Morphology of the mandibles and the first maxillae in the family Lithobiidae (Myriapoda, Chilopoda), with remarks on their phylogenetic significance

Abstract

The mandibles and the first maxillae of 37 species of the family Lithobiidae (Myriapoda, Chilopoda) were investigated and compared to provide a structural overview and evaluate their significance for the systematics of the family. The species sampling focused on the genus Lithobius, examining 33 species of four subgenera (Lithobius, Monotarsobius, Sigibius, Ezembius), as this genus represents about half of the known diversity of Lithobiidae, including more than 500 assigned species and subspecies. The microstructures on the mandibular gnathal edge and the first maxillary telopodites and coxal projections were studied using scanning electron microscopy.

Although having a similar structural pattern, we demonstrate that the microstructures are variable within and between species of adult specimens and commonly show intergradation. To check for intraspecific variability of microstructures and character stability, specimen sampling was extended for the two common Austrian species Lithobius dentatus and Lithobius validus, for which seven specimens depicted no major differences in the mandibular gnathal edge and the first maxillae. Our data suggest the presence of three characters in the mandibular gnathal edge and the first maxillae useful for lithobiid phylogeny. These characters were tested in a phylogenetic analysis together with previously described and novel morphological characters. Subgenera of Lithobius are mostly non‐monophyletic, and several other genera of Lithobiinae as well as other subfamilies group with particular species or clades of Lithobius. The results corroborate a close relationship between Disphaerobius loricatus and Lithobius (Ezembius) giganteus, strengthening the hypothesis that Pterygoterginae is nested within Lithobiinae and specifically within Lithobius, allied to L. (Ezembius) and Hessebius.

1. INTRODUCTION

The mandibles are a key character underpinning the Mandibulata hypothesis for arthropod interrelationships, unifying myriapods, hexapods, and crustaceans as a monophyletic group (Snodgrass, 1950). This monophyly is additionally supported by homologous structures on the most distal part of the mandible, the mandibular gnathal edge (Edgecombe, Richter, & Wilson, 2003). The gnathal edge mainly consists of the pars incisiva and the pars molaris, and variably by the lacinia mobilis, each of which bears several microstructures (Edgecombe et al., 2003). These microstructures provide phylogenetic information at finer taxon levels within the mandibulate subphyla (see Edgecombe et al., 2003 and references therein). For example, for representatives of the myriapod class Chilopoda, the microstructures of the mandibular gnathal edge have proven to be important in several studies addressing the morphology, taxonomy, systematics, and phylogeny of different orders (e.g., Edgecombe, 2004a; Edgecombe & Giribet, 2004; Edgecombe, Giribet, & Wheeler, 2002; Edgecombe & Hollington, 2002; Hollington & Edgecombe, 2004; Koch & Edgecombe, 2012; Koch, Edgecombe, & Shelley, 2010). In the order Lithobiomorpha, as in the three other chilopod orders that comprise the Pleurostigmophora, the mandibular gnathal edge consists principally of the pars incisiva, which bears aciculae (pectinate lamellae), the mandibular teeth (dentate lamina) and branching bristles. A well‐developed lobe equipped with trichomes dorsally viz. the pulvillus (sensu Crabill, 1960) or Haarpolster (sensu Verhoeff, 1918), was considered as the pars molaris (Edgecombe et al., 2002; Edgecombe et al., 2003). However, the pulvillus has been reinterpreted in recent studies as the dorsalmost part of the pars incisiva whereas the pars molaris is confined to a sclerite that bears the mandibular condyle, the lamina condylifera (Koch & Edgecombe, 2012).

In addition to the mandibles, the first maxillae of Lithobiomorpha have also been documented in some systematic works (e.g., Edgecombe, 2004a; Edgecombe et al., 2002; Edgecombe & Giribet, 2004; Edgecombe & Hollington, 2002; Hollington & Edgecombe, 2004). The first maxillae consist of paired coxosternites and two‐segmented telopodites, each with a basal and a distal article, and a coxal projection. The presence of plumose bristles along the inner margin of the distal article of the maxillary telopodite is autapomorphic for Lithobiomorpha (Edgecombe, 2004b).

The mandibles and the first maxillae have provided characters that were used in combination with molecular data for phylogenetic analyses on the lithobiomorph family Henicopidae (e.g., Edgecombe et al., 2002; Edgecombe & Giribet, 2003; Koch & Edgecombe, 2008). To date, only a few representatives (13 species and eight genera) of the family Lithobiidae were investigated in detail for their mandibular and first maxillary microstructures but mainly as an outgroup in analyses of henicopids, and the character coding was invariant (Edgecombe et al., 2002; Edgecombe & Giribet, 2004; Koch & Edgecombe, 2008). Therefore, it has remained unclear if the mandibles and the first maxillae of the Lithobiidae bear useful characters for systematic and phylogenetic analyses. This is especially the case for the species‐rich genus Lithobius Leach, 1814 known to accommodate more than 500 species/subspecies (see Bonato et al., 2016; Zapparoli & Edgecombe, 2011). Thus, any new morphological or molecular information is of potential value to help solve some of the unclear species‐interrelationships within the genus, its phylogeny and evolutionary history. The genus Lithobius was hitherto subject to one morphological phylogenetic analysis based on 40 characters resolving it as non‐monophyletic, with a few species being more closely related to the genera Australobius Chamberlin, 1920, Hessebius Verhoeff, 1941 and Pleurolithobius Verhoeff, 1899 (Koch & Edgecombe, 2008). A more recent study on the peristomatic structures (epipharynx and hypopharynx) of 32 species of the same genus highlighted eight new characters, which could also be systematically useful (Ganske, Edgecombe, & Akkari, 2018).

In this study, we focus on the mandibles and maxillae of the family Lithobiidae. Therefore, we examined and compared the mandibular gnathal edge and the first maxillae of 37 species using scanning electron microscopy. Among these, 33 species were from the genus Lithobius, including four subgenera: Lithobius Leach, 1814; Monotarsobius Verhoeff, 1905; Sigibius Chamberlin, 1913 and Ezembius Chamberlin, 1919, as well as four species from allied lithobiid genera, viz. Eupolybothrus Verhoeff, 1907; Disphaerobius Attems, 1926 and Neolithobius Stuxberg, 1875.

The aim of this work is twofold: First, we provide a descriptive and comparative overview on the morphology of the mandibular gnathal edge and the first maxillae of the studied species. Second, we evaluate the phylogenetic importance of the investigated microstructures together with other morphological characters for the systematics of Lithobiidae, especially within the genus Lithobius. Intraspecific variability and plasticity of the structures were checked with an extended specimen sampling for two Lithobius species. Phylogenetic analyses using maximum parsimony and maximum likelihood of the newly described characters together with characters from other morphological systems were performed, including characters extracted from other Lithobiidae, Henicopidae, and two representatives of Scutigeromorpha as outgroup. We further compared the mandibular gnathal edge of Disphaerobius loricatus (Sseliwanoff, 1881) and Lithobius (Ezembius) giganteus Sseliwanoff, 1881 and, together with the phylogenetic results, verified that it supports previous suggestions on their relationships based on external morphological and peristomatic characters (Farzalieva, Nefediev, & Tuf, 2017; Ganske et al., 2018). This is of particular systematic importance because Disphaerobius is usually classified separately from Lithobius, in the subfamilies Pterygoterginae and Lithobiinae, respectively.

2. MATERIAL & METHODS

2.1. Material

The studied material consists of 62 specimens belonging to 37 species preserved in 70 or 95% EtOH (Table 1), deposited at the Natural History Museum Vienna (NHMW), The Natural History Museum, London (BM/NHMUK), and the Hungarian Natural History Museum Budapest (HNHMB) (Supporting Information 1). The material was examined with light and scanning electron microscopy.

Table 1.

List of species where mandibles and first maxillae were studied (+) or not studied (−)

			Mandibles		First maxillae
Species	Voucher	Sex	Left	Right	Left	Right
Lithobius (Lithobius) agilis C.L. Koch, 1847	NHMW 9123	f	+	−	−	+
L. (L.) calcaratus C.L. Koch, 1844	NHMW 9132	m	+	−	−	−
L. (L.) carinatus L. Koch, 1862	NHMW 9125	f	−	+	+	+
L. (L.) castaneus Newport, 1844	NHMW 9194	f	+	−	−	+
L. (L.) cyrtopus Latzel, 1880	NHMW 1081	f	+	+	−	+
	NHMW 9345	m	+	+	−	−
L. (L.) dentatus C.L. Koch, 1844	NHMW 9135	f	+	+	+	+
	NHMW 9338	f	+	+	+	+
	NHMW 9339	f	+	+	+	+
	NHMW 9340	f	+	+	+	+
	NHMW 9341	f	+	+	+	+
	NHMW 9342	f	+	+	+	+
	NHMW 9343	m	+	+	−	−
L. (L.) erythrocephalus C.L. Koch, 1847	NHMW 9136	f	+	+	+	−
L. (L.) fagei Demange, 1961	NHMUK	m	+	−	+	+
	NHMW 9138	m	+	+	−	−
L. (L.) forficatus (Linnaeus, 1758)	NHMW 9139	f	−	−	+	+
	NHMW 9349	f	+	+	−	−
L. (L.) lapidicola Meinert, 1872	NHMW 9196	f	+	−	−	+
L. (L.) latro Meinert, 1872	NHMW 9141	f	+	+	+	+
L. (L.) lucifugus L. Koch, 1862	NHMW 9143	f	−	+	+	+
L. (L.) macilentus L. Koch, 1862	NHMW 9144	m	−	−	+	−
L. (L.) mutabilis L. Koch, 1862	NHMW 9126	f	+	+	−	−
	NHMW 9127	f	+	+	+	+
L. (L.) muticus C.L. Koch, 1847	NHMW 9145	m	+	+	−	−
L. (L.) nodulipes Latzel, 1880	NHMW 9146	f	+	−	−	+
	NHMW 9346	m	+	+	−	−
L. (L.) pelidnus Haase, 1880	NHMW 9149	m	+	+	+	+
L. (L.) peregrinus Latzel, 1880	NHMW 9129	m	−	+	−	+
L. (L.) piceus L. Koch, 1862	NHMW 9128	f	+	−	+	+
L. (L.) pyrenaicus Meinert, 1872	NHMW 9130	m	+	+	+	+
	NHMW 9131	f	+	+	−	−
	NHMW 9347	f	+	+	−	−
L. (L.) tenebrosus Meinert, 1872	NHMW 9152	f	+	−	+	+
L. (L.) tricuspis Meinert, 1872	NHMW 9153	f	+	+	+	+
	NHMW 9150	f	+	+	+	+
	NHMW 9332	m	+	+	+	+
	NHMW 9333	f	+	+	+	+
L. (L.) validus Meinert, 1872	NHMW 9334	f	+	+	+	+
	NHMW 9335	m	+	+	+	+
	NHMW 9336	f	+	+	+	+
	NHMW 9337	f	+	+	+	+
L. (Monotarsobius) aeruginosus L. Koch, 1862	NHMW 7546	f	+	+	−	−
	HNHMB 5980	f	+	+	+	+
L. (M.) austriacus (Verhoeff, 1937)	HNHMB 5984	m	−	+	+	+
	HNHMB 5983	m	+	+	−	−
L. (M.) crassipes L. Koch, 1862	NHMW 9157	f	+	+	+	+
	NHMW 9344	m	+	+	−	−
L. (M.) curtipes C.L. Koch, 1847	HNHMB 5985	f	−	+	+	+
L. (M.) franciscorum Dányi & Tuf, 2012	HNHMB 5987	m	+	−	−	+
	HNHMB 5988	m	−	+	−	−
L. (Sigibius) burzenlandicus Verhoeff, 1931	HNHMB 5989	m	−	+	+	+
	HNHMB 5990	m	+	+	−	+
L. (S.) microps Meinert, 1868	HNHMB 5991	f	+	−	+	+
L. (S.) trebinjanus Verhoeff, 1900	NHMW 9156	f	+	−	+	−
L. (Ezembius) electus Silvestri, 1935	NHMUK	f	−	+	+	+
L. (E.) giganteus Sseliwanoff, 1881	NHMW 9348	m	+	+	+	+
	NHMW 9368	m	+	+	+	+
Neolithobius aztecus (Humbert & Saussure, 1869)	NHMUK, BM1894.4.1.75–77	f	+	−	+	+
Disphaerobius loricatus (Sseliwanoff, 1881)	NHMW 9204	f	+	+	+	+
Eupolybothrus (Eupolybothrus) grossipes (C.L. Koch, 1847)	NHMW 9176	m	−	+	+	+
Eupolybothrus (Leptopolybothrus) tridentinus (Fanzago, 1874)	NHMW 9350	f	+	+	+	+

