Euglena: 2013 Evolutionary Divergence of Monodelphis domestica and Myrmecopius fasciatus through Complete Mitochondrial Genome Analysis Kelsey Hermick1, Gabrielle Van Nest1, Michael Terwilliger1, Shannon Wood2, and Victoria Le- gere1. Department of Biology1 and Ecology2, Susquehanna University, Selinsgrove, PA 17870. Abstract Marsupials are characterized by the presence of a marsupium; however, two marsupial species lack a fully developed marsupium. Myrmecopius fasciatus has a protective flap of skin and Monodelphis domestica does not possess a protective pouch. Through the analysis of the marsu- pium, location of mammae, number of mammae, epipubic bone, and the structure of the pseudo- vaginal canal, we classified Myrmecopius fasciatus and Monodelphis domestica to be no more closely related to the outgroup than any other marsupial. Using MEGA5, Maximum Parsimony and Maximum Likelihood trees were developed to show evolutionary placement of the 27 taxa of interest using complete mitochondrial genome sequences. We interpret our analyses to indicate that the absence of a marsupium is a derived character state for both Myrmecopius fasciatus and Monodelphis domestica. The purpose of this paper is to analyze five morphological characters of the reproductive system in marsupials by phylogenetic analysis, specifically as they relate to Myrmecopius fasciatus and Monodelphis domestica. Please cite this article as: Hermick, K., G. Van Nest, M. Terwilliger, S. Wood, and T. Legere. 2013. Evolutionary divergence of Monodelphis domestica and Myrmecopius fasciatus through complete mitochondrial genome analysis. Euglena. doi:/euglena. 1(1): 17-25. Introduction teats differ among several marsupial species (Tyn- Marsupials are an infraclass of Mammalia whose dale-Biscoe 1987). The mammary glands lactate a distinguishing character is the presence of a marsu- nutritious supply of milk to the teats, and the young pium, a protective pouch used to carry underdevel- stay latched onto the teat for several weeks, or in oped offspring (Tyndale-Biscoe 2005). Marsupials some cases months. After the young are attached to give birth to poorly developed fetuses (4-5 weeks the teat, they consume the nutritious milk (Tyndale- old) that then crawl from the birth canal to the mar- Biscoe 2005). Whether the mammae are concealed supium, where they live until they reach maturity by the marsupium or exposed are character states of (Dawson et al.1989). However, while Myrmecopius marsupials. The mammae are often concealed within fasciatus and Monodelphis domestica each lack a the marsupium; however, some mammae are epider- fully developed marsupium, Monodelphis domestica mally exposed. completely lacks a pouch. (Wesierska and Turlejski The epipubic bone is a common morphology 2000). According to Cooper et al. (2003), Myrme- among marsupials, although there are some species copius fasciatus is still considered a marsupial be- that do not exhibit this characteristic, for example, cause the pelt acts as an underdeveloped pouch Notoryctes typhlops (Szalay 1994). The epipubic (Cooper et al. 2003). The purpose of this paper is to bone, or “marsupial bone”, is found in modern mar- analyze five morphological characters of the repro- supials, and other mammals, and functions as a sup- ductive system in marsupials, specifically as they port for the marsupium (Kielan-Jaworowska relate to Myrmecopius fasciatus and Monodelphis 1975). Because this bone is also found in other domestica. mammals, Kielan-Jaworowska (1975) thought that The offspring latch on to a mamma of its mother the original purpose of the epipubic bone was not to until the young are developed enough to function support the marsupium, but to aid in locomotion without full dependence (Tyndale-Biscoe 2005). (Kielan-Jaworowska 1975). Marsupials do not possess the placenta that all other Female marsupials possess two ovaries, two uteri mammals, excluding montrema, possess (Nelson and two vaginal canals (Dawson et al. 1989). In ad- 1978). Therefore, the presence of the marsupium is dition, marsupials possess the character of a third necessary for the young to grow to maturity (Dawson vaginal canal called the pseudovaginal canal that is et al. 1989). The location of mammae and number of specifically for giving birth. Progesterone is a hor- 17 Euglena: 2013 mone secreted from the corpus luteum within the mammals. One major feature is that five tRNA genes uterus that prepares marsupials for parturition by around the origin of the light strand replication are softening the tissues of the pseudovaginal canal rearranged. Another is the anticodon of tRNAasp (Tyndale-Biscoe 2005). The pseudovaginal canal can which is post transcriptionally changed by an RNA either be permanent, meaning that after birth the ca- editing