Doryctobracon adaimei Marinho and Penteado-Dias, 2017

Marinho, Cláudia F., Cônsoli, Fernando L., Penteado-Dias, Angélica Maria & Zucchi, Roberto A., 2017, Description of two new species closely related to Doryctobracon areolatus (Szépligeti, 1911) (Hymenoptera, Braconidae), based on morphometric and molecular analyses, Zootaxa 4353 (3), pp. 467-484 : 473-477

Doryctobracon adaimei Marinho and Penteado-Dias sp. nov.

( Figs 1 C–C 2, D–D2, E–E2 , 3 B , 5 A–H )

http://www.zoobank.org/NomenclaturalActs/7 E53462 B-1C9E-4FEC-A205-4D9877675D4A

Doryctobracon sp. 2: Veloso et al. 1996 (Cerrados de Goiás, GO ex Anastrepha spp. in Pouteria gardneriana (DC.) Radlk and P. ramiflora Radlk ). Bomfim et al. 2007 (Tocantins, TO ex Anastrepha coronilli in Bellucia grassularioides ); Braga Filho et al. 2001, Leopoldo Bulhões, GO in Salacia crassifolia (Mart.) Peyr ; Deus et al. 2010 (Pedra Branca do Amapari, AP, ex Anastrepha atrigona in Geissospermum argenteum ); Silva et al. 2011 (Laranjal do Jari, ex Anastrepha atrigona in Geissospermum argenteum ; ex A. fraterculus in Psidium guajava ; Vitória do Jari, AP, ex A. atrigona in Geissospermum argenteum ; ex A. striata in Psidium guajava ).

Diagnosi s. Differs from D. areolatus in having the fore wing infumate, with a broad rounded hyaline spot from immediately after the stigma to the middle of R1a and not reaching the posterior margin of the fore wing, ending at the middle of the second subdiscal cell (2nd disc) ( Fig 3B ). Stigma dark brown ( Fig 3B ). In D. whartoni sp. nov. the fore wing has a hyaline spot in the anal area and a hyaline stripe in the central portion, and the stigma is yellow ( Fig 3A ). Hind legs with chromatic variations (orange, orange-brown, dark brown to black) on the entire leg or on part of it ( Fig 1C 2, D2, E2 ). Tegula yellowish orange as in D. whartoni sp. nov. (dark brown in D. areolatus ). Propodeum with one short basal transverse keel projecting laterally in the anterior portion of the areola reaching the longitudinal lateral keel, as in D. whartoni sp. nov. ( Fig 5D ). Ovipositor apex similar to D. areolatus , with a dorsal node and ventral serrations ( Fig 5G ). Ventral serrations morphologically similar to those of D. areolatus . In D. whartoni sp. nov., the serrations are more conspicuous and distinct ( Fig 4E ).

Description Female. Length of body, excluding ovipositor 5.5–6.3 mm.

Head. 1.4–1.2× wider than long; 1.5–1.3× wider than width of mesoscutum. Face polished, bright, and distinctly setose; midridge smoother, restricted between toruli ( Fig 5A ); distance between toruli equal to distance from toruli to eye ( Fig 5A ). Antenna longer than body, 8.4–8.7 mm in length, with 53 to 55 flagellomeres, first flagellomeres about 1.1–1.2× longer than second; 1.5–2.1× longer than wide. Eye large, 1.2–1.3× wider than high ( Fig 5C ); in dorsal view, eye 1.6–2.0× wider than temples; malar space 0.4–0.5× longer than height of eyes. Clypeus 2.6–3.9× wider than high, polished, slightly convex and lateral margin slightly curved with sparse setae two to three times longer than setae on face ( Fig 5A ); clypeus sinuate, similar to D. whartoni sp. nov., protruding as lobe medially on ventral margin; labrum partially covered by clypeus; distinct opening between clypeus and mandibles ( Fig 5A ).

Mesosoma. 1.2–1.5× longer than high; 1.7–2.0× longer than wide; 1.3–1.5× higher than wide; pronotum not visible dorsally; median lobe of mesoscutum and lateral lobes as in D. whartoni sp. nov., polished, shiny with few sparse setae; margins of lateral lobes with few setae and shorter than D. whartoni sp. nov. ( Fig 5B ); notaulus smooth, complete, deeper anterior to margin of mesoscutum and later shallower, reunited in broad and polished impression without midpit ( Fig 5B ); scutellar groove divided into two large pits by median longitudinal septum; scutellum smooth with few or weak punctures, few setae on margins, robust or wide in apical portion, similar to D. areolatus ( Fig 5B ); mesopleuron smooth; propodeum setose with median anterior basal keel (0.11–0.15) and complete posterior areola. As in D. whartoni sp. nov., laterally, in the anterior region of the areola, one short basal transverse keel extends to reach the lateral longitudinal keel, which is prominent and distinctly curved, from the posterior half of the propodeum ( Fig 5D ).

