TWO NEW SPECIES OF TYPHLOCHARIS DIECK , 1869 ( COLEOPTERA , CARABIDAE ) FROM PORTUGAL : DESCRIPTION AND NOTES ON RELATED SPECIES

The present study describes two endogean carabid species of Typhlocharis Dieck, Typhlocharis rochapitei n. sp. and Typhlocharis crespoi n. sp., belonging to the T. diecki species group and to the T. outereloi species group, respectively. The work provides diagnostic characters and in particular, the structure of male genitalia. Affinities to putative relatives and keys for the identification of the diecki (all species) and outereloi (Portugal) species groups are also given.


Introduction
Within the subtribe Anillina and according to the catalogue of the ground beetles of the Iberian Peninsula (Serrano, 2003) plus some more recent works (Ortuño, 2005;Serrano et al., 2005;Serrano & Aguiar, 2006a, 2006b;Andújar et al., 2008)  Typhlocharis are distributed throughout the Iberian Peninsula (Europe), Morocco and Tunisia (North Africa) (Jeannel, 1963).Representatives of this genus have been found in the vestiges of the Lusitanian, the Lionigurian, the Betic Riffian and the Numidian Massifs and the genus is considered a very old one (Jeanne, 1973).According to Jeannel (1963) the ancestral lucicolous of the Typhlocharis expanded from Africa to the Betic Riffian Massif.Taking into account the more recent knowledge of the structure of the Western Mediterranean from the end of the Mesozoic to the end of the Pleistocene (Oosterbroek & Arntzen, 1992;Andeweg, 2002), the land connections between the Iberian Península and North Africa occurred during the Oligocene and most probably during the Miocene.So, the present distribution of Typhlocharis may be dated from 25-30 MY onwards.Some comments on the origin and evolution of this genus can be found also in the recent work of Andújar et al. (2008).
All species of this genus are endogean carabids, wingless (apterous) and eyeless (anophtalmous), occurring and moving very slowly in soil.They may also be collected on the bottom surface of stones more or less deeply buried in the soil.The majority of species of Typhlocharis are local or regional endemisms with restricted distributions, probably because of isolation by physical barriers and low dispersal ability.Thus, taking into account their present distribution, they represent a great potential for phylogeographic studies.
The aim of this work is to describe two new species of the genus Typhlocharis for Portugal, one in the diecki group and the other in the outereloi group.Representatives of both groups were already known from Portugal (e.g.Coiffait, 1968;Serrano & Aguiar, 2000, 2006a).We also provide notes about relationships of these species with the closest forms.Moreover we give keys to all known species of the diecki species group and to the outereloi species group from Portugal.
According to Zaballos & Ruiz-Tapiador (1997) the adults belonging to the species of the diecki and outereloi groups are recognized by the following combination of characters: diecki group, a) umbilicate series of elytra with four setae in the anterior group and three setae in the posterior group (4+3) and b) apical edge of elytron with no more than two teeth; outereloi group, a) umbilicate series of elytra with four setae in the anterior group and one-two setae in the posterior group (4+1 or 4+2) and b) apical edge of elytron with no more than two teeth.The eight known species of the diecki Group were found in Spain, Tunisia and Portugal (Andújar et al., 2008).On the other hand, the fifteen known species of the outereloi group can be found in several regions of Portugal and Spain (e.g.Serrano, 2003).

Material and methods
Field work was conducted within the Province of Estremadura in the vicinity of Lisbon region and in some localities throughout the Province of Algarve (both Provinces of Portugal); this resulted in the collection of specimens of endogean beetle species of the subtribe Anillina in two localities (Alcabideche and Pincho, respectively in Estremadura and Algarve Provinces).After a careful study, we concluded that the specimens represent two new species of the genus Typhlocharis, belonging to the diecki and outereloi groups.The specimens were collected by direct hand collecting on the bottom surface of deeply buried stones in fragments of pristine-type Mediterranean forest habitats dominated by holmoak bushes (Quercus coccifera Linnaeus) (Alcabideche) or in fragments of habitats dominated by lentisk bushes (Cistus ladanifer Linnaeus) (Pincho).Additional specimens were obtained from samples Figs.1-8.-Typhlocharis rochapitei n. sp.: 1) head and anterior part of pronotum (dorsal view), 2) head (ventral view), 3) stridulatory organ (pars stridens), 4) head (part), pronotum and anterior part of elytra (dorsal view), 5) elytra (left latero-dorsal view), 6) elytra (apical view), 7) thorax (part) and abdomen (male, ventral view), 8) thorax (part) and abdomen (female, ventral view).
