Biological Sciences
Paya Sarraf

Phylogeny and Biogeography of Maclura

  • Faculty Advisor

    Nyree Zerega

Published On

May 2015

Originally Published

NURJ 2014-15


Maclura, a genus of plants in the mulberry family (Moraceae), is comprised of twelve species that inhabit every continent except for Antarctica. The widespread distribution of Maclura, despite its limited number of species, brings up inquiries about its origination, dissemination and evolution. The phylogeny of Maclura and its biogeography was examined through the use of molecular sequence data, fossil calibrations, and independent geological data. Previous studies indicate that most Moraceae lineages originated in the Old World. In order to test whether Maclura follows this trend, DNA from chloroplast gene regions as well as two nuclear gene regions, g3pdh and ITS, were sequenced. Bayesian inference methods were then used to analyze these sequences, along with age constraints from Maclura fossils. This dating analysis concluded that Maclura originated about 62 mya (95% interval [47,79 mya]) and that the split between New World and Old World Maclura species occurred about 46 mya (95% [33,60 mya]). By comparing these divergence times to the timeline of continental separation, we hypothesize that Maclura originated in South America, spread to North America, then Europe and finally to Africa and Asia. Additionally, we found that while Maclura is monophyletic, its sections are not, as Maclura africana, originally in Section Cardiogyne (New World), is nested within the clade containing section Plecospermum (Old World). This reclassification is crucial to our proposed biogeographical hypothesis because it indicates the presence of a clear New World-Old World split, which narrows the possible migratory paths of Maclura. These research findings are important because they not only help us understand the vicariance of Earth’s landmasses and Maclura’s biogeography, but also because they outline important evolutionary trends in Moraceae and contribute to a better understanding of its diversity.


The classification of biological organisms has long been a human pursuit as it aids in the understanding of our world, how we fit in it, and how we can utilize natural resources. Over time the guiding principles for the organization of biological organisms have changed. In the case of plants, for example, some of the earliest classification systems were developed by medical doctors because plants were the primary source of medicine (i.e. De Materia Medica by Dioscorides). Today, however, classifications strive to reflect evolutionary history (Judd et al 2007). Having phylogenetic classifications not only gives us insight into evolutionary processes, but also furthers scientific capabilities in a variety of pursuits. For example, phylogenetic approaches facilitate understanding of not only extant distributions of taxa, but also of extinct taxa. In addition, phylogenetic methods can provide insights into the earth’s paleoclimate because the evolution of biological organisms is closely related to changes in the environment. This is especially true of plants, which have great diversity and distributions throughout the globe and are often unable to migrate as quickly as animals. Therefore, they can provide valuable information about the planet’s history. The question of how plants came to be so diverse and occupy their current distributions are considerations that have important implications for plant conservation, biogeography and taxonomy, and are integral in the field of plant phylogenetics.

Maclura and the Moraceae family

The mulberry family (Moraceae) allows us to address a variety of interesting questions about evolution, classification, and biogeography. This large family, composed of 39 genera and approximately 1100 species, has a global distribution, but its greatest diversity lies in the tropics (Clement and Weiblen 2009). The Moraceae family is of great interest; at least 80 papers on Moraceae have been published since 2006 in areas such as ecology, conservation biology, coevolution and phytochemistry (Clement and Weiblen 2009).

Karren Wcisel,

Figure 1. Fruit of the Maclura pomifera tree. The tree, sometimes called the hedge-apple, is used by humans for a variety of purposes.

Moraceae is classified into six tribes, and the focus of this study is the tribe Maclureae. Like most of Moraceae, Maclureae is centered in the tropics, with a few species extending into temperate regions, such as North America. Maclureae is comprised of a single genus, Maclura, which is intriguing because it is one of the few examples of a small genus (consisting of 12 species divided into 5 sections) with a cosmopolitan distribution, inhabiting every continent except Antarctica. Maclura pomifera, the only North American member of the genus, is native to Texas and parts of Oklahoma (Fig. 1). The tree itself is often used as a hedge plant (the common name for the tree is hedge apple), and its dense wood makes for decay-resistant and durable tools (Culina 2002). The fruit of Maclura pomifera have medicinal properties and are used today as pesticides (Jaurdon 1997).

Table 1. Classification of the genus Maclura.

