The first sexual associations in the genus Darditilla Casal, 1965 (Hymenoptera, Mutillidae)

Abstract New sex associations are proposed for four species of Darditilla: Darditilla amabilis (Gerstaecker, 1874); Darditilla bejaranoi Casal, 1968; Darditilla debilis (Gerstaecker, 1874); and Darditilla felina (Burmeister, 1854). Darditilla botija Casal, 1965, syn. n. is the male of Darditilla amabilis; the other three males were previously unknown. Mutilla decorosa Kohl, 1882, syn. n. is conspecific with Darditilla felina. Descriptions and extended diagnoses are provided for previously unknown males and for females that were not adequately described. These represent the first sex associations for the genus Darditilla.


Introduction
The genus Darditilla Casal, 1965 was erected to include a single new species, D. botija Casal, 1965, which was known from males only (Casal 1965). In his study, Casal suggested that six described females may belong to Darditilla, and specifically suggested that D. amabilis (Gerstaecker, 1874) could be the female of D. botija. In 1968, Casal described 28 new Darditilla species or subspecies, each known only from the female, We use the abbreviations T2, T3, etc., to denote the second, third, etc., metasomal terga while S2, S3, etc., denote the second, third, etc., metasomal sterna. To compare mesosomal length and width, the distance between the anteromedial pronotal margin (excluding the anterior collar) and the scutellar-scale apex is divided by the distance between the extreme posterolateral pronotal margins, the maximum mesosomal width. The digitus or cuspis length relative to the free paramere length is used here to quantify differences in genitalic structure. For ease of comparison and to facilitate identification without dissecting the genitalic capsule, the cuspis, digitus and paramere measurements are taken in dorsal view from the apical margin of the parapenial lobe to the apex of each respective structure. Using this method, all measurements can be taken from the dorsal view and a single anchor point can be used for all three measurements. These are not actual measurements of structure length, but an index to compare relative lengths; all provided length ratios of genitalic structures are based on these indices.

Type species. Darditilla botija
Diagnosis. Male. Males of Darditilla can be separated from other South American mutillid genera by the apical row of parallel bristles on T2-4 or T2-5 (e.g. Fig. 1E) and by the ventral margin of the clypeus that is preceded by a transverse furrow (e.g. Figs. 3D, 7D) and is sometimes expanded into a broad plate-like structure over the mandibles (Fig. 1D). Additionally, Darditilla males have the scape bicarinate with a relatively flat or concave anterior surface between the carinae (although the dorsal carina is often obscure or obliterated); the axillae unarmed posteriorly; T1 broadly rounded into T2; the paramere downcurved apically; and the cuspis short and pad-like (e.g. Fig. 12).
Female. Females of Darditilla are most readily recognized by their granulate pygidium (e.g. Fig. 6E) and also have a unique combination of characters, wherein the clypeus is bidentate with the teeth slightly farther apart than the antennal tubercles (e.g. Fig. 8C); the mandible is acuminate apically and has its largest tooth situated in the basal half of the internal margin; the mesosoma is constricted anterior to the propodeal spiracles, lacks a scutellar scale, lacks a sharp dorsal tubercle directly anterior to the propodeal spiracle, and has the lateral mesonotal teeth small (e.g. Fig. 6A); T1 is broadly rounded into T2; and the metasomal setae are simple.
Distribution. Darditilla species are known from throughout South America, putative members of Darditilla are known throughout Central America as well.
Remarks. Casal (1965) described the genus from a single male specimen and used some synapomorphies of that species and its relatives in his generic description. The newly associated males described here match the diagnostic features listed by Casal and other authors in keys (e.g. Brothers 2006), but in two of the species: D. bejaranoi and D. debilis, the clypeus is less strongly modified. Rather than expanding forward to cover the mandibles, the ventral clypeal margin of these species is short, yet still has the ventral margin angled anteriorly.