Wings. Fore wing 5.3–6.3 mm length; stigma wide, 3.0–4.2× longer than wide, with vein r slightly projecting from midpoint; (RS+M)a slightly sinuate posteriorly, 1.0–1.6× longer than 3RSa; 2RS 0.7–1.3× longer than 3RSa, 1.2–1.5× longer than 1m-cu and 1.6–2.6× longer than r-m; 3RSa 1.9–3.0× longer than r; 3RSb ending almost at wing tip; 2M 1.6–1.9× longer than 3RSa; (RS+M)b absent; 1cu-a straight line separated from 1M by 0.17–0.30 in length. Fore wing with a rounded hyaline spot that begins from immediately after stigma and ends in the middle of R1a, and before the wing posterior margin at second subdiscal cell (2nd disc). Length of hind wing 3.4–5.0 mm with m-cu, curved, distinctly pigmented just beyond the half toward to the wing margin.

Metasoma. 1.2–2.3× longer than wide and 1.0–2.3× wider than high; T1 length 0.9–1.0 mm greater than apex width; metasoma not sculptured, smooth and bright; T1 apex about 1.2–1.4× width of base; T1 with two dorsal keels developed at the base that gradually decrease from the posterior half, becoming weak or indistinct ( Fig 5E ); spiracles in half of T1 modified in two oval structures that protrude from the smooth surface, with or without setae ( Fig 5E, F ); ovipositor about 5.5–6.3 mm long; ovipositor tip with a dorsal node and ventral serrations that are less developed than in D. whartoni sp. nov. ( Fig 5G ); ovipositor sheath twice metasoma length, with 4–5 rows of setae ( Fig 5H ).

General coloration. Yellowish orange; first and second pair of legs bright yellow, third pair quite variable, may have all segments with blackish or dark-brown spots ( Fig 1C 2 ); tibiae and tarsi brown ( Fig 1D 2 ) or coxa, trochanters, and femur yellow ( Fig 1E 2 ); tegula yellowish orange; ovipositor sheath and antenna dark brown; apices of mandibles black; T2, T3, and T 4 may have black stripes; wings infumate; fore wing with broad rounded hyaline spot that begins from immediately after stigma to middle of R1a and in middle of second subdiscal cell (2nd disc). Veins and setae yellow in the hyaline spot; stigma, veins, and bristles dark brown in infumate area ( Fig 3B ).

Male. Similar to females, but usually last tergite with dark-brown or black spots. Head in dorsal view 1.7× wider than mesoscutum width, 2.0× wider than face; in dorsal view, eye 1.85× wider than temple; face 1.5–1.7× wider than high; malar space 0.42–0.44× height of eyes; clypeus 2.9–3.2× wider than high; antennae with 50 to 51 flagellomeres; first flagellomere 1.1-1.2× longer than second, 1.8–1.9× longer than wide; mesosoma 1.9× longer than wide, 1.3–1.4× higher than wide; metasoma 2.4–2.7× longer than wide, 1.1–1.4 wider than high.

Type material. Holotype. Female (DCBU 270198), BRAZIL: Amapá, Serra do Navio, 00°55’57.0”N, and 051°55’14.8”W 01.ii.2006, reared from fruit fly larva in fruits of guava ( Psidium guajava ), collr. R. A. Silva. Paratypes, with same data as holotype, 2 females (DCBU 270199, DCBU 270200), 1 male (DCBU 270201), collr. R. A. Silva and 1 female (ESALQ); 1 male, Tocantins, 28.ii.2005, reared from fruit fly larva Anastrepha coronilli in “Goiaba-de-anta” ( Bellucia grossularioides L.), collr. M. A. Uchôa-Fernandes; 2 females, 1 male, Goiás, Leopoldo Bulhões, 14. ix. 1999, reared from fruit fly larva Anastrepha spp. In “bacupari” ( Salacia crassifolia ) collr. V.R.S Veloso, (ESALQ).

Etymology. This species is named after Ricardo Adaime, who has provided valuable information of many Anastrepha species, their host fruits, and parasitoids in Amapá, Brazil.

GenBank accession numbers. Doryctobracon adaimei sp. nov. Amapá, FJ560535 (ITS2) ; Tocantins, FJ560536 (ITS2) ; Goiás, FJ560537 (ITS2) ; Amapá, FJ560543 (28SD2) ; Tocantins, FJ560544 (28SD2) ; Goiás, FJ560545 (28SD2).