Objectives and Hypotheses

This study aims to use data from DNA sequences from the chloroplast and nuclear genome along with morphological, fossil, and spatial data in order to better understand the phylogeny and biogeography of the genus Maclura. Specific hypotheses that will be addressed are: 1) Maclura is a monophyletic genus, 2) The sections within Maclura are monophyletic, and 3) Maclura originated in the Old World and subsequently migrated to the New World. By reconstructing the phylogeny of the genus, an evolutionary framework will be in place to investigate questions about its biogeography.

Materials and Methods

Taxon Sampling

Thirty samples representing all 12 Maclura species were included in the study. In addition, 27 Moraceae species were used as outgroups. DNA was extracted (using DNeasy, Qiagen, Valencia, CA) and PCR amplified for seven regions from the nuclear and chloroplast genome (primers from: White et al. 1990, Strand et al. 1997, Shaw et al. 2005, 2007). DNA was cycle sequenced, cleaned and run on an ABI Prism 3700 following standard protocol (Zerega et al. 2010). Sequences were edited and aligned in Codon Aligner (Centerville, MA) and Mesquite (Maddison and Maddison 2011). Phylogenetic analyses and divergence date estimates using Bayesian analyses and fossil calibrations were set up in BEAUti, and run in BEAST (Drummond et al. 2007), analyzed by Tracer (Rambaut et al. 2003) and viewed in FigTree (Rambaut 2007).


Divergence Times

When the Maclura fossil was placed at the crown node of the genus, the estimated age of the genus was 137 [94,185] mya (95% Highest Posterior Density (HPD) (Table 2). Additionally, the divergence time of section Cudrania was 36 [24,47] mya.

Table 2. Divergence times of Maclura and other groups within Moraceae. An asterisk indicates that the node was not calibrated by a fossil and was instead estimated by BEAST.

Phylogenetic Tree

An analysis within BEAST estimated posterior values for the species and sections of Maclura. The genus Maclura is monophyletic as well as all of its sections, except for Section Cardiogyne. This is because M. africana is included in a clade with Section Plecospermum. Within Section Cudrania, one sample of M. cochichinensis is polyphyletic, although it is supported by a low posterior value of 0.4451 (Figure 2).

Figure 2. Phylogenetic tree of Maclura with labeled posterior support values. The branches are colored according to the section to which the species belong. The genus Maclura and all sections except section Cardiogyne (blue) are monophyletic.


Divergence Times of Old and New World Taxa

New World taxa form a basal grade relative to the monophyletic Old World taxa. This New World and Old World split is significant because it provides for opportunities to propose biogeographic hypotheses based on vicariance due to continental movements in the past. Specifically, it was found that M. tinctoria formed the most basal clade, and its most recent common ancestor (MRCA) diverged approximately 62 [47, 79] mya (Table 2). The finding that the South American species, M. tinctoria, forms the most basal clade within Maclura has important implications for its origins because it provides additional evidence that Maclura originated in the New World, specifically South America. Additionally, M. tinctoria has cystoliths, an ancestral character in Moraceae, which also points to a South American origin for Maclura.

On the other hand, the approach that assumes that the region with the greatest species diversity is the region of origin predicts an Old World origin for Maclura. This approach, while informative, has issues. First, a species could follow a speciation process where dispersal occurs before radiation. In this case, the region of the least species diversity would be the origin of the species in question. Second, even if one assumes that radiation occurred before dispersal, then there could be extinctions after the original radiation, reducing the extant species diversity in the regions with extinction events. Thus, one would be unable to conclude that the region of greatest species diversity is the origin of the plant in question.

Biogeography of Maclura

Divergence dates are not only important for estimating the dates of origin of a species or genus, but also serve as invaluable tools in testing biogeographical hypotheses and explaining extant distributions of species. This is because divergence dates help rule out certain geological vicariance events. The two different placements of the Maclura fossil lead to different divergence time estimates and thus different biogeographical hypotheses. Both results are discussed below.

Calibration with Maclura fossil placed at Maclura crown node

When the Maclura fossil is placed at the crown of Maclura, a few migratory paths of Maclura are eliminated. For example, at 62 mya, the approximate date for the divergence of Maclura, Africa would have long been separated from South America, so a direct land connection from South America to Africa could not explain Maclura’s origin in South America and subsequent radiation into the Old World. Instead, another explanation must be invoked to understand the mechanisms that may have allowed Maclura to disperse from the New to the Old World.