2.2. Sample preparation

The mandibles and/or first maxillae were detached from the head with forceps and dissecting needles in one to seven adult male or female individuals per species (see Table 1; Supporting Information 1). Manual multifocus images of the sclerotized parts of the mandibles and first maxillae were obtained with a Nikon Eclipse Ni compound microscope (LM) equipped with a Nikon DS‐Ri2 camera using NIS‐Elements Microscopic Imaging Software. For scanning electron microscopy (SEM), the specimens were: (a) cleaned in an ultrasonic bath (50–60 Hz) for 5 to 10 s (maximum); (b) dehydrated in an ascending alcohol series (70%, 80%, 90%, 96% EtOH, 2 × 10 min each); (c) covered with HMDS (hexamethyldisilazane) and air dried overnight. The mouthparts were mounted on sticky aluminium tape fixed on aluminium stubs, glued with conductive silver and coated with platinum (Leica EM SCD500). Then, they were studied and photographed with a JEOL JSM 6610‐LV at an accelerating voltage of 10–15 kV. LM and some SEM images were stacked and processed using Zerene Stacker (version 1.04) and Adobe Photoshop CS6. Figures were assembled in Adobe InDesign CS6.

Terminology follows Bonato et al. (2010), Edgecombe et al. (2002), and Edgecombe et al. (2003) where applicable.

2.3. Morphological characters and coding

The data matrix in Table 2 codes for 62 morphological characters described below under “Morphological characters and observations.” The character matrix (Table 2) was compiled using Mesquite version 3.40 (Maddison & Maddison, 2018) and is available as RTF‐file (Supporting Information 2).

Table 2.

Data matrix for 62 morphological characters as described under “Character description and observations”: ? = missing data; – = inapplicable character state; (xx) = polymorphism

	Characters
	0000000001	1111111112	2222222223	3333333334	4444444445	5555555556	66
Species	1234567890	1234567890	1234567890	1234567890	1234567890	1234567890	12
Scutigera coleoptrata	0––0–2–000	–10000000–	0100–00000	?–00010000	–––05300––	0000–––––0	–0
Scutigerina weberi	0––0–2–000	–10000000–	00–0–00000	?–00010000	–––0?300––	0000–––––0	–0
Anopsobius neozelanicus	3––1–00012	1211101101	1302111001	??01?00101	0001?110––	00001010?1	?0
Dichelobius bicuspis	3––1–10012	1211101101	1302111001	??01?00101	00013110––	0000101001	?0
Dzhungaria gigantea	3––1–20111	1110101102	10–2001000	0112100110	00210010––	0000100001	10
Henicops washpoolensis	2––1–20111	112010?103	1112001000	0012102010	00100010–0	00000000?1	10
Lamyctes emarginatus	2––1–20111	1122101101	1112101000	?111110011	00100010–0	0000100001	?0
Paralamyctes validus	2––1–20012	1120102101	1112001000	?111102010	00100010––	0000000001	?0
Zygethobius pontis	2––1–20111	100010?1??	1112101001	?111103110	00200010––	0000000001	?0
Australobius scabrior	121211110(12)	110111110?	0201011110	1003102100	10101200–0	0000–10102	10
Bothropolys multidentatus	1210110102	121211?1??	0200111010	??03?03100	1110?20??0	0000–00102	00
Disphaerobius loricatus	1010110101	1002112013	0202–11110	1103100100	1010120210	0010–00002	00
Eupolybothrus fasciatus	1210121102	1111112210	0200011010	1103?03100	1110120210	0010–000?0	00
Eupolybothrus grossipes	1210121102	1111112210	0200111010	1103103100	1110120220	0000–00010	00
Eupolybothrus tridentinus	1210121102	1111112110	02000??110	1103003100	1110120220	0000–00010	00
Harpolithobius anodus	1200121101	0122112101	1200?11110	?103?12100	1010050200	0000–00112	00
Hessebius plumatus	1210111101	1122112013	?202–11010	11031?0100	10100200–0	0000–000?2	?1
Neolithobius aztecus	1210121102	1121112110	1200011110	?103102100	1010420110	0000–00102	00
Pleurolithobius patriarchalis	1100121101	122111?1?0	1202111010	?103?10100	1010020220	0000–10012	00
Pseudolithobius megaloporus	1200110101	1022112100	0201011010	?103?02100	1020320120	0000–10002	00
Lithobius (Ezembius) electus	1210111101	1101112100	0202011110	1103100100	10103200–0	0000–10002	01
L. (E.) giganteus	1010111101	1002112013	0202–11110	1103110100	1010120210	0000–00002	01
L. (Lithobius) agilis	1210121101	1101112100	1201111010	0003112100	10102200–0	0000–00102	00
L. (L.) calcaratus	1000121101	0201112200	120211101?	00???10100	10100400–0	1000–00202	00
L. (L.) carinatus	1100121101	1101112100	0201111010	0?03010100	10100500–0	0000–10002	00
L. (L.) castaneus	1210121101	0221112100	0202011010	1103000100	10104200–0	0000–00202	00
L. (L.) cyrtopus	1210121101	1101011101	1200111110	1103111100	1010120120	0000–00112	00
L. (L.) dentatus	1210121101	1101112100	1202111110	1103012100	1010120200	0000–00112	00
L. (L.) erythrocephalus	1210121101	1101112100	1202111110	0103012100	10102200–0	0000–00102	00
L. (L.) fagei	1210121101	1201112100	0201011010	1003011100	10101200–0	0000–00102	00
L. (L.) forficatus	1210121102	1221112100	0200011110	1103002100	1010120210	0000–00102	10
L. (L.) lapidicola	1210121101	12010111?0	1??21?1110	1103011100	10100600–0	0000–00102	00
L. (L.) latro	1210121101	1101012100	1202111110	0103110100	1010120100	0000–00112	00
L. (L.) lucifugus	1210121101	1101112200	1202111110	0103010100	10101200–0	0000–00102	00
L. (L.) macilentus	1000021101	1121112100	12011111??	??0??12100	1010120220	0000–00112	00
L. (L.) mutabilis	1210121101	1101111100	1202111110	1103012100	1010120200	0000–00112	10
L. (L.) muticus	1210121101	0101111100	0200111110	01???10100	10101200–0	0001–(01)0112	10
L. (L.) nodulipes	1210121101	10211121?0	1??1111110	110??12100	10100500–0	1000–(01)0112	00
L. (L.) pelidnus	1210121101	1101012101	1200011110	1?0??12100	1010120220	0011–00112	00
L. (L.) peregrinus	1210121102	1221112102	0200011010	0103012100	1010220210	0000–00102	00
L. (L.) piceus	1210121101	1201112100	0202111010	0103012100	10102200–0	0000–10202	00
L. (L.) pyrenaicus	1210121101	1101111100	0200011110	1103100100	1010120200	0000–00012	00
L. (L.) tenebrosus	1210121101	1201011200	12021?1?10	0103112100	10101500–0	0000–00112	00
L. (L.) tricuspis	1210121101	12211121?0	0??2011010	0103012100	10102200–0	0000–10102	00
L. (L.) validus	1210121102	1201112100	0200011110	1103003100	1010120220	0000–00112	10
L. (Monotarsobius) aeruginosus	1100011101	1101111100	1202111110	0103000101	10101500–0	0000–00102	10
L. (M.) austriacus	1100111101	1101111100	1201111110	??03000101	10101500–1	0000–00102	10
L. (M.) crassipes	1210111101	1201112100	1202111110	1103010101	10101200–0	0000–00102	10
L. (M.) curtipes	1000111101	1121012100	020111111?	??03010101	10101200–0	1000–00112	00
L. (M.) franciscorum	1100011101	122101?100	1200111110	1103010101	10103200–0	0100–10002	00
L. (M.) holstii	1210?11101	1101?11101	0302111110	?103010101	10101200–0	0000–000?2	?0
L. (Sigibius) burzenlandicus	1(01)00021101	1221111100	120?111010	0103010101	10101400–0	0000–00102	10
L. (S.) microps	1100121101	1221111101	1202111010	0003?10101	10100500–0	0000–00102	00
L. (S.) trebinjanus	1100121101	1202112100	1201111010	000??10101	10100400–0	0000–00102	00

Information on the peristomatic structures of the species included in the phylogenetic analysis was obtained from Koch and Edgecombe (2008) and Ganske et al. (2018). Other morphological information was acquired from Koch and Edgecombe (2008), new observations, or from literature resources.

2.4. Phylogenetic analysis

Cladograms were rooted with Scutigeromorpha as an outgroup to Lithobiomorpha. Parsimony analyses under equal weights were conducted using TNT version 1.1 (Goloboff, Farris, & Nixon, 2003) with heuristic searches involving 1,000 random addition sequences and TBR branch swapping saving up to 100 trees per replicate. Nodal support was measured using Jackknife resampling (Farris, 1997) and Bremer support (Bremer, 1994), both with TNT. Jackknifing used 1,000 replicates with 36% deletion, each replicate involving a heuristic search. Bremer support was calculated from collections of suboptimal trees obtained by heuristic searches. Implied weighting implemented in TNT used the same heuristic search parameters as for equal weights, across concavity constants k = 2, 3, 4, and 5. Multistate characters 17 and 18 were ordered (additive), the others unordered (nonadditive). Character optimisations were examined using ASADO version 1.61 (Nixon, 2004).

The maximum likelihood analysis was performed with the default settings in W‐IQ‐TREE (IQ‐TREE version 1.6.6 [Nguyen, Schmidt, von Haeseler, & Minh, 2015]) using an ascertainment bias correction (ASC; Lewis, 2001) and ultrafast bootstrap approximation (1,000 iterations; Hoang, Chernomor, von Haeseler, Minh, & Vinh, 2018). The implemented ModelFinder (Kalyaanamoorthy, Minh, Wong, von Haeseler, & Jermiin, 2017) estimated ORDERED+FQ+ASC+G4 as the best‐fit substitution model according to BIC.

Trees were visualized using FigTree v1.4.2 (http://tree.bio.ed.ac.uk/software/figtree/) and adjusted in Adobe Illustrator CS6 and Adobe Photoshop CS6.

