The digenean complex life cycle: phylostratigraphy analysis of the molecular signatures

  • Maksim Nesterenko Department of Invertebrate Zoology, Faculty of Biology, Saint Petersburg State University, Universitetskaya nab., 7–9, Saint Petersburg, 199034, Russian Federation
  • Sergei Shchenkov Department of Invertebrate Zoology, Faculty of Biology, Saint Petersburg State University, Universitetskaya nab., 7–9, Saint Petersburg, 199034, Russian Federation
  • Sofia Denisova Department of Invertebrate Zoology, Faculty of Biology, Saint Petersburg State University, Universitetskaya nab., 7–9, Saint Petersburg, 199034, Russian Federation
  • Viktor Starunov Department of Invertebrate Zoology, Faculty of Biology, Saint Petersburg State University, Universitetskaya nab., 7–9, Saint Petersburg, 199034, Russian Federation; Zoological Institute, Russian Academy of Sciences, Universitetskaya nab., 1, 199034, Saint Petersburg, Russian Federation


The parasitic flatworms from Digenea group have been the object of numerous in-depth studies for several centuries. The question of the evolutionary origin and transformation of the digenean complex life cycle remains relevant and open due to the biodiversity of these parasites and the absence of fossil records. However, modern technologies and analysis methods allow to get closer to understanding the molecular basis of both the realization of the cycle and its complication. In the present study, we have applied phylostratigraphy and evolutionary transcriptomics approaches to the available digenean genomic and transcriptomic data and built ancestral genomes models. The comparison results of Platyhelminthes and Digenea ancestor genome models made it possible to identify which genes were gained and duplicated in the possible genome of digenean ancestor. Based on the bioprocesses enrichment analysis results, we assumed that the change in the regulation of many processes, including embryogenesis, served as a basis for the complication of the ancestor life cycle. The evolutionary transcriptomics results obtained revealed the “youngest” and “oldest” life cycle stages of Fasciola gigantica, F. hepatica, Psilotrema simillimum, Schistosoma mansoni, Trichobilharzia regenti, and T. szidati. Our results can serve as a basis for a more in-depth study of the molecular signatures of life cycle stages and the evolution transformation of individual organ systems and stage-specific traits.


flatworms, Digenea, complex life cycle, molecular signature, phylostratigraphy, evolutionary transcriptomics


Download data is not yet available.


Abu-Jamous, B. and Kelly, S. 2018. Clust: automatic extraction of optimal co-expressed gene clusters from gene expression data. Genome Biology 19(1):1–11.

Almagro Armenteros, J. J., Tsirigos, K. D., Sønderby, C. K., Petersen, T. N., Winther, O., Brunak, S., von Heijne, G., and Nielsen, H. 2019. SignalP 5.0 improves signal peptide predictions using deep neural networks. Nature Biotechnology 37(4):420–423.

Almudi, I., Vizueta, J., Wyatt, C. D. R., de Mendoza, A., Marlétaz, F., Firbas, P. N., Feuda, R., Masiero, G., Medina, P., Alcaina-Caro, A., Cruz, F., Gómez-Garrido, J., Gut, M., Alioto, T. S., Vargas-Chavez, C., Davie, K., Misof, B., González, J., Aerts, S., Lister, R., Paps, J., Rozas, J., SánchezGracia, A., Irimia, M., Maeso, I., and Casares, F. 2020. Genomic adaptations to aquatic and aerial life in mayflies and the origin of insect wings. Nature Communications 11(1):1–11.

Altenhoff, A. M., Levy, J., Zarowiecki, M., Tomiczek, B., Vesztrocy, A. W., Dalquen, D. A., Müller, S., Telford, M. J., Glover, N. M., Dylus, D., and Dessimoz, C. 2019. OMA standalone: Orthology inference among public and custom genomes and transcriptomes. Genome Research 29(7):1152–1163.

Arendsee, Z., Li, J., Singh, U., Seetharam, A., Dorman, K., and Wurtele, E. S. 2019. Phylostratr: A framework for phylostratigraphy. Bioinformatics 35(19):3617–3627.

Bendtsen, J. D., Jensen, L. J., Blom, N., Von Heijne, G., and Brunak, S. 2004. Feature-based prediction of nonclassical and leaderless protein secretion. Protein Engineering, Design and Selection 17(4):349–356.

