Ascetosporea: Ornate spore-bearers of doom

Yana Eglit

Ph.D. Candidate, Department of Biology, Dalhousie University, Halifax, Nova Scotia, Canada

If you studied any biology, you have probably heard of the three domains of life: the eukaryotes, i.e. a group of organisms, or taxon, with a true nucleus (plants, animals, fungi, and “protists”), as well as those without a nucleus, the bacteria and the lesser known archaea (latest data suggest eukaryotes are essentially a very derived ‘type of archaea’). It might surprise many, that plants, animals and fungi are but twigs on the eukaryote tree of life (Fig. 1; in blue). In fact, the eukaryotes are nothing but a small twig in the largely bacteria-dominated diversity of life (Simpson & Eglit 2016). Most posts in this Taxon of the Month series, most diversity of life you’re intuitively familiar with, most nature you see around you by the naked eye, exists entirely on these three ‘tiny’ twigs. All other eukaryotes are “protists”, and the vast majority of these “protists” are small, and thus invisible to the naked eye. However, in the ocean, protists can get big, in the form of seaweeds. Kelps, red algae, and green algae are all considered “protists” (see Phaeophyta, Rhodophytes, and Chlorophyceae in Fig. 1, respectively), and some of these can also be quite tasty!

  Figure 1 . Tree of Eukaryotes (modified from  Simpson & Eglit 2016 ) showing Plants, Animals, and Fungi in blue and Ascetosporea in red. Everything not in blue text is typically considered a ‘protist’.

Figure 1. Tree of Eukaryotes (modified from Simpson & Eglit 2016) showing Plants, Animals, and Fungi in blue and Ascetosporea in red. Everything not in blue text is typically considered a ‘protist’.

Now, the point isn’t that plant and animal diversity is somehow dull, but rather that if this is just a very small part of the overall diversity within eukaryotes. Just try to imagine the breathtaking diversity of the rest of it! And with new major branches of this tree found or established each year (Brown et al. 2013; Seenivasan et al. 2013Janouškovec et al. 2017), one can only dream of what alien-like lifeforms remain unknown to humankind! Developments in sequencing technology have accelerated this rate of discovery to unprecedented levels – and at least a handful more entirely new types of eukaryotes will be published in the next few years.

Today, we will look at one such neglected and poorly-known group that is so esoteric, its formal scientific name means “elaborately-crafted spore bearers” – Ascetosporea (from Greek asketos: ‘well-made, elaborate, ornate’). Their ornateness probably brings little comfort to the hundreds if not thousands of livelihoods that can be wiped out by these parasites – they are highly efficient parasites of economically-important shellfish, sometimes causing absolute mortality and total collapse of the fishery.

Ascetosporea (Fig. 1; in red) find their phylogenetic home within a little-known supergroupnamed Rhizaria. Previously a superficially eccentric assemblage of vastly different-looking organisms only supported by molecular data, this group is also becoming increasingly riddled with parasites. Ascetosporeans comprise two groups: Haplosporidia, known for spores like lidded jars, and Paramyxids, with an incomprehensible inward division resulting in a cellular matryoshka.

  Figure 2 . Haplosporidian spores:  A ) transmission electron microscopy (TEM) image showing a cross section of a  Haplosporidium  sp. spore ( Feist et al. 2009 );  B ) diagram of a mature  Haplosporidium montforti  spore ( Azevedo et al. 2006 );  C ) scanning electron microscopy image of a  Minchinia mercenariae  spore, arrow pointing to the opened “lid” and the opening below ( Ford et al. 2009 ).

Figure 2. Haplosporidian spores: A) transmission electron microscopy (TEM) image showing a cross section of a Haplosporidium sp. spore (Feist et al. 2009); B) diagram of a mature Haplosporidium montforti spore (Azevedo et al. 2006); C) scanning electron microscopy image of a Minchinia mercenariae spore, arrow pointing to the opened “lid” and the opening below (Ford et al. 2009).

Haplosporidia are parasites to a variety of marine invertebrates like molluscs (including the oysters), crustaceans, echinoderms, and even other parasites (Azevedo & Hine 2017). The infection starts out with solitary uninucleate cells invading and gradually forming a multinucleate ooze (plasmodium) growing among the host tissues. The plasmodium splits up into smaller pieces (sporonts), then forming walls of membrane centred on each nucleus (up to hundreds!), and (in most species) proceeds to build spores (Azevedo & Hine 2017). Haplosporidian spores feature jars or sometimes handle-less amphorae with hinged ‘lids’ and various surface decorations like long tails or tassels and surface reliefs (Fig. 2, Ford et al. 2009). Inside each vessel is a nucleus, haplosporosomes (some harpoon-like organelles of uncertain function) and a peculiar “spherulosome”, a complex clump of membranes. Once inside an appropriate next victim, the lid then opens (Fig. 2c, arrow), releasing newborn parasites to graze happily on their new host. Haplosporidians are mysterious and fascinating in their own right, just remember that somewhere out there are cellular oozes currently making little microscopic pottery in which to spit out its progeny.