The Eocene North Atlantic Landbridge Connection

The Eocene North Atlantic land connection, which featured North America and western Eurasia in close enough proximity to facilitate the exchange of biological organisms across land and water, has been thought to be “possible” by some researchers (Raven and Axelrod, 1974; Wolfe, 1975). However, much more attention has been given to the Beringian land connection, which was the land connection that existed between Asia and North America at various times since the beginning of the Cenozoic period, which began 66 mya and continues into the present (Tiffney, 1985). Evidence has been presented by some researchers that suggests that the Eocene North Atlantic land connection is much more important than previously thought. The evidence for these claims comes primarily from the evident paleofloristic link between North American and Eurasian plant species that existed during the Eocene (Tiffney, 1985). Other researchers, however, have suggested that these same Eocene faunal similarities were instead due to migration via trans-Asiatic migration routes such as the Beringian land connection (McKenna, 1972).

PHOTO | Ron Blakey, Professor of Geology, Northern Arizona University.

Figure 3. A proposed biogeographical hypothesis of Maclura given a divergence estimates based on the placement of a Maclura fossil at the crown node of the genus Maclura. The yellow X indicates the area of origin for the genus Maclura and the red X indicates the Eocene Cudrania fossil found in France. This map represents earth’s geology at about 65 mya, before Africa collided with Eurasia at about 60 mya. The dates on the graph indicate divergence date estimates. See text for more detailed explanation of dispersal route as associated with changing positions of the continents.

This analysis offers evidence that Maclura may have reached Eurasia through a North Atlantic land connection as opposed to other routes. This conclusion is supported in part from the placement of M. africana in the phylogenetic tree (Fig. 2). In the most current delimitation of Maclura outlined by Berg (1986), M. africana is in section Cardiogyne, a South American clade. In this analysis, however, M. africana is very strongly supported in a clade including M. andamanica and M. spinosa, which themselves form section Plecospermum and constitute an Old World clade. This is an important distinction because M. africana is the only African species of Maclura, and this more distant relationship between M. africana and M. brasiliensis (a South American species) makes it less likely that Maclura made its way to Eurasia directly via a South American-Africa land connection. The existence of the Eocene North Atlantic land connection, on the other hand, provides an alternative explanation for the placement of M. africana in section Chlorophora instead of section Cardiogyne, as the next section will outline.

A biogeographic explanation based on placing the Maclura fossil at the Maclura crown node is as follows. First, Maclura originated in South America, since M. tinctoria (native to regions from Bolivia to Argentina) was found to be the most basal species in the genus. Second, Maclura dispersed into North America across islands in close proximity between North and South America. Evidence for this trajectory is twofold: extant M. pomifera are native to regions in North America, and the clade containing M. pomifera is the second most basal clade in Maclura. Third, during the Eocene, Maclura dispersed across the North Atlantic landbridge into Eurasia. Lastly, the lineage of Maclura that dispersed into Eurasia diverged with one lineage dispersing into Africa and the other into Eurasia. The migration into Africa is supported not only by the placement of M. africana in the phylogenetic tree, but also because Africa collided with Eurasia about 60 mya, shortly before the Eocene, making for a straightforward and simple passage of Maclura into Africa from Eurasia (McLoughlin, 2001). The migration into eastern Eurasia is supported by fossil evidence. Known Maclura fossils seem to track an eastward movement. In order from west to east, these fossils are found in epochs including: the Eocene (56 to 34 mya) of France (fossil used in this analysis), the Miocene (23 to 5.3 mya) of Bulgaria, and the Pliocene (5.3 to 2.6 mya) of China (Martínez-Cabrera and Cervallos-Ferriz 2006). A diagram of Maclura’s biogeographical history under the assumption of a Maclura crown fossil calibration is given in Figure 3.


Maclura continues to be an intriguing genus within Moraceae for a variety of reasons. Unlike most Moraceae lineages that originated in the paleotropics (Zerega 2005), Maclura appears to have its origins in the neotropics. The Maclura phylogenies and divergence date estimations presented here were based on only a few fossils and provided enough information for a single biogeographic hypothesis. Due to limited fossil data, however, other biogeographic hypotheses, such as migration to Asia through the Beringian land connection were unable to be definitively ruled out. Nevertheless, this analysis provides the most complete phylogenetic analysis of Maclura to date and can serve as the framework for future studies on the Moraceae family.

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Paya Sarraf


Berg C. 1986. The delimitation and subdivision of the genus Maclura (Moraceae). Proc. K. Ned. Akad. van Wet. Ser. C, Biol. Med. Sci. 89:241–247.

