Comparative study of larval head structures of Megaloptera ( Hexapoda )

External and internal head structures of the larva of Neohermes are described in detail. The results are compared to conditions found in other representatives of Corydalidae, in Sialidae, and in Raphidioptera and Neuroptera. Corydalidae and Sialidae are mainly characterised by plesiomorphic features such as distinct frontal and coronal sutures, six stemmata, a movable labrum with a full set of muscles, a thin tentoriomandibular muscle, a distinct maxillary groove, 4-segmented maxillary palps, an oblique arrangement of the extrinsic maxillary muscles, a labium with all components except for the glossae and paraglossae, 3-segmented labial palps, and a largely complete muscle system. The partly reduced maxillary groove, the strongly elongated stipes, the apical membranous stipital collar, the close connection of the palp and galea, the subdivision of the galea, the strongly shortened palp, the bipartite tentoriocardinal and tentoriostipital muscles, the anterolateral submental notch, the lateral tentoriopharyngeal muscle, and the postgular plate are autapomorphies of Corydalidae. An additional antennomere is present in large corydalid species. The monophyly of the subfamilies Corydalinae and Chauliodinae is not supported by features of the larval head. The reduced condition of the anterior and dorsal tentorial arms and the antennal muscles, the transverse labial muscle, the loss of muscles of the salivary duct, and possibly the lateral origin of M. frontopharyngalis posterior are autapomorphies of Sialidae. The monophyly of Megaloptera is suggested by the insertion of a peg-like or spine-shaped sensillum on the antepenultimate antennomere, the vestigial salivary duct, and a verticopharyngeal muscle composed of several bundles. The distinct neck region, the parietal ridge, and the anterior position of the posterior tentorial grooves are features shared by Corydalidae and Raphidioptera. Arguments in favour of a clade comprising Megaloptera and Raphidioptera are the presence of a circular ridge anterad of the neck region, an increased number of Semper cells and retinula cells in the stemmata, the presence of a gula in adults, a similar cleaning behaviour, and molecular data. Potential autapomorphies of Neuropterida are the prognathism of the larvae and the absence of a mandibular mola. However, the polarity of these characters is unclear. A derived condition found in most groups of Endopterygota, but not in Hymenoptera, is the presence of one or two sensorial appendages on one of the intermediate antennomeres, usually the penultimate. Larval autapomorphies of Endopterygota suggested in earlier studies are confirmed for Corydalidae.

The short-lived adults are medium sized (Sialidae) to very large (Corydalidae).The larvae are predacious, exclusively aquatic and equipped with lateral abdominal gills (Neunzig & Baker, 1991).The order is subdivided into the distinctive families Sialidae (seven genera, ca.70 spp.)and Corydalidae (ca. 25 genera,ca. 200 spp.), the latter comprising the subfamilies Corydalinae and Chauliodinae (Glorioso, 1981;Aspöck & Aspöck, 2005).Sialidae occur on all continents with the exception of Antarctica, but are restricted to the east coast of Australia and to the southernmost and north eastern part of Africa.Chauliodinae occur in North America, Chile, Australia, New Zealand, Madagascar, southern Africa and Asia, and Corydalinae in the New World, southern Africa and Southeast Asia (Glorioso, 1981;Grimaldi & Engel, 2005).Adults of Corydalinae reach a wing span up to 175 mm (Grimaldi & Engel, 2005).Males of Corydalus are characterised by extremely elongate mandibles.The larvae of Corydalidae live in streams and breathe with lateral abdominal gills, and sometimes with additional ventral gill tufts (Corydalus) (Tarter et al., 1975(Tarter et al., , 1979;;Neunzig & Baker, 1991).
The morphological data presented here are discussed with respect to their phylogenetic implications.The aims include the evaluation of the monophyly of Megaloptera, the interrelationships within the order, and the phylogenetic affinities of the group within Neuropterida.The data are presented in a data matrix, but not analysed cladistically at this stage.Ultimately, they will contribute to an extensive future cladistic analysis of all orders of Endopterygota (see Beutel & Pohl, 2006).

Labrum (Figs 1A, 2A, 6A, 7B)
Small, connected with anterior clypeal margin by a membrane.Rounded laterally and anteriorly, with a very shallow anteromedian emargination.Dorsal side with a pair of longer setae.Shorter and longer fine setae inserted along anterior margin.Tormae at posterolateral edge well developed.