3. RESULTS

3.1. Mandibles

In all examined species, the mandibular gnathal edge (Figure 1) agrees with the previous descriptions provided for the same family (Edgecombe et al., 2002; Edgecombe et al., 2003).

• aAciculae (aci, aco) and pinnules (pi)

Figure 1.

Overview of the mandibular gnathal edge of the genus Lithobius. (a, b) Lithobius (Lithobius) forficatus (LM), (a) outside, right is dorsal, (b) inside, right is ventral; (c) Lithobius (Sigibius) microps, outside, right is dorsal (SEM); (d) Lithobius (Lithobius) erythrocephalus, inside, right is ventral (SEM). Abbreviations: aci—Inner row of mandibular aciculae; aco—Outer row of mandibular aciculae; ad—Accessory denticles on mandibular teeth; fr—Fringe of branching bristles on mandibular gnathal edge; pi—Pinnules on mandibular aciculae; pu—Pulvillus on mandible; tb—Mandibular bicuspid tooth/teeth; tpu—Trichomes on pulvillus; tt—Mandibular tricuspid tooth/teeth

The aciculae are present on the ventral part of the gnathal edge (Figure 1) and occur as an inner and an outer row (Figures 1 and 2a,b). They vary in number from eight to 21 between the species. The number may also vary between conspecifics, for example, L. pyrenaicus with 18 or 21 (N = 3), L. validus with 17 or 22 (N = 7) and L. dentatus with 14 to 16 (N = 6). Numbers of aciculae can be the same between left and right mandibles, for example, L. nodulipes (16, 16) or different, for example, L. cyrtopus (14, 15), L. crassipes (14, 13) and L. pyrenaicus (17, 18). It seems that there is no correlation between the number of aciculae and the body size of the adult individuals and species. There are also no sexual differences.

Figure 2.

Mandibular aciculae of the genus Lithobius (SEM). (a, b) Lithobius (Lithobius) validus, left mandible with inner and outer row of aciculae, (a) outside, right is dorsal, (b) inside, right is ventral; (c) Lithobius (Lithobius) cyrtopus, aciculae with median ridge (arrow) and bipinnulation, outside, bottom is ventral; (d) Lithobius (Lithobius) carinatus, aciculae with median ridge (arrow) and bipinnulation, outside, bottom is ventral, inset: Closeup of distal part of aciculae; (e) Lithobius (Monotarsobius) franciscorum, pinnules fringe three‐quarters of the aciculae’ length, outside, bottom is ventral; (f) Lithobius (Sigibius) microps, pinnules fringe three‐quarters of the aciculae’ length, stout pinnules with blunt tips, inside, bottom is ventral. Abbreviations: aci—Inner row of mandibular aciculae; aco—Outer row of mandibular aciculae; fr—Fringe of branching bristles on mandibular gnathal edge; pi—Pinnules on mandibular aciculae

The aciculae are mainly smooth and may show a weak surface striation with ridges running parallel to the edges of the pinnules (Figure 3c,d). The aciculae have a median ridge on the internal surface (Figures 2c,d and 3b,c) and are bipinnulate (Figures 2c,d and 3b,d,e), except for E. tridentinus, which has some apinnulate aciculae (Figure 3f). The pinnules fringe approximately half to three quarters of the length of the aciculae from distal to proximal (Figures 2d–f and 3a). The number of pinnules ranges from 15 to more than 50. The number of pinnules on the aciculae of a single individual is highly variable, although there are exceptions, with two or three aciculae bearing the same numbers in the same individual (e.g., L. franciscorum, L. tenebrosus). Pinnules may appear slender or stout, with pointed, blunt or minute teeth‐bearing tips (Figures 2c–f and 3). The number of minute teeth on the tips of the pinnules ranges from two to five (Figure 3d,e). The pinnules of the inner row of aciculae are usually less pronounced compared to those on the outer row (Figure 3a).

• bMandibular teeth (tb, tt) and accessory denticles (ad)

Figure 3.

Mandibular aciculae of Lithobiidae (SEM). (a) Eupolybothrus (Eupolybothrus) grossipes, pinnules fringe half of the aciculae’ length, pinnules on the inner row of aciculae are less pronounced than those on the outer row, right mandible, bottom is ventral; (b) Lithobius (Lithobius) forficatus, aciculae with a median ridge (arrow); (c) detail of Figure 3a, aciculae with median ridge (arrow) and slender pinnules, bottom is ventral; (d) Lithobius (Ezembius) electus, smooth aciculae with weak surface striation running parallel to the edges of the pinnules, pinnules are blunt and bear minute teeth; (e) Lithobius (Lithobius) cyrtopus, pinnules with minute teeth at their tips (arrows); (f) Eupolybothrus (Leptopolybothrus) tridentinus, bipinnulate and apinnulate (arrow) aciculae, left mandible, bottom is ventral. Abbreviations: aci—Inner row of mandibular aciculae; aco—Outer row of mandibular aciculae

The mandibles bear four teeth with the two or three ventralmost being tricuspid and the one or two dorsalmost bicuspid (Figures 1, 4a, and 8). Accessory denticles on the teeth are present in all examined lithobiid species (Figures 1c, 4, 5, and 6a,b). The denticles occur with different shapes and shape transitions, which are consistent between conspecifics. The accessory denticles are multifurcate or triangular scales presenting a spinous, tuberculate, or smooth surface. Species may display (a) only one type of scales, for example, smooth and triangular (e.g., D. loricatus, L. giganteus; Figure 5a,b) or multifurcate and structured scales (e.g., L. aeruginosus; Figure 4d), or (b) a transition of more than one scale type (e.g., E. grossipes, L. piceus, L. pyrenaicus, and L. castaneus; Figure 4b,c,e,f). The accessory denticles are interrupted by a more or less distinct ridge on the ventral edge of a tooth between two flanking teeth (Figures 1c, 4a–c, and 5a,b). This ridge is exceptionally pronounced in L. giganteus and D. loricatus (Figures 5a,b). The accessory denticles pass into the pulvillus area continuously (Figures 5a‐d,f, and 6a,b) or with a break (a trichome‐free strip) (Figures 1c and 5e). If continuous, it can be a progressive (Figure 5d) or abrupt transition (e.g.,Figures 5a,b,c,f and 6a,b).

• cThe pulvillus (pu)

Figure 4.

Details of mandibular gnathal edge of Lithobiidae (SEM). (a, b) Eupolybothrus (Eupolybothrus) grossipes, right mandible, left is ventral, (a) accessory denticles on the mandibular teeth are separated by a distinct median ridge (arrow) and pass abruptly into the trichomes of the pulvillus, the two dorsalmost teeth are bicuspid, (b) details from Figure 4a with accessory denticles of different scale types and a fringe of “scale‐like” branching bristles; (c) Lithobius (Lithobius) piceus, multifurcate and structured or smooth accessory denticles divided by a less distinct ridge (arrow), right mandible, right is dorsal; (d) Lithobius (Monotarsobius) aeruginosus, multifurcate and structured accessory dentilces, right mandible, right is ventral; (e) Lithobius (Lithobius) pyrenaicus, multifurcate or triangular and structured or smooth accessory denticles, right mandible, right is ventral; (f) Lithobius (Lithobius) castaneus, multifurcate or triangular and structured or smooth accessory denticles, right mandible, right is ventral. Abbreviations: ad—Accessory denticles on mandibular teeth; fr—Fringe of branching bristles on mandibular gnathal edge; pu—Pulvillus on mandible; tpu—Trichomes on pulvillus

Figure 8.

Inner side of mandibular gnathal edge of Lithobiidae (SEM). (a) Lithobius (Lithobius) muticus, internal surface without spinulation, left mandible, right is ventral; (b) Lithobius (Lithobius) mutabilis, internal surface with three branching bristles (arrow), left mandible, right is ventral; (c, d) Lithobius (Ezembius) giganteus, internal surface with simple and tapering spines, (c) three spines (arrow), right mandible, right is dorsal, (d) seven spines (arrow), left mandible, right is ventral; (e) Disphaerobius loricatus, internal surface with simple and tapering spines (arrow), left mandible, right is ventral; (f) Lithobius (Ezembius) electus, internal surface with simple and tapering spines (arrow), right mandible, left is dorsal

Figure 5.

Details of mandibular gnathal edge of Lithobiidae (SEM). (a, b) triangular and smooth accessory denticles that pass continuously into the trichomes of the pulvillus; the trichomes of the pulvillus transition continuously from branched to simple ones; the branching bristles of the fringe have a widened base, branch on the upper two‐thirds and are not in contact with the trichomes of the pulvillus, right mandible, top is ventral, (a) Lithobius (Ezembius) giganteus, (b) Disphaerobius loricatus; (c) Lithobius (Lithobius) nodulipes, the trichomes of the pulvillus transition from branching to simple ones with a break (arrow), right mandible, top is ventral; (d) Lithobius (Lithobius) dentatus, the transition of accessory denticles to the trichomes of the pulvillus is continuous, left mandible, right is dorsal; (e) Lithobius (Lithobius) fagei, the branching bristles are in contact with the trichomes of the pulvillus (arrow) and there is a trichome‐free strip between accessory denticles and trichomes of the pulvillus, left mandible, right is dorsal; (f) Lithobius (Lithobius) cyrtopus, the branching bristles of the fringe are not in contact with the trichomes of the pulvillus and the accessory denticles transition abruptly into the trichomes of the pulvillus, right mandible, top is ventral. Abbreviations: ad—Accessory denticles on mandibular teeth; fr—Fringe of branching bristles on mandibular gnathal edge; pu—Pulvillus on mandible; tpu—Trichomes on pulvillus

Figure 6.

Details of mandibular gnathal edge of Lithobiidae (SEM). (a) Lithobius (Lithobius) cyrtopus, the trichomes of the pulvillus transition from branching to simple with a break (arrow), right mandible; (b) Eupolybothrus (Eupolybothrus) grossipes, abrupt transition of accessory denticles into the trichomes of the pulvillus, right mandible, left is dorsal; (c) Lithobius (Lithobius) muticus, subtle increase in length of branching bristles ventrally, the branching bristles skirt the inner row of aciculae completely, left mandible; (d) Lithobius (Lithobius) erythrocephalus, subtle increase in length of branching bristles ventrally, the branching bristles skirt the inner row of aciculae until the penultimate acicula, left mandible; (e) Eupolybothrus (Eupolybothrus) grossipes, the bases of the ventralmost branching bristles are smooth without branches and skirt the inner row of aciculae until the penultimate acicula, right mandible, bottom is ventral; (f) Lithobius (Ezembius) giganteus, the branching bristles decrease in length from dorsal to ventral and skirt the inner row of aciculae until the penultimate acicula, left mandible, right is dorsal. Abbreviations: aci—Inner row of mandibular aciculae; aco—Outer row of mandibular aciculae; ad—Accessory denticles on mandibular teeth; fr—Fringe of branching bristles on mandibular gnathal edge; pu—Pulvillus on mandible; tpu—Trichomes on pulvillus

The pulvillus is a well‐differentiated lobe with trichomes (tpu), on the dorsalmost part of the mandibular gnathal edge (Figures 1c, 4a, 5, and 6a). The trichomes can be branched and/or simple (Figures 5 and 6a,b). If both of these types occur, there is a transition from branching to simple trichomes from proximal to distal (Figures 5a‐d,f and 6a). The transition is either continuous (Figures 5a,b,d,e) or interrupted (Figures 5c and 6a).