Brandl, H., Moon, H. K., Vila-Farré, M., Liu, S. Y., Henry, I., and Rink, J. C. 2016. PlanMine — A mineable resource of planarian biology and biodiversity. Nucleic Acids Research 44(D1):D764–D773.

Camacho, C., Coulouris, G., Avagyan, V., Ma, N., Papadopoulos, J., Bealer, K., and Madden, T. L. 2009. BLAST+: Architecture and applications. BMC Bioinformatics 10(1):1–9.

Cantacessi, C., Mulvenna, J., Young, N. D., Kasny, M., Horak, P., Aziz, A., Hofmann, A., Loukas, A., and Gasser, R. B. 2012. A deep exploration of the transcriptome and “ excretory /secretory ” proteome of adult Fascioloides magna. Molecular & Cellular Proteomics 11(11):1340–1353.

Cantalapiedra, C. P., Hernández-Plaza, A., Letunic, I., Bork, P., and Huerta-Cepas, J. 2021. eggNOG-mapper v2: functional annotation, orthology assignments, and domain prediction at the metagenomic scale. Molecular Biology and Evolution msab293.

Capella-Gutiérrez, S., Silla-Martínez, J. M., and Gabaldón, T. 2009. trimAl: A tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25(15):1972–1973.

Chen, S., Zhou, Y., Chen, Y., and Gu, J. 2018. fastp: an ultrafast all-in-one FASTQ preprocessor. Bioinformatics 34(17):i884–i890.

Choudhary, V., Garg, S., Chourasia, R., Hasnani, J. J., Patel, P. V, Shah, T. M., Bhatt, V. D., Mohapatra, A., Blake, D. P., and Joshi, C. G. 2015. Transcriptome analysis of the adult rumen fluke Paramphistomum cervi following next generation sequencing. Gene 570(1):64–70.

Cwiklinski, K., Dalton, J. P., Dufresne, P. J., Course, J. L., Williams, D. J. L., Hodgkinson, J., and Paterson, S. 2015. The Fasciola hepatica genome: gene duplication and polymorphism reveals adaptation to the host environment and the capacity for rapid evolution. Genome Biology 16(1):1–13.

Darriba, D., Taboada, G. L., Doallo, R., and Posada, D. 2011. ProtTest 3: Fast selection of best-fit models of protein evolution. Bioinformatics 27(8):1164–1165.

Domazet-Lošo, T., Brajković, J., and Tautz, D. 2007. A phylostratigraphy approach to uncover the genomic history of major adaptations in metazoan lineages. Trends in Genetics 23(11):531–533. tig.2007.07.007

Domazet-Lošo, T. and Tautz, D. 2010. A phylogenetically based transcriptome age index mirrors ontogenetic divergence patterns. Nature 468(7325):815–819.

Drost, H.-G., Gabel, A., Liu, J., Quint, M., and Grosse, I. 2018. MyTAI: Evolutionary transcriptomics with R. Bioinformatics 34(9):1589–1590.

Dylus, D., Nevers, Y., Altenhoff, A. M., Gürtler, A., Dessimoz, C., and Glover, N. M. 2020. How to build phylogenetic species trees with OMA. F1000Research 9:511.

Emanuelsson, O., Nielsen, H., Brunak, S., and Von Heijne, G. 2000. Predicting subcellular localization of proteins based on their N-terminal amino acid sequence. Journal of Molecular Biology 300(4):1005–1016.

Fernández, R. and Gabaldón, T. 2020. Gene gain and loss across the metazoan tree of life. Nature Ecology and Evolution 4(4):524–533.

Galaktionov, K. V. and Dobrovolskij, A. A. 2003. The biology and evolution of trematodes. Kluwer Academic Publ., St Petersbug.

Garg, G. and Ranganathan, S. 2011. In silico secretome analysis approach for next generation sequencing transcriptomic data. BMC Genomics 12(3):1–10.

Gibson, D. I. 1987. Questions in digenean systematics and evolution. Parasitology 95(2):429–460.

Guindon, S. and Gascuel, O. 2003. A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Systematic Biology 52(5):696–704.

