DOI: 10.5061/dryad.3n5tb2rg7
title: Character set and phylogenetic analyses of the living and fossil egerniine scincids of Australia

description: The diverse living Australian lizard fauna contrasts greatly with their
 limited Oligo-Miocene fossil record. New Oligo-Miocene fossil vertebrates
 from the Namba Formation (south of Lake Frome, South Australia) were
 uncovered from multiple expeditions from 2007–2018. Abundant
 disarticulated material of small vertebrates was concentrated in shallow
 lenses along the palaeo-lake edges, now exposed on the western shore. The
 fossiliferous lens occurring within the Namba Formation, also known from
 Billeroo Creek 2 km northeast of Lake Pinpa, includes abundant aquatic
 (such as fish, platypus Obdurodon, and waterfowl) and diverse terrestrial
 (such as possums, dasyuromorphs, and scincids) vertebrates and is
 hereafter recognised as the Fish Lens. The stratigraphic provenance of
 these deposits in relation to prior finds in the area is also established.
 A new egerniine scincid taxon Proegernia mikebulli sp. nov. described
 herein, is based on a near-complete reconstructed mandible, maxilla,
 premaxilla, and pterygoid. Postcranial scincid elements were also
 recovered with this material, but could not yet be confidently associated
 with P. mikebulli. This new taxon is recovered as the sister species to P.
 palankarinnensis, in a tip-dated total-evidence phylogenetic analysis,
 where both are recovered as stem Australian egerniines. These taxa also
 help pinpoint the timing of the arrival of scincids to Australia, with
 egerniines the first radiation to reach the continent.
description: In order to better understand the timing of the Australian
 colonisation by the Egerniinae, both molecular and morphological data
 (including fossils) are required to generate tip-dated phylogenies.
 Undated parsimony and tip-dated Bayesian analyses infer, respectively, the
 phylogeny with the least homoplasy, and the most probable dated
 phylogeny. Morphological characters
                 Morphological characters used in the following
 analyses consisted of 102 discrete and 48 continuous traits, forming an
 expanded matrix from Thorn et al. (Thorn, Hutchinson et al. 2019). The
 expanded character list is included. Continuous characters, derived from
 the measurements of the individual bones or teeth from the dentaries and
 maxillae, were taken from either Micro-CT scan data in Avizo Lite (v. 9.0)
 or SEM at Flinders Microscopy, to the nearest micrometre, or with digital
 callipers to the nearest ten micrometres. All measurements were converted
 to ratios of either dentary or maxilla length to standardise for size.
 Continuous character states were linearly scaled to values spanning 0–2 to
 replicate the mean number of discrete character states (three), for
 analyses in both TNT (Goloboff and Catalano 2016) and BEAST 1.8.3
 (Drummond, Suchard et al. 2012), so that they do not have a
 disproportionate weight. Molecular partitions
                 Molecular data sourced from Tonini et al.
 (Tonini, Beard et al. 2016) and Gardner et al. (Gardner, Hugall et al.
 2008) were analysed using Partition Finder 2 (Lanfear, Frandsen et al.
 2016) to find optimal partitions and substitution models [15]. The same
 six molecular (gene) partitions, 12s (412 base pairs [bps]), 16s (681
 bps), ND4 (693 bps), BDNF (699 bps), CMOS (835 bps) and B-fibrinogen (1051
 alignable bps) and substitution models chosen in that study (Thorn,
 Hutchinson et al. 2019) are used again here. Maximum
 Parsimony                 The parsimony analyses for
 the combined discrete morphological, continuous morphological, and
 molecular data were performed using TNT v.1.5 (Goloboff and Catalano
 2016). Eutropis multifasciata was set as the most distant outgroup
 following the phylogenetic interpretations of Gardner et al. (Gardner,
 Hugall et al. 2008) and Thorn et al. (Thorn, Hutchinson et al. 2019). The
 most parsimonious tree (MPT) for the combined data was found using 1000
 replicates of tree-bisection-reconnection (TBR) with up to 1000000 trees
 held. To assess clade support, 200 partitioned
 bootstrap replicates (with discrete characters, continuous characters, and
 each gene locus treated as a separate resampling partition), were
 performed using TNT, using new search methods (XMULT) with 1000 replicates
 and 1000000 trees held. The MPT and bootstrap trees from TNT were exported
 in nexus format, and continuous and discrete characters were traced (in
 Mesquite; Maddison and Maddison 2017). The executable files for finding
 the Most Parsimonious tree, and for performing 200 reps of Partitioned
 Bootstrap resamples can be found in the SI data files
 Namba_Egerniines_Topology.tnt (MPT file) and
 Namba_Egerniines_PartitionedBootstrap.tnt. Bayesian
 analysis                 The discrete and continuous
 morphological data, and molecular data were simultaneously analysed in
 BEAST v1.8.4 using tip-dated Bayesian approaches (Drummond, Suchard et al.
 2012). Eutropis multifasciata was again set as the furthest outgroup.