• dFringe of the mandibular branching bristles (fr)

The fringe of branching bristles extends along the entire margin of the gnathal edge in all examined lithobiid species (Figures 1c and 4a). From ventral (aciculae) to dorsal (pulvillus) there is at first a subtle increase in length of branching bristles followed by a decrease in length (Figures 1c, 4a, and 6c–f). Due to the decrease in length, the shaft gradually changes from relatively long and cylindrical to “scale‐like” (Figures 1c, 4a–e, 5c–e, 6a–e, 7b–e). This is not the case for L. giganteus and D. loricatus, in which the bases of the branching bristles only slightly broaden dorsally (Figures 5a,b and 7a,f). Dorsally, the branching bristles are in contact with the bases of the pulvillus' trichomes (Figure 5e) or are not (Figures 1c, 4a, 5a–d,f, and 6a,b). Ventrally, they usually fringe the inner row of aciculae completely (Figure 6c,d) or until the penultimate acicula of the inner row (Figure 6e,f). In some cases, a few branching bristles lie outside the fringe (e.g., L. forficatus, L. giganteus; Figure 7e,f). The branching bristles branch along the whole length of their shaft (Figures 1c, 2a, 4a, 6c‐e, and 7b‐e) or only partly, in the upper two‐thirds to one‐third, as displayed only in D. loricatus and L. giganteus (Figures 5a,b, 6f, and 7a,f). The back of the shaft is nonbranching and smooth (Figure 7b). In some species, the base of the ventralmost branching bristles is smooth (Figures 6d,e and 7c,d).

• eInternal surface of the mandibular gnathal edge

Figure 7.

Details of mandibular gnathal edge of Lithobiidae (SEM). (a) Disphaerobius loricatus, branching bristles branch along the upper third or two‐thirds of their shaft length, right mandible, right is ventral; (b) Lithobius (Monotarsobius) crassipes, branching bristles are relatively long and cylindrical ventrally, the back of the shaft is smooth without branches (arrows), right mandible, right is ventral; (c) Lithobius (Monotarsobius) burzenlandicus, branching bristles branch along their entire length (bases of ventralmost bristles smooth [arrows]), right mandible; (d) Lithobius (Ezembius) electus, left mandible, the bases of the ventralmost branching bristles are smooth (arrows); (e) Lithobius (Lithobius) forficatus, branching bristles decrease in length from ventral to dorsal, randomly distributed branching bristles (arrows), right mandible, right is ventral; (f) Lithobius (Ezembius) giganteus, arrow points to a randomly distributed branching bristle, left mandible, right is dorsal

The internal side of the mandibular gnathal edge may present (a) no spinulation (Figures 1d and 8a) or (b) a spinulation (sp) with branching bristles (Figures 8b and 9a‐d) or simple spines (Figures 8c–f and 9f) at the distal margin of the ventralmost tooth. Branching bristles branch mainly bilaterally (Figure 9a–d). Simple spines are long or stout and always tapering (Figures 8c–f and 9e,f). A strong similarity is noted for L. giganteus and D. loricatus with long simple spines clustering close together (Figure 8c–e). In E. grossipes, six of the nine spines display small branches uni‐ or bilaterally (Figure 9e). The number of branching bristles or simple spines varies between three (e.g., L. mutabilis; Figure 8b) and nine (e.g., E. grossipes; Figure 9e). Spinulation expands to the dorsal margin of the ventralmost tooth at most (Figure 8b‐f) except for L. forficatus, in which it reaches the neighbouring tooth as well (Figure 9a). The number of spines/bristles varies between specimens of the same species (e.g., L. dentatus from 3 to 7 [N = 6] and L. validus from 3 to 11 [N = 7]) as well as between right and left mandibles of an individual, for example, (3, 7) in L. giganteus (Figure 8c,d).

Figure 9.

Details of internal spinulation on mandibular gnathal edge of Lithobiidae (SEM). (a) Lithobius (Lithobius) forficatus, bilaterally branching bristles expanding beyond the ventralmost tooth (t1) to second ventralmost tooth (t2), right mandible, right is dorsal; (b) Lithobius (Lithobius) fagei, several bilaterally branching bristles, left mandible, right is ventral; (c) Lithobius (Lithobius) cyrtopus, two bilaterally branching bristles, left mandible, right is ventral; (d) Lithobius (Lithobius) mutabilis, three bilaterally branching bristles, left mandible, right is ventral; (e) Eupolybothrus (Eupolybothrus) grossipes, simple and branching spines, right mandible, right is dorsal, inset: Details of branching spines as highlighted with arrows in 9e; (f) Eupolybothrus (Leptopolybothrus) tridentinus, simple tapering spines, right mandible, right is dorsal

3.2. First maxillae

In all examined species, the first maxillae (Figure 10) correspond with the previous descriptions provided for the same family (Edgecombe et al., 2002; Edgecombe & Giribet, 2004; Koch & Edgecombe, 2008).

• aCoxosternites (cs)

Figure 10.

Overview of first maxillae of Lithobiidae. (a, b) Lithobius (Lithobius) dentatus (LM), (a) ventral, top is anterior, (b) dorsal, top is antero‐lateral; (c) Eupolybothrus (Eupolybothrus) grossipes, ventral view, top is anterior (SEM); (d) Lithobius (Lithobius) cyrtopus, left first maxillae, dorsal view, right is anterior (SEM). Abbreviations: Ba—basal article of the first maxillary telopodite; cp—First maxillary coxal projection; cs—First maxillary coxosternite; da—Distal article of the first maxillary telopodite; ms—Median suture of first maxillae; pb—Plumose bristles on first maxillae; sm—Sensilla microtrichodea on first maxillae; st—Sensilla trichodea on first maxillae; ti—Trichomes on inside of first maxillary distal article; tp—First maxillary telopodite

The median submargins of the maxillary coxosternites may show short and/or longer trichoid sensilla running parallel to the median suture (ms) (Figures 10c and 11). The trichoid sensilla can be randomly distributed (Figures 10c and 11a,b,d) or arranged in a single row (Figure 11c). The distribution is mainly asymmetric in terms of position, type and number of sensilla (Figure 10c and 11). The proximal part of the coxosternite may bear symmetrically arranged long trichoid sensilla (Figure 11c) or these may be absent (Figure 11a,b,d).

• bTelopodites

Figure 11.

First maxillary coxosternite of the genus Lithobius, ventral, top is anterior (SEM). (a) Lithobius (Ezembius) electus, randomly distributed short and relatively long trichoid sensilla on medial submargins of coxosternites; (b) Lithobius (Lithobius) forficatus, randomly distributed short and longer trichoid sensilla on medial submargins of coxosternites; (c) Lithobius (Lithobius) dentatus, short trichoid sensilla in a row along the medial submargins of coxosternites with two characteristically long proximal trichoid sensilla; (d) Lithobius (Lithobius) piceus, asymmetrically arranged short and longer trichoid sensilla at submargins of coxosternites. Abbreviations: ba—Basal article of the first maxillary telopodite; cp—First maxillary coxal projection; cs—First maxillary coxosternite; sm—Sensilla microtrichodea on first maxillae

The distal article (da) of the telopodite bears more than one row of plumose bristles (pb) (Figures 10, 12, and 16c) on the inner margins, and bears sensilla trichodea (st) on the submargins (Figures 10c,d, 12a‐e, and 16c). Some species present a few sensilla microtrichodea (sm) on the distal article, mainly dorso‐laterally (Figure 13a). These sensilla differ in number and location and show no bilateral symmetry (Figures 10c, 12a, and 16c). The inside of the distal article of the telopodite is equipped with several rows of mainly flattened and/or more rounded trichomes (ti) that may branch at the tip (Figures 10d and 13). The basal article (ba) may carry both sensilla trichodea and microtrichodea, most often on the dorsal side (Figures 10c, 12b‐d, and 16c). The articles of the telopodite and the coxal projection (cp) present a few pores placed individually or in clusters, mainly close to the socket of a sensillum trichodeum or microtrichodeum (Figures 12d, 14a,d, and 15c).

• cCoxal projections (cp)

Figure 12.

Overview and details of first maxillary telopodite of Lithobiidae (SEM). (a) Lithobius (Ezembius) electus, distal articles of telopodite bearing plumose bristles and randomly distributed abundant sensilla trichodea, antero‐ventral view; (b) Eupolybothrus (Eupolybothrus) grossipes, several rows of plumose bristles and sensilla trichodea on left distal article of telopodite, ventral view, right is posterior; (c, d) Lithobius (Monotarsobius) crassipes, (c) left distal article with plumose bristles and a few sensilla trichodea, arrow marks a sensillum microtrichodeum, ventral view, right is posterior; (d) closeup of 12c showing a pore (arrow); (e) Lithobius (Lithobius) cyrtopus, right distal article presenting plumose bristles and sensilla microtrichodea and trichodea, ventral view, right is posterior, Inset: Detail of sensilla microtrichodea; (f) Lithobius (Monotarsobius) franciscorum, plumose bristles on inner margin of telopodite with some s. trichodea. Abbreviations: ba—Basal article of the first maxillary telopodite; cp—First maxillary coxal projection; da—Distal article of the first maxillary telopodite; pb—Plumose bristles on first maxillae; sm—Sensilla microtrichodea on first maxillae; st—Sensilla trichodea on first maxillae

Figure 16.

Details of first maxillary coxal projection of the genus Lithobius (SEM). (a, b) Lithobius (Monotarsobius) crassipes, (a) distal margin with one row of plumose and feather‐like branching bristles and one row of sensilla trichodea, sensilla trichodea expanding further proximally compared to other trichomes, (b) plumose bristles and sensilla trichodea; (c, d) Lithobius (Lithobius) validus, (c) high number of different types of sensilla trichodea and sensilla microtrichodea and trichomes on the telopodite and coxal projection, ventro‐lateral view, (d) sensilla trichodea expand further proximally along the coxal projection. Abbreviations: ba—Basal article of the first maxillary telopodite; bb—Branching bristles on first maxillary coxal projection (feather‐/hassock‐like); cp—First maxillary coxal projection; cs—First maxillary coxosternite; da—Distal article of the first maxillary telopodite; pb—Plumose bristles on first maxillae; st—Sensilla trichodea on first maxillae

Figure 13.

Overview and details of the inner margin of first maxillary telopodite of Lithobiidae (SEM). (a) Lithobius (Lithobius) forficatus, flattened and tip‐branching trichomes; bottom part shows distal part of hypopharynx (lips forming median crest); sensilla microtrichodea are distributed on the dorso‐lateral margin of the distal article, lateral, right is anterior; (b) Lithobius (Lithobius) cyrtopus, detail of inner margin of distal article showing flattened and more rounded trichomes with a few branches at their tips (closeup of Figure 10d), dorsal, right is anterior; (c–g) flattened trichomes with branched tips, (c) Lithobius (Ezembius) electus, (d) Eupolybothrus (Eupolybothrus) tridentinus, (e) Lithobius (Lithobius) erythrocephalus, (f) Lithobius (Monotarsobius) austriacus, (g) Lithobius (Lithobius) tenebrosus. Abbreviations: da—Distal article of the first maxillary telopodite; sm—Sensilla microtrichodea on first maxillae; ti—Trichomes on inside of first maxillary distal article

Figure 14.