Antenna (Figs 1, 2, 3A, B)
Four-segmented, slender, less than half as long as length of head capsule.Inserted in a cylindrical membranous socket on a sclerotised articulating area immediately laterad of the secondary mandibular joint.First segment about twice as long as wide, slightly widening distally.Antennomere 2 more than 5 times as long as maximum width, slender basally, gradually widening towards the distal part, slightly narrowing apically; with cupuliform sensillum subapically on ventral side, accompanied by 2 small peg-shaped sensilla and one very small sensillum (Fig. 3B: sen).Antennomere 3 slender, slightly longer than 1.Distal segment very slender, cylindrical, rounded apically, with apical field of minute sensilla.

Mandible (Fig. 7C, D)
Primary (Fig. 7D: co) and secondary mandibular joint well developed.Surface smooth.Outer margin evenly rounded.Slightly asymmetric distally.Apical tooth strongly developed.Three additional teeth arranged in a row on right mandible; four on left mandible; subapical tooth of left mandible blade-like; following tooth strongly pointed.Mesal margin of proximal part of mandibles evenly curved.Prostheca, mola and brushes of hairs absent.

Maxilla (Figs
Not connected with labium and hypopharynx, movable in lateral direction.Slightly protracted, but inserted in distinct articulatory fossa formed by lateral submental wall and concave genal area below antennal articulatory area (Fig. 3C).Articulatory membrane not visible externally.Cardo large, undivided, with rounded lateral margin.Anterior margin nearly straight, connected with stipes by a hinge.Stipes very elongate, forming a tube-like structure together with a prominent mesal edge, which probably represents a partly reduced lacinia (Fig. 4: 1c); both parts entirely fused (stipitolacinia); slightly narrowing towards apex; longer and shorter setae distributed over entire surface; longer setae mainly inserted along lateral margin; apical part membranous.Presumptive lacinia represented by narrow but distinct and sclerotised flattened lobe distally; mesal edge set with long setae; distal setae slightly stronger and apically slightly curved; not recognisable as a separate element proximally (see above).Galea and palp adjacent, inserted on apical membranous part of stipes (Fig. 3D: col).Galea threesegmented; proximal galeomere broad and laterally broadly connected with palpifer; short second galeomere with strong mesally directed seta and a pair of anteriorly directed adjacent setae; other setae as shown in Fig 3C ; distal galeomere almost four times as long as wide basally; narrowing distally, apically rounded with a field of minute sensilla.Short, four-segmented palp borne by semicircular palpomere; membrane between palpifer and proximal palpomere widened laterally; palpomere 1 short, with two adjacent mesally directed stronger setae and a transverse row of thinner setae on mesal side; other setae as shown in Fig. 3C; palpomere 2 very short, especially on mesal side, not distinctly separated from following segment; palpomere 3 about as long as wide, slightly narrowing distally; palpomere 4 conical, with group of minute sensilla on flattened apex.

Labium (Figs 1B, 2B, 3C, 8, 9A)
Submentum laterally bordered by suture but integrated into ventral wall of head capsule; posteriorly not separated from gula; with two strongly developed lateral lobes, with an angular lateral margin bordering the maxillary groove, and deeply notched anterolateral apices (Figs 1B,3C).Large, laterally rounded, largely unsclerotised mentum inserted in deep median submental excavation; paired transverse sclerites present on posterior part of mentum, and two very small sclerites posteromesally, immediately close to anterior submental margin; anterior part of mentum membranous.Prementum slightly smaller than mentum, depending on degree of retraction (Figs 2B, 3C); largely sclerotised ventrally, with small posteromedian membranous area; anterior margin of sclerotised part with regular fringe of short spines.Dorsal side largely membranous, but with distinctly sclerotised lateral areas with short spine-like setae.Longitudinal dark sclerotisation with median fissure (longitudinal salivary sclerite in the following; Fig. 7B: lscl) separates dorsal prementum from anterior hypopharyngeal margin.Three-segmented palp inserted on membranous elevation; palpiger not present as sclerotised element; palpomere 1 short, about as long as wide; palpomere 2 elongate, cylindrical; palpomere 3 small, conical, with apical field of minute sensilla.Paraglossae absent.Medially divided, membranous ligula possibly homologous with glossae.
Musculature (Fig.   A strongly developed ventral transverse muscle (vtm) is present anterad of the anatomical mouth.
Posterior part roughly H-shaped in cross section, with trilobed dorsolateral folds and bilobed ventrolateral folds (Fig. 9E).Oesophagus distinctly widened, with a helical arrangement of deep longitudinal folds.

