Organelles that illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes


Many anaerobic microbial parasites possess highly modified mitochondria known as mitochondrion-related organelles (MROs). The best-studied of these are the hydrogenosomes of Trichomonas vaginalis and Spironucleus salmonicida, which produce ATP anaerobically through substrate-level phosphorylation with concomitant hydrogen production; and the mitosomes of Giardia intestinalis, which are functionally reduced and lack any role in ATP production. However, to understand the metabolic specializations that these MROs underwent in adaptation to parasitism, data from their free-living relatives are needed. Here, we present a large-scale comparative transcriptomic study of MROs across a major eukaryotic group, Metamonada, examining lineage-specific gain and loss of metabolic functions in the MROs of Trichomonas, Giardia, Spironucleus and their free-living relatives. Our analyses uncover a complex history of ATP production machinery in diplomonads such as Giardia, and their closest relative, Dysnectes; and a correlation between the glycine cleavage machinery and lifestyles. Our data further suggest the existence of a previously undescribed biochemical class of MRO that generates hydrogen but is incapable of ATP synthesis.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Phylogeny of Metamonada and distribution of MRO-localizing proteins.
Figure 2: Predicted mitochondrial protein import proteins in metamonads.
Figure 3: Evolutionary transitions of metabolic pathways of MROs from the common ancestor of Parabasalia and Fornicata to representative extant species.


  1. 1

    Stairs, C. W., Leger, M. M. & Roger, A. J. Diversity and origins of anaerobic metabolism in mitochondria and related organelles. Phil. Trans. R. Soc. Lond. B 370, 20140326 (2015).

    Article  Google Scholar 

  2. 2

    Lindmar, k, D. G. & Müller, M. Hydrogenosome, a cytoplasmic organelle of the anaerobic flagellate Tritrichomonas foetus, and its role in pyruvate metabolism. J. Biol. Chem. 248, 7724–7728 (1973).

    Google Scholar 

  3. 3

    Müller, M. et al. Biochemistry and evolution of anaerobic energy metabolism in eukaryotes. Microbiol. Mol. Biol. Rev. 76, 444–495 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  4. 4

    Jerlstrom-Hultqvist, J. et al. Hydrogenosomes in the diplomonad Spironucleus salmonicida. Nat. Commun. 4, 2493 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  5. 5

    Jedelsky, P. L. et al. The minimal proteome in the reduced mitochondrion of the parasitic protist Giardia intestinalis. PLoS ONE 6, e17285 (2011).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  6. 6

    Karnkowska, A. et al. A eukaryote without a mitochondrial organelle. Curr. Biol. 26, 1274–1284 (2016).

    CAS  Article  PubMed  Google Scholar 

  7. 7

    Takishita, K. et al. Multigene phylogenies of diverse Carpediemonas-like organisms identify the closest relatives of ‘amitochondriate’ diplomonads and retortamonads. Protist 163, 344–355 (2012).

    Article  PubMed  Google Scholar 

  8. 8

    Simpson, A. G. & Patterson, D. J. On core jakobids and excavate taxa: the ultrastructure of Jakoba incarcerata. J. Eukaryot. Microbiol. 48, 480–492 (2001).

    CAS  Article  PubMed  Google Scholar 

  9. 9

    Dolezal, P., Likic, V., Tachezy, J. & Lithgow, T. Evolution of the molecular machines for protein import into mitochondria. Science 313, 314–318 (2006).

    CAS  Article  PubMed  Google Scholar 

  10. 10

    Martincova, E. et al. Probing the biology of Giardia intestinalis mitosomes using in vivo enzymatic tagging. Mol. Cell. Biol. 35, 2864–2874 (2015).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  11. 11

    Neupert, W. & Herrmann, J. M. Translocation of proteins into mitochondria. Annu. Rev. Biochem. 76, 723–749 (2007).

    CAS  Article  PubMed  Google Scholar 

  12. 12

    Yaffe, M. P., Ohta, S. & Schatz, G. A yeast mutant temperature-sensitive for mitochondrial assembly is deficient in a mitochondrial protease activity that cleaves imported precursor polypeptides. EMBO J. 4, 2069–2074 (1985).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  13. 13

    Harris, S. R., Matus, A., Hrdy, I. & Kute , E. Reductive evolution of the mitochondrial processing peptidases of the unicellular parasites Trichomonas vaginalis and Giardia intestinalis. PLoS Pathog. 4, e1000243 (2008).

    Google Scholar 

  14. 14

    Garg, S. et al. Conservation of transit peptide-independent protein import into the mitochondrial and hydrogenosomal matrix. Genome Biol. Evol. 7, 2716–2726 (2015).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  15. 15

    Kikuchi, G. The glycine cleavage system: composition, reaction mechanism, and physiological significance. Mol. Cell. Biochem. 1, 169–187 (1973).

    CAS  Article  PubMed  Google Scholar 

  16. 16

    Mukherjee, M., Brown, M. T., McArthur, A. G. & Johnson, P. J. Proteins of the glycine decarboxylase complex in the hydrogenosome of Trichomonas vaginalis. Eukaryot. Cell 5, 2062–2071 (2006).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  17. 17

    Morrison, H. G. et al. Genomic minimalism in the early diverging intestinal parasite Giardia lamblia. Science 317, 1921–1926 (2007).

