz, M., Bragazza, L., Hofsommer, M.L., Mills, R.T.E., Buttler, A., Signarbieux, C., Robroek, B.J.M., 2016. Loss of testate amoeba functional diversity with increasing frost intensity across a continental gradient reduces microbial activity in peatlands. European Journal of Protistology 55, Part B, 190-202.

Jassey, V.E.J., Signarbieux, C., Haettenschwiler, S., Bragazza, L., Buttler, A., Delarue, F., Fournier, B., Gilbert, D., Laggoun-Defarge, F., Lara, E., Mills, R.T.E., Mitchell, E.A.D., Payne, R.J., Robroek, B.J.M., 2015. An unexpected role for mixotrophs in the response of peatland carbon cycling to climate warming. Scientific Reports 5, 16931.

Lamentowicz, M., Gałka, M., Lamentowicz, Ł., Obremska, M., Kühl, N., Lücke, A., Jassey, V.E.J., 2015. Reconstructing climate change and ombrotrophic bog development during the last 4000 years in northern Poland using biotic proxies, stable isotopes and trait-based approach. Palaeogeography, Palaeoclimatology, Palaeoecology 418, 261-277.

MacArthur, R.H., Wilson, E.O., 1967. The Theory of Island Biogeography. Princeton University Press.

Marcisz, K., Colombaroli, D., Jassey, V.E.J., Tinner, W., Kołaczek, P., Gałka, M., Karpińska-Kołaczek, M., Słowiński, M., Lamentowicz, M., 2016. A novel testate amoebae trait-based approach to infer environmental disturbance in Sphagnum peatlands. Scientific Reports 6, 33907.

Marcisz, K., Lamentowicz, L., Slowinska, S., Slowinski, M., Muszak, W., Lamentowicz, M., 2014. Seasonal changes in Sphagnum peatland testate amoeba communities along a hydrological gradient. European Journal of Protistology 50, 445-455.

Mitchell, E.A.D., Charman, D.J., Warner, B.G., 2008. Testate amoebae analysis in ecological and paleoecological studies of wetlands: past, present and future. Biodivers. Conserv. 17, 2115-2137.

van Bellen, S., Mauquoy, D., Payne, R.J., Roland, T.P., Hughes, P.D.M., Daley, T.J., Loader, N.J., Street-Perrott, F.A., Rice, E.M., Pancotto, V.A., 2017. An alternative approach to transfer functions? Testing the performance of a functional trait-based model for testate amoebae. Palaeogeography, Palaeoclimatology, Palaeoecology 468, 173-183.