Register      Login
Australian Systematic Botany Australian Systematic Botany Society
Taxonomy, biogeography and evolution of plants
RESEARCH ARTICLE

Genome skimming provides well resolved plastid and nuclear phylogenies, showing patterns of deep reticulate evolution in the tropical carnivorous plant genus Nepenthes (Caryophyllales)

Lars Nauheimer https://orcid.org/0000-0002-2847-0966 A B C G , Lujing Cui D E , Charles Clarke A , Darren M. Crayn https://orcid.org/0000-0001-6614-4216 A B C D , Greg Bourke F and Katharina Nargar https://orcid.org/0000-0002-0459-5991 A B C D
+ Author Affiliations
- Author Affiliations

A Australian Tropical Herbarium, James Cook University, PO Box 6811, Cairns, Qld 4878, Australia.

B Centre for Tropical Environmental Sustainability Science, James Cook University, McGregor Road, Smithfield, Qld 4878, Australia.

C Centre for Tropical Bioinformatics and Molecular Biology, James Cook University, McGregor Road, Smithfield, Qld 4878, Australia.

D National Research Collections Australia, Commonwealth Industrial and Scientific Research Organisation (CSIRO), GPO Box 1700, Canberra, ACT 2601, Australia.

E School of Computer Science and Engineering, University of New South Wales, NSW 2052, Australia.

F Blue Mountains Botanic Garden, Bells Line of Road, Mount Tomah, NSW 2758, Australia.

G Corresponding author. Email: lars.nauheimer@jcu.edu.au

Australian Systematic Botany 32(3) 243-254 https://doi.org/10.1071/SB18057
Submitted: 11 September 2018  Accepted: 2 May 2019   Published: 12 June 2019

Abstract

Nepenthes is a genus of carnivorous plants consisting of ~160 species that are distributed in the paleotropics. Molecular systematics has so far not been able to resolve evolutionary relationships of most species because of the limited genetic divergence in previous studies. In the present study, we used a genome-skimming approach to infer phylogenetic relationships on the basis of 81 plastid genes and the highly repetitive rRNA (external transcribed spacer (ETS)–26S) for 39 accessions representing 34 species from eight sections. Maximum-likelihood analysis and Bayesian inference were performed separately for the nuclear and the plastid datasets. Divergence-time estimations were conducted on the basis of a relaxed molecular-clock model, using secondary calibration points. The phylogenetic analyses of the nuclear and plastid datasets yielded well resolved and supported phylogenies. Incongruences between the two datasets were detected, suggesting multiple hybridisation events or incomplete lineage sorting in the deeper and more recent evolutionary history of the genus. The inclusion of several known and suspected hybrids in the phylogenetic analysis provided insights into their parentage. Divergence-time estimations placed the crown diversification of Nepenthes in the early Miocene, c. 20 million years ago. This study showed that genome skimming provides well resolved nuclear and plastid phylogenies that provide valuable insights into the complex evolutionary relationships of Nepenthes.

Additional keywords: divergence time estimation, infrageneric classification, molecular systematics, Nepenthaceae.


References

Akaike H (1974) A new look at the statistical model identification. IEEE Transactions on Automatic Control 19, 716–723.
A new look at the statistical model identification.Crossref | GoogleScholarGoogle Scholar |

Alamsyah F, Ito M (2013) Phylogenetic analysis of Nepenthaceae, based on internal transcribed spacer nuclear ribosomal DNA sequences. Acta Phytotaxonomica et Geobotanica 64, 113–126.
Phylogenetic analysis of Nepenthaceae, based on internal transcribed spacer nuclear ribosomal DNA sequences.Crossref | GoogleScholarGoogle Scholar |

Álvarez I, Wendel JF (2003) Ribosomal ITS sequences and plant phylogenetic inference. Molecular Phylogenetics and Evolution 29, 417–434.
Ribosomal ITS sequences and plant phylogenetic inference.Crossref | GoogleScholarGoogle Scholar | 14615184PubMed |

Baldwin BG, Markos S (1998) Phylogenetic utility of the external transcribed spacer (ETS) of 18S–26S rDNA: congruence of ETS and ITS trees of Calycadenia (Compositae). Molecular Phylogenetics and Evolution 10, 449–463.
Phylogenetic utility of the external transcribed spacer (ETS) of 18S–26S rDNA: congruence of ETS and ITS trees of Calycadenia (Compositae).Crossref | GoogleScholarGoogle Scholar | 10051397PubMed |

