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  • Hanxi Li, Xuena Huang, Aibin Zhan
    Marine Life Science & Technology, 2024, 6(2): 315-330. https://doi.org/10.1007/s42995-024-00228-y

    Marine ecosystems are facing escalating environmental fluctuations owing to climate change and human activities, imposing pressures on marine species. To withstand recurring environmental challenges, marine organisms, especially benthic species lacking behavioral choices to select optimal habitats, have to utilize well-established strategies such as the antioxidant defense system (ADS) to ensure their survival. Therefore, understanding of the mechanisms governing the ADS-based response is essential for gaining insights into adaptive strategies for managing environmental challenges. Here we conducted a comparative analysis of the physiological and transcriptional responses based on the ADS during two rounds of 'hypersalinity-recovery' challenges in two model congeneric invasive ascidians, Ciona robusta and C. savignyi. Our results demonstrated that C. savignyi exhibited higher tolerance and resistance to salinity stresses at the physiological level, while C. robusta demonstrated heightened responses at the transcriptional level. We observed distinct transcriptional responses, particularly in the utilization of two superoxide dismutase (SOD) isoforms. Both Ciona species developed physiological stress memory with elevated total SOD (T-SOD) and glutathione (GSH) responses, while only C. robusta demonstrated transcriptional stress memory. The regulatory distinctions within the Nrf2-Keap1 signalling pathway likely explain the formation disparity of transcriptional stress memory between both Ciona species. These findings support the 'context-dependent stress memory hypothesis', emphasizing the emergence of species-specific stress memory at diverse regulatory levels in response to recurrent environmental challenges. Our results enhance our understanding of the mechanisms of environmental challenge management in marine species, particularly those related to the ADS.

  • Kyung Hyun Kim, Jeong Min Kim, Ju Hye Baek, Sang Eun Jeong, Hocheol Kim, Hwan Su Yoon, Che Ok Jeon
    Marine Life Science & Technology, 2024, 6(2): 298-314. https://doi.org/10.1007/s42995-024-00227-z

    Mutualistic interactions between marine phototrophs and associated bacteria are an important strategy for their successful survival in the ocean, but little is known about their metabolic relationships. Here, bacterial communities in the algal sphere (AS) and bulk solution (BS) of nine marine red algal cultures were analyzed, and Roseibium and Phycisphaera were identified significantly more abundantly in AS than in BS. The metabolic features of Roseibium RMAR6-6 (isolated and genome-sequenced), Phycisphaera MAG 12 (obtained by metagenomic sequencing), and a marine red alga, Porphyridium purpureum CCMP1328 (from GenBank), were analyzed bioinformatically. RMAR6-6 has the genetic capability to fix nitrogen and produce B vitamins (B1, B2, B5, B6, B9, and B12), bacterioferritin, dimethylsulfoniopropionate (DMSP), and phenylacetate that may enhance algal growth, whereas MAG 12 may have a limited metabolic capability, not producing vitamins B9 and B12, DMSP, phenylacetate, and siderophores, but with the ability to produce bacitracin, possibly modulating algal microbiome. P. purpureum CCMP1328 lacks the genetic capability to fix nitrogen and produce vitamin B12, DMSP, phenylacetate, and siderophore. It was shown that the nitrogen-fixing ability of RMAR6-6 promoted the growth of P. purpureum, and DMSP reduced the oxidative stress of P. purpureum. The metabolic interactions between strain RMAR6-6 and P. purpureum CCMP1328 were also investigated by the transcriptomic analyses of their monoculture and co-culture. Taken together, potential metabolic relationships between Roseibium and P. purpureum were proposed. This study provides a better understanding of the metabolic relationships between marine algae and algae-associated bacteria for successful growth.

