DOI: 10.5061/dryad.2v6wwpznh
title: Data Matrices Nukukammophis

description: Reassessment of the anatomy of a three-dimensionally preserved, partial
 braincase from the Lower Cretaceous (lower Valanginian; ~139 mya) Kirkwood
 Formation of the Kirkwood Cliffs Quarry, South Africa, reveals it to be
 that of a new fossil snake species. The braincase presents a number of
 squamate skull features but also specific braincase features shared with
 early snakes such as Najash, Dinilysia, Sanajeh, and Menarana (e.g.,
 presence of a large fenestra ovalis; fenestra rotunda separated from
 fenestra ovalis by prominent crista interfenestralis; reduced basal tubera
 on basioccipital). Phylogenetic relationships were analyzed using two
 distinct data sets and under different optimality criteria, recovering
 this new snake as an early-evolving lineage with close affinities to other
 Cretaceous Gondwanan snakes including Najash, Dinilysia, and other typical
 madtsoiids. This three-dimensionally preserved braincase reveals important
 aspects of early snake cranial evolution that were previously unknown from
 the oldest known fossil snakes. This specimen extends the fossil record of
 Gondwanan snakes by at least 45 my, making it the oldest known African and
 Gondwanan fossil snake and supporting a much broader palaeobiogeographical
 distribution of early snakes across ancient Gondwanan continents during
 the Early Cretaceous.
description: (a) Phylogenetic data
 sets Two phylogenetic data sets were
 used to investigate the relationships of the new fossil snake with other
 snake and non-snake lizards. Data Set 1 is a recent snake phylogeny data
 set [3] with a number of updates made herein. For this analysis,
 we included only fossil taxa that preserve associated cranial
 materials; thus, chimaeric taxa such as <i>Coniophis </i>[22],
 or taxa lacking cranial remains of any kind such as <i>Xiaophis
 </i>[13], were excluded. Similarly, <i>Nanowana </i>was
 only scored for skull characters as the type or paratype do not include
 associated vertebrae [18,19]. In total, 35 taxa comprising early-evolving
 snakes (17 extinct and 18 extant) were scored for 247 osteological
 characters (see electronic supplementary materials). Two versions of Data
 Set 1 were used: one consisting of the complete data set and one excluding
 character 217 (absence/presence of parazygantral foramen) in order to test
 the effect of that character on retrieving, or not, a monophyletic
 Madtsoiidae. The presence of parazygantral foramina has been considered a
 strong synapomorphy of the Madtsoiidae [19,23–25]; however, their presence
 is not restricted to madtsoiids and they can also be absent in taxa
 considered closely related (e.g., <i>Dinilysia</i>),
 indicating that this character is variably distributed, possibly within a
 more inclusive clade than just the conventional Madtsoiidae
 [26]. Data Set 2 is a much broader early
 diapsid and squamate data set [12] that includes recently added fossil and
 extant snake taxa [3], and thus differs from the original matrix [12] (see
 electronic supplementary materials). The use of this data set is intended
 to investigate the placement of snakes among other squamates, and their
 divergence times, based on morphological and molecular
 data. (b) Maximum parsimony analyses
 (MP) Analyses
 were conducted in TNT 1.5-beta [27]. For Data Set 1, all heuristic
 searches were performed under equal weights and consisted of 1000 rounds
 of random addition sequence (RAS) of taxa followed by Tree Bisection
 Reconnection (TBR) branch swapping, holding ten trees per replication and
 collapsing branches of zero length after tree search. The resulting trees
 were subjected to a final round of TBR branch swapping and optimal
 topologies were rooted with <i>Varanus</i> (see electronic
 Supplementary Materials). Node support was calculated by the Bremer
 support and under absolute frequencies by jackknifing (p = 0.36, 1000
 pseudo-replicates). For Data Set 2, we utilized all New Technology Search
 algorithms available in TNT, following the same procedures as in
 [12]. (c)
 Non-clock Bayesian inference (BI) analyses Non-clock BI analyses were
 conducted for Data Set 1 and Data Set 2 using Mr. Bayes v. 3.2.7a [28] via
 the CIPRES Science Gateway v.3.3 [29]. For Data Set 2, as there were no
 changes to the molecular data set previously used, molecular partitions
 and models of evolution were the same as in [3]. The morphological
 partition was analyzed with the Mkv model [30]. (d) Time-calibrated relaxed-clock
 Bayesian inference analyses (TED-BI) For Data Set 2, we implemented
 “total-evidence-dating” using the fossilized birth-death (FBD) tree model
 allowing for sampled ancestors, under a relaxed-clock model in Mr. Bayes
 v.3.2.6 [31–34], following the same procedures as in [3,12]. We chose to
 use the Independent Gamma Rate (IGR) relaxed-clock model [31–34]. The base
 of the clock rate was given an informative prior based on the previous
 non-clock analysis: the median value for tree height in substitutions from
 posterior trees divided by the age of the tree based on the median of the
 distribution for the root prior: 29.4493/325.45 = 0.09048, in natural log
 scale = -2.402626. We chose to use the exponent of the mean to provide a
 broad standard deviation:
 <i>e</i><sup>0.09048</sup><i> </i>=
 1.094699. Sampling strategy was set to diversity, which is more
 appropriate when extant taxa are sampled in a way to maximizes diversity
 (as performed herein) and fossils are sampled randomly [31,32]. The vast
 majority of our calibrations were based on tip-dating, which accounts for
 the uncertainty in the placement of fossil taxa and avoids the issue of
 bound estimates for node-based age calibrations [31]. The fossil ages used
 for tip-dating correspond to the uniform prior distributions on the age
 range of the stratigraphic occurrence of the fossils (available in
 electronic supplementary materials). For the clades for which we lacked
 some of the oldest known fossils in our analysis, and for which there is
 overwhelming support in the literature (and in all our other analyses)
 regarding their monophyly, and for which the age of the oldest known
 fossil is well-established, we employed node age calibrations with a soft
 maximum age calibration (available in electronic supplementary materials).
 The age of the root was set with a soft lower bound to the same age as in
 [3,12]. Initial runs
 of Data Set 2 found two taxa, <i>Paliguana </i>and
 <i>Pamelina</i>, to behave as wildcards, as also previously
 detected in previous studies [12,35,36]; we therefore removed those two
 taxa for the final relaxed clock BI analysis reported here.
 For all
 BI analyses, convergence of independent runs was assessed using the
 average standard deviation of split frequencies (ASDSF ≈ 0.01), potential
 scale reduction factor (PSRF ≈ 1 for all parameters), and effective sample
 size (ESS &gt; 200 for each parameter). All input and output files
 (summary trees, log files, etc.) from all analyses (including Mr. Bayes
 blocks with all prior and analytical parameters) are available online as
 electronic supplementary materials.