Institutional abbreviations

AMNH, American Museum of Natural History, New York; DPC, Division of Fossil Primates, Duke Lemur Center, Duke University; BMNH, Natural History Museum, London; KNM/NMK, National Museums of Kenya, Nairobi, Kenya; MCZ, Museum of Comparative Zoology, Harvard University, Cambridge; MNHN, Muséum National d’Histoire Naturelle, Paris; RRBP, Rukwa Rift Basin Project (identifier used by the Tanzanian Antiquities Unit), Dar es Salaam, Tanzania.

Geological context

The type specimen was excavated from the Nsungwe 2 locality in the late Oligocene Songwe Member of the Nsungwe Formation in the Rukwa Rift Basin of southwestern Tanzania (Fig 1). The Nsungwe Formation represents a continental rift-fill sequence containing several fossil bearing localities in horizons interpreted as sheet flood deposits within a small, flashy discharge fluvial system that appears to have drained into a local wetland [27, 30]. Nsungwe 2 is one of the richest localities in the Nsungwe Formation, and is dated to ~25.2 Ma based on biostratigraphy, dated ash beds, and detrital zircon geochronology [27, 30, 37]. Sedimentological data and the presence of aquatic and semi-aquatic taxa (e.g., fishes, frogs, and aquatic invertebrates), suggest habitats that experienced seasonal or periodic climatic fluctuations, with perennially available water sources [27, 30, 38, 39].

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Fig 1. Geological context of Nsungwe 2 locality.

Nsungwe 2 (yellow star in all portions of the figure) is a fossiliferous locality in the Songwe Member of the Nsungwe Formation in the Rukwa Rift Basin of southwestern Tanzania. A, The context of the Rukwa Rift Basin in Africa and B, in Tanzania. C, Geological map of the Nsungwe Formation and surrounding outcrop. D, Stratigraphic column of the fossiliferous section of the Songwe Member of the Nsungwe Formation with magnetostratigraphic correlations mapped through the section. Radiometric date of 25.2 Ma inferred from U-Pb zircon dating (see [27, 37]). Ms, Mudstone; Ss, Sandstone; Cs, Conglomerate; Striped, Volcanic tuff; VGP lat, virtual geomagnetic pole latitude; Black, normal polarity; White; reverse polarity.

https://doi.org/10.1371/journal.pone.0185301.g001

Nomenclatural acts

The electronic edition of this article conforms to the requirements of the amended International Code of Zoological Nomenclature, and hence the new names contained herein are available under that Code from the electronic edition of this article. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix "http://zoobank.org/". The LSID for this publication is: urn:lsid:zoobank.org:pub:B03A9CC0-B057-42AB-A7A0-DEDC3BBF16F0. The electronic edition of this work was published in a journal with an ISSN, and has been archived and is available from the following digital repositories: PubMed Central, LOCKSS.

Permissions

All necessary permissions were obtained to undertake the described study, which complied with all relevant regulations. Fieldwork was conducted under permits issued by the Tanzanian Commission for Science and Technology (COSTECH), the Tanzania Antiquities Unit, and the Tanzanian Division of Immigration.

Specimen preparation, measurement, and body mass estimation

After mechanical preparation by K. Whitman at the Ohio University Fossil Preparation and Imaging Facility, the specimen was micro-CT scanned in the Duke MicroCT lab in the Shared Materials Instrumentation Facility at Duke University, Raleigh, NC on a Nikon XTH 225 ST scanner. A digital model of the specimen is available as a 3D PDF in the supplementary materials for this study and a PLY digital model of the specimen is available for viewing and download in Project 303 on MorphoSource, a repository for 3D scan data supported by the NSF < http://www.morphosource.org/Detail/SpecimenDetail/Show/specimen_id/8140>. The specimen was scanned with a voxel size of 0.02003 mm in each dimension, at a voltage of 114 kV and amperage of 127 μa. The digital model was constructed and visualized using volume rendering and isosurface rendering in Avizo 8.0.

Dental nomenclature used in this study follows Holroyd [40] and Borths et al. [29]. Specimen measurements from the new taxon were collected using hand calipers and ImageJ [41].

Body mass for the new taxon was estimated using equations developed for this study and by Van Valkenburgh [42] as detailed in S1 Appendix. We recognize mammalian dentition is not a perfect proxy for body mass [43], but it is possible to derive an estimate to ease communication about the new taxon using Van Valkenburgh’s [42] methods.

Phylogenetic analysis

A phylogenetic analysis was conducted to place the Nsungwe hyaenodont in a larger systematic context within hyaenodont evolution. Of interest in this analysis are the relationships between the Nsungwe taxon and hyaenodonts from the early Oligocene and early Miocene of Afro-Arabia (e.g., [29, 44]). The Nsungwe hyaenodont is relatively incomplete, hence phylogenetic hypotheses presented here should be interpreted with caution until additional materials become available.

A character taxon matrix, modified from Borths and Seiffert [44], included 81 OTUs and 143 characters including nine characters either modified from Bastl et al. [35] or new to this study to capture the morphology of dP³ and dP⁴. The character matrix is S1 Dataset and a list of characters, their states, and their sources is available as S2 Appendix. The character-taxon matrix was assembled in Mesquite [45]. Eighteen multistate characters were treated as ordered, using outgroup morphology as a point of reference for character order [46]. All characters were treated as equally weighted. Additional information is included in S3 Appendix on all OTUs, including their age, formation, locality, and specimens used to score the character-taxon matrix.

For the analysis of the character-taxon matrix, we employed a model-based, Bayesian approach, a phylogenetic method applied in many recent studies of phylogenetic relationships based on morphological character information [29, 47–55]. In this case, we use “tip-dating” Bayesian inference, which incorporates the character-taxon matrix, estimated age ranges for each OTU, and a clock model to simultaneously estimate the most likely topology, divergence times, and the rate of evolution for each lineage. This allows us not only to explore topologies that are likely based on the character-taxon matrix, but also to search for topologies that are likely, given the estimated rates of evolution implied by the fossil record. This powerful analytical method is particularly useful for understanding hyaenodont evolution, as the earliest fossil evidence of this clade is a highly specialized taxon, Lahimia [56], from the middle Paleocene. In contrast with results obtained using Bayesian tip-dating approaches, some parsimony analyses recover Lahimia in a somewhat unlikely position, deeply nested within Hyaenodonta [29, 57], implying many extensive ghost lineages in the early history of the clade.

The analysis was conducted in MrBayes 3.2.6 [58] and the MrBayes formatted nexus file is available as S2 Dataset. The stratigraphic age and the sources of the estimated ages for each OTU are included in S3 Appendix. The M_k likelihood model was used to model morphological evolution and an independent gamma rates (IGR) relaxed clock model [58, 59] was used to estimate divergence dates and infer evolutionary rates within the tree. Hyaenodonta was constrained as an ingroup with Tinerhodon, Altacreodus magnus, and Maelestes designated as part of the outgroup. A prior of 100–75 Ma was used to constrain the root to be consistent with estimates of the age of Eutheria [60]. A prior of 75–62 Ma was set for the origin of Hyaenodonta, bracketing the estimates for the emergence of crown placental mammals and the earliest known hyaenodont [60, 61]. The precision of the date associated with the Nsungwe 2 locality allows us to fix the age of the Nsungwe hyaenodont at 25 Ma [27]. All other members of the in-group were assigned age ranges based on literature review. The analysis was run for 20,000,000 generations, and performed with two runs simultaneously with four Markov chains, three of which were heated (temp = 0.02), sampling every 1000^th generation with the first 25% discarded as part of the burn-in period. The remaining generations are incorporated into the summary statistics for the analysis. After the analysis was completed, convergence was assessed using effective sample size and average standard deviation of split frequencies statistics from the final generation. The results are summarized using an “allcompat” tree with the discussion of evolutionary rates based on median rate estimates as suggested Beck and Lee [49].