  • Zhixin Zhang, Jinxin Zhou, Jorge García Molinos, Stefano Mammola, Ákos Bede-Fazekas, Xiao Feng, Daisuke Kitazawa, Jorge Assis, Tianlong Qiu, Qiang Lin
    Marine Life Science & Technology, 2024, 6(2): 349-362. https://doi.org/10.1007/s42995-024-00226-0

    Correlative species distribution models (SDMs) are important tools to estimate species’ geographic distribution across space and time, but their reliability heavily relies on the availability and quality of occurrence data. Estimations can be biased when occurrences do not fully represent the environmental requirement of a species. We tested to what extent species’ physiological knowledge might influence SDM estimations. Focusing on the Japanese sea cucumber Apostichopus japonicus within the coastal ocean of East Asia, we compiled a comprehensive dataset of occurrence records. We then explored the importance of incorporating physiological knowledge into SDMs by calibrating two types of correlative SDMs: a naïve model that solely depends on environmental correlates, and a physiologically informed model that further incorporates physiological information as priors. We further tested the models’ sensitivity to calibration area choices by fitting them with different buffered areas around known presences. Compared with naïve models, the physiologically informed models successfully captured the negative influence of high temperature on A. japonicus and were less sensitive to the choice of calibration area. The naïve models resulted in more optimistic prediction of the changes of potential distributions under climate change (i.e., larger range expansion and less contraction) than the physiologically informed models. Our findings highlight benefits from incorporating physiological information into correlative SDMs, namely mitigating the uncertainties associated with the choice of calibration area. Given these promising features, we encourage future SDM studies to consider species physiological information where available.

  • Hao Ling, Yongxin Lv, Yu Zhang, Ning-Yi Zhou, Ying Xu
    Marine Life Science & Technology, 2024, 6(2): 331-348. https://doi.org/10.1007/s42995-024-00224-2

    Phenolic compounds, as well as other aromatic compounds, have been reported to be abundant in hadal trenches. Although high-throughput sequencing studies have hinted at the potential of hadal microbes to degrade these compounds, direct microbiological, genetic and biochemical evidence under in situ pressures remain absent. Here, a microbial consortium and a pure culture of Pseudomonas, newly isolated from Mariana Trench sediments, efficiently degraded phenol under pressures up to 70 and 60 MPa, respectively, with concomitant increase in biomass. By analyzing a high-pressure (70 MPa) culture metatranscriptome, not only was the entire range of metabolic processes under high pressure generated, but also genes encoding complete phenol degradation via ortho- and meta-cleavage pathways were revealed. The isolate of Pseudomonas also contained genes encoding the complete degradation pathway. Six transcribed genes (dmpKLMNOP sed) were functionally identified to encode a multicomponent hydroxylase catalyzing the hydroxylation of phenol and its methylated derivatives by heterogeneous expression. In addition, key catabolic genes identified in the metatranscriptome of the high-pressure cultures and genomes of bacterial isolates were found to be all widely distributed in 22 published hadal microbial metagenomes. At microbiological, genetic, bioinformatics, and biochemical levels, this study found that microorganisms widely found in hadal trenches were able to effectively drive phenolic compound degradation under high hydrostatic pressures. This information will bridge a knowledge gap concerning the microbial aromatics degradation within hadal trenches.

  • Huijuan Hao, Yinjie Lian, Chenhui Ren, Sitong Yang, Min Zhao, Tao Bo, Jing Xu, Wei Wang
    Marine Life Science & Technology, 2024, 6(2): 183-197. https://doi.org/10.1007/s42995-024-00219-z

    Histone modification and nucleosome assembly play important roles in chromatin-related processes. Histone chaperones form different complexes and coordinate histone transportation and assembly. Various histone chaperone complexes have been identified in different organisms. The ciliate protozoa (ciliates) have various chromatin structures and different nuclear morphology. However, histone chaperone components and functions of different subunits remain unclear in ciliates. Tetrahymema thermophila contains a transcriptionally active macronucleus (MAC) and a transcriptionally inactive micronucleus (MIC) which exhibit multiple replication and various chromatin remodeling progresses during vegetative growth and sexual developmental stages. Here, we found histone chaperone RebL1 not only localized evenly in the transcriptionally active MAC but also dynamically changed in the MIC during vegetative growth and sexual developmental stages. REBL1 knockdown inhibited cellular proliferation. The macronuclear morphology became bigger in growing mutants. The abnormal macronuclear structure also occurred in the starvation stage. Furthermore, micronuclear meiosis was disturbed during sexual development, leading to a failure to generate new gametic nuclei. RebL1 potentially interacted with various factors involved in histone-modifying complexes and chromatin remodeling complexes in different developmental stages. REBL1 knockdown affected expression levels of the genes involved in chromatin organization and transcription. Taken together, RebL1 plays a vital role in maintaining macronuclear structure stability and gametogenesis in T. thermophila.