Morphometry. Geometric morphometry generated 36 relative deformation measurements (k=2n–4), where k represents the total number of relative deformations, and n the number of anatomical markers (20). The results of the multivariate analysis (CVA) of populations of male and female D. areolatus , D. whartoni sp. nov. and D. adaimei sp. nov. were statistically significant, Wilks’ Lambda (p <0.0001), Pillai’s Trace (p <0.0001), Hotelling- Lawley Trace (p <0.0001), and Roy’s Greatest Root (p <0.0001). The first two canonical axes explained 66.0% and 15.7% of the data variability, respectively. In the scatter plot, we observed the complete separation of the population groups in the space of canonical variables CV1 and CV2 ( Fig 6 ). Specimens termed D. whartoni sp. nov., collected only in the state of Amapá, formed an isolated group of D. adaimei sp. nov. and of D. areolatus . Doryctobracon adaimei sp. nov. separated completely from the groups formed by the other species and revealed total overlap of their populations collected in the states of Goiás and Amapá; among the populations of D. areolatus , there was also the formation of distinct grouping with D. whartoni sp. nov. and D. adaimei sp. nov., and total overlap of their populations in the states of Amapá, Tocantins, Goiás and São Paulo ( Fig 6 ). The Mahalanobis distance matrix revealed a lower degree of morphological similarity for D. whartoni sp. nov. in relation to the distinct populations of D. areolatus and D. adaimei sp. nov. We also observed the same result for population of D. adaimei sp. nov. in relation to the populations of D. areolatus ; however, the Mahalanobis distances between samples of D. whartoni sp. nov. and D. adaimei sp. nov. were fairly wide, i.e. D. whartoni sp. nov. (AP) - D. adaimei sp. nov. (AP) = 38.39%, and D. whartoni sp. nov. (AP) - D. adaimei sp. nov. (GO) = 34.96% ( Fig 7 , Table 2).

Molecular markers. We detected intraspecific size variation for both molecular markers tested when comparing populations of D. areolatus from different geographical regions. The size variation was larger for the ITS2, which ranged from 564 bp for D. areolatus from the state of Amapá to 590 bp for specimens from the state of São Paulo. ITS2 was 587 bp long in samples from the states of Goiás and Tocantins. The size of 28S-D2 from samples of D. areolatus differed by no more than 5 bp in length, ranging from 381 bp for specimens from Amapá to 386 bp for those from Goiás. Specimens from Tocantins and São Paulo had intermediate sizes (384 and 385 bp, respectively). Despite the variation in size, these markers shared very high sequence similarities ( Table 3). However, no size variation was observed for both markers for the species identified as D. whartoni sp. nov. and D. adaimei sp. nov. ITS2 for D. whartoni sp. nov. from Amapá was 585 bp long, while ITS2 for D. adaimei sp. nov. from Amapá, Tocantins, and Goiás was 567 bp in length. The 28S-D2 of D. whartoni sp. nov. from Amapá was 385 bp long, and 386 (Amapá and Tocantins) and 387 bp long (Goiás) for specimens of D. adaimei sp. nov. Pairwise sequence similarities of ITS2 and 28S-D2 among D. whartoni sp. nov., D. adaimei sp. nov., and D. areolatus were very low ( Table 3).

Phylogenetic reconstruction of specimens of Doryctobracon species based on the isolated and concatenated sequences of the molecular markers ITS2 and 28S-D2, using maximum likelihood (ML) and neighbor-joining (NJ) methods, produced trees sharing similar topologies and well-defined clades ( Fig 8 ). In all cases, molecular analysis consistently indicated variation in D. areolatus from different collection sites (Amapá, Goiás, Tocantins, and São Paulo), with specimens from Amapá being the most divergent. The most variation in clade definition between the ML and NJ methods was found for the positioning of D. areolatus from Amapá and D. whartoni sp. nov. ( Fig 8 ). Nevertheless, the topologies produced with the concatenated sequences left no doubts regarding the external grouping of specimens of D. areolatus from Amapá from a more internal clade of D. areolatus from the other states. In the concatenated analysis, D. whartoni sp. nov. resolved in a subclade with the divergent D. areolatus from Amapá, but was positioned on a long branch away from D. areolatus ( Fig 8C ). The remaining samples of D. areolatus grouped together in a more internal subclade ( Fig 8 ). Samples of D. adaimei sp. nov. always resolved in a defined clade, in the analysis with either the isolated or concatenated molecular markers. In the analysis with the concatenated sequences, the clade of D. adaimei sp. nov. was the first to branch out from all of the remaining samples. The NJ analysis resolved specimens belonging to D. adaimei sp. nov. from Amapá and Tocantins in a more internal subclade from specimens from Goiás, while ML placed specimens from all three populations in a single clade separated from each other by short branches ( Fig 8 ).