Biogeographic analysis

Hyaenodonts are the only known carnivorous mammals occupying Afro-Arabia during the Paleogene, but they are not limited to that landmass. Hyaenodonts are also found on the northern landmasses of Asia, North America, and Europe and multiple studies have discussed close phylogenetic connections among hyaenodonts from the northern continents and Afro-Arabia [62–66]. Borths et al. [29] specifically illustrated multiple likely dispersals of hyainailourine hyaenodonts (the clade that includes Pterodon and Megistotherium) from Afro-Arabia to Europe, North America, and Asia and two possible dispersals during the Oligocene from Europe to Afro-Arabia. With the new morphological and temporal data provided by the Nsungwe taxon, we performed a biogeographic analysis with the “allcompat” consensus tree to determine how the new taxon may affect our understanding of the complicated biogeographic history of Hyaenodonta.

We used Bayesian Binary MCMC (BBM) to reconstruct the likely origins of each clade given the distribution of the taxa included in the analysis and the branch lengths implied by the phylogenetic analysis. Each OTU was assigned to one of four continental areas: Afro-Arabia, Asia, Europe, or North America. The two hyaenodont OTUs known from India–Paratritemnodon and Indohyaenodon–were placed in the “Asia” continental land area for the purposes of this analysis which is focused on exchange between Afro-Arabia and the northern continents.

The biogeographic analysis was conducted in RASP version 3.2 [67]. We hypothesize that dispersal is a more likely mechanism to explain the distribution of hyaenodonts during the Cenozoic, so the number of continental areas from which a clade could disperse was limited to one in the analysis. The MCMC analysis ran for 1,000,000 generations with 10 Markov chains sampled every 100 generations and all chains set with a temperature of 0.1. The first 100 trees were discarded as part of the burn-in period and the Jukes-Cantor model was implemented with equal among-site variation.

Ecomorphological reconstruction

A study of Afro-Arabian carnivore diversity through the Paleogene and early Neogene was conducted to place the Nsungwe hyaenodont in a temporal and ecological context with other Afro-Arabian carnivores. In extant mammalian carnivores, the ratio of trigonid to total molar length on the lower carnassial can be used to infer diet [68, 69]. The trigonid is the portion of the tooth that bears the slicing carnassial blades. The talonid is a separate a portion of the molar that often forms a basin that occludes with the protocone. Food is ground between the protocone and talonid like a dental mortar and pestle. Carnivores that are hypercarnivores, acquiring 70% or more of their calories from meat, have a high trigonid ratio, with most of the mesiodistal length of the molar occupied by elongate carnassial blades. In contrast, generalist carnivores exhibit a lower trigonid ratio (in other words, the slicing trigonid is reduced relative to total molar length). This relatively longer grinding talonid reflects the fact that generalists acquire only 50% to 60% of their calories from meat and supplement the rest of their diet with plant matter and invertebrates [68, 70].

In this study, we calculate the trigonid ratio in all Paleogene (hyaenodonts) and early Miocene (hyaenodonts + carnivorans) carnivores. This allows us to explore the changing dietary diversity of hyaenodonts and carnivorans through the Paleogene-Neogene transition. Prionogale and Namasector, two taxa referred to the enigmatic lineage Prionogalidae from the early Miocene of Afro-Arabia, are also included in the analysis. In addition to trigonid ratio, we also collected a proxy for body mass for all carnivore taxa, using the mean mesiodistal length of M₁ as a proxy for carnivoran body mass and mean mesiodistal length of M₂ as a proxy for hyaenodont body mass. M₂ is used as a proxy for body mass in hyaenodonts because it is more functionally and developmentally homologous with the carnivoran M₁ than the often heavily worn hyaenodont M₁ [71, 72]. Specimens measured in this study are listed in S3 Appendix. Measurements were collected using ImageJ [41].

Relevant portions of the lower dentition are not known for all hyaenodonts (e.g., Koholia, Pterodon syrtos, Metapterodon kaiseri). In a series of correlation studies (detailed in S1 Appendix), we demonstrate the significant correlation between measurements of the upper carnassial to the lower carnassial. The Nsungwe specimen preserves only where dP₄ and M₁ would have occluded. In order to provide a preliminary assessment for the Nsungwe taxon to be tested upon recovery of more complete materials, we estimate M₁ size for RRBP 09088 based on the size of the alveoli of dP⁴ and M¹. Regression equations used to calculate the crown measurements of dP⁴ and M¹ are presented in S1 Appendix. We then used the regression equations calculated based on a large sample of carnassial-bearing carnivorous mammals to reconstruct dimensions of M₁. Using M₁ to reconstruct the body mass of the Nsungwe taxon likely underestimates the size of the animal, but is a best effort to reconstruct the body mass of the only carnivorous mammal known from the late Oligocene of Afro-Arabia. This body size estimate can help provide a backdrop for understanding the meat-eating fauna encountered by the first Afro-Arabian carnivorans that dispersed to the landmass from Eurasia.

Systematic paleontology

Systematic hierarchy

Placentalia Owen, 1837 [73]

Hyaenodonta Van Valen, 1967 [74]

Hyainailouroidea Borths, Holroyd & Seiffert, 2016 [29]

Genus Pakakali Borths & Stevens, gen. nov.

urn:lsid:zoobank.org:act:991B6709-BFFA-430C-AD11-C603C4F56E9B

Pakakali rukwaensis Borths & Stevens, sp. nov.

urn:lsid:zoobank.org:act:A16065CB-5BC1-4441-AA12-11E79D43FDB8

Fig 2

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Fig 2. Pakakali rukwaensis holotype (RRBP 09088).

Rostrum fragment from the right maxilla of Pakakali rukwaensis discovered at Nsungwe 2 locality, Nsungwe Formation, late Oligocene (~25.2 Ma) with dP³, alveoli of dP⁴ (or P⁴) and M¹ in (A) occlusal view, (B) occlusal-lingual view, (C) buccal (lateral) view, (D) lingual (medial) view, (E) dorsal view, (F) mesial (rostral) view, (G) distal (caudal) view. Digital model of the specimen is available as S4 Appendix and as Project 303 at Morphosource.

https://doi.org/10.1371/journal.pone.0185301.g002

Description

Specimen RRBP 09088 is a portion of the right maxilla and palatine bearing a complete dP³ and the alveoli of dP⁴ and M¹. A digital model of the specimen is available as S4 Appendix and is accessioned as part of Project 303 at morphosource.org. The rostral margin of the specimen contains the infraorbital foramen dorsal to the mesial root of dP³, a feature commonly observed in hyaenodonts. The medial edge of the specimen is fractured along the maxillo-palatine suture just lingual to the alveolus of M¹. A portion of the right palatine is preserved lingual to the protocone alveolus of dP⁴ and the protocone of dP³. The palatal process of the maxilla is embayed between the protocone of dP³ and the protocone alveolus of dP⁴ and between the protocone alveolus of dP⁴ and the protocone alveolus of M¹, providing space for the occluding trigonid of dP₄ and M₁. The maxillary process of the specimen is fractured caudal to the infraorbital foramen. The maxillary process was folded medially postmortem and it is preserved lying superior to the infraorbital canal, obscuring the maxillary recess.