Details of sensilla trichodea and microtrichodea on first maxillae of Lithobiidae (SEM). (a, b) Lithobius (Lithobius) validus, (a) sensillum trichodeum surrounded by a single pore or cluster of pores (arrows), (b) seventeen sensilla microtrichodea on the lateral margin of coxosternite; (c–f) cluster of sensilla microtrichodea with different numbers and arrangements on lateral margin of coxosternite, (c) Eupolybothrus (Eupolybothrus) grossipes, (d) Lithobius (Lithobius) agilis, pores (arrow) surrounding sensilla microtrichodea, (e) Lithobius (Lithobius) dentatus, (f) Lithobius (Lithobius) mutabilis. Abbreviation: st—Sensilla trichodea on first maxillae

Figure 15.

Details of first maxillary coxal projection of the genus Lithobius (SEM). (a) Lithobius (Lithobius) cyrtopus, a distal band of branching bristles changing from plumose (one row) to feather‐like (more rows) from medial to lateral, proximally following rows of sensilla trichodea and microtrichodea; (b) Lithobius (Ezembius) electus, plumose bristles change to feather‐like branching bristles from medial to lateral on the distal part of the coxal projection; (c) Lithobius (Lithobius) mutabilis, sensilla trichodea and pores (arrow), Insets: Left, broken sensillum trichodeum exposing hollow shaft lumen; right, sockets of broken sensilla trichodea; (d) Lithobius (Lithobius) validus; (e) Lithobius (Lithobius) erythrocephalus; (f) Lithobius (Monotarsobius) austriacus, hassock‐like branching bristle, Inset: Lithobius (Monotarsobius) aeruginosus, terminal shaft pore (arrow) of a sensillum trichodeum. Abbreviations: bb—Branching bristles on first maxillary coxal projection (feather‐/hassock‐like); cp—First maxillary coxal projection; pb—Plumose bristles on first maxillae; sm—Sensilla microtrichodea on first maxillae; st—Sensilla trichodea on first maxillae

The lateral borders of the coxal projections present some sensilla microtrichodea (sm) (Figures 10c, 11 a–c, and 14b–f). Their number varies between four (e.g., L. lapidicola) and seventeen (e.g., L. validus; Figure 14b). They are usually arranged in an off‐set pattern more like a cluster (Figure 14b–f). The coxal projection is equipped with different types of branching bristles and setae (Figure 10c). The distal margin is framed by branching bristles that are plumose on the lateral side (Figure 16b) and feather‐/hassock‐like (Figure 15d,e,f) on the medial side (see L. cyrtopus and L. electus as examples; Figure 15a,b). These branching bristles may occur in one or more rows (Figures 15a,b,d and 16). Submarginally are one or more rows of sensilla trichodea (Figures 15a–e and 16a,c,d) and sensilla microtrichodea (Figures 15a,b,e). Sensilla microtrichodea are absent in some species (e.g., L. validus and all investigated species of the subgenus Monotarsobius; Figure 16a,c,d). Smaller species tend to have fewer trichomes and setae in general (compare L. crassipes and L. validus; Figures 16a,d). In E. grossipes and E. tridentinus, the coxal projection is densely covered with multiple rows of plumose bristles, showing in between sensilla trichodea and microtrichodea (Figure 17b–d). In D. loricatus, the coxal projection bears multiple rows of stout plumose bristles and scattered sensilla trichodea and microtrichodea (Figure 17a). The sensilla trichodea expand more posteriorly along the coxal projection than the other types of trichomes and setae (Figures 15a,b,e, 16a,c,d, and 17a–c). The trichomes and setae mainly cover a half to three‐quarters of the coxal projection (Figures 15a‐e, 16a,c,d, and 17a–c).

Figure 17.

Details of first maxillary coxal projection of Lithobiidae (SEM). (a) Disphaerobius loricatus, several rows of stout plumose bristles and scattered sensilla trichodea/microtrichodea; (b) Eupolybothrus (Eupolybothrus) grossipes, coxal projection densely covered with plumose bristles and randomly distributed sensilla trichodea/microtrichodea; (c, d) Eupolybothrus (Leptopolybothrus) tridentinus, (c) several rows of plumose and feather‐like branching bristles with sensilla trichodea in between, (d) detail of Figure 17c. Abbreviations: bb—Branching bristles on first maxillary coxal projection (feather‐/hassock‐like); cp—First maxillary coxal projection; pb—Plumose bristles on first maxillae; st—Sensilla trichodea on first maxillae

3.3. Morphological characters and observations

3.3.1. Head

1. Ocelli: (0) compound eye; (1) cluster of ocelli, differentiated into posterior major ocellus and seriate ocelli; (2) single ocellus; (3) blind. (see Koch & Edgecombe, 2008, char. 1)

2. Arrangement of ocelli: (0) irregular; (1) one row; (2) two or more rows.

Ocelli may be arranged irregularly in D. loricatus (Farzalieva & Zalesskaja, 2002, Figure 12), L. giganteus (Sseliwanoff, 1881, p. 15), L. calcaratus (Eason, 1964, Figure 429), L. macilentus (Koren, 1992, Figure 20b) and L. curtipes (Eason, 1964, Figure 453). L. burzenlandicus is polymorphic, expressing both states (0) and (1) (Matic, 1966, p. 226). For all other Lithobiidae a regular arrangement in one or more rows is observed (states (1) and (2)). Coding is inapplicable for Scutigeromorpha and Henicopidae.

Figure 20.

Strict consensus of two shortest cladograms under implied weights (k = 4–5). Coloured boxes indicate the four subgenera of the genus Lithobius and the Ethopolyinae as follow: red—Lithobius; blue—Sigibius; grey—Monotarsobius; yellow—Ezembius; green—Ethopolyinae

3. Size of posterior ocellus relative to seriate ocelli: (0) posterior ocellus smaller or equal in size; (1) posterior ocellus larger.

State (0) is shared by species of the subgenera Sigibius, Monotarsobius (except for L. holstii [Eason, 1973, Figure 52] and L. crassipes [Koren, 1992, Figure 34b]), three species within the subgenus Lithobius (L. calcaratus, Eason, 1964, Figure 429; L. carinatus, Koch, 1862, Table II 42 and new observation; L. macilentus, Koren, 1992, Figure 20b), H. anodus (Koren, 1992, p. 28), P. patriarchalis and P. megaloporus (both new observation). Coding is inapplicable for Scutigeromorpha and Henicopidae.

4. Tömösvary organ: (0) anteroventral to ocellar cluster/compound eye; (1) on cephalic pleurite on ventral side of head; (2) below inferior row of ocelli. (combination of Koch & Edgecombe, 2008, char. 5 and Edgecombe & Giribet, 2004, char. 143).

5. Size of Tömösvary organ relative to posterior ocellus: (0) same size or larger than posterior ocellus; (1) smaller than posterior ocellus.

State (0) is observed for L. macilentus (Koren, 1992, Figure 20b), L. aeruignosus (Koren, 1992, Figure 36d,e), L. franciscorum (Dányi & Tuf, 2012, p. 18, Figure 2) and L. burzenlandicus (new observation) only. Coding is inapplicable for Scutigeromorpha and Henicopidae.

6. Antennal articulation: (0) 15 articles; (1) 17 to 24 articles; (2) 25 or more articles. (see Edgecombe et al., 2002, char. 3; adjusted)

State (0) from character 3 in Edgecombe et al. (2002) was divided into two states (1) and (2) to separate species of the subgenera Monotarsobius and Ezembius (all state (1)) from Lithobius and Sigibius (all state (2)). D. loricatus (Farzalieva & Zalesskaja, 2002, p. 267), P. megaloporus (Crabill, 1950, p. 10), H. plumatus (Zalesskaja, 1978, p. 47), A. scabrior (Edgecombe & Hollington, 2002, p. 107), B. multidentatus (new observation) and the henicopid D. biscupis (Ribaut, 1923, p. 24) possess 17 to 24 antennal articles (state (1)).

7. Lateral margin of the cephalic plate interrupted at anterior limit of marginal ridge: (0) not interrupted; (1) interrupted. (see Koch & Edgecombe, 2008, char. 3)

Absence of interruption of the lateral marginal ridge is also observed for D. loricatus (Farzalieva & Zalesskaja, 2002, Figure 1 and new observation) besides P. megaloporus and B. multidentatus (see Koch & Edgecombe, 2008). Coding is inapplicable for Scutigeromorpha.

8. Median furrow on cephalic plate: (0) deep and continuous to transverse suture; (1) terminating in front of transverse suture. (see Koch & Edgecombe, 2008, char. 2)

9. Pleurites of forcipular segment connected ventrally, forming a continuous band between forcipular coxosternite and sternite of first pedigerous segment: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 6)

10. Coxosternal teeth: (0) 4 + 4 spine bristles; (1) 2 + 2 to 4 + 4 teeth; (2) 5 + 5 or more teeth. (see Koch & Edgecombe, 2008, char. 7; adjusted)

The number of coxosternal teeth varies between 4 + 4 and 10 + 9 in adult individuals of A. scabrior (Edgecombe & Hollington, 2002, p. 110) resulting in a polymorphic character coding (1&2).

11. Size of coxosternal teeth: (0) vestigial to small; (1) well developed.

All Lithobiomorpha have well developed coxosternal teeth except for three species of the subgenus Lithobius (L. calcaratus, Eason, 1964, p. 232; L. castaneus, Koren, 1992, Figure 23c; L. muticus, Eason, 1964, p. 216) and H. anodus (Koren, 1992 Figure 10c,d). Coding is inapplicable for Scutigeromorpha.

12. Shape of dental margin of forcipular coxosternite: (0) convex; (1) straight, transverse; (2) V‐shaped.

A straight (1) or V‐shaped (2) dental margin is present in most Lithobiidae and Henicopidae whereas the convex state (0) is only shared by the species D. loricatus (Farzalieva & Zalesskaja, 2002, Figure 3), L. giganteus (Eason, 1986, Figures 3, 4) and L. nodulipes (Koren, 1992, Figure 14b).

13. Shape of anterior third of forcipular coxosternal lateral margins: (0) convex; (1) concave; (2) straight.

While the Henicopidae show concave or straight lateral margins (states (1) and (2)), most of the Lithobiidae have dominant coxosternal shoulders and if shoulders are absent, the margin is convex (state (0)). Within Lithobiidae, only species of the genus Eupolybothrus and B. multidentatus share a concave lateral margin (state (1), new observations).

14. Porodont: (0) absent; (1) translucent, seta‐like porodont; (2) thickened, tooth‐like. (see Koch & Edgecombe, 2008, char. 8)

L. giganteus and L. trebinjanus are the only Lithobius species showing a thickened, tooth‐like porodont (Eason, 1986, Figure 3; Eason, 1983, Figure 23).

15. Proportion of forcipular tarsungulum: (0) pretarsal section of approximately equal length to tarsal section; (1) pretarsal section much longer than tarsal section. (see Edgecombe et al., 2002, char. 15)

16. Notches on labral side piece: (0) absent (unidentate labrum); (1) present (tridentate labrum). (see Koch & Edgecombe, 2008, char. 4)

3.3.2. Epipharynx

17. “Bottle‐shaped” glandular shafts at the border between labral and clypeal part of epipharynx: (0) absent; (1) present with one distinct regular row; (2) present with more than one regular or irregular rows. (see Ganske et al., 2018, char. 1; adjusted)

State (0) was added to the original description to accommodate Scutigeromorpha in the present study.

