[en] Liverworts, with approximately 7300 species worldwide, exhibit remarkable morphological diversity in terms of growth form, ontogeny, and architecture. Their mitochondrial genome exhibits lower average substitution rates compared to their nuclear and plastid genomes, and shows less structural variation, suggesting its suitability for inferring relationships at higher taxonomic levels. In this study, we substantially expanded mitochondrial sampling in liverworts by adding complete mitochondrial gene sets from 97 species across 25 families, thereby increasing family-level coverage to 71%. Among these, we newly assembled 23 complete mitochondrial genomes. Although four species with structural variants were newly identified, the overall architecture of liverwort mitochondrial genomes remains highly conserved, with taxa that diverged over 470 million years ago still having collinearity. Phylogenetic inferences from mitochondrial genome sequences confirmed the monophyly of most suprafamilial taxa, with the exceptions of Porellales, Ptilidiales, and Pelliidae. Herzogianthus (Ptilidiales) was well-supported as a sister group to Jungermanniales sensu lato, rather than forming a monophyletic lineage with Ptilidium (Ptilidiales). This work provides an important resource for future genetic and phylogenetic studies of liverworts.
Huang, Dan; School of Life Sciences, Guizhou Normal University, Guiyang, China ; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Zhou, Xuping; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Dong, Shanshan ; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Sheng, Wei; Jinhua Academy of Agricultural Sciences, Jinhua, China
Zuo, Qin; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Zhang, Li; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Ma, Wen-Zhang; CAS Key Laboratory for Plant Biodiversity and Biogeography of East Asia, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming, China
Golinski, G. Karen; UBC Herbarium, The University of British Columbia, Vancouver, Canada
Vanderpoorten, Alain ; Université de Liège - ULiège > Département de Biologie, Ecologie et Evolution > Biologie de l'évolution et de la conservation - Unité aCREA-Ulg (Conseils et Recherches en Ecologie Appliquée)
Goffinet, Bernard; Department of Ecology and Evolutionary Biology, University of Connecticut, Storrs, United States
Liu, Yang; Laboratory of Southern Subtropical Plant Diversity, Fairy Lake Botanical Garden, Shenzhen & Chinese Academy of Sciences, Shenzhen, China
Peng, Tao; School of Life Sciences, Guizhou Normal University, Guiyang, China
Language :
English
Title :
A family-level phylogeny of liverworts (Marchantiophyta): New insights from mitochondrial sequences
The authors acknowledge Dr. Yin\u2010Long Qiu (Michigan University) for providing liverwort DNA. This study was supported by the Scientific Foundation of the Urban Management Bureau of Shenzhen (202005, 202203, 202403 to Yang Liu, and 202106, 202302 to Shanshan Dong).
Bechteler J, Peñaloza-Bojacá G, Bell D, Gordon Burleigh J, McDaniel SF, Christine Davis E, Sessa EB, Bippus A, Christine Cargill D, Chantanoarrapint S, Draper I, Endara L, Forrest LL, Garilleti R, Graham SW, Huttunen S, Lazo JJ, Lara F, Larraín J, Lewis LR, Long DG, Quandt D, Renzaglia K, Schäfer-Verwimp A, Lee GE, Sierra AM, von Konrat M, Zartman CE, Pereira MR, Goffinet B, Villarreal AJC. 2023. Comprehensive phylogenomic time tree of bryophytes reveals deep relationships and uncovers gene incongruences in the last 500 million years of diversification. American Journal of Botany 110: e16249.
Bell D, Lin Q, Gerelle WK, Joya S, Chang Y, Taylor ZN, Rothfels CJ, Larsson A, Villarreal JC, Li FW, Pokorny L, Szövényi P, Crandall-Stotler B, DeGironimo L, Floyd SK, Beerling DJ, Deyholos MK, von Konrat M, Ellis S, Shaw AJ, Chen T, Wong GK, Stevenson DW, Palmer JD, Graham SW. 2020. Organellomic data sets confirm a cryptic consensus on (unrooted) land-plant relationships and provide new insights into bryophyte molecular evolution. American Journal of Botany 107: 91–115.
Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: A flexible trimmer for Illumina sequence data. Bioinformatics 30: 2114–2120.
Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T. 2009. trimAl: A tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25: 1972–1973.
Chen C, Wu Y, Li J, Wang X, Zeng Z, Xu J, Liu Y, Feng J, Chen H, He Y, Xia R. 2023. TBtools-II: A “one for all, all for one” bioinformatics platform for biological big-data mining. Molecular Plant 16: 1733–1742.
Crandall-Stotler B, Stotler R, Long D. 2009. Phylogeny and classification of the Marchantiophyta. Edinburgh Journal of Botany 66: 155–198.