Darditilla is apparently closely related to Pseudomethoca and could be nested within that genus. Males of some Nearctic and Central American Pseudomethoca species have thickened setae on T2-4 that resemble the bristles of Darditilla and some females currently placed in Pseudomethoca have a granulate pygidium. Further complicating this situation, Casal'S, treatments of Darditilla focused on southern South America and the types of northern Neotropical Pseudomethoca species consistent with Darditilla were not available to him (Casal 1965(Casal , 1968a. Without phylogenetic analysis or careful study of both sexes of these species, we cannot determine which of these northern Neotropical Pseudomethoca should be transferred to Darditilla, or whether Darditilla is even a valid genus. We, therefore, maintain Darditilla using the aforementioned diagnoses and hope that this paper will facilitate the future studies needed to clarify the validity and limits of this genus.  (Fig. 1D), which is widely transverse, almost covering the mandibles, with the ventral margin raised broadly and medially coming to an obtuse point with a subapical brush of golden setae, and have the penis valve unidentate apically (Fig. 11).
Female. The female of D. amabilis can be recognized by having T1, T2, S1 and S6 entirely orange ( Fig. 2A), having an arcuate transverse band of recumbent pale golden setae on the vertex, and having a pair of longitudinal pale golden stripes in the mesosomal dorsum, which extend to the anterior margin of mesonotum ( Fig. 2A).
Extended female diagnosis. Body length 7.6 mm. Coloration. Body and appendages reddish-black, except T1, T2, S1 and S6 entirely orange. Tibial spurs whitish. Vertex with dense, arcuate transverse band of recumbent pale golden setae, front and remainder of vertex with recumbent black setae; genal setae silver. Mesosomal dorsum covered with recumbent black setae, except laterally, with a pair of longitudinal pale golden stripes, extending to anterior margin of mesonotum. Posterior fringes of T1 and T2 black; T2 setae black anteriorly and posteriorly, pale golden mixed with black laterally, and reddish orange on orange integumental spots; T3-T6 clothed with black setae laterally and pale golden setae medially. Head. Transverse, posterior margin flat, occipital carina weak, but distinct. Head width 1.2 × pronotal width. Eye slightly ovate transversely, ommatidia distinct. Front, vertex and gena densely punctate. Genal carina well-defined, terminating in slightly sharp angle posterior to hypostomal carina. Clypeus with transverse glabrous concavity, margined by dorsal and ventral carinae, between widely separated lateral teeth. Mandible slender, tapering, bidentate apically (subapical tooth minute, distant from apex and usually obliterated through wear), unarmed ventrally. Antennal scrobe with complete dorsal carina. Antennal tubercle punctate basally, with weak scratches on anterior face, glabrous dorsally. Scape simple, moderately punctate. Flagellomere 1 1,7 × pedicel length; flagellomere 2 1.3 × pedicel length. Mesosoma. Mesosomal length 1.4 × width. Mesosomal dorsum coarsely reticulate, propodeal reticulae broader and shallower. Lateral pronotal carina extending to epaulet, humeral angle with moderately sharp obtuse angle. Mesopleuron densely punctate and setose, posterior margin defined by vertical carina. Metapleuron and lateral face of propodeum smooth and shining dorsally with isolated fine setae, micropunctate and densely setose ventrally. In dorsal view, mesosoma broadened to anterior third, strongly narrowed at propodeal spiracle, propodeum abruptly broadened. Scutellar scale lacking. Propodeum convex, dorsal and lateral faces not obviously differentiated. Legs. Foreleg with a few long strong articulated spines on posterior/lateral margins of tarsomeres. Mid-and hind tibiae each with one rows of prominent spines, 5 spines in each row; apical spurs finely serrated laterally. Hind tibia with distinct secretory pore on inner/posterior surface near base of inner spur. Metasoma. T1gradually broadened from base, not constricted apically, sessile with T2, 0.6 × as wide as T2; anterior face moderately punctate and setose. T2 densely punctate, punctures slightly smaller and sparser on orange spots; felt line broad, 0.5 × as long as T2 laterally. T3-5 densely punctate. Pygidium broad and slightly convex, lateral margins defined by distinct weakly bowed carina, posterior margin rounded and defined by indistinct carina, finely granulate. S1 punctate, with weak darkened median longitudinal carina. S2 moderately punctate. S3-5 densely punctate. S6 moderately punctate.  Distribution. This species is widespread in Argentina and also occurs in Rio Grande do Sul, and Uruguay (Nonveiller 1990).