 Polymorphic discrete morphological data were treated exactly as coded
 rather than as unknown, i.e. if coded as states (0,1) it was treated as 0
 or 1, but not 2. The discrete character set was analysed using the
 Mkv-model with correction for non-sampling of constant characters (Lewis
 2001, Alekseyenko, Lee et al. 2008). Despite recent disputes over the
 effectiveness of this model (Goloboff, Pittman et al. 2018), it is
 well-tested (Wright and Hillis 2014, O'Reilly, Puttick et al. 2016)
 and is still widely accepted and applied to morphological data (Harmon
 2019). Continuous characters, transformed to span values between 0 and 2,
 were analysed with the Brownian motion model. Bayes factors were used to
 test the need to accommodate among-character rate variability for both
 discrete and continuous morphological characters (i.e. gamma
 parameter).                 The stratigraphic data used
 for tip-dating analyses were derived from fossil taxa and their associated
 stratigraphy noted in Table 1. No node age constraints were imposed in
 this analysis, all dates are retrieved from the morphological and
 stratigraphic age ranges from the noted fossil taxa (tips). The most
 appropriate available model in BEAST v.1.8.4, birth-death serial sampling
 (Stadler 2010), was applied. An uncorrelated relaxed clock  (Drummond, Ho
 et al. 2006) was separately applied to the molecular and morphological
 data.                 Each Bayesian analysis was run
 for 100,000,000 generations with a burn-in of 20%. The analysis was
 conducted four times to confirm stationarity. The post-burnin samples of
 all four runs were examined in Tracer 1.7.1 (Rambaut, Drummond et al.
 2018) to ensure convergence was achieved. All four runs were combined in
 LogCombiner, and the consensus tree produced by TreeAnnotator (Drummond,
 Suchard et al. 2012). The executable .xml file for BEAST, all output log
 files, and the final consensus tree file (.tree) are available as
 supplementary information. Alekseyenko, A. V., C. J.
 Lee and M. A. Suchard (2008). "Wagner and Dollo: a stochastic duet by
 composing two parsimonious solos." Systematic Biology 57(5):
 772–784. Drummond, A. J., S. Y. W. Ho, M. J. Phillips
 and A. Rambaut (2006). "Relaxed phylogenetics and dating with
 confidence." PLOS Biology 4(5): e88. Drummond, A.
 J., M. A. Suchard, D. Xie and A. Rambaut (2012). "Bayesian
 phylogenetics with BEAUti and the BEAST 1.7." Molecular Biology and
 Evolution 29(8): 1969–1973. Gardner, M. G., A. F.
 Hugall, S. C. Donnellan, M. N. Hutchinson and R. Foster (2008).
 "Molecular systematics of social skinks: phylogeny and taxonomy of
 the Egernia group (Reptilia: Scincidae)." Zoological Journal of the
 Linnean Society 154(4): 781–794. Goloboff, P. A. and S.
 A. Catalano (2016). "TNT version 1.5, including a full implementation
 of phylogenetic morphometrics." Cladistics 32(3): 221–238.
 Goloboff, P. A., M. Pittman, D. Pol and X. Xu (2018).
 "Morphological data sets fit a common mechanism much more poorly than
 DNA sequences and call into question the Mkv model." Systematic
 Biology 68(3): 494–504. Harmon, L. J. (2019).
 Phylogenetic comparative methods. Published Online, Luke Harmon.
 Lanfear, R., P. B. Frandsen, A. M. Wright, T. Senfeld and B.
 Calcott (2016). "PartitionFinder 2: new methods for selecting
 partitioned models of evolution for molecular and morphological
 phylogenetic analyses." Molecular Biology and Evolution 34(3):
 772–773. Lewis, P. O. (2001). "A likelihood
 approach to estimating phylogeny from discrete morphological character
 data." Systematic Biology 50(6): 913–925.
 Maddison, W. and D. Maddison (2017). Mesquite: a modular system
 for evolutionary analysis. Version 3.2. O'Reilly,
 J. E., M. N. Puttick, L. Parry, A. R. Tanner, J. E. Tarver, J. Fleming, D.
 Pisani and P. C. Donoghue (2016). "Bayesian methods outperform
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 and M. A. Suchard (2018). "Posterior summarisation in Bayesian
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description: See README_NambaEgerniinesRunFiles for detailed
 information. Phylo files are named
 accordingly: Run file:
 Project_numberofcharacters_numberoftaxa_numberofruns.file
 Consensus
 trees: Project_numberofcharacters_numberoftaxa_CONSENSUS.tree
 Log
 files: Project_numberofcharacters_numberoftaxa_clockmodels_calibrations.log The file 'Namba_matrices_for_Mesquite.txt' can be opened in the freely available Mesquite software (Maddison and Maddison, 2017), and allows the morphological characters to be traced across the MP tree. The Excel spreadsheet containing the continuous data (Namba_continuous_data.xlsx) includes the xml codes required to convert the morphological data set for import into xml file for analyses in BEAST. Maddison, W. and D. Maddison (2017). Mesquite: a modular system for evolutionary analysis. Version 3.2.