Crandall-Stotler B, Stotler RE, Long DG. 2008. Morphology and classification of the Marchantiophyta. In: Shaw AJ, Goffinet B eds. Bryophyte Biology. Cambridge: Cambridge University Press. 1–54.
Danecek P, Bonfield JK, Liddle J, Marshall J, Ohan V, Pollard MO, Whitwham A, Keane T, McCarthy SA, Davies RM, Li H. 2021. Twelve years of SAMtools and BCFtools. Gigascience 10: giab008.
Dierckxsens N, Mardulyn P, Smits G. 2017. NOVOPlasty: de novo assembly of organelle genomes from whole genome data. Nucleic Acids Research 45: e18.
Dong S, Liu Y. 2021. The mitochondrial genomes of bryophytes. Bryophyte Diversity and Evolution 43: 112–126.
Dong S, Wang S, Li L, Yu J, Zhang Y, Xue JY, Chen H, Ma J, Zeng Y, Cai Y, Huang W, Zhou X, Wu J, Li J, Yao Y, Hu R, Zhao T, Villarreal AJC, Dirick L, Liu L, Ignatov M, Jin M, Ruan J, He Y, Wang H, Xu B, Rozzi R, Wegrzyn J, Stevenson DW, Renzaglia KS, Chen H, Zhang L, Zhang S, Mackenzie R, Moreno JE, Melkonian M, Wei T, Gu Y, Xu X, Rensing SA, Huang J, Long M, Goffinet B, Bowman JL, Van de Peer Y, Liu H, Liu Y. 2025. Bryophytes hold a larger gene family space than vascular plants. Nature Genetics 57: 2562–2569.
Dong S, Yu J, Zhang L, Goffinet B, Liu Y. 2022. Phylotranscriptomics of liverworts: revisiting the backbone phylogeny and ancestral gene duplications. Annals of Botany 130: 951–964.
Dong S, Zhang S, Zhang L, Wu H, Goffinet B, Liu Y. 2021. Plastid genomes and phylogenomics of liverworts (Marchantiophyta): Conserved genome structure but highest relative plastid substitution rate in land plants. Molecular Phylogenetics and Evolution 161: 107171.
Dong S, Zhao C, Zhang S, Zhang L, Wu H, Liu H, Zhu R, Jia Y, Goffinet B, Liu Y. 2019. Mitochondrial genomes of the early land plant lineage liverworts (Marchantiophyta): Conserved genome structure, and ongoing low frequency recombination. BMC Genomics 20: 953.
Drouin G, Daoud H, Xia J. 2008. Relative rates of synonymous substitutions in the mitochondrial, chloroplast and nuclear genomes of seed plants. Molecular Phylogenetics and Evolution 49: 827–831.
Edgar RC. 2022. Muscle5: High-accuracy alignment ensembles enable unbiased assessments of sequence homology and phylogeny. Nature Communications 13: 6968.
Flores JR, Bippus AC, Suárez GM, Hyvönen J. 2021. Defying death: Incorporating fossils into the phylogeny of the complex thalloid liverworts (Marchantiidae, Marchantiophyta) confirms high order clades but reveals discrepancies in family-level relationships. Cladistics 37: 231–247.
Flores JR, Catalano SA, Muñoz J, Suárez GM. 2018. Combined phylogenetic analysis of the subclass Marchantiidae (Marchantiophyta): Towards a robustly diagnosed classification. Cladistics 34: 517–541.
Forrest L, Davis C, Long D, Crandall-Stotler B, Clark A, Hart ML. 2006. Unraveling the evolutionary history of the liverworts (Marchantiophyta): Multiple taxa, genomes and analyses. The Bryologist 109: 303–334.
Harris BJ, Clark JW, Schrempf D, Szöllősi GJ, Donoghue PCJ, Hetherington AM, Williams TA. 2022. Divergent evolutionary trajectories of bryophytes and tracheophytes from a complex common ancestor of land plants. Nature Ecology and Evolution 6: 1634–1643.
Heinrichs J, Gradstein SR, Wilson R, Schneider H. 2005. Towards a natural classification of liverworts (Marchantiopyta) based on the chloroplast gene rbcL. Cryptogamie, Bryologie 26: 131–150.
Hendry TA, Wang B, Yang Y, Davis EC, Braggins JE, Schuster RM, Qiu Y-L. 2007. Evaluating phylogenetic positions of four liverworts from New Zealand, Neogrollea notabilis, Jackiella curvata, Goebelobryum unguiculatum and Herzogianthus vaginatus, using three chloroplast genes. The Bryologist 110: 738–751.