Host. Unknown. Remarks. Gerstaecker (1874) described Mutilla amabilis based on two female specimens, one from Alegrete, Rio Grande do Sul State, Brazil and another from Paraná, Entre Ríos Province, Argentina. The specimen from Alegrete, deposited in the ZMB and bearing the labels cited above, was examined and is herein designated as a lectotype. Casal (1965) hypothesized that D. botija was the male of D. amabilis based on geographical distribution in northern Argentina. Yet another overlapping distribution was found in Pelotas, Rio Grande do Sul State, Brazil. Only two other Darditilla females are known from Rio Grande do Sul: D. debilis and D. infantilis (Burmeister, 1875). The male of D. debilis is described below; D. infantilis is structurally similar to D. bejaranoi and has a consistently small body size, precluding it from association with D. botija. The newly described male of D. felina has similar genitalia and clypeal modifications to D. botija. Likewise, females of D. felina are similar to those of D. amabilis in the integumental markings of T2 and the genal carina. Distribution, similar body size, and morphological similarities to both sexes of D. felina support the synonymy of D. botija under D. amabilis.
Burmeister (1875) had briefly described the supposed male of M. amabilis from Paraná, Entre Ríos, Argentina. Later, André (1908) studied and redescribed the male that Burmeister (1875) originally associated with M. amabilis; he pointed out that its identity and sexual association were doubtful. This male is clearly not conspecific with the male of D. amabilis described above, most notably differing in its reddish basal metasomal segments. The description of this putative D. amabilis male is, however, consistent with some Argentinean Pseudomethoca males. Casal, 1968 Figs 3-4, 13-16 Darditilla bejaronoi Casal, 1968 Diagnosis. Male. The male of D. bejaronoi can be recognized by having the ventral clypeal margin produced as a short transverse slightly upcurved impunctate lamella ( Fig. 3D), by having the tegula truncate with a flat posterior face, and by having the bidentate penis valve teeth widely separated (Fig. 15).

Darditilla bejaronoi
Female. This female has a reddish mesosoma, with distinct black areas on the lateral pronotal dorsum and the posterior half of the pleurae (Fig. 4A, B), has lateral circular to transversely ovate silver setal spots on T2 (Fig. 4D), and has a strong hyaline median carina on S1.

Diagnosis. Male.
Males are similar to D. bejaronoi, but have a simply convex tegula, have the ventral impuctate lamella of the clypeus less produced than the preceding species (Fig.  5D), and have the apical and preapical teeth of the penis valve coalescent (Fig. 19).
Host. Unknown. Remarks. Darditilla debilis is a widely distributed and common species. Over 200 additional specimens were examined in MZSP and others were studied from various North American collections.
Diagnosis. Male. The male of D. felina is easily recognized by having a unique clypeus, with the ventral margin, often hyaline, raised broadly lamellate (Fig. 7D), and the penis valve, which is unidentade apically (Fig. 23).
Female. This female can be separated from all other southern and southeastern Brazillian Darditila by the large, coalescing lateral orange spots of T2 (Fig. 8D), and the large coalescing lateral patches of silver to pale golden setae on the propodeum (Fig. 8A).