Paramyxids are parasites of various marine invertebrates ranging from molluscs and crustaceans, including many commercial ‘shellfish’, to polychaete worms and even tunicates. Once inside their host, paramyxids proliferate throughout tissues, devouring their host from within, and then sporulate primarily in digestive and reproductive organs (Fig. 3a, Lester & Hine 2017). This sporulation is distinctive for paramyxids, namely their remarkable formation of spores inside cells. The process is quite complicated and not at all how we learned mitosis in school, as the daughter cell is formed inside the mother, like inverse budding (Fig. 3b,c). As far as I know, the only other cells that like that are pollen forming cells in plants. Species differ in how many inverse buddings they undergo (Feist et al. 2009), and to keep things simple I’ll just go over the most complex case: Paramyxa paradoxa (Audemard et al. 2002; Stentiford et al. 2017; Fig. 4).

  Figure 3 . Paramyxid appearance:  A ) an infected oyster ( Desportes 1984 );  B ) general view (TEM) of  Paramyxa nephtys  cells in various stages of spore formation  within  polychaete gut lining cells – this species develops inside host cells rather than between them! Pink outlines roughly correspond to host cells, yellow/orange and blue concentric outlines show layers of paramyxid cells (modified from  Audemard et al. 2002 );  C ) detail of  P. nephtys  tertiary cell (C3) with C4-C5 internal cells nested inside ( Audemard et al. 2002 ). For images of mature spores see  Audemard et al. 2002  (free access).  D ) Primary cell (C1) of  Paramarteilia canceri  with sporonts and spores (colours corresponding to those in Fig. 4; modified from  Larsson & Køie 2005 ). Note the dark blob and rod-like structures in C3: those are thought to be related to the dark-staining structures in haplosporidia called haplosporosomes labelled ‘Hs’ in Fig. 2a.

Figure 3. Paramyxid appearance: A) an infected oyster (Desportes 1984); B) general view (TEM) of Paramyxa nephtys cells in various stages of spore formation within polychaete gut lining cells – this species develops inside host cells rather than between them! Pink outlines roughly correspond to host cells, yellow/orange and blue concentric outlines show layers of paramyxid cells (modified from Audemard et al. 2002); C) detail of P. nephtys tertiary cell (C3) with C4-C5 internal cells nested inside (Audemard et al. 2002). For images of mature spores see Audemard et al. 2002 (free access). D) Primary cell (C1) of Paramarteilia canceri with sporonts and spores (colours corresponding to those in Fig. 4; modified from Larsson & Køie 2005). Note the dark blob and rod-like structures in C3: those are thought to be related to the dark-staining structures in haplosporidia called haplosporosomes labelled ‘Hs’ in Fig. 2a.

An amoeboid primary cell (C1) crawls between the host cells and multiplies until it is ready to host the spore construction action. It then divides endogenously (inward) to form a secondary cell (C2), which itself divides ‘normally’ twice to form four (Fig. 4a). The precise mechanism of this idiosyncratic endogenous division remains mysterious!  Now, each of the secondary cells repeats the process above to form four internal tertiary cells (C3; Fig. 4a). From this point on, we will no longer see conventional mitosis, only inward budding: each of the tertiary cells divides endogenously four times, forming a four-cell-layered spore (C4-C6; Fig. 4a). To tally up the math: C1 + 4C2*4C3*4(C4-C6) = 4x(4x4+1)+1 = 69 cells and nuclei total! (Fig. 4b) And to top it all off, sometimes the primary cell can have a parasite of its own (Stentiford et al. 2017)!

The primary and secondary cells then decay away to release the C3s (spores) (Fig. 4c), which in some cases undergo further maturation by forming a cell wall and a striated “tail” (Fig. 4d; Larsson & Køie 2005). After such convoluted origins, where do these newborn spores go next? As the paramyxid life cycle progresses, our understanding of it drops approximately exponentially, and it was only in the oyster parasite Marteilia refringens when the next stage for only one species was determined to be a copepod – by cleverly taking advantage of a minimalist (low species-richness) lagoon ecosystem and molecularly testing each species of invertebrate for signs of the parasite (Audemard et al. 2002). For the rest of paramyxids – the few that are discovered – we have no idea. It is unclear how the spores from copepod return to the oyster, as experimental infection attempts have failed (Carrasco et al. 2008), and there does appear to be a seasonal component (Boyer et al. 2013).

  Figure 4 . Spore formation ( A , B ) and maturation ( C , D ) in among the most complicated known paramyxids,  Paramyxa (= Paramyxoides )  nephtys  (based on  Audemard et al. 2002 ;  Stentiford et al. 2017 ). Can you count all 69 cells in the final sporont?

Figure 4. Spore formation (A,B) and maturation (C,D) in among the most complicated known paramyxids, Paramyxa(=Paramyxoides) nephtys (based on Audemard et al. 2002; Stentiford et al. 2017). Can you count all 69 cells in the final sporont?

There are about a couple dozen or so people in the world who work on Paramyxids, largely government scientists who monitor the health of fisheries. Perhaps another dozen work on Haplosporidia. Despite obvious economic importance, obscure protist parasites do not receive much attention or funding. Even oomycetes, a group containing a plant pathogen from which the population of Ireland has yet to recover (Potato Famine), are an acquired taste.