The morphological study of adult specimens was done using a scanning electron microscope JEOL JSM-5200 LV.Measurements and drawings were done with a Wild M5 stereoscopic microscope equipped with a dissecting microscope ocular micrometer and a drawing tube.
The distribution of species in the descriptions is given in U.T.M. coordinates (1 km x 1 km).DIAGNOSIS: Anophtalmous, body parallel, depressed, brown, with integument microreticulate, with scattered pubescence.Elytron with six discal rows of setae, impair rows with setae bigger than the ones of the pair rows; seven (4+3) marginal umbilicate setae; apical edge not sinuate, with one tooth at the ending of seventh stria region, one slightly parasutural tooth (males and females).Hind trochanters round-shaped in both sexes.Abdominal female sterna II and III with one common faint lateral fovea on each side.Aedeagus (Figs. 17A and B) with the median lobe sickleshaped, basal lamina markedly arcuate, the apex bent to left (dorsal view); internal sac diffuse, with a twisted sclerite and tangled membranes; left and right parameres with two apical seta.DESCRIPTION: Length of Holotype: 1.8 mm.Length of paratypes: 1.6 mm (males), 1.7-1.8mm (females).Head (Figs. 1-2) slight wider than long [length: 0.31-0.34mm (males) and 0.32-0.34(females), width: 0.34-0.39mm (males) and 0.40-0.43mm (females)] with penta and hexagonal microsculpture; vertex with transversal microsculpture arranged in parallel ridges to form a file (pars stridens) in the area below the anterior margin of pronotum (Fig. 3).Cephalic chaetotaxy (large setae): Labrum with three pairs of setae (those on sides longer), one pair on sides of clypeus and one pair close to frontal sulcus, two pairs of supraocular setae (anterior and posterior) and 1-2 pairs of setae on the posterior region between vertex and lateral carinae.Antennae moniliform, mouth-parts (Fig. 2) with no special features, as for other members of the genus.Pronotum rectangular (Fig. 4), slightly longer than wide (1.0-1.1 times) [length: 0.43-0.50mm (males) and 0.50-0.54(females), width: 0.42-0.46mm (males) and 0.47-0.50mm (females)], with hexagonal microsculpture, slightly narrowed towards posterior angles which are dentate; disk flattened, with one central and one lateral slight sulcus on each side; anterior margin slightly arcuate; lateral margins with 2 or 3 minor denticles near the posterior angles; disk faintly depressed near the posterior margin, this slightly expanded on each side and strongly arcuate in the middle region.Chaetotaxy: 3-4 irregular longitudinal series of minute setae between midline and lateral margins directed anteriad and inward; one anterior seta on each side in anterior quarter, one posterior seta on hind angle; five-six pairs of setae near the anterior margin.Elytra (Fig. 5) 1.8-1.9times longer than wide [length: 0.80-0.91mm (males) and 0.90-0.96mm (females)], width: 0.43-0.50mm (males) and 0.50-0.53mm (females)], with hexagonal microsculpture, parallel and oval posteriorly, with a faint longitudinal carinae at the beginning of seventh stria; flattened on the disk; transverse scutellar organ present near the base of suture; scutellar region not punctured; humeral angles well marked and rounded, with a tooth in the base of carinae; lateral margins serrate, teeth decreasing in size posteriorly; apical margin not sinuate after the end of longitudinal carina, with one tooth coinciding with end of seventh stria, presence of a small parasutural denticle .Chaetotaxy: part of the pubescence of the disk is arranged in six lines since the sutural region, setae of impair rows are bigger than the ones of pair rows, all these setae are erect and slightly directed anteriad; none large discal setae are present; umbilicate series with 4 setae in the front group and 3 in the hind group (4+3) (Fig. 5).Legs with robust femora; males with inner margin of intermediate femur very slightly dentate (Fig. 7); trochanters of third pair similar in male and female, without special features (Figs.7-8), protarsus of male without dilated segments.Abdomen (Figs. 7-8) with sternum II of male without trace of lateral fovea (Fig. 7), female sterna II and III with one faintly joint lateral fovea on each side (Fig. 8).Male genitalia (Figs.17A and B) in lateral aspect with median lobe strongly sickle-shaped, basal lamina markedly arcuate (Fig. 17A); median lobe in dorsal aspect (Fig. 17B) with apex acute, bent to left; internal sac with one twisted sclerite and tangled membranes; left paramere with two apical Graellsia, 64(2), Diciembre 2008, pp.281-293 -ISSN: 0367-5041 Table 1.-Species of Typhlocharis of diecki group: comparison of total length and of some diagnostic character states from pronotum, legs, elytron and abdomen (including male and female genitalia).Tabla 1.-Especies de Typhlocharis del grupo diecki: comparación de la longitud total y de algunos caracteres del pronoto, patas, élitro y abdomen (incluso de las genitalias de los machos y de las hembras).setae, dorso-basal edge expanded; right paramere with two apical setae.Female genitalia (Fig. 18A) with gonocoxites of ovipositor weakly sclerotized, each one in ventral aspect with one apical seta and two medium setae, the proximal bigger than the basal one; internal genital tract with spermathecal duct short and spermatheca barrel shaped, spermathecal gland long, with proximal region membranous and apical region more or less sclerotized.