Extended female diagnosis. Body length 5.7-11.9 mm. Coloration. Body entirely black, except T2 with large, coalescing lateral orange spots and S2 often orange basomedially. Appendages variable, ranging from entirely black to entirely orangish. Tibial spurs whitish. Head with sparse erect black setae. Mesosoma with sparse erect black setae, except propodeum with coalescing large lateral patch of silver to pale golden setae. Posterior fringes of T1 and T2 black; T2 setae black anteriorly and posteriorly, pale golden laterally, and reddish orange on orange integumental spots; T3-6 clothed with silver to golden setae. Head. Transverse, posterior margin flat, occipital carina obscure. Head width 1.5 × pronotal width. Eye slightly ovate transversely, ommatidia distinct. Front, vertex and gena densely punctate. Genal carina well-defined, terminating in sharp angle posterior to hypostomal carina. Clypeus with transverse glabrous concavity, margined by dorsal and ventral carinae, between widely separated lateral teeth. Mandible tapering to apex, with tooth in basal third and in apical third, unarmed ventrally. Antennal scrobe with complete dorsal carina. Antennal tubercle punctate basally and on anterior face, glabrous dorsally. Scape simple, moderately punctate. Flagellomere 1 2.0 × pedicel length; flagellomere 2 1.25 × pedicel length. Mesosoma. Mesosomal length 1.4 × width. Mesosomal dorsum densely reticulate, propodeal reticulae broader. Lateral pronotal carina extending to epaulet, humeral angle obtusely angulate. Mesopleuron densely punctate and setose, posterior margin defined by vertical carina. Metapleuron and lateral face of propodeum smooth and shining dorsally with isolated fine setae, micropunctate and densely setose ventrally. In dorsal view, mesosoma broadened to anterior third, strongly narrowed at propodeal spiracle, propodeum abruptly broadened. Scutellar scale lacking. Propodeum convex, dorsal and lateral faces not obviously differentiated. Legs. Foreleg with a few long strong articulated spines on posterior/lateral margins of tarsomeres. Mid-and hind tibiae each with one rows of prominent spines, 5 spines in each row; apical spurs finely serrated laterally. Hind tibia with distinct secretory pore on inner/posterior surface near base of inner spur. Metasoma. T1gradually broadened from base, not constricted apically, sessile with T2, 0.5 × as wide as T2; anterior face moderately punctate and setose. T2 densely punctate, punctures slightly larger and sparser anterolaterally on orange spots; felt line broad, 0.6 × as long as T2 laterally. T3-5 densely punctate. Pygidium broad and slightly convex, lateral margins defined by distinct weakly bowed carina, posterior The male that Burmeister (1854) originally associated with D. felina was recognized as Ephuta inaurata (Smith, 1855) by Mickel (1964).
Both sexes of D. felina have been examined from throughout the Atlantic Rainforest. The subspecies, D. felina agatas Casal, 1968 differs from typical females of D. felina in setal coloration and is known from two specimes from the Chaco region of Bolivia. It is unclear whether this is a valid species, valid subspecies, or a synonym of D. felina without study of further specimens.
Females that key (Casal 1968a) to D. felina have been examined from Rondônia. Although these females match D. felina in coloration, their genal carina is different. Darditilla felina has the genal carina extending from the posterior head margin to below the eye, where it terminates at an angle; this putative new species has the genal carina extending nearly to the hypostomal carina where it gradually terminates. We refrain from describing the species at this time because the focus of this manuscript is southern and southeastern Brazillian Darditila and the male is yet unknown.
Females show extensive variation in coloration of the legs. Many specimens have the legs entirely orange, while other specimens, including the female from the mating pair, have the legs partially darkened or entirely black. In an unpublished key to female Pseudomethoca types, Mickel separated D. felina from Mutilla decorosa (Kohl, 1882) on the basis of leg color. Photographs of the type of M. decorosa were provided by Dominique Zimmermann (NMW) and it is a perfect match for the orange-legged form of D. felina. As such, we place M. decorosa as a junior synonym of D. felina.

Discussion
These are the first valid species-level sex associations in Darditilla. The sex associations presented here support Casal'S, (1965Casal'S, ( , 1968a initial genus-level associations. Because his male diagnosis, however, was based on a single species, it needed to be altered, as provided above. Seven additional South American genera are recognized from both sexes, but do not yet have any published species-level associations: Calomutilla Mickel, 1952(Quintero and Cambra 1996, Quintero and Cambra 2001; Limaytilla Casal, 1964(Brothers 2006; Neomutilla Reed, 1898 (Quintero and Cambra 2001); Pertyella Mickel, 1952 (Quintero andCambra 1996); Suareztilla Casal, 1968(Casal 1968b; Tobantilla Casal, 1965(Williams et al. 2011andVianatilla Casal, 1962 (Quintero andCambra 1996). Discovery of species-level sex associations in these genera will likely lead to broader diagnoses for the genera and can verify, or potentially invalidate, these genus-level associations.
The initial sex associations for this project came from a field observation of a mating pair (D. felina) and from a mating pair in a museum (D. bejaranoi). These rare events provided the data needed for further discoveries. The male morphology analyzed from the first two sex associations was compared to other male mutillids and allowed us to associate the sexes of two other species (D. amabilis & D. debilis). There remain 31 Darditilla species known only from females (Nonveiller 1990). Data pre-sented here could be vital for recognizing the males. Instead of relying on rare events, further progress can be made in mutillid sex associations by systematic study of the distribution and morphology of museum specimens.