Seven new species of Notiospathius ( Hymenoptera , Braconidae , Doryctinae ) from Northwest Venezuela

We describe seven new species of the doryctine wasp genus Notiospathius Matthews & Marsh from Northwest Venezuela: N. araguae sp. n., N. bolivari sp. n., N. curvilineatus sp. n., N. dantei sp. n., N. estradae sp. n., N. larensis sp. n., and N. venezuelae sp. n. These represent the first described species of the genus JHR 29: 37–61 (2012) doi: 10.3897/JHR.29.3555 www.pensoft.net/journals/jhr Copyright E. Karen López-Estrada et al. This is an open access article distributed under the terms of the Creative Commons Attribution License 3.0 (CC-BY), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. ReSeARCH ARtiCle E. Karen López-Estrada et al. / Journal of Hymenoptera Research 29: 37–62 (2012) 38 reported for this country. Species boundaries for the above taxa were confirmed using the General mixed Yule-coalescent method with 441 DNA barcode sequences from specimens assigned to Notiospathius and other closely related genera collected in different countries along the Neotropics.


Seven new species of Notiospathius (Hymenoptera, Braconidae, Doryctinae) from Northwest Venezuela introduction
The wasp family Braconidae is a group mainly composed of parasitoids that attack a wide range of insect larvae from different orders, but mostly Coleoptera, Diptera and Lepidoptera (Shaw and Huddleston 1991;Marsh 1997).One of the most speciose braconid subfamilies is Doryctinae, with around 200 currently recognised genera and over 1,300 described species (but minimum estimate of 3,000 undescribed species; Jones et al. 2009).Within this subfamily, Notiospathius Matthews & Marsh is probably the most speciose genus in the Neotropics after Heterospilus Haliday (Ceccarelli et al. 2012).Notiospathius was erected to contain 14 species previously placed within the cosmopolitan Spathius Nees (Matthews & Marsh, 1973).Currently, the genus comprises 34 Neotropical species (Zaldívar-Riverón and De Jesús-Bonilla 2010;De Jesús-Bonilla et al. 2011), though a considerable, undetermined number of undescribed species remains to be described (Ceccarelli et al. 2012).Notiospathius is principally characterised by a considerably enlarged and tubular first metasomal tergite, although this feature is also shared by members of other doryctine genera that are either closely (Tarasco Marsh, Masonius Marsh) or distantly related (e.g.Spathius) to it (Zaldívar-Riverón et al. 2007, 2008;Ceccarelli and Zaldívar-Riverón submitted).
Recently, a number of specimens assigned to Notiospathius, Tarasco and Masonius were collected by three of the authors (AZR, HC, RB) in different localities along northwest Venezuela.In this work, we describe seven new species of Notiospathius from this region following an integrative taxonomic approach (Schlick-Steiner et al. 2010;Padial and De la Riva 2010), where both morphological and DNA barcoding (Hebert et al. 2003) data consistently support our species limits.These represent the first described species of the genus recorded for Venezuela.

Examined specimens
A total of 243 specimens originally assigned to the genera Notiospathius, Masonius and Tarasco were obtained from several localities along four northwest Venezuelan states (Lara, Yaracuy, Carabobo and Aragua).All specimens were collected with yellow pan traps or sweeping nets and preserved in 100% ethanol until they were prepared for DNA extraction.Specimens are deposited in the following collections: Colección Nacional de Insectos, Instituto de Biología, Universidad Nacional Autónoma de México (IB-UNAM CNIN), Museo Entomológico "José M. Osorio", Decanato de Agronomía, Universidad Centroccidental "Lisandro Alvarado", Cabudare, Lara, Venezuela (UCOB), and Departamento de Ecologia e Biologia Evolutiva, Universidade Federal de São Carlos, São Carlos, SP, Brazil (DCBU).The wing venation and sculpture terminologies employed followed Marsh (2002).Digital SEM photographs were taken with a FEI Quanta TM 250 SEM in a low vacuum mode.Colour digital photographs were taken with a Leica ® Z16 APO-A stereoscopic microscope and a Leica ® DFC295/ DFC290 HD camera, and edited with the Leica application Suite ® program.

