First record of Apanteles hemara (N.) on Leucinodes orbonalis Guenée and biodiversity of Hymenoptera parasitoids on Brinjal

Insect sampling

This study was carried out to record and assess the potential of the natural enemies associated with the L. orbonalis Guenée during two consecutive seasons (November 2021–October 2022) in vegetable fields located at ICAR-Indian Agricultural Research Institute (ICAR-IARI), New Delhi, India.

The weather parameters during the study period such as rainfall, minimum and maximum temperature and relative humidity were recorded from an agrometeorological observatory, Division of Agricultural Physics, IARI, New Delhi and are provided in Table S1.

To calculate the pest infestation rate (%) in brinjal, the field was divided into four quarters, and 15 plants per quarter were randomly checked. The pest incidence was observed at 7 days’ intervals, and infestation (%) was calculated according to the following equation.

(1) $$\mathbf{F}\mathbf{r}\mathbf{u}\mathbf{i}\mathbf{t}\mathbf{i}\mathbf{n}\mathbf{f}\mathbf{e}\mathbf{s}\mathbf{t}\mathbf{a}\mathbf{t}\mathbf{i}\mathbf{o}\mathbf{n}\left(\mathrm{\%}\right)={\displaystyle \frac{\mathrm{N}\mathrm{o}.\mathrm{o}\mathrm{f}\mathrm{i}\mathrm{n}\mathrm{f}\mathrm{e}\mathrm{s}\mathrm{t}\mathrm{e}\mathrm{d}\mathrm{f}\mathrm{r}\mathrm{u}\mathrm{i}\mathrm{t}\mathrm{s}}{\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{f}\mathrm{r}\mathrm{u}\mathrm{i}\mathrm{t}\mathrm{s}\mathrm{o}\mathrm{b}\mathrm{s}\mathrm{e}\mathrm{r}\mathrm{v}\mathrm{e}\mathrm{d}}\times 100.}$$

Infested brinjal fruits were collected, and the larvae were reared for the emergence of parasitoids under laboratory conditions of 25 ± 0.5 °C, relative humidity 65 ± 5%, and a photoperiod of 12 light: 12 Dark h. The larvae, after pupation were separated and kept in plastic containers until parasitoid emergence. The parasitoids that emerged were preserved in 70% alcohol and card mounted for taxonomic studies. The parasitism percentage during each month was calculated according to the following equation (Van Driesche, 1983).

(2) $$\mathbf{P}\mathbf{a}\mathbf{r}\mathbf{a}\mathbf{s}\mathbf{i}\mathbf{t}\mathbf{i}\mathbf{s}\mathbf{m}\left(\mathrm{\%}\right)={\displaystyle \frac{\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{p}\mathrm{a}\mathrm{r}\mathrm{a}\mathrm{s}\mathrm{i}\mathrm{t}\mathrm{o}\mathrm{i}\mathrm{d}\mathrm{s}\mathrm{e}\mathrm{m}\mathrm{e}\mathrm{r}\mathrm{g}\mathrm{e}\mathrm{d}\mathrm{f}\mathrm{r}\mathrm{o}\mathrm{m}\mathrm{l}\mathrm{a}\mathrm{r}\mathrm{v}\mathrm{a}\mathrm{e}\mathrm{o}\mathrm{r}\mathrm{p}\mathrm{u}\mathrm{p}\mathrm{a}}{\mathrm{T}\mathrm{o}\mathrm{t}\mathrm{a}\mathrm{l}\mathrm{N}\mathrm{o}.\mathrm{o}\mathrm{f}\mathrm{l}\mathrm{a}\mathrm{r}\mathrm{v}\mathrm{a}\mathrm{e}\mathrm{o}\mathrm{r}\mathrm{p}\mathrm{u}\mathrm{p}\mathrm{a}\mathrm{c}\mathrm{o}\mathrm{l}\mathrm{l}\mathrm{e}\mathrm{c}\mathrm{t}\mathrm{e}\mathrm{d}\mathrm{f}\mathrm{r}\mathrm{o}\mathrm{m}\mathrm{f}\mathrm{i}\mathrm{e}\mathrm{l}\mathrm{d}}\times 100.}$$

In addition, a randomized weekly survey was carried out throughout two seasons to study the presence of Hymenoptera parasitoids in brinjal utilizing a sweep net (consists of a net with a 35 cm diameter and 40 cm length held on a circular aluminum frame which is connected to aluminum handle of 80 cm in length) and yellow pan trap (consists of a shallow yellow tray, about 5 cm deep and 15 cm diameter).