18. Number of transverse bulge(s) at border between labral and clypeal parts of epipharynx: (0) absent; (1) one; (2) two. (see Ganske et al., 2018, char. 3)

19. Lateral expansion of median sensilla cluster of epipharynx: (0) isolated from the lateral spine fields; (1) partly overlapping with the lateral spine fields. (see Ganske et al., 2018, char. 7)

Sensilla cluster 5 and spine cluster 1 of Scutigeromorpha (s5, sp1, Koch & Edgecombe, 2006) were homologized with the median sensilla cluster and the lateral field of spines in Lithobiomorpha (msc, lsp, Koch & Edgecombe, 2008). S5 is always isolated from sp1 in Scutigeromorpha (state (0)).

20. Shape of branching bristles on labral flap of epipharynx: (0) lateral to medial transition from plumose to “fan‐shaped” bristles; (1) “fan‐shaped” bristles only; (2) plumose bristles only; (3) simple bristles only. (see Ganske et al., 2018, char. 6)

21. Labral bristle bands of epipharynx: (0) bristle bands continuous across transverse bulge; (1) distinct break in bristle bands proximal and distal to transverse bulge. (see Koch & Edgecombe, 2008, char. 31 and Ganske et al., 2018, char. 2)

22. Lateral field of spines and pores on clypeal part of epipharynx: (0) absent; (1) cluster of numerous scattered spines; (2) oblique rows of single spines; (3) a few small groups of branching spines. (see Koch & Edgecombe, 2008, char. 32)

23. Length of spines on lateral clypeal part of epipharynx: (0) mostly long, tapering; (1) consistently short, conical. (see Koch & Edgecombe, 2008, char. 33)

24. Median field of branching spines immediately proximal to the border between labral and clypeal parts of epipharynx: (0) rhomboid, widening medially; (1) widening laterally; (2) subequal width medially and laterally. (see Ganske et al., 2018, char. 5)

25. Direction of distal and proximal furrowed margins of transverse bulge or transverse bulges on epipharynx: (0) parallel; (1) nonparallel. (see Ganske et al., 2018, char. 4)

Coding is inapplicable for species without a transverse bulge (char. 18, state (0)).

3.3.3. Hypopharynx

26. Cluster of proximomedially directed spines at lateral margin of pharyngeal plate: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 40)

27. Shape of hypopharynx: (0) elongate tongue with median excavation; (1) short outgrowth with median crest. (see Koch & Edgecombe, 2008, char. 38)

28. Differentiation of spines flanking median crest of hypopharynx: (0) intergrading rows of branching bristles; (1) single outer row of simple flattened spines with abrupt transition to multifurcating inner rows of branching bristles. (see Koch & Edgecombe, 2008, char. 39 and Ganske et al., 2018, char. 8)

3.3.4. Mandibles

29. Band or isolated cluster of trichomes extending from pulvillus onto outer face of mandible: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 13)

The presence of branching bristles or simple spines for Lithobiidae described by Koch and Edgecombe (2008) is verified by data of the present study (state (0); Figures 1, 4a, 5, and 6a,b).

30. Fringe of branching bristles on mandible: (0) extends along entire gnathal margin, skirting aciculae; (1) terminates at aciculae. (see Koch & Edgecombe, 2008, char. 11)

All examined Lithobius‐species show a fringe of branching bristles skirting the aciculae (state (0); Figures 1c, 2a, 4a, and 6c–f) as observed by Koch and Edgecombe (2008) for Lithobiomorpha (except for Anopsobius neozelanicus, Dichelobius bicuspis, Zygethobius pontis).

31. Spinulation on internal side of margin to mandibular teeth on mandible: (0) absent; (1) present (branching bristles, simple/branching spines).

All studied species of the subgenus Sigibius, some species of the subgenus Lithobius (Figures 1d, 8a) and Henicopidae (data available for D. gigantea and H. washpoolensis only) lack an internal spinulation (state (0)). For species of the subgenus Monotarsobius information are missing for L. austriacus, L. curtipes and L. holstii but state (0) applies to L. aeruginosus whereas L. crassipes and L. franciscorum possess an internal spinulation (state (1)).

32. Continuous transition of accessory denticles into trichomes of pulvillus: (0) absent (trichome‐free stripe); (1) present (either intergrading or with abrupt elongation). (Edgecombe et al., 2002, char. 28; revised, see discussion)

A trichome‐free stripe (state (0)) is present in a few lithobiids (e.g., L. fagei, Figure 5e) and the henicopid H. washpoolensis (Edgecombe & Hollington, 2005, Figure 29) only. Information is missing for L. carinatus, L. macilentus, L. austriacus, and L. curtipes. Coding is inapplicable for Scutigeromorpha.

3.3.5. First maxillae

33. Basal joint of telopodite of first maxillae fused on inner side to coxal projection: (0) telopodite distinctly demarcated; (1) telopodite fused to adjacent part of coxa. (see Koch & Edgecombe, 2008, char. 14)

All examined Lithobiidae (Figures 10a, c and 11), Anopsobiinae and Scutigeromorpha share state (0) (information missing for L. calcaratus and L. muticus).

34. Structure of setae on coxal projection of first maxillae: (0) mostly simple, with cluster of distally‐tufted setae at apex; (1) all simple; (2) a few lacinate setae amid mostly simple setae; (3) mix of plumose and simple setae. (see Koch & Edgecombe, 2008, char. 15)

State (3) is shared by all Lithobiidae (Figures 15, 16, 17; information missing for L. calcaratus, L. macilentus, L. muticus, L. nodulipes, L. pelidnus, L. trebinjanus).

35. Sensilla microtrichoidea on coxal projection of first maxillae: (0) absent; (1) present.

Sensilla microtrichodea are absent throughout the investigated species of the subgenus Monotarsobius (e.g., L. crassipes, Figure 16a). Information is missing for some species (see Table 1).

3.3.6. Segments

36. Tergite 1 narrower than head and tergite 3: (0) absent; (1) present.

Eason (1964) noted that the width proportion of T1 to the head and T3 differs between lithobiomorphs. A narrower tergite 1 (state (1)) is widely represented within Lithobiidae, whereas state (0) is rare, especially in species of the genus Lithobius.

37. Projections of posterolateral corners of tergites: (0) all rounded; (1) T9 rounded, T11, T13 with projections; (2) T9, T11, T13 with projections; (3) T6, T7, T9, T11, T13 with projections. (see Koch & Edgecombe, 2008, char. 9; adjusted)

States (1) to (3) were previously summarized in one state (“some angular or toothed”) but were defined now to separate groups within Lithobiidae and Lithobius. Three species of the genus Lithobius (all belonging to the subgenus Lithobius) share state (1) (L. cyrtopus, Latzel, 1880, p. 94; L. fagei, Demange, 1961, p. 279; L. lapidicola, Eason, 1964, p. 208). State (3) is shared by all examined species of the genus Eupolybothrus, B. multidentatus (new observation), L. validus (Koren, 1992, p. 38) and Z. pontis (Chamberlin, 1911, p. 34). Representatives of the subgenera Monotarsobius and Sigibius as well as Scutigeromorpha possess rounded tergites without projections (state (0)), and this is also the case of several species of the subgenus Lithobius and most Henicopidae.

38. Spiracle on first leg‐bearing segment: (0) present; (1) absent. (see Koch & Edgecombe, 2008, char. 10)

39. Anal pores in adult: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 29)

3.3.7. Legs

40. Tarsus of legs 1–12: (0) divided into two tarsal articles; (1) undivided. (see Koch & Edgecombe, 2008, char. 21)

41. Insertion of anterior accessory claw: (0) dorsolaterally on main claw; (1) ventrolaterally on main claw. (see Koch & Edgecombe, 2008, char. 22)

42. Coxal pore arrangement: (0) in single file; (1) scattered pore field. (see Koch & Edgecombe, 2008, char. 19)

43. Serial distribution of coxal pores: (0) present on segments 14 and 15 only; (1) present on segments 12–15; (2) present on segments 11–15. (see Koch & Edgecombe, 2008, char. 18)

44. Coxa of leg 15 with long, lobate process ending in a spine: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 16)

45. Coxa of leg 15 with distal spurs: (0) spurs absent; (1) dorsal anterior spur only; (2) dorsal and ventral anterior spurs only; (3) dorsal median spur only; (4) dorsal median and ventral anterior spurs only; (5) spur developed as spine bristle, one ventrally.

States (0) to (2) describe most of the Lithobiidae and Henicopini. A single dorsal median spur (state (3)) is shared by L. electus (Silvestri, 1935, p. 195 and new observation), L. franciscorum (Dányi & Tuf, 2012, Tables 1–3), P. megaloporus (new observation) and D. bicuspis (new observation). Both dorsal median and ventral anterior spurs are present in N. aztecus (new observation) and L. castaneus (Koren, 1992, p. 80) only (state (4)). State (5) applies to Scutigeromorpha only.

46. Prefemur of leg 15 with distal spurs: (0) spurs absent; (1) single ventral spur; (2) dorsal and ventral anterior, median, posterior spurs encircling distal end of prefemur; (3) spurs developed as spine bristles, one ventrally and two dorsally; (4) dorsal and ventral median, posterior spurs only; (5) dorsal median, posterior and ventral anterior, median, posterior spurs only; (6) dorsal and ventral anterior, median, posterior spurs with accessory posterior prefemoral spur. (see Koch & Edgecombe, 2008, char. 17; states (4) to (6) added)

State (4) to (6) are added to accommodate several species of the genus Lithobius with deviating spinulation to state (2) used in the original character description. State (4) is shared by L. calcaratus (Eason, 1964, p. 232), L. burzenlandicus (Matic, 1966, p. 228) and L. trebinjanus (Eason, 1983, p. 140). H. anodus (Koren, 1992, p. 30), L. carinatus (new observation), L. nodulipes (Koren, 1992, p. 51), L. tenebrosus (Koren, 1992, p. 54), L. aeruginosus (Koren, 1992, p. 127), L. austriacus (Koren, 1992, p. 126) and L. microps (Brolemann, 1930, p. 323) share state (5). State (6) occurs in L. lapidicola (Eason, 1964, p. 208) only.

47. Distal spinose projection on tibiae: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 20)

48. Sulci on 14th/15th legs of males: (0) absent; (1) present on 15th leg only; (2) present on 14th and 15th legs.

The absence of sulci (0) is shared by Scutigeromorpha, Henicopidae, species of the subgenera Monotarsobius and Sigibius and several species of the subgenus Lithobius. L. cyrtopus (Latzel, 1880, p. 94), L. latro (Koren, 1992, Figure 26a), N. aztecus and P. megaloporus (both new observations) present sulci only on 15th legs (state (1)). All representatives of the genus Eupolybothrus as well as H. anodus (Koren, 1992, p. 29), P. patriarchalis (new observation) and several species of the subgenus Lithobius as well as D. loricatus and L. giganteus (both Farzalieva & Zalesskaja, 2002, p. 266) share the presence of sulci on both 14th and 15th legs (state (2)).

49. Sulci on femora, tibiae of 14th/15th legs in male: (0) present on tibiae of 14th and/or 15th leg only; (1) present on femora of 14th and/or 15th leg only; (2) present on tibiae and femora of 14th and/or 15th leg.

If sulci are present on the 14th and/or 15th legs, they occur on both tibiae and femora or on one of those podomeres only. State (0) is recorded in H. anodus (Koren, 1992, p. 29) and L. mutabilis (Koren, 1992, Figure 33a). Sulci on femora only (state (1)) are for example observed in L. forficatus (Koren, 1992, Figure 13b), D. loricatus and L. giganteus (both Farzalieva & Zalesskaja, 2002, p. 266 and Farzalieva et al., 2017, Table 1). State (2) applies to L. pelidnus (Matic, 1966, p. 171) and E. grossipes (Koren, 1992, p. 20). Coded as inapplicable for species in which sulci are absent (char. 48, state (0)).