ETYMOLOGY: This new species is dedicated to Professora Maria Teresa Rocha Pité, a Portuguese entomologist and evolutionary ecologist, who greatly contributed to the taxonomic and ecological knowledge of the Diptera Drosophilidae, mainly of Portugal.
AFFINITIES (see also Table 1): The new belongs to the diecki group based on the umbilicate series setae pattern (4+3), presence of one parasutural tooth and anterior margin of clypeus not dentate.It is well differentiated from all the species within this group, among other features, by the shape of aedeagus (particularly median lobe, including apex in dorsal view).Within the diecki group the pronotum of T. rochapitei n. sp.presents one lateral slight longitudinal sulcus on each side like T. lunai Serrano & Aguiar, 2006 Apparently only T. farinosae and T. gonzaloi bear one medium seta in the gonocoxite while T. rochapitei n. sp.presents two setae (Fig. 18A).
Taking into account the analysed characters, it is very hard to say to which species T. rochapitei n. sp. is more akin within the diecki group.
The morphological study of adult specimens of the new species, using a scanning electron microscope, allowed us to verify that the internal edge of intermediate tibiae of males does not bear a long seta near the apical margin, as well as the elevation in posterior margin of abdominal sternum IV of females, features that were found in T. lunai (Serrano & Aguiar, 2006a).Abdominal sternum II in males are not tuberculate near the posterior margin either.
As a final remark, we would like to point out that T. rochapitei n. sp. is the second species recorded for Portugal, belonging to the diecki group.This group has a large distribution area that includes most parts of the Iberian Peninsula and Tunisia (Algeria and North Morocco are not yet well known concerning the Typhlocharis fauna).The finding of T. lunai and now T. rochapitei sp.n. in Portugal agrees with this large distribution (see Serrano & Aguiar, 2006a, Figs. 26 and 27).
DESCRIPTION: Length of Holotype: 1.2 mm.Length of paratypes: 1.0-1.2mm (males and females).Head (Figs. 9-10) slightly longer than wide [length: 0.22-0.26mm (males) and 0.27-0.29 mm (females), width: 0.20-0.26mm (males) and 0.26 mm (females)] with hexagonal microsculp-ture, not depressed in the middle of the frons; vertex without transversal microsculpture, absence of a file (pars stridens) (Fig. 11).Cephalic chaetotaxy (large setae): Labrum with three pairs of setae (those on sides longer), one pair on sides of clypeus and one pair close to frontal sulcus, a pair of supraocular setae present over each eye and one posterior pair of setae between vertex and lateral carinae.Antennae moniliform, mouth-parts (Fig. 10) with no special features, as for other members of the genus.Pronotum quadrangular (Fig. 12), as long as wide [length: 0.26-0.30mm (males) and 0.29-0.31mm (females), width: 0.24-0.29 mm (males) and 0.27-0.29 mm (females)], with hexagonal microsculpture, slightly narrowed towards posterior angles which are marked; disk flattened, with one central and two lateral slight sulci on each side; anterior margin slightly arcuate; lateral mar-  gins slightly rounded, without minor denticles near the posterior angles; disk faintly depressed near the posterior margin, this slightly arcuate.Chaetotaxy: three longitudinal series of minute setae between midline and lateral margins directed anteriad and inward; one anterior seta on each side in anterior quarter, one posterior seta on hind angle; five-six pairs of setae near the anterior margin.Elytra  twice longer than wide [length: 0.48-0.55mm (males) and 0.56-0.59mm (females), width: 0.25-0.29 mm (males) and 0.26-0.29 mm (females)], parallel and oval posteriorly, with hexagonal microsculpture and a slightly longitudinal carinae at the beginning of seventh stria; depressed on the disk; transverse scutellar organ present near the beginning of suture (Fig. 13); scutellar region not punctured; humeral angles rounded, with a minor tooth at the beginning of carinae; lateral margins slightly serrate, teeth decreasing in size posteriorly; apical margin (Fig. 14) with one tooth