Identification of parasitoids

From the collected specimens, large parasitoids were pinned and dry-preserved, while small ones were preserved in 70% ethanol. Parasitoids were identified with the help of Bouček (1951), Habu (1960), Stary (1975), Husain & Agarwal (1982), Greco (1997), Narendran, Galande & Mote (2001), Belokobylskij (2003), Jonathan (2006), Gibson (2009), Rousse, Villemant & Seyrig (2011), Rousse & Villemant (2012), Sheeba & Narendran (2013), Xu, Olmi & He (2013), Akhtar, Singh & Ramamurthy (2014), Ghafouri-Moghaddam, Lotfalizadeh & Rakhshani (2014), Amer, Zeya & Veenakumari (2016), Cao, LaSalle & Zhu (2017), Fernandez-Triana et al. (2017), Khalaim (2018), Ahmed et al. (2020), Zerova & Fursov (2020), Gull-e-Fareen et al. (2021), Talamas et al. (2021), Burks et al. (2022a). The morphological studies were carried out using a Leica S8AP0 stereo microscope and a LEICA M205 C stereozoom automountage microscope. Multi-focused montage images were taken using a LEICA MC190 HD digital camera attached to the LEICA M205 C stereozoom automountage microscope. The photographs and illustrations were processed with Adobe Photoshop CS5 and plates were then prepared. The morphological terminology and wing venation are based on Nixon (1965), Jonathan (2006), Gibson (2009), Sheeba & Narendran (2013). All the specimens have been deposited in the National Pusa Collection (NPC), ICAR-IARI, New Delhi, India.

Statistical analysis

The parasitism (%) was analyzed statistically, and the extent of parasitism (%) was also subjected to correlation analysis with brinjal fruit infestation (%) as well as with weather factors using Minitab^® Statistical Software (17.0).

Systematic study

The main diagnostic characteristics of five parasitic wasps that emerged from Brinjal fruit and shoot borer (L. orbonalis Guenée), as well as their hosts and distribution details, were highlighted as follows.

Apanteles hemara Nixon, 1965 (Hymenoptera: Braconidae) (Figs. 1A–1D)

Diagnosis: Body length 3 mm, Body colour black. Mandible brown, labrum orange testaceous, palpi pale yellow, clypeus dark brown and antennae black. Fore and mid legs yellowish orange, entire hind coxa densely punctate rugose; hind femur brown, yellow trochanter, hind tibia most often strikingly bicolor, yellow and brown on posterior with spurs white, hind tarsus infuscate, ovipositor yellow, ovipositor sheet dark brown. Wing hyaline, venation brown; pterostigma mostly brown. Head entirely densely setose. Clypeus ventral margin slightly concave. Face and clypeus moderately and shallowly punctate; antenna slightly shorter than body length, with 16 segments; OOL 1.5x POL. Mesosoma, mesoscutum and scutellum with relatively coarse and dense punctures (distance between punctures smaller than diameter of puncture). Propodeum with areola complete, propodeal areola strong, centrally smooth, apically pointed and basally petiolate to base of propodeum by two sub-median irregular carinas (Fig. 1D). Forewing with 2Rs more than twice shorter than r, Vein r length 1.7x vein 2RS, 1-R long, Vein 1-R length 1.2x pterostigma length. The areolet open (Fig. 1B). Legs hind coxae entirely punctate rugose. Hind femur slightly swollen. Metasoma T1 of metasoma much longer than wide with strong, longitudinal striation, its margins sub-parallel to strongly converging apically. Tergum 2 wider than long, short, transverse and apically widened. Tergum 3 longer than tergum 2 (Fig. 1C). Ovipositor sheath slightly shorter than tibia. Ovipositor large, usually slightly decurved and gradually tapered.

Material examined: 1♀ June; 2♀ July; 2♀, 1♂ August; 1♀ September 2022 emerged from L. orbonalis and 1♀ August; 2♀ September 2022 IARI, New Delhi, yellow pan trap on brinjal (HC).

Host records: Tebenna micalis (Choreutidae); Cnaphalocrocis trapezalis, Herpetogramma stultalis, Hydriris ornatalis, Omiodes indicatae, Spoladea recurvalis and Udea ferrugalis (Crambidae) (Fernandez-Triana et al., 2017), Hymenia fascialis, Pachyzancla stultalis (Pyralidae) (Nixon, 1965).

Distribution: Kenya, Madagascar, Cape Verde Islands, Mauritius, Saudi Arabia, Senegal, Republic of Congo, South Africa, Yemen, Australia, and India (new record established at New Delhi) (Fernandez-Triana et al., 2017).

Bracon greeni Ashmead 1896 (Hymenoptera: Braconidae) (Figs. 2A–2E)

Diagnosis: Body length 3.1 mm, body colour brownish-yellow. Disc of metasoma, extreme apex of second tergite and large dorsal blotches on third and fourth tergites black; wings hyaline, stigma and veins brown, ocellar triangle black (Figs. 2A, 2C). Head smooth, wider than long; antenna 24 segments, nearly as long as the body length. vertex rugose, anteriorly smooth, shiny posteriorly with setose; eyes glabrous, length of eye in dorsal view 2.2x temple; temple smooth, shiny; width of face 1.7x its height; clypeus smooth, intertentorial distance 1.8x tentorio-ocular distance (Fig. 2B). Mesosoma: pronotum and mesoscutum smooth, shiny with notauli weakly impressed, scutellum smooth, shiny; metanotum with anterior median carina; propleuron smooth, shiny with setose; propodeum smooth, shiny with setose laterally and posterior median longitudinal carina extending upto its middle; propodeal spiracle round, small and medially placed; fore wing vein 3-SR about 3.0x vein r (Fig. 2E). Metasoma oval and shagreen; second to fourth metasomal tergites subequal, remaining a little shorter. Large dorsal blotches on third and fourth tergites black; ovipositor nearly as long as metasoma (Fig. 2D).

Material examined: 2♀ November; 1♀ December 2021; 1♀ February 2022 emerged from L. orbonalis and 1♀ November; 2♀ December 2021 ICAR-IARI, New Delhi, yellow pan trap on brinjal (HC).

Host records: Adisura atkinsoni, Alcides affaber, Earias fabia, Heliothis obsolete, Rabila frontalis (Sheeba & Narendran, 2013) and L. orbonalis (Venkatraman, Gupta & Negi, 1948). In the present study, it emerged from L. orbonalis.

Distribution: India (Kerala, Uttar Pradesh), Bangladesh, China, and Sri Lanka.

Goryphus nursei (Cameron, 1907) (Hymenoptera: Ichneumonidae) (Figs. 3A–3F)

Diagnosis: Body length 8.2–10 mm, body colour dark orange except eighth to tenth flagellar segments, face and frons along the eye margin, apex of metasoma T1 and T5–T8 light yellow to white; T2–T4 entirely black. Legs in general reddish, except hind femur broadly at apex, whole of tibia and tarsus blackish; hind tibia with a subbasal white band (Figs. 3A, 3B). Head face and clypeus strongly punctate, clypeus tending to be smooth towards the apex, intertentorial distance 1.2x tentorio-ocular distance. Antennal scrobe shallow, smooth and shiny, temple broadly smooth and shiny, malar space weakly granulose. Frons below ocellar triangle weakly striate. Mandible thick with longitudinally striate with punctures in between the striae with teeth subequal, pointed, superior tooth 1.2x longest than the inferior. Mesosoma pronotum coarsely striate, Mesoscutum strongly punctate, striate along margins of each lobe. Scutellum, Metascutellum smooth and shiny (Fig. 3D). Mesopleuron rugose, tending to be wrinkled above, Propodeum largely reticulate, basad of basal carina wrinkled, both transverse carinae strong and complete, basal carina more or less straight, apical carina evenly and strongly arched, spiracle small and oval. Areolet of fore wing small, squares, about 3.0x as high as the width of bordering veins (Fig. 3F). Metasoma T1 short, about 1.3x as long as wide at apex, 2^nd and 3^rd tergite closely punctate; 4^th tergite less punctate. Ovipositor tip with distinct teeth ventrally (Fig. 3C).

Material examined: 1♀ December 2021; 2♀ February 2022 emerged from L. orbonalis and 1♀ November 2021; 1♀ January 2022, ICAR-IARI, New Delhi, yellow pan trap on brinjal (HC).

Host records: BFSB L. orbonalis (Alam et al., 2003).

Distribution: India (Bihar, Delhi, Gujarat, Haryana, Jharkhand, Maharashtra, Odisha, Punjab, Rajasthan, Tamil Nadu, Uttarakhand, and Uttar Pradesh), China and Pakistan (Jonathan, 2006).

Trathala flavoorbitalis (Cameron, 1907) (Hymenoptera: Ichneumonidae) (Figs. 4A–4F)

Diagnosis: Body length 7.2 mm, body colour brownish-yellow, Antenna scape and pedicel yellowish and flagellomere brownish, vertex (except ocelli area black) and around eye light yellow; notauli often well marked with yellow, tegula and scutellum yellow; legs yellow, hind tibia slightly infuscate basally and apically; wings hyaline, pterostigma brown; metasoma orange, tergites 1–2 and basal triangle on tergite III dark brown, sheath and ovipositor dark brown (Figs. 4A–4F). Head strongly constricted behind eyes. Face densely punctate, clypeus smooth, vertex and frons granulate, center of frons smooth. Temples short and slightly rounded; malar space 0.65 times as long as basal mandible width, occipital carina complete; mandible with equal size teeth (Fig. 4C); antenna filiform longer than head and mesosoma with 35 segments; scape length 1.3x pedicel length. POL 0.6x OOL. Mesonotum densely punctate-shagreened, scutellum and metanotum more sparsely punctate. Scutellum rounded without dorsal lateral carinae. Propodeum densely punctate-shagreened dorsally than laterally. Propodeal carination complete, area basalis small but distinct (Fig. 4D). Metasoma tergite I a little longer than tergite II and twice longer than tergite III. Tergite I almost smooth, slightly longitudinally strigose at apex. Tergite II three times longer than apically wide and longitudinally striated (Fig. 4F); ovipositor shorter than abdomen.

Material examined: 3♀, 1♂ November; 2♀ December 2021; 1♀, 1♂ January; 2♀, 1♂ February; 3♀, 2♂ March; 2♀, 1♂ June; 4♀, 2♂ July; 6♀, 2♂ August; 3♀, 1♂ September; 1♀, 1♂ October 2022 emerged from L. orbonalis and 2♀ July; 3♀ August 2022, ICAR-IARI, New Delhi, sweeping net, on brinjal (HC).

Host records: Seventy-eight host records, all Lepidoptera (Gelechoidea, Noctuoidea, Pyraloidea, Tineoidea and Tortricoidea) (Rousse, Villemant & Seyrig, 2011) brinjal shoot and fruit borer (Alam et al., 2003; Ranjith et al., 2020). In the present study, it emerged from L. orbonalis.

Distribution: Reunion, Madagascar (Rousse, Villemant & Seyrig, 2011). Widespread through Indo-Pacific and Eastern Oriental Areas (Rousse & Villemant, 2012).

Spalangia gemina Boucek, 1963 (Hymenoptera: Spalangiidae) (Figs. 5A–5E)

Diagnosis: Body length 3.2 mm, body colour dark. Legs dark except basal four tarsal segments yellow. Head capsule smooth and shiny with dense circular setiferous punctures; temple distinctly circular punctures. Gena with malar space distinctly shorter than eye height, scrobal depression with punctate-crenulate scrobes on either side of smooth and shiny interantennal region; the punctures separated by linear ridges and often multisided or more or less reticulate-rugose, gena densely punctate without malar sulcus, malar space about 0.8x eye height (Fig. 5B). Antenna with scape about 6.6x as long as greatest width, the inner and outer surfaces uniformly setose and strongly. Mesosoma pronotal collar in lateral view convex behind neck, with coarsely reticulate-rugose, except for a nearly triangular area close to the crenulate cross-line posteriorly; axillae smooth and shiny except for a few pinprick-like setiferous punctures. Scutellum smooth and shiny except for a few pinprick-like setiferous punctures laterally (Fig. 5D); mesopleuron with longitudinal carinae extending from subalar area ventrally over almost all of upper mesepimeron, subalar scrobe extending posteroventrally along transepisternal line, and epistern (Fig. 5C). Fore wing hyaline; bare behind submarginal vein (Fig. 5E). Propodeum with postspiracular sulcus; dorsal surface punctate-rugose anteriorly and posteriorly, sculptured, sometimes almost smooth; supracoxal; propodeal sides smooth and shiny. Metasoma smooth and shiny. Petiole about 1.7x as long as medial width; almost smooth to finely, transversely carinate between longitudinal carinae.

Material examined:1♀ June; 2♀ August; 1♀ September 2022 emerged from L. orbonalis and 2♀ July; 1♀ August 2022, ICAR-IARI, New Delhi, sweeping net trap on brinjal (HC).

Host records: Micropezidae, Noctuidae and Tortricidae (Lepidoptera) as primary hosts (Burks et al., 2022b) recorded on L. orbonalis (Murali et al., 2017).

Distribution: Afrotropical, Australasian, Oriental, and Neotropical region (Burks et al., 2022b).

Evaluation of parasitism extent by T. flavoorbitalis and other parasitoids on BFSB

During July and August 2022, T. flavoorbitalis showed higher parasitism of 15.55% and 18.46%, respectively. However, the average parasitism (%) of A. hemara, B. greeni, G. nursei, T. flavoorbitalis and S. gemina was 3.10%, 1.76%, 1.10%, 9.28% and 1.20% respectively. Throughout the study period, T. flavoorbitalis was the dominant parasitoid. Its parasitism in 1^st season peaked at 12% in November 2021, followed by a decline in December 2021. However, it experienced subsequent increases in February and March 2022. During the 2^nd season, its parasitism peaked at 18.50% in August 2022, and higher fruit infestation (%) was observed (Fig. S1). The parasitism (%) exhibited by B. greeni was 6.70% and 3.30% in November and December 2021, respectively; however, it was not recorded during the 2^nd season. The parasitism (%) of G. nursei was 3.30% and 2.90% in December and January, respectively but it was not observed during the 2^nd season (Fig. 6).

The higher parasitism (%) of S. gemina was 2.38% recorded in August 2022. Apanteles hemara, as a new parasitoid (Hymenoptera: Braconidae), was found to parasitizes on the larvae of BFSB. This species was recorded during the 2^nd season, and its higher parasitism (%) was 6.12% and 4.76% recorded in August and September, respectively (Fig. 6). The total parasitism (%) exhibited by all parasitoids, as well as the fruit infestation (%) reach their peaks in August (Fig. 7).

Correlation analysis of fruit infestation (%) and parasitism (%) under study conditions

Correlation analysis was performed to explore the interrelationship among the infestation (%) and parasitism (%) with the environmental parameters during the study period. A significant (p ≤ 0.01) strongly positive correlation (r = 0.7) was shown between fruit infestation (%) and total parasitism (%) of parasitoid species. Also, a significant (p ≤ 0.01) strongly positive correlation (r = 0.7) was recorded between infestation (%) and parasitism (%) of T. flavoorbitalis. Parasitism (%) of T. flavoorbitalis showed a significant (p ≤ 0.5) moderately negative correlation with rainfall (r = −0.4), moderately positive correlation with Tmin (r = 0.3), RH (r = 0.4) and not correlated with Tmax (r = −0.02). Fruit infestation (%) showed a significantly (p ≤ 0.1–0.5) moderately positive correlation with Tmax, (r = 0.3), Tmin (r = 0.5), RH (r = 0.4), and a moderate negative correlation with rainfall (r = −0.5). Total parasitism (%) showed a significant (p ≤ 0.5) moderately negative correlation with rainfall (r = −0.3), weakly positive correlation with Tmin (r = 0.2), moderately positive correlation with RH (r = 0.4) and not correlated with Tmax (r = −0.05).

Survey for study of associated Hymenoptera parasitoids

Parasitoids were collected weekly from brinjal during two seasons (November 2021–October 2022), utilizing a sweep net and yellow pan trap. The monthly distribution of different Hymenoptera parasitoid families (19 families) collected from brinjal is shown in (Fig. S2). In this study a total of 60 species were recorded, from which 48 were identified up to the species level, and 11 were identified up to the genus level. This provides novel insights into the distribution patterns of recently identified parasitoid species recorded on brinjal from Delhi. About 41.7% of these newly recorded species (25 out of the 60 species). The superfamily Chalcidoidea was the most dominant, followed by Ichneumonoidea, Platygastroidea and Ceraphronoidea. These 60 species belong to various families as follows: Braconidae (twelve species), Ichneumonidae (nine species), Chalcididae (seven species), Scelionidae (nine species), Pteromalidae (four species), Dryinidae (three species), two species each in the families (Diapriidae, Eulophidae, Eurytomidae, Figitidae, and Mymaridae) as well as one species was recorded in each of the families (Agonidae, Aphelinidae, Bethylidae, Ceraphronidae, Platygastridae, and Spalangiidae) (Table 1 and Figs. 8–11).