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Article << Previous     |     Next >>   Contents Vol 26(6)

DNA barcoding and morphological identification of Argentine species of Apanteles (Hymenoptera : Braconidae), parasitoids of cactus-feeding moths (Lepidoptera : Pyralidae : Phycitinae), with description of a new species

Juan José Martínez A F, Carolina Berta B, Laura Varone C, Guillermo Logarzo C, Paula Zamudio B D, Alejandro Zaldívar-Riverón E and R. Gabriela Aguilar-Velasco E

A CONICET–División Entomología, Museo Argentino de Ciencias Naturales ‘Bernardino Rivadavia’, Ángel Gallardo 470, C1405DJR, Buenos Aires, Argentina.
B Instituto Superior de Entomología ‘Dr. Abraham Willink’, Universidad Nacional de Tucumán y Fundación Miguel Lillo, San Miguel de Tucumán, Tucumán, Argentina.
C USDA/ARS/South American Biological Control Laboratory, Hurlingham, Buenos Aires, Argentina.
D Departamento de Ciencias Básicas y Tecnológicas, Universidad Nacional de Chilecito, La Rioja, Argentina.
E Colección Nacional de Insectos, Instituto de Biología, Universidad Nacional Autónoma de México, México DF, México.
F Corresponding author. Email: jjmartinez@macn.gov.ar



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Invertebrate Systematics 26(6) 435-444 http://dx.doi.org/10.1071/IS12060
Submitted: 8 August 2012  Accepted: 6 September 2012   Published: 19 December 2012


Abstract

The gregarious species of Apanteles that attack cactus-feeding phycitine moths (Lepidoptera : Pyralidae) from Argentina are identified using DNA barcodes and morphological data. Sequences of specimens from 10 different populations were generated. Corrected genetic divergences showed two clusters of specimens separated by COI divergences higher than 6%. Our morphological examinations were congruent with the molecular evidence and therefore two species were confidently identified: Apanteles alexanderi Brèthes and a new species, Apanteles opuntiarum Martínez & Berta, sp. nov., which we describe and illustrate. Despite these two taxa being considered to represent a single polyphagous species for at least 50 years, they can be easily distinguished by the length of their ovipositor sheaths. A key to the species of microgastrine parasitoids of cactus-feeding Lepidoptera in South America is provided. The recognition of the new species reveals a narrower host range for the species involved in this system, making it necessary to re-evaluate their potential as biocontrol agents of the prickly pear moth.

Additional keywords: Apanteles alexanderi, Apanteles opuntiarum, biological control, Cactoblastis cactorum, COI, Opuntia, prickly pear.



Introduction

The parasitoid wasp genus Apanteles is one of the largest genera of Microgastrinae (Hymenoptera : Braconidae), currently containing ~1000 described species (Yu et al. 2005). Accurate identification of Apanteles species is very difficult even for specialists, and currently there are no keys for the Neotropical Region (Whitfield 1997). Apanteles species are endoparasitoids, mostly of microlepidoptera (Whitfield 1997), with one specialised species group containing gregarious endoparasitoids of cactus-feeding moths (Lepidoptera : Pyralidae : Phycitinae) in the New World. Only one species of this group has been described for the Neotropical Region, Apanteles alexanderi Brèthes. This species has been associated with many cactus-feeding moth species, including the prickly pear moth, Cactoblastis cactorum (Berg) (Lepidoptera : Pyralidae) (Pemberton and Cordo 2001).

The introduction of Cactoblastis cactorum into Australia in the early 20th century to control prickly pear (Opuntia spp.) represents one of the most successful and remarkable cases in classical biological control of weeds (Dodd 1940; Moran and Zimmermann 1984; Zimmermann et al. 2000). The successful control of Opuntia encouraged the introduction of Cactoblastis into other countries, including some Caribbean islands (Simmonds and Bennett 1966). In 1989, the Caribbean populations of Cactoblastis reached the peninsula of Florida and since then the species has become a major concern for the conservation of native species of Opuntia in North America: Cactoblastis cactorum is spreading westwards at ~160 km per year along the Gulf of Mexico coast (Hight et al. 2002; Solis et al. 2004). More recently, this species has been recorded from Mexico on Isla Mujeres, a small island just 5 km off the coast of the Yucatan Peninsula, in the state of Quintana Roo (Zimmermann et al. 2007). This moth not only threatens to decimate the centre of Opuntia biodiversity in North America if it continues spreading westwards from its current range in eastern Louisiana, but might also become a serious pest affecting the Opuntia cultures that are widely consumed by Mexican people since prehispanic times.

During recent collecting trips conducted in Argentina to survey the parasitoid species associated with Cactoblastis in its native range, several gregarious specimens of Apanteles were obtained. All microgastrine species reared from Cactoblastis and other phycitine species on Opuntia during the 20th century were almost axiomatically assumed to be A. alexanderi without a thorough examination of the specimens. Zimmermann et al. (1979) reported that this species was responsible for 30% of larval mortality of C. cactorum, but because of its presumed polyphagy it has not been considered in biological control programs of the prickly pear moth. Nevertheless, we found that our recently collected specimens exhibited morphological variation, suggesting that more than one species is associated with phycitine moths in South America. The aim of this work was therefore to investigate the species boundaries among these specimens using DNA barcode sequences (Hebert et al. 2003), complemented by morphological comparison, and to describe any newly recognised species. The use of the COI marker to characterise economically important species has already shown its utility for identifing invasive insect species (e.g. Scheffer et al. 2006), host–parasitoid associations (e.g. Hrcek et al. 2011), as well as parasitoid wasp species employed in biological control of aphids and scale insects (e.g. Zhang et al. 2011; Derocles et al. 2012).


Material and methods

Extensive samplings were carried out from 2002 to 2012 in several localities along central and northern Argentina in order to survey the cactus-feeding phycitine moths and their parasitoids. Phycitine larvae were transported to the laboratory and fed with their original hosts. Once pupation occurred, parasitised larvae were placed in 5-mL plastic cups for emergence of adult parasitoids and were subsequently kept in 96% ethanol until they were processed for DNA sequencing. Specimens from more than 50 localities across 15 Argentine provinces were examined morphologically, including specimens obtained from various entomological collections. The examined material is listed alphabetically according to Argentine provinces.

Specimens from 10 different populations were selected to obtain barcode sequences to assess DNA sequence variation among populations (DNA voucher numbers CNIN 1106–25, GenBank accession numbers JX566772–90). DNA Barcode sequences (~650 bp of the cytochrome oxidase I (COI) mitochondrial DNA gene: Hebert et al. 2003) were generated using the same DNA extraction, amplification and sequencing protocols employed by Ceccarelli et al. (2012). Sequences from seven Costa Rican species belonging to the A. leuscostigmus complex (GenBank accession numbers DQ492267, DQ492269, DQ492272, DQ492274, DQ492275, DQ492283, DQ492286: Janzen et al. 2009) were included in the analysis for comparison. These species were chosen because our Argentine specimens are morphologically similar to A. lecuostigmus according to the available key to species (Muesebeck 1921). Corrected genetic divergences among the included sequences were calculated using the K2P distance model (Kimura 1980) and a Neighbour-Joining (NJ) tree was reconstructed with these distances using PAUP* ver. 4 (Swofford 2002).

Species descriptions are organised following the basic format of recent taxonomic descriptions of Apanteles species (Whitfield et al. 2001) in order to facilitate comparisons. Absolute measurements of body and fore wing length are given for primary types, followed by observed paratype variation in parentheses. Photographs were taken and edited using a Leica® Z16 APO-A stereoscopic microscope, a Leica® DFC295/DFC290 HD camera, and the Leica Application Suite® program. Scanning electronic microscope images were obtained using a PHILIPS xl 30 instrument. Specimens for SEM images were coated with gold–palladium with a Thermo VG Scientific SC7620 coater. Morphological terminology follows Sharkey and Wharton (1997), surface sculpture terminology follows Harris (1979), and illustrations and descriptions of wing veins follow Mason (1986). Material examined is deposited at Museo Argentino de Ciencias Naturales ‘Bernardino Rivadavia’, Buenos Aires, Argentina (MACN), Instituto y Fundación Miguel Lillo, San Miguel de Tucumán, Argentina (IFML) and Instituto de Biología de la Altura, Universidad Nacional de Jujuy, San Salvador de Jujuy, Argentina (INBIAL).


Results

Species boundaries using DNA barcodes

In total, 19 barcode sequences were generated for the specimens of Apanteles included in this work. A NJ tree based on corrected genetic distances among the included specimens and previously published sequences of the A. leucostigmus species complex is shown in Fig. 1. The sequenced specimens were collected from populations from most of the known geographic range of the Apanteles species group that attacks pyralid moths in Argentina (Fig. 2) and represent specimens covering all the observed morphological variation.


 
Fig. 1.  NJ tree showing corrected genetic distances among Apanteles alexanderi, Apanteles opuntiarum, sp. nov. and species belonging to the Apanteles leucostigmus species complex.
 
Click to zoom
 


 
Fig. 2.  Distributional records of populations of Apanteles alexanderi (squares) and Apanteles opuntiarum (circles). Arrows indicate populations from which DNA barcode sequences were obtained.
 
 

The NJ tree (Fig. 1) recovered two separate sequence clusters of Apanteles specimens originally assigned to A. alexanderi. This separation is congruent with the morphological variation observed among the two groups, demonstrating that two separate species were present, A. alexanderi and a new species that is described below, A. opuntiarum, sp. nov. Intraspecific genetic distances within A. alexanderi ranged from 0 to 0.98% (DNA voucher numbers CNIN 1114–1115, 1120–1122) whereas within A. opuntiarum, sp. nov, they varied from 0 to 0.16% (DNA voucher numbers CNIN 1106–1113, 1116–1119, 1123–1125). However, the interspecific variation between the above two species ranged from 6.8 to 8.1%, whereas variation among these two species and species of the A. leucostigmus complex ranged from 9.6 to 12.6%.


Taxonomy

Apanteles alexanderi Brèthes

(Figs 3–10)


 
Fig. 3.  Apanteles alexanderi, habitus of female in lateral view.
 
 


 
Figs 4–9.  Apanteles alexanderi. 4, 5, mesosoma in dorsal view; 6, 7, first and second metasomal median tergites in dorsolateral view; 8, hind leg and metasoma showing ovipositor sheaths in lateral view; 9, cocoon mass. Figs 4, 6: specimen from Blanchard’s collection collected in the type locality. Figs 5, 7: newly reared specimen. Arrows indicate rugosities on second metasomal tergite.
 
Click to zoom
 


 
Fig. 10.  Apanteles alexanderi, wings.
 
 

Apanteles (Pseudapanteles) alexanderi Brèthes, 1922: 19. Lectotype female (here designated), MACN, examined.

Material examined

Type series. Lectotype female (here designated): ARGENTINA, Buenos Aires, Carmen de Patagones, 23.i.1921, V. B. Alexander; Apanteles (Pseudapanteles) alexanderi Brèthes; Lectotype Apanteles (P.) alexanderi (Brethès) by P. Marsh ‘79 (MACN); Paralectotype, one female with the same data and on the same pin.

Other material examined. ARGENTINA. Buenos Aires: 4 females, Carmen de Patagones (one coated for SEM images), 17.i.1921, Blanchard col. (MACN). Catamarca: 2 females, Andalgalá, Mar. 1921 (MACN). Chaco: 8 females, 1 male, Puerto Tirol II, 27°21′49.7″S, 59°03′59.2″W, 12.xi.2010, Logarzo, Varone, from larvae of Tucumania sp. on O. cardiosperma (DNA voucher nos CNIN1120, CNIN1121, CNIN1122; GenBank accession nos JX566787–88, JX566790) (MACN, IFML). Formosa: 4 females, Laguna Yerma, 24°14′47.5″S, 61°14′01.2″W, 28.ii.2009, Varone, Logarzo, from larvae of Tucumania sp. on O. cardiosperma (DNA voucher nos CNIN1114, CNIN1115; GenBank accession nos JX566786, JX566789) (MACN, IFML); 6 females, 8 km N Pozo del Mortero, 24°21′42.2″S, 62°05′32.8″W, 28.ii.2009, Logarzo, Varone, from larvae of Tucumania sp. on O. anacantha (MACN, IFML). Jujuy: 2 females, 2 males, Pucará-Tilcara, 4.x.1990, E. Neder, on ‘oruga azul’ ex/Trichocereus pasacana (INBIAL). Santiago del Estero: 31 females, 6 males; no locality, 08.ii.1935, from larvae of Tucumania tapicola (MACN); 14 females, 17 males, Añatuya, Nov. 1933, from larvae of Tucumania tapicola (MACN).

Diagnosis

Apanteles alexanderi can be easily distinguished from other Apanteles species associated with cactus-feeding moths by the weak but distinct rugosity on the second metasomal tergite (Figs 6, 7).

Female (Fig. 3)

Body length (excluding ovipositor and sheaths) 2.9 mm (2.9–3.7 mm). Fore wing length 2.9 mm (2.9–3.8 mm).

Colour. General body colour black, except palpi which are whitish; antenna, tibia and tarsus of hind leg and ovipositor sheaths dark brown; fore and mid legs, trochanter, trochantellus and femur of hind leg and ovipositor yellowish brown. Laterotergites of anterior metasomal tergites dark brown. Tegula pale, light brown. Wings hyaline, most veins unpigmented except C-Sc-R, pterostigma, R1, r, 2RS, 2M, 1Cua-b, which are brownish. Pterostigma pigmented only marginally, pale translucent medially.

Head. Face shallowly punctate, wider than high. Malar space as short as, or shorter than, width of mandible base, with distinct malar suture. Vertex coarsely but indistinctly punctate, surface smoother between ocelli. Ocelli reddish, forming an obtuse angle, lateral ocelli more than 2 times as far from one another as either is from anterior ocellus. Antennae clearly shorter than body; first flagellomere 2.6 times longer than apically wide and about as long as second, distal flagellomeres 2.0 times as long as broad and with only a single rank of placodes.

Mesosoma (Figs 4, 5). Lateral areas of pronotum with two transverse slightly scrobiculate furrows, median area and dorsolateral and ventrolateral corners rugulose, otherwise smooth. Mesonotum densely and distinctly punctate, punctures becoming coarser in anterior and posterior courses of notauli. Scutoscutellar scrobe formed by 9–10 sometimes partially confluent pits. Scutellum triangular, largely smooth, except for hair punctures; lunules subsemicircular to subtriangular, entirely smooth and shining, delimited anteriorly by strongly scrobiculate curved furrows. Dorsolateral areas of propodeum weakly sculptured and densely setose, posterolateral areas rugulose and largely devoid of setae; areola well defined posteriorly, open anteriorly but indicated by convergent rugae; with transverse rugae or carinae on its inner surface. Hind tibiae on outer surface with scattered spines. Inner hind tibial spurs 1.4–1.5 times longer than outer and 0.4–0.5 times as long as hind basitarsus.

Wings. Fore wing (Fig. 10) with pterostigma 2.5–2.6 times longer than wide and mostly pale translucent, pigmented only on its margins. R1 ~0.8–0.9 times the length of pterostigma and 0.6 times the length between apex of pterostigma and end of 3RS fold. Vein r distinctly longer than 2Rs, 1.3–1.4 times its length and weakly curved, meeting 2RS at a distinct obtuse angle.

Metasoma. Tergite I approximately as long as broad, to only slightly longer, almost equally broad anteriorly as posteriorly, anteriorly smooth and rugose to rugulose posteromedially. Second tergite largely smooth, with a very weakly but distinctly sculptured curved area (Figs 6, 7), 3.1–3.2 times broader than medially long. Remaining tergites entirely smooth. Hypopygium about two-thirds as long as ovipositor sheaths, strongly acuminate apically, medially desclerotised over its entire length. Ovipositor sheaths (Fig. 8) about as long as metasoma and 1.4–1.5 times as long as hind tibia, weakly decurved and hairy over most of length.


Male

Similar to female but with slightly smaller body size, 2.3–3.2 mm long; with relatively longer antennae and first metasomal tergite slightly tapering apically.


Biology

Apanteles alexanderi has been recorded as parasitising several species of phycitine moths in South America, including C. cactorum. Almost all specimens assigned to this species during the current study were reared from larvae of the phycitine genus Tucumania, with a few exceptions for which there are no host associations. Larvae pupate in a whitish ovoid mass (Fig. 9). The previously reported associations of A. alexanderi with Cactoblastis doddi Heinrich, Salambona analamprella (Dyar), Plutella xylostella (Linné), Argyrotaenia loxonephes (Meyrick) and Argyrotaenia sphaleropa (Meyrick) (Pemberton and Cordo 2001) need to be revisited.


Distribution

Specimens of A. alexanderi studied here are distributed from central to northern Argentina (Fig. 2).


Comments

According to Brèthes (1922) the type series was collected in Carmen de Patagones, Argentina, in January 1921 and, based on his original labels, it was collected by V. B. Alexander. Later, Blanchard (1935) mentioned that he himself collected the type series sent to Brèthes for study. It is possible that both Blanchard and Alexander conducted the collecting trip together since both of them were involved in the survey of potential enemies of prickly pear in Argentina to be introduced into Australia. Fortunately, Blanchard kept some specimens collected in the same locality and probably during the same collecting trip in his personal collection. After Blanchard’s death, his collection (including these non-type specimens of A. alexanderi) was donated to the entomological collection of the Museo Argentino de Ciencias Naturales. These specimens, collected one week before the type series, were used for comparison with the new specimens applying methodologies which are unsuitable for type specimens (i.e. gold-palladium coating for SEM images). The types examined by Brèthes were labelled as lectotype and paralectotype by P. Marsh; however, to our knowledge the designation was never published.


Apanteles opuntiarum Martínez & Berta, sp. nov.

(Figs 11–16)


 
Fig. 11.  Apanteles opuntiarum, habitus of female in lateral view.
 
 


 
Figs 12–15.  Apanteles opuntiarum. 12, mesosoma in dorsal view; 13, first and second metasomal median tergites in dorsolateral view; 14, hind leg and metasoma showing ovipositor sheaths in lateral view; 15, cocoon mass. Arrow indicates the entirely smooth second metasomal tergite.
 
Click to zoom
 


 
Fig. 16.  Apanteles opuntiarum, wings.
 
 

Material examined

Holotype female. ARGENTINA, Corrientes, Rt Nac. 123, 32 km S San Salvador, 29°33′29.8″S, 57°30′07.2″W; 26.ii.2009; Varone, Logarzo, from C. cactorum larvae on O. elata var. elata (DNA voucher no. CNIN1107; GenBank accession no. JX566773) (MACN).

Paratypes. ARGENTINA. Catamarca: 2 females, Chumbicha, 28°51′25.6″S, 66°13′50.2″W, 27.viii.2007, Logarzo, Palottini, from larvae of C. cactorum on O. ficus-indica (DNA voucher nos CNIN1124, CNIN1125; GenBank accession nos JX566780–81) (MACN, IFML). Córdoba: 1 female, El Fortín, 31°57′45.3″S, 62°20′15.1″W, 03.ii.2008, Logarzo, Palottini, from larvae of C. cactorum on O. megapotamica (DNA voucher no. CNIN1123; GenBank accession no. JX566779) (MACN). Corrientes: 1 female, 1 male, same data as holotype (DNA voucher nos CNIN1106, CNIN1108; GenBank accession nos JX566772, JX566774) (MACN, IFML). Entre Ríos: 3 females, 9.1 km W Médanos, 33°23′09.8″S, 59°10′18.4″W, 24.ii.2009, Logarzo, Varone, from C. cactorum larvae on O. elata var. elata (DNA voucher nos CNIN1109, CNIN1110, CNIN1111; GenBank accession nos JX566775–76) (MACN, IFML); 1 female, Conquistadores, Rt 127, 30°36′01.7″S, 58°28′00.3″W, 25.ii.2009, Logarzo, Varone, from larvae of C. cactorum on O. elata var. elata (DNA voucher no. CNIN1116; GenBank accession no. JX566782) (MACN); 2 females, Nogoyá, 32°22′14.3″S, 59°37′34.0″W, 27.x.2009, Logarzo, Moore, from unidentified lepidopteran larvae on O. megapotamica (DNA voucher nos CNIN1118, CNIN1119; GenBank accession nos JX566784–85) (MACN, IFML). Salta: 1 female, 5 km NO El Carril, Rt 68, 25°02′58.5″S, 65°29′47.4″W, 15.xii.2008, Varone, Logarzo; from larvae of C. cactorum on O. ficus-indica amyclaea (DNA voucher no. CNIN1117; GenBank accession no. JX566783) (MACN). Tucumán: 2 females, near Concepción, 27°23′19.7″S, 65°35′45.3″W, 06.iii.1009, Logarzo, Varone, from larvae of C. cactorum on O. ficus-indica (DNA voucher nos CNIN1112, CNIN1113; GenBank accession nos JX566777–78) (MACN, IFML).

Other material examined. ARGENTINA. Catamarca: 6 females, 3 males, Chumbicha, 28°51′25.6″S, 66°13′50.2″W, 27.viii.2007, Logarzo, Palottini, from larvae of C. cactorum on O. ficus-indica (MACN). Córdoba: 4 females, Trompa de Elefante, 31°24′14″S, 64°35′26.9″W, 07.v.2009, Logarzo, Guala, from larvae of C. cactorum on O. megapotamica (MACN); 11 females, 2 males, Villa Quilino, 30°12′15″S, 64°28′32″W, 15–17.xii.2008, Varone, Logarzo, from larvae of C. cactorum on O. ficus-indica (MACN); 7 females, El Fortín, 31°57′45.3″S, 62°20′15.1″W, 03.ii.2008, Logarzo, Palottini, from larvae of C. cactorum on O. megapotamica (MACN); 16 females, 2 males, Santa Coloma, Tanti, 31°20′54″S, 64°31′56″W, 10.i.2008, Logarzo, from larvae of C. cactorum on O. ficus-indica amyclaea (MACN); 2 females, 2 males, between Athos Pampa and La Cumbrecita, 31°59′04″S, 64°40′15.3″W, 1062 m, 26.x.2008, Zamudio, from Cactoblastis sp. larvae on O. ficus-indica (IFML); 2 females, 2 males, Quilino, 30°12′54.8″S, 64°29′25.4″W, 416 m, 27.x.2008, Zamudio, from Cactoblastis sp. larvae from O. ficus-indica (IFML); 2 females, 2 males, Luyaba, 32°09.193′S, 65°03.761′W, 687 m, 15.iii.2011, Zamudio, from Cactoblastis sp. larvae, on O. ficus-indica (IFML). Corrientes: 20 females, 5 males, 32 km S San Salvador, Rt 123, 29°33′29.8″S, 57°30′07.2″W, 26.ii.2009, Varone, Logarzo, from larvae of C. cactorum on O. elata var. elata (MACN); 3 males, Rt Nac. 12 near Empedrado, 28°13′11″S, 58°43′37″W, 62 m, 3.v.2008, Zamudio, from Cactoblastis sp. larvae on O. discolor (IFML) Entre Ríos: 3 females, 2 males, Nogoyá, 32°22′14.3″S, 59°37′34″W, 27.x.2009, Logarzo, Moore, from unidentified lepidopteran larvae on O. megapotamica (MACN); 6 females, 17 km S Concordia, 31°33′10″S, 59°04′44.6″W, 10.xii.2008, Logarzo, Varone, from larvae of C. cactorum on O. cardiosperma (MACN); 27 females, 9 males, 9.1 km W Médanos, 33°23′09.8″S, 59°10′18.4″W, 24.ii.2009, Logarzo, Varone, from larvae of C. cactorum on O. elata var. elata (MACN); 10 females, 7 males, 2.2 km E El Pingo, 31°34′46.5″S, 59°52′16.7″W, 25.ii.2009, Logarzo, Varone, from larvae of C. cactorum on O. elata var. elata? (MACN); 10 females, 3 males, Conquistadores, Rt 127, 30°36′01.7″S, 58°28′00.3″W, 25.ii.2009, Logarzo, Varone, from larvae of C. cactorum on O. elata var. elata? (MACN); 18 females, 2 males, Diamante-Strobel, 19.vi.2003, Zamudio, Briz, from Cactoblastis sp. larvae on O. elata (IFML); 16 females, 1 male, Diamante, 32°3′43.7″S, 60°38′39.61″W, 27.iii–3.vi.2006, Campos, on C. cactorum (IFML). Jujuy: 6 females, 4 males, Rt9, 1 km S El Carmen, 24°23′37.7″S, 65°16′16.6″W, 15.xii.2008, Logarzo, Varone, from larvae of C. cactorum on O. ficus-indica (MACN); 7 females, 1 male, Purmamarca, 23°44′46.7″S, 65°28′10.1″W, 2225 m, 18.i.2012, Varone, Cuadra, from larvae of C. doddi on O. sulfurea (MACN); 2 females, 2 males, Los Alisos, Jan. 1991, Neder, Arce, from C. cactorum larvae (INBIAL-UNJu); 1 female, Hornillos, 19.vi.2010, Neder, from Cactoblastis sp. larvae on O. sulfurea (INBIAL-UNJu); 2 females, 2 males, near Tumbaya, 23°52′34.3″S, 65°27′12.3″W, 2031 m (INBIAL-UNJu); 2 females, 2 males, near Maimará, 23°39′40.6″S, 65°26′12.1″W, 2420 m (INBIAL-UNJu); 2 females, 2 males, near Purmamarca, 23°40′30″S, 65°26′40″W, 2369 m, 20–23.iii.2010, Zamudio, from Cactoblastis sp. larvae on O. sulfurea (IFML); 2 females, 2 males, El Carmen, A° Severino, 24°19′45.6″S, 65°14′58.7″W, 1196 m, 21.iii.2010, Zamudio, from Cactoblasitis sp. larvae on O. sulfurea (IFML). La Rioja: 7 females, 2 males, San Pedro, 28°39′49.6″S, 66°55′14″W, Varone, Logarzo, from larvae of C. cactorum on O. ficus-indica (MACN); 4 females, 5 males, Rt Nac 40 between Famatina and Chilecito, 29°00′05″S, 67°28′37″W, 1226 m, 10.iv.2008, Zamudio, from Cactoblastis sp. larvae on O. sulfurea (IFML). Salta: 1 female, Rt 9, 10 km SE Juramento, 25°12′27.6″S, 64°56′45.5″W, 13.xii.2008, Varone, Logarzo, from larvae of C. cactorum on O. ficus-indica (MACN); 6 females, 5 km NO El Caril Rt 68, 25°02′58.5″S, 65°29′47.4″W, 15.xii.2008, Varone, Logarzo, from larvae of C. cactorum on O. ficus-indica amyclaea (MACN); 5 females, 9 males, between Salta and Cafayate, Arroyo Ayuza, 25.viii.2002, Zamudio, from Cactoblastis sp. larvae on O. sulfurea (IFML). San Juan: 36 females, 10 males, Balde del Rosario, Rt Prov. 510, 30°18′58″S, 67°41′56″W, 1174 m, 11.iv.2008, Zamudio, from Cactoblastis sp. larvae on O. sulfurea (IFML). San Luis: 11 females, 25 km S Merlo, 24.xii.2008, Logarzo, from larvae of C. cactorum on O. ficus-indica (MACN). Santa Fe: 6 females, 6 males, 14 km NW El Trebol, 32°12′56.0″S, 61°41′19.2″W, 03.ii.2008, Logarzo, Palottini, from larvae of C. cactorum on O. ficus-indica amyclaea (MACN); 11 females, 2 males, Rt Nac. 11 between Las Garzas and Reconquista, 28°58′51″S, 59°34′52″W, 59 m, 4.v.2008, Zamudio, from Cactoblastis sp. larvae on O. elata (IFML); 5 females, 5 males, Sunchales, 30°54′49.1″S, 61°33′14.2″W, 61 m, 25.iii.2009, Zamudio, from Cactoblastis sp. larvae on O. elata (IFML); 5 females, 5 males, Curupaity, 30°23′38.7″S, 61°39′02.7″W, 93 m, 25.iii.2009, Zamudio, from Cactoblastis sp. larvae on O. ficus-indica (IFML). Santiago del Estero: 5 females, 1 male, El Virqui, 28°11′55.6″S, 65°28′06.7″W, 23.viii.2007, Logarzo, Palottini, from larvae of C. cactorum on O. ficus-indica (MACN); 4 females, 1 male, El Zanjón, 23.viii.2007, Logarzo, Palottini, from larvae of C. cactorum on O. ficus-indica (MACN); 8 females, 7 males, Rt Nac. 34, 2 Km east Vilmer, 27°47′25″S, 64°07′05″W, 175 m, 6.v.2008, Zamudio, from Cactoblastis sp. larvae on O. discolor (IFML); 8 females, 6 males, Robles, 27°46′30″S, 64°08′44″W, 178 m, 17.vii.2008, Zamudio, from Cactoblastis sp. larvae on O. ficus-indica (IFML); 2 females, 2 males, Vilmer, 27°45.967′S, 64°09.566′W, 190 m, 3.iv.2011, Zamudio, from C. cactorum larvae on O. ficus-indica (IFML). Tucumán: 6 females, near Concepción, 27°23′19.7″S, 65°35′45.3″W, 06.iii.2009, Logarzo, Varone, from larvae of C. cactorum on O. ficus-indica (MACN); 13 females, near La Cocha, 27°47′53.9″S, 65°35′17.3″W, 28.ii.2009, Logarzo, Varone, from larvae of C. cactorum on O. ficus-indica (MACN); 43 females, 46 males, Burruyacu, Gdor. Piedrabuena, 26°44′18.48″S, 64°38′26.57″W, 6.ii.2005, Zamudio, from C. cactorum larvae (IFML); 15 females, 15 males, Burruyacu, La Virginia, 17.xii.2002, Zamudio, from C. cactorum larvae on O. ficus-indica (IFML); 4 females, 3 males, Leales, Las Tusquitas, 27°7′0.46″S, 64°55′37.53″W, 7.vii.2007 from Cactoblastis sp. on O. ficus-indica (IFML); 29 females, 10 males, 11.i.–12.ii.2007, Zamudio (IFML); 3 females, Leales, Tres Pozos, 18.x.2006, Zamudio, from C. cactorum larvae (IFML).


Biology

Apanteles opuntiarum is a gregarious endoparasitoid of cactus-feeding moths, specially C. cactorum, with a single record from Cactoblastis doddi. Almost all specimens studied in this work were reared from larvae of Cactoblastis, except one sample which was reared from an unidentified lepidopteran larva. Larvae of Cactoblastis were collected from at least six species of Opuntia. Pupation occurs in ovoid yellowish cocoon masses that are slightly more compact than those of A. alexanderi (Fig. 15).


Distribution

Based on the specimens studied here, A. opuntiarum occurs in north and central Argentina (Fig. 2). However, this species is expected to occur more extensively within the distribution of C. cactorum in Argentina, Paraguay, Uruguay and southern Brazil.


Comments

The specimens chosen to be the type series of A. opuntiarum are those from which COI sequences were obtained.


Etymology

The specific epithet refers to the plant genus (Opuntia) where the host is commonly encountered, meaning ‘from the prickly pears’.

Key to Apanteles species parasitic on cactus-feeding moths in South America

  Ovipositor sheaths shorter than metasoma and about as long as hind tibia (Figs 11, 14), second metasomal tergite smooth and polished (Fig. 13) ....Apanteles opuntiarum Martínez & Berta, sp. nov.
Ovipositor sheaths about as long as metasoma and distinctly longer than hind tibia (Figs 3, 8), second metasomal tergite with weak but distinct rugosities (Figs 6, 7) ....A. alexanderi Brèthes


Discussion

According to the excellent review provided by Pemberton and Cordo (2001), A. alexanderi has been recorded as parasitising at least eight microlepidopteran species on Opuntia. Almost all specimens that were confidently identified in this work as A. alexanderi were reared from specimens of the genus Tucumania. The first mention of A. alexanderi associated with C. cactorum is that of Parker et al. (1953), who reported this association from Uruguay. Subsequent records mentioning this association (De Santis 1967; Mann 1969; Zimmermann et al. 1979) are species lists with no original information and are probably based on the first report. All specimens reared from Cactoblastis were identified as A. opuntiarum. These results suggest a strong host preference for Tucumania and Cactoblastis by A. alexanderi and A. opuntiarum, respectively.

Our morphological observations were entirely congruent with the information provided by the barcoding data, with ~6% interspecific and less than 1% intraspecific divergence for both species. The arbitrary limit of 2% divergence traditionally employed to separate ‘barcoding species’ (Hebert et al. 2003) thus seems to easily apply for this species complex.

The host range of both A. alexanderi and A. opuntiarum, as they relate to other cactus-feeding moths in South America, remains uncertain. The association of A. alexanderi with other phycitine moths needs to be confirmed. A recent study of what was thought to be a polyphagous species, Apanteles leucostigmus, revealed that it actually constitutes a complex of cryptic and oligophagous species associated with several hesperiid butterflies in tropical America (Janzen et al. 2009). Similar results have recently been obtained for the genus Anicetus (Hymenoptera : Encyrtidae) associated with scale insects (Hemiptera : Coccidae) (Zhang et al. 2011) and for tachinid parasitoid flies (Smith et al. 2006). It is possible that the complex of cactus-feeding Lepidoptera supports a similar and richer community of microgastrine parasitoids than has been previously assumed.

The finding of a new species, A opuntiarum, attacking the prickly pear moth, Cactoblastis cactorum, in Argentina is significant for our understanding of the potential of microgastrine species as possible control agents of C. cactorum outside its native distribution. A re-evaluation of the potential of these species to control the prickly pear moth where it has become a serious concern for endemic populations or economically important species of Opuntia is therefore strongly suggested.



Acknowledgements

We thank Robert Kula (National Museum of Natural History and Smithsonian Institution, Washington DC, USA) for the loan of type specimens of Nearctic species of Apanteles associated with cactus-feeding moths for comparisons. Grateful thanks are also extended to Fabián Tricárico (Museo Argentino de Ciencias Naturales, Buenos Aires) and Susana Guzmán Gómez (UNIBIO, Instituto de Biología, UNAM, Mexico) for their assistance in taking SEM micrographs and stereoscopic microscope photographs respectively. Thanks to Arabella Peard (USDA/ARS/South American Biological Control Laboratory) for her corrections to the English version of the manuscript. This work was partially financed by a grant given by the Agencia Nacional de Promoción Científica y Tecnológica, Argentina (PICT 2007/1238 – Préstamo BID) to JJM and by a grant given by the Programa de Cooperación Bilateral CONACyT-CONICET 2010 to JJM and AZR.


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