These off-the-beaten path protist parasites are likely relevant to a variety of lesser-studied crops outside Europe, North America and East Asia, and thus potentially important for food security. Without well-understood evolutionary contexts and basic biology, it would be difficult to manage these organisms when they turn to being our nightmares. I hope to have piqued a little bit of interest in these odd and diverse parasites, and maybe even in their just-as-fascinating free-living relatives too. Just remember that every bit of awe-some biodiversity you learn about comes with its own assortment of parasites, some perhaps equipped with their very own surrealist spores.

About the Author

Yana Eglit is a protistology Ph.D. Candidate in the lab of Alastair Simpson (Department of Biology, Dalhousie University, Canada) studying evolution and cell biology of understudied and novel deep lineages of eukaryotes (i.e., “very weird protists”).

Acknowledgements

I would like to thank Noèlia Carrasco (IRTA), IFREMER, and the paramyxid community for the 2015 workshop invite immersing me in the Paramyxean world, and Aaron L. Beek (University of Memphis) for clarifying the nuanced meanings of asketos in Ancient Greek.

References

Audemard, C. 2002. Needle in a haystack: involvement of the copepod Paracartia grani in the life-cycle of the oyster pathogen Marteilia refringens. Parasitol. 124: 315–323.

Azevedo, C. & P. M. Hine. 2017. Haplosporidia. In Handbook of the Protists (J. Archibald, A.G.B. Simpson, C. Slamovits, Eds.).

Azevedo, C., P. Balseiro, G. Casal, C. Gestal, R. Aranguren, N.A. Stokes, R.B. Carnegie, B. Novoa, E.M. Burreson & A. Figueras. 2006. Ultrastructural and molecular characterization of Haplosporidium montforti n. sp., parasite of the European abalone Haliotis tuberculate. J. Invert Pathol. 92: 23–32.

Boyer, S., B. Chollet, D. Bonnet & I. Arzul. 2013. New evidence for the involvement of Paracartia grani (Copepoda, Calanoida) in the life cycle of Marteilia refringens (Paramyxea). Int. J. Parasitol. 43: 1089–1099.

Brown, M.W., S.C. Sharpe, J.D. Silberman, A.A. Heiss, B.F. Lang, A.G. Simpson & A.J. Roger. 2013. Phylogenomics demonstrates that breviate flagellates are related to opisthokonts and apusomonads. Proc. Biol. Sci. 289: 20131755.

Carrasco, N., I. Arzul, I. B. Chollet, M. Robert, J.P. Joly, M.D. Furones & F.C.J. Berthe. 2008. Comparative experimental infection of the copepod Paracartia grani with Merteilia refringens and Marteilia maurini. J. Fish Dis. 31: 497–504.

Desportes, I. 1984. The Paramyxea Levine 1979: An original example of 10volution towards multicellularity. Origin of Life 13: 343–352.

Feist, S.W., P.M. Hine, K.S. Bateman, G.D. Stentiford & M. Longshaw. 2009. Paramerteilia canceri sp. n. (Cercozoa) in the European edible crab (Cancer pagurus) with a proposal for the revision of order Paramyxids Chatton, 1911. Folia Parasitol. 56: 73–85.

Ford, S.E., N.A. Stokes, E.M. Burreson, E. Scarpa, R.B. Carnegie, J.N. Kraeuter & D. Bushek. 2009. Minchinia mercenariae n. sp. (Haplosporidia) in the hard clam Mercenaria mercenaria: implications of a rare parasite in a commercially important host. J. Euk. Microbiol. 56: 542–551.

Janouškovec, J., D.V. Tikhonenkov, F. Burki, A.T. Howe, F.L. Rohwer, A.P. Mylnikov & P.J. Keeling. 2017. A New Lineage of Eukaryotes Illuminates Early Mitochondrial Genome Reduction. Curr. Biol. 27: p3717–3724.

Larsson, J.I.R. & M. Køie. 2005. Ultrastructural study and description of Paramyxoides nephtys gen. n., sp. n. a parasite of Nephtys caeca (Fabricius, 1780) (Polychaeta: Nephtyidae). Acta Protozool. 44: 175–187.

Lester, R.J.G. & P.M. Hine. 2017. Paramyxida. in Handbook of the Protists (eds. Archibald J., Simpson AGB., Slamovits C.).

Seenivasan, R., N. Sausen, L.K. Medlin & M. Melkonian. 2013. Picomonas judraskeda Gen. et sp. Nov: The first identified member of the Picozoa Phylum Nov., a widespread group of picoeukaryotes, formerly known as ‘Picobiliphytes’. PLoS ONE 8: e59565.

Simpson, A.G.B. &. Y. Eglit. 2016. Protist diversification. Encyclopedia of Evolutionary Biology 3: 344–360.

Stentiford, G.D., A. Ramilo, E. Abollo & R. Kerr. 2017. Hyperspora aquatica n.gn., n.sp. (Microsporidia), hyperparasitic in Marteilia cochillia (Paramyxida), is closely related to crustacean-infecting microspordian taxa. Parasitol. 144: 186–199.