Howe, K. L., Bolt, B. J., Shafie, M., Kersey, P., and Berriman, M. 2017. WormBase ParaSite — a comprehensive resource for helminth genomics. Molecular and Biochemical Parasitology 215:2–10.

Huerta-Cepas, J., Szklarczyk, D., Heller, D., Hernández-Plaza, A., Forslund, S. K., Cook, H., Mende, D. R., Letunic, I., Rattei, T., Jensen, L. J., Von Mering, C., and Bork, P. 2019. eggNOG 5.0: A hierarchical, functionally and phylogenetically annotated orthology resource based on 5090 organisms and 2502 viruses. Nucleic Acids Research 47(D1):D309–D314.

Katoh, K. and Standley, D. M. 2013. MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. Molecular Biology and Evolution 30(4):772–780.

Krogh, A., Larsson, B., Von Heijne, G., and Sonnhammer, E. L. L. 2001. Predicting transmembrane protein topology with a hidden markov model: application to complete genomes. Journal of Molecular Biology 305(3):567–580.

Leontovyč, R., Young, N. D., Korhonen, P. K., Hall, R. S., Bulantová, J., Jeřábková, V., Kašný, M., Gasser, R. B., and Horák, P. 2019. Molecular evidence for distinct modes of nutrient acquisition between visceral and neurotropic schistosomes of birds. Scientific Reports 9(1):1–12.

Leontovyč, R., Young, N. D., Korhonen, P. K., Hall, R. S., Tan, P., Mikeš, L., Kašný, M., Horák, P., and Gasser, R. B. 2016. Comparative transcriptomic exploration reveals unique molecular adaptations of neuropathogenic trichobilharzia to invade and parasitize its avian definitive host. PLoS Neglected Tropical Diseases 10(2):e0004406.

Li, B., McNulty, S. N., Rosa, B. A., Tyagi, R., Zeng, Q. R., Gu, K., Weil, G. J., and Mitreva, M. 2016. Conservation and diversification of the transcriptomes of adult Paragonimus westermani and P. skrjabini. Parasites & Vectors 9(1):497.

Littlewood, D. T. J. 2006. Parasitic flatworms: molecular biology, biochemistry, immunology and physiology. Cabi Publishing-C a B Int. 480 pp.

Liu, F., Li, Y., Yu, H., Zhang, L., Hu, J., Bao, Z., and Wang, S. 2021. MolluscDB: An integrated functional and evolutionary genomics database for the hyper-diverse animal phylum Mollusca. Nucleic Acids Research 49(D1):D988–D997.

Martín-Durán, J. M., Ryan, J. F., Vellutini, B. C., Pang, K., and Hejnol, A. 2017. Increased taxon sampling reveals thousands of hidden orthologs in flatworms. Genome Research 27(7):1263–1272.

McNulty, S. N., Tort, J. F., Rinaldi, G., Fischer, K., Rosa, B. A., Smircich, P., Fontenla, S., Choi, Y. J., Tyagi, R., Hallsworth-Pepin, K., Mann, V. H., Kammili, L., Latham, P. S., Dell’Oca, N., Dominguez, F., Carmona, C., Fischer, P. U., Brindley, P. J., and Mitreva, M. 2017. Genomes of Fasciola hepatica from the Americas reveal colonization with Neorickettsia endobacteria related to the agents of potomac horse and human sennetsu fevers. PLoS Genetics 13(1):e1006537. pgen.1006537

Minelli, A. and Fusco, G. 2010. Developmental plasticity and the evolution of animal complex life cycles. Philosophical Transactions of the Royal Society B 365(1540):631–640.

Minh, B. Q., Schmidt, H. A., Chernomor, O., Schrempf, D., Woodhams, M. D., Von Haeseler, A., and Lanfear, R. 2020. IQ-TREE 2: New models and efficient methods for phylogenetic inference in the genomic era. Molecular Biology and Evolution 37(5):1530–1534.

Mirdita, M., Steinegger, M., and Söding, J. 2019. MMseqs2 desktop and local web server app for fast, interactive sequence searches. Bioinformatics 35(16):2856–2858.