coinciding with end of seventh stria, presence of a parasutural tooth.Chaetotaxy: Part of the pubescence of the disk arranged in three lines, these short setae are erect and slightly directed anteriad; umbilicate series with 4 setae in the front group and 2 in the hind group (4+2) (Fig. 13).Legs with robust femora, mace shaped, tibiae dilated distally in both sexes; trochanters of third pair round-shaped, similar in both sexes , protarsus with segments not dilated.Abdominal sterna II and III (males and females) without fovea on each side .Male genitalia (Figs.17C and D) in lateral aspect (Fig. 17C) with median lobe almost right, basal lamina markedly arcuate; median lobe in dorsal aspect (Fig. 17D) with apex more or less large; internal sac in central area without sclerites, presence of a tubiforme membrane (Fig. 17C); parameres bisetulose apically.Female genitalia (Fig. 18B) with ovipositor gonocoxites weakly sclerotized, each one in ventral aspect with 1 apical seta and 1 medium seta; internal genital tract with spermathecal duct short and spermatheca barrel shaped.ETYMOLOGY: This new species is dedicated to Professor Eduardo Frias Crespo, who has greatly contributed to the taxonomic, phylogenetic and ecologic knowledge of the Portuguese amphibian and reptilian faunas, and become a remarkable academic reference to the senior author.

AFFINITIES (see also Table 2):
The new species belongs to the outereloi group based on umbilicate series of elytron with four setae in the anterior group and two setae in the posterior group (4+2) and the apical edge of elytron with two teeth.Within the outereloi species group, the new species as well as T. intermedia Zaballos, 1986 do not exhibit denticles in lateral margins of pronotun near the hind angles.The new species is akin to T. quadridentata Coiffait, 1968, T. navarica Zaballos & Wrase, 1998, T. atienzai Zaballos & Ruiz-Tapiador, 1997, T. estrellae Zaballos & Ruiz-Tapiador, 1997, T. bullaquensis Zaballos & Ruiz-Tapiador, 1997and T. singularis Serrano & Aguiar, 2000 because of the presence of two lateral sulci on each side of pronotum, to T. quadridentata, T. belenae Zaballos, 1983, T. intermedia, T. jeannei Zaballos, 1989, T. bullaquensis and T. elenae Serrano & Aguiar, 2002 because of the presence of two teeth in the apical margin of elytron, to T. outereloi Novoa, 1979, T. quadridentata, T. belenae, T. intermedia, T. navarica, T. singularis, T. gomesalvesi Serrano & Aguiar, 2002, T. laurentii Magrini, 2000and T. bazi Ortuño, 2000 because of the absence of thorn shaped hind trochanters in males, to T. outereloi, T. quadridentata, T. belenae, T. intermedia, T. toriboi Ortuño, 1988, T. jeannei, T. singularis, T. gomesalvesi, T. laurentii and T. bazi because of the absence of lateral fovea in abdominal sternum II (one or two) and to T. intermedia, T. jeannei, T. atienzai, T. estrellae, T. bullaquensis, T. singularis, T. elenae and T. gomesalvesi because of the pattern of apical and medium gonocoxite setae (1+1).Typhlocharis crespoi n. sp., T. outereloi, T. singularis, and T. laurentii exhibit a similar pattern of form (slightly arcuate and pointed apex) of the median lobe of the aedeagus in lateral view.However, differences in the shape of the apex of the median lobe (dorsal view), left paramere and internal sac separate the new species from all the other species belonging to this group.The new species together with T. quadridentata and T. singularis, by the known data (Magrini, 2000;Serrano & Aguiar, 2002), are the smallest ones of the outereloi group.
Taking into account the analysed characters, the new species seems to have a closer relationship with T. intermedia, T. quadridentata and T. singularis.The first species was found near Ciudad Real (Spain) and the latter two in several localities of Portugal (Coiffait, 1968;Serrano & Aguiar, 2002).

Fig. 17 .
Fig. 17.-Typhlocharis rochapitei n. sp.: median lobe of aedeagus and left paramere in lateral view (A) and median lobe of aedeagus in dorsal view (B); Typhlocharis crespoi n. sp.: median lobe of aedeagus and left paramere in lateral view (C) and median lobe of aedeagus in dorsal view (D).