Although germs of P³ and P⁴ are not visible on micro-CT scans of the specimen, the tooth crown on the Nsungwe specimen is consistent in morphology with dP³s preserved in other hyaenodont specimens. It has three distinct roots and is mesiodistally elongate (mesiodistal length = 7.0 mm; more than 1.5 times its buccolingual width = 4.2 mm). The elongate parastyle comprises over 20% of the entire mesiodistal length of dP³. The parastyle is buccolingually compressed into a blade-like crista that meets the preparacrista at a defined notch. The preparacrista slopes distally at a 45-degree angle to the apex of the paracone. Like the parastyle, the paracone is buccolingually compressed. The postparacrista slopes distally at a steeper angle than the preparacrista. The postparacrista meets a distinct metacone that is fused to the distal margin of the paracone. The metacone is compressed buccolingually into a blade-like cusp. The postmetacrista slopes distally to form a deep notch at the junction with the metastyle. The metastyle, like the parastyle and paracone, is buccolingually compressed into an elongate blade. The mesiodistal length of the metastyle (2.0 mm) accounts for nearly 30% of the total mesiodistal length of the tooth. The lingual margin of the metastyle is sheer with a distal wear facet that indicates the sharp upper carnassial of dP³ sliced past the preprotocristid and postparacristid of the occluding dP₄. The buccal margin of metastyle slopes to the buccal cingulum more gently than the metastylar blade slopes to the lingual margin of occlusion. A shallow basin is formed between the metastyle, buccal cingulum and the paracone. The buccal cingulum is narrow, but distinct, connecting the buccal metastyle to the buccal parastyle. Lingually, the metastyle is not connected to the protocone. Rather, the postprotocrista abuts the base of the paracone. The protocone projects lingually 1.3 mm from the base of the paracone. The distinct protocone forms the lingual-most point of a shallow talon basin that is mesially open along the preprotocrista. The preprotocrista traces the lingual-mesial face of the paracone to the lingual base of the parastyle. In occlusal view, the elongate parastyle, paracone, and metastyle are perpendicular to the lingual projection of the protocone.

The crown of dP⁴ (or possibly) P⁴ is not preserved, but the alveoli indicate the tooth had three distinct roots. The mesial root of dP⁴ is very close to the metastyle of dP³, indicating the metastyle of dP³ and parastyle of dP⁴ would have been closely packed. The lingual root of dP⁴ projects as far lingually as the lingual protocone of dP³. Unlike the protocone of dP³, the orientation of the lingual root of dP⁴ indicates the cusp was mesial relative to the paracone. The diameter of the alveolus for the lingual root of dP⁴ is equal to the diameter of the mesial root, indicating the protocone would have been a large, distinct cusp. The alveoli of M¹ also indicate the lingual alveolus held a large, deeply anchored root for the protocone. The diameter of the lingual alveolus of M¹ is nearly equal to the diameter of the distal alveolus. The mesial and distal alveoli of M¹ are aligned along the buccal margin of the maxilla and the lingual alveolus is more mesially oriented than the lingual alveolus of dP⁴. The lingual alveolus of M¹ is almost directly medial to the mesial root of M¹.

The mesiodistal distance from the distal protocone alveolus of M¹ to the distal protocone of dP⁴ is 7.91 mm. This measurement was used to infer the length of M₁ with the hyaenodont carnassial regression equation presented in S1 Appendix. The estimated length of M₁ is approximately 8.3 mm, yielding an estimated body mass of 5.8–10.1 kg with Van Valkenburgh’s [42] body mass equations as detailed in the S1 Appendix.

Hyaenodont comparisons

Few upper deciduous teeth of hyaenodonts have been formally described, though recent efforts [35] have presented specimens that can be compared directly with Pakakali. As noted by Zack [75], in hyaenodonts, the morphology of a deciduous premolar closely resembles the morphology of the permanent tooth in the next position distally. Hence, dP³ morphology in this taxon would be expected to more closely approximate that of a permanent P⁴ than that of a permanent P³. As such, we compare Pakakali to the more restricted record of hyaenodont dP³s, and also to a broader sample of Afro-Arabian hyaenodont P⁴s.

Bastl, Nagel, and Peigné [35] presented one of the only comparative descriptions of the “milk tooth” or deciduous dentition of several hyaenodonts. Their study focused on species in the genus Hyaenodon, a genus phylogenetically distant from Afro-Arabian hyaenodonts [29, 57]. Despite the phylogenetic distance, dP³ in Hyaenodon shares several basic features with Pakakali. Like Pakakali, the infraorbital foramen of Hyaenodon is dorsal to the roots of dP³. Further, like Pakakali, the dP³ of Hyaenodon has a distinct metacone that is lower than the paracone and the protocone is mesiodistally narrow and projects lingually and nearly perpendicular to the buccal margin of the tooth. The amount the protocone projects lingually varies among Hyaenodon species, but in all dP³ specimens identified by Bastl and colleagues, and in Pakakali, the parastyle is mesiodistally elongate compared to the parastyle of P³ and P⁴, giving dP³ a “T”-like morphology in occlusal view.

Among hyainailourines, dP³ is known in Leakitherium, Pterodon dasyuroides, and Paroxyaena (Fig 3). These three taxa are larger than Pakakali, and the dP³ in each is more distally inclined with the preparacrista sloping steeply back toward the metastyle. Protocone morphology differs among taxa, with the protocone of Pterodon and Leakitherium mesiodistally wide, and the protocone of Paroxyaena and Pakakali mesiodistally narrow. Among these hyainailourines, the metacone of Pakakali is the smallest and most fully fused to the paracone. Wear along the metacone may partially explain this morphology though the base of the metacone is wider and more prominent in Leakitherium, Paroxyaena, and Pterodon than it is in Pakakali, evidence that the metacone of Pakakali, even when unworn, was likely closely appressed to the paracone.

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Fig 3. Pakakali dP³ compared with dP³ of other hyainailouroids.

(A) Digital model of Pakakali rukwaensis right rostral fragment (RRBP 09088) with dP³ in (subscript 1) occlusal view (subscript 2) and buccal (lateral) view with measurements used to estimate the size of M₁ overlaid. Comparative specimens scaled to same size as Pakakali with individual scale bars between the occlusal and buccal view of each specimen: (B) Paroxyaena pavlovi (images of cast of GGM no. Ca-300 and (C) Pterodon dasyuroides (BSPG 1879 XV 642), a hyainailourines from the late Eocene of Europe; (D) Masrasector nananubis (DPC 20882), a teratodontine from the late Eocene of Afro-Arabia and (E) Leakitherium hiwegi (KNM-RU 2949), a hyainailourine from the early Miocene of Afro-Arabia. Note variation in the metacone, paracone, and parastyle in each specimen.

https://doi.org/10.1371/journal.pone.0185301.g003

Apterodontinae is the sister clade to Hyainailourinae. Like Leakitherium and Pterodon, the protocone of dP³ in Apterodon is mesiodistally wide and the talon basin shallow and not a narrow cusp like the protocone of dP³ of Pakakali. The metacone of dP³ in Apterodon is mesiodistally elongate, its base prominent, and its apex distinct from the paracone, all features that depart from the reduced, blade-like metacone of Pakakali.

Closer in size to Pakakali are several teratodontines known from dP³ including Masrasector, Metasinopa, Dissopsalis and Anasinopa. Metasinopa is closest in size to Pakakali. Like Pakakali, the parastyle and metastyle of dP³ in Metasinopa is gracile, though the protocone is relatively wider in Metasinopa, and the metacone is a lower, mesiodistally wider cusp than it is in Pakakali. Dissopsalis, like Metasinopa, has a mesiodistally elongate metacone with a prominent base. Dissopsalis further differs from Pakakali by having a relatively short metastyle that does not form a deep carnassial notch with the metacone. Anasinopa is close to the same size as Pakakali but has a mesiodistally wider protocone, a mesiodistally shorter parastyle and metastyle, and a lower paracone. Of the teratodontines known from dP³, Masrasector has the most gracile and Pakakali-like protocone. Masrasector and Pakakali also share a deep carnassial notch formed between the metastyle and metacone. Masrasector differs from Pakakali by having a paracone that rises significantly higher than the metacone and does not incline distally. Instead, the paracone of Masrasector forms a tall, acute triangle in buccal view. The paracone of Pakakali is lower than the paracone of Masrasector with a wider angle formed by the apex of the paracone.

The deciduous dentition of Teratodon, a taxon comparable in size to Pakakali, has not been described, but the morphology of Teratodon’s P⁴, a massive, block-like structure with difficult to discern cusps, suggests dP³ was not as gracile as the dP³ of Pakakali. Further, the distal alveolus of M¹ in Pakakali supported a mesiodistally extensive metastyle, which contrasts with the mesiodistally short, blunt M¹ metastyle of Teratodon.

The deciduous dentition of Metapterodon kaiseri, a hyaenodont from the early Miocene of Namibia and Kenya, is also not known, though the morphology of dP³ in Pakakali can be compared to the mid-sized hypercarnivorous Metapterodon P⁴. The dP³ of Pakakali has a long parastyle compared to the P⁴ of Metapterodon and has a buccolingually narrow metastyle. In buccal view, the paracone on dP³ in Pakakali is mesiodistally long, forming an equilateral triangle. This differs from the P⁴ paracone of Metapterodon, which has a narrower base and mesial and distal margins that meet at a more acute angle, like the apex formed by the dP³ paracone of Masrasector.

Two other genera of comparable size to Pakakali are known from the early Miocene, though neither have had deciduous upper dentition referred to them: Mlanyama and Isohyaenodon. The holotype of Mlanyama has thick, crenulated enamel and closely packed premolars that differ from the thin, smooth enamel of Pakakali. Further, the deciduous premolars of Mlanyama were likely more robust structures, more similar to dP³ of Leakitherium than the dP³ of Pakakali. Isohyaenodon is a complex genus, with at least three species: Isohyaenodon andrewsi, Isohyaenodon matthewi, and Isohyaenodon pilgrimi. Both I. andrewsi and I. matthewi are in similar size ranges as Pakakali. Based on measurements and observations, we support the hypothesis of Holroyd [40] that I. andrewsi is synonymous with Metapterodon kaiseri, or at least part of the same genus (though the Metapterodon OTU constructed for this analysis does not include material referred to I. andrewsi). Isohyaenodon matthewi is known from fragmentary rostral and dentary material. KNM-SO 8527 is a maxilla fragment with a portion of M¹ preserved. The protocone of I. matthewi is reduced and the root of the protocone is close to the parastylar root. This differs from the lingually projecting M¹ protocone alveolus of Pakakali. Further, the talonid of M₂ in I. matthewi is reduced to single cusp, an unlikely morphology to occlude with the more expansive protocones of Pakakali. Isohyaenodon pilgrimi is smaller than Pakakali and I. matthewi, but shares with I. matthewi reduced protocones, elongate metastyles, and reduced talonids, all morphology that is not consistent with the molar morphology inferred from Pakakali.

Carnivoran comparisons

Both the fossil record and molecular divergence estimates indicate carnivorans may be present in Afro-Arabia in the late Oligocene and may be preserved as part of the Nsungwe fauna [31, 32]. The oldest fossil evidence of Carnivora in Afro-Arabia is Mioprionodon hodopeus from the Nakwai locality in Kenya, a site that is likely early Miocene in age [76]. Molecular divergence estimates of Madagascar’s endemic carnivoran clade, Eupleridae, suggest that lineage diverged from Herpestidae about 25.5 million years ago [31]. The ancestors of euplerids have been suggested to have dispersed from Africa to Madagascar [31], indirect evidence that Carnivora may have been present in Africa at least 25.5 million years ago.

The single carnassial complex in Carnivora forms between dP³ and dP₄ in immature carnivorans and P⁴ and M₁ in mature carnivorans [8]. In all early Miocene carnivorans, including Mioprionodon, Kanuites, Legetetia, Africanictis, and Stenoplesictis, the mesiodistal length of P⁴ and M₁ is greater than the mesiodistal length of M¹ and M₂. In each of these taxa, the trigonid of M₁ is taller than the trigonid of M₂ and the mesiodistal distance between the protocone of P⁴ and M₁, the occlusal space for the trigonid of M₁, is longer than the mesiodistal distance between the protocone of M¹ and M², the occlusal space for the trigonid of M₂. Also in each of these carnivorans, the tall trigonid of M₁ occludes into a deep embayment in the palate that forms distal to the short, mesially oriented protocone of P₄. Further, the buccal margin of the maxilla distal to P⁴ is medially inflected in these carnivorans, the protocone root of M¹ is lingually rather than mesially oriented, and the M¹ parastyle and metastyle have similar mesiodistal lengths. In contrast to the early Miocene carnivorans of Afro-Arabia, the mesiodistal length of dP⁴ (~7.3 mm based on the alveolus to crown regressions presented in S1 Appendix) of Pakakali is shorter than M¹ (7.85 mm: see S1 Appendix). Further, in Pakakali, it appears based on alveolar morphology that the protocones of the missing dP⁴ and M¹ did not frame a space wider than that framed by the protocones of M¹ and M², indicating the trigonid of M₁ in Pakakali was not significantly larger than the trigonid of M₂. In Miocene Afro-Arabian carnivorans, the trigonid of M₂ is mesiodistally shorter and lower than the trigonid of M₁. The palate of Pakakali is concave between the protocone alveoli of dP⁴ and M¹ and distal to the protocone alveolus of M¹ (where M₂ would have occluded), further evidence that the trigonids of M₁ and M₂ were both tall and prominent and both were accommodated by the palate, a morphological condition not found on the palate of early Miocene carnivorans. Distal to dP⁴, the buccal margin of the maxilla continues the lateral trend of the maxilla, the protocone alveolus of M¹ was mesially positioned relative to the parastyle alveolus, and the M¹ metastyle alveolus is larger than the parastyle alveolus, features consistent with a hyaenodont maxilla.

Phylogenetic results

Pakakali rukwaensis is recovered within Hyainailouroidea, a clade that includes all Afro-Arabian hyaenodont taxa included in the analysis except Koholia, Lahimia, and Boualitomus. The relationship of Pakakali to other Afro-Arabian hyaenodonts is shown in Fig 4, which illustrates the portion of the “allcompat” (majority rule plus compatible groups) tree relevant to the taxonomy of the species at the center of this study and statistics relevant to major Afro-Arabian clades are shown in Table 1. The “allcompat” tree with all OTUs incorporated into the analysis is S1 Fig. and statistics for all branches in the analysis are part of S1 Table. The consensus tree file output from MrBayes is S3 Dataset and the PSTAT file with overall analysis statistics is S4 Dataset. Note that the nodes and tips in these trees are illustrated at the median age of origin. All mean dates in S1 Table are offset by 11.56 Ma and all median dates are offset by 11.37 Ma. In this analysis, the overall mean clock rate is 0.0095 change/Ma and the median clock rate is 0.0059 change/Ma.

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Fig 4. Phylogeny and biogeography of Afro-Arabian Hyaenodonta.

The portion of the “allcompat” Bayesian consensus tree containing all Afro-Arabian hyaenodonts included in the Bayesian phylogenetic analysis with BBM biogeographic results summarized in circle graphs over each node. The number in the center of the circle is the posterior probability for each node. The rectangles to the left of the OTU name represent the estimated age of each OTU based on a literature review. Taxa that span a long geological interval reflect either substantial specimen sampling or imprecise dates for localities, see S3 Appendix for details of each OTU. The black vertical line in each OTU rectangle is the median estimated age of the OTU based on the Bayesian analysis. The bar for Pakakali is narrow because the age of the Nsungwe 2 is known with great precision. The proportion of the circle filled by each color reflects the probability that the clade originated from the corresponding continental area (green, Afro-Arabia; purple, Asia; yellow, Europe; red, North America; grey (on map), continents without hyaenodonts). Branches are colored with the most likely origin for each clade and gradients indicate branches that are likely dispersal events. Pakakali is nested within Hyainailouroidea and Teratodontinae, both clades that most likely originated in Afro-Arabia. The light red vertical bar illustrates the likely interval when Carnivora dispersed to Afro-Arabia and it crosses the hyaenodont lineages that were likely extant on the continent when carnivorans arrived. S1 Fig. and S1 Table contain information on the expanded phylogeny of Hyaenodonta.

https://doi.org/10.1371/journal.pone.0185301.g004

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Table 1. Statistics related to major Afro-Arabian hyaenodont clades.

https://doi.org/10.1371/journal.pone.0185301.t001

Presently, the analysis is limited by the fragmentary nature of the specimen, but provides a starting point for interpreting the phylogenetic position of Pakakali in the context of broader hyaenodont relationships examined by Borths et al. [29]. Pakakali is here recovered within Hyainailouroidea, a clade defined by the node that represents the common ancestor of Teratodon and Hyainailouros. Specifically, Pakakali is weakly resolved as an early branching node in Teratodontinae, a clade of hyaenodonts that is Afro-Arabian in origin and predominately radiated on that continent. Teratodontinae includes taxa that are mesocarnivorous (e. g. Masrasector nananubis, Glibzegdouia tabelbalaensis) and a few taxa that are hypercarnivorous (e.g. Dissopsalis). The clade is estimated to have originated ~57.6 Ma. Furodon, an Eocene teratodontine from Algeria, is the sister taxon of all other teratodontines. After the divergence of the Furodon lineage, the Pakakali lineage diverges from the rest of Teratodontinae in the early Eocene (~52 Ma). The branch that unites Pakakali with all later diverging hyainailourines is weakly supported (22% posterior probability support). Three early Miocene teratodontines were part of the phylogenetic analysis: Teratodon, Anasinopa, and Dissopsalis pyroclasticus. Teratodon is weakly supported (40% posterior probability) as the sister taxon of the strongly supported clade Anasinopa + Dissopsalis (92% posterior probability).

The sister clade of Teratodontinae includes Hyainailouridae, a moderately supported clade (58% posterior probability) that includes Afro-Arabian, European, Asian, and North American taxa. Hyainailouridae likely originated between ~60.19 Ma and ~46.45 Ma. Within Hyainailouridae are the robustly supported clades Apterodontinae (98% posterior probability) and Hyainailourinae (58% posterior probability). Hyainailourinae includes six early Miocene hyainailourines: Metapterodon, weakly supported (27% posterior probability) as the sister taxon of a clade of Afro-Arabian OTUs that includes early Oligocene, Afro-Arabian “Pterodon”, and early Miocene Isohyaenodon, Hyainailouros, Leakitherium, Megistotherium, and Mlanyama, a taxon from the early Miocene that has not previously been incorporated into a phylogenetic analysis.

Biogeographic results

Results of the Bayesian binary MCMC biogeographic analysis of hyaenodonts most closely related to Pakakali are depicted in Fig 4, together each clade’s likelihood of having originated from each of the four designated biogeographic areas (Afro-Arabia, Asia, Europe, or North America) visualized as a proportion of the circle plot over each node. Area of origin results are detailed in S1 Table. Probabilities recovered in biogeographic analyses are visualized in S1 Fig.

Based on this analysis, the clade that includes all Afro-Arabian OTUs most likely reflects a dispersal event from Europe to Afro-Arabia between the Late Cretaceous or early Paleogene. After the dispersal of the common ancestor of Lahimia and Hyainailouros to Afro-Arabia, subsequent dispersals occurred from Afro-Arabia back to the northern continents. In any case, all nodes in Teratodontinae, including the node containing Pakakali, are resolved as likely Afro-Arabian in origin.

The most ambiguous biogeographic dispersal in Hyainailouroidea pertains to the series of nodes near the origin of Hyainailourinae. The most likely interpretation of the biogeographic history of that group is that their common ancestor dispersed from Afro-Arabia to Asia or Europe during the early Eocene. Later, the common ancestors of Akhnatenavus and the common ancestors of the Miocene hyaenodonts independently dispersed from Europe to Afro-Arabia, though future investigations into the phylogenetic relationships of Hyainailourinae may refine this biogeographic scenario.

Phylogenetic and biogeographic position of Pakakali

The Bayesian phylogenetic analysis resolved Pakakali as an early-branching lineage in the clade Teratodontinae, a clade that contains only Afro-Arabian taxa and that most likely originated in Afro-Arabia. Teratodontinae is nested within Hyainailouroidea, a clade that includes all Afro-Arabian hyaenodonts except the oldest hyaenodonts in Afro-Arabia: Lahimia, Boualitomus, and Koholia. Bastl et al. [35] first incorporated deciduous characters into an analysis of hyaenodont systematics with a focus on relationships within the Eurasian and North American taxon Hyaenodon. In this study, we expanded on the efforts of Bastl et al. [35]. Notably, even with the inclusion of these new characters, the relationships among Afro-Arabian OTUs found in this study are similar to the results of Borths et al. [29] and Borths and Seiffert [44]. Pertinent to this analysis, the placement of Pyrocyon/Preregidens as the sister clade to the group that contains all Afro-Arabian OTUs differs among these analyses, though the relationships of Pyrocyon and Preregidens are unstable, hence it is not unexpected that these taxa are weakly supported in this analysis. In summary, inclusion of Pakakali and the addition of characters describing variation in the upper deciduous dentition do not disrupt recent phylogenetic interpretations of relationships of Afro-Arabian hyaenodonts retrieved by earlier studies (e.g., [29, 44]) except by resolving Metapterodon kaiseri as the sister taxon of the clade that includes Fayum “Pterodon” and the Miocene hyainailourines, a position that differs from the results of Borths and Seiffert [44].

The only complete tooth preserved in Pakakali is dP³. As noted by Zack [75], deciduous teeth in carnivorans closely resemble and function in a way homologous to the immediately distal adult tooth. Hence, the dP³ of Pakakali likely offers insights into the morphology of P⁴. Both dP³ and P⁴ are part of the carnassial complex in hyaenodonts with dP³ shearing past dP₄ in sub-adult hyaenodonts and P⁴ shearing past M₁ in sub-adult and mature hyaenodonts. The dP³ of Pakakali bears a mesiodistally extensive metastyle and buccolingually compressed paracone, evidence of a well-developed carnassial shearing complex coupled with a well-developed protocone. Teratodontinae contains many Afro-Arabian taxa with extensive shearing crests and expansive protocones and trigon basins, thus the placement of Pakakali within this clade is consistent with the morphology preserved on the fragmentary holotype. Precise relationships resolved for Pakakali within Teratodontinae should be interpreted with caution until additional anatomical information of the taxon is recovered, particularly the early-branching position of the lineage within both Teratodontinae and Hyainailouroidea. Overall, there are relatively few OTUs in the analysis scored for the dP³ characters. Further, as noted by Bastl et al. [35] and Borths et al. [29], deciduous dental morphology can reveal phylogenetic affinities that may place a taxon known only from deciduous material in an earlier-branching phylogenetic position than might otherwise be resolved with the addition of the complete adult dentition.

Teratodontine and Hyainailourinae contain several lineages that persist across the Paleogene-Neogene boundary, an interval coincident with tectonic and climatic changes in Afro-Arabia. Both clades include lineages that diversify in the early Miocene including Dissopsalis, Mlanyama, Metapterodon, Isohyaenodon, and Hyainailouros. There may be several species of Hyainailouros [9] in the Miocene (though the diversity of the genus is contested, see [24]) and at least one, Hyainailouros sulzeri, dispersed from Afro-Arabia to Europe and southwestern Asia across the “Gomphothere landbridge” [21]. Dissopsalis and Metapterodon also dispersed from Afro-Arabia to southwestern Asia [77, 78] during the Miocene. These dispersal events further emphasize that the extinction of Hyaenodonta was not a simple replacement of the incumbent hyaenodonts by the newly-arrived carnivorans. Rather, Miocene hyaenodonts adapted to the changing landscape and were capable of moving into ecosystems on the northern continents previously dominated by carnivorans during the late Oligocene. Further study of Pakakali and its close relatives will be important for unraveling how biotic and abiotic factors affected the origination and morphological adaptations of hyainailouroids in Afro-Arabia and what biological factors may have enabled hyainailouroids to persist across three continents nearly to the end of the Miocene.

Dental development in Pakakali

The holotype of Pakakali is not a dentally mature individual, as it retains dP³. Discussions of hyaenodont deciduous dentition have predominantly focused on Hyaenodon, a genus known from Asia, Europe, and North America [33–35, 79]. In Hyaenodon the terminal M² is almost fully erupted into occlusion when the last deciduous tooth (dP³) is replaced. In Afro-Arabian hyainailouroids, dP³ is also replaced late in ontogeny. In many carnivoran lineages, dP³ is also the last deciduous tooth replaced [80], though there is variation in the eruption sequence across the clade. Hyaenodonts seem to differ from carnivorans by erupting adult dentition relatively slowly. Most carnivorans entirely replace deciduous teeth within one year [81]. Mellet [79] concluded Hyaenodon replaces deciduous teeth at a much slower rate than do carnivorans, allowing Hyaenodon to wear M₁ and M₂ while dP₄ and dP₃ were still present in the dentary. This hypothesis was tested by Bastl and Nagel [34] with a specimen of North American Hyaenodon; applying a forensic technique exploring cementum annulation rate, they support the hypothesis that Hyaenodon did not fully erupt its adult dentition until it was developmentally older, possibly between three to four years of age. Like Hyaenodon, the deciduous teeth of Pakakali likely did not erupt in a rapid sequence, a conclusion supported by a sample of hyainailouroid specimens that preserve the deciduous dentition. A specimen of the Afro-Arabian hyainailourine Leakitherium (KNM-RU 15182) preserves an unerupted P⁴, yet preserves no evidence of the bud of P³ in the maxilla above dP³. Brychotherium ephalmos, a teratodontine from the late Eocene of Egypt, is represented by a specimen (DPC 17627) that preserves the left dP⁴ along with M¹–M², and an almost fully erupted M³. The right side of the specimen preserves P⁴. This specimen reveals that dP⁴ was still in the process of replacement when M³ was almost fully erupted. Based on comparisons with other hyaenodonts, it is likely dP³ was present in the tooth row after all the molars erupted and P⁴ was replaced.

The dental eruption sequence and the timing of tooth replacement in Pakakali has important implications for the life history of the hyaenodonts that encountered the first Afro-Arabian carnivorans. The carnassial complex in hyaenodonts and carnivorans allows them to slice vertebrate tissues [70]. A functioning carnassial complex requires precise occlusion between the upper carnassial and lower carnassial. In sub-adult carnivorans dP³ and dP₄ form the first carnassial complex, and is lost within the first year of a carnivoran’s life. The adult carnassial complex, formed between P⁴ and M₁, must last the remainder of an individual’s life. Hyaenodonts also form the first carnassial complex between dP³ and dP₄ and the second is formed between dP⁴ and M₁. The late retention of dP³ and dP⁴ allows hyaenodonts to utilize the earliest two carnassials for a longer interval than carnivorans utilize their deciduous carnassial pair. After utilizing the carnassials formed between dP³/dP₄ and dP⁴/M₁, hyaenodonts had up to three additional adult complexes to use throughout life: P⁴/M₁, M¹/M₂, and M²/M₃. A dentally immature hyaenodont utilized its deciduous dentition for an extensive period of its behaviorally mature life, making deciduous dentition part of the adult phenotype with implications for the fitness of hyaenodonts on an evolutionary scale.

Carnivore ontogeny features discrete intervals of a juvenile, immature phase characterized by deciduous dentition rapidly followed by an adult, mature phase characterized by adult dentition. In contrast, it appears that hyaenodont ontogeny was a multi-step process of gradual maturation that allowed hyaenodonts to aggressively wear multiple carnassial pairs, extending the utility of the slicing teeth beyond the juvenile stage. Using the individual dental age stages (IDAS) system proposed by Anders et al. [82], hyaenodonts exhibited a long IDAS 2 phase compared to carnivorans. This may explain why taxa like the European proviverrine Lesmesodon and the early Miocene hyainailourine Leakitherium are apparently overrepresented by juvenile specimens. Prior to this study, only Bastl et al. [35] utilized deciduous dental characters in a phylogenetic analysis of hyaenodonts. Based on our observations of the hyaenodont dental record, we encourage the description of deciduous dentition to further reveal the complex life history and evolution of Hyaenodonta.

Ecomorphology of Afro-Arabian carnivores from the Paleocene through early Miocene with special reference to Pakakali

Results of the size and degree of carnassial specialization in Paleogene and early Miocene hyaenodonts and carnivorans are shown in Fig 5 and listed in Table 2. Specimens measured for this study are listed in S2 Appendix. These calculations aim to place Pakakali in temporal and ecological contexts relative to other Afro-Arabian carnivores. The mesiodistal length of a lower carnassial-bearing molar is used as a proxy for body size and the ratio of the mesiodistal length of the carnassial-baring trigonid is used as a proxy for dietary specialization. Larger trigonid ratios are found in taxa that are more specialized for carnivory. Based on the estimated size of M₁ (8.3 mm), Pakakali is closest in size to several early Miocene Afro-Arabian carnivorans including Herpestes aequatorialis, Leptoplesictis namibiensis, and Herpestides aegypticus and the early Eocene hyaenodont Furodon crocheti and Glibzegdouia tabelbalaensis, and the early Miocene hyaenodonts Teratodon and Metapterodon. The trigonid ratio of Pakakali is 0.65, a ratio similar to several hyaenodonts including early Eocene Boualitomus, late Eocene Brychotherium, early Oligocene Metasinopa, and the early Miocene Teratodon. The estimated trigonid ratio of Pakakali rukwaensis is also similar to several early Miocene carnivorans including the viverrid Ketketictis solida, the herpestids Leptoplesictis rangwai, Leptoplesictis mbitensis, and Legetetia nandii, and the amphicyonid Amphicyon giganteus.

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Fig 5. Afro-Arabian Paleogene and early Miocene carnivore morphospace occupation.

A comparison the trigonid ratio for the hyaenodont M₂ and carnivoran M₁ in Afro-Arabia from the late Eocene through early Miocene. The X-axis is average mesiodistal length (mm) of the molars for each taxon included in the analysis as a proxy for body size. Note the scale is not continuous along the X-axis. The Y-axis is geological time expressed as geological ages and absolute age in millions of years (Ma). The timescale is not proportional in the early Miocene to accommodate the dense taxon sample through this interval. Each hyaenodont taxon included in the phylogenetic analysis is placed at the median age estimated by the tip-dating Bayesian analysis. Carnivorans found in the same localities as hyaenodonts in the analysis are placed in the same temporal range. Some taxa are found at multiple localities and the full age range estimated for each taxon is listed in Table 2 and S2 Appendix. The proportion of the horizontal bar that is colored reflects the proportion of the tooth occupied by the slicing carnassial complex. The proportion of the horizontal bar that is black reflects the proportion of the tooth occupied by the talonid complex. The longer the carnassial complex, the more vertebrate prey was likely incorporated into the diet of the taxon. Orange, Hyaenodonta; Red, Carnivora. Yellow, Prionogalidae. Hyaenodonta: 1, Masrasector nananubis; 2, Brycotherium ephalmos; 3, Akhnatenavus nefertiticyon; 4, “Sinopa” ethiopica; 5, Metapterodon markgrafi; 6, Apterodon langebadreae; 7, Apterodon macrognathus; 8, "Pterodon" phiomensis; 9, "Pterodon" africanus; 10, Akhnatenavus leptognathus; 11, Masrasector ligabuei; 12, Metasinopa fraasi; 13, Metapterodon schlosseri; 14, Masrasector aegypticum; 15, Quasiapterodon minutus; 16, Pterodon syrtos; 17, Teratodon (Meswa Bridge); 18, Mlanyama sugu; 19, Hyainailouros spp.; 20, Isohyaenodon matthewi; 21, Teratodon (Rusinga); 22, Metapterodon kaiseri; 23, Isohyaenodon andrewsi; 24, Buhakia moghraensis; 25, Megistotherium osteothlastes; 26, Isohyaenodon pilgrimi; 27, Anasinopa libyca; 28, Leakitherium hiwegi; 29, Anasinopa leakeyi. Carnivora: 30, Mioprionodon hodopeus; 31, Mioprionodon pickfordi; 32, Legetetia nandii; 33, Leptoplesictis namibiensis; 34, Kenyalutra songhorensis; 35, Ginsburgsmilus napakensis; 36, Cynelos euryodon; 37, Africanictis schmidtkittleri; 38, Stenoplesictis muhoronii; 39, Afrosmilus turkanae; 40, Kichechia zamanae; 41, Leptoplesictis mbitensis; 42, Leptoplesictis rangwai; 43, Leptoplesictis senutae; 44, Herpestides aegypticus; 45, Luogale rusingensis; 46, Namibictis senuti; 47, Diamantofelis ferox; 48, Herpestides aequatorialis; 49, Ketketictis solida; 50, Moghradictis nedjema; 51, Afrocyon burolleti; 52, Africanictis meini; 53, Africanictis hyaenoides; 54, Amphicyon giganteus; 55, Ysengrinia ginsburgi; 56, Orangictis gariepensis; 57, Namafelis minor; 58, Syrtosmilus syrtensis. Prionogalidae: 59, Namasector soriae; 60, Prionogale breviceps.

https://doi.org/10.1371/journal.pone.0185301.g005

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Table 2. Tooth size and carnassialization in Afro-Arabian carnivora and Hyaenodonta.

https://doi.org/10.1371/journal.pone.0185301.t002

The smallest carnivorans in the sample are the herpestid Leptoplesictis rangwai and the viverrid Stenoplesictis muhoronii. The largest carnivorans in the sample are the amphicyonids Afrocyon burolleti from Libya and Ysengrinia ginsburgi from Namibia. The smallest hyaenodonts in the sample are Tinerhodon (if the taxon is confirmed as a hyaenodont with additional fossils) and Boualitomus from the early Eocene and Isohyaenodon pilgrim from the early Miocene. The largest hyaenodonts and carnivores in the sample are Hyainailouros and Megistotherium from the early Miocene with Pterodon africanus from the early Oligocene occupying a slightly smaller size range than the massive early Miocene hyaenodonts.

Body mass estimations for Pakakali (see S1 Appendix for calculations) suggest that this hyaenodont was between 5.8 kg and 10.1 kg, comparable in size to Lynx rufus (bobcat) and Cryptoprocta ferox (fosa). Building upon dietary categories and trigonid ratios generated for extant small-bodied carnivorans described by Friscia et al. [83], the inferred trigonid ratio of Pakakali is similar to the trigonid ratio of many carnivorous-to-omnivorous carnivorans including Ichneumon albicauda (0.64; white-tailed mongoose), a sub-Saharan insectivore to omnivore; Galidia elegans (0.65; ring-tailed mongoose), a Malagasy carnivore; and Urocyon cinereoargenteus (0.61; gray fox), a North American carnivore. Body mass estimates of Pakakali are well below 21.5 kg, a body size range occupied by carnivorans that mostly feed on prey that is 45% or less of their body mass [84].

The Nsungwe Formation is reconstructed as seasonal, wetland ecosystem [27, 30] with aquatic and semi-aquatic vertebrates such as freshwater fish, frogs, and anthracotheres, and arboreal taxa like the primates Nsungwepithecus and Rukwapithecus [37]. The presence of the Pakakali holotype with little evidence of postmortem transport in this depositional context suggests the carnivore could have inhabited this complex environment much like a modern gray fox (Urocyon cinereoargenteus) or ringtail (Bassariscus astutus), pursuing small vertebrate prey across any number of substrate/habitat types.

Paleogene-Neogene carnivore transition in Afro-Arabia

The arrival of Carnivora in Afro-Arabia is documented by fossil evidence from the earliest Miocene of Kenya [76], and potentially by the divergence of Malagasy Eupleridae from the rest of Carnivora based on molecular clock estimates [31]. The full diversity of the endemic Afro-Arabian hyaenodont fauna that preceded the arrival of Carnivora is best observed in the Fayum Depression of Egypt. The Afro-Arabian record of Hyaenodonta in the Paleocene and early to middle Eocene is improving [65, 85], but it is not until the extensive late Eocene deposits from the Fayum (Egypt) that the record reveals a complex hyaenodont fauna with a range of body sizes and dental specializations, from small, mesocarnivorous Masrasector to large, hypercarnivorous “Pterodon” africanus [29, 40]. The arrival of Carnivora in the late Oligocene or early Miocene is a natural ecological experiment in adaptation and niche replacement as the diverse hyaenodont fauna is injected with ecologically similar carnivorans. Examinations of carnivore competition, adaptation, or replacement (i.e. [72, 86, 87]) are feasible because, unlike omnivorous or herbivorous taxa that use soft tissue strategies in conjunction with dental adaptations to break down food, mammalian carnivory is relatively simple to extrapolate purely from teeth, which conveniently form most of the carnivorous mammal fossil record. Vertebrate muscle and bone is a material constant through the Cenozoic and multiple mammalian lineages evolved the same strategies for slicing muscle and breaking bone [70]. Using body size and the degree of carnassial specialization, both recorded in teeth, it is possible to bluntly model the niche occupied by a mammalian carnivore in modern or extinct ecosystems. The ecological niches occupied by carnivorans and hyaenodonts in Afro-Arabia can be approximated by dental size and specialization and possible interaction between carnivore communities can be explored using dental morphology. Unfortunately, the diversity of the hyaenodont fauna first encountered by carnivorans is not directly known. Before the discovery of Pakakali from the Nsungwe Formation, there was a substantial (~8MY) gap in the Afro-Arabian record from the late Rupelian through the Chattian (late Oligocene). The hyaenodont fauna encountered by invading Carnivora was inferred mainly from the presence of multiple hyaenodont lineages that appear to persist from the Paleogene into the Neogene. Pakakali rukwaensis is the first carnivorous hyaenodont described from the late Oligocene gap in Afro-Arabia. Until the recovery of a more complete record of late Oligocene hyaenodonts, this information on the relative size and inferred dental specialization of Pakakali affords a first step in examining adaptive trends in the Afro-Arabian carnivore community as two convergent carnivore groups encountered one another.

Pakakali is a relatively small carnivore (<10 kg) that likely consumed small vertebrate and invertebrate prey [80]. Hyaenodonts with similar body masses and trigonid ratios are known from the Fayum, such as Brychotherium ephalmos [29] and Masrasector aegypticum [88]. Direct comparisons are more difficult to draw between the Pakakali niche and early Miocene hyaenodont niches. Teratodon is close to Pakakali in body size and trigonid ratio and would seem to be an extension of the Pakakali niche into the Miocene if only the molars are examined. The premolars of Teratodon are globular and massive and their bizarre morphology reveal that this taxon was exploiting a very different niche than Pakakali, possibly as a durophagous carnivore [9, 89]. The other early Miocene hyaenodonts in the Pakakali size range, Mlanyama and Metapterodon kaiseri, are hypercarnivores with large carnassials and reduced talonid basins. Smaller Isohyaenodon pilgrimi is a specialized hypercarnivore like Metapterodon, but of the Afro-Arabian hyaenodonts it most similar in size to Boualitomus, a mesocarnivorous taxon from much older Eocene deposits [90]. Among early Miocene hyaenodonts, only Anasinopa has premolars and a trigonid ratio comparable to Pakakali, but at ~20 kg, Anasinopa is a much larger taxon.

Few hyaenodonts in the early Miocene are ecologically comparable to Pakakali, although many early Miocene carnivorans share broadly similar body size and carnassial specializations with Pakakali, including Kichechia zamanae, Leptoplesictis namibiensis, and Herpestides aegypticus. An abundance of small-bodied, mesocarnivorous carnivorans in African early Miocene faunas suggest that carnivorans rapidly dominated this niche soon after dispersal to the continent. Indeed, the oldest Afro-Arabian carnivoran known, Mioprionodon hodopeus [76], was a relatively small carnivore with a well-developed talonid basin. The precise relationships among early Miocene Afro-Arabian carnivorans and the number and timing of dispersal events to Afro-Arabia from the northern continents has not been placed within a larger phylogenetic context of carnivoran evolution, making it difficult to determine whether multiple lineages from Europe and Asia took advantage of small-bodied, mesocarnivorous niche space, or if there was a rapid endemic radiation of Afro-Arabian carnivores in the early Miocene seeded by relatively few dispersal events. Regardless, the early Miocene African carnivore fauna was diverse and populated by many small carnivorans with large talonid basins.

Hyaenodonts appear to have found other niches in the crowded early Miocene by assuming more hypercarnivorous roles. Borths et al. [29] noted a trend towards hypercarnivory among multiple lineages of Miocene hyaenodonts, but it was difficult to determine whether this trend had initiated prior to the arrival of Carnivora, or whether it occurred in response to the appearance of a new lineage of carnivorous mammals. The discovery of Pakakali reveals that small-bodied hyaenodonts with relatively diverse diets persisted up to the very close of the Paleogene. This suggests that the trend towards hypercarnivory may have coincided with the arrival of Carnivora, such that small-bodied mesocarnivorous carnivores helped to propel hyaenodonts toward hypercarnivory. Notably, Hyaenodonta also apparently accommodated the arrival of Carnivora by moving into novel body size niches. Isohyaenodon is one of the smallest meat-eaters from the early Miocene and is also among the most hypercarnivorous. At the other end of the scale, lineages that led to both Megistotherium and Hyainailouros became colossal and hypercarnivorous. Early Miocene amphicyonids like Afrocyon burolleti and Ysengrinia ginsburgi are also gigantic, yet these taxa are mesocarnivores compared to the large hyainailourines.

This model of the early Miocene carnivore community, characterized by hypercarnivorous hyaenodonts and mesocarnivorous carnivorans, is not a complete picture. Hypercarnivorous barbourofelids (e.g., Syrtosmilus, Afrosmilus, and Ginsburgsmilus) and felids (e.g., Diamantofelis, Namafelis) also found niche space in the early Miocene carnivore fauna of Africa. In addition, Morales et al. [23] noted that the reduction of hyaenodont diversity in the early Miocene of Namibia is coincident with the aridification of southwestern Africa. They propose that hyaenodonts were adapted to more densely forested environments than carnivorans and drier, more open localities would be expected to have more carnivorans than hyaenodonts. Future studies are needed to explore continent-wide patterns among environmental variables and carnivore faunal compositions through time.