Species boundaries based on DNA barcodes
We assessed species boundaries among the above Venezuelan specimens by obtaining DNA sequences from the standard animal DNA barcoding locus [cytochrome c oxidase I mitochondrial DNA gene (COI)].DNA extraction, amplification and sequencing for these specimens was carried out at the University of Guelph, Ontario, Canada (see laboratory procedures in Smith et al. 2009).The above sequences are deposited in GenBank (see accession numbers in Table 1) and are also available in the project file "Doryctinae of the world" (DORYC project) of the Barcode of Life Data Systems (BOLD; www.barcodinglife.org).Fourteen sequences of specimens belonging to Notiospathius and Masonius were also obtained from the BOLD project file "Parasitoid Wasps (Braconidae: Doryctinae) of Chamela-Cuixmala Biosphere Reserve" (ASDOR project; GenBank accession numbers HM434312, 324, 544, 1013-15, 1292, 1293, HQ548183, HQ926041, JF912317-20).
Moreover, we obtained 12 additional sequences of specimens collected in various Neotropical countries (GenBank accession numbers JX870412-23).These sequences were generated at the Instituto de Biología UNAM following the lab procedures described in Ceccarelli et al. (2012).All these sequences were pooled together with a previously published COI data set (Ceccarelli et al. 2012) containing 171 sequences of specimens assigned to Notiospathius, Masonius and Tarasco that were collected in different regions along the Neotropics, and a sequence of Spathius that was employed to root the tree.
The ultrametric tree generated was employed for species delimitation using the SPLITS package for the R statistical environment (http://r-forge.r-project.org/projects/splits) and running the single threshold optimisation (Pons et al. 2006;Monaghan et al. 2009).

Species boundaries analysis
The GMYC analysis based on the ultrametric tree reconstructed with the lognormal coalescent prior is shown in the Supplementary material.The analysis yielded 112 "GMYC" species (confidence interval 100-112; -lnL of null model = 3392.3;-lnL of GMYC model = 3415.7;threshold time = -0.03),52 of which were recovered as sequence clusters and 60 as singletons.The 267 Venezuelan specimens examined were segregated into 29 GMYC species, of which only one contained specimens from a different country (Costa Rica; N. venezuelae sp.n.).Two of these species actually belong to an undescribed genus that is morphologically similar but is distantly related to Notiospathius according to a recent multilocus phylogenetic study (Ceccarelli and Zaldívar-Riverón, submitted).Moreover, two separate clades contained most of the examined Venezuelan specimens (232 sequences), one comprising nine (spp.2-10 in Supplementary Material) and the other one two (spp.11 and 12) GMYC species, respectively.Species within each of these two clades were morphologically indistinguishable according to our examined features and therefore they will not be described until more evidence is gathered.Other three Venezuelan GMYC species assigned to Notiospathius were only represented by males or single, damaged, females, whereas five belong to Tarasco and one to Masonius.Below we describe the remaining seven GMYC species represented by Venezuelan specimens that also were delimited morphologically.A list with the species delimited in this study, the Venezuelan states where they were collected, number of representative specimens and their GenBank accession numbers is provided in Table 1.Description.Female.Colour: Head light brown; scape light brown to honey yellow; flagellomeres brown turning dark brown, the last 10 white; palpi pale yellow to white.Mesonotum brown; propodeum, metapleuron and first metasomal tergite black; second and third metasomal tergites brown; remaining metasomal tergites dark brown except two last metasomal tergite, which are honey yellow; ovipositor and sheaths brown.Fore and middle coxae, trochanter and trochantellus pale yellow; femora and tibiae light brown; tarsi dark brown; hind coxa black; trochanter and trochantellus pale yellow; femur, tibia and tarsi ligth brown to brown.Wings dusky; veins and stigma brown; tegula pale yellow.Body length: 3.2 mm (lateral view); ovipositor 1.5 mm.Head: Clypeus rugose; face, frons and vertex striate, temple and gena smooth; eye about 1.6 times higher than wide (lateral view); malar space 0.1 times eye height (lateral view); temple 0.4 times eye width (dorsal view); hypoclypeal depression elliptic; ocular-ocellar distance four times diameter