Biswal DK, Debnath M, Konhar R, Yanthan S, Tandon P (2018) Phylogeny and biogeography of carnivorous plant family Nepenthaceae with reference to the Indian pitcher plant Nepenthes khasiana reveals an Indian subcontinent origin of Nepenthes colonization in south east Asia during the Miocene epoch. Frontiers in Ecology and Evolution 6.,
Phylogeny and biogeography of carnivorous plant family Nepenthaceae with reference to the Indian pitcher plant Nepenthes khasiana reveals an Indian subcontinent origin of Nepenthes colonization in south east Asia during the Miocene epoch.Crossref | GoogleScholarGoogle Scholar |

Bouckaert R, Heled J, Kühnert D, Vaughan T, Wu C-H, Xie D, Suchard MA, Rambaut A, Drummond AJ (2014) BEAST 2: a software platform for Bayesian evolutionary analysis. PLoS Computational Biology 10, e1003537
BEAST 2: a software platform for Bayesian evolutionary analysis.Crossref | GoogleScholarGoogle Scholar | 24722319PubMed |

Buckler ES, Ippolito A, Holtsford TP (1997) The evolution of ribosomal DNA divergent paralogues and phylogenetic implications. Genetics 145, 821–832.

Bunawan H, Yen CC, Yaakop S, Noor NM (2017) Phylogenetic inferences of Nepenthes species in Peninsular Malaysia revealed by chloroplast (trnL intron) and nuclear (ITS) DNA sequences. BMC Research Notes 10, 67
Phylogenetic inferences of Nepenthes species in Peninsular Malaysia revealed by chloroplast (trnL intron) and nuclear (ITS) DNA sequences.Crossref | GoogleScholarGoogle Scholar | 28126013PubMed |

Charlesworth B (2009) Effective population size and patterns of molecular evolution and variation. Nature Reviews. Genetics 10, 195–205.
Effective population size and patterns of molecular evolution and variation.Crossref | GoogleScholarGoogle Scholar | 19204717PubMed |

Cheek M, Jebb M (2001) ‘Flora Malesiana. Series I, Spermatophyta. Vol. 15: Nepenthaceae.’ (Nationaal Herbarium Nederland)

Chen Y-S, Meseguer AS, Godefroid M, Zhou Z, Zhang J-W, Deng T, Kim J-H, Nie Z-L, Liu Y-S, Sun H (2017) Out-of-India dispersal of Paliurus (Rhamnaceae) indicated by combined molecular phylogenetic and fossil evidence. Taxon 66, 78–90.
Out-of-India dispersal of Paliurus (Rhamnaceae) indicated by combined molecular phylogenetic and fossil evidence.Crossref | GoogleScholarGoogle Scholar |

Chin L, Moran JA, Clarke C (2010) Trap geometry in three giant montane pitcher plant species from Borneo is a function of tree shrew body size. New Phytologist 186, 461–470.
Trap geometry in three giant montane pitcher plant species from Borneo is a function of tree shrew body size.Crossref | GoogleScholarGoogle Scholar | 20100203PubMed |

Clarke C (1997) ‘Nepenthes of Borneo.’ (Natural History Publications Borneo: Kota Kinabalu, Sabah, Malaysia)

Clarke C (2001) ‘Nepenthes of Sumatra and Peninsular Malaysia’. (Natural History Publications Borneo: Kota Kinabalu, Sabah, Malaysia)

Clarke C, Schlauer J, Moran J, Robinson A (2018) Systematics and evolution in Nepenthes. In ‘Carnivorous Plants: Physiology, Ecology and Evolution’. (Eds AM Ellison, L Adamec) pp. 58–69. (Oxford University Press: Oxford, UK) 10.1093/oso/9780198779841.003.0005

Conti E, Eriksson T, Schonenberger J, Sytsma KJ, Baum DA (2002) Early tertiary out-of-India dispersal of Crypteroniaceae: evidence from phylogeny and molecular dating. Evolution 56, 1931–1942.
Early tertiary out-of-India dispersal of Crypteroniaceae: evidence from phylogeny and molecular dating.Crossref | GoogleScholarGoogle Scholar | 12449480PubMed |

Crawley SS, Hilu KW (2012) Impact of missing data, gene choice, and taxon sampling on phylogenetic reconstruction: the Caryophyllales (angiosperms). Plant Systematics and Evolution 298, 297–312.
Impact of missing data, gene choice, and taxon sampling on phylogenetic reconstruction: the Caryophyllales (angiosperms).Crossref | GoogleScholarGoogle Scholar |

Crayn DM, Costion C, Harrington MG (2015) The Sahul–Sunda floristic exchange: dated molecular phylogenies document Cenozoic intercontinental dispersal dynamics. Journal of Biogeography 42, 11–24.
The Sahul–Sunda floristic exchange: dated molecular phylogenies document Cenozoic intercontinental dispersal dynamics.Crossref | GoogleScholarGoogle Scholar |

Danser BH (1928) Contributions a l’etude de la flores des Indes Neerlandaises. XV. Bulletin du Jardin de Botanique Buitenzorg, Serie III. Vol. IX. Livre. 3–4, 249–438.