  • Monika Tunjić-Cvitanić, Daniel García-Souto, Juan J. Pasantes, Eva Šatović-Vukšić
    Marine Life Science & Technology, 2024, 6(2): 236-251. https://doi.org/10.1007/s42995-024-00218-0

    Research on bivalves is fast-growing, including genome-wide analyses and genome sequencing. Several characteristics qualify oysters as a valuable model to explore repetitive DNA sequences and their genome organization. Here we characterize the satellitomes of five species in the family Ostreidae (Crassostrea angulata, C. virginica, C. hongkongensis, C. ariakensis, Ostrea edulis), revealing a substantial number of satellite DNAs (satDNAs) per genome (ranging between 33 and 61) and peculiarities in the composition of their satellitomes. Numerous satDNAs were either associated to or derived from transposable elements, displaying a scarcity of transposable element-unrelated satDNAs in these genomes. Due to the non-conventional satellitome constitution and dominance of Helitron-associated satDNAs, comparative satellitomics demanded more in-depth analyses than standardly employed. Comparative analyses (including C. gigas, the first bivalve species with a defined satellitome) revealed that 13 satDNAs occur in all six oyster genomes, with Cg170/HindIII satDNA being the most abundant in all of them. Evaluating the “satDNA library model” highlighted the necessity to adjust this term when studying tandem repeat evolution in organisms with such satellitomes. When repetitive sequences with potential variation in the organizational form and repeat-type affiliation are examined across related species, the introduction of the terms “TE library” and “repetitive DNA library” becomes essential.

  • Likang Lyu, Yijia Yao, Songyang Xie, Xiaojie Wang, Haishen Wen, Yun Li, Jianshuang Li, Chenpeng Zuo, Shaojing Yan, Jingyi Dong, Xin Qi
    Marine Life Science & Technology, 2024, 6(1): 15-30. https://doi.org/10.1007/s42995-023-00214-w

    Prostaglandins (PGs) are profound hormones in teleost sexual behavior, especially in mating. PGs act as pheromones that affect the olfactory sensory neurons of males, inducing the initiation of a series of mating behaviors. However, the molecular mechanism by which PGs trigger mating behavior in ovoviviparous teleosts is still unclear. In the present study, we employed the ovoviviparous black rockfish (Sebastes schlegelii), an economically important marine species whose reproductive production is limited by incomplete fertilization, as a model species. The results showed that when the dose of PGE2 was higher than 10 nmol/L, a significant (P < 0.05) increase in mating behaviors was observed. Dual-fluorescence in situ hybridization indicated that PGE2 could fire specific neurons in different brain regions and receptor cells in the olfactory sac. After combining with specific neurons in the central nervous system (CNS), a series of genes related to reproduction are activated. The intracerebroventricular administration of PGE2 significantly increased lhb levels (P < 0.05) in both sexes. Moreover, steroidogenesis in gonads was also affected, inducing an increase (P < 0.05) in E2 levels in males and T levels in females. PGE2 levels were also increased significantly (P < 0.05) in both sexes. The present study revealed that PGE2 can activate mating behavior in black rockfish in both hormone and pheromone pathways, leading to variations in sex steroid levels and activation of reproductive behaviors. Our results provide not only novel insight into the onset of mating behaviors in ovoviviparous teleosts but also solutions for the incomplete fertilization caused by natural mating in cage aquaculture.

  • Jiwen Liu, Da-Wei Li, Xinxin He, Ronghua Liu, Haojin Cheng, Chenglong Su, Mengna Chen, Yonghong Wang, Zhongsheng Zhao, Hanyue Xu, Zhangyu Cheng, Zicheng Wang, Nikolai Pedentchouk, David J. Lea-Smith, Jonathan D. Todd, Xiaoshou Liu, Meixun Zhao, Xiao-Hua Zhang
    Marine Life Science & Technology, 2024, 6(1): 168-181. https://doi.org/10.1007/s42995-023-00212-y