50. 15th prefemora with a ventral posterior spur developed as a small trifid spur: (0) absent; (1) present.

A small trifid spur is present in L. austriacus only (Koren, 1992, p. 126).

51. Male 15th tibiae or femora with dorsal/posterodorsal distal wart‐like outgrowth: (0) absent; (1) present.

State (1) is shared by L. calcaratus (Eason, 1964, Figure 433; Matic, 1966, Figure 101), L. nodulipes (Matic, 1966, Figure 51d; Koren, 1992, Figure 14a) and L. curtipes (Eason, 1964, Figure 456; Matic, 1966, Figure 86a‐c).

52. Male 15th femora with a dorsal distal group of club‐like setae: (0) absent; (1) present.

A group of club‐like setae is present in L. franciscorum only (Dányi & Tuf, 2012, Figures 26–27).

53. Male 15th femora or prefemora with distal knob: (0) absent; (1) present.

Present in D. loricatus (Farzalieva & Zalesskaja, 2002, Figure 4), E. fasciatus (Eason, 1970, Figure 1) and L. pelidnus (Koren, 1992, Figure 27f) only.

54. Male 14th/15th tibiae/femora with a circular protuberance covered with setae: (0) absent; (1) present.

Present in L. muticus (Koren, 1992, Figure 24b) and L. pelidnus (Koren, 1992, Figure 27d) only.

55. Length of posteroventral spine on pretarsus: (0) short, <25% length of main claw; (1) long, 50% length of main claw. (see Koch & Edgecombe, 2008, char. 23)

3.3.8. Genitalia

56. Number of spurs on female gonopod: (0) 2; (1) 3 or more. (see Edgecombe et al., 2002, char. 47; state (1) adjusted)

State (1) was adjusted to “3 or more spurs on female gonopod” compared to the original description due to lithobiid species which show also four or five spurs (e.g., L. franciscorum, Dányi & Tuf, 2012, Figure 14; A. scabrior, Edgecombe & Hollington, 2002, p. 114; L. piceus, Eason, 1964, p. 200). L. muticus (Koren, 1992, p. 83) and L. nodulipes (Koren, 1992, p. 51) may possess two or three spurs and were coded as polymorphic (0&1).

57. First article of female gonopod extended as a short process: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 27)

58. Claw of female gonopod: (0) simple, unipartite; (1) tripartite, with an accessory denticle on each side of main claw; (2) bipartite. (see Koch & Edgecombe, 2008, char. 28; state (2) added)

To include L. calcaratus (Eason, 1964, Figure 437), L. castaneus (Koren, 1992, p. 79) and L. piceus (Koren, 1992, Figure 12e; Matic, 1966, Figure 47E), state (2) was added to the original character description. Coding is inapplicable for Scutigeromorpha.

59. Row/group of dorsomedial setae on gonopodal first article of female: (0) absent; (1) present.

The subgenera Sigibius and Ezembius share the absence of dorsomedial setae (state (0)) together with several species of the subgenera Lithobius and Monotarsobius as well as A. scabrior (new observation), D. loricatus (Farzalieva & Zalesskaja, 2002, Figure 13), B. multidentatus, N. aztecus, P. megaloporus (all new observations) and Henicopidae (state (0)). Coding is inapplicable for Scutigeromorpha.

60. Segmentation of male gonopod on first genital segment: (0) style‐like, without apparent segmentation; (1) four segments with a seta‐like terminal process bearing spines at its base; (2) small gonopod with single segment or minute second segment. (see Koch & Edgecombe, 2008, char. 25)

61. Second genital sternite of male with setae: (0) absent; (1) present.

The presence of setae on the second genital sternite of males is rare within Lithobiidae (e.g., A. scabrior, Edgecombe & Hollington, 2002, p. 115) and Lithobius (e.g., L. muticus, Eason, 1964, p. 218; L. forficatus, Eason, 1964, p. 196). Coding is inapplicable for Scutigeromorpha.

62. Dense concentration of short setae on posterior part of sternite and coxa 15 of male: (0) absent; (1) present. (see Koch & Edgecombe, 2008, char. 26)

A dense concentration of short setae on the posterior part of the sternite and the 15th coxa of males was coded present (state (1)) for L. giganteus and H. plumatus in Koch and Edgecombe (2008). We observed the presence of those setae in L. electus (Silvestri, 1935, Figure 13 and new observation) as well. Interestingly, these setae are absent in males of D. loricatus, but were described for its congener Disphaerobius svenhedini (Verhoeff, 1934) (Farzalieva et al., 2017, Figure 8).

3.4. Phylogenetic analyses

One‐hundred and thirty‐eight shortest cladograms of 303 steps were obtained under equal weights (consistency index 0.265, retention index 0.541) using the matrix with 62 morphological characters (Table 2). The strict consensus tree is shown in Figure 18 and all characters are optimized on one of the 138 minimal‐length trees (unambiguous optimizations; Figure 19). Under implied weights, the same two shortest trees were retrieved for k = 4 and 5 (see consensus tree of k = 4 in Figure 20 with a fit of 21.2880). The consensus tree constructed from 1,000 bootstrap trees under maximum likelihood analysis with a log‐likelihood of −1,437.394991 is illustrated in Figure 21.

Figure 18.

Strict consensus of 138 minimal‐length cladograms under equal weights. Numbers above branches are jackknife frequencies ≥50% and numbers below branches are Bremer support values. Coloured boxes indicate the four subgenera of the genus Lithobius and the Ethopolyinae as following: red—Lithobius; blue—Sigibius; grey—Monotarsobius; yellow—Ezembius; green—Ethopolyinae

Figure 19.

One of 138 shortest cladograms under equal weights showing character optimisation. Only unambiguous changes are shown, with contradicted nodes collapsed after optimizing on a fully resolved cladogram. Black circles: Non‐homoplastic changes; white circles: Homoplastic changes. Characters and states are numbered as in the text. Mandibular and first maxillary characters are highlighted in bold and italics. Coloured boxes indicate the four subgenera of the genus Lithobius and the Ethopolyinae as follow: red—Lithobius; blue—Sigibius; grey—Monotarsobius; yellow—Ezembius; green—Ethopolyinae

Figure 21.

Maximum likelihood consensus tree. Numbers above nodes are ultrafast bootstrap support values ≥75%. Coloured boxes indicate the four subgenera of the genus Lithobius and the Ethopolyinae as follow: Red—Lithobius; blue—Sigibius; grey—Monotarsobius; yellow—Ezembius; green—Ethopolyinae

Although Lithobiidae and Henicopidae are rendered monophyletic in the ML analysis (Ultrafast Bootstrap 100%; Figure 21) and under implied weights (MP; Figure 20), the tree under equal weights parsimony shows a different topology in placing Anopsobiinae s. str. as sister group to Lithobiidae and thus recovering Henicopidae as paraphyletic (Figure 18). Monophyly of Lithobiidae is supported by the presence of trichomes on the pulvillus (character 29, state (1)) and a mix of plumose and simple setae on the coxal projection (character 34, state (3)) on the mandible (Figure 19).

The genus Lithobius and its subgenera are resolved as nonmonophyletic except for the subgenus Sigibus based on equal weights (Figure 18). Groups that conflict with monophyly of Lithobius and most of its subgenera are partly supported by characters of the mandibles (character 31, state (1); character 32, state (0)) and first maxillae (character 35, state (1); Figure 19). A close relationship of the two Monotarsobius‐species L. austriacus and L. aeruginosus is supported with 51% jackknife and 99% ultrafast bootstrap frequencies in the MP and ML consensus trees, respectively (Figures 18 and 21), but not under implied weights (Figure 20). Ten pairs of Lithobius‐species are resolved as sister taxa with rather high ultrafast bootstrap frequencies in the ML tree (L. crassipes/L. holstii 80%, L. lapidicola/L. tenebrosus 91%, L. curtipes/L. franciscorum 80%; L. cyrtopus/L. pelidnus 98%, L. pyrenaicus/L. muticus 85%; Figure 20). The clades L. cyrtopus/L. pelidnus and L. muticus/L. pyrenaicus were additionally supported under equal weights (Bremer support 1 each; Figure 18; character 35, state (1); Figure 19) and L. crassipes/L. holstii under implied weights (Figure 20). L. giganteus and D. loricatus are consistently resolved as sister taxa in all trees (MP: Jackknife 69%, Bremer support 3; ML: Ultrafast Bootstrap 99%; Figures 18 and 21), but not in an alternative hypothesis based on implied weights (Figure 20). H. plumatus being sister taxon to L. giganteus and D. loricatus is also supported (MP: Jackknife 64%, Bremer 4; ML: Ultrafast Bootstrap 99%; except for implied weighting; Figures 18, 20, 21).

Based on implied weighting Ethopolyinae is monophyletic, but this is not the case under equal weights or ML (Figures 18, 20, 21). The Eupolybothrus‐clade is supported as a monophylum in all trees obtained (MP: Jacknife 58%, Bremer 3; ML: Ultrafast Bootstrap 99%; Figures 18, 20, 21). Some taxa are notably sensitive to character weights, such as Pseudolithobius megaloporus, which is variably nested deeply within Lithobiidae/Lithobius (equal weights parsimony) or is sister group of Lithobiinae (implied weights parsimony) (Figures 18 and 20). The latter is consistent with the long‐standing classification of Pseudolithobiinae as a separate subfamily, whereas the former is not.

4. DISCUSSION

This study presents a descriptive and comparative overview of the mandibular and first maxillary microstructures of 33 species of the genus Lithobius and four species of the genera Eupolybothrus, Neolithobius and Disphaerobius. The extensive examination of the specimens with scanning electron microscopy revealed similarities and differences of the structures. Some of the differences are intergradational between the species and cannot be defined as separate character states. There are however a few exceptions, which we here discuss and compare with results from previous studies. Furthermore, we provide phylogenetic analyses including Henicopidae and Scutigeromorpha.

4.1. Mandibles

In contrast to the family Henicopidae, which displays either a single row or two rows of aciculae (Edgecombe, 2004a [char. 50]; Edgecombe & Giribet, 2004 [char. 151]), the examined species of the family Lithobiidae showed a consistent number of two rows of aciculae (see also Edgecombe & Giribet, 2004; Edgecombe & Hollington, 2002). As reported by Edgecombe et al. (2002) and Edgecombe (2004a), the number of aciculae varies in the henicopid species of genera Lamyctes Meinert, 1868; Paralymyctes Pocock, 1901 and species of the subgenus Haasiella Pocock, 1901. We observed the same for the examined lithobiid species and further noticed a variation between conspecifics, and even between left and right mandibles of the same individual. Previous studies revealed aciculae to be consistently bipinnulate for all Lithobiidae (Edgecombe et al., 2002; Edgecombe & Giribet, 2004; Edgecombe & Hollington, 2002) which is confirmed by data in the present study, except for Eupolybothrus tridentinus, which shows a few apinnulate aciculae. This is in contrast to Henicopidae which have bipinnulate, pinnulate (pinnules on dorsal edge only) or apinnulate aciculae (Edgecombe et al., 2002 [char. 22]). In any case, it is not thus far possible to define character states for Lithobiidae from the aciculae and pinnules.