Darriba D, Taboada GL, Doallo R, Posada D (2012) jModelTest 2: more models, new heuristics and parallel computing. Nature Methods 9, 772
jModelTest 2: more models, new heuristics and parallel computing.Crossref | GoogleScholarGoogle Scholar | 22847109PubMed |

Degnan JH, Rosenberg NA (2009) Gene tree discordance, phylogenetic inference and the multispecies coalescent. Trends in Ecology & Evolution 24, 332–340.
Gene tree discordance, phylogenetic inference and the multispecies coalescent.Crossref | GoogleScholarGoogle Scholar |

Dodsworth S (2015) Genome skimming for next-generation biodiversity analysis. Trends in Plant Science 20, 525–527.
Genome skimming for next-generation biodiversity analysis.Crossref | GoogleScholarGoogle Scholar | 26205170PubMed |

Feliner GN, Rosselló JA (2007) Better the devil you know? Guidelines for insightful utilization of nrDNA ITS in species-level evolutionary studies in plants. Molecular Phylogenetics and Evolution 44, 911–919.
Better the devil you know? Guidelines for insightful utilization of nrDNA ITS in species-level evolutionary studies in plants.Crossref | GoogleScholarGoogle Scholar |

Fleischmann A, Schlauer J, Smith SA, Givnish TJ (2018) Evolution of carnivory in angiosperms. In ‘Carnivorous Plants: Physiology, Ecology, and Evolution’. (Eds AM Ellison, L Adamec) pp. 22–42. (Oxford University Press: Oxford, UK). 10.1093/oso/9780198779841.003.0003

Gilbert KJ, Nitta JH, Talavera G, Pierce NE (2018) Keeping an eye on coloration: ecological correlates of the evolution of pitcher traits in the genus Nepenthes (Caryophyllales). Biological Journal of the Linnean Society. Linnean Society of London 123, 321–337.
Keeping an eye on coloration: ecological correlates of the evolution of pitcher traits in the genus Nepenthes (Caryophyllales).Crossref | GoogleScholarGoogle Scholar |

Guo X, Thomas DC, Saunders RMK (2018) Gene tree discordance and coalescent methods support ancient intergeneric hybridisation between Dasymaschalon and Friesodielsia (Annonaceae). Molecular Phylogenetics and Evolution 127, 14–29.
Gene tree discordance and coalescent methods support ancient intergeneric hybridisation between Dasymaschalon and Friesodielsia (Annonaceae).Crossref | GoogleScholarGoogle Scholar | 29678645PubMed |

Hall R (2009) The Eurasian SE Asian margin as a modern example of an accretionary orogen. Geological Society of London, Special Publications 318, 351–372.
The Eurasian SE Asian margin as a modern example of an accretionary orogen.Crossref | GoogleScholarGoogle Scholar |

Jebb M, Cheek M (1997) A skeletal revision of Nepenthes (Nepenthaceae). Blumea 42, 1–106.

Katoh K, Standley DM (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Molecular Biology and Evolution 30, 772–780.
MAFFT multiple sequence alignment software version 7: improvements in performance and usability.Crossref | GoogleScholarGoogle Scholar | 23329690PubMed |

Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, Buxton S, Cooper A, Markowitz S, Duran C, Thierer T, Ashton B, Meintjes P, Drummond A (2012) Geneious basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics 28, 1647–1649.
Geneious basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data.Crossref | GoogleScholarGoogle Scholar | 22543367PubMed |

Krutzsch W (1989) Paleogeography and historical phytogeography (paleochorology) in the Neophyticum. Plant Systematics and Evolution 162, 5–61.
Paleogeography and historical phytogeography (paleochorology) in the Neophyticum.Crossref | GoogleScholarGoogle Scholar |