of lateral ocellus; length of scape twice its width (frontal view); antenna with 26 flagellomeres.Mesosoma: Length of mesosoma two times its maximum height; pronotum rugose, lateral area of pronotum coriaceous; pronotal groove wide and scrobiculate; propleuron slightly striate; mesoscutal lobes coriaceous; notauli wide and scrobiculate, meeting in scutellum in a costate area; scutellar disc coriaceous; mesopleural and subalar sulcus continuous, both wide, deep and scrobiculate; mesopleuron porcate dorsally, coriaceous medially and ventrally; precoxal sulcus wide, deep and scrobiculated, as long as mesopleuron; venter of mesosoma slightly coriaceous; metapleuron and propodeum strongly rugose; apical lateral corners without distinct tubercles; spines over hind coxa indistinct.Wings: Fore wing length 4.4 times its maximum width; length of pterostigma 3.3 times its maximum width; vein r 0.4 length of vein 3RSa; vein m-cu antefurcal to vein 2RS; vein 1cu-a interstitial to vein 1M; hind wing vein M+CU about 0.3 times length of vein 1M.Legs: hind coxa rugose, without basoventral tubercle; middle and hind femora smooth.Metasoma: First metasomal tergite rugose, length five times its apical width (dorsal view); basal sternal plate (acrosternite) about 0.7 times length of tergum; second metasomal tergite mostly smooth, only indistinctly costate near basal suture; suture between second and third metasomal tergites indistinct; third metasomal tergite and remaining metasomal tergites smooth and polished; ovipositor about 0.9 times length of metasoma.Male.Smaller than female; body length 3 mm; head brown, orbit surrounding eyes yellow; antenna with 23 flagellomeres; second and third metasomal tergites light brown.
Etymology.The name of this species refers to the Venezuelan state where both type specimens were collected.

Biology. Unknown.
Etymology.This species is named in honour to Simón Bolívar, the South American military and political leader who led Venezuela, Colombia, Ecuador, Peru and Bolivia to independence.Description.Female.Colour: Head brown to reddish brown; scape pale yellow with a brown longitudinal stripe laterally; flagellomeres dark brown turning black to apex, apical 9 white; palpi white.Mesosoma and first metasomal tergite reddish brown to brown; second metasomal tergite dark brown, yellow basolaterally; third metasomal tergite yellow, dark brown apically; remaining metasomal tergites brown except the last one which is yellow; ovipositor and sheaths brown to reddish brown.Fore and middle coxae, trochanter and trochantellus pale yellow; femora pale yellow with a median and an apical dark brown transverse bands dorsally; tibiae pale yellow basally, light brown medially and apically, with a dark brown transverse band dorso-basally; tarsi light brown to brown; hind coxa pale yellow, brown apically; trochanter and trochantellus honey yellow; femur pale yellow on basal third, apical two thirds dark brown; tibia pale yellow basally, light brown medially and apically, with a dark brown transverse band dorso-basally; tarsi brown.Wings dusky; veins and stigma dark brown; tegula yellow.Body length: 5.8 mm (lateral view); ovipositor 6.8 mm.Head: Clypeus rugose; face straite; frons striate; vertex striate; temple and gena smooth; eye 1.5 times higher than wide (lateral view); malar space 0.4 times eye height (lateral view); temple 0.2 times eye width (dorsal view); hypoclypeal depression elliptic; ocular-ocellar distance 2.6 times diameter of lateral ocellus; length of scape 1.4 times its width (frontal view); antenna with 35 flagellomeres.Mesosoma: Length of mesosoma 1.9 times its maximum height; pronotum longitudinally costate-coriaceous; pronotal groove wide and slightly scrobiculate; propleuron slightly striate; lateral mesoscutal lobes coriaceous medially, transversally costate along notauli and edges; median mesocutal lobe transversally costate-rugose; notauli wide, deep and scrobiculate, meeting just before scutellum in a longitudinally costate-rugose area; scutellar disc smooth; mesopleural and subalar sulcus discontinuous, the first one deep and scrobiculate and the second one wide, deep and including the striate sculpture of mesopleuron; mesopleuron porcate-slightly coriaceous dorsally, slightly curved, diagonally costate from postero-median to medioventral region medially and ventrally; precoxal sulcus wide, deep and including the striate sculpture of mesopleuron, as long as mesopleuron; venter of mesosoma smooth and polished; metapleuron costate