We confirm the presence of a distinctive fringe of branching bristles, evenly grading in their structure, along the mandibular gnathal edge for the examined Lithobiidae (Koch & Edgecombe, 2008 [char. 18]; Edgecombe et al., 2002 [char. 25]; Edgecombe & Giribet, 2004 [char. 127]) (Figure 1c). The fringe decreases in width from ventral to dorsal (see also Edgecombe et al., 2002) but we also noticed a subtle increase in size of the ventralmost bristles. In contrast to species of the henicopid genera Anopsobius Silvestri, 1899; Zygethobius Chamberlin, 1903; Dichelobius Attems, 1911 and Shikokuobius Shinohara, 1982, in which the fringe of branching bristles terminates at the aciculae, it always skirts the inner row of aciculae completely or at least largely in Lithobiidae (Figure 6c–f; Edgecombe et al., 2002 [char. 23]; Edgecombe & Giribet, 2004 [char. 125]). The fringe narrows dorsally and is not developed along the whole width of the trichomes of the pulvillus in Lithobiidae and most Henicopidae (Edgecombe et al., 2002 [char. 26]), which is verified by our results. Edgecombe et al. (2002) described (but did not illustrate) smooth bases of ventral branching bristles for L. variegatus rubriceps Newport, 1845. This is here confirmed in other species of Lithobius (e.g., Figures 6d,e and 7c,d). In contrast to other lithobiids, the dorsal branching bristles of L. giganteus and D. loricatus are not “scale‐like” but show a broader base (Figure 5a,b) as was described and illustrated for the henicopid Zygethobius pontis Chamberlin, 1911 (Figure 6c in Edgecombe et al., 2002).

Edgecombe and Giribet (2004 [char. 153]) stated that accessory denticles are ubiquitous in Lithobiomorpha, though also present in the scolopendromorph family Cryptopidae. Drawing on the fairly large number of examined lithobiid species and specimens in this study, we provide additional information on the differences in shape (triangular/multifurcating, simple/structured), including intergrading forms. Accessory denticles as multifurcating scales were only described for the genus Lamyctes (Figure 7b in Edgecombe et al., 2002) while Edgecombe & Giribet (2004) coded these denticles (see char. 156) as simple and triangular only. Furthermore, the transition of multifurcating scales to simple scales was not described in detail before, although it was depicted in Lamyctes emarginatus (Newport, 1844) for example (Figure 7b in Edgecombe et al., 2002). If accessory denticles are structured, they can be equipped with tubercles or spines. Edgecombe (2004a) described “tuberculate accessory denticles” in character 51 (state (2)) for the henicopid subgenus Haasiella as well. However, the mandible of a studied moulting specimen of L. lapidicola showed tuberculate denticles on the old part and spinous multifurcating scales on the freshly moulted mandibles, indicating that tuberculate accessory denticles are perhaps the result of attrition (e.g., due to chewing). It could be worth cross‐checking individuals of different stadia to verify the state of this character and possible environmental influence.

The grooved ridge on the ventral edge of a mandibular tooth interrupts the accessory denticles in all examined lithobiid species, consistent with the findings of Edgecombe et al. (2002 [char. 27]) for the family. There are however slight differences in the expansion of the ridge along the tooth.

For the Lithobiidae, we revised the description of character 28 in Edgecombe et al. (2002), which describes the connection of the accessory denticles and the trichomes of the pulvillus. It now appears subjective to define the elongation as abrupt or progressive according to Edgecombe & Giribet (2004 [char. 155]). We now specify if there is a contact between the denticles and the trichomes of the pulvillus or not in character 32: continuous transition of accessory denticles into trichomes of pulvillus: (0) absent (trichome‐free strip); (1) present (either gradual or abrupt). This character conflicts with monophyly of the subgenera Lithobius and Sigibius (Figure 19).

As a new character for lithobiid systematics, we suggest the occurrence of an internal spinulation of the mandibular gnathal edge (Figures 8 and 9), as follows (char. 31): spinulation on internal side of mandibular gnathal edge posterior to margin of ventralmost mandibular tooth: (0) absent; (1) present (simple spines or branching bristles). The presence of an internal spinulation was hitherto illustrated in L. forficatus (see Rilling, 1968) and in the description of Australobius scabrior Chamberlin, 1920 (“three small curved spines”; Figure 8d in Edgecombe & Hollington, 2002). Both species are resolved in a clade also based on this character (Figure 19), which was already supported by the peristomatic structures (Koch & Edgecombe, 2008; Figure 12B). However, the internal spinulation appears to be absent on illustrations of Lithobius variegatus Leach, 1814 (Figure 2e in Edgecombe & Giribet, 2004), and in the henicopids Henicops dentatus Pocock, 1901, H. tropicanus Hollington & Edgecombe, 2004 and H. milledgei Hollington & Edgecombe, 2004 (Figures 5, 10, and 14 in Hollington & Edgecombe, 2004).

4.2. First maxillae

Edgecombe et al. (2002 [char. 31]) described a brushlike “setal” cluster on the first maxillary coxal projection including: (a) plumose setae, which we describe here as plumose bristles, as they have no sensory function (see Müller, Sombke, Hilken, & Rosenberg, 2011); (b) simple striated setae (here described as sensilla trichodea with a socket and shaft lumen [Figure 15c]; see Müller et al., 2011) and (c) complex setae composed of slender strands (here called feather−/hassock‐like branching bristles). Koch and Edgecombe (2008 [char. 15]) defined a mix of plumose and simple setae for Lithobiidae. These observations can be verified for most of the examined lithobiid species in this study. To our knowledge, previous studies do not provide information on the arrangement of the different types of bristles and trichoid sensilla or about the existence of sensilla microtrichodea (for terminology see Müller et al., 2011). Therefore, we propose a new character (char. 35) for the first maxillary coxal projection: sensilla microtrichodea on coxal projection of first maxillae: (0) absent; (1) present.

The paired rows of plumose bristles on the inner margin of the distal article of the first maxillary telopodite as an autapomorphic character of Lithobiomorpha (Edgecombe, 2004b) is verified by our data. However, our observations reveal the occurrence of more rounded and/or flattened bristles even further to the inner margin as illustrated for Lithobius obscurus Meinert, 1872 by Edgecombe and Giribet (2004, see Figure 7d therein), but not described in detail.

Sensilla microtrichodea occur as a cluster or row between the coxal projection and the telopodite (Figure 14b–f) and vary in number between species, conspecifics and within an individual. This high variation makes it impossible to define a character. A cluster of sensilla microtrichodea was described by Edgecombe and Hollington (2002) for Australobius scabrior as well and, being present in Henicopidae as well as Lithobiidae, was proposed as an autapomorphic character for Lithobiomorpha (Edgecombe, 2004b).

4.3. Comments on the variation of microstructures

Ecological influence may affect the expression of characters within a single species due to food availability, habitat choice or environmental conditions during development (e.g.,Tobias, 1969). For example, intraspecific variation of the aciculae was described for the scolopendromorph Ectonocryptoides quadrimeropus Shelley & Mercurio, 2005 by Koch et al. (2010; p. 56). However, the described mandibular and first maxillary characters are invariant between conspecifics of the genus Henicops Newport, 1845 from different populations (Edgecombe, Colgan, & Sharkey, 2006 [char. 10–13]). Our study of seven individuals of the species L. validus and L. dentatus from different locations in Austria showed a low intraspecific variation of microstructures. There are mainly differences in the number of bristles on the internal side of the mandibular gnathal edge and the number of aciculae, reported above in “Results”. Besides examining several individuals of the same species, the investigation of different stadia per species is necessary to check for structural variability and, in a more systematic context, character stability as already suggested by Tobias (1969).

There are also differences in number of internal spines/bristles and aciculae of left and right mandibles or sensilla microtrichodea (between the coxal projection and telopodite) of left and right first maxillae of a single individual. The same was observed for the number of aciculae and even number of mandibular teeth in Henicops washpoolensis (Edgecombe & Hollington, 2005).

4.4. Phylogenetic implications

As revealed by their cladistic analyses based on 40 morphological characters earlier, Koch and Edgecombe (2008) showed that the genus Lithobius is non‐monophyletic. The present phylogenetic results with an extended set of taxa of the genus Lithobius and more characters likewise recovers non‐monophyly with species of Ethopolyinae and other lithobids (consistently including at least Hessebius, Neolithobius and Australobius) clustering within the genus. This study further demonstrates that the subgeneric divisions within Lithobius, which mostly use combinations of the same set of characters rather than unique apomorphies, are likely artificial and do not represent natural groupings within the genus.

Based on a number of striking similarities in the peristomatic structures, Ganske et al. (2018) inferred a close relationship between the genus Disphaerobius (=Pterygotergum), on which the subfamily Pterygoterginae Verhoeff, 1933, is based, and the lithobiine (sub)genera Lithobius and Ezembius. These similarities are congruent with other shared characters from external morphology identified in a recent taxonomic work (Farzalieva et al., 2017). Here, we further compared the mandibular gnathal edge of Disphaerobius loricatus, Hessebius plumatus (Zalesskaja, 1978) and Lithobius (Ezembius) giganteus and noted strong similarities, especially in the presence of simple spines on the internal side; the low number of simple, triangular accessory denticles; and the similar pattern of the fringing bristles, these branching distally only and showing broad bases. These characters and the phylogenetic results, which depict strong support for the node grouping D. loricatus and L. (E.) giganteus, provide additional evidence in favour of the inference made by Ganske et al., (2018) that Pterygoterginae is an invalid subfamily because its recognition renders Lithobiinae paraphyletic.

Nevertheless, our data are sensitive to character weights, resulting in unstable groupings, and most nodes receive low support values. This is likely a consequence of the low number of characters in relation to the number of species, and a rather high degree of homoplasy, as measured by the Consistency Index (CI 0.265).

4.5. Conclusion/outlook

The mandibles and the first maxillae of Lithobius and more generally the Lithobiidae might be useful as a source for new morphological characters, but do not provide as much information as their homologous structures in Henicopidae (see Edgecombe et al., 2002). However, new information on microstructures, for example, from the mandibles, the first maxillae and the peristomatic structures (Ganske et al., 2018) might be useful for taxonomists using SEM data as a data source to delimit cryptic species (e.g., Pilz, Melzer, & Spelda, 2008).

As stated by Edgecombe (2007, p. 337), “… the taxonomic sampling in microanatomical studies is limited, generally involving detailed descriptions of single or few species within each of the major chilopod groups and as such these data are informative for questions involving relationships between chilopod orders but are generally neutral on taxonomic problems at finer levels, even between families.” The expanded taxon sampling in this study mainly based on the genus Lithobius showed that there are differences suggesting at least three new characters. The considerable degree of sensitivity of our results to character weights and optimality criteria (i.e., parsimony versus likelihood) and weak support for most deep nodes within Lithobiidae suggest that molecular data will be especially valuable for advancing lithobiid phylogeny. The morphological characters compiled in this study will be combined with sequence data in a forthcoming phylogenetic analysis focusing on the genus Lithobius.

AUTHOR CONTRIBUTIONS

All authors designed the study. ASG performed the sample preparation. All authors collected and interpreted the data. All authors wrote, edited and approved the manuscript for publication.

CONFLICT OF INTEREST

The authors have no conflicts of interest to disclose.

Supporting information

Appendix S1: Supplementary Material

Appendix S2: Supplementary Material

Ganske A‐S, Edgecombe GD, Akkari N. Morphology of the mandibles and the first maxillae in the family Lithobiidae (Myriapoda, Chilopoda), with remarks on their phylogenetic significance. Journal of Morphology. 2018;279:1798–1826. 10.1002/jmor.20902