Salivarium (Figs
Represented by fold between dorsal side of prementum and anterior hypopharyngeal margin, longitudinal salivary sclerite, and distinctly reduced salivary tube.Salivary tube opens on posterior part of fissure of salivary sclerite (see above).Distal unpaired part of duct vertical and sclerotised, with thick walls.Paired salivary tubes extremely thin (diameter ca. 10 µm) without recognisable lumen, difficult to identify on sections of the posterior head region.Salivary glands not present within head capsule (see below).

Glands
Not present within head capsule.

Fat body
Absent in specimens examined.

MEGALOPTERAN CHARACTERS OF PHYLOGENETIC RELEVANCE
The characters are listed in a morphology based sequence (see results).Presumably plesiomorphic character states are coded as (0).The polarity determination is preliminary as it is not based on a cladistic analysis.

DISCUSSION
Larvae of Corydalidae and Sialidae are mainly characterised by plesiomorphic features such as distinct frontal and coronal sutures, the presence of six stemmata, a movable labrum with a full set of extrinsic and intrinsic muscles, a thin but recognisable tentoriomandibular muscle, the presence of a maxillary groove, 4-segmented maxillary palps, the oblique or nearly vertical arrangement of the extrinsic maxillary muscles, a labium with all components except for the glossae and paraglossae, 3-segmented labial palps, and a largely complete muscle system.The presence of a nervus connectivus is also likely plesiomorphic.However, this feature needs more study in other groups.
Character states identified as potential larval autapomorphies of Endopterygota in earlier studies (e.g., Beutel & Ge, 2008;Beutel et al., 2008) are confirmed for Corydalidae.These features include the absence of intrinsic antennal muscles, the reduction of glossae and paraglossae, and the absence of M. praementoglossalis and M. praementoparaglossalis.A supposedly derived feature found in larvae of Neuropterida (Figs 1, 10) and larvae of almost all other groups of endopterygote insects is the presence of one or two sensorial appendages on the antepenultimate or penultimate antennomere, although homology is difficult to establish in larvae of Hymenoptera (P.Švácha, pers.comm.).
Few features investigated here support the monophyly of Neuropterida.Prognathism and the absence of a mandibular mola are potential autapomorphies of this lineage.However, the polarity of both characters is ambiguous as a prognathous head and mandibles without molae do also occur in other endopterygote groups (e.g., Nannochoristidae, Siphonaptera, Diptera; e.g., Pilgrim, 1972;Sharif, 1937;Cook, 1949).Besides this, the head of Sialis is subprognathous.Whether this is plesiomorphic or due to reversal is presently unclear.The monophyly of Megaloptera, which was often challenged (e.g., Achtelig, 1967;Büning, 1998), is supported by three presumably apomorphic character states of the larval head, the insertion of a peg-like or spine-shaped sensillum on the antepenultimate antennomere, the vestigial condition of the salivary duct, and a verticopharyngeal muscle composed of several subcomponents.The latter features also occur in other groups.The salivary duct is generally absent in beetle larvae and M. verticopharyngalis is also composed of three separate bundles in Nannochorista and Panorpa (Bierbrodt, 1942).The monophyly of Megaloptera is further corroborated by molecular analysis (Haring & Aspöck, 2004) and by a hitherto overlooked derived groundplan feature; the presence of eversible sacs within the complex of the fused gonocoxites 11 in Corydalidae and Sialidae (Aspöck & Aspöck, 2008).
Despite the overall similarity of megalopteran larvae, they differ in a considerable number of features of the head.Corydalus is plesiomorphic compared to Sialidae in several characters.The reduction of parts of the tentorium, the reduced condition of the antennal muscles, the presence of a transverse labial muscle, and the loss of muscles of the salivary duct are likely autapomorphies of Sialidae.It has to be stated in this context, that in contrast to Röber (1942), anterior tentorial arms are present in Sialis even though very thin (N.Kristensen, pers. comm.).Whether the presence of a sclerotised gular plate is plesiomorphic or apomorphic is unclear.The character state distribution is similar as in the case of the neck region (see below).It is conceivable that the absence in Sialis and several groups of Neuroptera (e.g., Osmylidae, Coniopterygidae, Chrysopidae; Rousset, 1966;Aspöck et al., 2001;Aspöck & Aspöck, 2007) is due to reversal.