ventrally, rugose-aerolate dorsally, slightly coriaceous on anterior first third; propodeum rugose; apical lateral corners without distinct tubercles; spines over hind coxa indistinct.Wings: Fore wing length 4.3 times its maximum width; length of pterostigma 2.8 times its maximum width; vein r 0.2 length of vein 3RSa; vein m-cu antefurcal to vein 2RS; vein 1cu-a distinctly postfurcal to vein 1M; hind wing vein M+CU about 0.4 times length of vein 1M.Legs: hind coxa striate, indistinct tubercule at the base; middle and hind femora slightly coriaceous.Metasoma: First metasomal tergite strongly costate with rugose microsculpture, length 3.1 times its apical width (dorsal view); basal sternal plate (acrosternite) about 0.6 times length of tergum; second metasomal tergite costate with rugose microsculpture; suture between second and third metasomal tergites distinct and sinuate; third metasomal tergite coriaceous on basal half, smooth on apical half; remaining metasomal tergites smooth and polished; ovipositor about 1.8 times length of metasoma.
Etymology.The name of this species refers to the curved, diagonally costate sculpture on the median and ventral areas of mesopleuron.

Biology. Unknown.
Etymology.This species is named in honour of the first author's father.Diagnosis.This species is similar to N. shawi Marsh, though it can be distinguished from the latter species by having the mesopleuron coriaceous-slightly rugose dorsally, coriaceous medially and ventrally (costate dorsally and coriaceous ventrally in N. shawi), and notauli obscuring before scutellum in a large porcate-rugose area (meeting before scutellum in a narrow rugose area in N. shawi).Description.Female.Colour: Head light brown to honey yellow; scape honey yellow; flagellomeres honey yellow turning dark brown to apex; palpi pale yellow.Mesosoma and first metasomal tergite black, pronotum light brown on half basal half, black on apical half; second metasomal tergite dark brown; third metasomal tergite and remaining metasomal tergites brown to dark brown; last one pale yellow; ovipositor brown; sheaths honey yellow.Fore and middle coxae light brown to honey yellow; trochanter and trochantellus pale yellow; femora brown; tibiae pale yellow basally, honey yellow medially and apically; tarsi honey yellow; hind coxa black; trochanter and trochantellus pale yellow; femur dark brown; tibia pale yellow basally turning light to apex; tarsi light brown.Wings dusky, veins and stigma dark brown, tegula dark brown to black.Body length: 4.4 mm (lateral view); ovipositor 3.8 mm.Head: Clypeus rugose; face, frons and vertex striate; temple and gena smooth; eye about 0.9 times higher than wide (lateral view); malar space 0.6 times eye height (lateral view); temple 0.2 times eye width (dorsal view); hypoclypeal depression elliptic; ocular-ocellar distance 1.5 times diameter of lateral ocellus; length of scape twice its width (frontal view); antenna with 29 flagellomeres.Mesosoma: Length of mesosoma 1.9 times its maximum height; pronotum coriaceous; pronotal groove narrow and slightly scrobiculate; propleuron slightly striate; mesoscutal lobes coriaceous; notauli wide and scrobiculate, not joining, obscuring in a large porcate-rugose area; scutellar disc coriaceous; mesopleural and subalar sulcus continuous, both deep and scrobiculate; mesopleuron coriaceousslightly rugose dorsally, coriaceous medially and ventrally; precoxal sulcus wide, deep and scrobiculate, as long as mesopleuron; venter of mesosoma coriaceous; metapleuron and propodeum strongly rugose with coriaceous microsculpture; apical lateral corners without distinct tubercles; spines over hind coxa indistinct.Wings: Fore wing length 4.3 times its maximum width; length of pterostigma 3.6 times its maximum width; vein r 0.3 length of vein 3RSa; vein m-cu intersticial to vein 2RS; vein 1cu-a distinctly postfurcal to vein 1M; hind wing vein M+CU about 0.5 times length of vein 1M.Legs: hind coxa coriaceous, with a distinct basoventral tubercle; middle and hind femora smooth.Metasoma: First metasomal tergite strongly costate with rugose microsculpture, length twice its apical width (dorsal view); basal sternal plate (acrosternite) about 0.5 times length of tergum; second metasomal tergite costate with rugose microsculpture; suture between second and third metasomal tergites distinct and sinuate; third metasomal tergite costate on basal half, smooth on apical half; remaining metasomal tergites smooth and polished; ovipositor about 1.8 times length of metasoma.