Nesterenko, M., Starunov, V., Shchenkov, S., Maslova, A., Denisova, S., Granovich, A., Dobrovolskij, A., and Khalturin, K. 2020. Molecular signatures of the rediae, cercariae and adult worm stages in the complex life cycles of parasitic flatworms (Psilostomatidae, Trematoda). Parasites & Vectors 13(1):1–21.

Nguyen, L. T., Schmidt, H. A., Von Haeseler, A., and Minh, B. Q. 2015. IQ-TREE: A fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Molecular Biology and Evolution 32(1):268–274.

Odening, K. 1974. Verwandtschaft, System und zyklo-ontogenetische Besonderheiten der Trematoden. Zoologischer Jahrbucher. Systematik 101(3):345–396.

Olson, P. D., Cribb, T. H., Tkach, V. V, Bray, R. A., and Littlewood, D. T. J. 2003. Phylogeny and classification of the Digenea (Platyhelminthes: Trematoda). International Journal for Parasitology 33(7):733–755.

Paradis, E. and Schliep, K. 2019. ape 5.0: an environment for modern phylogenetics and evolutionary analyses in R. Bioinformatics 35(3):526–528.

Patro, R., Duggal, G., Love, M. I., Irizarry, R. A., and Kingsford, C. 2017. Salmon provides fast and bias-aware quantification of transcript expression. Nature Methods 14(4):417–419.

Pérez-Ponce de León, G. and Hernández-Mena, D. I. 2019. Testing the higher-level phylogenetic classification of Digenea (Platyhelminthes, Trematoda) based on nuclear rDNA sequences before entering the age of the “next-generation” Tree of Life. Journal of Helminthology 93(3):260–276.

Picard, M. A. L., Boissier, J., Roquis, D., Grunau, C., Allienne, J.-F., Duval, D., Toulza, E., Arancibia, N., Caffrey, C., Long, T., Nidelet, S., Rohmer, M., and Cosseau, C. 2016. Sex-biased transcriptome of Schistosoma mansoni: hostparasite interaction, genetic determinants and epigenetic regulators are associated with sexual differentiation. PLoS Neglected Tropical Diseases 10(9):e0004930.

Protasio, A. V., Tsai, I. J., Babbage, A., Nichol, S., Hunt, M., Aslett, M. A., de Silva, N., Velarde, G. S., Anderson, T. J. C., Clark, R. C., Davidson, C., Dillon, G. P., Holroyd, N. E., LoVerde, P. T., Lloyd, C., McQuillan, J., Oliveira, G., Otto, T. D., Parker-Manuel, S. J., Quail, M. A., Wilson, R. A., Zerlotini, A., Dunne, D. W., and Berriman, M. 2012. A systematically improved high quality genome and transcriptome of the human blood fluke Schistosoma mansoni. PLoS Neglected Tropical Diseases 6(1):e1455.

Reinhard, E. G. 1957. Landmarks of parasitology I. The discovery of the life cycle of the liver fluke. Experimental Parasitology 6(2):208–232.

Shi, L., Derouiche, A., Pandit, S., Rahimi, S., Kalantari, A., Futo, M., Ravikumar, V., Jers, C., Mokkapati, V. R. S. S., Vlahovicek, K., and Mijakovic, I. 2020. Evolutionary analysis of the Bacillus subtilis genome reveals new genes involved in sporulation. Molecular Biology and Evolution 37(6):1667–1678.

Train, C. M., Pignatelli, M., Altenhoff, A., and Dessimoz, C. 2019. IHam and pyHam: Visualizing and processing hierarchical orthologous groups. Bioinformatics 35(14):2504– 2506.

Wagner, G. P., Kin, K., and Lynch, V. J. 2013. A model based criterion for gene expression calls using RNA-seq data. Theory in Biosciences 132(3):159–164.

Wang, B., Lee, J., Li, P., Saberi, A., Yang, H., Liu, C., Zhao, M., and Newmark, P. A. 2018. Stem cell heterogeneity drives the parasitic life cycle of Schistosoma mansoni. eLife 7:e35449.

Wang, J., Zhang, L., Lian, S., Qin, Z., Zhu, X., Dai, X., Huang, Z., Ke, C., Zhou, Z., Wei, J., Liu, P., Hu, N., Zeng, Q., Dong, B., Dong, Y., Kong, D., Zhang, Z., Liu, S., Xia, Y., Li, Y., Zhao, L., Xing, Q., Huang, X., Hu, X., Bao, Z., and Wang, S. 2020. Evolutionary transcriptomics of metazoan biphasic life cycle supports a single intercalation origin of metazoan larvae. Nature Ecology and Evolution 4(5):725–736.

Wood, D. E., Lu, J., and Langmead, B. 2019. Improved metagenomic analysis with Kraken 2. Genome Biology 20(1):1–13.

Wood, D. E. and Salzberg, S. L. 2014. Kraken: Ultrafast metagenomic sequence classification using exact alignments. Genome Biology 15(3):1–12.

Yates, A. D., Achuthan, P., Akanni, W., Allen, J., Allen, J., Alvarez-Jarreta, J., Amode, M. R., Armean, I. M., Azov, A. G., Bennett, R., Bhai, J., Billis, K., Boddu, S., Marugán, J. C., Cummins, C., Davidson, C., Dodiya, K., Fatima, R., Gall, A., Giron, C. G., Gil, L., Grego, T., Haggerty, L., Haskell, E., Hourlier, T., Izuogu, O. G., Janacek, S. H., Juettemann, T., Kay, M., Lavidas, I., Le, T., Lemos, D., Martinez, J. G., Maurel, T., McDowall, M., McMahon, A., Mohanan, S., Moore, B., Nuhn, M., Oheh, D. N., Parker, A., Parton, A., Patricio, M., Sakthivel, M. P., Abdul Salam, A. I., Schmitt, B. M., Schuilenburg, H., Sheppard, D., Sycheva, M., Szuba, M., Taylor, K., Thormann, A., Threadgold, G., Vullo, A., Walts, B., Winterbottom, A., Zadissa, A., Chakiachvili, M., Flint, B., Frankish, A., Hunt, S. E., Iisley, G., Kostadima, M., Langridge, N., Loveland, J. E., Martin, F. J., Morales, J., Mudge, J. M., Muffato, M., Perry, E., Ruffier, M., Trevanion, S. J., Cunningham, F., Howe, K. L., Zerbino, D. R., and Flicek, P. 2020. Ensembl 2020. Nucleic Acids Research 48(D1):D682–D688.

Young, N. D., Jex, A. R., Cantacessi, C., Hall, R. S., Campbell, B. E., Spithill, T. W., Tangkawattana, S., Tangkawattana, P., Laha, T., and Gasser, R. B. 2011. A portrait of the transcriptome of the neglected trematode, Fasciola gigantica — biological and biotechnological implications. PLoS Neglected Tropical Diseases 5(2):e1004.

Zajac, N., Zoller, S., Seppälä, K., Moi, D., Dessimoz, C., Jokela, J., Hartikainen, H., and Glover, N. 2021. Gene duplication and gain in the trematode Atriophallophorus winterbourni contributes to adaptation to parasitism. Genome Biology and Evolution 13(3):evab010.

Zakhvatkin, A. A. 1949. The comparative embryology of the low invertebrates. Sources and method of the origin of metazoan development. Moscow, Soviet Science. (In Russian)

Zambelli, F., Mastropasqua, F., Picardi, E., D’Erchia, A. M., Pesole, G., and Pavesi, G. 2018. RNentropy: An entropybased tool for the detection of significant variation of gene expression across multiple RNA-Seq experiments. Nucleic Acids Research 46(8):e46.

Zhang, X.-X., Cwiklinski, K., Hu, R.-S., Zheng, W.-B., Sheng, Z.-A., Zhang, F.-K., Elsheikha, H. M., Dalton, J. P., and Zhu, X.-Q. 2019. Complex and dynamic transcriptional changes allow the helminth Fasciola gigantica to adjust to its intermediate snail and definitive mammalian hosts. BMC Genomics 20(1):1–18.

Zhang, Y., Parmigiani, G., and Johnson, W. E. 2020. ComBatseq: batch effect adjustment for RNA-seq count data. NAR Genomics and Bioinformatics 2(3):lqaa078.

How to Cite
Nesterenko, M., Shchenkov, S., Denisova, S., & Starunov, V. (2022). The digenean complex life cycle: phylostratigraphy analysis of the molecular signatures. Biological Communications, 67(2), 65–87.
Full communications