Linder CR, Goertzen LR, Heuvel BV, Francisco-Ortega J, Jansen RK (2000) The complete external transcribed spacer of 18S–26S rDNA: amplification and phylogenetic utility at low taxonomic levels in Asteraceae and closely allied families. Molecular Phylogenetics and Evolution 14, 285–303.
The complete external transcribed spacer of 18S–26S rDNA: amplification and phylogenetic utility at low taxonomic levels in Asteraceae and closely allied families.Crossref | GoogleScholarGoogle Scholar | 10679161PubMed |

Magallón S, Gómez-Acevedo S, Sánchez-Reyes LL, Hernández-Hernández T (2015) A metacalibrated time-tree documents the early rise of flowering plant phylogenetic diversity. New Phytologist 207, 437–453.
A metacalibrated time-tree documents the early rise of flowering plant phylogenetic diversity.Crossref | GoogleScholarGoogle Scholar | 25615647PubMed |

Malé P-JG, Bardon L, Besnard G, Coissac E, Delsuc F, Engel J, Lhuillier E, Scotti-Saintagne C, Tinaut A, Chave J (2014) Genome skimming by shotgun sequencing helps resolve the phylogeny of a pantropical tree family. Molecular Ecology Resources 14, 966–975.
Genome skimming by shotgun sequencing helps resolve the phylogeny of a pantropical tree family.Crossref | GoogleScholarGoogle Scholar |

McLoughlin S (2001) The breakup history of Gondwana and its impact on pre-Cenozoic floristic provincialism. Australian Journal of Botany 49, 271–300.
The breakup history of Gondwana and its impact on pre-Cenozoic floristic provincialism.Crossref | GoogleScholarGoogle Scholar |

Meimberg H (2002) Molekular-systematische Untersuchungen an den Familien Nepenthaceae und Ancistrocladaceae sowie verwandte Taxa aus der Unterklasse Caryophyllidae s.l. PhD thesis, Ludwig Maximilian University of Munich, Munich, Germany.

Meimberg H, Heubl G (2006) Introduction of a nuclear marker for phylogenetic analysis of Nepenthaceae. Plant Biology 8, 831–840.
Introduction of a nuclear marker for phylogenetic analysis of Nepenthaceae.Crossref | GoogleScholarGoogle Scholar | 17203435PubMed |

Meimberg H, Wistuba A, Dittrich P, Heubl G (2001) Molecular phylogeny of Nepenthaceae based on cladistic analysis of plastid trnK intron sequence data. Plant Biology 3, 164–175.
Molecular phylogeny of Nepenthaceae based on cladistic analysis of plastid trnK intron sequence data.Crossref | GoogleScholarGoogle Scholar |

Meimberg H, Thalhammer S, Brachmann A, Heubl G (2006) Comparative analysis of a translocated copy of the trnK intron in carnivorous family Nepenthaceae. Molecular Phylogenetics and Evolution 39, 478–490.
Comparative analysis of a translocated copy of the trnK intron in carnivorous family Nepenthaceae.Crossref | GoogleScholarGoogle Scholar | 16414286PubMed |

Merckx VSFT, Hendriks KP, Beentjes KK, Mennes CB, Becking LE, Peijnenburg KTCA, Afendy A, Arumugam N, de Boer H, Biun A, Buang MM, Chen P-P, Chung AYC, Dow R, Feijen FAA, Feijen H, Soest CF, Geml J, Geurts R, Gravendeel B, Hovenkamp P, Imbun P, Ipor I, Janssens SB, Jocqué M, Kappes H, Khoo E, Koomen P, Lens F, Majapun RJ, Morgado LN, Neupane S, Nieser N, Pereira JT, Rahman H, Sabran S, Sawang A, Schwallier RM, Shim P-S, Smit H, Sol N, Spait M, Stech M, Stokvis F, Sugau JB, Suleiman M, Sumail S, Thomas DC, van Tol J, Tuh FYY, Yahya BE, Nais J, Repin R, Lakim M, Schilthuizen M (2015) Evolution of endemism on a young tropical mountain. Nature 524, 347–350.
Evolution of endemism on a young tropical mountain.Crossref | GoogleScholarGoogle Scholar |

Moran JA, Gray LK, Clarke C, Chin L (2013) Capture mechanism in Palaeotropical pitcher plants (Nepenthaceae) is constrained by climate. Annals of Botany 112, 1279–1291.
Capture mechanism in Palaeotropical pitcher plants (Nepenthaceae) is constrained by climate.Crossref | GoogleScholarGoogle Scholar | 23975653PubMed |

Moyle RG (2004) Calibration of molecular clocks and the biogeographic history of Crypteroniaceae. Evolution 58, 1871–1873.
Calibration of molecular clocks and the biogeographic history of Crypteroniaceae.Crossref | GoogleScholarGoogle Scholar | 15446441PubMed |

Mullins JT (2000) Molecular Cystematics of Nepenthaceae. PhD dissertation, University of Reading, Reading, UK.