Biology.Unknown.Etymology.We named this species in honour to the first author's mother, Margarita Estrada.
Description.Female.Colour: Head dark brown to black; orbit surrounding eyes yellow; scape light brown, with a brown stripe; flagellomeres dark brown, last 11 white; palpi white.Mesosoma and first metasomal tergite black except pronotum wich is brown; second metasomal tergite dark brown; third, fourth and fifth metasomal tergites brown with a light brown band at the middle; sixth metasomal tergite dark brown, pale yellow in apex; remaining metasomal tergites pale yellow; ovipositor brown; sheaths pale yellow turning dark brown.Fore and middle coxae and trochanter white, trochantellus brown, femora pale yellow on basal third, brown on apical two thirds, pale yellow apically, tibiae light brown, tarsi brown; hind coxa black, pale yellow apically; trochanter pale yellow; trochantellus brown; femur white to pale yellow on basal third, brown on apical two thirds, pale yellow at extreme apex; tibia pale yellow basally, turning light brown apically; tarsi brown.Wings dusky; veins and stigma brown; tegula light brown.Body length: 4.1 mm (lateral view), ovipositor 1.6 mm.Head: Clypeus rugose; face, frons and vertex striate; temple slightly striate; gena smooth; eye about 1.1 times higher than wide (lateral view); malar space 0.2 times eye height (lateral view); temple 0.2 times eye width (dorsal view); hypoclypeal depression elliptic; ocular-ocellar distance three times diameter of lateral ocellus; length of scape twice its width (frontal view); antenna with 28 flagellomeres.Mesosoma: Length of mesosoma 2.1 times its maximum height; pronotum coriaceous; pronotal groove wide and coriaceous; propleuron striate; mesoscutal lobes coriaceous; notauli narrow, scrobiculate and with coriaceous microsculpture, not joining, obscuring in a large costate area; scutellar disc coriaceous; mesopleural and subalar sulcus continuous, both scrobiculate; mesopleuron porcate-coriaceous dorsally, coriaceous medially and ventrally; precoxal sulcus narrow, deep and scrobiculated, as long as mesopleuron; venter of mesosoma coriaceous; metapleuron and propodeum costate with rugose microsculpture; apical lateral corners with distinct tubercles; spines over hind coxa distinct.Wings: Fore wing length 5.1 times its maximum width; length of pterostigma 3.3 times its maximum width; vein r 0.3 length of vein 3RSa; vein m-cu antefurcal to vein 2RS; vein 1cu-a distinctly antefurcal to vein 1M; hind wing vein M+CU about 0.5 times length of vein 1M.Legs: hind coxa rugose at the base, striate posteriorly, indistinct tubercule RBG, a grant given by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), fundação de amparo à Pesquisa do Estado de São Paulo (FAPESP) to AMPD, a grant given by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (Capes) to JFN and AMPD, and a postdoctoral fellowship given by DGPA-PA-UNAM (Mexico) to FSC.
Laboratory analyses on sequences generated in Guelph since 2009 were funded by the Government of Canada through Genome Canada and the Ontario Genomics Institute (2008-0GI-ICI-03).

table 1 .
List of the species delimited in this study, the Venezuelan states* where they were collected, number of representative specimens and GenBank accession numbers.