process, altering the coding capac- nal remains open, or transitional, meaning after birth, ity. tRNAasp codes for the anticodon GCC in mar- the tissues within the canal fuse and there is no indi- supials versus that of the GTC anticodon that monot- cation that a canal ever existed (Sweet 1907). remes and placentals code for. The mammalian mi- Marsupials today are found only on three conti- tochondrial genome codes for 22 tRNAs, 2 rRNAs, nents; however, fossil records show that at one point, and 13 proteins. This alteration likely contributed to marsupials were prevalent across the the evolutionary divergence of marsupials from globe. Continental drift during the Oligocene Era (34 eutherians (Janke et al. 1994). to 23 million years before the present) tore the conti- A single morphological character is not suffi- nents apart, and today, 235 species of marsupial can cient in distinguishing between two taxa. Monodel- be found in Australia and 99 species can be found in phis domestica and Myrmecopius fasciatus do not the Americas (Nelson 1978). Throughout the world, possess a physical marsupium; however, other char- marsupials could not compete with other placental acters that these species possess are synapomorphic mammals and began to die out, but in South America with those of species among the marsupial infraclass. and especially Australia, the marsupials had no major Therefore, Monodelphis domestica and Myrmecopius predators and little competition for survival, so they fasciatus both lack a marsupium, but are still classi- were able to flourish and evolve into many forms fied under marsupials because of their reproductive (Nelson 1978). structures and gestation periods (Cooper et al. 2003; There are two major features that distinguish the and Wesierska and Turlejski 2000). mitochondrial genome of marsupials from other Table 1: Taxa selected were identified by scientific name, order, and common name. Taxa were used for phyloge- netic analysis. The authority for each taxa is also included. There are seven extant orders being investigated from the marsupial infraclass and one order from the placental mammals, Cannis lupus. Complete mitochondrial genome sequences were collected from accession numbers through NCBI. (NCBI 2013) (Redlist 2012) Accession Taxa: Scientific Name Order Common Name Authority Number Lagostrophus fasciatus Diprotodontia Banded Hare-wallaby Péron & Lesueur, 1807 NC_008447 Macropus robustus Diprotodontia Common Wallaroo Gould, 1841 NC_001794 Lagorchestes hirsutus Diprotodontia Rufous Hare-wallaby Péron & Lesueur, 1807 NC_008136 Petaurus breviceps Diprotodontia Sugar Glider Waterhouse, 1838 NC_008135 Vombatus ursinus Diprotodontia Common Wombat Shaw, 1800 NC_003322 Phalanger vestitus Diprotodontia Stein's Cuscus Milne-Edwards, 1877 NC_008137 Potorous tridactylus Diprotodontia Long-Nosed Potoroo Kerr, 1792 NC_006524 Tarsipes rostratus Diprotodontia Honey Possum Gervais & Verreaux, 1842 NC_006518 Trichosurus vulpecula Diprotodontia Common Brushtail Possum Kerr, 1792 NC_003039 Phascolarctos cinereus Diprotodontia Koala Goldfuss, 1817 NC_008133 Pseudocheirus peregrinus Diprotodontia Common Ring-Tail possum Boddaert, 1785 NC_006519 Monodelphis domestica Didelphimorphia Gray Short-Tailed Opossum Wagner, 1842 NC_006299 Metachirus nudicaudatus Didelphimorphia Brown Four-Eyed Opossum E. Geottroy, 1854 NC_006516 delphis Virginiana Didelphimorphia Virginia Opossum Kerr, 1792 NC_001610 Myrmecobius fasciatus Dasyuromorphia Numbat Waterhouse, 1836 NC_011949 Thylacinus cynocephalus Dasyuromorphia Tasmanian tiger Harris, 1808 NC_011944 Dasyurus hallucatus Dasyuromorphia Northern Quoll Gould, 1842 NC_007630 Sminthopsis crassicau- Dasyuromorphia Fat-Tailed Dunnart Gould, 1844 NC_007631 data Sminthopsis douglasi Dasyuromorphia Julia Creek Dunnart Archer, 1979 NC_006517 Sarcophilus harrisii Dasyuromorphia Tasmanian Devil Boitard, 1841 NC_018788 Macrotis lagotis Peramelemorphia Greater Bilby Reid, 1837 NC_006520 Isoodon macrourus Peramelemorphia Northern Brown bandicoot Gould, 1842 NC_002746 Perameles gunnii Peramelemorphia Eastern Barred Bandicoot Gray, 1838 NC_006521 Caenolestes fuliginosus Paucituberculata Dusky Shrew Opossum Tomes, 1863 NC_005828 Canis lupus dingo Carnivora Dingo Meyer, 1793 NC_008092 Dromiciops gliroides Microbiotheria Monito del Monte Thomas, 1894 NC_005826 Notoryctes typhlops Notoryctemorphia Southern Marsupial Mole Stirling, 1889 NC_006522 18 Euglena: 2013 Materials and Methods Maximum Parsimony (MP), Figure 1-2, trees were The sequences collected from each taxon were run. mitochondrial complete genome (Table 1). The se- To construct Figures 1 and 2, MEGA 5 was used quences were acquired from the NCBI (National with a bootstrap of 1,000 replications (Tamura et. al Center for Biotechnology