    Hadal trenches are characterized by enhanced and infrequent high-rate episodic sedimentation events that likely introduce not only labile organic carbon and key nutrients but also new microbes that significantly alter the subseafloor microbiosphere. Currently, the role of high-rate episodic sedimentation in controlling the composition of the hadal subseafloor microbiosphere is unknown. Here, analyses of carbon isotope composition in a ~ 750 cm long sediment core from the Challenger Deep revealed noncontinuous deposition, with anomalous 14C ages likely caused by seismically driven mass transport and the funneling effect of trench geomorphology. Microbial community composition and diverse enzyme activities in the upper ~ 27 cm differed from those at lower depths, probably due to sudden sediment deposition and differences in redox condition and organic matter availability. At lower depths, microbial population numbers, and composition remained relatively constant, except at some discrete depths with altered enzyme activity and microbial phyla abundance, possibly due to additional sudden sedimentation events of different magnitude. Evidence is provided of a unique role for high-rate episodic sedimentation events in controlling the subsurface microbiosphere in Earth’s deepest ocean floor and highlight the need to perform thorough analysis over a large depth range to characterize hadal benthic populations. Such depositional processes are likely crucial in shaping deep-water geochemical environments and thereby the deep subseafloor biosphere.

  • Juanjuan Su, Hao Wu, Chengying Yin, Fengchao Zhang, Feng Han, Wengong Yu
    Marine Life Science & Technology, 2023, 6(1): 93-101. https://doi.org/10.1007/s42995-023-00201-1

    The application of chondroitinase requires consideration of the complex microenvironment of the target. Our previous research reported a marine-derived sodium dodecyl sulfate (SDS)-resistant chondroitinase VhChlABC. This study further investigated the mechanism of VhChlABC resistance to SDS. Focusing on the hydrophobic cluster on its strong hydrophilic surface, it was found that the reduction of hydrophobicity of surface residues Ala181, Met182, Met183, Ala184, Val185, and Ile305 significantly reduced the SDS resistance and stability. Molecular dynamics (MD) simulation and molecular docking analysis showed that I305G had more conformational flexibility around residue 305 than wild type (WT), which was more conducive to SDS insertion and binding. The affinity of A181G, M182A, M183A, V185A and I305G to SDS was significantly higher than that of WT. In conclusion, the surface hydrophobic microenvironment composed of six residues was the structural basis for SDS resistance. This feature could prevent the binding of SDS and the destruction of hydrophobic packaging by increasing the rigid conformation of protein and reducing the binding force of SDS-protein. The study provides a new idea for the rational design of SDS-resistant proteins and may further promote chondroitinase research in the targeted therapy of lung diseases under the pressure of pulmonary surfactant.

  • Weiguang Kong, Guangyi Ding, Gaofeng Cheng, Peng Yang, Zhen Xu
    Marine Life Science & Technology, 2023, 6(2): 266-279. https://doi.org/10.1007/s42995-023-00199-6

    The eye, as a specialized visual organ, is directly exposed to the external environment, and, therefore, it faces constant challenges from external pathogenic organisms and toxins. In the ocular mucosa (OM) of mammals, mucosal-associated lymphoid tissues (MALTs) constitute the primary line of defense. However, the immune defense role of the OM remains unknown in aquatic vertebrates. To gain insights into the immune processes within the OM of teleost fish, we developed an infection model of rainbow trout (Oncorhynchus mykiss) OM using a parasite, Ichthyophthirius multifiliis (Ich). Immunofluorescence, qPCR, and H&E staining revealed that Ich successfully infiltrates the OM of rainbow trout, leading to pathological structural changes, as evidenced by A&B staining. Importantly, the qPCR results indicate an up-regulation of immune-related genes following Ich infection in the OM. Moreover, transcriptome analyses were conducted to detect immune responses and impairments in eye function within the OM of rainbow trout with Ich infection. The results of the transcriptome analysis that Ich infection can cause an extensive immune response in the OM, ultimately affecting ocular function. To the best of our knowledge, our findings represent for the first time that the teleost OM could act as an invasion site for parasites and trigger a strong mucosal immune response to parasitic infection.