Berg, C.C. 2001. Flora Neotropica Monograph 83: Moreae, Artocarpeae, Dorstenia (Moraceae). With introductions to the family and Ficus and with additions and corrections to Flora Neotropica Monograph 7: 1-346.

Clement W.L., Weiblen G.D. 2009. Morphological Evolution in the Mulberry Family (Moraceae). Syst. Bot. 34:530–552.

Cohen, K.M., Finney, S., Gibbard, P.L. (2013), International Chronostratigraphic Chart, International Commission on Stratigraphy.

Corner E.J.H. 1962. The Classification of Moraceae. Gard. Bull. 19:187–252.

Cullina, William. Native Trees, Shrubs, & Vines: A Guide to Using, Growing, and Propagating North American Woody Plants. Boston: Houghton Mifflin, 2002.

Dupéron-Laudoueneix, M., 1980. Présence d' un bois fossile de Moraceae dans l'Éocène de la Charente. 105e Congrès national des Sociétés Savantes, Caen. Sciences, Fasc., vol. I, pp. 117–129.

Jauron, Richard. "Facts and Myths Associated with "Hedge Apples"". Iowa State University. Retrieved 27 March 2014.

Judd, W. et al. 2007. Plant Systematics: A Phylogenetic Approach. Sinauer Associates, Sunderland, Massachusetts, USA.

Maddison W.P., Maddison D.R. 2011. Mesquite: a modular system for Evolutionary Analysis.

Magallón, S., Sanderson, M.J., 2001. Absolute diversiWcation rates inangiosperm clades. Evolution 55, 1762–1780.

Martínez-Cabrera, H.I., Cevallos-Ferriz, S.R.S., 2006. Maclura (Moraceae) wood from the Miocene of the Baja California Peninsula, Mexico: Fossil and biogeographic history of its closer allies. Review of Palaeobotany and Palynology 140 (2006) 113–122.

McLoughlin, S., 2001. The breakup history of Gondwana and its impact on pre-Cenozoic Xoristic provincialism. Aust. J. Bot. 49, 271–300.

McKenna, M.C., 1972. Was Europe directly connected to North America prior to the Middle Eocene? Evolutionary Biology. 6, 179-189.

Raven, P.H., Axelrod, D.I. 1974. Angiosperm biogeography and past continental movements. Ann. Missouri Bot. Gard. 61: 539-673.

Scarborough, J; Nutton, V (1982). "The Preface of Dioscorides' Materia Medica: introduction, translation, and commentary". Transactions & studies of the College of Physicians of Philadelphia 4 (3): 187–227.

Shaw, J. et al. 2005. The tortoise and the hare II: relative utility of 21 noncoding chloroplast dna sequences for phylogenetic analysis. American Journal of Botany 92(1): 142–166

Shaw, J. et al. 2007. Comparison of whole chloroplast genome sequences to choose noncoding regions for phylogenetic studies in angiosperms: the tortoise and the hare III. American Journal of Botany 94(3): 275-288.

Sloan, L. C., 1994: Equable climates during the early Eocene: Significance of regional paleogeography for North American climate. Geology, 22, 881-884.

Strand, A.E. et al. 1997. Nuclear DNA-based markers for plant evolutionary biology. Molecular Ecology 6: 113-118.

Tiffney, B.H., 1985. Perspectives on the origin of the Xoristic similarity between eastern Asia and eastern North America. J. Arnold Arboretum 66, 73–94.

White, T. J., T. Bruns, S. Lee, and J. W. Taylor. 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Pp. 315-322 In: PCR Protocols: A Guide to Methods and Applications, eds. Innis, M. A., D. H. Gelfand, J. J. Sninsky, and T. J. White. Academic Press, Inc., San Diego, CA.

Wolfe, J.A., 1975. Some aspects of plant geography of the Northern Hemisphere during the Late Cretaceous and Tertiary. Ann. Missouri Bot. Gard. 62: 264-279.

Zerega N.J.C., Clement W.L., Datwyler S.L., Weiblen G.D. 2005. Biogeography and divergence times in the mulberry family (Moraceae). Mol. Phylogenet. Evol. 37:402–16.

Zerega N.J.C., Nur Supardi M.N., Motley T.J. 2010. Phylogeny and Recircumscription of Artocarpeae (Moraceae) with a Focus on Artocarpus. Syst. Bot. 35:766–782.