The monophyly of Corydalidae is strongly supported by features of the larval head, especially by characters related to the maxilla.Presumptive autapomorphies are the distinctly reduced maxillary groove, the strongly elongated stipes, the distal membranous stipital collar, the distinctly reduced lacinia, the close connection between palp and galea, the subdivision of the galea, the shortened palp, and the bipartite M. tentoriocardinalis and M. tentoriostipitalis.Additional autapomorphies are the postgular plate and the anterolateral submental notch.Derived conditions only found in some members of the family, especially in large species, are the 5-segmented antenna and the presence of a craniotentorial muscle described by Kramer (1955).The monophyly of the subfamilies Corydalinae and Chauliodinae is not supported by features of the larval head.
A closer phylogenetic affinity between Corydalidae and Raphidioptera was suggested by Achtelig (1967) based on characters of adults.A shared feature of larvae is the presence of a parietal ridge (Beutel & Ge, 2008).However, a similar structure does also occur in Coleoptera (Beutel, 1993), and the ridge may be secondarily absent in Sialis.Another presumably derived feature found in both groups is the anterior position of the posterior tentorial grooves.This condition has independently evolved in many groups of Coleoptera with predacious larvae (Beutel, 1993(Beutel, , 1999;;Beutel & Molenda, 1997), and is also present in Nevrorthidae, Myrmeleontiformia and Siphonaptera (Sharif, 1937).A distinct neck region is another feature shared by Corydalidae and Raphidioptera.As it is also present in Nevrorthidae and Myrmeleontiformia ("head-cervix-articulation" in Aspöck, 1992: Fig. 7), it is conceivable that this is a derived groundplan feature of Neuropterida.A neck region occurs in few groups of Coleoptera (e.g., Gyrinidae [partim]; Beutel, 1993), but is almost certainly absent in the groundplan of the order (Beutel & Hörnschemeyer, 2002a, b;Beutel et al., 1998;Beutel & Molenda, 1997).
A closer relationship between Sialidae and Raphidioptera, based on the presence of specific telotrophic ovarioles, as suggested by Štys & Bili ski (1990) and Kubrakiewicz (1998), is not supported by the results of our study.This hypothesis is also in contrast to Büning (1998), who pointed out specific conditions of the somatic ovarian tissues shared by Corydalidae and Sialidae.This suggests that the panoistic ovarioles of Corydalidae may be a secondary feature.As already pointed out by Beutel & Ge (2008), in this case the specialised telotrophic condition would be a strong argument for a clade comprising Raphidioptera and a monophylum Megaloptera (see also Kristensen, 1999).This more inclusive lineage was also suggested by Achtelig & Kristensen (1973) and Achtelig (1978) based on the presence of a gula in adults and the cleaning behaviour, and by Beutel & Gorb (2006) based on the presence of hairy tarsal attachment devices with a specific type of tenent setae.A potential synapomorphy of the larval head is the presence of a circular ridge anterad of the neck region (Figs 1, 10A, C: cri).The increased number of eight Semper cells and 20-45 retinula cells in the stemmata (Paulus, 1986) are further potential synapomorphies of the two orders.A clade comprising Raphidioptera and Megaloptera was also suggested in a molecular study based on 18S rDNA of 182 endopterygote terminal taxa (Whiting, 2002).However, as already pointed out by Beutel & Ge (2008), Sialis was the only megalopteran representative included and the bootstrap value for the clade is low (61).Megaloptera turned out as paraphyletic in another analysis of 18S rDNA sequence data (Winterton, 2003), and Raphidioptera as the sistergroup of Neuroptera.
A megalopteran-raphidiopteran clade is in contrast to a sistergroup relationship between Megaloptera and Neuroptera, which was proposed by Aspöck (1995Aspöck ( , 2002) ) based on several potential synapomorphies such as a specific rosette-like arrangement of trichobothria on the ectoproct, primarily aquatic habits of larvae, the elongation of the larval stipites, and the "integration of the larval cardines in the head capsule" (Aspöck et al., 2001;Aspöck, 2002: p. 52).Megaloptera + Neuroptera were also corroborated by a molecular analysis (Haring & Aspöck, 2004) and by Aspöck & Aspöck (2008).As already pointed by Beutel & Ge (2008), the stipes is not elongated in Sialis and the retracted position of the maxillae, i.e. the presence of a distinct fossa maxillaris, is likely a plesiomorphic feature.Apparently, analyses of an extensive combined data set will be required for the solution of this crucial controversy in the systematics of Neuropterida.