    CAS  Article  PubMed  Google Scholar 

  18. 18

    Zubacova, Z. et al. The mitochondrion-like organelle of Trimastix pyriformis contains the complete glycine cleavage system. PLoS ONE 8, e55417 (2013).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. 19

    Hampl, V. et al. Genetic evidence for a mitochondriate ancestry in the ‘amitochondriate’ flagellate Trimastix pyriformis. PLoS ONE 3, e1383 (2008).

    Article  PubMed  PubMed Central  Google Scholar 

  20. 20

    Nývltová, E., Smutná, T., Tachezy, J. & Hrdý, I. OsmC and incomplete glycine decarboxylase complex mediate reductive detoxification of peroxides in hydrogenosomes of Trichomonas vaginalis. Mol. Biochem. Parasitol. 206, 29–38 (2016).

    Article  PubMed  Google Scholar 

  21. 21

    Xu, F. et al. On the reversibility of parasitism: adaptation to a free-living lifestyle via gene acquisitions in the diplomonad Trepomonas sp. PC1. BMC Biol. 14, 62 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  22. 22

    Hampson, R. K., Barron, L. L. & Olson, M. S. Regulation of the glycine cleavage system in isolated rat liver mitochondria. J. Biol. Chem. 258, 2993–2999 (1983).

    CAS  PubMed  Google Scholar 

  23. 23

    Steinbüchel, A. & Müller, M. Anaerobic pyruvate metabolism of Tritrichomonas foetus and Trichomonas vaginalis hydrogenosomes. Mol. Biochem. Parasitol. 20, 57–65 (1986).

    Article  PubMed  Google Scholar 

  24. 24

    Van Hellemond, J. J., Klockiewicz, M., Gaasenbeek, C. P., Roos, M. H. & Tielens, A. G. Rhodoquinone and complex II of the electron transport chain in anaerobically functioning eukaryotes. J. Biol. Chem. 270, 31065–31070 (1995).

    CAS  Article  PubMed  Google Scholar 

  25. 25

    Tielens, A. G. M., van Grinsven, K. W. A., Henze, K., van Hellemond, J. J. & Martin, W. Acetate formation in the energy metabolism of parasitic helminths and protists. Int. J. Parasitol. 40, 387–397 (2010).

    CAS  Article  PubMed  Google Scholar 

  26. 26

    Sanchez, L. B. & Müller, M. Purification and characterization of the acetate forming enzyme, acetyl-CoA synthetase (ADP-forming) from the amitochondriate protist, Giardia lamblia. FEBS Lett. 378, 240–244 (1996).

    CAS  Article  PubMed  Google Scholar 

  27. 27

    Noguchi, F. et al. Metabolic capacity of mitochondrion-related organelles in the free-living anaerobic stramenopile Cantina marsupialis. Protist 166, 534–550 (2015).

    CAS  Article  PubMed  Google Scholar 

  28. 28

    Field, J., Rosenthal, B. & Samuelson, J. Early lateral transfer of genes encoding malic enzyme, acetyl-CoA synthetase and alcohol dehydrogenases from anaerobic prokaryotes to Entamoeba histolytica. Mol. Microbiol. 38, 446–455 (2000).

    CAS  Article  PubMed  Google Scholar 

  29. 29

    Nývltová, E. et al. Lateral gene transfer and gene duplication played a key role in the evolution of Mastigamoeba balamuthi hydrogenosomes. Mol. Biol. Evol. 32, 1039–1055 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  30. 30

    Schneider, R. E. et al. The Trichomonas vaginalis hydrogenosome proteome is highly reduced relative to mitochondria, yet complex compared with mitosomes. Int. J. Parasitol. 41, 1421–1434 (2011).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  31. 31

    Murcha, M. W., Narsai, R., Devenish, J., Kubiszewski-Jakubiak, S. & Whelan, J. MPIC: a mitochondrial protein import components database for plant and non-plant species. Plant Cell Physiol. 56, e10 (2015).

    Article  PubMed  Google Scholar 

  32. 32

    Zhang, Q. et al. Marine isolates of Trimastix marina form a plesiomorphic deep-branching lineage within Preaxostyla, separate from other known Trimastigids (Paratrimastix n. gen.). Protist 166, 468–491 (2015).

    Article  PubMed  Google Scholar 

  33. 33

    Kolisko, M. et al. A wide diversity of previously undetected free-living relatives of diplomonads isolated from marine/saline habitats. Env. Microbiol. 12, 2700–2710 (2010).

    CAS  Google Scholar 

  34. 34

    Chevreux, B. et al. Using the miraEST assembler for reliable and automated mRNA transcript assembly and SNP detection in sequenced ESTs. Genome Res. 14, 1147–1159 (2004).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  35. 35

    Grabherr, M. G. et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat. Biotechnol. 29, 644–652 (2011).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  36. 36

    Gentekaki, E. et al. Large-scale phylogenomic analysis reveals the phylogenetic position of the problematic taxon Protocruzia and unravels the deep phylogenetic affinities of the ciliate lineages. Mol. Phylogenet. Evol. 78, 36–42 (2014).