  • Branko Glamuzina, Lorenzo Vilizzi, Marina Piria, Ante Žuljević, Ana Bratoš Cetinić, Ana Pešić, Branko Dragičević, Lovrenc Lipej, Marijana Pećarević, Vlasta Bartulović, Sanja Grđan, Ivan Cvitković, Tatjana Dobroslavić, Ana Fortič, Luka Glamuzina, Borut Mavrič, Jovana Tomanić, Marija Despalatović, Domen Trkov, Marina Brailo Šćepanović, Zoran Vidović, Predrag Simonović, Sanja Matić-Skoko, Pero Tutman
    Marine Life Science & Technology, 2023, 6(1): 143-154. https://doi.org/10.1007/s42995-023-00196-9

    Globally, marine bioinvasions threaten marine ecosystem structure and function, with the Mediterranean Sea being one of the most affected regions. Such invasions are expected to increase due to climate change. We conducted a risk screening of marine organisms (37 fishes, 38 invertebrates, and 9 plants), both extant and ‘horizon’ (i.e., not present in the area but likely to enter it). Based on expert knowledge for the Eastern Adriatic Sea coasts of Slovenia, Croatia, and Montenegro, screenings were conducted under both current and predicted climate conditions indicating with an increase in sea surface temperature and salinity of the Adriatic Sea together with changes in precipitation regime. Our aims were to: (1) identify non-native extant and horizon marine species that may pose threats to native biodiversity and (2) evaluate the risk of invasiveness of the selected species under current and predicted climate conditions. Of the 84 species screened, there was an increase in those ranked as ‘high risk’ from 33 (39.3%) under current climate conditions and to 47 (56.0%) under global warming scenarios. For those ranked as ‘very high’ risk, the increase was from 6 (7.1%) to 21 (25.0%). Amongst the screened species, the already established high-risk species Pacific oyster Magallana gigas and Atlantic blue crab Callinectes sapidus represent a threat to ecosystem services. Given the under-representation of marine species in the current European Union List, the species we have ranked as high to very high risk should be included.

  • Sanqiang Gong, Jiayuan Liang, Gang Li, Lijia Xu, Yehui Tan, Xinqing Zheng, Xuejie Jin, Kefu Yu, Xiaomin Xia
    Marine Life Science & Technology, 2023, 6(1): 155-167. https://doi.org/10.1007/s42995-023-00190-1

    Coral fluorescence phenotypes have been suggested as an adaptation to a broad range of environmental conditions, yet the mechanisms linking thermal bleaching tolerance in reef-building coral populations, associated with fluorescence phenotypes due to GFP-like proteins, remains unclear. In this study, the relationship between the thermal sensitivity and phenotypic plasticity of corals was investigated using two phenotypes of Galaxea fascicularis, green and brown. The results reveal that brown G. fascicularis was more susceptible to bleaching than green G. fascicularis when exposed to a higher growth temperature of 32 °C. Both phenotypes of G. fascicularis were associated with the thermotolerant Symbiodiniaceae symbiont, Durusdinium trenchii. However, the brown G. fascicularis showed a significant decrease in Symbiodiniaceae cell density and a significant increase in pathogenic bacteria abundance when the growth temperature was raised from 29 to 32 °C. The physiological traits and transcriptomic profiles of Symbiodiniaceae were not notably affected, but there were differences in the transcriptional levels of certain genes between the two phenotype hosts of G. fascicularis. Under heat stress of 32 °C, the gene encoding green fluorescent protein (GFP)-like and chromosome-associated proteins, as well as genes related to oxidative phosphorylation, cell growth and death showed lower transcriptional levels in the brown G. fascicularis compared to the green G. fascicularis. Overall, the results demonstrate that the green form of G. fascicularis is better able to tolerate ocean warming and defend against pathogenic bacteria, likely due to higher gene transcription levels and defense ability.

  • Shuhao Liu, Yang Liu, Katharina Teschke, Mark A. Hindell, Rachel Downey, Briannyn Woods, Bin Kang, Shuyang Ma, Chi Zhang, Jianchao Li, Zhenjiang Ye, Peng Sun, Jianfeng He, Yongjun Tian
    Marine Life Science & Technology, 2023, 6(1): 68-83. https://doi.org/10.1007/s42995-023-00188-9