Nauheimer L, Boyce PC, Renner SS (2012) Giant taro and its relatives: a phylogeny of the large genus Alocasia (Araceae) sheds light on Miocene floristic exchange in the Malesian region. Molecular Phylogenetics and Evolution 63, 43–51.
Giant taro and its relatives: a phylogeny of the large genus Alocasia (Araceae) sheds light on Miocene floristic exchange in the Malesian region.Crossref | GoogleScholarGoogle Scholar | 22209857PubMed |

Pelser PB, Kennedy AH, Tepe EJ, Shidler JB, Nordenstam B, Kadereit JW, Watson LE (2010) Patterns and causes of incongruence between plastid and nuclear Senecioneae (Asteraceae) phylogenies. American Journal of Botany 97, 856–873.
Patterns and causes of incongruence between plastid and nuclear Senecioneae (Asteraceae) phylogenies.Crossref | GoogleScholarGoogle Scholar | 21622451PubMed |

Reeves C (2014) The position of Madagascar within Gondwana and its movements during Gondwana dispersal. Journal of African Earth Sciences 94, 45–57.
The position of Madagascar within Gondwana and its movements during Gondwana dispersal.Crossref | GoogleScholarGoogle Scholar |

Rieseberg LH, Soltis DE (1991) Phylogenetic consequences of cytoplasmic gene flow in plants. Evolutionary Trends in Plants 5, 65–84.

Ripma LA, Simpson MG, Hasenstab‐Lehman K (2014) Geneious! Simplified genome skimming methods for phylogenetic systematic studies: a case study in Oreocarya (Boraginaceae). Applications in Plant Sciences 2, 1400062
Geneious! Simplified genome skimming methods for phylogenetic systematic studies: a case study in Oreocarya (Boraginaceae).Crossref | GoogleScholarGoogle Scholar | 25506521PubMed |

Ronquist F, Huelsenbeck JP (2003) MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19, 1572–1574.
MrBayes 3: Bayesian phylogenetic inference under mixed models.Crossref | GoogleScholarGoogle Scholar | 12912839PubMed |

Schwallier R, Raes N, Boer HJ, Vos RA, Vugt RR, Gravendeel B, Beaumont L (2016) Phylogenetic analysis of niche divergence reveals distinct evolutionary histories and climate change implications for tropical carnivorous pitcher plants. Diversity & Distributions 22, 97–110.
Phylogenetic analysis of niche divergence reveals distinct evolutionary histories and climate change implications for tropical carnivorous pitcher plants.Crossref | GoogleScholarGoogle Scholar |

Simpson MG, Guilliams CM, Hasenstab-Lehman KE, Mabry ME, Ripma L (2017) Phylogeny of the popcorn flowers: use of genome skimming to evaluate monophyly and interrelationships in subtribe Amsinckiinae (Boraginaceae). Taxon 66, 1406–1420.
Phylogeny of the popcorn flowers: use of genome skimming to evaluate monophyly and interrelationships in subtribe Amsinckiinae (Boraginaceae).Crossref | GoogleScholarGoogle Scholar |

Stamatakis A (2014) RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30, 1312–1313.
RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies.Crossref | GoogleScholarGoogle Scholar | 24451623PubMed |

Straub SCK, Parks M, Weitemier K, Fishbein M, Cronn RC, Liston A (2012) Navigating the tip of the genomic iceberg: next‐generation sequencing for plant systematics. American Journal of Botany 99, 349–364.
Navigating the tip of the genomic iceberg: next‐generation sequencing for plant systematics.Crossref | GoogleScholarGoogle Scholar |

Tsitrone A, Kirkpatrick M, Levin DA, Morgan M (2003) A model for chloroplast capture. Evolution 57, 1776–1782.
A model for chloroplast capture.Crossref | GoogleScholarGoogle Scholar | 14503619PubMed |

Zimmer EA, Wen J (2015) Using nuclear gene data for plant phylogenetics: progress and prospects II. Next‐gen approaches. Journal of Systematics and Evolution 53, 371–379.
Using nuclear gene data for plant phylogenetics: progress and prospects II. Next‐gen approaches.Crossref | GoogleScholarGoogle Scholar |