Figures

FIGURE 1. Chromatic variation on the wings and legs. Doryctobracon areolatus (A-A2), Doryctobracon whartoni sp. nov. (yellow stigma – YS) (B-B2), Doryctobracon adaimei sp. nov. (Goiás) (C-C2), D. adaimei sp. nov. (Tocantins) (D-D2), D. adaimei sp. nov. (Amapá) (brown stigma – BS) (E-E2).(the figures are not on the same scale).

FIGURE 3. Fore wing. (A) Doryctobracon whartoni sp. nov. (yellow stigma); (B) Doryctobracon adaimei sp. nov. (brown stigma).

FIGURE 4. Doryctobracon whartoni sp. nov. head, front view (A), mesosoma, smooth notaulices, dorsal view (B), head and mesosoma, smooth mesopleura (C), setaceous propodeum, dorsal view (D), ovipositor apex (E), pattern of ovipositor sheath bristles (F).

FIGURE 5. Doryctobracon adaimei sp. nov. head, front view (A), mesosoma smooth notaulices, dorsal view (B), head and mesosoma smooth mesopleura (C), propodeum, dorsal view (D), dorsal view of the petiole (E), modified spiracle (F), ovipositor apex (G), pattern of ovipositor sheath bristles (H).

FIGURE 6. Dispersion graphic of Doryctobracon areolatus males and females from Amapá (DAAP), Tocantins (DATO), Goiás (DAGO) and São Paulo (DASP); Doryctobracon whartoni sp. nov. yellow stigma, Amapá (YSAP); Doryctobracon adaimei sp. nov. brown stigma, Amapá (BSAP) and Goiás (BSGO) in the bidimensional space of canonical variables VC1 and VC2.The deformations diagrams indicate the presumable wing conformations for individuals in the superior and inferior ends of the canonical variables. Deformation magnitudes were amplified 3x for visualization.

FIGURE 7. Comparison concatenated tree ITS2 – 28S rDNA D2 (UPGMA) produced from the nucleotide sequences (A), cluster analysis of the Mahalanobis distance (UPGMA) calculated from the shape of the wings components (B). (DAAP = Doryctobracon areolatus from Amapá, DAGO = D. areolatus from Goiás, DATO = D. areolatus from Tocantins, DASP = D. areolatus from São Paulo, YSAP = Doryctobracon whartoni sp. nov. (yellow stigma) from Amapá; BSAP = Doryctobracon adaimei sp. nov. (brown stigma) from Amapá, BSGO = D. adaimei sp. nov. from Goiás, BSTO = D. adaimei sp. nov. from Tocantins.

FIGURE 8. Phylogeny reconstruction for Doryctobracon species inferred by using the maximum likelihood and Neighborjoining methods using sequences from the molecular markers ITS2 (A), 28S-D2 (B) and their concatenated sequences (C).ML tree with the highest log likelihood (-1065.5986) and the optimal NJ tree with the sum of branch length (=0.13197667) shown for the ITS2 sequences were based on the Tamura 3-parameter model (Tamura, 1992); ML tree with the highest log likelihood (-719.5043) and the optimal NJ tree with the sum of branch length (=0.11870754) shown for the 28S-D2 sequences were based on the Tamura 3-parameter model; the ML tree with the highest log likelihood (-1598.9138) and the optimal NJ tree with the sum of branch length (=0.076130) for the concatenated ITS2 and 28S-D2 sequences was based, respectively, on the Tamura 3- parameter model+G (=0.1096)+I (=0.001%) and Tamura 3-parameter model+G (=0.1).

FIGURE2. Landmarks on fore wing of Doryctobracon areolatus.1. base of veincostal; 2. Intersection of the parastigma, 1RS and wing margin; 3. Intersection of the end of stigma and R1a; 4. Intersection of the radial sector 3RSb and wing margin; 5. Intersection of vein 3M and wing margin; 6. Intersection of vein 3CU and wing margin; 7. Intersection of vein 3-1A and wing margin; 8. Intersection of the veins 3-1A and 2cu-a; 9. Intersection of the veins 2CUa, 3CU and 2cu-a; 10. Intersection of the veins 1-1A, 1cu-a and 2-1A; 11. Intersection of the veins 1-1A and base of wing; 12. Intersection of the veins M+CU, 1M and 1CU; 13. Intersection of the veins M+CU, 1cu-a and 1CU; 14 Intersection of the veins 2CUa, 1CU and 1m-cu; 15. Intersection of the veins 2M, r-m and 3M; 16. Intersection of the veins 1m-cu, (RS+M)a, 2RS and 2M; 17. Intersection of the veins 1M, (RS+M)a and 1RS; 18. Intersection of the veins 3RSa, 3RSb and r-m; 19. Intersection of the veins 3RSa, r and 2RS; 20. Intersectionof the vein r and base of stigma (venational terminologyaccording to Sharkey & Wharton 1997).