    Mesopelagic fish (meso-fish) are central species within the Southern Ocean (SO). However, their ecosystem role and adaptive capacity to climate change are rarely integrated into protected areas assessments. This is a pity given their importance as crucial prey and predators in food webs, coupled with the impacts of climate change. Here, we estimate the habitat distribution of nine meso-fish using an ensemble model approach (MAXENT, random forest, and boosted regression tree). Four climate model simulations were used to project their distribution under two representative concentration pathways (RCP4.5 and RCP8.5) for short-term (2006–2055) and long-term (2050–2099) periods. In addition, we assess the ecological representativeness of protected areas under climate change scenarios using meso-fish as indicator species. Our models show that all species shift poleward in the future. Lanternfishes (family Myctophidae) are predicted to migrate poleward more than other families (Paralepididae, Nototheniidae, Bathylagidae, and Gonostomatidae). In comparison, lanternfishes were projected to increase habitat area in the eastern SO but lose area in the western SO; the opposite was projected for species in other families. Important areas (IAs) of meso-fish are mainly distributed near the Antarctic Peninsula and East Antarctica. Negotiated protected area cover 23% of IAs at present and 38% of IAs in the future (RCP8.5, long-term future). Many IAs of meso-fish still need to be included in protected areas, such as the Prydz Bay and the seas around the Antarctic Peninsula. Our results provide a framework for evaluating protected areas incorporating climate change adaptation strategies for protected areas management.

  • Tao Li, Tengyue Zhang, Mingjian Liu, Zhe Zhang, Jiachen Zhang, Junhua Niu, Xiangrui Chen, Saleh A. Al-Farraj, Weibo Song
    Marine Life Science & Technology, 2024, 6(2): 212-235. https://doi.org/10.1007/s42995-024-00230-4

    Species of the ciliate genera Myxophyllum and Conchophthirus are found as endocommensals of terrestrial and freshwater mollusks, respectively. So far, there have been few studies of these genera and morphological data for most members are often incomplete. In the present work, two new species, Myxophyllum weishanense sp. nov. and Conchophthirus paracurtus sp. nov., and a known species, Conchophthirus lamellidens, were isolated from hosts in Lake Weishan Wetland, China. Taxonomic studies indicate that M. weishanense sp. nov. can be recognized mainly by the combination of about 60 somatic kineties on both ventral and dorsal sides and the presence of caudal cilia. Conchophthirus paracurtus sp. nov. differs from congeners in its body shape and size, having a glabrous area on the posterior right side, and having fewer somatic kineties. In addition, differences in their ITS2 (Internally Transcribed Spacer 2) secondary structures support the discrimination of the two new species from their highly similar congeners. An improved diagnosis for the poorly known species, C. lamellidens is also provided. Phylogenetic analyses reveal that members of the genus Myxophyllum belong to a fully supported clade that is sister to a large, poorly supported clade consisting of Hemispeiridae, Ancistridae, and several lineages of the nonmonophyletic Cyclidiidae. The Myxophyllum clade also includes Protophyra ovicola JQ956552, a possible misidentification. Sequences of the two new Conchophthirus species cluster with other congeners in a fully supported clade that is unrelated to either the ‘typical’ thigmotrichs or to pleuronematids, thus conflicting with the traditional classification, and may represent an orphan scuticociliate lineage.

  • Weicai Song, Wenbo Shi, Huan Wang, Zirui Zhang, Ruiqing Tao, Jin Liu, Shuo Wang, Michael S. Engel, Chao Shi
    Marine Life Science & Technology, 2024, 6(3): 425-441. https://doi.org/10.1007/s42995-024-00242-0

    The aquatic plant Nymphaea, a model genus of the early flowering plant lineage Nymphaeales and family Nymphaeaceae, has been extensively studied. However, the availability of chloroplast genome data for this genus is incomplete, and phylogenetic relationships within the order Nymphaeales remain controversial. In this study, 12 chloroplast genomes of Nymphaea were assembled and analyzed for the first time. These genomes were 158,290–160,042 bp in size and contained 113 non-repeat genes, including 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. We also report on codon usage, RNA editing sites, microsatellite structures, and new repetitive sequences in this genus. Comparative genomics revealed that expansion and contraction of IR regions can lead to changes in the gene numbers. Additionally, it was observed that the highly variable regions of the chloroplast genome were mainly located in intergenic regions. Furthermore, the phylogenetic tree showed the order Nymphaeales was divided into three families, and the genus Nymphaea can be divided into five (or three) subgenera, with the subgenus Nymphaea being the oldest. The divergence times of nymphaealean taxa were analyzed, with origins of the order Nymphaeales and family Nymphaeaceae being about 194 and 131 million years, respectively. The results of the phylogenetic analysis and estimated divergence times will be useful for future evolutionary studies of basal angiosperm lineages.