He-Nygrén X, Juslén A, Ahonen I, Glenny D, Piippo S. 2006. Illuminating the evolutionary history of liverworts (Marchantiophyta)—towards a natural classification. Cladistics 22: 1–31.
Hu SY, Shi G, Yang CA, Van de Peer Y, Li Z, Xue JY. 2025. Comprehensive sampling from mitochondrial genomes substantiates the Neoproterozoic origin of land plants. Plant Communications 6: 101497.
Johnson MG, Gardner EM, Liu Y, Medina R, Goffinet B, Shaw AJ, Zerega NJ, Wickett NJ. 2016. HybPiper: Extracting coding sequence and introns for phylogenetics from high-throughput sequencing reads using target enrichment. Applications in Plant Sciences 4: 1600016.
Kück P, Struck TH. 2014. BaCoCa–a heuristic software tool for the parallel assessment of sequence biases in hundreds of gene and taxon partitions. Molecular Phylogenetics and Evolution 70: 94–98.
Kwon W, Yongsung K, Park J. 2019. The complete mitochondrial genome of Dumortiera hirsuta (Sw.) Nees (Dumortieraceae, Marchantiophyta). Mitochondrial DNA Part B 4: 1586–1587.
Laenen B, Shaw B, Schneider H, Goffinet B, Paradis E, Désamoré A, Heinrichs J, Villarreal JC, Gradstein SR, McDaniel SF, Long DG, Forrest LL, Hollingsworth ML, Crandall-Stotler B, Davis EC, Engel J, Von Konrat M, Cooper ED, Patiño J, Cox CJ, Vanderpoorten A, Shaw AJ. 2014. Extant diversity of bryophytes emerged from successive post-Mesozoic diversification bursts. Nature Communications 5: 5134.
Lartillot N, Rodrigue N, Stubbs D, Richer J. 2013. PhyloBayes MPI: Phylogenetic reconstruction with infinite mixtures of profiles in a parallel environment. Systematic Biology 62: 611–615.
Li H, Durbin R. 2009. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25: 1754–1760.
Li YF, Luo L, Liu Y, He Q, Yu NN, Gaowa N, Yi ZQ, Wang JJ, Han W, Peng T, Ho BC, He X, Zhang L, Chen ZD, Jia Y, Wang QH. 2024. The bryophyte phylogeny group: A revised familial classification system based on plastid phylogenomic data. Journal of Systematics and Evolution 62: 577–588.
Liu Y, Cox CJ, Wang W, Goffinet B. 2014a. Mitochondrial phylogenomics of early land plants: Mitigating the effects of saturation, compositional heterogeneity, and codon-usage bias. Systematic Biology 63: 862–878.
Liu Y, Johnson MG, Cox CJ, Medina R, Devos N, Vanderpoorten A, Hedenäs L, Bell NE, Shevock JR, Aguero B, Quandt D, Wickett NJ, Shaw AJ, Goffinet B. 2019. Resolution of the ordinal phylogeny of mosses using targeted exons from organellar and nuclear genomes. Nature Communications 10: 1485.
Liu Y, Medina R, Goffinet B. 2014b. 350 my of mitochondrial genome stasis in mosses, an early land plant lineage. Molecular Biology and Evolution 31: 2586–2591.
Lanfear R, Frandsen PB, Wright AM, Senfeld T, Calcott B. 2017. PartitionFinder 2: New methods for selecting partitioned models of evolution for molecular and morphological phylogenetic analyses. Molecular Biology and Evolution 34: 772–773.
McCauley DE. 2013. Paternal leakage, heteroplasmy, and the evolution of plant mitochondrial genomes. New Phytologist 200: 966–977.
Myszczyński K, Górski P, Ślipiko M, Sawicki J. 2018. Sequencing of organellar genomes of Gymnomitrion concinnatum (Jungermanniales) revealed the first exception in the structure and gene order of evolutionary stable liverworts mitogenomes. BMC Plant Biology 18: 321.
Porebski S, Bailey LG, Baum BR. 1997. Modification of a CTAB DNA extraction protocol for plants containing high polysaccharide and polyphenol components. Plant Molecular Biology Reporter 15: 8–15.
Renner MAM, Salter J. 2020. Unique shoot architecture in the leafy liverwort Herzogianthus vaginatus (Herzogianthaceae): Insights into novel growth patterns in Jungermanniidae. Botanical Journal of the Linnean Society 193: 207–227.
Rydin C, Wikström N, Bremer B. 2017. Conflicting results from mitochondrial genomic data challenge current views of Rubiaceae phylogeny. American Journal of Botany 104: 1522–1532.