    CAS  Article  PubMed  Google Scholar 

  37. 37

    Katoh, K. & Standley, D. M. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol. Biol. Evol. 30, 772–780 (2013).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. 38

    Criscuolo, A. & Gribaldo, S. BMGE (Block Mapping and Gathering with Entropy): a new software for selection of phylogenetic informative regions from multiple sequence alignments. BMC Evol. Biol. 10, 210 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  39. 39

    Price, M. N., Dehal, P. S. & Arkin, A. P. FastTree: computing large minimum evolution trees with profiles instead of a distance matrix. Mol. Biol. Evol. 26, 1641–1650 (2009).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. 40

    Stamatakis, A. RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30, 1312–1313 (2014).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  41. 41

    Nguyen, L. T., Schmidt, H. A., von Haeseler, A. & Minh, B. Q. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol. Biol. Evol. 32, 268–274 (2015).

    CAS  Article  Google Scholar 

  42. 42

    Minh, B. Q., Nguyen, M. A. & von Haeseler, A. Ultrafast approximation for phylogenetic bootstrap. Mol. Biol. Evol. 30, 1188–1195 (2013).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  43. 43

    Lartillot, N., Rodrigue, N., Stubbs, D. & Richer, J. PhyloBayes MPI: phylogenetic reconstruction with infinite mixtures of profiles in a parallel environment. Syst. Biol. 62, 611–615 (2013).

    CAS  Article  PubMed  Google Scholar 

  44. 44

    Fu, L., Niu, B., Zhu, Z., Wu, S. & Li, W. CD-HIT: accelerated for clustering the next-generation sequencing data. Bioinformatics 28, 3150–3152 (2012).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  45. 45

    Capella-Gutierrez, S., Silla-Martinez, J. M. & Gabaldon, T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25, 1972–1973 (2009).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  46. 46

    Katoh, K. & Toh, H. Recent developments in the MAFFT multiple sequence alignment program. Br. Bioinform. 9, 286–298 (2008).

    CAS  Article  Google Scholar 

  47. 47

    Noguchi, F., Tanifuji, G., Brown, M. W., Fujikura, K. & Takishita, K. Complex evolution of two types of cardiolipin synthase in the eukaryotic lineage stramenopiles. Mol. Phylogenet. Evol. 101, 133–141 (2016).

    CAS  Article  PubMed  Google Scholar 

  48. 48

    Claros, M. G. MitoProt, a Macintosh application for studying mitochondrial proteins. Comput. Appl. Biosci. 11, 441–447 (1995).

    CAS  PubMed  Google Scholar 

  49. 49

    Emanuelsson, O., Brunak, S., von Heijne, G. & Nielsen, H. Locating proteins in the cell using TargetP, SignalP and related tools. Nat. Protoc. 2, 953–971 (2007).

    CAS  Article  Google Scholar 

  50. 50

    Fukasawa, Y. et al. MitoFates: improved prediction of mitochondrial targeting sequences and their cleavage sites. Mol. Cell. Proteomics 14, 1113–1126 (2015).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

Download references


M.K., M.M.L. and C.W.S. were supported by a grant (MOP-142349) from the Canadian Institutes of Health Research awarded to A.J.R. This work was also supported, in part, by a grant from the JSPS Strategic Young Researcher Overseas Visits Program (awarded to R.K.), by NSERC Grant 298366-2009 to A.G.B.S., by a Czech Science Foundation grant to I.Č. (project GA14-14105S) and by grants from the Japan Society for the Promotion of Science (JSPS; nos 15H05606 and 15K14591 awarded to R.K., 23117005 and 15H05231 awarded to T.H., and 23117006 awarded to Y.I.). We thank A. A. Heiss for his help with Trimastix marina data generation, and N. Ros for her comments on the manuscript.

Author information




M.K., R.K., J.O.A., Y.I., A.G.B.S., T.H. and A.J.R. conceived and designed the experiments; M.K., K.K., I.Č., J.D.S., F.X., A.Y. and Q.Z. performed the experiments; M.M.L., M.K., R.K., C.W.S., K.K., L.E. and Y.I. analysed the data; R.K., C.W.S., J.D.S., K.T., Y.I., A.G.B.S., T.H. and A.J.R. contributed materials and/or analysis tools; and M.M.L., M.K., R.K., A.G.B.S. and A.J.R. wrote the paper.

Corresponding authors

Correspondence to Tetsuo Hashimoto or Andrew J. Roger.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Figures 1–20, Supplementary Table 1 (PDF 1444 kb)

Supplementary Data 1

Predicted mitochondrion-related organelle proteins, and selected predicted cytosolic proteins, in metamonads. (XLSX 149 kb)

Supplementary Data 2

File containing the raw phylogenetic trees depicted in Supplementary Figs 4–20, including bootstrap support values, in Newick format. (TXT 539 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Leger, M., Kolisko, M., Kamikawa, R. et al. Organelles that illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes. Nat Ecol Evol 1, 0092 (2017).

Download citation

Further reading


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing