Redescription of Phymolepis  cuifengshanensis (Antiarcha: Yunnanolepididae) using high-resolution computed tomography and new insights into anatomical details of the endocranium in antiarchs

Material

The specimens of P. cuifengshanensis in this study are housed at the Institute of Vertebrate Paleontology and Paleoanthropology (IVPP), Chinese Academy of Sciences (CAS). The material were all collected from muddy limestone of the Xitun Formation in Cuifengshan, Qujing, Yunnan Province.

The Xishancun, Xitun, Guijiatun and Xujiachong formations (in ascending chronological order) represent the Early Devonian non-marine strata in Qujing District (Liu & Wang, 1973; P’an et al., 1978; Zhu, Wang & Fan, 1994; Liu, Gai & Zhu, 2018). The Xitun Formation consists mainly of grayish-green muddy limestone and mudstone (Fang et al., 1985; Xue, 2012), yielding a rich biota (Cuifengshan Assemblage) characterized by the diversification of sarcopterygians (Chang & Yu, 1981; Chang & Yu, 1984; Zhu, Yu & Janvier, 1999; Zhu, Yu & Ahlberg, 2001; Zhu & Yu, 2002) and primitive antiarchs including Yunnanolepis, Parayunnanolepis, Zhanjilepis, Chuchinolepis and Phymolepis. Other fishes in the Xitun Formation include the endemic agnathans (Liu et al., 2015), arthrodires (Dupret, Zhu & Wang, 2017), actinopterygians (Zhu et al., 2006; Lu et al., 2016), acanthodians and chondrichthyans, the latter two of which are mostly known from microvertebtrate remains (Wang, 1984). The Xitun Formation has been dated as late Lochkovian (c. 410–415 million years ago) with evidence from fossil assemblages (Gao, 1981; Fang et al., 1994; Zhao et al., 2011; Xue, 2012).

CT analysis

V4425.2, which preserved the only head shield material for Phymolepis in addition to the almost complete trunk shield, was CT scanned at the Key Laboratory of Vertebrate Evolution and Human Origins of Chinese Academy of Science, Beijing, using the 225 KV micro-tomography scanner (developed by Institute of High Energy Physics, CAS) with following parameters: 150 kV voltage; 100 mA current; 32.93 µm voxel size. All scans were conducted using a 720°rotation with a step size of 0.5°and an unfiltered aluminium reflection target. A total of 1,440 transmission images were reconstructed in a 2,048 × 2,048 matrix of 1,536 slices. The software Mimics v. 18.0 was applied for the three-dimensional reconstruction (segmentation and rendering).

Phylogenetic analyses

The character matrix of antiarchs herein consists of 42 ingroup taxa, two outgroup taxa (Kujdanowiaspis and Romundina), and 79 morphological characters. The matrix is modified from those of Zhu (1996), Jia, Zhu & Zhao (2010) and Pan et al. (2018), with revised codings and the addition of 10 cranial characters. More details including additional references and character re-formulations are provided in the Supplementary Information.

We performed a traditional search in TNT v 1.5 (Goloboff et al., 2008), using 1,000 random addition sequence replicates, saving 100 trees per replication. We assessed nodal supports through bootstrap values with 100 pseudoreplicates and Bremer decay indices. All characters were treated as equally weighted, and unordered (except Characters 19, 49 and 50). Character state transformations to the nodes of one of the most parsimonious trees (MPTs) were reconstructed in PAUP*4.0a (Swofford, 2003) adopting DELTRAN and ACCTRAN optimizations respectively. Character mapping was performed in MacClade 4.0 (Maddison & Maddison, 2000).

Systematic paleontology

Type species. Phymolepis cuifengshanensis Chang, 1978

Included species. Phymolepis guoruii Zhu, 1996

Emended diagnosis. Yunnanolepididae in which the posterior median dorsal plate bears a strong posterior process; the anterior median dorsal plate with anterior division longer than posterior division; anterior ventral process and pit situated just below a conspicuous tergal angle and at the same level of the lateral corners of the anterior median dorsal plate; sharp median dorsal ridge between the tergal and posterior dorsal angles.

Remarks. The diagnosis of this genus follows Zhu (1996) with a minor revision. While examining V4425.2 based on high-resolution CT, we noticed that the anterior ventral process and pit are situated at the level of the lateral corners of the anterior median dorsal plate rather than behind it.

Holotype. IVPP V4425.3, a relatively complete trunk shield (Figs. 1A–Fig. 1C).

Paratype. IVPP V4425.6, a posterior median dorsal plate (Figs. 1D and 1E).

Referred specimens. IVPP V4425.1 (Fig. 2), trunk shield; V4425.2 (Fig. 3), nearly complete dermal shield only missing the anteriormost portion of head shield and the posterior median dorsal plate; V10500.1, V10508.1–3, posterior median dorsal plate; V10500.2, left anterior dorsolateral plate; V10500.3, left posterior dorsolateral plate; V10500.4, right posterior lateral plate; V10500.5, left anterior ventrolateral plate; V10500.6, right posterior ventrolateral plate.

Occurrence. The material was collected from two sites (Cuifengshan and Liaokuoshan) in Qujing city, eastern Yunnan, southwestern China.

Emended diagnosis. Phymolepis species in which the posterior process of the posterior median dorsal plate reaches one third of the plate length; the median dorsal ridge of trunk shield developed as a blunt elevation in front of the tergal angle and as a sharp crest behind the tergal angle.

Remarks. The diagnosis follows Zhu (1996) with an addition of the shape of the median dorsal ridge.

Description

Reconstruction and ornamentation

Using the complete specimen of Yunnanolepis chii (Zhang, 1978: V4423.101, fig.1) as a reference, Phymolepis cuifengshanensis could reach 84 mm in the dermal shield length, and represents the largest known species among Yunnanolepididae.

This re-investigation of P. cuifengshanensis brings together all referred specimens and leads to a new reconstruction (Figs. 5 and 6). The tentative restoration for the missing pre-orbital portion of head shield follows that of Y. chii (Zhang, 1978: fig. 1).

Small, round tubercles are densely distributed on the dorsal surface of the head and trunk shields. The tubercles are generally larger on various ridges and along outer margins of head shield than elsewhere. They are aligned parallel to the sutures between dermal plates, or radiated from the angles on the dorsal wall of trunk shield. In addition, they tend to form the rows along the sensory grooves. The tubercles on the lateral wall of the trunk shield are weakly developed and finer than those elsewhere. The ventral wall of the trunk shield is sparsely covered with tubercles that are slightly larger than the rest of the dermal shield.

Anatomical comparisons of several cranial characters in antiarchs

The restoration of the endocranium in antiarchs was mainly based on the imprints of its dorsal aspect on the visceral surface of the head shield (Stensiö, 1948; Stensiö, 1969; Miles, 1971; Denison, 1978; Young, 1984). The exception was Minicrania, which preserved the internal cast of the endocranial canals and part of the cranial cavity, thus providing information on its deeper endocranial structures (Zhu & Janvier, 1996).

In yunnanolepidoids, the visceral surface of head shield was known in Yunnanolepis (Liu, 1963: fig. 1) and Chuchinolepis (Tông-Dzuy & Janvier, 1990: fig. 17). The digital visualization of Phymolepis shows not only its visceral surface of head shield but also some internal architecture within the dermal plates, such as the trajectory of the endolymphatic duct and the cavity for the cranio-spinal process. We also re-examine the holotype (V2690.1, Fig. 12A) and one referred specimen (V4423.3, Fig. 12B) of Y. chii from the Early Devonian of Qujing, and provide more details for the visceral surface of head shield in Yunnanolepis. Based on these new data, we make comparisons in antiarchs, and show a high degree of morphological disparity with respect to the endocranium.

Anterior postorbital process. The endocrania of gnathostomes share a developed lateral projection where the orbits meet the otic capsules (Brazeau & Friedman, 2014). This process was termed the ‘anterior postorbital process’ in placoderms, and deemed as supporting the hyoid arch articulation and delimiting the posterior boundary of the spiracular chamber by its anterior surface (Young & Zhang, 1996; Brazeau & Friedman, 2014). The positional differences of the anterior postorbital process (and associated cranial nerves) along the longitudinal axis of dermal shield were thought to be informative for phylogenetic analysis (Carr, Johanson & Ritchie, 2009; Dupret et al., 2017).

Antiarchs have a well-developed anterior postorbital process. The process extending in front of the anterior border of the orbital notch, has been considered as one of the synapomorphies uniting Bothriolepis and Grossilepis (Zhang & Young, 1992; Zhu, 1996; Jia, Zhu & Zhao, 2010; Pan et al., 2018). Accordingly, the process behind the anterior border of the orbital notch is referred to a plesiomorphy of antiarchs and this state has been simply summarized as “anterior postorbital process short” in previous phylogenetic analyses (Zhang & Young, 1992).

When examining the short anterior postorbital process in antiarchs, we recognized that this state can be subdivided into two conditions: the anterior postorbital process at the same level with the posterior border of the orbital notch in yunnanolepidoids, Minicrania (Zhu & Janvier, 1996), and probably Sinolepis (Liu & P’an, 1958; Long, 1983); the process anteriorly beyond the posterior border of the orbital notch, but behind its anterior border in euantiarchs excluding Grossilepis and Bothriolepis. In this case, the distinction between these two conditions can be added to the transformation series of the anterior postorbital process.

Postorbital crista. The postorbital crista in antiarchs separates the otico-occipital depression from the orbital region in front. In several euantiarchs, the crista extends obliquely from the lateral plate to the nuchal plate as a mesial wall of the semicircular depression, such as in Bothriolepis (Stensiö, 1948), Monarolepis (Young & Gorter, 1981), Pterichthyodes (Hemmings, 1978) and Wufengshania (Pan et al., 2018). For the rest of antiarchs including yunnanolepidoids, the postorbital crista runs from the lateral plate to the postpineal plate rather than the nuchal plate as a transversely directed crest embracing the suborbital fenestra posteriorly.

Supraotic thickening. The supraotic thickening (Young, 1983: sot, fig. 3D) is bounded posteriorly by the transverse nuchal crista and extensively developed at its connection with the crista. As the supraotic thickening is porous, different from the rest of dermal skeleton in microstructure, it was considered as a junction that is co-ossified with both the endocranium and overlying head shield (Stensiö, 1948; Karatajūte-Talimaa, 1963; Moloshnikov, 2004; Moloshnikov, 2008). Euantiarchs have a persistent supraotic thickening with the exception of Microbrachius, which bears a deep groove throughout the whole length of the otico-occipital depression (Hemmings, 1978: figs. 25C–F). The presence of this thickening on the visceral surface of euantiarchs is in stark contrast to the condition in yunnanolepidoids, Minicrania and sinolepids, which lack any thickening in the corresponding area.

Median occipital crista. This crista was first identified and named by Stensiö (1931): cro, figs. 11 and 12) in Bothriolepis, and was also termed the ‘posterior median process’ by Hemmings (1978) in Pterichthyodes. Lying on the descending lamina of occipital part of the head shield, it is separated from the otico-occipital depression by the transverse nuchal crista in euantiarchs. A shallow depression of levator muscles (termed the ‘insertion fossa on head shield for levator muscles’) usually flanks on each side of the crista. In sinolepids, such as Grenfellaspis (Ritchie et al., 1992), the insertion fossa is elongated as that in euantiarchs but lacks the median crista. In yunnanolepidoids, the insertion fossa is either very short (Yunnanolepis, Fig. 12B), or totally absent (Phymolepis, Fig. 7B).

Posterior process of head shield. The posterior process of head shield (prnm, see Young (1988): figs. 7B, 37C and 44A) in euantiarchs, also termed the ‘nuchal process’ or ‘posterior median process’ (Long & Werdelin, 1986; Moloshnikov, 2004; Moloshnikov, 2008; Moloshnikov, 2010), was first identified and named by Stensiö (1931: figs. 4, 9 and 12). Although the median occipital crista is usually continuous with the posterior process of head shield, the process is apparently independent of the crista in development as evidenced by Asterolepis and Remigolepis, which possess the process but lack the crista. The process is usually developed in euantiarchs, in contrast to its absence in yunnanolepidoids and sinolepids.

Among non-antiarch placoderms, the posterior process is also known in petalichthyids (Liu, 1991; Pan et al., 2015) and arthrodires (Wang & Wang, 1983; Gardiner & Miles, 1990; Young, 2005; Carr & Hlavin, 2010; Rücklin, Long & Trinajstic, 2015).

The cavity for cranio-spinal process. The cranio-spinal process was named by Nielsen (1942), and also termed the ‘supravagal process’ by Stensiö (1969) and the ‘paroccipital process’ by Eaton (1939). It is widely developed in early gnathostomes, including arthrodires (Young, 1979), petalichthyids (Stensiö, 1925), acanthodians (Miles, 1973), actinopterygians (Patterson, 1975) and dipnoans (Miles, 1977). However, the cavity for cranio-spinal process on the visceral surface of head shield, which might function for fixing the endocranium to the external bony shield, is only found in primitive antiarchs and some arthrodires.

The cranio-spinal process in yunnanolepidoids is strongly developed, as indicated by the large cavity for the process. The process and the corresponding cavity in euantiarchs were either reduced or absent (Young, 1984).

Supraoccipital pit. The supraoccipital pit of the head shield is present for housing the endocranial supraocciptial process. It is bounded posteriorly by the transverse nuchal crista in yunnanolepidoids. The same condition is also seen in Grenfellaspis (Ritchie et al., 1992) and Minicrania, despite the supraoccipital pit in the latter has ever been interpreted as impression of endolymphatic sac (Zhu & Janvier, 1996; Dupret et al., 2017). In euantiarchs, the supraoccipital pit is only seen in few Bothriolepis species with different positions: either immediately anterior to the transverse nuchal crista as exemplified by B. tatongensis (Long & Werdelin, 1986), or on the transverse nuchal crista as in B. macphersoni and B. portalensis (Young, 1988).

In non-antiarch placoderms, the supraoccipital pit has been observed in petalichthyids (Liu, 1991: cv.ifNu, fig. 2), and most arthrodires, including Holonematidae (Miles, 1971: tf; figs. 53 and 117; Young, 2005: if.pt, fig. 2C), Buchanosteidae (Young, 1981b: if.pt, fig. 6), Coccosteoidea (Miles & Westoll, 1968: p.pts.Nu, fig. 2A), Dunkleosteoidea (Zhu, Zhu & Wang, 2016: f.pt.u, fig. 5; Carr & Hlavin, 2010: pt.u, fig. 6A) and Dinichthyidae (Carr & Hlavin, 2010: pt.u, fig. 1A).

Trajectory of endolymphatic duct. The trajectory of the endolymphatic duct through the dermal bone in respect of length and orientation mainly depends on the relative position between the internal and external pores. This character was considered informative for the resolution of placoderm interrelationships. The trajectory had ever been decomposed into two states: vertical (a trait in most non-arthrodire placoderms), long and oblique (a trait shared by arthrodires) by Goujet & Young (1995). Coates & Sequeira (1998) considered the posteriorly oriented duct as a primitive character of gnathostomes as it is shared by agnathans, placoderms and osteichthyans. Brazeau (2009) suggested the presence of posterodorsally angled trajectory as an arthrodire character and the absence of oblique trajectory of endolymphatic duct as a character shared by antiarchs, Brindabellaspis and petalichthyids. Our study herein shows the condition in antiarchs is more complicated than previously thought.

In antiarchs, the distance between the internal pores is usually greater than that of external ones (Stensiö, 1948; Karatajūte-Talimaa, 1966; Long, 1983), and the endolymphatic duct extends dorsomesially. As the external pore of endolymphatic duct is always positioned close to the posterior edge of the nuchal plate in antiarchs, the relative position of the internal pore along the antero-posterior axis reflects the relative length and orientation of the endolymphatic duct.

In yunnanolepidoids, the internal pore of endolymphatic duct is located far in front of the transverse nuchal crista, and thus far from the external pore. As such, the endolymphatic duct is elongated through the nuchal plate and obliquely oriented. Sinolepids (Ritchie et al., 1992; Janvier, 1996) and euantiarchs differ in having a short, slight oblique endolymphatic duct as the internal pore is positioned just anterior to the external one.

In non-antiarch placoderms, the elongated endolymphatic duct is also present in arthrodires (Young, 2010; Dupret et al., 2017). However, the endolymphatic duct of arthrodires is directed dorsolaterally, not dorsomesially as in antiarchs.

Occipital portion of endocranium. The internal pore for the endolymphatic duct in antiarchs, is located roughly at the posterior boundary of the semicircular depression on the visceral surface of head shield as that in arthrodires (Zhu, Zhu & Wang, 2016), hence we use this pore as a proxy to denote the otic-occipital boundary. Taking the length of the otic-occipital depression as the constant variable in antiarchs, the length between the internal pore of endolymphatic duct and the posterior border of otic-occipital depression represents the occipital proportion in endocranium.

In yunnanolepidoids, the internal pore on the visceral surface of the head shield is far from the transverse nuchal crista as described above, implying the occipital portion of the endocranium is elongated as in arthrodires (Zhu, Zhu & Wang, 2016). By comparison, the occipital portion is short in other antiarchs (Stensiö, 1948; Ritchie et al., 1992).

Confluence of anterior and posterior semicircular canals. The anterior and posterior semicircular canals meet at the confluence in the medial part of the inner ear (Dupret et al., 2017). As the posterior border of orbital notch and the transverse nuchal crsita roughly border the anterior and posterior margins of the otic-occipital depression respectively, we can use them as references to estimate the relative position of the confluence in endocranium.

In yunnanolepids and Minicrania (Zhu, 1996), the confluence is halfway between the posterior border of the orbital notch and the transverse nuchal crista. By comparison, the confluence is closer to the transverse nuchal crista than to the posterior border of orbital notch in sinolepids and euantiarchs.

Phylogenetic results

The maximum parsimony analysis yields 726 MPTs of 179 steps each (consistency index = 0.4693; retention index = 0.8045). All the MPTs are summarized as a strict consensus tree (Fig. 13A) and a 50% majority-rule consensus tree (Fig. 13B). One MPT is selected to illustrate the character transformations at nodes (Fig. 14A), and the list of synapomorphies defining various nodes is shown in Supplementary Information.

Antiarchs (Fig. 14A: node 1) are characterized by up to 10 synapomorphies including two newly proposed cranial features (Character 27⁰, absence of posterior process of head shield; Character 38¹, presence of supraoccipital pit). Character 27 shows a reversal in euantiarchs (Fig. 14A: node 15). Character 38 is a highly homoplastic character, and shows a reversal in euantiarchs and a parallelism in Bothriolepis.

Yunnanolepidoids (Fig. 14A: node 2) form the basal members of antiarchs, consistent with the position as they were first phylogenetically analysed (Zhu, 1996). However in new scenario, Chuchinolepis, Vanchienolepis and a clade formed by yunnanolepids, Zhanjilepis and Heteroyunnanolepis (Wang, 1994) fall into a polytomy with remaining antiarchs. In yunnanolepids, Yunnanolepis is the sister group of a polytomic clade comprising Phymolepis, Mizia and Parayunnanolepis.

Four new cranial characters provide further support the monophyly of euantiarchs (Fig. 14A: node 15), including one uniquely shared character (Character 36¹, anterior postorbital process lying in front of posterior level of orbital notch) and three homoplastic characters (Character 26¹, presence of median occipital crista of head shield; Character 27¹, presence of posterior process of head shield and Character 38⁰, absence of supraoccipital pit of head shield).

Microbrachiids (Fig. 14A: node 16) are resolved as the sister group of the remaining euantiarchs, and the conventional bothriolepidoids are resolved as a paraphyletic assemblage. These results are congruent with previous analyses of Zhu (1996) and Pan et al. (2018). Relationships of the remaining bothriolepidoids (Fig. 14A: node 19) are completely unresolved in the strict consensus tree, which may be related to the large number of missing data in some of them. Euantiarchs excluding microbrachiids bear one uniquely shared endocranial character (Character 25¹, presence of supraotic thickening of head shield).

Our analysis that incorporate new cranial characters yields resultant trees, which are consistent with previous resolutions of Zhu (1996), Jia, Zhu & Zhao (2010) and Pan et al. (2018) in broad phylogenetic pattern. Under the new phylogenetic scenario, we can trace the character transformations relating to the dorsal aspect of endocranium in antiarchs.

Yunnanolepidoids (Figs. 7 and 12 and 14A ) and Minicrania show primitive character states, such as the anterior postorbital process being posteriorly positioned (Character 36⁰), presence of cranio-spinal process (Character 37¹) and supraoccipital process (Character 38¹), anterior and posterior semicircular canals being anteriorly positioned (Character 39⁰), long endolymphatic duct (Character 40⁰), and long occipital portion (Character 41⁰).

At the node comprising sinolepids and euantiarchs (Fig. 14A: node 10), there are two derived endocranial character states: short endolymphatic duct (Character 40¹) and short occipital region (Character 41¹). Euantiarchs differ from sinolepids in possessing the following derived states: the anterior postorbital process lying in front of the posterior level of orbital notch (Character 36¹), and the absence of the supraoccipital process (Character 38⁰). In short, there exists a large morphological disparity relating to the dorsal aspect of endocranium between yunnanolepidoids, sinolepids and euantiarchs.