Schuster RM. 1960. Studies on Hepaticae. II. The new family Chaetophylliopsidaceae. The Journal of the Hattori Botanical Laboratory 23: 68–76.
Schuster RM. 1972. Phylogenetic and taxonomic studies on Jungermanniidae. The Journal of the Hattori Botanical Laboratory 36: 321–405.
Schuster RM. 1984. Evolution, phylogeny and classification of the Hepaticae. Nichinan: Hattori Botanical Laboratory.
Shen C, Li H, Shu L, Huang WZ, Zhu RL. 2025. Ancient large-scale gene duplications and diversification in bryophytes illuminate the plant terrestrialization. New Phytologist 245: 2292–2308.
Siu-Ting K, Torres-Sánchez M, San Mauro D, Wilcockson D, Wilkinson M, Pisani D, O′Connell MJ, Creevey CJ. 2019. Inadvertent paralog inclusion drives artifactual topologies and timetree estimates in pylogenomics. Molecular Biology and Evolution 36: 1344–1356.
Söderström L, Hagborg A, von Konrat M, Bartholomew-Began S, Bell D, Briscoe L, Brown E, Cargill DC, Costa DP, Crandall-Stotler BJ, Cooper ED, Dauphin G, Engel JJ, Feldberg K, Glenny D, Gradstein SR, He X, Heinrichs J, Hentschel J, Ilkiu-Borges AL, Katagiri T, Konstantinova NA, Larraín J, Long DG, Nebel M, Pócs T, Puche F, Reiner-Drehwald E, Renner MA, Sass-Gyarmati A, Schäfer-Verwimp A, Moragues JG, Stotler RE, Sukkharak P, Thiers BM, Uribe J, Váňa J, Villarreal JC, Wigginton M, Zhang L, Zhu RL. 2016. World checklist of hornworts and liverworts. PhytoKeys 59: 1–828.
Soltis DE, Soltis PS. 2019. Nuclear genomes of two magnoliids. Nature Plants 5: 6–7.
Soto Gomez M, Lin Q, da Silva Leal E, Gallaher TJ, Scherberich D, Mennes CB, Smith SY, Graham SW. 2020. A bi-organellar phylogenomic study of Pandanales: Inference of higher-order relationships and unusual rate-variation patterns. Cladistics 36: 481–504.
Stamatakis A. 2014. RAxML version 8: A tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30: 1312–1313.
Su D, Yang L, Shi X, Ma X, Zhou X, Hedges SB, Zhong B. 2021. Large-scale phylogenomic analyses reveal the monophyly of bryophytes and Neoproterozoic origin of land plants. Molecular Biology and Evolution 38: 3332–3344.
Sun M, Soltis DE, Soltis PS, Zhu X, Burleigh JG, Chen Z. 2015. Deep phylogenetic incongruence in the angiosperm clade Rosidae. Molecular Phylogenetics and Evolution 83: 156–166.
Villarreal AJC, Crandall-Stotler BJ, Hart ML, Long DG, Forrest LL. 2016. Divergence times and the evolution of morphological complexity in an early land plant lineage (Marchantiopsida) with a slow molecular rate. New Phytologist 209: 1734–1746.
Wolfe KH, Li WH, Sharp PM. 1987. Rates of nucleotide substitution vary greatly among plant mitochondrial, chloroplast, and nuclear DNAs. Proceedings of the National Academy of Sciences of the United States of America 84: 9054–9058.
Xia X. 2013. DAMBE5: A comprehensive software package for data analysis in molecular biology and evolution. Molecular Biology and Evolution 30: 1720–1728.
Xiang YL, Jin XJ, Shen C, Cheng XF, Shu L, Zhu RL. 2022. New insights into the phylogeny of the complex thalloid liverworts (Marchantiopsida) based on chloroplast genomes. Cladistics 38: 649–662.
Xue TT, Janssens SB, Liu BB, Yu SX. 2024. Phylogenomic conflict analyses of the plastid and mitochondrial genomes via deep genome skimming highlight their independent evolutionary histories: a case study in the cinquefoil genus Potentilla sensu lato (Potentilleae, Rosaceae). Molecular Phylogenetics and Evolution 190: 107956.
Yu Y, Yang JB, Ma WZ, Pressel S, Liu HM, Wu YH, Schneider H. 2020. Chloroplast phylogenomics of liverworts: A reappraisal of the backbone phylogeny of liverworts with emphasis on Ptilidiales. Cladistics 36: 184–193.
Zhou W, Soghigian J, Xiang QJ. 2022. A new pipeline for removing paralogs in target enrichment data. Systematic Biology 71: 410–425.
